key: cord-033064-3b4jv1zb authors: larsen, reinhard title: sepsis und septischer schock date: 2016-06-14 journal: an&#x000e4;sthesie und intensivmedizin f&#x000fc;r die fachpflege doi: 10.1007/978-3-662-50444-4_66 sha: doc_id: 33064 cord_uid: 3b4jv1zb die sepsis ist eine lebensbedrohliche organfunktionsstörung aufgrund einer fehlregulierten reaktion des körpers auf eine infektion. hinweise sind ein abfall des systolischen blutdrucks auf unter 100 mm hg, bewusstseinsatörungen und ein anstieg der atemfrequenz auf über 22/min (qsofa-score) beim septischen schock, einer unterform der sepsis, muss die herz-kreislauf-funktion mit kardiovaskulären medikamenten und volumenersatz gestützt werden. das serumlaktat ist auf mehr als 2 mmol/l erhöht. trotz intensiver bemühungen ist die letalität der sepsis unverändert hoch. eine frühzeitige diagnosestellung und therapie ist für das überleben der patienten von entscheidender bedeutung. die sepsis-konsensuskonferenz von 2016 bewertet die sirs-kriterien als zu ungenau für die diagnose einer sepsis und empfiehlt daher, den begriff nicht mehr zu verwenden. daher wird der begriff in den aktuellen sepsis-definitionen von 2016 nicht mehr aufgeführt. sepsis ist eine systemische entzündungsreaktion des organismus auf eine infektion durch mikroorganismen (bakterien, viren, pilze, rickettsien, protozoen). entsprechend müssen bei einer sepsis mikroorganismen oder deren toxine im blut nachweisbar sein. da aber die mikroorganismen auch intermittierend in das blut gelangen, entgehen sie häufig dem nachweis (z. b. durch blutkulturen oder toxinnachweis) und die diagnose muss anhand klinischer zeichen gestellt werden. die sepsis ist folge einer komplexen generalisierten entzündungsreaktion durch eine infektion. sie führt zu lebensbedrohlichen organfunktionsstörungen und gehört zu den häufigsten todesursachen auf intensivstationen. trotz intensiver bemühungen ist die letalität der sepsis weiterhin unverändert hoch. eine frühzeitige diagnosestellung und therapie ist für das überleben der patienten von entscheidender bedeutung. zur früherkennung der sepsis wird als "schnelltest" der qsofa-score (q = quick) eingesetzt. der qsofa-score umfasst folgende 3 kriterien: surviving sepsis campaign: international guidelines for management of severe sepsis and septic shock the third international consensus definitions for sepsis and septic shock (sepsis-3) sepsis und mods. 5. aufl anwendungsbeschränkungen für hes. rote hand brief www leitlinie diagnostik und therapie der sepsis. www.awmf.org/leitlinien u key: cord-014658-oeuvelb1 authors: martin, greg s. title: optimal fluid management in sepsis date: 2019-11-07 journal: qatar med j doi: 10.5339/qmj.2019.qccc.40 sha: doc_id: 14658 cord_uid: oeuvelb1 sepsis clinically manifests as life-threatening organ dysfunction due to a dysregulated host response to infection.(1) optimal fluid resuscitation is relevant for all sepsis patients, and perhaps it is most important for those with septic shock. septic shock is defined as a subset of sepsis in which particularly profound circulatory, cellular, and metabolic abnormalities are associated with a greatest risk of mortality, and septic shock is clinically identified as sepsis patients with serum lactate level >2 mmol/l and who require vasopressor infusion to maintain a mean arterial pressure ≥ 65 mm hg in the absence of hypovolemia. sepsis is among the most common conditions in the intensive care unit (icu), accounting for up to half of all hospital deaths and being the third leading cause of death overall in the united states.(2) sepsis and septic shock are medical emergencies for which treatment and resuscitation should begin immediately. the goals of fluid resuscitation for these patients are: a) to rapidly replace intravascular volume and restore tissue perfusion, and b) to minimize organ dysfunction through timely interventions that either halt or reverse the physiologic derangements. if hypoperfusion is present, at least 30 ml/kg of iv crystalloid fluid should be given rapidly, and additional fluids should be guided by frequent reassessment of hemodynamic status, preferably using dynamic indices to indicate the likelihood of a beneficial response to fluid administration. fluid administration should be targeted to achieve a map of at least 65 mm hg, and to normalize lactate in patients with elevated lactate due to hypoperfusion.(3) balanced crystalloids are the fluid of first choice for sepsis resuscitation based on ready availability and taking medication costs into account. use of 0.9% saline compared to a balanced crystalloid, such as lactated ringer's or plasmalyte, produces more kidney dysfunction and with a greater risk of dying.(4) the individual side effect profiles may best differentiate the natural and synthetic colloids. albumin may be considered for administration to sepsis patients with refractory shock or who have received substantial amounts of crystalloid fluids, but should not be administered to patients with severe traumatic brain injury.(5) hydroxyethyl starch (hes) products should not be administered to patients with sepsis because of increased risk of acute kidney injury and death. gelatin solutions are not recommended in sepsis. norepinephrine is the vasopressor of first choice for patients with septic shock, and should be administered to achieve a mean arterial pressure of at least 65 mm hg after excluding hypovolemia as a cause for hypotension. the selection of a second line vasopressor, such as vasopressin, dopamine, phenylephrine, epinephrine or angiotensin-2, depends on patient factors such as underlying cardiac dysfunction, presence of arrhythmias, and current response to vasoconstrictor or inotropic agents. dopamine should not be used for renal perfusion or protection and it should be avoided in patients with tachyarrhythmias. sepsis and septic shock are medical emergencies for which treatment and resuscitation should begin immediately. the goals of fluid resuscitation for these patients are: a) to rapidly replace intravascular volume and restore tissue perfusion, and b) to minimize organ dysfunction through timely interventions that either halt or reverse the physiologic derangements. if hypoperfusion is present, at least 30 ml/kg of iv crystalloid fluid should be given rapidly, and additional fluids should be guided by frequent reassessment of hemodynamic status, preferably using dynamic indices to indicate the likelihood of a beneficial response to fluid administration. fluid administration should be targeted to achieve a map of at least 65 mm hg, and to normalize lactate in patients with elevated lactate due to hypoperfusion. 3 balanced crystalloids are the fluid of first choice for sepsis resuscitation based on ready availability and taking medication costs into account. use of 0.9% saline compared to a balanced crystalloid, such as lactated ringer's or plasmalyte, produces more kidney dysfunction and with a greater risk of dying. 4 the individual side effect profiles may best differentiate the natural and synthetic colloids. albumin may be considered for administration to sepsis patients with refractory shock or who have received substantial amounts of crystalloid fluids, but should not be administered to patients with severe traumatic brain injury. 5 hydroxyethyl starch (hes) products should not be administered to patients with sepsis because of increased risk of acute kidney injury and death. gelatin solutions are not recommended in sepsis. norepinephrine is the vasopressor of first choice for patients with septic shock, and should be administered to achieve a mean arterial pressure of at least 65 mm hg after excluding hypovolemia as a cause for hypotension. the selection of a second line vasopressor, such as vasopressin, dopamine, phenylephrine, epinephrine or angiotensin-2, depends on patient factors such as underlying cardiac dysfunction, presence of arrhythmias, and current response to vasoconstrictor or inotropic agents. dopamine should not be used for renal perfusion or protection and it should be avoided in patients with tachyarrhythmias. the third international consensus definitions for sepsis and septic shock (sepsis-3) incidence and trends of sepsis in us hospitals using clinical vs claims data surviving sepsis campaign: international guidelines for management of sepsis and septic shock: 2016 balanced crystalloids versus saline in critically ill adults evidence-based colloid use in the critically ill key: cord-018284-grvj99eh authors: fresenius, michael title: sirs, sepsis und multiorganversagen date: 2014-10-04 journal: repetitorium intensivmedizin doi: 10.1007/978-3-642-44933-8_24 sha: doc_id: 18284 cord_uid: grvj99eh nan unter sepsis versteht man die systemische inflammatorische antwort (sirs) auf eine vermutete oder nachgewiesene infektionsquelle bzw. eine überschießende aktivierung von primär protektiven defensivsystemen des septischen patienten im sinne einer »host defence failure disease«. neben veränderungen des immunsystems kommt es zu einer imbalance von prokoagulatorischen und antikoagulatorischen faktoren. durch lipopolysaccharidstimulation bilden die endothelzellen vermehrt »tissue factor«, wodurch die gerinnung aktiviert wird störung der mikrozirkulation durch bildung von mikrothromben mit ge-. abb. 24.1 pathogenese der sepsis: beeinflussung des zellulären und humoralen immunsystems . abb. 24.2 pathogenese der sepsis: beeinflussung der gerinnung fahr der/des mods/mov und störung der makrozirkulation aufgrund einer reduzierten kardialen pumpfunktion, eines erniedrigten systemvaskulären widerstandes und einer ausgeprägten hypovolämie infolge »capillary leak syndrome«. in der sepsis kommt es zu einer reduktion des protein-c-spiegels störung der fibrinolyse und thrombenbildung aufgrund einer fehlenden thrombin-thrombomodulin-stimulation (fehlender abbau von faktor v und viii; . abb. 24.2). nach einer initialen proinflammatorischen phase kommt es zu in einer gegenreaktion mit einer kompensatorischen antiinflammatorischen antwort (cars). diese antwort kann überschießend sein und zur immunsuppression bzw. anergie führen. der immunstatus kann anhand folgender faktoren bestimmt werden: 4 interleukin-6-spiegel 4 ausmaß der hla-dr-expression (immunstimulation durch gm-csf-gabe zur steigerung der immunantwort oder gabe von apc im rahmen der hyperfibrinolyse) 4 ex vivo tnf-anstieg nach lps-gabe verzicht auf intravenöse selen-gabe 2c verzicht auf die gabe von aktivierten protein c (rhapc) drotrecogin alfa (activated) for adults with severe sepsis and low risk of death empfehlungen zur diagnostik und therapie der schockformen der iag schock der divi critical illness polyneuropathie und criticall illness myopathy epidemiology of sepsis and infection in icu patients from international multicentre cohort study intravenous immunoglobuline for treating sepsis and septic shock. (cochrane review). the cochrane library of critical care medicine (accp/sccm) concensus conference (1992) definition for sepsis and multiple organ failure, and guidelines for the use of innovative therapies in sepsis selenium in intensive care: results of a prospective randomized, placebo-controlled, multicenter study in patients with severe systemic inflammatory response syndrome, sepsis and septic shock the riro concept: p is for predisposition effect of treatment with low doses of hydrocortisone and fludrocortisone on mortality in patients with septic shock clinical equipoise remains for issues of adrenocorticotropic hormone administration, cortisol testing, and therapeutic use of hydrocortisone effect of treatment with low doses of hydrocortisone and fludrocortisones on mortality in patients with septic shock ventilation with lower tidal volumes as compared with traditional tidal volumes for acute lung injury and the acute respiratory distress syndrome norepinephrine plus dobutamine versus epinephrine alone for management of septic shock: a randomised trial mikrozirkulatorisches monitoring in der sepsis efficacy and safety of recombinant human activated protein c for severe sepsis empfehlungen zur kalkulierten parenteralen initialtherapie bakterieller erkrankungen bei erwachsenen. update a controlled clinical trial of high-dose methylprednisolone in the treatment of severe sepsis and septic shock stress doses of hydrocortisone reverse hyperdynamic septic shock: a prospective, randomized, double-blind, single-centre study higher versus lower positive end-expiratory pressures in patients with acute respiratory distress syndrome diagnostic approach to sepsis -state of the art aktuelle aspekte und zukunft der sepsisdiagnose intensive insulin therapy and pentastarch resuscitation in severe sepsis diagnosis of sepsis: an evidence-based review effects of drotrecogin alfa activated on microcirculation alterations in patients with severe sepsis surviving sepsis campaign: international guidelines for management of severe sepsis and septic shock cardiovascular management of septic shock glucose control in critically ill patients effects of high dosis of selenium, as sodium selenit, in septic shock: a placebo-controlled, randomized, double-blind, phase ii study aktivated protein c (xigris) treatment in sepsis: d drug in trouble sepsis: state of the art crit care 16 r94 und annane djilllali crystmas study adds to concerns about renal safety and increased mortality in sepsis patients diagnostic value of procalcitonin, interleukin 6 and interleukin-8 in critically ill patients admitted with suspected sepsis the pathophysiology and treatment of sepsis immunologic and hemodynamic effects of »low-dose« hydrocortisone in septic shock: a doubleblind, randomized, placebo-controlled, crossover study high dose vasopressin is not superior to norepinephrine in septic shock immunmodulatory therapies in sepsis early antimicrobial therapy in severe sepsis and septic shock sccm/esicm/accp/ats/sis international sepsis definitions conference multiple-center, randomized, placebocontrolled, double-blind study of the nitric oxide synthase inhibitor 546c88: effect on survival in patients with septic shock the epidemiology of sepsis in the united states from 1979 through 2000 definition of sepsis ventilation strategy using low tidal volumes, recruitment maneuvers, and high positive end-expiratory pressure for acute lung injury and acute respiratory distress syndrome: a randomized controlled trial hydroxyethyl starch or saline for fluid resuscitation in intensive care the problems and challenges of immunotherapy in sepsis lactate in the critically ill patients: an outcome marker with the times terlipressin for norepinephrin-resistant septic shock beneficial effects of shortterm vasopressin infusion during severe septic shock hydroxyethyl starch 130/0.42 versus ringer's acetate in severe sepsis hydroxyethylstarch and ringer's lactate for fluid resuscitation in patients with severe sepsis -results from the visep-study diagnose und therapie der sepsis therapie und nachsorge der sepsis (2010) 1. revision der s-2k-leitlinien der deutschen sepsis-gesellschaft e.v. (dsg) und der deutschen interdisziplinären vereinigung der intensiv und notfallmedizin (divi) early goal-directed therapy in the treatment of severe sepsis and septic shock drug therapy: management of sepsis vasopressin versus norepinephrine infusion in patients with septic shock effects of hydroxyethyl starch and gelatine on renal function in severe sepsis: a multicentre randomised study blutkulturdiagnostik sepsis, endokarditis, katheterinfektionen, teil 1 deutscher anästhesiecongress pro &contra-sitzung: »selensubstitution bei sepsis?« münchen 16 hydrocortisone therapy for patients with septic shock guidelines for the management of severe sepsis and septic shock intensive insulin therapy in critically ill patients outcome benefit of intensive insulin therapy in the critically ill: insulin dose versus glycaemic control high-dose antithrombin iii in severe sepsis. a randomized controlled trial klinisches management bei patienten mit sepsis sepsis management -antibiotic therapy key: cord-006414-60lpjg09 authors: engelmann, l. title: die diagnose der sepsis date: 2006 journal: intensivmed notfallmed doi: 10.1007/s00390-006-0741-y sha: doc_id: 6414 cord_uid: 60lpjg09 the early diagnosis of sepsis is mandatory for the further reduction of mortality due to sepsis. current findings exist that accentuate the role of the time factor, comparable with acute myocardial infarction or with ischemic stroke. on the other hand, there are no generally accepted diagnostics for sepsis, realizing the demands of early diagnosis and based on the physician’s experience. the diagnostics start with the recognition of the inflammatory reaction caused by infection (at least 2 of 4 criteria of inflammatory reaction have to be fulfilled). this definition has high sensitivity, but remarkably lower specificity and it leads either to too frequent admissions or only to hospitalization in case of a complicating organ failure. making a careful history and knowledge about sepsis are essential for the out-patient department physicians. in addition to the varying pictures of sepsis, the clinicians have laboratory findings available, most of all procalcitonin. patients have to be considered as septic with a serum pct level higher than 1 ng/ml particularly when clinical signs do not exclude sepsis and in cases of positive blood cultures. initially pct is a product of macrophages if the defense reaction starts, but it becomes an infection marker, when the serum pct level declines less than the half life falls. the diagnosis of sepsis " summary the early diagnosis of sepsis is mandatory for the further reduction of mortality due to sepsis. current findings exist that accentuate the role of the time factor, comparable with acute myocardial infarction or with ischemic stroke. on the other hand, there are no generally accepted diagnostics for sepsis, realizing the demands of early diagnosis and based on the physician's experience. the diagnostics start with the recognition of the inflammatory reaction caused by infection (at least 2 of 4 criteria of inflammatory reaction have to be fulfilled). this definition has high sensitivity, but remarkably lower specificity and it leads either to too frequent admissions or only to hospitalization in case of a complicating organ failure. making a careful history and knowledge about sepsis are essential for the out-patient department physicians. in addition to the varying pictures of sepsis, the clinicians have laboratory findings available, most of all procalcitonin. patients have to be considered as septic with a serum pct level higher than 1 ng/ml particularly when clinical signs do not exclude sepsis and in cases of positive blood cultures. initially pct is a product of macrophages if the defense reaction starts, but it becomes an infection marker, when the serum pct level declines less than the half life falls. " key words sepsis -diagnosisinfection marker -procalcitonininflammatory response " zusammenfassung die frühe diagnostik der sepsis ist das nadelöhr für die weitere senkung der sepsissterblichkeit. inzwischen liegen befunde vor, dass, vergleichbar mit akutem myokardinfarkt und schlaganfall, dem faktor zeit eine wichtige rolle zukommt. demgegenüber gibt es keine allgemein akzeptierte sepsisdiagnostik, die der forderung nach früher diagnose und der unabhängigkeit vom erfahrungsstand des jeweiligen arztes rechnung tragen könnte. ausgangspunkt der diagnose ist die durch infektion hervorgerufene inflammatorische reaktion (mindestens 2 von 4 sirs-kriterien erfüllt), die eine hohe sensitivität, aber eine geringe spezifität besitzt. hier besteht das dilemma für die ambulant tätigen ärzte, die dann entweder zu häufig einweisen oder erst einweisen, wenn die sepsis zum organversagen geführt hat. der ambulante arzt kommt um das sorgfältige erheben der anamnese und das wissen der befunde bei sepsis nicht herum. ärzten von notaufnahmen und klinisch tätigen ärzten steht neben dem wissen um die variable klinik des septischen patienten vor allem der laborparameter procalcitonin zur verfügung. patienten mit einem pct ≥ 1 ng/ml sollten als septisch betrachtet werden, wenn die klinik die sepsis nicht ausschließt und erst recht dann, wenn positive blutkulturbefunde vorliegen. pct ist zunächst das produkt der makrophagen am be-ginn der abwehrreaktion, wird aber zum infektionsmarker, wenn der abfall des serum-pct nicht dessen halbwertzeit folgt. die von uns benutzte werteinterpretation für pct folgt dabei weniger irgendeiner statistik als tatsächlich der jahrelangen klinischen beobachtung (tab. 8). die sirs-kriterien als ausdruck der inflammation enthalten neben den mehr akut reagierenden variablen wie tachypnoe und tachkardie die weniger schnell reagierenden parameter temperatur und den laborparameter leukozytenzahl. in den deutschen kliniken wird von 90,1% der intensivmediziner das crp als sepsisparameter genutzt [4] . crp ist das produkt der durch zytokine stimulierten hepatozyten und somit das endergebnis der inflammationsreaktion, damit ein akut-phase-protein [48] . es kann dann als sepsisparameter genutzt werden, wenn die sepsis diagnostiziert, also die infektiöse ursache gesichert ist. damit unterscheidet es sich grundsätzlich vom pct, das die infektion beschreibt. sinkt aber das pct und nachfolgend die infektionsgetriggerte inflammation, dann fällt auch das crp. dessen halbwertzeit liegt bei 48 stunden [49] , damit ist crp ein eher träge reagierender parameter der inflammation [50] . patienten mit inflammationreaktion septischer und nicht-septischer genese unterscheiden sich nicht im crp-verlauf (abb. 4), während bei gesicherter sepsis dem crp-verhalten eine prognostische bedeutung zukommt (abb. 5; [41, 50] ) die inflammationsreaktion ist eine weitestgehend uniforme reaktion des organismus auf schädigung. initial steht die aktivierung des makrophagensystems, die u.a. über die produktion "proximaler" zytokine wie il-1 und tnf-alpha die inflammationsreaktion triggert und die die produktion "distaler" proinflammatorische zytokine (z. b. il-6, il-8) in allen zellen des körpers induziert. die proinflammatorische reaktion ist unerlässlich für die elimination der schädigung. der organismus wird aber versucht sein, die entzündungsreaktion durch gleichzeitige antiinflammation (z. b.il-10) zu steuern, um sich selbst zu erhalten (abb. 6). gelingt die kontrolle der inflammatorischen reaktion nicht, was von einer reihe von faktoren abhängt (stärke der schädigung, genetische prädisposition, immunstatus), führt die inflammationsreaktion zu immunparalyse, multiorganversagen und tod. vor diesem hintergrund kann die bestimmung ausgewählter "proximaler" sowie pro-und antiinflammatorischer "distaler" zytokine wichtige infor procalcitonin, soluble interleukin-2 receptor, and soluble e-selectin inpredicting the severity of acute pancreatitis predictive value of procalcitonin and interleukin-6 in critically ill patients with suspected sepsis serum procalcitonin and c-reactive protein levels as marker of bacterial infections: a systematic review and meta-analysis usefulness of procalcitonin serum levels for the diagnosis of bacteremia procalcitonin serum levels in tertian malaria usefulness of procalcitonin to differentiate typical from atypical community-aquired pneumonia usfulness of procalcitonin levels in community-aquired pneumonia according to the patients outcome research team pneumonia team severity index procalcitonin kinetics in the prognosis of severe communityaquired pneumonia potential use of procalcitonin as a diagnostic criterion in febrile neutropenia: experience from a multicentre study c-reaktives protein (crp) in: thomas l (hrsg) labor und diagnose, 5. aufl. th-books verlagsgesellschaft mbh spsis und sepsismarker-update comparison of procalcitonin (pct) and c-reactive protein (crp) plasma concentrations at different sofa scores during the course of sepsis and mods high circulating levels of interleukin-6 in patients with septic shock: evolution during sepsis, prognostic value, and interplay with other cytikines prognostic value of serum cyteokines in septic shock circulating interleukin-8 concentrations in patients with multiple organ failure of septic and nonseptic origin early dedection and markers of sepsis balance of inflammatory cytokines related to severity and mortality of murine sepsis pro-versus anti-inflammatory cytokine profile in patients with severe sepsis: a marker for prognosis and future therapeutic options circulating endithelin-1 and tumor necrosis factor-a: early predictors of mortality in patients with septic shock interleukin-10: a complex role in the pathogenesis of sepsis syndroms and ist potential as an anti-inflammatory drug hla-dr-expression auf monozyten bei peritonitis und sepsis -möglichkeiten eines therapeutischen ansatzes verbesserung der monozytenfunktion -ein therapeutischer ansatz? in: update in intensive care and emergency medicine bd 18 is low monocyte hla-dr expression helpful to predict outcome in severe sepsis? hla-dr as a marker for increased risk for systemic inflammation and septic complications after cardiac surgery key: cord-005697-l1zmrq4p authors: pène, frédéric; pickkers, peter; hotchkiss, richard s. title: is this critically ill patient immunocompromised? date: 2015-12-02 journal: intensive care med doi: 10.1007/s00134-015-4161-y sha: doc_id: 5697 cord_uid: l1zmrq4p nan in a very real sense, sepsis illustrates the multiple dimensions of the immune response of critically ill patients. on the one hand, sepsis can be viewed as the simple result of the interplay between a pathogen and the immune response to that pathogen. on the other hand, it is becoming clearer that this interplay is much more complex than it appeared. it often leads to major defects in immunity during recovery, conferring increased susceptibility to secondary infections and leading to worsened outcomes [1, 2] . the generic entity of sepsis, as defined by the pathological host response to infection, encompasses dichotomous clinical presentations (fig. 1) . a robust and tightly regulated early response to invading organisms is required to contain and eliminate the microbes. both a too strong or too mild response may adversely affect outcome. a subset of septic patients infected with highly virulent pathogens may die rapidly from refractory shock, disseminated intravascular coagulation, and intractable multiple organ failure as a result of hyper-cytokinemia and uncontrolled inflammatory response. however, with improvements in acute care and resuscitation therapies, this classical exuberant presentation is relatively rare nowadays, and the physician is more often challenged by a blunted clinical response to infection, with subtle findings including lethargy or depressed mental status, glucose intolerance and hyperglycemia, hypothermia, and/or a change in the white blood cell count or cell differential. the nature and intensity of the inflammatory response is mainly determined by the patient's background [1, 2] . sepsis is increasingly a problem for patients who have impaired immunity and who then tend to offer a blunted clinical presentation. in addition to overt immunosuppressive conditions (e.g., cancer and hematological malignancies, solid organ transplant, autoimmune and systemic diseases, hiv, use of immunosuppressive drugs), many septic patients commonly exhibit additional risk factors affecting immune status [3] . for example, chronic alcoholism, malnourishment, renal insufficiency, and liver failure all impair immunity. another particularly important risk factor is age, since the elderly are known to have blunted immunity, a condition termed ''immunosenescence''. furthermore, genetic predisposition and epigenetic changes are also hidden determinants of the host's response to infection [4, 5] . numerous clinical and laboratory findings are consistent with new-onset or worsening of pre-existing immunosuppression in critically ill patients: loss of body temperature regulation and/or new-onset hypothermia [6] ; hypoalbuminemia despite adequate nutritional support; persistent low absolute lymphocyte counts [7] . most importantly is a failure to clear the primary infection despite source control and adequate antibiotics as well as development of icu-acquired infections with opportunistic pathogens such as acinetobacter, stenotrophomonas, enterococci, or fungal organisms [1, 2, 8] . illustratively, over 40 % of septic patients have evidence of reactivation of latent viruses, through detection of hsv in upper or lower respiratory tract with or without oropharyngeal herpetic lesions, or increased plasma hsv or cytomegalovirus dnaemia [9] . though rarely responsible for specific organ failures, viral reactivation may add to the risk for further bacterial or fungal infections by subverting the functions of immune cells. on top of immunosuppressed patients being more vulnerable to sepsis, sepsis itself is profoundly immunosuppressive, and it may worsen immunity in both previously healthy and already immunocompromised patients. sepsis induces multiple defects in innate and adaptive immunity including apoptosis-induced depletion of immune effector cells (lymphocytes and dendritic cells), monocyte deactivation, t cell exhaustion, increased myeloid-derived suppressor cells, and increased t regulatory cells [8] . it is noteworthy that similar immune defects have also been reported in patients suffering from severe non-septic tissue damage related to trauma, or even to major surgery, and numerous icu therapies likely impair local or systemic immunity (e.g., mechanical ventilation, red blood cell transfusion, corticosteroids, catecholamines) [10] . the net result of these immuno-enhancing therapies: gm-csf, il-7, anti pd-1/pd-l1, ifn-γ, thymosin alpha-1 how should development of immunosuppression impact clinical management? regardless of underlying comorbidities and primary injuries responsible for icu admission, a significant proportion of critically ill patients can reasonably be considered to be immunocompromised and at risk for icu-acquired infections, especially if they develop overt signs of immunosuppression. direct clinical implications include a meticulous diagnostic work-up for common sources of infection such as infected central lines, wound infections, ventilator-associated pneumonia, as well as appraising the probability of invasive fungal infections and investigating cytomegalovirus or herpes simplex virus reactivations. depending upon the level of suspicion and previous known colonization, it may be reasonable to start empirical antimicrobial therapy while awaiting microbial cultures. whether a preemptive treatment of cytomegalovirus or herpes simplex virus reactivations with antivirals may improve the outcome of critically ill patients is currently being investigated. an attractive therapeutic goal lies in the maintenance or restoration of immune response in septic patients who are recovering from the primary infectious episode. when feasible, discontinuation or reducing immunosuppressive medications is a theoretical key management principle in this setting, but may hardly be achieved in some clinical situations. this includes judicious use of corticosteroids which are still commonly used in septic shock patients, although we advocate avoiding their use whenever possible. future implications: immuno-adjuvants to the rescue? the burden of nosocomial infections and the spread of highly resistant bacteria call for innovative preventive and therapeutic approaches in critically ill patients. although there is abundant circumstantial evidence that immunosuppression is responsible for, or at least contributing to, the morbidity and mortality in sepsis, definitive evidence of its true role awaits results of clinical trials of immunoadjuvant therapy. case series and small trials indicate that immunostimulating compounds are able to restore immune function and may exert beneficial clinical effects in sepsis patients, while no overt safety issues have been reported [12, 13] . importantly, the identification of accurate biomarkers to detect immunosuppression in septic patients will be key to success of these trials [14, 15] . currently, the expression of hla-dr on monocytes as a feature of monocyte deactivation seems to be the most reliable biomarker to assess the immune status of critically ill patients. a lower expression and especially failure of restoration of hla-dr expression appear indicative of immunoparalysis, as it is related to the susceptibility to opportunistic infections and outcome in sepsis patients [15] . several promising immuno-adjuvant agents are slated for testing in the near future, including gm-csf (clinicaltrials.gov identifier: nct02361528), il-7, anti-pd-l1 (clinicaltrials.gov identifier: nct02576457), and thymosin-alpha-1 [2, 8, 12, 13, 16, 17] . given the remarkable success of immunotherapy in cancer and the similarities in the immune defects in cancer and sepsis patients [18] , it appears plausible that immunotherapy may represent a major advance in the treatment of this highly lethal disease. conflicts of interest none. the pathophysiology and treatment of sepsis immunotherapy for the adjunctive treatment of sepsis: from immunosuppression to immunostimulation. time for a paradigm change? timing and causes of death in septic shock genetic influences on incidence and case-fatality of infectious disease epigenetic programming of monocyteto-macrophage differentiation and trained innate immunity the presence of hypothermia within 24 hours of sepsis diagnosis predicts persistent lymphopenia specific mait cell behaviour among innate-like t lymphocytes in critically ill patients with severe infections sepsis-induced immunosuppression: from cellular dysfunctions to immunotherapy reactivation of multiple viruses in patients with sepsis innate immune dysfunction in trauma patients: from pathophysiology to treatment temporary withdrawal of immunosuppression for life-threatening infections after liver transplantation monocytic hla-dr expression in intensive care patients: interest for prognosis and secondary infection prediction granulocyte-macrophage colony-stimulating factor to reverse sepsis-associated immunosuppression: a double-blind, randomized, placebocontrolled multicenter trial elevated plasmatic level of soluble il-7 receptor is associated with increased mortality in septic shock patients low monocyte human leukocyte antigen-dr is independently associated with nosocomial infections after septic shock il-7 restores lymphocyte functions in septic patients the efficacy of thymosin alpha 1 for severe sepsis (etass): a multicenter, single-blind, randomized and controlled trial parallels between cancer and infectious disease key: cord-104180-f3hoz9bu authors: kirk-bayley, justin; venn, richard title: recently published papers: inflammation, elucidation, manipulation? date: 2003-07-03 journal: crit care doi: nan sha: doc_id: 104180 cord_uid: f3hoz9bu nan the search for the marker in sepsis and inflammation continues. perhaps when we do find it we may be able to alter and influence the underlying pathophysiology of sepsis. many reports from those who believe that this may lie in the hypothalamic-pituitary-adrenal axis have recently been published. manglik and colleagues [1] looked into secondary adrenocortical insufficiency in patients who present with severe sepsis. measuring maximal cortisol secretion after stimulation with adrenocorticotrophic hormone, those investigators found that 9% of their population failed an adrenocorticotrophic hormone stimulation test. four per cent had previously undiagnosed pituitary disease and 5% were suffering from sepsis-related adrenal dysfunction. they used absolute cortisol levels and not delta cortisol to define adrenocortical insufficiency, but until we have an established definition for adrenocortical insufficiency in sepsis the studies will continue to yield disparate results. looking at reduced endogenous steroid levels in sepsis, marx and coworkers [2] focused on the androgens dehydroepiandrosterone (dhea) and its sulphated precursor (dheas) and looked at disparity between survivors and nonsurvivors from severe sepsis. the cortisol levels of survivors reached upper normal limits and decreased significantly toward late sepsis. however, nonsurvivors had persistently lower cortisol levels (but within the normal physiological range) throughout sepsis. dheas paralleled this in survivors, with normal early levels reducing in late sepsis, but in nonsurvivors levels were persistently low throughout and to a significant degree. survivors had persistently elevated levels of dhea as compared with nonsurvivors. marx and coworkers showed that relative adrenocortical insufficiency extends to androgens, that dhea and dheas changes do not parallel each other, and that dheas levels may even predict survival in severe sepsis, which apache (acute physiology and chronic health evaluation) scores do not. so will stress doses of hydrocortisone attenuate severe systemic inflammatory response syndrome after cardiac surgery with cardiopulmonary bypass? kilger and coworkers [3] would suggest that in high-risk patients this is so. they found that severe systemic inflammatory response syndrome (serum interleukin-6 concentration >1000 pg/ml) was best predicted in their population by a bypass time in excess of 97 min and cardiac ejection fraction below 40%. thus, by targeting their predefined high-risk patients to receive hydrocortisone, they showed recipients to have significantly lower interleukin-6 levels, reduced duration of ventilation and catecholamine support, and reduced duration of stay in the intensive care unit and the hospital. nothing yet suggests a mortality benefit though. increased tissue factor is produced as part of the septic inflammatory process because the coagulation cascade is activated too. carraway and coworkers [4] have already shown that pretreating baboons with active site-inactivated factor vii and tissue factor pathway inhibitor at the onset of sepsis attenuates organ injury and is protective for lung and kidney. they have now shown that similarly blocking tissue factor in established gram-negative sepsis will attenuate this damage [5] . using a baboon model again, they blocked coagulation cascade initiation using active site-inactivated factor vii at the time of first antibiotic therapy after inducing sepsis. this resulted in reduced acute lung injury, renal injury, metabolic acidosis and sepsis-induced coagulopathy. can we manipulate this therapeutically in human sepsis? time and more research will tell. moving away from direct involvement of the inflammation/ coagulation process, soliman and colleagues [6] looked at how the ionized portion of serum magnesium varies in the critically ill. they found that ionized magnesium level at presentation did not affect outcomes (two-thirds of their population had normal levels), but that the development of reduced levels of ionized magnesium while in intensive care was associated with higher mortality and more severe organ dysfunction. sepsis was an independent risk factor for ionized hypomagnesaemia, but soliman and colleagues postulated that prolonged disease and diuretic administration may also be contributory. it may be that low levels contribute to critical illness, or that just the converse is true. which is true is unclear. what we need now is to discover how magnesium supplementation will alter these findings. inappropriate ventilatory strategies for patients with acute respiratory distress syndrome (ards) may have more consequences for the recipient than just lung abnormalities. imai and colleagues [7] indicated that remote cellular damage also occurs and that this may contribute to the multiple organ dysfunction that often accompanies ards. they looked at end-organ epithelial cell apoptosis in a rabbit model of ards and at the effects of plasma on epithelial cells from recipients of the injurious ventilatory strategy, and analyzed samples from a previous trial into lung protective ventilation [8] . they showed that kidney and small intestine damage occurred when ventilation with high tidal volumes and low positive end-expiratory pressures were used, and that plasma from rabbits in this group would induce apoptosis in cells in vitro. the fas-fas ligand system is involved in cell apoptosis, and they found elevated levels of soluble fas ligand in human patients who had not received a protective ventilatory strategy, which correlated well with elevations in creatinine levels. choosing the right ventilation strategy for ards patients has more benefits than just lung protection, and therapeutic targeting of these factors that induce end organ apoptosis may be the next step. choosing the right ventilation strategy may not be the only way to avoid lung problems in patients with ards. desirable strategies involve the prevention of over-distended alveoli and their de-recruitment. thus, it is not unreasonable to consider that periodic endotracheal suctioning might undermine these benefits. maggiore and colleagues [9] set out to find out whether this was so in patients with acute lung injury, and considered preventative measures. they found that closed suctioning systems reduced large lung volume falls and preserved positive end-expiratory pressure induced recruitment. recruitment was augmented by performing recruitment manoeuvres at the time of suctioning. repetitive shearing stresses on alveoli are injurious, and this study shows some ways to prevent this. using inhaled nitric oxide (no) in patients with ards improves oxygenation, but it has not been proven to improve outcome. why is this so? gerlach and coworkers [10] looked at dose-response characteristics when long-term no is used and found that these characteristics changed. patients could become sensitized to no, such that the number of responders to low no doses increased and some became nonresponders at higher no doses. it would seem that constant dose no is not right for all patients throughout their treatment, and that there is more inter-patient and intrapatient no dose-response variability than we had thought. titrating doses at each step may improve long-term benefits from no therapy. with respect to inotropes, which one should we use? the debate continues. de backer and colleagues [11] compared inotropes and their effects on the splanchnic circulation in sepsis. dopamine was used and then substituted for either adrenaline (epinephrine) or noradrenaline (norepinephrine) to achieve or maintain adequate arterial pressures depending on shock severity. the three inotropes were comparable in moderate shock, and dopamine would actually appear to have modest benefits for splanchnic circulation. however, in more severe septic shock adrenaline impaired splanchnic circulation, and de backer and colleagues suggested that it should be avoided in high doses in these patients. a larger, truly randomized trial should determine whether they are correct. however, if you use low dose dopamine in critically ill patients for renal protection, holmes and walley [12] tell us that the practice should be, "relegated to the place of hightidal volume ventilation and liberal transfusion practices". glucocorticoid insufficiency in patients who present to the hospital with severe sepsis: a prospective clinical trial adrenocortical hormones in survivors and nonsurvivors of severe sepsis: diverse time course of dehydroepiandrosterone, dehydroepiandrosterone-sulfate, and cortisol stress doses of hydrocortisone reduce severe systemic inflammatory response syndrome and improve early outcome in a risk group of patients after cardiac surgery coagulation blockade prevents sepsis-induced respiratory and renal failure in baboons blockade of tissue factor: treatment for organ injury in established sepsis development of ionized hypomagnesemia is associated with higher mortality rates injurious mechanical ventilation and end-organ epithelial cell apoptosis and organ dysfunction in an experi-critical care effect of mechanical ventilation on inflammatory mediators in patients with acute respiratory distress syndrome: a randomized controlled trial prevention of endotracheal suctioning-induced alveolar derecruitment in acute lung injury dose-response characteristics during long-term inhalation of nitric oxide in patients with severe acute respiratory distress syndrome: a prospective, randomized, controlled study effects of dopamine, norepinephrine, and epinephrine on the splanchnic circulation in septic shock: which is best? bad medicine: low-dose dopamine in the icu on the physiologic and clinical relevance of lung-borne cytokines during ventilator-induced lung injury discrepancies between perceptions by physicians and nursing staff of intensive care unit end-of-life decisions a randomized trial of inhaled nitric oxide to prevent ischemia-reperfusion injury after lung transplantation activated protein c inhibits the expression of platelet-derived growth factor in the lung treatment of acute respiratory distress syndrome with recombinant surfactant protein c surfactant impact of randomized trial results on acute lung injury ventilator therapy in teaching hospitals none declared. as well as the references cited in the text, i also recommend the following. key: cord-270213-ygb64yxc authors: williams, alexander t.; muller, cynthia r.; govender, krianthan; navati, mahantesh s.; friedman, adam j.; friedman, joel m.; cabrales, pedro title: control of systemic inflammation throughearly nitric oxide supplementation with nitric oxide releasing nanoparticles date: 2020-10-02 journal: free radic biol med doi: 10.1016/j.freeradbiomed.2020.09.025 sha: doc_id: 270213 cord_uid: ygb64yxc amelioration of immune overactivity during sepsis is key to restoring hemodynamics, microvascular blood flow, and tissue oxygenation, and in preventing multi-organ dysfunction syndrome. the systemic inflammatory response syndrome that results from sepsis ultimately leads to degradation of the endothelial glycocalyx and subsequently increased vascular leakage. current fluid resuscitation techniques only transiently improve outcomes in sepsis, and can cause edema. nitric oxide (no) treatment for sepsis has shown promise in the past, but implementation is difficult due to the challenges associated with delivery and the transient nature of no. to address this, we tested the anti-inflammatory efficacy of sustained delivery of exogenous no using iv infused no releasing nanoparticles (no-np). the impact of no-np on microhemodynamics and immune response in a lipopolysaccharide (lps) induced endotoxemia mouse model was evaluated. no-np treatment significantly attenuated the pro-inflammatory response by promoting m2 macrophage repolarization, which reduced the presence of pro-inflammatory cytokines in the serum and slowed vascular extravasation. combined, this resulted in significantly improved microvascular blood flow and 72-hour survival of animals treated with no-np. the results from this study suggest that sustained supplementation of endogenous no ameliorates and may prevent the morbidities of acute systemic inflammatory conditions. given that endothelial dysfunction is a common denominator in many acute inflammatory conditions, it is likely that no enhancement strategies may be useful for the treatment of sepsis and other acute inflammatory insults that trigger severe systemic pro-inflammatory responses and often result in a cytokine storm, as seen in covid-19. sepsis represents a dynamic progression of host-pathogen interactions that progressively 64 causes a systemic inflammatory response syndrome (sirs) and ultimately leads to multi-65 organ dysfunction syndrome (mods) even after the initial insult has been controlled. 1 the 66 complications associated with sepsis are the most common cause of death in non-coronary 67 intensive care units (icus) worldwide. medical care costs related to sepsis treatment add up to 68 approximately $20 billion in the united states. 2,3 however, the development of a 69 comprehensive study to target key aspects of sepsis development and progression has been 70 challenging and most of the results obtained from bench top experiments are hard to translate 71 to the bedside. the difficulty to generate translatable data in experimental studies comes 72 mainly from two reasons. first, the large number of variables that play a role in the 73 deterioration of cell and tissue function during sirs and sepsis makes it almost impossible to 74 pinpoint a single therapeutic target. 3,4 second, the complexity and diverse sources of human 75 sepsis makes the development of an animal model that is reproducibly translatable, in which 76 different theories could be tested, almost impossible. thus, a middle ground should be set in 77 which individual components of sepsis can be used to understand the response of the insulted 78 organism. 79 80 a well-described hallmark of sepsis is endothelial dysfunction in response to a cytokine 81 'storm', which is associated with an increase in a series of negative consequences arising from 82 overproduction of reactive oxygen species (ros), disruption of the glycocalyx, and endothelial 83 nitric oxide synthase (enos) uncoupling, all contributing to increased adhesion of red blood 84 cells (rbcs), white blood cells (wbcs), and platelets to the endothelium lining, enhanced 85 platelet activation, blood stagnation, decreased tissue perfusion and increased vascular 86 permeability. 1, 5, 6 experimental therapies designed to target this aspect have been promising 87 and represent pathways that could be targeted to increase survival. 5,6 from a clinical 88 standpoint, several studies have highlighted that microvascular function is drastically impaired 89 in patients in different stages of sepsis. 7 the evidence indicates that the gold standard, fluid 90 replacement therapies, fail to recover microvascular blood flow causing poor perfusion, which 91 has been associated with increased mortality. 8, 9 routinely monitored clinical variables poorly 92 reflect the true state of the microcirculation in central and peripheral tissues. 10 recently it has 93 been proposed that in order to properly restore cardiovascular homeostasis, the 94 microcirculation should be targeted as a functional unit. intravital microscopy and microvascular measurements. the window chamber was 169 studied using transillumination on a custom intravital microscope. briefly, the animals were 170 restrained on a plexiglass tube with a longitudinal opening from which the window protruded 171 and then they were fixed to the stage of an upright microscope (bx51wi, olympus, new hyde 172 park, ny) as described and depicted elsewhere. 20 measurements were carried out using a 40ˣ 173 (lumpfl-wir, numerical aperture 0.8, olympus) water immersion objective. the microscope 174 was equipped with a high-speed video camera (fastcam 1024 pci, photron usa), which was 175 used to record videos of the microvascular blood flow at 1,000 frames per second (fps). 176 briefly survival. in addition to measuring 297 functional parameters, we also assessed 298 survival of mice dosed with lps over 72 299 hours (figure 4) cd206, indicating an m1-like macrophage phenotype, which is considered to be inflammatory. 323 in fact, over 72% of macrophages tested via facs presented an m1-like phenotype for 324 animals treated with control-np, but only 16% of macrophages represented an m1-like 325 phenotype for animals treated with no-np. however, animals treated with no-np showed a 326 larger population of m2-like macrophages (representing 33% of macrophages harvested), 327 which are typically considered to be anti-inflammatory, and are associated with tissue repair. 328 to supplement these macrophage phenotypes, we also measured the cytokine profile of 329 treated mice after lps injection. cytokine profile. cytokines were measured 24 and 48 hours after lps injection (figure 6) . 339 after 24 hours, animals treated with no-np showed significantly higher levels of cytokines 340 traditionally considered to be anti-inflammatory (interleukin [il]-10, tgf-beta), and significantly 341 lower levels of cytokines associated with a proinflammatory response (il-1, 6, 12, mcp, and 342 tnf alpha) than animals treated with control-np. however, after 48 hours, all cytokines 343 measured were elevated in animals dosed with control-np compared to no-np, but not all of 344 these comparisons were statistically significant. the principle finding of this study is that sustained delivery of exogenous no using no 350 releasing nanoparticles improved microvascular flow and capillary transit compared to animals 351 treated with control nanoparticles during lps-induced endotoxemia. additionally, this study 352 demonstrates that the adverse microcirculatory changes from lps-induced endotoxemia there are a number of mechanisms that may be responsible for the improved survival and 372 maintenance of microvascular perfusion seen with no-np treatment in this study, but the exact 373 mechanism or set of mechanisms is unclear. as we saw, and as others have demonstrated, 22 374 no treatment decreased m1, and increased m2 polarization of macrophages. this can have a 375 multitude of downstream effects as m1 macrophages produce proinflammatory cytokines, 376 upregulate inducible nitric oxide synthase (inos), and enhance production of ros and 377 reactive nitrogen species, which disrupt the glycocalyx, expose the endothelial layer, decrease 378 no production from enos, and destroy endothelial cells. 23,24 379 380 disruption of the glycocalyx undermines vascular integrity and eliminates the shear stress-381 based trigger for production of no from enos. mice treated with control-np in our study 382 experienced significantly increased vascular permeability, as shown in figure 4 , suggesting 383 endothelial cell and glycocalyx disruption in these animals. the glycocalyx has a number of 384 vital physiological roles, including acting as a barrier to preserve intravascular oncotic 385 pressure, 23 promoting rbc marginalization and the presence of a red cell-free layer, 25,26 386 reducing leukocyte adhesion and infiltration, 26 and preventing thrombus formation and 387 attachment, 27 so its disruption is likely a primary determinant for many of the clinical 388 presentations of sepsis, such as edema. as such, it is logical that the glycocalyx has been a 389 common drug target in improving outcomes from sepsis. others have explored individual 390 mechanisms as a method to prevent glycocalyx degradation during sepsis, but these 391 j o u r n a l p r e -p r o o f strategies have not shown much clinical success. 23 however, no-np treatment seems 392 particularly promising as it attempts to treat a more upstream target than previous therapies by 393 preventing the initial insult to the glycocalyx, and thus preserving endogenous no signaling. 394 endogenous no may function to upregulate and activate nuclear factor erythroid 2-related 395 factor 2 (nfr2) which represents a potent signaling pathway to counter an lps-induced 396 inflammatory cascade, and to down regulate pro-inflammatory pathways and thus decrease 397 ros production. 28 limitations: lps induced endotoxemia does not fully replicate the cascade of events that 464 occurs during septic shock, but no study can fully replicate the morbidity of septic shock due to 465 its distinct etiologies. however, this study did demonstrate that this mouse model replicates 466 many of the early changes that occur in septic shock, including increased vascular 467 permeability, and impaired systemic oxygen transport, and allowed us to observe these 468 changes in an awake, unanesthetized model. since anesthetics have a poorly characterized 469 influence on inflammation, the awake unanesthetized state is most representative of changes 470 that may occur during sepsis. unfortunately, this study did not measure any parameters 471 examining the status of the vascular endothelial glycocalyx, since sepsis damages the 472 glycocalyx, and exogenous no could help protect the glycocalyx. future studies should 473 examine changes in the glycocalyx by measuring its breakdown molecules, such as heparan 474 sulfate and sydecan-1, in the plasma, or measure glycocalyx integrity and thickness via 475 fluorescent lectin binding, in order to directly study glycocalyx disruption during lps-induced 476 endotoxemia, and its protection by no-nps. 477 478 conclusion 479 pathogenetic mechanisms of septic shock 506 epidemiology of severe sepsis in the united states: analysis of incidence, outcome, and 507 associated costs of care epidemiology of severe sepsis severe sepsis and septic shock sepsis and endothelial permeability novel therapies for microvascular permeability in 515 sepsis the microcirculation is the motor of sepsis severe abnormalities in microvascular perfused vessel density are associated 520 to organ dysfunctions and mortality and can be predicted by hyperlactatemia and 521 norepinephrine requirements in septic shock patients microvascular resuscitation as a therapeutic goal in 523 severe sepsis 525 early difference in tissue ph and microvascular hemodynamics in hemorrhagic shock 526 resuscitation using polyethylene glycol-albumin-and hydroxyethyl starch-based plasma 527 expanders b. & intaglietta, m. microcirculation: its significance in clinical and molecular microcirculation: physiology, pathophysiology, and 532 clinical application clinical review: clinical imaging of the 534 sublingual microcirculation in the critically ill -where do we stand? lps/tlr4 signal transduction pathway animal models of sepsis: setting the stage sustained release nitric oxide releasing nanoparticles: characterization of a novel 542 delivery platform based on nitrite containing hydrogel/glass composites a nanoparticle delivery vehicle for s-nitroso-n-acetyl cysteine: 546 sustained vascular response 548 sustained release nitric oxide from long-lived circulating nanoparticles. free radical 549 biology & medicine 49 technical report-a new chamber 551 technique for microvascular studies in unanesthetized hamsters a. & leunig, m. dorsal skinfold chamber preparation in mice: studying 554 microhemodynamic parameters quantification 556 from intravital microscopy videos macrophage polarization mediates anti-inflammatory effects of endothelial nitric oxide 560 signaling glycocalyx and 562 sepsis-induced alterations in vascular permeability the m1 and m2 paradigm of macrophage activation: time 564 for reassessment implications enzymatic degradation 566 of the endothelial glycocalyx on the microvascular hemodynamics and the arteriolar red 567 cell free layer of the rat cremaster muscle modulates immobilization of leukocytes at the endothelial surface the glycocalyx: a novel diagnostic and 572 therapeutic target in sepsis activation of 574 endothelial nitric oxide synthase by dietary isoflavones: role of no in nrf2-mediated 575 antioxidant gene expression the coagulopathy of acute sepsis the role of microvascular thrombosis in sepsis endogenous and exogenous nitric oxide protect against intracoronary 582 thrombosis and reocclusion after thrombolysis nitric oxide regulates exocytosis by s-nitrosylation of n-586 ethylmaleimide-sensitive factor nitric oxide and its relationship to thrombotic disorders 590 exposure of fibrinogen and thrombin to nitric oxide donor prolinonoate affects fibrin clot 591 properties the role of 593 the microcirculation in multiple organ dysfunction syndrome (mods): a review and 594 perspective nitroglycerin in septic shock after intravascular volume resuscitation nitrite protects against morbidity and mortality associated with tnf-or lps-induced 600 shock in a soluble guanylate cyclase-dependent manner reducing ischemia/reperfusion injury by the targeted delivery of nitric 604 oxide from magnetic-field-induced localization of s-nitrosothiol-coated paramagnetic 605 nanoparticles 607 covid-19: consider cytokine storm syndromes and immunosuppression interactions of coronaviruses with ace2, angiotensin ii, and ras 610 inhibitors-lessons from available evidence and insights into covid-19 cov-2 cell entry depends on ace2 and tmprss2 and is blocked 615 by a clinically proven protease inhibitor antigenicity of the sars-cov-2 spike glycoprotein a 622 crucial role of angiotensin converting enzyme 2 (ace2) in sars coronavirus-induced 623 lung injury ace2-angiotensin-(1-7)-mas axis and oxidative 625 stress in cardiovascular disease harnessing nitric oxide 627 for preventing, limiting and treating the severe pulmonary consequences of covid-19 inflammatory response was reduced with nitric oxide releasing nanoparticles (no-nps) endotoxemia-induced microvascular deficits were alleviated by no-nps no-nps promoted m2 macrophage repolarization, and reduced m1 macrophage population no-nps could improve outcomes of insults that cause a cytokine storm key: cord-348785-f67amppy authors: kapicibaşi, hasan oğuz; kiraz, hasan ali; demir, emin tunç; adali, yasemen; elmas, sait title: pulmonary effects of ozone therapy at different doses combined with antibioticotherapy in experimental sepsis model date: 2020-07-13 journal: acta cirurgica brasileira doi: 10.1590/s0102-865020200060000004 sha: doc_id: 348785 cord_uid: f67amppy purpose: this experimental sepsis model created with escherichia coli aimed to investigate the histopathological effects of two different doses of ozone combined with antibiotherapy on lung tissue. methods: rats were divided into 5 groups. then sepsis was induced intraperitoneally in the first 4 groups. the 1(st) group was treated with cefepime, the 2(nd) and 3(rd) groups were treated with cefepime combined with ozone at a dose of 0.6 mg/kg and 1.1 mg/kg. lung tissue sections were stained with hematoxylin-eosin and assessed under light microscope and scored between 0-4 in terms of histopathological findings. results: in the comparisons between group 1 and group 4 in terms of cellular damage (p=0.030), inflammation (p=0.000) and overall score (p=0.007), statistically significant positive effects were observed in favor of group 1. in the comparisons of groups 2 and 3 with group 4, only positive effects were observed in terms of inflammation (p=0.020, p=0.012, respectively). conclusion: although negative histopathological effects of ozone on tissue injury were detected, it was noteworthy that the increase in the ozone dose reduced the number of damaged parameters. (file registration number: 2018/1800080971 & decision number: 2018/06-06). our study included 40 male sprague-dawley rats with mean weight 350-400 g in appropriate condition. during the study, rats were kept in çomü experimental research center in wire cages with 12-hour night-12-hour day circadian rhythm, environmental temperature 24-26°c and humidity 50-60%. rats were fed with standard commercial feed and municipal drinking water. rat feed was stopped 12 hours before the study; however, water was given freely during this time. all rat care was performed in accordance with the "regulation on the welfare and protection of animals used for experimental and other scientific purposes" (13.12.2011-28141) prepared by the ministry of food, agriculture and livestock. experimental animals were randomly divided into 5 groups with equal numbers (n=8). sepsis was induced in rats in the first 4 of these groups with 2.1x10 9 cfu/ml escherichia coli atcc 25922 administered in 1 ml saline via the intraperitoneal route in 32 rats. rats in the 5 th group only had 1 ml intraperitoneal saline administered. twenty-four hours after escherichia coli administration, rats in the 1 st , 2 nd and 3 rd groups were arranged in groups following skin cleaning for asepsis and the following treatments began. care was taken to administer treatments at the same time every day. additionally, an ozone generator (device name and model: turkozone blue s, device serial no: bg-19144427046) was used to produce the ozone/ oxygen mixture for ozone therapy. sepsis is defined as life-threatening organ dysfunction caused by an unregulated host response to infection 1 . the pathology of sepsis involves complex interactions between host organs and invading pathogens. ultimately tissue injury and organ failure are due to negative effects of systemic activation of host immunity 2,3 . sepsis forms a large problem in intensive care and is the main cause of death occurring there 4,5 . according to a report published by the global burden of disease, each year nearly 10 million people die from infections and this is much greater than the number of people who die from cancer annually 6 . lungs are the first organs affected by sepsis and sepsis causes severe injury to lung tissue 7 . in spite of advanced antibiotherapy, supportive treatments and all technological opportunities, sepsis continues to be a situation progressing with morbidity and mortality 8 . the earliest target-directed basic treatment principles for sepsis comprise determining high-risk patients, ensuring appropriate cultures and source control, and beginning appropriate antibiotherapy without delay 9 . medical ozone therapy is an alternative treatment model linked to the administration of ozone and gas mixture with oxygen to body fluids or cavities. to date, clinical and experimental studies have shown that ozone therapy is beneficial for inflammation-mediated diseases like infected wounds, chronic skin ulcers, burns and advanced ischemic diseases 10 . ozone/ oxygen mixtures are reported to display a variety of effects on the immune system like phagocytic activity modulation 11 . additionally, ozone therapy is proposed to cause increased antioxidant enzyme expression 12 . additionally, it is known that ozone is also an immune system modulator. though antiseptic properties of ozone are well known, it may harm not just the pathogen but also the patient linked to dose and exposure duration 13 . ozone is one of the strongest oxidants and may show unwanted effects through a variety of mechanisms like free radical formation, lipid peroxidation, enzymatic activity loss, changes in membrane permeability, direct organ inflammation and injury 14 . based on the known positive and negative effects of ozone, in our study we aimed to assess the effect of two different doses of ozone therapy added to antibiotic treatment in an experimental sepsis model induced with escherichia coli on the histopathologic findings observed in the inflammatory process in the lungs. pulmonary effects of ozone therapy at different doses combined with antibioticotherapy in experimental sepsis model kapicibaşi ho et al. acta cir bras. 2020;35(6):e202000604 3 4 th group: rats in this group were not given any treatment after administration of escherichia coli by the intraperitoneal route with the aim of inducing sepsis. rats in this group did not have a sepsis model induced, but were administered 1 ml saline intraperitoneal at the start and were only administered subcutaneous saline at equivalent times to the treatments in the other groups. twenty-four hours after the final treatment in our study, rats were administered high-dose anesthetics (xylazine -10 mg/kg and ketamine -80 mg/kg) and lung tissue samples and blood samples were taken by accessing the thorax cavity with median sternotomy and rats were sacrificed. in the blood samples, the diagnosis of sepsis was confirmed by working with the rats escherichia coli protein (e.coli p) enzyme-linked immunosorbent assays (elisa) kits (shangai coon koon biotech co, china). in view of the data on the researched serums, the very low level of e.coli p in the rats of the control group was construed and reported as the absence of sepsis. the statistically significant differences concerning the presence of e.coli p in the first four groups in comparison with the control groups signify that sepsis was successfully induced in these groups (fig. 1) . lung tissue samples were fixated for 24 hours in 10% buffered formalin solution. later, tissue monitoring procedures were performed appropriate to groups and tissues were submerged in paraffin. sections with 4-micron thickness were taken from the prepared paraffin blocks and stained with hematoxylin & eosin (h&e) before being investigated with a light microscope (olympus bx46, japan) at magnification above 10 fields 15 . the tissues were assessed by the parameters used by yamanel et al. 16 with some modification and their scoring system was used. hemorrhage, cellular injury, alveoloseptal thickening and inflammation were scored from 0-4 and a general injury score (total score) was obtained by summing all scores per case. maximum care was taken to use the lowest number of animals necessary to gain scientific quality from the methods applied in the study and the obtained results. analysis of data used the statistical package for the social sciences for windows evaluation v. 15 .0 (spss, chicago, il, usa) program. results are given as median and minimum-maximum values. comparisons of groups used the mann-whitney u test. tests were completed in the 95% confidence interval and values with p lower than 0.05 were accepted as statistically significant. as the histopathologic findings observed in the study are categoric data, the median and interval values are presented in table 1 . when group 4 is compared with the control group of group 5, group 4 was identified to have more statistically significant injury in terms of cellular damage (p=0.030), alveoloseptal thickening (p=0.037), inflammation (p=0.001) and total score (p=0.009), with no significant difference between the groups in terms of hemorrhage (p=0.663). when group 4 is compared with group 1, there was no statistical significance for bleeding (p=0.428) and alveoloseptal thickening (p=0.085), while there were clear positive effects in group 1 for cellular injury (p=0.030), inflammation (p=0.000) and total score (p=0.007). when group 4 is compared with group 2 and group 3, positive effects were only observed in groups 2 and 3 for inflammation (p=0.020, p=0.012, respectively), with no statistical significance for the other parameters (p>0.05). while there was no statistical significance for hemorrhage between group 1 and group 2 (p=0.655), there were clear positive effects in group 1 for cellular damage (p=0.002), alveoloseptal thickening (p=0.044), inflammation (p=0.002) and total score (p=0.017). when group 1 is compared with group 3, there was no statistical significance for bleeding (p=0.901), alveoloseptal thickening (p=0.081) and inflammation (p=0.054), but positive effects were pronounced in group 1 for cellular injury (p=0.002) and total score (p=0.035). histopathologic findings of the groups are demonstrated in figure 2 . comparison of groups 2 and 3 did not have any statistical significance (p>0.05). bleeding 2/2 (1-3) 1/1 (1-4) 2 (1-3) 2/2 (1-4) 2/2 (1-4) cellular damage 0/0 (0-1) b 1/1 (1-3) a,c 2 (1-3) a,c 1/1 (0-3) a,c 0/0 (0-1) b alveoloseptal thickening 0/0 (0-1) 1/2 (0-3) a,c 1 (0-3) a 1/1 (0-3) a 0/0 (0-1) b inflammation 0/0 (0-1) b 1/1 (1-3) a,b,c 1 (0-3) b 2/3 (2-4) a, sepsis-linked acute lung injury is an important cause of morbidity and mortality in adults and children and significantly contributes to intensive care costs [17] [18] [19] [20] . many studies have created pioneering ideas for the use of ozone as a strong antimicrobial agent [21] [22] [23] [24] . ozone is a strong oxidizing agent and important disinfectant. according to literature data, exposure of bacteria, spores and viruses to ozone for only a few minutes causes inactivation [25] [26] . studies by ricevudi et al. 27 proposed oxygen-ozone treatment as a new immunotherapeutic treatment method for the viral agent covid-19 causing this pandemic and that patients with covid-19 using this in combination with other treatment methods would benefit from the assisting and synergic effects of ozone therapy. again, as stated by ricevudi et al. 27 the need for more studies is clear. the bactericidal effect of ozone is linked to attacking the biological material in microorganisms through the oxidation pathway. in fact, the antibacterial effect of ozone is said to be more effective than iodine and chlorine 23, 28 . previous experimental studies have shown that ozone therapy has experimental benefits on pathologic processes 16, 23, 29 . in many organs like pancreas, peritoneum, liver, mesenteric lymph nodes and cecum, ozone therapy has proven reducing effects on bacterial translocation 16 . in an experimental necrotizing pancreatitis model, ozone therapy was seen to be more effective to reduce oxidative stress levels, tissue injury and bacterial translocation rates compared to hyperbaric oxygen treatment 30 . schulz et al. 31 observed a decrease in polymicrobial peritonitis after they administered intraperitoneal ozone. çakır et al. 32 reported the systemic inflammatory response markers of tnf-alpha and il-1b levels reduced after ozone treatment. an experimental sepsis study by yamanel et al. 16 compared hyperbaric oxygen treatment with ozone therapy and identified that both methods lowered oxidative stress indices, myeloperoxidase activity and serum proinflammatory cytokine levels. as a result, histopathologic injury reduced in the lung tissue of septic rats and ozone therapy showed to provide more benefits according to histopathologic injury scores and il-1b levels. as a result of the experimental study, they proposed that ozone therapy may be a complementary medicine treatment method that could be applied together with antibiotherapy for sepsis. additionally, studies have shown that ozone causes clear air pollution, and irritation and injury when taken in with immune and inflammatory cells in the lungs. this event progresses with bronchial epithelium desquamation and alveolar septal injury results in emphysema and airway hyperresponsiveness 33,34 . ozone-linked injury and inflammation is linked to the dose and frequency of ozone exposure 35 . in our study, positive effects were observed in terms of inflammation in group 2 and group 3 with added ozone therapy (p=0.020, p=0.012, respectively). however, it is considered that these effects are due more to antibiotic effects rather than to ozone therapy. the source point for this thought is that the cellular injury, inflammation, alveoloseptal thickening and general histopathologic scores in group 1, only administered antibiotics, were at statistically significantly low levels compared to group 2 (p=0.002, p=0.002, p=0.044, p=0.0017, respectively). similarly, the cellular injury and general histopathologic score were observed at statistically significantly low levels in group 1, administered only with antibiotic, compared to group 3, administered with high-dose ozone (p=0.002, p=0.035, respectively). when the results of our study are investigated, it was identified that ozone increased the negative histopathologic effects induced in the lung by inflammation. however, as the dose of ozone increased, the fall in the number of parameters with statistical significance confirms that positive effects may occur if the dose of ozone used was increased. additionally, a limitation of our study is that there was no group administered with ozone therapy alone after sepsis, so the histopathologic effects of ozone on lung tissue in sepsis could not be assessed. data obtained as a result of our study lead to the consideration that ozone therapy administered in addition to antibiotherapy may cause negative effects on lung tissue damaged due to sepsis. sepsis 2018: definitions and guideline changes the pathophysiology and treatment of sepsis extravascular lung water and sepsis sepsis-induced long-term immune paralysis-results of a descriptive, explorative study epidemiology of severe sepsis in the united states: analysis of incidence, outcome, and associated costs of care allcause mortality, and cause-specific mortality for 249 causes of death, 1980-2015. a systematic analysis for the global burden of disease study does leflunomide attenuate the sepsis-induced acute lung injury? optimizing intensive care management in paediatric sepsis prospective trial of supranormal values of survivors as therapeutic goals in high-risk surgical patients scientific and medical aspects of ozone therapy. state of the art ozone as janus: this controversial gas can be either toxic or medically useful does ozone therapy normalize the cellular redox balance? implications for therapy of human immunodeficiency virus infection and several other diseases biological and clinical effects of ozone. has ozone therapy a future in medicine? biochemical basis of ozone toxicity a novel animal model of sepsis after acute lung injury in sheep ozone therapy and hyperbaric oxygen treatment in lung injury in septic rats has mortality from acute respiratory distress syndrome decreased over time? a systematic review recent trends in acute lung injury mortality management of acute lung injury and acute respiratory distress syndrome in children gruppo italiano per la valutazione degli interventi in terapia intensiva. variable costs of icu patients: a multicenter prospective study antibacterial activity of ozonized sunflower oil (oleozon) evaluation of ozonated oxygen in an experimental animal model of osteomyelitis as a further treatment option for skull-base osteomyelitis analysis of the bactericidal effect of ozone pneumoperitoneum ozone killing action against bacterial and fungal species; microbiological testing of a domestic ozone generator studies on the biological effects of ozone 1. induction of interferon gamma on human leucocytes ozone as a bioregulator. pharmacology and toxicology of ozonetherapy today oxygen-ozone immunoceutical therapy in covid-19 outbreak: facts and figures toxicity and biochemical mechanisms of ozone gaseous ozone treatment inactivates listeria innocua in vitro efficacy of hyperbaric oxygen therapy and medical ozone therapy in experimental acute necrotizing pancreatitis repetitive pneumoperitoneum with ozonized oxygen as a preventive in lethal polymicrobial sepsis in rats pulmonary effects of ozone therapy at different doses combined with antibioticotherapy in experimental sepsis model il-33 drives augmented responses to ozone in obese mice γδ t cells are required for m2 macrophage polarization and resolution of ozone-induced pulmonary inflammation in mice role of neutralizing anti-murine interleukin-17a monoclonal antibody on chronic ozone-induced airway inflammation in mice key: cord-028164-yn53209z authors: abe, toshikazu; yamakawa, kazuma; ogura, hiroshi; kushimoto, shigeki; saitoh, daizoh; fujishima, seitaro; otomo, yasuhiro; kotani, joji; umemura, yutaka; sakamoto, yuichiro; sasaki, junichi; shiino, yasukazu; takeyama, naoshi; tarui, takehiko; shiraishi, shin-ichiro; tsuruta, ryosuke; nakada, taka-aki; hifumi, toru; hagiwara, akiyoshi; ueyama, masashi; yamashita, norio; masuno, tomohiko; ikeda, hiroto; komori, akira; iriyama, hiroki; gando, satoshi title: epidemiology of sepsis and septic shock in intensive care units between sepsis-2 and sepsis-3 populations: sepsis prognostication in intensive care unit and emergency room (spice-icu) date: 2020-06-30 journal: j intensive care doi: 10.1186/s40560-020-00465-0 sha: doc_id: 28164 cord_uid: yn53209z background: diagnosing sepsis remains difficult because it is not a single disease but a syndrome with various pathogenand host factor-associated symptoms. sepsis-3 was established to improve risk stratification among patients with infection based on organ failures, but it has been still controversial compared with previous definitions. therefore, we aimed to describe characteristics of patients who met sepsis-2 (severe sepsis) and sepsis-3 definitions. methods: this was a multicenter, prospective cohort study conducted by 22 intensive care units (icus) in japan. adult patients (≥ 16 years) with newly suspected infection from december 2017 to may 2018 were included. those without infection at final diagnosis were excluded. patient’s characteristics and outcomes were described according to whether they met each definition or not. results: in total, 618 patients with suspected infection were admitted to 22 icus during the study, of whom 530 (85.8%) met the sepsis-2 definition and 569 (92.1%) met the sepsis-3 definition. the two groups comprised different individuals, and 501 (81.1%) patients met both definitions. in-hospital mortality of study population was 19.1%. in-hospital mortality among patients with sepsis-2 and sepsis-3 patients was comparable (21.7% and 19.8%, respectively). patients exclusively identified with sepsis-2 or sepsis-3 had a lower mortality (17.2% vs. 4.4%, respectively). no patients died if they did not meet any definitions. patients who met sepsis-3 shock definition had higher in-hospital mortality than those who met sepsis-2 shock definition. conclusions: most patients with infection admitted to icu meet sepsis-2 and sepsis-3 criteria. however, in-hospital mortality did not occur if patients did not meet any criteria. better criteria might be developed by better selection and combination of elements in both definitions. trial registration: umin000027452 sepsis is an aberrant or dysregulated host response resulting in organ dysfunctions and is different from infection [1] . it is not a single disease but a syndrome exhibiting with various symptoms caused by pathogens and host factors. sepsis should be immediately recognized because it is the primary cause of death from infection, especially if not diagnosed and treated promptly. sepsis-2 has high sensitivity [2] but captures mild infection and not infectious diseases. sepsis-3 was established to improve risk stratification among patients with a suspected infection focusing on organ failures [1] . when considering previous studies about the diagnosis and taxonomy of sepsis to date [1, 3, 4] , nearly all of them just defined sepsis as cases of high mortality due to infectious diseases. sepsis studies may be controversial because they were unable to differentiate an aberrant or dysregulated host response itself from infection. the definitions of sepsis-2 and sepsis-3 have still been inadequate to accurately capture sepsis. therefore, both definitions may have misclassified patients with sepsis as patients with infectious diseases. although the true nature of sepsis remains to be identified, we should clearly know what the definitions of sepsis-2 and sepsis-3 indicate because different definitions could change its epidemiology to identify the clinical care, future research, and healthcare planning. such information would facilitate the definition criteria of the next sepsis. therefore, this study aimed to describe characteristics of patients who met sepsis-2 and sepsis-3 definitions. this multicenter, prospective cohort study was conducted in an intensive care unit (icu) subset of the japanese association for acute medicine sepsis prognostication in intensive care unit and emergency room (jaam spice-icu), including 22 icus in japan from december 2017 to may 2018. adult patients (≥ 16 years) with newly suspected infection were included. suspected infection was defined by the administration of any kind of antibiotic, and thereby a culture of body fluids or imaging should be conducted to identify the infectious pathogen. all patients were admitted to the icus in study hospitals. exclusion criteria included patients who were not transferred from other hospitals and those without infection at the final diagnosis. data were extracted from the spice database, compiled by spice investigators. collected variables included relevant patient information, such as demographics, comorbidities, degree of clinical frailty, vital signs, and site of infection. in-hospital mortality was identified as the primary outcome. secondary outcomes were ventilator-free days (vfd), intensive care unit-free days (icu-free days), length of hospital stay (los), and condition at discharge. data collection was conducted as part of the clinical routine workup. spice site investigators recorded all data throughout the patient's hospital stays. if case of missing data, the spice committee requested a reconfirmation of data extraction from spice investigators. sepsis-2 was defined as having a suspected site of infection, ≥ 2 systemic inflammatory response syndrome criteria (sirs) [5] and ≥ 1 organ dysfunction criteria [6] . severe sepsis was actually defied as sepsis-2 according to the sepsis-2 definition [3] . sepsis-3 was defined as having a suspected site of infection and organ dysfunction (an acute change in the total sequential organ failure assessment (sofa) score of ≥ 2 points consequent to the infection) [1] . regarding shock, sepsis-2 and sepsis-3 shocks were defined according to the sepsis-2 [6] and sepsis-3 definitions, respectively [1] (supplemental file 1). frailty was defined according to the clinical frailty scale (cfs), an easy and intuitive determinable categorization tool based on simple visual descriptions [7] . patients' status for cfs before hospital admission was obtained from patients themselves or their relatives. infection sites at final diagnosis included the lung, intraabdominal, urinary tract, soft tissue, central nervous system (cns), osteoarticular, endocardium, wound, catheter-related, implant device-related, others, or unidentified infections. the diagnosis of the infection site was recorded at discharge. acute physiology and chronic health evaluation ii (apache ii) score was calculated at the initial examination instead of the worst data within 24 h. if apache ii score was missing, zero was used instead of missing data. sofa score was calculated similarly as the apache ii score. vfd was defined as the number of days within the first 28 days postadmission that the patient can breathe without a ventilator. the vfd of patients who died during the study period was set as zero. icu-free days were calculated similarly with vfd. status at discharge was categorized as home, transfer to another facility (including longterm care and nursing homes), or death. patients with infection in icus were compared according to whether they met sepsis-2 or sepsis-3 definition. patients were divided into five groups: sepsis-2; sepsis-3; sepsis-2 and sepsis-3; sepsis-2 and no sepsis-3; no sepsis-2 and sepsis-3; and no sepsis-2 and no sepsis-3. descriptive statistics included proportions for categorical variables, and medians (interquartile range [iqr]) of continuous variables were calculated because not all variables were normally distributed. a few missing data was considered missing randomly. no assumptions were made on these data. all statistical analyses were performed using the stata software version 15.1 (statacorp, tx, usa). a total of 618 patients with suspected infection admitted to 22 icus during the study period were included in this study. among them, 530 (85.8%) patients had sepsis-2, and 569 (92.1%) had sepsis-3; most patients were overlapped; however, patients with sepsis-2 and sepsis-3 were different individuals. a total of 501 (81.1%) met both definitions, and 29 exclusively met the sepsis-2 (only baseline sofa for sepsis-3 glasgow coma scale 12 (7-14) 12 (7-15) 11 (7) (8) (9) (10) (11) (12) (13) (14) 14 (11sepsis-2), and 68 exclusively met the sepsis-3 (only sepsis-3) definition. a total of 20 patients did not meet either of the definitions (fig. 1) . majority of patients were admitted to the icus directly from the emergency departments (eds) (56.6%). a total of 592 (95.8%) patients were positive for sirs, and 368 (59.6%) were positive for qsofa. a total of 297 (48.1%) patients met the sepsis-2 shock criteria, and 109 (17.6%) met the sepsis-3 shock criteria. table 1 shows characteristics of patients with infection in icus according to sepsis definitions. the distributions of baseline characteristics such as age, sex, and comorbidities were comparable between sepsis-2 and sepsis-3. the baseline sofa score for the sepsis-2 only group was 4 (iqr, 0-8) although the baseline sofa for other groups was 0 or 1. lactate and blood culture positivity were lower if they did not meet the definitions. the sofa score was lower if they did not meet any definitions. the trend in pathogens and antibiotics according to sepsis definitions was also nonspecific; however, blood culture positivity was lower if patients did not meet any definitions (sepsis-2 or sepsis-3), and carbapenem was more frequently used in patients who met any definitions (table 2) . in-hospital mortality of study population was 19.1%. in-hospital mortality among patients with sepsis-2 and sepsis-3 patients was comparable (21.7% and 19.8%, respectively) ( table 3 ). patients exclusively identified by sepsis-2 or sepsis-3 had a lower mortality (17.2% vs. 4.4%, respectively). no patients died if they did not meet any definitions. patients who met sepsis-3 shock criteria had higher in-hospital mortality than those who met sepsis-2 shock criteria. characteristics and in-hospital mortality were compared according to sepsis-2 and sepsis-3 definitions in this prospective observational cohort of icu patients. almost all patients presenting to icu with infection fulfill both definitions, but what each definition identified is different. however, in-hospital mortality was zero if patients did not meet any definitions. better criteria might be developed by better selection and combination of elements in both definitions. nearly all patients admitted to icu with suspected infection fulfill both definitions in our study as well as in previous studies [8, 9] . in a retrospective cohort study of icus in england, with similar setting as in our study, sepsis-2 and sepsis-3 definitions identified similar populations (92% overlapped), which is consistent with that of our study (81% overlapped). actually, 95% of patients overlapped if sepsis-3 definition was evaluated in sepsis-2 (severe sepsis) population in our cohort. their severity scores, such as the sofa, were derived from an estimation such as a receipt of organ support and could have been over-or underestimated in the study [9] . another study also virtually calculated the sofa score, even though it is one of the most important elements in sepsis-3 definition [8] . previous studies reported some variations of epidemiology by data sources, data acquisition timing, and interpretation of organ failure criteria in sepsis criteria [8, 10, 11] . our prospective study was designed to compare sepsis-2 and sepsis-3 among patients with suspected infection and directly confirmed results of previous studies [8, 9] . however, these minor variations in the precise interpretation of definitions may have not affect characteristic and mortality differences, especially in the icu setting [8] . patients exclusively identified with sepsis-2 or sepsis-3 had different characteristics when compared to patients with both sepsis-2 and sepsis-3. although a total of 63% patients were diagnosed with sepsis-3 using qsofa, only 29% of patients with sepsis-2(−)/sepsis-3(+) were diagnosed with sepsis-3 using qsofa. patients exclusively identified with sepsis-2 or sepsis-3 presumably included those who have clinically unmeasured features such as vague symptoms [12] . patients who meet only one definition may need more attention because their symptoms were not prominent. since sepsis-2 captures high level of inflammation, a patient with sepsis-2 would still need further attention; however, a negative sepsis-3 does not meet the current definition of sepsis. in-hospital mortality differed by approximately four times between the sepsis-2(+)/sepsis-3(−) group and the sepsis-2(−)/sepsis-3(+) group (17% vs. 4%). although the number of patients was small, judging by one definition alone may cause misclassification of poor outcome patients who may be identified by the other definition. however, since they were actually in the icu, the physician did not misclassify the patients. this highlights the limitations of both the definitions. since sepsis-2 does not include an increased acute sofa score from the baseline, any chronic organ failure may possibly be regarded as an acute organ failure. sofa score was not identified even though sepsis-2 was defined based on this score, except for chronic organ failures. in our cohort, 96 (16%) patients had "not available" (na) sepsis-3 baseline sofa, which was indicated as zero according to the sepsis-3 definition, although all data of chronic organ failures were tried to obtain. therefore, a number of patients with unknown chronic organ failures at baseline should have been included in those with acute organ failure in any definitions. either way, the sepsis-3 definition has become more clinically objectively understandable than the sepsis-2 definition. moreover, the sepsis-3 definition was originally easier to evaluate than the sepsis-2 definition. excluding sirs as the starting point for sepsis-3 did not affect the incidence as majority of patients with organ failures also tend to have sirs. the sepsis-3 shock was associated with a higher risk of death than sepsis-2 [13] , because the sepsis-3 shock requires the presence of elevated serum lactate levels in addition to fluidresistant hypotension [14] . the problem of sepsis diagnosis has been a little arbitrary, with differences in epidemiology. the sepsis-3 definition may be advantageous because it may increase the comparability of sepsis incidence and related mortality among studies by possibly reducing the subjective interpretation [8] . a consistent diagnosis of sepsis and septic shock between institutions should be considered not only for research purposes but also for quality measurement. generally, severity scores such as sofa were better used for clinical research and quality measurement rather than risk assessment. therefore, the definition of sepsis has undoubtedly dramatically advanced the research due to enhanced medical research efficiency when agreed disease and outcome definitions are used. however, definitions are still insufficient and have not been used beyond as tools of research and quality measurement. when a patient was admitted to icu due to an infection, his or her mortality rate was approximately 20% in this cohort. this will make little contribution even if new criteria are used because 20% is one of the highest mortality in icu diseases. the advancement of sepsis definitions may lead to the concept that similar conditions were caused by infections, despite the different backgrounds and triggers. however, risk stratifications and predictions should be investigated in detail in the future. for example, the upgrade or downgrade type of sepsis should be assessed based on the immune response, a subgroup for site of infection, or a phenotype of treatment responsiveness. "one size fits all approach" has reached its limits. this study has several limitations. first, organ failure data before the icu admission were missing in some patients, which was also noticed in the original sepsis-3 study [1] . second, regarding to apache ii score, data at initial diagnosis were used instead of the worst data within 24 h of icu admission because of availability. this may have led to underestimation of the severity of patient conditions. third, missing data were indicated as zero in the apache ii and sofa scores, if some elements were missing. this would have been used to identify any underestimation of the variance of patient's severity. however, effects of missing data should be small because missing data of elements were few. a majority of the patients who were admitted to the icu with suspected infection met sepsis-2 and sepsis-3 definitions. in-hospital mortality was indicated as zero if patients did not meet any sepsis definitions. supplementary information accompanies this paper at https://doi.org/10. 1186/s40560-020-00465-0. the third international consensus definitions for sepsis and septic shock (sepsis-3) classification of sepsis, severe sepsis and septic shock: the impact of minor variations in data capture and definition of sirs criteria sis international sepsis definitions conference of critical care medicine consensus conference: definitions for sepsis and organ failure and guidelines for the use of innovative therapies in sepsis the surviving sepsis campaign: results of an international guidelinebased performance improvement program targeting severe sepsis surviving sepsis campaign: international guidelines for management of severe sepsis and septic shock: 2012 clinical frailty scale (cfs) reliably stratifies octogenarians in german icus: a multicentre prospective cohort study robustness of sepsis-3 criteria in critically ill patients epidemiology of sepsis and septic shock in critical care units: comparison between sepsis-2 and sepsis-3 populations using a national critical care database severe sepsis cohorts derived from claims-based strategies appear to be biased toward a more severely ill patient population variation in identifying sepsis and organ dysfunction using administrative versus electronic clinical data and impact on hospital outcome comparisons presenting symptoms independently predict mortality in septic shock: importance of a previously unmeasured confounder the influence of a change in septic shock definitions on intensive care epidemiology and outcome: comparison of sepsis-2 and sepsis-3 definitions sepsis definitions task f, et al. developing a new definition and assessing new clinical criteria for septic shock: for the third international consensus definitions for sepsis and septic shock (sepsis-3) publisher's note springer nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations we thank the jaam spice study group for the contribution to this study. we would like to thank enago (https://www.enago.jp) for english language editing. this work was supported by jsps kakenhi grant number jp19k19376.notation of prior abstract publication/presentation we will present this research at 33rd annual congress european society of intensive care medicine (esicm) 2020. authors' contributions ta contributed to the acquisition of data, conceived of and designed this study, interpreted the data, drafted the manuscript, and revised the manuscript for important intellectual content. ky contributed to the acquisition of data, conducted data cleaning, interpreted the data, and revised the manuscript for important intellectual content. all of the authors contributed to the acquisition of data, and reviewed, discussed, and approved the final manuscript. this study was supported by the japanese association for acute medicine (2014-01). the datasets generated during and/or analyzed during the current study are available from the corresponding author on reasonable request. the study protocol was reviewed and approved by the research ethics committee of all participating institutions at the japanese association for acute medicine (jaam) spice study group. given the retrospective and anonymized nature of this study in the routine care, the ethics committees waived the need for informed consent from the study participants. the institutional review board of hokkaido university, a leading institution in spice, approved this study (approval no. 016-0386). competing interests all authors declare that they have no competing interests. key: cord-302379-jh6jxwyn authors: jevon, phil; abdelrahman, ahmed; pigadas, nick title: management of odontogenic infections and sepsis: an update date: 2020-09-25 journal: br dent j doi: 10.1038/s41415-020-2114-5 sha: doc_id: 302379 cord_uid: jh6jxwyn the management of odontogenic infections has improved over recent decades, but further improvements are still required. the ongoing education of gdps and their dental teams on this issue continues to be important, especially during the current covid-19 pandemic, where remote triage poses additional difficulties and challenges. odontogenic infections can lead to sepsis, a potentially life-threatening condition caused by the body's immune system responding in an abnormal way. this can lead to tissue damage, organ failure and death. a patient with non-odontogenic-related infection could also present with sepsis at a dental practice. early recognition and prompt management of sepsis improves outcomes. gdps and their dental teams should be trained in the recognition and management of sepsis. age-specific sepsis decision support tools have been developed by the uk sepsis trust to help dental staff recognise and manage patients with suspected sepsis. the aim of this article is to provide an update on the management of odontogenic infections and sepsis. although the management of odontogenic infections has improved over recent decades, further improvements are needed and the ongoing education of gdps and their dental teams on this issue is essential. in addition, the covid-19 pandemic has imposed new difficulties and challenges; for example, telephone triage and prescription of antibiotics, and it is important to be up-to-date with current guidelines. 1, 2 odontogenic infections can lead to sepsis, 3,4 a potentially life-threatening condition caused by the body's immune system responding abnormally. this can lead to tissue damage, organ failure and death. 5 a patient with nonodontogenic-related infection could also present with sepsis at a dental practice. early recognition and prompt effective treatment of sepsis improves outcomes. 5 the dental team should be trained in the principles of the management of sepsis. 6 agespecific sepsis decision support tools have been developed by the uk sepsis trust 7 to assist the dental team to recognise and manage patients with suspected sepsis. the aim of this article is to provide an update on the management of odontogenic infections and sepsis in the dental practice. the morbidity and mortality rate of odontogenic infections has dropped significantly over the past 70 years. 8, 9, 10 this dramatic drop is undoubtedly linked to the discovery of antibiotics, the improvement of the general population health standards, and a better understanding of appropriate medical and surgical management of these cases. further improvements are needed and ongoing education of the dental team on this issue is very important. odontogenic infections pass through three key stages: 11 • stage 1: 1-3 days; soft and mildly tender swelling • stage 2: 2-5 days; hard, red and severely sore swelling • stage 3: 5-7 days; abscess formation. there is a strong belief that once the abscess has formed, surgical drainage is mandatory to achieve resolution. 12 medical management has a role in selected cases. 13 seven principles have been proposed to achieve the best outcome in managing odontogenic infections: 11 1. establish the severity of the infection 2. assess host defences 3. elect the setting of care 4. surgical intervention 5. medical support 6. antibiotic therapy 7. frequently evaluate the patient. a careful history and thorough clinical examinations are essential to determine the severity of any infection. history-taking will highlight factors like immune system competence and the level of systemic reserves to fight infections. a physical examination can identify clinical observations outside normal limits. several clinical and haematological parameters have been used as prognostic indicators for the severity of the infection. c-reactive protein (crp), fever and anatomical locations have been investigated for the assessment of the extent of odontogenic infections and presumed duration of hospital stay. 14 additional factors must be considered to establish the infection severity: • anatomical location • airway compromise. there are a number of potential spaces between the musculoskeletal head and neck structures and the regional fasciae and organs better known as fascial spaces. a summary of these spaces and their level of risk 11, 12 is found in table 1 . in healthy and systemically well patients without trismus, infections of low-risk spaces can be initially treated in a primary care dental practice, while infections spreading to higher risk spaces should be managed more aggressively and may need to be treated in a secondary care centre. ludwig's angina was described by karl friedrich wilhelm von ludwig in 1836 as a rapidly and frequently fatal progressive gangrenous cellulitis and ooedema of the soft tissues of the neck and floor of the mouth. 15 thankfully, mortality rates have reduced significantly with the introduction of antibiotics, improved oral and dental hygiene, and timely surgical intervention. 15 the majority of ludwig's angina infections are odontogenic; 16 peritonsillar or parapharyngeal abscesses, mandibular fractures, oral lacerations/piercing and submandibular sialoadenitis are other recognised causes. 15 a compromised airway is synonymous with ludwig's angina and the initial assessment of a patient with ludwig's angina should follow the familiar 'airway, breathing, circulation, disability, exposure' (abcde) approach. signs of a compromised airway in these patients could include noisy (gurgling) breathing with drooling saliva, stridor, dyspnoea, tachypnoea, tachycardia, dysphagia and trismus. the initial immediate management usually includes positioning the patient in an upright position and administering oxygen 15 litres/minute, 15 while colleagues call 999 for an ambulance. a healthy immune system is essential to the maintenance of host defence against infection. multiple medical conditions can affect it. 17 box 1 summarises the common factors that can cause immune system compromise. the concept of 'physiologic reserve' represents a significant driver of outcome in patients fighting infection. this can be defined as the capability of an organ to carry out its activity under stress. 18 age is an essential factor that is inversely related to the physiologic reserve; that is, decreased respiratory, cardiovascular and metabolic reserve. 19 elect the setting of care an uncomplicated localised dental abscess in a healthy young person, who does not show signs and symptoms of a worsening immune response, can be safely treated in a dental practice. early and adequate intervention is essential in order to prevent avoidable deterioration with invasion of adjusted anatomical spaces and symptoms of sepsis ( fig. 1) . similarly, severe neck infection in an immunocompromised elderly person warrants treatment in a secondary care setting. the clinical decision to choose the setting of care is not always straightforward though, prompting the need for clear secondary care referral criteria. although there are no agreed national guidelines on when to admit to a secondary care setting, criteria for hospital admission have been proposed 2,20 (box 2). a careful history, thorough clinical examinations and a high index of suspicion will enable the gdp to diagnose and appropriately manage patients presenting with odontogenic sepsis. early surgical intervention has been advocated to improve the clinical outcome of odontogenic infection. the dramatic improvement in the outcome of sever odontogenic infection is directly linked to the immediate establishment of a safe airway, followed by early surgical intervention. once the airway has been deemed patent and not at risk of being compromised, in either a hospital setting or dental practice, the principles of management are very similar. thorough knowledge of head and neck anatomy will enable the surgeon to access the abscess cavities using incisions in safe places without damaging any vital structures like blood vessels or nerves. most of the odontogenic infections can be drained through intraoral access. five principles must be followed: 11 21 although abscess formation takes place between the fifth and seventh days, early elimination of the infection source and surgical intervention will decompress the involved anatomical spaces. 14 relying on antibiotics only in relieving dental infection is likely to be less effective and can cause antimicrobial resistance. 22 two of the challenges to performing adequate drainage of any odontogenic infection in dental practice are: 23 1. achieving adequate local anaesthesia 2. risk of spreading the infection to other anatomical spaces. the ability to deliver safe, adequate local anaesthesia is essential for any dental procedure. the mechanism of action of the local anaesthetic solution depends on the tissue ph. in the presence of infection, tissue ph becomes more acidic, which slows down the degree of ionisation, resulting in less optimal or failed anaesthesia. 23 to overcome this problem, the injection of the anaesthetic solution at a distance from the inflammatory site is required (nerve blocks). it will also avoid infection spread to different tissue spaces. although surgical drainage is the classic approach to most of the odontogenic infection, medical support has a critical role in controlling the disease. 24 adequate hydration, nutrition and control of fever are essential to optimise the medical care for patients presenting with odontogenic infections. stabilisation of any underlying systemic disease (for example, uncontrolled diabetes) is extremely important. 24 box 2 criteria for referral to secondary care 2, 20 • difficulty in swallowing and dehydration first-line antibiotics for dental abscess in dental practices (adults and children more than 12 years) odontogenic infections are multi-microbial with a combination of facultative and anaerobes species. facultative streptococcus viridans group are commensal gram-positive bacteria and include s. anginosus, s. intermedius and s. constellatus. these organisms are abundant in the mouth and most frequently associated with orofacial cellulitis and abscess. after a few days, the anaerobes (prevotella and porphyromonas) predominate. the majority of the facultative streptococci that cause odontogenic infections are sensitive to penicillin. 25 approximately a quarter of strains of prevotella and porphyromonas are penicillin-resistant. 26 the scottish dental clinical effectiveness programme (sdcep) has published evidencebased guidance on antibiotic prescription in dental practice. penicillin-based antibiotics remain the first line for the treatment of odontogenic infections. metronidazole is effective against anaerobic bacteria. 1, 27, 28 the antibiotic doses recommended in the sdcep's guidance are based on the doses recommended by the british national formulary (bnf) 13 (table 2 ). in secondary care settings, the antibiotics are prescribed in accordance with the local hospital antimicrobial therapy. consultation with the on-call microbiologist is a common practice for severe cases and cases which are not responding to first-line treatment. the last principle, but as vital as the previous ones, is the periodic re-evaluation of these patients. in outpatient settings, the recommended follow-up is after two days. 29 forty-eight hours will allow the drainage to cease and the immune system to overcome the initial insult from the infection. if no improvement or deterioration of symptoms is noted, further escalation in care must be provided. the review interval, however, depends on the clinical course of the infection. a patient with a rapidly developing swelling and mild temperature may need review within 24 hours, but a patient with a chronic abscess and no systemic symptoms will need to be reviewed at the end of the antibiotic treatment. causes of treatment failure include: • failure to remove the source of infection • underlying systemic disease; for example, uncontrolled diabetes • antibiotic-related factors -patient non-compliance, drug not reaching site secondary to inadequate drainage, wrong antibiotic choice or incorrect dose. in hospital settings, more frequent evaluations are essential as the disease is expected to be more aggressive. the covid-19 pandemic has dramatically changed dental practice since march 2020. guidance on the management of acute dental problems is available. 2, 30 this is likely to change as the situation evolves. advice, analgesia and antimicrobials (when indicated) should form the basis of primary care dental triage when using remote consultation (telephone call or video call). 1 while assessing the patient, covid-19 status should be established and documented, as this will determine how the patient's care will be managed should referral to an urgent dental care centre or secondary care be required. patients should be advised that dental treatment options are currently severely restricted and that they should call back in 48-72 hours if their symptoms have not resolved. 2 the sdcep's flowchart (fig. 2) helps the remote management of patients by guiding the gdp to categorise the patient into one of three management groups. 2 the sdcep has also recently updated their drugs for the management of dental problems worryingly, there has been a rise in anecdotal reports of antibiotics apparently being overprescribed for dental pain since the outbreak of covid-19. 31 this pandemic has demonstrated the havoc a pathogen can unleash when we have no protection against it. inappropriate use of antibiotics increases the likelihood that resistant bacteria will evolve 31 and it is essential that gdps remain guardians against antimicrobial resistance. 1,32 antibiotics should only be prescribed if it is likely that the patient has a bacterial infection, and the principles of prescribing and follow-up (as detailed earlier) should be followed. it is estimated that 234,000 patients develop sepsis in the uk every year, 5 with 70% of sepsis cases originating in the primary care setting. annually, there are approximately 44,000 deaths from sepsis in the uk 5 and six million deaths worldwide. 33 although deaths from sepsis due to odontogenic infection are very rare, they have been reported. 34 the incidence of sepsis is on the increase, possibly due to: 35 • a growing elderly population • an increased use of invasive surgery • an increased incidence of bacterial resistance • an increased number of immunocompromised patients. a localised infection which progresses into an uncontrolled systemic response is usually the cause of sepsis. progression to acute physiological deterioration with the risk of multiple organ failure and death can be swift. • pneumonia: 50% • urinary tract: 20% • abdomen: 15% • skin, soft tissue, bone and joint: 10% • endocarditis: 1% • device-related infection: 1% • meningitis: 1% • others: 2%. in normal circumstances, the body's immune system will prevent or fight infection (bacteria, viruses, fungi). however, the immune system can sometimes go into overdrive, resulting in vital organs and other tissues being targeted. this can result from any injury or infection in the body. although a wide variety of different microorganisms (for example, streptococcus, e. coli, mrsa or clostridium difficile) can cause sepsis, it is usually caused by common bacteria that don't normally make patients ill. 36 any infection can lead to sepsis (box 3), though pneumonia (commonly referred to as chest sepsis) is the cause in half of the cases. 5 the national institute for health and care excellence (nice) 6 the uk sepsis trust has developed age-specific sepsis decision support tools to assist the dental team to assess both adult and paediatric patients who may have sepsis. 7 utilisation of these sepsis decision tools will help determine if red flag sepsis (see below) is present, prompting appropriate timely action. the prompt transfer of patients presenting with orofacial infections suspected of sepsis to an acute hospital setting for early treatment should ultimately improve sepsis survival rates. 37 the care quality commission (cqc) 38 endorses these sepsis decision tools and, ideally, all three should be readily available in the dental practice. the 'gdp sepsis decision support tool for primary dental care' (fig. 3) should be applied to all adults and young people aged 12 years and over with fever (or recent fever), symptoms presenting with a source of orofacial/dental infection (including post-operative infection) or have clinical observations outside normal limits. 7 it details what to look out for if the patient has presumed infection and, in particular, what constitutes red flag sepsis. red flag sepsis is a definition from the uk sepsis trust which lists a set of easyto-assess clinical parameters, the presence of one of which in the context of infection identifies sepsis with a high risk of death and a requirement for urgent treatment (fig. 3) . 7 if red flag sepsis is present: there are two paediatric sepsis decision tools, one for children aged 5-11 years (fig. 4) and one for children <5 years (fig. 5) . these should be used in children who have a suspected source of orofacial/dental infection (including post-operative infection) or have clinical observations outside the normal range. 7 the paediatric sepsis decision tools take into account paediatric considerations, including differences in paediatric physiology. covid-19 infection can cause sepsis on its own. 5 unfortunately, the sepsis signs and symptoms for a number of initial conditions can be very similar. this stresses the importance for dental teams to be familiar with sepsis and the decision tools described here for safe management of such patients. in addition, evidence suggests that, for a period of time following sepsis, patients may be vulnerable and develop further infections including covid-19; therefore, they have an increased risk of readmission with infective complications (including sepsis). 5 the nice advises that patients with suspected sepsis are assessed following a structured set of observations to stratify the risk of acute illness or death. 6 the royal college of physicians' national early warning score (news) 2 is widely used by the ambulance service and in hospitals, and reliably detects deterioration in adults, triggering review, treatment and escalation of care, particularly sepsis. 41 although news2 hasn't yet been validated for use in primary care, nhs england is encouraging its widespread use in this sector. 42 the cqc has created a webpage titled 'dental mythbuster 25: sepsis' on its website, 38 providing helpful information relating to the management of sepsis in the dental practice, including online links to professional guidelines (nice and uk sepsis trust) as well as what to expect from the cqc, relating to sepsis, when they review dental practices to determine whether they are safe and well-led. when reviewing dental practices, the cqc will ask dental staff what systems and processes are in place to manage a patient with a bacterial infection, including procedures for follow-up and referral for specialist care when necessary. this will include treating patients who: • are not responding to conventional oral antibiotic treatment • cannot have their infection drained at an initial appointment. the cqc will also ask what advice is given to patients, including when they should seek emergency advice or treatment, if symptoms worsen or when the dental surgery is closed. odontogenic infections can lead to sepsis, which can result in tissue damage, organ failure and death. this article has outlined the management of odontogenic infections, including the latest covid-19 guidelines. drugs for the management of dental problems during covid-19 pandemic management of acute dental problems during covid-19 pandemic severe odontogenic infections with septic progress -a constant and increasing challenge: a retrospective analysis medical emergencies: sepsis in primary dental care professional resources sepsis: recognition, diagnosis and early management nice guideline uk sepsis trust. clinical tools. 2020. available online at changing trends in deep neck abscess. a retrospective study of 110 patients deep neck abscesseschanging trends life-threatening oro-facial infections peterson's principles of oral and maxillofacial surgery deep space neck infection: principles of surgical management prescribing in dental practice increasing frequency and severity of odontogenic infection requiring hospital admission and surgical management ludwig's angina ludwig's angina management of head and neck infections in the immunocompromised patient organ reserve, excess metabolic capacity, and aging influence of aging and environment on presentation of infection in older adults criteria for admission of odontogenic infections at high risk of deep neck space infection irrigating drains for severe odontogenic infections do not improve outcome antibiotic selection in head and neck infections pharmacology of local anaesthetics used in oral surgery is conservative treatment of deep neck space infections appropriate? evaluation of bacterial spectrum of orofacial infections and their antibiotic susceptibility severe odontogenic infections. part 2. prospective outcomes study management of acute dental problems: guidance for healthcare professionals sdcep. drug prescribing for dentistry: dental clinical guidance deep neck infection covid-19 guidance and standard operating procedure when to prescribe antibiotics royal college of surgeons, faculty of general dental practice (uk) & bda. open letter on prescribing antibiotics during covid-19 assessment of global incidence and mortality of hospital-treated sepsis. current estimates and limitations death from overwhelming odontogenic sepsis: a case report sepsis guidance implementation advice for adults sepsis decision support tool for primary dental care dental mythbuster 25: sepsis resuscitation council uk. the abcde approach basic guide to medical emergencies in the dental practice: second edition news) 2: standardising the assessment of acute-illness severity in the nhs national early warning score (news) the recognition and management of sepsis in the dental practice has also been discussed, including the age-specific sepsis decision support tools developed by the uk sepsis trust. key: cord-001319-mlkaowqr authors: giamarellos-bourboulis, evangelos j; apostolidou, efterpi; lada, malvina; perdios, ioannis; gatselis, nikolaos k; tsangaris, iraklis; georgitsi, marianna; bristianou, magdalini; kanni, theodora; sereti, kalliopi; kyprianou, miltiades a; kotanidou, anastasia; armaganidis, apostolos title: kinetics of circulating immunoglobulin m in sepsis: relationship with final outcome date: 2013-10-21 journal: crit care doi: 10.1186/cc13073 sha: doc_id: 1319 cord_uid: mlkaowqr introduction: the aim of this study was to investigate the kinetics of immunoglobulin m (igm) during the different stages of sepsis. methods: in this prospective multicenter study, blood sampling for igm measurement was done within the first 24 hours from diagnosis in 332 critically ill patients; in 83 patients this was repeated upon progression to more severe stages. among these 83 patients, 30 patients with severe sepsis progressed into shock and igm was monitored daily for seven consecutive days. peripheral blood mononuclear cells (pbmcs) were isolated from 55 patients and stimulated for igm production. results: serum igm was decreased in septic shock compared to patients with systemic inflammatory response syndrome (sirs) and patients with severe sepsis. paired comparisons at distinct time points of the sepsis course showed that igm was decreased only when patients deteriorated from severe sepsis to septic shock. serial measurements in these patients, beginning from the early start of vasopressors, showed that the distribution of igm over time was significantly greater for survivors than for non-survivors. production of igm by pbmcs was significantly lower at all stages of sepsis compared with healthy controls. conclusions: specific changes of circulating igm occur when patients with severe sepsis progress into septic shock. the distribution of igm is lower among non-survivors. although initially considered a state of hyperactivity of the innate and adaptive immune systems, it is currently understood that severe sepsis and septic shock are characterized by a functional state of immunoparalysis [1] . this involves not only monocytes and macrophages, but also cd4 lymphocytes and b lymphocytes [2] . under normal conditions, cd4 lymphocytes orchestrate b lymphocyte responses for the secretion of the polyvalent immunoglobulin m (igm) antibodies that are of crucial importance for the opsonization and the subsequent rapid clearance of the invading microorganisms [3] . immunoparalysis of sepsis is characterized by defective b-lymphocyte responses toward low immunoglobulin production [2] . to this end, it was expected that the intravenous administration of immunoglobulin preparations enriched in igm would be beneficial for patients with severe sepsis and septic shock. on the contrary, most of the conducted randomized clinical trials (rct) yielded contradictory results [4, 5] , despite one meta-analysis indicating that igm preparations significantly decrease the relative risk of death in both adult and child populations [4] . the existing controversies of conducted rcts may derive from our incomplete understanding of the kinetics of igm over the time course of sepsis. the current study was designed in order to embed into the changes of circulating igm levels of patients upon progression to the more severe stages of sepsis in relation with the production of igm from circulating lymphocytes and with the final outcome. this prospective multicenter study was conducted from january 2010 to december 2010 in 27 departments across greece participating in the hellenic sepsis study group. the participating departments were 15 intensive care units (icus), seven departments of internal medicine, two departments of pulmonary medicine, two departments of surgery and one department of urology. patients with signs of systemic inflammatory response syndrome (sirs) either admitted to the emergency department or hospitalized in the general ward or in the icu were eligible. written informed consent was provided by the patients or by their first-degree relatives for patients unable to consent. inclusion criteria were: (a) age ≥18 years; (b) diagnosis of sirs, sepsis, severe sepsis or septic shock; and (c) sirs due to acute pancreatitis or sepsis due to specific infections. these infections were: community-acquired pneumonia (cap), ventilator-associated pneumonia (vap), acute pyelonephritis (uti), acute intra-abdominal infection (iai) and primary bacteremia (bsi); and (d) first blood sampling within 24 hours from diagnosis. exclusion criteria were (a) infection by the human immunodeficiency virus type 1; (b) neutropenia defined as less than 1,000 neutrophils/mm 3 ; (c) chronic intake of corticosteroids defined as systemic intake of more than 1 mg/kg of equivalent prednisone for more than one month; and (d) other types of immunodeficiency like organ transplantation, hematologic malignancies and intake of chemotherapy. sirs was diagnosed by the presence of at least two of the following [6] : (a) core temperature >38°c or <36°c, (b) p co2 <32 mmhg or more than 20 breaths/min, (c) pulse rate >90/min, and (d) white blood cells >12,000/mm 3 or <4,000/mm 3 or >10% of band forms. sepsis was defined as any microbiologically or clinically documented infection complicated by sirs. patients with sepsis were classified as suffering from uncomplicated sepsis, severe sepsis or septic shock, according to standard definitions [6] . multiple organ dysfunctions syndrome (mods) was defined by the same criteria [6] . acute pancreatitis, cap, vap, uti, iai and bsi were defined according to standard definitions [7] [8] [9] [10] [11] . for each patient a complete diagnostic workup was performed comprising history, thorough physical examination, white blood cell (wbc) count, blood biochemistry, arterial blood gas, blood cultures from peripheral veins and central lines, urine cultures, chest x-ray and chest and abdominal computed tomography if appropriate. if necessary, quantitative cultures of tracheobronchial secretions (tbs) or bronchoalveolar lavage (bal) were performed and evaluated as previously described [9] . survival was recorded for 28 days and at hospital discharge. clinical and demographic data were recorded on a case report form (crf). all crfs were monitored by an independent monitor blinded to the study design. for all enrolled patients and for 35 healthy volunteers 5 ml of blood was sampled within the first 24 hours from diagnosis. from this volume: (a) 2 ml was collected into sterile, pyrogen-and anticoagulant-free tubes (vacutainer, becton dickinson, cockeysville, md, usa) for quantitative measurement of igm; and (b) 3 ml was collected into edta-coated tubes (vacutainer) for the measurement of the absolute counts of b lymphocytes. from 55 patients and 20 healthy volunteers another 8 ml was collected into heparin-coated tubes (vacutainer) and used for the isolation of peripheral blood mononuclear cells (pbmcs). for 83 patients, blood sampling was repeated on the day of worsening of sepsis stage. for 30 patients who progressed into septic shock, blood was sampled daily for seven days starting immediately after the start of vasopressors. tubes were transported by a courier service within the same day to the laboratory of immunology of infectious diseases of the 4 th department of internal medicine at attikon university hospital of athens. tubes were centrifuged and serum was kept frozen at -70°c until assayed. igm was estimated in duplicate by an enzyme-linked immunosorbent assay (e-bioscience inc., san diego, ca, usa) following the manufacturer's instructions; the lower detection limit was 20 ng/ml. all estimations were performed and reported by two technicians who were blinded to clinical information. the central laboratory of the study participates in the uk neqas quality control system for leukocyte immunophenotyping (registration number 40926). in this laboratory, the absolute count of b lymphocytes was measured as described elsewhere [12] . briefly, red blood cells were lysed with ammonium chloride 1.0 mm. white blood cells were washed three times with phosphate-buffered saline (pbs) (ph 7.2) (merck, darmstadt, germany) and subsequently incubated for 15 minutes in the dark with the monoclonal antibody anti-cd19 at the flurochrome fluorescein isothiocyanate (fitc, emission 525 nm, immunotech, marseille, france) using fluorospheres (immunotech) for the determination of absolute counts. one igg isotypic negative control at the fluorocolor fitc was analyzed for every patient. cells were analyzed after running through the epics xl/msl flow cytometer (beckman coulter, inc., miami, fl, usa) with gating for mononuclear cells based on their characteristic forward scatter/side scatter (fs/ss) scattering. the isolation of pbmcs was limited to 55 patients because these samples should come from patients hospitalized at study sites close to the central laboratory. this allowed the time from blood collection until processing to be less than 30 minutes. as such, pbmcs were studied from patients hospitalized at the attikon university hospital that is close to the central laboratory of the study. production of igm was studied according to a procedure described elsewhere [13] . heparinized venous blood was layered over ficoll hypaque (biochrom, berlin, germany) and centrifuged for 20 minutes at 1400 g. separated pbmcs were washed three times with ice-cold pbs (ph: 7.2) (biochrom) and counted in a neubauer chamber. their viability was more than 99% as assessed by trypan blue exclusion of dead cells. they were then diluted in rpmi 1640 enriched with 2 mm of l-glutamine, 10% fetal bovine serum (biochrom), 100 u/ml of penicillin g, 100 μg/ml of gentamicin and 10 mm of pyruvate and suspended into wells of a 96-well plate (greiner, alphen a/d rijn, the netherlands). the final volume per well was 200 μl with a density of 2 ×10 6 cells/ml. pbmcs were incubated in the absence or presence of 5 μg/ml of the lymphocyte agonist phytohemagglutin (pha) of phaseolus vulgaris (pha-l, roche diagnostics gmbh, mannheim, germany) for 24 or 72 hours at 37°c in 5% co 2 atmosphere. at the end of the incubation, the plates were centrifuged. supernatants were kept stored at -70°c until assayed. concentrations of igm were measured at the end of the 72-hour incubation period; those of tumor necrosis factor alpha (tnfα) were measured at the end of the 24-hour incubation period. measurements were done in duplicate and igm was measured as described above. tnfα was measured by an enzyme-linked immunosorbent assay (r&d minneapolis, mi, usa). the lower detection limit was 40 pg/ml. the primary endpoint was the over-time changes of igm serum levels of patients upon progression to septic shock in relation with the final outcome that is survival or 28-day mortality. the secondary study endpoint was the impact of sepsis on production of igm from circulating lymphocytes. demographic characteristics of enrolled patients were provided as percentages for qualitative variables and as means ± standard error of the mean (se) or medians and interquartile ranges for quantitative variables. comparisons between groups were done by the x 2 test for qualitative variables and by anova with post hoc bonferroni corrections for quantitative variables. serum concentrations of igm were expressed as medians and 95% confidence intervals (ci). comparisons between groups were done by the mann-whitney u test with corrections by bonferroni for multiple testing. paired comparisons of serum igm at baseline and upon progression of the same patients to a more severe stage were done by the wilcoxon's rank sum test. for every patient with septic shock, the area under the curve (auc) of igm over time for seven days was measured by the linear trapezoidal rule. comparison between survivors and non-survivors was done by student's t test. concentrations of igm and of tnfα in supernatants of pbmcs were expressed as means ± se. comparisons between groups were done by the kruskal-wallis test with corrections by bonferroni for multiple testing. patients were also divided into 'non-igm' and 'igm-producers' if the concentrations of igm in supernatants of pbmcs were below the limit of detection or not. comparisons were done by the x 2 test. values of p below 0.05 were considered significant. a total of 351 patients were screened and 332 were enrolled ( figure 1 ). the demographic characteristics of enrolled patients in relation with the severity of critical illness are shown in table 1 . as expected, acute physiology and chronic health evaluation ii (apache ii) score, wbc counts, c-reactive protein and mortality were greater in the more severe stages of sepsis. the study end point was the kinetics of serum igm upon progression from severe sepsis to septic shock in relation with final outcome. to reach this end point, a three-step approach was followed ( figure 1 ) (a) circulating igm was compared between critically ill patients with varying severity; (b) circulating igm was measured at baseline and on the first day of worsening; and (c) distribution of igm was compared over time between survivors and nonsurvivors from septic shock. median igm was 44.1 mg/dl in healthy volunteers; 34.9 mg/dl in sirs; 23.0 mg/dl in sepsis; 36.2 mg/dl in severe sepsis; and 21.9 mg/dl in septic shock. statistical analysis showed that serum igm was decreased in septic shock compared to healthy volunteers (p = 0.001), to patients with sirs (p = 0.028) and to patients with severe sepsis (p <0.0001) but not to patients with uncomplicated sepsis (p = 0.754) (figure 2 ). paired comparisons of igm were done for 83 patients at two specific time points: on initial diagnosis and on worsening. these measurements involved: 13 patients initially diagnosed with uncomplicated sepsis who worsened into severe sepsis; 16 patients initially diagnosed with uncomplicated sepsis who worsened into septic shock; 49 patients initially diagnosed with severe sepsis who worsened into septic shock; and five patients initially diagnosed with septic shock who worsened into mods (figure 3 ). from all these paired comparisons, significant changes of circulating igm were found only between severe sepsis and septic shock; igm was significantly decreased upon worsening from severe sepsis into septic shock (p = 0.039). within the enrolled population with septic shock, serum igm did not differ between survivors and nonsurvivors (median igm of survivors 23.1 mg/dl and of non-survivors 20.7 mg/dl, p = 0.442). the time curves of igm were designed for 30 patients with severe sepsis who progressed into septic shock. sampling was started on the day of the start of vasopressors and lasted for seven consecutive days. separate curves were designed for survivors and non-survivors. these curves suggested that circulating igm remained stable and at low levels in non-survivors whereas igm of survivors increased to an early peak and then gradually decreased. as a consequence, the distribution of igm expressed by the auc of serum igm over time was significantly greater for survivors than for non-survivors. this finding was similar both when outcome was assessed after 28 days and at hospital discharge ( figure 4a and 4b). production of igm by pbmcs of 55 patients was also studied. from these patients, 24 had uncomplicated sepsis, 20 severe sepsis and 21 septic shock. respective mean ± sd age was 66.5 ± 18.7, 76.4 ± 9.2 and 60.0 ± 21.4 years; mean ± sd apache ii score was 10.7 ± 5.4, 17.4 ± 4.3 and 23.6 ± 4.8; and mean ± sd white blood cell count was 12,226.4 ± 5,262.0, 16,384.0 ± 11,294.0 and 17,130.9 ± 9,793.7/mm 3 . high production of both igm and tnfα was found by the pbmcs of healthy volunteers after stimulation with the selective lymphocyte agonist pha. production of igm and of tnfα was significantly lower at all stages of sepsis compared with healthy controls (figure 5a and 5b). furthermore, the rate of 'igm producers' was significantly lower among patients with septic shock than among patients at all other sepsis stages ( figure 5c ). the present study is the largest cohort to the best of our knowledge that describes the kinetics of circulating igm in sepsis. analysis indicates that the decrease of igm is a predominant characteristic when a patient with severe sepsis develops septic shock. close monitoring from the start of vasopressors shows that the distribution of igm is greater in survivors than in non-survivors from septic shock. these conclusions are based on the multilevel approach of the current study; at first, comparisons between sirs, sepsis, severe sepsis and septic shock indicated that septic shock is the stage of critical illness with the lower circulating igm; then measurements at distinct time points that is, upon initial diagnosis and upon worsening showed that circulating igm decreases specifically upon progression from severe sepsis to septic shock; and finally, intense monitoring of igm after the start of vasopressors revealed a relationship between lacking distribution of igm and unfavorable prognosis. igm levels in patients with septic shock are reported in two more studies. in the first study [14] , igm was decreased in 21 patients with septic shock. patients were followed up every 48 hours for five days and they were divided into those with hypo igm concentrations and normal igm concentrations. no differences were found between survivors and non-survivors. in the second study [15] , low igm levels were reported in the plasma of 62 patients with septic shock. the igm concentrations reported by the authors of this study were within the range of concentrations reported in our study. however, the authors failed to define any differences in circulating igm between survivors and nonsurvivors [15] . in their study, blood samplings of days 1 and 2, of days 3 and 4 and of days 5 to 7 were reported together which did not allow measurement of the distribution of circulating igm as this was done in our study. igm is a polyvalent immunoglobulin circulating as a pentamer [3] . it opsonizes bacteria and primes phagocytosis by neutrophils; it binds and inactivates endotoxins of gram-negative bacteria and exotoxins of gram-positive cocci; and it also binds and inactivates proinflammatory host mediators like cytokines. its role is underscored by models of experimental sepsis in mice; survival is prolonged after induction of sepsis through cecal ligation and puncture (clp) within the animals that possess the highest potential for igm-primed phagocytosis [16] . recent data coming both from rodents and humans suggest that release of igm is primed by a new subset of b lymphocytes known as ira (innate response activator) b cells. these cells belong to the innate defense system, they contain large cytoplasmic stores of igm antibodies and they are the main effectors of the rapid release of igm. ira b cells are depleted in experimental sepsis and this leads to early death [17] . the evidence coming from experimental animal data may help explain the importance of the ex vivo production of igm from our patient population. all patients produced much lower igm than healthy volunteers; this defect was exaggerated in septic shock. our findings lead to the hypothesis that during severe sepsis lymphocytes are hypofunctional for igm production but high circulating igm compensates for the patient's needs; once septic shock develops circulating igm is fully consumed and lymphocytes are completely anergic for any igm production. two major limitations of the current study should, however, be acknowledged: (a) the lack of explanation why septic shock is a specific condition where circulating igm is depleted. it is most probable that this is related with the consumption of circulating igm during sepsis worsening and with the inability of b lymphocytes for igm production; and (b) the lack of explanation from our findings why circulating igm does not differ between uncomplicated sepsis and septic shock. the present study managed to identify specific changes of the kinetics of circulating igm that are related with final outcome. these occur when patients with severe sepsis progress to septic shock. in these patients, the distribution of igm is lower among non-survivors. these findings may guide the design of future rcts for the management of septic shock. serum levels of igm are significantly decreased in septic shock but not in severe sepsis. dramatic changes of serum igm occur when patients at severe sepsis progress into septic shock. in these cases, the distribution of igm is lower among non-survivors. circulating lymphocytes of patients render anergic for the production of igm. the immune response to severe bacterial infections: consequences for therapy hotchkiss rs: immunosuppression in patients who die of sepsis and multiple organ failure igm in microbial infections: taken for granted? use of polyclonal immunoglobulins as adjunctive therapy for sepsis or septic shock polyclonal intravenous immunoglobulin for the treatment of severe sepsis and septic shock in critically ill adults: a systematic review and meta-analysis sccm/esicm/accp/ats/sis international sepsis definitions conference acute pancreatitis procalcitonin guidance of antibiotic therapy in community-acquired pneumonia. a randomized trial international conference for the development of consensus on the diagnosis and treatment of ventilator-associated pneumonia fever in the clinical diagnosis of acute pyelonephritis the international sepsis forum consensus conference definitions of infections in the intensive care unit early alterations of the innate and adaptive immune statuses in sepsis according to the type of underlying infection grouping of patients with common variable immunodeficiency based on immunoglobulin biosynthesis: comparison with a classification system on cd4-naïve cells γ-globulin levels in patients with community-acquired septic shock assessment of plasmatic immunoglobulin g, a and m levels in septic shock patients presence of pre-existing antibodies mediate survival in sepsis innate response activator b cells protect against microbial sepsis submit your next manuscript to biomed central and take full advantage of: • convenient online submission • thorough peer review • no space constraints or color figure charges • immediate publication on acceptance • inclusion in pubmed, cas, scopus and google scholar • research which is freely available for redistribution the study was funded (a) in part by the hellenic institute for the study of sepsis; and (b) in part by an unrestricted educational grant by biotest ag, dreieich, germany. abbreviations apache ii: acute physiology and chronic health evaluation ii; ards: acute respiratory distress syndrome; bsi: primary bacteremia; cap: communityacquired pneumonia; iai: intra-abdominal infection; igm: immunoglobulin m; ira: innate response activator; mods: multiple organ dysfunction syndrome; pbmc: peripheral blood mononuclear cell; pha: phytohemagglutin; rct: randomized controlled trial; sirs: systemic inflammatory response syndrome; tnfα: tumor necrosis factor alpha; uti: acute pyelonephritis; vap: ventilator-associated pneumonia; wbc: white blood cell. the authors declare that they have no competing interests related to this submission. key: cord-017337-vq3edhxn authors: vincent, jean-louis title: piro: the key to success? date: 2009 journal: management of sepsis: the piro approach doi: 10.1007/978-3-642-00479-7_1 sha: doc_id: 17337 cord_uid: vq3edhxn sepsis continues to represent a major problem in intensive care units worldwide. diagnosis and management are often complex due in part to the remarkably diverse nature of the septic patient. indeed, sepsis can range in severity from mild systemic inflammation of little clinical importance through to a widespread severe inflammatory response with multiple organ failure and a mortality rate in excess of 50%. sepsis can affect individuals of any age group, with no or multiple co-morbidities, and with many different ongoing diagnoses. it can occur as the result of infection by one or more of a multitude of microbial pathogens impacting on any of numerous different sites within the body. given the huge complexity of sepsis and the diverse populations of patients it affects, simple definitions are of relatively little use and a more detailed framework which can be used to better characterize patients with sepsis has been proposed, much as the tnm classification (tumor size, nodal spread, metastases) has been successfully used in clinical oncology. in this chapter, we discuss the development of this piro system, and suggest how it may be used in the future to aid diagnosis, guide therapy, and improve prognostication. over the years, there has been considerable confusion regarding the defi nition of sepsis, with terms such as "infection" and "sepsis" often being used interchangeably. while obviously related, these elements are not exact synonyms; sepsis is the host response to an infection by an invading microorganism, be it virus, bacteria, or fungus. in 1992, as the links between infl ammation and sepsis were becoming increasingly clear, a consensus conference on sepsis defi nitions introduced the term sirs (systemic infl ammatory response syndrome) in an attempt to clarify and simplify the defi nitions of sepsis [1] . a patient was classifi ed as having sirs if he/she had at least two of four parameters (temperature >38 or <36° c; heart rate >90 beats/min; respiratory rate >20 breaths per minute or pco 2 < 32 mmhg; white blood cell count >12 or <4 × 10 9 /l). sepsis was defi ned as sirs plus infection. however, it soon became apparent that nearly all intensive care unit (icu) patients meet the sirs criteria at some point during their icu stay [2, 3] , making this approach too sensitive to be useful in diagnosing sepsis [4] . almost 10 years later, a second consensus conference on sepsis defi nitions was convened, sponsored by the society of critical care medicine (sccm), the european society of intensive care medicine (esicm), the american college of chest physicians (accp), the american thoracic society (ats), and the surgical infection societies (sis) [5] . the participants at this meeting agreed that the sirs concept was not helpful and should no longer be used per se, but that the sirs criteria be incorporated into a longer list of signs of sepsis that could be employed to support a diagnosis of sepsis. this list includes biologic signs of infl ammation (e.g., increased serum concentrations of c-reactive protein [crp] or procalcitonin), hemodynamic parameters (e.g., increased cardiac output, low systemic vascular resistance [svr], low oxygen extraction ratio), signs of altered tissue perfusion (e.g., altered skin perfusion, reduced urine output), and signs of organ dysfunction (e.g., increased urea and creatinine, low platelet count or other coagulation abnormalities, 1 hyperbilirubinemia). the participants also suggested that as the defi nitions did not allow for precise characterization and staging of patients with sepsis, a clinically useful staging system that could stratify patients by both their baseline risk of an adverse outcome and their potential to respond to therapy was needed. building on a system that had emerged at the fifth toronto sepsis roundtable held in toronto, canada, in 2000 [6] , the sepsis defi nitions conference participants, therefore, proposed the piro system [5] , which can classify patients on the basis of their predisposing conditions, the nature and extent of the infection, the nature and magnitude of the host response, and the degree of concomitant organ dysfunction. disease stratifi cations systems are widely used in clinical medicine, but perhaps the most familiar and frequently employed is the tnm system, which was developed by pierre denoix in the 1940s [7] , and is universally recognized as a standard for classifying patients with cancer. the tnm system classifi es malignant tumors based on descriptors of the extent of the primary tumor (t), on the presence, absence, and extent of metastases to regional lymph nodes (n), and on the presence or absence of distant metastases (m) ( table 1) . each patient with a tumor will, therefore, receive a specifi c classifi cation, e.g., t1, n0, m0, for that tumor. tnm classifi cations are then grouped into stages, usually from i to iv, which provide valuable prognostic information. importantly, staging systems in cancer stratify patients not only according to prognosis, but also according to the probability that they will respond to a particular therapy. sepsis is in many ways very similar to cancer. both disease processes are common, with high mortality rates. both are the result of a complex pathophysiological process involving cellular dysregulation. both can develop in (almost) any organ, and both frequently require surgical and medical therapies. treatments for both are expensive and often involve several pharmacological agents. finally, when treatment is successful, it is associated with slow step-by-step improvement. size and/or extent of the primary tumor regional lymph nodes (n) nx regional lymph nodes not evaluated n0 no regional lymph node involvement n1, 2, 3 number and extent of regional lymph node involvement these similarities between sepsis and cancer led to the suggestion that a disease stratifi cation system, similar to the tnm system for cancer, could be developed for sepsis [5] . the piro system for the grading of sepsis uses clinical and laboratory parameters to aid diagnosis and patient classifi cation, with each element being divided according to the degree of involvement (e.g., infection can be classifi ed as localized, extended, or generalized; immune response can be classifi ed as limited, extensive, or excessive; organ dysfunction can be classifi ed as mild, moderate, severe). as with the tnm system, it has been proposed that points could be allocated such that a patient with sepsis could, for example, be staged as p 1 i 2 r 1 o 0 [6] , depending on the features present for each of the four piro components. all aspects of the four components of the piro system impact on outcome and can infl uence therapeutic choices. as the tnm system is divided into clinical (ctcncm) and pathological (ptpnpm) classifi cations, so each component of piro can be considered to have potentially relevant clinical and laboratory variables ( table 2 ). predisposition can include multiple factors such as age, sex, presence of certain premorbid diseases, prolonged immunosuppressant or antimicrobial medication, even cultural and religious beliefs [8] . all these factors individually and collectively can impact on outcome, modifying both the disease process and the approach to therapy. recent advances in genetic techniques have enabled several factors associated with an increased risk of infection and of mortality from sepsis to be identifi ed. single nucleotide polymorphisms, microsatellites, insertion and deletion polymorphisms are all forms of genetic variation that can characterize an individual's risk for sepsis, organ dysfunction, or death [9] . most genetic traits associated with severe infection are associated with defects in innate immune responses. for example, a polymorphism of the tumor necrosis factor (tnf)-α gene, the tnf-2 allele, is associated with increased serum levels of tnf and a greater risk of mortality from septic shock [10] . a polymorphism within intron 2 of the interleukin-1 receptor antagonist (il-1ra) gene (il-1rn * 2) has been associated with reduced il-1ra production and increased mortality rates [11] . recently, polymorphisms in the toll-like receptor 1 gene were reported to be associated with increased susceptibility to organ dysfunction, death, and gram-positive infection in sepsis [12] . sex differences are another area of interest with several studies reporting that women are less likely to develop sepsis than men [13, 14] . however, women who do develop sepsis, particularly older women, may have worse outcomes than men [15, 16] . studies have also suggested racial differences in susceptibility to and outcomes from sepsis [17] , and older patients are known to be at an increased risk of developing sepsis and succumbing to it [18] . certain chronic diseases, such as cirrhosis, diabetes, and chronic obstructive pulmonary disease (copd), as well as chronic use of immunosuppressant medication may also predispose to sepsis and a worse outcome. moreover, each factor may have a different impact on the other three piro components [5] . for example, chronic immunosuppression may increase a person's risk of infection, but may decrease the magnitude of that person's infl ammatory response. undoubtedly these are complex relationships with multiple confounding factors and further research is needed to clearly defi ne which factors should be taken into account when considering the impact of predisposition on prognosis, to determine which carry most weight, and to identify how knowledge of increased risks can be translated into improved clinical outcomes. advances in genetics technology now enable investigators to create glass slides (chips) with minute quantities of short, gene-specifi c nucleotides. these gene-specifi c probe nucleotides, ideally one for each gene in the genome, are arrayed onto the chip surface to produce a dna microarray. these can be used to generate an expression profi le, the transcriptome, for the cell or tissue of interest. genomics, and the broader fi eld of proteomics, is likely to be increasingly used in routine patient management in hospitals of the future and will facilitate the task of assessing predisposition. four key aspects related to the underlying infection can infl uence management and prognosis in patients with sepsis: source, degree, hospital-acquired versus community-acquired, and microorganism [19] . in terms of source, for example, infections of the urinary tract are usually less severe than intra-abdominal or pulmonary infections. in the protein c worldwide evaluation in severe sepsis (prowess) trial [20] , patients with urinary tract infections as a source of severe sepsis had a 28-day all-cause mortality of 21% compared with patients with a pulmonary source of sepsis who had a mortality rate of 34% (p < .01). the size of the inoculum, virulence, and sensitivity of the infecting organisms are also important in determining outcomes. in the sepsis occurrence in acutely ill patients (soap) study, infection with pseudomonas spp. was independently associated with increased icu mortality (or: 1.62 [95% ci 1.09-2.42], p = 0.017) [16] . in a multicenter study from china, gram-positive bacterial infection and invasive fungal infections were risk factors for hospital mortality [21] . however, classifying the relative importance of infections on outcome can be diffi cult. cohen et al. [22] recently generated specifi c risk codes for the six most common infections: bacteremia, meningitis, pneumonia, skin and soft tissue infections, peritonitis, and urinary tract infections. for each infection site and organism, a two-digit code was generated according to the mortality rate associated with that infection (from 1: ≤5% to 4: >30%), and the level of evidence available to support the mortality risk (level a representing evidence from more than fi ve studies with greater than 100 patients, through to level e where there was insuffi cient evidence from case reports). this grading system for site and severity of infection (gsssi) needs to be validated, but could be a useful means of characterizing the risks associated with infections caused by various organisms in different sites. the timing of the onset of infection may also infl uence outcomes. one study showed that patients who developed septic shock within 24 h of icu admission were more severely ill, but had better outcomes, than patients who became hypotensive later during their icu stay [23] . sepsis is defi ned as the host response to infection, yet that host response has proved difficult to characterize [24] . various approaches have been proposed, including the presence of characteristic signs and symptoms or the degree of elevation of biological markers, such as procalcitonin or c-reactive protein, but as yet, none of the suggested markers is specifi c for sepsis. importantly, the initial theory that sepsis was simply an uncontrolled infl ammatory response and could be treated by blocking or removing any or several of the proinfl ammatory cytokines has been replaced by the realization that the infl ammatory response is a normal and necessary response to infection, and interrupting that response at any point may do more harm than good. indeed, the early hyperinfl ammatory phase of sepsis is soon replaced by a hypoinfl ammatory state. the host response to infection thus varies between patients and with time in the same patient [25] . this differentiation is important for therapeutic decisions, as antiinfl ammatory therapies may be harmful if given to a patient who is already in the hypoinfl ammatory phase; such a patient may benefi t rather from a proinfl ammatory therapy to boost their immune system. as with genomics, technological advances now enable multiple markers to be assessed simultaneously from small blood samples. this approach could provide clinicians with an immune profi le for individual patients. again, considerable research is needed to indentify which markers should be included on such microarrays. furthermore, the optimal set of biologic markers for 1 any patient may depend on the therapy being proposed [5] . for example, an indicator of dysregulation of the coagulation system might be more valuable when deciding whether or not to give drotrecogin alfa (activated), whereas a marker of adrenal dysfunction might be more useful for determining whether to give hydrocortisone. organ dysfunction in severe sepsis is not a simple "present" or "absent" variable, but presents a continuous spectrum of varying severity in different organs over time [26] . the degree of organ involvement can be assessed with various scoring systems, such as the sequential organ failure assessment (sofa) [27] . this system uses parameters that are routinely available in all icus to assess the degree of dysfunction for six organ systems: respiratory, cardiovascular, renal, coagulation, neurologic, and hepatic, with a scale of 0 (no dysfunction) to 4 for each organ. importantly, organ dysfunction can be recorded for each organ separately or a composite score can be calculated. thus with repeated scores, a dynamic picture of the effects of sepsis on individual or global organ dysfunction can be developed. sequential assessment of the sofa score during the fi rst few days of icu admission has been shown to be a good indicator of prognosis, with an increase in sofa score during the fi rst 48 h in the icu predicting a mortality rate of at least 50% [28] . levy et al. reported that early improvement in cardiovascular, renal, or respiratory function from baseline to day 1 was signifi cantly related to survival [29] . continued improvement in cardiovascular function before the start of day 2 and start of day 3 was associated with further improvement in survival for patients who improved compared with those who worsened. in the future, organ dysfunction scores may be replaced by or combined with more direct assessment of cellular stress and injury, for example, measures of mitochondrial dysfunction, apoptosis, or cytopathic hypoxia. the piro concept at its simplest provides a means of putting some order to the various aspects of sepsis. further work is needed to determine exactly which factors should be included in each of the four components and whether or how they should be measured and weighted to achieve a quantitative measure by which heterogeneous groups of septic patients could be characterized and categorized. once validated, it is possible that patients could receive a piro grade or stage, e.g., p 3 i 2 r 1 o 2 , which would help direct treatment and indicate prognosis. in addition to characterizing individual patients, such grades would facilitate comparison of patient populations for clinical trial purposes and help focus clinical research. several groups have already attempted to apply the piro system clinically and the results of these studies will be discussed in more detail in later chapters. moreno et al. used the saps iii database to assess whether the piro system could be useful for predicting mortality in patients with sepsis [30] . for each of the four piro components, multivariate analysis was used to select variables signifi cantly associated with hospital mortality, which were then weighted and allocated points. the authors felt it was not possible to separate host response from the resulting organ dysfunction, so they combined these two components. for predisposition, the fi nal variables were age, location of patient prior to icu admission, length of stay before icu admission, certain comorbidities (cancer, cirrhosis, acquired immunodefi ciency syndrome [aids]), and cardiac arrest as the reason for icu admission; for infection, the variables were nosocomial infection, respiratory infection, and infections by candida species or other fungi; for response/organ dysfunction, the variables were renal or coagulation dysfunction, and failure of the cardiovascular, renal, respiratory, coagulation, or central nervous systems. the authors suggested that, although further prospective validation is needed, the proposed saps iii piro system could be used to stratify patients at or shortly after icu admission to enable better selection of management according to the risk of death [30] . in a prospective, observational study, lisboa et al. [31] applied the piro concept to patients with ventilator-associated pneumonia (vap), again using multivariate logistic regression to identify variables independently associated with icu mortality for inclusion in the piro model. in this study in vap patients, the variables for predisposition were comorbidities (copd, immunocompromise, heart failure, cirrhosis, chronic renal failure); for infection, the variable was bacteremia; for response, the variable was systolic blood pressure <90 mmhg; and for organ dysfunction, the variable was acute respiratory distress syndrome (ards). a four-point score was thus developed, with one point for each component. mortality increased with increasing score: a score of 0 was associated with a mortality rate of 9.8%, increasing to 93.3% for patients with a score of 4. these authors suggested that the vap-piro score could thus be a useful practical tool to predict disease severity in patients with vap. the two studies discussed briefl y above are just two clinical examples of how the piro system could be adopted for use clinically. mortality remains high in patients with severe sepsis (around 40%) and septic shock (around 60%) and is closely associated with the degree of multiple organ failure. results from studies of proposed new interventions in severe sepsis have largely been disappointing with few demonstrating any positive effect on outcomes. one of the possible reasons for the multiple "failed" trials is that the groups of patients studied have been too heterogeneous and that global results have masked any potential benefi t in specifi c subgroups of patients [32] . better targeting of proposed interventions by better characterization of septic patients with the piro system may lead to better outcomes. improved classifi cation of septic patients using the piro system may, thus, facilitate the development and evaluation of clinical trials of sepsis therapies and will also encourage further study into the pathophysiology and epidemiology of sepsis. importantly, just as the tnm system is adjusted to specifi c cancers [33] , so the piro system will need to be adapted to fi t specifi c patient 1 groups, local practice, purpose (e.g., clinical trial inclusion, prognostication, patient management), or proposed therapies. for example, if the planned intervention is an anticoagulant then evidence of coagulopathy is likely to be more relevant than presence of respiratory failure, while if considering hemodialysis, the presence and degree of renal failure are likely to be most pertinent [24] . however, despite general acceptance of the piro concept and belief that it may contribute to improving outcomes in patients with sepsis, many questions remain unanswered. for example, in patients with cancer, correct staging is critical because treatment is directly related to disease stage. thus, incorrect staging can lead to improper treatment and to reduced patient outcomes. whether the same would hold true for patients with sepsis is unknown. clearly, considerable work remains to be done in testing and validating the piro system, but it represents an important step toward more successful management of the patient with severe sepsis. defi nitions of sepsis and multiple organ failure and guidelines for the use of innovative therapies in sepsis systemic infl ammatory response syndrome, sepsis, severe sepsis and septic shock: incidence, morbidities and outcomes in surgical icu patients the italian sepsis study: preliminary results on the incidence and evolution of sirs, sepsis, severe sepsis and septic shock dear sirs, i'm sorry to say that i don't like you sccm/esicm/accp/ats/sis international sepsis defi nitions conference measures, markers, and mediators: toward a staging system for clinical sepsis. a report of the fifth toronto sepsis roundtable enquete permanent dans les centres anticancereaux the piro concept: p is for predisposition injury research in the genomic era association of tnf2, a tnfα promoter polymorphism, with plasma tnfα levels and mortality in septic shock interleukin-1 receptor antagonist gene polymorphism and mortality in patients with severe sepsis toll-like receptor 1 polymorphisms affect innate immune responses and outcomes in sepsis incidence and mortality of severe sepsis in surgical intensive care patients: the infl uence of patient gender on disease process and outcome the epidemiology of sepsis in the united states from 1979 through 2000 effect of patient sex on intensive care unit survival sepsis in european intensive care units: results of the soap study racial variation in the incidence, care, and outcomes of severe sepsis: analysis of population, patient, and hospital characteristics the effect of age on the development and outcome of adult sepsis the piro concept: i is for infection effi cacy and safety of recombinant human activated protein c for severe sepsis epidemiology of severe sepsis in critically ill surgical patients in ten university hospitals in china new method of classifying infections in critically ill patients septic shock of early or late onset: does it matter? the piro concept: r is for response monocyte deactivation -rationale for a new therapeutic strategy in sepsis the piro concept: o is for organ dysfunction use of the sofa score to assess the incidence of organ dysfunction/failure in intensive care units: results of a multicentric, prospective study serial evaluation of the sofa score to predict outcome early changes in organ function predict eventual survival in severe sepsis sepsis mortality prediction based on predisposition, infection and response the ventilator-associated pneumonia piro score: a tool for predicting icu mortality and health-care resources use in ventilator-associated pneumonia clinical trials of immunomodulatory therapies in severe sepsis and septic shock tnm classifi cation of malignant tumours key: cord-347833-b3yrxkt0 authors: ahlström, björn; larsson, ing-marie; strandberg, gunnar; lipcsey, miklos title: a nationwide study of the long-term prevalence of dementia and its risk factors in the swedish intensive care cohort date: 2020-09-04 journal: crit care doi: 10.1186/s13054-020-03203-y sha: doc_id: 347833 cord_uid: b3yrxkt0 background: developing dementia is feared by many for its detrimental effects on cognition and independence. experimental and clinical evidence suggests that sepsis is a risk factor for the later development of dementia. we aimed to investigate whether intensive care-treated sepsis is an independent risk factor for a later diagnosis of dementia in a large cohort of intensive care unit (icu) patients. methods: we identified adult patients admitted to an icu in 2005 to 2015 and who survived without a dementia diagnosis 1 year after intensive care admission using the swedish intensive care registry, collecting data from all swedish general icus. comorbidity, the diagnosis of dementia and mortality, was retrieved from the swedish national patient registry, the swedish dementia registry, and the cause of death registry. sepsis during intensive care served as a covariate in an extended cox model together with age, sex, and variables describing comorbidities and acute disease severity. results: one year after icu admission 210,334 patients were alive and without a diagnosis of dementia; of these, 16,115 (7.7%) had a diagnosis of sepsis during intensive care. the median age of the cohort was 61 years (interquartile range, iqr 43–72). the patients were followed for up to 11 years (median 3.9 years, iqr 1.7–6.6). during the follow-up, 6312 (3%) patients were diagnosed with dementia. dementia was more common in individuals diagnosed with sepsis during their icu stay (log-rank p < 0.001), however diagnosis of sepsis during critical care was not an independent risk factor for a later dementia diagnosis in an extended cox model: hazard ratio (hr) 1.01 (95% confidence interval 0.91–1.11, p = 0.873). renal replacement therapy and ventilator therapy during the icu stay were protective. high age was a strong risk factor for later dementia, as was increasing severity of acute illness, although to a lesser extent. however, the severity of comorbidities and the length of icu and hospital stay were not independent risk factors in the model. conclusion: although dementia is more common among patients treated with sepsis in the icu, sepsis was not an independent risk factor for later dementia in the swedish national critical care cohort. trial registration: this study was registered a priori with the australian and new zeeland clinical trials registry (registration no. actrn12618000533291). dementia is a common and often detrimental group of diseases with a sharply increasing incidence in the elderly [1, 2] . in general, the disease is characterized by memory loss, disturbances in language, altered perception, and other psychological and psychiatric changes. together, these symptoms cause impaired daily functioning [3] and may severely affect health-related quality of life [4] . in hospitalized patients, and especially in the intensive care population, sepsis is a common syndrome [5] defined by a dysregulated host response to an infection [6] . the systemic inflammation in sepsis has been suggested to have a long-term negative impact on the brain [7] . both short-and long-term effects of experimental sepsis on brain cells and behavior have been described in rodents [8] [9] [10] . septic encephalopathy and persisting cognitive disturbances have also been reported in human studies [11] [12] [13] . moreover, in patients with an assessment of cognitive function before and after hospitalization, a decline in cognitive function was more pronounced after hospitalization for severe sepsis than after admission for other reasons [14] . sepsis diagnosis has also been reported to be more common in the history of patients with dementia than age-and sex-matched controls from the health care system [15] . finally, sepsis is an independent risk factor for dementia in observational studies [16, 17] . based on these data, we hypothesized that dementia would develop more commonly in patients admitted for, or developing, sepsis in intensive care compared with other patient groups. however, the prevalence of dementia in the population is relatively low [1] . additionally, dementia is usually a slowly developing syndrome with a long subclinical period before diagnosis [18] . dementia also increases the risk of sepsis [19] , and several comorbidities are risk factors for both dementia and sepsis. we therefore set out to investigate our hypothesis in a large nationwide database with an extended follow-up and accounting for comorbidities. our primary endpoint was the hazard ratio (hr) of sepsis for a diagnosis of dementia adjusting for several potential risk factors. we also investigated the impact of these risk factors and crude dementia incidence in this cohort. this study was approved by the regional ethics committee of uppsala (approval no. 2016/421). since this is a registry-based study, informed consent was waived. the protocol of the study was registered a priori with the australian and new zeeland clinical trials registry (registration no. actrn12618000533291). reporting strictly follows the strobe statement [20] . the swedish intensive care registry (sir) is a national registry to which all general icus in sweden are reporting data on all admissions [21, 22] . the national patient register (npr) includes data from all in-patient hospital visits in sweden, and the cause of death registry includes deaths of all swedish residents, including all deaths abroad [23] . both registries are run by the swedish national board of health and welfare. the swedish dementia registry (svedem) is a national quality registry started in 2007, with primary care and specialized memory units reporting cases in sweden. the svedem had an estimated coverage ratio of 27-35% of dementia incidence in sweden in 2017, partly overlapping with the npr [24, 25] . all adult patients aged > 17 years who had at least one episode of intensive care in the sir in 2005 to 2016 were included. we excluded patients with a diagnosis of dementia at icu admission and patients who died or acquired a diagnosis of dementia during the first year after icu admission. patients in the sepsis diagnosis code cohort (henceforth referred to as the sepsis cohort) were identified by sepsis diagnosis codes registered in the sir. the diagnosis of severe sepsis and septic shock, represented as icd-10 a41.9 (2005-2010), r65.1 or r57.9 (2011-2016), has to be confirmed or negated when registering a patient in the sir. those without sepsis diagnosis codes in sir were included in the no sepsis diagnosis code cohort (hereafter referred to as the no sepsis cohort). during the entire study period, the sir defined the diagnosis of sepsis, severe sepsis, and septic shock according to the second international sepsis definitions conference of 2001 [26] . from the sir, we extracted data on the severity of illness at admission, invasive ventilator support, renal replacement therapy (rrt), icu length of stay (icu-los), and diagnoses relevant to the icu episode. for patients with repeated admissions, we used the first icu episode with a sepsis diagnosis code or, for patients without a sepsis diagnosis code, the first icu episode in the sir. we treated overlapping icu episodes as one episode. the severity of illness was initially reported as acute physiology, age, chronic health evaluation ii (apache ii) in the sir [27] and, during 2010, gradually substituted with the simplified acute physiology score 3 (saps3) [28] . death date was extracted from the cause of death registry. from the npr, we derived icd-10 [29] diagnosis codes for all inpatient care episodes from 5 years before the icu care episode to december 31, 2016. the revised charlson comorbidity index (cci) [30] was calculated using diagnoses from all health care contacts preceding or coinciding with the first icu episode. we defined dementia using the following icd-10 codes according to the cci [31] : f00x-f03x, f051, and g30x-31x. because the inpatient care diagnoses database only includes diagnoses from inpatient care, we incorporated data from the svedem to track down patients with dementia not admitted to the hospital. the date of dementia diagnosis was the first occurrence of the condition in the npr or the svedem. the swedish dementia centre, commissioned by the swedish national board of health and welfare, provides recommendations on the diagnostic process in suspected dementia. in both primary and specialist care, the recommendation is to use the diagnostic criteria of the icd-10, especially identifying the importance of symptom stability (6 months) and the exclusion of co-existing confusion [32] . for descriptive statistics, we used counts with percentages, means with standard deviations and medians and interquartile ranges (iqr) as appropriate. we assessed the crude incidence of dementia with kaplan-meier curves using the log-rank test. for the primary analysis, hrs for the risk of dementia were calculated in a cox regression model with mortality censored. the following covariates were chosen from available variables through directed acyclic graph analysis and a literature review: sepsis; age [1] and sex, all of which have been previously described as independent risk factors for dementia [33] ; cci; saps3 box 2+3; hospital length of stay (h-los); icu-los; invasive ventilator therapy; and rrt. missing data were substituted by redundancy between data sources where possible. missing saps3 box 2+3 was substituted by multiple imputations into five datasets using the multivariate imputation by chained equations (mice) package in r [34] . the results from the analyses on the imputation datasets were pooled using the harrel miscellaneous (hmisc) package in r. the proportional hazards assumption was deemed fulfilled after visual inspection of plots of scaled schoenfeld residuals against time and the covariates treated as continuous were evaluated for linearity by plots of martingale residuals against the covariate. because of nonlinearity for all continuous covariates, we used cubic splines in the cox model [35] . seven sensitivity analyses were performed according to the description in additional file 1. we defined statistical significance as p < 0.05. hrs for which cubic splines were applied were calculated between the 25th and 75th percentiles. data management and descriptive statistics were performed in spss for windows version 24 (microsoft inc., il, usa). for inference tests (i.e., regression analyses) and multiple imputations, we used r software version 3.5.3 (the r foundation for statistical computing, vienna, austria; https://www.r-project.org). of 315,155 patients, 210,334 (67%) were still alive and without dementia 1 year after icu admission (fig. 1) . of those 210,334 patients, 16,115 (8%) had a sepsis diagnosis code in icu care. the patients were followed for a median of 3.9 years (iqr 1.7-6.6). saps3 data were completely missing in 23.8% of the patients in the sepsis cohort and 45.2% in the no sepsis cohort. of patients with missing saps3, 46% had a registration of an apac he ii score in the sepsis cohort and 23% in the no sepsis cohort. missing saps3 data were imputed. of all patients admitted to the icu in 2005-2016, 8495 (4.0%) emigrated at least once from sweden, and of these, 5358 (63.1%) had at least one listing in the npr or the sve-dem > 1 year after icu admission. patients in the sepsis cohort were older, had higher saps3, and had more comorbidities, expressed as a higher cci score. they also had longer icu-los and h-los and were more commonly treated with invasive ventilation and rrt (table 1) . during follow-up, the sepsis cohort had a mean of 3.9 episodes of inpatient care, whereas 32% of these patients had no such episode. the no sepsis cohort had a mean of 3.4 episodes of inpatient care, and 38% of these patients were without any such episode. the 6312 patients ultimately developing dementia were older and predominantly female and had higher saps3 and longer icu-los and h-los than those without dementia ( table 2 ). in addition, those patients who developed dementia were less frequently treated with rrt and invasive ventilator therapy. finally, patients with dementia had a higher rate of acute surgical admissions, but a lower rate of planned surgical admissions. the size of these differences was generally small, however. dementia prevalence and 1-year mortality increased with age in patients admitted to the icu during the study and alive on the last day of follow-up (december 31, 2016) (fig. 2) . in the unadjusted analysis, dementia was more common in individuals diagnosed with sepsis during their icu stay (log-rank p < 0.001) as depicted in the kaplan-meier survival curve (fig. 3) . however, after adjusting for age, sex, cci score, saps3 box 2+3, h-los, icu-los, invasive ventilator therapy, and rrt, sepsis was no longer an independent risk factor for dementia (hr 1.01, 95% ci 0.91-1.11) (fig. 4) in any of the pre-specified sensitivity analyses, sepsis was not an independent risk factor for dementia (additional file 2). in our nationwide swedish cohort of 210,334 patients alive without dementia 1 year after icu admission, sepsis was found to be a crude risk factor for a later diagnosis of dementia. however, after adjusting for baseline characteristics of the patients in our cohort, sepsis was no longer an independent risk factor for dementia. this finding was consistent in all performed sensitivity analyses. in a previous study [36] , 516 patients having survived a sepsis episode were compared with 4517 patients having survived a hospitalization without sepsis. all included individuals underwent at least one prospective cognitive testing. when followed for up to 1 year, the sepsis patients performed worse on repeated cognitive testing. however, it is not clear whether this condition evolves into fulfilling the diagnostic criteria of dementia, and the authors did not control for age, comorbidities, or the degree of acute illness. in the present study, we controlled for several factors expressing both comorbidity and the degree of acute illness besides age. guerra et al. performed two studies on a 2005 medicare cohort. in their first study [16] , the findings seem to confirm the hypothesis that the higher rate of observed cognitive dysfunction after sepsis is evolving into a higher rate of clinically diagnosed dementia in patients treated in the icu with sepsis than in those treated in the icu without sepsis during their hospitalization. however, they were only able to adjust for comorbidities diagnosed in the year preceding the index hospitalization, thereby risking underestimating the comorbidities. in addition, the authors run the risk of overlooking the presence of dementia diagnoses in earlier years that was not registered in the year preceding hospital admission. furthermore, because dementia is a syndrome of a slowly evolving disease [18] , dementia diagnosed early after the index hospitalization might be an example of a clinically overt disease coming to the attention of the medical system in the convalescence period after hospitalization rather than a consequence of the sepsis episode or acute illness per se. dementia may also be a risk factor for sepsis [19] . we sought to lessen the effect of both over-diagnosis due to hospitalization and causality problems between sepsis and dementia by excluding dementia diagnoses registered during the first year after icu admission. in the second study by guerra et al. [17] , patients admitted to the icu with sepsis were compared with non-hospitalized patients matched on age group, sex, and race. sepsis was a significant risk factor for dementia. however, in a model adjusting for comorbid diagnoses associated with dementia during the index hospitalization, the effect of sepsis decreased compared with using the same comorbidities diagnosed before the index hospitalization. moreover, when using comorbidities diagnosed during the index quarter in the model, the effect of sepsis on the risk of dementia disappeared in line with our findings. in a casecontrol study [15] , the odds ratio for having had a sepsis diagnosis in 5955 patients with a dementia diagnosis was higher than in age-and sex-matched controls without dementia. in their design, the authors did not account for the amount of time elapsed from sepsis to the diagnosis of dementia, nor did they adjust for comorbidities diagnosed during the index hospitalization. despite that the patients in the sepsis and the no sepsis cohorts are from the same icu cohort, they were not comparable, i.e., sepsis cohort patients had more chronic illnesses, were older, and had a higher degree of acute illness. hence, it was essential to adjust to these specific factors. our study used the revised cci as a composite fig. 2 dementia prevalence by age interval in patients of the icu cohort alive at 31st of december 2016 and mortality 1 year from icu admission. line thickness represents the number of patients at risk. icu, intensive care unit; n_mort, number at risk of mortality; n_dem, number at risk of dementia measure of the comorbid burden of each patient. we also adjusted for the severity of acute illness, as we presumed that it might mediate the effect of sepsis on dementia development. saps3 box 2+3, i.e., acute illness severity, was an independent risk factor for dementia. however, in a sensitivity analysis in which saps3 box 2+3 was excluded from the model, sepsis was not a significant risk factor for dementia. this finding implies that saps3 box 2+3 does not modulate the effect of sepsis in the model. as expected, age was a strong risk factor for developing dementia during follow-up in our cohort of icufor variables treated as continuous: age, icu-los, hospital los, cci score, and saps3 box 2+3, the hr is of the difference between the 25th and 75th percentiles. hr, hazard ratio; ci, confidence interval; icu, intensive care unit; los, length of stay; cci, revised charlson comorbidity index; saps3, simplified acute physiology score 3; rrt, renal replacement therapy treated patients. intriguingly, in the oldest age category (> 90 years), we observed a lower prevalence of dementia compared with another study on a swedish cohort [33] . this lower prevalence may be related, in part, to the very high mortality rates in this elderly patient group that have been treated in the icu. surprisingly, the cci score was not an independent risk factor for dementia in the model. we speculate that this finding is due to the small difference between the 25th and 75th percentiles of the cci score. a wider range of cci might have yielded another result. for the variables in which cubic splines were applied, the hr was calculated for the difference between values at the 25th and 75th percentiles to reduce the risk of false conclusions from the hrs of these splined variables. still, despite this adjustment, results for the splined variables need to be interpreted cautiously. however, their validity as covariates of sepsisthe primary endpoint of the study-in the model is higher after cubic spline application than if we would have categorized these nonlinear variables [37] . a major strength of this study is that our sample contains virtually all intensive care patients in sweden over 11 years (2005 to 2015) , which represents the different socioeconomic groups of a high-income country. another strength is the possibility to follow the patients for an extended period before icu admission. such an approach allows for an accurate assessment of comorbidities in general and pre icu dementia in particular. we believe that the long mean follow-up of > 4 years from 1year post-icu admission is also an important asset of the study in that most types of dementia are gradually developing diseases [38] . our study also has by far the largest cohort of icu survivors to study the association between sepsis and dementia. finally, we were able to control for the severity of acute illness. our study has some limitations of note. the major limitation is that we define dementia as a dementia diagnosis at an inpatient visit at a hospital or a dementia diagnosis in the svedem. we thus expect to miss a proportion of patients despite using data from several registries to detect dementia in both hospital-admitted patients and outpatients. however, we have no reason to believe that dementia would be unreported to a larger extent in patients treated for sepsis in the icu as the sepsis cohort patients had more hospital inpatient visits than the no sepsis cohort patients during follow-up. this observation is possibly due to the higher comorbidity burden and higher age in the sepsis cohort patients. moreover, we choose not to include patients that did not live up to 1 year after icu admission, resulting in excluding almost one third of the patients in the analysis. however, we found in a sensitivity analysis that our results did not change after including those patients not surviving up to 1 year after icu admission. another potential shortcoming of the data is the possibility the patients die before they develop dementia, implying that the findings of this study could potentially need to be verified in cohorts with much lower mortality. in a sensitivity analysis, we included only patients with saps3 in the lowest quartile expecting a lower mortality and thus a smaller risk of dying acting as a competing event, but sepsis was not an independent risk factor in this analysis. finally, the present study is limited because 4% of the patients emigrated from sweden at some point during the study. however, 63% of these patients had at least one listing in our data sources during the follow-up, which also were the case in the complete cohort. thus, we chose not to exclude emigrated patients from the main analysis; however, doing so in a sensitivity analysis did not affect our results. because the effect of sepsis on the risk of later dementia development has been shown to be minimal in previous studies and not present at all in our study, we recommend that further research on outcome after sepsis be directed in other directions. in conclusion, although dementia was more common in the whole nation swedish icu cohort for 2005-2015 treatment for sepsis in the icu, sepsis was not a risk factor for later dementia after adjustment for pre-specified, relevant, baseline variables. in our sample, acute illness severity altered the risk of dementia, which might account for a fraction of the apparent causality between sepsis and dementia in icu patients. supplementary information accompanies this paper at https://doi.org/10. 1186/s13054-020-03203-y. dementia in western europe: epidemiological evidence and implications for policy making the public's perceptions about cognitive health and alzheimer's disease among the u.s. population: a national review dementia: timely diagnosis and early intervention what do we know about quality of life in dementia? a review of the emerging evidence on the predictive and explanatory value of disease specific measures of health related quality of life in people with dementia epidemiology of severe sepsis the third international consensus definitions for sepsis and septic shock (sepsis-3) does infection-induced immune activation contribute to dementia? deterioration of spatial learning performances in lipopolysaccharide-treated mice sepsis causes neuroinflammation and concomitant decrease of cerebral metabolism the additive effect of aging on sepsis-induced cognitive impairment and neuroinflammation septic encephalopathy: relationship to serum and cerebrospinal fluid levels of adhesion molecules, lipid peroxides and s-100b protein cognitive impairment in the septic brain persistent cognitive impairment, hippocampal atrophy and eeg changes in sepsis survivors long-term cognitive impairment and functional disability among survivors of severe sepsis association between sepsis and dementia risk factors for dementia after critical illness in elderly medicare beneficiaries risk of a diagnosis of dementia for elderly medicare beneficiaries after intensive care alzheimer' s disease dementia increases the risks of acute organ dysfunction, severe sepsis and mortality in hospitalized older patients: a national population-based study strengthening the reporting of observational studies in epidemiology (strobe) long-term mortality and cause of death for patients treated in intensive care units due to poisoning the swedish board of health and welfare. the national patient register. 1-2 no significant difference in cognitive decline and mortality between parkinson's disease dementia and dementia with lewy bodies: naturalistic longitudinal data from the swedish dementia registry sccm/esicm/accp/ats/sis international sepsis definitions conference apache ii: a severity of disease classification system.pdf saps 3-from evaluation of the patient to evaluation of the intensive care unit. part 1: objectives, methods and cohort description the swedish board of health and welfare available from: www. socialstyrelsen.se accessed 3 updating and validating the charlson comorbidity index and score for risk adjustment in hospital discharge abstracts using data from 6 countries coding algorithms for defining comorbidities in icd-9-cm and icd-10 administrative data the swedish dementia centre twenty-year changes in dementia occurrence suggest decreasing incidence in central mice: multivariate imputation by chained equations in r flexible smoothing with b -splines and penalties spurious inferences about longterm outcomes: the case of severe sepsis and geriatric conditions regression modelling strategies. 2. springer international publishing swizerland measuring cognition and function in the preclinical stage of alzheimer's disease. alzheimer's dement preferred reporting items for systematic reviews and meta-analyses: the prisma statement publisher's note springer nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations henrik renlund, the statistician, is gratefully acknowledged for statistical support. authors' contributions ba, gs, iml, and ml conceived and designed the study; ba and ml acquired and analyzed the data; ba and ml drafted the manuscript; and ba, gs, iml, and ml finalized the manuscript. all authors approved the final manuscript as submitted. uppsala university hospital research fund and the centre for clinical research at region dalarna, sweden, funded this research. open access funding was provided by uppsala university. the data used in this study are available from the sir, the npr, and the svedem. however, privacy or ethical restrictions apply to the availability of these data, which were used under license for the current study. thus, these data are not publicly available. the data, however, are available from the authors upon reasonable request and with permission of the sir, the npr, and the svedem. this study was approved by the regional ethics committee of uppsala (approval no. 2016/421). because this is a registry-based study, informed consent was waived by the same ethics committee. not applicable. the authors declare that they have no competing interests. key: cord-103081-k7ev5qkn authors: janosevic, danielle; myslinski, jered; mccarthy, thomas; zollman, amy; syed, farooq; xuei, xiaoling; gao, hongyu; liu, yunlong; collins, kimberly s.; cheng, ying-hua; winfree, seth; el-achkar, tarek m.; maier, bernhard; ferreira, ricardo melo; eadon, michael t.; hato, takashi; dagher, pierre c. title: the orchestrated cellular and molecular responses of the kidney to endotoxin define the sepsis timeline date: 2020-05-30 journal: biorxiv doi: 10.1101/2020.05.27.118620 sha: doc_id: 103081 cord_uid: k7ev5qkn clinical sepsis is a highly dynamic state that progresses at variable rates and has life-threatening consequences. staging patients along the sepsis timeline requires a thorough knowledge of the evolution of cellular and molecular events at the tissue level. here, we investigated the kidney, an organ central to the pathophysiology of sepsis. single cell rna sequencing revealed the involvement of various cell populations in injury and repair to be temporally organized and highly orchestrated. we identified key changes in gene expression that altered cellular functions and can explain features of clinical sepsis. these changes converged towards a remarkable global cell-cell communication failure and organ shutdown at a well-defined point in the sepsis timeline. importantly, this time point was also a transition towards the emergence of recovery pathways. this rigorous spatial and temporal definition of murine sepsis will uncover precise biomarkers and targets that can help stage and treat human sepsis. acute kidney injury (aki) is a common complication of sepsis that doubles the mortality risk. in 30 addition to failed homeostasis, kidney injury can contribute to multi-organ dysfunction through 31 distant effects. indeed, the injured kidney is a significant mediator of inflammatory chemokines, 32 cytokines, and reactive oxygen species that can have both local as well as remote deleterious 33 effects 1-4 . therefore, understanding the complex pathophysiology of kidney injury is crucial for 34 the comprehensive treatment of sepsis and its complications. 35 we have recently shown that renal injury in sepsis progresses through multiple phases. these 36 include an early inflammatory burst followed by a broad antiviral response and culminating in 37 translation shutdown and organ failure 5 . in a non-lethal and reversible model of endotoxemia, 38 organ failure was followed by spontaneous recovery. the exact cellular and molecular 39 contributors to this multifaceted response remain unknown. indeed, the kidney is architecturally 40 a highly complex organ in which epithelial, endothelial, immune and stromal cells are at 41 constant interplay. therefore, we now examined the spatial and temporal progression of 42 endotoxin injury to the kidney using single cell rna sequencing (scrnaseq). our data revealed 43 that cell-cell communication failure is a major contributor to organ dysfunction in sepsis. 44 remarkably, this phase of communication failure was also a transition point where recovery 45 pathways were activated. we believe this spatially and temporally anchored approach to sepsis 46 pathophysiology is crucial for identifying potential biomarkers and therapeutic targets. we harvested a cumulative amount of 63,287 renal cells obtained at 0, 1, 4, 16, 27, 36 and 48 55 hours after endotoxin (lps) administration. the majority of renal epithelial, immune and 56 endothelial cell types were represented (fig. 1a) . note the absence of podocyte and mesangial 57 cells, which can be a limitation of single cell rnaseq renal dissociation procedures 6 . cluster 58 identities were assigned and grouped using known classical phenotypic markers (fig. 1b, 59 supplementary fig. 1a ) [7] [8] [9] [10] [11] . interestingly, the umap-based computational layout of epithelial 60 clusters recapitulated the normal tubular segmental order in the nephron. this indicates that 61 gene expression gradually changes among neighboring tubular segments along the nephron. 62 note that the expression of cluster-defining markers varied significantly during the injury and 63 recovery phases of sepsis ( fig. s1b; supplementary table 1 ). therefore, we also identified a 64 set of genes that are conserved across time for a given cell type (fig. s1c) . 65 in the integrated umap (fig. 1a) , we noted the presence of a proliferative cell cluster (cdk1 and 66 ki67 expression). by back mapping to time-specific unintegrated umaps, we determined that 67 these proliferating cells could be traced to specific cell types at various points along the sepsis 68 timeline (fig. 1c) . for example, within the first hour after lps, these proliferative indices were 69 expressed primarily in s1 cells. these cells are the site of lps uptake in the kidney as we have 70 previously shown 12-14 . at later time points, proliferative indices are seen in macrophages (4 71 hours) and s3 cells (36 hours) (fig. 1c) . these proliferative indices reflect cell cycle activity 72 which may be involved in injury, repair or recovery processes 15 . 73 we also noted the presence of a proximal tubular cluster expressing unique gene identifiers: 74 agt, rnf24, slc22a7 and slc22a13 (fig. 2a) . this is likely the proximal tubular s3-type 2 75 (s3t2) reported by others 16 . this cluster maintained a separate and distinct identity throughout 76 the sepsis timeline (fig. 1c) . because the location of s3t2 is currently unknown, we performed 77 in-situ spatial transcriptomics on septic mouse kidneys 17 . we then integrated our scrnaseq 78 with the in-situ rnaseq in order to map our scrnaseq clusters onto the tissue (supplementary 79 fig. 2a, s2b) . we found that the classical s3 cluster localizes to the cortex while s3t2 is in the 80 outer stripe of the outer medulla (fig. 2b, supplementary fig. 2b) . we confirmed the location 81 of s3t2 to the os-om with single molecular fish (supplementary fig. 2c) . the differential 82 gene expression between s3 and s3t2 is likely dictated by regional differences in the 83 microenvironments of the cortex and the outer stripe. 84 because angiotensinogen (agt) was strongly expressed in s3t2, we examined the expression 85 of other components of the renin-angiotensin system (ras). we first noted the absence of ace 86 expression in s1 tubular cells (fig. 2c, supplementary fig. 3) . in contrast, ace2 was strongly 87 expressed in s1, s3 and s3t2 cells. there is currently great interest in understanding the 88 biology of ace2 because of its role in sars-cov-2 cellular invasion. other essential 89 components of the sars-cov-2 entry mechanism include tmprss2 and slc6a19 18-22 . while 90 tmprss2 was expressed in all proximal tubular segments, slc6a19 was more strongly 91 expressed in s1 throughout the sepsis timeline. this may point to the s1 tubular segment as 92 one point of entry of sars-cov-2 into the kidney. 93 the immune cell profile in the septic kidney was time-dependent and showed a five-fold 96 increase in immune cells, primarily macrophages (fig. 3a, 3b) . we noted two distinct 97 macrophage clusters denoted as macrophage a and macrophage b (mϕ-a, mϕ-b). both of 98 these clusters expressed classical macrophage markers such as cd11b (itgam) (fig. 3c) . 99 accumulated macrophages were predominantly mϕ-a. we noted the absence of proliferation 101 markers (cdk1, ki67) in this cluster, raising the possibility that this may be an infiltrative 102 macrophage type (fig. 3d) . the mϕ-b cluster, located between mϕ-a and conventional 103 dendritic cells (cdc) expressed also cdc markers such as mhc-ii subunit genes (h2-ab1) and 104 cd11c (itgax) indicating that it is an intermediary macrophage type (fig. 3c) . this continuum 105 between macrophages and dendritic cells in the kidney has been reported 23-26 . interestingly, 106 mϕ-b cells expressed proliferation markers (cdk1, ki67) and thus, may be differentiating 107 towards a mϕ-a or cdc phenotype (fig. 3c) . pseudotime and velocity field analysis suggested 108 that at earlier time points (1 hour) mϕ-b was differentiating toward mϕ-a phenotype. at later 109 time points (36 hours) the velocity field suggested that mϕ-b was differentiating towards cdc 110 but pseudotime analysis was inconclusive (fig. 3e) . similarly, the mϕ-a cluster also showed two 111 subclusters on the rna velocity map (supplementary fig. 4a) . one of the subclusters showed 112 increased expression of alternatively activated macrophages (m2) markers such as arg1 113 (arginase 1) and mrc1 (cd206) 27 at later time points (36 hours, supplementary fig. 4b) . 114 therefore, rna velocity analysis may be a useful tool in distinguishing macrophage subtypes in 115 scrnaseq data. 116 in t-cells, while cd4 expression was minimal at all time points, the expression of cd8 was 117 robust and relatively preserved over time (fig. s4c) . we also noted an increase of a distinct 118 plasmacytoid dendritic cell cluster at one hour (pdc). these pdcs, along with natural killer (nk) 119 cells, are known to signal through the interferon-gamma pathway and stimulate cd8 expression 120 28, 29 . this supports the early antiviral response we have previously reported in this sepsis model defined with pseudotime analysis. we note that at any given time point, directional progression 128 of states along pseudotime correlated well with real time state changes (fig. 4a) . note that the 129 endothelium exhibited changes in states as early as 1 hour, while s1 showed changes at later 130 time points (4 hours). these sequential state changes may reflect the spatial and temporal 131 propagation of lps signaling in the kidney. as sepsis progressed, many cell types lost function-132 defining markers while acquiring novel ones. for example, s1 and s3 lost classical markers like 133 slc5a2 (sglt2) and aqp1 and expressed new genes involved in antigen presentation such as 134 h2-ab1 (mhc-ii) and cd74 (fig. 4b) . moreover, the highly distinct phenotypes that 135 differentiated s1 from s2/s3 at baseline merged into one phenotype for all three sub-segments 136 by 16 hours after lps (fig. 4c) . however, despite the apparent convergent phenotype at 16 137 hours, additional analytical approaches such as rna velocity revealed significant differences in 138 rna splicing kinetics between s1 and s3 segments at this time point. in addition, rna velocity 139 revealed the presence of two subclusters within the s3 segment at 16 hours (fig. 4d) . these 140 two velocity subclusters did not correlate with the two states seen in pseudotime analysis. this 141 indicates that multiple analytic approaches are needed to fully characterize cellular changes 142 along the sepsis timeline. 143 we next show gene expression profiles in select cell types along the sepsis timeline. in this 145 analysis, we included endothelial cells, pericyte/stromal cells, macrophages and s1 tubular 146 cells. within 1 hour of lps exposure, most cell types showed decreased expression of select 147 genes involved in ribosomal function, translation and mitochondrial processes such as eef2 and rpl genes (fig. 5a, supplementary fig. 5a ). this reduction peaked at 16 hours and recovered 149 by 27 hours. concomitantly, most cell types exhibited increased expression of several genes 150 involved in inflammatory and antiviral responses such as tnfsf9, cxcl1, ifit1, and irf7. however, 151 this increase was not synchronized among all cell populations. indeed, it occurred as early as 1 152 hour in endothelial cells, macrophages and pericyte/stromal cells, all acting as first responders. 153 in contrast, epithelial cells were late responders, with increases in inflammatory and antiviral 154 responses occurring between 4 and 16 hours. in fact, four hours after lps administration, 155 cluster-specific go terms were indistinguishable among the majority of cell types with 156 enrichment in terms related to defense, immune and bacterium responses (fig. 5b) . one noted 157 exception was the s3t2 cells (outer stripe s3) which did not enrich as robustly as other cell 158 types in these terms. it mostly maintained an expression profile related to ribosomes, translation 159 and drug transport throughout the sepsis timeline (supplementary fig. 6 ). other players of 160 interest in sepsis pathophysiology such as prostaglandin and coagulation factors are described 161 in supplementary figure 5b . 162 at the 48-hour time point, while s1 cells partially recovered to baseline, the macrophages 163 showed increased expression of genes involved in phagocytosis, cell motility and leukotrienes, 164 broadly representative of activated macrophages (e.g. csf1r, lst1, capzb, s100a4, cotl1, 165 alox5ap, fig. 5a) . intriguingly, at this late time point, the pericyte/stromal cells are enriched in 166 unique terms related to specific leukocyte and immune cell types such as lymphocyte-mediated 167 immunity, t cell mediated cytotoxicity and antigen processing and presentation. this suggests 168 that the pericyte may function as a transducer between epithelia and other immune cells. 169 therefore, we next examined comprehensively cell-cell communication along the sepsis 171 timeline. we show select examples of cell type-specific receptor ligand pairs. for example, we 172 found that s1 and endothelial cells communicate with the angpt1 (angiopoetin 1) and tek (tie2) ligand-receptor pair at baseline and throughout the sepsis timeline ( fig. 6a-b, supplementary 174 fig. 7a ). in contrast, c3 was strongly expressed in pericyte/stromal cells, while its receptor 175 c3ar1 localized to macrophage/dcs. this communication, present at baseline, did increase 176 along the sepsis timeline with additional players such as s1 participating in the cross talk 177 (supplementary fig. 7) . another strong communication was noted between endothelial cells 178 and macrophage/lymphocytes using the ccl2 and ccr2 receptor-ligand pair. the architectural 179 layout of these four cell types, with pericytes and endothelial cells residing between proximal 180 tubule and macrophage/dcs may explain these complex communication patterns 6 . such 181 communication patterns among these four cell types may also explain macrophage clustering 182 around s1 tubules at later time points in sepsis as we previously reported 13 . 183 when examined comprehensively, receptor-ligand signaling progressed from a broad pattern at 184 baseline into a more discrete and specialized one 4 hours after lps (fig. 6c, supplementary 185 fig. 7b-c) the murine sepsis timeline allows staging of human sepsis 201 finally, we asked whether our mouse sepsis timeline could be used to stratify human sepsis 202 aki. to this end, we selected the differentially expressed genes from all cells combined (pseudo 203 bulk) for each time point across the mouse sepsis timeline (supplementary table 4) . we then 204 examined the orthologues of these defining genes in human kidney biopsies of patients with 205 sepsis and aki. the clinical data associated with these human biopsies did not allow further 206 stratification or staging of the sepsis timeline (supplementary table 5 ). as shown in figure 7d , 207 our approach using the mouse data succeeded in partially stratifying the human biopsies into 208 early, mid and late sepsis-related aki. these findings suggest that underlying injury 209 mechanisms are conserved, and the mouse timeline may be valuable in staging and defining 210 biomarkers and therapeutics in human sepsis. 211 in this work, we provide comprehensive transcriptomic profiling of the kidney in a murine sepsis 213 model. to our knowledge, this is the first description of spatial and temporal transcriptomic 214 changes in the septic kidney that extend from early injury well into the recovery phase. our data 215 cover nearly all renal cell types and are time-anchored, thus providing a detailed and precise 216 view of the evolution of sepsis in the kidney at the cellular and molecular level. 217 using a combination of analytical approaches, we identified marked phenotypic changes in 218 multiple cell populations along the sepsis timeline. the proximal tubular s1 segment exhibited 219 significant alterations consisting of early loss of traditional function-defining markers (e.g., 220 sglt2). similar losses of function-defining markers along the nephron may explain the 221 profound derangement in solute and fluid homeostasis seen in sepsis. concomitantly, we 222 observed novel epithelial expression of immune-related genes such as those involved in antigen 223 presentation. this indicates a dramatic switch in epithelial function from transport and 224 homeostasis to immunity and defense. these phenotypic changes were reversible, thus 225 underscoring the remarkable resilience and plasticity of the renal epithelium. 226 in addition, our combined analytical tools clearly identified unique subclusters within each 227 epithelial cell population (e.g., cortical s3 and os s3). these subclusters likely represent novel 228 populations that may be in part influenced by the complex microenvironments in the kidney. it is 229 likely that such microenvironments define unique features in epithelial subpopulations such as 230 the expression of complete sars-cov-2 machinery in s1. 231 similarly, we also identified unique features in immune-cell populations. for example, the 232 combined use of rna velocity field and pseudotime analyses uncovered differences in 233 macrophage subtypes relating to rna kinetics and cell differentiation trajectories. of note is that 234 these subtypes only partially matched the traditional flow cytometry-based classification of 235 macrophages (e.g., m1/m2). therefore, the use of single-cell rna seq is a powerful approach 236 that will add to and complement our current understanding of the immune cell repertoire in the 237 additional approaches such as receptor-ligand crosstalk and gene regulatory network analyses 239 identified unique cell-and time-dependent players involved in sepsis pathophysiology. our work points to the urgent need for defining a more accurate and precise timeline for human 250 sepsis. such definition will guide the development of biomarkers and therapies that are cell and 251 time specific. we show evidence supporting the relevance of murine models and their 252 usefulness in staging human sepsis. these precisely time-and space-anchored data will 253 provide the community with rich and comprehensive foundations that will propel further 254 investigations into human sepsis. 255 animal model: male c57bl/6j mice were obtained from the jackson laboratory. mice were 8biopsies were used in this study, the institutional review board determined that informed 269 consent was not required. 270 murine kidneys were transported in rpmi1640 (corning), on ice immediately after surgical 273 procurement. kidneys were rinsed with pbs (thermofisher) and minced into eight sections. 274 each sample was then enzymatically and mechanically digested with reagents from multi-275 tissue dissociation kit 2 and gentlemacs dissociator/tube rotator (miltenyi biotec). the 276 samples were prepared per protocol "dissociation of mouse kidney using the multi tissue 277 dissociation kit 2" with the following modifications: after termination of the program "multi_e_2", 278 we added 10 ml rpmi1640 (corning) and 10% bsa (sigma-aldrich) to the mixture, filtered and 279 homogenate was centrifuged (300 g for 5 minutes at 4°c). cell pellet was resuspended in 1 ml 280 of rbc lysis buffer (sigma), incubated on ice for 3 minutes, and cell pellet washed three times 281 (300 g for 5 minutes at 4°c ). annexin v dead cell removal was performed using magnetic bead 282 separation after final wash, and the pellet resuspended in rpmi1640/bsa 0.04%. viability and 283 counts were assessed using trypan blue (gibco) and brought to a final concentration of 1 284 million cells/ml, exceeding 80% viability as specified by 10x genomics processing platform. 285 286 the sample was targeted to 10,000 cell recovery and applied to a single cell master mix with 288 lysis buffer and reverse transcription reagents, following the chromium single cell 3' reagent 289 kits v3 user guide, cg000183 rev a (10x genomics, inc.). this was followed by cdna 290 synthesis and library preparation. all libraries were sequenced in illumina novaseq6000 291 platform in paired-end mode (28bp + 91bp). fifty thousand reads per cell were generated and 292 the 10x genomics cellranger (v. 2.1.0) pipeline was utilized to demultiplex raw base call files to 300 fastq files and reads aligned to the mm10 murine genome using star 30 . cellranger 301 computational output was then analyzed in r (v.3.5.0) using the seurat package v. 3.0.0.9999, 302 31 . seurat objects were created for non-integrated and integrated (inclusive of all time points) 303 using the following filtering metrics: gene counts were set between 200-3000 and mitochondrial 304 gene percentages less than 50 to exclude doublets and poor quality cells. gene counts were log 305 transformed and scaled to 10 4 . the top 20 principle components were used to perform 306 unsupervised clustering analysis, and visualized using umap dimensionality reduction 307 (resolution 1.0). using the seurat package, annotation and grouping of clusters to cell type was 308 performed manually by inspection of differentially expressed genes (degs) for each cluster, 309 based on canonical marker genes in the literature 8-10,32,33 . in some experiments, we used 310 edger negative binomial regression to model gene counts and performed differential gene 311 expression and pathway enrichment analyses (topkegg, topgo, fig. 5, supplementary fig. 312 5a, supplementary fig. 6 , and david 6.8 fig. 7b . 34, 35 . 313 the immune cell subset was derived from the filtered, integrated seurat object and included the 314 macrophage/dc (cluster 10), neutrophil (cluster 19) and lymphocyte (cluster 13) cells. gene 315 counts were log transformed, scaled and principle component analysis performed as for the 316 integrated object above. umap resolution was set to 0.4, which yielded 14 clusters. the 317 clusters were manually assigned based on inspection of degs for each cluster, and cells 318 grouped if canonical markers were biologically redundant. we confirmed manual labeling with 319 an automated labeling program in r, singler 36 . 320 scenic analysis 37 was performed using the default setting and mm9-500bp-upstream-322 7species.mc9nr.feather database was used for data display. 323 we performed pseudotime analysis on the integrated seurat object containing all cell types as 325 well as the immune cell subset. cells from each of the seven time points were included and 326 were split into individual gene expression data files organized by previously defined cell type. a septic mouse kidney was immediately frozen in optimal cutting temperature media. a 10 µm 376 frozen tissue section was cut and affixed to a visium spatial gene expression library 377 preparation slide (10x genomics). the specimen was fixed in methanol and stained with 378 hematoxylin-eosin reagents. images of hematoxylin-eosin-labeled tissues were collected as 379 mosaics of 10x fields using a keyence bz-x810 fluorescence microscope equipped with a 380 nikon 10x cfi plan fluor objective. the tissue was then permeabilized for 12 minutes and rna 381 was isolated. the cdna libraries were prepared and then sequenced on an illumina novaseq 382 6000. using seurat 3.1.4, we identified anchors between the integrated single cell object and 383 the spatial transcriptomics datasets and used those to transfer the cluster data from the single 384 cell to the spatial transcriptomics. for each spatial transcriptomics spot, this transfer assigns a 385 score to each single cell cluster. we selected the cluster with the highest score in each spot to 386 represent its single cell associated cluster. using a loupe browser, expression data was 387 visualized overlying the hematoxylin-eosin image. 388 formalin-fixed paraffin-embedded cross sections were prepared with a thickness of 5µm. the 390 slides were baked for 60 minutes at 60 °c. tissues were incubated with xylene for 5 minutes 391 x2, 100% etoh for 2 minutes x2, and dried at room temperature. rna in situ hybridization was fluorescein plus evaluation kit (perkinelmer, inc) was used as secondary probes for the 396 detection of rna signals. all slides were counterstained with dapi and coverslips were 397 mounted using fluorescent mounting media (prolong gold antifade reagent, life 398 technologies). the images were collected with a lsm800 confocal microscope (carl zeiss). 399 no blinding was used for animal experiments. all data were analyzed using r software 401 packages, with relevant statistics described in results, methods and fig. legends . 402 data will be deposited to ncbi geo. the authors declare that all relevant data supporting the 404 findings of this study are available on request. 405 r scripts for performing the main steps of analysis are available from the lead contact on 407 request. 408 correspondence and requests for resources and reagents should be directed to and will be 410 fulfilled by the lead contact takashi hato (thato@iu.edu). 411 412 supplemental fig. 1-7 : refer to "supplemental_fig 1-7.pdf" 414 supplemental table 1 : cell-type specific differentially expressed genes from 0-48 hours, related 415 to fig. 1, supplemental fig. 1. lung-kidney cross-talk in the critically ill patient distant organ dysfunction in acute kidney injury: a 442 review sepsis: current dogma and new perspectives sepsis associated acute kidney injury bacterial sepsis triggers an antiviral response that causes translation shutdown rna sequencing of adult kidney: rare cell types and novel cell states revealed in fibrosis representation and relative abundance of cell-type selective markers in whole-452 kidney rna-seq data a single-nucleus rna-sequencing pipeline to decipher the molecular anatomy 454 and pathophysiology of human kidneys deep sequencing in microdissected renal tubules identifies nephron segment-specific transcriptomes single-cell transcriptomics of the mouse kidney reveals potential cellular targets of 458 kidney disease single-cell profiling reveals sex, lineage, and regional diversity in the mouse 460 kidney the macrophage mediates the renoprotective effects of endotoxin 462 preconditioning endotoxin preconditioning reprograms s1 tubules and macrophages to protect 464 the kidney endotoxin uptake by s1 proximal tubular segment causes oxidative stress in 466 the downstream s2 segment epithelial cell cycle arrest 468 in g2/m mediates kidney fibrosis after injury joint profiling of chromatin accessibility and gene expression in thousands of single 470 cells visualization and analysis of gene expression in tissue sections by spatial 472 transcriptomics structural basis for the recognition of sars-cov-2 by full-length human ace2 sars-cov-2 cell entry depends on ace2 and tmprss2 and is blocked by a 476 clinically proven protease inhibitor multiorgan and renal tropism of sars-cov-2 renal histopathological analysis of 26 postmortem findings of patients with covid-479 19 in china sars-cov-2 entry factors are highly expressed in nasal epithelial cells 481 together with innate immune genes identification and functional characterization of dendritic cells in the healthy 483 murine kidney and in experimental glomerulonephritis quantification of dendritic cell subsets in human renal tissue under 485 normal and pathological conditions macrophages in renal injury and repair the debate about dendritic cells and macrophages in the kidney distinct macrophage phenotypes contribute to kidney injury and repair pivotal role of plasmacytoid dendritic cells in inflammation and nk-cell 493 responses after tlr9 triggering in mice interferon-lambda modulates dendritic cells to facilitate t cell immunity 495 during infection with influenza a virus star: ultrafast universal rna-seq aligner comprehensive integration of single-cell data a transcriptional map of the renal tubule: linking structure to function single-cell transcriptomics of a human kidney allograft biopsy specimen defines a 503 diverse inflammatory response bioinformatics enrichment tools: paths toward the 507 comprehensive functional analysis of large gene lists reference-based analysis of lung single-cell sequencing reveals a transitional 509 profibrotic macrophage scenic: single-cell regulatory network inference and clustering the dynamics and regulators of cell fate decisions are revealed by 513 pseudotemporal ordering of single cells characterization of cell fate probabilities in single-cell data with palantir rna velocity of single cells cellphonedb: inferring cell-518 cell communication from combined expression of multi-subunit ligand-receptor complexes circlize implements and enhances circular 521 visualization in r we thank the kidney precision medicine project for making data available for human kidney 528 reference nephrectomy specimens. we thank daria barwinska for assistance with specimen 529 validation. this work was supported by nih k08-dk113223 to th, nih r01-dk080063, key: cord-016127-tbot0fc9 authors: hurtado, f. j.; buroni, m.; tenzi, j. title: sepsis: clinical approach, evidence-based at the bedside date: 2009-11-19 journal: intensive and critical care medicine doi: 10.1007/978-88-470-1436-7_25 sha: doc_id: 16127 cord_uid: tbot0fc9 sepsis is a common disease in intensive care medicine representing almost one third of patient admissions. its incidence has substantially increased over the past decades and overall mortality has declined during this period of time. it was reported that sepsis incidence increased from 82.7 to 240.4 per 100,000 population between 1979–2000. at the same time, sepsis global mortality decreased from 27.8 to 17.9% [1–3]. however, the absolute number of deaths significantly increased from 21.9 to 43.9 per 100,000 population. male gender, some chronic diseases like diabetes, immunosuppressive states, human immunodeficiency virus infections, and malignancies are factors that increase the risk for sepsis. some particular conditions like progressive number of organ dysfunctions, in-hospital-acquired infections and increasing age are associated with higher risk of death [1,4]. on the other hand, septic shock mortality only diminished from 61.6 to 53.1% [5]. this slight decline in mortality observed during recent decades could be attributable to improvements in supportive care and/or avoidance of iatrogenic complications. for example, the instrumentation of early goal resuscitation protocols not aiming at supranormal targets for cardiac output and oxygen delivery, and the use of lung protective strategies could explain at least in part this favorable change. other strategies directed to treat the pathophysiological mechanisms involved in the septic process like recombinant human-activated protein-c (rhapc), have also contributed to improve survival. however, mortality remains unacceptably high and further improvement in sepsis management is needed. sepsis is a common disease in intensive care medicine representing almost one third of patient admissions. its incidence has substantially increased over the past decades and overall mortality has declined during this period of time. it was reported that sepsis incidence increased from 82.7 to 240.4 per 100,000 population between 1979-2000. at the same time, sepsis global mortality decreased from 27.8 to 17.9% [1] [2] [3] . however, the absolute number of deaths significantly increased from 21.9 to 43.9 per 100,000 population. male gender, some chronic diseases like diabetes, immunosuppressive states, human immunodeficiency virus infections, and malignancies are factors that increase the risk for sepsis. some particular conditions like progressive number of organ dysfunctions, in-hospital-acquired infections and increasing age are associated with higher risk of death [1, 4] . on the other hand, septic shock mortality only diminished from 61.6 to 53.1% [5] . this slight decline in mortality observed during recent decades could be attributable to improvements in supportive care and/or avoidance of iatrogenic complications. for example, the instrumentation of early goal resuscitation protocols not aiming at supranormal targets for cardiac output and oxygen delivery, and the use of lung protective strategies could explain at least in part this favorable change. other strategies directed to treat the pathophysiological mechanisms involved in the septic process like recombinant human-activated protein-c (rhapc), have also contributed to improve survival. however, mortality remains unacceptably high and further improvement in sepsis management is needed. novel therapeutic approaches are under investigation and will probably be incorporated in the clinical practice in the near future. since 2002 the surviving sepsis campaign was introduced with the initial goal of increasing clinicians' awareness about severe sepsis mortality and to improve outcome in this patient population. it was pursued to generate a change in the standard of care that could finally result in a significant mortality reduction. a consensus committee from several international organizations was created and evidence-based guidelines were elaborated [6] . despite the fact that most of these recommendations were not supported by high levels of evidence, they represented the international consensus on the best available standards of care for the management of sepsis. these guidelines were recently updated and continue to be the core of the surviving sepsis campaign [7] . the clinical practice needs clear and concise recommendations based on the best available level of evidence. sepsis is defined as the host response to infection. in other terms, it is the clinical syndrome that results from the inflammatory response to infection. in the clinical setting, sepsis is diagnosed when an evident or suspected infection courses with a systemic response called the systemic inflammatory response syndrome (sirs). according to the 1991 north american consensus conference, sirs was defined by the presence of at least two of the following signs: body temperature >38ºc or <36ºc, heart rate >90 beats/min, respiratory rate >20 breaths/min (or paco2 <30 torr), and/or white blood cells count >12,000 or <3,000/mm 3 [8] . however, these signs are too sensitive and nonspecific for sepsis and could occur in many other different situations not related to infection. in an attempt to better reflect the systemic response to infection, the clinical manifestations described by bone et al. were expanded by the 2001 consensus conference [9] . other possible signs, symptoms, and laboratory findings were summarized (table 25.1) . again, most of them are also nonspecific for sepsis. it is well known that infection and sepsis are sometimes difficult to confirm. in an attempt to improve diagnostic capabilities, some biological markers were developed. procalcitonin (pct) and c-reactive protein (c-rp) have been proposed but it is considered that there is still no ideal biological marker for sepsis diagnosis [10, 11] . none of the mentioned biomarkers are absolutely specific, meaning that diagnosis or prognosis cannot be made solely on this basis. an infection probability score (ips) was also proposed to be calculated from several variables: body temperature, heart rate, respiratory rate, white blood cells count, c-reactive protein, and sequential organ failure assessment score (sofa). the potential role of such an index was recently evaluated [12] . some concepts and definitions remained unchanged after the last consensus conferences and should be emphasized. the following terms are widely accepted. infection is the pathologic process caused by the invasion of normally sterile tissues, fluids, or cavities by pathogenic microorganisms. sepsis is the clinical syndrome defined by the presence of infection and a systemic inflammatory response syndrome. severe sepsis relates to the presence of sepsis and one or more related organ dysfunctions. septic shock should be diagnosed when severe sepsis courses with acute circulatory failure. cardiovascular compromise becomes evident when arterial hypotension remains after adequate fluid infusion or there is need for vasopressor therapy. systemic hypotension is defined when arterial systolic pressure remains <90 mm hg, mean arterial pressure <60 mm hg, or there is a decrease in blood pressure >40 mm hg from previous values. different processes could occur during severe sepsis and septic shock at the same time. hypovolemia, maldistribution of blood flow within or between organs, vasoreghowever, cytokine release from the inflammatory reaction or prolonged tissue hypoxia is followed by severe microcirculatory abnormalities that become a central protagonist of organ dysfunction/failure [13] . the main significance of microvascular dysfunction has been studied during sepsis and major changes like decrease of both capillary density and microvascular blood flow were documented in vivo by video microscopy [14] . thus, peripheral gas exchange becomes impaired and tissue dysoxia ensues. these abnormalities also occur despite normal or even supranormal hemodynamic variables. in terms of peripheral oxygen metabolism, severe heterogeneity of oxygen distribution within the tissues is characteristic during septic shock. under-and overperfuse areas coexist within the same tissue resulting in an inhomogeneous tissue oxygen partial pressure distribution (pto2). in these conditions, metabolic demands are not met by microvascular oxygen delivery, making peripheral shunting and tissue dysoxia the cause of organ failure. however, metabolic abnormalities also occur at the cellular level. mitochondria dysfunction is secondary to oxidative and nitrative stress initiated by the inflammatory reaction. energetic failure develops and less high energy compounds are available for cellular function. this situation was referred to as cytopathic hypoxia and leads to multiple organ failure and death. mitochondria dysfunction and decreased atp production was documented during the course of sepsis in experimental and clinical situations. efforts should be made to preserve mitochondrial functioning and improve the cellular energetic state [15] [16] [17] . early and accurate recognition of the signs and symptoms of sepsis is mandatory after patient admission. risk factors like age, gender, race, immunocompromised states, presence of invasive instrumentation maneuvers, or any other condition that could represent a via for bacterial colonization. clinical presentation and laboratory findings are essential. fever is the hallmark of infection, but hypothermia is also possible in some patients. other nonspecific signs like tachycardia, tachypnea, and hypotension should also be documented. when looking for the source of infection a careful physical examination should be complemented with x-rays images, ct scans, ultrasound, etc. finally, it is necessary to investigate the presence and severity of organ dysfunction. in the vast majority of the cases this information is easily collected and diagnosis becomes straightforward. however, this is not always the case. it is important to realize that the septic patient is always at risk of death and some clinical signs may be indicative of disease severity. clinical demonstration of acute respiratory and/or circulatory failure, or any other organ dysfunction are indicative of the aggressive host response to the septic insult [8, 9] . since organ failure is an integral part of severe sepsis, a brief summary of major organ dysfunctions will follow. pulmonary or extrapulmonary ali is present in about 60-70% of severe sepsis. it is defined by pulmonary infiltrates in the chest x-ray and the absence of left ventricular failure (pulmonary wedge pressure <18 mm hg). pulmonary gas exchange is impaired showing a pao2/fio2 ratio under 300 for ali or below 200 for ards. most of the time, the severity of ali/ards determines mechanical ventilation. while mechanical ventilation will restore pulmonary gas exchange and decrease systemic metabolic demands, detrimental effects should be avoided by a rational application of protective ventilatory strategies. when the focus of infection is located outside the central nervous system (cns), the neurologic compromise could be attributable to septic encephalopathy. some other conditions may add secondary effects such as hypoxemia, metabolic and electrolytical disorders, and cerebral hypoperfusion during shock states. symptoms may vary from agitation, confusion, delirium, and coma. no focal neurologic deficits are present but myoclonias and seizures are possible [18] . severe cns derangement requires airway protection and ventilatory support. liver dysfunction is characterized by some degree of hepatomegalia and total bilirubin plasma levels >2 mg/dl. higher conjugate bilirubin concentration is characteristic and increased gamma glutamil transferase is frequently observed. moderate levels of aminotransferases generally <200 ui can also be found. decreased red blood cells without bleeding evidence and platelets <100,000/mm 3 are frequent findings. coagulation cascade has been widely studied. sepsis enhances coagulation and impairs fibrinolysis. endogenous-activated protein c that prevents microvascular thrombosis is decreased during sepsis. when small and medium microvessels become occluded, the disrupted microcirculation generates tissue dysoxia. given the context of severe sepsis, rhapc could contribute to ameliorate coagulation disorders [19] . renal dysfunction could course with normal or decreased urine output. increase in creatinine level >0.3 mg/dl from previous values or a percentage increase >50%, or a reduction in urine output (oliguria <0.5 ml/kg/h for more than 6 h) defines acute renal injury and is associated with poor outcome. arterial hypotension unresponsive to volume expansion defines septic shock. variable degrees of hemodynamic dysfunction may vary from hypodynamic to hyperdynamic shock. mortality increases according to the presence of shock, and metabolic markers like arterial lactate are useful to characterize disease severity and the response to treatment [8] . despite the fact that lactate concentration depends on the balance between tissue production and metabolism, a plasma level >4 mmol/l should be considered as indicative of circulatory failure. some other organ compromise could also be part of the multiple organ dysfunction syndrome. splanchnic ischemia and intramucosal acidosis ensue early during the course of sepsis. clinical expression includes changes in smooth muscle function like ileum or diarrhea. gastrointestinal bleeding because of stress ulcer or acute gastritis may also be a manifestation of sepsis. gastric intramucosal ph monitoring was used to identify and guide resuscitation therapy. increased levels of intraluminal pco2 are associated with tissue ischemia and mucosal acidosis. skeletal muscles are also affected by inflammatory mediators and reactive oxygen species. there is simultaneous decrease in protein synthesis and proteolysis. in conjunction, these factors explain decreased muscular force. respiratory muscles are involved and respiratory pump failure may aggravate or precipitate an acute respiratory failure. multiple organ dysfunction is part of the severe sepsis syndrome. poor prognosis is related to increased number of organ failures. technical resources for the management of organ dysfunction have improved in recent years and consume a substantial part of the therapeutic effort. most of the described dysfunctions are reversible as long as the infectious disease becomes controlled. however, the additive effects of the different failures may initiate a series of independent processes that may aggravate the patient status and be the cause of death. prognostic scores are helpful to predict mortality and some organ failure scores were proposed to evaluate severity and to follow the evolution of septic patients. the multiple organ dysfunction score (mods) and the sequential organ failure assessment (sofa) are frequently used for this purpose [20, 21] . to identify the source of infection and the microbial agent is crucial during sepsis. microbiological investigation is mandatory and adequate antibiotic therapy must be initiated as early as possible [7] . most of the time the diagnosis results from a correct anamnesis and clinical examination. suspicion of sepsis must be followed by complete bacteriological cultures, taking samples from blood and other possible foci of infection. some other special exams should not be deferred and may add complementary information. positive blood cultures are only confirmed in about 50% of the cases [22] . no bacterial etiology is identified in 20-30% of septic patients. almost 45% of the initial antibiotic selection should be changed or adjusted after blood cultures are informed. decreased mortality is related to prompt bacteriological identification [23] . despite the fact that infection is generally caused by bacterial agents, virus and fungal agents are possible, especially in immunocompromised patients. epidemiological data coming from each icu or hospital could be helpful when hospital-acquired infections are under study. infection is a frequent complication in polytrauma and in the critically ill patient who was subject to invasive procedures. increased life expectancy and special situations like organ transplant create further opportunities for microbial invasion and sepsis development. as mentioned before, biochemical markers of infection could be helpful in particular situations where diagnosis is not straightforward. procalcitonin (pct), c-reactive protein (crp), and some interleukins like interleukin-6 (il-6) have been proposed to contribute to diagnosis. however, further evidence is needed before these biomarkers are incorporated in the clinical practice. actually, some authors have considered pct as a good indicator for severe sepsis and septic shock [24, 25] . research on new biomarkers continues with the aim of early detection of patients at risk of severe sepsis [26] . the current management of severe sepsis and septic shock aims to control infection, achieve hemodynamic stabilization, modulate the immune response, and provide metabolic and organ support. evidence-based medicine has become the cornerstone of medical practice but it is difficult to apply in patients with sepsis. the ssc is a global initiative that involves several international organizations with the common objective of elaborating evidence-based guidelines and recommendations for the management of severe sepsis and septic shock. lack of high-level evidence coming from large rct is a severe limitation in sepsis. to accomplish these goals, experts determined that improving patient care was a possible task and could lead to a significant decrease in mortality. despite the fact that only a few of the guidelines were supported by high levels of evidence, it was agreed that they represent the best available evidence for the management of the septic patient. during the last consensus conference a grade system was agreed upon by the participants. guidelines were classified from a to d based on the levels of evidence; however, at the same time a strong or weak recommendation was introduced by the panel of experts [7] . the international guidelines of the surviving sepsis campaign will be briefly discussed below. this group of measurements should be accomplished within the first 6 h of patient admission. this could probably happen in the emergency department before icu admission. early identification and comprehensive resuscitation of septic patients will have a significant impact on outcome. the first 6 ''golden hours'' constitute a critical opportunity for the patient [27, 28] . resuscitation should be started immediately when hypotension or elevated serum lactate (>4 mmol/l) are detected and treatment should not be delayed until icu admission. initial resuscitation not only includes hemodynamic stabilization but also simultaneous administration of empiric antimicrobial drugs and actions directed toward the control of infection [7] . early resuscitation is initially based on aggressive volume expansion. it could be administered via a peripheral vascular access while a central venous line and central venous pressure (cvp) measurement are instrumented somewhat later within the initial hours. when fluid therapy does not restore arterial blood pressure or lactate remains elevated, administration of vasopressors becomes mandatory. the resuscitation goals are based on easily obtainable physiologic variables. treatment targets cvp pressures between 8 and 12 mm hg, mean arterial pressure ≥65 mm hg, urine output ≥0.5 ml/kg/h, and superior cava vein oxygen saturation ≥70% or mixed venous oxygen saturation ≥65% [7, 29] . no difference between crystalloids and colloids fluid was demonstrated [30] . however, it is mentioned that resuscitation with crystalloids is less expensive but requires more fluid to achieve the same end points and may result in more edema formation. fluid challenges of 1,000 ml of crystalloids or 300-500 ml of colloids over 30 min are recommended, but larger volumes or infusion rates could be required [7] . when resuscitation goals are not rapidly achieved vasopressor therapy must be started. there is no high-quality primary evidence to recommend norepinephrine over dopamine. however, norepinephrine could be more effective in reversing hypotension in patients with septic shock. the selected vasopressor, either norepinephrine or dopamine, should be titrated until map ≥65 mm hg. epinephrine is another alternative vasoactive agent when blood pressure is poorly responsive to norepinephrine or dopamine. low-dose dopamine should not be used for renal protection [31] . in patients requiring vasopressors, an arterial catheter and continuous arterial pressure monitoring must be instrumented. if central venous oxygen saturation remains <70% further fluid infusion and/or packed red blood cells transfusion should be considered. hematocrit ≥30% is desirable to assure systemic oxygen delivery. increase in cardiac index by the effect of dobutamine infusion to a maximum 20 μg/kg/min is recommended. dobutamine is the first line inotrope for patients with measured or suspected low cardiac output and adequate or high left ventricular filling pressures. a combination of inotropes/vasopressors, such as norepinephrine and dobutamine, is recommended if cardiac output is not directly measured [7] . antibiotics should be administered during the first hour of the initial resuscitation. the time taken to initiate effective antimicrobial therapy is one of the strongest predictors of outcome in septic shock [32] . initial antimicrobial selection should be wide enough to cover likely pathogens. there is evidence that failure to initiate appropriate antimicrobial therapy within this period of time correlates with increased mortality [33] . source control includes an appropriate diagnosis of the specific site of infection within the first 6 h. surgical procedures aimed at abscess draining, debridement of infected necrotic tissue, or removal of potentially infected devices should be instrumented without delay [7] . these practices are believed to be important for infection control but no randomized trials support them [34] . most of the measurements initiated in the previous stage will continue during the following hours. at the same time, some other therapeutic interventions could be started earlier, during the initial resuscitation phase. two big trials of patients with vasopressor-unresponsive septic shock showed a significant and faster resolution of shock when steroid therapy was associated [35, 36] . thus, low-dose intravenous hydrocortisone (≤300 mg/day) should be considered for adult septic patients when hypotension is poorly responsive to fluid resuscitation and vasopressors. on the other hand, an adrenocorticotrophic hormone (acth) stimulation test is not recommended. steroid therapy must be weaned once vasopressors are no longer required [7] . the importance of lung-protective strategies for patients with ali/ards is supported by clinical trials and has been widely accepted [37] . low tidal volume (6 ml/kg) and upper limit plateau pressure ≤30 cm h2o are desirable in patients with ali/ards. this respiratory pattern may result in paco2 increase above normal or permissive hypercapnia. a prone position should be considered when potentially injurious levels of fio2 or plateau pressure cannot be controlled. titration of positive end expiratory pressure (peep) should be made according to bedside measurements in an attempt to reach optimal levels of respiratory system compliance [7] . several randomized, observational clinical trials showed reductions in icu mortality when intensive insulin therapy was utilized [38, 39] . a large randomized trial recently showed that intense glucose control increased mortality. these authors found that a blood glucose target <180 mg/dl resulted in lower mortality than a target between 81 to 108 mg/dl. higher episodes of hypoglycemia were reported in the tight glucose control group [40] . based on this report, intense insulin therapy is questionable. however, the ssc recommendation is to maintain blood glucose levels below 150 mg/dl [7] . recent studies reported nonsignificant mortality reduction after rhapc administration in patients with a low risk of death or in the pediatric population. furthermore, rhapc administration is associated with increased risk of bleeding. the evidence concerning rhapc use in adults is primarily based on two rcts: the prowess and the address (stopped early for futility) [41, 42] . additional information comes from an open-label observational study, the enhance that suggested that early administration of rhapc was associated with better outcomes [43] . as a result, the latest recommendation of the ssc is to consider rhapc only for adult patients at high risk of death (apache ii ≥25 or multiple organ failure). during patient selection, possible contraindications for rhapc administration should be discharged. red blood cell transfusion should be administered when haemoglobin decreases below 7.0 g/dl. a hemoglobin target between 7.0 to 9.0 g/dl in adult septic patients is recommended. do not use fresh frozen plasma to correct laboratory clotting abnormalities unless there is bleeding evidence or planned invasive procedures. administer platelet concentrates when platelet counts are <5,000/mm 3 regardless of bleeding. platelet counts between 5,000 to 30,000/mm 3 do not call for platelet administration unless there is a significant risk of bleeding [7] . • sedation and analgesia in sepsis. it is recommended to use sedation protocols with daily interruption/lightening to produce awakening [7] . • renal replacement therapy. current evidence is insufficient to draw strong conclusions regarding the best replacement therapy method for ari in septic patients [44, 45] . it is not clear whether high doses of renal replacement may influence patient outcome [46] . intermittent hemodialysis and continuous veno-venous hemodiafiltration (cvvh) are considered equivalent for septic patients. however, cvvh and sustained low-efficiency dialysis will probably offer a safer and easier management in hemodynamically unstable patients. • bicarbonate therapy. sodium bicarbonate infusion must not be used for the purpose of improving hemodynamics or reducing vasopressor requirements when treating hypoperfusion-induced lactic acidemia with a ph ≥7.15 [7] . • deep vein thrombosis prophylaxis. use either low-dose unfractioned heparin (ufh) or low-molecular weight heparin (lmwh), unless contraindicated. use a mechanical prophylactic device when heparin is contraindicated [7] . • stress ulcer prophylaxis. stress ulcer prophylaxis based on h2 blockers or proton pump inhibitors could be used for septic patients [7] . • nutritional support. it is very important to initiate early nutritional support in critically ill patients. enteral nutrition is generally safer and more effective than total parenteral nutrition. immunonutrition needs to be further studied before clear recommendations can be made. as mentioned before, these guidelines were based on the best available evidence. ongoing and future studies will provide further valuable information and changes in these recommendations will become necessary. the use of these guidelines is not easy in clinical practice. it was an objective of the ssc to facilitate the instrumentation of these recommendations [47] . in the last phase of the ssc the concept of sepsis bundles was introduced. this idea aimed to create a series of simple recommendations easily applicable as packages of measurements for the clinical setting [48] . conceptually, a bundle is a group of interventions that when implemented together will result in better outcomes. the bundles were developed in conjunction with the institutes for health care improvement (ihci). different tools were created to assist clinicians at the bedside. these clinical tools and databases are available on the ssc web page. treatment of severe sepsis can be organized into two groups of interventions known as the initial resuscitation bundle (initial 6 h) and the management bundle (24 h bundle) [49] . table 25 .2 summarizes this particular approach. some recent works have studied bundles compliance and new favorable results on outcomes are coming. it has been shown that bundles compliance is associated with a reduction in icu mortality and length of stay [50, 51] . the epidemiology of sepsis in the united states from 1979 through 2000 epidemiology of severe sepsis in the united states: analysis of incidence, outcome, and associated cost of care is the mortality rate for septic shock really decreasing? epidemiology of sepsis and infection in icu patients from an international multicentre cohort study has the mortality of septic shock changed with time surviving sepsis campaign: guidelines for management of severe sepsis and septic shock surviving sepsis campaign: international guidelines for management of severe sepsis and septic shock definitions for sepsis and organ failure and guidelines for the use of innovative therapies in sepsis sccm/esicm/accp/ats/sis. international sepsis definitions conference procalcitonin as a marker of bacterial infection in the emergency department: an observational study c-reactive protein levels correlate with mortality and organ failure in critically ill patients a prospective evaluation of the infection probability score (ips) in the intensive care unit the microcirculation is the motor of sepsis bench-to-bedside review: sepsis is a disease of the microcirculation cytopathic hypoxia. is oxygen use impaired in sepsis as a result of an acquired intrinsic derangement in cellular respiration? cellular dysfunction in sepsis septic diaphragmatic dysfunction is prevented by mn(iii)porphyrin therapy and inducible nitric oxide synthase inhibition update in intensive care and emergency medicine. brain failure coagulation in sepsis multiple organ dysfunction score: a reliable descriptor of a complex clinical outcome the sofa (sepsis related organ failure assessement) score to describe organ dysfunction failure incidence, risk factors, and outcome of severe sepsis and septic shock in adults decreased mortality associated with prompt gram staining of blood cultures procalcitonin as a diagnostic test for sepsis in critically ill adults and alter surgery or trauma: a systematic review and meta-analysis the diagnostic value of procalcitonin in severe sepsis a prospective, multicenter derivation of a biomarker panel to assess risk of organ dysfunction, shock, and death in emergency department patients with suspected sepsis early goal-directed therapy in the treatment of severe sepsis and septic shock early goal-directed therapy in severe sepsis and septic shock: a contemporary review of the literature management of sepsis: early resuscitation colloid use for fluid resuscitation: evidence and spin use of dopamine in acute renal failure: a meta-analysis duration of hypotension prior to initiation of effective antimicrobial therapy is the critical determinant of survival in human septic shock the influence of inadequate antimicrobial treatment of bloodstream infections on patient outcomes in the icu setting approach to the patient with sepsis effect of treatment with low doses of hydrocortisone and fludrocortisone on mortality in patients with septic shock hydrocortisone therapy for patients with septic shock ventilation with lower tidal volumes as compared with traditional tidal volumes for acute lung injury and the acute respiratory distress syndrome intensive insulin therapy in critically ill patients intensive insulin therapy in the medical icu intensive versus conventional glucose control in critically ill patients (2009) the nice-sug-ar study investigators efficacy and safety of recombinant human activated protein c for severe sepsis drotrecogin alfa (activated) for adults with severe sepsis and a low risk of death drotrecogin alfa (activated) treatment in severe sepsis from the global open-label trial enhance: further evidence for survival and safety and implications for early treatment continuous versus intermittent renal replacement therapy: a meta-analysis acute renal failure in the intensive care unit: a systematic review of the impact of dialytic modality on mortality and renal recovery the va/nih acute renal failure trial network: intensity of renal support in critically ill patients with acute kidney injury sepsis change bundles: converting guidelines into meaningful change in behavior and clinical outcome the impact of compliance with 6-hour and 24-hour sepsis bundle on hospital mortality in patients with severe sepsis: a prospective observational study implementation of the surviving sepsis campaign guidelines for severe sepsis and septic shock: we could go faster key: cord-015946-biu5zxd1 authors: peng, daizhi; liu, xiao title: research advances in biomarker for sepsis date: 2016-11-16 journal: advanced trauma and surgery doi: 10.1007/978-981-10-2425-2_15 sha: doc_id: 15946 cord_uid: biu5zxd1 sepsis is one of the most common causes of death in severely injured patients worldwide. the early detection of sepsis still has to be solved in clinical practice. the delayed diagnosis often contributes to inappropriate antimicrobial treatment and subsequent high mortality. sepsis biomarkers are produced during the host response to infection. traditional biomarkers are polypeptides and/or proteins derived from this response. omics-based biomarkers are screening out from all kinds of molecules of host response while high-throughout omics technologies are emerging. this review describes traditional and potential omics-based sepsis biomarkers from currently available literatures. the combination of these biomarkers would refine the identification of sepsis for further clinical and experimental sepsis studies. sepsis is one of the leading fatal causes of critical injured and ill patients all over the world. despite recent advances in comprehensive management of trauma patients, sepsis is still a life-threatening condition with poor outcome. the risk factors of patients for the development of sepsis usually refer to their physiological characteristics, underlying illnesses and clinical treatment backgrounds. the patients with one or more of these factors are susceptible to sepsis. the most vulnerable populations for sepsis are the elderly and infants, patients with chronic diseases, patients with severe trauma and burns, those who are immunocompromised or receiving immunosuppressive therapy, and malnourished and debilitated biomarkers are molecular indicators that help doctors diagnose illnesses, predict the outcome or identify which people are susceptible. biomarkers, should be some quantifiable measurements of biological homeostasis and by defining the normal status can provide a frame of reference for predicting abnormal or pathogenic processes [2] , and make an impact on clinical decision making in time. most commonly proposed sepsis and infection biomarkers including c-reactive protein (crp), procalcitonin (pct) [5, 6] , cytokines (tnf-α, il-1, il-6, il-10, osteopontin) [7, 8] , chemokines [macrophage migration inhibitory factor (mif), high-mobility-group box 1 (hmgb1)] [9, 10] , soluble receptor [soluble triggering receptor expressed on myeloid cells 1 (strem-1), soluble urokinase-type plasminogen activator receptor (supar)] [11, 12] etc. given the complicated pathophysiology in sepsis which involves hundreds of mediators or single molecules, it is unlikely to identify one single biomarker which is able to satisfy all the existing needs and expectations in sepsis research and management. crp, for example, is frequently used to assess the presence of infection and sepsis [2] , and there is a positive correlation between its plasma level and the risk of organ dysfunction and death [13, 14] . however, plasma concentrations of crp may increase during minor infection and do not adequately reflect the severity of infection, or remain at a high level for several days, even overstay the infection [2] . besides, crp may experience an increase during the inflammation caused by noninfectious etiologies, such as tumor, tissue necrosis or operation, which explains its nonspecificity as an early stage sepsis biomarker. meanwhile, pct, perceived as the most potential biomarker, is listed as one of the diagnostic criteria for sepsis [15] . a recent meta-analysis of pct that included 30 studies found mean sensitivity and specificity of 0.77 and 0.79 respectively, with the area under the receiver operating characteristic curve (auc) was 0.85 [16] . although pct was thought as a helpful biomarker for early diagnosis of sepsis in critically ill patients, it was not suitable to be recommended as the single definitive diagnostic test [16] . therefore, some researchers have put forward that combinations of biomarkers may overcome the limitations mentioned above. in a prospective cohort study including 151 patients with systemic inflammatory response syndrome (sirs), kofoed et al. [17] found that the aucs of six biomarkers-supar (0.5), strem-1 (0.61), mif (0.63), pct (0.72), neutrophil count (0.74), crp (0.81)-for detection of a bacterial cause of inflammation had ranged from 0.5 to 0.81. with method reported by xiong et al. [18] , which discussed the statistical estimation of the optimum linear combination test and the associated maximum area under the roc curve, kofoed got the combined auc of the six-marker test at 0.88. consequently, the six-marker test had a better diagnostic accuracy in detecting bacterial versus nonbacterial causes of inflammation, and significantly greater than that of each single marker. similarly, in another prospective research among the critically ill patients [19] , a 'bioscore' combining the polymorphonuclear neutrophil (pmn) cd64 index together with pct and strem-1 serum levels was put forward to diagnose sepsis and had a better performance with an auc of 0.97 than that of each individual biomarker, and was externally confirmed in the validation cohort with 90.9 % of patients being correctly classified by the very model. although the combination of biomarkers do improve diagnostic sensitivity and specificity, due to the factors of time-consuming, economic cost, the amount of sample, the feasibility of biomarker detection method and so on, it has a limited application in clinical practice and still needs further prospective studies conducted in multicenter on cost-effectiveness. circulating dna, including nuclear dna (ndna) and mitochondrial dna (mtdna), can either actively release or be released passively into the blood stream after rupture or necrosis of host cells [20, 21] . these nucleic acids released in the plasma during sepsis could serve as danger associated molecular patterns (damps) [22] , which make them potential biomarkers for the very condition [23, 24] . a clinical study [22] found that plasma cytokine concentrations, as well as ndna and mtdna levels of septic shock patients were increased at the onset of septic shock and remained elevated. and during the first 5 days of septic shock, ndna levels consistently correlated with plasma cytokine concentrations as well as with the shock-related parameter norepinephrine infusion rate and markers of organ damage (total bilirubin and creatinine). these findings not only indicate a relationship between plasma ndna levels and the inflammatory response, but also demonstrate that ndna levels are associated with markers of shock and organ damage in septic shock patients. genomics, a discipline in genetics and an emerging field, explains physiological or pathophysiological events from the point of view of complete set of dna, including recombinant dna, dna sequencing methods, and bioinformatics to sequence, assemble, and analyze the function and structure of genomes. sepsis can be regarded as a polygenic syndrome initiated by infection. genetics plays a crucial role in both susceptibility and response to infection [25] , and genetic predisposition influences clinical outcomes of infectious diseases [26, 27] . a gene polymorphism is defined as regular occurrence (>1 %) of two or more alleles at a particular chromosome location. several polymorphisms of genes broadly involved in inflammation, immunity, and coagulation have been linked with susceptibility to sepsis, or outcome of sepsis [25] , and have become the focus of most gene association studies of sepsis as well. snp, the most common type of polymorphisms, is a substitution, deletion, or insertion of a single nucleotide occurring in approximately 1 per 1000 base pairs of human dna. snp can lead to an altered protein, a change in the amount of normal protein expression, or no discernible change in protein function [28] . study of snp genotypes in sepsis helps identify potential markers of susceptibility, severity, and clinical outcome. extensive researches on snp genotyping of main genes cd14 [29] [30] [31] [32] , toll like receptors (tlrs) [27, 33] , lipopolysaccharide-binding protein (lbp) [34] , cytokines [26, 33, 35, 36] and coagulation factors [37, 38] -have provided valuable information for sepsis. for example, burn patients with tlr4 and tnf-α polymorphisms were 1.8 times more likely to develop severe sepsis, but none of them were significantly associated with mortality [33] . tlr1 snps are associated with increased mortality in patients with gram-positive sepsis after traumatic injury, which may represent a novel marker of risk for death in critically injured patients [27] . and most recently, it was found that there was a new association between vascular endothelial growth factor (vegf) +936 cc genotype and the risk to develop acute kidney injury (aki) in severe sepsis patients [35] . genome-wide snp genotyping assays allow to detect hundreds of thousands of snps accurately in a single experiment [39, 40] and are expected to be of great applicative prospect in finding novel sepsis susceptibility-associated snp genotypes. in order to evaluate the validity of these studies and translate this concept to the bedside, several important factors have to be kept in mind: potential confounding variables should be recognized and matched; positive association studies and replicate studies should be validated and analyzed on the basis of the primary hypothesis other than multiple comparisons; large scale collaborations and studies on sepsis susceptibility-associated snp genotypes need to be performed for the sake of possible new risk factors at the genetic backgrounds of sepsis development [25, 41] . genes concerning immunity and inflammation are subject to epigenetic regulation [25] , which refers to heritable changes in gene expression that are not related to direct dna sequence changes [42] . dna methylation and histone post-translational modifications play vital roles in the epigenetic control [43] and gene expression, and strongly impact on the host defense responses. sepsis induces epigenetic changes in dendritic cells and lymphocytes rendering the host immune deficiency for a long period after the initial sepsis challenge [44] [45] [46] . late-phase immunosuppression of sepsis is strengthened by a postmortem study [47] . by suppression of proinflammatory gene products and subsequent immune cell activation and proliferation, epigenetic mechanisms are put forward to have an influence on the very stage of sepsis, which may not only provide a better understanding of septic mechanism but also yield important biomarkers [48] . dna methylation, the addition of methyl group to cytosine or adenine nucleotide, is now replacing the biological markers with their high specificity, sensitivity and prognostic efficacy. a retrospective investigation has showed that calcitonin-related polypeptide α (calca) gene promoter methylation varied with different types of preterm bacterial sepsis [49] . based on this finding, another study demonstrated that global dna methylation varied significantly among newborns with sepsis and those without sepsis [50] . more specifically, a recent study analyzed the cpg methylation status in the epigenome of septic and non-septic babies [51] or icu patients [52] . given biological and clinical significance, they found 81 differentially methylated cpgs located in 64 genes, and a panel of differentially methylated protocadherin β (pcdhb) genes that play vital role in leukocyte cell adhesion and wnt signaling pathway. these genes play vital role in calcium dependent cell to cell adhesion and other immunological processes like antigen processing and presentation. in sepsis, suppression of leukocyte cell adhesion and migration may exaggerate disease severity and poor outcome due to multiple organ dysfunctions. therefore, this study provides some novel insights into the role of dna methylation in neonatal sepsis. however, further studies are called for exploring the clinical relevance as well as related therapeutic approaches of the observed findings. all the above mentioned sepsis biomarkers are derived from the development of genomics technology and summarized in table 1 . the immune responses involved in sepsis are so complicated that the exact molecular mechanism remains to be fully elucidated [53] . the balance between pro-inflammatory responses and anti-inflammatory responses is closely related to the expression and regulation of relevant genes [54] . hence, evaluating the key gene expression profiles by high throughput dna chip may reveal the immune status of septic patients. researchers have found specific changes of gene expression with microarray in certain organs and tissues of septic mice model, which including heart [55] , liver, spleen [56] , leucocytes [57] and so on. accordingly, lukaszewsk recruited 92 icu patients who had the risk of developing into sepsis [58] . the mrna expression levels of il-1β, il-6, il-8, il-10, tnf-a, fasl and ccl2 in their blood leukocytes were measured on a daily basis by means of real-time reverse transcription pcr (rt-pcr), and analyzed with a nonlinear technique (neural network analysis). the data correctly predict the onset of sepsis in an average of 83.09 % of patient cases between 1 and 4 days before clinical diagnosis with high sensitivity and selectivity (91.43 and 80.20 %, respectively). sutherland et al. [59] evaluated transcriptional profiles in circulating white blood cells of icu sepsis patients, post-surgical patients and healthy controls with a microarray and multiplex tandem (mt)-pcr. a panel of 42-gene expression markers was identified, by which the prediction of sepsis within a mixed inflammatory population had an auc between 86 and 92 %. sepsis has a unique gene expression profile that is different from uninfected inflammation and becomes apparent prior to the clinical manifestations of sepsis for 0-48 h [60] . in that case, the specific gene expression profile, which may involve the function of innate immunity, cytokine receptors, t cell differentiation as well as the protein synthesis, may make a reference for early diagnosis of sepsis. however, an important limitation of transcriptomics is that it only partially reflects the steady-state mrna abundance, and the degree of mrna abundance is influenced by multiple factors, and does not provide any direct information about gene end products (proteins), nor post-translational modifiers of protein function [61] . when it comes to the sample used as rna source, there is a contradiction. for the whole blood approach, it may be difficult to interpret the confounded data of rna for the reason of the heterogeneity among blood cell populations. as for the cell-specific approach, there is a possibility to miss relevant expression information from other cells due to the complexity of clinical sepsis [61] . it highlights the necessity of linking theory to clinical practice. micrornas (mirnas) is a class of short rnas with 18-25 nucleotides in length which regulate gene expression in a post-transcriptional manner via sequence-specific interaction with target sites in mrna [62] , associated with various physiological and pathological processes. the levels of mirna in serum and plasma are consistent among individuals of the same species, resistant to rnase a digestion, and stable even after the freeze-and-thaw and a long term of storage [63, 64] . the stability of mirna makes it a potentially useful candidate for diagnostic and other clinical applications. although the source of circulating mirna is still unclear, it has been proved that there is a link among a range of diseases, such as circulating mirna and cancer [65, 66] , trauma [67, 68] , acute pancreatitis [69] , and hepatitis [70] . when it comes to sepsis, by using genome-wide mirna profiling with microarray in peripheral blood leukocytes and quantitative rt-pcr, vasilescu [71] found that mir-150 levels were significantly reduced in both leukocytes and plasma of sepsis patients and had a negative correlation with the level of disease severity measured by the sequential organ failure assessment (sofa) score, which made it a biomarker of early sepsis. similarly, zeng [72, 73] investigated the levels of mir-150 and mir-143 in peripheral blood leukocytes in sepsis patients with rt-pcr, and found that the expression levels of mir-150 and mir-143 were significantly decreased in sepsis patients and could reflect the severity of sepsis in certain degree, which not only made it a marker to reflect the situation of inflammatory response, but also made it a prognostic marker in sepsis. recently, higher serum mir-133a levels were found among sepsis patients in icu [74] . as they were significantly correlated with disease severity, classical markers of inflammation and bacterial infection, as well as organ failure, high mir-133a levels were considered as independent biomarkers for unfavorable prognosis of critically ill patients. however, given that the pathophysiological process of sepsis involves a variety of tissues and organs, a simple screen for mirna differentially expressed in leukocytes may omit those secreted by other cells. wang et al. [75] used genome-wide microarray to identify differential serum mirnas in survival and non-survival sepsis patients, and then further validated the differential expressions of mir-297 and mir-574-5p by rt-pcr in a larger group. the serum mir-574-5p together with sepsis stage and sepsis-related organ failure assessment scores has a better predictive capability for the death of sepsis patients. in addition, serum mir-146a and mir-223 were found significantly reduced in septic patients compared with sirs patients and healthy controls which might serve as new biomarkers for sepsis with high specificity and sensitivity [76] . due to our knowledge on serum mirnas is still at a primary stage, the expression level of circulating mirnas at different stages of sepsis and their potential correlation with injured organs need further investigation. to sum up, from the point of view of gene transcription, mirna may undertake the task of diagnosing sepsis in an early stage and evaluating the prognosis, as well as becoming the new target for sepsis therapy. as discussed above, epigenetic factors not only include histone modifications and dna methylation, but also contain non-coding rnas(ncrnas), which have diverse size and can be generated from intergenic regions, introns, or enhancers [45] . lncrnas are transcripts longer than 200 nucleotides and lack protein-coding capacity. peng et al. [77] first discovered the widespread differential expression of lncrnas in response to severe acute respiratory syndrome coronavirus (sars-cov) virus infection. accordingly, there is a possible link between lncrnas and the host defense response against infection. lncrna has the potential to become new class of biomarkers and new therapeutic target for infectious diseases. however, as the functions of lncrnas remain largely unexplored, there is a need for future studies on their regulatory role in infection. all the transcriptomics-based biomarkers mentioned above are outlined in table 2 . proteome is the complete set of proteins that can be expressed by the genetic material of an organism. proteomics is the analysis of the expression, localizations, functions, and interactions of proteomes. compared to other immunologic tests, proteomics is a novel method with advantages of high throughput, high sensitivity and specificity. the development of proteomics has allowed for a better understanding of the molecular bases concerning the identification of cell signaling, modifying protein, post-translation modification pathway, as well as the characterization of specific biological markers [78] . proteomics has irreplaceable clinical significance and an expansive application prospect in studies of sepsis biomarkers. in a rabbit sepsis model by intravenous injection of pseudomonas aeruginosa at 24 h after scald, 11 discrepant expression proteins from lymphocyte were found by matrix-assisted laser desorption/ionization time of flight mass spectrometry(maldi-tof ms). they are related with the folding, assembling, transportation and degradation of proteins, signal transmission, inflammation, immunization, energy metabolism, the proliferation, differentiation and apoptosis of cells [79] . in a recent research, 41 differential expressed proteins in the neutrophils from acinetobacter baumannii sepsis rats were identified using two-dimensional electrophoresis and mass spectrometry [80] . they included antioxidant proteins, signaling proteins, cytoskeleton and regulatory proteins, energy metabolism and protease protein, which may play a key role in such kind of sepsis and provide potential clues in early diagnosis and treatment of sepsis. in clinic, ykl-40 was identified with proteomics analysis on a significantly higher expression level in serum samples from sepsis patients and considered as a possible biomarker of sepsis [81] . paugam-burtz et al. [82] used plasma profiling coupling proteinchip array with surface-enhanced laser desorption ionization time-of-fly mass spectrometry (seldi-tof ms) to analyze the plasma of postoperative patients, and found that a combination of five plasma protein peaks may have potential as diagnostic biomarkers of postoperative sepsis in patients undergoing liver transplantation. even so, these proteins remain to be identified and validated in more clinical trials. although many potential sepsis biomarkers have been revealed by genomics, transcriptomics, and proteomics, the changes of cellular metabolism in sepsis should be paid attention. metabolomics is an emerging omics technology following genomics and proteomics and focuses on the metabolic products with a molecular weight less than 1000 kd under the physiological or pathological status. it can analyze the biochemical events of cells, tissues or organs and evaluate the disease and its severity. the research methods of metabolomics mainly include nuclear magnetic resonance (nmr), gas chromatography/mass spectra (gc/ms), high performance liquid chromatography/mass spectra (hplc/ms). the development of sepsis involves the reactions of multiple systems on various levels, which has a significant influence on the expression levels and activities of metabolic enzymes. and by detecting the concentration and ratio changes of those metabolites involved, a better understanding of condition and prognosis of sepsis may be achieved at an early stage [83] . metabolic profile of the serum from septic rats with cecal ligation and puncture was achieved with the help of nmr and lc/ms [83] . in the septic rats, especially the non-survivors, many free fatty acids showed a lower level which may be consumed greatly for energy supply in sepsis and may be related with the prognosis of sepsis. moreover, there was a rise of some polyunsaturated fatty acids in the very group, which may have a relationship with the increased anti-inflammatory effect. based on the metabolic profile analysis, a model for outcome predication was built with high sensitivity and specificity, which provided a novel method for sepsis prognosis judgment. nmr-based metabolic profiling revealed the difference of metabolites of energy metabolism and inflammation in lung tissue, bronchoalveolar lavage (bal) fluid, and serum samples between the septic rat and the control rat [84] . in septic rats, creatine concentration increased in all the three types of samples, whereas alanine and phosphoethanolamine concentrations increased only both in lung tissue and in serum. myoinositol increased in lung tissue but decreased in bal fluid. in addition, acetoacetate increased whereas formate decreased in serum. and with the construction of a predictive model for diagnosis of sepsis using partial least-squares discriminant analysis, the preliminary goal of sepsis diagnosis was achieved. the possible sepsis biomarkers screened out from the application of proteomics and metabolomics technologies are summarized in table 3 . sepsis involves sophisticated pathophysiologic changes among various organs and different systems, so that comprehensively identifying sepsis biomarkers and understanding sepsis molecular mechanism from the perspectives of omics may provide valuable information about a more macroscopical and authentic state following infection. the most important issue involved in the researches of sepsis biomarkers is the criteria of septic cases. the golden standard of infection, currently, still depends on the results of clinical microbiology laboratory, which, however, due to the severity of the disease, the load and type or growth capacity of pathogens, and the use of antibiotic treatment may show a negative result while patients has clinical manifestation of infection [85] . or false positive results may present for the reason of contamination. therefore, whether this gold standard is appropriate in sepsis needs further reflection and investigation. there is a lack of effective and united assessment methods for sepsis biomarkers, especially in the multi-centered or multi-index researches. valid evaluation needs to be performed to pick out the ideal diagnostic indicators, those both of high sensitivity and specificity in the field of data statistics and of great practical use in clinical practice. majority studies of sepsis only focus on a single set of omics technology (table 1 ) rather than apply the combination of multiple omics approaches. since different omics may display sepsis mechanism at various levels of a specific molecule or a particular group of sepsis-associated molecules, the comprehensive application of two or more omics may provide integrated information of potential sepsis biomarkers. mapit algorithm (multi analyte pathway inference tool), for example, enables principled integration of epigenomics, transcriptomics, and proteomics data for cancer diagnosis, prognosis, and biomarker discovery [86] . last but not the least, as mentioned above, the lack of a clear insight of the pathophysiology of sepsis process will, to some extent, put off the sepsis biomarker researches [2] . but with the application of state-of-art technology and exploration in a novel view, the progress in sepsis biomarkers will promote awareness and understanding of sepsis. duration of hypotension before initiation of effective antimicrobial therapy is the critical determinant of survival in human septic shock new approaches to sepsis: molecular diagnostics and biomarkers the third international consensus definitions for sepsis and septic shock (sepsis-3) the immunopathogenesis of sepsis diagnostic value of procalcitonin, interleukin-6, and interleukin-8 in critically ill patients admitted with suspected sepsis serum procalcitonin and c-reactive protein levels as markers of bacterial infection-a systematic review and meta-analysis cytokine profiles as markers of disease severity in sepsis: a multiplex analysis serum levels of osteopontin are increased in sirs and sepsis redox responses in patients with sepsis: high correlation of thioredoxin-1 and macrophage migration inhibitory factor plasma levels role of high mobility group box 1 in inflammatory disease: focus on sepsis accuracy of plasma strem-1 for sepsis diagnosis in systemic inflammatory patients-a systematic review and meta-analysis usefulness of supar as a biological marker in patients with systemic inflammation or infection: a systematic review c-reactive protein levels correlate with mortality and organ failure in critically ill patients plasma c-reactive protein levels are associated with mortality in elderly with acute lung injury sccm/esicm/accp/ats/sis international sepsis definitions conference procalcitonin as a diagnostic marker for sepsis: a systematic review and meta-analysis use of plasma c-reactive protein, procalcitonin, neutrophils, macrophage migration inhibitory factor, soluble urokinase-type plasminogen activator receptor, and soluble triggering receptor expressed on myeloid cells-1 in combination to diagnose infections: a prospective study combining correlated diagnostic tests: application to neuropathologic diagnosis of alzheimer's disease combination biomarkers to diagnose sepsis in the critically ill patient circulating cell-free dna: an up-coming molecular marker in exercise physiology circulating dna. its origin and fluctuation plasma nuclear and mitochondrial dna levels, and markers of inflammation, shock, and organ damage in patients with septic shock neutrophil-derived circulating free dna (cf-dna/nets): a potential prognostic marker for posttraumatic development of inflammatory second hit and sepsis diagnostic accuracy of neutrophil-derived circulating free dna (cf-dna/nets) for septic arthritis genetics and genomics in pediatric septic shock the association of interleukin 6 haplotype clades with mortality in critically ill adults toll-like receptor 1 polymorphisms and associated outcomes in sepsis after traumatic injury: a candidate gene association study mechanisms and regulation of the gene-expression response to sepsis cd14 expression in the first 24 h of sepsis: effect of −260c > t cd14 snp the influences of cd14 −260c > t polymorphism on survival in icu critically ill patients the −260 c-> t promoter polymorphism of the lipopolysaccharide receptor cd14 and severe sepsis in trauma patients innate immunity snps are associated with risk for severe sepsis after burn injury tlr4 and tnf-alpha polymorphisms are associated with an increased risk for severe sepsis following burn injury a functional variant of lipopolysaccharide binding protein predisposes to sepsis and organ dysfunction in patients with major trauma genetic predisposition to acute kidney injury induced by severe sepsis il-10, il-6 and cd14 polymorphisms and sepsis outcome in ventilated very low birth weight infants the single nucleotide polymorphism ser128arg in the e-selectin gene is associated with enhanced coagulation during human endotoxemia 4g/5g promoter polymorphism in the plasminogen-activator-inhibitor-1 gene in children with systemic meningococcaemia a review of clinical competence assessment in nursing next generation genome-wide association tool: design and coverage of a high-throughput european-optimized snp array genetic polymorphisms and sepsis an operational definition of epigenetics epigenetic control epigenetic regulation of dendritic cell-derived interleukin-12 facilitates immunosuppression after a severe innate immune response epigenetics and bacterial infections. cold spring harb perspect med adoptive transfer of naive dendritic cells in resolving post-sepsis long-term immunosuppression immunosuppression in patients who die of sepsis and multiple organ failure epigenetic regulation of immune cell functions during post-septic immunosuppression dna methylation pattern of calca in preterm neonates with bacterial sepsis as a putative epigenetic biomarker global dna methylation in neonatal sepsis comparison of genomic dna methylation pattern among septic and non-septic newborns-an epigenome wide association study methylation metabolism in sepsis and systemic inflammatory response syndrome changes in the inositol lipid signal system and effects on the secretion of tnf-alpha by macrophages in severely scalded mice mechanism of altered tnf-alpha expression by macrophage and the modulatory effect of panax notoginseng saponins in scald mice an investigation of changes in gene expression profile of heart tissue in a rat sepsis model sepsis gene expression profiling: murine splenic compared with hepatic responses determined by using complementary dna microarrays change of gene expression spectra of leucocyte in sepsis mice presymptomatic prediction of sepsis in intensive care unit patients development and validation of a novel molecular biomarker diagnostic test for the early detection of sepsis gene expression profiles differentiate between sterile sirs and early sepsis clinical review: sepsis and septic shock-the potential of gene arrays circulating micrornas: association with disease and potential use as biomarkers characterization of micrornas in serum: a novel class of biomarkers for diagnosis of cancer and other diseases circulating micrornas as stable blood-based markers for cancer detection detection of elevated levels of tumour-associated micrornas in serum of patients with diffuse large b-cell lymphoma serum microrna signatures identified in a genome-wide serum microrna expression profiling predict survival of non-small-cell lung cancer altered expression levels of mirnas in serum as sensitive biomarkers for early diagnosis of traumatic injury circulating mir-210 predicts survival in critically ill patients with acute kidney injury plasma mir-216a as a potential marker of pancreatic injury in a rat model of acute pancreatitis circulating micrornas in patients with chronic hepatitis c and non-alcoholic fatty liver disease microrna fingerprints identify mir-150 as a plasma prognostic marker in patients with sepsis expression of microrna-150 in peripheral blood leukocytes in sepsis patients and its clinical significance expression of microrna-143 in sepsis and its clinical significance levels of circulating mir-133a are elevated in sepsis and predict mortality in critically ill patients serum mir-574-5p: a prognostic predictor of sepsis patients serum mir-146a and mir-223 as potential new biomarkers for sepsis unique signatures of long noncoding rna expression in response to virus infection and altered innate immune signaling proteomic updates on sepsis proteomic study of peripheral blood lymphocytes of rabbits with severe burn and pseudomonas aeruginosa sepsis proteomic analysis of neutrophils of rats with acinetobacter baumannii sepsis: bengbu medical college ykl-40 identified by proteomic analysis as a biomarker of sepsis plasma proteome to look for diagnostic biomarkers of early bacterial sepsis after liver transplantation a metabonomic approach to early prognostic evaluation of experimental sepsis a metabolomic approach for diagnosis of experimental sepsis a multiplex real-time pcr assay for rapid detection and differentiation of 25 bacterial and fungal pathogens from whole blood samples multi-analyte network markers for tumor prognosis key: cord-312197-d5d8amk7 authors: edmond, karen; zaidi, anita title: new approaches to preventing, diagnosing, and treating neonatal sepsis date: 2010-03-09 journal: plos med doi: 10.1371/journal.pmed.1000213 sha: doc_id: 312197 cord_uid: d5d8amk7 karen edmond and anita zaidi highlight new approaches that could reduce the burden of neonatal sepsis worldwide. neonatal sepsis or septicaemia is a clinical syndrome characterized by systemic signs of circulatory compromise (e.g., poor peripheral perfusion, pallor, hypotonia, poor responsiveness) caused by invasion of the bloodstream by bacteria in the first month of life. in the pre-antibiotic era neonatal sepsis was usually fatal. case fatality rates in antibiotic treated infants now range between 5% and 60% with the highest rates reported from the lowestincome countries [1] . the world health organization (who) estimates that 1 million deaths per year (10% of all under-five mortality) are due to neonatal sepsis and that 42% of these deaths occur in the first week of life [2] . there are wide disparities in neonatal care between highand low-income countries. in high-income countries the major concern is the increasing numbers of extremely premature infants with high nosocomial infection rates due to multiresistant organisms in intensive care units. health facility infections are also a major problem in lowincome countries, but the more pressing issues are the high proportion of home deliveries in unclean environments predisposing to sepsis and ensuring that all neonates have access to effective interventions from health care providers in the first days of life 2 . indeed, new strategies that can prevent, diagnose, and treat neonates with sepsis are needed in both low-and high-income settings. distal risk factors for neonatal sepsis include poverty and poor environmental conditions. proximate factors include prolonged rupture of membranes, preterm labour, maternal pyrexia, unhygienic intrapartum and postnatal care, low birth weight, and prelacteal feeding of contaminated foods and fluids [3] [4] [5] . the bacteria that cause neonatal sepsis are acquired shortly before, during, and after delivery (figure 1 ). they can be obtained directly from mother's blood, skin, or vaginal tract before or during delivery or from the environment during and after delivery. streptococcus agalactiae (group b streptococcus, gbs) is the most common cause of neonatal sepsis in many countries, though low rates are reported from many low-income countries, especially those in south asia. [6] [7] [8] ; gramnegative bacilli (escherichia coli, klebsiella spp., pseudomonas spp., acinetobacter spp.) and gram-positive cocci (such as staphylococcus aureus and staphylococcus epidermidis) are other important causes [6] [7] [8] . however, there are many difficulties in interpreting aetiological neonatal sepsis data, because many studies report selected populations of high-risk infants. specimens from infants in the first 24 hours of life are also seriously under-represented, especially those from low birth-weight babies and babies born outside health facilities [6, [9] [10] [11] . intrapartum antibiotic prophylaxis against s. agalactiae has also led to a substantial change in the bacteria responsible for early onset neonatal sepsis; gramnegative bacilli and staphylococcus spp. predominate in countries implementing these programs [12] . there are also many other important neonatal infectious disease pathogens that are not associated with the sepsis syndrome including: treponema pallidum, rubella virus, herpes simplex virus, cytomegalovirus, toxoplasmosis, clostridium tetani, hiv, hepatitis b virus, and bordetella pertussis ( figure 1 ) [1, 7, 13] . these infectious pathogens cause serious morbidities in young infants and multifaceted disease syndromes including congenital anomalies, developmental disabilities, chronic liver disease, neonatal tetanus, and apnoea. they are also important causes of morbidity and mortality in older age groups. however, only pathogens that cause neonatal sepsis are discussed in this paper. neonates have a functionally immature immune system. they have extremely low immunoglobulin (ig) levels except for igg to specific maternal antigens transferred passively across the placenta during the last trimester of pregnancy [14, 15] . t cell function is relatively unimpaired but complement activity is half that of healthy adults. neonates have a low neutrophil storage pool, and their existing neutrophils have impaired capacity to migrate from the blood to sites of infection [16] . the basal expression of toll-like receptors (tlrs, receptors that detect the presence of microbes) is similar in the neonate and adult [17] . however, innate immune responses of neonatal mononuclear cells are characterised by markedly reduced release of the proinflammatory th1-polarizing cytokines tumour necrosis factor-alpha (tnf-a) and interferon-gamma (ifn-c) with relative preservation of anti-inflammatory th2-polarizing cytokines such as interleukin 6 (il6) [18] . these findings may reflect in utero requirements, including the avoidance of harmful inflammatory immune reactions [19] . these immunological problems are reflected in the clinical presentation of neonatal sepsis. neonates have a rapid and fulminant progression of septicaemic disease, nonspecific clinical signs of infection, and difficult-to-interpret laboratory results including haematological and immunological biomarkers of infection and inflammation. low birth-weight (preterm and small for gestational age) infants have even poorer functional immunity, and are especially at risk of sepsis [19] . however, neonates do have well-functioning cationic membrane-active antimicrobial proteins and peptides (apps) which have microbicidal properties [15, 19] . these apps can be found in the vernix caseosa covering the skin at birth, and in the neonatal gastrointestinal and respiratory tracts. many older studies have demonstrated that improving maternal health and nutri-tion before delivery is directly associated with improved neonatal health outcomes [3] . randomised controlled trials (rcts) of maternal protein-calorie and multiple micronutrient and supplementation have demonstrated significant improvements in rates of prematurity and birth weight and variable impact on mortality; but no studies have examined their impact on rates of neonatal sepsis [20, 21] . maternal immunisation is an important method of providing neonates with appropriate antibodies as soon as they are born [22] . this approach is less sensitive to obstacles in accessing the health care system than are other approaches, and examples of successful interventions include maternal tetanus toxoid and influenza immunisations [23, 24] . studies of maternal immunisation with s. agalactiae type iii conjugate vaccine have demonstrated excellent placental transfer and persistence of protective levels in 2month-old infants [22] . phase i and ii trials of other serotypes in nonpregnant women have also demonstrated safety and immunogenicity. a recent modelling study estimated that vaccination with s. agalactiae vaccine would prevent 4% of us preterm births and 60%-70% of neonatal s. agalactiae infections [25] . encouraging results are also emerging from studies of maternal immunisation with pneumococcal polysaccharide and conjugate vaccines [22, 26] . the vaccines all have excellent safety profiles. however, barriers to maternal immunisation include: liability issues for vaccine manufacturers in developed countries; education of the public and health care providers regarding the benefits of maternal immunisation; and poor ascertainment of data from lowincome countries [22] . there is strong evidence that clean delivery practices and handwashing during delivery reduces rates of neonatal sepsis in both home and health facility settings [27] [28] [29] . interventions to improve handwashing rates have been remarkably successful in research settings [30, 31] . the reasons for lack of successful scaleup of handwashing interventions into policy, programs, and behaviour change are less clear [32] . new studies from malawi and nepal indicate that maternal antisepsis interventions such as vaginal chlorhexidine during labour may have a significant impact on rates of neonatal mortality and sepsis in developing countries [33] . however, other studies from high-income countries have demonstrated little effect on rates of hiv or neonatal infections [34] . intrapartum antibiotic prophylaxis has been highly effective in reducing both early-onset neonatal bacterial and maternal sepsis in developed countries [35] . chemoprophylaxis in the us has halved the incidence of early-onset neonatal bacterial sepsis caused by s. agalactiae from 1.7 per 1,000 live births in 1993 to 0.6 per 1,000 in 1998 [36] . clear protocols are in place in high-income countries for the management of women with risk factors for neonatal sepsis [37] . risk factors for early-onset neonatal bacterial sepsis in low-income settings are probably similar to resource-rich settings, but have not been evaluated in the context of high rates of maternal undernutrition, anaemia, hiv, and malaria. there is also strong evidence that handwashing by health care providers after delivery can reduce neonatal sepsis and infection rates, especially in hospitals [27, 28] . there is less evidence for the importance of rigorous handwashing and use of antiseptics in mothers of their own infants. five key papers on preventing, diagnosing, and treating neonatal sepsis in high-income settings, studies have not shown an advantage of antibiotics or antiseptics over simply keeping the umbilical cord clean [2] . however, umbilical stump chlorhexidine cleansing has recently been shown to substantially reduce neonatal deaths in nepal [38] . other studies investigating the effects of chlorhexidine on prevention of omphalitis are currently underway in several countries [39] . there is emerging evidence that neonatal skin antisepsis preparations such as sunflower seed oil provides cheap, safe, and effective protection against nosocomial infections in hospitalized preterm neonates and infants in studies in south asia. application of chlorhexidine to neonatal skin has also been shown to be effective in reducing neonatal sepsis in studies from south asia [39, 40] . neonatal immunisation has long been considered an important method of reducing neonatal infections. however, response varies according to the antigen [15] . bcg, polio, and hepatitis b vaccines are highly immunogenic when given at birth [41] . however, maternal antibodies interfere with a neonate's response to measles vaccine when administered under six months. protein antigen vaccines (e.g., pertussis and tetanus toxoid) given at birth have been shown to produce poor responses compared to the same antigen given at two months of age and are associated with later tolerance [41] . studies also indicate that s. agalactiae and streptococcus pneumoniae vaccines are both likely to be ineffective when given in the neonatal period [15] . breastmilk contains secretory iga, lysozymes, white blood cells, and lactoferrin and has been shown to encourage the growth of healthy lactobacilli and reduce the growth of e. coli and other gramnegative pathogenic bacteria [15] . rcts that focused on increasing early initiation and exclusive breastfeeding rates demonstrated significant reductions in diarrhoea and acute respiratory infections in neonates and older infants in india [42] . other observational studies have demonstrated impact on infection specific mortality rates and all-cause mortality during the neonatal period [43] [44] [45] . neonatal micronutrient supplementation trials have focused on vitamin a supplementation. older studies have shown significant reductions in respiratory disease in low birth-weight infants after the administration of parenteral vitamin a [46] . more recently, trials of newborn vitamin a supplementation have shown encouraging reductions in neonatal mortality, and more trials are underway [47] . in high-income countries, clinical trials of immune stimulants such as granulocyte/ monocyte colony stimulating factor (gm-csf) to enhance the quantity and quality of neonatal neutrophils and monocytes appear promising but have not yet shown a significant clinical benefit [15] . the evaluation of recombinant apps as adjunctive therapy for neonatal infection are still under evaluation. the impact of tlr agonists to improve defences against microorganisms are also being evaluated [15] . neonatal clinical sepsis syndrome identification is difficult as the clinical signs of neonatal septicaemia can be very similar to those of other life-threatening diseases such as necrotising enterocolitis, hyaline membrane disease, and perinatal asphyxia [48, 49] . however, recent studies in middle-and low-income countries have provided seven danger signs which can be used to identify infants with very severe disease including neonatal sepsis (table 1 ) [49] . these signs provide high sensitivity and moderate specificity for detecting serious illness in newborns in low-resource settings and have now been incorporated into the new neonatal who integrated management of childhood illness (n-imci) guidelines. identification of neonatal sepsis before delivery also remains challenging. a combination of maternal risk factors and clinical signs and symptoms is currently used [50] . however, peripartum proteomic analysis of the amniotic fluid is now offering the opportunity for early and accurate diagnosis of early-onset neonatal sepsis in the select population of women undergoing amniocentesis in high-risk pregnancies [51, 52] . confirmation of pathogenic organisms allows targeted antibiotic therapy. however, identification of pathogenic organisms in neonates with sepsis syndrome is fraught with difficulties. bacterial load may be low due to mothers receiving antepartum or intrapartum antibiotics and because only small amounts of blood can often be taken from newborns [53] . contamination rates may also be very high due to the technical difficulties of sterile venipuncture in small babies. there may also be misinterpretation of the role of coagulase-negative staphylococci (e.g., s. epidermidis), as these organisms are both normal skin flora and pathogenic organisms in preterms and infants with indwelling blood vessel catheters [54] . automated blood culture systems have long been considered the gold standard for microbiological diagnosis. however, despite improvements in growth media and instrumentation, results of blood culture can be delayed by up to 48 hours [53, 55] . the condition of a neonate with true sepsis can deteriorate quickly, thus the most common approach is to initiate empiric broad-spectrum antibiotic therapy in all young infants with suspected bacterial infection [49] . a negative blood culture after 48 hours may allow cessation of antibiotic therapy in a well infant. while appropriately cautious, this practice leads to antibiotic exposure in a large number of newborns for whom antibiotic treatment may be unnecessary since blood cultures are positive in only 5%-10% of suspected sepsis cases, even at highly resourced facilities [56] . antigen detection techniques allow rapid detection and identification of microorganisms without culturing. the most commonly used commercially available test is the latex agglutination assay, which is based on specific agglutination by bacterial cell wall antigens of antibodycoated latex particles. however, these tests can only detect specific organisms such as s. agalactiae and are associated with high false positive and negative rates [57] . new urinary antigen tests for pneumococcus are more encouraging but are also associated with false positives from pneumococcal carriage [58] . the polymerase chain reaction (pcr) has been widely used in biomedical research laboratories for pathogen identification in neonatal sepsis and in some clinical hospital laboratories. the high sensitivity of pcr allows detection of bacterial dna even when concentrations are low [57] . conventional assays are being replaced by a newer ''real-time'' system, which is faster and associated with lower contamination rates because amplification and detection occur simultaneously in a closed system [59] . the real-time pcr is based on the measurement of a fluorescent signal generated during each amplification cycle. it produces quantitative results within 30 minutes and calculates bacterial load. broad-range real-time pcr uses a single primer to detect the universal bacterial genome (16s rna or 23s rna) which is a conserved ribosomal genome sequence across all bacterial genera [60] . broadrange real-time pcr can be used to distinguish bacterial septicaemic disease from other causes of neonatal illness such as asphyxia or complications of prematurity. however, it has been used with varying success in the analysis of whole blood for neonatal sepsis; specificity is generally high but sensitivity can be as low as 40% [60, 61] . in contrast, multiplex pcr involves the parallel amplification of different targets but is focused only on specific pathogens, and false negatives can occur if the aetiologic agent of interest is not included in the database [62] . real-time pcr is now often used to screen for microbial load, followed by sequence-based targeting and identification of pcr amplicons (pyrosequencing) [62] . this process can detect very small copy numbers of specific nucleic acid sequences. there is also a new commercially available multiplex pyrosequencing pcr assay which can identify up to 40 different bacterial and fungal pathogens directly from whole blood [63] . realtime pcr and pyrosequencing of the universal 23s rrna gene has also recently been used successfully in neonatal blood culture samples [64] . further tests on neonatal whole blood have been planned by a number of different research groups. the biggest problem with real time pcr testing is that the specimen must be collected with a sterile venipuncture, which may be difficult in young neonates. neonatal capillary heel prick specimens are easier to collect but highly contaminated by skin flora. there is also high potential for contamination of enrichment media, reagents, or the sample during collection and processing [61] other problems include low sensitivity due to competition from human dna in whole blood, especially if white cell counts are high. also, bacterial organisms require lysis before their dna can be available for analysis, and gram-positive organisms are difficult to lyse because of their resilient cell wall [61] . real-time pcr technologies are also expensive and currently can be used only by highly trained staff. important haematological tests include microscopic examination of the blood for white cells (total leucocyte count, differential, neutrophil count, and immature neutrophil to total neutrophil ratio). advantages are that these specimens do not require sterility and a heel prick specimen can be used. however many of these indices are falsely low in a septic neonate. biological biomarkers are human blood components that increase in response to infection. the most commonly used acute phase reactant is the c-reactive protein (crp). however, the crp takes 12-24 hours to increase to measurable levels; its half life is very long and it takes 5-7 days to normalize after eradication of the infectious agent. cytokines such as il6, il8, tnf-a, and procalcitonin have also been extensively studied [65, 66] . cytokines rise quickly after infection even in neonates, and are more sensitive to low concentrations of pathogens than crp [66] . however, cord and postnatal blood cytokine concentrations can be depressed in the presence of pregnancy-induced hypertension and can rise after induced vaginal or urgent cesarean delivery, delivery room intubation, muscular damage, and inflammation from other causes [57] . simultaneous measurement of multiple biomarkers may improve both sensitivity and specificity [66, 67] . however, biomarker assays are likely to be less acceptable to physicians who often place higher value on tests that confirm biological agents and allow targeting of antibiotic therapy [57] . microtechnologies, especially microfluidics, have provided the greatest recent contribution to the diagnosis of neonatal sepsis. microfluidics is the study of the behaviour, precise control, and manipulation of fluids geometrically constrained to submillimetre (nanolitre or picolitre) channels [68] . microfluidic technology uses the unique proprieties of continuous flow micro-volume channels: viscosity, surface tension, energy dissipation, and fluidic resistance, and also includes micro pneumatic pump and valve systems. one specific application of microfluidics is bacterial dna protein microarray hybridization [69] . in this test, dna probes specific to selected targets are spotted on a glass or silicon slide in a known order. target dna fragments are labelled with a reporter molecule, combined into a single hybrid, and measured using fluorescent signals [62, 68] . this technique has been used in the identification of the specific sepsis pathogen in bacterial meningitis, acute viral respiratory tract infections, and neonatal sepsis, and also in the detection of their antimicrobial resistance and virulence genes in research settings [63] . microfluidic technology has also allowed sample preparation and a number of different assays to be combined in small, disposable, single-use diagnostic cartridges or cards that have been called a ''lab on-achip'' or loc (figure 2 ) [68] . some locs have combined sample preparation, biomarkers, real-time pcr, and dna microarrays to provide information about indices of inflammation, pathogen identification, and antimicrobial susceptibility patterns at the point of care [68, 70] . locs have been reported to perform assays at sensitivity, specificity, and reproducibility levels similar to those of central laboratory analysers, but yet require little user input other than the insertion of the sample. single drops of blood, faeces, and saliva have all been tested with encouraging results. locs are currently being evaluated for use in sepsis, endocarditis, hiv, tuberculosis, severe acute respiratory syndrome (sars), and pneumonia [68] . however, they are not yet in clinical use nor licensed by regulatory authorities. as neonatal sepsis can be rapidly fatal if left untreated, highly effective antibiotic therapy must be used and delays in the provision of care must be minimised. treatment must be effective against the causative pathogen, safe for the newborn, and feasible to deliver reliably in the hospital or community setting. parenteral (intravenous or intramuscular) regimens for neonatal sepsis currently recommended by national paediatric associations are a combination of penicillin/ ampicillin and gentamicin, or third-generation cephalosporins (e.g., ceftriaxone or cefotaxime) for 10-14 days. these antibiotics are safe and retain efficacy when administered at extended intervals (e.g., twice daily or daily dosing) [56] . these regimens are very effective against streptococcus spp., but staphylococcus spp. can be highly resistant [71] . gram-negative antimicrobial susceptibility to ampicillin and gentamicin can also be poor, especially for klebsiella spp. [8, 71] . emerging e. coli resistance to ampicillin, gentamicin, and third-generation cephalosporins in hospital nurseries in both developed and developing countries is also causing increasing concern [8] . the potential for significant life-threatening toxicity among neonates associated with chloramphenicol makes it the least preferred empiric parenteral therapy [56] . oral antibiotic therapy must be considered in settings where referral is not possible and there are no health care providers trained to give parenteral antibiotics [72] . the incremental benefit of injectable over oral antibiotics is not known, and oral antibiotic therapy is better than no antibiotic therapy at all. a series of trials are currently evaluating the impact of home and clinic-based short course (7 days) intramuscular and oral antibiotic therapy for neonatal sepsis in low-income countries [72] . most data are available on the effect of oral cotrimoxazole in community-based treatment of serious neonatal bacterial infections from nepal and india. however, there are concerns about high resistance rates, and side effects such as neonatal jaundice have been reported [71] . oral amoxicillin is highly efficacious against streptococcus spp. and some gram-negative bacilli and has an excellent safety record. however, it has no anti-staphylococcus coverage and resistance is emerging in gram-negative bacilli such as e. coli. new, better-absorbed oral antibiotics are also being considered. the new second-generation cephalosporins (e.g., cefadroxil and cefuroxime) have an excellent safety profile, a spectrum of activity similar to cotrimoxazole, and may be more effective given the high resistance of neonatal pathogens to cotrimoxazole. ciprofloxacin also is increasingly accepted as safe in neonates and warrants further investigation for treatment of infections in newborns. however, the current cost of these agents and potential for exacerbating antimicrobial resistance may limit widespread use in developing countries [72] . poor maternal-neonatal health systems, low levels of care-seeking, and lack of access to sick newborns during the first day of life, when mortality risks are highest, are also important concerns [73] . recent studies have shown that community health workers can deliver antibiotic treatment to neonates with very severe infections at home safely and acceptably when hospitalization is not feasible [74] . trials are currently evaluating the effectiveness, quality of care, and coverage of these community health worker programmes in asia and africa [73] . barriers to large-scale implementation include high cost, poor staff training and retention, and difficulties with referral (e.g., lack of ambulances and poor institutional links). newborn sepsis is a major cause of child mortality across the world. industrialized countries have made remarkable progress in reducing newborn sepsis and sepsisrelated mortality by providing access to hygienic skilled delivery for all women, risk-based intrapartum antibiotic prophylaxis, and high-quality intensive care for newborns that need it. although resource constraints preclude whole-scale adoption of these strategies in developing countries, there are a number of low-cost proven interventions and promising approaches that have the potential to significantly reduce the burden of neonatal sepsis worldwide (table 2) . however, practicability of implementing these new advances must be considered. skilled attendance at delivery is increasing in low-and middle-income countries. thus, intrapartum approaches such as risk-based antibiotic prophylaxis and improved hand washing during delivery are likely to be both cost-effective and feasible in these countries. more challenges face the implementation of diagnostic technologies. it may take many years for technologies such as the ''lab-on-a-chip'' to be sufficiently robust and affordable for scale-up to low-income countries. homebased antibiotic treatment of neonatal sepsis also faces major obstacles to largescale implementation. concerns such as ''one law for the rich and another for the poor'' have already been raised. a careful assessment of the risks and benefits of new technologies and interventions is clearly needed. in low-income settings there are also difficulties with care-seeking for neonatal illnesses, and home visiting programs are needed to identify sick newborns early in life. neonatal sepsis is also one of the most rapidly fulminating clinical diseases, and many practitioners, including experi-enced neonatologists, administer parenteral antibiotics rather than wait for the results of any diagnostic tests. these practitioners rightly consider that the individual patient's health is more important than the potential risks of emerging antibiotic resistance. thus, front-line health workers and families must be partners in all research and evaluation planning. detailed assessment of end-user attitudes and preferences using formative and qualitative research methods must be included in the development of programs to reduce morbidity and mortality from neonatal sepsis. finally, advocacy for equitable resource allocation across and within countries must be a priority and modelling techniques to assess public health impact of neonatal sepsis interventions must be developed and used more widely. icmje criteria for authorship read and met: ke az. agree with the manuscript's results and conclusions: ke az. designed the experiments/the study: ke az. analyzed the data: ke az. collected data/did experiments for the study: ke az. wrote the first draft of the paper: ke. contributed to the writing of the paper: ke az. burden of neonatal infections in developing countries: a review of evidence from community-based studies million neonatal deaths: when? where? why? evidencebased, cost-effective interventions: how many newborn babies can we save? research priorities to reduce global mortality from newborn infections by 2015 risk 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markers of bacterial infections predictive value of soluble immunological mediators in neonatal infection microfluidic diagnostic technologies for global public health microfluidic dna amplification-a review enabling a microfluidic immunoassay for the developing world by integration of on-card dry reagent storage antimicrobial resistance among neonatal pathogens in developing countries oral antibiotics in the management of serious neonatal bacterial infections in developing country communities management of newborn infections in primary care settings: a review of the evidence and implications for policy? effect of home-based neonatal care and management of sepsis on neonatal mortality: field trial in rural india key: cord-252859-zir02q69 authors: chung, t. philip; laramie, jason m.; meyer, donald j.; downey, thomas; tam, laurence h.y.; ding, huashi; buchman, timothy g.; karl, irene; stormo, gary d.; hotchkiss, richard s.; cobb, j. perren title: molecular diagnostics in sepsis: from bedside to bench date: 2006-09-11 journal: j am coll surg doi: 10.1016/j.jamcollsurg.2006.06.028 sha: doc_id: 252859 cord_uid: zir02q69 background: based on recent in vitro data, we tested the hypothesis that microarray expression profiles can be used to diagnose sepsis, distinguishing in vivo between sterile and infectious causes of systemic inflammation. study design: exploratory studies were conducted using spleens from septic patients and from mice with abdominal sepsis. seven patients with sepsis after injury were identified retrospectively and compared with six injured patients. c57bl/6 male mice were subjected to cecal ligation and puncture, or to ip lipopolysaccharide. control mice had sham laparotomy or injection of ip saline, respectively. a sepsis classification model was created and tested on blood samples from septic mice. results: accuracy of sepsis prediction was obtained using cross-validation of gene expression data from 12 human spleen samples and from 16 mouse spleen samples. for blood studies, classifiers were constructed using data from a training data set of 26 microarrays. the error rate of the classifiers was estimated on seven de-identified microarrays, and then on a subsequent cross-validation for all 33 blood microarrays. estimates of classification accuracy of sepsis in human spleen were 67.1%; in mouse spleen, 96%; and in mouse blood, 94.4% (all estimates were based on nested cross-validation). lists of genes with substantial changes in expression between study and control groups were used to identify nine mouse common inflammatory response genes, six of which were mapped into a single pathway using contemporary pathway analysis tools. conclusions: sepsis induces changes in mouse leukocyte gene expression that can be used to diagnose sepsis apart from systemic inflammation. the incidence of sepsis and the number of sepsisrelated deaths is increasing at substantial cost ($17 billion annually in the us), 1 as reported recently. the cornerstone of successful therapy is early, accurate diagnosis, coupled with eradication of the source of infection and appropriate antibiotic therapy. 2 unfortunately, efforts to identify specific and sensitive diagnostic markers for sepsis have met with limited success. 3 the diagnosis of sepsis in icus is especially challenging because it is frequently difficult to distinguish between systemic inflammation and systemic infection. clinicians are unable to identify the pathogen responsible for sepsis in up to 50% of patients or to determine whether patients are responding to antibiotic therapy 1 ; and the traditional means of identifying the organism responsible for bacterial infections are nonspecific (gram stain), slow (culture), or insensitive (both gram stain and culture). there is compelling evidence that efforts to identify clinically important biomarkers in sepsis will prove successful ultimately, 4-6 especially given the recent successes of high-throughput, genomic technologies in other fields that face diagnostic challenges (eg, leukemia, 7 breast 8, 9 and colon cancer 10 ). this molecular classification strategy involves searching for expression patterns in a subset of genes from tissues of known phenotype (the "training" data set), constructing a prediction rule, and then using these "biomarker" genes to predict the phenotype of new samples (the "test" data set). studies that apply these genome-wide technologies to inflammation and sepsis in animal models and patients are now underway, [11] [12] [13] [14] as reviewed recently. 15, 16 these reports suggest that genome-wide profiling of gene expression holds promise as a molecular diagnostic tool, capable of generating profiles from leukocytes that are sensitive, specific, and timely for pathogen detection. 13 despite provocative in vitro findings, 4, 5 there are few reports of using microarrays to study sepsis in vivo. these reports indicate that the transcriptional response during polymicrobial sepsis is organ-specific in mice 12 and rats. 11 there are no reports of using microarrays to make a diagnosis of sepsis, although very recent reports suggest that this will be feasible in patients. 17, 18 we hypothesized that leukocyte gene expression profiles obtained using dna microarrays could be used to predict septic states; in particular, distinguishing between sterile and infectious sources of systemic inflammation, a common conundrum in caring for the critically ill or injured. tissue samples from both septic patients and clinically relevant mouse models of sepsis were tested. using an investigational protocol approved by the washington university human studies committee, informed consent was obtained to collect samples of splenic tissue intraoperatively or immediately postmor-tem as described previously. 19 seven specimens for expression profiling were chosen retrospectively from patients with injury (trauma or operation) complicated by sepsis and organ dysfunction (sepsis group). these were compared with six age-and gender-matched control specimens from patients with injury (trauma) requiring splenectomy (injury group). a total of 13 patient spleen samples were collected. care and use of laboratory animals were conducted in accordance with a protocol approved by the washington university animal studies committee. seven-to nineweek-old, male c57bl/6 mice were purchased (harlan, inc) and allowed to acclimatize for at least 1 week. male mice were used to avoid the confounding effect of the female estrous cycle and the well-characterized sexual dimorphism of the adaptive response to cecal ligation and puncture (clp). 20 the seven experimental groups were designed to make classification difficult, reflecting clinically important distinctions relevant to care of icu patients: lethal abdominal inflammation from a sterile source, lethal abdominal infection, and antibiotictreated abdominal infection. mice were assigned to the seven treatments listed in table 1 , grouped into those with no deaths (controls) and those that were "sick" with substantial associated mortality (sterile or infectious causes of systemic inflammation). normal animals were untreated. previously reported protocols were used to perform clp and sham laparotomy. 21, 22 the ceca of some animals were punctured twice using a 25-gauge needle (clp), and others were punctured using a 27gauge needle to produce a "milder" sepsis that caused much lower mortality (mild clp). animals that had laparotomy and cecal manipulation without ligation or puncture were included in the sham laparotomy group (sham). another group of animals had clp and treatment with a standard antibiotic regimen for mouse abdominal sepsis: ip ceftriaxone 6.4 mg/kg and metronidazole 3.5 mg/kg, delivered 1, 12, and 24 hours after clp (clp ϩ antibiotics group). 23 to induce severe systemic inflammation without infection, ip injection of escherichia coli lipopolysaccharide (lps) serotype 0111:b4 20 mg/kg (sigma, inc) was performed (lps group). 24 the dose of lps (20 mg/kg) was used because it reliably produces death over several days in the animals; smaller doses in pilot experiments tended to produce morbidity without mortality. mice injected ip with abbreviations and acronyms clp ϭ cecal ligation and puncture fdr ϭ false discovery rate est ϭ expressed sequence tag ip ϭ intraperitoneal lps ϭ lipopolysaccharide pca ϭ principal components analysis normal saline vehicle (saline group) acted as the concurrent control group for lps treatment. the census of surviving mice was recorded at 24-hour intervals for 7 days. in a separate cohort, mice were sacrificed at 24 hours after injury by cervical dislocation under halothane general anesthesia. the 24-hours time point after injury was chosen because pilot experiments showed that the ability to distinguish between spleen expression profiles was greater at 24 hours than at 48 or 72 hours after injury. 25 whole spleen tissue from eight clp and eight sham mice was harvested and flash-frozen in liquid nitrogen and stored at ϫ80°c. in another cohort of animals, intracardiac blood from eight animals per group was pooled using the paxgene blood rna system to derive total rna from whole blood (preanalytix gmbh). while 5 genechips from blood for each of the seven groups were prepared, there were a total of 35 mouse blood genechips from 280 animals: 28 gene-chips were in the training data set and 7 genechips were in the test (de-identified) data set. whole blood was collected for automated white blood cell counts from eight animals in each group over two replicates, each run performed with samples from concurrent control animals. a hemavet multispecies hematology analyzer (cdc technologies) provided counts and an automated differential. data were expressed as the mean ϯ sem. total rna from human and mouse spleens was extracted per trizol protocol (life technologies, inc). total rna from mouse blood was extracted using the paxgene kit as recommended by the manufacturer. crna target for genechip hybridization was prepared from total rna using the protocol recommended (affymetrix, inc). the human spleen crna was hybridized with the human full-length genechip (approximately 7,000 probe sets). mouse spleen and blood crna were hybridized with the u74av2 genechip (approximately 12,400 probe sets). fluorescent hybridization signal was detected using a genearray scanner (hewlett-packard, inc) at the washington university alvin j siteman cancer center and, for the prospective mouse cohort, the genechip scanner 3000 (affymetrix). the data discussed in this publication have been deposited in ncbi's gene expression omnibus (geo, http://www.ncbi.nlm.nih.gov/ geo/) and are accessible through geo series accession number gse5663. tests for differential expression, class prediction, and pathway analysis expression values were calculated from genechip.cel files using dna chip analyzer software (version 1.3) and default settings (only perfect match probes were used). 26 principal components analysis (pca) was used to visually explore global effects for genome-wide trends, unexpected effects, and outliers in the expression data (partek software). differentially expressed genes were identified using a mixed-model anova and linear contrasts. we next determined whether it was possible to accurately distinguish tissue samples resulting from models of sterile systemic inflammation or models of systemic infection in human spleen, mouse spleen, and mouse blood. to produce unbiased estimates of prediction accuracy while also optimizing the set of predictor genes, classifier type, and classifier parameters, we used a two-level, nested cross-validation procedure that produces prediction estimates that are not biased by gene, classifier, and parameter selection. this procedure makes use of an "outer" cross-validation to produce the estimate of accuracy, and an "inner" cross-validation to perform classifier and gene selection. 27 for class prediction, we compared k-nearest neighbor, nearest centroid, and linear and quadratic discriminant analysis classifiers. two complementary programs were used to query regulatory networks: pathwayassist (ariadne genomics) and pathway analysis (ingenuity systems). both path-wayassist and pathway analysis use the published literature or publicly available databases to identify relationships between genes, small molecules, or other objects. this information in turn is used to map de novo, putative, interaction networks from a given list of input gene survival data were calculated using the kaplan-meier method and survival curves were compared using logrank test (prism v3.03, graphpad software, inc). cbc and cell differential data were analyzed by one-way anova using the tukey-kramer post-test to correct for multiple comparisons (prism v3.03). modified multiple organ dysfunction syndrome (mods) score 28 are reported as the mean ϯ sem and analyzed using the mann-whitney u test for nonparametric data (prism v3.03). the significance of change in mouse blood relative rna abundance was measured across all seven groups using two-way anova for the effects of treatment and batch (time). from this analysis, a step-up false discovery rate (fdr) of 0.1 was used to identify a subset of discriminating genes for treatment effect, visualized using pca. 29 for pair-wise group comparisons across mouse blood, a two-way anova for the effects of treatment and batch was used. because there was no batch effect for spleen, one-way anova was used. an fdr of 0.1 was applied to the raw p value for each pair-wise comparison, giving a list of informational genes for each comparison. the characteristics of the 13 patients sampled are found in table 2 . none of the injury (control) patients had positive cultures or signs of infection or sepsis at the time of sample collection, and all but one recovered uneventfully after splenectomy. in contrast, all sepsis patients had positive bacterial or fungal cultures or obvious signs of infection (pus or necrotizing fasciitis) complicated by sepsis-induced organ dysfunction at the time of operation (modified mods score, p ϭ 0.001 versus injury). dna chip analyzer hybridization signal analysis using default filters flagged 1 of 13 human genechips as a statistical outlier (ͼ 15% probe pair expression values calculated as statistical outliers, suggesting unreliable hybridization signal). this sample (patient 7) was omitted from additional analysis. pca was used to explore differences in gene expression for all genes across the remaining 12 microarrays, demonstrating considerable differences between the sepsis and injury classes (data not shown). consistent with the large variance in expression observed across subjects, there were no genes identified as differentially expressed between these two classes using a fdr of 0.1. to estimate sepsis class prediction accuracy among these samples, a 12 ϫ 11 leaveone-sample-out nested cross-validation was performed. the average predictive accuracy was 67.1%. all control group animals (saline, sham, normal) survived the duration of the experiment. in contrast, "sick" animals with systemic infection (clp, clp ϩ antibiotics, mild clp, and lps) exhibited typical signs of piloerection, anorexia, and lethargy followed by 7-day mortality rates of 78%, 25%, 44%, and 100%, respectively ( fig. 1) . absolute white blood cell counts varied among the seven experimental groups (p ͻ 0.0001), in particular between control and sick animals (fig. 2) . there was no difference among animals with sterile or infectious causes of systemic inflammation (lps, clp, mild clp, or clp ϩ antibiotics). likewise, there was no difference in cell differentials among controls or among animals with sterile or infectious causes of systemic inflammation, except for lps versus clp ϩ antibiotics (neutrophils and lymphocytes, p ͻ 0.05). mouse differential gene expression dna chip analyzer hybridization signal analysis flagged 2 of the 35 mouse blood genechips as statistical outliers: one saline and one sham genechip from the training data set. these mouse outliers genechips were omitted from additional analysis. thirty-three gene-chips remained in the mouse blood data set. pca was used to visualize treatment differences in expression for all genes in blood and spleen, demonstrating batch and any-clp treatment effects (p ͻ 0.05, fig. 3 ). we applied a stringent multiple test correction (bonferroni, 0.05) to the p values from the two-way anova to identify a small set of genes in blood differentially expressed between the seven treatment groups (25 probe sets corresponding to 24 genes, as probe sets for lipocalin 2 appeared twice [ table 3 ]). 30 pca analysis of these 25 probe sets revealed that the seven experimental groups were clustered into three apparent phenotypes (fig. 4) : control animals, lps-treated animals (sterile source of systemic inflammation), and those that had any clp treatment (sepsis). comparisons of gene expression across groups generated several informational gene lists (fdr ϭ 0.1); each indicated apparent increases and decreases in gene expression induced by clp or lps (fig. 5 ). using mouse spleen samples for gene expression analysis, we were able to classify the samples as clp or sham with 96.0% accuracy, estimated using cross-validation. for the blood data, the experimental design dictated that the first four replicates (batches) were used to train the classifiers. the best classifiers were trained on all 26 training samples (batches 1 to 4) and used to predict the seven de-identified test samples (batch 5). all seven test samples were predicted correctly as any-clp (septic) versus non-clp. we also performed fivefold leave-onebatch-out cross-validation, which produced an overall accuracy estimate of 94.4% (fig. 6) . the prediction accuracy differentiating lps from controls was 93.2%. the ability to predict low versus high mortality after clp was substantially lower (62.4%). to obtain the final mouse blood classifier, leave-one-batch-out crossvalidation was performed for the purpose of classifier selection. sixty-four classifiers tied for best prediction. the median number of genes for these 64 classifiers was 450. of these 450 genes, only 61 genes showed changes greater than twofold, and the majority (86.4%) were altered by twofold or less. there were nine genes that demonstrated increased rna abundance across spleen and blood and across clp and lps (fig. 5) . we call this cluster of genes the "common inflammatory response cluster" (table 4 ). pathway analysis tools also were used to put this list into a biologic (functional) context. all but one of these genes is annotated and has been associated previously with a gene product, small molecule, or cellular process linked to inflammation (fig. 7) . a single expressed sequence tag (est) completed the list of nine genes (genechip identifier, 99849_at). a blast search identified this sequence as retroviral. a search of the ncbi gene expression omnibus microarray database showed that rna for this affymetrix probe set is increased in a number of models of inflammation, both animal and human. the ability to diagnose sepsis more accurately would allow appropriate treatment to be instituted earlier, thereby improving the likelihood of survival. 31, 32 we hypothesized that expression profiles could distinguish between septic and nonseptic states in vivo, and that expression profiles could define lists of common responder genes using a systematic, unbiased approach. 15,16 recent , saline, sham, and normal) is apparent, but less notable than the batch effect (appearing on pc3 and pc12, explaining 10% ϩ 2% of variance, respectively). no pcs besides pc3 and pc12 showed a substantial difference between any-clp and non-clp groups (determined by t-test on each pc). note that the genechip scanner used for batches 1 to 4 was different than that used for batch 5 (latest generation), which is one explanation why batch 5 is most different from the other four batches. reports using microarray technology in vitro indicated that inflammatory and infectious insults produce distinct transcriptional signatures. 4, 5 the current study is the first examining the ability of microarray gene expression profiles to distinguish sterile from infectious causes of systemic inflammation in vivo. we examined gene expression profiles of splenic tissue from patients with injury versus those with injury complicated by sepsis and organ dysfunction. differences in apparent gene expression between the sepsis and injury (control) phenotypes were used to construct a classifier, the accuracy estimate of which was 67.1%. this small, exploratory clinical study provided "bedside" proof of feasibility using human transcriptional profiles to model the septic phenotype, but also demonstrated a large degree of variance in gene expression between subjects, because of both technical and biologic differences. to control more of this variance we moved from the bedside to the bench, performing a systematic examination of the diagnostic potential of spleen and blood gene expression profiling in inbred mice. consistent with the human data, spleen samples from mice after clp exhibited microarray patterns that could be modeled to predict the septic phenotype. the nested cross-validation accuracy estimate of 96.0% for sepsis prediction using mouse spleen was considerably better than that found using human spleen, likely because of the mouse experimental design that exploited fresh tissues and identical age, gender, and genotype across subjects. because the clinical use of gene expression analysis using splenic tissue is severely limited, we explored next the use of circulating blood for class prediction in our mouse models. the combined accuracy of the predictions for any-clp versus non-clp and lps versus controls was high at 94.4% and 93.2%, respectively. the accuracy estimate for distinguishing between the high and low mortality clp groups (clp versus mild clp and clp plus antibiotics) at this 24-hour time point was much less at 62.4%. these conclusions are consistent with the pca analysis in figure 4 , in which we sought differences in apparent table 3 . abx, antibiotics. gene expression across the seven experimental groups. interestingly, the only samples that were cluster outliers were in the clp plus antibiotics group, consistent with an effect of antibiotics to change the septic phenotype toward the control phenotypes. we conclude that circulating blood gene expression profiles can be used to predict clp and non-clp phenotypes in prospective cohorts, in particular, distinguishing controls from lethal endotoxicosis (lps) from lethal infection (clp). it is important to note that at this 24-hour time point, there were no substantial differences between lps or any of the clp groups in clinical presentation or complete blood counts. microarray analysis could make the diagnosis of sepsis (distinguishing between sterile and infectious sources of systemic inflammation) when clinical criteria and white blood cell counts could not, a frequent occurrence in icus. 33 in addition, the differences between groups as measured by absolute fold-changes in individual gene expression were small (eg, in the final model for blood, 86.4% of genes were altered less than twofold), yet the changes in the patterns of expression across hundreds of genes were robust predictors of phenotype. to discover changes in expression generic to the inflammatory and septic responses, we used the intersection of gene lists identified by the pair-wise group comparisons (fig. 5) . nine genes were commonly increased, validated across two tissue types (spleen and blood) and two insults (clp and lps). in contrast, there were no genes that were commonly decreased. given that this list of nine genes was based on changes in relative rna abundance across a number of cell types, the network analysis performed served as an exploratory tool, validating in silico the role of six of nine genes in canonical pathways for inflammation, apoptosis, and signal transduction: inhibitor of dna binding 2, calgranulin a and b, interferon regulatory factor 7, lipocalin 2, and formyl peptide receptor-like 1 ( table 4) . several characteristics of these nine common inflammatory response genes are notable. inhibitor of dna binding 2 is required for normal mouse immune development, especially of lymph nodes and peyer's patches. 34 calgranulin a and b, which belong to a recently described group of proinflammatory molecules, form extracellular complexes that bind to and activate endothelial cells, promoting chemotaxis and phagocytic adhesion in a positive feedback manner. 35, 36 interferon regulatory factor 7 is a key regulatory of monocyte development, essential to differentiation of monocytes to macrophages. 37 lipocalin 2 is a secreted protein that undergoes transcriptional induction after cytokine withdrawal and induces leukocyte-specific apoptosis. 38 lipocalin 2 transcription, translation, and secretion are induced by ligation of toll-like receptors on leukocytes, with secreted lipocalin 2 acting to sequester siderophores, thereby limiting bacterial growth. 39 formyl peptide receptor-like 1 is a member of the chemoattractant subfamily of g proteincoupled receptors that are involved in controlling leukocyte migration. 40 the other two annotated genes not listed in the network, neutrophilic granule protein and serum amyloid a3, have also been associated with inflammation and cellular defense, although less is known about their functions and protein interactions. neutrophilic granule protein is a cysteine protease inhibitor that has been associated with myeloid differentiation. 41 serum amyloid a3 is a highdensity apolipoprotein, the only amyloid made by both hepatocytes and peripheral monocytes and macrophages. 42 it is believed to function by retargeting transported lipids, including cholesterol, in the disposal of toxins. 42 the function of the ninth gene, a retroviral species that is increased in a number of different models of infection and inflammation, is not known. we compared this common inflammatory response cluster with the list of proteins recently reported to diagnose intra-amniotic infection in patients, 18 as another means of validating the importance of these nine genes. of the 11 proteins and polypeptides detected in that study, 3 were also identified in our study, specifically calgranulin a and b and lipocalin 2 (neutrophil gelatinase-associated lipocalin). calgranulin a (s100a8) was also the most differentially expressed gene in blood in a small microarray study comparing eight septic patients with four surgical controls with-out systemic inflammation. 17 serum amyloid a protein was identified as a plasma proteome biomarker in patients with coronavirus (severe ards). 43 together, these studies support our hypothesis that there are a group of common inflammatory response genes that can be used as novel biomarkers to diagnose inflammation across species, tissue, and different types of infecting organisms, at either the rna or protein level. because of the large degree of variation (noise) in human spleen gene expression, no individual genes surveyed passed the fdr filter. nevertheless, the data from these patients have proved invaluable for the study of immune dysfunction in human sepsis 2 and provided proof-ofprinciple here that molecular profiles of human lymphoid tissues could be used to distinguish between septic and nonseptic phenotypes. because cellular populations of mammalian tissues are heterogeneous, use of microarray profiles to study the cellular response to a given stimulus must be understood in the context of changes in cell populations. this substantially limits conclusions about whole spleen data, given our reports and those of others that sepsis accelerates splenocyte apoptosis. 2, 19, 44 in contrast, changes in blood cellular heterogeneity are measured routinely. although clp and lps stimuli changed both absolute wbcs and cell differentials compared with the controls, among the four groups of "sick" animals cell counts were indistinguishable (fig. 2) . this mirrors the clinical situation where differentiating between sterile (lps) and infectious (any-clp) sources of systemic inflammation is not possible based on clinical grounds or cell counts. microarray profiles, as we have discussed, were very successful at making this distinction. many questions remain unanswered. what are the optimal computational methods to identify robust predictors from microarray or proteomic data? can gene or protein expression profiles be used to diagnose sepsis in other animal models? if so, are there predictive gene sets that are common to different types of infection (eg, gram-positive versus gram-negative infections)? once the diagnosis has been made, are there markers that indicate response to therapy or prognosis? there are sufficient preclinical and preliminary patient data to justify testing these hypotheses. because of the heterogeneity of expression profiles, large-scale collaborative studies will be required to enroll sufficient patients to identify robust sepsis markers, and in the process, untangle the biology of infection from inflammation. 13 back to the bedside-the promise of molecular profiling for sepsis diagnosis in conclusion, our in vivo data corroborate in vitro findings indicating that microarray analysis holds promise as a means of identifying distinct expression profiles ("molecular fingerprints") that can diagnose the septic phenotype. our human spleen data join recent blood data from septic patients 17 and serum and amniotic fluid data from patients with intra-amniotic fluid infection, 18 indicating that transcriptome and proteome studies will deliver on the promise of novel inflammation diagnostics. 15, 16 a single inflammation gene, calgranulin a (s100a8), was detected in all three studies at either the rna or protein level. our data are unique in that they show that transcriptome molecular profiles can distinguish between sterile and infectious causes of systemic inflammation and can make a diagnosis of sepsis in prospective cohorts. importantly, we observed that the magnitude of change in gene expression that was needed to predict the septic phenotype was very small. it was the pattern of these small changes in expression that were predictive, not the magnitude of any single change. we and others reported recently validated clinical protocols for blood gene expression profiling used to characterize the human systemic inflammatory response. 13, 14 the data presented here suggest that these protocols should be extended to clinical trials, testing the efficacy of microarray gene expression profiling to diagnose human sepsis. we expect that these studies will provide new insight into how specific pathogens uniquely perturb the physiology of cir-culating leukocytes and how the host successfully mounts pathogen-specific defenses. . mouse common inflammatory response cluster. nine probe sets (red genes) were commonly altered regardless of tissue or insult. a contemporary pathway analysis tool was used to automatically create a network of interactions among these nine genes, based on previously reported interactions in the literature. eight of the nine probe sets are known inflammation genes; the ninth probe set is an expressed sequence tag. this network validates in silico that six of these genes are involved in canonical pathways of inflammation, apoptosis, and regulation of signal transduction. the epidemiology of sepsis in the united states from 1979 through 2000 the pathophysiology and treatment of sepsis sccm/esicm/ accp/ats/sis international sepsis definitions conference the plasticity of dendritic cell responses to pathogens and their components human macrophage activation programs induced by bacterial pathogens measures, markers, and mediators: toward a staging system for clinical sepsis. a report of the fifth toronto sepsis roundtable molecular classification of cancer: class discovery and class prediction by gene expression monitoring molecular portraits of human breast tumours gene expression profiles in hereditary breast cancer broad patterns of gene expression revealed by clustering analysis of tumor and normal colon tissues probed by oligonucleotide arrays molecular signatures of sepsis: multiorgan gene expression profiles of systemic inflammation sepsis gene expression profiling: murine splenic compared to hepatic responses determined using cdna microarrays application of genome-wide expression analysis to human health and disease a network-based analysis of systemic inflammation in humans functional genomics of critical illness and injury injury research in the genomic era expression profiling: toward an application in sepsis diagnostics diagnosis of intraamniotic infection by proteomic profiling and identification of novel biomarkers apoptotic cell death in patients with sepsis, shock, and multiple organ dysfunction effect of gender and sex hormones on immune responses following shock evaluation of factors affecting mortality rate after sepsis in a murine cecal ligation and puncture model inducible nitric oxide synthase (inos) gene deficiency increases the mortality of sepsis in mice overexpression of bcl-2 in transgenic mice decreases apoptosis and improves survival in sepsis sequence makes a difference: paradoxical effects of stress in vivo physiological genomics model-based analysis of oligonucleotide arrays: expression index computation and outlier detection primary and secondary transcriptional effects in the developing human down syndrome brain and heart multiple organ dysfunction score: a reliable descriptor of a complex clinical outcome controlling the false discovery rate: a practical and powerful approach to multiple testing simultaneous statistical inference treating sepsis reducing mortality in sepsis: new directions determination of infection probability versus the diagnosis and treatment of antibiotic-responsive diseases development of peripheral lymphoid organs and natural killer cells depends on the helix-loop-helix inhibitor id2 s100a8: emerging functions and regulation phagocyte-specific s100 proteins: a novel group of proinflammatory molecules monocyte differentiation to macrophage requires interferon regulatory factor 7 induction of apoptosis by a secreted lipocalin that is transcriptionally regulated by il-3 deprivation lipocalin 2 mediates an innate immune response to bacterial infection by sequestrating iron agonist-induced trafficking of the low-affinity formyl peptide receptor fprl1 identification of a series of differentiation-associated gene sequences from gm-csf stimulated bone marrow murine serum amyloid a3 is a high density apolipoprotein and is secreted by macrophages plasma proteome of severe acute respiratory syndrome analyzed by two-dimensional gel electrophoresis and mass spectrometry apoptosis in lymphoid and parenchymal cell during sepsis: findings in normal and t & b cell deficient mice we thank ms alice tong, ms sandra k macmillan, and ms tracey h wagner for expert technical assistance. critical revision: chung, downey, buchman, stormo, hotchkiss, cobb tests for differential expression, class prediction, and pathway analysis: for the mouse blood study, a mixed-model anova was used to detect differential expression between treatment groups, with a linear contrast between the any-clp and non-clp groups. the anova model was chosen to partition treatment group and technical batch variability from variability due to biological and experimental noise. the following linear mixed model was used to detect differential expression on a gene-bygene basis in the mouse blood data:where y gij is the expression of the gth gene for ith treatment and jth batch. the mean expression for the gth gene is given by g . the symbols t and b represent effects due to treatment and batch respectively. the error for the gth gene for sample ij is designated as gij . for the mixed-model analysis of variance, treatment is a fixed effect and batch is a random effect. a batch constitutes 7 samples (one from each treatment group) which were processed and hybridized at the same time. in the case of the last batch (batch 5), the genechips were scanned on a different scanner. for the mouse and human spleen studies samples were processed in a single batch, so a simple one-way analysis of variance with a contrast between any-clp and non-clp was used to identify differentially expressed genes. the linear contrast between any-clp and non-clp is given by:where clp is the mean of the clp group, clpϩabx is the mean of the clpϩabx group, etc.where possible, the following competing classifiers were considered for all tasks, and the optimal classifier was selected: number of genes (1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50 , 60, 70, 80, 90, 100, 150, 200, 250, 300, 350, 400, 450, 500, 600, 700, 800, 900, 1000, 2000, 5000, and 10000), prior probabilities for nearest centroid (equal and proportional), functions for discriminant analysis (liner and quadratic), prior probabilities for discriminant analysis (equal and proportional), number of neighbors (k) for k-nn (1, 3, and 5), and distance functions for k-nn (euclidean distance, pearson's linear correlation, and absolute value distance). thus as many as 426 classification models were considered for each classification task.for the mouse blood data, we used a leave-one-batchout (5-fold, one for each of the 5 batches) outer crossvalidation, while the inner cross-validation is leave-onesample-out. we refer to this as nested cross-validation with an outer "leave-one-batch-out" layer and an inner "leave-one-sample-out" layer. (27) using this method, the determination of how many and which genes to use for classification were determined using only the training samples. in addition, all additional classifier parameters (e.g., number of neighbors and distance measure) were determined using only the training samples. for each held-out batch in the outer 5-fold cross-validation, the classifier and genes that performed best on inner cross-validation were selected and applied to the 6 or 7 held-out test samples (two batches only had 6 samples due to removal of an outlier sample). for the mouse and human spleen data where all samples were processed in a single batch, both the outer and inner cross-validation used full leave-one-sample-out. 598 .e1 key: cord-017420-tjwxec77 authors: stephens, r. scott title: neutropenic fever in the intensive care unit date: 2019-07-09 journal: oncologic critical care doi: 10.1007/978-3-319-74588-6_118 sha: doc_id: 17420 cord_uid: tjwxec77 neutropenic fever is a common and potentially life-threatening condition in patients treated for cancer. rapid initiation of appropriate antimicrobial therapy is necessary to decrease the risk of mortality. most infections are due to gram-positive organisms, but the mortality rate is higher for gram-negative infections. multidrug-resistant organisms are an emerging threat to neutropenic patients. increasing data suggest that the pathophysiology of neutropenic fever and neutropenic sepsis is substantially different from non-neutropenic fever and sepsis. additional research is needed to both further elucidate the pathogenesis of neutropenic fever and to develop additional effective antimicrobials. neutropenic fever is one of the defining conditions of oncologic critical care. neutropenic fever is defined as a single temperature higher than 38.3 c or a sustained temperature greater than 38.0 c for more than 1 h, in the presence of an absolute neutrophil count [anc]) less than 1500 cells/mm 3 , though many centers and guidelines use an anc cutoff of less than 500 cells/mm 3 [28, 33] . exceedingly common in patients receiving cytotoxic chemotherapy, neutropenic fever is a medical emergency which requires urgent initiation of broad-spectrum antibiotics. any oncologic patient receiving myelosuppressive chemotherapy is at risk of developing neutropenia and opportunistic infections, but profound neutropenia with life-threatening infectious complications is most commonly seen in patients with hematologic malignancies. this can occur during aplasia and before engraftment in patients receiving hematopoietic stem cell transplant (hsct) or from disease-related or treatment-induced cytopenias in other hematologic malignancy patients. while there are many potential causes of fevers in neutropenic hosts, both infectious and noninfectious, fevers must be presumed to be infectious in origin. potential infectious agents and sources of infection are legion and include viruses, bacteria, and fungus. viral pathogens include respiratory viruses (e.g., respiratory syncytial virus, rhinovirus, adenovirus, coronavirus, influenza, parainfluenza), reactivated or de novo herpes viruses (e.g., herpes simplex, cytomegalovirus, epstein-barr virus), and many other potential candidates. bacterial infections may arise from gut translocation, oral mucosal translocation, infection of indwelling vascular catheters, skin and soft tissue infections, pneumonias, and urinary sources. fungal infections typically arise from gut translocation, fungal pneumonias (e.g., aspergillosis), and vascular catheters. drug fevers or "tumor fevers" are the most common example of noninfectious fevers, but these are diagnoses of exclusion. neutropenic fever occurs in up to 50% of patients with solid tumors receiving cytotoxic chemotherapy and in more than 80% of patients receiving chemotherapy for hematologic malignancies or undergoing hsct [28] . in 2012, more than 90,000 adults were hospitalized for cancer-related neutropenia, with a total cost of $2.3 billion [75] . in-hospital mortality for all patients admitted with neutropenic fever is nearly 10%; this increases to a hospital mortality rate of more than 15% for patients with leukemia admitted for neutropenic fever [37] . bacteremia is documented in up to 25% of neutropenic fever patients [28] . whereas in the past gram-negative organisms were commonly cultured, gram-positive organisms, including staphylococci, enterococci, and streptococci, are currently the most commonly isolated bacteria [28, 50] (table 1) . this shift is presumably due to the increased use of long-term indwelling vascular catheters and may also be affected by increased use of prophylaxis against gram[50, 59] . though grampositive infections are more common, gramnegative infections confer a higher risk of mortality [77] . fungal infections, particularly candida and aspergillus, are also frequent, especially in patients with prolonged or profound neutropenia [33] . respiratory viruses can be isolated in approximately 20% of patients [34] . despite best efforts, no causative organism can be identified in about 50% of cases of neutropenic fever [28, 33] . sepsis has been most recently defined as "lifethreatening organ dysfunction caused by a dysregulated host response to infection" [70] . a premium is placed on identification of organ dysfunction using either the sequential organ failure assessment (sofa) score or the quick sofa (qsofa) score; the presence of organ dysfunction in the setting of a suspected or proven infection is sufficient to diagnose sepsis (tables 2 and 3 ). septic shock is defined by the need for vasopressors to maintain a mean arterial pressure (map) !65 mmhg or a serum lactate >2 mmol/l despite adequate volume resuscitation. there is no specific consensus definition for neutropenic sepsis, other than sepsis occurring in the presence of neutropenia. any evidence of organ dysfunction, including an elevated lactate, in the presence of neutropenia should be treated as a potential indicator of sepsis. there are few reliable data on the incidence of neutropenic sepsis or neutropenic septic shock. it has been estimated that 50% of patients with neutropenic fever will develop sepsis, and up to 10% of patients with neutropenic fever will progress to septic shock [33] . among neutropenic allogeneic hsct patients, approximately 10% will develop severe sepsis during the engraftment period [38] . cytotoxic chemotherapy or cytotoxic radiation, whether given as an antitumor agent or conditioning regimen for hsct, induces neutropenia by injuring or destroying hematopoietic precursor cells within the bone marrow as well as injuring the bone marrow structure itself [44] . circulating [43] , so neutropenia develops rapidly after myelosuppressive therapy. depending on the chemotherapy employed, circulating neutrophils and other white blood cells may also be damaged or destroyed, causing a more rapid leukopenia than regimens which only affect the marrow. some agents will injure progenitor cells but not primitive stem cells (which re-populate the progenitor cells), whereas other agents will injure all cells in the marrow [44] . in the normal patient with a localized infection, neutrophils are rapidly recruited at the onset of focal infection and are essential to microbial killing and infection control [40, 45] . neutropenic fever and neutropenic sepsis have historically been thought of as variants of non-neutropenic fever and sepsis, just without an intact immune system, akin to a "fire without firefighters." however, increasing data suggest that the pathophysiology of neutropenic patients is much more complicated than simply an uncontrolled infection [36] . in addition to their key role in the response to localized infections (e.g., pneumonia), in sepsis neutrophils are rapidly recruited to organs (lungs, kidneys, liver) and are thought to contribute to tissue damage and organ dysfunction [45] . neutrophils also contribute to resolution of injury and tissue repair [36] , as well as regulation of the adaptive immune response [40, 76] . thus neutropenic patients are not only deficient in the immediate response to infection but also have an altered physiology regarding the development of systemic inflammation and sepsis and in the regulation of the immune response to injury and tissue repair. it is increasingly unclear whether neutropenic sepsis (and the organ damage sequelae) is the same disease as non-neutropenic sepsis. because cytotoxic chemotherapy is not selective, neutropenic patients are also lymphopenic, thrombocytopenic, and anemic. lymphocytes, including t cells, b cells, and natural killer (nk) cells, have protean roles in the immune system and a broad effect on the response to infection and septic physiology [20, 43] . these cells are also drastically reduced after cytotoxic chemotherapy, and lymphopenia has been associated with an increased risk of death [67, 71] . adding to the complexity, the kinetics of count recovery and engraftment vary, and reconstitution of all cell lines does not happen simultaneously [11] . though the total effect of lymphopenia on the pathophysiology of neutropenic fever and infection remains to be fully determined, patients with neutropenic sepsis have different cytokine profiles than patients with non-neutropenic sepsis: neutropenic patients have higher levels of il-6, il-8, and g-csf than non-neutropenic patients [60] . monocytes are also dysfunctional in neutropenic sepsis, with evidence not only of deactivation of monocytes in peripheral blood but also of deactivation of differentiated pulmonary macrophages in patients with neutropenic sepsis [48, 49] . the implications of pancytopenia for the response to infection extend beyond white blood cells. most neutropenic patients are thrombocytopenic, and platelets are increasingly recognized to play an important role in the immune response [23, 74] . thrombocytopenia is associated with poor outcomes in critical illness, including in sepsis [18] . in addition to secreting mediators and regulators of inflammation, platelets interact with neutrophils and monocytes and play a vital role in the defense against bacterial, viral, and fungal infections via the formation of neutrophil extracellular traps (nets) [16, 21, 23] . platelets also play roles in the development and resolution of organ failure in inflammatory states, including sepsis. in particular, thrombocytopenia has been demonstrated to potentiate lung injury [46, 86] , and platelets may play a role in the pathophysiology of acute kidney injury [23] . while the role of thrombocytopenia in infection remains incompletely elucidated, it seems clear that [70] thrombocytopenia is an important factor in the pathophysiology of neutropenic infection. other organ systems with relevance to neutropenic infections are also affected by cytotoxic chemotherapy. the most important of these is injury to the mucosal barrier of the intestinal tract [13, 14, 78] . disruption of this barrier, which can occur throughout the gastrointestinal (gi) tract, creates portals through which enteric pathogens, including bacteria and yeast, can enter the bloodstream. this is an important source of gram-positive, gram-negative, and fungal infections in neutropenic patients. the respiratory system is also affected in neutropenia. not only are pulmonary macrophages known to be qualitatively dysfunctional in neutropenic sepsis [48, 49] , but quantitative cell counts of alveolar macrophages, lymphocytes, and neutrophils are decreased during neutropenia [19] . neutropenia also appears to adversely affect lung repair after injury [12] . taken together, these data strongly support the hypothesis that neutropenic sepsis has a significantly different pathophysiology than non-neutropenic sepsis. as defined above, the diagnosis of neutropenic fever requires a single temperature higher than 38.3 c or a sustained temperature greater than 38.0 c for more than 1 h in the presence of neutropenia [28, 33] . while some patients are asymptomatic in the presence of neutropenic fever, many describe non-specific symptoms (e.g., cough, anorexia, nausea, fatigue, dizziness, myalgias, confusion, behavioral changes). patients may also present with respiratory symptoms (cough, shortness of breath, sinus pain or drainage) or abdominal symptoms such as pain or diarrhea. fewer than half of patients will feel feverish, shiver, or have rigors [17] . presentation to the hospital can be delayed, with one study suggesting a mean delay in presentation of 11 h and nearly 40% of patients delaying presentation for more than 12 h [17] . though neutropenic patients have higher fevers than non-neutropenic patients, there is no association between peak temperature and mortality. hypothermia during neutropenic sepsis is associated with worse outcomes [84] . neutropenic patients with septic shock tend to have more frequently positive blood cultures, more fungal infections, more multidrug-resistant bacterial infections, and higher mortality rates than immunocompetent patients. compared to non-neutropenic patients, patients with neutropenic sepsis have higher rates of shock and are at higher risk to sustain acute kidney injury [60] . early recognition of neutropenic fever is essential. regardless of whether neutropenic patients are hospitalized or not, frequent temperature checks are essential to detect fever; outpatients must particularly be educated on the importance of monitoring temperature. similarly, interventions that might mask fevers (e.g., antipyretics such as acetaminophen) should generally be avoided in neutropenic patients, and clinicians should be aware of other interventions (e.g., steroids, continuous renal replacement therapy) which may suppress fevers. once a fever is detected, blood cultures should be obtained without delay, and antibiotics initiated as quickly as possible. diagnostic measures must not interfere with the timely administration of antibiotics. a comprehensive physical exam should be performed and may uncover potential sources of infection (e.g., mouth sores, skin lesions, pulmonary findings, abdominal tenderness or pain). blood cultures are essential to the evaluation of neutropenic patients with fever. a minimum of two sets of blood cultures should be drawn upon presentation. current recommendations suggest obtaining two sets of cultures, including both peripheral blood cultures and cultures from a central venous catheter, if present [28, 33] . additional laboratory studies at presentation should include complete blood count with differential, electrolytes, and markers of renal and hepatic function (creatinine, blood urea nitrogen, transaminases). depending on the clinical scenario, strong consideration should be given to obtaining an arterial blood gas, coagulation studies, and lactate level [33] . lactate is of particular potential interest, as an elevated level may help detect early evidence of sepsis-induced malperfusion [33, 70] . other microbiologic studies can be targeted toward patient-specific indicators. for example, urinary symptoms or an abnormal urinalysis should prompt urine cultures. diarrhea, especially in a patient treated with antibiotics, should prompt evaluation for clostridium difficile colitis. fungal markers such as galactomannan or beta-d-glucan may be useful in some patients [33] . respiratory symptoms or abnormalities on chest imaging should be evaluated with testing for respiratory viruses and sputum culture [28] . symptom-guided imaging studies comprise an important part of the evaluation of neutropenic fever. chest computed tomography (ct) scanning should be performed in patients with respiratory symptoms, and potentially asymptomatic patients with cryptic fevers [33] . plain chest radiographs are of limited utility in this population and should not be routinely obtained in lieu of ct scans [87] . sinus, head, and abdominal imaging should be performed as indicated [28, 33] . the use of nuclear medicine techniques such as fdg-pet/ ct to identify foci of infections in febrile neutropenia has been described, but the utility of these techniques has not yet been proven, and remains impractical for current clinical use [80] . as noted above, respiratory symptoms and/or the presence of abnormalities on chest imaging should prompt evaluation for a respiratory infection. in most cases, this can be done noninvasively [6, 8, 10] . bronchoscopy may be indicated in some patients, but the benefits of potential diagnosis must be weighed against the risk of requiring endotracheal intubation during bronchoscopy. abdominal symptoms (pain, diarrhea) should lead to consideration of neutropenic enterocolitis, also known as typhlitis, which is an incompletely understood condition of ileocolonic inflammation which can lead to intestinal necrosis and perforation and is associated with a high mortality rate [63, 64] . diagnosis requires the presence of neutropenia, bowel wall thickening >4 mm over a >30 mm longitudinal distance, fever >38 c, and abdominal pain [31, 51, 63] . specific diagnosis is elusive in many patients, and diagnostic steps should be repeated if no source is found and fevers persist for 72-96 h. repeat blood cultures and repeat or expanded imaging studies may provide additional diagnostic information. risk stratification is an essential part of diagnosis, as it informs immediate management. the multinational association of supportive care in cancer (mascc) score (table 4 ) [35] was developed to predict which patients with neutropenic fever may be safely treated as outpatients. a score !21 identifies a standard risk patient, whereas a score <21 indicates a high-risk patient. additional criteria for outpatient treatment have been enumerated by heinz et al. [33] ; these focus on signs of clinical and social stability, with particular emphasis placed on expected good adherence to oral medications, adequate social support (the patient does not live alone), and the ability to present to the hospital within 60 min. it should be noted that the mascc score has limited utility in predicting either the risk of critical illness or icu outcomes. other factors which influence risk stratification include the depth and duration of neutropenia, with an anc 100 cells/mm 3 and >7 days duration, respectively, being markers of a high-risk patient [28] . accordingly, neutropenic modified from [35] patients with a hematologic malignancy or status post hsct are almost always higher risk than patients who become neutropenic during the course of cyclic treatment for a solid malignancy. comorbid conditions should also be integrated into any clinical risk assessment. pharmacologic prophylaxis in neutropenic patients before neutropenic fever develops due to the high risk of infection in neutropenia, many centers provide routine antimicrobial prophylaxis to patients deemed to be at high risk for infection and who are expected to have prolonged periods of profound neutropenia (e.g., anc 500 cells/mm 3 for >7 days). the rationale for antibacterial prophylaxis is to reduce the risk of gram-negative infections and streptococcal infection from oral mucositis. in high-risk patients, current guidelines suggest antibacterial prophylaxis with a fluoroquinolone such as ciprofloxacin or levofloxacin, with the latter preferred if severe mucositis is anticipated [29, 30] . antifungal prophylaxis against yeast is recommended in highrisk patients (hsct, chemotherapy for leukemia) with fluconazole, itraconazole, voriconazole, posaconazole, or an echinocandin (caspofungin or micafungin). anti-mold (aspergillus) prophylaxis (with voriconazole or posaconazole) is recommended in patients undergoing chemotherapy for leukemia or patients with anticipated very prolonged neutropenia or prior invasive mold infection [28] . antiviral prophylaxis acyclovir or valacyclovir is recommended for patients who are seropositive for herpes simplex virus (hsv) and for varicella zoster virus (vzv)-seropositive hsct patients. finally, leukemia and hsct patients should receive prophylaxis against pneumocystis jirovecii pneumonia. these prophylactic antimicrobials may lessen, but do not eliminate the risk infections while neutropenic; thus, patients and caregivers must remain vigilant for the development of fever. moreover, attention must be paid to the prophylactic regimen, as it will affect the choice of empiric antibiotics for neutropenic fever. neutropenic fever is a medical emergency, and appropriate empiric antibiotics must be started without delay: within 60 min of presentation [28, 33, 57] . some data suggest that even delays in antibiotic administration beyond 30 min are associated with increased mortality [62] . as noted above, it is desirable to obtain blood cultures if possible before antibiotic initiation, so long as this does not delay antibiotic administration. no other diagnostic maneuvers should be attempted before antibiotic initiation. empiric antibiotics must cover the common organisms discussed above (pseudomonas aeruginosa, staphylococcus aureus, streptococcal species) and should also be tailored according to prior patient-specific culture data and institutional epidemiology [28, 33] . appropriate empiric antibiotics include an antipseudomonal penicillin (e.g., piperacillin/ tazobactam), or antipseudomonal cephalosporin (e.g., cefepime), or a carbapenem (imipenem, meropenem) [28, 33] . some data suggest improved outcomes with prolonged antibiotic infusion times [58] though these data require confirmation. fluoroquinolones, which are frequently used as prophylactic therapy in neutropenia, should not be used as empiric monotherapy in neutropenic fever due to the possibility of resistance. though gram-positive organisms are common causes of neutropenic fever, vancomycin is not routinely indicated, but should be added in the presence of suspected catheter-related infection, soft tissue infection, oral mucositis, radiographically proven pneumonia, known colonization with resistant gram-positive organisms (e.g., methicillin-resistant staphylococcus aureus, penicillin-resistant streptococcus), or hemodynamic instability [28, 33, 57] . the use of combination antibiotic regimens (defined as dual gram-negative coverage with an antipseudomonal beta-lactam and an aminoglycoside) in neutropenic fever is controversial. though some studies have suggested a mortality benefit to combination antibiotics [39] , this has not been a consistent finding, and 2 metaanalyses have shown no benefit to the addition of an aminoglycoside to a beta-lactam and a higher risk of renal failure with combination therapy [55, 56] . accordingly, current guidelines for the management of neutropenic fever and sepsis recommend monotherapy with an antipseudomonal beta-lactam unless otherwise dictated by circumstances such as patient allergies, the presence of resistant organisms, or refractory hemodynamic instability [28, 57, 61] . it should be noted, however, that this recommendation does not preclude the use of vancomycin or antifungals; it is only directed at combination therapy targeted against gram-negative bacteria. even with appropriate antibiotics, fever in neutropenic patients typically persists for a median of 5 days; thus, ongoing fevers, unless accompanied by clinical instability, should not necessarily be viewed as evidence of failure of antibiotic therapy [73] . in patients with persistent or recurrent fevers after 3-5 days, a modification of antibiotic regimen is reasonable, especially if guided by new or changing clinical data. earlier antibiotic escalation is necessary in some patients, most commonly in patients with hemodynamic instability. in the face of hemodynamic instability, vancomycin should be added if not already part of the regimen. additionally, antipseudomonal cephalosporins or penicillins should be escalated to a carbapenem (e.g., meropenem), and strong consideration should be given to the addition of an aminoglycoside, fluoroquinolone, or aztreonam [28, 33, 57, 61] . drug-resistant and multidrug-resistant (mdr) organisms are an increasing problem in neutropenic infections [26, 29, 50] . vancomycin-resistant enterococcus (vre) bacteremia affects a substantial portion (10-35%) of patients during induction therapy for leukemia or after hsct and is associated with significantly worse outcomes [3, 53, 85] . early treatment with agents active against vre such as linezolid or daptomycin may improve outcomes. predictive models are being developed to assist with early identification of patients who might benefit from early initiation of antibiotics active against vre [83] . similarly, mdr gram-negative infections, particularly carbapenem-resistant enterobacteriaceae, are associated with high mortality rates, especially among allogeneic hsct patients [66] . successful treatment of these infections requires early use of multidrug antibiotic regimens, typically including aminoglycosides, carbapenems, and polymyxins. the use of surveillance rectal cultures, performed pre-transplant and then weekly after hsct, to identify patients with mdr infections and allow immediate initiation of antibiotic therapy targeted against mdr organisms may result in better outcomes [26] . fungal pathogens are a constant threat in neutropenic patients, and consideration must be given to the use of antifungal agents in all patients with neutropenic fever. in general, antifungal therapy should be initiated in the setting of persistent fever after 5-7 days of appropriate antibacterials [28, 33] . appropriate antifungals should have activity against molds, especially aspergillosis; examples include liposomal amphotericin, caspofungin, and voriconazole, though the data are more robust in favor of the former two [33, 81, 82] . fluconazole should not be used as empiric therapy. an increasing number of neutropenic patients are on antifungal prophylaxis with voriconazole prior to developing fever; the utility of changing antifungal agents upon fever development in this setting is unclear. antifungals should be strongly considered as early therapy in all patients who are hemodynamically unstable [28, 33] . with the advent of multidrug-resistant organisms, antibiotic stewardship is increasingly important, even in neutropenic patients. the optimum duration of antimicrobial therapy in the neutropenic patient and whether antimicrobials may be safely de-escalated in the face of clinical stability are ongoing areas of investigation. one randomized controlled trial suggests that empiric antibiotics may be safely discontinued after a patient has defervesced and remained afebrile for 72 h, regardless of whether neutrophil recovery has occurred [2] . another recent paper suggested that it is safe to withhold antibacterial therapy in children with neutropenic fever in whom infection with a respiratory virus has been proven [65] . neither of these tactics has become standard of care, but both highlight the increasingly realized importance of antibiotic stewardship, even in neutropenic patients. hematopoietic growth factors may be considered in select cases of neutropenic fever, as they have been shown to shorten the duration of neutropenia, but do not impact mortality [22] . according, due to a lack of proven mortality benefit, current guidelines recommend against the routine use of hematopoietic growth factors in neutropenic fever or neutropenic sepsis [29, 72] . granulocyte transfusions have been used to support patients with neutropenia, both to prevent infections and to help treat established infections. very few studies have been performed to evaluate this intervention, and those studies that are available are small. two recent cochrane metaanalyses have examined the use of granulocyte transfusions in neutropenic patients to prevent and treat infections, respectively. both concluded that there was insufficient evidence to determine whether granulocyte transfusions conferred any mortality benefit [24, 25] . early and appropriate administration of antimicrobials is essential to preventing death from neutropenic fever and neutropenic sepsis. good outcomes also depend on successfully managing the hemodynamic derangements and organ failure of sepsis. the intensive care unit (icu) is the bestsuited location to care for neutropenic patients with sepsis and septic shock, and earlier icu admission has been associated with improved survival rates [7, 9] . recent guidelines have been published for the management of sepsis and septic shock [61] ; these guidelines are also applicable to the management of neutropenic sepsis. key points include initial resuscitation with at least 30 ml/kg of intravenous crystalloid with additional fluid resuscitation as indicated and using norepinephrine as a first-line vasopressor to target a mean arterial pressure !65 mm hg. either vasopressin or epinephrine may be added if the response to norepinephrine is inadequate. there may be benefit to using balanced crystalloid solutions such as lactated ringer's or plasmalyte rather than normal saline [68, 69] . source control should be obtained if possible. though occasionally an abscess may be present and feasible to drain, most commonly, an indwelling central venous catheter is the only addressable source of infection. in the hemodynamically unstable patient with a suspected catheter infection, early catheter removal is associated with improved survival [39] ; accordingly, infected or potentially infected catheters should be removed without delay. patients with neutropenia and sepsis are at high risk of developing multi-organ failure, particularly the acute respiratory distress syndrome (ards) [4, 5] . the use of noninvasive ventilation in immunosuppressed patients with hypoxemic respiratory failure is increasingly controversial, and heated humidified high-flow oxygen may be a better option [5, 15, 27, 41, 42, 52] . once patients are intubated, low tidal volume ventilation should be used to maximize lung protection and minimize ventilator-induced lung injury [1] . adjuncts such as neuromuscular blockade and prone positioning should be used in patients with moderate to severe ards (pao2: fio2 < 150) [32, 54] . mortality for patients who develop ards in this context remains high, but outcomes are improving. attention to best practices for mechanical ventilation is essential. an algorithm for the empiric management of neutropenic fever can be found in fig. 1 . the risk of neutropenic fever and neutropenic sepsis resolves once the bone marrow recovers and neutrophil counts return to normal. if treated appropriately, neutropenic fever is a common, predictable, and manageable complication of cytotoxic therapy or hsct. neutropenic sepsis continues to confer a poor prognosis, with recent data suggesting an approximate 46% mortality rate in patients with hematologic malignancies who develop septic shock [7, 39, 47] . predictors of mortality include sepsis after allogeneic hsct, the presence of graft vs host disease, respiratory failure requiring mechanical ventilation, positive blood cultures, cardiac failure, renal failure, and hepatic failure [38, 47] . younger age (<70 years) and the presence of neutropenic enterocolitis are associated with improved survival [47] . encouragingly, survival in neutropenic sepsis and septic shock appears to be improving but is still worse than in non-neutropenic septic patients [9, 10, 39] . the emergence of multidrug-resistant organisms is a major concern, and there is an urgent need for novel antibiotics to address this threat. along these lines, additional work needs to be done to identify patients in whom antibiotic therapy can be safely de-escalated. the 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impact of neutropenia on the outcomes of critically ill patients with cancer: a matched case-control study small but mighty: platelets as central effectors of host defense cost of cancer-related neutropenia or fever hospitalizations, united states neutrophil-mediated suppression of influenza hemabis registry-seifem group, i. current epidemiology and antimicrobial resistance data for bacterial bloodstream infections in patients with hematologic malignancies: an italian multicentre prospective survey mucosal barrier injury, fever and infection in neutropenic patients with cancer: introducing the paradigm febrile mucositis the sofa (sepsis-related organ failure assessment) score to describe organ dysfunction/failure. on behalf of the working group on sepsis-related problems of the european society of intensive care medicine the use of fdg-pet/ct in patients with febrile neutropenia liposomal amphotericin b for empirical therapy in patients with persistent fever and neutropenia. national institute of allergy and infectious diseases mycoses study group caspofungin versus liposomal amphotericin b for empirical antifungal therapy in patients with persistent fever and neutropenia prediction of bloodstream infection due to vancomycin-resistant enterococcus in patients undergoing leukemia induction or hematopoietic stem-cell transplantation association between early peak temperature and mortality in neutropenic sepsis colonization, bloodstream infection, and mortality caused by vancomycin-resistant enterococcus early after allogeneic hematopoietic stem cell transplant platelets in the pathogenesis of acute respiratory distress syndrome the utility of routine chest radiography in the initial evaluation of adult patients with febrile neutropenia patients undergoing hsct key: cord-030385-btf502ju authors: sun, zhiheng; pan, yuchen; qu, junxing; xu, yujun; dou, huan; hou, yayi title: 17β-estradiol promotes trained immunity in females against sepsis via regulating nucleus translocation of relb date: 2020-07-22 journal: front immunol doi: 10.3389/fimmu.2020.01591 sha: doc_id: 30385 cord_uid: btf502ju sepsis is more common among males than females, and the unequal estrogen levels have been suspected to play a vital role in gender differences. recently, trained immunity is reported to be a novel strategy for the innate immune system to fight infection. however, it has not been clarified whether β-glucan-induced trained immunity causes different responses to early sepsis between male and female mice. in this study, sepsis was induced in mice by intraperitoneal injection of escherichia coli (e. coli). the changes of inflammatory cytokines expression, and macrophage polarization in male, female, and ovariectomized c57bl/6 mice in sepsis model were investigated. for in vitro studies, different macrophages were treated with lps. the function of estradiol (e2) on macrophage cell lines was verified and the mechanism of e2 affecting trained immunity was explored. we demonstrated that β-glucan-induced trained immunity was more resistant to sepsis in female than male mice. macrophage polarization toward the m1 phenotype, which exhibited enhanced trained immunity, was related to the difference in sepsis resistance between female and male mice. moreover, ovariectomized (ovx) mice manifested serious sepsis consequences with a weaker trained immunity effect than female mice. female bone marrow-derived macrophages (bmdms) were also apt to be polarized to the m1 phenotype in response to trained immunity in vitro. furthermore, e2 promoted trained immunity in macrophage cell lines j774 and raw264.7. e2 was also verified to facilitate trained immunity in primary bmdms from female and male mice. mechanistically, we found that e2 inhibited the nuclear translocation of relb, which is a member of non-canonical pathway of nfκb and contributes to macrophage polarization to change the intensity of trained immunity. this study is the first to indicate the role of e2 in the trained immunity induced by β-glucan to protect against e. coli-induced sepsis via the non-canonical nfκb pathway. these results improve our understanding of the molecular mechanisms governing trained immunity in gender differences. sepsis is a systemic reaction, which can even be caused by an ordinary infection (1) . the infection is most commonly bacterial but also can be fungal, viral, or parasitic. since many organs are affected, the mortality rate of sepsis is 30-50% (2) . with the development of modern medicine, the mortality rates have declined to about 30% (3), but this is still far from acceptable. the biomarker of sepsis is widely discussed by a number of reviews (4) (5) (6) (7) . several important signs of sepsis are damage to the kidney, liver, and lung, as well as an increased bacterial load in the kidney and elevated levels of transaminase and lactate in serum (8) . moreover, the common model of sepsis is an immune response to escherichia coli or endotoxin, lipopolysaccharide (lps), found in the cell wall of gram-negative bacteria. endotoxin is an excellent example of a pathogen-associated molecular pattern (pamp). of note, a number of studies indicate gender dimorphism in terms of response to sepsis (5) . the intuitive result is that the incidence of female sepsis is much lower than that of males (9, 10) . furthermore, some studies attributed these differences to the prevailing hormonal milieu of the victim (11) (12) (13) . thus, it is necessary to explore the mechanism of estrogen in modulating immunity, which allows females to resist sepsis. the nfκb (nuclear factor kappa-b) family of transcription factors constitutes five members (rela or p65, relb, crel, nfκb1 or p50, and nfκb2 or p52), all of which play important roles in cell homeostasis, especially in the immune process (14) . apart from the well-known canonical signaling pathway, the noncanonical pathway is also involved in vital immune processes, which could be triggered by signaling from a subset of tnfr members. this pathway mediates the persistent activation of relb/p52 complex with the ability to modulate a series of gene expression, including macrophage polarization-associated cytokines. to date, a growing number of studies indicated that estrogen is involved in some immune processes. estrogen can bind to and activate estrogen receptors (ers), which regulate the expression of downstream genes (15) . the interaction between ers and nfκb is mainly discussed in breast cancer articles (16) . a highly significant negative correlation between the expression of nfκb target genes and er activation was found. some studies abbreviations: e 2 , 17-β-estradiol; ti, trained immunity; lps, lipopolysaccharide; pbs, phosphate buffer saline; ast, aspartate aminotransferase; alt, alanine aminotransferase; il-6, interleukin-6; tnfα, tumor necrosis factor α; il-10, interleukin-10; il-4, interleukin-4; m-csf, macrophage colony-stimulating factor. demonstrated the interaction of er with nfκb; ers can compete with nfκb for binding to transcriptional coactivators (ie, creb) or ers to recruit co-suppressors into nfκb complexes (17) . ers can inhibit de novo relb synthesis in breast cancer tissues and cell lines. in addition, e 2 inhibits the nucleus translocation of p65, c-rel, and relb without affecting p50 in mouse splenocytes (18) . these data suggest that estrogen-er signaling regulates the nfκb pathway at the transcriptional level of its constituents. the host's immune defense mechanisms can be divided into innate immunity and adaptive immunity. adaptive immunity, although slower than the innate immune response, has good specificity and produces immunological memory (19) . recently, researchers discovered an ability in innate immunity, similar to immune memory in adaptive immunity, called trained immunity (20) . trained immunity is found in mammals and its basic features have been identified by several researchers. trained immunity mainly involves a set of cells (myeloid cells, natural killer cells, and innate lymphoid cells) (21) . antituberculosis vaccine bacillus calmette-guérin (bcg) is a wellknown immune modulator that induces trained immunity. it was reported that estrogen did not influence the induction of trained immunity by bcg, and did not induce training or tolerance in monocytes themselves, indicating that the estrogen is unlikely to explain the sex-differential effects after bcg vaccination (22) . in fact, β-glucan is a major cell wall component of c. albicans and can induce trained immunity in monocytes. the initiation of trained immunity is associated with enhanced signaling of the akt (protein kinase b)-mtor (mammalian target of rapamycin)-hif-1α (hypoxia-inducible factor-1α) pathway (23) , modifications in metabolic pathways (conversion to glycolysis), and epigenetic rewriting (24, 25) . however, the effect of estrogen on β-glucan-induced trained immunity to resist sepsis has not been clarified. β-glucan (g-59303) and β-estradiol (50-28-2) were purchased from xiensi biotechnology company (tianjin, china). m-csf (#cb34), ifn-γ (#c746), and tnfα (#cf09) were purchased from novoprotein (shanghai, china). e 2 and lps was purchased from sigma (st. louis, mo, usa). antibody against p-akt (ser473, #4060), akt (#4691), p-4ebp1 (#2855), 4ebp1 (thr37/46, #9644t), p-s6 (ser235/236, #4858), s6 (#2217), pcna (#13110), p65 (#8242), estrogen receptor α (13258s), gapdh (#5174), and β-actin (#3700) were purchased from cell signaling (boston, mo, usa). relb (sc-166416) was purchased from santa cruz biotechnology (dallas, tx, usa). f4/80-apc, f4/80-fitc, cd116-pe, cd206-apc, cd206-fitc, inos-pe were all from biolegend (san diego, ca, usa). raw264.7, j774, and hek293t cells were obtained from the type culture collection of the chinese academy of sciences, shanghai, china. cells were cultured in phenol red-free dmem with 10% fbs, 1% (100 u/ml penicillin and 100 ug/ml streptomycin) at 37 • c in an atmosphere of 95% air and 5% co 2 . cells were seeded onto different types of plates for further experiments when the cell density reached ∼80%. the cells were used within a maximum of five passages. the in vitro trained immunity model was established with raw264.7 and j774. cells were challenged with 5 µg/ml βglucan for 24 h. the cells were then washed and rested in culture medium for 5 days. next, cells were treated with 100 ng/ml lps for 6 h, and then the cytokines were measured in mrna level. 1 × 10 5 bmdms were seeded in 96-well plates (200 µl final volume; corning) and stimulated with 100 µg/ml β-glucan for 24 h. then cells were washed and rested for 3 days in culture medium. on day 4, bmdms were washed again and treated with 25 ng/ml ifn-γ for 24 h. on day 5, a final wash was performed, and cells were primed with 1 µg/ml lps. the supernatants were harvested for elisa assay after 24 h of lps stimulation. other experiments were all based on the same model but with different numbers of bmdms. to assess mrna expression and cell viability, 6 × 10 5 bmdms were plated in non-treated 24well plates (1,200 ml final volume; corning) and followed the training scheme described above. for western blotting (wb) assays 3 × 10 6 bmdms were plated in six-well plates (3 ml final volume; corning). male and female c57bl/b mice were purchased from the model animal research center of nanjing university. mice used in the model of trained immunity and sepsis were 10 weeks old. all animal procedures were performed in accordance with guidelines of the us nih with specific pathogen free conditions. soyfree standard rodent chow and water were provided ad libitum. female mice were anesthetized with 4% chloral hydrate and then underwent ovariectomy (ovx) at 6 weeks of age. when they were 10 weeks old, the same model of trained immunity and sepsis was established. peripheral blood mononuclear cells (pbmcs) were separated from mouse plasma by ficoll centrifugation using lymphocyte separation medium from mp biomedicals (solon, usa) according to the standard procedures. mice were sacrificed via cervical dislocation and sterilized by soaking in 75% ethanol. dissected the legs and bone marrow was extracted from tibia and femur bones by using a 25-gauge needle and a 1 ml syringe filled with pbs following removal of surrounding muscle. blow the bone marrow gently and spread it through a 70 µm cell strainer. the cell suspension was centrifuged at 300 g for 5 min at room temperature. cells were cultured in phenol red-free dmem with 10% ultra-low endotoxin fbs and m-csf (20 ng/ml). after changing the fresh medium on the third and fifth day, bmdm was induced. the e. coli 15597 strains were purchased from atcc, collected, and identified by the medical laboratory center of zhongda hospital in nanjing jiangsu, china, and stored at −80 • c. bacterial strains were prepared in lb medium. a nuclear protein extraction kit was purchased from biyuntian (wuhan, china) and used according to the manufacturer's instructions. sirna transfection sirna was transfected according to the product instructions (ruibo company, china). the concentration of sirna used in the study was 50 nm. the erα sirna target sequence is tgcacattgaagatgctga. the target sequence of non-coding (nc) sirna was a random sequence with no biological effects. to assess the effect of e 2 on cell viability, a cck-8 assay was used according to the manufacturer's instructions (bioss company, china). raw264.7/j774 were seeded onto 96-well plates at a concentration of ∼5 × 10 4 cells/well. different concentrations of e 2 were used to treat cells for different length of time as described in the article. total rna was extracted from cells using trizol reagent, and reverse transcriptions were performed in a 20 µl mixture with 1 µg of total rna according to the manufacturer's instructions (vazyme company, china). the oligonucleotide primers used for pcr amplification are listed in table 1 . pcr amplification consisted of 30 cycles of denaturation at 95 • c for 2 min, annealing at 60 • c for 45 s, and extension at 72 • c for 2 min. all reactions were run in triplicate. the gene expression levels were normalized to ß-actin. the protein samples were obtained from lysis buffer treated cells. cell lysates were put on ice for 15 min and then centrifuged at 12,000 × g for 10 min. subsequently, 30 µg of protein per lane was separated on 10% polyacrylamide gels and transferred onto polyvinylidene difluoride membranes (millipore, billerica, ma, usa). membranes were blocked with 5% bovine serum albumin (bsa) in tris-buffered saline containing 0.1% tween 20, and then the membranes were incubated with specific antibodies. the values were normalized to the β-actin/gapdh intensity levels. mice were trained with two intraperitoneal (i.p.) injections of 1 mg β-glucan particles on days −7 and −4. sterile pbs was used as a control. on day 0, mice were challenged with 1.77 × 10 7 e. coli. the lung, kidney, and serum were harvested 24 h after e. coli treatment. the fresh lung tissues were fixed in 4% paraformaldehyde (pfa). then, the samples were gradually dehydrated and embedded in paraffin. after that, the samples were cut into 3 µm sections and stained with hematoxylin and eosin for further light microscopy observation. scores were evaluated by a pathologist based on the lung tissue integrity, alveolar integrity, and mononuclear infiltration (0 = none; 1 = mild; 2 = moderate; 3 = severe). cultured cells were seeded on glass coverslips in six-well plates. after three pbs washes, the samples were fixed for 15 min at room temperature with 4% paraformaldehyde. fixed cells were rinsed with pbs and then incubated for 10 min at 4 • c with 0.2% triton x-100 and 0.2% bsa in pbs. following permeabilization, non-specific binding in the cells was blocked by 5% bsa in pbs for 1 h at room temperature. cell samples were incubated with anti-erα, anti-relb, and anti-p65 primary antibodies at a 1:200 dilution for 2 h at room temperature. samples were further incubated with alexa fluor 488-conjugated and alexa 647conjugated secondary antibody at a 1:400 dilution for 1.5 h in the dark. after washed with pbs, the nuclei were stained by dapi. slides were visualized using a nikon eclipse ti-u fluorescence microscope equipped with a digital camera (ds-ri1, nikon). the protein concentration of il-6, lactate, tnfα, and estrogen in cell supernatant or mouse serum were detected using the corresponding mouse enzyme-linked immunosorbent assay (elisa) kit according to the manufacturer's instructions (biolegend, china). the kidney e. coli burden at indicated time points was measured by plating organ homogenates obtained mechanically over 70 µm cell strainers (bd biosciences) following slicing the tissue, in serial dilutions on lb agar plates; colony-forming units (cfus) were counted after growth at 37 • c for 24 h, and data are shown as cfus in total kidney. pbmcs or bmdms were filtered through a 70 µm cell strainer and then washed with complete rpmi medium to generate single-cell suspensions. an fc-receptor blocker (cd16/32, ebioscience) was used to reduce non-specific antibody binding. antibodies used in these experiments included f4/80-apc, f4/80-fitc, inos-pe, cd206-apc, and cd11b-pe. stained samples were detected by a facs calibur flow cytometer (bd bioscience) and data were analyzed using flowjo software (treestar, ashland, or). the statistical analysis was performed using prism (prism 5 for windows, graphpad software inc., usa). unless specified, statistical significance for comparison between two sample groups with a normal distribution (shapiro-wilk test for normality) was determined using two-tailed paired or unpaired student's t-test. differences were considered significant at p < 0.05 as indicated. except when specified, only significant differences are shown. as indicated in figure legends, either a representative experiment or a pool is shown, and the number of repetitions of each experiment and number of experimental units (either cultures or mice) is indicated. the results are presented as the means ± standard error (sem). several studies have shown that women have better survival and tolerance to sepsis than men (26, 27) . thus, we further verified this phenomenon by establishing a sepsis model in mice ( figure 1a) . the difference in lung injury between male and female mice in sepsis was determined by h&e analysis of lung sections (figure 1ba) . the histochemical scores (figure 1bb ) indicated that male mice had more severe lung damage than female mice; trained immunity prevented lung injury in mice and the protective effect of trained immunity was better for females than males. notably, we observed a decreased renal e. coli burden in females than males. in addition, in trained immunity mice, females had less renal e. coli burden than males (figures 1c,d) . in addition, liver damage markers, aspartate aminotransferase (ast), and alanine aminotransferase (alt) were increased in males than females; and trained immunity markedly decreased ast and alt concentration, with a greater decrease in females than in males (figures 1e,f) . we found that in the sepsis model, the serum lactate concentration of the male mice was higher than that of the female mice. in the trained immunity group, serum lactate was decreased, with the level in the females lower than males ( figure 1g ). as expected, male serum e 2 content is much lower than female mice ( figure 1h) . our results showed that females expressed higher il-6 and tnfα than males in sepsis, and trained immunity exacerbated this trend (figures 1i,j) . macrophages are the most significant cells in the body in regard to trained immunity (28) . the promoted trained immunity is related to the increased m1 phenotype macrophage. in order to understand the different response to sepsis between the different genders, we further investigated the polarization of macrophages during the process. thus, we focused on macrophage content and polarization. by detecting the percentage of macrophages in the spleen, we found that macrophages in female mice are more abundant than male mice, and trained immunity significantly increased the content of female macrophages (figure 2a) . sepsis promoted the polarization of macrophages to m1 and trained immunity aggravated this trend. notably, females showed more changes than males (figure 2b) . on the other hand, females inhibited the polarization of m2 macrophages ( figure 2c) . trained immunity simulated m1type polarization of macrophages, and this effect was more pronounced for females. by analyzing the experiments above, we found that the difference between male and female responses to sepsis was due to the difference in macrophage polarization in vivo, and the difference in estrogen content in the different genders was very significant. to verify that differences in the estrogen level play a vital role, ovx mice were made, since the ovary is the main organ to produce e 2 . the same sepsis model was established with ovx mice (figure 1a) . firstly, the serum e 2 concentration was significantly decreased (figure 3a) . the degree of lung injury was much higher than that of female mice as well as ti + e. coli group (figure 3b) . the kidney e. coli burden was also evaluated. the results showed that ovx mice also had more severe kidney injury ( figure 3c ) as well as lung injury, as measured by serum alt (figure 3d ) and ast ( figure 3e ) concentrations, in comparison to female mice. with a lower serum il-6 and tnfα concentration in ovx mice in the ti + e. coli group, the data suggested that ovx mice have a lower trained immunity response than female mice (figures 3f,g) . finally, similar to male mice, ovx mouse pbmcs began polarizing to m2-type at 24 h after i.p. e. coli ( figure 3h ). taken together, these results demonstrated that e 2 does facilitate trained immunity in the body to resist sepsis, and simultaneously inhibits macrophage polarization to m2. since the macrophages are mostly derived from bone marrow cells, and also the cytokines up-regulated in trained immunity are pro-inflammatory cytokines. although the upregulation of inflammatory cytokines after trained immunity of macrophages is determined, no research has studied the polarization of macrophages during this process. therefore, exploring the polarization of bmdm as an important primary macrophage in trained immunity is very significant and necessary to understand the nature of trained immunity. to determine whether bmdms are similar to macrophage polarization in pbmcs, we tested the polarization of microphages from male or female mice when challenged with lps or trained immunity plus lps. we considered f4/80 + inos + bmdms as m1-type bmdms and f4/80 + cd206 + bmdms as m2-type bmdms. the results demonstrated that female bmdms are more polarized to m1 in both the lps group (7.04-5.12%) and ti + lps group (28.8-17.9%) ( figure 4a) . also, male bmdms began to transform to m2 in 24 h, which did not occur in female bmdms. trained immunity also prevented both male and female bmdms from m2 polarization as well as pbmcs (figure 4b ). macrophage cell lines j774 and raw264.7 in order to examine the effect of estradiol on trained immunity, we decided to pre-treat the macrophage cell lines with estradiol before trained immunity model. since the concentration of estradiol that can stimulate the macrophage cell lines raw264.7 and j774 is unknown, we made a concentration gradient of estradiol on the activity of the macrophage cell lines. the results showed that the response of the macrophage cell lines to the stimulation of estradiol is not particularly sensitive, and the viability of j774 begins to increase significantly under the stimulation of 50 nm estradiol. thus, the concentration of e 2 used in cell experiments was verified as 50 nm (figures 5a,b) . the in vitro trained immunity model was established with raw264.7 and j774 ( figure 5c ) cell lines derived from male and female mice, respectively. using tnfα and il-1β as the marker of inflammation, our data suggested that e 2 can induce stronger trained immunity both in raw264.7 and j774 (figures 5d,e) . we also made a trained immunity model of bmdms (figure 6a ). m-csf was used to induce bmdm with a purity of over 95% from both male and female mice ( figure 6b) . then, the mrna levels ( figure 6c ) and protein levels ( figure 6d ) of tnfα and il-6 were measured. the results suggested that female bmdms can have a more intensive response to lps as well as a trained immunity response than male bmdms. phosphorylation of akt and mtor targets (s6 and 4ebp1) are considered to be key hallmarks of trained immunity (23) . to clarify the effect of e 2 on trained immunity, these hallmarks are checked by western blot, which indicated that e 2 as well as β-glucan induced trained immunity in bmdms ( figure 6e ). in addition, the data showed that e 2 further boosted the polarization of m1-type macrophage in trained immunity. consistently, this effect of e 2 is more apparent in female bmdms, and e 2 inhibited m2 polarization in trained immunity (figure 6f ). nfκb signaling pathway is involved in the regulation of many cellular physiological processes. previous articles have reported that relb −/− mice, or inhibiting the nucleus translocation of relb, led to a higher basal inflammatory level (29, 30) . other reviews declared that e 2 could regulate nuclear translocation of nfκb through ers. since it has been widely studied that tnfα can combine with tnfr on the cell membrane surface to promote nfκb translocate into the nucleus. also, because tnfα as a representative inflammatory cytokine is secreted by macrophages and will stimulate back to macrophage. we use tnfα as a positive control to study nfκb entry into the nucleus. we use immunofluorescence to explore the intracellular localization of nfκb protein. in immunofluorescence, the results indicated that when tnfα is used to stimulate cells, it does not affect the colocalization of erα and the nucleus but increases the colocalization area of p65 and relb with the nucleus. on the contrary, e 2 treatment appeared to induce nuclear positioning of erα and kept relb distribution in the cytoplasm in both hek293t and bmdms (figures 7a,b) . however, stimulation of e 2 did not affect the distribution of p65 in cells. next, we used the rna interference system to further verify the impact of e 2 on the nuclear translocation of relb. the silencing performance of interfering small rna sequence was evaluated both at the mrna level ( figure 7c ) and protein level ( figure 7d) . by extracting nuclear protein, we found that estrogen could reduce the content of relb protein in the nucleus; by silencing the expression of erα, the content of relb protein in the nucleus was also restored ( figure 7e ). as markers of m2 macrophage polarization, il-4 and il-10 are also the downstream genes regulated by relb (30, 31) . we found that e 2 inhibited the mrna expression level of il-10 and il-4 by regulating nuclear translocation of relb ( figure 7f ). in conclusion, these results indicated that e 2 could block relb translocation to the nucleus; to further inhibit m2-associated gene expression, e 2 suppressed bmdms m2 polarization. during the development of sepsis, many systemic organ abnormalities occur including damage to the lung, liver and kidney; the normal operation of the body's neurosensory system may even be affected (32) . since lps is a component in the cell wall of gram-negative bacteria, it is wildly used as an ideal stimulus to establish sepsis models. when tlr4 on figure 4 | female bmdms are apt to be polarized to m1 phenotype in responses to trained immunity in vitro. (a) female bmdms m1 polarization was stronger than male bmdms treated with lps or ti + lps groups. (b) very few female bmdms to polarized to m2 in lps group, while male bmdms showed the polarization of m2 phenotype. ti suppressed bmdms to m2 polarization from both male and female mice (n ≥ 3/group). ***p < 0.001, paired student's t-test comparing lps group and ti + lps group in the same gender. # p < 0.05, ### p < 0.001, paired student's t-test comparing same group between different genders. the macrophage cell membrane recognizes lps, a series of signaling pathways will be initiated to stimulate macrophages to produce pro-inflammatory cytokines, which can induce potent systemic inflammatory responses in vivo. therefore, tnfα, il-1β, and il-6 are currently considered to be markers of the early phase of sepsis (33) . the development of sepsis begins with systemic inflammatory response syndrome (sirs) (34) , which is considered as the early phase of sepsis. elevated expression of pro-inflammatory cytokines could help the body return to a normal state in early sepsis (35) . after sirs, the body enters the cars (compensatory anti-inflammatory response syndrome) stage. in the cars stage, the immune system will undergo the decline-regulation (36) . the hyper-inflammatory response of sirs will normally be restored by the subsequent cars stage. the patients can return to normal state after the cars stage (37) . however, if the cars phase fails, it can cause death from sepsis. as one of the important ways to protect the body from foreign microbes and antigens, the role of trained immunity is very significant. this study focused on the effects of e 2 on early sepsis by promoting trained immunity. in the development of early sepsis, estrogen induced a stronger trained immunity response of macrophages to remove antigens and microorganisms from the body. simultaneously, estrogen promoted the polarization of macrophages to m1 in vivo and inhibited the polarization of m2, which is considered anti-inflammatory in the early phase of sepsis. in summary, estrogen can maintain the higher proinflammatory state of macrophages. this also illustrates the reason why females are more tolerant to sepsis than males in one aspect. however, further research is needed to understand the role of estrogen in the later stages of sepsis. the mrna levels of tnfα and il-1β in raw264.7 were detected by qpcr to determine the trained immunity effect with or without e 2 (n ≥ 3/group). # p < 0.05, ### p < 0.001, paired student's t-test comparing β-glucan + lps group and lps group. *p < 0.05, ***p < 0.001, paired student's t-test comparing with control group. ∧p < 0.05, ∧∧p < 0.01, and ∧ ∧ ∧p < 0.001, paired student's t-test comparing between β-glucan + lps groups with or without e 2 . here, we demonstrated that β-glucan-induced trained immunity was more resistant to sepsis in female than male mice. ovx mice manifested serious sepsis consequences with a weaker trained immunity effect than female mice. macrophage polarization toward m1 phenotype is thought to exhibit the enhanced trained immunity. e 2 promoted trained immunity in vitro and in vivo. inhibition of e2 on the nucleus translocation of relb contributed to macrophage polarization to change the intensity of trained immunity. our results indicated that e 2 is involved in the trained immunity in gender differences. since trained immunity was discovered in 2014, although there are many articles about trained immunity, there is no article to find the fundamental way of trained immunity activation but only about the changes of immune cell characterization after trained immunity activation. such as epigenetic changes, upregulation of glycolysis, autophagy level, akt phosphorylation, and so on. in this study, we found that e 2 promotes trained immunity by suppressing relb entry into nucleus, while inhibiting macrophage polarization to m2 in inflammation (a way reverse the inflammatory response). it is possible that e 2 can change the epigenetics of macrophages through estrogen receptors to affect the trained immunity. this hypothesis needs further researches. trained immunity is a way of immunization that occurs in the innate immune cells through the first stimulation to fight the later secondary infections. by obtaining trained immunity, the body can fight various infections from bacteria, fungi and even viruses timely. it is worth mentioning that in countries and regions with underdeveloped scientific research capabilities, it may be an economical and quick means of protection against covid-19 to enhance people's trained immunity ability. sepsis is a clinically common infectious disease with obvious gender differences. in this article, sepsis model is used as an infection model to evaluate the role of estradiol in enhancing trained immunity. the mechanism and effects of estradiol in promoting trained immunity also need subsequent study in other types of infectious diseases. the purpose of this paper was to explore the reasons why different genders have different tolerance effects on sepsis. an article reported that estradiol and dht (dihydrotestosterone) did not influence bcg-induced trained immunity, and the article only focused on the inflammatory cytokines of monocytes induced by bcg in vitro (22) . apart from this, almost no articles study the effects of e 2 on trained immunity and its biological significance. our research attempted to explain its effects on trained immunity and understand how it causes women to be more tolerant of sepsis than men. however, in terms of gender differences, many factors, such as androgen concentration, genetic differences, metabolic differences, and physiological differences may all be the result of different tolerances for sepsis between genders (12, 38, 39) . gender differences are a very complex and exquisite scientific issue (40) , and all of the factors that contribute to gender differences are worth exploring for subsequent experiments. in summary, we demonstrated that as one of the key factors for gender differences, e 2 plays a vital role in the stronger tolerance of females to sepsis than males. one the one hand, e 2 can better facilitate trained immunity by increasing expression of inflammatory cytokines and inducing macrophage m1-type polarization; on the other hand, e 2 can inhibit or delay the macrophage m2-type polarization by regulating nucleus translocation of relb and its down-stream m2-associated genes. thus, this may provide a new idea for the clinical treatment of sepsis. drugs with a structure similar to estrogen may be developed to treat sepsis. more importantly, the treatment of some diseases may able to find better solutions by addressing the different characteristics of different genders. the datasets generated for this study are available on request to the corresponding author. the animal study was reviewed and approved by model animal research center of nanjing university. all authors reviewed the manuscript, contributed to data analysis, drafting and revising the article, gave 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and transcriptional activity are regulated by p100 year in review in intensive care medicine, 2008. i. brain injury and neurology, renal failure and endocrinology, metabolism and nutrition, sepsis, infections and pneumonia sepsis and septic shock: a review sepsis: a review of advances in management review: immunology of sinusitis, trauma, asthma, and sepsis review on sepsis in children did not mention important trial clinical outcome and risk factors of neonatal sepsis among neonates in felege hiwot referral hospital systematic review of gender differences in sepsis management and outcomes the influence of gender on the epidemiology of and outcome from severe sepsis age and sex influence the hippocampal response and recovery following sepsis the authors appreciate the warm-hearted help from all the co-authors and the professor. key: cord-261633-r4qlbnc5 authors: xie, guo-hao; chen, qi-xing; cheng, bao-li; fang, xiang-ming title: defensins and sepsis date: 2014-08-19 journal: biomed res int doi: 10.1155/2014/180109 sha: doc_id: 261633 cord_uid: r4qlbnc5 sepsis is a leading cause of mortality and morbidity in the critical illness. multiple immune inflammatory processes take part in the pathogenesis of sepsis. defensins are endogenous antimicrobial peptides with three disulphide bonds created by six cysteine residues. besides the intrinsic microbicidal properties, defensins are active players which modulate both innate and adaptive immunity against various infections. defensins can recruit neutrophils, enhance phagocytosis, chemoattract t cells and dendritic cells, promote complement activation, and induce il-1β production and pyrotosis. previous publications have documented that defensins play important roles in a series of immune inflammatory diseases including sepsis. this review aims to briefly summarize in vitro, in vivo, and genetic studies on defensins' effects as well as corresponding mechanisms within sepsis and highlights their promising findings which may be potential targets in future therapies of sepsis. sepsis, severe sepsis, and septic shock represent a continuum of clinical syndromes which are common complications observed in patients with infection, trauma, and major surgeries [1] [2] [3] . these syndromes start with infection induced systemic inflammatory response syndrome (sirs) and evolve to sepsis induced acute organ dysfunction and cardiovascular collapse. epidemiology studies demonstrated that severe sepsis has a population prevalence of 300/100 000 in the united states and counts for 10-30% of the intensive care unit (icu) patients [4] [5] [6] . and severe sepsis has already been acknowledged as the first cause of death in noncoronary icus with a high mortality rate of approximately 25-50% [7] . in the past one or two decades, steady progresses in treatment of sepsis have been made due to the advanced supportive care in icu and the implementation of bundle therapies [7] . however, searching for specific remedies and reliable predictors within the pathophysiological mechanisms of sepsis is still the emphasis of today's studies [8, 9] . defensins are classified as a subfamily of cationic antimicrobial peptides, which are major components of the human innate immunity. they are small endogenous peptides with three disulphide bonds created by six cysteine residues. defensins are categorized into three subtypes, -, -, and -defensin, based on the spatial structure and the locations of three disulphide bonds within the peptide. in the past decade, cumulative evidences have suggested that defensins play an important role and may be a potential intervention target in sepsis. this review hereby will summarize in vitro, in vivo, and genetic studies on defensins' effects as well as corresponding mechanisms within sepsis and its sequential syndromes. defensins have broad spectrum antimicrobial activities against most pathogens in sepsis. [10] [11] [12] . they can inhibit a large variety of gram-positive bacteria, gram-negative bacteria, and some species of fungi and viruses [11] . the -defensins are mainly distributed in the epithelial cells of the respiratory system, digestive system, and genitourinary system [10] [11] [12] . they can effectively kill a number of gram-negative bacteria, such as e. coli and p. aeruginosa, gram-positive bacteria, such as s. aureus and streptococcus pyogenes, and candida albicans. -defensin-3 even has bactericidal effect towards multiresistant s. aureus and vancomycin-resistant enterococcus [11, 13] . the -defensins, which have a unique macrocyclic structure, are isolated from leukocytes from some species of monkey and have not been detected in humans [14] . they are also reported to have antimicrobial activity against a spectrum of pathogens including e. coli, s. aureus, and c. albicans [15] . also, they are found to have protective effect in a mouse model from a lethal pulmonary infection by a mouse adapted strain of sars-coronavirus [16] . the classic mechanism of defensins' bactericidal effect is the "pore formation" theory. these positively charged antimicrobial peptides target negatively charged bacterial membrane components, such as lipopolysaccharides, teichoic acids, or phospholipids. then they form transmembrane pores, disrupt cell integrity, and lead to bacteria lysis [10, 11] . recently, another mechanism has been reported that defensins kill bacteria by inhibiting the synthesis of bacterial cell wall through interaction with certain precursors such as lipid ii [17] . defensins' bactericidal effect can be limited by high salt concentration of local environment where they encounter with the pathogens [18, 19] . also, the antimicrobial action appears to be regulated by the redox response, as -defensin-1 become more potent after reduction of disulfide bridges by thioredoxin or a reducing environment [20, 21] . defensins are also reported to have modulating effects on both innate and adaptive immune response. it is well known that hnp1-3 participate in the host immune defense via multiple mechanisms, including enhancing macrophage phagocytosis, facilitating neutrophil recruitment, modulating complement activation, and chemoattracting immature t cells and dendritic cells [12, 22] . in vitro studies showed that -defensins have potent chemotactic effects, leading to the recruitment and maturation of naive dendritic cells and memory t cells in the inflammatory sites and the triggering of specific immune response in the host [23] . as the endogenous ligand of tlr-4, -defensins interact with tlr-4 of the immune cells and regulate the expression of inflammatory mediators via the nf-b pathway [18] . in vivo researches have revealed that the abnormal expression of -defensins is associated with sepsis and various infectious diseases, as levels ofdefensins in both plasma and bronchoalveolar lavage fluid in patients with pulmonary infections are elevated [24] [25] [26] , transcription of -defensin-2 in leukocytes of severe septic patients is suppressed [27] , expression of -defensins in burn wound is reduced [28] , and impaired expression of -defensins is associated with inflammatory bowel diseases [29, 30] . in a mouse model of acute lung injury, shu et al. expressed recombinant -defensin-2 in lung tissue via recombinant adenovirus to study its protective effect against p. aeruginosa infection. compared with control mice, they found considerably less p. aeruginosa in the transinfected lung tissue, as well as alleviated alveolar impairment, interstitial edema, and neutrophil infiltration [31, 32] . in subsequent studies, mice transinfected by adenovirus with or withoutdefensin-2 genes received cecal ligation and puncture (clp) twice to generate sepsis models. the impact of -defensin-2 on the inflammatory response (e.g., the level of icam-1 expression), the severity of lung injury, and the sepsis outcome (7-day survival rate) were observed and evaluated. it was found that recombinant -defensin-2 could downregulate the expression of icam-1 in lung tissue 24 h, 36 h, and 72 h after clp and significantly raised the 7-day survival rate in sepsis mice [31, 33] . in the clinical setting, olbrich et al. found preterm neonates had lower level of -defensin-2 in cord blood when compared to term neonates [34] . and among these preterm neonates, lower -defensin-2 level was associated with late-onset sepsis. these studies indicate that -defensin-2 may play an important role in the immune inflammatory response in sepsis and might influence the outcome of sepsis. among the -defensins, rhesus macaque -defensin (rtd), which has six subtypes, has been extensively studied. though not expressed in humans, rtds were reported to significantly reduce levels of tnf-, il-1 , il-6, il-8, mip1, and so on, in human peripheral blood leukocytes that are preincubated with various toll-like receptor agonists [35] . furthermore, in vivo study showed that subcutaneously administration of 5 mg/kg rtd-1 could improve the survival rate and suppress the levels of a number of inflammatory cytokines and chemokines in two sepsis mouse models (received either intraperitoneal injection of e. coli or clp). although detailed mechanisms of the protective effect of rtd-1 have not been illuminated, the authors suggested that the interaction between rtd-1 and leukocyte is the critical determinant of tnf-blockade [35] . the latter is a major proinflammatory cytokine and influences the consequent inflammatory cascades. these results indicate that -defensins may be a potential immune adjuvant in the treatment of sepsis, though they are not expressed in human. in sepsis and other inflammatory disorders, defensins are among a group of rapidly-released host endogenous molecules, which are capable of both recruiting and activating apcs and are also termed the alarmins. recently, in vitro studies have shown that alarmin hnp1-3 have the ability to boost host inflammatory response by promoting macrophage il-1 production and pyroptosis via purinergic p2x7 receptor [36] . however, this effect is a double-edged sword in sepsis since it can promote pathogen elimination as well as mediate organ dysfunction such as acute lung injury [22, 37] . in molecular genetics and molecular biology, knock-out animal model is one of the most convincing means to determine the role of a specific molecule in the physiopathology of a certain disease. however, as members of the defensin family have overlapped biological functions, the function of the knock-out gene in animal models may probably be compensated by other defensins. since the gene cluster coding for the entire defensin family cannot be fully knocked out using the present techniques of molecular biology and genetics as well as human defensins lack of absolute animal analogues, genetic association analysis is a good alternative that can effectively explore the relationship between genetic polymorphism and sepsis. in normal peripheral blood cells, mrna levels of both -defensin-1 and -defensin-2 raise remarkably when stimulated by lps or p. aeruginosa [23] . however, the upregulation of -defensin-1 and -defensin-2 varies among individuals, resulting in interindividual differences in host defense capacity and hence influencing the clinical progression of sepsis. previous studies showed that single nucleotide polymorphism (snp) of -defensin-1 gene (defb1) correlates with chronic obstructive pulmonary disease, asthma, genetic allergy, hiv infection, and pseudomonas species infection in oral mucosa [38] [39] [40] [41] [42] . since sepsis is a multifactorial disease caused by both environmental factors (pathogenic microbes) and host factors (comorbidities and genetic background), its occurrence and outcome are influenced with individual genetic background [43] . chen et al. selected 5 snps in the promote region of defb-1 (-1816a/g, -390a/t, -52a/g, -44c/g, and -20a/g) and one in its extron (1654g/a) as candidate loci and studied 211 patients with severe sepsis and 157 healthy controls [44] . distribution of alleles, gene types, and haplotypes associating with these loci were studied and compared between septic patients and controls, as well as between survivals and victims of severe sepsis. association analysis, logistic regression, and linkage disequilibrium study showed that -44g allele was closely related with susceptibility to severe sepsis and poor outcome. and severe septic patients with haplotype -20g/-44g/-52g had even poorer outcome, while individuals with haplotype -20a/-44c/-52g were less susceptible to severe sepsis. the reason why -44c/g is correlated with the occurrence and outcome of severe sepsis may attribute to the following points. it located in the 5 untranslated region of defb1 and its polymorphism may result in changes in the space conformation of mrna, which would alter the stability of mrna and the efficiency of translation. and its impact on the protein function is more significant than nonsynonymous snp in coding region [45] , as the quantity of protein would change dramatically. however, as any other genetic association analysis, defb1 −44c/g may be only a surface marker of some unknown real genetic marker of sepsis in linkage disequilibrium. although these hypothesis need to be proved by further researches, the above-mentioned study indicated that -defensin-1 might be an influential factor in the process of immune defense and inflammation regulation in sepsis, and the locus of −44c/g may be an important genetic warning indicator of susceptibility to severe sepsis and its outcomes. copy number variation (cnv) is a kind of genetic polymorphism that accounts for approximately 12% of human genomic dna. it refers to a large-scale duplication or deletion of certain dna sections, which causes a variation in the number of copies of one or more genes. previous publications reported that cnv is present in -defensin-2 gene (defb4), -defensin-3 gene (defb103), -defensin-4 gene (defb104), -defensin-1 gene (defa1), and -defensin-3 gene (defa3) [18, [46] [47] [48] . and copy number of defb4 has a positive correlation with its mrna level [35, 45] . recently, chen et al. screened 179 severe sepsis and 233 healthy controls for defa1 and defa3 [49] . an average defa1/defa3 copy number of 7 per genome was observed in the studied population, with a range of 2 to 15. the authors found that patients with high copy number of defa1/defa3 were predisposed to severe sepsis and tended to have lower level of plasma hnp1-3 as well as cytokines such as tnf-, il-6, and il-10. they further validated their findings in an independent cohort. these results indicated that cnvs in the defensin gene may be potential genetic markers for identifying high risk patients or providing individual treatment in sepsis. defensins are emerging therapeutic molecules against pathogens in sepsis because of their broad spectrum antimicrobial properties. in the past decade or two, a number of potent and salt insensitive defensins and their analogs have been screened, structurally modified, and synthesized. however, most of these studies are performed in vitro and not much is known about the in vivo roles of these molecules. in fact, chemoattracting and immunomodulating effects make defensins a double-edged sword in the pathogenesis of sepsis, which leads to facilitation of pathogen clearance as well as exacerbation of inflammation and injury of self-tissues. recently, several investigations showed that the chemoattractant and antimicrobial activities of defensins could be separated, which shed light on the design of defensin-derived pharmaceuticals [50] . in addition, genetic studies help identify high risk patients with susceptibility to sepsis or its adverse outcome, which provides foundation for future individualized sepsis treatments that are targeting defensins. the pathophysiology 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neutrophil granule microbicidal activities rhesus theta-defensin prevents death in a mouse model of severe acute respiratory syndrome coronavirus pulmonary disease turning defensin mimetics novel antibiotics targeting lipid ii defensins: antimicrobial peptides of innate immunity isolation and characterization of human -defensin-3, a novel human inducible peptide antibiotic reduction of disulphide bonds unmasks potent antimicrobial activity of human 2-defensin 1 cellmediated reduction of human -defensin 1: major role for mucosal thioredoxin amped up immunity: how antimicrobial peptides have multiple roles in immune defense multiple roles of antimicrobial defensins, cathelicidins, and eosinophil-derived neurotoxin in host defense increased levels of antimicrobial peptides in tracheal aspirates of newborn infants during infection elevated balf concentrations of -and -defensins in patients with pulmonary alveolar proteinosis expression and activity of -defensins and ll-37 in the developing human lung inducibility of the endogenous antibiotic peptide -defensin 2 is impaired in patients with severe sepsis reduced antimicrobial peptide expression in human burn wounds a chromosome 8 gene-cluster polymorphism with low human geta-defensin 2 gene copy number predisposes to crohn disease of the colon hammarström, " -defensin-3 and -4 in intestinal epithelial cells display increased mrna expression in ulcerative colitis protection against pseudomonas aeruginosa pneumonia and sepsis-induced lung injury by overexpression of -defensin-2 in rats the protective effect of beta-defensin 2 in acute lung injury induced by respiratory pseudomonas aeruginosa infection in rats the effect of pretreatment of recombinant beta-defensin 2 on the icam-1 expression in lung of rats with acute lung injury association of human beta-defensin-2 serum levels and sepsis in preterm neonates rhesus macaque theta defensins suppress inflammatory cytokines and enhance survival in mouse models of bacteremic sepsis alarmin hnp-1 promotes pyroptosis and il-1 release through different roles of nlrp3 inflammasome via p2x7 in lps-primed macrophages neutrophil alpha-defensins cause lung injury by disrupting the capillaryepithelial barrier genetic variants of human -defensin-1 and chronic obstructive pulmonary disease association of defensin beta-1 gene polymorphism with asthma a single-nucleotide polymorphism in the human beta-defensin 1 gene as associated with hiv-1 infection in italian children single-nucleotide polymorphisms (snps) in humandefensin 1: high-throughput snp assays and association with candida carriage in type i diabetics and nondiabetic controls contribution of alpha-and beta-defensins to lung function decline and infection in smokers: an association study genomic polymorphisms in sepsis genomic variations within defb1 are associated with the susceptibility to and the fatal outcome of severe sepsis in chinese han population human catechol-o-methyltransferase haplotypes modulate protein expression by altering mrna secondary structure extensive normal copy number variation of a -defensin antimicrobialgene cluster copy number polymorphism and expression level variation of the humandefensin genes defa1 and defa3 screening of copy number polymorphisms in human -defensin genes using modified real-time quantitative pcr increased genomic copy number of defa1/defa3 is associated with susceptibility to severe sepsis in chinese han population analysis and separation of residues important for the chemoattractant and antimicrobial activities of beta-defensin 3 the authors declare that there is no conflict of interests regarding the publication of this paper. this work was supported by the national natural science foundation of china (no. 81201495) and the research project of the department of education of zhejiang province (no. y200909678). key: cord-005603-kjcbbgse authors: brun-buisson, c. title: the epidemiology of the systemic inflammatory response date: 2000 journal: intensive care med doi: 10.1007/s001340051121 sha: doc_id: 5603 cord_uid: kjcbbgse objective: to examine the incidence, risk factors, aetiologies and outcome of the various forms of the septic syndromes (the systemic inflammatory response syndrome [sirs] sepsis, severe sepsis, and septic shock) and their relationships with infection.¶design: review of published cohort studies examining the epidemiology of the septic syndromes, with emphasis on intensive care unit (icu) patients.¶results: the prevalence of sirs is very high, affecting one-third of all in-hospital patients, and > 50 % of all icu patients; in surgical icu patients, sirs occurs in > 80 % patients. trauma patients are at particularly high risk of sirs, and most these patients do not have infection documented. the prevalence of infection and bacteraemia increases with the number of sirs criteria met, and with increasing severity of the septic syndromes. about one-third of patients with sirs have or evolve to sepsis. sepsis may occur in approximately 25 % of icu patients, and bacteraemic sepsis in 10 %. in such patients, sepsis evolves to severe sepsis in > 50 % of cases, whereas evolution to severe sepsis in non-icu patients is about 25 %. severe sepsis and septic shock occur in 2 %–3 % of ward patients and 10 %–15 % or more icu patients, depending on the case-mix; 25 % of patients with severe sepsis have shock. there is a graded severity from sirs to sepsis, severe sepsis and septic shock, with an associated 28-d mortality of approximately 10 %, 20 %, 20 %–40 %, and 40 %–60 %, respectively. mortality rates are similar within each stage, whether infection is documented or not, and microbiological characteristics of infection do not substantially influence outcome, although the source of infection does. while about three of four deaths occur during the first months after sepsis, the septic syndromes significantly impact on long-term outcome, with an estimated 50 % reduction of life expectancy over the following five years. the major determinants of outcome, both short-term and long-term, of patients with sepsis are the severity of underlying diseases and comorbidities, the presence of shock and organ failures at onset of sepsis or evolving thereafter. it has been estimated that two-thirds of the overall mortality can be attributed to sepsis.¶conclusions: the prevalence of sepsis in icu patients is very high, and most patients have clinically or microbiologically documented infection, except in specific subset of patients. the prognosis of septic syndromes is related to underlying diseases and the severity of the inflammatory response and its sequelae, reflected in shock and organ dysfunction/failures. abstract objective: to examine the incidence, risk factors, aetiologies and outcome of the various forms of the septic syndromes (the systemic inflammatory response syndrome [sirs] sepsis, severe sepsis, and septic shock) and their relationships with infection. design: review of published cohort studies examining the epidemiology of the septic syndromes, with emphasis on intensive care unit (icu) patients. results: the prevalence of sirs is very high, affecting one-third of all in-hospital patients, and > 50 % of all icu patients; in surgical icu patients, sirs occurs in > 80 % patients. trauma patients are at particularly high risk of sirs, and most these patients do not have infection documented. the prevalence of infection and bacteraemia increases with the number of sirs criteria met, and with increasing severity of the septic syndromes. about onethird of patients with sirs have or evolve to sepsis. sepsis may occur in approximately 25 % of icu patients, and bacteraemic sepsis in 10 %. in such patients, sepsis evolves to severe sepsis in > 50 % of cases, whereas evolution to severe sepsis in non-icu patients is about 25 %. severe sepsis and septic shock occur in 2 %±3 % of ward patients and 10 %±15 % or more icu patients, depending on the case-mix; 25 % of patients with severe sepsis have shock. there is a graded severity from sirs to sepsis, severe sepsis and septic shock, with an associated 28-d mortality of approximately 10 %, 20 %, 20 %±40 %, and 40 %±60 %, respectively. mortality rates are similar within each stage, whether infection is documented or not, and microbiological characteristics of infection do not substantially influence outcome, although the source of infection does. while about three of four deaths occur during the first months after sepsis, the septic syndromes significantly impact on long-term outcome, with an estimated 50 % reduction of life expectancy over the following five years. the major determinants of outcome, both short-term and long-term, of patients with sepsis are the severity of underlying diseases and comorbidities, the presence of shock and organ failures at onset of sepsis or evolving thereafter. it has been estimated that two-thirds of the overall mortality can be attributed to sepsis. conclusions: the prevalence of sepsis in icu patients is very high, and most patients have clinically or microbiologically documented infection, except in specific subset of patients. the prognosis of septic syndromes is related to underlying diseases and the severity of the inflammatory response and its sequelae, reflected in shock and organ dysfunction/failures. similarly to ards [1] the definition of septic syndromes has caused much controversy and debate in the past decade. many of these controversies have stemmed from the frustration accumulated following the repeatedly negative results of new therapeutic interventions aiming at controlling the inflammatory response associated with infection. hence the suggestions that new definitions were needed, that would allow a quicker and simpler identification of septic patients for accrual into randomised trials of new therapies [2] and that would help derive more consistent and comparable results from epidemiological studies and clinical trials. while the ultimate goal of showing the efficacy of these pharmacological interventions remains elusive, the definitions elaborated then and now in widespread use did provide the impetus for conducting several epidemiological studies aimed at better characterising the septic syndromes and their sequelae. in this paper, we shall review these studies and summarise the current understanding of the clinical and microbiological epidemiology of the septic syndromes, their interplay, and outcomes of patients affected. finally, we shall discuss the implications of this information for conducting clinical trials. the term ªsepsisº has long been used interchangeably with bacteraemia, severe sepsis or even septic shock, undoubtedly a source of some confusion and difficulty in putting together results from published studies. in 1992 the us expert panel from the american college of chest physicians and the society of critical care medicine [2] produced a consensus statement on the suggested definitions to characterise the various stages of the associated inflammatory response and help in differentiating infectious from non-infectious processes. while the recent definitions are centred on the documentation of infection, they aim at encompassing all potential clinical presentations of infection and its consequences. the principles followed in elaborating the definitions were that: (1) infectious (and some non-infectious) processes, whatever their cause, elicit a common systemic response which, although of variable intensity, is the expression of common pathophysiologic pathways resulting from the expression and interaction of various humoral and cellular mediators and cytokines; (2) sepsis and related terms should be reserved for infectious processes; and (3) there is a continuum between the various stages of this response to infection (table 1) . although the definitions do provide a framework for classifying patients ± a useful achievement for enrolling patients into clinical trials ± a persisting and unresolved problem facing clinicians in clinical practice is that the definitions are in part retrospective (based on the documentation of infection) and do not actually help them solve the major clinical issue when faced with a septic patient, which is to differentiate infectious from non-infectious processes. another critique of this classification has been that its broad-based approach, intended to identify patients early in the course of the infectious process, did not in fact help physicians, and especially intensivists, to better characterise patients exhibiting the least severe presentations of the septic syndromes. in other words, the high sensitivity of the definition is counterbalanced by a rather low specificity. finally, even the sensitivity of the definitions has been questioned, as there are unquestionably infected patients that do not meet sepsis criteria. nevertheless, several large epidemiological studies conducted after this conference have contributed to s 65 sepsis-induced hypotension, persisting despite adequate fluid resuscitation, (2, 3) and manifestations of hypoperfusion as listed in 5 7. hypotension, sepsis-induced a decrease in systolic blood pressure to < 90 mm hg, or of > 40 mm hg from baseline, in the absence of other cause for hypotension (3) (1) the sirs may be caused by a variety of insults in addition to infection, including but not limited to trauma and status post-major surgery, acute pancreatitis, and burns (2) an adequate fluid challenge is usually considered as at least 500 ml fluid infused rapidly, and persisting hypotension as one persisting for > 1 hour (3) patients on inotropic/vasoactive agents may not be hypotensive at time of evaluation our better understanding and characterisation of the epidemiology and relationships to infection of the various stages of the inflammatory response, and of their outcome. in reviewing these, we will use definitions recommended by the expert panel [2] and will use the term ªseptic syndromesº to refer to all stages of the inflammatory response to infection. evaluations of the incidence of sepsis have initially focused on the most indisputable evidence for infection, i. e., bacteraemia. the incidence of bacteraemia has been increasing steadily over the years. in 1990, the us national center for health statistics reported that the rate of bacteraemia had increased from 0.74/1,000 to 1.76/1,000 hospital discharges between 1979 to 1987 [3] . much ± if not all ± this change is caused by the increasing importance of nosocomial infection. at one tertiary-care institution, the rate of nosocomial bacteraemia has increased steadily from 6.7/1,000 to 18.4/1,000 discharges between 1980 and 1992 [4] . in the french multi-center study conducted in 1993 in 24 public or public-affiliated hospitals [5] we recorded an overall incidence rate of bacteraemia of 9.8 (95 ci 9.2 to 10.5) per 1,000 admissions; this rate was more than eight-fold higher in icus (69/1,000) than in wards (8.2/1,000) ( table 2) . of the 842 bacteraemic episodes recorded in this study, 19 % occurred in icus, 63 % in medical wards, and 18 % in surgical wards. extrapolating these results to the whole country would give a figure of approximately 67,500 bacteraemic episodes occurring annually, of which 13,500 would occur in icus. so far, the most comprehensive study on the clinical significance of the early stages of the septic syndromes was conducted by rangel-fausto et al. at the university of iowa hospital and clinics [6] . this study was performed in three icus (medical, surgical, and cardiovascular) and 3 wards of a 900-bed teaching hospital including 200 icu beds. the incidence of sirs, sepsis, and severe sepsis and septic shock was assessed during a 9-month period, including a follow-up period of up to 28 days. of the 3708 patients admitted during the study period, s 66 data are reported with 95 % confidence intervals in brackets, when available. na = not available (1) 2-month incidence study in 24 hospitals in france; (2) survey at 8 academic medical centres in the us; (3) incidence study at one academic centre in the netherlands; (4) prevalence survey at one academic medical centre in the us; (5) prevalence survey at n hospitals in italy; (6) incidence of bacteremic sepsis only at one academic medical centre in switzerland a incidence/1,000 admissions; b incidence/1,000 patient-days 2527 (68 %) met at least two criteria for sirs at some point in their hospital stay [6 < ]. the major finding from this study was that medical or surgical icu patients met 2 or more sirs criteria during > 80 % of their unit stay, whereas patients in the cardiovascular icu met such criteria during slightly over one-half of their unit stay, and patients from other wards from 32 % to 67 % of their stay. it should be noted however, that the wards surveyed likely housed a population at unusually high risk of sepsis, as indicated by the two prevalence surveys done to complement the incidence study. in these surveys including all hospital wards, the prevalence of sirs was about twice higher (64 % and 61 %) in the 3 wards participating in the incidence study than that recorded in the 27 other wards (25 % and 27 %) ( table 2 ). the prevalence of sepsis was of 6 %±8 % in the latter wards, and that of severe sepsis and shock was similar in both surveys in the two categories of wards, respectively at 2 %±3 %, and 0 %. of the 2729 episodes of septic syndromes (i. e., at least sirs) recorded in the incidence study, 1541 (56.4 %) were classified as sepsis, 994 (36.4 %) as severe sepsis, and 194 (7.1 %) as septic shock [6] . patients with infection were classified as having culture-proven or culture-negative sepsis. it is noteworthy that less than 50 % of all episodes were microbiologically documented, although this proportion increased from 42 % when patients only met criteria for sirs, to 57 % in patients presenting with shock. importantly, this study confirmed the expected natural progression between the different stages of septic syndromes: 32 % and 36 % of patients having 2 or 3 sirs criteria, respectively developed culture-proven sepsis by day 14, and 45 % of those with 4 criteria subsequently developed sepsis, while 64 % of those with sepsis developed severe sepsis, a median of only 1 day after sepsis; conversely, only 23 % of patients presenting with severe sepsis developed septic shock, and this occurrence was delayed by a median of 28 days after severe sepsis [6, 7] . the authors also noted an increasing prevalence of eventual organ dysfunctions (respiratory, renal, disseminated intravascular coagulation and shock) with increasing number of sirs criteria. of note, 27 % of patients meeting four sirs criteria developed shock at some point in time. although there were some minor differences in risk of organ failures depending on the stage examined, the overall rate of organ failures was similar within each stage (as well as mortality) whether infection was confirmed or not, to the notable exception of acute renal failure, which was more frequent at all stages in the presence of confirmed infection. several conclusions can be drawn from this study: 1. the incidence of sirs is very high in icu patients, and its recognition cannot help in accurately identifying patients who will prove to be infected or those at higher risk of the more severe stages. this is confirmed by the fact that only about one-third to onehalf of patients meeting sirs criteria were subsequently proven to have confirmed (i. e., microbiologically proven) sepsis; the prevalence of infection, however, increases with the number of sirs criteria met. however, this conclusion must be tempered by the fact that many patients with sirs were thought to have infection, and were thus administered empirical antimicrobial therapy, which likely interfered with the documentation of infection; the actual proportion of non-infectious sirs or ªsevere sirsº in this study is unknown. 2. there is indeed a continuum between the different stages of the inflammatory response from sirs to sepsis, severe sepsis and shock. however, only about one-third of patients presenting with sirs have confirmed sepsis and about one-fourth will evolve to severe sepsis. conversely, sepsis (microbiologically confirmed) appears at high risk of evolving rapidly to severe sepsis, as shown by the 64 % proportion of cases subsequently developing severe sepsis, of which one-half will occur within one day of sepsis. 3. whether infection is confirmed or not, the outcomes are similar in terms of organ dysfunctions and mortality, within each corresponding stage (with the possible exception of renal failure). in another large study, sands [10] have evaluated 168 patients with severe trauma during the first 48 hours in the sicu; 95 (56 %) had sirs. when stratifying patients on the presence of multiple organ dysfunction (mods), these authors found that the rate of sirs was much higher in patients with mods (22/27, 81 %) than in patients without (73/136, 54 %). the occurrence of mods appeared related to the severity of injury, the volume of blood and fluid replacement, but not to the presence of infection: infection rates were 9 % and 4 %, respectively, in patients with and without mods. as expected in this particular population, sirs appeared as an extremely frequent and non-specific finding, irrespective of the presence of infection. likewise, muckart and bhangwanjee assessed the incidence of the septic syndromes in penetrating or blunt trauma [11] . of 450 patients followed-up, 399 (88 %) fulfilled sirs criteria: 22 % had sirs only and 14 % sepsis; 14 % had severe sepsis and 8 % severe sirs; and 20 % had septic shock and 9 % non-documented septic shock. documentation of infection was more frequent with penetrating trauma. in the italian multicenter study conducted in 1993±1994 in 99 icus [12] , 52 % of 1101 patients had sirs on admission; at any time during the study, sirs only was recorded in 58 % of patients, sepsis in another 16 %, severe sepsis in 5.5 %, and septic shock in 6 %. overall, 85 % of patients had one of the septic syndromes, of which more than two-thirds were non-microbiologically documented sirs. similarly to the study from iowa [6] the investigators noted that a substantial proportion of patients evolved from an earlier stage on admission to a more severe one: 15 % of patients with sirs evolved to sepsis, but only < 5 % to severe sepsis, while 30 % of patients with sepsis evolved to severe sepsis or shock. severe sepsis and septic shock these syndromes are of much more concern to intensivists than sirs, given their more severe outcome, and the poor specificity (and suboptimal sensitivity) of the latter. a closer view of the overall incidence of these two severe syndromes, which are easier to characterise in the icu, has been provided by two multicenter multi-institutional hospital-wide studies. in the french bacteraemia/sepsis study, including 24 hospitals on the one hand [5] and 170 icus on the other [13] both surveyed during a 2-month period, the overall incidence of severe sepsis and shock (including clinically and microbiologically documented infection) was of 6/1000 of all hospital admissions, but only of 2.9/ 1000 in medical/surgical wards and 119/1000 in icus ( table 2 ). of note, nearly half the episodes were of nosocomial origin. in the parallel larger icu survey, severe sepsis or shock occurred in 9 % icu admissions; 71 % of the 1064 episodes were microbiologically docu-mented. the attack rate was higher in larger ( > 400 beds) than smaller hospitals (10.3 vs. 6.7/1000 admissions). septic shock occurred in 6.3/1000 icu admissions. in the study by sands et al. [8] sepsis was noted in 20 % icu patients and severe sepsis (defined in that study as sepsis + one of seven criteria for organ dysfunction) occurred in 10 % of icu admissions, a figure very close to the rate recorded in france in large hospitals. while nearly 41 % sepsis episodes occurred in non-icu patients, only 24 % episodes of severe sepsis occurred in such patients, and 76 % were recorded in icu patients [14] . the relationship between bacteraemia and sepsis has been specifically studied in the french bacteraemia/sepsis multicenter survey [5] . data from this study indicate that only 19 % bacteraemic episodes occurred in icus, while 81 % occurred in wards, including 63 % in medical wards, and 18 % in surgical wards. sepsis was recorded in 74 % of bacteraemic episodes, severe sepsis in 26 %, of which 60 % were associated with septic shock (16 % of all bacteraemic episodes). however, the prevalence of severe sepsis during bacteraemia was much higher in icus than in medical/surgical wards (65 % vs. 17 % of bacteraemic episodes, p < 0.001), emphasising the much higher risk of organ dysfunction and shock during bacteraemia in critically ill patients. conversely, the prevalence of bacteraemia during severe sepsis was estimated at 38 % by kieft et al. [15] at 43 % in the french multicenter study [5] and at 32.5 % of episodes in the study conducted by sands et al. [ 8] . rangel-frausto et al., recorded bacteraemia in 17 % of patients with sepsis, 25 % of those with severe sepsis, and 69 % of patients with septic shock [6] . in non-icu patients, the prevalence of bacteraemia during severe sepsis was estimated at 48 %, compared to 38 % in icu patients [13] . likewise, sands et al., found a prevalence of bacteraemia during severe sepsis of 50 % in non-icu patients and of 26 % in icu patients [8, 14] . these data indicate that, although icu patients are at much higher risk of severe sepsis than ward patients, bacteraemic severe sepsis is proportionally less often encountered in icu than in non-icu patients. factors associated with bacteraemia at the onset of sepsis have been examined in the study conducted by sands et al. [14] . independent predictors of bacteraemia during sepsis were: a suspected or documented focal infection, absence of antibiotic therapy, presence of liver disease, of a hickman catheter, altered mental status, and focal abdominal signs. infection caused by staphylococci were associated with hemodialysis and mechanical ventilation, while gram-negative infection were associated with the use of tpn, the absence of antibiotic therapy, the presence of a hickman catheter, of focal abdominal signs, and of chills [14] . the prediction rules derived from these data performed reasonably well, although the rates of bacteraemia in the highest-risk groups varied from only 20 % to 60 %, depending on the source and micro-organisms involved. host factors identified (by multivariate analysis) as independently associated with severe sepsis among the 11 740 admissions to the 170 icus participating in the french bacteraemia/sepsis study were: age, male sex, admission to a large ( > 400 beds) hospital, a medical or unscheduled surgical admission, presence of chronic liver insufficiency, of immunodepression, and of severe underlying disease [13] . there was no difference in these risk factors when excluding patients with non-documented severe sepsis (i. e.,`severe sirs') from the cohort. this question was specifically addressed in the french multicenter study of 832 patients with bacteraemia [5] . by cox regression analysis, independent factors associated with severe sepsis during bacteraemia were increasing age ( > 50 years), sources other than the urinary tract, an intravascular catheter, or primary bacteraemia. organisms involved were not associated with the occurrence of severe sepsis, nor was the severity of the underlying disease. in another retrospective study of 1505 patients that had been included in the va corticosteroids trials [16] (of whom 40 % had uncomplicated sepsis, 45 % met criteria for severe sepsis and 15 % for septic shock), independent risk factors for the development of severe sepsis or shock were age, gastro-intestinal tract disease, liver disease, haematological disorders, spinal cord injury, and drug abuse. as already mentioned, only a limited fraction of patients presenting with one of the septic syndromes have microbiologically documented infection. in patients meeting criteria for sirs, only 42 % were found by rangel-fausto et al., to have documented infection (i. e., sepsis), and 58 % had culture-negative, but clinically documented, infection; the proportion of documented infection rose to only 47 % in patients with severe sepsis, and to 57 % in patients with septic shock [6] . higher rates of infection were similarly found in the more severe forms of septic syndromes in other studies: clinically or microbiologically documented infection was recorded in 92 % of episodes in patients meeting clinical criteria for severe sepsis by sands et al. [8] and in 95 % of episodes recorded by brun-buisson et al. [13] ; in these two studies, 70 % and 71 % of patients had microbiologically documented infection, and 30 % and 29 % had clinically documented infection, respectively. therefore, only a small fraction of patients presenting with clinically suspected severe sepsis (5 %±10 %) had no infection clinically or microbiologically documented. the four major sources of infection in patients with severe sepsis, in descending order, are the respiratory tract, the abdomen, the urinary tract, and primary bacteraemia [8, 13] ; these sources account for > 75 % of cases of severe sepsis (table 3 a) . this distribution differs somewhat from that observed in patients with bacteraemic sepsis, where the urinary tract is the major source of infection (table 3 b), reflecting the lower risk associated with this source in causing severe sepsis, as already mentioned. of note, there is no major difference in the distribution of sources of infection when one compares microbiologically documented cases to clinically documented cases, except for a higher proportion of urinary tract infection and catheter infection in the former group. microbial epidemiology of sepsis, severe sepsis or shock the microbiological features of the septic syndromes may depend in part on the population studied and setting. bacteraemia may be taken as the reference syndrome for looking at microbial aetiologies of sepsis. it should be recalled that major changes have occurred in the past two decades in the epidemiology of bacteraemia. these include increasing rates overall, and a growing importance of gram-positive organisms over the years, especially among nosocomial episodes, which account for most of the recent increased rates [4] . much of this increasing role of gram-positive organisms is due to catheter-related infections and primary bacteraemia. as a result, gram-positives now outweigh gramnegative among bacteraemic episodes (55 % vs 45 %), as shown in the french multicenter study (table 4 ) [5] . in severe sepsis, however, the proportion of grampositives and gram-negatives appear similar, reflecting the lower risk of severe sepsis associated with infection caused by coagulase-negative staphylococci [13] ; in non-bacteraemic severe sepsis, however, gram-negative organisms appear to predominate [8] . again, there was no major difference in the distribution of organisms when comparing bacteraemic episodes associated with sepsis only or with severe sepsis, except for a marginally higher proportion of polymicrobial infection (table 4 ). these data suggest that the microbiologic characteristics of infection are not a major determinant of the clinical presentation and intensity of the host response to infection. this notion is also consistent with the fact that it appears quite difficult to predict bacteraemia in patients presenting with clinical sepsis [14] . it is apparent that the classification into three major syndromes (sepsis, severe sepsis, and sepsis shock) reflects a grading in prognosis of patients affected, and this is clearly an important outcome of the current classification. there are, however, wide variations in mortality rates reported in cohorts of patients with septic s 70 * p < 0.001 for comparison of sources between episodes of bacteremic sepsis and bacteremic severe sepsis. data from [5] ; reproduced with permission syndromes, especially for hospital-wide data. in the study by rangel-fausto et al. [6, 7] the 28-day mortality of the different stages from sirs to septic shock was 7 %, 16 %, 20 %, and 46 % for sirs, sepsis, severe sepsis and septic shock, respectively. in the study conducted by sands et al. [8, 14] the 28-day mortality of patients with severe sepsis and septic shock was 34 %. in the french multicenter study, the 28-day mortality was 25 % in patients with bacteraemic sepsis (19 % in ward patients), and of 54 % in patients with bacteraemic severe sepsis or shock [5] . in studies restricted to icu patients, mortality rates were slightly more consistent across studies. pittet et al., reported a 28-day and hospital mortality rate of bacteraemic sepsis of 35 % and 43 %, respectively [17] ; 77 % of these patients had severe sepsis, as assessed by the presence of organ dysfunction at onset or secondarily. brun-buisson et al., reported a 28-day mortality of 55 % in icu patients with bacteraemic sepsis, 65 % of whom had severe sepsis or shock [5] ; overall, mortality was 56 % at 28 days after severe sepsis among 1052 icu patients, of whom 71 % had septic shock [13] . it is apparent that the mortality rate for a given stage upon inclusion is dependent in large part on the proportion of patients rapidly evolving to a more severe stage. knaus et al., have shown that a wide range of mortality risk could be observed in patients classified as having sepsis or even shock [18] . further insight into a better characterisation of outcome for septic patients has been provided by studies looking at mortality risk adjustment through risk factors analyses and models specific to septic patients. as for all icu patients, there are two major determinants of outcome for septic patients: the severity of underlying disease, and the severity of acute illness. severity of underlying disease has been assessed via several indexes or systems, such as the simple (and robust, but somewhat subjective) three-classes index developed by maccabe et al., for bacteraemic patients [19, 20] or a comorbidity scoring index, a system primarily developed for adjusting the risk of nosocomial infection [21] ; finally, general scoring systems such as the apache ii include comorbidities, expressed as pre-existing organ dysfunction for the four major organ systems (respiratory, cardiovascular, renal, and liver), or include a few major comorbidities such as in the saps ii (aids, metastatic cancer, and haematological malignancy) [22] . a more complete assessment of pre-existing conditions can probably be obtained by ascribing a weighted score to diagnosis and comorbidities, as used in the apache ii and iii scoring systems [18] . stratifying patients by risk class according to one of the general scores provides a more accurate risk prediction of outcome for septic patients than a simple stratification in one of the stages of sepsis [18] . further analysis of the performance of these scores in septic patients have led to the development of customised scores, either through the integration of additional variables [23] or through modification of the weighting of variables included in the original score [24] . the use of such customised models in risk prediction in the context of clinical trials to accurately compare predicted to observed mortality and derive the efficacy of new therapies in subgroups of patients has, however, been disappointing [23, 25] . it is apparent that the general scores, whether or not customised, principally reflect the severity of acute physiologic disturbances when measured at time of sepsis; in other words, they reflect the severity of organ dysfunction associated with sepsis. two studies have examined risk factors for short-term mortality in a predefined cohort of icu patients with sepsis or severe sepsis [13, 17] . in the french multicenter study [13] several factors recorded at onset of sepsis were found associated with early mortality of patients with severe sepsis: the maccabe index (or = 1.5 and 2 for ultimately and rapidly fatal underlying disease, respectively), and bacteraemia (or = 1.7); however, the three most important independent risk factors for early mortality were the saps ii (or = 1.03 per unit of score, p = 0.003), the presence of shock (or = 2.9, p = 0.01), and the presence of more than one organ system failures, as defined according to knaus et al. [26] (or = 8.4 and or = 9.4 for 2 or > 3 organ failures, respectively; p = 0.002). later mortality after severe sepsis was also associated with the saps ii, and the number of organ system failures, but also with other factors related to underlying diseases, such as pre-existing liver or cardiovascular failure or the maccabe index, the admission category, and presence of multiple sources of infection. in 176 patients with bacteraemic sepsis, pittet et al. [17] also found that the apache ii score measured at the time of sepsis was highly predictive of mortality (and a better predictor than apache ii measured on icu admission). these authors also noted that prior antimicrobial therapy and hypothermia were associated with a poorer prognosis. organ dysfunctions were also strongly associated with mortality; however, only those recorded after the onset of sepsis were associated with mortality in that study [17] . this surprising finding may have been due to the limited power of the study. therefore, in addition to a general severity score, and one assessing prior comorbidities such as the mac-cabe score, organ dysfunctions at onset of sepsis and developing after its onset appear as the major determinants of the short-term outcome of septic patients. whether customised models for sepsis perform better than the above combination of three major determinants has not been formally tested on a large cohort of patients, and remains unknown. an interesting approach would be to incorporate a score of organ dysfunction in the prognostic assessment of patients with sepsis. these refined scoring systems for organ dysfunction/failures [27, 28, 29] which allows identification of organ dysfunction in a graded manner and at an earlier stage, would likely allow a more precise description of prognostic factors and of the interrelations between the various organ dysfunctions and their respective impact on outcome. however, from the viewpoint of mortality prediction, it is unlikely that these scores perform better than the general (or customised) scoring systems. most studies and clinical trials have focused on 28-day or hospital mortality, a relatively short-term view, which does not provide a complete picture of the impact of sepsis on life expectancy, an important consideration for cost-benefit studies of the impact of new therapies. admittedly, most deaths usually occur early in severe sepsis. for example, 77 % and 71 % of all deaths had occurred by day 14 and by day 21, respectively, in the french multicenter study [13] and in the study by pittet et al. [17] . however, 16 % of patients remained in the hospital for more than 30 days in the former study, and the median hospital stay of survivors was 34 days, with lengths of stay ranging from 1 to 87 days [13] . these data suggest that it would be advisable to assess the outcome of sepsis at least after 3 rather than 1 months after sepsis. similarly, sands et al. reported that the crude mortality of patients with severe sepsis was 34 % at 28 days and 45 % at five months post-discharge [8] . sasse et al. [30] reported a crude 28-day and hospital mortality rate of 40 % and 51 %, respectively, in 153 icu patients with bacteraemic sepsis, and of 65 % and 72 % respectively at 6 months and 1 year after admission; it should be emphasised that in this particular study, 25 % of patients each had hiv infection or malignancy. finally, rangel-frausto et al., reported a 28-day crude mortality of 9 % in the cohort of 2527 patients meeting at least sirs criteria; an additional 111 patients died during the ensuing 3 months, and 113 more between 3 and 6 months follow-up [6] . the overall crude mortality rate at 6 months was therefore of 17 % in patients meeting at least sirs criteria. however, the relative part of sepsis and other host factors in the overall mortality is unknown. perl et al., have addressed part of the problem by examining factors associated with late mortality after sepsis in a cohort of 100 patients with severe sepsis entered in one clinical trial of anti-endotoxin antibodies [20] . in that study, the crude mortality of patients was 32 % at 1 month, 37 % at 3 months, and 43 % at 6 months; after a mean follow-up of 30 months, 60 % patients had died. when examining factors associated with mortality at those different points in time after sepsis, they found that all models included the severity of underlying disease (maccabe classification) and a combined index of comorbidities, in addition to vasopressors (or shock) and ventilator use (or ards). therefore, pre-existing illness and comorbidities, in addition to shock and organ failures (i. e., ards), are also confirmed as important predictors of long-term outcome in this study. an elegant study by quartin et al. [16] has provided some more insight into the problem of long-term mortality attributable to sepsis. these authors have estimated the increased risk of mortality attributable to sepsis over a 6-year follow-up period in patients qualifying for sepsis and entered into a clinical trial of corticosteroid therapy, as compared to a control cohort of non-septic hospitalised patients, after adjustment on risk factors for death in the control cohort. after 8 years, 1229 (82 %) of the 1505 septic patients had died. the authors estimated that septic patients had an increased mortality risk persisting beyond one month and over the five years following sepsis; the increased risk was apparent in all stages of septic syndromes. the median predicted life expectancy was 5 years among septic patients. the average life expectancy cost of sepsis was estimated at 2.4 years and the median survival among 30-day survivors was reduced from 6.2 to 2.3 years. in the septic population, there were 452 (30 %) more deaths than predicted from controls within the first month, 192 (13 %) more within one year, and 61 (4 %) more within 5 years; thus, the overall mortality attributable to sepsis was 43 % at one year. sepsis also appears to significantly affect the quality of life of survivors. in the study by perl et al. [20] survivors had lower scores than normal by functional status and general health perception scales. it is quite clear from the above epidemiological information that sirs criteria are much too non-specific to be used for the selection of patients. including patients characterised only by these criteria would only result in augmenting the`background noise', by introducing a large population having a < 50 % risk of sepsis, and a low risk of mortality. one would like sepsis or its more severe forms to be used as criteria for inclusion. however, physicians remain with the dilemma that there is no reliable method for identifying patients having sepsis among those presenting with clinical criteria for sirs/ sepsis. in this respect, the new classification has not provided a significant advance in identifying at-risk patients. it is noteworthy that all recent clinical trials have actually used criteria for severe sepsis or shock in their inclusion criteria. if this classification is used, and there is a need for studying patients at an early stage of infection, then a more in-depth analysis of risk factors for sepsis or its more severe forms in patients with sirs patients is needed. septic shock remains a major prognostic factor and, importantly, is readily available for stratification at inclusion of patients into clinical trials. a general severity score (original or customised) could also be used, or better, an organ dysfunction score, depending on objectives and end-points pursued in the trial. consideration should also be given to major underlying conditions, using a simple index, such as the maccabe score. it is apparent that the systemic response to infection, not infection itself, is the major determinant of the outcome of patients. mortality remains the reference endpoint. this simple and robust end-point is validated by the fact that mortality remains high, at least in the most severe forms of the septic syndromes, and by the estimated attributable mortality which is also very high, and likely accounts for two-thirds of the overall mortality, especially of the short-term mortality. since sepsis has also substantial effects on long-term survival, a longer than usual follow-up (i. e., 6 months to 1 year) should be used, at least if some possible delayed effects of therapy on survival are expected. using mortality as an end-point implies that factors other than sepsis itself, which have a significant impact on patients' survival, are accounted for in the survival analysis. these factors include, (but may not be limited to) the severity of the underlying disease, the presence of comorbidities, and the severity of haemodynamic disturbances and other organ dysfunction at inclusion. the importance of underlying conditions is highlighted by the relatively low spontaneous life expectancy associated with underlying illness and comorbidities in most patients with sepsis. it has been suggested that the assessment of organ dysfunction/failure, especially via a grading score could be used as a substitute for mortality. this debate is at present unsettled. clearly, organ dysfunctions are strongly (linearly) related to mortality, and there is no obvious advantage from using such a score instead of mortality. using organ failure-free days, as suggested when dealing with one organ system dysfunction such as ards, may be misleading, as death may still occur relatively late after onset of sepsis. at present, organ failure scores are best used as adjustment variables at onset of sepsis and their assessment over time should be viewed as explanatory observations in an attempt to provide better insight into the physiological effects of interventions. the american-european consensus conference on ards: definitions, mechanisms, relevant outcomes and clinical trials coordination definitions for sepsis and organ failure and guidelines for the use of innovative therapies in sepsis increase in national hospital discharge survey rates for septicemia ± united states nosocomial bloodstream infections. secular trends in rates, mortality, and contribution to total hospital deaths bacteremia and severe sepsis in adults: a multicenter prospective survey in icus and wards of 24 hospitals the natural history of the systemic inflammatory response syndrome the dynamics of disease progression in sepsis: markov modeling describing the natural history and the likely impact of effective antisepsis agents epidemiology of sepsis syndrome in 8 academic medical centers systemic inflammatory response syndrome, sepsis, severe sepsis and septic shock: incidence, morbidities and outcomes in surgical icu patients role of systemic inflammatory response syndrome and infection in the occurrence of early multiple organ dysfunction syndrome following severe trauma american college of chest physicians/ society of critical care medicine consensus conference definitions of the systemic inflammatory response syndrome and allied disorders in relation to critically injured patients the italian sepsis study: preliminary results on the incidence and evolution of sirs, sepsis, severe sepsis and septic shock incidence, risk factors, and outcome of severe sepsis and septic shock in adults. a multicenter prospective study in intensive care units predicting bacteremia in patients with sepsis syndrome the sepsis syndrome in a dutch university hospital. clinical observations the departement of veterans affairs sepsis cooperative studies group (1997) magnitude and duration of the effect of sepsis on survival bedside prediction of mortality from bacteremic sepsis. a dynamic analysis of icu patients wagner dp (1992) evaluation of definitions for sepsis gram-negative bacteremia.iv. re-evaluation of clinical features and treatment in 612 patients long-term survival and function after suspected gram-negative sepsis importance of pre-existing comorbidities for prognosis of septicemia in critically ill patients a new simplified acute physiology score (saps ii) based on a european-north american multicenter study the clinical evaluation of new drugs for sepsis: a prospective study design based on survival analysis the icu scoring group (1995) customized probability models for early severe sepsis in adult intensive care patients the phase iii rhil-1ra sepsis study group (1994) recombinant human interleukin-1 receptor antagonist in the treatment of patients with sepsis syndrome: results from a randomized prognosis in acute organ system failure the icu scoring group (1996) the logistic organ dysfunction system: a new way to assess organ dysfunction in the intensive care unit multiple organ dysfunction score: a reliable descriptor of a complex clinical outcome use of the sofa score to assess the incidence of organ dysfunction/failure in intensive care units: results of a multicenter, prospective study long-term survival after intensive care unit admission with sepsis key: cord-001293-dfaxj3bv authors: cavaillon, jean-marc; eisen, damon; annane, djilalli title: is boosting the immune system in sepsis appropriate? date: 2014-03-24 journal: crit care doi: 10.1186/cc13787 sha: doc_id: 1293 cord_uid: dfaxj3bv a relative immunosuppression is observed in patients after sepsis, trauma, burns, or any severe insults. it is currently proposed that selected patients will benefit from treatment aimed at boosting their immune systems. however, the host immune response needs to be considered in context with pathogen-type, timing, and mainly tissue specificity. indeed, the immune status of leukocytes is not universally decreased and their activated status in tissues contributes to organ failure. accordingly, any new immune-stimulatory therapeutic intervention should take into consideration potentially deleterious effects in some situations. refinements of supportive care of patients with severe sepsis have decreased their overall mortality, but no adjuvant drug therapy has emerged despite strenuous efforts in the field. twenty years have passed since the first patients with sepsis were included in clinical trials based on the understanding that tnf orchestrates the inflammatory response and should be the target for therapeutic intervention. in response to the failure of therapies aiming to target either the up-stream microbial activators or the effector molecules of the inflammatory cascade, a new concept has emerged of boosting the immune system to counter immunosuppression that develops in patients who survive the initial, hyperinflammatory period of sepsis [1] . inflammation is a highly sophisticated and complex response that fundamentally 'aims' to protect the host. in this review, we argue against the promulgation of what we believe is a misleading perception of sepsis inducing secondary immunosuppression. the possible negative consequences of immune system-boosting therapy are paradoxical properties have also been reported for il-10, the prototypic anti-inflammatory cytokine. its proinflammatory activity been established in human volunteers receiving endotoxin injection [6] . our own in vitro studies showed that adherence of human monocytes modulated the effect of il-10 on expression of 16 genes, including 'suppressor of cytokine stimulation' (socs) molecules, in the opposite direction as compared with non-adherent cells [7] . these observations illustrate the statement by moore and colleagues that 'il-10 can effect very different outcomes depending on timing, dose, and location of expression. in some scenarios, the expected immuno-suppressive activities are observed, while in others, il-10 enhances immune or inflammatory responses' [8] . among other cytokines classified as antiinflammatory, transforming growth factor-beta (tgfβ) may behave as pro-inflammatory mediator as tgfβtransgenic mice are more sensitive to lps-induced shock [9] and some of its inflammatory activities reflect its capacity to favor the differentiation of t helper (th) 17 and production of the pro-inflammatory il-17. the classification of non-cytokine inflammatory mediators also relies on an overly simplistic division between proand anti-inflammatory properties. this is well illustrated by prostaglandin e 2 (pge 2 ), a key mediator of infectious immunopathology. on one hand, pge 2 induces fever, increases vascular permeability, increases vasodilatation, and causes pain while also inhibiting production of tnf, increasing production of il-6, inhibiting 5-lipoxygenase and leukotriene a4 generation, and inducing 15lipoxygenase and the generation of the lipoxins involved in inflammation resolution. on the other hand, pge 2 has inhibitory properties on macrophages, neutrophils, th1 lymphocytes, natural killer (nk) cells, and cytotoxic lymphocytes but activates mast cells, th2, th17, and regulatory t lymphocytes (t reg ) [10] . this panoply of pge 2 -stimulated events amply demonstrates the inability to simply characterize the activities of this and the other molecules mentioned as pro-or anti-inflammatory. in an early anti-tnf monoclonal antibody intervention study, a significant improvement in day 3 survival was observed between the antibody-treated group and the placebo group [11] . although this was not a prespecified primary outcome, it is interesting to see that the treatment targeting tnf consisting of a single early injection was beneficial within a short period of time after sepsis onset, reinforcing the idea that tnf plays a key deleterious role in the early events of sepsis. once anti-tnf treatments were better targeted to the sickest patients by adding biological inclusion parameters (plasma il-6 level), survival was significantly improved on day 28 [12] . synergistic effects between immune modulators are a key characteristic of their effect. this explains how a non-lethal dose of one cytokine can lead to mortality when injected with a non-lethal dose of another cytokine. similarly, it may explain how the removal of some inflammatory mediators by coupled plasma filtrationadsorption was protective in an endotoxin-shock model while levels of circulating bio-active tnf were unaffected [13] . clear demonstrations of cytokine-mediated tissue damage exist. nevertheless, because of their ambiguous role mentioned above, identification of their precise role during sepsis has led to controversy. in animal models of sepsis, the role of tnf may vary depending upon the type of infection [14] . many model parameters influence conclusions of the relative role of the different mediators studied. identical cytokines have been found to be protective or deleterious depending upon the model. this has been the case for ifnγ [15] and granulocytemacrophage colony-stimulating factor (gm-csf) [16] among others (for example, il-17, il-33, 'tnf-related apoptosis-inducing ligand' (trail), and tgfβ). host-protective innate immune responses and consequent inflammation are inextricably linked and overlapping. consequently, the same cellular actors are key elements defending the host against infection while simultaneously contributing to deleterious events. for example, neutrophil extracellular traps that catch and kill bacteria and fungi are associated with the release of elements such as histones and mitochondria that behave like damage-associated molecular patterns perpetuating the inflammatory process. beneficial or deleterious roles of the same leukocyte subset have been reported depending upon the experimental model. for example, a peritonitis model using nude mice (lacking t cells) suggested that t lymphocytes contribute to protective immune responses [17] . by contrast, in an escherichia coli sepsis murine model, t lymphocytes markedly contributed to severity [18] . similarly, t reg improved survival in polymicrobial sepsis [19] whereas, in another report, reduced t reg activity led to improved survival [20] . the 'half angel/half devil' role of nk cells during severe infection is also described. nk cells contribute to systemic inflammation during polymicrobial sepsis but play a critical protective role in host defense against staphylococcus aureus lung infection (as reviewed in [21] ). although apoptosis of dendritic cells (dcs) is particularly increased during sepsis, they are protective in murine polymicrobial sepsis [22] . transcriptomic analysis of dcs in trauma patients shows a large number of upregulated inflammatory genes, suggesting their contribution to systemic inflammation and organ failure [23] . apoptosis of lymphocytes, dcs, and nk cells is a hallmark of sepsis. hotchkiss and colleagues [24] provided key experiments demonstrating that lymphocyte apoptosis was deleterious and its prevention highly protective. in addition to the depletion of apoptotic lymphocytes that contribute to the alteration of the immune status, apoptotic t cells themselves can further produce an immunosuppressive milieu following their release of tgfβ [25] . in contrast, the apoptosis of neutrophils is reduced. interestingly, injection of apoptotic neutrophils in lps-challenged mice with or undergoing cecal ligature puncture improved outcomes [26] . this may be due to the capacity of apoptotic neutrophils to limit the production of il-1 and tnf by lps-activated monocytes and to favor the production of il-10 and tgfβ [27] . favoring neutrophil apoptosis while differentially preventing that of lymphocytes and dcs would represent a considerable interventional challenge! in sepsis, apoptosis does not only affect immune cells. apoptosis of epithelial cells, endothelial cells, neurons, and cardiac myocytes is reported with crucial effects of loss of altered barrier function ( figure 1 ): in the lungs -acute lung injury and adult respiratory response syndrome [28] ; in the kidneys -acute kidney injury [29] . enhanced translocation of bacteria and bacterial products occurs consequent on intestinal epithelial cell apoptosis [30] , contributing to the concept of the gut as the motor of multiple organ failure (mof). sepsis-induced cardiac myocyte apoptosis produces altered contractility and cardiac dysfunction [31] . apoptosis of endothelial cells [32] induces vascular leakage. finally, microglial and neuronal apoptosis may follow autonomic failure that precedes shock and mof [33] . in addition to epithelial apoptosis, tight junction alterations enhance organ dysfunction. it has been demonstrated that nitric oxide favors disruption of epithelial cell tight junctions in numerous organs, including liver, gut, and lung. leukotrienes favor protein extravasation as shown in the kidney of septic mice [34] . still, cytokines remain the main orchestrators of these tissue injuries. in a model of acute kidney injury, it was nicely demonstrated that inflammatory cytokines, including tnf and il-17, cause small intestine and liver injury [35] . among others, il-17a is critical for generation of intestinal ischemia/reperfusion injury and subsequent liver and kidney injury [36] . all together, the altered functions of epithelial cells, endothelial cells, neurons, and cardiac myocytes contribute to mof that may influence outcomes in sepsis more than altered immune status. the concomitant occurrence of inflammation, anti-infectious response, and altered immune status in sepsis when roger bone coined the concepts of systemic inflammatory response syndrome (sirs) and compensatory anti-inflammatory response syndrome (cars), he conceived that one or the other would be predominating figure 1 during sepsis, many types of cells (but not neutrophils) display enhanced apoptosis, leading to various deleterious consequences. aki, acute kidney injury; ali/ards, acute lung injury/acute respiratory distress syndrome; nk, natural killer. [37] . however, we contend that cars should be considered an adapted compartmentalized response with the aim of silencing some acute pro-inflammatory genes and maintaining the expression of certain genes involved in the anti-infectious process. despite our views [38] , authors still propose a two-wave concept with sirs appearing before cars, although they admit that 'rigorous examination of previous studies provides evidence that both proinflammatory and opposing anti-inflammatory response(s) occur concomitantly in sepsis' [1] . tamayo and colleagues [39] studied a large panel of circulating cytokines in patients with sirs or sepsis, concluding that both pro-and anti-inflammatory mediators play roles from the very beginning of this life-threatening condition. similarly, meta-analysis of 12 transcriptomic studies including 784 individuals led to the conclusion that 'the arbitrary distinction of separating sepsis into proinflammatory and anti-inflammatory phases is not supported by gene-expression data' [40] . immune status has been studied frequently by measuring tnf or other inflammatory cytokine production by circulating monocytes in response to lps [41] . we studied patients undergoing abdominal aortic surgery, showing that reduced expression of human leukocyte antigen (hla)-dr on cd14 high monocytes occurs during surgery [42] . similarly, hla-dr expression was already reduced on monocytes taken very soon after severe trauma at accident scenes [43] . altered tnf production capacity of circulating cells in response to tlr2 or tlr4 agonists is also observed very soon after injurious insults, such as on admission of patients after cardiac arrest [44] . even if soon after the initial insult the intensity of the inflammatory response reaches its peak, there is a persistent inflammation associated with altered immune status in surviving patients [45] . the more severe the insult, the more profound is the alteration and the more chance the patients have to develop adverse clinical outcome. the immune status of leukocytes during sepsis and sirs varies depending on the compartment in which they reside terms such as 'immunoparalysis, immunosuppression, and anergy' are far too extreme to describe the immune status of circulating leukocytes in patients with sepsis or sirs. altered immune status of circulating leukocytes is not globally present. indeed, some functions like phagocytosis remain unaltered [46] , and ex vivo cytokine production in response to heat-killed s. aureus (hksa) remains unchanged in patients with sepsis [47] compared with healthy controls. this is in full agreement with the observation that lps primes hksa-induced tnf production in macrophage cell lines instead of leading to cross-tolerance [48] . while the concept of endotoxin tolerance is considered to partially mimic the alteration of immune status in sepsis, it is worth mentioning that cross-tolerance between microbial agonists is not invariant. for example, candida albicans and fungal cell wall β-glucan also prime lps-induced proinflammatory cytokine production [49] . these observations led us to propose the concept of leukocyte reprogramming [50] to explain the fact that tolerised macrophages retain anti-infectious properties. in addition, in tissues, there are numerous examples to illustrate the hyper-activity of these cells. for example, in mice with polymicrobial sepsis alone or as a 'second hit' after traumatic hemorrhage, it was nicely demonstrated by chaudry's group [51] that the ex vivo production of tnf or il-6 after lps activation was significantly reduced among peripheral blood mononuclear cells and splenic macrophages but that it was enhanced in alveolar and kupffer cells. similarly, in a murine model of trauma, the cytokine productive capacity of kupffer cells and alveolar macrophages was enhanced [52] . indeed, macrophage functions differ depending on the compartment from which they derive. we established [53] that the specific cytokine and cellular microenvironment within the lung was responsible for this particular resistance of alveolar macrophages to endotoxin tolerance, which can also be observed in human alveolar macrophages [54] . similarly, in kidneys, in response to a second challenge with lps, the expression of tnf and inducible nitric oxide synthase was further enhanced [55] . this may explain why unilateral nephrectomy could be protective in a murine peritonitis model and after lps injection [56] . most importantly, despite the fashionable concept of m1/m2 macrophages, the response of macrophages to il-4 and ifnγ is in fact completely different depending upon their origin [57] . as a consequence of this great heterogeneity of immune cells within different compartments, each tissue behaves independently, contributing to the global inflammatory response with a specific pattern, as illustrated by differential cytokine expression in liver, lungs, heart, brain, muscle, kidney, intestine, and spleen [58] . another example of the different behavior of leukocytes in various compartments is the frequent occurrence of hemophagocytosis (>60%) directly observed in the bone marrow of the critically ill [59] . this phenomenon is associated with extreme production of inflammatory cytokines. accordingly, it has been proposed that when hemophagocytosis is diagnosed in critical care patients, aggressive immunosuppressive therapy be undertaken without delay [59] . differences between cells harvested from different compartments after sepsis have also been reported for spleen and peritoneal myeloid dcs [60] . the major differences between compartments are further illustrated by the fact that gene deficiency may differentially affect outcomes of infection. for example, il-10 deficiency protects against francisella tularensis pulmonary infection but aggravates cutaneous infection [61] . similarly, we showed that scavenger receptor-a (sr-a), 'macrophage associated receptor with a collagenous base' (marco), cd36, or tlr2 deficiency protect mice against peritoneal s. aureus infection while these deficiencies aggravated pneumonia [62] . interestingly, when streptococcus pneumoniae was the pathogen used to colonize the murine nasopharynx, marco ko mice (but not sr-a ko mice) had significantly impaired clearance of pneumococcal colonization [63] . furthermore, inflammatory foci cells may not behave similarly to cells from other healthy compartments. for example, it was shown that neutrophils derived from sputum of patients with chronic bronchitis or cystic fibrosis are insensitive to inhibitory effects of il-10 in contrast to circulating neutrophils [64] . the concomitant presence of inflammation within tissues and altered immune status within the hematopoietic compartment is short-lived in murine models rendering them inappropriate to study patients with concomitant sepsis and cars [65] . in addition, mice are highly resistant to bacteria like s. aureus and their serum contains factors that limit inflammatory response intensity as compared with human serum [66] . a most provocative report comparing transcriptomic patterns of circulating cells from trauma patients, human endotoxemia-model participants, and murine-model equivalents revealed total absence of correlation [67] . when most therapeutic approaches have been validated in preclinical studies performed with murine models, one understands why those were not the most appropriate ones. the scientific community needs to reconsider models used to validate therapeutic approaches. if murine responses do not resemble human processes, maybe other species, like the pig, should be preferred. porcine monocytes and lps-activated macrophages are closer to their human counterparts than murine cells [68] . of course, murine models remain valuable to further decipher the mechanisms of sepsis. the best example is the two-hit model, which demonstrated that the nature of the first hit and its severity, the nature of the infection, and the route of infection may influence the outcome in a completely opposite direction [69] . are patients with sepsis dying of immune failure -dissecting the arguments used to describe compensatory immunosuppression occurring after sepsis? the clinical observations used to argue that immunosuppression occurs in sepsis patients surviving the initial inflammatory cascade [1] are in essence that patients develop nosocomial infections due to opportunistic pathogens, including reactivated chronic viral infections, and that patients who die after sepsis have unresolved foci of infection. these underpinning observations require further consideration. representing bacteria such as enterococcus faecium, stentrophomonas maltophilia, and pseudomonas aeruginosa along with candida as 'opportunistic pathogens' overstates the role of sepsis-induced immune dysregulation as the primary cause of nosocomial infection in these patients. these multiply-instrumented, high-intensity care, bed-bound, vulnerable patients often have breaches in their integument and mucous membranes (airways, surgical sites, indwelling catheters) and perturbed microbiomes from antibiotic treatments. overgrowth of antibioticresistant microorganisms and barrier defects predispose them to secondary infections, even without overt defects in their immune defenses [70] . these are all organisms of normal virulence that cause nosocomial infections in sepsis patients because of the selection pressure of potent antibiotics and the presence of biofilm affected/colonized intravascular and urinary catheters. additionally, reactivation of herpes simplex virus (hsv) and cytomegalovirus (cmv) may have some clinical relevance in critically ill patients. cmv-emia is quite common in patients with sepsis (30% in some studies) and is at least associated with worse outcome in icu patients in recent meta-analyses. whether cmv could cause immune compromise itself, be a reflection of immune compromise, or simply be an indicator of poor outcome in patients with sepsis remains unclear [71] . reactivation of oro-labial hsv is extremely common in sepsis, and hsv can frequently be detected in respiratory secretions. however, only one study has reliably investigated lower respiratory tract infection in critically ill, immunocompetent patients, showing that 21% of patients with ventilator-associated pneumonia (vap) had bronchopneumonitis due to hsv [72] . in 55% of these patients, the vap appeared to be due to hsv alone. however, acyclovir treatment had no impact on the outcome in patients with hsv bronchopneumonitis [72] . of greater relevance to predisposition to nosocomial infection in sepsis patients remaining in icus for prolonged periods are physical breaches in innate immune system barriers. intravascular catheters, endotracheal tubes with consequently increased dead space, and increased gastric ph due to peptic ulcer prophylaxis regimens are all, along with broad-spectrum antibiotics, potent promoters of nosocomial infection. post-mortems (pms) identifying unresolved infection foci are not reliable proof that patients are dying of sepsis. pneumonia is frequently present in patients in whom supportive care is withdrawn due to failure to thrive. where pneumonia has been found more frequently at pm than was appreciated ante-mortem, the extent of pulmonary involvement was not quantified [73] . in this series, there was clear agreement by clinical and pm assessment that mof was the commonest cause of death [73] . these data call into question the relevance of unresolved, pm infection in patients dying in the icu as a direct indicator of immunosuppression following as a direct consequence of previous sepsis. if the patients die with infectious foci and altered immune status, it does not mean they die because of them. because of the monocyte deactivation in sepsis, it was proposed to restore it with the use of either ifnγ or gm-csf, two cytokines that counteract endotoxin tolerance. the first attempt was successfully performed in nine septic patients who received subcutaneous ifnγ that restored ex vivo cytokine production and hla-dr expression by monocytes [74] . the authors claimed that overall mortality was lower in the treated group compared with historical controls. in mechanically ventilated trauma patients, ifnγ was aerosolized. however, in a previous phase iii study in burn patients, ifnγ had failed to protect patients from infection or decrease mortality [75] . we must recall that ifnγ injection increases mortality in animal models of polymicrobial infection [15] . all together, these data have limited the routine use of ifnγ in icu patients, although a dutch clinical trial is ongoing. gm-csf has been demonstrated to be able to restore some immune status parameters. however, a metaanalysis concluded that gm-csf did not significantly reduce in-hospital mortality, although it significantly increased infection recovery [76] . although no adverse effects were reported, it is worth recalling a case report of a patient who developed a fatal adult respiratory distress syndrome after gm-csf treatment [77] . in animal models, gm-csf favors lps-induced lung inflammation, amplifying lps-induced bronchoconstriction [78] . gm-csf favors production of tnf and il-1. in a recent study, it was confirmed that gm-csf synergizes with lps, promoting il-1β secretion [79] . lethal injection of lps in gm-csf receptor ko mice led to far lower mortality among these mice as compared to wild type mice. given all the efforts made by some authors to convince the scientific community of the use of gm-csf, it is challenging to read the conclusion of this present paper given that gm-csf has been previously underestimated as a target for therapeutic intervention in many bacterial infections and inflammatory disorders associated with the production of il-1β. il-7 is another cytokine that is promoted for the treatment of sepsis and that is supported by murine and human ex vivo tissue data [1, 80] . one can conjecture that systemic treatment with il-7 may act in undesired places, as illustrated by the following: il-7 worsens graft-versus-host-induced tissue inflammation [81] ; favors inflammation in colitis [82] , contributes to arthritis severity [83] ; upregulates chemokines, ifnγ, macrophage recruitment, and lung inflammation [84] ; and, finally, increases production of inflammatory cytokines by monocytes and t cells [85] . many other cytokines (for example, il-2, il-12, il-15, and tnf) can boost the immune system and are reported to be beneficial in murine sepsis models. however, one wonders whether systemic treatment with any immunostimulating cytokine may act on tissue leukocytes boosting the inflammatory process while boosting immune status as well. in this perspective, the attempt to treat peripheral mononuclear cells of sepsis patients ex vivo with il-2 before re-injecting them is an interesting approach that prevents the delivery of this cytokine to the bloodstream, allowing it to act strictly on the desired cells [86] . in this study, the mortality was 8% in the extracorporeally treated group of patients (n = 121) but was 21% in the patients receiving standard treatment (n = 52). rather than repeating the mistakes of past experimental treatments for sepsis in which therapies were developed after successful preclinical models that may be far from mimicking human disease, it would be ideal to proceed in the future with new treatments in which extensive human data are available prior to embarking on expensive licensure studies. furthermore, identifying currently licensed drugs with tolerable safety profiles as potential sepsis agents leap-frogs costly drug development and early-phase human studies. in animal models, extant licensed drugs, such as chloroquine [87] and androstenenediol [51] , have successfully restored immune status. most interestingly, in the latter case, the treatment protected mice against polymicrobial sepsis and boosted altered ex vivo cytokine production observed with peripheral blood cells and spleen macrophages, dampening production observed with alveolar macrophages and kupffer cells. a similar compartmentalized adapted specificity was reported with estradiol [88] . other approaches involve pro-resolving lipid mediators [89] , although it is uncertain whether they may also adversely boost immune status. the recently recognized aspirin-triggered lipoxins, anti-inflammatory mediators of inflammation resolution, make aspirin a possible inexpensive agent for both prevention and treatment of sepsis. considerable observational cohort data show improvements in mortality in patients with sepsis pretreated with aspirin [90] . this approach is being prospectively studied as part of an aspirin primary prevention trial. could other immunomodulatory approaches be considered with less putative dangerous consequences on inflamed tissues. this may be the case of thymosin-α1. indeed, a very promising study demonstrated its efficiency to improve clinical outcome in patients with severe sepsis [91] , after a preliminary investigation had demonstrated a better performance with respect to organ failure scores in thymosin-α1-treated patients with sepsis arising from intra-abdominal infection due to carbapenem-resistant bacteria [92] . however, one must call for caution since thymosin-α1 can also favor the production of inflammatory cytokines and nitric oxide and further increases the percentage of t reg cells [93, 94] . still, very little is known of its effect on leukocytes present in different compartments. the cell surface molecules containing in their intracytoplasmic domain an immunoreceptor tyrosine-based inhibition motif -such as programmed death-1 (pd-1), b and t lymphocyte attenuator (btla), and cytotoxic t-lymphocyte antigen 4 (ctla-4) -could also be interesting targets for new therapeutic approaches. the expression of pd-1 on t cells and its ligand (pd-l1) on monocytes is upregulated in critically ill [95] or septic shock [96] patients. increased expressions were associated with increased occurrence of secondary nosocomial infections and mortality after septic shock [97] . not only are pd-1-deficient mice markedly protected from the lethality of sepsis, accompanied by a decreased bacterial burden and suppressed inflammatory cytokine response [98] , but also blockade of pd-1 or pd-l1 improves survival in a murine model of sepsis, reverses immune dysfunction, inhibits lymphocyte apoptosis, and attenuates organ dysfunction [99] [100] [101] . the relevance of these observations in human settings is still needed. ctla-4 is a high-avidity receptor for cd80 and cd86. enhanced ctla-4 expression was demonstrated more frequently in patients with sepsis than in non-infected critically ill patients or control subjects [102] , and blocking ctla-4 improved survival in bacterial and fungal experimental sepsis [103, 104] . however, the use of such an approach seems tricky since, in animal models at high dose, anti-ctla-4 could worsen survival [103] , and the use of abatacept (a soluble ctla-4 dimerized with an fc fragment of immunoglobulin) led to increased survival in invasive pneumococcal infection [105] . similarly, btla expression is enhanced in patients with sirs or sepsis [106] and, in a murine model of sepsis, btla-deficient mice displayed an enhanced resistance [107] . in contrast, these mice displayed enhanced susceptibility to endotoxin-induced shock [108] . accordingly, the exact role of btla needs to be further deciphered before strategies targeting btla could be proposed to treat patients with sepsis. new therapeutic approaches to treat sepsis should take into consideration that the immune status of leukocytes in the peripheral blood might be quite different from those present in inflamed tissues. we believe that a systemic approach to immune stimulation is not appropriate if immune cells are boosted generally, independent of their location. an ideal drug would limit the overzealous inflammatory process that leads to organ failure and favor homeostatic responsiveness of leukocytes ( figure 2 ). this is the challenge we have to address if we wish to avoid further decades of disillusionment. figure 2 new therapeutic interventions should address both the events in the tissues that lead to organ failure and the altered immune status of leukocytes restricted to some specific compartments. cars: compensatory anti-inflammatory response syndrome; cmv: cytomegalovirus; ctla-4: cytotoxic t-lymphocyte antigen 4; dc: dendritic cell; gm-csf: granulocyte-macrophage colony-stimulating factor; hksa: heat-killed staphylococcus aureus hsv: herpes simplex virus; ifnγ: interferon-gamma lps: lipopolysaccharide; marco: macrophage-associated receptor with a collagenous base; mof: multiple organ failure; nk: natural killer pd-1: programmed death-1; pd-l1: programmed death-1 ligand prostaglandin e 2 ; pm: post-mortem; sirs: systemic inflammatory response syndrome; sr-a: scavenger receptor-a; tgfβ: transforming growth factor-beta th: t helper; tlr: toll-like receptor; tnf: tumor necrosis factor; t reg : regulatory t lymphocyte; vap: ventilator-associated pneumonia immunosuppression in sepsis: a novel understanding of the disorder and a new therapeutic approach paradoxical anti-inflammatory actions of tnf-alpha: inhibition of il-12 and il-23 via tnf receptor 1 in macrophages and dendritic cells tnf suppresses acute intestinal inflammation by inducing local glucocorticoid synthesis interleukin 1 receptor signaling regulates duba expression and facilitates toll-like receptor 9-driven antiinflammatory cytokine production systemic interferon-gamma suppresses the development of endotoxin-induced uveitis in mice proinflammatory effects of il-10 during human endotoxemia adib-conquy m: adherence modifies the regulation of gene expression induced by interleukin-10 a: interleukin-10 and the interleukin-10 receptor hepatic over-expression of tgf-beta1 promotes lps-induced inflammatory cytokine secretion by liver cells and endotoxemic shock regulation of immune responses by prostaglandin e2 efficacy and safety of monoclonal antibody to human tumor necrosis factor alpha in patients with sepsis syndrome. a randomized, controlled, double-blind, multicenter clinical trial. tnf-alpha mab sepsis study group assessment of the safety and efficacy of the monoclonal anti-tumor necrosis factor antibody-fragment, mak 195 f, in patients with sepsis and septic shock: a multicenter, randomized, placebo-controlled, dose-ranging study continuous plasmafiltration coupled with sorbent adsorption in a rabbit model of endotoxic shock neutralization of tumor necrosis factor in preclinical models of sepsis interferon-gamma increases mortality following cecal ligation and puncture granulocytemacrophage-colony-stimulating factor-dependent peritoneal macrophage responses determine survival in experimentally induced peritonitis and sepsis in mice role of t cells for cytokine production and outcome in a model of acute septic peritonitis role of t cells in a murine model of escherichia coli sepsis adoptive transfer of in vitro-stimulated cd4 + cd25+ regulatory t cells increases bacterial clearance and improves survival in polymicrobial sepsis neutralization of interleukin-10 or transforming growth factor-beta decreases the percentages of cd4+ cd25+ foxp3+ regulatory t cells in septic mice, thereby leading to an improved survival natural killer (nk) cells in antibacterial innate immunity: angels or devils cd11c + dendritic cells are required for survival in murine polymicrobial sepsis altered gene expression patterns in dendritic cells after severe trauma: implications for systemic inflammation and organ injury overexpression of bcl-2 in transgenic mice decreases apoptosis and improves survival in sepsis tgf-beta released by apoptotic t cells contributes to an immunosuppressive milieu apoptotic cells protect mice 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experimental sepsis by inhibiting lymphocyte apoptosis and reversing monocyte dysfunction pd-l1 blockade attenuated sepsis-induced liver injury in a mouse cecal ligation and puncture model decreased response to recall antigens is associated with depressed costimulatory receptor expression in septic critically ill patients dose-dependent effect of anti-ctla-4 on survival in sepsis blockade of the negative co-stimulatory molecules pd-1 and ctla-4 improves survival in primary and secondary fungal sepsis inhibition of t cells provides protection against early invasive pneumococcal disease b and t lymphocyte attenuator expression on cd4+ t-cells associates with sepsis and subsequent infections in icu patients btla expression contributes to septic morbidity and mortality by inducing innate inflammatory cell dysfunction b and t lymphocyte attenuator inhibits lps-induced endotoxic shock by suppressing toll-like receptor 4 signaling in innate immune cells cite this article as: cavaillon et al.: is boosting the immune system in sepsis appropriate? critical care key: cord-017376-wrhkfcff authors: liu, yongjian; hao, yu; bhandari, suwas; jin, shengwei title: pro-resolution of inflammation: new hints to manage sepsis? date: 2019-05-28 journal: severe trauma and sepsis doi: 10.1007/978-981-13-3353-8_8 sha: doc_id: 17376 cord_uid: wrhkfcff sepsis is newly defined as life-threatening organ dysfunction caused by a dysregulated host response to infection. the pathophysiological mechanism of sepsis is highly complex, and the mortality of in-patients suffering from sepsis is more than 10%. severe unmanaged inflammation and inappropriate immune response characterize sepsis. anti-inflammation therapies alone are not successful for the reason that disbalance of anti-inflammatory and pro-resolving agents. in the recent researches, the host responses during the course of self-resolving infections are found to have the involvements of specialized pro-resolution mediators (spms), namely, lipoxins, resolvins, protectins and maresins. these endogenous lipid metabolites are core signal molecules in the resolution of inflammation, playing a key role in regulating the inflammation and promoting return to homeostasis. besides, heme oxygenase-1 (ho-1, a sensitive marker for oxidative stress) is also known for upregulation in inflammation profiling. carbon monoxide, synthesized by ho-1, performs multiple stances of anti-inflammation and pro-resolution along with the spms. if the potentially beneficial effects of these mediators would be well evaluated in clinical trials, they present encouraging new hints in managing infectious maladies especially sepsis. sepsis is a complicated series of pathophysiological and biochemical disbalance due to the uncontrolled inflammatory response to the invasive pathogens. its newest definition is defined as organ dysfunction due to infection, which remains a major public health concern that is associated with poor clinical outcomes (in-hospital mortality is more than 10%) and substantial healthcare expenditures [1] . data showed that the cost of medical care for sepsis patients was more than $20 billion (5.2%) of total us hospital costs in 2011 [2] . although the accurate incidence of sepsis is unknown, the reported data are increasing [3, 4] , and conservative estimates indicate that sepsis is a leading cause of mortality in the us hospitals and critical illness in the world [5, 6] . the inflammatory host response was long considered as a passive process, which is terminated by the clearance of pro-inflammatory mediators. recently, it is recognized that the initiation and cessation of immune and inflammatory responses is an initiative and symphony process. the pathogenesis of sepsis is intricate. the early stages of sepsis are characterized by excessive generation of inflammatory mediators; however, as sepsis develops into severe chronic sepsis, immunosuppression dominates. this demonstrates that excessive uncontrolled inflammation and inappropriate immune response characterize sepsis [7] . the new lights into sepsis contribute to the explanation of failure in current treatment strategies, including inhibition of the activation phase of the acute inflammatory response to infection (e.g. glucocorticoids, nonsteroidal anti-inflammatory drugs and anti-tnf-α drugs) [8] . therefore, it's urgent and beneficial to explore new paradigms in managing sepsis. as of recent studies relating the host response to a self-resolving infection, it is brought into light that a genus of endogenous bioactive lipid mediators (spms) is produced by the innate immune system cells. a process of stereoselective enzyme conversion of essential fatty acids including arachidonic acid (aa), eicosapentaenoic acid (epa), docosahexaenoic acid (dha) and n-3 docosapentaenoic acid (n-3 dpa) produces these molecules. the synthetases include cyclooxygenase (cox), lipoxygenase (lox) and cytochrome p-450 monooxygenase. the spms can further be subdivided into lipoxins, resolvins, protectins, maresins and immunoresolvents (rvts, pctrs and mctrs) ( fig. 8.1 ) [9] [10] [11] [12] . through the g-protein-coupled receptor-dependent manner, these spms exhibit the fundamental bioactivities in the maintaining of host responses. hence, the productions of pro-inflammatory cytokines (tnf-α and il-1β) and chemokines are inhibited, and the downregulation of inflammation-initiating eicosanoids (prostaglandins and leukotrienes) is promoted. this as well regulates the leukocyte infiltration and stimulates the macrophage efferocytosis of apoptotic cells and debris [10, 11] . recent studies have shown that the spms exhibit beneficial impacts at the site and in course of sepsis while maintaining the homeostasis between the abundance of microorganisms and the host. in this chapter, we aim at: (1) the functions of spms in sepsis. (2) because aspirin partakes in the synthesis of lipoxins and resolvins, studies related to aspirin-triggered lipid mediators are included [13] . (3) protective effects of carbon monoxide (co) on inflammation and its impacts of enhanced productions of pro-resolving lipids [14, 15] . therefore carbon monoxide and its synthetase ho-1 are summarized. (4) since annexin (a1) is a stress response protein, the current evidence for beneficial pro-resolving effects of this mediator in sepsis is also addressed in this chapter [16] . lipoxins (trihydroxy-tetraene-containing eicosanoids) are derived from the omega-6 polyunsaturated fatty acid arachidonic acid (aa) through sequential reactions involving lipoxygenase enzymes, including 5-lipoxygenase (5-lox), 12-lipoxygenase (12-lox) and 15-lipoxygenase type 1 (15-lox-1). there are two series of lipoxins, lipoxin-a4 (lxa4) and lipoxin-b4 (lxb4) [17] . we mainly summarize the activities of lxa4 in this chapter. lipoxins exert effects through a g-protein-coupled receptor, formyl peptide receptor-2 (fpr2/alx). this receptor mainly expresses on the membrane of cells in the immune system along with the resident fibroblasts and epithelial cells [18] . in general, lipoxins are potent inhibitors of inflammatory activity, inducing the resolution of leukocyte activity. they block the response to leukotrienes and chemotactic factors such as leukotriene b4 (ltb4) and platelet-activating factor [19] . these effects inhibit the infiltration of pro-inflammatory cells. the resolution of inflammation involves reduction of neutrophil recruitment, promotion of macrophage migration and augmentation of apoptotic cells efferocytosis. lipoxins exert immune-modulatory activity as well [17, 18] . lxa4, treated after injury, was proved to limit inhaled lps-induced lung injury [20] . in other experimental models, 5 h after caecal ligation and puncture (clp), lxa4 was administered to rats which resulted in decreasing of plasma il-6, chemokine (c-c motif) ligand 2 (ccl2) and il-10 and reducing nf-κb activity in peritoneal macrophages. lxa4 further enhanced the phagocytosis of macrophage recruited to the peritoneum and reduced bacterial load in the blood. hence, lxa4 improved the mortality of this clp model [21] . further study showed that flavocoxid, a dual inhibitor of cox-2 and 5-lox, reduces the expression of nf-κb, cox-2 and 5-lox which improves survival rate in a murine clp sepsis model. plasma il-10 and lxa4 concentrations are increased, while tumour necrosis factor-α (tnf-α), il-6, ltb4 and pge2 are decreased [22] . neutrophil phagocytosis involving the fc receptor i (cd64) is increased and enhanced by lxa4. meanwhile, lxa4 decreases the release of the exotoxin pyocyanin by pseudomonas aeruginosa, reducing its pathogenicity [23] . moreover, lxa4 blocks trafficking of neutrophils, inhibits their adhesion and their release of azurophilic granules. and in activated t-cells, lxa4 lowers tnf release [24] . the finding shows that lxa4 not only modulates the host response but also affects bacterial toxicity [23] . a test was run to investigate the role of receptor fpr2/alx in mediating the protective effects in sepsis using the receptor agonist bml-111. it ameliorates intestinal inflammation in septic rats. the anti-inflammatory cytokine transforming growth factor-β (tgf-β) which is supposed to protect intestinal cells from apoptotic cell death was induced [25] . in a murine model of non-lethal polymicrobial sepsis, fpr2/alx-deficient animals developed more serious disease and exhibit higher cytokine levels and reduced recruitment of monocytes in peritoneal lavages. treatment with an fpr2/alx agonist protected wild type but not the knock-out mice from cardiac dysfunction [26] . these findings prove that lxa4 protect from organ dysfunction, a major cause of mortality in sepsis [27] . as known to all, acute lung injury (ali) and the acute respiratory distress syndrome (ards) are major common complications of severe sepsis [27] . with the inoculation of klebsiella pneumoniae in mice, pulmonary sepsis occurred and induced lxa4 and fpr2/alx expression in the lung. later, the treatment of receptor antagonists and inhibition of 5-lipoxygenase and 15-lipoxygenase in early sepsis (1 h postinfection) even increased leukocyte migration to the infected tissues, and survival rate increased. on the contrary, receptor agonist and lxa4 application consequently worsened early infection and reduced migration of leukocytes. but, 24 h postinfection, lxa4 improved animal survival. here, this research demonstrates the dual role of lxa4 and highlights the time dependence when targeting the lxa4 pathway in pulmonary infection [28] . in the case of septic patients, all the mediators identified in preclinical studies and tested for the treatment in clinical trials have failed [29] . pro-resolving lipid analysis in critically ill patients may reveal a novel orientation for treatment and bring in further insights into the pathways playing a role in the pathophysiology of sepsis. while comparing to 27 non-survival septic patients for 28 days of admission to the intensive care unit, lxa4 was significantly reduced in 39 patients that survived, but levels of this lipid were not associated with death [30] . resolvins are also derived from omega-3 polyunsaturated fatty acids and exist as two series (d and e). e-series resolvins (rve1 and rve2) are products of eicosapentaenoic acid (epa) involving 5-lox, cytochrome p450 and aspirin-acetylated cox-2 as well. d-series resolvins (rvd1-rvd6) are synthesized from docosahexaenoic acid (dha) metabolism involving enzymes 5-lox/15-lox [13, 31] . the biological activities of resolvins are similar to lipoxins. rve1 and its analogues are more potent than lxa4 on a molar basis. rve1 binds to the leukotriene receptor blt1 and blocks tnf-α-stimulated nf-κb activation at the chemr23 receptor [32] . in a mouse model of aspiration pneumonia and subsequent involvement of one lobe with e. coli infection, the function of rve1 in acute lung injury was analysed and found that the rve1, when injected before the acid injury, reduced pulmonary neutrophil infiltration and enhanced bacterial clearance. this was accompanied by lower levels of inflammatory cytokines and chemokines and marginally improved survival rate [33] . in other two murine models of acute lung injury, rve1 enhanced cell death of neutrophils arising from the phagocytosis of opsonized e. coli or yeast and is mediated by the leukotriene b4 receptor blt1. consequently, rve1 enhanced the resolution of the established pulmonary inflammation [34] . lxa4, rve1 and protectin d1 increase levels of the c-c chemokine receptor 5 (ccr5) on apoptotic polymorphonuclear cells (pmns) and thereby terminate chemokine signalling [35] . rve1 and 15-epi-lipoxin protect macrophages from oxidative stress-associated apoptotic cell death, and this contributes to the removal of cytotoxic debris and the inflammation resolution [36, 37] . d-galactosamine-sensitized mouse endotoxin shock mode was also tested for the effects of rvd1, which counteracted the induction of high-mobility group box-1 (hmgb1) and pro-inflammatory cytokines. hepatocyte apoptosis was suppressed, and also neutrophil immigration to the peritoneum was reduced by the effects of rvd1 [38] . in a mouse model of intraperitoneal e. coli peritonitis, rvd5 enhanced phagocytosis of bacteria compared to the control group. rvd1 had a similar but smaller effect. both rvd1 and rvd5 reduced significantly titre of viable bacteria in peritoneal exudates and blood and lowered degree of hypothermia as well. plasma levels of pro-inflammatory cytokines (tnf-α and il-1β) were reduced by rvd1 and rvd5. interestingly, it was found that rvd1 enhanced the antimicrobial effect of ciprofloxacin in resolving e. coli peritonitis and increased survival rate in this model [39] . administration of resolvin d2 could improve outcomes of burn-related sepsis by regulating pmn chemotaxis. in a rat model of burn-related sepsis, rvd2 restored the chemotaxis of pmn to almost normal level. furthermore, when burned rats received intravenous lps 9 days after their burn injury, with intravenous pretreated rvd2, the survival of rats improved significantly. similarly, rvd2 pretreatment increased survival, following caecal ligation after burn injury [40] . rvd1 injected after clp model of sepsis increased bacterial clearance and mice survival. the numbers of peritoneal neutrophils were decreased, while cd3 t-lymphocytes apoptosis in thymus got significant improvement [41] . when at-rvd1 was administered 1 h after the toxin in a lps-induced acute kidney injury mouse model, renal function was improved. lower expression of adhesion molecules, less activation of nf-κb and reduced infiltration of neutrophils have been reported as well [42] . previous studies have shown that iv administration of rvd2 on a clp sepsis model exhibits the following protective pro-resolution effects and increases survival rate: (1) reduce viable aerobic bacterial load in peritoneal exudates and blood; (2) reduce pmn migration into the peritoneum; (3) reduce plasma levels of il-10 and il-17; (4) reduce pro-inflammatory cytokine (il-6, il-1β, il-23 and tnf-α) levels in plasma and peritoneum; (5) reduce concentrations of the pro-inflammatory lipids pge2 and ltb4; while (6) increase clearance of bacteria by phagocytes in inguinal lymph nodes and in vitro; (7) enhance phagocytosis of e. coli by human pmn and also increase intracellular production of reactive oxygen species; and (8) increase survival as a result [43] . inflammatory pain is mainly caused due to the activation of transient receptor potential subtype vanilloid 1 (trpv1) and trp ankyryn 1 (trpa1). rvd2 is a potent inhibitor of both channels in primary sensory neurons, while rve1 inhibits trpv1 and rvd1 inhibits trpa1, respectively. hence, these lipids contribute to pain-relieving activities [44] . the potential therapeutic uses of exogenous resolvins are currently under investigation. in a recent phase 2 clinical trial involving patients with dry eye syndrome, an rve1 analogue significantly improved signs and symptoms of corneal inflammation. this is the first trial to show the clinical efficacy of the novel class of resolvins therapeutics that stimulate resolution rather than inhibit inflammatory mediators [14] . protectins are also omega-3 polyunsaturated fatty acid derivatives, generated from docosahexaenoic acid (dha) through 12-lox/15-lox-mediated pathways. neuroprotectin d1 (npd1) and protectin d1 (pd1) are included [45] . in a severe human influenza model, h1n1-a virus was introduced through the intratracheal route in mice, and pd1 levels were found to be reduced, suggesting that endogenous production was suppressed by the virus. when other influenza a virus strains were tested, the reduction of pd1 was found to inversely correlate with the virulence of the virus strain used. in the same model of severe human influenza, intravenous administration of pd1 reduced in vitro replication of the h1n1 influenza a virus and increased survival. besides, when pd1 was administered in combination with the antiviral agent peramivir, a dramatic increase in survival was found compared to use of peramivir alone [46] . protectin dx (pdx, an isomer of protectin d1) also exhibits protective effects of anti-inflammation and pro-resolution. in a clp sepsis mice model, pdx increased overall survival rate and attenuated multiple organ injury. in addition, pdx reduced pro-inflammatory cytokines and bacterial load 24 h after clp. moreover, pdx promoted phagocytosis of peritoneal macrophages and increased the percentage of m2 macrophages in peritoneum of septic mice [47] . in a recent report encompassing 22 sepsis patients, it was shown that plasma levels of inflammation-initiating mediators including pgf2α and ltb4 and proresolving mediators, including rve1, rvd5 and 17r-pd1, were significantly higher in non-survivors than in surviving sepsis subjects. further analysis revealed increased respiratory failure in non-survivors. these results indicate that peripheral blood lipid mediator profiles (rve1, rvd5 and 17r-pd1) in sepsis correlate with survival and ards development, thus suggesting plausible novel biomarkers and biological targets for critical illness [48] . maresins are a new family of anti-inflammatory and pro-resolving lipid mediators derived from docosahexaenoic acid (dha) by macrophages via human 12-lipoxygenase (12-lox). the first member of this family, termed as maresin 1 (mar1), exhibits potent phagocyte-directed actions that include inhibition of neutrophil infiltration and stimulation of macrophage efferocytosis by dihydroxyl products in this pathway [10, 49, 50] . in a murine model of ards, lipid mediator metabololipidomics discovered that mar1 was temporally generated and regulated in vivo. early intravascular mar1 was organ-protective, and mar1 production was dependent on platelet-neutrophil interactions leading to reduced lung neutrophils, oedema, tissue hypoxia and inflammatory mediators [51] . pro-resolution effects are activated to terminate inflammation as soon as the inflammatory response initiates. in a peritonitis model, mar1 was one of the first spms upregulated in the peritoneum during self-resolving infections. acting as a partial agonist/antagonist to the ltb4 receptor (blt1), mar1 supressed the activity of ltb4 so as to promote the uptake and clearance of apoptotic cells and bacteria [52] . interestingly, levels of mar1 and ltb4 reached a maximum level in the early stages of the inflammatory response, suggesting that early mar1 production impacts leukocyte infiltration to the inflammatory site [39, 52] . in a lps-induced ali mice model, high dose of mar1 exhibited protective activities by mitigating patho-histological changes, attenuating pulmonary oedema and restoring oxygenation. besides, high-dose mar1 inhibited the increasement of pro-inflammatory cytokines (tnf-α, il-1β and il-6) and chemokines, while antiinflammatory cytokine il-10 was upregulated. moreover, mar1 lowered lpsinduced neutrophil adhesions and suppressed the expression of intercellular adhesion molecule (icam)-1, p-selection and cd24 [53, 54] . another study reported that mar1 can maintain the permeability of lung epithelial cells by upregulating the expression of claudin-1 and zo-1 in lps-induced ali [55] . in a clp sepsis mice model, it was found that mar1 markedly mitigated the levels of pro-inflammatory cytokines (tnf-α, il-1β and il-6). intervention of mar1 lowered the lps level in serum and enhanced the bacterial clearance. furthermore, mar1 attenuated lung injury and decreased serum level of alanine transaminase (alt), aspartate transaminase (ast), creatinine (cre) and blood urea nitrogen (bun) in this sepsis model. consequently, the survival rate was improved. inhibition of nf-κb activation pathway by mar1 is the possible protective mechanism [56] . maresin 2 (mar2) was identified later. mar2 exhibits similar potency to mar1 in limiting pmn recruitment but has an apparent optimal concentration 2-3 log orders lower than mar1 in enhancing human macrophage phagocytosis of zymosan. mar2 also enhanced human macrophage uptake of apoptotic pmn but was less potent than mar1 [57] . aspirin is a classic anti-platelet anti-inflammatory agent. acetylating cox-2 facilitates aspirin to induce a shift from the synthesis of pro-inflammatory to pro-resolving lipid mediators termed as aspirin-triggered lipoxins (at-lx) and aspirin-triggered resolvins (at-rv) [13, 58] . at-lx and at-rv share the pro-resolution effects of lxa4 and rvd1, respectively, and act by the same intracellular pathways [59] . the aspirin-triggered lipoxin, also known as 15-epi-lxa4 (at-lxa4), was found to be increased with administration of aspirin in both in vitro and in vivo models of infection; with infection alone increase in 15-epi-lxa4 levels was observed [60] . in murine models of sepsis and ards, aspirin increased the survival rate effectively [61] . low dosage of aspirin administered 30 min prior to the endotoxin model of sepsis improved the survival and reduced the levels of thromboxane and prostaglandins derived from arachidonic acid [62] . another research indicated that low-dose aspirin, however, did not reduce cytokine and prostaglandin levels while enhancing 15-epi-lxa4 synthesis [63] . in an e. coli sepsis model, 15-epi-lxa4 injected 24 h after the injury lowered pmn number in broncho-alveolar lavage by stimulating the apoptosis [64] . several observational clinical studies have confirmed the benefits of aspirin in those who took it prior to medical consult in sepsis patients. in a group of 1149 critically ill patients, 25% of them with preclinical aspirin use had a decreased risk of developing to ards and tended to a lower mortality [65] . a different study with 5523 patients suffering from systemic inflammatory response syndrome or sepsis suggested that a lower mortality was found in 2082 patients when given aspirin within 24 h after diagnosis [66] . similarly, in a study comprising 1005 patients with community-acquired pneumonia, 100 mg/day intake of aspirin was associated with a lower mortality rate within 30 days [67] . hence, a clinical trial indicates that the aspirin and essential fatty acid supplemented in healthy human volunteers increase the endogenous production of pro-resolving mediators including 17-r-pd1 and augment bacterial clearance of leukocytes [68] . however, a multicentre study with 3855 patients did not pinpoint a significant association between preclinical aspirin therapy and progression to ards [69] . a prospective observational study with 972 patients indicated that preclinical aspirin therapy was neither associated with the development of organ failure nor shock nor 90-day mortality up to 90 days after hospitalization [70] . evidence above does not make a solid statement of conclusion on exact beneficial prognoses of aspirin therapy in sepsis. further studies are required to evaluate a potential protective effect of aspirin in sepsis patients. a novel family of pro-resolving mediators termed 13 series resolvins (rvts) were identified recently in the very early stages (≤4 h) of self-resolving e. coli infections model. these rvts mediators include rvt1, rvt2, rvt3 and rvt4, which are derived from n-3 docosapentaenoic acid (n-3 dpa) through sequential reactions involving cox-2/5-lox in the crosstalk of vascular endothelia and neutrophils. each of these molecules activates the host immunity in a dose-dependent manner for bacterial clearance and counter-regulates the production of pro-inflammatory molecules including endothelin-1, plasminogen activator inhibitor-1 and inflammatory eicosanoids. moreover, concentrations of rvts in peripheral blood increased rapidly after exercise (a self-resolving inflammatory state) in healthy volunteers and were significantly higher in patients with sepsis than in healthy subjects. these results indicate that rvts biosynthesis occurs in a coordinated manner in correlation with acute activation of the immune response. therefore, inability to form these pathways leads to delayed resolution responses and an impaired ability to clear bacterial infections [71] . recent studies suggest that the formation of the immunoresolvent protectin conjugate in tissue regenerations (pctrs: pctr1, pctr2 and pctr3) by group 3 innate lymphoid cells (ilc-3), which in turn regulates peritoneal macrophage responses to bacterial infections, is promoted by the vagus nerve [72] . pctrs are derived from dha through 15-lox-mediated pathways in leukocytes. since pctrs actively promote the termination of bacterial infections by stimulating the uptake and killing the bacteria as well as the repair and regeneration of damaged tissues, these mediators are immunoresolvents [73] . pctrs parallelly promote a macrophage phenotype shift, downregulating the production of pro-inflammatory cytokines including tnf-α and il-8 and increase the production of regenerative molecules including tgf-β [74] . any form of damage to the vagus reflex dysregulates pctrs formation and macrophage phenotype leading to an impaired ability of the recruited leukocytes to efficiently clear pathogens causing the delayed resolution of inflammatory infections [72] . further researches into mediators formed during the late phases of resolution of e. coli infections pointed out a huge group of molecules that are peptide-lipid conjugates. these molecules termed as maresin conjugates in tissue regeneration (mctrs: mctr1, mctr2 and mctr3) are produced by 14-lipoxygenation of dha through 12-lox-mediated pathways in human macrophages. mctrs exhibit some potential benefits in regulating bacterial phagocytosis, promoting tissue repair and regeneration [74, 75] . both pctrs and mctrs were found in inflammatory exudates and spleens from infected mice as well as in human plasma, serum and spleens. the levels of these mediators were increased during the later stages of infectious inflammation in mice [73, 74] . carbon monoxide (co) enhances the immune cell function. co is produced by the constitutively expressed heme oxygenase-2 (ho)-2, and by ho-1, which is upregulated upon cellular stress. ho-1 catalyses the degradation of heme to biliverdin, co and iron (which binds to ferritin), all acting as anti-oxidative and anti-inflammatory agents [76] . endogenously formed co acts as a signalling molecule and induces antioxidant genes [76, 77] . co downregulates inflammatory prostaglandins and thromboxanes. on the other hand, it enhances the expression of lipoxygenases which are key synthetases of spms [15] . resolvins and lipoxins in turn upregulate ho-1 in macrophages demonstrating mutual amplification of these two proresolving pathways [15] . co contributes to active pathways on the elimination of microorganisms, promote killing of bacteria and enhance their clearance by macrophages in a protective manner [78] . furthermore, pro-inflammatory cytokines including tlr2, -4, -5 and -9 are repressed, while anti-inflammatory cytokines such as il-10 are initiated by co in macrophages [79] . enhanced endocytosis of apoptotic cells by efferocytosis is also known [15] . in murine sepsis model, higher mortality and hepatic necrosis in ho-1-deficient mice group were found pointing out the protective function of this enzyme [80] . hepatoprotective effects were reported with increasing levels of co in endotoxic rats [81] . hepatic accumulation of pmn, expression of the intercellular adhesion molecule-1 (icam-1) and activation of nf-κb in murine polymicrobial sepsis are reduced by co. in endotoxin-activated human umbilical vein, the endothelial cells cocultured with co, the production of reactive oxygen species (ros), nitric oxide (no), activation of nf-κb, induction of inducible no synthase and icam-1 and pmn adhesion were reduced [82] . in clp mice model, levels of circulating inflammatory cytokines and the number of bacteria in blood and organs were decreased by the inhalation of co which resulted in the increased survival rate of the mice [83] . co inhalation, 2 h prior to initiation of peritonitis, lowered the number of infiltrating pmn. monocyte numbers remained unchanged, while the clearance of microbial pathogens and dead pmn significantly raised [15] . inflammation resolved nearly two times faster than co nonexposed group. lipid mediators derived from arachidonic acid, ltb4 and pge were low in the early stage of inflammation and pro-resolving lipids including rvd1, rve2 and maresin, were markedly high [15] . the clinical patients have been observed for the protective roles of co and ho-1. patients with lower respiratory tract infection indeed had higher co in their breath, and levels descended in those patients recovering from disease after antibiotic treatment [84] . when compared to 30 healthy neonates, co in plasma of 7 neonates with sepsis was significantly increased [85] . when 36 patients with severe sepsis or septic shock were compared to 21 patients without sepsis, arterial blood co and ho-1 protein levels in monocytes were ascended. these two molecules were positively related to survival [86] . exhaled co is higher in severely ill patients compared to the control group [87] . in mechanically ventilated patients with severe sepsis or septic shock, exhaled co was nearly threefold higher compared to controls and declined along with therapy, and high amount of co in exhaled air on the first day during treatment was associated with better outcome [88] . co is reported to have beneficial impacts in different patient groups; therefore, inhalation of co would be a potential policy guide for the treatment of sepsis [77] . annexin a1 is a 37 kda monomeric protein, existing abundantly in some pivotal cell types of the innate and adaptive immune systems and the neuroendocrine system. the synthesis and release of annexin a1 is regulated by glucocorticoids (gcs) and expresses as a stress response protein through binding to formyl peptide family receptors (fprs) [89] . an increasing number of experimental evidence in present days indicate that annexin a1 is crucial to many of the acute actions of gcs in several systems relevant to the stress response, including the innate and adaptive host immune systems [89, 90] and the hpa axis [91] . in annexin a1-null mice models of inflammation, treating with anti-annexin a1 antibodies or antisense constructs of annexin a1, have confirmed that annexin a1 exhibits as an endogenous regulator of antiinflammatory and pro-resolving as well as a mediator of gc action [89, 90] . annexin a1, binding to fpr in the innate immune system, exerts general suppressive activities including the reduction of the amounts of pro-inflammatory eicosanoids molecules generated and the release of histamine and preformed cytokines from mast cells. in contrast, anx-a1 may induce the release of pro-resolving mediators, such as il-10. furthermore, annexin a1 highly downregulates pmn migration into inflammatory sites and accelerates their apoptosis, while the migration of monocytes into inflammatory sites is promoted by annexin a1 which eventually transform the microenvironment into a resolving procedure [92] . in an in vivo experiment, the absence of annexin a1 or its major receptor (fpr2) amplifies greatly the duration and intensity of acute and chronic inflammation. consequently, the acute antiinflammatory actions of gcs are greatly reduced or even abrogated. conversely, human recombinant annexin a1 and peptides derived from the n-terminal domain can rescue this phenotype and exert high anti-inflammatory and pro-resolution effects in many models of inflammation [90, 92] . sepsis is a severe organ dysfunction syndrome due to uncontrolled infection with a high mortality risk. despite of inspiring results in preclinical models, none of the presumptive therapeutic agents tested so far succeed in clinical trials. excessive uncontrolled inflammation and inappropriate immune responses are both characteristic features of sepsis that complicate identification of suitable drug targets. inhibition of inflammation may delay the resolution because of an improper induction of anti-inflammatory and pro-resolving pathways. overcoming sepsis, there is still a long way to run. an accumulating evidence decodes the role that endogenous resolutions triggered by spms demonstrate the ensured internal homeostasis in host body by activating the ultimate cardinal signs of resolution that include pathogen and cellular debris lavage, analgesia and restoration of organ functions. latest studies point out that monitoring the concentrations of these spms in infected patients may provide a better understanding of the inflammation-resolution procedures and hence reflect a positive outcome [48] . yet, requiring further prospective studies to confirm these observations. the points and the hints that are mentioned in this review indicate that spms act as potential biomarkers and, more importantly, hold a great promise as novel therapeutics in inflammatory diseases. latest studies reveal that the majority of end-stage fatal sepsis patients are immunosuppressed rather than hyperinflammation [93] . given, the potential anti-inflammatory and pro-resolution benefits of these spms (including co, ho-1 and annexin a1) mentioned above, endogenous or exogenous supplement of these mediators analogues or natural extracts may be a new paradigm in treating sepsis. the third international consensus definitions for sepsis and septic shock (sepsis-3) national inpatient hospital costs: the most expensive conditions by payer population burden of long-term survivorship after severe sepsis in older americans benchmarking the incidence and mortality of severe sepsis in the united states assessment of the worldwide burden of critical illness: the intensive care over nations (icon) audit international forum of acute care trialists. assessment of global incidence and mortality of hospital-treated sepsis: current estimates and limitations is sepsis a pro-resolution deficiency disorder? med hypotheses resolvin infectious inflammation by targeting the host response pro-resolving lipid mediators are leads for resolution physiology protectins and maresins: new pro-resolving families of mediators in acute inflammation and resolution bioactive metabolome treating inflammation and infection in the 21st century: new hints from decoding resolution mediators and mechanisms does promoting resolution instead of inhibiting inflammation represent the new paradigm in treating infections? resolvins: a family of bioactive products of omega-3 fatty acid transformation circuits initiated by aspirin treatment that counter proinflammation signals novel endogenous proresolving molecules: essential fatty acid-derived and gaseous mediators in the resolution of inflammation inhaled carbon monoxide accelerates resolution of inflammation via unique proresolving mediator-heme oxygenase-1 circuits annexin a1 and resolution of inflammation: tissue repairing properties and signalling signature lipoxins and aspirin-triggered 15-epi-lipoxins are the first lipid mediators of endogenous anti-inflammation and resolution novel anti-inflammatory--pro-resolving mediators and their receptors the human serum metabolome posttreatment with aspirin-triggered lipoxin a4 analog attenuates lipopolysaccharideinduced acute lung injury in mice: the role of heme oxygenase-1 lipoxin a4 increases survival by decreasing systemic inflammation and bacterial load in sepsis flavocoxid, a dual inhibitor of cox-2 and 5-lox of natural origin, attenuates the inflammatory response and protects mice from sepsis lipoxin a4 augments host defense in sepsis and reduces pseudomonas aeruginosa virulence through quorum sensing inhibition aspirin-triggered lipoxin a4 and b4 analogs block extracellular signal-regulated kinase-dependent tnf-alpha secretion from human t cells effect of bml-111 on the intestinal mucosal barrier in sepsis and its mechanism of action nonredundant protective properties of fpr2/alx in polymicrobial murine sepsis ards and the multiple organ dysfunction syndrome: common mechanisms of a common systemic process dual role of lipoxin a4 in pneumosepsis pathogenesis why have clinical trials in sepsis failed? plasma levels in sepsis patients of annexin a1, lipoxin a4, macrophage inflammatory protein-3a, and neutrophil gelatinase-associated lipocalin resolvins and protectins in inflammation resolution protective effect of resolvin e1 on the development of asthmatic airway inflammation the anti-inflammatory and proresolving mediator resolvin e1 protects mice from bacterial pneumonia and acute lung injury resolvin e1 promotes phagocytosis-induced neutrophil apoptosis and accelerates resolution of pulmonary inflammation apoptotic neutrophils and t cells sequester chemokines during immune response resolution through modulation of ccr5 expression resolvin d1-mediated nox2 inactivation rescues macrophages undertaking efferocytosis from oxidative stress-induced apoptosis activation of autophagy in macrophages by pro-resolving lipid mediators suppressive action of resolvin d1 on the production and release of septic mediators in d-galactosamine-sensitized endotoxin shock mice infection regulates pro-resolving mediators that lower antibiotic requirements resolvin d2 restores neutrophil directionality and improves survival after burns resolvin d1 improves survival in experimental sepsis through reducing bacterial load and preventing excessive activation of inflammatory response aspirin-triggered resolvin d1 down-regulates inflammatory responses and protects against endotoxin-induced acute kidney injury resolvin d2 is a potent regulator of leukocytes and controls microbial sepsis resolvin d2 is a potent endogenous inhibitor fortransient receptor potential subtype v1/a1, inflammatory pain, and spinal cord synaptic plasticity in mice: distinct roles of resolvin d1, d2, and e1 anti-inflammatory actions of neuroprotectin d1/protectin d1 and its natural stereoisomers: assignments of dihydroxy-containing docosatrienes the lipid mediator protectin d1 inhibits influenza virus replication and improves severe influenza protectin dx increases survival in a mouse model of sepsis by ameliorating inflammation and modulating macrophage phenotype human sepsis eicosanoid and proresolving lipid mediator temporal profiles: correlations with survival and clinical outcomes maresins: novel macrophage mediators with potent anti-inflammatory and proresolving actions total synthesis and bioactivities of two proposed structures of maresin maresin 1 biosynthesis during platelet-neutrophil interactions is organprotective identification and actions of the maresin 1 metabolome in infectious inflammation maresin 1 mitigates lps-induced acute lung injury in mice maresin 1 prevents lipopolysaccharide-induced neutrophil survival and accelerates resolution of acute lung injury maresin 1 maintains the permeability of lung epithelial cells in vitro and in vivo maresin 1 mitigates inflammatory response and protects mice from sepsis maresin biosynthesis and identification of maresin 2, a new anti-inflammatory and pro-resolving mediator from human macrophages novel lipid mediators promote resolution of acute inflammation: impact of aspirin and statins resolvin d1 and aspirin-triggered resolvin d1 promote resolution of allergic airways responses protective role of acetylsalicylic acid in experimental trypanosoma cruzi infection: evidence of a 15-epi-lipoxin a 4 -mediated effect aspirin as a potential treatment in sepsis or acute respiratory distress syndrome studies on the beneficial effects of aspirin in endotoxic shock: relationship to inhibition of arachidonic acid metabolism effects of low-dose aspirin on acute inflammatory responses in humans 15-epi-lipoxin a4 inhibits myeloperoxidase signaling and enhances resolution of acute lung injury prehospital aspirin use is associated with reduced risk of acute respiratory distress syndrome in critically ill patients: a propensity-adjusted analysis acetyl salicylic acid usage and mortality in critically ill patients with the systemic inflammatory response syndrome and sepsis lower mortality rate in elderly patients with community-onset pneumonia on treatment with aspirin identification and signature profiles for pro-resolving and inflammatory lipid mediators in human tissue association of prehospitalization aspirin therapy and acute lung injury: results of a multicenter international observational study of at-risk patients chronic antiplatelet therapy is not associated with alterations in the presentation, outcome, or host response biomarkers during sepsis: a propensity-matched analysis elucidation of novel 13-series resolvins that increase with atorvastatin and clear infections vagal regulation of group 3 innate lymphoid cells and the immunoresolvent pctr1 controls infection resolution novel proresolving and tissue-regenerative resolvin and protectinsulfido-conjugated pathways the protectin pctr1 is produced by human m2 macrophages and enhances resolution of infectious inflammation maresin conjugates in tissue regeneration biosynthesis enzymes in human macrophages eets and ho-1 cross-talk carbon monoxide in the treatment of sepsis macrophages sense and kill bacteria through carbon monoxide-dependent inflammasome activation carbon monoxide differentially inhibits tlr signaling pathways by regulating ros-induced trafficking of tlrs to lipid rafts reduced stress defense in heme oxygenase 1-deficient cells carbon monoxide from heme catabolism protects against hepatobiliary dysfunction in endotoxin-treated rat liver carbon monoxide liberated from carbon monoxide-releasing molecule corm-2 attenuates inflammation in the liver of septic mice carbon monoxide confers protection in sepsis by enhancing beclin 1-dependent autophagy and phagocytosis exhaled carbon monoxide in patients with lower respiratory tract infection plasma carbon monoxide levels in term newborn infants with sepsis beneficial effects of the heme oxygenase-1/carbon monoxide system in patients with severe sepsis/septic shock increased heme catabolism in critically ill patients: correlation among exhaled carbon monoxide, arterial carboxyhemoglobin, and serum bilirubin ix alpha concentrations increased endogenous carbon monoxide production in severe sepsis annexin-a1: a pivotal regulator of the innate and adaptive immune systems annexin a1 and glucocorticoids as effectors of the resolution of inflammation annexin 1, glucocorticoids, and the neuroendocrine-immune interface annexin i is stored within gelatinase granules of human neutrophil and mobilized on the cell surface upon adhesion but not phagocytosis sepsis-induced immunosuppression: from cellular dysfunctions to immunotherapy key: cord-018840-ts2g1ux7 authors: katragkou, aspasia; roilides, emmanuel; walsh, thomas j. title: role of immunoglobulin therapy to prevent and treat infections date: 2018-06-19 journal: management of infections in the immunocompromised host doi: 10.1007/978-3-319-77674-3_17 sha: doc_id: 18840 cord_uid: ts2g1ux7 immunoglobulins have been used widely in medicine for a variety of diseases including infectious diseases. while the main clinical applications of immunoglobulin therapy concern their use as replacement for patients with primary immunodeficiencies, or as treatment for autoimmune and inflammatory disorders, their role in infectious disease is limited largely to viral and toxin neutralization and replacement therapy in patients with immunoglobulin deficiencies. many aspects of the therapeutic regimen of immunoglobulins even in the established indications remain open. recently, due to the worldwide surge of immunosuppression caused by aids, organ transplantation, cancer, and autoimmune therapies, as well as the emergence of multidrug-resistant bacteria, there has been renewed interest in the use of antibody preparation to prevent infections in high-risk groups. knowing the limitations of the current anti-infective armamentarium, approaches that target the host through manipulations to augment the host immune response provide a helpful aid to conventional treatment options. a substantial body of evidence has demonstrated that strategies aiming to support or stimulate immune response could be feasible approaches that would benefit immunocompromised patients. in the present chapter, we present contemporary indications of immunoglobulin administration for therapy and prophylaxis of infections in the immunocompromised population. salmonella species, no consistently effective sera were produced [28] . with the discovery of antibiotics in 1940s, the interest in "serum therapy" for the treatment of infectious disease waned. the initial interest in using combination therapies with antibiotics and serum was abandoned, as the potential benefits were marginal. antimicrobial chemotherapy proved to be less toxic and more effective than serum therapy in the treatment of infections. however, dr. cohn's discovery of purified antibodies through cold ethanol fractionation of plasma during the second world war revived the interest in antibody treatment especially for infections not able to be treated with antibiotics. the fractionation procedure stabilizes the product, denatures most viruses, and assures a more uniform antibody content. cohn fraction (igg from plasma after cold alcohol fractionation) was initially used for prophylaxis against prevalent and life-threatening infections, such as measles. it was not until 1952 that bruton reported for the first time the use of immunoglobulin preparation injected subcutaneously for the treatment of a young boy with agammaglobulinemia [21] . thereafter, the use of immunoglobulin injected intramuscularly became established as the standard therapy for primary immunodeficiencies, lasting until the development of purer and safer intravenous immunoglobulin preparation in the early 1980s [86] . the advent of hybridoma technology, which allows continuous generation of large quantities of monoclonal antibodies specific to antigens of interest and the generation of humanized antibodies, revolutionized antibody therapeutics [63] . monoclonal antibody technology offers supply advantage, reduces the risks of adverse events, and decreases lot-to-lot variation. in the mid-1980s a monoclonal antibody (mab) to cd3 was introduced into clinical practice to prevent organ rejection. almost a decade later, the humanized mab palivizumab (synagis®, a humanized mouse monoclonal antibody to prevent rsv pulmonary infections in high-risk patients, especially infants) was licensed. palivizumab was 50-fold more potent than the polyclonal product, resulting in reduced volume of administration and intramuscular use [45] . during the last three decades, 30 therapeutic mabs have been licensed, mainly for treatment of malignancies and rheumatic or autoimmune diseases, but only two were licensed for infectious diseases (palivizumab and raxibacumab: human mab to anthrax toxin). although the use of mabs to treat infectious diseases does not depend on discrimination between self-antigen as there are large antigenic differences between the microorganism and the host, the pace of discovery and development of new mabs against infectious disease is limited. currently, the areas of mabs development have been focused on viral diseases without available vaccines [hiv, ebola, severe acute respiratory syndrome (sars), middle east respiratory syndrome, marburg virus], viral disease with limited effective antiviral drugs (influenza, rabies), and bacterial toxinmediated disease (anthrax, clostridium difficile colitis). in the clinical setting, therapeutic mabs can be used when there are nosocomial/iatrogenic outbreaks. for example, a new approach to the prevention of recurrent c. difficile infection is the administration of mabs against c. difficile toxins (in addition to antibiotic therapy) as a form of passive immunity. actoxumab and bezlotoxumab are fully human monoclonal antibodies that bind and neutralize c. difficile toxins a and b, respectively. a double-blind randomized placebo-controlled phase 3 trial showed that a single intravenous dose of bezlotoxumab when given with standard-of-care antibiotics provided protection against recurrent c. difficile infection for up to 12 weeks that was superior to that provided by treatment with standard-of-care antibiotics alone [114] . other therapeutic mabs can be applied in drug resistance (staphylococcus aureus, vrsa), pandemic outbreaks (ebola virus), bioterrorism attacks (bacillus anthracis), emerging infectious diseases (nipah or hendra virus), and use in high-risk host groups or in severe diseases (respiratory syncytial virus, cytomegalovirus retinitis in hiv patients, hepatitis c virus, influenza virus). another application of therapeutic mabs concerns their use as adjunct therapies that have anti-inflammatory or immune modulatory roles (mabs against tnf-α and other immune mediators) [45, 55] . immunoglobulins are glycoprotein molecules that are produced by plasma cells in response to antigens or immunogens and which function as antibodies. serum contains a heterogeneous immunoglobulin pool that reflects the host response to endogenous microbiota and the immunological memory of the host for a variety of acquired microbial agents [26] . different immunoglobulins can differ structurally; however, they are built from the same basic units. there are five classes of immunoglobulin, classified according to the type of heavy chain they possess (table 17 .1) [70] . each class of immunoglobulins has a specific function, and deficiency of each class leads to particular dysfunction of immune system. serum igm predominates in the acute immune response to most antigens and is the most efficient complementfixing immunoglobulin. immunoglobulin class switching subsequently occurs, leading to a predominance of igg, which is responsible for protection during the first infectious attack and long-term protection via memory b cells. secretory iga, due to its abundance in mucosal secretions, provides primary defense mechanism against some mucosal infections. ige primarily defends against parasitic invasion [2] . immunoglobulins together with t cells are the key mediators of adaptive immunity, and deficiencies in either of these two arms of the adaptive immune system can result in higher host susceptibility to bacterial, fungal, or viral infections [76] . immunoglobulins interact with the [38, 102] . understanding the mechanisms of interactions between immunoglobulins, immunomodulatory molecules, and cells of the immune system, both innate and adaptive, is the basis for understanding the future therapeutic perspectives of immunoglobulins [38] . immunoglobulins, upon binding of a specific antigen, stimulate significant direct and indirect "effector functions." classically, in bacterial disease, immunoglobulins neutralize toxins, facilitate opsonization, and, with complement, promote bacteriolysis. in viral diseases, immunoglobulins block viral entry into uninfected cells, promote antibody-directed cell-mediated cytotoxicity by natural killer cells, and neutralize virus alone or with the participation with the complement [62] . furthermore, more recent studies have demonstrated the immunomodulatory functions of antibodies, including the potential for antibody therapy to reduce damage from the host inflammatory response to major infections [24, 25] . notably, igg can exert pro-and antiinflammatory activities depending on its concentration. low dose of igg has pro-inflammatory activity and requires complement activation or binding of the fc fragment from igg to iggspecific receptors (fcγr) on innate immune effector cells. this results in receptor clustering, recruitment of secondary effector functions, and subsequent activation of signaling pathways, leading to an increase in intracellular calcium levels and cell activation. by comparison, high concentrations of igg have anti-inflammatory properties. the mechanisms proposed for this mode of action are modulation of the expression and function of fcγrs, interference with activation of the complement cascade and the cytokine network, neutralization of autoantibodies, and regulation of cell proliferation [38] . the immunoglobulin preparations used in passive immunization are the standard human serum immunoglobulin, which is available in three forms: immune globulin (ig) for intramuscular use (imig), intravenous use (ivig), and subcutaneous use (scig). imig is used primarily for the prevention of certain infections, such as hepatitis a, measles, and rubella, and less commonly for the treatment of antibody immunodeficiencies. ivig is used in the treatment of primary and secondary antibody deficiencies, many immunoregulatory disorders (e.g., immune thrombocytopenic purpura, kawasaki disease), and neurologic disorders (e.g., guillain-barré syndrome, peripheral neuritis). igsc is used exclusively for the antibody deficiencies. ivig preparations comprise the pooled fraction of serum from ~3000 to 60,000 donors, which is generated by a cold ethanol precipitation, providing, thus, a broad spectrum of opsonic and neutralizing igg antibodies. opsonic and neutralizing igg antibody content varies with each product batch, primarily due to differences in the local pathogen ecology of donor exposure. igg and complement proteins are the principal classes of opsonins contributing to bacterial clearance. in addition to igg, varying amounts of immunoglobulin isotypes, especially iga, can be found in the ivig preparation. regarding the different human igg subclasses (igg1-igg4), ivig preparations reflect the hierarchy present in the serum, consisting mainly of igg1 and igg2 and containing much smaller amounts of the other igg subclasses. only the product pentaglobin® (biotest, germany) is igm-enriched [93] . the clinical use of ivig can be distinguished by the infused amount [92] . the principal manufacturing process in all current igg preparations is cold ethanol fractionation with product-specific additional processes for manufacturing. the commonest processes for virus reduction include solvents/detergent, low ph (ph 4), incubation, nanofiltration, and chromatography [93] . other major quality control practices in the production process, besides viral reduction, include the depletion of blood coagulation factors and the removal of igg aggregates, since these aggregates could result in a cytokine release syndrome owing to the ubiquitous activation of innate immune effector cells via activating fcγrs. igg aggregations are absent from the majority of ivig preparation; however, depending on the provider and batch, up to 1-10% of igg can be found in dimeric form in most ivig preparations [92] . the two major indications for which immunoglobulins are used are igg replacement therapy and anti-inflammatory therapy in a variety of acute and chronic autoimmune diseases. apart from immunoglobulin replacement therapy, currently licensed application of immunoglobulin (ivig) administration includes guillain-barré syndrome, kawasaki disease, and chronic inflammatory demyelinating polyneuropathy. licensed indications, however, only account for approximately 40-50% of the worldwide immunoglobulin sales, as most immunoglobulin administrations are "off-label" [76] . the use of immunoglobulins for infectious disease can involve the passive transfer of antibodies for pre-/postexposure prophylaxis or for treatment. passive immunization provides temporal immunity to unimmunized individuals either prophylactically or therapeuti-cally. the different forms of passive immunotherapies are shown in table 17 .2 [96] . the technology of ethanol fractionation of plasma resulted in products used for the treatment and prophylaxis of infectious diseases (table 17. 3). human immune sera have fewer adverse effects, but there are concerns about availability, potency, and consistency. table 17 .4 summarizes the adverse reactions of immunoglobulin used in the prevention and treatment of infectious diseases. administration of immunoglobulins is indicated for the majority of patients with primary immune deficiencies and for patients with combined immune deficiencies and for those with secondary immune deficiency with significant antibody deficiency. the benefits of replacement immunoglobulin therapy for the prevention of infections in patients with antibody deficiencies are well established and pertain to the reduction of the incidence and the severity of infections and prevention long-term deterioration in organ function [12, 13] . primary immune deficiencies (pidd) are one of the us food and drug administration (fda)approved indications for immunoglobulin therapy. over 80% of all pidd involve antibody-mediated immunity; however, each individual disorder has a different immunopathogenesis in terms of the number of b cells in the blood and b-cell function. moreover, any persisting endogenous antibody production varies both between specific conditions and within individual disorders [10, 35, 97] . table 17 .5 describes the pidd for which immunoglobulin replacement is or may be efficacious. the recommendations for immunoglobulin replacement treatment in primary and secondary immune deficiencies are shown in table 17 .6 [32, 84, 97] . the main indications are primary antibody deficiencies including agammaglobulinemia (autosomal recessive or x-linked) and common variable immunodeficiency disorders. rarely, other antibody deficiencies, such as igg subclass deficiency, may be managed by immunoglobulin replacement. in these immunodeficiencies, a trial of 12 months may be indicated if there is a substantial infection burden. on the contrary, for selective iga deficiency, immunoglobulin replacement is not required or recommended, as anaphylactic reactions may occur during ivig infusions. combined immunodeficiencies with antibody deficiency also benefit from immunoglobulin therapy until the defects in cell-mediated immunity are corrected by hematopoietic stem cell transplantation. however, b-cell function is not restored universally after transplantation, and immunoglobulin therapy may be continued [86] . it is important that each patient receives a thorough evaluation before starting immunoglobulin therapy especially those with partial antibody defects. the american academy of allergy, asthma, and immunology, based on a 2006 review of evidence, recommends for pidd the dose of 400-600 mg/kg of ivig every 4 weeks, titrating the dose and interval between infusions to achieve a trough igg level at least greater than 500 mg/dl in agammaglobulinemic patients [84] . however, recent evidence suggests that the goal of igg replacement therapy should be to reduce or prevent serious or recurrent infections instead of aiming to achieve a specific igg level. the clinicians should identify for each patient with pidd an individual "biological" igg level with which the patient achieves the best clinical outcome instead of trying to reach a specific igg level [16, 17] . the two modes of igg replacement (ivig and scig) have significant pharmacokinetic differences, which are important to know when choosing the mode of igg delivery or switching from ivig to scig. scig causes sustained release of igg and thus attains higher igg trough levels; this mode of delivery may benefit the 10-15% of patients who show increased risk of infection during the 3rd and 4th weeks after receiving ivig or who experience extreme lethargy during the same period. ivig achieves higher peak levels (160% higher than that obtained by sc infusion), and this mode of delivery is usually initially preferred for patients with pidd who are very symptomatic (present with pneumonia or other serious infectious such as sepsis) and who present with pneumonia or for those with other medical problems such as sepsis [16] . ivig has also been used in a number of diseases that cause secondary humoral immunodeficiency. while for the majority of secondary immunodeficiencies, the use of ivig was supported only by anecdotal reports, and b-cell chronic lymphocytic leukemia (cll) and pediatric hiv infection are fda-approved indications. for both, cll and hiv, infections are the most common complications. ivig has been shown to be a useful prophylactic therapy against infections in such patients [29, 53, 54, 72, 95, 110] . administration of ivig in cll patients with hypogammaglobulinemia has been shown to decrease the rate of bacterial infections; however, decision analysis modeling showed that this decrease might not improve the length or quality of treated patients' lives, and, furthermore, it is extraordinarily expensive [110] . the prophylactic administration of ivig in cll patients has not been studied extensively, and, thus, there are no guidelines to define the patient population that would benefit from this treatment; also the optimal dosing and timing of ivig administration remained to be defined. some experts support the use of ivig in selected cases, depending on the history of the patient and especially in patients that ivig has been shown to work in the past. ivig therapy together with antiviral therapy was beneficial in infants and children with aids and hypogammaglobulinemia or two or more bacterial infections in the previous year. other indications for ivig therapy in hiv-infected patients include those with severe parvovirus b19 or measles infection [72, 95, 119] . however, it is important to note that these studies occurred before the era of highly active antiretroviral treatment for hiv [84] . ivig has been utilized in allogeneic bone marrow transplantation (bmt) in an attempt to decrease the incidence of cytomegalovirus (cmv) infection, infections due to other pathogens, and graft-versus-host disease (gvhd). immunoglobulin use in the setting of bmt is fda approved. the rationale for using ivig in transplantation is that the administration of passive antibodies may prevent infections in these immunocompromised patients and especially infections caused by cmv [84] . several randomized controlled trials provided the basis to recommend ivig after allogeneic bmt [19, 31, 46, 87, 98, 116, 117] . meta-analysis of these trials found significant reduction of fatal cmv infections, cmv pneumonia, non-cmv interstitial pneumonia, and transplant-related mortality among patients receiving prophylactic ivig [11] . while an improvement in survival was reported in some studies [46, 49, 118] , a more recent meta-analysis showed that ivig or hyperimmune cmv-ivig had no effect on the reduction of allcause mortality [90] . collectively, the data regarding the benefit of prophylactic administration of ivig after bmt remain controversial and contradictory. in addition, until currently, there is no consensus on the type, schedule, dose, and patients benefiting from ivig. subsequent studies suggested that double prophylaxis consisting of high-dose ivig and ganciclovir was more successful than either treatment alone in reversing cmv pneumonia in patients after bmt [39, 69] . the american society for blood and marrow transplantation does not recommend the routine use of ivig to hematopoietic cell transplant recipients for prophylaxis for cmv disease or for bacterial infections within the first 100 days after transplantation. for patients with severe hypogammaglobulinemia (igg <400 mg/dl), ivig prophylaxis of bacterial infections may be considered. ivig dose and frequency for these patients should be individualized to maintain trough serum igg concentrations >400 mg/dl [103] . routine use of ivig appears to offer little benefit to patients with malignancies undergoing hla-identical sibling bmt [84] . given that the landscape of patients receiving bmt is evolving, it is likely that the available data are outdated, and more updated randomized trials are warranted to inform clinical practice. gvhd and infection are major complications of allogeneic bmt. in vitro and in vivo experimental models showed that the prevention of acute gvhd by ivig is mediated by the induction of apoptosis of activated alloreactive cd4+ expressing cd134+ donor t cells and reducing the amount of ifn-γ produced by donor t cells [22] . ivig was shown to decrease the severity of acute gvhd in recipients of allogeneic bmt [98, 116, 117] . on the contrary, administration of ivig prophylaxis has no effect on the incidence or mortality of chronic gvhd on bmt [99] . while there is no consensus on the optimal dose of ivig, it appears that the incidence of acute gvhd is less in patients receiving higher doses of ivig. the benefits of ivig appear to correlate with igg trough levels where acute gvhd was less frequent among patients achieving maximum serum igg levels ≥3000 mg/dl after the administration of ivig. trough serum igg levels >1200 mg/dl were associated with less severe acute gvhd [1, 33, 40] . over the last decade, ivig usage in solid organ transplantation has increased significantly. there are encouraging data on the role of ivig for the treatment of antibody-mediated rejection, desensitization to hla and/or abo antigens, as well as prevention and treatment of infectious complications for patients undergoing solid organ transplantation [74, 94] . there is also some evidence that ivig may be useful for the treatment of autoimmune cytopenias after solid organ transplantation [91] . dosing of ivig is empiric although higher than those for replacement therapy. especially for the treatment of antibodymediated rejection, the dose is 1-2 gm/kg [23, 38, 58, 60, 61, 76] . the use of higher doses of ivig is related to higher rates of adverse events. these include aseptic meningitis thrombotic events and bronchospasm [59] . sepsis is the systemic inflammatory response of the host to an infectious insult. severe sepsis is characterized by acute organ dysfunction, while septic shock is characterized by hypotension, which is refractory to fluid replacement, or by hyperlactatemia [9] . severe sepsis and septic shock represent one of the oldest and most pressing problems in medicine. care of patients with sepsis has improved over the last decades; however, the incidence of sepsis is increasing along with morbidity and mortality rates especially in critically ill adults. worldwide, the annual incidence of severe sepsis lies between 100 and 300 cases per 100,000 population, and mortality for severe sepsis and septic shock reaches 30% and 50%, respectively [8, 56, 73, 109] . while our understanding of the underlying biologic features of sepsis has made significant progress, the clinical assessment of several new strategies for implementation for sepsis treatment has led to disappointing results [3, 7, 15, 18, 108] . there have been more than 100 randomized clinical trials of strategies to modify the systemic inflammatory response during sepsis; however, no strategy showed to improve dramatically the survival of patients with sepsis [71] . the development of highly purified human plasma-derived polyclonal ivigs presented a very compelling therapy for severe infections including sepsis and septic shock. ivigs have broad and potent activity against microorganisms, their extracellular products, and potent immunomodulatory effects [78] . ivig preparations, in particular igm-enriched preparations, contain antibodies against lipopolysaccharides of escherichia coli, pseudomonas aeruginosa, and klebsiella spp. [104] . the effects of ivigs on the sepsis-induced host response seem to be pleotropic, not yet completely clarified, and are likely to be secondary to both suppression of synthesis and direct scavenging of upstream and downstream mediators of the host response and complex immunomodulatory effects [93] . the cellular effects of immunoglobulins are mediated through the igg constant fragment (fc). immunoglobulin acts as an adaptor between the innate and adaptive immune system by interacting with fc, which mediate both pro-and antiinflammatory signals. ivigs have direct antibacterial effects through pathogen recognition and increased clearance. ivigs also have anti-inflammatory properties mediated by the scavenging of bacterial toxins and proinflammatory cytokines, by immune cell depletion, by the blockade of activating receptors, and by modulating fcγr expression, dendritic cell activity, and t-cell expansion [77, 92, 93] . the challenging pathobiology of sepsis is associated with acquired hypogammaglobulinemia, which seems to prevent optimal pathogen clearance and pathogen toxin scavenging [100, 107, 113] . furthermore, sepsis, by causing endothelial dysfunction and capillary leak together with the iatrogenic fluid resuscitationrelated increase in extravascular volume, eventually causes an alteration in the distribution of immunoglobulins [93] . consequently, it is logical to predict that the administration of ivig during sepsis would be of benefit. in the clinical setting, the role of ivig as an adjunctive treatment in sepsis has been controversial for years. a number of randomized placebo-controlled clinical trials in adult critical care patients evaluating standard polyclonal ivig-or igm-enriched polyclonal adjunctive therapy in severe sepsis as well as the metaanalyses of these trials have been published [5, 65, 67, 75, 88, 105] . positive findings of controlled trials and anecdotal reports have been criticized for methodological weakness including the small number of the patients and adequacy of blinding. the more recent studies, which were more meticulously designed, have shown much less effect of ivig than older, smaller, and less well-designed studies [50] . of note, the studies that used albumin as control showed less benefit of ivig than those that did not [41] . the score-based immunoglobulin g treatment in sepsis (sbits) study, one carefully designed, large study representing almost half of all the adults studied to date, showed no reduction in mortality by ivig in patients with score-defined sepsis and sepsis-induced multi-organ failure [113] . the first clinical trial, which evaluated the effect of igma-enriched immunoglobulin preparation (7.8 g igm, 7.8 g iga, and 49.4 g igg), which have shown to contain superior antibody content against bacterial lipopolysaccharides, in an appreciable number of neutropenic patients with hematologic malignancies and sepsis or septic shock, showed that immunoglobulins had no beneficial effects [51] . however, as the editor comments, the study, with a high evidence level, demonstrates that neutropenic patients with malignancies and low-grade sepsis with no or only one organ failure will not benefit from adjunctive ivig treatment [111] . the prophylaxis and treatment of neonatal sepsis has been a major global priority, and large international trials have been carried out testing ivig ( [57, [79] [80] [81] [82] [83] ; group et al. 2011 ). mortality during hospital stay in infants with clinically suspected infection at trial entry was not significantly different after ivig treatment [81] . the results of the international neonatal immunotherapy study (inis) and recent metaanalyses showed that ivig did not reduce mortality during hospital stay or major disability at 2 years of age in infants with sepsis [48, 83] . based on the results of the inis trial (3493 subjects), routine administration of ivig to prevent mortality in infants with sepsis is not currently recommended [48] . when considering the administration of ivig during sepsis, important aspects that should be taken into account are the dose, the type, the timing, and pharmacokinetics of ivig [6, 50] . while dose-ranging studies have not been completed, studies that used high (>1 g/kg body weight) doses of ivig demonstrated better effects. this seems plausible given the clinical observations in other inflammatory conditions, such as kawasaki disease, where greater effect was noted with higher doses [52] . the type of ivig may have an important effect, possibly in favor of a greater pooled effect of igma-enriched compared with standard preparations of ivig. igma-enriched preparations are associated with greater complement inactivation and improvement in microvascular perfusion in experimental models [112] . however, collectively, the results from animal models and in vitro experiments show contradictory results and do not allow for a definite conclusion regarding the superiority of one specific immunoglobulin preparation in patients with sepsis. in an efficacy study, administration of polyvalent igg versus igma in selected patients at high risk for sepsis was associated with a comparable improvement in disease severity [89] . regarding the timing of ivig administration during sepsis, there is probably a "window of opportunity" in the first days that follow clinical presentation of sepsis [14] . if this window is missed, probabilities of success could be greatly diminished [6] . pharmacokinetic studies of ivig in sepsis have not been performed yet. data for dosage selection in current practice are primarily derived from studies in volunteers and in patients with primary immune deficiencies and other indications for immunomodulation. existing pharmacokinetic studies also do not address immunoglobulin clearance or area under the curve parameters and target serum immunoglobulin concentrations [64] . in addition, it is still unknown whether the main goal of ivig in sepsis is to refill low levels of endogenous immunoglobulins or alternatively whether ivig could exert a beneficial effect regardless of these levels [6] . most studies evaluating the use of ivig for sepsis are small; some have methodological flaws and high-quality, large studies showed no effect [48, 113] . given immunoglobulin high-cost, limited supply and the lack of strong evidence to support their beneficial effect, widely used guidelines either neglect or grade as a weak recommendation the use of polyclonal ivig in sepsis [36] . while clinical judgment may guide immunoglobulin use in individual cases, particularly those due to gram-negative etiologies or streptococcal toxic shock syndrome, these practices are based largely on theoretical rationale, anecdotal, and retrospective clinical observations [50, 66, 106] . the effect of monoclonal antibodies against tumor necrosis factor (tnf)-α has been evaluated in a series of trials on different anti-tnf-αdirected therapies [4, 30, 37, 42] . the long-anticipated sepsis trial (monarcs [monoclonal anti-tnf, a randomized controlled sepsis trial]) reported that afelimomab, which is made up of the fab component of a monoclonal antibody against tnf-α, in patients with severe sepsis and elevated il-6 levels decreased mortality and had a safety profile similar to placebo [85] . however, combining the results of these studies, a small improvement in mortality can be detected [34, 47] . as sepsis is increasingly being considered as an exaggerated, poorly regulated innate immune response to microbial products, by the time of diagnosis, an entire network of cytokines has already been activated. in this regard, the results of the previous studies would have been anticipated, as it seems unlikely that therapy aimed at only one cytokine would by itself have the highly significantly impact on sepsis mortality [34] . immunoglobulins have been used widely in medicine for a variety of diseases including infectious diseases. while the two major indications for immunoglobulin use are as replacement and antiinflammatory therapy in a variety of acute and chronic autoimmune diseases, their use in the prevention and treatment of infectious diseases is emerging as an attractive option especially in the era of multi-antibiotic resistance. many aspects of immunoglobulin therapy remain controversial and contradictory. consequently, immunoglobulin use is sometimes determined by clinical judgment or expert opinion, which is based largely on theoretical rationale, anecdotal, and retrospective clinical observations. gaps of knowledge that need to be addressed are certain categories of patient populations that would benefit from immunoglobulin treatment or prophylaxis, the optimal immunoglobulin dosing, and duration, as well as timing of administration. monoclonal antibody technology has opened a new era in antibody therapy. on many occasions, human monoclonal antibodies have better therapeutic properties than immunoglobulins including low toxicity, longer protective immunity, higher than natural protection, and high specificity. several antibodies for the treatment of bacterial and viral infections have been developed [101] . however, some challenges need to be overcome before they become preferred agents for the treatment and prophylaxis against infectious diseases. biofilms are now acknowledged to contribute to a plethora of chronic and recurrent infections. while treatment or eradication of 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a streptococcal infections veterans administration systemic sepsis cooperative study, g. effect of high-dose glucocorticoid therapy on mortality in patients with clinical signs of systemic sepsis international study of the prevalence and outcomes of infection in intensive care units cost effectiveness of prophylactic intravenous immune globulin in chronic lymphocytic leukemia immunoglobulin treatment in sepsis -is the answer "no"? ccm.0000216189.80613.bb mirror, mirror on the wall, which is the fairest meta-analysis of all? score-based immunoglobulin g therapy of patients with sepsis: the sbits study bezlotoxumab for prevention of recurrent clostridium difficile infection emil von behring and serum therapy intravenous immunoglobulin and cmvseronegative blood products for prevention of cmv infection and disease in bone marrow transplant recipients intravenous immune globulin for prevention of cytomegalovirus infection and interstitial pneumonia after bone marrow transplantation high-dose weekly intravenous immunoglobulin to prevent infections in patients undergoing autologous bone marrow transplantation or severe myelosuppressive therapy. a study of the american bone marrow transplant group does intravenous immune globulin have a role in hiv-infected patients? textbook of pediatric infectious diseases key: cord-298505-r7ihqb96 authors: górski, andrzej; borysowski, jan; międzybrodzki, ryszard title: sepsis, phages, and covid-19 date: 2020-10-15 journal: pathogens doi: 10.3390/pathogens9100844 sha: doc_id: 298505 cord_uid: r7ihqb96 phage therapy has emerged as a potential novel treatment of sepsis for which no decisive progress has been achieved thus far. obviously, phages can help eradicate local bacterial infection and bacteremia that may occur in a syndrome. for example, phages may be helpful in correcting excessive inflammatory responses and aberrant immunity that occur in sepsis. data from animal studies strongly suggest that phages may indeed be an efficient means of therapy for experimentally induced sepsis. in recent years, a number of reports have appeared describing the successful treatment of patients with sepsis. moreover, novel data on the anti-viral potential of phages may be interpreted as suggesting that phages could be used as an adjunct therapy in severe covid-19. thus, clinical trials assessing the value of phage therapy in sepsis, including viral sepsis, are urgently needed. the world health organization (who) considers sepsis to be a global health priority. recently, it has been emphasized that, despite some success in preclinical studies on experimental sepsis, no significant progress has been achieved in clinical therapy [1] . although more than a hundred clinical trials have focused on sepsis, none of them have provided data that could be used for improvements or a cure [2] . furthermore, the most recent data indicate that, despite best efforts to provide protocol-based care pathways, mortality from sepsis may reach nearly 40% [3] . gaidelyte et al. showed that most of the sepsis-causing bacteria carry functional phages that are released and circulate in the blood of septicemic patients [4] . those phages can lyse other isolates of the same bacterial strain but not the pathogenic strain in sepsis. thus, those prophages play a role in clonal selection of pathogens in this disorder. at the same time, it appears reasonable that phage application may potentially be used in the treatment of sepsis based on both well-known anti-bacterial as well as non-bacterial activities of phages, especially those related to their anti-inflammatory and immunomodulating effects [2, 5] . in fact, in addition to data obtained in experimental animals, there are already reports of successful phage therapy in patients with sepsis [2] . in this review, we summarize the progress in treating sepsis with phage therapy over the last three years. leshkasheli et al. demonstrated the therapeutic efficacy of phage therapy in galleria mallonella larvae and in a mouse model of sepsis caused by acinobacter baumanii [6] . similar data were obtained by wu et al., who showed that a phage effective against that pathogen can rescue lethal sepsis mice [7] . interestingly, the effect of a single phage treatment (1 ml ip at a dose of 10 9 ) was comparable to the effect of a phage cocktail containing 14 phages. in experiments performed by wang et al., phage therapy applied concurrently with the inoculation of the pathogen rescued 100% of mice, whilst phage administration applied 1 h after inoculation with a. baumanii reduced survival to approx. 50% [8] . phage therapy efficacy has also been studied in a mouse model of neonatal sepsis caused by escherichia coli, klebsiella pneumoniae, haemophilus influenzae, pseudomonas aeruginosa, citrobacter freundii and moraxella catarrhalis. a single intraperitoneal (ip) phage dose rescued 60-100% of mice depending on the phage dose. more detailed studies have revealed that a concentration as low as 0.01 moi (multiplicity of infection) was effective in rescuing 80% of mice, whilst a 0.001 moi dose rescued only 20% of mice. interestingly, the mice could be rescued even when phage administration was delayed for 24 h after pathogen inoculation. while both a single phage and phage cocktails were effective, optimal results have been achieved with cocktails [9] . high effectiveness of phage therapy in the treatment of experimental sepsis induced by multidrug resistant p. aeruginosa was also confirmed by alvi et al. [10] . phage-treated bacteremic mice had a survival rate of almost 100%, and no viable pathogen could be detected at 96 h post inoculation. the authors estimate that, in order for therapy of sepsis to be efficient, phages should persist in the blood for at least 3-5 h. a native agarose gel electrophoresis was applied to assess phage surface associated with high blood persistence [11] . phages have been shown to upregulate gene expression of an anti-inflammatory cytokine il-r antagonist [12] and downregulate nf-kappa b signaling [13] . interestingly, suppressing nf-kappa b signaling may be beneficial in an experimental mouse model of sepsis [14] . a 2-year-old boy with digeorge syndrome, recalcitrant p. aeruginosa bacteremia, and an allergy to antibiotics received a cocktail of two phages every 6 h intravenously (iv) for 36 h. treatment with antibiotics, including meropenem, tobramycin, and polymyxin b, was also continued. blood cultures turned negative and then positive again after cessation of phage therapy. the therapy was subsequently resumed, and blood cultures were reverted to negative again. thus, in this patient, phage therapy resulted in sterilization of the blood [15] . jennes et al. described a patient with acute kidney injury complicated by septicemia caused by colistin-only sensitive p. aeruginosa and acute kidney failure. the patient received a 50 ml cocktail of two phages in a 6 h iv infusion for 10 days. blood cultures turned negative immediately, c-reactive protein (crp) dropped, fever disappeared, and renal function recovered [16] . recently, australian authors have reported results of adjunctive phage therapy (combined with antibiotics) of s. aureus bacteremia in 13 patients, most of whom suffered from infective endocarditis. a phage cocktail composed of three phages was administered twice daily for 14 days. a 10 9 pfu infused phage dose yielded approx. 2 × 10 5 pfu/ml of blood, which suggests that such a dose may be adequate to achieve a desired therapeutic effect. the iv infusions were well tolerated; no fever, rashes, hypotension or other adverse reactions were observed. clinical improvement was evident in eight of the 13 patients, and inflammation markers declined during or soon after the therapy [17] . further progress in phage therapy of sepsis has recently been achieved by introducing engineered phages used to treat a patient with a disseminated drug resistant mycobacterial infection. genome engineering and forward genetics were applied to obtain lytic phage derivatives that were infused iv as a three-phage cocktail (10 9 pfu per dose of each phage) twice daily for 32 weeks. phage infusion was well tolerated, without significant side effects. serum phage titers reached levels exceeding 10 9 /ml. weak anti-phage protein antibody responses were noted, but no evidence of phage neutralization was observed. the clinical condition of the patient improved while sterilization of the blood and sputum was achieved [18] . sepsis is one of the principal causes of morbidity and mortality in neonates and young children in low and middle-income countries. recently, scientists in iraq have developed a cocktail of phages against pathogens implicated in neonatal sepsis in a bagdad teaching hospital (e. coli, k. pneumoniae, h. influenzae, p. aeruginosa, c. freundii, and m. catarrhalis). the cocktail containing 29 phages showed activity against all bacterial hosts in vitro and should be further examined for its in vivo efficacy in experimental and clinical sepsis [19] . in summary, in animal studies, phages were administered ip at a dose of 10 9 -10 10 , and the outcome was assessed as animal rescue and reduction of bacterial burden in organs or blood. in human studies, the outcome was assessed as a clinical improvement combined with laboratory signs (e.g., disappearance of fever, improvement in general patient well-being, improvement in organ function (e.g., renal function), drop of crp, negative blood cultures). no significant adverse effects were observed in treated patients, which is in line with our data on large cohorts of patients [20] . phage treatments elicit antibody responses; however, those serum antibodies do not appear to significantly influence the outcome of therapy [21] . on the other hand, it cannot be excluded that phage therapy also induces the formation of a phage-antibody immune complex. therefore, the clinical significance of a humoral response to phages during the therapy requires further studies. furthermore, more detailed studies on phage pharmacokinetics would be helpful for the advancement of phage therapy. recent data suggest that disturbances in the microbiome can enhance susceptibility to sepsis [1] . interestingly, fecal microbiota transplantation (fmt) may rescue mice from human pathogen-mediated sepsis [22] . additionally, fmt improves survival in sepsis induced in rats [23] . notably, it has been suggested that transfer of phages may play a role in the efficacy of fmt [24, 25] . interestingly, a clinical trial of fmt for patients with covid-19 is ongoing [26] . moreover, promising results have been achieved in subgroups of septic patients treated with the il-r antagonist (28 day mortality 34.6% vs. 64.7% placebo) [1] . in addition to the data presented earlier [2] , these findings seem to strengthen arguments for the potential application of phages in the treatment of sepsis. in addition to phages, phage-derived lytic enzymes (lysins) have also been studied as a potential weapon against multi-drug resistant bacteria. recently, the results of a first placebo-controlled clinical trial involving an antistaphylococcal lysin (exebacase) administered in conjunction with antibiotics in patients with s. aureus septicemia (most with infectious endocarditis) have been published. in comparison to the control (antibiotics only), the exebacase group showed higher responder rates as well as a reduction in length of stay and readmission rates. no hypersensitivity reactions to exebacase were reported. although preexisting anti-lysin antibodies were detectable in some patients, this did not affect the efficacy and the safety of treatment. these results offer the first tangible opportunity to improve clinical results and reduce mortality in staphylococcal sepsis using phage-derived lysin [27] . table 1 summarizes the recent progress in successfully applying phages and lysin in experimental and clinical sepsis. severe acute respiratory syndrome coronavirus (sars-cov-2) probably originated from a virus that has been circulating in horseshoe bats for several decades. it was first detected in wuhan, china, which suggests the existence of an intermediary (pangolin?) that facilitated transmission to humans. there were also hints that the virus escaped or was deliberately released from a local institute [28] . as of 30 september 2020 there have been almost 34 million covid-19 cases worldwide with >1 million deaths, and >25 million patients have recovered [29] . it has been noted that severe covid-19 patients may develop typical manifestations of septic shock with blood and respiratory tract cultures testing negative for bacteria and fungus. therefore, viral sepsis could be responsible for clinical manifestations in those patients in whom systemic cytokine storm, lymphopenia, and thrombotic complications are usually detectable. effective antiviral therapy combined with attempts to modulate the innate immune response and upgrade the adaptive immune response are recommended to improve the outcome [30] [31] [32] . in fact, antiviral and anti-inflammatory treatments may be effective early in the disease [33, 34] . the lungs-the primary target organ of the sars-cov-2 virus-are relatively accessible to phages delivered by different routes, including oral administration. however, nasal or tracheal delivery are preferred to achieve sufficient in situ concentrations. aerosol phage preparations as well as nebulized preparations may serve as the most efficient therapeutic applications [35] . growing data suggests that phages may interfere with the pathogenic action of eukaryotic viruses [36] . this historical data has been supported by new findings indicating a protective action of the t4 phage on human lung epithelial cells infected with human adenovirus (adv); furthermore, adsorption of adv to human lung and kidney epithelial cells as well as viral replication was also inhibited [37] . new data suggest that cell layers of the body may be the major sink for administered phages; interestingly, lung epithelial cells show the highest accumulation of phages [38] . moreover, the expression of adv genes and synthesis of adv dna may also be downregulated by t4 and staphylococcal phages [39] . we hypothesized that phage therapy might be helpful in combatting covid-19 [40] . thus, it is known that cov-expressing cells display markedly upregulated levels of reactive oxygen species (ros), and high levels of ros are observed in the lungs of patients with covid-19 [41, 42] . phages downregulate ros production induced by bacteria and endotoxins [20] . lymphocytopenia is frequently found in covid-19, while the virus is known to induce apoptosis [41, 43] . moreover, autopsies have revealed atrophy of the spleen and the lymph nodes [44] . interestingly, phages may reduce apoptosis of human airway epithelial cells when cultured in vitro [45] . in addition, sweere et al. demonstrated that pf phages cause upregulation of interferon (ifn) alpha and il-12, thus promoting an antiviral signature in the lungs of mice [46] ; gogokhia et al. found that phages of lactobacillus, e. coli and bacteroides stimulate production of another potent antiviral cytokine, ifn gamma [47] . recently, we showed that the t4 phage induces upregulation of the human defensin 2 gene (hbd2) [48] . this peptide, which is exposed primarily by epithelial cells, reduces viral replication and may enhance pathways responsible for other anti-microbial effects, both anti-bacterial and anti-viral. hbd2 activates primary anti-viral innate immune responses [49] . it suppresses hiv infection of hela cells in vitro [50] , drastically reduces human respiratory syncytial virus (hrsv) infection of human lung epithelial cells [51] , and inhibits the infectivity of hiv virions of human tonsil epithelial cells [52] . thus, defensins have been shown to participate in antimicrobial defenses in the human respiratory tract, and the up-regulation of hbd2 may enhance those defenses [53] . therefore, t4-induced hbd2 could also be engaged in mediating anti-sars-cov-2 defenses, which requires experimental confirmation. an inflammatory response causes activation of the hemostatic system (endothelial and platelet activation and coagulation promoting thrombosis)-a syndrome also referred to as thromboinflammation which is relevant in covid-19 [54] . in fact, thrombocytopenia is associated with increased risk of severe disease [55] . platelets are known to interact with viruses and have recently been shown to be transient carriers of hiv, thus contributing to hiv dissemination by propagating the virus to macrophages. this process could be prevented by the anti-integrin alphaiib/beta3 antibody [56] . coronaviruses may also infect bone marrow cells [55] . sars-cov rna may be present in platelets of covid-19 patients, which suggests that platelets can participate in the dissemination of the virus [57] . it would be of interest to determine if this phenomenon could also be blocked by the anti-alphaiib/beta3 antibody, which has been effective in preventing hiv dissemination. the alphaiib/beta3 integrin binds specifically to a kgd (lys-gly-asp) sequence motif exposed on the gp24 capsid protein of t4 phages. therefore, t4 phages could interfere with platelet-dependent sars-cov dissemination, mimicking the effect of the anti-alphaiib/beta3 antibody. in fact, it has been demonstrated that such interference could enable t4 phages to reduce the adhesion of platelets to fibrinogen [13] . interestingly, there appears to be another target for anti-covid-19 effects of phages. sars-cov-2 binds to its receptor, angiotensin-converting enzyme 2 (ace2), through the receptor-binding domain (rbd) present in its major structural protein spike. recently, an exposed kgd motif has been identified in ace2, thus enabling it to interact with integrin alphaiib/beta3 [58] . therefore, platelets could associate with the sars-cov-2-ace2 complex using their alphaiib/beta3 integrin receptor targeting the kgd sequence present within the ace2 molecule. this phenomenon could contribute to sars-cov-2 dissemination and upregulate platelet-mediated coagulopathy and tissue injury. the presence of platelet-fibrin thrombi is common in lung lesions in patients with covid-19 and is considered to be the main target of therapy [59] . occupation of the platelet alphaiib/beta 3 integrin receptor by the phage kgd could inhibit platelet engagement with the complex formed by the virus and its receptor and prevent the ensuing pathology. in recent years, a number of reports derived from experimental studies in animals and human clinics have suggested the potential value of phage therapy in the treatment of sepsis. the activity of both anti-bacterial and non-bacterial phages is relevant for successful phage therapy. the anti-inflammatory and the immunomodulating properties of phages could also be useful in the treatment of severe covid-19 syndrome including viral sepsis (table 2) . a recent article from china concludes that phage therapy in sepsis treatment can be expected in the near future [60] . the data discussed in this review support this assumption. as pointed out, relevant clinical trials assessing the therapeutic value of phage therapy in those clinical settings are urgently needed [40] . 1. phages can interact with epithelial cells and protect those cells from virus-induced damage and apoptosis; this could be especially relevant for lung epithelial cells. 2. phages may prevent viral adsorption to epithelial cells and downregulate viral replication in those cells. 3. phages may induce production of cellular chaperones protecting cells from viral injury (e.g., induction of hsp70 in human alveolar cells). 4. phages inhibit inflammation (downregulation of nuclear factor (nf) kappa b and reactive oxygen species (ros) production). 5. phages induce anti-viral immunity (e.g., induction of interferon (ifn)-alpha and ifn-gamma, defensin 2 and inhibition of hsp90). 6. phages may interfere with severe acute respiratory syndrome coronavirus (sars-cov)-b2 binding to angiotensin-converting enzyme 2 (ace2). author contributions: a.g. wrote the major part of the manuscript. a.g., j.b., and r.m. contributed to the conceptualization and reviewed the manuscript. all authors have read and agreed to the published version of the manuscript. funding: this work was supported by statutory funds of the institute of immunology and experimental therapy, wrocław, poland (501-16). a.g., r.m., and j.b. are co-inventors of patents owned by the institute and covering phage preparations. sepsis therapies: learning from 30 years of failure of translational research to propose new leads the potential of phage therapy in sepsis sepsis and septic shock-basics of diagnosis, pathophysiology and clinical decision making bacteria, phages and septicemia phage therapy: current status and perspectives efficacy of newly isolated and highly potent bacteriophages in a mouse model of extensively drug-resistant acinetobacter baumannii bacteraemia a novel phage pd-6a3, and its endolysin ply6a3, with extended lytic activity against acinetobacter baumannii efficacy of ϕkm18p phage therapy in a murine model of extensively drug-resistant acinetobacter baumannii infection an experimental mouse model for phage therapy of bacterial pathogens causing bacteremia. microb. path rehman ur, s. rlp, a bacteriophage of the family 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key: cord-000522-d498qj2b authors: vincent, jean-louis; abraham, edward; annane, djillali; bernard, gordon; rivers, emanuel; van den berghe, greet title: reducing mortality in sepsis: new directions date: 2002-12-05 journal: crit care doi: 10.1186/cc1860 sha: doc_id: 522 cord_uid: d498qj2b considerable progress has been made in the past few years in the development of therapeutic interventions that can reduce mortality in sepsis. however, encouraging physicians to put the results of new studies into practice is not always simple. a roundtable was thus convened to provide guidance for clinicians on the integration and implementation of new interventions into the intensive care unit (icu). five topics were selected that have been shown in randomized, controlled trials to reduce mortality: limiting the tidal volume in acute lung injury or acute respiratory distress syndrome, early goal-directed therapy, use of drotrecogin alfa (activated), use of moderate doses of steroids, and tight control of blood sugar. one of the principal investigators for each study was invited to participate in the roundtable. the discussions and questions that followed the presentation of data by each panel member enabled a consensus recommendation to be derived regarding when each intervention should be used. each new intervention has a place in the management of patients with sepsis. furthermore, and importantly, the therapies are not mutually exclusive; many patients will need a combination of several approaches – an 'icu package'. the present article provides guidelines from experts in the field on optimal patient selection and timing for each intervention, and provides advice on how to integrate new therapies into icu practice, including protocol development, so that mortality rates from this disease process can be reduced. sepsis is the tenth most common cause of death in the us [1] . a recent us study reported that severe sepsis accounts for in excess of 215,000 deaths annually from a total population of approximately 750,000 patients-a mortality rate of approximately 29% (with published studies quoting a range of 28-50%) [2] . this persistent, high mortality rate is clearly unacceptable, given that it ranks sepsis above some of the higher profile causes of in-hospital death, including stroke (12-19% risk of death in the first 30 days) and acute myocardial infarction (ami) (8% risk of death in the first 30 days) [3] . moreover, the actual number of deaths associated with the condition may be even higher than current estimates suggest. many sepsis patients have at least one comorbidity and deaths are often attributed to these conditions rather than to sepsis [4] [5] [6] . unfamiliarity with the signs and symptoms of sepsis may further hinder accurate diagnosis. there are many possible reasons for this high mortality. sepsis is certainly a complex disease state; the pathophysiology is only now beginning to be unraveled, and it is complicated by heterogeneous presentation (possible signs of sepsis are presented in table 1 ). while none of these signs alone is specific for sepsis, the otherwise unexplained presence of these signs should signal the possibility of a septic response. many cases of sepsis are recognized late, and patients are often inappropriately treated before entering the intensive care unit (icu) by physicians unfamiliar with the signs and symptoms of the condition. furthermore, treatment may be initiated by any of a number of physicians (anesthetists, hematologists, intensivists, infectious disease specialists, pulmonologists, and emergency physicians). there are presently various defined supportive strategies for treating patients with sepsis, but improvements are needed to reduce the unacceptably high mortality rate. moreover, as with other areas of medicine, the application and integration of new but proven strategies for reducing morbidity and mortality into clinical practice has been slow. encouraging new data have recently been presented on new approaches to the management of patients with sepsis. many of these approaches attempt to modulate or interrupt the sepsis cascade and to address the cause of multiorgan dysfunction. although many of these approaches are in early phases of development (e.g. antibodies to tumor necrosis factor [tnf] alpha, bactericidal permeability increasing protein, high-flow hemofiltration to remove circulating inflam-matory mediators, platelet-activating factor acetyl hydrolase, and antielastases), other approaches are more advanced and are already beginning to impact on outcomes in the icu. at a roundtable discussion in london in june 2002, professor jean-louis vincent brought together five experts to discuss more effective implementation of five exciting new interventions in the icu setting to decrease the unacceptable burden of mortality in patients with severe sepsis. each of the roundtable panelists is a highly respected physician in the world of sepsis and critical care medicine. the interventions discussed encompassed low tidal volume in patients with acute lung injury (ali)/acute respiratory distress syndrome (ards) (edward abraham), early goal-directed therapy (egdt) (emanuel rivers), drotrecogin alfa (activated) (gordon bernard), moderate-dose corticosteroids (djillali annane), and tight control of blood sugar (greet van den berghe). the purpose of the roundtable discussion was to provide guidance for clinicians on the integration of new interventions into the icu to reduce the mortality in sepsis, on appropriate patient selection for these interventions, and on appropriate timing of these interventions. the present review reports the discussions and recommendations of the panel. the overall 30-day mortality in the icu is typically ~20% [8] . the 30-day mortality in the population with severe sepsis, defined as sepsis with organ dysfunction, is 30-50%. it is clear from this figure that severe sepsis contributes disproportionately to the overall 30-day mortality in the icu and compares unfavorably with some of the higher profile acute killers in hospital (e.g. stroke and ami) [3] . despite the general improvements in medicine overall, this mortality rate has remained essentially unchanged for the past 25 years. this has contributed to a feeling of pessimism among table 1 possible signs of sepsis (adapted from [ intensivists and other medical professionals regarding treatment prospects for severe sepsis, and a reluctance to rapidly incorporate new interventions into clinical practice [9] . although the sepsis mortality rates are unacceptable, they camouflage some significant developments that are and have been occurring for hospital patients, for the general icu population and, particularly, for those with severe sepsis. direct comparison of mortality rates among patients with identical acute pysiology and chronic health evaluation (apache) scores in the placebo arm of anti-tnf or anti-endotoxin studies published 10-15 years ago [10] [11] [12] with more recent studies [13, 14] , demonstrates that the mortality rate is much lower in more recent studies. interestingly, this decrease was apparent even before the five interventions discussed in the present article were published, reflecting improvements in the general supportive care of sepsis patients. indeed, the panel contends that mortality from septic shock has already been reduced. some patients who in the recent past would have died from severe sepsis or septic shock do not reach the icu now because they are well managed on the wards, in the emergency department, and even during preoperative and postoperative care. for example, those sepsis patients that receive prompt antibiotic therapy have a 10-15% lower mortality rate than those who receive antibiotic therapy later in their care [15] . progress is also being made in diagnosing sepsis: more patients are being tested to identify the source of infection and the pathogens involved, supportive care measures have been improved (e.g. hemodynamic support), and other measures have been put in place to reduce the incidence of nosocomial infections (e.g. reducing the need for pulmonary artery catheters by using echo techniques to assess cardiac function). there has also been a realization of the importance of specially trained intensive care physicians in the icu. it has been internationally recognized that changing the icu from an 'open format', whereby patients are cared for by their admitting physician, to a 'closed format', whereby patients are managed by appointed intensivists, reduces mortality rates [16] . although the mortality rate is beginning to decline, it still remains unacceptably high. furthermore, the number of patients with severe sepsis and septic shock is increasing; people are living longer, and there has been a rise in the number of immunocompromised patients due to aggressive cancer therapy and the increased prevalence of hiv. in-hospital ami-associated mortality rates averaged approximately 25-30% in the 1960s [3] . this clearly unacceptable mortality rate was addressed by the development of a number of new pharmacological and mechanical interventions together with improvements in supportive care. in the landmark second international study of infarct survival trial, published in 1988, 17,187 suspected ami patients were treated with either streptokinase or aspirin, with both drugs, or with neither. the mortality rate in the combination group of this trial was 8%, compared with 13.2% in those patients given neither streptokinase nor aspirin [17] . cardiologists have effectively implemented multiple pharmacologic and supportive care interventions to reduce mortality in ami from 25-30% to 8% and lower. not satisfied with this already remarkable figure, they are trying to reduce it further. physicians treating patients with sepsis are clearly faced with a very different situation to those treating patients with ami, and so direct comparisons are not possible. however, several factors have contributed to the success of ami therapy and possibly to the lack of such success in sepsis (table 2) . sepsis is undoubtedly complicated. however, many of the lessons that have been learned through effective application of therapies in other disease states can be applied to severe sepsis. furthermore, the encouraging data that are beginning to appear in the literature indicate that sepsis may not be as intractable to treat as once thought. the following sections provide salient information on five interventions that have shown a significant positive impact on mortality rates in sepsis, severe sepsis, septic shock, or sepsis-related diseases in recent clinical trials. the interventions were presented at the roundtable by one of the principal investigators of the key trial of the intervention. each section concludes with recommendations for the integration of the particular intervention into clinical practice. the traditional approach in patients with ali/ards is to ventilate using tidal volumes between 10 and 15 ml/kg body weight, almost twice the average tidal volume at rest (7-8 ml/kg body weight), and to maintain a low positive endexpiratory pressure (peep). the purpose of this approach is to achieve normal values for the ph and partial pressure of arterial carbon dioxide. however, this method leads to high inspiratory airway pressures and to excessive stretch of the aerated lung. in 1997, tremblay et al. examined the effect of ventilation strategy on lung inflammatory mediators in the presence and absence of a pre-existing inflammatory stimulus in sprague-dawley rats [18] . in both stimulated and nonstimulated groups, the presence of inflammatory mediators (tnf-α, il-1β, il-6, il-10, macrophage inflammatory protein 2, and ifn-γ) was highest in those rats ventilated with a large tidal volume and zero peep. furthermore, in a study by ranieri et al. in 1999 [19] , the concentration of inflammatory mediators 36 hours after randomization of the groups was significantly lower in the lung-protective strategy group (tidal volume, 7.6 ± 1.1 ml/kg) than in the control group (tidal volume, 11.1 ± 1.3 ml/kg) (p < 0.05). following on from the positive results in the tremblay et al. trial [18] , a small study (53 patients) was carried out by amato et al. in brazil [20] . the mortality rate was 38% in patients given 'protective' ventilation (peep above the lower inflection point on the static pressure-volume curve, tidal volume < 6 ml/kg ideal body weight, driving pressures < 20 cmh 2 o above the peep value, permissive hypercapnia, and preferential use of pressurelimited ventilatory modes) compared with 71% in patients on conventional ventilation (p < 0.001). this impressive reduction in mortality was tempered by the higher than normal mortality level in the control group, prompting the national institutes of health-funded acute respiratory distress syndrome network to set up a similar, larger (861 patients), prospective, multicenter, randomized trial in the us [21] . for a summary of the protocol used in this study, see appendix 1. the trial was stopped after the fourth interim analysis because the use of lower tidal volumes was found to be associated with a significantly reduced mortality (p = 0.005 for the difference in mortality between groups). the primary endpoints were mortality prior to hospital discharge with unassisted breathing and ventilator-free days (days alive, off mechanical ventilation, between enrollment and day 28). both of these endpoints were achieved (figs 1-3 ). in addition, patients receiving a tidal volume of 6 ml/kg ideal body weight had increased organ failure free days and lower il-6 levels. ali is seen in 25-42% of patients with sepsis [22] . although the approach has only been tested in patients with ali/ards, a tidal volume of 6 ml/kg ideal body weight is at the lower end of the range of physiologic ventilation. hence, this approach should be suitable for most patients in the icu setting. furthermore, as many patients with severe sepsis or septic shock progress to frank ali/ards, the panel believes that low tidal volume therapy is a valid option in these patients, and an option that may indeed prevent the development of ali/ards. although patient selection in the clinical trial specified both blood gas and lung infiltrate criteria, at least 90% of patients in the general icu setting meet the criteria for blood gas but table 2 a comparison of acute myocardial infarction (ami) and sepsis market issues significant publicity surrounding and general awareness lack of understanding among physicians and the of the condition; large trials general public diagnosis a relatively straightforward and relatively common complicated by a long list of signs and symptoms diagnosis (electrocardiogram, enzymes, troponin), and and few objective tools for validation one that can be made by generalists, not just cardiology specialists generally single organ disease (notable exception when often chronic or acute comorbidities complicated by cardiogenic shock) generalists have been taught to recognize the signs sepsis patients often come 'second hand' from a and symptoms of ami; initial treatment is usually specialist who may not be appropriately trained to provided by emergency physicians, who are trained diagnose, manage, and refer patients with sepsis to treat these patients mortality prior to hospital discharge in patients receiving a tidal volume of 6 and 12 ml/kg ideal body weight. acidosis is more likely to develop in patients with severe lung problems rather than in those exhibiting milder disease when tidal volumes are kept low. however, acidosis is seldom a clinical problem and rarely requires administration of bicarbonates. one of the issues with low tidal volume therapy is that the patients are often more uncomfortable, at least initially, when they are being ventilated with a tidal volume of 6 ml/kg ideal body weight. the patients tend to exhibit tachypnea and may become more agitated. sedation is generally required, but the ventilator setting can be maintained. of more concern is that icu staff may consider a respiration rate of 40/min to be a sign of something more serious and may attempt to terminate the intervention. education of staff is clearly essential. the strategy assessed in this trial not only includes ventilation with a low tidal volume, but also the provision of extrinsic peep. there may be some concern that an increased respiratory rate may result in intrinsic peep and hemodynamic problems (e.g. decreased cardiac filling, decreased cardiac output, and diminished blood pressure). the panel believes that auto peep was not an issue in the acute respiratory distress syndrome network study. in addition, in the groups with low tidal volume, at least 10% more oxygen was required to maintain the fraction of inspired oxygen (fio 2 ), suggesting that there was very little auto peep occurring. when mechanical ventilation is indicated for treatment of patients with ali/ards, the tidal volume should be limited tõ 6 ml/kg ideal body weight. goal-directed therapy represents an attempt to adjust the cardiac preload, afterload, and contractility to balance systemic oxygen delivery with oxygen demand. in patients with severe sepsis and septic shock, such an approach would seem eminently reasonable as part of general supportive measures to restore and maintain adequate cellular perfusion and to prevent organ dysfunction. in the setting of the icu, however, supranormal and normal approaches have met with little or no success [23, 24] . it is possible that, by the time these therapies are applied in the icu, any such intervention may have been too late. hence, the focus has shifted towards hemodynamic optimization in the early presentation of disease, such as in the emergency department. a prospective, randomized, predominantly blinded study was initiated by the early goal-directed therapy collaborative group to examine the results of hemodynamic interventions in the emergency department [25] . in this study, patients were randomly assigned to either 6 hours of egdt or to standard therapy prior to admission to the icu. baseline characteristics (including the adequacy and duration of antibiotic therapy) in the egdt and standard therapy groups were not significantly different. the vital signs, resuscitation endpoints, organ dysfunction scores, and coagulation-related variables were similar in these groups at baseline [25] . however, there were some important differences between the treatment groups (see table 3 ). available online http://ccforum.com/content/6/s3/s1 proportion of patients alive and off the ventilator having been ventilated with a tidal volume of 6 and 12 ml/kg ideal body weight. median number of ventilator-free days in patients receiving a tidal volume of 6 and 12 ml/kg ideal body weight. patients randomized to egdt received the same therapy but, in addition, were monitored for the endpoint of central venous oxygen saturation (scvo 2 ) > 70%. egdt patients were given more intravenous fluids (including blood transfusions) and more inotropic support (mostly dobutamine). for more information on the protocol used in this study, see appendix 2. key data are presented in table 4 . the in-hospital mortality was 30.5% in the group assigned to egdt and was 46.5% in the group assigned to standard therapy (p = 0.009), indicating that egdt provides significant benefits in improving outcomes in patients with severe sepsis and septic shock. during the interval from 7 to 72 hours, patients assigned to egdt exhibited a more significant improvement in mean scvo 2 (70.4 ± 10.7% versus 65.3 ± 11.4%), in lactate concentration (3.0 ± 4.4 mmol/l versus 3.9 ± 4.4 mmol/), in base deficit (2.0 ± 6.6 mmol/l versus 5.1 ± 6.7 mmol/l), and in ph (7.40 ± 0.12 versus 7.36 ± 0.12) than patients assigned to standard therapy (p ≤ 0.02 for all comparisons). during the same period, the mean apache ii scores were significantly lower, indicating less severe organ dysfunction, in the patients assigned to egdt than in those patients assigned to standard therapy (13.0 ± 6.3 versus 15.9 ± 6.4, p < 0.001). the protocol was based predominantly on guidelines published in 1999 by the society of critical care medicine [26] . however, these guidelines have not been universally followed in clinical practice since their publication. an increasing number of critically ill patients are presenting to, and being treated in, emergency departments [27, 28] . this is present-ing significant resource challenges in the emergency department environment. the inability to institute egdt may thus not be a conscious decision by the clinician not to follow the society of critical care medicine guidelines. emergency medicine in general may have to develop and formulate the cost-benefit analysis to support or implement such care in this environment in order to improve outcomes. there are sufficient evidence-based data to recommend that all patients with severe sepsis or septic shock should receive early and aggressive resuscitation based on this egdt protocol (see appendix 2) . it is important that the interventions are individualized to each patient. a negative or positive value indicates how the control group therapy compares with the treatment group. a p < 0.001, b p = 0.01, c p = 0.02, d p = 0.03, e p = 0.04. egdt, early goal-directed therapy. it is possible to identify patients with profound global myocardial dysfunction who are hence at risk of impaired perfusion. these patients, almost 15% of those in the egdt group, received dobutamine during the first 6 hours because myocardial suppression was diagnosed. once myocardial dysfunction is corrected (and compliance improved), these patients become more suitable for volume loading, so this group received almost 3.5 liters more fluids in the first 6 hours than the control patients. therefore, although vasopressor use was similar in the first 6 hours, patients in the egdt group were more aggressively weaned off these agents during this period, resulting in fewer patients in this group entering the icu on vasopressors than in the control group. the lack of aggressive volume loading in the control group led to greater use of vasopressors in patients over the subsequent 72 hours. in spite of more volume loading, the egdt group received less mechanical ventilation over the subsequent 72 hours than in the standard treatment group. why was cardiovascular collapse a significant cause of death in the control group? cryptic shock (shock with normal vital signs) is a frequent occurrence in early severe sepsis and septic shock. despite resuscitation to the goals for mean arterial blood pressure and cvp, almost 40% of control patients continued to exhibit global tissue hypoxia (decreased scvo 2 and increased lactate levels); in these patients, there was a twofold increase in hemodynamic deterioration, requiring more mechanical ventilation, pulmonary artery catheterization, and vasopressor use in the subsequent 72 hours. how do severe sepsis and septic shock differ hemodynamically in the early stages compared with that classically described in the icu? patients presenting with early sepsis and septic shock are characterized by hypovolemia (low cvp), normal to increased blood pressures, and decreased cardiac output (decreased central venous oxygen saturation and low cardiac index). this is in contrast to icu patients who are euvolemic, have high scvo 2 , and have elevated cardiac indices [29] . what are the most important ways in which egdt can improve outcomes? the key factors are early detection of high-risk patients in cryptic shock, early reversal of hemodynamic perturbations and global tissue hypoxia, prevention of acute cardiovascular collapse, and the possibility of preventing the inflammatory aspects of global tissue hypoxia that accompany the inflammation or infection. severe sepsis and septic shock patients should receive early aggressive therapy to restore and maintain oxygen availability to the cells. there should also be generous use of fluids and inotropic agents titrated by appropriate hemodynamic monitoring. background a large number of observational studies have shown that patients with sepsis have severe depletion of protein c [30, 31] . a number of studies have also shown the association of protein c depletion with high mortality in sepsis [32] [33] [34] . furthermore, baboon studies have demonstrated that treatment with activated protein c prevents death from live escherichia coli infusions [35, 36] . activated protein c exerts a number of actions. anticoagulant action includes the inactivation of coagulation factors va and viiia, and the inhibition of the formation of thrombin. profibrinolytic action allows the activity of tissue plasminogen activator (endogenous tissue plasminogen activator), by inactivating plasminogen activator inhibitor 1 and thrombin activatable fibrinolysis inhibitor. finally, anti-inflammatory action reduces il-6 (in vivo) and proinflammatory cytokines (in vitro). the specific mechanisms by which drotrecogin alfa (activated) exerts its effect on survival in patients with severe sepsis are not completely understood. the efficacy of drotrecogin alfa (activated) (recombinant human activated protein c) in reducing mortality in patients with severe sepsis was investigated in a large multicenter, blinded, placebo-controlled, randomized, phase iii clinical trial, the protein c worldwide evaluation in severe sepsis (prowess) trial [14] . all patients in the prowess trial received standard supportive care in addition to either drotrecogin alfa (activated) or placebo. for a summary of the protocol used in the prowess study, see appendix 3. the overall mortality in patients treated with drotrecogin alfa (activated) was 24.7% compared with 30.8% in patients receiving placebo, an absolute risk reduction of 6.1% (p = 0.006) (see fig. 4 ). the absolute risk reduction in patients with high risk of death defined by an apache ii score ≥ 25 was 12.8% (p < 0.001). the absolute risk reduction in patients with high risk of death defined by multiple organ failure was 7.4% (p = 0.006). no substantial differences in drotrecogin alfa (activated) treatment effects were observed in subgroups defined by gender, ethnic origin, or infectious agent. can drotrecogin alfa (activated) be used in patients on dialysis for pre-existing renal failure, a category that was specifically excluded in the prowess trial? no pharmacokinetic data were available on drotrecogin alfa (activated) in patients on chronic dialysis when the prowess trial began, so such patients were excluded from the trial. subsequent research has shown that the pharmacokinetics of drotrecogin alfa (activated) are not substantially changed in patients on chronic dialysis. the design of the prowess trial allowed a maximum of 48 hours between the onset of first organ dysfunction and the receipt of drotrecogin alfa (activated) (a 24-hour window was allowed for receipt of the drug following the first confirmation of first organ dysfunction, which in turn had to have been present for no more than 24 hours). the treatment effect of drotrecogin alfa (activated) was consistent across all time intervals from meeting the entry criteria to the receipt of the study drug. treatment with drotrecogin alfa (activated) thus does not appear to be as time critical as interventions such as tissue plasminogen activator in stroke or myocardial infarction. because most of the experience with drotrecogin alfa (activated) was based on organ failure times less than 48 hours, treatment should not be delayed when an appropriate candidate is identified. the time window employed in the prowess trial should allow a full history to be taken and other tests to be performed to determine the bleeding risk. as with all anticoagulants, drotrecogin alfa (activated) is associated with a risk of severe bleeding. during the infusion period in the prowess trial, the bleeding rates were 2.4% in the drotrecogin alfa (activated) group versus 1.0% in the placebo group (p = 0.024). the risk of bleeding was fairly constant across most subgroups. however, severe thrombocytopenia (< 30,000/mm 3 ) was commonly associated with serious bleeding and intracerebral hemorrhage. patients at high risk of death in the prowess trial were most likely to benefit from drotrecogin alfa (activated). in the prowess trial, the apache ii score was the most effective predictor of risk of death and likelihood of benefit from drotrecogin alfa (activated), particularly in those patients with an apache ii score ≥ 25. in the prowess trial, the number of organ dysfunctions was also an important indicator that supported an association between likelihood of benefit from drotrecogin alfa (activated) and risk of death. two or more organ dysfunctions identify a population that responds well to therapy, and is a practical measurement. the panel believes that acute respiratory failure or hypotension unresponsive to fluid challenge should suggest the use of drotrecogin alfa (activated). however, coagulopathy, a platelet count < 80,000/mm 3 , acidosis, or low urine output alone should not suggest its use. a very large international study of 11,500 patients will be started in late 2002 to investigate the efficacy of drotrecogin alfa (activated) in patients with a single organ failure and/or apache scores < 25. the decision on whether to administer the drug should ultimately depend on whether the patient meets the selection criteria. a patient presenting in the emergency room with acute respiratory failure or acute cardiovascular decompensation should receive appropriate treatment there. the drawback to treatment in the emergency room is that there may not be sufficient time in which to evaluate the patient's bleeding risks. delaying treatment for a few hours will enable more tests to be performed and a fuller history to be taken, both of which will provide a better indication of whether drotrecogin alfa (activated) is appropriate. the dose is always the same (24 µg/kg/hour), regardless of the type of organ failure or the degree of sepsis severity. in addition, the 96-hour window of treatment is always the same so that interruptions of treatment are made up at the end to maintain a total of 96 hours of treatment. twenty-eight-day survival in patients treated with drotrecogin alfa (activated) or placebo: all-cause mortality. do patients require any laboratory testing before they receive drotrecogin alfa (activated)? no laboratory testing was carried out in the prowess trial, and subgroup analysis identified no biochemical marker that conclusively indicates treatment. for example, treatment-associated reductions in mortality were observed in patients with normal protein c levels and in those with low protein c levels. clinical criteria are recommended for the initiation of therapy. aspirin (650 mg/day) was allowed in the prowess trial. patients on glycoprotein iib/iiia inhibitors were excluded because no data were available regarding drug interactions and pharmacokinetics. use of these types of agents is likely to increase the risk of bleeding with drotrecogin alfa (activated) therapy. the anticipated benefits must therefore be weighed against the potential risks. in the prowess trial, efforts were made to correct the international normalized ratio towards normal if it was greater than 3 at any time during infusion of drotrecogin alfa (activated). approximately one-third of patients in the prowess trial received steroids at the same time as drotrecogin alfa (activated). there was no interaction with steroid use, presumably because the mechanism of action of steroids is so different from that of activated protein c. hence, steroids should be used if they are needed, and if the patient qualifies for drotrecogin alfa (activated) the two should be used together. drotrecogin alfa (activated) should be considered for use in all adult patients with recent onset severe sepsis or septic shock, and a high risk of death. the value of steroids in the treatment of patients with severe sepsis and septic shock has been fiercely debated for some time. although a number of well-designed, randomized, controlled trials failed to show any benefits of steroid therapy in terms of improved survival in patients with severe sepsis (reviewed in [37, 38] ), with mortality increased in many as a result of an increased incidence of nosocomial infections, these trials were primarily investigating the efficacy of short courses of high-dose steroids. the question of whether lower doses of steroids may provide benefit in these patients has only recently been addressed. there is a relatively strong rationale for considering the use of steroids in patients with refractory septic shock. relative adrenal insufficiency is common in patients with refractory septic shock (50-75% of patients) [39] . in addition to such relative adrenal insufficiency and the blunted response to corticotrophin, a large body of evidence indicates that sepsis and refractory septic shock are characterized by peripheral tissue resistance to corticosteroids [40, 41] . in septic patients, this can be evidenced in a variety of ways. first, global cortisol binding, which carries cortisol from the adrenal glands to the tissues, decreases in patients with severe sepsis [42] . second, the number and binding affinity of glucocorticosteroid receptors may be reduced in patients with sepsis and severe sepsis [43] , leading to a decrease in the conversion of cortisone to its active form, cortisol, particularly by il-2 levels in the tissues. finally, data have been published demonstrating that moderate doses of steroids may restore cell sensitivity to vasopressors [44] . this may reduce the intensity of the inflammatory response and decrease organ dysfunction. low-dose steroid treatment is also well tolerated [40] . this body of evidence prompted the initiation of a phase iii randomized, controlled trial performed in 19 centers in france with 300 patients [45] . the aim of the trial was to determine whether moderate-dose corticosteroid therapy affected survival in patients with refractory septic shock and adrenal insufficiency. all patients had to be treated with vasopressor agents and mechanical ventilation. for a summary of the protocol used in this study, see appendix 4. patients were stratified according to their response to the adrenocorticotrophic hormone (acth) test. nonresponders were defined by an increment in cortisol levels < 9 µg/dl or < 250 nm/l after challenge with 250 µg cosyntropin. of the 300 patients included, there were 229 nonresponders to the corticotropin test (placebo, 115 patients; steroids, 114 patients). a significant survival benefit was demonstrated among nonresponders receiving moderate-dose corticosteroids. there were 73 deaths in the placebo group (63%) and 60 deaths in the steroid group (53%) (hazard ratio, 0.67; 95% confidence interval, 0.47-0.95; p = 0.023). no beneficial effects were observed in the subset of patients who were classified as responders. hence, in this paradigm, the acth test serves as a useful prognostic measure. since a beneficial effect was observed in the total population, however, the need for an acth test can be challenged and further studies are required. if an acth test is performed, corticosteroid administration can be started before results are received. moderate-dose corticosteroids should be administered to patients with established refractory septic shock. what is the optimal dose for this intervention? hydrocortisone should be given daily at a dose of 200-300 mg. fludrocortisone should be given daily at a dose of 50 µg. what is the optimal duration for this intervention? moderate doses of steroids should be given for 7 days. hydrocortisone can be administered as serial boluses or as a continuous infusion. it may be that rebound phenomena at treatment discontinuation are more frequent when hydrocortisone is given as a continuous infusion. in addition, in the phase iii randomized trial, hydrocortisone was given as serial boluses. the phase iii randomized trial has shown that the combination of hydrocortisone and fludrocortisone increased survival. in addition, sepsis is more frequently associated with a mineralocorticoid deficiency than a glucocorticoid deficiency. hence, fludrocortisone should be added to hydrocortisone. administration of moderate-dose corticosteroids should be considered in cases of refractory septic shock, particularly in those with relative adrenal insufficiency. it is recommended that an acth test be carried out before starting the intervention. hyperglycemia, caused by insulin resistance in the liver and muscle, is a common finding in icu patients. it can be considered an adaptive response, providing glucose for the brain, red cells, and wound healing, and is generally only treated when blood glucose increases to > 215 mg/dl (> 12 mmol/l). previous studies have shown that high levels of insulin-like growth factor binding protein 1 (a very good marker of lack of hepatic insulin effect) predict mortality [46, 47] . patients with high insulin-like growth factor binding protein 1 also tend to have the lowest insulin levels, indicating that beta cell function is impaired and, therefore, not enough insulin is being produced. these results indicate that hyperglycemia may not always be adaptive and that it should be treated to avoid the onset of specific complications. nevertheless, conventional wisdom in the icu has been that hyperglycemia is beneficial and that hypoglycemia should be avoided. the hypothesis that hyperglycemia (> 110 mg/dl, > 6.1 mmol/l) predisposes to specific icu complications, prolonged intensive care dependency and death was tested in a prospective, randomized, controlled trial [48] . for a summary of the protocol used in this study, see appendix 5. thirty-five of the 765 patients (4.6%) in the intensive insulin group died in the icu, compared with 63 patients (8.0%) in the conventional therapy group. for further mortality data on both the length of hospital stay and the cause of death, see tables 5 and 6 . for morbidity data, see figure 5 . tight control of blood sugar, as outlined in appendix 5, requires a strict protocol for insulin administration and repeated determination of blood sugar. this is yet to be proven, and is the subject of an ongoing study. because medical patients tend to stay in the icu longer than surgical patients, the results from this study indicate that this intervention would be even more favorable to medical icu patients. however, one needs to be careful with application of the algorithm in certain disease states, especially severe hepatic dysfunction and renal failure. no, all carbohydrates are included. see appendix 5 for guidelines on feeding. the level was chosen because it is in the physiologic range for healthy people. as well as its effect on glycemia, insulin has been shown to inhibit tnf-α and macrophage inhibitory factor (when infused concomitantly with glucose). this has led to some doubts as to whether the effect in this study was due to normalization of blood glucose levels. however, multivariate analysis of all the risk factors for mortality, including severity of illness on admission, indicated that blood glucose determines the outcome; there was a 75% increase in risk of death per 50 mg/dl increase in blood glucose. it is not yet possible to determine this. although it was blood glucose levels that were measured, the effects of insulin may in fact be on free fatty acids, as they change in parallel with s11 blood glucose. one of the key mechanisms may be prevention of hypertriglyceridemia and high concentrations of free fatty acids. it is strongly advisable to tightly control blood sugar close to physiologic levels, especially in surgical patients. implemen-tation of this recommendation requires a well-defined icu protocol. the interventions discussed in the present article have been applied in different patient populations and at different times in the course of the disease (see table 7 ). it is essential for physicians to understand that these therapies are not mutually exclusive. optimal patient management may require a combination of approaches: mechanical ventilation to preserve lung function, hemodynamic support to maintain adequate scvo 2 , intensive insulin therapy to normalize blood sugar, steroids to provide adequate immunosuppression, and drotrecogin alfa (activated) to prevent the systemic coagulopathy characteristic of severe sepsis and, hence, to preserve organ function. a sound understanding of the indications and contraindications of these interventions will guide appropriate intervention. similarly, the timing of therapy needs to be closely monitored. education in the signs and symptoms of sepsis and severe sepsis should prompt early initiation of therapy. many of the interventions discussed in this article were tested at specific available online http://ccforum.com/content/6/s3/s1 multiple organ failure, no sepsis focus 18 14 multiple organ failure, with sepsis focus 33 8 most important effects on morbidity [46] . cvvh, continuous venovenous hemofiltration; icu, intensive care unit; nnt, number needed to treat; rrr, relative risk reduction. despite the wealth of data to support the approaches discussed, it is clear that uptake of these interventions into clinical practice has been slow. although there may be practical reasons for this, it would appear in many cases to involve either unfamiliarity with the data or a reluctance, or at least inertia, to change established practices (witness the necessity of proving that hypoglycemia is beneficial in icu patients despite no good evidence to the contrary). the icu has changed in the past 30 years; there are more tools to use and more interventions to implement. despite application of new methods, however, outcomes have changed very little and certainly not in proportion to the changes that were expected based on the results from clinical trials. efficient integration of new interventions into the wider icu population is clearly essential. the panel believes that optimal use of existing therapies and the integration of proven new therapies will reduce mortality rates. further positive results from new trials with improved trial designs should encourage intensivists to incorporate new interventions into their practice. protocols are essential to ensure efficient integration of new therapies and to improve outcomes on the wards. morris predicted in a recent paper that an increase in compliance with evidence-based recommendations through the use of protocols would decrease error and would enhance patient safety [49] . however, a complete treatment protocol is only effective when each ward (inside and outside of the icu) has the trained staff to implement it, and when a skilled intensive care physician is available to lead the team. training and education of staff is essential. all five of the interventions discussed in this article have generated convincing evidence for their use, and they hold out hope for reducing mortality in patients with sepsis, severe sepsis and septic shock. yet, despite compelling data, the application of these interventions has yet to become routine practice in most icus. it is our hope that this article will enable physicians to understand how best to apply these therapies in clinical practice; from appropriate patient selection and timing of therapy, to combining different approaches for optimal patient management. a willingness to embrace new interventions, coupled with the development and implementation of rigorous protocols to ensure appropriate use, will improve outcomes and lead to a substantial reduction in mortality in these patients. • a respiratory rate ≥ 20 breaths/min or a partial pressure of arterial carbon dioxide ≤ 32 mmhg, or the use of mechanical ventilation for an acute respiratory process. • a white cell count ≥ 12,000/mm 3 or ≤ 4000/mm 3 , or a differential count showing >10% immature neutrophils. patients should meet at least one of the following five criteria: • pregnancy or breastfeeding. • aged younger than 18 years or weight >135 kg. • platelet count < 30,000/mm 3 . • conditions that increase the risk of bleeding: • surgery requiring general or spinal anesthesia within 12 hours before the infusion, the potential need for such surgery during the infusion, or evidence of active bleeding postoperatively; • a history of severe head trauma requiring hospitalization, intracranial surgery, or stroke within 3 months before the study, or any history of intracerebral arteriovenous malformation, cerebral aneurysm, or mass lesions of the central nervous system; • a history of congenital bleeding diatheses; gastrointestinal bleeding within 6 weeks before the study unless corrective surgery had been performed; or • trauma considered to increase the risk of bleeding. • a known hypercoagualable condition including: • resistance to activated protein c; • hereditary deficiency of protein c, protein s, or antithrombin iii; • presence of anticardiolipin antibody, antiphospholipid antibody, lupus anticoagulant, or homocysteinemia; or • recently documented (within 3 months) or highly suspected deep-vein thrombosis or pulmonary embolism. • patient's family or physician, or both, not in favor of aggressive treatment of the patient, or the presence of an advanced directive to withhold life-sustaining treatment. • patient not expected to survive 28 days because of an uncorrectable medical condition, such as poorly controlled neoplasm or other end-stage disease. • moribund state in which death is perceived to be imminent. • human immunodeficiency virus infection in association with a last known cd4 cell count ≤ 50/mm 3 . • history of bone marrow, lung, liver, pancreas, or smallbowel transplantation. • chronic renal failure requiring hemodialysis or peritoneal dialysis (acute renal failure was not an exclusion criterion). • known or suspected portosystemic hypertension, chronic jaundice, cirrhosis, or chronic ascites. • acute pancreatitis with no established source of infection. • participation in an investigational study within 30 days before treatment. • use of any of the following medications or treatment regimens: • unfractionated heparin to treat an active thrombotic event within 8 hours before the infusion (prophylactic treatment with a dose of unfractionated heparin of up to 15,000 u/day was permitted); • low molecular weight heparin at a higher dose than recommended for prophylactic use (as specified in the package insert) within 12 hours before the infusion; • warfarin (if used within 7 days before study entry and if the prothrombin time exceeded the upper limit of the normal range for the institution); • acetylsalicylic acid at a dose of more than 650 mg/day within 3 days before the study; • thrombolytic therapy within 3 days before the study (thrombolytic agents permitted for the treatment of thromboses within a catheter); • glycoprotein iib/iiia antagonists within 7 days before study entry; • antithrombin iii at a dose of more than 10,000 u within 12 hours before the study; • protein c within 24 hours before the study. drotrecogin alfa (activated) should be given at a dose of 24 µg/kg/hour for 96 hours. infusion should be interrupted 1 hour prior to any percutaneous procedure or major surgery, and should be resumed 1 and 12 hours later, respectively, in the absence of bleeding complications. there was an 8-hour time window from shock onset to check for eligibility and to perform a short acth test (blood samples before and 30 and 60 min after a 250 µg intravenous bolus of tetracosactrin). patients were then randomly assigned to receive 50 mg hydrocortisone as an intravenous bolus every 6 hours and one 50 µg tablet of fludrocortisone through a nasogastric tube once a day, or their respective placebos. treatments were given for 7 days, and patients were followed up for 1 year. on admission, patients should receive continuous intravenous glucose (200-300 g over 24 hours). after 24 hours, total parenteral, combined parenteral and enteral, or total enteral feeding should be instituted: 20-30 nonprotein kcal/kg/day with a balanced composition (0.13-0.26 g nitrogen/kg/day and 20-40% nonprotein calories in the form of lipids). total enteral feeding should be attempted as early as possible. national vital statistics reports epidemiology of severe sepsis in the united states: analysis of incidence, outcome, and associated costs of care the task force on the management of acute myocardial infarction of the european society of cardiology epidemiology of sepsis: an update severe sepsis and septic shock. definitions, epidemiology, and clinical manifestations gram-negative sepsis: a dilemma of modern medicine sepsis definitions. lancet infect dis use of the sofa score to assess the incidence of organ dysfunction/failure in intensive care units: results of a multicenter, prospective study improved survival of patients with acute respiratory distress syndrome (ards): 1983-1993 [concepts in emergency and critical care treatment of gram-negative bacteremia and septic shock with ha-1a human monoclonal antibody against endotoxin. a randomized, double-blind, placebo-controlled trial treatment of gram-negative bacteremia and shock with human antiserum to a mutant escherichia coli the tnfα α mab sepsis study group: monoclonal antibody to human tumor necrosis factor alpha (tnfα α mab): efficacy and safety in patients with the sepsis syndrome lenercept study group: lenercept (p55-tumor necrosis factor receptor fusion protein, ro 45-2081, tenefuse) patients with severe sepsis or early septic shock. a randomized double-blind placebo-controlled multicenter phase iii trial with 1342 patients recombinant human protein c worldwide evaluation in severe sepsis (prowess) study group: efficacy and safety of recombinant human activated protein c for severe sepsis current concepts: treating patients with severe sepsis need for intensivists in intensive care units randomised trial of intravenous streptokinase, oral aspirin, both, or neither among 17,187 cases of suspected acute myocardial infarction: isis-2 (second international study of infarct survival) collaborative group injurious ventilatory strategies increase cytokines and c-fos m-rna expression in an isolated rat lung model effect of mechanical ventilation on inflammatory mediators in patients with acute respiratory distress syndrome: a randomized controlled trial effect of a protective-ventilation strategy on mortality in the acute respiratory distress syndrome ventilation with lower tidal volumes as compared with traditional tidal volumes for acute lung injury and the acute respiratory distress syndrome. the acute respiratory distress syndrome network airway and lung in sepsis elevation of systemic oxygen delivery in the treatment of critically ill patients a trial of goal-oriented hemodynamic therapy in critically ill patients. svo 2 collaborative group early goal-directed therapy collaborative group: early goal-directed therapy in the treatment of severe sepsis and septic shock task force of the american college of critical care medicine, society of critical care medicine: practice parameters for hemodynamic support of sepsis in adult patients in sepsis critical care in the emergency department: a physiologic assessment and outcome evaluation critical care provided in an urban emergency department a hemodynamic comparison of early and late phase severe sepsis and septic shock low levels of protein c are associated with poor outcomes in severe sepsis protein c, protein s, c4b-binding protein in severe infection and septic shock prognostic value of protein c concentrations in neutropenic patients at high risk of severe septic complications van der voort e: protein c and s deficiency in severe infectious purpura of children: a collaborative study of 40 cases. intensive care med epidemic meningioccemia and purpura fulminans with induced protein c deficiency protein c prevents the coagulopathic and lethal effects of escherichia coli infusion in the baboon the endothelial cell protein c receptor aids in host defense against escherichia coli sepsis corticosteroid treatment for sepsis: a critical appraisal and meta-analysis of the literature steroid controversy in sepsis and septic shock: a meta-analysis a 3-level prognostic classification in septic shock based on cortisol levels and cortisol response to corticotropin stress doses of hydrocortisone reverse hyperdynamic septic shock: a prospective, randomized, double-blinded, singlecenter study prolonged methylprednisolone treatment suppresses systemic inflammation in patients with unresolving acute respiratory distress syndrome: evidence for inadequate endogenous glucocorticoid secretion and inflammation-induced immune cell resistance to glucocorticoids patterns of corticosteroidbinding globulin and the free cortisol index during septic shock and multitrauma adrenal insufficiency during the late stage of polymicrobial sepsis reversal of late septic shock with supraphysiologic doses of hydrocortisone effect of a treatment with low doses of hydrocortisone and fludrocortisone on mortality in patients with septic shock reactivation of pituitary hormone release and metabolic improvement by infusion of growth hormone-releasing peptide and thyrotropin-releasing hormone in patients with protracted critical illness a paradoxical gender dissociation within the growth hormone/insulin-like growth factor i axis during protracted critical illness bouillon r: intensive insulin therapy in the critically ill patients decision support and safety of clinical environments. qual saf health care the roundtable discussion was supported by an unrestricted educational grant from eli lilly and company. jlv, ea, gb and er are consultants to eli lilly and company. all authors received an honorarium/grant for participating in this meeting. inclusion criteria • partial pressure of arterial oxygen/fio 2 ≤ 300 mmhg. • bilateral infiltrates consistent with pulmonary edema on frontal chest radiograph.• no clinical evidence of left atrial hypertension, pulmonary artery wedge pressure ≤18 mmhg if measured. • positive pressure ventilation via endotracheal tube. inclusion criteria patients must have a known infection or a suspected infection, as evidenced by one or more of the following:• white cells in a normally sterile body fluid.• perforated viscus.• radiographic evidence of pneumonia in association with the production of purulent sputum. • a syndrome associated with a high risk of infection (e.g. ascending cholangitis). patients should meet at least three of the following four criteria:• a core temperature ≥ 38°c (100.4°f) or ≤ 36°c (96.8°f).• a heart rate ≥ 90 beats/min, except in patients with a medical condition known to increase the heart rate or those receiving treatment that would prevent tachycardia.available online http://ccforum.com/content/6/s3/s1 the study included all mechanically ventilated patients entering the icu: predominantly surgical patients, with some neu-rological patients (the icu in which the trial took place also sees such patients). medical icu patients (e.g. those with chronic obstructive pulmonary disease or oncologic or hematological disorders) were not included as they are not treated in the unit where the study was conducted. however, septic patients that were initially surgical but then came back from the ward with sepsis were included.only those patients who were moribund or had do-notresuscitate status at icu admission were excluded from the trial. if blood glucose ≥110 mg/dl (≥ 6.1 mmol/l), infuse with insulin to maintain normoglycemia (80-110 mg/dl, 4.4-6.1 mmol/l). do not exceed 50 iu/hour. adjust insulin dose based on measurements of whole-blood glucose in undiluted arterial blood, performed at 1-4 hour intervals, based on the following algorithm: adjust the dose in proportion to the observed change in blood glucose level (if blood glucose decreases by 50% then the insulin dose should be decreased by 50% and checked within the next hour). appendix 5 table 1 provides information on the appropriate action depending on the blood glucose level. the numerical instructions provided in appendix 5 table 1 are a guide; insulin dosage should always be done with common sense, proportionate to the previous changes in blood glucose observed upon previous changes in dosage.available online http://ccforum.com/content/6/s3/s1appendix 5 table 1 appropriate action depending on blood glucose level key: cord-006460-3ayc0hne authors: baue, arthur e. title: multiple organ failure – the discrepancy between our scientific knowledge and understanding and the management of our patients date: 2000-10-19 journal: langenbecks arch surg doi: 10.1007/s004230000162 sha: doc_id: 6460 cord_uid: 3ayc0hne the excitement of molecular biology and of genetic knowledge and their possibilities must be balanced against our limitations in using this information for the care of our patients. there is a great discrepancy between what we know and what we can do. there are many reasons for this. a major one is that science must simplify/reduce the variables in experimentation and then generalize in terms of a specific factor or effect, whereas patients are complex with variables that we do not yet understand completely. this powerful science is now teaching us about the genetic diversity in both susceptibility and outcome of disease, and the diversity in life experiences and antigen exposures. clinicians have tried to lump together and treat in a similar way many diverse human diseases. this has not worked well. pancreatitis and perforated diverticulitis both produce inflammation and sepsis, but they are different processes and may both lead to multiple organ failure. this lumping together has contributed to the failure of so-called magic bullets. there are new contributors to organ damage. gender, lifestyle and prior disease differences also complicate the care of patients. despite this, we are slowly and gradually improving the care of our surgical patients by careful pre-, intraand postoperative support and better, simpler and safer operations. there is a tremendous difference between what we know in science and what we can do for our patients. the extraordinary explosion of knowledge about inflammation, mediators and critically ill patients has greatly increased our knowledge of these processes. therapy has improved, but at a slower and more gradual pace, primarily by better monitoring and support of organ function. a classic example of this phenomenon is demonstrated by the presentation of the 1999 nobel prize in physiology and medicine to robert furchgott, louis ignarro and ferid murad [1] . these scientists demonstrated that vascular endothelial cells form nitric oxide (no) through nitric oxide synthase (nos), which in turn stimulates cyclic guanosine monophosphate (cgmp) synthesis in the underlying vascular smooth muscle causing relaxation or vasodilatation. this exciting contribution opened up the entire world of no and its many biologic functions. it is an important mechanism whose activities are still being mapped out. this led to the study of excess vasodilatation in septic shock, which was thought to be due to the overproduction of no. many animal and clinical studies suggested that this phenomenon was occurring. it was then proposed that a nos inhibitor would help prevent this problem of excess vasodilatation and decreased blood flow in septic shock [2, 3] . many trials and experiments of nos inhibitors were performed in animals and patients. however, in a recent issue of critical care medicine, grover et al. [4] reported that the randomized, double-blind clinical trial of an nos inhibitor was discontinued due to increased mortality and adverse outcomes in patients who received the drug. there is now a moratorium against the clinical use of such agents until more is learned about them. here, then, is an example of a nobel prize being awarded for the study of endothelial a.e. baue ( ✉ ) cell-produced no on the one hand, and the concept of excess vasodilatation in septic shock, the hypothesis that this was due to no and a clinical trial blocking no that increased mortality on the other. some would say that a specific nos, such as inducible nos, would have more promise. bloom [5] made a comparison of the last millennium from st. thomas aquinas to newton and his principia. "st. thomas aquinas' view was that knowledge is of two types -that which man could know and that which was 'higher than man's knowledge' and not to be sought through reason." newton's principia, on the other hand, stated "that our universe and all within it are indeed knowable." i add to this that all things may be knowable, but perhaps not doable. the tremendous amount of information generated by molecular biology -the techniques of dna identification; the human genome project; mediators; signal transduction; the immune system; the response to injury; organ, cell, endothelium and membrane function -all stretch the imagination. there have been improvements in the way we care for patients, but they have been modest and slow and have not been easy to document. it has not been possible to apply most of the information of molecular biology to patient care. i believe that there are three major reasons for this discrepancy. you may have others. 1. there are differences between individuals (diversity) which may not be measurable and which are not taken into consideration in therapy and clinical trials. we do not look, act or respond alike. our genetic differences dictate how we will respond to injury, an operation, infection or stress. genetic studies are now beginning to describe these extensive genomic differences. sir william osler once said, "as no two faces, so no two cases are alike in all respects, and unfortunately it is not only the disease itself which is so varied but the subjects themselves have peculiarities which modify its action" [6] . 2. our experiences in life are all different -our exposure to antigens, prior illness and injury; our immune history; and our group experiences. we are not genetically pure animals living in the same cage for our lifetimes before an insult occurs. there is also much evidence for these powerful and complex acquired differences and exposures. 3. we have tried to lump together and treat human abnormalities according to symptoms and signs rather than to the basic causes of their diseases -we have tried to treat inflammation, sepsis, systemic inflammatory response syndrome (sirs), and multiple organ dysfunction syndrome (mods) rather than what caused them; perforated diverticulitis is not acute pancreatitis and neither are ventilator-associated pneumonia and appendicitis. all of these differences and factors complicate our scientific understanding and make developing appropriate therapy for patients based on molecular biology more complex. the wonders of science are revealing the complexities of human disease. the picture is becoming clearer -genetic polymorphisms, antigen exposure, etc. [7] . patient care is improving, but slowly. this should not be discouraging, but we must try harder. next, a word about mof, its meaning and its relationship to these complexities. in 1975, i wrote an editorial entitled "multiple, progressive or sequential systems failure: a syndrome of the 1970s" [8] . in it, i described three examples of patients that i observed who developed multiple organ problems. one patient had acute hemorrhagic pancreatitis. another had a colon resection and developed an anastomotic leak leading to peritonitis. a third was a patient of mine in whom i had performed a double valve replacement (aortic and mitral valves); the patient had an extremely low cardiac output for the next 6 weeks or so and died. i reviewed the clinical course of these patients and then the autopsy findings. in one patient, for example, the autopsy indicated bronchopneumonia, necrotizing bacterial arteritis, hyaline membranes in the lung, healed and acute renal infarcts and acute tubular necrosis in the kidneys, hemorrhagic foci in the gut, pseudomembranous enterocolitis in the colon, and massive acute central lobular necrosis of the liver. thus, in this patient there was pathologic evidence of severe multiple organ dysfunction and/or failure. this led to the development of the concept of mof. during a sabbatical in munich, germany, i worked with dr eugen faist. together we reviewed the experience at the klinikum grosshadern, ludwigs maximillians universität of the problem of mof after injury. this was reported by faist at a meeting of the american association for the surgery of trauma in colorado springs, usa [9] . in 433 consecutive patients with multiple injuries, 50 patients had single organ failure (sof) and 34 had mof. mortality for mof patients was 56%. there were 78 deaths overall (18%). faist described two patterns: a rapid single-phase pattern of development of mof due to trauma and shock; and a delayed second-phase mof due to trauma, shock and then sepsis. the temporal sequence of events of organ failure was the lung first, then the clotting system, the kidney and the liver, with sepsis ultimately being the cause of death in a number of patients. definitions of failure of each organ system were also given. for example, cardiovascular failure was defined as the presence of one or more of the following: (a) a heart rate of <54/min; (b) mean arterial pressure of <49 mmhg; (c) occurrence of ventricular tachycardia and/or ventricular fibrillation; (d) ph<7.24 with a paco 2 of <49 mmhg. respiratory failure was described as one or more of the following: (a) a respiratory rate of <5 or >49/min; (b) paco 2 >50 mmhg; (c) a a po 2 >350 mmhg, fio 2 >0.40, positive end-expiratory pressure (peep) >5; (d) dependence on a ventilator on the fourth day of organ system failure. in the same way, failure of all organs and systems -the kidney, liver, gastrointestinal tract, nervous system, etc. -were defined. these figures were also updated and reported by faist et al. [10] in the german literature. following this presentation, many joined in the battle. beale and bihari [11] stated that "the search for a unifying mechanism and, hence, perhaps an effective therapy for mof has been intense." i reviewed our experience with mof at st. louis university in almost 6000 patients who underwent cardiac operations over several years [12] . mof occurred in 128 patients (2.1%), primarily after emergency operations and in high-risk patients. however, 78% of these mof patients died. mortality in all patients was 4.7%. thus, most were elective cardiac surgical procedures and the risk was low. recently, a new proposal was developed by a consensus conference of physicians, intensivists and surgeons [13] . they defined the concept of sirs and mods. sirs is defined as the response to a variety of severe clinical insults with two or more of the following conditions: a temperature >38 or <36°c; a heart rate >90 beats/min; a respiratory rate of >20 breaths/min; or a paco 2 <32 torr (<4.3 kpa) or a white blood count of >12,000 or <4000 cells/mm 3 , or >10% immature or band forms. mods was described simply as the presence of altered organ function in an acutely ill patient such that homeostasis cannot be maintained without intervention. since that time, many have continued to use sirs and mods as a concept [14, 15] ; however, this has been criticized by many [16] , including myself. i stated then that "our ingenuity in developing terminology exceeds our abilities to take care of these patients" [17] . for example, jones and lowes [18] found that sirs is not a very good predictor of bacteremia, the positive predictive value being only 7%. sirs, mods and mof are constructs or concepts. they are not diseases or even syndromes. they cannot be treated other than to support organ function. mof is the final common pathway to death in the modern organsupporting intensive care unit (icu). blakiston's gould medical dictionary (3rd edition) defines a syndrome as a group of symptoms and signs which, when considered together, characterize a disease or lesion [19] . thus, none of these concepts are diseases and, therefore, are not treatable as such. in a recent article, i described the concept (shown in figure 1 ) that sirs to mods to mof is simply going from sick to sicker to very sick [20] . some have focused on sirs indicating an inflammatory process. as goris et al. [21] demonstrated that inflammation alone, as produced by zymosan in the peritoneal cavity of animals, can lead to mof. nor is inflammation a disease. it is a complex biologic process of host defense and repair of injury. many years ago, john hunter stated that, "inflammation in itself is not to be considered as a disease -and in disease, where it can alter the disease mode of action, it likewise leads to a cure; but where it cannot accomplish that salutary purpose, it does mischief" [22] . the scientific advances of molecular biology developed a number of substances and approaches that, in terms of animal experimentation, suggested possible help for patients. these were called "magic bullets" after the concept developed by ehrlich and morgenroth [23] at the turn of the twentieth century. there are a number of socalled magic bullets that have failed to improve overall mortality (table 1) . for details and references to these trials and reviews of them, i refer the reader to papers by opal [24] , eidelman and sprung [25] , the late roger bone [26] , and zeni et al. [27] . i have also reviewed the reasons why i think many of these trials have failed [28] . there have been more recent clinical trials using what some thought might be magic bullets and these have also failed. one such trial used diaspirin cross-linked hgb, which was found to increase mortality [29] . trials were halted. n-acetylcysteine, an antioxidant, was found to be of no help in patients and actually may be harmful despite early encouraging results in adult respiratory distress syndrome (ards) [30] . the use of leptin to control obesity has not been impressive in clinical trials [31] and the nos inhibitor 546c88 increased mortality [4] . ibuprofen did not decrease shock, ards or mortality in septic patients [32] . antithrombin iii treatment in severely injured patients did not attenuate the inflammatory response or improve outcome [33] . another study of a platelet-activating factor (paf) receptor (bb-882) antagonist in septic patients also offered no advantages [34] . why have there been failures of so many potential magic bullets? many reasons have been proposed by a number of authors [24, 25, 26, 27, 28] . the three major reasons that i described earlier for the discrepancy between science and patient care also apply here. in addition, specific problems affect trials of "magic bullets", including: 1. the causality of disease: many different complex diseases are being lumped together and treated with one of these magic bullets. sirs has been used as the entry criteria in some studies. it is no wonder that they have failed to improve 28-day mortality, which is the usual endpoint for success. i do not believe that there is a single treatment or agent which will help all or many different complex disease processes in patients. 2. redundancy and overlap of mediators: there is considerable overlap of the activities of cytokines and other mediators. 3. there is no single factor that acts as the lethal activator. 4. the timing of treatment is extremely important. in experimental studies in animals, pretreatment before the insult will help. obviously, this cannot be performed for injured, sick or septic patients. 5. there is the problem of modulation of an essential biologic function -inflammation. can we control inflammation without doing harm to the host? that is a real question that has not been resolved. 6. is there immune deficiency or immune excess? perhaps both occur. t-cell immune deficiency occurs after injury, whereas immune excess of the non-specific immune system such as neutrophil activation may be harmful. thus, both may be at work. how do we sort out which is most important and which should be treated or blocked? another example is the treatment of patients with sepsis syndrome with a soluble tnf receptor (stnfr) [35] . this was a phase ii controlled trial of a novel p75 soluble receptor fusion (fused with igfc fragment) protein. at higher doses there was increased mortality, and in lower doses there was no difference from the controls. some have proposed that physiologic improvement in a critically ill patient should be used as an endpoint for a clinical trial rather than 28-day mortality. the problem with this is, what if there is some physiologic improvement such as ventilation function but the patient dies anyway. what difference does it make? goldfarb [36] described investigations into the polydeterminate nature of sepsis. he stated that, "a fundamental concept of modern pharmacology is: for each pathologic condition (disease) a single pharmaceutical can be found to correct 444 i believe we must go back to the suggestions of stehbens [37, 38] . he stated that, "treatment of a disease, when based on symptoms or clinical manifestations is at best palliative and nonspecific." all of us use or give therapy based on symptoms or clinical manifestations. when we get a cold or the flu, we may take aspirin or a non-steroidal anti-inflammatory agent. this is for palliation, to help us feel better, but it will not alter the basic disease process. only specific treatment of a disease will improve the mortality and alter the disease process. how do you treat an injury severity score (iss) of 26, an apache iii score of 30, sirs, mof, the sepsis syndrome, or a consensus conference definition of a human state. it is impossible to treat such abnormalities. fortunately, some of the gurus of critical care agree that it is time for a reevaluation of these definitions [39] . solbach et al. [40] wrote that "lymphocytes play the music, but the macrophages call the tune." they stated that, "dealing with a single cytokine on an isolated population of cells (or an organism) may be a gross oversimplification of events in vivo." i believe that they have hit the nail on the head. i have now reviewed the third reason for the discrepancy between excellent science and patient care. i will now go back to reasons 1 and 2. first, the differences between individuals. these variations include: 1. genetic differences in cytokine response to sepsis, including dna polymorphism of tumor necrosis factor (tnf), interleukin (il) -1 and il-1ra. some seem to protect, while others are associated with a poor prognosis. some of these have been described by zehnbauer et al. [41] . an il-1ra polymorphism may increase susceptibility to sepsis [42] . there is the possibility of another il-1 predisposition to sepsis that may also vary in populations in the uk from those in australia [43] . a tnf2 allele is strongly associated with death from septic shock [44] and another tnfb gene is associated with sepsis in trauma patients [45] . 2. there are differences in preoperative, pre-illness immunity and prior immune history. an example of this was described by bennett-guererro et al. [46] , who measured the endotoxin core antibody endocab. a low preoperative level of endocab in patients undergoing cardiac operations predicted an adverse outcome, whereas high levels suggesting previous expo-sure to either gram-negative bacteria or endotoxin predicted an improved outcome. this finding has been confirmed by hamilton-davies et al. [47] . 3. there are male/female gender differences in both animals and patients as described by chaudry et al. [48] and others. 4. the age of the patient makes a big difference. 5. prior illness, such as chronic obstructive pulmonary disease (copd), coronary artery disease, etc., make a difference. 6. lifestyle, obesity, poor nutrition, lack of exercise, etc. are all factors! there are other variations in patients, including cytokine pleiotropy and redundancy, as described by sánchez-cuenca et al. [49] . examples of this are that the blocking of tnf-does not alter other products [50] , the polymorphisms of interferon-microsatellites [51] , discordant tnf-super family gene expression [52] , il-1 receptor antagonist polymorphism [42] , leukocyte receptors for tnf differences [53] , different polymorphic heat shock proteins (hsps) and variations [54] , and variations in different populations, such as the difference in uk and australian population responses to various substances [43] . aasen et al. [55] studied the immunologic and inflammatory consequences of surgical trauma and found extensive immunologic and inflammatory responses taking place in the closed surgical wound that would not be recognized in blood samples. other complexities include pleiotropy and redundancy in cytokines, genetic variations in the response to lipopolysaccharide (lps) [56] , polymorphic hsps, and differential receptor subtypes as cited above. also present are discordant super family gene expression, a gene promoter polymorphism and receptor antagonist polymorphism and, of course, cytokine microsatellites [57] . an example was described by sh et al. [7] . the association of tnf microsatellites with outcome in sepsis indicated that tnf-6 and tnf-1 levels increase in septic patients; however, tnf-10 increased in non-survivors [44, 45] . there is a tremendous disparity between scientific knowledge about endotoxin and the clinical usefulness of that knowledge. there is nothing at the present time that seems clinically useful about the knowledge on endotoxin. we know that it is present in the walls of gramnegative organisms. we know that it does all sorts of bad things when it is injected into animals and people, but we still have no information about what it does in various sick patients and what we might do about it. new developments may help [58] . o'reilly et al. [59] reported on endotoxin sepsis and the primrose path. they reviewed how the use of endo-toxin models has failed to identify a clinically useful therapeutic agent for the treatment of sepsis. archibald philip primrose was the fifth earl of roseberg. he promoted the concept that the aristocracy and monarchy are no longer necessary. this was labeled the "primrose path, a tempting but hazardous course." endotoxin models of sepsis may be the primrose path of research. there are a number of new and confusing concepts relating to organ damage that have been heretofore unknown. one of these is that arteriosclerosis may be the result of infection, a possibility suggested by the finding of organisms in arteriosclerotic plaques. hatch [60] reported finding the organism chlamydia pneumoniae in 70% of arteriosclerotic plaques. herpes simplex virus (hsv) has been implicated, as has cytomegalovirus (cmv) [61, 62] . others have suggested that helicobacter pylori may be involved [63] . dental infection and remote infection elsewhere in the extremities have been implicated in rapid occlusion of coronary arteries [64] . are these the causes or are they associations that are not totally worked out? the circumstantial evidence is very interesting. there are also tremendous genetic differences in disease susceptibility and these are now being studied vigorously. chaudry et al. [48] found in animals with trauma that infection was more prevalent in males and that this was related to testosterone. they performed a number of studies supporting this relationship [65] . the male swagger and ornamentation in animals suggests a decreased humoral immune response as the basis for this [66] . eachempati et al. [67] , however, described increased mortality from sepsis in female patients, as did crabtree et al. [68] . however, offner et al. [69] found increased infections in males after trauma. in autoimmune diseases, estrogen and prolactin seem to be the problem [70] . in comparison with males, multiple sclerosis occurs two times more commonly in females, rheumatoid arthritis occurs three times more frequently, and lupus erythematosus occurs nine times more often. what is the answer to this gender gap? it seems that if you are injured, you are better off being female; however, if you develop an infection, then you are better off being male. what should you do if you are injured and develop an infection? another new and confusing concept of organ damage is recent evidence that the mesenteric lymph of animals in shock is toxic and, when it returns to the central circulation, it activates neutrophils which produce cell dam-age [71] . in a clinical study, thoracic lymph was diverted and sampled in patients with mof and no toxicity was found [72] . others found that mesenteric lymph is a problem and this must be explored further in order to ascertain what its consequences are in man [73, 74] . in experimental animals, there seems little doubt about this phenomenon [75] . postimplantation inflammation has been described by zimmer et al. [76] . the implantation of a device in a patient produces an inflammatory reaction that can be quite severe. there is also a question as to whether we should replace rather than block various mediators. for example, increasing the level of protein c could be helpful. there is a question as to whether transgenic knockout animals, which provide fascinating information, will explain the redundancies, the overlap, and the interactions between cytokines and various other mediators. they may not be able to indicate the problems of cell-cell interaction such as the interactions between neutrophils and platelets [77] . polymerase chain reaction (pcr) will contribute greatly to our understanding of human disease [78] . pcr detects microbial dna in blood and is much more sensitive than blood cultures. a consensus pcr will provide information about shared conserved genetic sequences. there is also representational difference analysis (rda) and pathogen detection chips. dna microarray techniques and gene expression immune cells will provide similar responses. it is possible, in the future, that everyone with sepsis or sirs may be found to have an infection of some sort, a virus which cannot be cultured, bacteria which cannot be cultured, or some other microbial factor [79, 80, 81] . other new concepts of organ damage include superantigen-mediated lethal shock [82] . the bacteria h. pylori produces gastric ulcer and duodenal ulcer disease, and perhaps gastric cancer. does h. pylori modulate the response to injury and is it a nosocomial infection [83, 84] ? it is known that neutrophils in the inflammatory response are both a culprit and a solution with delayed apoptosis, cytotoxicity, activation of neutrophils, and the production and release of elastase and superoxide anions, which damage cells [87, 88] . the effect of granulocyte colony-stimulating factor (g-csf) is important. for example, the use of neupogen, a g-csf compound, is encouraging [89] , whereas root [90] reported that rg-csf (filgrastin) did not alter morbidity or mortality in the treatment of pneumonia, sepsis and septic shock [89] . there is considerable information about these factors, but what to do about them is far from being established. these are presently imponderables. the timing of treatments is critical. for example, knox et al. [91] found that recombinant human growth hormone (rhgh) can be helpful in burns when given at an appropriate time; however, in other studies, it was found to increase mortality. the relationships of infec-tion and cancer are becoming increasingly apparent. hepatitis b, h. pylori, the lyme disease spirochete and other organisms are associated with increased incidence of certain malignancies. patients with short-bowel syndrome have benefited from rhgh [92] , as have patients with burns [93] . a multicenter phase iii trial in europe administered rhgh to patients after operations, trauma or with ards and after 5 days of intensive care. the company allegedly called off the trial because of a twofold increase in mortality in the treated group. this has not been reported yet (d. wilmore, pers. comm.). there are other differences that must be considered. for example, the neutrophil response to trauma depends on the bacterial species. in addition, septic serum signaling is diverse and complex [94] . genes are expressed in the serum of septic patients which were not previously associated with sepsis or known to have any relationship with sepsis. other differences include: ab t-cell receptor diversity differences in species sensitivity of hormones; variability in the g-csf response; and lps stress variability in various species. an additional problem is that of autoimmunity, which may be related to molecular mimicry. albert et al. [95] recently described the relationship that may pertain. for example, if the host acquires an infection with an agent that has an antigen similar to host antigens, tolerance to the autoantigens may break down. the pathogen response then cross-reacts to cause tissue damage and an autoimmune response. are we improving patient care and, if so, how? prevention of mof is the goal. we know that we cannot treat specifically but can only support mof, mods and sirs; therefore, we must prevent them by means of better patient care. are we improving patient care, are we winning the battle [96, 97] , and is mof disappearing? in levine et al. [98] we reviewed this recently, raising the question: mof -is it disappearing? our answer to this question was no: mof is not disappearing and, as we help more severely injured and sick patients to survive, mof may actually increase and the mortality will remain high. i will now review the evidence for this. zimmerman et al. [99] compared the risks and outcomes for patients with organ system failure from 1982 to 1990. they found that "the incidence and overall outcome of organ system failure have not changed significantly over the past eight years. there has been significant improvement in survival of patients with persistent severe organ system failure." thus, this report confirms again (1) that better intensive care is helping, and (2) my insistence that the secret to mof is prevention. regel et al. [101] reported on the results of patients with multiple trauma (3406 patients) treated between 1972 and 1991 at a german level one trauma center. they found increased head and thoracic injuries in the second decade. in the second decade, there was a decrease in renal failure and ards, but an increase in mof from 15.4 to 28.2%. lethal mof increased from 13.8 to 18.6%; however, overall mortality decreased from 37 to 22%. regel et al. believed that more severely injured patients stayed alive longer to develop mof and then die. christou et al. [101] reported on the delayed hypersensitivity response and host resistance in surgical patients, and on the results of surgical care over a 20-year period. they found that overall surgical mortality decreased from 11.4% in the 1970s to 10.2% in the 1980s and to 2.4% in the 1990s. they believe that this was due to improved pre-, post-and intra-operative care. however, there was no decrease in surgical icu deaths. the icu remains a problem. o'keefe et al. [102] reported 10-year trends in costs, resource utilization and survival in an established trauma center. they found no change in mean age or iss over the 10-year period. crude mortality did not change. it stayed at 8%. length of stay decreased from 9.5 to 6.8 days, while costs increased by 16.7%. adjusting for iss and accidental injury score, the mortality decreased by 3%/year in patients with an iss >16. thus, to answer the question -is mof disappearing? -, in the last decade the incidence varied from 2 to 25%, depending on the clinical setting, with a mortality rate of 40-80%. regel et al. [100] found that the mortality from trauma decreased, but the mortality from mof increased. christou et al. [101] found that overall surgical mortality decreased, but icu mortality did not change. i found that, in cardiac surgery, the overall incidence of mof was unchanged, but more re-operations were performed recently where the incidence of mof was higher. o'keefe and maier [103] concluded that there are documented improvements in the treatment of septic shock, trauma, ards, and that these are "due to overall and cumulative improvements in care in the icu rather than to any isolated specific intervention." they predicted, "as survival rates improve, the ability to test specific interventions and document relevant improvements will become more difficult." the promissory note in scientific research i have described the results of wonderful studies of science and injury and how some investigators concluded, by predicting usefulness in patient care, beyond what their studies allow [104] . this phenomenon continues and is unfortunate. extending the results beyond what the study actually shows is a form of "hype". for example, an author may state that these biologic effects suggest a promising candidate for the treatment of sepsis in humans from a study based on 30 rats. recent examples of this include a paper by forceville et al. [105] , from paris, where it was stated, "in severely ill icu patients with sirs, we observed an early 40% decrease in plasma selenium." they then went on to state "… this could explain the threefold increase in morbidity and mortality rates in these patients …" this is ridiculous of course. wong [106] stated that, "in molecular terms, mods may occur when endothelial cells undergo disparate patterns of gene expression either simultaneously or consecutively." that does not help me understand mods. in 1960, i sat at a dinner of the american heart association with the great pioneer wilhelm kolff. kolff was the developer of the artificial kidney, an immense contribution from a man who dedicated his life to this concept and the science behind it. at this dinner, kolff talked about how science and technology would advance. he stated that, "advances in science and technology will easily be able to develop a satisfactory long-term implantable artificial heart." he said that it would be just like going to the moon, a simple scientific problem. i asked him whether he thought there would be any biologic problems. he said no, those were surmountable. it is now the year 2000, 40 years later, and there is still no satisfactory, long-term implantable artificial heart, nor is one on the immediate horizon. have there been any clinical advances in the care of patients in the past 20 years? there have been a number of advances, but they are difficult to document on a patient-by-patient basis. they can only be documented by the types of experience that were cited earlier in the section entitled "are we improving patient care and, if so, how?: "is multiple organ failure disappearing?"are we winning the battle? one such advance is better resuscitation and initial care. there certainly have been improvements in monitoring. early definitive operations for fracture immobilization allow patients to move about and decreases the need for ventilatory support and the development of ards [107] . better circulatory, pulmonary and gut support are provided by various agents, ventilator strategies and enteral and immune nutritional support [96, 97] . immune-enhancing diets seem to make a difference [108] . hemodiafiltration has been helpful in certain circumstances [109, 110] . in addition, immunomodulation has made a contribution. some of the new therapeutic agents, such as glucan, ketoconazole, and antithrombin iii, remain controversial, depending on how they are used. other advances include some approximation of microcirculatory blood flow by measuring intercellular ph in the stomach [111] , by minimal surgical procedures, the widespread use of epidural block and other adjuncts. there remains the contrast between molecular biology and minimal surgery and the disparity between them [112] . minimal surgery has certainly contributed to better patient care. there are other procedures on the horizon such as the measurement of hemodynamic patterns during high-risk elective operations, as performed by shoemaker et al. [113] . they found that lethal circulatory dysfunction may occur during the operation. during an operation, non-survivors were found to have a decreasing cardiac index and stroke index and stroke work, a decrease in oxygen delivery, and a decrease in oxygen consumption [114] . identifying these factors during surgery may allow immediate corrective measures. we found, for example, that using oxygen consumption and delivery as endpoints for resuscitation in critically ill patients is not particularly helpful. oxygen parameters are more useful as predictors of outcome than as endpoints for resuscitation [115] . we have known for many years that surgical and injured patients must increase oxygen transport, cardiac index and oxygen consumption to survive. goodwin [116] stated that, "the great gift of chaos theory to the practice of medicine has been the simple but profound negative statement: 'traditional science cannot predict complex systems'." people (patients) are such complex systems. another reason for the discrepancy between our science and patient care is that much of science today is based on traditional concepts. the "milieu interieur" of bernard [117] is important, as is the contribution to homeostasis by cannon [118] . now there is a new science of non-linear integration, which, according to schultz [119], may be another emerging scientific revolution. he stated that, "molecular biology provides parts of a puzzle which are dynamic or plastic. putting them together is the problem." it is our job to put them together by learning the rules of association or integration. schultz described non-linearity as "the shape of the 'whole' cannot be predicted by knowing only the shapes of the separated parts." buchman stated that "if nested, nonlinear models are better representatives of human physiology, than cannon's collection of negative feedback servomechanisms, then therapy should be redirected toward transitions to a basal range -not therapeutically manipulating things such as cytokines or nitric oxide" [120] . focusing on the phases of inflammation (pro-anti, etc.) would be cannon's approach. blocking one mediator may change not only the effects of that mediator, but may also disturb other mediators and the entire system. sir karl popper, in his medawar lecture in 1986 to the royal society in london, stated that "biology cannot be reduced to physics because biochemistry cannot be reduced to chemistry. reductionism is not possible in biologic systems. an organism cannot be reduced to a series of systems" [121] . as physicians, we were taught that stability is healthy. now we are learning that regularity and stability are more characteristic of the beginning of disease. for example, injection of endotoxin into normal human volunteers increased cardiac regularity [122] . godin and buchman stated that "a nonlinear system is one whose behavior is not a simple sum or multiples of the inputs to the system. chaos is irregular behavior of some deterministic, nonlinear systems" [123] . they proposed a theory of uncoupling of biologic oscillators to progress from sirs to mods. biologic systems are always coupling and uncoupling (multiple stable states). disease is a failure to recouple. thus, the physiologic states described by siegel et al. [124] may be a better way to describe and treat human problems. in the meantime, we have much to learn about how non-linear integrative biology will help us to solve clinical problems. the discrepancy between what we are learning in molecular biology and what we can do for our patients is great. the science is exciting, very worthwhile, and should be encouraged. eventually it will make contributions to health and patient care. in the meantime, we must focus on better and safer operations, better treatment of injury, and clinical research to improve care and decrease morbidity and mortality. we must avoid exaggerating the results and significance of animal research because it may not apply to man. all of us remember exciting experiments in the laboratory that have never helped patients. our research may eliminate the need for an operation. professor edward d. churchill [125] at harvard and the massachusetts general hospital in boston once stated that, "surgery began with trauma and congenital anomalies and therein shall it end." even congenital anomalies may be prevented or treated in utero. angioplasty with stents may eventually take the place of coronary bypass grafting. many years ago, antibiotics eliminated the need for mastoidectomies, but otolaryngologists went on to other things. we must be cautious about so-called "magic bullets" and remember the differences in individuals, both in terms of genetics and in life experiences. we must treat disease, not concepts of inflammation or of being sick. we must be careful about "academically-correct biology," as described by vogel [126] : "(1) it seeks molecular explanations; (2) it views scientific progress as incremental accretion of detail; (3) the immediate goal is human therapy; (4) its operation is unabashedly entrepreneurial." latour [127] stated that, "science does not enter a chaotic society to put order into it anymore but to add new, uncertain ingredients -to the collective process." claude lenfant [128] , director of the national heart, lung and blood institute of the national institutes of health (nhl&b), stated that, "the real challenge of the new millennium may indeed be to strike an appropriate balance between the pursuit of exciting new knowledge and the full application of strategies that are already known to be extremely effective but considerably underused." finally, i have paraphrased a section of the old testament, ecclesiastes 3:1 [129] : there is a time for everything and a season for every activity under heaven. "a time to be born and a time to die, a time to plant and a time to pluck up, a time to weep and a time to laugh, a time to mourn and a time to dance, a time of war and a time of peace," a time to give cytokines and a time to block cytokines, a time to supplement and a time to modulate, and a long time to figure out the time for any of these. then, if successful, the lord will invite you to stockholm and there will be a time for honor. on 1 and 2 march 2000, the fifth world congress on trauma, shock, inflammation and sepsis took place at the klinikum grosshadern, the ludwig maximillians universität, munich, hosted by professor eugen faist and professor wilhelm schildberg. thousands attended from more than 60 countries all over the world. i suggest to you that a major theme of this congress, although not stated, was the theme of this presentation -the discrepancy between understanding and management. at this meeting, basic scientists presented the results of their exciting research into molecular biology on the leading edge of scientific knowledge. clinical investigators presented their equally exciting findings about sick, injured and septic patients. some of the experiences presented will help us to take better care of our patients. all of these studies are very worthwhile; however, there is a widening gap between our knowledge of human biology, our understanding of it, and our ability to help patients. why is this? 1 449 nitric oxide leads to prized nobility prolonged inhibition of nitric oxide synthesis in severe septic shock: a clinical study effect of nitric oxide synthase inhibitors on hypotension in patients with septic shock multicenter, randomized placebo-controlled, double-blind study of the nitric oxide synthase inhibitor 546c88 the endless pathways of discovery the principles and practice of medicine association of tumor necrosis factor microsatellites with incidence and outcome of severe sepsis multiple, progressive or sequential systems failure: a syndrome of the 1970s multiple organ failure in polytrauma patients multiple organ failure: the pilgrim's progress the role of the gut in the development of multiple organ dysfunction in cardiothoracic patients definitions for sepsis and organ failure and guidelines for the use of innovative therapies in sepsis mods/sirs: result of an overwhelming inflammatory response to sirs with love dear sirs, i'm sorry to say i don't like you what's in a name? an acronym or a response? the systemic inflammatory response syndrome as a predictor of bacteraemia and outcome from sepsis blakiston's gould medical dictionary (1972) blakiston's gould medical dictionary sirs, mods, mofwhat's in a name? multiple organ failure: generalized and destructive inflammation treatise on the blood inflammation and gunshot wounds ueber hämolysins. fünfte mittei lung clinical trials for severe sepsis: past failures and future hopes why have new effective therapies for sepsis not been developed? why sepsis trials fail anti-inflammatory therapies to treat sepsis and septic shock: a reassessment mof, mods and sirs -why no magic bullets? blood substitutes n-acetylcysteine enhances recovery from acute lung injury in man: a randomized, double-blind, placebo-controlled clinical study tracing leptins partners in regulating body weight swindell bb (1997) the effects of ibuprofen on the physiology and survival of patients with sepsis high-dose antithrombin iii treatment of severely injured patients: results of a prospective study phase ii multicenter clinical study of the platelet-activating factor receptor antagonist bb-882 in the treatment of sepsis treatment of patients with sepsis syndrome with soluble tnf receptor (stnfr) investigations into the polydeterminant nature of sepsis causality in medical science with particular reference to heart disease and atherosclerosis. perspect the concept of cause in disease consensus conference definitions for sepsis, septic shock, acute lung injury, and acute respiratory distress syndrome: time for a reevaluation lymphocytes play the music but the macrophages call the tune genetic predisposition to sepsis and organ failure comparison of two polymorphisms of the interleukin-1 gene family: interleukin-1 receptor antagonist polymorphism contributes to susceptibility to severe sepsis is there an interleukin-1 genetic predisposition to developing severe sepsis? association of tnf2, a tnf-α promoter polymorphism, with septic shock susceptibility and mortality, a multicenter study relation of a tnf gene polymorphism to severe sepsis in trauma patients relationship of preoperative antiendotoxin core antibodies and adverse outcomes following cardiac surgery relationship between preoperative endotoxin immune status, gut perfusion, and outcome from cardiac valve replacement surgery hemorrhage and resuscitation; immunological aspects cytokine pleiotropy and redundancy -gp130 cytokines in human implantation attenuating tumor necrosis factor α does not ameliorate other cytokine and peroxidase products during sepsis microsatellite marker of interferon-γ receptor 1 gene correlates with infection following major trauma discordant tumor necrosis factor-α superfamily gene expression in bacterial peritonitis and endotoxemic shock differential expression of tumor necrosis factor receptor subtypes on leukocytes in systemic inflammatory response syndrome analysis of two human leukocyte antigen-linked polymorphic heat shock protein 70 genes in patients with severe sepsis immunological and inflammatory consequences of surgical trauma genetic component in the inflammatory response induced by bacterial lipopolysaccharide genetic factors in septic shock new developments in pathophysiology of endotoxins and opportunities for treatment endotoxin, sepsis, and the primrose path chlamydia: old ideas crushed, new mysteries bared inflammation, infection, and coronary artery disease does chlamydia pneumoniae cause atherosclerosis? helicobacter pylori and critical illness: a passive bystander or cause of disease? antibiotics and risk of subsequent first-time acute myocardial infarction gender dimorphism in trauma-hemorrhage-induced thymocyte apoptosis the male's dilemma gender-based differences in outcome with patients with sepsis genderdependent differences in outcome after the treatment of infection in hospitalized patients male gender is a risk factor for major infections after surgery multiple sclerosis and autoimmunity (1999) a gender gap in autoimmunity post-hemorrhagic shock mesenteric lymph is cytotoxic to endothelial cells and activates neutrophils thoracic duct in patients with multiple organ failure: no major route of bacterial translocation mesenteric lymph: the critical bridge between dysfunctional gut and multiple organ failure translocating gut bacteria reach the pulmonary artery by means of the thoracic duct during hemorrhagic shock gut-derived mesenteric lymph: a link between burn and lung injury inflammatorisches syndrom nach endovaskulären aortenstentimplatationen -eine vergleichende studie neutrophil-platelet interactions in sepsis the impact of the polymerase chain reaction in clinical medicine polymerase chain reaction surveillance of fungal dna in critically ill patients with systemic inflammatory response syndrome detection of bacterial dna in the plasma of patients with sepsis, severe sepsis and multiple organ dysfunction syndrome septic shock without documented infection: an uncommon entity with a high mortality superantigen-mediated lethal shock: the functional state of ligand-reactive t cells helicobacter pylori virulence and genetic geography helicobacter pylori infection in intensive care: increased prevalence and a new nosocomial infection circulating postinjury neutrophils are primed for the release of proinflammatory cytokines early neutrophil sequestration after injury: a pathogenic mechanism for multiple organ failure sequential systemic platelet-activating factor and interleukin 8 primes neutrophils in patients with trauma at risk of multiple organ failure does programmed cell death (apoptosis) play a role in the development of multiple organ dysfunction in critically ill patients? a review and a theoretical framework gcsf treatment of normal volunteers induces a sustained immunomodulation characterized by an anti-inflammatory cytokine response granulocyte colonystimulating factor (gcsf) in the treatment of severe pneumonia, sepsis and septic shock increased survival after major thermal injury: the effect of growth hormone therapy in adults a new treatment for patients with shortbowel syndrome: growth hormone, glutamine, and a modified diet characterization of growth hormone enhanced donor site healing in patients with large cutaneous burns priming of the neutrophil respiratory burst is species-dependent and involves map kinase activation molecular mimicry and autoimmunity systemic inflammatory response syndrome (sirs), multiple organ dysfunction syndrome (mods), multiple organ failure (mof): are we winning the battle? multiple organ failure, multiple organ dysfunction syndrome, and the systemic inflammatory response syndrome -where do we stand multiple organ failure: is it disappearing? a comparison of risks and outcomes of patients with organ system failure: 1982-1990 treatment results of patients with multiple trauma: an analysis of 3,406 cases treated between 1972 and 1991 in a german level one trauma center the delayed hypersensitivity response and host resistance in surgical patients -20 years later the complications of trauma and their associated costs in a level i trauma center are we winning the battle in the surgical intensive care unit? the promissory note in scientific research selenium systemic immune response syndrome, sepsis, and outcome in critically ill patients nuclear factorskappa β and nitric oxide regulating life and death: nonsense or harsh reality early osteosynthesis and prophylactic ventilation in the multi-trauma patient an immune-enhancing enteral diet reduces mortality rate and episodes of bacteremia in septic intensive care unit patients clinical benefits of cytokine removal with continuous hemodiafiltration (chdf) in critical care plasmafiltration in sepsis study group (1999) continuous plasmafiltration in sepsis syndrome persistent occult hypoperfusion is associated with a significant increase in infection rate and mortality in major trauma patients molecular biology and minimal surgery hemodynamic patterns of survivors and nonsurvivors during high risk elective surgical operations intraoperative evaluation of tissue perfusion in high-risk patients by invasive and noninvasive hemodynamic monitoring the use of oxygen consumption and delivery as end points for resuscitation in critically ill patients chaos and the limits of modern medicine an introduction to the study of experimental medicine (translated by hc green) the wisdom of the body. norton, new york 119. schultz sg (1996) homeostasis, humpty dumpty and integrative biology news physiologic stability and physiologic state reductionism and biologic systems experimental human endotoxemia increases cardiac regularity: results from a prospective, randomized crossover trial uncoupling of biological oscillators: a complementary hypothesis concerning the pathogenesis of multiple organ dysfunction syndrome physiologic classification and stratification of illness severity of post-trauma "sepsis" patients as a basis for randomization of clinical trials surgeon to soldiers academically correct biology from the world of science to the world of research medicine and the new millennium ecclesiastes 3:1. the holy bible of king james version 1 an address at an honorary degree presentation to dr. baue by ludwig maximillians universität, munich, germany, 28 february 2000. key: cord-005872-w1x1i0im authors: volk, t.; kox, w.j. title: endothelium function in sepsis date: 2000 journal: inflamm res doi: 10.1007/s000110050579 sha: doc_id: 5872 cord_uid: w1x1i0im endothelial cells can be the prime target for an infection and infected endothelial cells may serve as an initiating system for a systemic response as these cells are able to secrete many mediators known to be of paramount importance. endothelial cell functions in turn are regulated by these circulating mediators. cellular interactions with leukocytes revealed protective and destructive functions. single cell and animal studies indicate that endothelial permeability is increased and apart from clinical obvious edema formation in septic patients, the endothelial component remains unknown. endothelial coagulation activation has been shown in vitro, however human data supporting an endothelial procoagulatory state are lacking. defects in endothelium dependent vasoregulation in animal models are well known and again human studies are largely missing.¶an imbalanced production of reactive oxygen species including nitric oxide has been found to be involved in all endothelial functions and may provide a common link which at present can be supported only in animal studies. sepsis is considered the leading cause of death in noncoronary intensive care units. it has been defined as a systemic inflammatory reaction to an infection. among many cellular disturbances endothelial cells play a major role in the pathogenesis of this disease as these cells are critically involved in maintaining a delicate balance between vasoconstriction and vasodilation, blood cell adherence and nonadherence, anticoagulation and procoagulation, permeability and thightness. all these functions are believed to be imbalanced and impairment is believed to precede clinically recognizable alterations (e.g. bleeding, edema, organ dysfunction and shock) but it is by no means clear whether these changes are the cause or consequence of endothelial dysfunction in sepsis. endothelial functions have largely been studied in vitro from the first successful attempts of culturing isolated human endothelial cells dating back to the early 70ties. endothelial cells from and within different organs or different species behave differently and tend to change properties the longer they are kept in culture. to overcome isolation variabilities several endothelial cell lines are currently available which have retained at least some features. however, experiments from single cell cultures are flawed in many ways and results from these may never be of any relevance to medical practice. it is inherent to any cell culturing that data obtained from these experiments often are restricted to cell type, time of culture and general working conditions. in vitro stressed endothelial cells almost uniquely tend to activate a functional program leading to a proinflammatory, procoagulatory and hyperpermeable phenotype. in this review we want to summarize investigations of disturbed endothelial functions including infection, mediator and cellular interactions, permeability, coagulation and vasoactivc properties from molecular findings to patient care. staphylococcus aureus is the most prevalent bacterial pathogen isolated from patients with blood stream infection in north america [1] . s. aureus has been reported to directly infect human umbilical vein endothelial cells (huvec) thereby inducing secretion of cytokines and functional upregulation of adhesion molecules [2] . internalized s. aureus may lead to apoptosis in huvec [3] or persistence as small colony variants [4] . group a streptococci can enter huvec [5] , a process which may render these cells particularly sen-sitive to otherwise subtoxic concentrations of hydrogen peroxide [6] . group b streptococci (gbs) are the most common cause of neonatal sepsis and pneumonia. gbs-induced endothelial cell injury can be confirmed by histological findings at autopsy, in animal studies and in vitro [7] . gbs invasion and subsequent damage of endothelial cells may be inhibited by cytochalasin d in huvec implicating that cytoskeletal interactions are important for toxicity. however, invasion of brain microvascular endothelial cells by gbs may be dose dependently cytotoxic due to beta-hemolysin production [8] . streptococcus pneumoniae may enter activated endothelial cells using paf-receptors [9] . this receptor engagement may also serve as a sorting signal for endothelial transcytosis [10] . infection and activation of endothelial cells by listeria monocytogenes is believed to be a critical component of the pathogenesis of this disease and includes ceramide generation, transcription factor activation and increases in adhesion molecule expression on huvec [11] . listeria have been described to enter huvec either directly via internalin b or by a cell-to-cell spread from infected monocytes [12] . gram negative bacteria have lipopolysaccarides (lps) within their cell wall. cellular binding of lps usually is accomplished by cd14. endothelial cells lack cd14 receptors and lps effects on endothelial cells generally require the presence of cd14 in the serum. lps effects from gram negative live bacteria (b. fragilis, e. cloacae, h. influenzae, k pneumoniae) on endothelial cells have been demonstrated by transcription factor activation and subsequent surface expression of e-selectin and tissue factor which was not seen from viable or heat-killed gram-positive bacteria (s. aureus, e. faecalis, s. pneumoniae) [13] . neisseria meningitidis adherence on endothelial cells has been reported to be influenced by pilus protein c expression and cd66 on endothelial surfaces [14, 15] and may cause tissue factor expression due to the presence of lps within the cell wall. haemophilus influenzae generally is not toxic to endothelial cells except three clones of biogroup aegyptius causing brazilian purpuric fever [16] . toxicity has been reported to be independent of endotoxin, phagocytosis and replication since irradiation, cycloheximide, cytochalasin d and methylamine have no effect on the ability of the bacterium to invade and cause a cytotoxic response. piliated pseudomonas aeruginosa adheres to and enters human endothelial cells leading to progressive damage [17] or may persist by lysis of endosomal membranes [18] . escherichia coli may invade human brain microvascular endothelial cells involving specialized proteins [19, 20] . endothelial infection by chlamydia pneumoniae also activates endothelial cells to produce cytokines and adhesion molecules and become procoagulant [21] [22] [23] . bartonella quintana, the cause of trench fever transmitted by the body louse, has recently been implicated in culture-negative endocarditis and bacteraemia amongst homeless people [24] . infection and damage of endothelial cells by b. quintana has been demonstrated in vitro and in vivo [25] . interaction of bartonella henselae with endothelial cells may result in bacterial aggregation on the cell surface and the subsequent internalisation of the bacterial aggregate by a unique struc-ture, the invasome [26] . rickettsia rickettsii, an obligate intracellular gram-negative bacterium which causes rocky mountain spotted fever, inhibits endothelial apoptosis allowing to remain inside the host endothelium [27] but is also known to cause a proinflammatory endothelial phenotype. rickettsia conorii causes mediterranean spotted fever which causes endothelial infection with secretion of cytokines, increases in adhesion molecules and induction of surface tissue factor [28, 29] . attachment of borrelia burgdorferi, the agent inducing lyme disease, to endothelial cells may be accomplished by cd14, alpha(v)beta3 and alpha5beta1 integrins or different classes of proteoglycans [30] [31] [32] . b. burgdorferi activates the transcription of chemokine and adhesion in molecule gene expression in endothelial cells [33] . plosmodium infected erythrocytes may bind to endothelial cells via p-selectin, cd36, intercellular adhesion molecule 1 (icam-1) or platelet endothelial cell adhesion molecule 1 (pecam-1) on the endothelial surface [34] [35] [36] . phagocytosed live candida albicans stimulates cytokine secretion and inducible cyclooxygenase expression in endothelial cells [37, 38] . some viral diseases are known to primarily infect endothelial cells and alter their function. dengue virus infection of huvec leads to production of chemoattractant proteins rantes and il-8 [39] , herpes and measles virus infection of brain microvascular endothelial cells increases lymphocyte adhesion by increasing icam-1 [40] and measles virus and cytomegalovirus increases tissue factor expression on huvec [41, 42] . hemorrhagic fever caused by hantaviruses may be accomplished by integrin mediated endothelial infection [43] . on the other hand, ebola virus infects endothelial cells with a transmembrane glycoprotein and inhibits inflammatory responses [44] . exotoxins are known to directly activate endothelial cells. platelet activating factor (paf), no˙and pgi 2 secretion from huvec has been demonstrated by e. coli hemolysin via inositolphosphate/diacylglycerol formation and by s. aureus alpha-toxin via transmembrane ca 2+ entry [45] . there is increasing evidence that hemolytic uremic syndrome results from the systemic action of verocytotoxin producing e. coli on vascular endothelial cells [46] . alpha toxin from clostridium perfringens is a phospholipase c and has been reported to induce adhesion molecule expression and secretion of chemokines from endothelial cells [47] . brain capillary endothelial cells have been reported to express mbec1, a protein that may serve as the c. perfringens enterotoxin receptor [48, 49] . the activity of small gtpase rho has been shown to be altered by c. difficile toxin b [50] and pasteurella mulrocida toxin [51] , which leads to alterations of endothelial permeability. p. aeruginosa exotoxin a may directly injure endothelial cells by a motif shared by many toxins [52] . listeriolysin and phosphatidylinositol-specific phospholipase c secreted from l. monocytogenes has been shown to induce phosphatidyinositol metabolism and diacylglycerol formation in the absence of bacterial uptake by the endothelial cells [53] . some bacterial exotoxins have been used for years as pharmacological tools like pertussis toxin for studying g-protein dependent endothelial cell functions. lipoteichoic acid and peptidoglycan from cell wall components of gram positive bacteria have been shown to induce sepsis in animal models. while lipoteichoic acid has been reported to directly activate endothelial cells [54] , peptidoglycan seems monocyte dependent [55] . most in vitro experiments, however, were performed with different sources of lps as a surrogate activator modelling gram negative bacterial infection. endothelial stimulation using lps in relevant concentrations are usually performed in the presence of serum containing soluble cd14-receptor because endothelial cells generally lack cd14. alternatively, proinflammatory cytokines including tumor necrosis factor alpha (tnf-a), interleukins (il-1, il-4) and interferon gamma (ifng) acting on endothelial cells have extensively been investigated and shown to alter endothelial in vitro functions [56, 57] . the inflammatory response in endothelial cells has been linked to an alteration in reactive oxygen production including superoxide (o 2 -˙) , hydrogen peroxide (h 2 o 2 ), nitric oxide (no˙), hydroxyl radicals (oh˙) and secondary reaction products thereof. o 2 -˙i s believed to be present in unstressed conditions in less than nanomolar quantities within cells. mitochondrial and cytoplasmatic superoxide dismutase readily reacts with o 2 -˙f orming h 2 o 2 which has been measured in micromolar concentrations in human blood. sources of endothelial o 2 -˙p roduction apart from mitochondrial leakage may include metabolism of cytp450 or other metabolic byproducts and production by a nadh oxidase system or by nitric oxide synthase in the absence of l-arginine. some bacterial pathogens are known to be able to produce h 2 o 2 by themselves, however interaction with the endothelium may greatly enhance cellular alterations. streptococcal hemolysin, streptolysin s, is capable of interacting with h 2 o 2 to injure vascular endothelial cells [6] . iron bound to the p. aeruginosa siderophore, pyochelin, augments oxidantmediated endothelial cell injury by modification of transferrin to form iron complexes capable of catalyzing the formation of oh˙from o 2 -˙a nd h 2 o 2 [58] . r. rickettsii infection of the endothelial cell line ea.hy 926 and huvec has been demonstrated to cause glutathione depletion, a major intracellular antioxidant, and reduced glutathione peroxidase activity leading to increased amounts of intracellular peroxide [59] . an increase in reactive oxygen species production has been shown in endothelial cells after incubation with lps, il-1, tnf-a and ifn-g [60] [61] [62] [63] [64] [65] . tnf-a has many times been reported to increase, e.g., adhesion molecule expression inhibitable by various antioxidants [62, [66] [67] [68] . nitric oxide, like o 2 -˙, is chemically not very reactive at all. endothelial production has been established by either constitutive no-synthase (nos) iii in a ca 2+ and phosphorylation dependent manner or by inducible nos ii. direct biochemical actions of no˙include metalcomplex containing proteins, other radical species, oxygen and oxygen derivatives leading to oxidation, nitrosation and nitration. relevant concentrations in vivo have been reported to range from nm to 100 mm. no˙directly reacts with oxyhemoglobin (fe 2 -o 2 ) leading to methemoglobin (fe 3+ ) and nitrate (no 3 -). no˙reacts with o 2 forming nitrite (no 2 -) via intermediate no 2 and n 2 o 3 . mainly n 2 o 3 is participating in n-or s-nitrosylations leading to nitrosamines or nitrosothioles, the latter may serve as a circulating source or as a transporter across cell membranes. reduced glutathione has a high affinity to n 2 o 3 and may also be important in toxicity related to no˙autoxidation. toxicity related cellular targets of no˙may include inhibition of cytochrome c oxidase, inhibition of catalase or dna damage. on the other hand, no˙has been reported to inhibit iron catalysed fenton reaction and to inhibit lipid peroxidation. iron nitrosyl formation in heme containing proteins are the best characterized reactions for no˙in biology. this type of interaction includes very sensitive stimulation of guanylate cyclase and inhibition of cytochrome c oxidase. e. coli hemolysin and s. aureus alpha toxin induce no˙formation in cultured porcine pulmonary endothelial cells [69] . transformed mouse endothelial cells stimulated by the combination of ifn-g and tnf-a killed intracellular r.conorii by a mechanism that required the synthesis of no˙ [70] . ifn-g, tnf-a and il-1 stimulated murine endothelial cells have been shown to kill schistosoma mansoni through the production of nitric oxide [71] . both no˙and o 2 -˙p roduction may have a limited influence on endothelial viability under resting conditions. cultured bovine and porcine aortic endothelial cells showed decreased no˙production after 1 h of lps incubation which was related to decreases in capacitative ca 2+ signals [72] . posttranscriptional destabilization of nos iii mrna may have accounted for this early decrease in no˙production [73] . nos ii, which is believed to produce larger amounts of no˙is also known to be regulated by many proinflammatory stimuli with significant cell type variablilities. at sites of endothelial involvement in an inflammatory process both vascular non-endothelial and non-resident cells are known to produce no˙. as the amount and physiological consequences of no˙produced from noss at the microcirculatory level is not known, it may well serve also to reduce inflammatory processes as recently implicated from a coculture experiment [74] . that an increase in reactive oxygen species is present in human sepsis has been documented by almost any study trying to quantify secondary reaction products by several methods, however the cellular sources remain speculative ( table 2 ). in 1990 beckman et al. showed that the presence of both no˙and o 2 -˙p roduced peroxynitrite (onoo -) which may decompose to produce ho˙like molecules and thereby kill endothelial cells [75] . at ph 7 its lifetime is in the order of a second. half of the onoo formed is rapidly equilibrated to peroxynitrous acid (onooh) and breaks down to no 3 11 sepsis immunohistochemical endothelial nitrotyrosine≠ [176] 3 septic shock no 2 -/no 3 -, plasmatic nitrotyrosine≠ [177] 3 septic lung injury immunohistochemical endothelial nitrotyrosine≠ antioxidant capacity was defined as the ability of plasma to inhibit * ferryl myoglobin production by hydrogen peroxide addition to metmyoglobin or ** oxo-iron induced damage to deoxyribose, phospholipids and dna. tbars, thiobarbituric acid reactive substances; xod, xanthine oxidase. models and therefore question the view that this molecule solely is detrimental. toxicity induced by an interaction between hydrogen peroxide and nitric oxide has led to conflicting results. rat lung microvascular endothelial cells and porcine pulmonary artery endothelial cells exposed to h 2 o 2 have been reported to be protected by no˙donors [76, 77] , whereas toxicity in bovine aortic endothelial cells [78] and in rat liver microvascular endothelial cells [79] was increased in the presence of no˙donors. no˙in the presence of h 2 o 2 may also produce oh˙like molecules independent of the presence of iron [80] . neutrophils may add to the complexity of toxic reactions. myeloperoxidase from activated neutrophils, which produces hocl and oh˙molecules in the presence of o 2 -˙, has recently been demonstrated to convert no 2 into no 2 , thereby damaging endothelial cells [81] . high doses of reactive oxygen species (including no˙) have been shown to cause apoptosis of endothelial cells, whereas low doses were protective [82] . in mice disseminated endothelial apoptosis has been suggested to be responsible for organ failure and shock induced by endotoxin or tnf-a [83] . both lps or tnf-a usually do not cause endothelial cell death unless protein synthesis is blocked probably due to simultaneous increases in antiapoptotic protein synthesis [84] . however, postmortal investigation in humans deceased from or with sepsis did not confirm these results [85] . most of the genes activated in vitro during the endothelial stress response are controlled by at least two transcription factor families: activator protein 1 (ap-1) and nuclear factor kappa b (nfkb). nfkb has gained wide interest as a target in inflammatory diseases as it seems to be invariably upregulated [86] . a variety of agents including cytokines and reactive oxygen stress cause ikb to dissociate from the complex after phosphorylation by ikb-kinase complex. h 2 o 2 has been reported to activate transcription factor nfkb in porcine aortic endothelial cells [87] whereas huvec were unresponsive [88] . no˙has in most cases been shown to inhibit transcription factor nfkb and its influence on transcription factor ap-1 is unclear. inhibition of nfk-b as a therapeutic means has been suggested. however, as this transcription factor is also involved in protective gene regulation, its inhibition can make cells sensitive to e.g. tnf-a [89] . data on activated intracellular signalling pathways in sepsis patients are scarce but include nfkb within mononuclear cells [90] . usually any blood cell is kept off endothelial surfaces. this is believed to be accomplished by net electrical charge, biomechanical characteristics of flowing blood and the secretion of no˙. if blood cells touch the endothelium a ca 2+ -signal is induced [91] , but it is unclear at present whether this has any functional consequence. it is tempting to speculate that ca 2+signals may then in a context sensitive manner augment proadhesive processes or antiadhesive endothelial properties. activated endothelial cells are potent producers of cytokines like il-1, a major proinflammatory cytokine, and chemotactic peptides including il-8 for neutrophils, macrophage inflammatory protein-1 alpha (mip-1a), monocyte chemoattractant proteins 1-4 and rantes (regulated on activa-tion, normal t cell expressed and secreted) for monocytes, t-lymphocytes and dendritic cells, growth related protein (gro) and gamma-interferon-inducible protein (ip-10) for activated t-lymphocytes, epithelial neutrophil activating peptide 78 (ena-78), vascular monocyte adhesion-associated protein (vmap-1) and endothelial monocyteactivating polypeptide ii (emap-ii) for monocytes [92] . many adhesion molecules are expressed on the surface of endothelial cells in a highly complex yet regulated manner. p-selectin, e-selectin, icam-1, vascular cell adhesion molecule 1 (vcam-1), pecam-1 are well known for their stimulus-, cell-, time-and organ specific dependence of expression and their importance in regulation of leukocyteendothelial interactions. moreover, apart from being passive adhesion molecules, all of the above mentioned molecules have been shown to signal inside endothelial cells upon receptor engagement. shed receptors from endothelial surfaces may also serve as endogeneous antiadhesive molecules demonstrating even more the dynamic and complex nature of these processes. soluble forms of adhesion molecules have been shown to be present in high amounts in human sepsis (table 3) , however whether this is beneficial or detrimental is not known and the assumption that these parameters may serve as practical indexes remains to be established. particularly ifn-g has been repeatedly shown to increase endothelial hla-dr expression rendering them capable of mhc class ii restricted interactions with cd4 + t-cells. costimulatory cd80 (b7-1) and cd86 (b7-2) are usually not present on endothelial surfaces leading to the conventional view that endothelial cells are semiprofessional antigen presenting cells. however, under certain conditions both costimulatory molecules and cd40 can be upregulated on endothelial surfaces indicating excessive antigen presentation [93] . migration of lymphocytes into inflamed mouse tis-vol. 49, 2000 endothelial function in sepsis 189 sues has been reported to depend on psgl-1 and esl-1 binding endothelial p-selectin and e-selectin, respectively [94] . this phenotype was found to be restricted to th1 lymphocytes, but human sepsis seems to be predominated by a th2 type [95] . both tnf-a and ifn-g are able to promote transmigration of leukocytes, whereas coapplication of both cytokines inhibit this process [96] . t-cells migrating through the endothelial barrier in a pecam-1-dependent manner are subject to inhibitory signals which may limit their activation in tissues [97] . tnf bound to monocytes has been shown to inhibit endothelial apoptosis, whereas this process is promoted in the presence of lymphocytes [98] . unperturbed endothelial cells express fas-ligand which has been implicated as a means of signalling apoptosis to constitutively fas receptor (cd95) bearing cells, whereas tnf-a treated endothelial cells decrease fas ligand expression thereby allowing leukocyte survival during extravasation [99] . how the endothelium might interact specifically with lymphocytes or monocytes in septic patients can only be speculated on. neutrophils interacting with endothelial cells have repeatedly been shown to cause toxicity due to the release of enzymes and reactive oxygen species. if uncontrolled, these cells are believed to mediate significant tissue damage. however whether uncontrolled activation or rather deactivation is present in human sepsis is a matter of debate. endothelial cells have been reported to inhibit some neutrophil functions [100] . transmigrated neutrophils may actively participate in the endothelial resealing process by the secretion of adenosin precursors [101] . however, massive leukocyte extravasation as one would expect from most animal studies has never been shown in septic patients [102] . endothelium is known to regulate transvascular fluid flux, flux of nutrients, mediators and cells by either paracellular or transcellular vacuolar channel related pathways. lateral junction proteins including the vascular endothelial cadherin-complex, platelet-endothelial cell adhesion molecule-1, occludin, zona occludens-1, recently described junctional adhesion protein, cd151/platelet endothelial tetraspan antigen and cd81/target of antiproliferative antigen are known to participate in this process. paracellular permeability is achieved by either an active contraction or the controlled release of an intrinsic tone mediated in most cases by the action of myosin light chain kinase (mlck) acting on non muscle myosin. generally an increase in the concentration of camp keeps cultured endothelial monolayers tight and cgmp has been reported to assist this function in human aortic and foreskin vessels [103] . endothelial retraction may be initiated by increases in intracellular ca 2+ concentration, but elevation of ca 2+ in the presence of maintained camp-kinase dependent phosphorylation is not edemagenic. these antagonistic effects of ca 2+ and camp in endothelial permeability regulation have recently been reviewed by moore et al. [104] . many reports documented increases in endothelial permeability involving exotoxins like s. aureus alpha-toxin and p. aeruginosa cytotoxin [105, 106] or endotoxins [107] . for example, p. multocida toxin has been shown to activate rho/rho kinase, which inactivates mlc phosphatase. the resulting increase in mlc phosphorylation caused endothelial cell retraction and a rise in endothelial permeability [51] . lps induced increases in paracellular permeability by caspase activated cleavage of adherens junction proteins [108] . counteracting lipid peroxidation during lps activation may inhibit increases in permeability [109] . tnf-a stimulated endothelial cadherin complex is disrupted in a proteasome dependent manner [110] . the resulting increase in permeability has been reported to lower camp and activate phosphediesterase ii and iv [111] . h 2 o 2 induced hyperpermeability of porcine pulmonary endothelial cells has been reported to be effectively reduced by cgmp elevating drugs including phosphodiesterase ii inhibition or no˙donators [112] . the electroneutral na-k-cl cotransport system is thought to function in the maintenance of a selective permeability. il-1, tnf-a and lps upregulate the expression of a bumetanide-sensitive na-k-cl cotransporter subtype in huvec and in murine lung and kidney endothelial cells [113] . septic rats show different increases in albumin flux accross several endothelial beds [114] . increases in venular permeability have been shown to be preventable by the antioxidants n-acetyl-cystein or tirilazad mesylate in e. coli infused rats [115, 116] . il-10, an antiinflammatory cytokine not produced by endothelial cells, was shown to participate as an inhibitor of endothelial permeability induced by lps in mice [117] . clinically, increases in endothelial permeability may be obvious in many septic patients, but only recently venous congestion plethysmography showed a selectively elevated filtration capacity as a measure of endothelial dysfunction in septic patients [118] . which of the many pathways of increased permeability might be turned on and whether it persists remains unknown. in principle endothelial cells are believed to be anticoagulatory by virtue of their surface expression of glycosaminoglycan-antithrombin iii complex, thrombomodulin, heparin releasable tissue factor pathway inhibitor and production of adenosine by ecto-adpases, their secretion of protein s, prostacyclin and no˙. no˙production was shown to participate in heparan sulfate preservation in porcine aortic endothelial cells [119] and may be responsible for prostacyclin secretion by activating cyclooxygenase-1 under resting conditions [120] . endothelial release of plasminogen activator (t-pa) and plasminogen activator inhibitor 1 (pai-1) may determine the fibrinolytic potential of plasma. endothelial cells specifically bind coagulation factors xii, iia, ix, viia and xa. xa was shown to bind to endothelial effector cell protease receptor-1 and thereby cause release of no˙, il-6, il-8, mcp-1 and functional upregulation of icam-1, e-selectin and vcam-1 [121] . when endothelium is perturbed by physical or chemical factors transformation to a prothrombotic surface is invariably seen in in vitro models. thrombomodulin surface expression on huvec can easily be downregulated by lps, il-1 and tnf-a and upregulated by increasing camp [122] . a tnf-a induced decrease in surface thrombomodulin has been suggested via activation of phosphodiesterase ii and iv thereby decreasing camp in baec [111] . vascular endothelial growth factor may counteract il-1, tgf-b and lps induced suppression of both thrombomodulin surface antigen and mrna [123] . adenosin nucleotides are released from damaged as well as lps stimulated and shear stressed huvec [124] . endothelial cells have atp diphosphohydrolase (cd 39) on their surface to degrade atp via adp and amp to adenosine. adenosine is known to have antiaggregatory properties due to stimulation of prostacyelin and no˙production. however, activating endothelial cells with tnf-a has been reported to cause loss of atp diphosphohydrolase activity, which was preventable in the presence of antioxidants [125] . tissue factor (tf) expression on endothelial cells by bacteria, lps, il-1 and tnf-a is well known. tissue factor pathway inhibitor (tfpi), a serine protease inhibitor of xa and xa/viia/tf complex on endothelial surfaces, which immediately blocks tissue factor activation, has been shown to be decreased under proinflammatory conditions. another counterbalancing mechanism includes shear stress in tnf-a stimulated huvec [126] . however, an anticipated increase in endothelial tissue factor expression has not convincingly been demonstrated in animal or human sepsis [127, 128] . fibrinolytic systems on endothelial surfaces are also believed to be altered in sepsis. increases in pai-1 has been reported after stimulation with il-1 or lps [129, 130] . however soluble pai-1 in septic patients was not found to be different from nonseptic patients [131] . once thrombin formation has occured, its cleavage of endothelial proteinase activated receptor (par) in turn may lead to secretion of il-8 and il-6 [132] , upregulation of icam-1 and vcam-1 [133] , relaxation via no˙production or to vasoconstriction by an as yet unidentified factor [134] . these data may demonstrate an interconnection between coagulation activation, inflammation and vasoregulation mediated by the endothelium. coagulation activation is clearly present in septic patients, however endothelial participation in this process is unclear. potent procoagulatorv sources may well include bacterial surfaces per se [135] or monocytes [128] . up to the late 60ies the endothelium was viewed as a passive organ, which at best was able to remove vasoactive hormones in the lung. in 1976-1978 sir john vane's group (noble laureate 1982) reported that endothelial cells can synthesize i series prostaglandins (like prostacyclin) and thereby relax arteries and inhibit platelet aggregation. however, whether pgi 2 regulates basal vascular tone is unclear. after a new technician used an unintended vessel preparation robert furchgott realized after a series of contradicting results that acetylcholine (ach) was no longer able to relax precontracted arteries when endothelium was removed and termed the nonprostanoid mediator an endothelium derived relaxing factor (edrf). in 1986 superoxide was shown to participate in vasoregulation. the presence of superoxide dismutase prolonged the action of edrf whereas addition of o 2 -˙i nactivated edrf. even direct effects of several reactive oxygen species have been suggested to be relevant in cerebral or coronary circulation. collectively robert furchgott, louis ignarro and ferrid murad received the noble laureate in 1998 for demonstrating that no˙is a major edrf. endothelial cells produce more relaxing factors which pharmacologically can be separated from nitric oxide action and these were termed endothelium derived hyperpolarizing factors (edhfs). these factors may particularly be important in coronary and gastrointestinal vessels. epoxyeicosatrienoic acids, anandamide, the endogenous ligand of cannabinoid receptors or simply the release of k + may constitute edhfs. conceptually shear may in larger vessels primarily determine production of no˙, whereas cyclic strain determines physiological edhf release. soon after the discovery of edrf endothelin-1 (et-1), a vasoconstricting peptide produced from endothelial cells was isolated. low doses of et-1 can induce no˙release and subsequent relaxation via et breceptors on endothelial cells. endothelin secretion is thought to occur abluminally leading to et a -receptor activation on smooth muscle cells and subsequent vasoconstriction. many other factors clearly contribute to endothelial control of vasoregulation by e.g. transcellular production of vasoconstricting prostanoids like thromboxane a 2 or prostaglandin h 2 or the enzymatic conversion of angiotensin i to angiotensin ii by angiotensin converting enzyme. a hallmark of sepsis is the heterogeneous pattern of vasoconstriction and vasodilatation in different organs, culminating in a fall in total peripheral vascular resistance concomitant with regional maldistribution of blood flow. vasoactive substances produced by the endothelium under experimental septic conditions are known to be altered by factors such as no˙, pgi 2 , angiotensin converting enzyme (ace) activity, endothelin and adrenomedullin. endothelium dependent vasoregulation has largely been studied in animal models (table 4 ). in 1985 endothelium dependent vasoregulatory failure was seen as a defect in reactive hyperemia related vasodilator release [136] and decreased dilatation of arterioles induced by ach [137] . in a rat model of cecal ligation and puncture decreased vasoconstriction was found after administration of norepinephrine in septic animals which was largely reversible by removal of the endothelium [138] . parker et al. [139] showed in explanted coronary arteries and aortas of guinea pigs treated with lps intraperitoneally for 16 h that endothelium dependent relaxation induced by acetylcholine and adp was depressed, whereas relaxation induced by substance p or receptor independent relaxations by ca 2+ -ionophore a23187 was unaffected. edrf release and bioactivity from explanted aortas of these animals was decreased after adp or ach stimulation, whereas a23187 induced edrf release was unaltered [140] . this group also demonstrated reduced adp and ach responses after 4 h of lps endotoxemia in guinea pigs and that adp may produce constricting thromboxane in septic animals [141] . in contrast, coronary arteries of rabbits treated for 5 weeks with low doses of lps stimulation with ach but not adp showed increased relaxation of explanted vessels [142] . wang et al. [143] isolated subepicardial arterioles from rats treated 48 h intraperitoneally with feces containing life e. coli and showed in a pressurized no-flow chamber that relaxation by alpha 2 agonist clonidine and adp was reduced in an endothelium dependent manner, which could be inhibited by the nos inhibitor lnma. also, in this model mesenteric arter-vol. 49, 2000 endothelial function in sepsis iolar relaxation after adp and clonidine was decreased, whereas in skeletal muscle these agonists caused vasoconstriction [144] . pulmonary arteries of rats treated with lps showed depressed endothelin-1 induced contractions which were even augmented in endothelium denuded vessels. the authors [145] concluded that a vasoconstrictor eicosanoid is produced in lps treated animals by pulmonary endothelium upon et-1 stimulation. swine infused with live p. aeruginosa showed no alteration in endothelium dependent bradykinin and endothelium independent nitroprusside relaxation, whereas ach induced relaxation was found to be reduced in explanted peripheral arteries [146] . chaudry's group investigated endothelium dependent relaxation in rats treated with cecal ligation and puncture. a time dependent alteration was demonstrated with increased ach induced vasorelaxation early after challenge whereas depressed vasodilatation after 5-20 h was found in explanted aortas with no alteration in nitroglycerine induced relaxation [147] . the decreased endothelium dependent response to ach was also found in superior mesenteric arteries and small intestinal arteries [148] . in the same model this group demonstrated a reduction of immunodetectable nos iii in explanted aortas [149] . porcine coronary arterioles incubated for 20 h with e. coli lps (100 mg/ml) decreased bradykinin induced edhf secretion [150] . carotid and coronary arteries from rabbits also showed decreases in edhf-release in an ex-vivo assay after treatment with lps, tnf-a and il-1 [151] . collectively these data indicate that the majority of sepsis models consistently show a disruption of receptor coupled relaxation mechanism leading to an intraendothelial signalling deficit. to quantify endothelial function in humans several methods are available. endothelium dependent relaxation after pharmacological stimulation or flow dependent relaxation after vessel obstruction are frequently quantified by high resolution ultrasound techniques [152] , alternatively flow and size can be determined angiographically. however, definite functional measurements in human sepsis are scarce. endothelium dependent relaxation has been investigated in isolated superficial hand veins of healthy volunteers after lps exposure. reduced vasorelaxation by bradykinin and arachidonic acid in noradrenalin precontracted vessels were noted which persisted for more than 2 days [153] . reactive hyperemia is believed to mainly test endothelial no˙production upon shear stress if vessel diameters and flow is monitored. implications from indirect measurements support an endothelial dysfunction in septic patients [154] [155] [156] . however, data on pharmacological stimulation of endothelium dependent relaxation in humans are currently not available. 192 t. volk +/-ec: presence or absence of endothelium; ne: norepinephrine; ach: acetylcholine; adp, adenosine diphosphate; sp, substance p; a23187, ca 2 + -ionophore; cox, cycloogygenase; snp, sodium nitroprusside; ntg, nitroglycerine; et, endothelin; bk, bradykinin; edhf, endothelium derived hyperpolarizing factor; pres., preserved. table 4 . endothelium dependent relaxation is impaired in animal sepsis models. a normal response to infection or other insults is a self limiting process that through temporal expression of regulators and effector molecules causes resolution. the failure to resolve the causative infection may lead to sepsis. cellular and animal models of sepsis using bacteria, endotoxins, exotoxins, cytokines or some peptides all consistently produce endothelial impairment which is usually regarded as dysfunctional. blocking the majority of pathways used by these inducing agents has often lead to the inhibition of such endothelial alterations. many aspects of these induced alterations can be expected to reveal exciting new pathways and complex interactions at various molecular and cellular level. the past has taught us that inhibitors of presumably activated pathways consistently failed to improve survival in septic patients. this has stimulated many researchers to reconcile the results of experimental and clinical models in sepsis. compared to activating pathways, considerably less is known about how an inflammatory response is endogenously counterregulated. cytokines induce a whole host of signal inhibiting proteins and endogenous counterregulating systems are just beginning to be elucidated. activation of endogenous counterregulatory systems may become the predominant feature of the so-called compensatory antiinflammatory response syndrome. endothelial responses to endogenously present antiinflammatory mediators have hardly been investigated in sepsis models. almost all endothelial cell studies in sepsis indicated that an imbalance in reactive oxygen species production is associated with the above described dysfunctions. animal studies in which endothelial function could be improved pharmacologically also consistently indicate that reversal of imbalanced reactive oxygen production may be a common link (table 4 ). it seems that at times an adequate production of nitric oxide is lacking whereas superoxide and/or derivatives are overproduced. however, as endothelial functional measurements in septic humans become available, we will hopefully get a clearer picture of what might happen in our patients. [199] clp, cecal ligation and puncture; lpo, lipid peroxidation; ros, reactive oxygen species. table 5 . treatment of endothelial vasoregulatory dysfunction in animal sepsis models. bacterial pathogens isolated from patients wit bloodstream infection: frequencies of occurrence and antimicrobial susceptibility patterns from the sentry antimicrobial surveillance program (united states and canada, 1997) staphylococcus aureus infections internalization of staphylococcus aureus by endothelial cells induces apoptosis staphylococcus aureus small colony variants are induced by the endothelial cell intracellular milieu genetic inactivation of the extracellular cysteine protease enhances in vitro internalization of group a streptococci by human epithelial and endothelial cells interaction of viable group a streptococci and hydrogen peroxide in killing of vascular endothelial cells group b streptococci invade endothelial cells: type iii capsular polysaccharide attenuates invasion invasion of brain microvascular endothelial cells by group b streptococci streptococcus pneumoniae anchor to activated human cells by the receptor for platelet-activating factor pneumococcal trafficking across the blood-brain barrier. molecular analysis of a novel bidirectional pathway two distinct phospholipases c of listeria monocytogenes induce ceramide generation, nuclear factor-kappa acivation, and e-selectin expression in human endothelial cells internalin b is essential for adhesion and mediates the invasion of listeria monocytogenes into human endothelial cells activation of human endothelial cells by viable or heat-killed gram-negative bacteria requires soluble cd14 interaction of neisseria maningitidis with the components of the blood-brain barrier correlates with an increased expression of pilc the ndomain of the human cd66a adhesion molecule is a target for opa proteins of neisseria meningitidis and neisseria gonorrhoeae human microvascular endothelial tissue culture cell model for studying pathogenesis of brazilian purpuric fever pseudomonas aeruginosa selective adherence to and entry into human endothelial cells endothelial function in sepsis 193 cincomitant endosome-phagosome fusion and lysis of endosomal membranes account for pseudomonas aeruginosa survival in human endothelial cells endothelial cell glcnac beta 1-4glcnac epitopes for outer membrane protein a enhance traversal of escherichia coli across the blood-brain barrier escherichia coli invasion of brain microvascular endothelial cells in vitro and in vivo: molecular cloning and characterization of invasion gene ibe10 characterization of a strain of chlamydia pneumoniae isolated from a coronary atheroma by analysis of the omp1 gene and biological activity in human endothelial cells chlamydia species infect human vascular endothelial cells and induce procoagulant activity signal transduction pathways activated in endothelial cells following infection with chlamydia pneumoniae bartonella (rochalimaea) quintana endocarditis in three homeless men bartonella quintana invades and multiplies within endothelial cells in vitro and in vivo and forms intracellular blebs interaction of bartonella henselae with endothelial cells results in bacterial aggregation on the cell surface and the subsequent engulfment and internalisation of the bacterial aggregate by a unique structure, the invasome nf-kappa b-dependent inhibition of apoptosis is essential for host cellsurvival during rickettsia rickettsii infection rickettsia conorii infection enhances vascular cell adhesion molecule-1-and intercellular adhesion molecule-1-dependent mononuclear cell adherence to endothelial cells il-6 and il-8 production from cultured human endothelial cells stimulated by infect on with rickettsia conorii via a cell-associated il-1 alpha-dependent pathway the role of cd14 in signaling mediated by outer membrane lipoproteins of borrelia burgdorferi integrins alpha(v)beta3 and alpha5beta1 mediate attachment of lyme disease spirochetes to human cells different classes of proteoglycans contribute to the attachment of borrelia burgdorferi to cultured endothelial and brain cells borrelia burgdorferi upregulates the adhesion molecules e-selectin, p-selectin, icam-1 and vcam-1 on mouse endothelioma cells in vitro characterization of plasmodium falciparum-infected erythrocyte and p-selectin interaction under flow conditions intercellular adhesion molecule-1 and cd36 synergize to mediate adherence of plasmodium falciparum-infected erythrocytes to cultured human microvascular endothelial cells pecam-1/cd31, an endothelial receptor for binding plasmodium falciparum-infected erythrocytes candida albicans stimulates cytokine production and leukocyte adhesion molecule expression by endothelial cells secreted aspartyl proteinases and interactions of candida albicans with human endothelial cells dengue virus infection of human endothelial cells leads to chemokine production, complement activation, and apoptosis adhesion molecule expression and lymphocyte adhesion to cerebral endothelium: effects of measles virus and herpes simplex 1 virus measles virus induction of human endothelial cell tissue factor procoagulant activity in vitro effects of viral activation of the vessel wall on inflammation and thrombosis cellular entry of hantaviruses which cause hemorrhagic fever with renal syndrome is mediated by beta3 integrins ebola virus inhibits induction of genes by double-stranded rna in endothelial cells human endothelial cell activation and mediator release in response to the bacterial exotoxins escherichia coli hemolysin and staphylococcal alpha-toxin infection by verocytotoxin-producing escherichia coli alpha toxin from clostridium perfringens induces proinflammatory changes in endothelial cells brain capillary endothelial cells express mbec1, a protein that is related to the clostridium perfringens enterotoxin receptors phospholipase c and perfringolysin o from clostridium perfringens 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standardization committee of the isth effects of lipopolysaccharide on the expression of fibrinolytic factors in an established cell line from human endothelial cells induction of plasminogen activator inhibitor type 1 and type 1 collagen expression in rat cardiac microvascular endothelial cells by interleukin-1 and its dependence on oxygencentered free radicals plasminogen: an important hemostatic parameter in septic patients potential mechanisms for a proinflammatory vascular cytokine response to coagulation activation thrombin-activated human endothelial cells support monocyte adhesion in vitro following expression intercellular adhesion molecule-1 cd54) and vascular cell adhesion molecule-1 (vcam-1; cd106) dual endothelium-dependent vascular activities of proteinase-activated receptor-2-activating peptides: evidence for receptor heterogeneity activation of the contact-phase system on bacterial surfaces -a clue to serious complications in infectious diseases reactive hyperemic 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plasma von willebrand factor in the systemic inflammatory response syndrome is derived from generalized endothelial cell activation blood levels of endothelin-1 and thrombomodulin in patients with disseminated intravascular coagulation and sepsis plasma levels of endothelial cell protein c receptor are elevated in patients with sepsis and systemic lupus or erythematosus: lack of correlation with thrombomodulin suggests involvement of different pathological processes demonstration of rickettsia conorii-induced endothelial injury in vivo by measuring circulating endothelial cells, thrombomodulin, and von willebrand factor in patients with mediterranean spotted fever influence of angiotensin-converting enzyme inhibitor enalaprilat on endothelial-derived substances in the critically ill mesenteric and skeletal muscle microvascular responsiveness in subacute sepsis influence of group b streptococci on piglet pulmonary artery response to bradykinin effects of antisense oligonucleotide to inos on hemodynamic and vascular changes induced by lps pentoxifylline maintains vascular endothelial cell function during hyperdynamic and hypodynamic sepsis a novel nonanticoagulant heparin prevents vascular endothelial cell dysfunction during hyperdynamic sepsis effect of endotoxin-enhanced hepatic reperfusion injury on endonthelium-dependent relaxation in rat aorta splanchnic vascular endothelial dysfunction in rat endotoxemia: role of superoxide radicals endothelial dysfunction in a rat model of endotoxic shock. importance of the activation of poly (adp-ribose) synthetase by peroxynitrite endothelial cell dysfunction in septic shock key: cord-002956-e5ihpe4i authors: chang, ya-chun; huang, kuo-tung; chen, yu-mu; wang, chin-chou; wang, yi-hsi; tseng, chia-cheng; lin, meng-chih; fang, wen-feng title: ventilator dependence risk score for the prediction of prolonged mechanical ventilation in patients who survive sepsis/septic shock with respiratory failure date: 2018-04-04 journal: sci rep doi: 10.1038/s41598-018-24028-4 sha: doc_id: 2956 cord_uid: e5ihpe4i we intended to develop a scoring system to predict mechanical ventilator dependence in patients who survive sepsis/septic shock with respiratory failure. this study evaluated 251 adult patients in medical intensive care units (icus) between august 2013 to october 2015, who had survived for over 21 days and received aggressive treatment. the risk factors for ventilator dependence were determined. we then constructed a ventilator dependence (vd) risk score using the identified risk factors. the ventilator dependence risk score was calculated as the sum of the following four variables after being adjusted by proportion to the beta coefficient. we assigned a history of previous stroke, a score of one point, platelet count less than 150,000/μl a score of one point, ph value less than 7.35 a score of two points, and the fraction of inspired oxygen on admission day 7 over 39% as two points. the area under the curve in the derivation group was 0.725 (p < 0.001). we then applied the vd risk score for validation on 175 patients. the area under the curve in the validation group was 0.658 (p = 0.001). vd risk score could be applied to predict prolonged mechanical ventilation in patients who survive sepsis/septic shock. imposes a financial burden on patients, families, and the healthcare facility. furthermore, it decreases the patient's quality of life. we sought to determine the risk factors for ventilator dependence in patients who survive sepsis and septic shock. patient characteristics and findings. a total of 379 patients with sepsis or septic shock and acute respiratory failure requiring mechanical ventilation were admitted to the medical intensive care unit in the kaohsiung chang gung memorial hospital from august 2013 to october 2015. of the 379 patients, 251 patients were enrolled in the study (fig. 1) . for validation, we collected data from sepsis/septic shock patients with mechanical ventilation admitted to the medical icu from november 2015 to november 2016 and 175 patients were included in the study (fig. 2) . among the 251 patients studied, 69.3% were admitted from the emergency room. the site of suspected infection was of pulmonary origin in 60.6% (table 1) . 72 patients (29%) were ventilator-dependent (on the ventilator for at least 21 days), and 179 (71%) patients were ventilator-independent (needing ventilator support for less than 21 days). the mean age (standard deviation (sd)) of ventilator-dependent patients was older (64.59 (15.67) vs. 69. 40 (15.65) , p < 0.05) ( table 2) . no statistical differences were observed in the apache ii score (p = 0.664) or the initial sofa score (p = 0.184) between the ventilator-dependent and ventilator-independent groups. however, there was a significant difference between the ventilator-dependent and ventilator-independent groups regarding previous stroke (p = 0.027 in the univariate analysis) ( table 2) . hematology, biochemistry, fio 2 , and pao 2 /fio 2 were collected for further analysis ( table 2 ). the platelet count was lower on admission day 7 in the ventilator-dependent group compared with the ventilator-independent ventilator dependence risk score. we constructed a ventilator dependence risk score using individual risk factors, which were first identified from the univariate analysis. risk factors on admission day 1, day 3 and day 7 in univariate analysis included white blood cells, hemoglobin, platelet, prothrombin time, inr, ast, alt, bun, creatinine, na, k, c-reactive protein, albumin, lactate, procalcitonin, ph, paco 2 , bicarbonate, fio 2 , pao 2 / fio 2 , etc. variables on admission day 1, admission day 3 and admission day 7, which were possibly associated with ventilator dependence in the univariate analysis (p < 0.1), were included in a multivariate analysis model. a total of 11 variables were included in the multivariate analysis model, such as age >69 years, fio 2 >39%, history of stroke, ph < 7.35, platelet <150,000/μl, history of coronary artery disease, hemoglobin, bun, c-reactive protein, pao 2 /fio 2 and paco 2 on admission day 7. after stepwise method, the independent factors associated with ventilator dependence were identified to build a score. a clinical score (vd risk score) was calculated based on four variables independently associated with ventilator dependence in the multivariate analysis, including previous stroke, thrombocytopenia, acidosis, and higher fio 2 ( table 3 ). the ventilator dependence risk score was calculated as the sum of these four variables after adjusting by proportion to beta coefficient. we assigned a history of stroke one point, platelet count on admission day 7 of less than 150,000/μl one point, ph value on admission day 7 of less than 7.35 two points, and the fraction of inspired oxygen on admission day 7 over 39% two points (table 4) . receiver operating characteristic curves were plotted in fig. 3 . the area under the curve (auc) of the ventilator dependence risk score was 0.725 with p value < 0.001. a ventilator dependence risk score equal to or more than one point yielded 80.5% sensitivity and 50.2% specificity. sofa score for ventilator dependence prediction. we tested sequential organ failure assessment (sofa) score 19 on admission day 1 and day 7 to predict ventilator dependence on sepsis and septic shock patients with respiratory failure. the area under the curve (auc) of the sofa score on admission day 1 was 0.441 and the auc of the sofa score on admission day 7 was 0.662. we also tested and found that sofa pao 2 /fio 2 subscore and gcs subscore on admission day 7 could help predict ventilator dependence on sepsis and septic shock patients with significant difference in univariate analysis ( table 5 ). the area under the curve of the pao 2 /fio 2 subscore on admission day 7 was 0.668 and the auc of the gcs subscore on admission day 7 was 0.673 (fig. 3 ). validation ventilator dependence risk score. for validation, we collected data from sepsis/septic shock patients with mechanical ventilation admitted to the medical icu from november 2015 to november 2016. we used the ventilator dependence risk score to predict ventilator dependence in these 175 patients. patient characteristics and underlying disease are collected in revealed as table 6 . the auc of the ventilator dependence risk score was 0.658 and the p-value was 0.001 (fig. 4) . a ventilator dependence risk score equal to or more than one point yielded 69.0% sensitivity and 53.0% specificity. we also found the auc of ventilator dependence risk score was 0.745 in the cancer group and p-value was 0.009. auc of ventilator dependence risk score was 0.723 in the chronic kidney disease group (p = 0.009) (fig. 5 ). in this study, we identified risk factors for prolonged mechanical ventilation in patients who survived sepsis and septic shock. these included a history of stroke, and data collected on day 7 (thrombocytopenia, acidosis, and a higher fraction of inspired oxygen). the ventilator dependence risk score can help easily predict prolonged mechanical ventilation. we chose biochemical and physiological variables from day 7 to incorporate into our score, as opposed to day 1 or day 21, which would each have advantages and disadvantages. for example, it is too late to predict ventilator dependency using day 21 data. on the other hand, with multiple factors and different treatment response, it is difficult to predict ventilator dependency from day 1 data. with aggressive treatment in the first week, day 7 data can help identify which patients face a substantial risk of becoming long-term ventilator-dependent. patients who have suffered a stroke in the past often have respiratory dysfunction due to respiratory drive impairment. according to a previous study, respiratory function depends on numerous neurologic structures, which extend from the cerebral cortex to the medulla; complications after an injury to the respiratory center could lead to prolonged mechanical ventilation 20, 21 . therefore, a previous stroke is an independent risk factor for predicting prolonged ventilator use. sepsis is a life-threatening organ dysfunction caused by a disproportionate host response to infection and it involves complex mechanisms 22 . during sepsis, platelet numbers decrease due to increased platelet destruction. sepsis may result in hypercoagulation due to fibrin deposition and platelet activation. this leads to the formation of micro-thrombi as a host defense mechanism against pathogens, in which platelets play a crucial role. in extreme situations, this may progress to disseminated intravascular coagulation (dic), with severe thrombocytopenia and coagulation system impairment [23] [24] [25] . platelet dysfunction during sepsis correlates with a poorer prognosis. thus, the morphology, number, and function of platelets may be used as biomarkers for the risk stratification of patients with sepsis 25 . although we excluded very ill patients with decreased platelet counts who expired within 21 days (in our series, average 152*10 3 /μl), the platelet count on day 7 could differentiate the ventilator-dependent and independent groups on day 21. a relatively low platelet count on admission day 7 suggests that a septic patient has not completely recovered and may have greater risk of ventilator dependence. although there was a significantly lower hemoglobin level in the ventilator-dependent group, it is hard to suggest that bleeding caused by thrombocytopenia is causing weaning failure. the hemoglobin level in both groups was greater than 10 g/dl. acidosis is increased acidity (hydrogen ion concentration) in the blood and other body tissues. it occurs when the arterial ph falls below 7.35. sepsis can cause tissue hypoperfusion and the accumulation of lactate, which causes metabolic acidosis 26 . acidosis resolution in survivors was attributable to a decrease in strong ion gap and lactate levels 26 . additionally, respiratory acidosis can be due to the accumulation of carbon dioxide in the lungs, which indicates poor lung functioning 27 . our data revealed that arterial blood gas acidosis on day 7 was one of the independent risk factors predicting ventilator dependence. we did not find statistical differences between groups on higher lactate levels or vasopressor use trends in ventilator dependent patients. acidosis could be non gap metabolic acidosis from hyperchloremia and fluid overload. in addition, either sepsis progression or poor lung functioning may have caused the resulting acidosis. fraction of inspired oxygen (fio 2 ) is the fraction or percentage of oxygen in the volume being measured. it is used to represent the percentage of oxygen participating in gas exchange. according to a study by diniz et al., fio 2 levels sufficient to ensure a spo 2 ≥92% do not alter breathing patterns or trigger clinical changes in weaning patients 28 . the fio 2 level was enough to represent the oxygen status of the ventilated patient. our data revealed that a higher fraction of inspired oxygen demand was associated with greater risk of ventilator dependence in patients with sepsis or septic shock. applying the ventilator dependence risk score to predict prolonged ventilator dependence can help us communicate with the family, enable quick adjustment of the treatment strategy, and ensure more efficient allocation of medical resources. in addition, it is clinically applicable. the score includes two components. one component is uncorrectable, such as previous stroke history; the other component is correctable if treatment is successful, such as thrombocytopenia, acidosis, and fraction of inspired oxygen. we do not suggest correction of thrombocytopenia and acidosis by blood transfusion and bicarbonate use, as there are inherent risks with platelet transfusion and bicarbonate infusion. however, the clinical physician should make the best efforts in correcting underlying progressive sepsis to avoid prolonged ventilator use. we do not routinely use subcutaneous heparin for prophylaxis of deep vein thrombosis or pulmonary embolism in taiwan. therefore, we seldom have heparin induced thrombocytopenia patients. in our study group, we had no patients with sepsis and pulmonary embolism concurrently. however, we should keep the possibility in mind. as some components of our ventilator dependency risk score are similar to sofa values, we tested sofa score for ventilator dependence prediction. we found the area under the curve (auc) of the ventilator dependence risk score (0.725) was better than the sofa score on admission day 1 and day 7. however, 2 components of sofa score (pulmonary subscore: pao 2 /fio 2 and gcs subscore) on admission day 7 were significant for predicting ventilator dependence in univariate analysis (p < 0.001). despite these findings, the pao 2 /fio 2 and gcs auc were not better than the ventilator dependence risk score auc (fig. 3) . in fact, we have previously described an immune dysfunction scoring system for predicting 28-day mortality in septic patients, with better discrimination than sofa score; this system was valid and reproducible. the above cases were from part of the current sepsis cohort, who agreed for immune function assessment 29 . however, in the present study, we are focused on ventilator dependency amongst patients who survive sepsis more than 21 days. combining those 2 tools, we can predict long term ventilator dependence and predict survival. the area under the curve (auc) of the ventilator dependence risk score was 0.725 in our study group and the auc of the ventilator dependence risk score was 0.658 in the validation group. after further analysis of the validation group, we found the auc of ventilator dependence risk score was 0.745 for sepsis with cancer group and the auc was 0.723 for sepsis with chronic kidney disease group. we are actively studying the effect of co-morbidity on the outcomes of patients with sepsis, although it is out of the scope of this study. our previous study revealed that among patients admitted to the icu with sepsis, those with underlying active cancer had higher baseline levels of plasma il-10, higher trend of g-csf and higher mortality rate than those without active cancer 30 . our ventilator dependence risk score could help predict who will need prolonged mechanical ventilation. we did not exclude patients with tuberculosis or severe immunosuppression (human immunodeficiency virus (hiv), oncologic, solid-organ or bone marrow transplantation). our score can also be used for these patients. septic patients admitted to the hospital or the intensive care unit are usually screened for contamination with multi-resistant bacteria and subjected to collection of blood cultures and respiratory secretions. as in our previous study 31 , multi-resistant bacteria or specific pathogens influence survival in patients with ventilator associated pneumonia. the phenomenon was not shown for ventilator dependency 14 . most of our patients came from er (69.3%) and most of our blood culture showed no growth. we suggest that multi-resistant bacteria may not influence prediction of prolonged mechanical ventilation. however, further study may be needed to determine the effect. renal replacement therapy could be a risk factor. however, there was no statistical significance in univariate analysis. in addition, the sofa renal subscores did not differ between ventilator dependent and independent patients. therefore, renal replacement therapy was not used in the scoring system. in 2011, sellares j et al. 32 described that copd, increased heart rate and paco 2 during the spontaneous breathing trial independently predicted prolonged weaning. however, our studied group had small proportion of copd (9.7% in ventilator-dependent group and 12.8% in ventilator-independent group) ( table 2 ). in addition, we did not routinely record heart rate and paco 2 during the spontaneous breathing trial. therefore, paco 2 and heart rate during the spontaneous breathing trial were not included in our scoring model. extubation failure before day 7 may be an additional prognostic parameter for ventilator dependence. however, in our study population, no extubation failure before day 7 was noted. the limitations of the study include the retrospective study design and possible selection bias. however, first, we used prospectively collected data and screened consecutive patients. second, we excluded patients who died within 21 days, which may have masked some predictors associated with both mortality and ventilator dependence. however, mortality prediction was beyond the scope of this study. the application of the score focused on patients who survived sepsis/septic shock with acute respiratory failure on admission day 7. this patient group was not completely recovered and needed further treatment and strategic decision making. from our results, the data from day 7 is enough to calculate the score, which makes it feasible to use for predicting ventilator dependence. patients require mechanical ventilation due to either pulmonary function problems or neurological function problems. in patients with sepsis, both components may co-exist. it is difficult to delineate what proportion of patients requiring prolonged mechanical ventilation is attributed to pulmonary or neurological problems. we did not incorporate any data on the patients' pulmonary system mechanics or respiratory muscle strength (respiratory system compliance or resistance, maximal exhaled tidal volume, negative inspiratory force, rapid shallow breathing index) that are typically studied during weaning from mechanical ventilation 33 . it is partially because of some missing data owing to the retrospective characteristic of the study, which makes it difficult to analyze. most importantly, obtaining parameters like static compliance requires an additional procedure such as paralysis and muscle relaxant, which may add risk to those patients with unstable severe sepsis. for easy application to patients with sepsis and septic shock, we chose to incorporate data easily checked in clinical practice. it is now well known that sepsis and multi organ failure can cause neurological dysfunction by way of critical illness neuropathy and myopathy (i.e., icu acquired weakness), which can cause difficulty weaning from mechanical ventilation due to diaphragmatic weakness. sepsis and multiple organ dysfunction are the most common and well accepted risk factors for icu acquired weakness. some other risk factors like ards, neuromuscular blockade, glucose control, and steroid use are missing from the analysis due to the retrospective study design. those particular entities deserve attention. the diagnosis of icu acquired weakness is often clinical with emg support, which is not often conducted in routine clinical practice. with respect to neurological function, we note a significant difference in the groups with the history of prior stroke. we did not have complete data differentiating hemorrhagic or ischemic strokes. in addition, the functional status or delirium data were also lacking. we attempted to use gcs (the required data are already present within the apache and sofa scores) but the results showed poor discrimination. those issues need to be explored further in the future. a valuable tool to predict which septic patients will need prolonged mechanical ventilation may have not only therapeutic ramifications, but also significant financial and social implications. as shown in table 1 , patients requiring long term mechanical ventilation have significantly longer icu stay and in hospital mortality. it is primarily due to the medical acuity. however, in part, it is also due to a paucity of ventilator weaning facilities. patients who require long term mechanical ventilation are often difficult to place, leading to longer hospital stays than expected for their given illness. we did not discuss diagnosis of ards in this study. the pao 2 /fio 2 were comparable between the two groups. in the same period, our colleagues participated in a multiple center study showing the effects of ards and fluid balance on outcomes. over resuscitation leads to fluid overload and pulmonary edema, and hypoxia, which may influence ventilator dependence. we found a negative day 1-4 cumulative fluid balance was associated with a lower mortality rate in critically ill patients with influenza 34 . we are now exploring whether cumulative fluid balance predicts ventilator dependency. we need to further evaluate an association between over resuscitation and ventilator dependence in the future. ventilator dependence risk score, including a history of stroke and data from day 7 (thrombocytopenia, acidosis, and the higher fraction of inspired oxygen), can be applied to predict prolonged mechanical ventilation in patients who survive sepsis and septic shock. setting and study design. this retrospective study was conducted in three medical icus, including 34 beds at kaohsiung chang gung memorial hospital, a 2,700-bed tertiary teaching hospital in southern taiwan. consecutive adult patients (aged ≥18 years) with acute respiratory failure on admission to the medical icu with sepsis/septic shock were surveyed from august 2013 to october 2015 through chart review. we excluded patients who passed away within 21 days, those whose families requested palliative treatment before day 21, and those who were already long-term mechanical ventilator dependent. the enrolled patients were divided into two groups, i.e., ventilator-independent and ventilator-dependent, according to their ventilator status at the time of ventilator use on day 21 from the chart review (fig. 1) . we also collected data from sepsis/septic shock patients with respiratory failure who were admitted to the medical icu from november 2015 to november 2016 as the validation group (fig. 2) . the study was approved by the institutional review board (irb) of chang gung memorial hospital and the requirements to obtain informed consent from patients were waived by irb (105-6824c). we confirmed that all methods were performed in accordance with the relevant guidelines and regulations. definitions. long-term ventilator dependence in patients was defined as the need for mechanical ventilation for more than six hours per day for more than 21 days 16 . sepsis was defined as a life-threatening organ dysfunction due to a disproportionate host response to infection 35 . patients with septic shock were identified by a vasopressor requirement to maintain a mean arterial pressure of >65 mmhg in the clinical condition 36, 37 . all enrolled patients met the new criteria for sepsis and required mechanical ventilation at the time of admission to the icu. moreover, they survived for at least 21 days after admission to the icu. iv titratable sedation was applied if the patient's condition required and the titration protocol was standardized by medical intensive care unit. the standard clinical practices for weaning the patient from mechanical ventilation were performed during the study period (i.e., pressure support and spontaneous breathing trials). clinical data were retrieved from medical records and included age, gender, sequential organ failure assessment (sofa) score 19 , acute physiological assessment and chronic health evaluation ii (apache ii) score 38 , charlson index and underlying comorbidities 39, 40 , and other clinical factors possibly related to prolonged ventilator use. we also collected hematology, biochemistry, fraction of inspired oxygen (fio 2 ) and pao 2 /fio 2 on admission day 7 to follow up on the patient's condition. all variables were evaluated as possible risk factors of prolonged ventilator use. score construction and calculation. categorical variables were analyzed using the chi-squared test, and continuous variables were compared using the student's t-test. a two-tailed p value of <0.05 was considered to indicate a significant result. univariate analysis was used to identify significant risk factors associated with ventilator dependence. variables associated with ventilator dependence in the univariate analysis (p < 0.1) were included in a multivariate analysis model. using the stepwise method, the independent factors associated with ventilator dependence were identified to build a score using the hosmer-lemeshow goodness-of-fit test. a clinical score (vd risk score) was calculated based on four variables independently associated with ventilator dependence in the multivariate analysis. the number of points assigned to each variable in the vd score was adjusted according to proportion to beta coefficient in the regression model. the vd risk score is the sum of the points for these four variables. the receiver operating characteristic (roc) curve was used to evaluate the performance of the vd risk score. all statistical analysis was performed using the spss 22.0 software package (spss inc., chicago, il, usa). sepsis: pathophysiology and clinical management healthcare cost and utilization project (hcup) statistical briefs epidemiology of severe sepsis in the united states: analysis of incidence, outcome, and associated costs of care management of severe sepsis in patients admitted to asian intensive care units: prospective cohort study assessment of the worldwide burden of critical illness: the intensive care over nations (icon) audit assessment of global incidence and mortality of hospital-treated sepsis. current estimates and limitations population burden of long-term survivorship after severe sepsis in older americans benchmarking the incidence and mortality of severe sepsis in the united states surviving sepsis campaign: international guidelines for management of severe sepsis and septic shock initial resuscitation guided by the surviving sepsis campaign recommendations and early echocardiographic assessment of hemodynamics in intensive care unit septic patients: a pilot study management of patients requiring prolonged mechanical ventilation: report of a namdrc consensus conference weaning from prolonged mechanical ventilation prolonged mechanical ventilation in critically ill patients: epidemiology, outcomes and modelling the potential cost consequences of establishing a regional weaning unit factors predicting ventilator dependence in patients with ventilator-associated pneumonia physiologic determinants of ventilator dependence in long-term mechanically ventilated patients chronic ventilator dependence in elderly patients prediction of prolonged ventilator dependence in children by respiratory function measurements early prediction of prolonged ventilator dependence in thermally injured patients application of the sequential organ failure assessment (sofa) score to bacteremic icu patients weaning failure in mechanical ventilation. 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respiratory icu a prospective study of indexes predicting the outcome of trials of weaning from mechanical ventilation association of day 4 cumulative fluid balance with mortality in critically ill patients with influenza: a multicenter retrospective cohort study in taiwan assessment of clinical criteria for sepsis: for the third international consensus definitions for sepsis and septic shock (sepsis-3) developing a new definition and assessing new clinical criteria for septic shock: for the third international consensus definitions for sepsis and septic shock (sepsis-3) the third international consensus definitions for sepsis and septic shock (sepsis-3) comparison of the sequential organ failure assessment, acute physiology and chronic health evaluation ii scoring system, and trauma and injury severity score method for predicting the outcomes of intensive care unit trauma patients importance of pre-existing co-morbidities for prognosis of septicemia in critically ill patients impact of different measures of comorbid disease on predicted mortality of intensive care unit patients the work was supported in part by a grant from the chang gung medical foundation (chang gung memorial hospital) grant (cmrpg8b1061, cmrpg8b1062, cmrpg8b1063, cmrpg8f1331, and cmrpg8f0821 to w.-f. fang). we would like to thank all the staff and clinicians in the intensive care unit who participated in this study, for their support. we also thank the biostatistics center, kaohsiung chang gung memorial hospital, for statistics consultation. competing interests: the authors declare no competing interests.publisher's note: springer nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations. license, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the creative commons license, and indicate if changes were made. the images or other third party material in this article are included in the article's creative commons license, unless indicated otherwise in a credit line to the material. if material is not included in the article's creative commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. to view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/. key: cord-006426-baf2d47y authors: kimura, fumio; shimizu, hiroaki; yoshidome, hiroyuki; ohtsuka, masayuki; miyazaki, masaru title: immunosuppression following surgical and traumatic injury date: 2010-08-26 journal: surg today doi: 10.1007/s00595-010-4323-z sha: doc_id: 6426 cord_uid: baf2d47y severe sepsis and organ failure are still the major causes of postoperative morbidity and mortality after major hepatobiliary pancreatic surgery. despite recent progress in understanding the immune conditions of abdominal sepsis, the postoperative incidence of septic complications after major visceral surgery remains high. this review focuses on the clinical and immunological parameters that determine the risk of the development and lethal outcome of postoperative septic complication following major surgery and trauma. a review of the literature indicates that surgical and traumatic injury profoundly affects the innate and adaptive immune responses, and that a marked suppression in cell-mediated immunity following an excessive inflammatory response appears to be responsible for the increased susceptibility to subsequent sepsis. the innate and adaptive immune responses are initiated and modulated by pathogen-associated molecular-pattern molecules and by damage-associated molecular-pattern molecules through the pattern-recognition receptors. suppression of cell-mediated immunity may be caused by multifaceted cytokine/inhibitor profiles in the circulation and other compartments of the host, excessive activation and dysregulated recruitment of polymorphonuclear neutrophils, induction of alternatively activated or regulatory macrophages that have anti-inflammatory properties, a shift in the t-helper (th)1/th2 balance toward th2, appearance of regulatory t cells, which are potent suppressors of the innate and adaptive immune system, and lymphocyte apoptosis in patients with sepsis. recent basic and clinical studies have elucidated the functional effects of surgical and traumatic injury on the immune system. the research studies of interest may in future aid in the selection of appropriate therapeutic protocols. severe sepsis is still a major cause of postoperative morbidity and mortality after major abdominal surgery. in particular, aggressive hepatobiliary pancreatic surgery, including an extended liver resection and radical pancreaticoduodectomy, has been associated with high complication rates of 40%-50%. [1] [2] [3] [4] the most common complications following major hepatobiliary pancreatic surgery are septic complications, including cholangitis, wound infection, pneumonia, intra-abdominal abscess, fi stula, and septicemia. [1] [2] [3] [4] a recent clinical trial has reported that although organ-preserving pancreatic surgery potentially decreased the incidence of postoperative delayed gastric emptying in comparison with a pancreatoduodenectomy, there were no signifi cant differences between the two groups in the incidence of pancreatic fi stulas or other complications. 5 despite recent progress in understanding the immune conditions of abdominal sepsis, the postoperative incidence of septic complications after major visceral surgery remains high. [1] [2] [3] [4] during the past decade, a number of experimental and clinical studies have provided evidence that surgical and trauma injury markedly affects the immune system, including both the specifi c and the nonspecifi c immune responses. [6] [7] [8] [9] the protective immunity of the hosts may critically depend on an appropriate cytokine balance, a proper activation and recruitment of polymorphonuclear neutrophils (pmns) and monocytes/macrophages, an intact macrophage-t-cell interaction, and an adequate t-helper (th)1/th2 conception of t-helper cell activation. the surgical and trauma injury potentially disintegrates these complex regulatory systems and induces the deterioration of immune function. [6] [7] [8] [9] this review focuses on the clinical and immunologic parameters that determine the risk of the development and lethal outcome of postoperative septic complication following major surgery and trauma. the elucidation of these mechanisms is a prerequisite subject for the introduction of preventive and therapeutic strategies into clinical practice. major injury due to surgical or major trauma produces potentially profound immunological dysfunction resulting in tissue injury, postoperative infection, and multiple organ dysfunction syndrome (mods) (fig. 1 ). the immune system consists of an early innate and a late adaptive response. the initial proinfl ammatory immune response, or systemic infl ammatory response syndrome (sirs), is mediated primarily by the cells of the innate immune system. this is followed by a compensatory anti-infl ammatory response syndrome (cars) that is primarily mediated by cells of the adaptive immune system. [6] [7] [8] [9] [10] moore and moore have described a model of early and late mods depending on the initial degree of injury severity. 11 an initial massive traumatic insult can create an early vigorous proinfl ammatory response and severe sirs independent of infection ("one-hit" model), resulting in early mods. in the "two-hit" scenario, initially less severely-injured patients eventually develop late mods as a result of the reactivation of their infl am-matory response caused by an adverse and often minor intercurrent event, such as additional surgical stress, bacterial infections, or ischemia/reperfusion injury. late mods is often accompanied by cars. 12 an unbalanced systemic compensatory anti-infl ammatory response can result in anergy and immunosuppression, which predisposes the host to the development of opportunistic infection. cytokines, chemokines, stress hormones, and many other humoral mediators have been implicated in the pathogenesis of sirs, cars, and mods in patients with severe surgical or traumatic injuries. 13, 14 in response to major tissue injury and/or bacterial infection, endothelial and epithelial cells, as well as neutrophils, macrophages, and lymphocytes, produce powerful proinfl ammatory cytokines, especially tumor necrosis factor-α (tnf-α), interleukin (il)-1β, and il-6. 13 polymorphonuclear neutrophils and macrophages respond to many of these mediators by releasing granular enzymes and producing reactive oxygen species (ros) that ultimately lead to organ dysfunction. 14 chemokines have the ability to favor neutrophil and monocyte recruitment to the infl ammatory site and to stimulate their subsequent activation. 15 according to the alignment of the sequences, chemokines can be classifi ed into two families, one with the fi rst two cysteines separated by one residue (cxc chemokines) and the other with the fi rst two cysteines adjacent (cc chemokines). the cxc chemokines are neutrophil chemoattractants and include il-8 and growth-related oncogenes in humans. 15 the cc chemokines are predominantly monocyte chemoattractants, and include macrophage chemoattractant protein (mcp)-1 and macrophage an initial massive insult can create an early vigorous proinfl ammatory response and severe sirs independent of infection (one-hit model), resulting in early mods. in the "two-hit" scenario, initially less severely-injured patients eventually develop late mods as a result of the reactivation of their infl ammatory response caused by additional surgical stress, ischemia/reperfusion injury, or bacterial infections. patients surviving the early proinfl ammatory sirs response to major injury may develop a counterinfl ammatory cars response, which is associated with postoperative infection. adapted from ni choileain and redmond 7 and moore and moore 11 infl ammatory protein (mip)-1α. 15 the postoperative serum levels of il-8 and mcp-1 have been reported to correlate with surgical insult in patients who have undergone cardiovascular surgery. 16 anti-infl ammatory cytokines potentially suppress various innate immune functions and thereby render patients susceptible to postoperative infection. 17, 18 in mouse models the rapid release of endogenous il-10 has been reported to be an essential anti-infl ammatory response controlling cytokine production during both gram-negative and grampositive infection. 19, 20 however, excessive production of il-10 renders patients susceptible to infection. high serum il-10 levels have also been reported to be associated with fatalities among patients with infection. 17 interleukin-10 may also be involved in immune depression associated with hemorrhage. 21 interleukin-4 also inhibits monocyte/macrophage function, including the ability to suppress monocyte-generated cytokines. 22 stress hormones may participate in the pathogenesis of postinjury infection and organ dysfunction. 23 although activation of the hypothalamic-pituitary-adrenal axis is essential for response to severe stress, 24 excessive glucocorticoids may delay wound healing by promoting catabolism and inhibiting the immune system, leading to postoperative immunosuppression and infection. 25 macrophage migration inhibitory factor (mif), which is a proinfl ammatory pituitary and macrophage cytokine and a contraregulator of endogenous glucocorticoids, plays a critical part in the pathogenesis of septic shock. 26 leptin, an adipocyte-derived hormone that acts centrally in hypothalamus to regulate body weight and peripheral energy expenditure, thereby helps to regulate the immune response. 27, 28 in humans, il-6, il-10, il-8, mcp-1, cortisol, and leptin have been reported to be released after a liver resection in response to surgical stress, and are correlated with postoperative infection and organ dysfunction. 29 cytokine antagonists also appear in the circulation, and the soluble tnf receptor p55 correlates with postoperative infection following surgery. 30 in these studies, increased plasma concentrations of proinfl ammatory and anti-infl ammatory mediators were observed immediately after surgery or on postoperative day 1. these elevations occurred simultaneously for both the proand anti-infl ammatory mediators. similar circulating cytokine/inhibitor profi les have been observed in a mouse model of sepsis. 31 this multifaceted response questions the use of a simple proinfl ammatory cytokine measurement for classifying the infl ammatory status of the patients with septic complications. 31 in addition, plasma levels should be carefully interpreted because they do not necessarily refl ect the immune status in other compartments of the host. nevertheless, due to a lack of more clinically useful markers, determination of pro-and anti-infl ammatory cytokine levels in plasma has become more and more important in the intensive care unit when dealing with surgical patients. 6 the pattern-recognition receptors (prrs) play a central role in the initiation of the innate and adaptive immune response to infection (fig. 2 ). membrane-bound or vesicular (endosomal) prrs, including the toll-like receptors (tlrs), nucleotide-binding oligomerization domain (nod)-like receptors (nlrs), and rig-i-like receptors (rlrs) recognize microorganisms derived products, so-called pathogen-associated molecular-pattern molecules (pamps) and activate intracellular cascades. 32, 33 toll-like receptors sense the presence of pamps extracellularly and in phagosomes, whereas the cytosolic nlrs recognize pamps in intracellular compartments. 34 rig-i-like receptors are cytoplasmic proteins that recognize viral rna. 35 in humans, extracellular bacterial products are sensed by fi ve tlrs. lipopolysaccharide (lps) is the main bacterial ligand for tlr4; lipotechoic acid and diacylated lipopeptides are sensed by a tlr2-tlr6 dimer; triacylated lipopeptides are sensed by a tlr2-tlr1 dimer; cpg motifs are sensed by tlr9, and fl agellin is sensed by tlr5. for antifungal responses, a tlr2-tlr6 dimer senses zymosan. five tlrs are involved in antiviral responses: tlr4 senses f protein from respiratory syncytial virus; double-stranded rna is sensed by tlr3; tlr9 senses viral cpg dna; and tlr7 and tlr8 sense single-stranded viral rna. protozoal products such as glycosylphosphatidylinositol-anchor proteins are also sensed by tlr2. therefore, almost all pathogens that infect humans will be sensed by tlrs. 33 toll-like receptor signaling potentially activates nuclear factor-κb (nf-κb) through myeloid differentiation primary-response protein 88 (myd88). myd88 is a tlr signaling adaptor protein that is used by all tlrs except tlr3. it interacts with the il-1 receptor-associated kinase (irak) family, leading to interaction with tumor necrosis factor receptor-associated factor 6 (traf6), which ultimately leads to activation of nf-κb and mitogen-activated protein (map) kinases (fig. 2) . these pathways lead to the production of such cytokines as tnf and other proinfl ammatory proteins. 33 in addition, monocytes and pmns from patients with sepsis have been reported to exhibit signifi cantly higher tlr-2 and tlr-4 expression levels than controls. 36 furthermore, in severely injured patients, the carriage of the variant tlr4 896 g allele has been reported to be associated with a decreased risk of complicated sepsis. 37 the nlr family is divided into subfamilies on the basis of their signal transduction domains (table 1) , and recent studies have highlighted the role of certain nlrs, including nod1, nod2, nalp, and ipaf in the detection of intracellular microbes. 38 nod1 and nod2 can detect muropeptides derived from peptidoglycan (pg). nod2 detects muramyl dipeptide, a motif that is present in the pgs of both gram-positive and gram-negative bacteria, whereas the recognition of bacterial pg by nod1 is dependent on the presence of the meso-dap, an amino acid characteristic of most gram-negative and some gram-positive bacteria. therefore, nod1 is involved in the immune response to escherichia coli, chlamydia pneumoniae, campylobacter jejuni, salmonella typhimurium, pseudomonas aeruginosa, and helicobacter pylori, whereas nod2 recognizes streptococcus pneumoniae, mycobacterium tuberculosis, and listeria monocytogenes. the nod proteins recruit the serinethreonine kinase rip2, which in turn leads to nf-κb activation. 38 other nlrs activate caspase-1, which results in the processing and release of the proinfl ammatory cytokines il-1β and il-18 (fig. 2) . in response to specifi c microbial or bacterial factors, the nlr proteins assemble a caspase-1-activating infl ammasome complex. several infl ammasomes have been defi ned by the nlr protein that they contain: the nalp1 infl ammasome, the nalp3 infl ammasome, and the ipaf infl ammasome [38] [39] [40] (table 1) . gram-positive bacteria such as l. monocytogenes, s. aureus, and gramnegative aeromonas hydrophila activate nalp3 using pore-forming toxins. activated nalp3 recruits apoptosis-associated speck-like protein (asc) through a homophilic pyrin domain interaction. apoptosis-associated speck-like protein in turn recruits procaspase-1 via homophilic caspase recruitment domain (card)-card interaction, which leads to activation of caspase-1, and active caspase-1 can then process pro-il-1β and pro-il-18. 39 ipaf recognizes fl agellin secreted by gramnegative pathogens, and can interact directly with the caspase-1 card domain. 40 nod2 leucine-rich repeat variants have been reported to be closely associated with susceptibility to crohn's disease. 41 recently, the monocyte expression of the infl ammasome mrnas for nalp1, asc, and caspase-1 has been reported to be signifi cantly lower in patients with septic shock compared with critically ill control subjects. 42 furthermore, the nalp1 mrna pathogen-associated molecular-pattern molecules (pamps) and damage-associated molecular-pattern molecules (damps). toll-like receptors (tlrs) sense the extracellular presence of so-called pamps, whereas nucleotide-binding oligomerization domain (nod)-like receptors (nlrs) recognize pamps in intracellular compartments and activate intracellular cascades. nod1, nod2, nalp, and ipaf are included in nlr family. extracellular adenosine triphosphate (atp) potentially contributes to the onset of acute infl ammation through p2 receptors. conversely, extracellular adenosine, a metabolite of atp, induces an anti-infl ammatory response through p1 receptors. hmgb1, high-mobility group box 1; hsps, heat shock proteins; irak, il-1 receptor-associated kinase; traf6, tumor necrosis factor receptor-associated factor 6; nf-κb, nuclear factor κb; rip2, receptor-interacting protein 2 levels are linked to survival in patients with sepsis, thus suggesting that the nalp1 infl ammasome plays a critical role in the pathogenesis of sepsis in humans. however, in that study, the monocyte expression of nod1 and nod2 is not affected by sepsis. pattern-recognition receptors can recognize some endogenous ligands released from damaged tissues and activate immune response of the host (fig. 2) . these endogenous ligands released by damaged or dying cells have been termed damage-associated molecular-pattern molecules (damps) because they contribute to the induction of infl ammation through prrs. 43 damps include the chromatin-associated protein high-mobility group box 1 (hmgb1), heat shock proteins (hsps), s100 proteins, products of purine metabolism (uric acid, adenosine triphosphate [atp], and adenosine), and others. 44 hmgb1, a typical damp, is a ubiquitous nuclear protein that binds to nucleosomes and promotes dna bending. there are two ways in which hmgb1 can be released into the extracellular environment: active secretion from cells of the innate immune system or passive release from necrotic cells. similar to proinfl ammatory cytokines, the active secretion of hmgb1 from monocytes or macrophages follows activation by micro-bial and proinfl ammatory stimuli. 44, 45 in a standardized mouse model of endotoxemia, hmgb1 secreted by activated macrophages acts as a late mediator of infl ammation. the serum hmgb1 levels begin to increase 12-18 h after tnf levels peak, which occurs at 2 h, and after il-1 levels peak, which occurs at 4-6 h. 46 in addition, hmgb1 is passively released from cells undergoing nonprogrammed cell death, and initiates infl ammation. in contrast, apoptotic cells modify their chromatin so that hmgb1 irreversibly binds and is therefore not released. hmgb1 has been reported to transduce cellular signals by interacting with at least three receptors: tlr4, tlr2, and the receptor of advanced glycation endproducts (rage). 44, 45 it has been reported that hmgb1 acts as a mediator of infl ammation and organ damage in experimental models of lps-induced acute lung injury (ali), 47 hemorrhage-induced ali, 48 and hepatic ischemia reperfusion injury. 49 in humans, hmg1 may also be involved in the pathogenesis of sepsis, 50 hemorrhagic shock, 51 and sepsis-induced ali. 52 however, no predictable correlation between serum levels of hmgb1 and severity of infection has been found in patients with sepsis. 53 heat shock proteins are stress-inducible proteins that protect against cellular injury as a molecular chaperone, playing an essential role in mediating protein folding, assembly, transport, and degradation. it has been docu38 martinon, 39 and sutterwala et al. 40 the lrr domains of the nlrs can sense the activating signals. the core structure of the nalp infl ammasome consists of a nalp, the adaptor asc, and caspase-1. the nacht-nad region initiates oligomerization of nalp. pyd-pyd, and card-card homotypic interactions (*) are essential for the recruitment and activation of the adaptor asc and the infl ammatory caspases, respectively. ipaf can interact directly with the caspase-1 card domain asc, apoptosis-associated speck-like protein; card, caspase recruitment domain; ipaf, interleukin-1 converting enzyme (ice) proteaseactivating factor; lrr, leucine-rich repeat; nacht, domain present neuronal apoptosis inhibitory protein; nad, nacht-associated domain; nalp, nacht, lrr, and pyd containing proteins; pyd, pyrin domain mented that extracellular hsps potentially act as an endogenous ligand for the cd14, tlr-4, and md2 complex, which mediates the activation of nf-κb and the synthesis of proinfl ammatory cytokines. 54, 55 furthermore, it has been reported that circulating hsp70 plays a pivotal role in postoperative infl ammatory response after open-heart surgery 56 and after liver resection. 57 therefore, even in the absence of pathogens, disrupted or injured cells recruit innate infl ammatory cells by releasing damps. 43 the purinergic receptors may also play an important role in regulation of immune response to tissue injury. the purinergic receptors determine the variety of effects induced by extracellular atp and adenosine released from injured tissue. 58 two families of purinergic receptors have been defi ned to date, namely the p1 and p2 receptors (fig. 2) . the p1 receptors are subdivided into the a1, a2a, a2b, and a3 receptor subtypes. upon injury or infection, damaged tissue cells release intracellular atp into their extracellular microenvironment. extracellular atp potentially contributes to the onset of acute infl ammation through p2 receptors expressed by neutrophils and macrophages. the activation of the p2x7 receptor by atp triggers the assembly of the nalp3 infl ammasome, thus resulting in il-1β secretion. conversely, extracellular adenosine, a metabolite of atp, contributes to alternative macrophage activation through p1 receptors and induces an anti-infl ammatory response. the activation of the a2 receptor by adenosine leads to an increase in intracellular cyclic adenosine monophosphate (amp), which inhibits the intracellular signaling of proinfl ammatory pathways in immune cells. adenosine inhibits virtually all effector functions of neutrophils, macrophages, and t lymphocytes. 58 most immune cells coexpress both p1 and p2 receptor subtypes, which suggests the dual regulation of cell function through purinergic signaling. furthermore, external atp is quickly converted to adenosine by ectoenzymes. therefore, the nature of the effects induced by extracellular atp and adenosine may shift from immunostimulatory to immunoregulatory, depending on the mechanisms that control atp release, expression of the ectoenzymes, and the availability of the p2 and p1 receptors. 58 a2 adenosine receptors have been reported to potentially be able to counteract collateral tissue damage due to excessive infl ammation. 59 in the infl ammatory tissue, activated immune cells might cause direct tissue damage. in addition, the increased expression of adhesion molecules might result in the augmented binding of neutrophils to the blood vessel walls, leading to vascular occlusions, local tissue hypoxia, and indirect collateral tissue damage. tissue hypoxia is conducive to the accumulation of extracellular adenosine. suffi ciently high extracellular adenosine levels will trigger the maximal activation of high-affi nity a2a and low-affi nity a2b adenosine receptors, reducing excessive infl ammation and collateral tissue damage. 59 it has also been reported that adenosine generation catalyzed by the ecto-apyrase (cd39) and 5′-nucleotidase (cd73) expressed on regulatory t cells (tregs) mediates immune suppression. 60 cd39 degrades atp and adenosine diphosphate (adp) into adenosine monophosphates and cd73 catalyzes the conversion of amp into adenosine. therefore, cd39-mediated removal of the proinfl ammatory atp and its conversion into immunosuppressive adenosine by cd73 represent an additional mechanism by which tregs can suppress immune response. 61 the clinical signifi cance of the purinergic receptors, however, remains to be investigated. polymorphonuclear neutrophils play a central role in the innate immune response as the archetypical phagocytic cells. pmns actively seek out, ingest, and destroy pathogenic microorganisms by means of ros, proteinases, and antimicrobial peptides. 62, 63 ordered recruitment and activation of pmns requires the presence of chemoattractants and the leukocyte adhesion molecules of the integrin family. the β2 integrins (cd11/cd18) in particular are critically involved in fi rm adhesion and migration of pmns. 62 it has been reported that cd11b (the adhesion/complement receptor) expression and ros generation of pmns are increased soon after surgical injury 64, 65 and after traumatic injury. 66, 67 the activation of pmns is closely associated with visceral ischemia, 68 lps absorption, 69 sirs, 65 and postoperative sepsis. 66 polymorphonuclear neutrophils are also activated for increased elastase release following major surgery or trauma. 70, 71 furthermore, spontaneous apoptosis of pmns has been reported to be signifi cantly delayed in patients who had undergone elective surgery. 67 these reports suggest that activation of pmns is induced by surgical or trauma injury according to the severity of tissue damage, and is potentially accelerated by lps and bacterial challenge. the activated functionality and prolonged survival of pmns may represent an appropriate adaptive response to injury to eliminate invading pathogens. in contrast, the presence of activated and apoptosis-resistant pmns may play a major role in the pathogenesis of ali and acute respiratory distress syndrome (ards). 72, 73 moreover, the accumulation of primed pmns into tissues, accompanied with systemic activation of complement and coagulation system, can lead to distant organ damage and mods. 74, 75 acute lung injury, acute respiratory distress syndrome, and pneumonia the two most important chemokines for pmn recruitment are il-8/cxcl8 and growth-related oncogene α (gro-α)/cxcl1. growth-related oncogene α is present in greater concentration in the lung than il-8 in patients with pneumonia and ards. [76] [77] [78] furthermore, il-8 binds to both cxcr1 and cxcr2, whereas gro-α only binds to the cxcr2. 77 therefore, gro-α and cxcr2 can participate in the pathogenesis of ards and pneumonia in humans. in a mouse model of pseudomonas pneumonia, the neutralization of cxcr2 results in a striking increase in mortality, which is associated with a marked decrease in neutrophil recruitment and bacterial clearance. conversely, the site-specifi c transgenic expression of keratinocyte-derived chemokine (kc)/cxcl1 results in enhanced clearance of bacteria after pseudomonas challenge. 79 it has been reported that in major trauma patients, cxcr2 responses are markedly diminished in the pmns of patients who progress to sepsis and pneumonia, but are elevated in pmns from patients who progress to ards. 80, 81 these reports suggest that high cxcr2 activity may correlate with pmn priming and outcomes such as ali and ards (fig. 3) , whereas the suppression of cxcr2 function in infl ammatory environments may impair pmn recruitment to the lung and predispose patients to pneumonia and sepsis. in the nonpulmonary organs, the augmented binding of pmns to blood vessel walls might cause indirect tissue damage (fig. 3) . the activated pmns adhere so strongly to the endothelium of postcapillary venules that they produce vascular occlusions, leading to tissue hypoxia and hypoperfusion. alternatively, pmns primed by circulating infl ammatory factors bind tightly to the endothelium, and are readily activated by chemokines expressed on the endothelial surface in response to an underlying infl ammatory or infective lesion. this untoward activation of pmns results in the release of lytic factors that induce endothelial dysfunction, the opening of intercellular junctions, and increase vascular permeability. 82 therefore, vascular occlusions and endothelial dysfunction due to activated pmns might be the major causes of organ dysfunction in sepsis. however, the role of the effector function of neutrophils in patients with sepsis has been poorly investigated. there are data showing that neutrophil adherence and transmigration are impaired in septic patients. 83 although cxcr1 expression is maintained on neutrophils during severe sepsis, desensitization of g-proteincoupled receptors (gpcrs), caused by the steric hindrance of the receptors due to receptor phosphorylation by gpcrs kinases (grk), might be one of the mechanisms for pmn dysfunction. 84, 85 endogenous mediators produced during sepsis might continually activate circulating neutrophils and induce grk activation, leading to gpcr phosphorylation. 84 desensitization of gpcrs, including cxcr1, may impair chemoattractant-induced tyrosine kinase activity and the subsequent rearrangement of the actin network, therefore compromising the ability of neutrophils from patients with sepsis to migrate. 84, 85 fig. 3. models of tissue damage and organ dysfunction in patients with major injury and sepsis. in nonpulmonary organs, vascular occlusions and endothelial dysfunction due to activated neutrophils and endothelium appear to be the major causes of organ dysfunction. activation of the coagulation and complement system might also be involved in this process. in the lung, infi ltrating neutrophils can induce direct damage resulting in acute lung injury and acute respiratory distress syndrome. accumulation of neutrophils in the lung depends on the chemokines that are produced by alveolar macrophages. suppression of chemokine functions may in turn impair neutrophil recruitment to the lung and predispose the lung to pneumonia and sepsis martins et al. reported that ros generation and phagocytosis of pmns are upregulated in patients with sepsis. 86 in contrast, kaufmann et al. have reported that phagocytosis of zymosan and the associated ros production are signifi cantly decreased in patients with septic shock. 87 these reports suggest that the development of pmn dysfunction depends on sepsis severity. danikas et al. evaluated the impact of the phagocytic activity of pmns on the outcome of patients with severe sepsis, and have reported that a reduced phagocytic activity of pmns during the fi rst 24 h after admission is a negative predictor for survival and that phagocytic activity of pmns is strongly correlated with the expression of cd64, 88 the high-affi nity receptor for igg1 and igg3 expressed by mononuclear phagocytes and activated neutrophils. 89 it has been reported that the phagocytic capacity of immature neutrophils is lower than in mature neutrophils. 90 an increase in immature neutrophils in severe sepsis may therefore undermine the overall phagocytic effi cacy of a host, despite the observed leukocytosis. the mononuclear phagocyte system (mps) displays a remarkable functional diversity, allowing cells to perform multiple defense functions from pathogen elimination by phagocytosis to the induction of antigenspecifi c t-cell responses. therefore, mps plays a central role in the innate immunity and orchestrates the adaptive immunity. 91, 92 monocytes circulating blood monocytes supply peripheral tissues with macrophage and dendritic cell precursors and, in the setting of infection, also directly contribute to immune defense against microbial pathogens. in humans, circulating monocytes are divided into two subsets on the basis of the expression of cd14. cd14highcd16-monocytes, which consist of a majority of circulating monocytes and are often called "infl ammatory monocytes," express high levels of ccr2, and traffi c to sites of microbial infection in response to mcp-1/ccl2 secretion. 91, 92 in contrast, cd14lowcd16+ monocytes, which are called "resident monocytes," express higher amounts of mhc class ii molecules than cd14highcd16-monocytes, high levels of cxcr1, and low levels of ccr2. these cells have been suggested to resemble mature tissue macrophages such as splenic macrophages, kupffer cells, alveolar macrophages, microglia, and osteoclasts. 91, 92 macrophages macrophages are heterogeneous cells and have been broadly classifi ed into two groups according to functional polarization: classically activated macrophages and alternatively activated macrophages. 93 however, recent experimental studies have shown that macrophage activation is plastic, rapid, and fully reversible in response to environmental cues, and that there might be at least three macrophage populations based on different physiological activities [92] [93] [94] (table 2) . classically activated macrophages arise in response to interferon-γ (ifn-γ), which can be produced during an adaptive immune response by th1 cells or during an innate immune response by natural killer (nk) cells; and in response to tnf, which is produced by antigenpresenting cells. classically activated macrophages have microbicidal activity, produce high levels of il-12, modest levels of il-10, and release reactive oxygen and nitrogen intermediates. [92] [93] [94] wound-healing (alternatively activated) macrophages arise in response to il-4, which can be produced during an adaptive immune response by th2 cells or during an innate immune response by granulocytes. wound-healing macrophages produce low levels of il-12 and il-10, and are involved in tissue repair. [92] [93] [94] the third macrophage population is adapted from gordon and taylor, 92 benoit et al., 93 and mosser and edwards 94 ifn-γ, interferon-γ; th, t helper; nk, natural killer; apcs, antigen-presenting cells; lps, lipopolysaccharide; ros, reactive oxygen species; tregs, regulatory t cells; gpcr, g-protein-coupled receptor; tgf-β, transforming growth factor-β regulatory macrophages, which are generated in response to various stimuli including immune complexes, prostaglandins, gpcr ligands, glucocorticoids, apoptotic cells, adenosine, or il-10. regulatory macrophages produce high levels of il-10 and low levels of il-12 to suppress immune responses. [92] [93] [94] [95] regulatory macrophages also produce transforming growth factorβ (tgf-β) and prostaglandin e2, and show reduced expression of mhc class ii molecules. following surgical or traumatic injury, these macrophage populations may be highly dynamic. classically activated macrophages and regulatory macrophages can fi rst take part in sirs and cars, respectively, and then wound-healing macrophages participate in the resolution of infl ammation and tissue repair. 92-94 in patients with infection following surgical, trauma, or burn injury, circulating monocytes show reduced production of il-12 96, 97 and suppressed expression of human leukocyte antigen (hla)-dr. [98] [99] [100] [101] depressed il-12producing activity by monocytes correlates with an adverse clinical course in severely injured trauma patients. 96 also, preoperative impaired monocyte il-12 production has been observed in patients with the lethal outcome of postoperative sepsis. 97 expression of hla-dr on circulating blood monocytes has been shown to be depressed in patients following trauma, 98 major surgery, 99,100 and burn injury. 101 in these reports, the suppression of hla-dr expression correlates with severity of infectious complication and poor outcome. [98] [99] [100] [101] furthermore, a reduction in hla-dr expression rate is a sensitive indicator of poor outcome in cases of sepsis, severe sepsis, or septic shock. 102, 103 these reports suggest that monocyte deactivation such as suppression of il-12 production and hla-dr expression is closely related to immuno-paralysis following major injury and in severe sepsis. in patients with sepsis, a reduction of monocyte hla-dr expression is inversely affected by serum il-10 levels or il-10 mrna expression in peripheral leukocytes. [104] [105] [106] interleukin-10 has been shown in vitro to potentially suppress the hla-dr expression on human monocytes 107,108 and il-12 production by human monocyte at the transcriptional level. 109 moreover, circulating il-10 levels are a remarkable predictor of postoperative infection and fatal outcome of sepsis. 106 therefore, il-10 plays a major role in the pathophysiological mechanism of monocyte deactivation following major injury and in sepsis. transforming growth factor-β has also been described in vitro to play a role in monocyte deactivation. 110, 111 however, the circulating tgf-β level is not observed to increase in patients with sepsis. it tends to either be lower in severe sepsis 105, 112 or is not signifi cantly different from controls. 103, 113 therefore, il-10 is likely more important than tgf-β regarding the pathophysiology of monocyte deactivation following major injury and in severe sepsis. toll-like receptor signaling might be another key factor in monocyte deactivation during severe sepsis. it has been reported that tlr-2 and -4 expression levels are signifi cantly increased in monocytes from both septic and surgical patients. 114, 115 continuous microbial stimulation during bacterial sepsis with a number of different antigenic structures might result in cell activation, inducing receptor upregulation. 116 causative factors may include the release of cytokines during sepsis, such as il-6, which has been shown to upregulate tlr4 on human monocytes. 117 however, tsujimoto et al. have reported that despite increased expression of tlrs, il-1β production from lps-stimulated peripheral blood mononuclear cells (pbmcs) is signifi cantly reduced in patients with sepsis compared to surgical and control patients. 114 recently, salomao et al. have revealed that tlr signaling gene expression in mononuclear cells is dynamically modulated across the stages of sepsis, and is decreased in more severe forms of the disease. in contrast, broad gene upregulation is present throughout the stages of the disease in pmns. 118 the phagocytic function and ros generation of monocytes in sepsis has been poorly investigated. it has been reported that increased phagocytic function of monocytes, as estimated by expression of cd64 antigen, was favorably correlated to patient survival. 88 reactive oxygen species generation is upregulated in monocytes from septic patients and it is differentially modulated depending on the stage of the disease and the stimuli. 119 moreover, ros generation of monocytes signifi cantly correlates with sepsis-associated organ failure assessment score in patients with severe sepsis and septic shock. 119 these reports suggest that early in the disease process, increased phagocytic function and a vigorous ros generation may be desirable and important to restrain the infecting microorganisms, and that later in this process the persistence of increased ros generation may be deleterious, promoting sepsis-associated organ failure. this upregulated ros generation of monocytes contrasts with monocytes dysfunction such as the downregulation of cytokine production and hla-dr expression. it has been reported that antimicrobial peptides such as defensins α and lps amplify ros release in a tlr4-independent manner, possibly by exerting a prolonged catalytic effect on the ros generating enzymes, whereas antimicrobial peptides inhibit cytokine and nitric oxide (no) induction by lps in a tlr4-dependent manner. 120 nitric oxide has well-known vasodilatory effects in sepsis, and has pro-and anti-infl ammatory and oxidant and antioxidant properties. 121 previous experimental studies support a role for inducible no synthase (inos) in the pathogenesis of severe sepsis. 121 the vasodilatory effect of no is clearly involved in the development of hypotension during septic shock. nevertheless, nos inhibition in animal models and septic shock patients could not improve and even aggravated the patient outcome, suggesting a bivalent role for no. although excessive no production provokes lethal hypotension, it also has an important antioxidant function, protecting organs from oxidative stress and lipid peroxidation. 122 it has been accepted that macrophages and other hematopoietic cells such as pmns are the principal source of high systemic no levels during septic shock. however, it has been shown that a wide range of nonhematopoietic cells such as hepatocytes, epithelial, vascular smooth muscle, and endothelial cells have the ability to express inos in response to lps or cytokines in rodents. 123 in humans, there is far less evidence for increased nos induction during sepsis. plasma nitrite/nitrate concentrations increase during sepsis, and inversely correlate with mean arterial pressure and systemic vascular resistance. 124 serum nitrite/nitrate concentrations are also increased in patients with postoperative sepsis. 125 the increased serum nitrite/nitrate levels that are found during postoperative sepsis correlate with the severity of the septic course. 126 therefore, inos and no might play a role in the pathophysiology of patients with sepsis or septic shock. the cell sources of no in human sepsis remain unclear. in patients with ards following sepsis, signifi cant expression of inos has been shown in alveolar macrophages, and nitrites/nitrates are elevated in the supernatant of bronchoalveolar lavage fl uid. 127 the frequency of inos expression in pmns is observed to increase in sepsis and sirs patients compared to non-sirs patients. 128 these reports suggest that macrophages and pmns are the principal source of no during sepsis. however, it has been reported that although human mononuclear phagocytes can produce inos mrna and protein in vitro, their abilities to generate no are very low. 129 in patients with septic shock, inos activity is increased in putrescent areas, but is compartmentalized at the very site of infection. 124 these reports suggest that in humans, no synthesis is more restricted than in other species. early studies have reported that the mitogenic response of circulating lymphocytes to the t-cell mitogen phytohemagglutinin and/or concanavalin a is potentially altered by major surgery, multiple trauma, and thermal injury. [130] [131] [132] [133] in these reports, the degree of lymphocyte suppression correlates with the complexity of surgery 130 or severity of injury. 133 in addition, the degree of lymphocyte suppression correlates with the subsequent development of infectious complications and mortality. [131] [132] [133] these reports suggest that major surgical injury can lead to depression of the mitogenic response of lymphocytes, resulting in subsequent development of infectious complications. the th1/th2 balance hypothesis, which was fi rst described in the late 1980s, 134 has since been applied to human immunity and has become a major focus of the attempt to clarify the pathophysiology underlying the postsurgical and post-traumatic immune response. [135] [136] [137] currently, much of the literature elevates the th1/th2 balance concept to the level of a paradigm. 138 uncommitted (naïve) cd4+ t-helper cells (th0) can be induced to differentiate toward th1 and th2 phenotypes depending on the local cytokine milieu (fig. 4) . the presence of interleukin il-12 skews toward th1, while il-4 tends toward th2. the differentiation processes of th0 to th1 or th2 effector cells require the action of two opposing transcription factors, t-bet and gata-3, respectively. t-bet is essential for the development of th1 cells, and gata-3 performs an equivalent role in th2 development. 139 th1 cells drive the cellular immunity against viruses and other intracellular pathogens, eliminate cancerous cells, and stimulate delayedtype hypersensitivity skin reactions, whereas th2 cells drive the humoral immunity and upregulate antibody production against extracellular organisms. it has been reported that after major injury, th1 response is suppressed as illustrated by diminished il-2, ifn-γ, and il-12 levels, while the enhancement of the th2 response is marked by elevated il-10 and il-4. 140 decker et al. have reported that in pbmcs derived from patients undergoing cholecystectomy, ifn-γ secretion, the index cytokine of th1 cells, is increased, while il-4 production, the index cytokine of th2 cells, is decreased following surgery. 135 heidecke et al. also reported that in patients with lethal intra-abdominal infection following surgery, t-cell proliferation and il-2 and tnf production are severely suppressed, thus correlating with sepsis mortality, while t-cell production of il-4 and il-10 is not affected by postoperative intraabdominal infections. 136 furthermore, zedler et al. reported that in pbmcs from patients with major burn injuries, the production of il-4 is excessively upregulated whereas the levels of ifn-γ are only slightly increased. 137 these reports suggest that surgical or burn injury potentially induce a shift in the th1/th2 balance toward th2, and that the suppression of t-cell effector functions may defi ne a state of impaired defense against pathogens and increase susceptibility to infection and septic complications. these alterations of t-cell response following injury have been explained at least in part by monocytederived cytokine il-12. 141, 142 hensler et al. reported a prospective study of 184 patients undergoing major elective surgery of the upper and lower gastrointestinal tract, and estimated a critical role for il-12 in human sepsis. 143 in that study, monocyte il-12 production was severely and selectively impaired in patients developing postoperative sepsis in contrast to patients showing uneventful recovery. moreover, the extent of monocyte il-12 suppression correlated with the severity of postoperative sepsis. major trauma also resulted in early and marked decrease in monocyte cytokine-producing activity. 96 furthermore, the degree of depressed capacity of monocyte il-12 production was statistically and significantly correlated with the development of adult respiratory distress syndrome, sepsis, or infections. these reports suggest that depression in il-12 production by the mps potentially promotes t-cell commitment toward a th2 pattern resulting in postinjury septic complications, and that il-12 is a potent immunoregulatory cytokine that is essential for the development of protective immunity. tregs and th17 cells also seem to modulate the host immune response following injury. [144] [145] [146] th0 can differentiate not only toward th1 and th2, but also toward the th17 and treg phenotypes on the basis of the cytokine environment. the presence of il-10 and tgf-β promotes skewing toward treg, and tgf-β in the presence of il-6 or il21 promotes skewing toward th17. treg and th17 are characterized by the expression of specifi c transcription factors: forkhead box p3 (foxp3) for tregs, and rorγt for th17 cells (fig. 4) . tregs are potent suppressors of the adaptive immune system, and th17 cells produce a strong proinfl ammatory response. the skewing of murine th0 toward th17 and treg is mutually antagonistic. in humans, however, there is no direct evidence for the existence of mutually exclusive development of th17 cells and tregs. [144] [145] [146] tregs have been reported to play a role in the suppression of immune reactions in patients with chronic infl ammation or viral infection. in patients with crohn's disease, foxp3(+)cd4(+) treg cells are expanded in mucosal lymphoid tissues and accumulate in areas of active infl ammation, including granulomas and retain potent regulatory activity ex vivo. 147 circulating and liver resident cd4+cd25+ tregs actively infl uence the antiviral immune response and disease progression in patients with hepatitis b. 148 macconmara et al. reported for the fi rst time that in trauma patients, increased cd4+ cd25+ tregs activity depresses protective th1 cytokine production. 149 furthermore, it has been reported that human cd4+cd25+foxp3+ tregs can induce alternative activation of monocytes/macrophages, which have strong anti-infl ammatory potential involved in immune regulation, tissue remodeling, and tumor promotion. 150 therefore, it is presumed that tregs are potent suppressors of the innate and adaptive immune system, and play a central role in the pathogenesis of immunosuppression following surgical injury and trauma. t-bet, gata-3, rorγt, and foxp3 are special transcription factors of th1, th2, th17, and tregs, respectively. interleukin (il)-12 enhances the expression of t-bet and promotes development of th1 cells, which secrete interferon-γ (ifnγ) and mediate immunity to intracellular pathogens. il-4 enhances expression of gata-3 and promotes the development of th2 cells, which secrete il-4, mediate immunity to helminths, and enhance allergy. transforming growth factor β (tgfβ) with il-6 or il-21 enhances expression of rorγt and promotes development of th17 cells, which secrete il-17, mediate immunity to extracellular pathogens, and enhance autoimmunity. il-10 and tgfβ promote induction of regulatory t cells (tregs), which express foxp3 and suppress immune responses. adapted from mills 145 many reports have provided convincing evidence that il-17-producing t cells have been implicated in the pathogenesis of experimental and human autoimmune diseases, allograft rejection, and chronic infl ammatory conditions. 144, [151] [152] [153] [154] th17 cells function with th1 cells to control immunity to bacteria. the major function of th17 cells is to promote chemokine and proinfl ammatory cytokine production, and the subsequent recruitment and activation of neutrophils and macrophages. 145 emerging data have suggested that in contrast to th1 and th2 cells, which protect against intracellular bacteria and helminths, th17 plays an essential role in the host defense against extracellular bacteria and fungi. 146 however, the role of th17 and il-17 in the immune dysfunction following injury still remains poorly understood. 155 in patients who died of sepsis and multiple organ dysfunction, caspase-3-mediated apoptosis has been reported to cause extensive lymphocyte apoptosis, thus contributing to an impaired immune response. 156 in addition, studies have reported that prolonged lymphopenia and apoptosis-associated depletion of lymphoid organs are involved in nosocomial sepsis-related death in critically ill children. 157 furthermore, lymphocyte apoptosis is increased in cd4 and cd8 t cells, b cells (cd20), and nk cells (cd56) in septic patients compared to nonseptic patients. 158 the authors also demonstrated that apoptotic lymphocytes are positive for activated caspases 8 and 9, consistent with cell death occurring by both mitochondrial-mediated and receptor-mediated pathways. these reports suggest that severe infection can induce apoptosis in a broad range of lymphocyte subsets, and that both of the intrinsic/ mitochondrial and extrinsic/death receptor-mediated pathways may contribute to the immune hyporesponsiveness that is seen in septic patients. 159, 160 conclusions a review of the literature indicates that surgical and trauma injury profoundly affects the innate and adaptive immune responses, and that marked suppression in cell-mediated immunity following an excessive infl ammatory response appears to be responsible for the increased susceptibility to subsequent sepsis. the innate and adaptive immune responses are initiated and modulated not only by pamps, but also by damps through prrs. the suppression of cell-mediated immunity may be caused by multifaceted cytokine/inhibitor profi les in the circulation and other compartments of the host, excessive activation and dysregulated recruitment of pmns, induction of alternatively activated or regulatory macrophages that have anti-infl ammatory properties, a shift in the th1/th2 balance toward th2, appearance of tregs which are potent suppressors of the innate and adaptive immune system, and lymphocyte apoptosis in patients with sepsis. recent basic and clinical studies have explored the functional effects of surgical and traumatic injury on the immune system. future studies will likely contribute further valuable information that will 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hepatitis b increased cd4+ cd25+ t regulatory cell activity in trauma patients depresses protective th1 immunity cd4+cd25+foxp3+ regulatory t cells induce alternative activation of human monocytes/macrophages the dynamics of effector t cells and foxp3+ regulatory t cells in the promotion and regulation of autoimmune encephalomyelitis interleukin-22, a t(h)17 cytokine, mediates il-23-induced dermal infl ammation and acanthosis evidence for the early involvement of interleukin 17 in human and experimental renal allograft rejection increased expression of interleukin 17 in infl ammatory bowel disease systemic il-17 after severe injuries sepsis-induced apoptosis causes progressive profound depletion of b and cd4+ t lymphocytes in humans prolonged lymphopenia, lymphoid depletion, and hypoprolactinemia in children with nosocomial sepsis and multiple organ failure accelerated lymphocyte death in sepsis occurs by both the death receptor and mitochondrial pathways leukocyte apoptosis and its signifi cance in sepsis and shock apoptosis and caspases regulate death and infl ammation in sepsis key: cord-005569-9d51l6bn authors: antonelli, massimo; azoulay, elie; bonten, marc; chastre, jean; citerio, giuseppe; conti, giorgio; de backer, daniel; lemaire, françois; gerlach, herwig; groeneveld, johan; hedenstierna, goran; macrae, duncan; mancebo, jordi; maggiore, salvatore m.; mebazaa, alexandre; metnitz, philipp; pugin, jerôme; wernerman, jan; zhang, haibo title: year in review in intensive care medicine, 2008: i. brain injury and neurology, renal failure and endocrinology, metabolism and nutrition, sepsis, infections and pneumonia date: 2008-12-09 journal: intensive care med doi: 10.1007/s00134-008-1371-6 sha: doc_id: 5569 cord_uid: 9d51l6bn nan recent experimental evidence suggests that matrix metalloproteinases (mmps) are implicated in the pathophysiology of traumatic brain injury (tbi) by increasing blood-brain barrier permeability and exacerbating posttraumatic edema. mmps are zinc-dependent and cell surface-associated endopeptidases that cleave all extracellular matrix (ecm) components, including collagen, laminin, and proteoglycans. the mmps and their potential deleterious effects are tightly regulated at transcriptional and post-transcriptional levels through proform activation and by mmp tissue inhibitors (timps) two members of this family have a very specific and marked activity against gelatin and are termed gelatinases. in vitro studies have demonstrated that the secretion of gelatinases is significantly increased in cortical cultures when mechanical injury is simulated. in human tbi, data about the presence of gelatinases in the brain extracellular fluid (ecf) and their temporal profile, both in plasma and ecf, are still lacking. sahuquillo's group [1] examined the acute profile of mmp-2 and mmp-9 in the plasma of patients with moderate or severe tbi and, more interestingly, in the brain extracellular fluid (ecf). high levels of gelatinases were found in plasma and brain ecf in the early phase of tbi, indicating that both local and systemic trauma-induced upregulation of gelatinases in the acute phase might play an important role in the pathophysiology of tbi and could be a future therapeutic target. after admission in intensive care units (icu), clinicians are often challenged with the contemporary management of intracranial priorities and extra-cranial complications. the incidence and severity of non-neurological organ dysfunction in acute neurological patients has been studied analyzing the database from the observational sepsis occurrence in acutely ill patients (soap) study in 198 icu [2] . the data comes from 373 cases, 12% of the 3,147 patients in the soap database, admitted with a neurological diagnosis, 41% with traumatic brain injury and 50% with cerebrovascular accident. neurological patients developed icu-acquired sepsis and respiratory failure more frequently than the other patients, and length of stay, icu and hospital mortality were higher compared with non-neurological patients. multivariate logistic analysis showed that cardiovascular failure, hepatic failure, and ali/ards were factors independently associated with a higher risk of death in the icu. one of the main intracranial therapy targets in the acute phase is high intracranial pressure (hicp). mauritz [3] described icp use in a sample of 1,856 severe tbi from a registry of patients admitted to austrian icus between 1998 and 2004. mauritz aimed to investigate reasons why patients did or did not receive icp monitoring and to describe factors influencing hospital mortality after severe tbi. the ad hoc created statistical model explains only 37% of the variance of the use of icp monitoring, but clearly showed that severity and age are importantly associated with icp monitoring. the more severe, as the less injured, cases were less likely to be submitted to icp. that is likely to reflect a clinical judgment concerning who can benefit from invasive monitoring. an evaluation of potential salvageability is part of the decisional analysis. interestingly, no clear, standardized definition of ''salvageability'' is available. older patients are less often monitored. the second aim of the paper was to analyze the impact of several predictors, including icp monitoring, on mortality at hospital discharge. the study confirms a clear association between the number of cases treated by center per year and better outcome. this supports the benefit of centralization, as shown in tbi and other neurological non-traumatic pathologies. finally, an additional result was that the subgroup with the highest rate of icp monitoring had the lowest mortality suggesting a utility of monitoring and treating high icp. due to potential complication icp related, even if minor in good hands, non-invasive methods of icp measuring are appealing. between others, the relationship between optic nerve sheath diameter (onsd) and icp in neurocritical care patients has been explored [4] . a significant relationship between onsd and icp was recorded. changes in icp were strongly correlated with changes in onsd. enlarged onsd was a suitable predictor of elevated icp. even if the studied cohort was limited, non-invasive measurements were correlated with invasive icp. this method, once validated in a wider number of patients, could be used as a screening test when raised icp is suspected. stocchetti [5] quantified, in a prospective study of 407 consecutive tbi patients, the occurrence of hicp refractory to conventional medical therapy. in this tbi subset more aggressive therapies, as profound hyperventilation, barbiturates, decompressive craniectomy, are currently used. hicp is frequent; 153 patients had at least 1 day of icp [20 mmhg . early surgery was necessary for 221 cases, and standard medical therapy [sedation, mannitol, cerebrospinal fluid (csf) withdrawal, paco 2 [30] [31] [32] [33] [34] [35] was used in 135 patients. reinforced treatment (paco 2 25-29 mmhg, induced arterial hypertension, muscle relaxants) was used in 179 cases (44%), and second-tier therapies in 80 (20%). surgical decompression and/or barbiturates were used in 28 of 407 cases (7%). six-month outcome was favorable (good recovery or moderate disability) in 195 cases (53%). hicp was associated with worse outcome. outcome for cases who had received second-tier therapies was significantly worse (43% favorable at 6 months, p = 0.03). therefore, hicp is frequent and is associated with worse outcome. the indications for surgical decompression and/or barbiturates seem restricted to less than 10% of severe tbi. the use of hyperventilation and the adherence to international guidelines after tbi have been studied [6] analyzing data coming from 22 european centers, participating in the brainit initiative (http://www.brainit.org). one hundred and fifty-one patients and 7,703 abgs, representing 2,269 ventilation episodes (ve), were included in the analysis. patients without elevated intracranial pressure (icp \20 mmhg) manifested a statistically significant higher paco 2 (36 ± 5.7 mmhg) in comparison to patients with elevated icp (c20 mmhg (34 ± 5.4 mmhg). intensified forced hyperventilation (paco 2 b 25 mmhg) in the absence of elevated icp was found in only 49 ve (2%). early prophylactic hyperventilation was used in 1,224 ve (54%). during forced hyperventilation simultaneous monitoring of cerebral oxygenation was used in only 9%. overall adherence to current guidelines seems common, but early prophylactic hyperventilation and the use of additional cerebral oxygenation monitoring during forced hyperventilation are not followed exposing the patient to potential iatrogenic complications. after the acute phase, hormonal disturbances have been described both in tbi and in subarachnoid hemorrhage patients. maiya [7] retrospectively studied acute anatomical changes in the pituitary gland in 41 tbi patients undergoing magnetic resonance imaging (mri) during the acute phase. mri scans from 43 normal healthy volunteers were used as controls. the pituitary glands were significantly enlarged in the tbi group. twelve of the 41 cases (30%) demonstrated focal changes. in approximately 30% of patients acute tbi was associated with pituitary gland enlargement with specific lesions. mri of the pituitary may provide useful information about the mechanisms involved in post-traumatic hypopituitarism. the neuro-intensive care and emergency medicine (nicem) section of the european society of intensive care medicine (esicm) developed a document on neuromonitoring in neuro-intensive care [8] . the questions discussed and addressed in this manuscript were: (1) who should have icp monitoring and for how long? (2) what icp technologies are available and what are their relative advantages/disadvantages? (3) should cpp monitoring and autoregulation testing be used? (4) when should brain tissue oxygen tension [pbro(2)] be monitored? (5) should structurally normal or abnormal tissue be monitored with pbro(2)? (6) should microdialysis be considered in complex cases? this articulated paper provide useful information to clinicians working in nicu and also to those developing specialist nicu services within their hospital practice. the paper by schlenk [9] in patients with subarachnoid hemorrhage suggests, as indicated by strong [10] in the accompanying editorial, that great caution is needed in choice of a target range for plasma glucose if tight glycemic control with insulin is undertaken. the authors in a prospective, nonrandomized, single-center study, explored whether hyperglycemia exerts deleterious effects via cerebral energy metabolism and the effects of cerebral high/low glucose in patients with aneurysmal subarachnoid hemorrhage. in all patients a microdialysis catheter was inserted. cerebral low-glucose episodes and high-glucose episodes occurred independently of blood glucose levels. during high-glucose episodes cerebral microdialysate levels were normal, while cerebral low glucose, occurring more frequently in symptomatic patients, was associated with severe cellular distress, i.e., increase in lactate/pyruvate ratio, glutamate, glycerol and with unfavorable outcome. cerebral low glucose was associated with severe metabolic distress and may present a target for therapy to improve clinical outcome. tight glycemic control could be deleterious in this neurological population. electrical activity in icu/epilepsy legriel [11] studied 140 icu patients with status epilepticus (se). median seizure time was 60 min and 58 patients had seizures longer than 30 min. the most common causes of se were cerebral insult in 53% and anticonvulsant drug withdrawal in 20% of patients. no cause was identified in 35% of patients. median time from se to treatment was 5. the se was refractory in 25% patients. hospital mortality was 21%. by multivariate analysis, independent predictors of 30-day mortality were age, gcs at scene, continuous se, symptomatic se and refractory se. further studies are needed to evaluate the possible impact of early maximal anticonvulsant treatment on outcomes. an increasing interest is rising re-evaluating the utility of monitoring cerebral electrical activity. this is done, usually, with continuous eeg system even if other, simpler, approaches are explored. walsh [12] prospectively assessed whether the entropy module, a device to measure hypnosis in anesthesia, is a valid measure of sedation state in critically ill patients. four hundred and seventyfive trained observer assessments were made and compared with concurrent entropy numbers. entropy of the frontal eeg does not discriminate sedation state adequately for clinical use in icu patients. morandi [13] and coworkers undertook a multinational effort to identify conflicts in terminology and phenomenology of delirium to facilitate communication across medical disciplines and languages. the evaluation of the terminology used for acute brain dysfunction was determined conducting communications with 24 authors from academic communities throughout countries/regions that speak the 13 variants of the romanic languages included into this manuscript. interestingly only 54% use the term delirium to indicate the disorder as defined by the dsm-iv as an acute change in mental status, inattention, disorganized thinking and altered level of consciousness. attempts towards standardization in terminology, or at least awareness of differences across languages and specialties, will help cross-talk among clinicians and researchers. cheung [14] surveyed the same topic in canadian icus with a response rate of 58.3%. when an etiological cognitive dysfunction diagnosis was obvious, 83-85% responded with the medical diagnosis to explain the cognitive abnormalities; only 43-55% used the term ''delirium''. in contrast, where an underlying medical problem was lacking, 74% of respondents diagnosed ''delirium''. non-pharmacological and pharmacological management varied considerably. commonly selected pharmacological agents were antipsychotics and benzodiazepines, followed by narcotics, non-narcotic analgesics, and other sedatives. canadian intensivists diagnose delirium based upon the presence or absence of an obvious medical etiology. looking for delirium scoring methods, plaschke [15] assessed the agreement between the delirium ratings of two independent delirium assessment methods: the confusion assessment method for the icu and the intensive care delirium screening checklist (icdsc). after excluding permanently unconscious patients with b4 on the richmond agitation sedations scale (rass), delirium was identified in 71 of the 174 patients (41%). the patients who were included were tested in 374 paired but researcher-independent ratings of delirium by both scoring methods. the kappa coefficient determined over 7 days of icu stay was 0.80, indicating good agreement. soja [16] implemented delirium monitoring, test reliability, and monitor compliance of performing the confusion assessment method for the intensive care unit (cam-icu) in trauma patients. following a webbased teaching module, bedside nurses evaluated patients daily for depth of sedation with the rass and for the presence of delirium with the cam-icu. on randomly assigned days, evaluations by nursing staff were followed by evaluations by an expert evaluator to assess compliance and reliability of the cam-icu in trauma patients. overall agreement (kappa) between nurses and expert evaluator was 0.77. the survey revealed that nurses were confident in performing the cam-icu, realized the importance of delirium, and were satisfied with the training that they received. defining the etiology of hyponatremia in acute neurological patients is a sometime an intriguing challenge. brimioulle [17] assessed whether hyponatremia is associated with the syndrome of inappropriate antidiuretic hormone secretion (siadh) or with cerebral salt-wasting syndrome (csws). measurement of blood, plasma, and red blood cell volumes to discriminate siadh and csws. renal, adrenal and thyroid functions were normal in all patients. average blood, plasma, and red blood cell volumes were 54, 37 and 17 ml/kg in control patients and 54, 37 and 18 ml/kg in hyponatremic patients, respectively. the adequate blood volumes in hyponatremic patients confirmed the diagnosis of siadh and did not support the concept of csws. hausfater [18] studied the effect of non-exertional heatstroke on serum procalcitonin (pct) levels in 53 patients with defined heatstroke during the august 2003 heat wave in france were analyzed; 30-day mortality was recorded. among the 53 patients included, 14 (26%) were admitted to an intensive care unit (icu). at 30 days, 24 patients (45%) had died. median pct value was 0.58 lg/l and 31 (58%) patients had pct above 0.2 lg/l (pct?). temperature above or equal to 40°c was the only variable significantly associated with fatal outcome. median pct values were similar between survivors and non-survivors (p = 0.22). high serum pct levels was observed in heatstroke without concomitant bacterial infection. the pct was not a valid mortality predictor in heatstroke, but could be an indicator of the severity of illness. the use of a single dose of etomidate to facilitate laryngeal intubation is still widespread in spite of the demonstration of the potential of the drug to inhibit the last step in cortisol synthesis in the adrenal cortex. the recent corticus study also shows the in vivo capability of the drug to suppress acth-cortisol responses, at least in septic patients. the clinical consequences, however, remain unclear, in spite of awareness of the problem for decades. the paper by vinclair et al. [19] somewhat adds to the evidence in describing the in vivo effects of etomidate in non-septic patients in the course of time, in terms of ratios of circulating 11b-deoxycortiol to cortisol, reflecting in vivo activity of the last enzymatic step in cortisol synthesis potentially inhibited by etomidate. the results suggest, in the absence of a control group, that the ratio increases after etomidate administration and is associated with a diminished rise in circulating cortisol upon acth. the etomidate effect has waned in 91% of patients after 48 h. again, the clinical consequences remain unclear. adrenal insufficiency has been recently observed in critically ill patients with liver cirrhosis and severe sepsis. du cheyron et al. [20] thus postulated that hyperreninemic hypoaldosteronism may be common and may have an impact on outcome in patients with acute on chronic liver failure. the authors thus investigated the relation between the adrenal production of gluco-and mineralo-corticoids, inflammatory status and outcome in critically ill patients with liver cirrhosis. they included 50 consecutive patients with liver cirrhosis and applied a corticotropin stimulation test within 12 h following icu admission. hyperreninemic hypoaldosteronism syndrome was defined as basal renin over aldosterone ratio (rra) higher than 2. they found impaired adrenal function in 41 (82%) patients. 26 patients (52%) presented with an rra [2. patients with rra [2 exhibited greater disease severity and organ dysfunction scores at baseline, but risk-adjusted mortality rates were not different between the two groups. renin and il-6 plasma concentrations were positively correlated. a cox regression analysis revealed hyperreninemic hypoaldosteronism syndrome, il-6 higher than 400 pg/ml and severe renal failure to be independent predictors of 30-day mortality. the authors concluded that adrenal dysfunction was common in critically ill cirrhotic patients. hyperreninemic hypoaldosteronism syndrome was related to a greater pro-inflammatory status and degree of acute organ failure, and was independently associated with a worse prognosis. in attempts to prevent harmful acute kidney injury (aki) and acute renal failure (arf) in the critically ill, control of intraabdominal hypertension may contribute. to this end, accurate measurements of intraabdominal pressure are warranted. malbrain and colleagues [21] compared 315 (15 observers) manual foley catheter, and automated spiegelberg catheter and cimon ò -balloon catheter measurements with the applied hydrostatic pressure in a 3-l container. the measurements (between 0-30 cm h 2 o) were very close to each other. it is therefore suggested that manual are as good as automated measurements (and vice versa). other preventive measures may include drug treatments and in this context heemskerk et al. [22] report on the interesting potential of inhibiting no-induced soluble guanylate cyclase by methylene blue (mb) administration. in a small (n=9, uncontrolled) series of (refractory) septic shock patients, they suggested on the basis of a decrease in no metabolites and excretion of renal tubular injury markers in the urine that mb (4 mg/kg over 4 h) was able to decrease renal tubular damage and increase renal function parameters. however, the outcome benefits of such treatment warrant further study. the results obtained may thus increase our mechanistic rather than therapeutic insight. when aki/arf has nonetheless developed, renal replacement therapy must be initiated, the type, timing and dosing of which continues to be subject of debate. in a randomized study on septic patients (n=10/9) with need for renal replacement, high volume (65 ml/kg per h) was compared with low volume (35 ml/kg per h) hemofiltration [23] . the former treatment resulted in less norepinephrine requirements to maintain mean arterial blood pressure and urine output tended to increase. there was no effect on renal recovery/survival in this small, but unique (randomized!) study, while the effect on hemodynamics confirms observational data. extracorporeal therapy with polymyxin-b to scavenge endotoxin in severely septic patients has been tried before. in the study by cantaluppi et al. [24] however, the technique was used to evaluate its effect on circulating proapoptotic factors in gram-negative sepsis (n=8 pmx-b, n=8 standard care) that could play a role, among others, in the development of organ damage such as aki/ arf. there were some indications that this therapy indeed worked in this preliminary mechanistic study. circulating proapoptotic factors on cultured renal cells diminished, as well as sofa/rifle scores, proteinuria and tubular enzymes. the data suggest a role of apoptosis in the development of sepsis-related aki/arf. when clinicians have decided on the basis of likelihood of resuming endogenous renal function, to discontinue renal replacement therapy, some patients later again need such treatment. this issue has been addressed retrospectively by wu et al. [25] from taiwan, who, not unexpectedly, found that, among others, oliguria, high age and sofa, and a long period of prior need for renal replacement predicted its recurrent need (in 30%, within 30 days). indeed, some patients may have only partial recovery of renal function or remain on renal replacement therapy even months after the initial insult. moreover, aki/arf is a risk factor for chronic renal failure over decades. the paper thus again nicely illustrates that aki/ arf with need for renal replacement therapy is not a benign disorder and the future challenge, of course, is to find ways to promote full recovery and limit (recurrent) need for renal replacement therapy. two physiological reviews and notes have dealt with renal failure, as part of a cardio-renal syndrome and as a basis for renal replacement therapy [26, 27] . the authors extend the link between heart and kidney beyond congestive heart failure and also indicate a more aggressive approach in the use of different renal therapies. hemofilter circuits used for extracorporal renal replacement therapy have an expected lifespan of 48-72 h. however, they often clot prematurely which impairs azotemic control and has also effects on costs and nursing workload. antithrombin (at) deficiency has been associated with hemofilter thrombosis (ht) in patients with sepsis. lanquetot et al. [28] investigated whether there was an association between at level activity and ht occurrence during early continuous hemofiltration. they included 48 consecutive patients following cardiopulmonary bypass. subjects were grouped according to the appearance of one or more episodes of hemofilter thrombosis. morbidity and mortality did not differ significantly between the two groups. the authors found initial at activity to be low in both groups. on the following days at activity was lower in the ht group but was not found to be a predictor of ht in multivariate analysis. the authors concluded that the potential interest of monitoring at levels to adapt anticoagulant strategy needs to be analyzed in larger series of patients. during 2008 there has been a number of studies over tight glucose control. honiden et al. [29] reported in a prospective observational study that early tight glucose control, within 48 h from icu admission, was associated with a better outcome in terms of mortality, ventilator free days, and length of stay. also after the use of stepwise regression analysis, the differences between the two groups remained associated with the timing of the tight glucose control. the result calls for a prospective randomized trial to address the timing of tight glucose control. two studies report of the advantages of a computerized algorithm to facilitate and to increase success rate of tight glucose control. pachler et al. [30] evaluated their algorithm in a small randomized study. they found a mean blood glucose value within the target range and a lower hyperglycemic index in the group using a computerized algorithm, but also a shorter sampling interval. safety was evaluated by hypoglycemia, which was inconclusive, with overall only one hypoglycemic event in the 25 ? 25 patients. in a questionnaire, nurses were in favor of the computerized algorithm. vogelzang et al. [31] report from a large observational study that the hypoglycemia rate was low (0.85%) and that target level was reached faster and with fewer samples when the computerized algorithm was used. lapichino et al. [32] investigated if tight glucose control affected no-metabolism as reflected by the circulating levels of asymmetricdimethylarginine. in a prospective study of 72 patients in septic shock they found no difference attributable to tight glucose control in clinical outcome parameters or in asymmetric-dimethylarginine. mean blood glucose were 110 versus 163 mg/dl, respectively, in the two groups. ornithine transcarbamylase deficiency is a rare disorder usually diagnosed in childhood, which may also have an adult onset and which gives encephalopathy and obscure unconsciousness. panlaqui et al. [33] report of a case and point out the possible treatment with hemodia-filtration, protein elimination, and ammonia scavenging medications. the possibility of recovery with correct diagnosis and treatment makes awareness of such a rare condition important. nguyen et al. [34] used manometry to study gastric motility. in a small case-control study they showed that critically ill patients have an impaired motility both in the antrum and pyloric region. furthermore, the synchronization between the two regions was also impaired in the critically ill patients. using gastric scintigraphic data nguyen et al. [35] also report from a small observational study that the mode of sedation may influence gastric emptying. patients kept on midazolam and morphine had a significantly slower gastric emptying as compared to patients kept om propofol. the result clearly calls for a larger prospective randomized study. the cumulated energy deficit in icu patients primarily on enteral nutrition, which has been reported in several studies, is mainly related to underfeeding during the initial week of icu stay. desachy et al. [36] report of a prospective randomized study in 100 consecutive patients given enteral nutrition introduced stepwise or all at once. success rate in terms of calories delivered was significantly better in the groups given the full dose of enteral nutrition from start. that group had a larger fraction of gastric residues [300 ml, but the rate of adverse events necessitating complementary parenteral nutrition was similar in the two groups. adverse effects related to the use of prokinetic therapy was reported by nguyen et al. [37] in an observational study of 180 consecutive patients receiving erythromycin, metoclopramide or the combination of the two. patients given the combination had a higher rate of diarrhea, but no single patient had a positive toxin test for clostridium difficile. the role of immuno-modulating diets (imds) in critically ill patients is controversial. marik and colleagues [38] in the november issue published a meta-analysis to determine the impact of imds on hospital mortality, nosocomial infections and length of stay (los) in critically ill patients. by using the medline, embase, cochrane register of controlled trials as data source, were selected randomized control trials (rcts) that compared the outcome of critically ill patients receiving an imd or a control diet. twenty-four studies (with a total of 3,013 patients) were included in the meta-analysis; 12 studies included icu patients, 5 burn patients and 7 trauma patients. four of the studies used formulas supplemented with arginine, two with arginine and glutamine, nine with arginine and fish oil (fo), two with arginine, glutamine and fo, six with glutamine alone and three studies used a formula supplemented with fo alone. overall imds had no effect on mortality or los, but reduced the number of infections. mortality, infections and los were significantly lower only in the icu patients receiving the fo imd. the authors concluded that an imd supplemented with fo improved the outcome of medical icu patients (with sirs/sepsis/ards). imds supplemented with arginine with/without additional glutamine or fo do not appear to offer an advantage over standard enteral formulas in icu, trauma and burn patients. the ratio of omega-3 to omega-6 polyunsaturated fatty acids in the diet is suggested to have implications on the severity of the inflammatory response. friesecke et al. [39] randomized 177 consecutive patients on parenteral nutrition, also stratified for sirs or not, to receive a standard fat emulsion or a fish-oil enriched emulsion rich in omega-3 polyunsaturated fatty acids. endpoints were interleukin 6 and monocyte hla-dr as well as a number of morbidity/mortality parameters. no differences in any of the parameters studied were seen, which illustrates the difficulty to reproduce the promising results from animal studies in a relevant clinical material. regulation of the activity of transcription factor nf-kappab is an important therapeutic effect of the major omega-3 fatty acids in fo, eicosapentaenoic and docosahexaenoic acid (epa and dha). using the articles obtained by a pubmed research, singer and colleagues [40] reviewed three aspects of nf-kappab/inflammatory inhibition by fish oil: (1) the inhibition of the nf-kap-pab pathway at different levels, (2) the production of resolvin d1 and protectin d1 that are potent, endogenous, dha-derived lipid mediators that attenuate neutrophil migration and tissue injury in peritonitis and ischemia-reperfusion injury, (3) the modulation of vagal tone with potential anti-inflammatory effects. the authors concluded that whether the pleiotropic actions of epa/dha contribute to fo's therapeutic effect in sepsis remained to be shown. in the january issue, in conjunction with critical care medicine was published the controversial guidelines for management of severe sepsis and septic shock with the intent of providing an update to the original surviving sepsis campaign clinical management guidelines, ''surviving sepsis campaign guidelines for management of severe sepsis and septic shock,'' published in 2004 [41] . the authors used the grade system to guide assessment of quality of evidence from high (a) to very low (d) and to determine the strength of recommendations. key recommendations, listed by category, include: early goal-directed resuscitation of the septic patient during the first 6 h after recognition (1c); blood cultures prior to antibiotic therapy (1c); imaging studies performed promptly to confirm potential source of infection (1c); administration of broad-spectrum antibiotic therapy within 1 h of diagnosis of septic shock (1b) and severe sepsis without septic shock (1d); reassessment of antibiotic therapy with microbiology and clinical data to narrow coverage, when appropriate (1c); a usual 7-10 days of antibiotic therapy guided by clinical response (1d); source control with attention to the balance of risks and benefits of the chosen method (1c); administration of either crystalloid or colloid fluid resuscitation (1b); fluid challenge to restore mean circulating filling pressure (1c); reduction in rate of fluid administration with rising filling pressures and no improvement in tissue perfusion (1d); vasopressor preference for norepinephrine or dopamine to maintain an initial target of mean arterial pressure [ or =65 mm hg (1c); dobutamine inotropic therapy when cardiac output remains low despite fluid resuscitation and combined inotropic/vasopressor therapy (1c); stress-dose steroid therapy given only in septic shock after blood pressure is identified to be poorly responsive to fluid and vasopressor therapy (2c); recombinant activated protein c in patients with severe sepsis and clinical assessment of high risk for death (2b except 2c for post-operative patients). in the absence of tissue hypoperfusion, coronary artery disease, or acute hemorrhage, target a hemoglobin of 7-9 g/dl (1b); a low tidal volume (1b) and limitation of inspiratory plateau pressure strategy (1c) for acute lung injury (ali)/acute respiratory distress syndrome (ards); application of at least a minimal amount of positive endexpiratory pressure in acute lung injury (1c); head of bed elevation in mechanically ventilated patients unless contraindicated (1b); avoiding routine use of pulmonary artery catheters in ali/ards (1a); to decrease days of mechanical ventilation and icu length of stay, a conservative fluid strategy for patients with established ali/ ards who are not in shock (1c); protocols for weaning and sedation/analgesia (1b); using either intermittent bolus sedation or continuous infusion sedation with daily interruptions or lightening (1b); avoidance of neuromuscular blockers, if at all possible (1b); institution of glycemic control (1b) targeting a blood glucose \150 mg/dl after initial stabilization (2c); equivalency of continuous veno-veno hemofiltration or intermittent hemodialysis (2b); prophylaxis for deep vein thrombosis (1a); use of stress ulcer prophylaxis to prevent upper gi bleeding using h2 blockers (1a) or proton pump inhibitors (1b); and consideration of limitation of support where appropriate (1d). recommendations specific to pediatric severe sepsis include: greater use of physical examination therapeutic end points (2c); dopamine as the first drug of choice for hypotension (2c); steroids only in children with suspected or proven adrenal insufficiency (2c); a recommendation against the use of recombinant activated protein c in children (1b). the authors concluded that evidenced-based recommendations regarding the acute management of sepsis and septic shock are the first step toward improved outcomes for this important group of critically ill patients. the field of biomarkers for systemic inflammatory response syndrome (sirs) and sepsis is continuously evolving. the role of osteopontin, a protein with cell signaling functions in the interstitium upon inflammation, has been studied as a sepsis marker, by vaschetto et al. [42] in 56 sirs/sepsis patients (vs. 56 controls). the investigators demonstrated that circulating osteopontin could be used as a marker of il-6 release and of sepsis (vs. sirs), even though the levels were elevated in sirs (vs. control) as well. osteopontin appeared capable of increasing il-6 secretion by macrophages in vitro. it is unclear how specific the findings were for microbial infection, however. dulhunty et al. [43] reported on a multi-center study (n = 3,543 patients) done in australia and new zealand to answer the question whether severe non-infectious sirs differs from severe sepsis in the icu, regarding epidemiological issues. even though mortality rates were similar, severe non-septic sirs was more common and more often associated with neurological abnormalities and causes of death, than severe sepsis. this study adds to the idea that infection per se is not a major determinant of patient outcome, in contrast to the response of the body to either non-infectious or infectious threats. interactions of inflammatory mediators (or biomarkers) in sepsis remain immensely complex. de kruif et al. [44] demonstrated that some of these biomarkers are dose-dependently inhibited by corticosteroid treatment, and variations in the gene expression of interleukins 23 and 27 appeared to be different between septic icu patients, non-septic bacteremic patients and healthy controls [45] . and patients undergoing elective cardiac surgery, preoperative levels of apolipoprotein ci were associated with increased perioperative levels of tnfalpha in patients experiencing endotoxemia [46] . payen and colleagues [47] assessed blood leucocytes gene profiling in the course of the septic shock recovery period and tested the relation between encoding gene expression and protein level in 17 septic shock patients. gene expression levels were studied on a dedicated microarray of 340 genes involved in inflammatory processes. the time-related gene expression study showed significant changes in ten genes. among them, s100a8 and s100a12 had a reduced expression over time compared with d0, whereas cd74's expression increased. by rt-qpcr, the s100a8 plasma levels decrease in parallel with the gene expression decrease. the cd74 gene expression evolution significantly correlated with hla-dr monocyte expression. moreno and coworkers [48] wished to evaluate the value of the piro concept in septic patients. piro stands for predisposition, infection, response and outcome. the predictive value for outcome of premorbid factors, together with infectious features in the icu and the subsequent changes in sequential organ failure assessment (sofa) were evaluated. the saps 3 database was used for this purpose, on 2,628 patients with signs of infection in the icu [48 h. sepsis, severe sepsis and septic shock were defined according to standard criteria. hospital mortality was 41%. predictive factors were scored as the saps 3 piro score which appeared to have excellent (even somewhat better than the saps 3 model) predictive value for mortality. the conclusion of the authors is that piro components independently contribute to outcome prediction. thereby, the study is one of the first to argue in favor of the clinical validity as well as the practicability or usefulness of the piro concept. early identification of septic patients at high risk of death may provide an opportunity to change the treatment strategy to improve the outcome. because plasma midregional pro-atrial natriuretic peptide (mr-proanp) concentrations have recently emerged as a valuable tool for individual risk assessment in such a setting, seligman et al. investigated the prognostic role of this marker in 71 consecutive patients who developed vap [49] . the study demonstrated that mr-proanp levels were significantly higher in vap patients dying within 28 days when compared to survivors. in multivariate logistic regression models of predictors of death including age, sex, apache ii, and blood creatinine, mr-proanp levels on the day of diagnosis of vap and on day 4 turned out to be the only parameters that remained as independent predictors. lipoproteins, and in particular high-density lipoproteins (hdl), have been demonstrated to play an important role in modulating inflammation and the response to infection. because apolipoprotein ci (apoci) protects against the development of murine bacterial sepsis and is virtually completely depleted in human sepsis, its timecourse in patients with severe sepsis may predict survival. this hypothesis was confirmed in a small pilot study performed by berbee et al. [50] . upon hospitalization, apoci levels were approximately 5 times lower than normal values in 16 septic patients. remarkably, apoci levels remained low in non-survivors, whereas apoci levels gradually increased to normal levels in survivors. during the host response to infection and the development of sepsis and shock, vasopressor-insensitive hypotension (shock) may supervene for which the clinician may consider adjunctive therapy with hydrocortisone. kaufmann et al. [51] again demonstrated in 30 patients with septic shock that hydrocortisone treatment decreases vasopressor requirements and circulating il-6. moreover, this therapy preserved opsonization (zymosan)-dependent neutrophil functions, while suppressing spontaneous hydrogen peroxide release (flow cytometry). the (small) study was placebo-controlled. these data may help our understanding of the mechanism of action of steroid therapy in septic shock and suggest that hydrocortisone has no major immunosuppressive effects on neutrophils limiting host defences. severe sepsis and septic shock are serious complications of hematological malignancies. vandijck and colleagues [52] compared characteristics and outcomes in icu patients with hematological malignancies and severe sepsis/septic shock who had or had not received recent intravenous chemotherapy. among the 186 patients, there were 77 patients with severe sepsis and 109 with septic shock; 91 (49%) had received recent intravenous chemotherapy. in-hospital, and 6-month mortality rates were 45.1 versus 58.9%, and 50.5 versus 63.2% in patients with and without recent chemotherapy, respectively. by multivariate analysis, previous chemotherapy was protective. after adjustment with a propensity score for recent chemotherapy, chemotherapy was not associated with outcome. activated protein c for severe sepsis patients is a controversial therapy. in the study of kalil and sun [53] , the authors aimed to respond to two questions. the first was, what is the current probability that activated protein c is not better than the control? and, if the current probability is not small, then the second question was, how many patients will be needed for the activated protein c confirmatory trial? to give an adequate response to these questions, the authors used a bayesian statistical approach. the p value commonly used in frequentist statistical methods only tell us how likely we will observe the reported data when the null hypothesis is true, but does not tell us the actual probability of treatment effect. bayesian methodology, however, can provide this actual (or current) probability. to do this, all available data is selected for the prior probability. the prior distributions were defined as severe skeptic, moderate skeptic, mild skeptic and enthusiastic. the authors found that, except for the enthusiastic analysis, the current probabilities that activated protein c is not better than the control are not small (range, 0.14-0.48). the number of patients needed for a confirmatory trial ranged between 0 and 8,350. the authors concluded that a confirmatory trial with about 600 patients with severe sepsis and high risk of death can provide a convincing answer for moderate and mild skeptic physicians regarding the efficacy of the drug. in a special article published in the november issue finfer et al. [54] report on the ongoing new trial of activated protein c for persistent septic shock. the authors, members of the steering committee of the trial, fully discuss the potential benefits and harm of this drug and provide in-depth explanations about organizational issues and the sponsor's role, the design and the goals of this ongoing study, the safety monitoring and the analysis and report of the data. a full disclosure of the conflicts of interest is provided in the esm file accompanying this article, together with the complete study protocol. because its particular characteristics, this article is accompanied by editors' comments [55] , and two editorials: one providing an european view on this issue [56] , the other providing a north-american view [57] . intravascular-device related infections remain among the most frequent infectious complications of intensive care treatment. yet, the absolute risk on infection per catheter per day is low, hampering the analysis of intervention studies aimed to reduce this incidence. in a large study by gowardman et al. [58] , the incidence density of tip colonization (15.1/1,000 catheter days) was almost ten times as high as that of catheter-related infection (1.8/1,000 catheter days). in another observational study, the incidence density of arterial catheter colonization was 19.9 per 1000 catheter days, and the relative risk of colonization increased in time [59] , as did the risk for central venous catheter-related infection in a spanish multi-center study [60] . indeed, bacterial growth on the tip of catheters has been used as a proxy for catheter-related infections. in a prospective study, souweine and coworkers [61] determined that in patients without a clinical suspicion of catheter-related infection, the chance of bacterial growth on the tip was 4.7% (8.0 per 1,000 catheter days) and that administration of antibiotics at the time of removal was associated with a lower risk of tip colonization. therefore, proportions of patients on antibiotics at the time of line removal should be taken into account when using this proxy as an endpoint. all predictions are difficult, especially when they involve the outcome of infections and sepsis. yet, several studies aimed to identify accurate prognostic factors. an increment of c-reactive protein of 10 mg/l increased the odds of death after icu-discharge with 1.09 (95% ci 1.03-1.16) after adjustment for age, apache ii score predicted mortality and delta sofa with an area under the receiver operating characteristic curve of 0.85 (95% ci 0.73-0.96) [62] . in icu patients, high loads of herpes simplex type i virus in bronchoalveolar lavage fluid were associated with mortality. boer and coworkers [63] developed a prognostic model for the presence of posttraumatic stress syndrome after abdominal sepsis, that included age, length of icu stay and having traumatic memories of the icu or hospital stay [64] . the numbers of studies evaluating genetic associations with outcome is now rapidly increasing. angiotensinconverting enzyme insertion/deletion polymorphism was not associated with outcome in sepsis and ards (villar et al. [65] ). selective decontamination of the digestive tract (sdd) is a frequently used infection prevention measure in some countries. in sdd, non-absorbable antibiotics are applied topically and it has been assumed that detectable systemic levels of these agents, such as tobramycin, will not be reached. however, detectable serum levels of tobramycin were measured in 63% of patients receiving both sdd and cvvh. one patient had a toxic level of 3 mg/l [66] . in a randomized cross-over study, langgartner and coworkers [67] demonstrated that appropriate bacterial concentrations of meropenem in patients on continuous renal replacement therapy could be achieved with continuous infusion, with similar areas under the curve but longer times above the mic as with intermittent bolus injection. water sinks are considered as relevant sources for many bacterial species, including pseudomonas aeruginosa. in a non-outbreak setting, these bacteria were isolated from 86% of samples taken from u-bends and in 5% of those from tap water. yet, based upon genotyping, only 1 of 14 patients was colonized by a p. aeruginosa clone also isolated from water samples [68] . candida infections remain a crucial issue in the icu. in a 1-year national perspective observational study conducted in 24 adult french icus bougnoux and colleagues [69] determined the concomitant incidence, molecular diversity, management and outcome. the study enrolled 262 with nosocomial candidemia and/or candiduria. the mean incidences of candidemia and candiduria were 6.7 and 27.4/1,000 admissions, respectively. eight percent of candiduric patients developed candidemia with the same species. the mean interval between icu admission and candidemia was 19.0 ±2.9 days, and 17.2 ±1.1 days for candiduria. c. albicans and c. glabrata were isolated in 54.2 and 17% of blood and 66.5 and 21.6% of urine candida-positive cultures, respectively. fluconazole was the most frequently prescribed agent. crude icu mortality was 61.8% for candidemic and 31.3% for candiduric patients. seventy-five percent of the patients were infected with a unique c. albicans strain; cross-transmission between seven patients was suggested in one hospital. no difference in susceptibility and genetic background were found between blood and urine strains of candida species. it is difficult to judge the quality of care in different icus. objective and reproducible criteria are badly needed, but hampered by the absence of a gold standard. najjar-pellet and coworkers [70] proposed a scoring system based upon 95 variables that might serve as a tool for quality assessment in future studies. one quality variable, not included in the previously mentioned scoring system, is the recognition of rare diseases, such as, for instance acute disseminated encephalomyelitis (adem). in a retrospective multi-center study of 6 years sonneville and coworkers [71] describe the characteristics of 20 adem patients needing icu admission. adem is not a benign disease, as 25% of the patients died and 35% had persistent functional sequelae. p damas and colleagues [72] in an observational single-center study assessed the temporal relationship between icu-acquired infection (iai) and the prevalence and severity of organ dysfunction or failure (od/ f). almost 2,200 patients hospitalized for more than 2 days during a 2-year observation period were studied: 845 did not acquire iai, 306 of whom had infection on admission (ioa); 346 did acquire iai, 125 of whom had ioa. the saps ii and sofa score of the first 24 h were significantly higher in patients with than in those without iai. sofapreinf of iai patients was also higher than the sofamax of patients without iai both in patients with (12.1 ± 4.6 vs. 8.9 ± 4.7) and those without ioa (9.2 ± 4.0 vs. 6.7 ± 3.5). sofapreinf represented 85.7% of the value of sofamax in patients with iai. sofapreinf increased significantly with the occurrence of sepsis, severe sepsis, or septic shock during icu stay. the authors concluded that icuacquired infections are significantly associated with hospital mortality; but thir contribution to od/f sems minor. aminoglycosides are broad-spectrum antibiotics active against most pathogens responsible for ventilator-associated pneumonia (vap), even those with multidrugresistance patterns. however, the systemic use of this antibiotic class is limited by its toxicity and poor penetration into the lung. aerosol administration offers the theoretical advantage of achieving high antibiotic concentrations at the infection site and low systemic absorption, thereby avoiding renal toxicity. however, some uncertainties persist regarding the real usefulness of such a mode of administration, since during mechanical ventilation (mv), high amounts of the particles dispersed by conventional nebulizers remain in the ventilatory circuits and the tracheobronchial tree before reaching the distal lung. in a study carried out in subjects with healthy lungs, ehrmann et al. [73] showed that, using an optimized nebulization technique with a vertical spacer placed underneath a vibrating mesh nebulizer, doses of 60 mg/kg amikacin were associated with serum concentrations equal to or less than those obtained after intravenous infusion of 15 mg/kg amikacin. because amikacin systemic pharmacokinetics reflect deposition of the nebulized drug in the distal pulmonary parenchyma, these data strongly support the hypothesis that optimized nebulization of antimicrobial agents may permit to obtain very high lung parenchyma concentrations during a sufficiently long period of time for achieving bactericidal activity. despite advances in prophylactic perioperative antibiotic therapy, post-operative pneumonia is a feared complication following major surgery and is associated with an icu mortality of 10-20%. one of the major predisposing factors is insufficient target-site concentration of antibiotics used for prophylaxis. because ventilation/perfusion mismatch due to atelectasis may also influence antibiotic distribution to lung tissue, hence increasing the risk of post-operative pneumonia, hutschala et al. compared the penetration of levofloxacin into the lung of two groups of patients, using microdialysis probes to sequentially determine in vivo lung tissue levels. the first group consisted of five patients who underwent coronary artery bypass grafting (cabg) with cardiopulmonary bypass (cpb) (atelectasis model), and the second one, of five patients operated with the offpump coronary artery bypass grafting (opcab)-technique [74] . in the opcab-group, the median of the maximum concentration of levofloxacin in lung tissue was significantly higher compared with the cpb-group, establishing that atelectasis formations lead to critically lower lung tissue concentrations of levofloxacin in nondependent parts of lung tissue. such data emphasizes the necessity of direct interstitial antibiotic measurement to re-evaluate commonly accepted prophylactic and therapeutic antibiotic dosages in various clinical settings and pulmonary diseases associated with the formation of atelectasis. to favorably impact the outcome of patients with severe nosocomial sepsis, antibiotic therapy covering the offending pathogen has to be initiated without delay, which implies administration within 24 h of clinical deterioration. in patients at risk for infection with multidrug resistant (mdr) pathogens, the clinician has to resort to broad-spectrum antimicrobials, which are themselves linked with the emergence of multi-drug resistance. the potential value of systematic endotracheal tracheal surveillance cultures as a tool to predict involvement of mdr microorganisms in vap was studied by two independent groups of investigators [75, 76] bacterial colonization of the respiratory tract frequently persists, even when a patient receives antimicrobial treatment, and even though the colonizing bacteria are, in vitro, susceptible to the antibiotics. to test the hypothesis that antibiotics with presumed efficacy, based on in vitro susceptibility testing, reduce the likelihood of persistence of respiratory tract colonization, compared to antibiotics presumed to be ineffective or when no antibiotics were administered at all, visscher et al. [78] analyzed endotracheal aspirate cultures performed during icu stay in a large cohort of 715 mechanically ventilated icu patients. systemic antibiotics were administered on 7,102 (61%) of patient days. antibiotic use was associated with non-persistence for all pathogens, except acinetobacter species and p. aeruginosa. relative risks for non-persistence (as compared to ineffective or no antibiotics) ranged from 3.1 (95% ci 1.4-6.6) for h. influenzae to 0.5 (0.3-1.0) for acinetobacter species. pathogen-specific characteristics, such as the ability of biofilm formation of p. aeruginosa, or patient-specific characteristics, such as the severity of underlying disease or immune paralysis, could both be involved, but further studies are needed to elucidate this matter. unlike mycobacterium tuberculosis, nontuberculous mycobacteria (ntm) exist in the environment and can be isolated from clinical specimens in the absence of true infection. in patients with complicated and critical conditions, such as those admitted to icu, the clinical significance of ntm in respiratory specimens and the prognostic impact of ntm pulmonary infection are even more difficult to understand than in stable patients. therefore, shu et al. conducted a retrospective study including all medical icu patients with ntm being isolated from respiratory specimens within a period of 8.5 years to evaluate the clinical significance of the presence of ntm and compare the demographic characteristics, clinical manifestations, and outcome in patients with ntm pulmonary infection with those with ntm colonization and control subjects whose respiratory samples were culture-negative for mycobacteria [79] . among the 5,378 patients admitted to medical icus, ntm were isolated from 169 (5.8%) patients. of them, 47 (28%) were considered ntm pulmonary infection. within 100 days after icu admission, significantly more patients with ntm infection died than those with ntm colonization and control subjects (47 vs. 8 vs. 14%, p \ 0.001). therefore, keeping a high suspicion when ntm is isolated and using careful consideration when starting anti-ntm treatment should be emphasized. in the absence of a clinically available gold standard, vap is usually diagnosed according to a combination of criteria, such as systemic signs of infection, abnormalities on chest radiograph, and microbiological identification of pathogens; however, each of these criterions combines high sensitivity with low specificity. in an attempt to raise diagnostic accuracy, luyt et al. [80] assesses the predictive capacity for the diagnosis of vap of serum procalcitonin levels before and on the day it is suspected. among the 73 suspected episodes vap was confirmed by quantitative bronchoalveolar lavage cultures in 32 and refuted in 41. on day 1 a 0.5 ng/ml procalcitonin threshold had 72% sensitivity but only 24% specificity for diagnosing vap. between ''before'' and day 1, procalcitonin increased in 41 and 15% of patients with and without vap, respectively. thus, crude values and procalcitonin rise had poor diagnostic value for vap in this particular setting and should not be used to initiate antibiotics when vap is clinically suspected. mixed results were also observed by el solh et al. [81] when they examined the potential role of serum and alveolar soluble triggering receptor expressed on myeloid cells (strem-1) as a biological marker of pulmonary aspiration syndromes. while circulating levels of strem-1 were comparable between those with aspiration syndromes and controls, the alveolar levels of strem-1 were higher in patients with culture-positive pulmonary aspiration compared with those culture-negative pulmonary aspiration (p \ 0.001). a cut-off value of 250 pg/ml for alveolar strem-1 achieved a sensitivity of 66% and a specificity of 92% with an area under the curve of 0.87. because cells and secretions recovered by bronchoalveolar lavage (bal) can be microscopically examined immediately after the procedure to detect the presence or absence of intracellular or extracellular bacteria in the lower respiratory tract, this technique is particularly well suited to provide rapid identification of patients with pneumonia. however, it is commonly assumed that prior antimicrobial therapy can dramatically decrease its sensitivity. in order to re-assess the influence of antibiotics on the value of various bal cytological parameters in diagnosing vap, linssen et al. [82] studied 335 episodes of clinically suspected vap in 282 patients. there was no difference in areas under the curve (aucs) of receiver operating characteristic curves between patients with and without antibiotic therapy for any parameter studied. the most prominent aucs were: total cell count, 0.65; percentage polymorphonuclear neutrophils, 0.71; and percentage infected cells, 0.90. based on these data, it appears that the percentage of infected cells in balf can be reliably used for diagnosing vap in patients receiving antibiotic therapy, provided that the introduction of the new antibiotics was recent (less than 72 h). prevention vap remains a major problem in intensive care units and effective preventive measures are eagerly searched for. the presence of an endotracheal tube (ett) not only compromises the natural barrier between the oropharynx and trachea, but also provides a protected environment for pathogens since a biofilm develops on its inner and outer surfaces. in an attempt to prevent bacterial colonization of the lower respiratory tract in patients requiring mechanical ventilation, berra et al. [83] developed a polyurethane ett coated with silver sulfadiazine. in a second study, the same group of investigators randomized 46 adult patients to be intubated with a standard non-coated ett, or with a silver sulfadiazine-coated ett [84] . coating with silver sulfadiazine prevented bacterial colonization of the ett and was associated with a thinner mucus layer. although preliminary, these data support the hypothesis that an endotracheal tube coated externally and internally with a potent antiseptic product such as silver could exert a sustained antimicrobial effect within the proximal airways and block biofilm formation at its surface. moderate and severe traumatic brain injury induce early overexpression of systemic and brain gelatinases extracranial complications in patients with acute brain injury: a post-hoc analysis of the soap study monitoring of intracranial pressure in patients with severe traumatic brain injury: an austrian prospective multicenter study noninvasive 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units: proposal for a scoring system in terms of structure and process acute disseminated encephalomyelitisin the intensive care unit: clinical features and outcome of 20 adults intensive care unit acquired infection and organ failure pharmacokinetics of high-dose nebulized amikacin in mechanically ventilated healthy subjects the impact of perioperative atelectasis on antibiotic penetration into lung tissue: an in vivo microdialysis study systematic surveillance cultures as a tool to predict involvement of multidrug antibiotic resistant bacteria in ventilator-associated pneumonia screening for resistant gram-negative microorganisms to guide empiric therapy of subsequent infection throat and rectal swabs may have an important role in mrsa screening of critically ill patients effects of systemic antibiotic therapy on bacterial persistence in the respiratory tract of mechanically ventilated patients nontuberculous mycobacteria pulmonary infection in medical intensive care unit: the incidence, patient characteristics, and clinical significance usefulness of procalcitonin for the diagnosis of ventilator-associated pneumonia triggering receptors expressed on myeloid cells in pulmonary aspiration syndromes influence of antibiotic therapy on the cytological diagnosis of ventilator-associated pneumonia antimicrobialcoated endotracheal tubes: an experimental study internally coated endotracheal tubes with silver sulfadiazine in polyurethane to prevent bacterial colonization: a clinical trial key: cord-302295-nblmshni authors: savva, athina; roger, thierry title: targeting toll-like receptors: promising therapeutic strategies for the management of sepsis-associated pathology and infectious diseases date: 2013-11-18 journal: front immunol doi: 10.3389/fimmu.2013.00387 sha: doc_id: 302295 cord_uid: nblmshni toll-like receptors (tlrs) are pattern recognition receptors playing a fundamental role in sensing microbial invasion and initiating innate and adaptive immune responses. tlrs are also triggered by danger signals released by injured or stressed cells during sepsis. here we focus on studies developing tlr agonists and antagonists for the treatment of infectious diseases and sepsis. positioned at the cell surface, tlr4 is essential for sensing lipopolysaccharide of gram-negative bacteria, tlr2 is involved in the recognition of a large panel of microbial ligands, while tlr5 recognizes flagellin. endosomal tlr3, tlr7, tlr8, tlr9 are specialized in the sensing of nucleic acids produced notably during viral infections. tlr4 and tlr2 are favorite targets for developing anti-sepsis drugs, and antagonistic compounds have shown efficient protection from septic shock in pre-clinical models. results from clinical trials evaluating anti-tlr4 and anti-tlr2 approaches are presented, discussing the challenges of study design in sepsis and future exploitation of these agents in infectious diseases. we also report results from studies suggesting that the tlr5 agonist flagellin may protect from infections of the gastrointestinal tract and that agonists of endosomal tlrs are very promising for treating chronic viral infections. altogether, tlr-targeted therapies have a strong potential for prevention and intervention in infectious diseases, notably sepsis. sepsis is one of the leading causes of death worldwide. incidence of severe sepsis is increasing and mortality rates remain significantly high despite early care management (1) . moreover, more than 30% of survivors develop long-term functional disabilities and cognitive impairments (2) . the surviving sepsis campaign is a global initiative incepted in early 2000s with the aim to improve sepsis diagnosis and treatment in order to enhance the awareness of sequelae and to decrease high mortality rates associated with sepsis 1 . in collaboration with many countries in europe and the united states, the surviving sepsis campaign suggests evidencebased guidelines and bundles. the most recent guidelines recommend acute resuscitation of septic patients, administration of antibiotics and support of organ failure. yet, no treatment targeting the underlying mechanism of sepsis is actually available (3) . recombinant human activated protein c (rhapc, xigris®, eli lilly), the only drug specifically registered for sepsis, has recently been withdrawn from the market following the negative results from the prowess-shock study that did not show reduction in mortality at 28 or 90 days in patients with septic shock (4) . it is generally admitted that sepsis results from a dysregulated host response to an initial insult, characterized by inflammation mediating tissue damage and organ failure and an immune suppression state responsible for the development of secondary infections (5) (6) (7) . the immune response to an infection is initiated by the sensing of microbial structures through families of receptors collectively called pattern recognition receptors (prrs). the most well-described families comprise toll-like receptors (tlrs), nucleotide binding oligomerization domains (nods)-like receptors (nlrs), c-type lectin receptors (clrs, such as dectin-1, dectin-2, dc-sign), rig-i-like receptors (rlrs, rig-i, and mda5), and intra-cytosolic dna sensors. prrs are expressed by innate immune cells like dendritic cells and macrophages. the binding of microbial ligands to prrs promotes the release of mediators, among which cytokines, that initiate and regulate the inflammatory response necessary to eliminate invasive pathogens and coordinate the development of the adaptive immune response (8, 9) . innate immune cells are also triggered by damage (or danger)associated molecular patterns (damps), known as alarmins. damps are endogenous components commonly released by injured or stressed cells, such as nucleic acids, histones, uric acid crystals, atp, cytochrome c, s100 molecules, and hmgb1. damps are primarily sensed through the nlrp3 inflammasome, which controls the secretion of il-1β and il-18 (10) . the concept of prrs sensing microbial-associated molecular patterns (mamps) and discriminating self from non-self molecular structures was proposed by janeway (11) . two major cornerstone discoveries largely confirmed janeway's concept. the first one was the demonstration of the essential antifungal role of www.frontiersin.org the toll protein in drosophila (12) . the second one arose from the positional cloning linking lps (commonly called endotoxin) unresponsive phenotype of c3h/hej and c57bl/10sccr strains of mice to missense and null mutations of the toll-like receptor 4 (tlr4) gene (13) . the importance of these discoveries and of the role of dendritic cells as central regulators of innate and adaptive immunity has been acknowledged by the 2011 nobel prize in physiology or medicine attributed to bruce a. beutler and jules a. hoffmann "for their discoveries concerning the activation of innate immunity" and to ralph m. steinman "for his discovery of the dendritic cell and its role in adaptive immunity" (14) . toll-like receptors belong to the most studied family of prrs, due to their central role in host defenses and involvement in a number of pathological processes that include sepsis. tlrs are type i trans-membrane proteins composed of an extracellular leucine-rich repeat (lrr) domain involved in ligand recognition, a trans-membrane domain, and a toll-interleukin 1 receptor (tir) domain involved in signaling (9, 15) . about 10 functional human tlrs (tlr1-10) and 12 functional mouse tlrs (tlr1-9, tlr11-13) have been described, each one being involved in the sensing of distinct microbial products ( table 1) . expressed at the cell surface, tlr4 detects lps from gram-negative bacteria. tlr4 shuttles to late endosome to induce alternative signaling following lps sensing. tlr2 as heterodimers in association with either tlr1 or tlr6 (and possibly tlr10) senses a variety of microbial products, such as lipopeptides, lipoproteins, peptidoglycan, porins, β-glucan, glycosylphosphatidylinositol (gpi) anchors, and glycoproteins from gram-positive bacteria, gram-negative bacteria, mycoplasma, mycobacteria, fungi, parasites, and viruses. tlr5 senses flagellin of bacterial flagella. tlr3, tlr7, tlr8, and tlr9 are strategically expressed in endosomal compartments to recognize microbial nucleic acids: double-stranded rna (dsrna) by tlr3, single-stranded rna (ssrna) by tlr7 and tlr8, and unmethylated cpg motif containing dna by tlr9 ( table 1) . it is therefore not surprising that endosomal tlrs have been primarily involved in host defenses against viruses, whereas tlr1, tlr2, and tlr4-6 have been mainly involved in host response to bacterial and fungal infection. tlrs cooperate with other molecules to recognize microbial ligands. for example, tlr2 requires cd14, cd36, and dectin-1 for the recognition of peptidoglycan, lipopeptides, and β-glucan, respectively. of note, tlrs are also triggered by damps released by injured or stressed cells during infection (16) ( table 1) . tlr activation enables pathogen elimination by promoting bactericidal activity of leukocytes, and maturation and function of antigen presenting cells, thus orchestrating the development of adaptive immune responses (17) . the signaling pathways resulting from tlr triggering engage adaptors that are recruited by tir/tir domain interactions ( table 1) : myeloid differentiation primary response gene (18) (myd88), tir domain-containing adaptor protein (tirap, also known as mal), tir domain-containing adaptor inducing interferon (ifn)β (trif), and trif related adaptor molecule (tram) (19) . myd88 is essential for signaling through all tlrs except tlr3 and is involved in early nuclear factor-κb (nf-κb) and mitogen-activated protein kinases (mapks) activation and pro-inflammatory gene expression. tirap serves as a bridge to recruit myd88 to tlr2 and tlr4. trif initiates myd88independent ifn regulatory factor 3 (irf3) and late nf-κb activation involved in the production of type i ifns and ifn-inducible genes (19, 20) . trif is recruited to the cytoplasmic domain of tlr3 and, in late endosome, through tram that bridges trif to tlr4. a fifth tir domain-containing adaptor, sterile α-, and armadillo-motif containing protein (sarm) acts as a negative regulator of tlr3 and tlr4 signaling. sarm interacts with trif and inhibits the induction trif-dependent genes. the signaling pathways activated downstream tlrs have some redundancy. yet, the engagement of multiple tlrs, especially myd88 and trif-dependent tlrs, have synergistic effects on host responses (21, 22) . intracellular cross talk between signaling pathways may also occur when different families of prrs are involved. for example, dectin-1 synergizes with tlr2 and tlr4 and increases cytokine production through canonical and noncanonical nf-κb pathways (23, 24) . moreover, a single mamp can be detected by different prrs. this differential sensing is primarily depending on the localization of the mamp. indeed, flagellin is sensed by tlr5 expressed at the cell surface and by the naip5/nlrc4 (also known as ipaf) inflammasome when localized in the cytoplasm (25) (26) (27) . similarly, peptidoglycans stimulate membrane tlr2 and intra-cytosolic nod1/nod2dependent cell activation (28) . interestingly, some cpg and non-cpg oligodeoxynucleotides directly stimulate and polarize t-cells through tlr9 and myd88-independent mechanisms (29), possibly through intracellular dna sensors. recently, hagan et al. and kayagaki et al. demonstrated non-canonical tlr4-independent recognition of intracellular lps through an uncharacterized receptor (30, 31) . this unconventional mode of lps sensing activates caspase-11-dependent il-1β secretion and sensitizes mice to endotoxic shock. all these observations indicate that the host has evolved different strategies to sense invading microorganisms. ideally, all possible interactions should be characterized and/or anticipated, so that the effect of treatment application can be predicted and/or translated. this mandates carefully planned experiments that represent real-life conditions and a detailed knowledge of compound's mode of action. obviously, the redundancy of microbial sensing pathways should be taken into consideration when developing or applying targeted-treatment strategies to a single prr. experimental animal models and human clinical studies support a crucial role for tlrs in infectious diseases. the first evidence came from the observation that tlr4 defective c3h/hej and c57bl/10sccr mice are hyporesponsive to lps and susceptible to otherwise non-lethal infection with escherichia coli and salmonella typhimurium. subsequent studies with mice knockout in tlrs or tlr adaptor molecules have demonstrated the importance of the tlr pathway in host defenses. for example, tlr2 knockout mice are highly susceptible to infections by staphylococcus aureus and streptococcus pneumoniae (32) . more recently, human association studies have linked polymorphisms affecting tlr expression or tlr structure with an augmented propensity to develop infections (32) (33) (34) (35) . the discovery of tlrs and their involvement in innate immune responses has attracted much interest into the development of drugs for controlling infections and improving sepsis management. this field of research has been very dynamic, and (36) . these two particular aspects of the tlr-targeting field will not be addressed in this review. herein, we will review the most popular agonist (tlr3, tlr5, tlr7, tlr8, tlr9) and antagonist (tlr2, tlr3, tlr4, tlr9) agents used in pre-clinical and clinical models of acute and chronic infections, including sepsis. relevant registered clinical trials 2 are listed in table 2 . frontiers in immunology | microbial immunology lps is the main pro-inflammatory molecule anchored in the outermembrane of gram-negative bacteria (51) . neutralization of bacterial lps, inhibition of its recognition by host cells or inhibition of signaling downstream lps binding to its receptor has long been considered a promising approach for the treatment of severe sepsis and septic shock. interestingly, endotoxemia is prevalent in septic patients, not only in those with gram-negative infection. indeed, translocation of viable bacteria and lps from the gastrointestinal tract has been proposed to participate in the pathophysiology of sepsis. tlr4 was identified 15 years ago as the signal-transducing molecule of the lps receptor complex (13) , which also comprises md-2 and cd14. thus, tlr4 is regarded as a primary target for treating sepsis (52) . tlr4 expression is increased in human monocytes of healthy volunteers challenged with lps (53), as well as in patients with sepsis (54) . moreover, polymorphisms in the tlr4 gene have been associated with gramnegative sepsis (33, 35) . in the following sections, we present the most advanced tlr4 antagonists developed for the treatment of sepsis. strategies to inhibit lps-mediated toxic effects have been initiated years before the discovery of tlr4 (13) and the unraveling of the crystal structure of the tlr4-md-2-lps complex (55) . lipid a, the toxic moiety of lps, is highly conserved among endotoxins and constitutes an ideal therapeutic target (56) . e5531, developed by eisai research institute of boston (andover, ma, usa), was the first-generation lipid a antagonist derived from rhodobacter capsulatus endotoxin. e5531 conferred protection in experimental models of endotoxemia and lethal infection with e. coli (57) . the protective effect likely occurred through the binding of e5531 to the tlr4-md-2 complex and the inhibition of the interaction between lps and tlr4-md-2 (58) . e5531 also blocked endotoxin response in human healthy volunteers challenged intravenously with lps (59). e5531 development went through phase 2 clinical trial, but was stopped due to issues of bioavailability. a second-generation lps antagonist drug candidate developed by eisai is eritoran tetrasodium (known as eritoran or e5564), a synthetic lipid a analog of rhodobacter sphaeroides (60) . eritoran blocked lps-induced cytokines in vitro and in experimental animal models (61) (62) (63) . in a phase 1 clinical trial enrolling healthy volunteers challenged with lps, eritoran inhibited pro-inflammatory cytokine production and diminished clinical symptoms of sepsis, including fever, chills, tachycardia, and headache. additionally, creactive protein levels and white blood counts were significantly decreased (64) (65) (66) (67) . the only adverse event observed was a dosedependent phlebitis, due to the fact that high doses of eritoran were used to achieve stable activity of the drug over time. a phase 2 randomized control trial recruiting critically ill septic patients as assessed by the acute physiology and chronic health evaluation ii (apache ii) score disclosed a trend toward decreased mortality in the eritoran treated group (37) . phase 3 access (a controlled comparison of eritoran and placebo in patients with severe sepsis) clinical trial for severe sepsis started in 2006, and results were published in 2013. about 1304 patients were treated with eritoran and 657 patients with placebo within 12 h after the onset of the first organ dysfunction. unfortunately, analyses did not reveal reduced all-cause mortality in primary and secondary end-points (i.e., 28 days and 1 year mortality) (38) . eisai (tokyo, japan) waived to submit eritoran to marketing authorization for the treatment of severe sepsis in january 2011, based on preliminary results of the access trial. several reasons may account for the lack of efficacy of eritoran (68) (69) (70) . for instance, patients were not enrolled or monitored based on the circulating levels of lps, questioning about the appropriateness of inclusion criteria. it is also possible that eritoran would be more efficient if administrated rapidly, before septic shock is underway, pointing the early and aggressive sepsis management as a possible interfering factor. other factors to take into account include the heterogeneity of patients for genetic background, underlying diseases, inflammatory and immune status, sepsis severity, infectious agent, and site of infection. as mentioned earlier, intracellular lps sensed in a tlr4-independent manner sensitizes mice to endotoxic shock (30, 31) . this non-canonical lps detection may have limited the efficacy of the anti-tlr4 strategy. moreover, upon infection, innate immune cells will likely sense several mamps via several tlrs and non-tlr prrs. for example, gram-negative bacteria express mamps that may trigger redundant inflammatory pathways through tlr2 (lipopeptides), tlr4 (lps), tlr5 (flagellin), tlr7 (ssrna), and tlr9 (bacterial dna). all these observations suggest that blocking one single pathway may be insufficient to interfere with the deleterious cascade of events observed in sepsis. the positive side of the access trial failure was a rethink of the design of sepsis clinical trials (68) (69) (70) . clearly, a drug like eritoran should be tested in selected patients and treatment efficacy examined and adjusted according to predefined appropriate biomarkers (such as lps blood levels and genetic polymorphisms affecting the tlr4 pathway). a rigorous approach combining the power of "omics" technologies would allow the selection of homogeneous cohorts and the follow-up of the response to treatment, both of which are mandatory for the successful development of anti-sepsis drugs. another anti-sepsis agent that exhibited promising therapeutic properties is tak-242 [ethyl-(6r)-[n -(2-chloro-4-fluorophenyl) sulfamoyl] or resatorvid] from takeda pharmaceutical company (osaka, japan). tak-242 was originally characterized as a suppressor of nitric oxide (no) and cytokine production by lpsstimulated macrophages and during endotoxic shock in mice (71) . tak-242 binds to cysteine 747 in the intracellular domain of tlr4, thereby inhibiting both myd88-dependent and myd88independent pathways activated by lps (72) . when administered in conscious guinea pigs following lps challenge, tak-242 significantly improved septic shock symptoms, decreasing hmgb1 systemic levels, and increasing survival in a dose-dependent manner (73) . tak-242 also increased survival rates from 17 to 50% and improved organ dysfunction when co-administered with antibiotics in a mouse model of cecal ligation and puncture (clp). no effect on circulating bacterial counts was observed (74) . a double-blind, randomized, placebo-controlled trial was initiated with tak-242 (39) . inclusion criteria comprised symptoms of severe sepsis accompanied with either shock and/or respiratory frontiers in immunology | microbial immunology failure. the study was stopped prematurely due to failure to achieve significant decrease of systemic cytokine levels at stage 1 of the analysis (39) . a phase 3 clinical study was designed but never launched based on business decision and not due to safety or efficacy concerns. antibodies directed against tlr4 or the tlr4-md-2 complex have been generated and showed promising results in several pre-clinical studies. we engineered a soluble chimeric protein composed of the n-terminal and central domains of mouse tlr4 (amino acid 1-334) fused to the fc domain of human igg1 (75) . the chimeric molecule was used to generate high titer anti-mouse tlr4 rabbit polyclonal antibodies. the anti-tlr4 antibodies powerfully inhibited nf-κb and mapk activation and cytokine production by lps-stimulated cells in vitro. the antibodies also hampered cytokine production and protected mice from lethal endotoxemia when administered both prophylactically and therapeutically 4 h after lps. prophylactic administration of anti-tlr4 antibodies blunted tnf production and strikingly increased survival in e. coli sepsis, from 0% in the control antibody group to 80% in the anti-tlr4 group. even more impressive, anti-tlr4 therapy initiated as much as 13 h after the onset of infection in a model of e. coli peritoneal infection improved survival from 30 to 75% (75) . our studies demonstrate that anti-tlr4 antibodies are efficient as adjunctive therapy for e. coli sepsis, with a window of clinical application comprising prophylactic and therapeutic intervention opportunities. several anti-tlr4 monoclonal antibodies have been produced. the group of miyake (university of tokyo, japan) reported in 2000 the generation of mts510, the first rat monoclonal antibody specific of the mouse tlr4-md-2 complex (76) . mts510 was shown to inhibit lps-induced nf-κb activation and tnf production by macrophages. 5e3 is a rat monoclonal antibody produced by novimmune sa (geneva, switzerland) that reacts with the tlr4-md-2 complex (77). 5e3 inhibited lps-induced cell activation, and protected mice from lethal endotoxemia when injected up to 7 h after lps challenge. moreover, administration of 5e3 at the time of surgery improved the outcome of mice with colon ascendens stent peritonitis, a model of polymicrobial abdominal sepsis. finally, the rat monoclonal antibody 1a6, that recognizes both mouse and human tlr4-md-2 complexes, conferred protection in a model of e. coli sepsis, but not salmonella enterica, sepsis (78) . although some tlr4 inhibitors have entered clinical and pre-clinical trials, others remain in the developmental stage. lps-trap-fc antibodies (comprising the extracellular domain of mouse tlr4 fused with md-2 and linked to human igg fc) dose-dependently decreased il-6 release by macrophages, opsonized gram-negative bacteria, and enhanced phagocytosis and complement-mediated bacterial killing (79) . cell-penetrating peptides comprising the translocating segment of drosophila antennapedia homeodomain fused with bb loop sequences of tlr4 (i.e., tlr4-bb peptides) inhibited lps-induced nf-κb and mapk activation and cytokine production (80, 81) . further studies will be required before advancing these products toward the clinical level. altogether, the experimental data reported above provided strong support for the concept of tlr4-targeted therapy for gram-negative sepsis. in the gloomy context following the withdrawn of rhapc and eritoran from the sepsis field, it is hopeful that ni-0101 has entered clinical development. ni-0101 is an anti-tlr4 monoclonal antibody produced by novimmune able to block tlr4 dimerization and tlr4-mediated signaling triggered by lps and endogenous and chemical ligands of tlr4. data from pre-clinical studies in models of arthritis, respiratory inflammation, and organ injury have highlighted the potential favorable action of this agent 3 . a phase 1 clinical study is currently recruiting participants to evaluate drug safety and tolerance in healthy volunteers before and after ex vivo and in vivo lps challenge. pharmacokinetics and pharmacodynamics will also be assessed. results from these studies are eagerly awaited. albeit less well characterized, tlr4 has been implicated in the sensing of non-bacterial microorganisms such as viruses and fungi. tlr4 recognizes o-linked mannan from candida albicans, and human studies have linked asp229gly tlr4 polymorphism with susceptibility to bloodstream candidiasis and pulmonary aspergillosis (82) (83) (84) . in a model of disseminated infection with c. albicans, c3h/hej tlr4-deficient mice exhibited a 10-fold increased fungal load in the kidneys, which was associated with reduced production of the chemokines kc and mip-2 and an impaired recruitment of neutrophils (85) . treatment with hta 125, an anti-human tlr4 mouse monoclonal antibody (86), interfered with neutrophil-mediated protection against c. albicans invasion and cell injury in an in vitro epithelial model of oral candidiasis (87) and inhibited tnf production by human pbmcs stimulated with aspergillus hyphae (88) . it is still unclear whether targeting tlr4 may be beneficial in the context of fungal infections. a more clear yet unexpected picture has arisen from viral infection studies. reactive oxygen species (ros) produced by the nadph oxidase generates oxidized host phospholipids that stimulate tlr4 and the production of cytokines involved in acute lung injury (89) . using a mouse model of lethal infection with influenza, the group of stephanie vogel (university of maryland, baltimore, ma, usa) reported that eritoran significantly increases survival in a dose-dependent manner even when administered 6 days after viral challenge. lung pathology and clinical symptoms were improved while viral titers and influenza-induced cytokine gene expression in lung homogenates were decreased compared to the placebo-treated group. these data suggest that the therapeutic effect of eritoran in a more practical timing of severe sepsis treatment remains substantial (90) . they also suggest that, despite the failure of eritoran in the access trial, new therapeutic potentials might still emerge for this agent. toll-like receptor 2 has been implicated in the recognition of an amazingly broad spectrum of microbial ligands originating from bacteria, fungi, viruses, and parasites (91) . this property is at least partly due to the fact that tlr2 forms heterodimers with tlr1, tlr6, and possibly tlr10 (9, 15, 92) . the biological relevance of tlr2 homodimers is controversial. indeed, some ligands have been reported to trigger cells through tlr2 independently of tlr1 and tlr6. yet, only tlr2/tlr1 and tlr2/tlr6 heterodimers have been successfully crystallized (93, 94) . tlr2 represents an interesting target for numerous conditions, but clinical development of tlr2-targeting drugs has been less extensive than that of tlr4. t2.5 is a tlr2 neutralizing mouse monoclonal antibody. t2.5 blocked pam 3 csk 4 lipopeptide (a tlr1/tlr2-ligand)stimulated nf-κb nuclear translocation and mapk phosphorylation in vitro. in models of pam 3 csk 4 -induced toxic shock and microbial challenge with a high inoculum of heat-inactivated bacillus subtilis, t2.5 prevented lethal shock-like syndrome and increased survival when administered 1 h before or up to 3 h after infection (95) . furthermore, t2.5 used in combination with the 1a6 anti-tlr4/md-2 antibody and antimicrobial therapy protected mice from sepsis caused by s. enterica and e. coli (78) . intracellular antibodies, i.e., intrabodies, have been designed to block the intracellular translocation of tlrs from the endoplasmatic reticulum to the cell surface. αt2ib is a functional anti-tlr2 scfv intrabody comprising the variable domains of the heavy and light chains of t2.5 linked together by a synthetic (gly 4 ser) 3 amino acid sequence. αt2ib bound intracellularly to tlr2 and led to retention and accumulation of tlr2 inside the endoplasmatic reticulum. adenovirus-mediated expression of αt2ib in raw 264.7 macrophages and mouse bone marrow derived macrophages inhibited tlr2 surface expression and tlr2-ligand-driven tnf production (96) . these data suggest for a therapeutic potential of t2.5 or αt2ib in microbial infections. many studies have attempted to elucidate the pathogenesis of acute kidney injury associated with sepsis, which involves mechanisms similar to those occurring during ischemia/reperfusion (97) . damps released during infection are detected through tlr2 by immune cells recruited to the ischemic tissue and/or by cells of the ischemic tissue itself, amplifying the inflammatory response and inducing injury upon reperfusion (98, 99) . blocking tlr2 under these conditions may be cytoprotective (18) . opn-305 is a humanized anti-tlr2 igg4 monoclonal antibody [derived from opn-301 (98) developed by opsona therapeutics (dublin, ireland)]. opn-305 reduced tlr2-driven pro-inflammatory cytokine production through blocking of tlr2/1 and tlr2/6 mediated signaling. in a porcine model of myocardial ischemia/reperfusion injury, pretreatment with opn-305 or administration of opn-305 1 h after ischemia was associated with a 50% decrease in infarct size (100) . results from a first in human phase 1 trial evaluating safety, tolerability, pharmacokinetics, and pharmacodynamics of ascending doses of opn-305 given intravenously in healthy adult subjects have just been released (101) . tlr2 occupancy and inhibition of il-6 secretion induced by heat-killed listeria monocytogenes were assessed in whole blood collected up to 90 days after treatment with either the antibody or placebo. opn-305 was well tolerated, with no significant toxicity even at the highest dose tested. impressively, opn-305 at doses of 0.5 and 10 mg/kg occupied 100% of tlr2 molecules expressed on monocytes collected 14 and 90 days after challenge, respectively. il-6 release was inhibited in a parallel manner. these results suggest that treatment with opn-305 could provide short-term protection against ischemia/reperfusion and be adjusted to confer long-lasting blockage in the case of tlr2-mediated chronic diseases (101) . a phase 2 trial assessing safety, tolerability, and efficacy of opn-305 in kidney transplant patients has been initiated (nct01794663). new techniques are continuously implemented to facilitate the identification of therapeutic targets for adjunctive treatment in sepsis. immunoprecipitation with systematic evolution of ligands by exponential enrichment (selex) was developed to screen and identify high-affinity dna and rna molecules that bind to tlr2 and could be used to detect other molecules influencing tlrdriven activity. a most promising candidate, ap-177, was shown to interact with tlr2, thereby obstructing ligand binding to the receptor and inhibiting tlr2-ligand-induced nf-κb activity and il-6 and il-8 production in thp-1 and hek 293 cells (102) . cellpenetrating tlr2-bb peptides have been generated and shown to interfere with tlr2-ligand-induced activation of nf-κb and mapk and cytokine production (80) . whether these compounds will undergo clinical evaluation is unknown. toll-like receptor 3 is an endosomal prr that senses dsrna typically produced during viral infection (103) . experimental models comparing tlr3 wild-type and tlr3 knockout mice revealed either a protective role (west nile virus, encephalomyocarditis virus, poliovirus, coxsackievirus, murine cytomegalovirus, herpes simplex virus), a deleterious role (west nile virus, influenza a virus, phlebovirus), or no influence (lymphocytic choriomeningitis virus, vesicular stomatitis virus, murine cytomegalovirus, reovirus) of tlr3 on anti-viral responses (104) . therefore, tlr3 agonists and antagonists might be efficient adjunctive therapies for viral infections depending on the context. in the following sections, we describe the development of synthetic dsrna tlr3 agonists (see tlr3 agonists) and of synthetic ssdna tlr3 antagonists and anti-tlr3 neutralizing antibodies (see tlr3 antagonists). the dsrna synthetic analog polyinosinic:polycytidylic acid [poly(i:c)] is a potent immunostimulant. for clinical development, poly(i:c) was stabilized with polylysine and carboxymethylcellulose (poly-iclc) (hiltonol, oncovir, washington, dc, usa) and used to generate poly(i:c 12 u) (rintatolimod, tradename ampligen, hemispherx biopharma, philadelphia, pa, usa) by substituting an uridylic acid at a molar ratio of 12:1 in the synthesis of the polycytidylic acid strand (105) . poly(i:c) and its derivatives have been tested in several clinical trials as adjuvants for vaccines (for both infectious diseases and cancer) and complement to haart (highly active anti-retroviral therapy) in human immunodeficiency virus (hiv) infected patients, topics that we do not discuss here. poly(i:c 12 u) is highly specific for tlr3 and, unlike its parental molecule poly(i:c), does not require melanoma differentiationassociated protein 5 (mda5, a cytosolic prr for viruses), for the induction of the signaling cascade leading to type i ifn production (106) . poly(i:c 12 u) has some anti-viral activity against hiv, hepatitis b virus (hbv), coxsackie b3 virus, and several flaviviruses. results from animal models of lethal respiratory viral infection by severe acute respiratory syndrome coronavirus (sars-cov) and punta toro virus highlighted the favorable impact of intranasal treatment with poly(i:c) or poly(i:c 12 u) on survival and viral loads in infected mice (107, 108) . the mode of action of poly(i:c) in the respiratory tract was linked to the induction of caspase-mediated apoptosis and ros, which are involved in the cleavage and shedding of soluble tnf receptor blocking tnf bioactivity (109) . interestingly, poly(i:c) protected against bacterial infections in the respiratory tract as well as in the central nervous system (cns). in a mouse model of pseudomonas aeruginosa pneumonia secondary to clp, intranasal administration of poly(i:c) improved immune activation and lowered bacterial load in the lungs compared to the untreated animals (110) . corroborating results showing enhanced phagocytosis and killing of e. coli by microglial cells suggest that tlr3 activation is crucial for the immune response of cns against invading pathogens (111, 112) . these data support the development of tlr3 agonists as adjuvant therapies to prevent or reduce the severity of respiratory tract infections caused by viruses and possibly bacteria. in connection with that particular field, a phase 1 safety, tolerability, and pharmacokinetic trial of nasally applied poly-iclc in human volunteers is ongoing and will explore immune activation markers. a phase 1/2 clinical trial is assessing the immunogenicity and safety of flumist®(live attenuated influenza vaccine, med-immune, gaithersburg, md, usa) intranasal influenza vaccine administered with and without a poly(i:c 12 u). more recently, tlr3 antagonists have been developed taking into consideration that tlr3 over-activation by viral dsrna may have detrimental consequences in some situations. indeed, it has been reported that administration of poly(i:c) in mice prior to intratracheal challenge with s. pneumoniae impaired bacterial clearance and increased mortality. excessive production of type i ifn was involved in this phenomenon (113) . single-stranded dna oligonucleotides (ssdna odns) efficiently competed with dsrna for binding to tlr3, thus inhibiting cytokine production and costimulatory molecule expression by epithelial cells, pbmcs, and dendritic cells (114, 115) . the efficacy of ssdna odns was demonstrated in cynomolgus macaques, where intranasal injection of ssdnas odns inhibited poly(i:c)-induced cytokine production in nasal secretions (115) . in addition to microbial ligands, tlr3 senses damps released from injured tissue during inflammation, for example rna from necrotic cells, promoting an excessive inflammatory response. interestingly, administration of a tlr3 neutralizing antibody to mice reduced cecal damage induced by gut ischemia and improved survival of animals with polymicrobial sepsis when the antibody was given 6 and 24 h after clp surgery (116) . collectively, these data demonstrate that tlr3 works as an endogenous sensor of necrosis and a regulator of the immune response, pointing to receptor modulation as a possible adjuvant therapy for sepsis. work remains to be done to clearly delineate the precise role of tlr3 in viral and bacterial infections and to appraise the benefit afforded by tlr3 agonistic or antagonistic strategies for infectious diseases, especially septic shock. a single tlr5 agonist has had clinical development, namely cblb502. flagellin is the only ligand of tlr5 described to date. detailed structural basis of flagellin recognition by tlr5 has been obtained through crystallographic analyses, unraveling a unique mode of interaction between the two molecules as depicted from stoichiometry, ligand arrangement, and binding interfaces (117) . the role of structural constraints for induction of the nf-κb signaling cascade downstream tlr5 was supported by structure-guided mutagenesis and deletion analyses on cblb502 (entolimob), a therapeutic agent derivative of s. enterica flagellin implemented by cleveland biolabs (buffalo, ny, usa). cblb502 is currently tested in a phase 1 trial in late stage cancer patients (nct01527136). several clinical trials have investigated the safety and adjuvant efficacy of recombinant flagellin in a number of vaccine settings (against influenza virus, helicobacter pylori, campylobacter, yersinia pestis, west nile virus, etc) (118) . cblb502 plays a protective role against radiation-induced tissue injury, probably by suppressing apoptosis, attenuating ros generation, and promoting tissue regeneration (119) . these properties could explain the beneficial effect of this agonist in a murine model of acute ischemic renal failure when administered 30 min after reperfusion (120) . highlighting the favorable role of flagellin against tissue damage, results from two mouse studies suggested that protection and repair of the intestinal mucosa that serves as a first line defense barrier is the key mode of action of flagellin. in the first study, flagellin was shown to induce the expression of regiiiγ, a c-type lectin with bactericidal activity, and to restrict small intestine colonization with vancomycin-resistant enterococcus (vre) in animals inoculated with vre via oral gavage (121) . in the second study, treatment with flagellin reduced intestinal epithelium destruction induced by dextran sodium sulfate (a chemical used to induce severe acute colitis) and increased survival of mice inoculated with s. typhimurium by oral gavage (122) . these data suggest that flagellin or tlr5 agonists may represent attractive tools for treating pathologies that injure the intestinal tract, including severe sepsis. no tlr5 antagonists have been reported. toll-like receptor 9 agonists tested in clinical trials are synthetic cpg oligodeoxynucleotides (cpg odns) among which cpg10101, imo-2125, sd-101, and cpg 7909 that mimic unmethylated cpg dinucleotide-rich sequences enriched in microbial dna. cpg odns are powerful immunostimulants, exploited for their adjuvant properties in vaccines against infectious diseases (flu, malaria, hiv infection, pneumococcal and meningococcal diseases) and cancer (melanoma, leukemia, glioblastoma, and colorectal, prostate and breast cancer) (123) . the adjuvant properties of cpg odns have been used to enhance www.frontiersin.org the phagocytosis and the killing of bacteria (s. typhimurium and s. pneumoniae) by phagocytic cells (124, 125) . interestingly, a recent study showed that cpg odn given 1 h prior to clp surgery prevented clp-induced cardiac dysfunction in mice. the authors proposed that targeting tlr9 could be a useful approach for the management of cardiovascular dysfunction in severe sepsis patients (126) . therapeutic strategies focusing on tlr9-mediated immunomodulation are currently being implemented for chronic viral infections, such as chronic hepatitis c (hcv). plasmacytoid dendritic cells are the main cells producing type i ifn and are therefore considered to play an important role in viral infections. tlr9 agonists stimulate plasmacytoid dendritic cells to produce large amounts of type i ifn, especially ifnα, which is the backbone of therapy for hcv. indeed, ifnα powerfully inhibits viral replication and promotes innate and adaptive host immune responses. moreover, ifn production appears to be impaired in plasmacytoid dendritic cells of hcv patients (127, 128) . cpg10101 was originally developed under the trade name actilon by coley pharmaceuticals (wellesley, ma, usa), a company recently incorporated by pfizer (new york city, ny, usa). cpg10101 has undergone two phase 1 studies with promising results. a phase 1a study for drug safety and pharmacokinetics conducted in 48 healthy volunteers revealed well tolerated immunostimulatory effects without serious adverse events even when using high doses of cpg10101 (129) . cpg10101 was also tested in 60 hcv patients, 50 infected with genotype 1 hcv. cpg10101 was administered subcutaneously to four randomized groups at different doses twice per week for 4 weeks alone or in combination with pegylated ifnα and ribavirin. the tlr9 agonistic effect of cpg10101 was associated with the induction of ifnγ and ifnα and the decrease of viral loads. the only serious adverse events were urticaria and pruritis, without manifestation of respiratory complications (130) . a phase 2 study enrolling 113 non-responders genotype 1 hcv patients has been completed, but results have not been released yet. imo-2125, manufactured by idera pharmaceuticals (cambridge, ma, usa), has undergone two phase 1 trials: one for dose estimation and one for safety, pharmacokinetics, and pharmacodynamics, enrolling 60 and 40 treatment-naïve genotype 1 hcv patients, respectively. imo-2125 administration dosedependently decreased viral loads, increased the production of anti-viral cytokines and chemokines especially ifnα, and activated nk and t-cell responses (41, 42) . a phase 2 trial was planned, but in april 2011 the company postponed its initiation. the decision was made based on histological data from a 26-week non-clinical toxicology study of imo-2125 in rodents and non-human primates 4 . preliminary analyses suggested evidence of atypical lymphocytic proliferation, although no adverse events were reported in humans. thorough analysis results are pending. a phase 1b study sponsored by dynavax (berkeley, ca, usa) investigated the safety and efficacy of sd-101 in chronic hcv. sd-101 was administered as monotherapy or in combination with ribavirin to 34 chronically infected, treatment-naïve, genotype 1 hcv patients. results released in 2010 indicated that sd-101 was well tolerated and safe without any serious adverse events. the drug had significant anti-viral activity based on dose-dependent anti-viral response, with 100% of patients at the highest dose showing more than one log reduction in viral load, and increased expression of type i ifn-dependent anti-viral genes (ip-10, mcp-1 mx-b, isg-54k). these data comfort results from in vitro studies showing that sd-101 stimulated human pbmcs to produce 20-fold higher levels of both ifnα and ifnλ in comparison with first-generation tlr9 agonists (40) . bacterial dna released during infection is a mamp, and exuberant activation of tlr9 may participate to the sepsis pathophysiology. hence, drug inhibitors of tlr9 may have therapeutic potential in human sepsis. as a proof of concept, administration up to 12 h after surgery of a single dose of an inhibitory cpg odn blocking tlr9 signaling protected mice from polymicrobial sepsis following clp (131) . hmgb proteins are essential for triggering nucleic acid receptor-mediated innate immune responses. hmgb-binding non-immunogenic-odns have been designed to inhibit hmgb-mediated pathologies. a non-immunogenic odn termed ism odn was tested in a mouse model of endotoxemia. impressively, 70% of mice treated with ism odn survived up to 72 h after lps challenge, while all mice from the control group died within 48 h (132). these data argue for a possible use of non-immunogenic-odns in therapeutic interventions. antimalarial drugs such as chloroquine are well known for their anti-inflammatory properties in autoimmune diseases such as rheumatoid arthritis and systemic lupus erythematosus (133) . chloroquine blunted cytokine production and protected mice from toxic shock induced by cpg odn and lps. chloroquine down-regulated the expression of both tlr9 and tlr4, suggesting that it acts at multiple levels to inhibit inflammation (134, 135) . additionally, when administered 6 h after clp in elderly mice treated with fluids and antimicrobials, chloroquine significantly improved survival, strengthened renal function and protected from multiple organ dysfunction (136) . these results support clinical evaluation of chloroquine in patients with severe sepsis, especially those presenting with acute renal failure. based on its anti-inflammatory activity and because it inhibits endosomal acidification which are important for cell infection by viruses, chloroquine is also tested in multiple trials for prevention and/or treatment of viral infections (hiv, influenza, dengue, chikungunya). yet, the mode of action of chloroquine is multi-factorial and not only through tlr9. toll-like receptor 7 and 8 are closely related tlrs well known for their capacity to recognize ssrna from viruses such as hcv, hbv, hiv, influenza virus, herpes simplex virus, epstein-barr virus, vesicular stomatitis virus, papilloma virus, respiratory syncytial virus, and sendai virus. in agreement, tlr7 is primarily expressed in plasmacytoid dendritic cells. more recently, tlr7 and tlr8 were shown to sense bacterial rna released within phagosomal vacuoles (137) . tlr7 and tlr8 triggering induces potent antiviral immune responses characterized by the production of type frontiers in immunology | microbial immunology i ifns and nf-κb-dependent cytokines. tlr7/8 agonists are primarily developed for treating viral diseases, but also as adjuvants for cancer and infectious disease vaccines. imiquimod (aldara, originally developed by 3m pharmaceuticals, maplewood, mn, usa) is the only tlr7 agonist marketed for anti-viral treatment, i.e., external ano-genital warts caused by human papilloma virus. numerous tlr7/8 agonists are in clinical development, like cl097, isatoribine, ana975, ana773, pf-04878691 (852a), r-848 (resiquimod), and gs-9620. although therapeutic strategies for hcv have evolved in the recent years, quest of new immunomodulatory targets remains mandatory. treatment with new agents such as protease inhibitors appears to be efficient but presents with issues of resistance in the long run (138) . moreover, current protocol therapy with ifnα provides replenishment with a specific subtype of this cytokine. however, use of tlr7 agonists is able to induce a variety of ifn subtypes, possibly providing a more radical and integrated anti-viral activity (139) . finally, administration of tlr7/8 agonists may overcome the adverse events caused by ifnα, like the suppression of granulocyte colony stimulating factor (g-csf) leading to neutropenia. indeed, cl097 reversed ifnα-mediated inhibition of g-csf production by pbmcs obtained from hcv patients and healthy volunteers (140) . cl097 also restored defective cytokine production by myeloid dendritic cells from hiv patients (141) . isatoribine (anadys pharmaceuticals, san diego, ca, usa) was one of the first guanosin analog selective tlr7 agonists to be implemented and tested on humans (142) . intravenous administration of isatoribine to 12 hcv patients during a 7day treatment plan resulted in reduced plasma concentration of hcv rna, regardless virus genotype. adverse events comprised dose-dependent joint pain, decreased white blood cells, and platelets counts, insomnia, and headache (143) . the development of an oral prodrug that could lack the detrimental effects of isatoribine, especially in the gastrointestinal tract, pointed to a new candidate, ana975. preliminary results from a study conducted with ana975 were promising. oral administration of ana975 presented with elevated plasma levels of isatoribine at a concentration able to reduce hcv rna in the plasma of infected patients (144) . unfortunately, anadys pharmaceuticals and novartis (basel, switzerland) announced in 2007 discontinuation of drug development due to unacceptable toxicity in pre-clinical animal studies (145) . subsequent elaboration of an oral prodrug of isatoribine by anadys pharmaceuticals led to the generation of ana773. ana773 exhibits efficient induction of endogenous type i ifns. a double-blind, placebo-controlled study was conducted in 34 patients with chronic hcv, either treatment-naïve or relapsed from ifn-based therapy. interestingly, ana773 was safe and well tolerated and presented only with grade 1 and 2 adverse events. moreover, ana773 dose-dependently decreased hcv rna levels (48) . in another study, repeated treatment with ana773 was associated with transient decrease of myeloid and plasmacytoid dendritic cells and increased levels of ifnα and ip-10 in the blood of patients achieving a reduction in the viral load, suggesting an impairment in ifnα production in the case of non-responders (49) . pf-04878691, formerly known as 852a, is a tlr7 agonist generated by pfizer and implemented so that repeated low doses of the drug would be accompanied with the benefits of the agonistic activity without adverse events. a proof of concept study was conducted to evaluate safety and tolerability of the drug and to determine pharmacokinetics and pharmacodynamics. twentyfour healthy volunteers received orally increasing doses of pf-04878691. pf-04878691 induced immune response in a dosedependent and frequency-related manner. however, two of the subjects exhibited severe lymphopenia along with flu-like symptoms and hypotension (50) . in an attempt to decide on the future perspectives of this compound, a model was used to predict the safety and efficacy of pf-04878691 in hcv patients. this model exploited clinical results from the former study in healthy volunteers along with those reported from the use of cpg10101. further optimization will be required before entering the drug in a phase 2 study (146) . other tlr7/8 agonists have reached phase 3 trials, but demonstrated lack of efficacy and serious adverse effects. when monocytes from hiv patients were stimulated with resiquimod, il-12 secretion was augmented while tnf production was decreased compared to the control group. additionally, hiv replication in cultured monocytes was inhibited (147) . these promising in vitro results were reproduced in a phase 2 trial that enrolled patients with herpes simplex virus type 2. topical application of resiquimod protected from viral lesion spreading (148) . however, a phase 3 trial disclosed lack of efficacy of the drug and, although a phase 2 study for the treatment of hcv demonstrated decreased viral loads, adverse side effects similar to those resulting from ifnα treatment were of serious concern (149). two phase 1 clinical trials for the safety and pharmacokinetics of a novel compound, gs-9620 (gilead sciences, foster city, ca, usa), are currently enrolling treatment-naïve and viral suppressed hbv patients respectively, while another one enrolling hcv patients has been recently approved. gs-9620 is a tlr7 agonist tested originally in hbv infected chimpanzees. drug administration was associated with reduced viral loads both in plasma and liver, probably through enhanced apoptosis of hepatocytes (150) . testing of ascending dosages of the drug in healthy volunteers presented with flu-like adverse events at a dose of 8-12 mg, but cytokine induction was achieved even at administration of 2 mg pointing to a promising adjunctive treatment for chronic viral infections (151). all the above data illustrate the efforts devoted to the development of tlr7/8 agonists for treating viral pathologies. taking into account that bacterial rna triggers tlr7 and tlr8 pathways, one may speculate that tlr7/8 agonists would impact on bacterial sepsis. unfortunately, there are almost no data available on that subject. in one report, intravenous injection of r-848 prior to sepsis induction using the colon ascendens stent peritonitis model increased cytokine release and bacterial clearance, but the effect on survival was not reported (152) . finally, considering that excessive tlr activation induced by viruses or bacteria may have www.frontiersin.org detrimental effects for the host, it might be of interest to generate tlr7/8 antagonists to counteract overwhelming immune activation in conditions of severe infections. targeting tlrs is a promising field for sepsis management and infection control. tlr agonists are widely used to optimize vaccine efficacy, taking advantage of their powerful adjuvancity. whether tlr agonists and antagonists will have such success for treatment therapies, especially for sepsis, has to be established. agonists of tlr3 and tlr7-9 yielded very promising results for treating viral infections. only few studies tested antagonistic drugs. mainly for historical reasons and because of their ligand specificity, tlr4 and to a lesser extent tlr2 are the favorite targets for developing antisepsis drugs. this domain of research has monopolized important resources, but unfortunately many drugs tested in animal models have not entered human studies. those that proceeded, like eritoran and tak-242, did not achieve primary endpoint goal and/or were accompanied with manifestation of adverse events. animal models provide invaluable information about the role of tlrs and the mechanism underlying infection pathogenesis, but lack complexity in terms of comorbidities and host response compared to human disease (153) . so, how to explore more efficiently new treatment strategies? improving the design of clinical studies is mandatory. we should discriminate those patients that could benefit from therapy based on their genotype (for example affecting tlr pathways) and the expression of the targeted molecule, and select homogeneous, well-described population bearing the same underlying conditions and disease severity (68) (69) (70) 154) . ideally, sepsis studies should use specific biomarkers to help patient enrollment and weigh treatment efficacy in realistic conditions (155) . the failure of the recent trials in patients with severe sepsis has taught us valuable lessons regarding patient selection, time of intervention, and follow-up (68) (69) (70) . blocking one single mediator may also not be sufficient to interfere with sepsis. developing sepsis-specific tlr-targeted therapies for patients is a path strewn with obstacles, but an exciting and promising area of 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6-(n-substituted sulfamoyl)cyclohex-1-ene-1-carboxylate analysis of binding site for the novel small-molecule tlr4 signal transduction inhibitor tak-242 and its therapeutic effect on mouse sepsis model the novel selective toll-like receptor 4 signal transduction inhibitor tak-242 prevents endotoxaemia in conscious guinea-pigs combination of imipenem and tak-242, a tolllike receptor 4 signal transduction inhibitor, improves survival in a murine model of polymicrobial sepsis protection from lethal gram-negative bacterial sepsis by targeting toll-like receptor 4 cutting edge: cell surface expression and lipopolysaccharide signaling via the toll-like receptor 4-md-2 complex on mouse peritoneal macrophages tlr4/md-2 monoclonal antibody therapy affords protection in experimental models of septic shock tlr4-induced ifn-gamma production increases tlr2 sensitivity and drives gramnegative sepsis in mice lipopolysaccharide-trap-fc, a multifunctional agent to battle gram-negative bacteria cutting edge: differential inhibition of tlr signaling pathways by cell-permeable peptides representing bb loops of tlrs targeting tlr4 signaling by tlr4 toll/il-1 receptor domain-derived decoy peptides: identification of the tlr4 toll/il-1 receptor domain dimerization interface immunity to fungal infections interplay between candida albicans and the mammalian innate host defense genetic susceptibility to candida infections the role of toll-like receptor (tlr) 2 and tlr4 in the host defense against disseminated candidiasis md-2, a molecule that confers lipopolysaccharide responsiveness on toll-like receptor human epithelial cells establish direct antifungal defense through tlr4-mediated signaling involvement of cd14 and toll-like receptors in activation of human monocytes by aspergillus fumigatus hyphae identification of oxidative stress and toll-like receptor 4 signaling as a key pathway of acute lung injury the tlr4 antagonist eritoran protects mice from lethal influenza infection the role of tlr2 in infection and immunity human tlr10 is a functional receptor, expressed by b cells and plasmacytoid dendritic cells, which activates gene transcription through myd88 crystal structure of the tlr1-tlr2 heterodimer induced by binding of a tri-acylated lipopeptide recognition of lipopeptide patterns by toll-like receptor 2-toll-like receptor 6 heterodimer antagonistic antibody prevents toll-like receptor 2-driven lethal shock-like syndromes generation of anti-tlr2 intrabody mediating inhibition of macrophage surface tlr2 expression and tlr2-driven cell activation searching for mechanisms that matter in early septic acute kidney injury: an experimental study myocardial ischemia/reperfusion injury is mediated by leukocytic tolllike receptor-2 and reduced by systemic administration of a novel anti-toll-like receptor-2 antibody ischaemia-induced up-regulation of toll-like receptor 2 in circulating monocytes in cardiogenic shock treatment with opn-305, a humanized anti-toll-like receptor-2 antibody, reduces myocardial ischemia/reperfusion injury in pigs randomized, double-blind, placebo-controlled, dose-escalating phase i, healthy subjects study of intravenous opn-305, a humanized anti-tlr-2 antibody identification and characterization of oligonucleotides that inhibit toll-like receptor 2-associated immune responses toll-like receptor, rig-i-like receptors and the nlrp3 inflammasome: key modulators of innate immune responses to double-stranded rna viruses antiviral responses induced by the tlr3 pathway structural requirements of the ri n-rc n complex for induction of human interferon essential role of mda-5 in type i ifn responses to polyriboinosinic:polyribocytidylic acid and encephalomyocarditis picornavirus tlr3 is essential for the induction of protective immunity against punta toro virus infection by the double-stranded rna (dsrna), poly(i:c12u), but not poly(i:c): differential recognition of synthetic dsrna molecules intranasal treatment with poly(i*c) 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an adjuvant: cellular mechanisms and potential an agonist of toll-like receptor 5 has radioprotective activity in mouse and primate models tlr5 agonist inhibits acute renal ischemic failure bacterial flagellin stimulates toll-like receptor 5-dependent defense against vancomycin-resistant enterococcus infection flagellin treatment protects against chemicals, bacteria, viruses, and radiation cpg still rocks! update on an accidental drug tlr 9 activation in dendritic cells enhances salmonella killing and antigen presentation via involvement of the reactive oxygen species toll-like receptor stimulation enhances phagocytosis and intracellular killing of nonencapsulated and encapsulated streptococcus pneumoniae by murine microglia the toll-like receptor 9 ligand, cpg oligodeoxynucleotide, attenuates cardiac dysfunction in polymicrobial sepsis, involving activation of both phosphoinositide 3 kinase/akt and extracellular-signal-related kinase signaling rational design of new cpg 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proinflammatory cytokine release chloroquine and inhibition of toll-like receptor 9 protect from sepsisinduced acute kidney injury detection of prokaryotic mrna signifies microbial viability and promotes immunity telaprevir and pegylated interferon-alpha-2a inhibit wild-type and resistant genotype 1 hepatitis c virus replication in patients stimulation of interferon and cytokine gene expression by imiquimod and stimulation by sendai virus utilize similar signal transduction pathways interferon-alpha suppressed granulocyte colony stimulating factor production is reversed by cl097, a tlr7/8 agonist tlr7/tlr8 activation restores defective cytokine secretion by myeloid dendritic cells but not by plasmacytoid dendritic cells in hiv-infected pregnant women and newborns molecular basis for the immunostimulatory activity of guanine nucleoside analogs: activation of toll-like receptor 7 isatoribine, an agonist of tlr7, reduces plasma virus concentration in chronic hepatitis c infection discovery of ana975: an oral prodrug of the tlr-7 agonist isatoribine masked oral prodrugs of toll-like receptor 7 agonists: a new approach for the treatment of infectious disease use of modelling and simulation techniques to support decision making on the progression of pf-04878691, a tlr7 agonist being developed for hepatitis c r-848 triggers the expression of tlr7/8 and suppresses hiv replication in monocytes topical resiquimod 0.01% gel decreases herpes simplex virus type 2 genital shedding: a randomized, controlled trial oral resiquimod in chronic hcv infection: safety and efficacy in 2 placebocontrolled, double-blind phase iia studies gs-9620, an oral agonist of toll-like receptor-7, induces prolonged suppression of hepatitis b virus in chronically infected chimpanzees safety, pharmacokinetics and pharmacodynamics of gs-9620, an oral toll-like receptor 7 agonist stimulation of tlr7 prior to polymicrobial sepsis improves the immune control of the inflammatory response in adult mice the search for effective therapy for sepsis: back to the drawing board? sepsis: time to reconsider the concept biomarkers in sepsis we would like to thank all our collaborators that participated in our studies. thierry roger is supported through grants from the swiss national science foundation (310000_114073, 310030_132744, 310030_138488, and 310030_146838), an interdisciplinary grant from the faculty of biology and medicine of the university of lausanne (switzerland) and the european com the authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. frontiers in immunology | microbial immunology key: cord-297039-vfuem6bk authors: beltrán-garcía, jesús; osca-verdegal, rebeca; nacher-sendra, elena; pallardó, federico v.; garcía-giménez, josé luis title: circular rnas in sepsis: biogenesis, function, and clinical significance date: 2020-06-25 journal: cells doi: 10.3390/cells9061544 sha: doc_id: 297039 cord_uid: vfuem6bk sepsis is a life-threatening condition that occurs when the body responds to an infection that damages it is own tissues. the major problem in sepsis is rapid, vital status deterioration in patients, which can progress to septic shock with multiple organ failure if not properly treated. as there are no specific treatments, early diagnosis is mandatory to reduce high mortality. despite more than 170 different biomarkers being postulated, early sepsis diagnosis and prognosis remain a challenge for clinicians. recent findings propose that circular rnas (circrnas) may play a prominent role in regulating the patients’ immune system against different pathogens, including bacteria and viruses. mounting evidence also suggests that the misregulation of circrnas is an early event in a wide range of diseases, including sepsis. despite circrna levels being altered in sepsis, the specific mechanisms controlling the dysregulation of these noncoding rnas are not completely elucidated, although many factors are known to affect circrna biogenesis. therefore, there is a need to explore the molecular pathways that lead to this disorder. this review describes the role of this new class of regulatory rnas in sepsis and the feasibility of using circrnas as diagnostic biomarkers for sepsis, opening up new avenues for circrna-based medicine. sepsis is a syndrome caused by the altered regulation of the host's immune response to infection, which can quickly evolve into a syndrome of multi-organ dysfunction, and finally death, if effective treatments are not immediately applied. in the third international consensus on sepsis and septic shock (ss), it was defined as "a life-threatening condition that arises when the body's response to infection damages the host's own tissues." ss is "a subset of sepsis in which particularly profound circulatory, cellular, and metabolic abnormalities are associated with a higher risk of mortality than sepsis" [1] . sepsis is one of the most common illnesses worldwide. its incidence was 677.5 (535.7-876.1) cases per 100,000 in the world in 2017 [2] , and it is increasing at a rate of 9% per year. approximately 2% of hospitalized patients and up to 75% of patients in intensive care units (icu) develop sepsis, of which around 30% enter into ss [3] . despite advances in antibiotic therapy and treatments applied in icu, sepsis has become a global problem, and is the leading cause of death in all icus worldwide. this review analyzes how these regulatory rnas play an important role in the pathophysiology of sepsis, and we propose some circrnas as feasible clinical biomarkers. furthermore, circrnas have a dual role, acting as therapeutic agents and as therapeutic targets. the multimodal functioning of circrnas opens new avenues to improve diagnosis and prognosis and increase therapeutic strategies against sepsis, thereby reducing the high sepsis-associated mortality. circrnas are produced mainly by the transcription of protein-coding gene' exons by rna polymerase ii (rna-pol ii) [22, 43] , but they can also contain introns [44, 45] . this kind of rna is not usually generated by the same canonical rna splicing as linear rna [43, [46] [47] [48] [49] . circrnas are produced by a pre-mrna back-splicing process, which is mediated by a spliceosome, and is able to link an upstream acceptor splice site (3 splice site) to a downstream splice donor site (5 splice site) on the same exon or others [12, 15, 29, 43, [49] [50] [51] (figure 1 ). circrnas biogenesis is typically done at the expense of canonical mrna isoforms, which suggests that circrnas compete with the maturation of their linear counterparts. so, circrnas appear to be important regulators of mrna production [29, 43, 51] . in addition, a single gene locus is able to generate many circrnas through alternative back-splice site selection, compared to canonical rna splicing [18] . this fact contributes to producing lower concentrations of linear rnas with the original function ("parental linear rna"). therefore, due to the back-splicing process, a different molecule of linear rna forms with a changing function that has a substantial biological impact [20, 52, 53] . jeck et al. have shown this to be the case in human fibroblasts, which express 10-fold more circrnas than their linear rna counterparts [14] . have a dual role, acting as therapeutic agents and as therapeutic targets. the multimodal functioning of circrnas opens new avenues to improve diagnosis and prognosis and increase therapeutic strategies against sepsis, thereby reducing the high sepsis-associated mortality. circrnas are produced mainly by the transcription of protein-coding gene' exons by rna polymerase ii (rna-pol ii) [22, 43] , but they can also contain introns [44, 45] . this kind of rna is not usually generated by the same canonical rna splicing as linear rna [43, [46] [47] [48] [49] . circrnas are produced by a pre-mrna back-splicing process, which is mediated by a spliceosome, and is able to link an upstream acceptor splice site (3′ splice site) to a downstream splice donor site (5′ splice site) on the same exon or others [12, 15, 29, 43, [49] [50] [51] (figure 1 ). circrnas biogenesis is typically done at the expense of canonical mrna isoforms, which suggests that circrnas compete with the maturation of their linear counterparts. so, circrnas appear to be important regulators of mrna production [29, 43, 51] . in addition, a single gene locus is able to generate many circrnas through alternative back-splice site selection, compared to canonical rna splicing [18] . this fact contributes to producing lower concentrations of linear rnas with the original function ("parental linear rna"). therefore, due to the back-splicing process, a different molecule of linear rna forms with a changing function that has a substantial biological impact [20, 52, 53] . jeck et al. have shown this to be the case in human fibroblasts, which express 10-fold more circrnas than their linear rna counterparts [14] . there are three classes of circrnas, depending on the pre-mrna material from which circrnas are made: (1) exonic circrna, which derives from back-spliced exons (ecircrna); (2) circular intronic rna (cirna), which originates from spliced introns; and (3) exon-intron circrna (eicirna), which come from circrna containing both exons and introns [54] . it has been reported that ecircrnas are exported to the cytoplasm (where they are most often detected), while some cirna and eicirna are retained in the nucleus [47, 55, 56] . jeck et al. proposed that circrnas can be produced through two different pathways. the first is called lariat-driven circularization (figure 2a) , and is associated with an "exon skipping" process, which consists of a covalent splice from the 5 end site (donor) to the 3 end site (acceptor) [14, 15, 29, 43, 49, 50, 57] . afterwards, the lariat is joined to the spliceosome by removing introns and forming an exonic circle [10] . the second pathway proposed by jeck et al. is known as intron-pairing driven circularization (figure 2b) , and is based on pairing complementary motifs in transcripts. alu elements are suggested as important regulators of circrna biogenesis, but other inversed repetitive sequences are also adequate to drive rna circularization [10, 14, 49] . some studies indicate that the intron-pairing-driven model might occur more frequently than lariat-driven circularization [44] . another process exists and is able to form circrnas by joining rna molecules flanking introns through rbps. among these proteins are, for example, quaking protein (qki) and muscleblind protein (mbl) [28, 43] (figure 2c) . thus, the qki protein is responsible for regulating circrna abundance in human cells, through the binding of the 3 and 5 ends of circularized exons, and is also responsible for facilitating dimerization and mediating their splicing [28] . other studies in human cells have shown that circrnas can be formed by intron flanking joining mechanisms because of debranching failure [58] ( figure 2d ). it has been demonstrated that during sepsis, there is an alteration in the alternative rna splicing patterns, which results in the disturbance of the patient's immune response, mainly due to the attenuation of b and t lymphocytes [59] , contributing to critical phenotypes. although it is not very clear how the alteration in the alternative splicing process occurs during sepsis, it is known that alternative splicing is a key process in the generation of circrnas. moreover, it has been shown that small changes in the process critically affect their biogenesis, affecting the concentration of some circrnas by modulating their expression [60] . furthermore, changes in alternative splicing patterns can also give rise to different circrnas, modulating a wide range of molecular mechanisms and altering their physiological state [43, 61] . due to the role that circrnas play in the modulation of different cytokines and immune proteins [62, 63] , altered states of alternative splicing in sepsis may alter the expression of circrnas, which could partially explain the changes in the immune response of septic patients. however, this is still an unexplored field, so, although it has been attracting a lot of interest in recent years, more studies are required to demonstrate how the alteration of the alternative splicing may affect the biogenesis of circrna during sepsis. in this regard, the fact that global transcription and translation profiles are altered during sepsis [64] lends support to the idea that changes in alternative splicing patterns may play a key role in sepsis, perhaps by modulating circrnas biogenesis. interestingly, circrnas are aberrantly expressed in many diseases and exhibit roles, such as mirna sponges, protein decoys, transcription regulators, and regulators of translation. despite the specific mechanisms involved in the dysregulation of circrnas in different pathologies like sepsis, they are not completely understood, because distinct factors contribute to this dysregulation, such as their biogenesis from parental genes, export from the nucleus to the cytoplasm, and cell removal, among others. in line with this, aberrant cis-elements seem to be important regulatory components in circrnas production, especially in humans [43] . abnormal spliceosomal machinery and aberrant transactivating factors have also been postulated as central players in the biogenesis of circrnas [28, 43, 65] . in this regard, despite the initial research concluding that splicing events occur co-transcriptionally in most cells and tissues [66, 67] , recent research demonstrates that the majority of circularizations occur post-transcriptionally [18, 68] , and splicing and transcription elongation are mutually dependent [69] . furthermore, several mechanisms have been put forward to explain the role of the epigenetic aberrations involved in the dysregulation of circrnas. accordingly, dna methylation, chromatin remodeling, and post-translational modifications of histones directly impact circrnas production [18, 68] , for example by controlling different alternative splicing events during circrnas biogenesis [12] . the biogenesis process of circrnas inherently lowers parental rna levels, which may lead to a reduction in mrna and, in turn, to low levels of translated proteins [59] , thereby producing the deregulation of a wide range of cellular processes. finally, circrnas can also be produced from intronic sequences with no clearly defined function, which may not lead to the physiological worsening associated with low specific linear rna levels, but then again, may do so, because it is extremely uncommon, and this biogenesis type is expected to occur in a lower proportion than others. e2 the biogenesis process of circrnas inherently lowers parental rna levels, which may lead to a reduction in mrna and, in turn, to low levels of translated proteins [59] , thereby producing the deregulation of a wide range of cellular processes. finally, circrnas can also be produced from intronic sequences with no clearly defined function, which may not lead to the physiological worsening associated with low specific linear rna levels, but then again, may do so, because it is extremely uncommon, and this biogenesis type is expected to occur in a lower proportion than others. as some of the above-described mechanisms are altered in sepsis, it is plausible to hypothesize that abnormal levels of circrnas and low levels of linear rnas concentrations due to the inherent biogenesis process of circrnas may affect the abundance of the various immune mediators and transcriptional factors involved in the inflammation and immune response. we will consider this below. one of the main reasons for the late discovery of circrnas lies in the extreme difficulty in finding them, mainly because it is hard to distinguish them from other small rnas, such as mirnas. fortunately, rna-seq and bioinformatics may help us to understand the pathways that produce circrnas and how these pathways modulate the different molecular responses in a wide range of diseases [14, 15, 22, 53] . current detection methodologies require circrnas to lose their circularity in order to detect them. since circrnas were discovered, several tools have been developed to analyze their expression, and to validate that circrnas indeed exist [70] . in parallel to molecular strategies, bioinformatic tools have also been developed to identify new circrnas and to quantify their expression with high fidelity. jeck et al. published a new protocol in nat biotechnol [44] called circle-seq, which consists of using the rnase r enzyme to process linear rnas, while circrnas remain intact. it has recently been shown that it is very difficult to determine the circularity of an rna transcript by using only this treatment, because some circrnas are sensitive to this enzyme [33] , which could cause false-negatives and lead to biases. the use of other, additional methodologies to isolate circrnas has also been proposed, such as 2d (two-dimensional) denaturing polyacrylamide gel electrophoresis or ribosomal rna (rrna) depletion and poly(a) depletion, to increase the amount of circrnas in samples for rna-seq [53] . nonetheless, their actual efficacy in clinical practice is unknown. the most widely used method to validate and quantify circrnas is reverse transcription-pcr (rt-qpcr). this method is implemented by using, for example, a strategy based on rna treatment with and without rnase r, followed by a step of reverse transcription to cdna. the cdnas from rna with or without rnase r treatment are then analyzed by pcr amplification, with primers specifically designed for each isoform by a pcr reaction to detect the presence of both circrnas and specific circrnas through the design of specific primers, which are not generated by the normal splicing occurring during mrna and other small regulatory rnas processing [71] . the great advantage of this method is that it may be widely used in clinical practice, and its implementation as a diagnostic biomarker detection tool is simple and cheap. the use of a microarray as a diagnostic method is also possible, thanks to its sensitivity and specificity. however, there is some concern about this approach, as only the circrnas included in the array can be evaluated, which means that newly discovered circrnas cannot be included with microarray technology. thus, it is extremely difficult to determine the absolute amount of circrnas. hence, microarrays are a good approach for determining the relative expression levels of circrnas in comparisons made of different exploratory groups. rna-sequencing (rna-seq), coupled with directed bioinformatic analysis, has notably contributed to discovering and characterizing circrnas. in fact, rna-seq has yielded many circrnas and contributed to the discovery of the intrinsic characteristics of circrnas [14, 29, 44, 72, 73] . more importantly, disease-relevant circrnas can be detected in human peripheral whole blood by rna-seq [74, 75] . in order to overcome the challenge of discovering new circrnas that contribute to comprehend and diagnose sepsis, a number of bioinformatic and statistical methods have been described. moreover, many tools have been designed to decipher whether resulting circrnas are exonic, intergenic, intronic, or utr [15, 29, 76] . the results have been deposited in specialized databases like circbase [77] and circpedia [18] . some currently used bioinformatic tools are designed to process rna-seq data and to identify circrnas [78, 79] . however, despite the central role that different predictive software and algorithms can play in discovering circrnas, their implementation in clinical practice is still a distant solution. however, their vast potential is undeniable, and they are expected to be implemented in clinical routine in the coming years, with the support of the latest technological advances. moreover, several tools and databases are appearing which contribute to the understanding of the different functions of circrnas and the role they can play in different diseases [80] . these tools help to identify the circrnas that competitively sequester mirnas, by preventing them from interacting with their natural targets. this process is highly relevant in sepsis, because mirnas are key regulators in inflammation, endothelial dysfunction, and immunosuppression during sepsis [81, 82] . furthermore, there is an evident need to understand the role that circrnas play in sepsis. future research is required to uncover not only the molecular role of different circrnas related directly and indirectly to sepsis, but also the new circrnas that modulate the great heterogeneity of sepsis, which can then be used as biomarkers of diagnosis, prognosis, and/or theragnosis. very little information is available about the role of circrnas in sepsis, because the elucidation of the role that these molecules play in human diseases has become relevant only in recent years. however, circrnas may play a key role in sepsis because of their ability to modulate different molecular mechanisms [10] , including inflammation [83] and immune response [62] , and to control multiple biological processes in metabolic organs (i.e., liver, pancreas [84] ) ( figure 3 and table 1 ). moreover, the identification of the mechanism by which host circrnas can bind viral mrnas merits special attention, because it indicates that circrnas are likely to resist viral infection [85] (figure 3 ). therefore, circrnas may play a key role in host defense against viruses. it is known that one of the first molecular responses in sepsis is the "cytokine storm". during sepsis, the "cytokine storm" mediates the initial pro-inflammatory phase by releasing proinflammatory cytokines, such as il-1α, il-1β, tumor necrosis factor-α (tnf-α), il-6, and interferon gamma (ifn-γ), among others [86] . nevertheless, during a septic process, anti-inflammatory cytokines (i.e., il-10, il-30, transforming growth factor-β (tgf-β), etc.) are also produced and continuously released, strongly influencing the sepsis progression and outcome [87] . the importance of mirnas in controlling sepsis pathophysiology has been demonstrated. in fact, both host mirnas and dna virus-encoded mirnas are involved in the sepsis-induced cytokine storm, leading to increased inflammation, and even to subsequent immunosuppression. in fact, it has been widely demonstrated that mirnas are able to regulate different key cytokines expressed during sepsis and significantly mediate their expression as tnf-α [88] [89] [90] , il-6 [91, 92] , nfκb [93] [94] [95] , il-10 [87] , il-18 [96, 97] , il-27 [98, 99] , and other pro-inflammatory and anti-inflammatory cytokines with differential expressions in sepsis. although very little information exists about circrnas in sepsis, it may be hypothesized that the regulation exerted by circrnas plays a fundamental role in different sepsis stages (figure 3 ). in fact, circrnas control the expression of the key proteins and cytokines that participate in sepsis ( figure 3 , table 1 ). for example, circ-4099 is induced by inflammatory mediators, such as tnf-α [100] and circrna_0038644, and has been demonstrated to regulate the expression of nf-κb in sepsis [101] , while il-6 is regulated by circ_007893 [102] . moreover, circrna_0005105 facilitates the expression of inflammatory cytokines [103] , thereby mediating a pro-inflammatory phenotype which is critical in sepsis pathophysiology. the findings of zhang et al. support the role of circrnas in mediating inflammation, by showing how circ_0012919 was abnormally up-regulated in cd4 + t cells in patients with a hyper-inflammatory syndrome, such as systemic lupus erythematosus (sle). circ_0012919 also increases the expression of dna methyl-transferase 1 (dnmt1), modulating the immune response by reducing the expression of cd70 and cd11 in cd4 + t [104] . interestingly, circrnas may function as "molecular sponges", by controlling the expression of different types of non-coding rnas, such as mirnas, involved in regulating different processes in sepsis [105] [106] [107] [108] . one of the best characterized mirnas in sepsis is inflamma-mir mir-223, which participates in the regulation of innate immunity by controlling the activation and differentiation of neutrophils and macrophages [109] . the down-regulation of mir-223-3p induces the expression of il-6, il-1β, and tnf-α, which supports the key role played by mir-223 in innate immunity regulation [110] . interestingly, circ_0003159 has recently been shown to be able to regulate the expression of mir-223 [111] , providing a new mechanism to explain the low levels of this mirna in sepsis patients [112] . another inflamma-mir described in sepsis is mir-146a, which is involved in several key processes in sepsis, such as the control of innate immunity, endotoxin tolerance and immunosuppression, inflammatory response, antiviral pathways, toll-like receptors (tlrs), and cytokine signaling [113] . importantly, mir-146a has proven its ability to predict 30-day mortality in septic patients [114] . in this scenario, two circrnas, namely circ-102685 and circ-rsf1, modulate the expression of mir-146a [115, 116] . the two circrnas may be postulated to be modulators of inflamma-mirs, thereby suggesting their role in inflammation during sepsis. chen j. et al. have demonstrated that circ-pvt1 binds mir-125 family mirnas to inhibit their function [117] . this family is especially interesting in sepsis, because mir-125b correlates with sepsis severity, inflammation, and increased mortality in septic patients [118] . in fact, circ-gli2 is a circrna that specifically targets mir-125b-5p [119] . mir-192-5p, mir-26a, and mir-191-5p have been postulated as key biomarkers in sepsis, for their ability to discriminate between sepsis and other severe inflammatory cases, for instance severe systemic inflammatory response syndrome (sirs) [120] . zhongrong z. et al. revealed that in intervertebral disc degeneration, circ-msr regulates the expression of mir-27 [121] , which is up-regulated and promotes an inflammatory response in sepsis [122] . different non-coding rnas have been postulated as key mediators of sepsis, for their ability to control the innate and adaptive immune system, in addition to simultaneous pro-and anti-inflammatory phenotypes ( figure 3 and table 1 ). besides the role of some circrnas in regulating inflammation, other mirnas may also regulate the immune system. for example, circ_0005075 is involved in the regulation of mir-23a-5p and mir-23b-5p, which control key events in sepsis pathophysiology. moreover, mir-23a-5p is up-regulated in sepsis, contributing to acute respiratory distress syndrome induced by lipopolysaccharide (lps) [123] . furthermore, mir-23b-5p up-regulation controls t-cell apoptosis through nf-κb signaling, as demonstrated in a mouse model of sepsis [124] . interestingly, the inhibition of mir-23b-5p causes down-regulation in programmed death ligand 1 (pd-l1) expression in splenic t-lymphocytes from septic mice, by reducing late-sepsis-induced immunosuppression and improving survival [124] . these results suggest that circ_0005075 could be used in a possible therapeutic approach in sepsis, as suggested previously for cancer therapy [125] . a recent study explored the expression of circrnas in macrophages under two different polarization conditions: m1 macrophages induced by ifn-γ and lps, and m2 macrophages induced by interleukin-4 (il-4). the results showed 189 circrnas with differential expression in the m1 compared to m2 macrophages, which have a relevant role in producing anti-inflammatory cytokines such as il-10 and il-13. of the 189 identified circrnas, circ-010231 was the most overexpressed circrna in m1 after lps stimulation [126] , which contributed to m1 to m2 polarization of macrophages and suggests that this circrna is a good candidate to be explored in human sepsis. therefore, knowledge of the entire molecular process contributing to m1 to m2 transition is extremely important, because it opens new therapeutic possibilities by controlling immune responses and immunosuppression. another circrna, circ_0005785, can bind to mir-181a and mir-181b-two mirnas which have been previously postulated as promoters of immunosuppression in late sepsis [127] . finally, circ_man2b2 regulates s100a8, which is another important protein involved in immunosuppressive states in sepsis [128] . impairment of the endothelial function is one of the most important physiopathological hallmarks of sepsis [129] [130] [131] . endothelial function is critical for maintaining vascular homeostasis and activating different processes, such as thrombosis, inflammation, and vascular remodeling [132] . hence, it has been shown that the outcome of sepsis substantially improves when endothelial dysfunction is avoided [133] [134] [135] . regarding inflammatory phenotypes and the endothelium, liu et al. found that the expression of circ-cer was up-regulated in chondrocyte with catabolic stimulators like il-1 and tnf-α [136] . moreover, wu et al. observed how circ-0005105 stimulated the expression of nicotinamide phosphoribosyltransferase (nampt), and the generation of il-6, il-8 and prostaglandin e2, due to the down-regulation of mir-26a expression [103] , while il-1β promotes circ-0005105 expression in a chondrocyte extracellular matrix degradation model [103] (figure 3 and table 1 ). in a recent study, tie-ning et al. counted 11 circrnas with associated differential expressions in an lps-induced rat ss model with septic myocardial depression [137] , one of the main causes of death associated with sepsis when the endothelium plays a key role. one of the most plausible biomarkers for sepsis is mcircrasgef1b. this circrna regulates the stability of mature icam1 mrna and can protect cells against microbial infection (figure 3) . in fact, a recent study revealed that mcircrasgef1b is induced after lps stimulus during microbial infection [138] . the mechanisms in endothelial cells may be similar to those in lps-stimulated mouse macrophages, in which ng et al. observed that only mcircrasgef1b was regulated by the tlr4 pathway [138] . interacts with the mrna of pro-inflammatory cytokines induces a pro-inflammatory phenotype [103] circ rna-cer induced in chondrocytes by il-1 and tnfα mediates an inflammatory response through interaction with il-1 and tnfα [136] circ_0028644 regulates the expression of nf-κb modulates different pro-inflammatory phenotypes [101] circ_4099 modulates the expression of mir-616-5p closely related to inflammatory phenotypes through tnf-α [100] circ_0003159 regulates mir-223 induces an inflammatory response due to increased expression of il-6, il-1β, and tnf-α [111] circ_rsf1 regulates the expression of inflammatory cytokines represses the interactions of mir-146a with rac1 by eliminating its inhibitory effect on the rac1 pathway [115] circ_102685 modulates the expression of mir-146a plays a role in endotoxin tolerance, immunosuppression, inflammatory response, and antiviral pathways [116] circ_0005075 regulates mir-23a-5p and mir-23b-5p suppresses the expression of mir-23b-5p in cancer and is related to immune response [123] [124] [125] circ-pvt1 interacts with the mir-125 family exhibits a possible correlation with sepsis severity, inflammation, and increased mortality [117] circ-gli2 negatively regulates the expression of mir-125b-5p involved in inflammation and immune response pathways [119] circ-mylk and circ_ctdp1 regulates mir-29a-3p feasible predictive biomarker for assessing 28-day mortality of sepsis patients [139, 140] regulates mir-29a-3p and mir-29c-3p, and matrix metrix metalloproteinases mmp-9 and mmp-2 increases mirnas stability [141] circ_nsd2 regulates different processes through sponge mir-199b related to the low mir-199b-5p levels found in sepsis patients [142] circ_0005785 regulates mir-181a and mir-181b possible role in sepsis by promoting immunosuppression in late sepsis [127] circ_0000096 regulates the expression of mir-224 and mir-200a modulates the immune response through cyclin d1, cdk6, mmp-2, and mmp-9 [143] circ_001569 modulates the expression of mir-145 involved in the immune response of host to pathogens [139, 140] circ_hipk3 modulates the expression of mir-193a-3 and mir-124 mediates a pro-inflammatory state by modeling the inflammatory response through sponge mir-124 (inhibitor of il-6) [144] circ_0003528, circ_0007196 and circ_0078738 interacts with mir-192-5p related to the low levels found in sepsis patients [145] circ rna-9119 modulates mir-26a increases the expression of ptgs2 by modulating the response of endothelium [146] circ_trim33 and circ_foxo3 modulates the expression of mir-191 and induces the expression of tet1 induces proliferation, migration, and immune regulation [147] [148] [149] circ rna_007893 regulates the expression of il-6 regulates the expression of il-6, through sponging and endogenous mir-485-5p [102] circ man2b2 regulates the expression of s1000a8 modulates immunosuppressive states [128] circ rna-msr modulates mir-27 induces pro-inflammatory phenotype [121] circ_0012919 increases the expression of dnmt1 modulates immune response by reducing the expression of cd70 and cd11a in cd4 + t cells [104, 150] infections are very common in young and old people worldwide. in most people, the host's immune response suffices to deal with a potential threat, but in some cases, infection may be associated with an inadequate or inappropriate host response, by mediating the development of sepsis [1, 148, 151, 152] . as there is no specific treatment for septic patients, their management is based on attempting to control infection and supporting the different organs whose functions may be compromised [153] . in fact, one of the biggest problems is that patients deteriorate quickly, progressing to ss and multiorgan failure if not treated promptly and effectively. it is noteworthy that early treatment has been shown to improve patient outcomes [153] [154] [155] , but early treatment depends on promptly recognizing and diagnosing sepsis, which may contribute to rapidly starting the appropriate therapy [153] . nevertheless, the early diagnosis of septic patients remains a challenge for clinicians and researchers around the world. because of all the aforementioned difficulties in making a correct early sepsis diagnosis, the availability of precise biomarkers would be extremely useful to allow proper and timely treatment to start, thus maximizing the possibilities of patient survival. to date, more than 170 biomarkers have been proposed and clinically evaluated [7] , including various cytokines, receptors, cell surface markers, coagulation factors, complement factors, and acute phase reactants, among many others [156] [157] [158] , but none offer anything near 100% specificity for sepsis. apparently, circrna modulates a wide range of molecular responses related with immune system control, inflammation, and endothelial function, which are relevant biological processes altered in sepsis. in this regard, the dysregulation of specific circrnas has been related to the development and progression of sepsis [126, 137, 138] . a number of studies report associations between circrnas and almost all the mirnas are postulated as biomarkers in sepsis, most notably with the different cytokines that have an abnormal expression in sepsis, inducing the characteristic "cytokine storm" which contributes to multiorgan failure. in light of this, circ_001569 directly inhibits the transcription of mir-145 [140] , thereby controlling tgfbr2 levels in lung tissues [159] and playing a direct role in the molecular signaling responsible for sepsis-induced acute lung injury. other circrnas, such as circ_hiat1, bind to mir-29a-3p and mir-29c-3p. in this case, circ-hiat1, also known as circ_0000096, performs a "mir reservoir" function by increasing mir stability in human atherosclerosis and some cancers [141] , unlike the classic function of circrnas that act as an "mir sponge". interestingly, circ-hiat1 also targets matrix metalloproteinase (mpp)-2, and mmp-9 in solid tumors [160] , which are elevated in the plasma [161] and lung tissue [162] of patients with severe sepsis. notably, circ-mylk and circ-ctdp1 are also able to target mir-29a-3p [163, 164] . the targeting of mir-29a-3p is of the utmost importance, because high serum levels of this mirna produced by immune cells have a good predictive value when assessing the 28-day mortality of sepsis patients (table 1) . circ_nsd2 is another interesting circrna to be explored in sepsis, because it regulates different processes through the sponging of mir-199b [142] , a mirna with higher levels found in total serum and blood cells (leukocytes, erythrocytes, platelets), and previously demonstrated to possess the early ability to discriminate ss from sepsis patients [165] . another important point to consider is that sepsis is usually associated with bacterial infections, but it can also be induced by viral and fungal infections, although the inflammatory response is generally less marked in these cases. in particular, viral sepsis lacks a definite diagnostic criterion [166] . so, circrnas could represent a major achievement in viral sepsis diagnosis and offer several advantages over other biomarkers. for example, circrnas remain stable in the presence of viral infections. in fact, competitive binding has been demonstrated between host circrna and viral mrnas, which indicates that circrnas participate in the host defense against viral infections [85] . these findings seem to indicate that specific circrnas are expressed in the presence of viral infections by postulating circrna as good biomarkers to diagnose the origin of sepsis and establish a prognosis for septic patients. moreover, circrnas are both tissue-specific and developmental stage-specific; their expression has been related to the initiation and progression of many disease types, such as neurological, cardiovascular, and cancer [38] [39] [40] . given their structural conformation (covalently closed loop that lacks free 3 and 5 ends), circrnas are very stable in blood [74] and resistant to exonucleases. indeed, circrnas rnas show an average half-life of about 48 h compared to the 10 h of linear rnas in plasma [15, 31, 44] . this confers upon them the capacity, not only to be feasible biomarkers, but also to act as theranostic tools, which means that circrnas may serve as circrna-based diagnostic and therapeutic agents in sepsis patients. nevertheless, sepsis pathophysiology is very complex, and understanding the molecular 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regulation glucocorticoid-induced microrna-511 protects against tnf by down-regulating tnfr1 multiplex cytokine profiling in patients with sepsis dysregulation in microrna expression in peripheral blood mononuclear cells of sepsis patients is associated with immunopathology attenuation of cardiac dysfunction in polymicrobial sepsis by microrna-146a is mediated via targeting of irak1 and traf6 expression toll-like receptor-mediated nf-kappab activation: a phylogenetically conserved paradigm in innate immunity a human homologue of the drosophila toll protein signals activation of adaptive immunity elevation of il-18 in human sepsis il-18) levels in patients with sepsis il-27 enhances lps-induced proinflammatory cytokine production via upregulation of tlr4 expression and signaling in human monocytes il-27, a heterodimeric cytokine composed of ebi3 and p28 protein, induces proliferation of naive cd4+ t cells circular rna circ-4099 is induced by tnf-α and regulates ecm synthesis by blocking mir-616-5p inhibition of sox9 in intervertebral disc degeneration follow-up study identifies two novel susceptibility loci prkcb and 8p11.21 for systemic lupus erythematosus calcitonin gene-related peptide induces il-6 expression in raw264.7 macrophages mediated by mmu_circrna_007893 circrna hsa_circ_0005105 upregulates nampt expression and promotes chondrocyte extracellular matrix degradation by sponging mir-26a the down-regulation of hsa_circ_0012919, the sponge for mir-125a-3p, contributes to dna methylation of cd11a and cd70 in cd4+ t cells of systemic lupus erythematous persistent inflammation and immunosuppression: a common syndrome and new horizon for surgical intensive care myd88-dependent expansion of an immature gr-1(+)cd11b(+) population induces t cell suppression and th2 polarization in sepsis microrna-155 promotes autoimmune inflammation by enhancing inflammatory t cell development microrna control in the immune system: bbasic principles microrna mir-223 as regulator of innate immunity impact of mir-223-3p and mir-2909 on inflammatory factors il-6, il-1ß, and tnf-α, and the tlr4/tlr2/nf-κb/stat3 signaling pathway induced by lipopolysaccharide in human adipose stem cells hsa_circ_0003159 inhibits gastric cancer progression by regulating mir-223-3p/ndrg1 axis serum mir-146a and mir-223 as potential new biomarkers for sepsis nf-kappab-dependent induction of microrna mir-146, an inhibitor targeted to signaling proteins of innate immune responses the prognostic value of plasma microrna-155 and microrna-146a level in severe sepsis and sepsis-induced acute lung injury patients circular rna rsf1 promotes inflammatory and fibrotic phenotypes of irradiated hepatic stellate cell by modulating mir-146a-5p circular rna expression alterations in colon tissues of crohn's disease patients circular rna profile identifies circpvt1 as a proliferative factor and prognostic marker in gastric cancer mir-125b but not mir-125a is upregulated and exhibits a trend to correlate with enhanced disease severity, inflammation, and increased mortality in sepsis patients circular rna gli2 promotes osteosarcoma cell proliferation, migration, and invasion by targeting mir-125b-5p circulating plasma micrornas can differentiate human sepsis and systemic inflammatory response syndrome (sirs) emerging roles of circrna related to the mechanical stress in human cartilage degradation of osteoarthritis mir-27a is up regulated and promotes inflammatory response in sepsis microrna-23a-5p acts as a potential biomarker for sepsis-induced acute respiratory distress syndrome in early stage inhibition of microrna-23b attenuates immunosuppression during late sepsis through nik, traf1 and xiap challenges in the analysis of epigenetic biomarkers in clinical samples long non-coding rnas discriminate the stages and gene regulatory states of human humoral immune response microrna 21 (mir-21) and mir-181b couple with nfi-a to generate myeloid-derived suppressor cells and promote 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circular rna and microrna profiles in septic myocardial depression: a lipopolysaccharide-induced rat septic shock model inducible rasgef1b circular rna is a positive regulator of icam-1 in the tlr4/lps pathway 0: decoding mirna-cerna, mirna-ncrna and protein-rna interaction networks from large-scale clip-seq data emerging roles of circrna_001569 targeting mir-145 in the proliferation and invasion of colorectal cancer circular non-coding rna anril modulates ribosomal rna maturation and atherosclerosis in humans nsd2 circular rna promotes metastasis of colorectal cancer by targeting mir-199b-5p-mediated ddr1 and jag1 signalling circular rna 0000096 affects cell growth and migration in gastric cancer circular rna circhipk3 promotes the proliferation and differentiation of chicken myoblast cells by sponging mir-30a-3p circular rna signature in hepatocellular carcinoma circrna-9119 regulates the expression of prostaglandin-endoperoxide synthase 2 (ptgs2) by sponging mir-26a in the endometrial epithelial cells of dairy goat circular rna circtrim33-12 acts as the sponge of microrna-191 to suppress hepatocellular carcinoma progression sepsis: older and newer concepts circular rna circ-foxo3 induced cell apoptosis in urothelial carcinoma via interaction with mir-191-5p basophil recruitment to skin lesions of patients with systemic lupus erythematosus mediated by ccr1 and ccr2 sepsis definitions: time for change the clinical challenge of sepsis identification and monitoring surviving sepsis campaign: international guidelines for management of severe sepsis and septic shock early versus delayed administration of norepinephrine in patients with septic shock empiric antibiotic treatment reduces mortality in severe sepsis and septic shock from the first hour: results from a guideline-based performance improvement program diagnostic and prognostic markers in sepsis new approaches to sepsis: molecular diagnostics and biomarkers mir-145 negatively regulates tgfbr2 signaling responsible for sepsis-induced acute lung injury function and clinical significance of circrnas in solid tumors matrix-metalloproteinases and their inhibitors are elevated in severe sepsis: prognostic value of timp-1 in severe sepsis matrix metalloproteinase-14 triggers an anti-inflammatory proteolytic cascade in endotoxemia predictive value of mirna-29a and mirna-10a-5p for 28-day mortality in patients with sepsis-induced acute kidney injury. nan fang yi ke da xue xue bao comprehensive analysis of differentially expressed profiles of lncrnas and circrnas with associated co-expression and cerna networks in bladder carcinoma cellular and extracellular mirnas are blood-compartment-specific diagnostic targets in sepsis epidemiology and immune pathogenesis of viral sepsis this article is an open access article distributed under the terms and conditions of the creative commons attribution (cc by) license key: cord-354384-bshj0w3o authors: tanak, ambalika s.; muthukumar, sriram; krishnan, subramaniam; schully, kevin l.; clark, danielle v.; prasad, shalini title: multiplexed cytokine detection using electrochemical point-of-care sensing device towards rapid sepsis endotyping date: 2020-10-19 journal: biosens bioelectron doi: 10.1016/j.bios.2020.112726 sha: doc_id: 354384 cord_uid: bshj0w3o the implementation of endotype-driven effective intervention strategies is now considered as an important component for sepsis management. rapid screening and frequent monitoring of immune responses are critical for evidence-based informed decisions in the early hours of patient arrival. current technologies focus on pathogen identification that lacks rapid testing of the patient's immune response, impeding clinicians from providing appropriate sepsis treatment. herein, we demonstrate a first-of-its-kind novel point-of-care device that uses a unique approach by directly monitoring a panel of five cytokine biomarkers (il-6, il-8, il-10, trail & ip-10), that is attributed as a sign of the body's host immune response to sepsis. the developed point-of-care device encompasses a disposable sensor cartridge attached to an electrochemical reader. high sensitivity is achieved owing to the unique sensor design with an array of nanofilm semiconducting/metal electrode interface that is functionalized with specific capture probes to measure target biomarkers simultaneously using non-faradaic electrochemical impedance spectroscopy. the sensor has a detection limit of ∼1 pg/ml and provides results in less than 5 min from a single drop of an undiluted plasma sample. furthermore, the sensor demonstrates an excellent correlation (pearson's r > 0.90) with the reference method for a total n = 40 clinical samples and the sensor's performance is ∼30 times faster compared to the standard reference technique. we have demonstrated the sensor's effectiveness to enhance diagnosis with a mechanistic biomarker-guided approach which can be helpful towards disease endotypying for effective clinical management of sepsis at the patient bedside. endotypes are biological subtypes characterized by distinct pathophysiological functions, described by specific biomarkers. the interpretation and verification of sepsis endotypes can save lives, by encouraging early identification of patient groups for accurate therapy. sepsis endotyping enables physicians to provide critical care and precision medicine as it showcases the patient's immune and treatment response to sepsis. thus, there has been a promising transition from predicting the outcome to pathobiology driven understanding of host response heterogeneity to sepsis, leveraging innovative high-performance translational techniques and analytical methods to identify distinct biomarker subgroups of the host response. medical communities have further acknowledged the value of biological markers as they continue to enhance sepsis diagnosis, which allows the classification of patients within the specific clinical category (akdis, 2012; burke, 2016) . sepsis is recognized as a global health crisis affecting more than 49 million people every year (rudd et al., 2020) . it is a life-threatening condition that represents the systemic immunological reaction of the body to an infectious incident that leads to death (singer et al., 2016) . it is well known that sepsisrelated fatality is not directly caused by the invading pathogen rather, the clinical complexity is triggered by the dysregulated host immune response that leads to multiple organ dysfunction (schouten et al., 2008) . the pathogen or the causative agent triggering sepsis may differ the host's immune response being a key indicator in assessing fatality and the need for complex medical treatments. ultimately, the combination of pathogen load, infection site, and host susceptibility leads to clinical presentation and course of the disease. sepsis may be caused by any form of infection; the most common cause is a bacterial infection (pneumonia or urinary tract infection) that affects the body and triggers bacterial sepsis. whereas, viral sepsis is caused by a viral infection (flu) and more cases of viral sepsis are triggered by covid-19, which has caused a pandemic in 2020. additionally, recent studies have reported mortality in covid-19 patients triggered by sepsis, especially for elderly patients with pre-existing chronic illness (alhazzani et al., 2020; arentz et al., 2020; bhatraju et al., 2020; phua et al., 2020) . currently, rapid diagnostics exist mainly for pathogen identification such as those highlighted in supplementary (table s1) while a traditional clinical workflow is illustrated in fig 1a. briefly, technologies such as septifast (roche) identifies multiple pathogens in six hours, hyplex (bag) relies on the pcr technique to recognize relevant pathogens within three hours, and the film array device (biofire) requires an hour for pathogen detection. these commercially available and those in development focus solely on detecting sepsis based on the pathogen. however, it is crucial to understand the nature of sepsis and its form of representation in the patient by tracking the host immune response. as sepsis initiates, both pro-inflammatory and anti-inflammatory mechanisms start promptly with a predominant initial hyper-inflammatory phase as shown in fig 1b. in most of the cases, the innate immune response destroys the invading pathogen, but occasionally the pathogen prevails, and the host response may become unbalanced and destructive. the increased production of cytokines and chemokines is attributed to the severity and prevalence in sepsis, implying that a chain of unregulated inflammation has initiated displaying signs of both excessive inflammations as well as immune suppression, the severity of which varies from patient to patient. this perplexing phenomenon of the host immune response has been thought to be a race to the death between the invading microbes and the host immune response, and the pathogens seek and benefit by suppressing various facets of host immunity according to hotchkiss and coworkers (hotchkiss et al., 2013) therefore, monitoring cytokine biomarkers can assist in clinical decisionmaking and forecast sepsis-related outcomes to treatments (dupuy et al., 2013) . for instance, measuring the cytokines levels near-patient at different time points would help in administering specific types of drugs, where, immunosuppressants may be prescribed early in the disease etiology addressing the hyperimmune state of the patient and immunomodulators at a later stage in conjunction with antimicrobial therapy. hence, it is critical to additionally track immune response imbalance triggered by inflammatory & anti-inflammatory cytokine immune response to channelize appropriate treatment strategy. as a point-of-care treatment option, to date, no molecular host biomarker panel is available which makes an informed decision on the specific intervention based on the diagnosis of the immune response or the ability to detect improvements in the status of patients with sepsis (albert-vega et al., 2018; gunsolus et al., 2019) . this gives rise to a major diagnostic gap in near-patient testing capabilities. to address this technological gap this work demonstrates first-of-a-kind near-patient testing 'detect sepsis' (direct electrochemical technique targeting sepsis) sensor, which directly measures a panel of five host immune biomarkers in <5 minutes to guide the physician with active feedback on patient immune status for better therapeutic administration. detect sepsis sensor enables a mechanistic approach for sepsis stratification by leveraging the use of endotypes as defined by specific biomarkers to classify based on pathophysiological process rather than the clinical representation of sepsis which is a step towards precision medicine. electrochemical sensing modality in conjunction with affinity-based capture probes specifically quantifies levels of pro-and anti-inflammatory biomarkers (il-6, il-8, il-10, trail, and ip-10) using minimally acquired blood plasma samples. fig. 1a ) illustration of the current clinical timeline vs. detect sepsis approach as rapid nearpatient testing for the screening of disease severity based on biomarker levels. image is created with biorender.com b) hyper-inflammatory and immunosuppressed phase of sepsis hypothesis c) proof of feasibility towards establishing evidence-based clinical management approach using n=40 patient samples. 1c) . our point-of-treatment technology allows rapid detection of multiple host-immune response sepsis cytokine biomarkers with ease of sample handling coupled with low sample volume (~40 μl) to facilitate near-patient bedside monitoring towards enabling biomarker-guided patient stratification, endotyping, and improving treatment response within the critical golden hour post sepsis detection. the advantages of the detect sepsis sensor over existing point-of-care tests are: (i) direct hassle-free measurement from a single drop of undiluted blood plasma; (ii) allows sepsis stratification based on the body's hyper and hypo immune response; (iii) specifically surface engineered sensor design facilitates high sensitivity and specificity; (iv) portable handheld format enables multi-measure capabilities at near-patient testing. such a point-of-care-testing device would allow clinicians on making an evidence-based decision on immune-modulating treatments customized to the patient's inflammatory response within the "golden-hour" as illustrated in fig 1c. the objective of our study was to establish a robust sensing performance of the developed biosensing detect sepsis sensor using enlisense's read platform for the detection of sepsis. the sensor was designed to allow simultaneous detection of cytokine biomarker panel using minimal sample fluid (<40 ul) in human blood plasma. sensor performance metrics (sensitivity, specificity, dynamic range, detection limit, precision, and accuracy) were tested for il-6, il-8, il-10, trail & ip-10 in plasma. detect sepsis sensor was further validated for clinical translation by testing 20 patient samples tested positive for sepsis at the time of admission compared to the 20 healthy cohorts. the performance of the developed sensor was compared with the luminex standard as a reference method. dithiobis (succinimidyl propionate) (dsp) and the dimethyl sulphoxide (dmso) solvent was purchased from thermo fisher scientific (usa) along with the phosphate buffer saline (pbs) and superblock. the antibodies and their specific antigens for il-6, il-8, il-10, trail, and ip-10 were purchased from abcam. pooled human plasma was obtained from innovative research, inc. (usa) for sensor characteristic studies. plasma from sepsis patients was obtained from the austereenvironments consortium for enhanced sepsis outcomes (aceso), a consortium consisting of us government, non-profit, academic, and industry partners. all the stock proteins and patient samples were stored at -20°c or according to their storage conditions until further use. none of the proteins underwent more than 3 freeze-thaw cycles to avoid denaturing of the proteins. the antibodies were diluted in pbs to bring them to their optimized concentration while their respective antigens were spiked in pooled human plasma in varying concentrations to perform calibrated response curves for each target biomarker. the detect (direct electrochemical technique targeting sepsis) sensor uses enlisense's rapid electroanalytical device (read) platform that comprises of the following: (1) a disposable, single-use sensor cartridge with an array of sensing electrodes that are individually configured to detect multiple biomarkers simultaneously from the sample specimen in real-time. (2) a handheld, palm-sized form-factor electronic reader onto which the sensor is mounted, which transduces the electrical outputs resulting from the sample specimen to other electronic devices/data server through a software interface (configurable to support both wired and wireless communication). the subtle changes between antibody-antigen affinity interaction result in an electrochemical impedance signal response. briefly, the detection mechanism is based on non-faradaic electrochemical impedance spectroscopy (eis). herein, a small input voltage (10 mv) is applied to the sensor over a frequency range and the resulting impedance response is measured by the portable electronic device. the functioning of the electronic reader has been previously demonstrated elsewhere from our group (sankhala et al., 2018) . eis is a powerful technique that captures subtle interaction at the functionalized electrode surface. when a sample is introduced on the electrode surface and the electrode is polarized, the rearrangement of charges occurs at the electrode-solution interface. this results in a local built-up of excessive ions of opposite charge. the extent to which the exponential charge built-up decays, forms the electrical double layer (or double-layer capacitance). the target analyte binds to the specific capture probe antibody within this double layer leveraging the antibody-antigen affinity mechanism across each working electrode and impedance is measured (jagannath et al., 2018; tanak et al., 2019a tanak et al., , 2019b . advantages of non-faradaic eis over faradaic method includes (i) label-free technique that can directly measure the subtle binding interactions without the need for a redox label for the measuring impedance response, thus, making non-faradaic eis considerably more compatible in point-of-care applications (ii) non-faradaic impedance measurement eliminates the need for a dc potential; thus, it does not denature the biomolecules immobilized on the sensing electrode surface. the sensing layer was surface engineered through a standard sputter fabrication technique using rf magnetron to deposit a 200 nm thickness of semiconducting thin film on the gold electrodes. before deposition, a solvent cleaning strategy was applied where the surface was thoroughly cleaned with isopropyl alcohol (ipa), acetone, and di water to eliminate any impurities. modulating the surface chemistry of the metal oxide layer helps to improve the rearrangement of charges near the electrode-solution interface. furthermore, zinc oxide semiconductor has unique properties including a large bandgap (3.367 ev), is non-toxic, and has high excitation binding energy (60 ev) that assists to increase overall sensitivity. additionally, due to its high adsorption capability owing to its high isoelectric point (∼9.5), chemical stability and good electrical conductivity enhance its use for sensitive electrochemical biosensing applications (tanak et al., 2019a) . nanoscale dimensions of the semiconducting thin film allow size-based matching to the target analyte, which effectively increases surface area to volume ratio. additionally, the structural morphology of the nanofilm offers selective biomolecular binding for the functionalized capture probes. with the increase in the surface-to-volume ratio, the surface structures of nanofilms can significantly modulate the charge carrier densities within the material and increase band bending (cho et al., 2020; kim et al., 2013) . previously, our group has experimentally demonstrated a three-fold increase in sensitivity for the nanofilm as compared to planar nontextured microelectrodes (shanmugam et al., 2016) . thus, leveraging the unique properties of the zno nanofilm surface engineered layer, we have demonstrated sensitive electrochemical biosensing for the detection of multiplexed biomarkers. the sensor surface was immobilized with 10mm dsp dissolved in dmso and incubated in dark at room temperature. specific capture antibodies (il-6, il-8, il-10, trail, and ip-10, 10 µg/ml each) were individually functionalized on each working electrode of the sensors. superblock was used to hydrolyze unbound linker sites to avoid non-specific interaction. a calibrated response was established for each of the pentaplex biomarkers against varying dose concentrations spiked in pooled human plasma. data were represented as a percentage change in impedance with respect to baseline (plasma blank without the target biomarkers), using the equation s1 (supplementary information). cross-reactive study was tested by preparing a cocktail of non-specific biomolecules in low (10 pg/ml) and high concentrations (1000 pg/ml) with the absence of the target biomarker. individual response for the cross-reacting molecules was measured for each sensor functionalized with the target capture antibody. to test the specificity of the sensor in the presence of interfering biomarkers, the target biomolecule was spiked to the previously prepared cocktail solution, and the response was measured for each of the pentaplex biomarkers. all the data represented is measured from n=3 replicates. the limit of detection (lod) described as the lowest measured concentration was calculated as 3 times sd of blank plasma. plasma samples were collected under written informed consent as part of an ongoing observational trial of sepsis in resource-limited settings conducted by aceso (schully et al., 2017; schully and clark, 2019) . briefly, patients presenting to the emergency department of a participating hospital with at least two sirs features and a suspected infection (sepsis-2 criteria) were eligible for enrollment. for this work, plasma samples collected 24 hours after enrollment from sepsis patients were used. samples were provided stripped of all identifiers by aceso to biomedical microdevices and nanotechnology laboratory, ut dallas following the material transfer agreement (mta), approved by the institutional review board (irb# 19mro151) at the university of texas at dallas. the samples were stored at -20°c immediately on arrival until further use and did not undergo more than two freeze-thaw cycles. statistical analysis was performed using graphpad prism software (graphpad software inc., la jolla, ca). ns: non-significant, *p<0.05, **p<0.01, ***p<0.001, ****p<0.0001. data represented as mean ±sem for n=3 replicates unless stated otherwise. one-way analysis of variance (anova) was used for the comparison between three or more groups. t-test was used to compare significance for specific signal against non-specific interferons in the cross-reactive study. differences between healthy and septic cohort were assessed using non-parametric (unpaired mann-whitney tests). there is an immense unmet need for a rapid diagnostic to enable patient stratification in sepsis towards effective disease management. therefore, the point-of-care-tests should be highly sensitive with a wide dynamic range for a panel of host response biomarkers, that can be leveraged for assessing the patient's immune state towards stratification and disease management. based on the wide body of scientific literature, and clinical evidence, our approach was to target a combination of pro and anti-inflammatory cytokines in conjunction with cell apoptosis monitoring protein. therefore, we selected il-6, il-8, ip-10, il-10, and trail for this study as the levels of these biomarkers provide a composite snapshot into a patient's immune response state towards establishing disease severity and mortality risk in the patient. trail and ip-10 were included for discriminating between the viral and bacterial loadings on the host (chaudhry et al., 2013) . detect sensor device was calibrated for each of the pentaplex biomarker panels in pooled human plasma (control set with no infections). herein, varying dose concentrations were measured for specific analytes to establish a calibrated dose-response on the multiplexed affinity capture probe functionalized sensor array. signal impedance response between antibody and the target analyte was captured using electrochemical impedance spectroscopy and represented as a percentage change in impedance with respect to the baseline (described in the methods section) in pooled plasma as shown in fig 2(a-e) . a dose-dependent increasing trend in impedance signal response was observed for all the biomarkers. the dynamic range for each biomarker was aimed to capture the healthy as well as a diseased state within the physiologically relevant range of clinical samples. il-6 demonstrated a wide dynamic range of 0.01 pg/ml to 10 ng/ml with a limit of detection (lod) of 0.1 pg/ml in spiked plasma samples (fig 2b) . the box plots for each biomarker display no overlapping interquartile ranges with a minimum variation for each concentration, indicating good repeatability with the least variance. the signal impedance response is reflective of the affinity binding mechanism between the specific capture probe and the target analyte that indicates the biomarker concentration in pooled plasma. the unique multiplexed sensor design coupled with specific surface functionalization augments the signal response and has previously been described elsewhere (tanak et al., 2019b (tanak et al., , 2019a . il-8 demonstrated a dynamic range of 0.1 pg/ml to 5 ng/ml with a detection limit of 0.1 pg/ml (fig 2c) . il-6 and il-8 are known to be major mediators of an inflammatory response, and their levels elevate in patients with sepsis, which act as key indicators during the development of severe sepsis. similarly, the dynamic range for il-10 was observed to be from 0.1 pg/ml to 1 ng/ml (fig 2d) with an lod of 1 pg/ml in pooled plasma. il-10 belongs to the group of immunoregulatory molecules called anti-inflammatory cytokines that prevents the body from the adverse effects of excess inflammatory immune reactions. the key risk factor for sepsis severity and the fatal outcome is the chronic overproduction of il-10 which indicates patients with sepsis are in a deep immunosuppression state (gogos et al., 2000) . data in fig 2e shows the dynamic range of trail from 1pg/ml to 1 ng/ml with a detection limit of 1 pg/ml whereas ip-10 displayed a dynamic range of 1 pg/ml to 2 ng/ml (fig 2f) respectively. additionally, each biomarker demonstrated a statically significant difference between concentrations as determined by one-way anova with 95% confidence intervals (table s2). the developed detect sepsis sensor demonstrated sensitive detection for biomarkers below normal threshold levels as well as the dynamic ranges extended beyond elevated levels predicted in disease states. the surface engineered semi-conducting nanofilm allows sensitive detection by potentially increasing the surface to volume ratio, allowing plenty of biomolecules to be immobilized onto the electrode surface. after establishing a sensitive and robust calibration response, the selectivity and specificity of the detect sepsis sensor on enlisense's read platform were evaluated. every analyte was tested with a series of non-specific markers, starting with the lowest concentration of the cross-reacting molecule, followed by the highest concentration as seen in fig 2 (g-k) . selectivity of the biosensor is extremely important while testing actual clinical samples where the concentrations of the analyte can be much lower than that of the non-specific molecule. thus, to mimic realistic scenarios, the sensor was additionally tested with a cocktail mixture of low and high non-specific molecules along with the target analyte spiked plasma sample to validate sensor device platform specificity. nonspecific biomarkers (represented as bar's a and b in fig 2 (f-j) showed less than 10 % reactivity compared to the specific response of the target biomarker. additionally, despite the presence of varying concentrations of cross-reacting molecules (represented as bar c and d in fig 2 (f-j) along with the target biomarker, the developed sensor demonstrated similar results (100% reactivity) to that of the specific individual analyte. detect sepsis sensor's capability to selectively bind to the functionalized antibody is attributed to the surface-functionalized highly specific monoclonal antibody combined with the effect of blocking buffer. by blocking the active functional groups on the electrode surface, the blocking buffer (superblock) prevents non-specific binding and can help stabilize the biomolecule attached to the electrode surface, thereby increasing the specificity of the biosensor (xiao and isaacs, 2012) . specificity is an important sensor metric to reduce false-positive results and provide accurate detection capability with increased resolution in distinguishing disease state. overall, the data confirmed the developed deteect sepsis sensor demonstrated high sensitivity coupled with specific and selective response despite the presence of non-specific biomolecules for the multiplexed cytokine biomarker panel in plasma using eis as the detection technique. repeatability, reproducibility, accuracy, and stability are the main considerations that need to be assessed while evaluating the sensing platform's effectiveness. the coefficient of variation (%cv) was calculated for all the study biomarkers as a measure to assess the dispersion within each reported concentration from n=10 sensors, as shown in fig 3 (a-e) . generally, lower concentration tends to show higher variability and the developed sensor demonstrates the cv range between 3-16%. the results displayed in fig 3 ( a-e) exhibit a cv < 20% which is clinically accepted as per the guidelines set by the clinical and laboratory standards institute (clsi), thereby demonstrating the repeatability of detect sepsis sensor within a wide dynamic range (clsi, 2014) . fig 3 (f-j) demonstrates the reproducibility of the electrochemical response for 12 identical sensors with an average concentration indicated by the dotted line for each target analyte. the relative standard deviation (rsd) across all twelve sensors was ~ 10%. the value of rsd indicates good reproducibility and repeatability of the detect sepsis sensor platform. fig. 3 (a-e) coefficient of variation plot examining the precision of detect sepsis sensor for il-6, il-8, il-10, trail and ip-10 in pooled plasma. the dotted line at 20% represents the acceptable limit according to clsi guidelines (f-j) repeatability and reproducibility of detect sepsis sensor across 12 sensors for il-6, il-8, il-10, trail and ip-10 in pooled plasma respectively. the dotted line represents average concentration across 12 replicates. next, detect sepsis sensor performance was tested for its accuracy by the spike and recovery study. known concentrations (actual) spiked in triplicate correlated with an r2 value of 0.99 with the measured concentration calculated based on the previously established calibration curve. fig 4 ( a-e) demonstrates reliable detection of actual spiked concentration across all five target biomarkers. the recovery percentage was then calculated along with the accuracy of the sensor as represented in fig 4 (f-j) . as observed from the results, the percent recovery was between ~89-110% for il-6, il-8, il-10 trail, and ip-10, which lies well within the acceptable range for assay validation according to clsi standards (andreasson et al., 2015) . once the accuracy was established, the operational stability of detect sepsis sensor was tested for up to five weeks (35 days) with the sensors stored in 4°c. 1 pg/ml concentration for trail and ip-10 was measured thrice (n=3). no significant change in response was observed for trail with only a 3% loss of signal at the end of five weeks as seen in supplementary fig s1. similarly, a loss of only 5% of the signal response was seen at the end of five weeks specific to ip-10 as seen in supplementary fig s2. in summary, both trail and ip-10 retained 95-97% of its original activity post five weeks of storage in 4°c, indicating excellent operational stability. to our knowledge, this is the first demonstration of a pentaplex biomarker sensor that allows simultaneous quantification of pro-and anti-inflammatory biomarkers with a single drop of plasma sample with reliable sensor performance metrics. clinical translation for the multiplexed poct technologies requires validation with patient samples. for this study, 20 septic patient plasma samples along with 20 control (non-septic) healthy plasma samples (table s3) were evaluated using the detect sepsis sensor and luminex as the reference standard. the onset of sepsis was confirmed at 24 hr time point from the hospital. all the 40 samples were tested for il-6, il-8, il-10, trail, and ip-10 using the developed sensor device. concentrations measured using the sensor correlated well with a pearson's r ≥ 0.90 for all the five test biomarkers as seen in fig 5(a-e) . fig 5 (f-j) represents the detect sepsis sensor's capability to distinguish healthy vs. sepsis patient cohorts for il-6, il-8 il-10, trail, and ip-10. we observed that levels of il-6, il-8, il-10, trail, and ip-10 of the healthy cohort were significantly different from the septic cohort as confirmed using mann-whitney u statistical analysis test. mean plasma il-6 levels for the septic cohort were 44.05±74.32 pg/ml as compared to 2.2±0.83 pg/ml of the healthy cohort. il-8 levels for the healthy cohort were all below 2.44±0.88 pg/ml while mean levels in the septic cohort were around 11.65±16.16 pg/ml. similarly, for anti-inflammatory il-10 biomarkers, the mean level established for the healthy cohort was 2.17±0.84 pg/ml and septic patients' mean levels were 15.47±18.18 pg/ml. as seen in fig 5d, trail levels were significantly lower in septic patients with a mean value of 27.62±18.62 pg/ml as compared to the healthy cohort mean levels of 47.69±18.74 pg/ml. studies have shown to correlate lower levels of trail to poor patient outcomes, thus indicating the overall severity of illness (schenck et al., 2019; tian et al., 2013) . mean healthy concentrations for ip-10 were 45.27±34.57 pg/ml whereas septic sample mean levels were measured to be 397.3±572.5 pg/ml. the detect sepsis method was also able to distinguish all the five biomarkers in healthy and septic patient samples with good statistical significance of p<0.001 (table s4) . trail, detect sepsis sensors capability to significantly distinguish healthy vs. sepsis patient cohorts for il-6, il-8 il-10, trail and ip-10. note: **p < 0.01, ***p < 0.001, ****p < 0.0001. patients with sepsis can be stratified based on evaluating specific immunological response patterns by a promising approach of cytokine profiling (marshall, 2006; ulloa et al., n.d.) . research has been proposed that a combination of biomarkers may yield better results, as no single biomarker exhibits an accuracy of 100% to predict a reliable outcome. accordingly, in this work, we developed a unique strategy targeting a combination of five pro and anti-inflammatory cytokine biomarkers to rapidly detect sepsis using the detect sepsis sensor to assess the patient's host response useful in a clinical setting. the empirical research approach adopted for this work was based on the evolving knowledge of host response to infection during sepsis. it is well-identified that sepsis fatality is not specifically induced by infectious microorganisms or pathogens; instead, the subsequent pathological outcome is triggered by dysregulation of the host immune response with a combination of pro and anti-inflammatory processes, contributing to multiple organ failure. detect sepsis sensor provides a descriptive understanding of the host immune response with the pentaplex biomarker strategy enabling patient stratification to predict timely evidence on the arc of sepsis. the sensitive and specific aspect combined with a wide dynamic range of the detect sensor device platform allows sepsis stratification to differentiate the patient state enabling appropriate therapeutic interventions. the combination of pro and anti-inflammatory markers (il-6, il-8, and il-10) reveals host immune response during the early stages of sepsis whereas trail and ip-10 provide information to differentiate between bacterial or viral sources of infection. each biomarker provides key information based on the specific pathophysiology. timing of cytokines release and the symbiosis between pro-and anti-inflammatory agents determines the degree of infection, and their excessive production can be associated with deleterious effects. a clinical investigation detected a high concentration of cytokines in plasma of critically ill patients affected with covid-19, suggesting that cytokine storm was associated with the severity of the disease as well (huang et al., 2020) . cytokine mediators are elevated in both pediatric and adult patients and are responsible for illicit symptoms including fever, hypotension, and production of acute-phase proteins (banyer et al., 2000; nedeva et al., 2019) . they are likely to be prognostic as primary regulators during the early stage of sepsis (damas et al., 1992; viallon et al., 2000) . additionally, elevated il-6 and il-8 levels are linked with early 48 hours and 28-day mortality in sepsis patients (bozza et al., 2007) . moreover, il-8 is a known mediator of the inflammatory response which plays a major role in neutrophil activation. our results are in agreement with previous research which illustrated the increase of il-6 and il-8 during the first 24 hours of hospital admission as compared to the healthy cohort (bozza et al., 2007; hou et al., 2015; thao et al., 2018) , thus, demonstrating the role of cytokines biomarkers in identifying sepsis prognosis at an early stage. the role of antiinflammatory biomarker il-10 during sepsis is complicated as it depends on the time of intervention of either being protective or destructive to the host. overproduction of il-10 in septic patients is an indicator of severity and fatal outcome as the body spirals in a state of immunoparalysis. therefore, il-10 in the detect pentaplex panel acts as an indicator of immune suppression, and thereby reflects the severity of the patient's condition for sepsis-induced immunosuppression. since the patient samples tested were collected at a 24-hour time point, the il-10 levels weren't as significant compared to the healthy cohort, thereby indicating an early stage of sepsis. our results demonstrating the down-regulated trail profile support the hypothesis that trail participates in sepsis by controlling inflammatory cell apoptosis and promotes inflammation resolution (renshaw et al., 2003) . the presence of trail and ip-10 has been independently used to differentiate viral from bacterial infection. (oved et al., 2015; van der does et al., 2018 van der does et al., , 2016 van houten et al., 2017) . moreover, the hypothesis that trail participates in sepsis by controlling inflammatory cell apoptosis and promotes inflammation resolution (renshaw et al., 2003) . broadly, each selected biomarker demonstrates key elements in activating immune response during sepsis. the approach of multiplexed cytokine profiling would enable rapid sepsis endotyping based on biomarker levels using the detect sepsis sensing platform. physicians can actively monitor patient status for better prognosis and provide enhanced therapeutic interventions, with multiple measures at patient bedside. clinical management of sepsis can be divided into three phases: (i) patient screening; (ii) patient stratification based on evidence-based clinical management and response to treatment therapies; and (iii) prognostic monitoring. as discussed earlier, much effort has been utilized in the first and third phases, while very little progress has been reported with patient stratification methods. existing methodology lacks the potential to deliver rapid poc results deprived of demanding postprocessing to facilitate adequate diagnosis strategies for septic patients. to date, very little work has been done on enabling multiplexed biomarker detection at point-of-care-testing allowing timely treatment. moreover, transitioning these findings into a clinically feasible test requires a rapid, convenient method that can resolve the lengthy testing process (1-8 hours) while providing accurate results. the developed self-integrated sensing device improvements over existing techniques include (i) direct patient sample measurement without the need for sample preparation or dilution (ii) low sample volume utilization (~40 µl) of blood plasma in a portable hand-held format, that could be used to collect data over multiple time points with (iii) rapid results achieved within ~5 mins (iv) simultaneous multiplexed detection of cytokine panel biomarkers, for classifying patients depending on the levels reflecting severity of illness (v) sensitive, selective, specific and stable biosensing response enhances the reliability of the detecting mechanism. many researchers have leveraged the use of il-6, il-8, il-10, trail, and ip-10 individually, but to our knowledge, this work is the first demonstration of a simultaneous pentaplex biomarker panel for early sepsis diagnosis and monitoring using low sample volume (< 40 µl) achievable rapidly. detect sepsis sensor can be used effectively in the emergency department for early sepsis screening, or it can be used to monitor sepsis prognosis for patients as a bedside monitoring device. biomarkers linked with sepsis are attributed to the complex immune response pathways, therefore rapid multiplexed detection would enable early therapeutic intervention and improve patient outcomes. additionally, the sample-to-detection time, measured for all five biomarkers from time of sampling until sensor readout, was ~5 mins, which is >30 times faster than the standard reference method (~5 hours). the repeatable and reproducible results demonstrated by the detect sepsis sensor shows evidence of an accurate and reliable electrochemical biosensing mechanism. when tested against common interferants, the specific sensing capability for the developed pentaplex sensor was not affected. moreover, the sensor displayed a stable response for over five weeks. to our knowledge, no multiplex point-of-treatment device is available for sepsis detection for nearpatient testing without sample dilution with rapid response time. our data shows the first demonstration of a truly novel multiplexed platform capable of monitoring host response using pentaplex biomarkers for sepsis detection that would enable patient stratification and sepsis endotypying. this rapid sample-to-result detection capability demonstrated by the detect sepsis sensor establishes it as a value-added point-of-care testing approach in hospitals and emergency departments towards risk stratification of sepsis severity and responses to treatment. in summary, detect sepsis device platform provides the first proof of concept for rapid diagnostic screening of sepsis leveraging a host immune response biomarker pane. the developed sensor showed comparable results to the reference standard as shown with pearson's r>0.97 for all the five biomarkers. however, the developed detect sepsis sensor achieved quicker response time with lesser completed assay procedure, leveraging a combination of unique surface engineered sensing strategy coupled with an affinity biosensing principle. the results of this research demonstrate a robust, sensitive, specific, and stable performance by the detect sepsis sensor which is highly expected from a point-of-care device. moreover, it possesses three major advantages over current detection methodology. firstly, a specific affinity-based transduction mechanism allows the simultaneous detection of il-6, il-8, il-10, trail, and ip-10. multiplexed detection capability will help provide a precise molecular fingerprint of every patient encouraging initiatives towards precision medicine. secondly, rapid response time offered by the detect sensor enables faster decision-making for physicians to operate within the "golden hour" and initiate required treatment thus, avoiding the dynamic sequence of irreversible organ failure and subsequent death caused due to delayed response time. finally, small form factor and ease in handling allow flexibility in using the device in a versatile environment (in an emergency department, or for bed-side monitoring) while ultra-low sample volume (< 40 µl) encourages physicians to collect multiple measurements within a day to monitor patients host immune response and assess the severity of sepsis. the study was limited to a small sample size of septic patients. however, these observations have several implications for research to integrate different host immune response biomarkers with selfintegrated point-of-care devices for a better patient outcome. we are currently in the prosses of expanding our multiplexed capabilities and investigating sepsis pathophysiology via a larger patient cohort. in conclusion, this work has pioneered a potential solution to the existing sepsis dilemma, by providing host response strategy to address complexity, shifting the paradigm of the on-going sepsis diagnostic approach. target marker along with a high concentration of non-specific biomarker mixture fig. 3 (a-e) coefficient of variation plot examining the precision of the detect sepsis sensor for il-6, il-8, il-10, trail, and ip-10 in pooled plasma. the dotted line at 20% represents the acceptable limit according to clsi guidelines (f-j) repeatability and reproducibility of detect sepsis sensor across 12 sensors for il-6, il-8, il-10, trail, and ip-10 in pooled plasma respectively. the dotted line represents average concentration across 12 replicates. j o u r n a l p r e -p r o o f (a-e) actual and measured concentration in pooled plasma represented as correlation plots using the detect sepsis sensor. (f-j) repeatable and reliable sensing capability highlighting the high recovery rate and accuracy of the developed sensing platform. (a-e) correlation between reference method (luminex) and detect sepsis sensor platform obtained by analyzing n= 40 patient blood plasma samples with a pearson's r ≥ 0.90 for detect sepsis sensors capability to significantly distinguish healthy vs. sepsis patient cohorts for il-6, il-8 il-10, trail and ip-10. note: **p < 0.01, ***p < 0.001, ****p < 0.0001. j o u r n a l p r e -p r o o f badrinath jagannath for providing feedback on the manuscript. declaration of competing interests shalini prasad and sriram muthukumar have a significant interest in enlisense llc, a company that may have a commercial interest in the results of this research and technology. the potential individual conflict of interest has been reviewed and managed by the university of texas at dallas, and played no role in the study design; in the collection, analysis, and interpretation of data the views expressed in this article are those of the authors and do not necessarily reflect the official policy or position of the henry m. jackson foundation for the advancement of military medicine title 17 u.s.c. 101 defines a u.s. government work as a work prepared by a military service member or employee of the u.s. government as part of a person's official duties. the views expressed in this article are those of the authors and do not necessarily reflect the official policy or position of the department of the navy, defense threat reduction agency writing-review & editing danielle v clark: recourses (patient samples), writing-review & editing reference akdis evaluation of precision of quantitative measurement procedures pro-versus anti-inflammatory molecular austere environments consortium for enhanced sepsis outcomes (aceso), henry m. jackson foundation for the advancement of military medicine we would like to thank yashaswee tamrakar, jad moumen, thien nguyen, fayha khan, raqeeb ali and bayan hamed, for their support in performing experiments. we would also like to thank address: 800 w. campbell rd. bsb 11, richardson, tx, usa 75080 email: shalini.prasad@utdallas.edu highlights:• developed biosensor simultaneous detects il-6, il-8, il-10, trail and ip-10 in undiluted plasma • multiplexed sensor design and specific antibodies facilitates robust, sensitive and specific detection using non-faradaic impedance spectroscopy drs. shalini prasad and sriram muthukumar have a significant interest in enlisense llc, a company that may have a commercial interest in the results of this research and technology. the potential individual conflict of interest has been reviewed and managed by the university of texas at dallas, and played no role in the study design; in the collection, analysis, and interpretation of data; in the writing of the report, or in the decision to submit the report for publication. key: cord-349076-x3rjasg0 authors: jarczak, dominik; kluge, stefan; nierhaus, axel title: use of intravenous immunoglobulins in sepsis therapy—a clinical view date: 2020-08-03 journal: int j mol sci doi: 10.3390/ijms21155543 sha: doc_id: 349076 cord_uid: x3rjasg0 sepsis is a life-threatening organ dysfunction, defined by a dysregulated host immune response to infection. during sepsis, the finely tuned system of immunity, inflammation and anti-inflammation is disturbed in a variety of ways. both pro-inflammatory and anti-inflammatory pathways are upregulated, activation of the coagulation cascade and complement and sepsis-induced lymphopenia occur. due to the manifold interactions in this network, the use of igm-enriched intravenous immunoglobulins seems to be a promising therapeutic approach. unfortunately, there is still a lack of evidence-based data to answer the important questions of appropriate patient populations, optimal timing and dosage of intravenous immunoglobulins. with this review, we aim to provide an overview of the role of immunoglobulins, with emphasis on igm-enriched formulations, in the therapy of adult patients with sepsis and septic shock. sepsis is a life-threatening, dysregulated immune response that occurs, when the body's defensive reactions against infection damage its own tissues and organs [1] . in 2017, an estimated 48.9 million cases of sepsis were recorded worldwide with 11.0 million sepsis-related deaths, representing 19 .7% of all global deaths [2] . due to continuous progress in the understanding of the underlying pathology and immunological mechanisms, the definition of sepsis as a clinical syndrome is subject to constant development. the current consensus definition ("sepsis-3") emphasizes for the first time the crucial role of the innate and adaptive immune response in the development of the clinical syndrome. despite the enormous efforts made during the last three decades of clinical and experimental research, the available therapeutic armamentarium to positively affect the course of the disease remains restricted. even today, the mortality of septic shock, the most severe subgroup of sepsis, lies in the range of more than 50% in north america and europe [3] . in contrast to an uncomplicated and often localized infection, sepsis leads to a system-wide release of cytokines, mediators and pathogen-related molecules ("cytokine storm") [4] . the starting signal for the activation of numerous signal cascades is given by the recognition of pathogen-derived molecules (pathogen-associated molecular patterns, pamps-e.g., endo-and exotoxins, lipids, dna) or endogenous host-derived danger signals (damage-associated molecular patterns, damps) via specific receptors (toll-like receptors, tlr) on the surface of monocytes and antigen-presenting cells (apcs). this initiates the clinical syndrome of sepsis through expression of genes involved in inflammation, cellular metabolism and adaptive immunity [5] . pro-and anti-inflammatory pathways are upregulated, leading to inflammation and progressive tissue damage, ultimately causing multi-organ dysfunction. simultaneous immunosuppression by downregulation of activating cell surface molecules, t cell exhaustion and increased apoptosis of immune cells invariably accounts for "immunoparalysis" during the later stages of the disease, making the affected patients susceptible for nosocomial infections, viral reactivation and opportunistic pathogens ( figure 1 ) [6, 7] . pro-and anti-inflammatory pathways are upregulated, leading to inflammation and progressive tissue damage, ultimately causing multi-organ dysfunction. simultaneous immunosuppression by downregulation of activating cell surface molecules, t cell exhaustion and increased apoptosis of immune cells invariably accounts for "immunoparalysis" during the later stages of the disease, making the affected patients susceptible for nosocomial infections, viral reactivation and opportunistic pathogens ( figure 1 ) [6, 7] . pro-and anti-inflammatory changes of the immune system during the course of sepsis and septic shock. hla-dr, human leukocyte antigen-d related; igm/g, immunoglobulin m/g; il, interleukin; ifn-γ, interferon gamma; pamps, pathogen-associated molecular patterns; tnf-α, tumor necrosis factor alpha; tlr, toll-like receptor. binding of pamps and damps to tlrs on monocytes and apcs causes signal transduction and induces the translocation of nuclear factor kappa-light-chain-enhancer of activated b cells (nf-κb) to the nucleus. in consequence, so-called "early activation genes" are expressed, including various proinflammatory interleukins (il), e.g., il-1, il-12, il-18, tumor necrosis factor alpha (tnf-α) and interferons (ifns). these changes lead to activation of further cascades of inflammatory cytokines (e.g., il-6, il-8, ifn-γ), the coagulation cascade and complement in addition to a downregulation of adaptive immunity components [8] . as a result, increased levels of both pro-inflammatory and antiinflammatory cytokines can be detected in the early stages of sepsis [7, 9, 10] . even though the early systemic inflammatory response is generally considered the hallmark of sepsis, there is also a significant component of immunosuppression that occurs both early and late in the host sepsis response [8] . the role of b-lymphocytes in sepsis exceeds the production and secretion of immunoglobulins, they also modulate the innate immune response, produce cytokines and act as figure 1 . pro-and anti-inflammatory changes of the immune system during the course of sepsis and septic shock. hla-dr, human leukocyte antigen-d related; igm/g, immunoglobulin m/g; il, interleukin; ifn-γ, interferon gamma; pamps, pathogen-associated molecular patterns; tnf-α, tumor necrosis factor alpha; tlr, toll-like receptor. binding of pamps and damps to tlrs on monocytes and apcs causes signal transduction and induces the translocation of nuclear factor kappa-light-chain-enhancer of activated b cells (nf-κb) to the nucleus. in consequence, so-called "early activation genes" are expressed, including various pro-inflammatory interleukins (il), e.g., il-1, il-12, il-18, tumor necrosis factor alpha (tnf-α) and interferons (ifns). these changes lead to activation of further cascades of inflammatory cytokines (e.g., il-6, il-8, ifn-γ), the coagulation cascade and complement in addition to a downregulation of adaptive immunity components [8] . as a result, increased levels of both pro-inflammatory and anti-inflammatory cytokines can be detected in the early stages of sepsis [7, 9, 10] . even though the early systemic inflammatory response is generally considered the hallmark of sepsis, there is also a significant component of immunosuppression that occurs both early and late in the host sepsis response [8] . the role of b-lymphocytes in sepsis exceeds the production and secretion of immunoglobulins, they also modulate the innate immune response, produce cytokines and act as apcs [11, 12] . in the early course of sepsis, a decrease in lymphocytes, monocytes and antigen-presenting dendritic cells has been observed [13, 14] . the delicate mechanisms underlying sepsis-induced lymphopenia have not yet been conclusively explained. as a possible explanation, recruitment of lymphocytes from the peripheral circulation into areas of inflammation and infection is discussed, but most data suggest that apoptosis causes sepsis-induced lymphopenia [15] [16] [17] . the persistence of lymphopenia and therefore also the lower levels of immunoglobulins over the course of sepsis is closely associated with increased mortality [17, 18] . in addition to sepsis-induced lymphopenia, an increase in apoptosis of monocytes and apcs could be shown, which is accompanied by a significant loss of pro-inflammatory cytokine production [13, 14, [19] [20] [21] [22] [23] [24] [25] [26] [27] . the remaining cells also present with a decreased expression of the human leukocyte antigen dr (hla-dr) on their surface, resulting in a diminished ability to recognize pathogens and to interact with t cell receptors via opsonisation. as a consequence, this leads to a lack of th1-and th2-response as a component of the adaptive immune response [28] . the inability of monocytes to restore normal expression levels of hla-dr has been shown to predict poor outcome in sepsis [29, 30] . furthermore, it has also been shown that endotoxin tolerance in the early course of sepsis is also associated with further clinical deterioration [31] . as part of the innate immune system, neutrophils belong to the first line of defence against pathogens. severe bacterial infections result in emergency granulocyte formation and the release of both mature and immature forms of neutrophils from the bone marrow. when activated via pamps and damps in septic patients, these cells show reduced phagocytosis and oxidative burst capacity [32] . high levels of immature granulocytes in sepsis are associated with clinical deterioration, since these have shown increased spontaneous production and liberation of neutrophil extracellular traps (nets) [33, 34] . nets are diffuse extracellular structures of decondensed chromatin with granular and nuclear proteins [35, 36] . these structures have the potential to capture a wide range of pathogens, including gram-positive and gram-negative bacteria, viruses, yeasts, but also protozoa and parasites that cannot be phagocytized due to their size [37] . it is known that cytokines, chemokines, platelet agonists and antibodies can also trigger their release. the pronounced presence of nets in tissues or vessels due to insufficient removal or overproduction is linked to hypercoagulation and endothelial injury [34, 38, 39] . during granulopoiesis as a response to acute infection, immature myeloid cells can shift into the peripheral blood and become functionally active. these myeloid-derived suppressor cells (mdscs) release various anti-inflammatory cytokines (e.g., il-10 and transforming growth factor β, tgf-β), leading to further immunosuppression [8, 40] . these immunosuppressing properties have been the focus of extensive research in the context of malignant diseases, but the knowledge in relation to sepsis is still scarce [41] . although many details are known about the function of b-lymphocytes in the context of sepsis, their clinical relevance cannot yet be conclusively determined. for an effective host protection, b cell function and antibody expression are decisive factors. the production of antibodies is one of the crucial functions of b cells after differentiation into high-affinity antibody-secreting plasma cells [42] . within the framework of the adaptive immune response, interaction with dendritic cells, macrophages, t-and other b-cells leads to clonal expansion and finally to the production of specific antibodies. however, in the early phase of sepsis, b cells can also be activated by pathogens themselves via pathogen recognition receptors (prrs), which leads to an initial immune response by innate like b cells [11, 43, 44] . just at the beginning of sepsis, the number of b-lymphocytes in the peripheral blood is often reduced [45, 46] . in addition to increased apoptosis, increased migration from the circulation into the tissue or reduced production of these b cells in favour of the production of monocytes and neutrophils as part of an emergency haematopoiesis is possible as a cause. a recent study by dong et al. showed that in septic shock, a severe functional impairment of b cells is present in non-survivors, resulting in both lower serum immunoglobulin m (igm) levels and in lower igm production upon b cell stimulation. examining five subsets of peripheral blood b cells (immature/transitional b cells, naive b cells, tissue-like memory b cells, resting memory b cells, and activated memory b cells), the authors demonstrated a distinct redistribution of these subsets in septic shock patients when compared to healthy controls [47] . a meta-analysis could show that the number of circulating b cells is significantly higher in sepsis survivors than in sepsis non-survivors, especially within the first 24 h after the onset of sepsis [48] . the mechanism presumably responsible for this protective effect seems to be the release of natural antibody igm, which plays a key role in the fight against gram-negative infections in particular [11] . furthermore, it could be shown that igm plasma levels in survivors of sepsis or septic shock were higher in the first 24 h than in non-survivors, which supports the hypothesis of a protective function of b cells through the production of igm [48] . interestingly, a similar relationship was also shown in non-septic critically ill patients [49] . however, current data are not yet sufficient to use the measurement of b cells or igm levels in early sepsis as a prognostic factor for outcome. immunoglobulins are glycoproteins secreted by differentiated b cells, so called plasma cells. each immunoglobulin molecule monomer consists of identical light and heavy chain pairs, held together by disulphide bonds and electrostatic forces. based on the heavy chain, there are five isotypes of immunoglobulins, iga, igd, ige, igg and igm, respectively [50] . the variable regions of immunoglobulins enable non-covalent cross-linking to bacterial and other antigens, whereby the constant region transduces signals in response to antigen-binding. the most important classes within the human humoral immune system are iga, igg and igm. iga has two subclasses (iga1, iga2) and the main function is mucosal immunity. igg has four subclasses (igg1-igg4) and the key functions besides secondary antibody responses are opsonisation and complement activation. the main functions of igm are complement activation and primary antibody responses. two types (kappa and lambda) of light chains are also present in the circulation independent of whole immunoglobulin molecules, referred to as free light chains (flc), which can be detected at abnormal high levels in adult patients with sepsis [51] . the concept that acquired immunosuppression is a significant event in sepsis and septic shock leads to the hypothesis that stimulation of the immune response and/or substitution of individual immune system components could be a promising therapeutic approach. within the intricate web of interacting and regulating factors of the immune system and the inflammatory response, polyvalent intravenous immunoglobulins (ivig) might be a tool for modulating both pro-and anti-inflammatory processes ( figure 2) . the clinical rationale for ivig therapy in sepsis can be categorized as follows: the role of immunoglobulins in (i) recognition and clearance of pathogens and toxins, (ii) scavenging and inhibition of up-and downstream mediator gene transcription, and (iii) anti-apoptotic effects on immune cells. the clinical rationale for ivig therapy in sepsis can be categorized as follows: the role of immunoglobulins in (i) recognition and clearance of pathogens and toxins, (ii) scavenging and inhibition of up-and downstream mediator gene transcription, and (iii) anti-apoptotic effects on immune cells. recognition of pamps is based on naturally occurring antibodies, which can also act as innate immune receptors. igg and the complement proteins are the most important opsonins for bacterial clearance. the activation of the classical pathway of the complement system is activated by the interaction of c1 complex with immunoglobulins, acute phase proteins and non-specific activators [52] . human neutrophils express multiple fcγ -type cell surface receptors, that are capable of binding igg, resulting in neutrophil activation via tyrosine kinase pathways and thereby upregulating the expression of phagocytic receptors, that are able to identify and phagocytose pathogens opsonized with igg and complement proteins [53, 54] . due to igg deficit, the activation of neutrophils as well as phagocytosis signals may be affected in septic patients, so this population seems to be an optimal cohort for ivig therapy. monocytes and t cells also become activated by superantigen exotoxins released by staphylococci and streptococci. igg molecules contained in ivig preparations can inhibit or neutralise these superantigens, thereby preventing superantigen-mediated t cell and monocyte activation [55] [56] [57] . a key mechanism for regulating host response in inflammatory situations like sepsis or septic shock (as well as other inflammatory diseases) is nf-κb-mediated up-regulation of il-1 and the il-1 receptors system [58] . it has been shown, that these components decrease, if ivig is supplemented in case of hypogammaglobulinaemia or sepsis, as well as il-1 mediated activity of mononuclear cells in peripheral blood reduces and il-1 receptor antagonist (il-1ra) becomes induced [58, 59] . recognition of pamps is based on naturally occurring antibodies, which can also act as innate immune receptors. igg and the complement proteins are the most important opsonins for bacterial clearance. the activation of the classical pathway of the complement system is activated by the interaction of c1 complex with immunoglobulins, acute phase proteins and non-specific activators [52] . human neutrophils express multiple fcγ -type cell surface receptors, that are capable of binding igg, resulting in neutrophil activation via tyrosine kinase pathways and thereby upregulating the expression of phagocytic receptors, that are able to identify and phagocytose pathogens opsonized with igg and complement proteins [53, 54] . due to igg deficit, the activation of neutrophils as well as phagocytosis signals may be affected in septic patients, so this population seems to be an optimal cohort for ivig therapy. monocytes and t cells also become activated by superantigen exotoxins released by staphylococci and streptococci. igg molecules contained in ivig preparations can inhibit or neutralise these superantigens, thereby preventing superantigen-mediated t cell and monocyte activation [55] [56] [57] . a key mechanism for regulating host response in inflammatory situations like sepsis or septic shock (as well as other inflammatory diseases) is nf-κb-mediated up-regulation of il-1 and the il-1 receptors system [58] . it has been shown, that these components decrease, if ivig is supplemented in case of hypogammaglobulinaemia or sepsis, as well as il-1 mediated activity of mononuclear cells in peripheral blood reduces and il-1 receptor antagonist (il-1ra) becomes induced [58, 59] . in addition to neutralising antibodies in ivig preparations, naturally occurring auto-antibodies, antiidiotype antibodies and immune proteins in ivig preparations may also contribute to its immunomodulatory properties [60] . in healthy individuals, autoantibodies neutralising cytokines such as ifn-α, -β and -γ, il-1α, -2, -4, -6, -8, -10, tnf-α and -β, and soluble tnf receptors have been regularly detected. therefore, polyclonal ivig formulations are likely to contain these antibodies, which contribute to cytokine modulation as another important component of anti-inflammatory ivig activity [61] [62] [63] . alas, despite the numerous theoretical advantages clinical studies have shown that the administration of preparations containing only igg did not lead to a reduction in mortality in patients with sepsis [64] [65] [66] . however, systematic reviews and meta-analyses suggest that the use of preparations with enriched iga and igm (iggam) is associated with a higher survival rate [67] [68] [69] [70] [71] . a recent meta-analysis, including 19 trials with more than 1500 patients showed a significant reduction in mortality when using igm-and iga-enriched immunoglobulins compared to human albumin solution or no special treatment as a control intervention [67] . in addition, a post-hoc analysis of the cigma trial demonstrated a significant relative reduction in all-cause mortality of 54-68% using igm-and iga enriched immunoglobulins in patients with severe pneumonia and had high c-reactive protein (crp), low igm and high crp/low igm ratios at baseline compared to placebo [72] . conversely, patients with normal igm levels showed a tendency towards higher mortality in the treatment group (10/25 vs. 5/24, p = 0.2165), which might suggest that precise identification of the target population is essential. further, a recent clinical trial, in which patients were treated with either iggam or placebo (nacl), demonstrated a significant decrease in il-6 and il-10 levels after 72 h only in the iggam group [73] . a positive effect of igm administration on microvascular perfusion parameters could be demonstrated in humans, confirming previous studies in an animal model of endotoxemia [73, 74] . igm has also been shown to have a positive effect on the integrity of the blood-brain barrier and on septic encephalopathy in rats [75, 76] . the decrease of systemic endotoxin levels associated with iggam has been shown in animal models. human studies have focused on neonates and preterm infants, or on neutropenic patients [77] [78] [79] . recently, a small controlled study in adult patients with sepsis and septic shock and elevated endotoxin levels demonstrated that iggam significantly attenuated lps levels and had a beneficial effect on sepsis-related coagulopathy in terms of platelet count and fibrinogen concentrations [80] . the precise mechanisms by which iggam brings about these effects in patients with sepsis or septic shock are not conclusively clarified until now. in addition to the opsonisation of pathogens and the neutralization of bacterial endotoxins and exotoxins, iggam modulates the immune response by attenuating an excessive inflammatory response [80] [81] [82] [83] . the reduced production of il-2 when iggam is used is considered to be one of the proven mechanisms of action [84] . il-2 (also t cell growth factor, tcgf) is mainly produced by activated t cells and is released after mhc-ii-mediated recognition of an antigen. it is mainly autocrine and serves two opposite functions: on the one hand, il-2 enhances the proliferative response of effector t cells (t eff cells) and natural killer cells (nk cells), on the other hand il-2 simultaneously controls immune homeostasis by influencing proliferation, differentiation and function of regulatory t cells (t reg cells) [85] . overall, this leads both in vitro and in lectin-stimulated peripheral blood mononuclear cells to a significant inhibition of the alloproliferative response of human t-lymphocytes [84, 86] . immunoglobulin-induced downregulation of il-2 also causes a lack of activation of b-lymphocytes and decreases the production of pro-inflammatory tnf-α as well as the cytokines of the type 2 t-helper cell response, il-4 and il-5 [87] . up to now, the use of intravenous ig as supportive therapy in sepsis is controversial and not entirely without risk. in some patients, serious adverse reactions consist of the development of a hyperviscosity syndrome with thromboembolic events. further, acute renal failure has been observed, which was presumably associated with stabilizers contained in the ivig preparations. ivig-associated renal failure is most common in patients with pre-existing conditions such as renal impairment, diabetes mellitus, advanced age, volume depletion or concomitant use of other substances known to cause renal toxicity [88] . however, most of these potential complications can be prevented by taking appropriate countermeasures. for example, slow infusion rates and adequate hydration may help to avoid renal failure as well as thromboembolic events [89] . nevertheless, due to their pleiotropic effects and the potential modulation of both pro-and anti-inflammatory processes, polyvalent intravenous immunoglobulins offer a promising strategy within the context of inflammation and immunity. experimental studies have shown that polyvalent immunoglobulins can improve phagocytosis of pathogens via opsonisation, neutralize exo-and endotoxins and interact with complement factors, thereby preventing nonspecific activation [80, [82] [83] [84] . nevertheless, the surviving sepsis campaign (ssc) guidelines suggested against ivig use in sepsis therapy 2016 due to lack of sufficient evidence of efficacy for preparations containing only igg [90] . prepared from a pool of donors, ivig are widely used in the treatment of haematological, immunological, and neurological diseases. classic ivig-preparations contain more than 90% igg. since human plasma contains all three immunoglobulin classes, iggam preparations are considered physiological [91] . pentaglobin is currently available as a preparation in which the proportion of igm and iga is enriched to 12% each, with an igg proportion of 76%. pentaglobin also contains toxin-binding and neutralising antibodies against numerous gram-positive and gram-negative bacteria and influences the effect of further pro-inflammatory (ifn-γ, il-6) and anti-inflammatory (il-10) cytokines during the lymphocyte response [84, 92, 93] . another preparation with an even higher proportion of igm (23%) and iga (21%) is in clinical development under the working name trimodulin [72] . most of the evidence currently available on the use of iggam in sepsis or septic shock is based primarily on the use of pentaglobin in clinical trials. unfortunately, these data suffer from the fact that, in addition to a large number of different protocols (various dosages, heterogeneous patients), the laboratory parameters analysed are also inconsistent [94] . the questions of "who?", "when?" and "how much?" understanding which patients can generally benefit most from therapy with iggam is of high clinical relevance. previous studies investigating immunomodulatory approaches in the treatment of sepsis often lacked a precise characterization of suitable patients [95] [96] [97] [98] [99] . according to the current state of knowledge, the identification of a suitable target population for iggam therapy is important. in particular, the idea of "one size fits all" therapy must be critically examined under the current concept of "personalised medicine" and also the lack of cost-effectiveness data [100] [101] [102] [103] [104] . in 2005, a meta-analysis investigated 9 trials including 435 patients, and showed an increase of direct icu costs for the treatment of adult patients with severe sepsis and septic shock with iggam by €2037, but this was offset by costs per life saved of €10,565 [105] . this analysis demonstrated also a significantly reduced mortality risk but showed no effect on icu length of stay. as many other analyses in this context, this was based on small trials of variable quality. as a complex syndrome, sepsis is characterized by a variety of pathogen-and host-specific factors that show a very dynamic behaviour over time [1] . especially patients with the appearance of an excessive immune reaction, but also those with the appearance of an immune paralysis (hypoinflammation), seem to be qualified for the use of iggam. as a consequence of sepsis-related hyperinflammation, all organs can be affected and numerous biomarkers can be influenced, including crp, procalcitonin (pct) and il-6 [106] . to date there is no clear predictor for the use of iggam. the shock index and the qsofa score (quick sepsis-related organ failure) are also readily available and cost-effective tools for the initial assessment of high-risk patients [107, 108] . the measurement of circulating immunoglobulins as biomarkers in sepsis has increasingly become the focus of research within the last ten years. different approaches are pursued with regard to the development of immune scores as a tool for predicting outcome. a meta-analysis could show that the prevalence of igg hypogammaglobulinaemia in heterogeneous sepsis cohorts at the time of sepsis diagnosis was as high as 70%, however, a single subnormal measurement of igg on the day of sepsis diagnosis could not help to identify patients with a higher risk of death [109] . in another study, the combination of several ig parameters was found to be significantly associated with an unfavourable outcome when levels of iga, igg and igm were below specified cut offs [110] . this finding is consistent with the evidence from a retrospective analysis of 129 patients with septic shock treated with iggam, where a survival benefit was associated with the start of treatment within the first 23 h [111] . in addition to the excessive, pro-inflammatory immune response, the syndrome of acquired septic immune paralysis has become the focus of attention, as it seems to be predominantly associated with morbidity and mortality [112] . although the clinical appearance of hypoinflammation is not as detailed and comprehensive as the phenotype of hyperinflammation, the increased risk of secondary infection in these patients is well known [112] . patients often develop a chronic critical illness after initial survival of septic shock and cannot leave the intensive care unit. chronic immune paralysis can lead to a renewed increase in the mortality rate after about 30 days; patients often suffer from deleterious nosocomial infections: in addition to the occurrence of multi-resistant bacteria (mdr), viral infections (de novo or reactivations) or infections with fungi are also observed, the successful treatment of which remains one of the major challenges of modern intensive care medicine [113] [114] [115] [116] [117] . possibly, the detection of a low hla-dr expression on monocytes could serve as an indication of (continued) immune dysfunction and thus indicate increased mortality [118] [119] [120] . in addition to the identification and selection of suitable patient groups for therapy with iggam ("who?"), the questions of "when?" and "how much?" remain open. there are no clear answers for either aspect. in general, it could be shown that an early start of iggam therapy (i.e., 36 h vs. 66 h, respectively) in the presence of an infection leads to a higher probability of survival, which makes the time factor a key influencing variable [121, 122] . there is also a lack of evidence on the question of optimal dosage, which is the basis of current studies. in addition to the pros and cons of an initial bolus application, the type of administration (continuous vs. intermittent), the quantity (determined by serum measurements of ig or without adaptation), but also that of the target value to be achieved ("normal" or "supranormal") are not sufficiently clarified. in 2019 the "severe acute respiratory syndrome coronavirus-2" (sars-cov-2) caused a pandemic with an unprecedented global crisis, affecting healthcare and research systems, but also infrastructure sectors, including education, politics and economy. according to current knowledge, there is a link between severity of coronavirus disease 2019 (covid-19), viral production and the severe dysregulation of the inflammatory immune reaction ("cytokine storm"). however, up to date it is still unclear which molecular mechanisms trigger the onset of the immune dysbalance and why it can rapidly progress to multiorgan dysfunction or ards with a fatal outcome in a considerable subset of patients [123, 124] . clinical observation of fatal courses of covid-19 often includes severe acute respiratory distress syndrome (ards), which is caused by alveolar injury, and multiple organ failure-both of which are associated with hyperproduction of cytokines [123] [124] [125] . both mild and severe/fatal cases display changes in cytokine production, particularly il-1β, il-1ra, il-6, il-10, tnf, gm-csf, il-17, and pathological shifts of circulating leukocyte subsets [126, 127] . as a consequence, this leads to the disturbed development of protective immunity against the infection. the most severe complications of covid-19 include sepsis-like inflammation, pulmonary or cardiovascular complications, and coagulopathy [128] [129] [130] . as discussed above, the innate immune system of the host is activated in response to the virus to limit infection. subsequently, the adaptive immune system develops specific immunoglobulins and activates t cells in direct response to the virus. however, if this inflammation is unmodulated or excessive, there is a risk of chronic hyperinflammation resulting in functional inhibition of the adaptive immune system. in addition to virus-induced lymphopenia, this can result in progressive tissue and organ damage and the failure of the adaptive immune system to develop functional immunoglobulins, thereby clearing the virus [131] . therefore, the use of iggam in patients showing signs of both hyper-and hypoinflammation could be an effective therapeutic strategy. to this day, the cornerstones of sepsis and septic shock therapy still consist of timely focus control, the administration of anti-infective drugs and haemodynamic stabilisation through early and sufficient administration of fluid and vasopressors. the clinical understanding of sepsis is evolving towards an immunological perspective and remarkable progress has been made in recent decades. shedding light on the complex pro-and anti-inflammatory pathways and the disorders of the complementary and coagulation systems, thereby demonstrating the complexity and heterogeneity of the syndrome, has yet not led to transfer this knowledge into evidence-based approaches to treat sepsis and develop effective therapies. despite progress in the (further) development of innovative therapeutic approaches, such as methods for extracorporeal blood purification, the use of novel anti-infective substances or targeted immune modulation, there are still no therapeutic measures backed up by sufficient evidence that lead to a convincingly reduced mortality in the therapy of sepsis [132] . therapy with ivig is a treatment option that is associated with relatively high (but internationally disparate) costs and variable availability of treatment. a targeted selection process of potentially profiting patients on evidence-based criteria in terms of personalized medicine is preferable to an indiscriminate approach. unfortunately, a reliable and validated assessment of the cost-effectiveness in relation to the total costs of therapy based on available data currently remains difficult to obtain. since the approach of using purely anti-inflammatory therapies has been disappointing, the investigation of strategies aiming at balancing the immune system appears to be more appropriate [133] . given their multiple effects on inflammatory and immune mechanisms, the use of polyclonal intravenous immunoglobulins seems to be a promising approach to both modulate proand anti-inflammatory pathways. however, further clinical studies and research are clearly needed to substantiate the rationale for the use of immunoglobulins presented here, and to target the interventions to the right patient subset, at the right time, at the appropriate dose and for an optimal duration. summary: • the use of ivig in sepsis and septic shock appears to be safe. the use of iggam shows beneficial effects on sepsis-related inflammation and coagulopathy. for adjunctive sepsis therapy with ivig, igm-enriched formulations may be advantageous for specific patients. further clinical data are urgently needed to be able to make definitive statements on the cost-benefit ratio. funding: this research received no external funding. the third international consensus definitions for sepsis and septic shock global, regional, and national sepsis incidence and mortality, 19902-017: analysis for the global burden of disease study frequency and mortality of septic shock in europe and north america: a systematic review and meta-analysis cytokine storm and sepsis disease pathogenesis current gaps in sepsis immunology: new opportunities for translational research genome-wide transcription profiling of human sepsis: a systematic review pro-and anti-inflammatory responses are regulated simultaneously from the first moments of septic 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creative commons attribution (cc by) license key: cord-023935-o2ffxgnn authors: lorts, angela; cornell, timothy t.; shanley, thomas p. title: sepsis date: 2011-12-16 journal: pediatric critical care study guide doi: 10.1007/978-0-85729-923-9_27 sha: doc_id: 23935 cord_uid: o2ffxgnn the health care provider faced with the management of a child with septic shock relies on a comprehensive understanding of the numerous disciplines embodied in the practice of pediatric critical care medicine. the child with septic shock may have simultaneous derangements in the function of virtually every system of the body including: cardiovascular, respiratory, immune, renal, coagulation, hepatic, metabolic and neurologic. the degree to which physiologic alterations are manifest in a given patient is variable and influenced by multiple host and non-host factors including: the developmental stage, the presence of co-morbidities, pathogen-related factors, and genetic influences on both the host inflammatory response as well as the response to pharmacologic agents, all combining to have a profound influence on outcome. the clinician must possess a systematic and multifaceted approach to these critically ill patients. the goal of this chapter is to provide a comprehensive description of the epidemiology, biology and pathophysiology (at both the cellular and organ level) of sepsis, as well as outlining the current principles of managing septic shock. it will be apparent that optimal management requires a strong working knowledge of cardiovascular physiology, infectious diseases, multiple organ interactions, immunity, coagulation, pharmacology, and the molecular biology of inflammation. the health care provider faced with the management of a child with septic shock relies on a comprehensive understanding of the numerous disciplines embodied in the practice of pediatric critical care medicine. the child with septic shock may have simultaneous derangements in the function of virtually every system of the body including: cardiovascular, respiratory, immune, renal, coagulation, hepatic, metabolic and neurologic. the degree to which physiologic alterations are manifest in a given patient is variable and infl uenced by multiple host and non-host factors including: the developmental stage, the presence of comorbidities, pathogen-related factors, and genetic infl uences on both the host infl ammatory angela lorts , timothy t. cornell, and thomas p. shanley response as well as the response to pharmacologic agents, all combining to have a profound infl uence on outcome. the clinician must possess a systematic and multifaceted approach to these critically ill patients. the goal of this chapter is to provide a comprehensive description of the epidemiology, biology and pathophysiology (at both the cellular and organ level) of sepsis, as well as outlining the current principles of managing septic shock. it will be apparent that optimal management requires a strong working knowledge of cardiovascular physiology, infectious diseases, multiple organ interactions, immunity, coagulation, pharmacology, and the molecular biology of infl ammation. before reviewing the epidemiology of pediatric sepsis, it must be appreciated that the conclusions of prevalence studies have been obscured in the past by several factors including a lack of a reliable case defi nition. it has only been in the 1990s that consensus defi nitions for sepsis and septic shock were achieved. it was hoped that the development of standard defi nitions would not only enable accurate characterization of the epidemiology of sepsis, but also serve to stratify patients early in the course of sepsis for the purpose of clinical studies aimed at testing novel therapies. the most widely used defi nition of pediatric sepsis/septic shock is based on the 1992 american college of chest physicians/society of critical care medicine (accp/sccm) consensus conference, with adaptations for the pediatric population. the following four defi nitions resulted from these discussions: sirs , sepsis , septic shock , and severe sepsis . although there is overlap between some of these terms (particularly between septic shock and severe sepsis), each is intended to defi ne a particular patient population. longstanding clinical observations have identifi ed the presence of tachycardia, tachypnea, hyperthermia and leukocytosis as signs of infection, though these responses may also be present in the absence of any apparent infectious source. as a result, this physiologic response was defi ned as the systemic infl ammatory response syndrome (sirs). sirs defi nes a state of infl ammation/immune activation in a child and is based on the presence of at least two of the four criteria listed in table 27 -1 . thus, patients with diverse clinical conditions such as sepsis, pancreatitis, burns, or severe trauma can meet criteria for sirs. it has been argued that the sirs defi nition is non-specifi c and that too broad a range of patients are ultimately classifi ed as having sirs. nevertheless, the criteria have been widely used in both prescriptive and interventional studies to enhance the "capture" of all patients at risk for the subsequent development of severe sepsis or septic shock. sirs i s a state of infl ammatory/ immune activation and is based on the presence of at least two of the four following clinical criteria: temperature >38°c or <36°c, heart rate >90th percentile for age, respiratory rate >90th percentile for age, or hyperventilation to paco 2 < 32 mm hg. the defi nition attempts to "capture" all patients at risk for the subsequent development of severe sepsis or septic shock. sepsis is defi ned as a sirs response which is secondary to an infection, either documented by microbiology cultures or other clinical evidence of infection. severe sepsis is defi ned by sepsis criteria plus evidence of insuffi cient end organ perfusion (table 27-1 ) . finally, septic shock is defi ned by sepsis criteria plus hypotension (two distinct measurements <3rd percentile for age) after the administration of at least 20 ml/kg of crystalloid or colloid, in addition to the criteria listed for severe sepsis (table 27-1 ) . these criteria have been used extensively for conducting clinical investigations and have proven to be of value despite criticism for lack of both sensitivity and specifi city. the latest consensus conference was convened in 2007 to further refi ne the diagnostic criteria and therapeutic recommendations, with specifi c considerations for the pediatric population. published in 2008, the surviving sepsis campaign aims to improve the outcome in sepsis worldwide. the refi nement of pediatric-specifi c criteria for septic shock is also intended to aid future clinical trials and epidemiologic investigations in pediatric sepsis. the few published pediatric-specifi c studies illustrate the importance of sepsis in this age range. proulx analyzed the incidence and outcome of sirs, sepsis, severe sepsis, and septic shock in a single institution. over 1,000 admissions were analyzed over a 1-year period. sirs was present in 82% of patients, while 23% had sepsis, 4% had severe sepsis, and 2% had septic shock. the overall mortality for this population was 6% with a majority of deaths occurring in patients with multiple organ dysfunction syndrome (mods). an epidemiologic study using discharge international classifi cation of disease, 9th revision (icd-9) codes reviewed hospital records from seven large states representing nearly one-quarter of the united states population. while the criteria used for inpatient coding at discharge are not identical to accp/sccm consensus conference criteria, the study estimated an incidence of 42,371 cases of severe sepsis in individuals less than 20 years of age (0.6 cases/1,000 population). the highest incidence was in neonates (5.2 cases/1,000 population), compared to children ages 5-14 who had an incidence of 0.2 cases/1,000 population. the overall mortality rate was 10.3% (4,364 deaths nationally) consistent with the frequent observation that the mortality rate remains lower than comparable adult data. the study estimated an annual national health care cost of $1.7 billion associated with severe sepsis in children. a follow-up study with the same methodology appeared to show a 13% increase in the absolute number of cases of severe sepsis from 1995 and 1999 with the majority of this increase accounted for by severe sepsis in children less than 1 year of age. the mortality rate had decreased to 9.0% during this time period. collectively, these data illustrate that sepsis is a major health problem on the basis of incidence, mortality, and health care costs. there remains a need for further, well-designed epidemiologic studies of pediatric sepsis. future studies will enhance our understanding of not only epidemiology, but also the impact of new diagnostic and therapeutic approaches resulting from improved design of interventional trials specifi c to the pediatric population. sepsis is a systemic disease and can impact the functioning of all organ systems. the most common clinical manifestations of sepsis include: fever or hypothermia, tachypnea, tachycardia, leukocytosis or leukopenia, thrombocytopenia, and change in mental status. one of the earliest signs of infection is fever which results from the pyrogenic effect of cytokines, particularly interleukin (il)-1 b and tumor necrosis factor (tnf)-a . presentation with hypothermia can also occur, but is more common in infants. one traditional classifi cation of shock states divides this clinical state into three broad categories: hypovolemic, cardiogenic and distributive shock. the shock associated with 555 c hapter 27 • s eps is sepsis is unique in that all three forms are likely to be present. hypovolemic shock results from capillary leak, increased insensible losses, and decreased effective blood volume secondary to venodilation. cardiogenic shock is related to direct myocardial depression, the cause(s) of which remains the focus of investigation. finally, distributive shock is often apparent as brisk capillary refi ll, widened pulse pressure and bounding peripheral pulses and is caused by abnormally decreased systemic vascular resistance from pathologic vasodilation. the particular pattern of these hemodynamic physiologic perturbations manifested by any individual patient can be variable. some children have increased cardiac output with diminished systemic vascular resistance characteristic of distributive shock or the so-called "warm" shock state. in stark contrast to adults, in which this hemodynamic profi le (increased cardiac output/decreased systemic vascular resistance) is most common, children more frequently present with depressed cardiac output and elevated systemic vascular resistance. these patients appear cool with diminished pulses and poor capillary refi ll that is characteristic of the "cold" shock state. while important to recognize that patients may transition from one state to another, the presence of hypotension is often a late and particularly ominous sign that requires prompt intervention as its presence is associated with increased mortality. patients with sepsis often present with alterations in their respiratory system, notably tachypnea that refl ects a compensatory respiratory alkalosis aimed at neutralizing a metabolic acidosis related to hypoperfusion and anaerobic metabolism. chest x-ray fi ndings can reveal a small heart in the presence of hypovolemia with few vascular markings. alternatively, the combination of capillary leak, decreased myocardial function and the result of fl uid resuscitation in some children with sepsis can result in pulmonary edema. rapid progression to acute respiratory failure from ards is not uncommon. all organ systems and ultimately cellular functions are affected by poor perfusion and decreased oxygen delivery related to depressed cardiac and respiratory function. in addition, there may be direct injurious effects of bacterial toxins and circulating cytokines such as triggering of programmed cell death or apoptosis. the neurologic state of a child with sepsis is frequently altered and can range from agitation or irritability to frank obtundation. this depressed mental status can be present even in the absence of meningitis as a manifestation of cerebral hypoperfusion. skin manifestations are not uncommon and can include petechiae and purpura that are ominous signs of disseminated intravascular coagulation (dic) and purpura fulminans secondary to meningococcemia. diffuse erythema secondary to toxic shock syndromes can be present. there is also an increasing appreciation of sepsis-induced microvascular angiopathy contributing to distal skin and organ ischemia. an initial thorough and detailed physical exam provides both important clues to the diagnostic possibilities of pediatric septic shock and the underlying hemodynamic profi le. however, serial exams are imperative to follow pathophysiologic changes and to gauge the impact of therapeutic interventions in reversing the manifestations of shock. data from both clinical and basic science studies have supported the hypothesis that pathogens and/or their products initiate a host immune response that triggers widespread infl ammation causing tissue injury and organ dysfunction. potential initiating pathogens include gram-negative and gram-positive bacteria, viruses, fungi and protozoa. in some cases, overwhelming spread of pathogens (e.g. bacteremia) with release of toxins (e.g. endo-or exotoxins) may directly injure the host resulting in organ dysfunction. higher order organisms have evolved an immune system to eradicate pathogens which has evolved to include two systems: the innate or natural immune system and the acquired or adaptive immune system. the innate immune system is responsible for the highly conserved function of recognizing pathogens and mounting an effector response. it includes a series of molecules located on the cell surface termed pattern-recognition receptors (prr) which are capable of recognizing a broad array of conserved structures on a variety of children with sepsis may have hemodynamic characteristics that transcend traditional classifi cation. they often have elements of hypovolemia, cardiac dysfunction and abnormal vascular tone. in the septic child, the combination of capillary leak, decreased myocardial function and the result of fl uid resuscitation may result in rapid progression to acute respiratory failure. pathogens (so-called pathogen-associated molecular patterns, or pamp's). examples of pamp's include: lipopolysaccharide (lps) on gram-negative bacteria, lipoteichoic acid on gram-positive bacteria, mannans on yeast, double-stranded rna of rna viruses and unmethylated, cpg dna from bacteria. the effector responses that are regulated by the innate immune system (e.g. phagocytes, complement) are activated immediately upon infection and are designed to rapidly inhibit the replication of microorganisms. these cell surface pattern-recognition receptors (prr) are expressed on most antigen presenting cells of the innate immune system and represent diverse families of proteins. one group of prrs, the toll-like receptors (tlr), has been identifi ed as perhaps the most critical pathogen recognition receptor family in the context of sepsis biology. other families of prr include the c-type coll agenous lectins (collectins) that bind to a variety of carbohydrate moieties on cells, bacteria and viruses. most members of this family share structural homology to the complement protein c1q and can functionally substitute for c1q in activating the complement cascade. another family of prr possesses leucine-rich regions critical for protein-protein interactions that are necessary for immune recognition. examples of these leucine rich receptors include cd14, a receptor on the cell surface of macrophages that binds to lps and the macrophage scavenger receptor that binds to bacterial cell walls. unbound circulating prrs exist and include pentraxins, such as c-reactive protein, an acute phase reactant synthesized by the liver and lipopolysaccharide-binding protein (lbp) which binds to lps to optimize its binding to the cd14/toll-like receptor cellular complex. another key component of innate immunity is the complement system. the complement system is a complex cascade of proteins that possesses a broad array of anti-pathogen activities including: opsonization (c3), neutrophil chemotaxis (c5a), perforating cytotoxicity (c6-9, mac complex) and the ability to bind to and directly lyse viruses (c1). an in depth discussion of the role of complement in the response to infection is beyond the scope of this chapter, but has been recently summarized. in summary, the host possesses a ubiquitous and diverse set of pathogen recognition receptors which function to protect the host from infectious challenges, but at the expense of triggering powerful effector responses. paramount to effector responses of the innate immune system is a proinfl ammatory action of numerous cytokines and chemokines. these biologically active proteins are critical to the activation and recruitment of cellular components of the adaptive immune system. while necessary for pathogen clearance, this acute, proinfl ammatory immune response must also ultimately subside in order to reestablish homeostasis and avoid cellular and tissue damage. a key pathophysiologic feature of sepsis is that this immune response often appears to become unregulated resulting in an overwhelming proinfl ammatory response and host autodestruction. this characteristic systemic infl ammatory response seen frequently in response to infection can also be observed in association with non-infectious triggers (e.g. trauma, burns, pancreatitis, cardiopulmonary bypass). lps recognition: recent epidemiologic surveys of the causative agents of sepsis have indicated an increase in the incidence of gram-positive organisms such that there is a roughly equivalent prevalence between these and gram-negative organisms. historically, sepsis research has focused on the role of gram-negative bacteria in evoking a pathologic response. the structure of endotoxin shows three domains: an outer polysaccharide hydrophilic chain which determines the o-antigenicity, an acidic core region, and a lipid-rich region. gramnegative organisms possess endotoxins with variable repeats of mono-and heteropolysaccharides with complex side chain structure to provide a basis for distinct antigenicity. the o-region is linked via an acidic core to the lipid a region that is highly conserved and responsible for much of the toxicity attributed to intact lps. a series of seminal observations have determined the molecular mechanisms by which the classic pamp, lps, initiates a proinfl ammatory response. first, a strain of lps-resistant mice, the c3h/hej strain, was identifi ed and its resistance was found to be attributed to a single genetic mutation. second, it was shown that the lethal effects of endotoxin could be conferred by transfer of hematopoietic cells. endotoxin tolerant mice could be rendered the cells of the innate immune system contain cell surface molecules termed patternrecognition receptors (prr). these receptors are capable of recognizing a broad array of conserved structures on a variety of pathogens (so-called pathogenassociated molecular patterns, or pamp's). examples of pamp's include: lipopolysaccharide, lipoteichoic acid, viral rna and bacterial dna. toll-like receptors (tlr) are pathogen recognition receptors that have a critical role in sepsis. tlr4 is active in recognition of lps on gram-negative bacteria whereas tlr2 is active in the recognition of lipotechoic acid on gram-positive bacteria. a hallmark of sepsis is an immune response that appears to become unregulated resulting in an overwhelming proinfl ammatory response and host autodestruction. this characteristic systemic infl ammatory response is seen frequently in response to infection, but can also be observed in association with non-infectious triggers (e.g. trauma, burns, pancreatitis, cardiopulmonary bypass). c hapter 27 • s eps is lps-sensitive after reconstitution with hematopoietic cells derived from the monocyte/macrophage lineage from an lps-sensitive strain. third, stimulation of monocyte-derived cells with endotoxin resulted in production of several cytokines and chemokines critical to the systemic infl ammatory response. among these, tnf and il-1 were shown to be critical initiators of the septic response and could in fact mimic the endotoxin response. finally, the elucidation of the lps receptor assisted the identifi cation of those signal transduction pathways by which endotoxin triggers infl ammatory gene expression. lps receptor: membrane bound cd-14 was shown to be required for lps signaling. however, it lacked a transmembrane extension required for cytoplasmic signaling indicating the presence of additional components of the receptor complex. investigators working with drosophila had identifi ed a gene, toll, which was responsible for dorsoventral polarization in embryonic development. when toll was functionally mutated, it was demonstrated to play a key role in host defense against aspergillus fumigatus . homology between the toll-like receptors and the mammalian il-1 family of receptors was discovered and provided additional evidence that this family was crucial to the human innate immune response. finally, it was determined that the c3h/hej mouse strain which is hyporesponsive to lps possessed a mutation in toll-like receptor 4 (tlr4), providing further evidence that this receptor was necessary for lps signaling. tlr4 is one of ten mammalian toll-like receptors that have been cloned to date, each being activated by a specifi c set of ligands. since these discoveries, other members of the lps-receptor complex have been elucidated and include both md-2 and myd88. it is also known that circulating lpsbinding protein (lbp) facilitates lps binding to the cell surface receptor complex. together these components are able to "sense" lps at the cell surface and transmit this signal via a series of complex pathways. similarly, the products of gram-positive organisms, notably the cell wall component lipotechoic acid, activate cell activation through the related toll-like receptor 2 (tlr2). after engagement of cell surface receptors (e.g. tlr2 and tlr4), several important signal transduction pathways are activated that elicit a number of transcriptional factors responsible for infl ammatory gene expression. among these, the nuclear factor-k b (nf-k b ) and the mitogen activated protein kinase (mapk) pathways play a prominent role in regulating the expression of a number of infl ammatory gene products key to propagating the sepsis response. in the case of nf-k b, stimulation of the lps receptor causes phosphorylation of the inhibitor of k b kinases (i k k) which in turn phosphorylates the intracellular inhibitor of nf-k b, i kappa b (i k b). upon phosphorylation, i k b undergoes poly-ubiquination followed by proteosomal degradation. the removal of i k b effectively unmasks a nuclear translocation sequence on nf-k b enabling it to proceed into the nucleus to bind to nf-k b consensus sequences present on the promoter regions of many infl ammatory genes: cytokines including tnf, chemokines including il-8, adhesion molecules including e-selectin and others such as inos (see fig. 27 -1 ). the role of nf-k b in sepsis is supported by studies demonstrating that survivors and non-survivors of sepsis are distinguishable on the basis of nf-k b binding activity in peripheral blood mononuclear cells. in addition, in sepsis-induced ards, increased activation of nf-k b in macrophages obtained by bal is found in ards patients when compared to icu controls. to a similar degree, the mapk signaling pathways are important in mediating the septic response. three mapk pathways exist: p38 protein kinase, extracellular-regulated protein kinase (erk), and c-jun-terminal kinase (jnk). evidence exists for the role of each of these signaling pathways in sepsis. tnf production by neutrophils and macrophages is dependent on p38 activation. lps stimulation of monocytes activates jnk with downstream activation of activating protein-1 (ap-1) and subsequent il-1 b production. lps induction of tnf is in part dependent on erk pathway activation. together, these two pathways, nf-k b and mapk's, appear to be critical to the propagation of signals from the cell surface to the nucleus where expression of infl ammatory gene products occurs. as such, these pathways remain valid targets for future strategies in modulating the septic response. nuclear factork b (nfk b ) and the mitogen activated protein kinase (mapk) pathways play a prominent role in regulating the expression of a number of infl ammatory gene products key to propagating the sepsis response. while numerous proteins have been shown to play a role in the septic response, a full review of each protein's function is beyond the scope of this study guide. instead, we aim to highlight some known principle mediators in this cascade. evidence that tnf mediates the septic response stems from numerous observations: it is produced by hematopoietic cells, its expression is temporally related to the development of septic shock, recombinant tnf induces experimental septic shock in animals, and passive immunization against tnf attenuates endotoxin-mediated responses. tnf possesses numerous functions in infl ammation such as driving adhesion molecule and chemokine expression to facilitate leukocyte-endothelial cell adhesion; upregulating tissue factor and inhibition of protein c to create a pathologic procoagulant state in the vasculature; and inducing nitric oxide synthase (inos) which mediates pathologic vasodilation. in human studies, levels of tnf have been shown to correlate with mortality, with the development of shock and purpura fulminans and with the development of sepsis-induced ards and shock. the name, il-1, is now used to describe the family of proteins including two agonists (il-1 a and il-1 b ) and one antagonist, the il-1 receptor antagonist protein (il-1ra). il-1 b which is secreted, mediates much of the systemic effects attributed to il-1 release in sepsis. synthesized as a propeptide, il-1 b requires proteolytic cleavage by the il-1 converting enzyme (ice) to become bioactive. il-1 b utilizes the 80-kda type i receptor which is tnf possesses numerous functions in infl ammation such as inducing adhesion molecules and chemokines that facilitate leukocyte-endothelial cell adhesion and inducing nitric oxide synthase (inos) which mediates pathologic vasodilation. tnf also upregulates tissue factor and inhibits protein c to create a pathologic procoagulant state in the vasculature. clinically, levels of tnf correlate with mortality, the development of shock and purpura fulminans and with the development of sepsis-induced ards and shock. p p iκ κ κ κk associated with a number of adapter proteins (e.g. myd88,tnf receptor-associated factor 6 (traf6) and interleukin-1 receptor-associated kinase (irak)) to propagate signals through both the nf-k b and ap-1 pathways. il-1 b infusion elicits fever, hypotension and leukocytic infi ltration to the lungs. in a manner similar to tnf, il-1 stimulates monocyte activation and phagocytosis, increases adhesion molecule expression, and increases tissue factor expression while inhibiting thrombomodulin secretion, thus creating a procoagulant state. when detected in the circulation of septic patients, il-1 levels also correlate with mortality. of note, the il-1ra is a circulating inhibitor of il-1 b that binds to the il-1 receptor without initiating a signal. the expression of il-1ra has been shown to follow peak expression of il-1. it is speculated that il-1ra is an endogenous regulator of il-1 effects. however, in clinical trials, il-1ra infusion failed to improve mortality in sepsis. furthering our molecular understanding of sepsis-induced organ dysfunction was the identifi cation of the "leukocyte-endothelial cell adhesion cascade". this cascade is characterized by cytokine activation of the selectin family of adhesion molecules (e.g. e-selectin) on the endothelium which initiate a process of neutrophil "rolling" via interaction with sialyated moieties constitutively present on circulating neutrophils. activation of the "rolling" neutrophil results in both increased expression and activation of the integrins which in turn bind to intercellular adhesion molecule (icam)-1 that is upregulated on the endothelial cell surface by tnf and il-1 b . this integrin-icam-1 interaction mediates fi rm adhesion of the neutrophil to the endothelial cell surface. finally, in response to various chemotactic cytokines or chemokines, neutrophils migrate to the site of infl ammation. release of both oxygen-and nitrogen-based radical species and proteases by the neutrophils may ultimately contribute to cellular injury and organ dysfunction. nitric oxide (no) is responsible for endothelium-derived relaxation of blood vessels. three isoforms of nitric oxide synthase are responsible for production of no: type i, a neuronal isoform (nnos); type ii, an inducible isoform (inos) and type iii, a constitutive, endothelial isoform (enos). tnf and il-1 b are capable of inducing inos and increased levels of circulating stable byproducts of no are found in both septic adults and children who simultaneously display low systemic vascular tone. this supports the hypothesis that no plays a principal role in septic shock via pathologic vasodilation. it has also been suggested that tnf and il-1 b may be the so-called "myocardial depressant factors" by increasing circulating no through induction of inos ; however, it is not clear that no is the exclusive mediator of these effects. in light of the evidence supporting the role of no in septic shock, clinical trials employing no synthesis inhibitors in septic shock were initiated. though early clinical reports and small studies reported that nos inhibitors could signifi cantly improve blood pressure, this was at the expense of decreasing cardiac output secondary to increased afterload. as a not uncommon hemodynamic profi le in pediatric septic shock is decreased cardiac output and elevated systemic vascular resistance, it is not cleat that nos inhibitors will have a therapeutic role in pediatric (or adult) sepsis in the future. studies employing agents directed against the early mediators of the septic response have been mostly ineffectual. this has led to the hypothesis that additional molecules with delayed kinetics of expression may infl uence the outcome in sepsis. as an example, it was observed that lps-challenged mice often die long after peak expressions of tnf and il-1 b suggesting that late-acting proteins may contribute to endotoxin-induced mortality. investigators searching for late expressed proteins identifi ed a member of the high mobility group (hmg)-1 non-histone chromosomal protein family in conditioned media 16 h after lps-stimulation of macrophages. this protein renamed hmgb1 is a known ligand for the receptor for advanced glycation end products (rage). the rage receptor is expressed on monocytes and vascular smooth muscle. binding by hmgb1 activates both the nf-k b and mapk pathways. increased expression of hmgb1 was found in endotoxemic mice and in critically ill patients with surgical sepsis where increased levels correlated with non-survival. in animal models, the blockade of hmgb1 can inhibit the infl ammation associated with endotoxemia, cecal ligation and puncture and lps-triggered acute lung injury. identifi cation of hmgb1 and similar "late" mediators may provide a broader therapeutic window for successful immune modulating therapy in sepsis. regulatory processes and mediators exist for the purpose of modulation and eventual resolution of infl ammation and the septic response. an absence in the decline of proinfl ammatory mediators such as tnf and il-6 over the course of sepsis is an associated risk factor for mortality. monocyte activation results not only in production of proinfl ammatory cytokines, but also expression of a number of endogenous cytokine antagonists including soluble tnf receptors, the il-1ra and additional anti-infl ammatory cytokines, such as il-10 and transforming growth factor-β (tgf-b ). il-10 has multiple anti-infl ammatory properties including inhibition of cytokine production from activated monocytes. il-10 inhibits expression of those cytokines known to contribute to sepsis, as well as important chemokines, including il-8. in addition, il-10 increases expression of other anti-infl ammatory molecules such as il-1ra and soluble tnf receptors. exogenous administration of il-10 in various experimental models has been used in an attempt to decrease infl ammatory cytokines and diminish organ injury. human studies showed that patients who did not survive ards had lower levels of il-10 in their bal fl uid compared to survivors. furthermore, the inability to increase il-10 in response to meningococcal infection was associated with increased mortality. thus, il-10 and additional regulatory cytokines (e.g. tgf-b , il-13) possess a number of anti-infl ammatory properties and are important contributors to the endogenous regulation of the acute septic response. dysregulation of the coagulation cascade occurs in sepsis as refl ected by activation of procoagulant pathways, consumption of clotting factors, alterations in fi brinolysis, and reduced anticoagulant activity. a common hematologic alteration in sepsis is the development of disseminated intravascular coagulation (dic) which is an acquired state of activation of coagulation and intravascular fi brin formation resulting in vascular thrombosis. in addition to proinfl ammatory cytokines, tissue factor (tf) activation also plays a prominent role in activating the coagulation cascade, initiating fi brin formation and contributing to the development of dic. concurrent with enhanced production of fi brin, there is decreased fi brinolysis related to increased plasminogen activator inhibitor type 1 (pai-1), as well as dysfunction and/or depletion of antithrombin iii, protein c, protein s and tissue factor pathway inhibitor (tfpi). at iii which inhibits thrombin by forming thrombin-antithrombin (tat) complexes is decreased in sepsis related to degradation by elastases from activated neutrophils, dilution secondary to volume resuscitation, and impaired hepatic synthesis. despite a correlation between low at iii levels and mortality in patients with sepsis, replacement trials of at iii have failed to show a signifi cant effect on improving mortality. protein c is also noted to be depleted among patients with sepsis and septic shock. regulation of activation of protein c to activated protein c (apc) in the coagulation cascade is mediated in a complex manner and will not be discussed here. apc, upon dissociation from its receptor, binds to its co-factor, protein s, to subsequently inactivate factors va or viiia, thus playing a key role in inhibiting coagulation. it is both antithrombotic and profibrinolytic. apc also possesses anti-infl ammatory activity. in models of endotoxemia, apc infusion decreases cytokine production and attenuates neutrophil activation. these antiinfl ammatory effects appear to be independent of apc's anticoagulant effect. following an absence in the decline of proinfl ammatory mediators such as tnf and il-6 over the course of sepsis is an associated risk factor for mortality. monocyte activation results not only in production of proinfl ammatory cytokines, but also expression of a number of endogenous anti-infl ammatory cytokines including soluble tnf receptors, the il-1ra and additional anti-infl ammatory cytokines, such as il-10 and tgfb . important anti-infl ammatory molecules include il-10, il-1ra and soluble tnf receptors. il-10 inhibits expression of proinfl ammatory cytokines known to contribute to sepsis, as well as important chemokines, including il-8. in addition, il-10 increases expression of other anti-infl ammatory molecules such as il-1ra and soluble tnf receptors. at iii inhibits thrombin by forming thrombin-antithrombin (tat) complexes. at iii is decreased in sepsis due to degradation by elastases from activated neutrophils, dilution secondary to volume resuscitation, and impaired hepatic synthesis. despite a correlation between low at iii levels and mortality in patients with sepsis, replacement trials of at iii have failed to show a signifi cant effect on improving mortality. these encouraging pre-clinical studies, clinical trials examining the effect of apc on mortality from sepsis were commenced culminating in the protein c worldwide evaluation in severe sepsis (prowess) trial. in this study, apc was associated with a statistically signifi cant reduction in 28-day mortality in septic adults. however, a recent pediatric study was stopped after an interim analysis showed that apc administration was highly unlikely to show improvement in outcome ( fig. 27-2 ). it is not uncommon to observe that patients exposed to seemingly identical pathogen insults display strikingly different pathophysiology and outcomes. it is believed that genetic differences among hosts are at least in part responsible for this variability in sepsis responses. as mentioned previously, the insensitivity to lps in the c3h/hej mouse line was mediated by a mutation in the coding sequence for tlr4. similar fi ndings of an attenuated response to pathogen stimulation have now been reported in patients with mutations in both the tlr4 and tlr2 gene. the polymorphism in tlr2 appears to confer an increased predisposition to severe gram-positive bacterial infections. more recently, a polymorphism within the cd14 promoter gene (c to t transition at base pair -159) was identifi ed with a particular genotype over-represented among septic shock patients compared to healthy controls. among the septic patients, the presence of this genotype also was associated with a signifi cantly higher mortality (71% versus 48%). these studies support the concept that genetic alterations in those genes known to participate in the septic response affect the host immune response and likelihood of survival. for a more complete review of the numerous examples of genetic alterations in key infl ammatory genes, the reader is directed to the suggested readings. the administration of activated protein c was associated with a statistically signifi cant reduction in 28-day mortality in septic adults. however, a recent pediatric study was stopped after an interim analysis showed that apc administration was highly unlikely to show improvement in outcome. the coagulation cascade in sepsis (reprinted with permission from bernard 2001 ) as the cellular response to sepsis has become better understood, the approach to treatment of sepsis has become broader. the treatment of sepsis involves four important components: initial resuscitation, elimination of pathogen, maintenance of oxygen delivery, and carefully directed regulation of the infl ammatory response. as reviewed above, sepsis is an immunologically complex response to an invasive pathogen necessitating tight physiologic regulation in order to eradicate the organism while maintaining cellular and organ homeostasis. in cases where the immunologic and infl ammatory responses continue to escalate, numerous pathways are altered and may ultimately prove amenable to immune modulating therapy, but this approach has been unsuccessful to date. the initial priority in the treatment of the septic child is respiratory and cardiovascular stabilization. the primary goals of therapy in those initial hours following clinical presentation are to maintain oxygenation and ventilation, achieve normal perfusion and blood pressure, and re-establish appropriate urine output for age. children with sepsis may have altered mental status which, if profound, raises concern about the ability to protect the airway. tachypnea associated with a primary or compensatory respiratory alkalosis is commonly present. the combination of increased lung vascular permeability and aggressive fl uid resuscitation to restore intravascular volume and maintain blood pressure may contribute to the subsequent development of pulmonary edema. in children with lung edema, the related changes in lung compliance and loss of functional residual capacity can dramatically increase the work of breathing ultimately necessitating tracheal intubation and mechanical ventilatory support. arterial blood gas analysis may show hypoxemia and metabolic acidosis; however, the decision to provide mechanical ventilatory support should not be based solely on laboratory fi ndings. the presence of increased work of breathing, hypoventilation or obtundation are all indications for instituting mechanical ventilatory support which holds additional benefi t in decreasing the overall oxygen consumption, especially when combined with sedation and paralysis. it should be stated, however, that children with warm shock can commonly be managed without endotracheal intubation so long as they are not obtunded or fl uid overloaded. disorientation or lethargy with intact responsiveness does not require placement of an artifi cial airway as many institutions manage these patients without intubation. the work of breathing associated with hyperventilation in the absence of pulmonary edema is not clinically signifi cant. furthermore, there is no evidence that decreasing work of breathing in the presence of distributive shock will result in redistribution of nutrient fl ow to vital organs, the very nature of distributive shock. however, it is more common for infants to present with cardiac dysfunction and pulmonary edema or seriously altered mental status requiring endotracheal intubation and mechanical ventilation. correction of intravascular volume depletion should be made prior to the institution of positive pressure ventilation. the decrease in venous return after the initiation of positive pressure ventilation may lead to further hemodynamic compromise in the child with intravascular volume depletion. caution should also be taken in choosing sedative agents for intubation, using agents that have the least impact on tenuous hemodynamics (e.g. ketamine). controversy exists over the adrenal suppressive effect of a single dose of etomidate when used for intubation in septic children and adults. the pediatric critical care clinician should be aware of the concern for adrenal suppression following a single dose of etomidate used for the intubation of children with septic shock and the published guidelines which do not recommend its use in this setting. following intubation, attention must be paid to matching the mechanically provided minute ventilation to that which was present during spontaneous respiratory effort so that respiratory compensation of acidemia is preserved. if it is deemed that positive pressure ventilation is not needed, supplemental oxygen should be provided to maintain normal oxygen saturations. treatment of sepsis involves four important components: initial resuscitation, elimination of pathogen, maintenance of oxygen delivery, and carefully directed regulation of the infl ammatory response. with regard to fl uid status, septic children have decreased effective intravascular volume related to a number of causes. poor oral intake of fl uid for a period of time prior to clinical presentation is common. increased vascular permeability leads to intravascular volume loss due to extravasation of fl uid from the vascular space, so-called "third spacing". finally, the no-mediated vasodilation reviewed above increases vascular capacitance thereby decreasing the effective circulating volume. thus, when sepsis is suspected, it is imperative to expeditiously achieve vascular access and initiate fl uid resuscitation with 20 ml/kg of isotonic fl uid as quickly as possible. while debate continues as to the most effective fl uid for resuscitation, no pediatric literature exists to support colloid over crystalloid, the latter of which was recently demonstrated to be equally effective in a large adult icu trial. there is some support for using colloid fl uid in patients with a narrow pulse pressure; however, this practice is not supported by any large, well-designed clinical studies. while following the clinical exam for signs of intravascular volume overload (new onset of rales, increased work of breathing, development of a gallop, or hepatomegaly), fl uid should be administered quickly with the goal of monitoring heart rate response, urine output, capillary refi ll time and level of consciousness. initial fl uid resuscitation of the child with septic shock commonly requires a volume of up to or greater than 60 ml/kg in the fi rst hour and one retrospective study demonstrated an increased survival in children given fl uid volumes of 40 ml/kg or more within the fi rst hour. the accm clinical practice parameters for hemodynamic support of pediatric and neonatal septic shock recommend that the child with septic shock be repeatedly examined for the development of "rales, gallop rhythm, hepatomegaly, and increased work of breathing" during volume loading, and that in the absence of such fi ndings, volumes up to 200 ml/kg can be administered in the fi rst hour. these guidelines further state that "the rate of fl uid administration should be reduced substantially when there are clinical signs of adequate cardiac fi lling without hemodynamic improvement." despite on-going fl uid resuscitation, hypotension and inadequate organ perfusion may persist requiring the initiation of inotropes and/or vasopressors. in children, vasoactive medicines should only be given in addition to fl uid resuscitation. however, consensus guidelines recommend that vasoactive infusions may be necessary in some cases to sustain perfusion pressure even when hypovolemia is not yet resolved. dopamine is the most common fi rst choice agent selected for hemodynamic support in those patients with fl uid-refractory shock. dopamine provides inotropic support at lower concentrations; however, it is often necessary to increase it to higher doses that provide vasopressor activity (up to 20 m g/kg/min) to maintain adequate tissue perfusion. the decision of which agent to add in the setting of dopamine-refractory shock should be based on the underlying cause of cardiovascular compromise. for example, if hemodynamic instability is related to low cardiac output from direct cardio-depressant effects then increased inotropy from dobutamine or low-dose epinephrine may be indicated. if hypotension persists secondary to decreased vascular tone, then agents such as epinephrine and norepinephrine dosed in the alpha agonist range should be considered. finally, in children who demonstrate low cardiac output and/or increased afterload from vasoconstriction (i.e. increased systemic vascular resistance), agents with primarily inotropic or vasodilator function including milrinone, dobutamine or short-acting nitrovasodilators can been considered in the fl uid-resuscitated, normotensive child. as in evaluating the adequacy of fl uid resuscitation, similar clinical parameters should be followed in titrating vasoactive medications. appropriate endpoints include: capillary refi ll time less than 2 s, normal peripheral pulses, warm extremities, urine output greater than 0.5 ml/kg/h, improved mentation, resolving acidemia, decreasing serum lactate and when available, superior vena cava (svc) oxygen saturation greater than 70%. invasive monitoring can further assist the clinician with goal directed endpoints. although frequent beside examination remains integral to the care of the child in septic shock, an additional task in the initial resuscitation phase is placement of appropriate and necessary vascular access and monitors. central venous access is a necessity for the child with fl uid refractory shock to provide for delivery of vasoactive medicines and large volumes of fl uid. these catheters can be useful for following the central venous pressure (cvp) during fl uid adequate volume resuscitation of the child with septic shock commonly requires a volume of up to or greater than 60 ml/kg in the fi rst hour. appropriate endpoints of sepsis resuscitation include: capillary refi ll time less than 2 s, normal peripheral pulses, warm extremities, urine output greater than 0.5 ml/kg/h, improved mentation, resolving acidemia, decreasing serum lactate and when available, superior vena cava oxygen saturation greater than 70%. administration. finally, when the tip is located in the superior vena cava, blood sampling can provide an approximate measure of the mixed venous oxygen saturation which has been validated as a critical target in adult shock resuscitation. the decision regarding the access site for a central venous catheter is dictated by a number of mitigating factors such as the experience level of the operator and the presence of coagulopathy. femoral catheters, in the absence of abdominal pathology, can be used to estimate cvp with good correlation. the cvp measured in the abdominal inferior vena cava must be assessed carefully as a low cvp can be a reliable indicator of hypovolemia, however, a normal or high cvp in the presence of abdominal distention does not automatically exclude the presence of hypovolemia. multiple adult studies have demonstrated that even an accurate intra-thoracic cvp may be a poor approximation of left ventricular end diastolic pressure and volume. the cvp can be elevated despite the presence of hypovolemia if pulmonary hypertension, right ventricular dysfunction with poor diastolic compliance, tricuspid regurgitation, cardiac tamponade or an intracardiac left-to-right shunt exists. even though precise determination of the true mixed venous saturation requires the presence of a pulmonary artery catheter, the approximations derived from the svc saturation have proven a useful target in septic adults. in contrast, because of differences in oxygen extraction between the upper extremities, abdomen, and lower extremities, venous oxygen saturations from a low lying femoral line do not accurately correlate with those measured in the pulmonary artery. consensus guidelines recommend therapeutic endpoints of superior vena cava oxygen saturation >70% or mixed venous (pulmonary artery) oxygen saturation >65%. placement of an intra-arterial catheter provides continuous monitoring of systemic blood pressure, pulse pressure and hemodynamic variation with respiration, as well as a means for drawing arterial blood gases, lactate levels and additional laboratory studies. the arterial blood also provides the most accurate measure of arterial oxygen content and can be used to both assess the function of the lungs and to maximize oxygen delivery. in the ventilated patient, variation in the amplitude of the arterial waveform has been found to correlate closely with intravascular volume status (see also chapter 3 ). systolic pressure variation (spv), also referred to as "reverse pulsus paradoxus," is the variation in beat-to-beat amplitude of the arterial pulse during positive pressure ventilation. a single positive pressure breath normally affects the arterial pressure in a biphasic manner. the initial response to a positive pressure breath is to "squeeze" pulmonary vascular blood into the left atrium (the opposite, "pooling" of blood, occurs with negative pressure inspiration) leading to a rise in systolic pressure. in addition, positive intrathoracic pressure reduces the afterload on the left ventricle by virtue of the pressure gradient from the thorax outward further augmenting this early rise in arterial pressure. these two effects produce an upward movement of the systolic blood pressure coincident with the positive pressure breath, referred to as the d up component of spv. following this d up, a fall in systolic pressure occurs a few beats later as the decreased venous return (preload) to the right ventricle that occurred during positive pressure inspiration is now evident as decreased preload to the left ventricle after a few cardiac cycles. the transient reduction in right ventricular volume and output leads to a smaller left ventricular stroke volume and a brief reduction in arterial pressure that occurs later in the ventilator cycle ( d down). an exaggerated spv (>10 mm hg) has been seen early in the setting of hypovolemia. this is due to a greater d down component. several studies have shown that an increase in the spv occurs prior to a fall in arterial pressure and may a better predictor of hypovolemia than a low pulmonary capillary wedge pressure (pcwp) (<10 mm hg). an increase in the spv due to a greater d down component can also occur due to high airway pressures causing decreased venous return. recently, pulse pressure variation (ppv) has also been found to be a sensitive indicator of preload, and more importantly, fl uid responsiveness. ppv is defi ned as the maximal pulse pressure (systolic minus diastolic blood pressure) less the minimum pulse pressure divided by the average of these two pressures. the use of systolic pressure variation and pulse pressure variation is limited to those patients on mechanical ventilation. these measurements should occur when there is no spontaneous breathing. in the presence of a consistent delivered tidal volume, systolic pressure variation can be used to track the adequacy of intravascular volume over time (fig. 27-3 ) . the decision to use a pulmonary catheter (pac) with the goals of optimizing left ventricular preload, monitoring cardiac index and measuring oxygen delivery remains controversial. caveats to interpretation of pac data include the presence of an intracardiac shunt and an abnormally functioning mitral valve or other obstructed left heart lesion as either shunting, regurgitation, or inaccurate pressure determinations will alter cardiac index and/or the pulmonary capillary wedge pressure measurements. as previous data in adults have shown no benefi t of pac use, a recent consensus statement regarding their use stated that the role of pac remains unclear. studies of children with septic shock have shown that the information obtained from a pac aided in identifying hemodynamic profi les different from those presumed by care givers and has directly infl uenced care decisions. it was concluded by a recent consensus panel to be of potential benefi t in improving the management of pediatric patients. pulmonary artery catheter placement should be considered for pediatric patients who remain in shock after resuscitation and initiation of the usual vasoactive agents in whom the fl uid status and cardiac function remains unclear. in this setting, therapeutic endpoints are a cardiac index of >3.3 and <6.0 l/min/m 2 and systemic and pulmonary vascular resistances within the normal range. early identifi cation of a possible offending pathogen and aggressive source control represent a crucial component of septic shock therapy. prompt initiation of appropriate antimicrobial therapy against the causative pathogen has been shown to be one of the most important predictors of outcome. in a 1980's study of over 1,100 adults, providing appropriate systolic pressure variation (spv) and pulse pressure variation (ppv) can accurately predict which patients will be responsive to further fl uid resuscitation (gunn and pinsky 2001 ) . antimicrobial coverage at least 1 day prior to identifi cation of the organism was associated with improved survival. the pathogen itself has prognostic signifi cance. fungal infections, while accounting for only a minority of sepsis cases, carry the lowest survival rate, followed by gram-positive and gram-negative bacteria. survival rate has been reported to be the highest in patients in whom no pathogen was identifi ed. because of the importance of appropriate antimicrobial therapy, the decision of which agents to empirically start must balance potential side effects versus maximizing coverage. in this respect, it is important to be familiar not only with the most common causative pathogens, but also the local icu nosocomial risks and pathogen resistance patterns. initially, broad antibiotic coverage is initiated. neonates are most frequently placed on ampicillin and an aminoglycoside (e.g. gentamicin) or a third generation cephalosporin such as cefotaxime. in infants and children over the age of 4-6 weeks, the decision to start vancomycin empirically should be considered in light of the increasing antibiotic resistance of streptococcus pneumoniae and rising incidence of community acquired methicillin resistant staphylococcus aureus (mrsa). in addition, a 3rd or 4th generation cephalosporin (e.g. ceftriaxone) should be used. suspicion of a gram-negative infection or nosocomial infection requires additional coverage, usually in the form of an aminoglycoside, for the possibility of pseudmonas species and other resistant gram-negative organisms. because of its broad coverage, including many anaerobic species, and low renal toxicity, piperacillin/tazobactam is empirically administered with increasing frequency. the antiviral agent, acyclovir, should be administered if there is suspicion of a herpes virus infection. in immunocompetent children, the decision to start empiric antifungal therapy remains controversial. in the child who is not improving over the initial days of empiric coverage or in whom there is a higher risk for fungal infection (e.g. presence of indwelling devices, immunosuppression or other signifi cant co-morbidities), antifungal coverage may be indicated. the development of agents equally as effective as amphotericin, but with substantially reduced nephrotoxicity such as fl uconazole and caspofungin, may ultimately sway the risk/benefi t analysis towards more aggressive, earlier initiation of empiric antifungal coverage in select, high risk populations. the ability to narrow the spectrum of treatment once the causative organism has been identifi ed will reduce the number of potential side effects and curtail the development of pathogen resistance related to imprudent use of broad spectrum antibiotics. the current mainstay of supportive care in sepsis remains the maintenance of adequate oxygen delivery in the face of myocardial depression, capillary leak, acidosis, and massive cytokine release. while some early adult studies have suggested improved outcomes when achieving supra-normal levels of oxygen delivery, this approach in pediatric sepsis remains unproven. this is likely due to the fact that septic patients may have a perturbed ability to extract oxygen in addition to suboptimal oxygen delivery. clinically, this impairment in cellular oxygen uptake may be refl ected by an inappropriately high central venous oxygen saturation (s cv o 2 ) in the face of a progressive and therapy refractory acidosis. optimizing appropriate oxygen delivery remains a clinical goal and incorporates the need for maximizing oxygen carrying capacity. while there is no recommended hemoglobin level for children, the most recent nih consensus conference suggested a hemoglobin concentration of 10 g/dl for adults with cardiopulmonary compromise as part of a protocol toward achieving the therapeutic goal of s cv o 2 >70%, with improved outcomes demonstrated when this goal was achieved during initial resuscitation. in the context of fl uid loading with blood transfusion, empiric administration of diuretics to eliminate extra fl uid should be avoided until hemodynamic stability has been achieved or if the child exhibits signs of intravascular volume overload defi ned earlier in this chapter. similar clinical parameters can be assessed in response to blood transfusion. perhaps the best assessments are determinations that provide indirect evidence of the balance between oxygen delivery and consumption such as lactate levels and mixed venous oxygen saturation. in the context of fl uid loading with blood transfusion, empiric administration of diuretics to eliminate extra fl uid should be avoided until hemodynamic stability has been achieved or if the child exhibits signs of intravascular volume overload. finally, the nutritional status of the septic child must be addressed. patients with sepsis often have poor nutrition prior to admission to the picu and often may not be fed in the fi rst few days of illness. this state combined with the increased metabolic rate associated with sepsis place the septic patient at risk for protein calorie malnutrition. intestinal hypoperfusion in combination with absence of local enterocyte nutrition can cause mucosal barrier dysfunction and may contribute to translocation of bacteria and endotoxin from the intestine into the blood stream. while the use of enteral feeding in critical illness has been shown to improve survival and decrease hospital stay, its use must be balanced with the risk of stressing intestinal function in the face of poor splanchnic perfusion, especially in the child requiring the use of vasopressors such as epinephrine and norepinephrine. regardless of which mode of nutrition is chosen, the goal of achieving nitrogen balance is important for allowing recovery and return to physiologic homeostasis. in the absence of enteral feedings, protection from stress-related gastrointestinal ulcer formation is advised. because poor outcome in sepsis has been attributed to a dysregulated proinfl ammatory state, anti-infl ammatory agents, such as corticosteroids, have long been proposed as a potential therapeutic strategy. anecdotally, it has been observed that some patients treated with antibiotics appear to acutely worsen in a time frame consistent with the onset of antibiotic activity. this observation has been attributed to massive release of bacterial products following the lysis of high numbers of bacteria. to this end, investigators had shown that animals treated with anti-infl ammatory drugs prior to receiving antibiotics demonstrated a less severe response to bacterial lysis. despite encouraging preclinical studies, two subsequent large adult trials using high dose steroids early in sepsis showed no improvement in mortality. more recently, studies using lower doses of steroids over a longer period of time have suggested a possible benefi t including a reduced time to cessation of vasopressor therapy. these more recent observations have stimulated a resurgence in the use of corticosteroids in sepsis. adrenal insuffi ciency is frequently unrecognized in children with septic shock and a low basal circulating cortisol level, especially in association with an abnormal corticotropin stimulation test, has been associated with higher mortality rates. therefore, identifying "at risk" patients with a corticotropin stimulation test and treating this group may improve outcome in sepsis. a study in which adult septic shock patients who were classifi ed as "nonresponders" based on corticotropin stimulation results were treated with hydrocortisone and fl udrocortisone for 7 days and had a signifi cantly reduced risk of death. however, this initial observation was not observed in larger follow-up studies. thus, the use of hydrocortisone and the application of a corticotropin stimulation test in septic patients remains highly controversial. in pediatrics, it is currently recommended that any child with fl uid-and catecholamine-refractory shock (inadequate response to two or more vasoactive agents), has a known history of adrenal insuffi ciency, or has previously received exogenous steroids should be considered for steroid replacement with hydrocortisone (usual dose between 50 and 100 mg/m 2 /day divided every 6 h). because the host response to sepsis is mediated by circulating infl ammatory molecules, it has been hypothesized that extracorporeal removal of these mediators via hemofi ltration or exchange transfusion may affect outcome. case reports suggest that arterial oxygenation and hemodynamics can be improved with use of hemofi ltration during sepsis and multiple organ failure. however, there exist many mitigating factors in evaluating the pediatric experience and the effi cacy of hemofi ltration remains unproven. challenges with instituting extracorporeal hemofi ltration include diffi culty with vascular access in smaller children, potential fl uid and electrolyte imbalance, hypothermia, anticoagulation requirements and acutely compromised hemodynamics during initiation. in addition, it is not known whether benefi cial proteins such as albumin, immunoglobulins, clotting factors and counter-regulatory cytokines are removed during this process. while experience shows that hemofi ltration can be safely performed in children with sepsis, it remains unclear if it will improve outcome. part of the infl ammatory response involves cytokines that cause widespread activation of the coagulation cascade with suppression of fi brinolysis as reviewed above. it is encouraging adrenal insuffi ciency is frequently unrecognized in children with septic shock. a low basal circulating cortisol level, especially in association with an abnormal corticotropin stimulation test, has been associated with higher mortality rates. that administration of activated protein c in adults with septic shock was associated with a signifi cant decrease in 28-day mortality. though activated protein c should not be routinely used in pediatric sepsis, indications for its use may be determined from further trials in pediatric sepsis. in the meantime, many other potential immune modulating therapeutic agents have been identifi ed and are currently under investigation. unfortunately, many of the antiinfl ammatory agents tried to date (anti-il-1, anti-bradykinin, anti-endotoxin, anti-tnf-a , soluble tnf receptor and anti-platelet activating factor) have not shown any benefi t in large, randomized clinical trials. it is hoped that improvements in study design which include thoughtful stratifi cation of patients, timely identifi cation of the presence or absence of a pathogen and consideration of genetic factors that infl uence outcome will eventually assist in discovering and targeting pharmacologic agents that ultimately improve the outcome of the pediatric patient with septic shock. vascular bed-specifi c hemostasis: role of endothelium in sepsis pathogenesis corticosteroids for severe sepsis and septic shock: a systematic review and meta-analysis immunological therapy of sepsis: experimental therapies effi cacy and safety of recombinant human activated protein c for severe sepsis signal transduction during innate and adaptive immunity role of nfkappab in the mortality of sepsis new insights into its pathogenesis and treatment the accp-sccm consensus conference on sepsis and organ failure stress doses of hydrocortisone reverse hyperdynamic septic shock: a prospective, randomized, double-blind, single-center study clinical practice parameters for hemodynamic support of pediatric and neonatal septic shock: 2007 update from the american college of critical care medicine toll-like receptors: molecular mechanisms of the mammalian immune response prognostic values of tumor necrosis factor/cachectin, interleukin-1, interferon-alpha, and interferon-gamma in the serum of patients with septic shock. swiss-dutch j5 immunoglobulin study group hemodynamic support in fl uid-refractory pediatric septic shock surviving sepsis campaign guidelines for management of severe sepsis and septic shock surviving sepsis campaign: international guidelines for management of severe sepsis and septic shock organization and regulation of mitogenactivated protein kinase signaling pathways lps induction of gene expression in human monocytes implications of arterial pressure variation in patients in the intensive care unit signal transduction by the c-jun n-terminal kinase (jnk) -from infl ammation to development the i kappa b kinase (ikk) and nf-kappa b: key elements of proinfl ammatory signalling use of corticosteroid therapy in patients with sepsis and septic shock: an evidence-based review myocardial dysfunction in septic shock: part ii. role of cytokines and nitric oxide rationale for restoration of physiological anticoagulant pathways in patients with sepsis and disseminated intravascular coagulation genomic polymorphisms in sepsis the contrasting effects of dopamine and norepinephrine on systemic and splanchnic oxygen utilization in hyperdynamic sepsis early enteral nutrition in acutely ill patients: a systematic review source control in the management of severe sepsis and septic shock: an evidence-based review role of the coagulation system in the local and systemic infl ammatory response new rationale for glucocorticoid treatment in septic shock anti-infl ammatory cytokines pediatric considerations epidemiology of sepsis and multiple organ dysfunction syndrome in children pulmonary artery catheter consensus conference: consensus statement drotrecogin alfa continuous plasmafi ltration in sepsis syndrome. plasmafi ltration in sepsis study group early goal-directed therapy in the treatment of severe sepsis and septic shock innate immune mechanisms triggering lung injury effects of nitric oxide in septic shock signal transduction pathways in acute lung injury: nf-k b and ap-1 sepsis remains one of the most pressing clinical challenges for the pediatric intensivist. it is apparent that while a great deal is now understood about the biological and molecular mechanisms involved in sepsis, this knowledge has not yet had a dramatic impact on improving outcome. at present, therapeutic modalities for sepsis remain largely supportive and founded on the fundamental physiologic principle of providing adequate oxygen delivery. with this approach, mortality in pediatric sepsis improved modestly over the past decades. however, the fact that over 4,000 children per year continue to die in association with severe sepsis argues that further advances be made. realization of the goal of improving survival requires investigators committed to achieving further mechanistic insights into the physiologic, molecular, and genetic biology of sepsis, in concert with large pediatric-specifi c interventional trials. a 5 year old female is admitted to the pediatric intensive care unit with septic shock. she is well oxygenated on a 40% oxygen face mask. she has already received 60 ml/kg of 0.9% normal saline and has been started on a dopamine infusion at a rate of 5 mcg/kg/min. in monitoring her response to these interventions, which of the following should not be used as a therapeutic endpoint to monitor her progress? a. capillary refi ll time b. echocardiographically measured ejection fraction c. mental status d. serum bicarbonate level e. urine output ³ 1 ml/kg/h 10. a 12 year old male with acute lymphocytic leukemia is admitted to the pediatric intensive care unit with vancomycin resistant enterococcus bacteremia. his vital signs reveal a temperature of 39.6°c, a heart rate of 145 bpm, a respiratory rate of 20 breaths/min, and a blood pressure of 108/35 mm hg. he is lethargic, but arousable. his pulses are bounding and his capillary refi ll is brisk. an arterial blood gas reveals a ph 7.31, a paco 2 33 mm hg, a pao 2 65 mm hg, an oxygen saturation of 93%, and a base defi cit of (−10). the oxygen saturation of venous blood sampled from the superior vena cava is 88%. a. the young man is bacteremic, but not in shock as evidenced by his bounding pulses, brisk refi ll, and normal systolic blood pressure. b. the young man is in shock with inadequate oxygen extraction at the tissue level evidenced by the elevated superior vena cava saturation. c. the young man has a primary metabolic acidosis, but has a normal oxygen extraction as oxygen saturation in the superior vena cava is normally higher than elsewhere in the body. d. the young man has a primary metabolic acidosis, but is not in shock, evidenced by his high superior vena cava saturation. e. the young man has a primary respiratory alkalosis that would benefi t from supplemental oxygen therapy.11. there is suffi cient data to justify the use of which of the following adjuvant therapies in pediatric sepsis? a. the administration of activated protein c to a child in septic shock without thrombocytopenia or coagulopathy b. the administration of anti-tnf-a monoclonal antibodies to a septic patient to decrease the proinfl ammatory response c. the administration of stress dose hydrocortisone to a septic patient whose serum cortisol level fails to increase sufficiently in response to a corticotropin stimulation test d. the early initiation of high volume, continuous veno-venous hemofi ltration to remove proinfl ammatory cytokines e. the transfusion of packed red blood cells to maintain a hemoglobin ³ 12 g/dl in order to provide supranormal oxygen delivery 12. it has become clear that dysregulation of the coagulation cascade occurs in sepsis as refl ected by activation of procoagulant pathways, consumption of clotting factors, alterations in fi brinolysis, and reduced anticoagulant activity. which of the following components of coagulation is increased during sepsis? a. antithrombin iii (at iii) b. plasminogen activator inhibitor type 1 (pai-1) c. protein c d. protein s e. tissue factor pathway inhibitor (tfpi) key: cord-017470-sjk7a34u authors: arlati, sergio title: pathophysiology of acute illness and injury date: 2018-06-14 journal: operative techniques and recent advances in acute care and emergency surgery doi: 10.1007/978-3-319-95114-0_2 sha: doc_id: 17470 cord_uid: sjk7a34u the pathophysiology of acute illness and injury recognizes three main effectors: infection, trauma, and ischemia-reperfusion injury. each of them can act by itself or in combination with the other two in developing a systemic inflammatory reaction syndrome (sirs) that is a generalized reaction to the morbid event. the time course of sirs is variable and influenced by the number and severity of subsequent insults (e.g., reparative surgery, acquired hospital infections). it occurs simultaneously with a complex of counter-regulatory mechanisms (compensatory anti-inflammatory response syndrome, cars) that limit the aggressive effects of sirs. in adjunct, a progressive dysfunction of the acquired (lymphocytes) immune system develops with increased risk for immunoparalysis and associated infectious complications. both humoral and cellular effectors participate to the development of sirs and cars. the most important humoral mediators are pro-inflammatory (il-1β, il-6, il-8, il-12) and anti-inflammatory (il-4, il-10) cytokines and chemokines, complement, leukotrienes, and paf. effector cells include neutrophils, monocytes, macrophages, lymphocytes, and endothelial cells. the endothelium is a key factor for production of remote organ damage as it exerts potent chemo-attracting effects on inflammatory cells, allows for leukocyte trafficking into tissues and organs, and promotes further inflammation by cytokines release. moreover, the loss of vasoregulatory properties and the increased permeability contribute to the development of hypotension and tissue edema. finally, the disseminated activation of the coagulation cascade causes the widespread deposition of microthrombi with resulting maldistribution of capillary blood flow and ultimately hypoxic cellular damage. this mechanism together with increased vascular permeability and vasodilation is responsible for the development of the multiple organ dysfunction syndrome (mods). • the inflammatory reaction is a highly adaptive, integrated response with a global protective effect against microbial pathogens or tissue damage. it provides for the elimination of pathogens, removal of cellular debris, and promotes tissue repair and healing. • multiple trauma or severe infection causes a widespread inflammatory reaction that causes diffuse endothelial activation (chemotaxis), damage (permeability edema), and vasodilation (hypotension-shock). • inflammation and coagulation are strictly coupled. as a general rule hyperinflammation means hypercoagulability. coagulation has beneficial effects when inflammation is localized, but it becomes catastrophic when disseminated intravascular coagulation ensues. • humoral inflammatory mediators include cytokines, complement, thrombin, acute phase proteins, kinins, and paf. endothelial cells, monocytes, antigenpresenting cells (macrophages and dendritic cells), and neutrophils are the main effector cells. • neutrophils are responsible for tissue damage. their widespread activation accounts for the noxious effects to innocent tissues with multiple organ damage. • a counter-inflammatory response mounts immediately after the hyperinflammatory reaction. such response is proportionate to inflammation along the whole course of disease. • an immediate decrease of the acquired (lymphocytic) immune response occurs in parallel with inflammation leading to dysfunction and progressive immunoparalysis. • apoptosis of the cells of the immune system is the main responsible of immunoparalysis. this exposes to increased risks for opportunistic infections and sepsis/septic shock. (compensatory anti-inflammatory response syndrome, cars) [1] . although cars opposes to the systemic inflammatory response syndrome (sirs) [2] , this is a double-edged sword because the risk of septic complications is increased. if unresolved, sirs and cars become the underlying players of a catabolic syndrome that leads to mods and ultimately death [3] . in the past sirs was viewed as an exaggerated response to inflammatory stimuli, but the latest experimental and observational data indicate that it is a rather predictable side effect of especially severe morbid events. in practice, sirs and cars result from the growing sophistication of icu care that keeps patients alive during the early (acute) phase of traumatic and septic diseases. the protracted survival of formerly rapid lethal conditions makes now appreciable their natural evolution. recent acquisitions also suggest that sirs and cars develop simultaneously rather than in sequence as previously believed. as a result a mixed antagonist response syndrome (mars) was coined to reflect the balance between sirs and cars [4] (fig. 2.1 ). however, the phenotypic predominance of the hyperinflammatory state is the rule in early sepsis, hypoxia, or trauma as influenced by antigenic load, microbial virulence, host genetic factors, age, nutritional status, and comorbidities [5] . in the past cars was believed to develop after sirs exhaustion by repeated noxious stimuli (second hit theory) [5, 6] . according to this theory, recurrent morbid insults augment the inflammatory response by repeated stimulation of the inflammatory cascade. in this sense, sirs would no longer depend by the initial insult but rather by the intensity and frequency of subsequent hits. however, the continuous challenge of the inflammatory system mounts an antiinflammatory response that ultimately becomes predominant. so cars does not develop simultaneously with sirs, but only a minimal overlap would exist between the two phe-nomena. this pathophysiological view is derived from the observation that cars prevails in the later stages of disease when the increased susceptibility to infections is associated with a weakened pro-inflammatory response (sirs exhaustion). this concept translated into the linear transition from acute (early) sirs to chronic (late) cars with possible alternating recurrence of the two phases (mars), (fig. 2.2) . however, this view is no longer accepted as the historical belief of the "cytokine storm" after a catastrophic acute illness (e.g., meningococcal sepsis) giving the spectacular inflammatory reaction is not the rule [7] . instead, the most common picture is by far a patient over 65 years of age with sepsis or recovering from multiple trauma/surgery and evidence of immunosuppression without the typical exaggerated acute phase inflammatory response [7] . in the past "cytokine storm" was synonymous of sirs that is hyperinflammation defined by excessive release of classical proinflammatory cytokines including il1, il6, il8, and tnfα. however, this concept is too narrow as it was quickly noted that "cytokine storm" is not the typical occurrence in late (chronic) sepsis or even in acutely septic patients with a weakened immune system [8, 9] . similarly, it seems incorrect to define cars on the basis of elevated release of antiinflammatory cytokines in the blood. the current concept is rather that the magnitude of cytokine release depends on the premorbid immune-inflammatory status of the patient [10] . otherwise stated the healthier the patient, the stronger will be the release of cytokines after stimulus. as a corollary, the more protracted is the disease, the more faded will be the inflammatory response over time (e.g., recurrent sepsis in postsurgical or trauma patient). however, an acute inflammatory response although typical of the acute phase may occur at any time of the disease profile if the host is sufficiently immunologically responsive. this view holds for both the pro-inflammatory and anti-inflammatory cytokines, so it is incorrect to define the patient's inflammatory status on the basis of his/her cytokine profile [10] . therefore, the mixed cytokine response pattern better represents the patient's inflammatory status leading to the paradigm "sepsis: always in mars" [11] . thus a hyperinflammatory status at the onset of sepsis or multiple trauma reflects the ability of the host to release a great amount of proinflammatory and anti-inflammatory mediators. such ability is destined to fade over time with progression to a late (chronic) inflammatory status. in recent years, the immunocompetent cells have emerged as a new relevant player for the appearance of immunosuppression or immunoparalysis that often characterizes the host's response during the more chronic disease stages. at present, the process of immunosuppression (decreased t-cell proliferation and production of il2, decreased monocyte and macrophage function) is believed to occur in parallel with the hyperinflammatory status of early sepsis of traumatic or surgical origin [12, 13] . animal studies indicate that in non-survivors, the immune cell suppression progresses indefinitely up to anergy from the very beginning to the more chronic stages in a timeindependent manner [10] . as a result, the phenotype of immunosuppression does not often correspond to the cytokine pattern of peripheral blood. prior to early deaths, cellular immunosuppression develops rapidly together with high pro-inflammatory and antiinflammatory cytokine release (mars-like) [14] . conversely, the chronic state is preceded by a progressive (subacute or chronic) impairment of immune cells function with robust pre-lethal signs of anergy and a deteriorating but mars-like cytokine profile (simultaneous presence of both pro-inflammatory and anti-inflammatory mediators in the blood) [10] . the spread of the inflammatory process starting from a single organ or tissue is by far the most frequent event in the pathophysiology of acute diseases. the inflammatory response is a highly coordinated process, which has evolved to limit the spread of noxious stimuli, eliminates pathogens and necrotic cellular debris, and promotes the healing of damaged tissues. it is subjected to multiple activations and control mechanisms and whose efficiency is largely dependent on genetic predisposition, age, and neurovegetative and hormonal milieu derived from the stress response. finally, inflammation and immunity are tightly related in a complex network of multiple interconnections and reverberating loops. however, extremely intense or repeated stimulations may disturb their tuned response so that the inflammatory mediators spill over the anatomical barriers and multiple organs dysfunction syndrome ensues. cardinal inflammation phenomena are local vasodilation, increased endothelial permeability, and chemotactic cells activation from the natural (granulocytes and monocytes) and acquired immune system (lymphocytes). the vascular mechanisms that lead to the four cardinal signs of inflammation are resumed in fig. 2. 3. neutrophils and monocytes are activated to infiltrate the site of infection with subsequent phagocytosis and lysis of bacterial products or cellular debris. in the meanwhile, the activated coagulation system seals the site of inflammation and provides a meshwork of fibrin that helps in the reparation process. therefore, increased membrane permeability, tnfα, il-1β, il-6 il-8, il-12, ifγ cytokine profile il-4, il-10,tgf-β sirs cars mars fig. 2. 2 temporal profile of pro-inflammatory and anti-inflammatory cytokines according to the hypothesis of sequential development of sirs and cars. mars is a transitional state in-between them capillary vasodilation, chemotaxis, and phagocytosis are defensive mechanisms that act in concert to ensure the maximum of protection against any threat or danger. a multiplicity of cell types (e.g., endothelium, monocytes, platelets) as well as humoral factors (complement, leukotrienes, kinins) acts following a synergistic and often redundant logic to activate, propagate, and maintain the inflammation so that the host defense is guaranteed. nevertheless, the uncontrolled diffusion of inflammatory mediators causes hypotension and tissue edema by generalized vasodilation and increased endothelial permeability. furthermore, the diffuse deposition of microthrombi by disseminated intravascular coagulation worsens the oxygen supply to tissues this, in turn, contributing to ischemic cells damage, further inflammation (ischemia/reperfusion injury), and mods. the local action of the inflammatory system is similar to a well-refined military strategy. after an enemy attack (e.g., trauma) that engages the local garrison (resident macrophages and glial cells), the combat zone is rapidly enclosed and sealed by reinforcement troops (chemotactic activation of pmns and intravascular coagulation). thereafter, the soldiers get into the battlefield (increased membrane permeability and leukocytes migration) and shoot at the enemy destroying him (phagocytosis, proteases, and toxic oxygen products). however, after a massive attack, the hurried and often disorganized mobilization of the reserve troops makes difficult or even impossible to implement an effective defense. the recruitment of soldiers is often chaotic and uncoordinated (widespread chemotactic activation), the effective concentration of troops is impossible (generalized increase of endothelial permeability), and military patrols often shoot at innocent people (organ damage). thus, pneumococcal pneumonia can transform into severe sepsis or septic shock if a generalized inflammatory reaction develops by either cellular (neutrophils, monocytes, macrophages, endothelium) or humoral effectors (complement, contact phase proteins, leukotrienes, cytokines, chemokines) resulting into increased capillary permeability (tissue edema), vasodilation (hypotension), and coagulation activation (ischemic organ damage). generally, the pathophysiological mechanisms that lead to a systemic inflammatory reaction are infections, trauma, and ischemiareperfusion damage. each of them can act by itself or in combination with the other two. for example, multiple trauma causes the activation of immune-inflammatory mechanisms by itself (tissue necrosis), or as a consequence of ischemia-reperfusion damage (e.g., gut ischemia by posttraumatic mesenteric hematoma or post-ischemic muscular tissue reperfusion after hemorrhagic shock). apart from the abovementioned mechanisms, the uncontrolled activation of the immune system (autoimmune diseases), massive cytokines production (metastatic cancer, leukemia, or lymphoma), and the unrestrained activation of serum proteases (acute pancreatitis) are less frequent causes of generalized inflammatory activation. the widespread activation of the inflammatory system (systemic inflammatory reaction syndrome, sirs) originates from the site of trauma, infection, or hypoxic cell damage (ischemia/reperfusion). infection, traumatic or hypoxic injury, causes the release of a heterogeneous pattern of endogenous and exogenous molecules that trigger the innate immune system as chemoattractants and activators of antigen-presenting cells. infection from bacterial, viral, and fungal agents releases signaling substances that are recognized by the innate immune system due to their characteristic molecular pattern (pathogen-associated molecular patterns, pamps) [15] . conversely, traumatic or hypoxic cell injury releases the so-called damage-associated molecular patterns (damps) [16, 17] which represent the correlate of pamp for danger signals of endogenous origin. pamps and damps are grouped into the larger family of "alarmins" in assignment to the term danger signals [15] . otherwise stated pamps and damps constitute a physiologic signaling system that alerts the body to the presence of foreign invaders or noxious stimuli. "alarmins" activate specific receptors of the superfamily of the toll-like receptors (tlrs) [18, 19] , expressed on endothelial and innate immune cells like macrophages, dendritic cells (antigenpresenting cells, apcs), monocytes, and pmns [20] . apcs act as an intermediate between innate and acquired immune system. their main function is to process antigen material and to present it to effector t cells of the immune system. tlr receptors recognize a variety of peptides that are important signaling molecules for activation and production of a multiplicity of inflammatory mediators. in addition, damps are potent activators of the complement system [21] [22] [23] whose anaphylatoxins attract monocytes and pmns on the endothelium. the high mobility group box protein (hmgb) is one of the most studied "alarmin." hmbg is a protein molecule derived from the nucleus of damaged cells [15, 24] . it is released by activated myeloid cells (e.g., neutrophils) [20] , macrophages, dendritic cells [25] [26] [27] , or necrotic cells [26] and acts as a chemoattractant for monocytes, macrophages, dendritic cells, neutrophils, and ϒδ cells [3] . it also participates to the secretion of proinflammatory cytokines [28] and mediates the monocyteendothelial interaction by increasing vascular leakage [24, 29] . fragments of dna and histones are other well-known potent "alarmins." they originate from damaged tissues and microbial digestion by resident tissue macrophages or activated neutrophils. peptides and mitochondrial dna are vigorous alert molecules probably because of their vestigial origin from intracellular bacteria [3] . in addition to "alarmins," the release of phospholipids by damaged cell membranes (cellular hypoxia and trauma) or exogenous lipopolysaccharides (e.g., lps-endotoxin) and polymers (lipoteichoic acid and peptidoglycans) alert the innate immune system by activating the complement and the contact phase proteins system (fxii, kallikrein, and kininogen). phospholipids activate the phospholipases a 2 and c [30] with the production of arachidonic acid metabolites as leukotriene b 4 , prostaglandin e 2 , and thromboxane a 2 [31] . the activation pathway of phospholipase a 2 and c is detailed in fig. 2. 4. in addition, mast cells release histamine and bradykinin with resulting vasodilation and increased capillary permeability and edema [32] . just 20-30 min after trauma or microbial invasion, the innate immune cells become activated, fig. 2 .5. the first line of the innate immune defense is represented by pmns, mostly neutrophils, and monocytes. pmns are chemoattracted by locally produced cytokines (e.g., tnf-α), leukotrienes, platelet-activating factor (paf), and complement fragments (c5a) [33] [34] [35] [36] . these inflammatory mediators also activate pmns to express adhesion surface molecules for appropriate ligands on the activated endothelium [37] . this receptor-ligand interaction allows for adhesion of leukocytes to the capillary wall. thereafter pmns migrate through the endothelial barrier into the tissues by opportune signaling receptors on the inner surface of the endothelium. in the meanwhile, resident macrophages secrete cytokines (tnfα and il-1) and chemokines (il-8) which help the immune and inflammatory cells in selfregulating and crosstalking each other. cytokines and chemokines are pleiotropic molecules with a great variety of effects that act in a paracrine and autocrine fashion. among the most important cytokines secreted in the hyper-acute phase, tnf-α and il-1β act within 1-2 h after trauma or sepsis, while il-6, il-10, and the chemokine il-8 are subacute mediators [38] . among the most relevant properties of cytokines, there is the ability to activate monocytes (innate immune system) and lymphocytes (acquired immune system) [39] . under the appropriate cytokine pattern, the circulating monocytes differentiate into resident macrophages, while lymphocytes mature into different cell lines having immune-stimulatory (th-1), immune-depressant (th-2), and immune-modulatory (treg) action [40] . pro-inflammatory cytokines trigger both the recruitment and the phagocytic activity of leukocytes [41] . they also stimulate the release of proteases and increase the production of free oxygen radicals from activated neutrophils [41] . 1. one of the best characterized cytokines is tnf which can produce sirs when injected experimentally in high doses [39] . its actions include the stimulation of many cell types and survival or apoptosis [42] , cytokine secretion (il-6, il-8, ifϒ, il-10), activation of the arachidonic acid pathways (thromboxane a2 and prostaglandin e2) and nitric oxide, induction of fever and production of selectins, and paf (see below). tnf secretion is stimulated by many physiological stresses (hemorrhage, hypoxemia) as well as ischemia-reperfusion, endotoxin, and complement system. macrophages and monocytes are the main tnf producers, although activated t cells are also implicated. tumor necrosis factor acts via its receptors tnfr1 and tnfr2 with effects that depend on the specific receptor binding and environmental factors influence. binding of tnf-α to tnfr1 leads to transcription of pro-inflammatory genes via the nf-kb transcription factor. however, tnfr1 possesses a death domain at the cytoplasmic tail so that its stimulation also induces apoptosis via the caspase enzymes cascade (see below). although these two effects seem contradictory, they might represent an adaptive mechanism that has evolved to protect cells against excessive stimulation (see below: kidney dysfunction). in contrast to the almost ubiquitous tnf1r, tnfr2 is expressed on immune and endothelial cells only and becomes activated by membrane-bound tnf-α [42, 43] . binding to tnfr2 leads to activation and proliferation of neutrophils and many immune cells as nk cells, b cells, and peripheral t cells [44] . 2. interleukin-1-beta is the other well-studied cytokine that acts synergistically with tnf-α. its actions include fever induction, cytokine and chemokine secretion [45] , t cell and macrophage stimulation and various inflammatory mediators (including adhesion molecules), acute phase proteins, and no synthesis. it also induces the release of neutrophils from the bone marrow and stimulates the expression of proteases and metalloproteases by tissues. il-1β is secreted by macrophages, monocytes, and endothelial cells. stimuli for il-1β include tnf, complement, endotoxin, hemorrhage, and ischemia. the effects of il-1β are mediated by the type il-1 receptor (il-1ri) which belongs to the il-1 receptor superfamily. besides il-1β, another molecule that binds il-1ri is the so-called soluble il-1 receptor antagonist (il1-ra) suggesting a controlling role in il-1-mediated immune response. 3. the third important cytokine in the early inflammatory response is il-6. its secretion is induced by tnf and il-1 [46] . although it does not induce sirs when injected into experimental animals, il-6 has vasodilatory properties as it induces the production of nitric oxide by the endothelial cells. il-6 is a subacute cytokine with marked regulatory properties on immune cell growth and differentiation. it inhibits apoptosis of neutrophils and stimulates the hepatic acute phase protein synthesis but also exhibits an anti-inflammatory action [47] by inducing the release of soluble tnfr and il-1ra [48] . it seems, therefore, to play a dual role in the inflammatory response by acting both as a pro-inflammatory and anti-inflammatory mediator [49, 50] . clinical studies demonstrated increased plasma levels in non-survivor septic patients so that il-6 has been proposed as a prognostic marker [51] [52] [53] . 4. the chemokine il-8 is the most powerful chemoattractant for pmns and monocytes [54] . chemokines control the traffic of the immune cells, which are the main effectors of the immune system. their binding to g-protein-coupled receptors leads to dose-dependent effects including chemotaxis and activation of immune cells. after stimulation by il-1, tnf-α, complement, microbial products (e.g., lps), hypoxia, and reperfusion, il-8 increases neutrophils degranulation, adherence, and chemotaxis. interestingly, chemokines act as chemoattractants in lower doses and potent activators of immune cells function in higher doses. recently a group of so-called silent chemokine receptors has been described suggesting a role as decoy or scavenger receptors. the duffy antigen receptors for chemokines (darc), first described as blood group antigen, have been shown to bind il-8 and other chemokines thus suggesting a regulatory action against excessive leukocyte activation in the systemic circulation [39] . the cellular source and the main effects of pro-inflammatory cytokines are resumed in table 2 .1. apart from cytokines, other important mediators of the hyper-acute phase are the arachidonic acid metabolites (leukotrienes, prostaglandins, and thromboxane). they are responsible for the activation of pmns and endothelium and platelet aggregation. these substances are generated by activation of the phospholipases a2 and c (pla 2 and plc) which follows the entry of ca 2+ ions into damaged cells. pla 2 also induces the release of paf from the endothelium. paf is an important inflammatory mediator with marked effects on macrophages, endothelial cells, and platelets aggregation. the inflammatory response is promptly propagated and amplified by important circulating mediators as the complement system, the contact phase proteins (kallikrein-kinins), and the coagulation cascade. 1. the complement cascade is a series of enzymatic cleavage reactions that are activated via the classical route by antigen-antibody complexes (igm and igg) or alternatively by bacterial degradation products (e.g., lps, endotoxin) [32, 55, 56] . the latter pathway does not require prior immunization and thus represents an immediate defense mechanism against microbes. conversely, the "natural antibodies" activate the complement by forming antigen-antibody complexes with different molecular species from exogenous (microbial) or endogenous (necrotic cells) origin [57] [58] [59] . "natural antibodies" are physiologically circulating antibodies that arise in serum independently from prior host immunization [60] . they are mainly produced by a subpopulation of cd5 + b cells and are poly-reactive with different antigens as peptides, phospholipids, and polysaccharides [60] [61] [62] . in musculoskeletal trauma and ischemia-reperfusion injury, the intense antigenic stimulation leads to binding of natural igm antibodies and development of post-traumatic "innate autoimmunity" [63] . complement activation leads to the formation of opsonins, anaphylatoxins, and the membrane attack complex, a multiprotein complex responsible for the increased capillary permeability [32] . anaphylatoxins and opsonins are involved in important inflammatory processes such as opsonization, chemotaxis, and neutrophils degranulation. complement also promotes the synthesis of acute phase proteins by the liver and stimulates the degranulation of mast cells with histamine release (vasodilation) [32, 64] . in practice, the activated complement is involved in all the most relevant inflammatory processes as immune cells attraction to the site of trauma or infection, microbial phagocytosis and lysis, antigens opsonization, platelets activation, and histamine release. the complement cascade is, therefore, a fundamental component of the innate host defense as it can be rapidly activated in a non-specific manner after injury or infection. it is phylogenetically ancient due to its basic role in the host response to bacterial pathogens. however, the systemic activation of the complement leads to severely deleterious effects such as shock and tissue edema due to massive vasodilation and increased membrane permeability. 2. coagulation factor xii and kallikrein, together with kininogen, represent the contact phase system. these proteins activate each other by forming an integrated, highly amplified system for activation of the coagulation cascade. kallikrein also stimulates the formation of the vasodilator bradykinin from kininogen. kinins are potent vasodilators and vigorous effectors of increased vascular permeability [65] . a close interconnection exists between hemostasis and the inflammatory system. for example, the activated platelets aggregate with circulating leukocytes to stimulate the immune cells system with production of endothelial damage [66] . platelets also release pro-inflammatory mediators that excite the immune system [67] thus creating a vicious circle that produces more and more sirs. the activated coagulation cascade after trauma or infection is primarily intended to stop the loss of blood and to seal the lesion. thereafter, phagocytic and immune cells eliminate necrotic cells and invading microbes. however, the uncontrolled activation of the coagulation cascade causes the widespread deposition of microthrombi, this in turn causing hypoxic cellular damage and increased risk for hemorrhagic complications due to consumption of platelets and coagulation factors [68] [69] [70] [71] . disseminated intravascular coagulation (dic) is a pathophysiological mechanism that frequently occurs after multiple trauma [68, 70, 71] and severe infections [68, 70] . dic is responsible for diffuse microthrombi deposition, capillary blockade with heterogeneous microcirculatory perfusion, tissue ischemia, and production of hypoxic cellular damage [68, 72] . 3. acute phase proteins are synthesized in the liver by induction of the locally (kupffer cells) produced cytokines (tnf-α, il1-β, and il6). this process is amplified by circulating cytokines that spillover from injured or infected tissue (overflow theory). acute phase proteins include c-reactive protein (crp), α-1-antitrypsin, α-2macroglobulin, ceruloplasmin, lipopolysaccharide-binding protein (lpb), fibrinogen, and prothrombin [73] . these proteins have several inflammatory and anti-inflammatory properties. for example, crp increases the surface expression of tissue factor (tf) [74] , a procoagulant membranebound 4.5 kd protein on pmns, monocytes, and tissue macrophages (see below). conversely, α-1-antitrypsin neutralizes leukocyte proteases and free oxygen radicals, while α-2-macroglobulin and ceruloplasmin are inhibitory cofactors for the synthesis of cytokines [74] . finally, high concentrations of lpb suppress lps activity [75] . at this step of the inflammatory process, the injured or infected tissue is invaded by activated pmns, mainly neutrophils that release proteases (elastase), exert their phagocytic and digestive properties (phagolysosomes), and produce free oxygen radicals (e.g., hydrogen peroxide) [76] and reactive no species (e.g., peroxynitrite) [77] . both neutrophils and macrophages kill bacteria. the first step involves phagocytosis of opsonized microbes. opsonization means that bacteria are covered with host proteins (e.g., antibodies and complement fragments) and appropriately recognized by cell surface receptors such as tlr and receptors for the fc portion of the immunoglobulins. the phagocytosed bacteria are typically inserted into a vacuole, the phagosome, which fuses with primary (azurophilic) or secondary granules to form phagolysosomes. azurophilic granules contain antimicrobial proteases such as elastase and bactericidal/permeability increasing proteins, while secondary granules contain antiseptic peptides as lysozyme, lactoferrin, and metalloproteases. fusion of granules with phagosomes creates a hostile environment that kills the pathogen ( fig. 2.6 ). at the same time, the resident macrophages and circulating monocytes begin to phagocyte cellular and microbial opsonized bacteria bind to complement receptors (crs) and fc receptors (fcrs) on the surface of phagocytic cells. thereafter the bacterium is taken into the cytoplasm and included into a phagosome. the fusion of phagosome with specific and azurophilic granules gives a phagolysosome that destroys the bacterium by toxic and enzymatic digestion debris. these cells produce cytokines and express membrane surface antigens that originate from the digestion process (see fig. 2 such molecules activate the endothelium to express adhesion molecules for further neutrophils chemoattraction. in the meanwhile, macrophages interact with t helper cells (cd4 + ) that secrete interferon ϒ (ifϒ) and other cytokines for phagocytic cells activation. finally, their interaction with b lymphocytes produces antimicrobial antibodies. this is a highly coordinated process, able to eliminate foreign bodies or cellular debris so ensuring the most effective protection against any insult of whatever origin. besides the plethora of pro-inflammatory mediators, many anti-inflammatory substances play a counter-regulatory role in the development of cars. il-4, il-10, il-13, and transforming-growth-factor β (tgf-β) are among the most important anti-inflammatory and immunosuppressant mediators. the main actions of anti-inflammatory cytokines are shown in table 2 .2. besides these anti-inflammatory cytokines, several immune effector cells develop their actions immediately after the onset of the acute inflammatory response. the nature of their response is largely determined by the expressed membrane antigenic pattern as it determines their subsequent cellular interactions and biochemical reactions. from the native, multipotent cd4 + t helper 0 cell (th0), two main cell lines differentiate depending on cytokine environment, either into th1 helper cell by il-12 or th2 helper cell by il-4 [78] . thereafter il-12 is produced, and opportune co-stimulatory molecules (cd80/86) are expressed together with mhcii antigens that bind to corresponding t-cell ligands. this binding results into t-cell activation and proliferation. the production of interferon γ stimulates the macrophages to kill intracellular bacteria [79] . th1 helper cells are therefore involved in the classical inflammatory response. overactivation of th1-type cells produces type 4 delayed hypersensitivity. 2. conversely, th2-type lymphocytes show marked antiinflammatory properties by producing anti-inflammatory cytokines as il-4, il-5, il-10, and il-13 [80] . overactivation causes type 1 ige-mediated allergy and hypersensitivity. other important players of the cell-mediated immune response in trauma and sepsis are the regulatory t (tregtype) cells and t17-type cells. 3. treg-type cells formerly known as suppressor t cells are modulator and deactivator of the immune response. they are immunosuppressive and generally downregulate the activation and proliferation of effector t cells (t cells that responds to a stimulus including helper and killer lymphocytes). treg-type cells produce il-10 and tgfβ a potent anti-inflammatory cytokine. elevated levels of circulating treg cells have been observed in the blood of immune-paralyzed infected patients [81, 82] . 4. the th17-type cells are a subset of t helper cells so named because of production of the cytokine il-17. they exert protective effects on the gastrointestinal mucosal barrier function [83] . there is general agreement that th17-type lymphocytes are protective against extracellular bacterial and fungal infections [84, 85] . the decrease of th17 cells population contributes to immunoparalysis [86] . it is widely accepted that the development of a successful immune response requires the balance between th1 and th2 helper cells. in the early stage of sepsis and trauma, a phenotypic th1-type cells profile predominates with ifϒ production which activates the bactericidal action of macrophages and induces b cell production of opsonizing and complement-fixing antibodies. conversely, later (chronic) stages are characterized by a shift from the th1-type profile into a predominantly th2-type response leading to immunoparalysis, reduced antigen recognition and cellular anergy (see below) [87] . other more complex anti-inflammatory actions include the decreased production of monocytederived cytokines by reduced transcriptional factors (nf-kb) for the encoding genes [88] . also, the reduced expression of membrane surface receptor cd14 on monocytes (lps-receptor) blunts their pro-inflammatory activity. also the expression of mhc (major histocompatibility complex) class ii molecules hla-dr (human leukocyte antigen) is downregulated [89] with interference in the elimination of infected cells and maturation of several immune cell lines. these anti-inflammatory effects play an important compensatory role in the development of the counter antiinflammatory response syndrome (cars). ideally cars is the physiological response to the risks of tissue damage by uncontrolled inflammation. the main sirs and cars events are summarized in fig. 2 .8. the highly integrated, redundant, and coordinated immune-inflammatory system makes easy its comparison with a great classical orchestra. as a classical orchestra, there are players (neutrophils), soloists (monocytes and macrophages), and the musical score (humoral mediators). this score may be harmoniously classical (locally coordinated inflammatory response) or dissonantly dodecaphonic (sirs). but who is the conductor? this fundamental role is covered by the endothelium. the acquisition that the endothelium orchestrates a complex, often redundant network of the immune-inflammatory players is quite recent. the endothelium is the interface between coagulation and inflammation in sepsis and trauma. endothelial cells line the vessels in every organ by forming a barrier with organ-specific properties. so the vascular permeability is virtually null in capillaries of the brain and almost complete in the kidney with intermediate levels in the liver, muscle, and lung. the main properties of the endothelium are: 1. lining the vascular system thus separating the blood from the cells and interstitium 2. regulating the vascular tone with a net vasodilatory effect after trauma, hypoxic damage, or sepsis, the injured endothelium becomes prothrombotic and anti-fibrinolytic ( fig. 2.9 ). the endothelium is involved in all the three major pathogenic pathways of septic and traumatic coagulopathy: tissue factor-mediated thrombin generation, dysfunctional anticoagulation, and fibrinolysis impairment [90] . moreover, its activation by inflammatory stimulus leads to platelet and leukocyte adhesion and inhibition of vasodilation. endothelial injury can be documented microscopically by a visible change of cell shape or damage and defects of endothelial lining. indirect evidence of the damaged endothelium is the finding of elevated soluble markers of cell injury notably thrombomodulin, intercellular adhesion molecule, and e-selectin and von willebrand factor [91, 92] . after endothelial damage, inflammatory fluids and cells can shift from the blood into the interstitial space. endothelial injury is sustained over time so that its property loss is long-standing [93, 94] . experimental animal and human studies demonstrated that after injury, the full recovery of endothelial lining occurs within 21 days [93] . the denudation of the vessel wall exposes tissue factor (tf), the principal activator of the extrinsic coagulation pathway with the risk of intravascular coagulation [93, 94] . normally the outer endothelium expresses various membrane-bound molecules with anticoagulants properties among which are cell surface heparin. antithrombin iii (atiii) further inhibits thrombin activity by preventing its binding with receptor (trr). also tissue factor (tf) is not expressed on the endothelium, while its main physiological inhibitor, tissue factor pathway inhibitor (tfpi), is normally released by the intact endothelium. finally the tis-sue plasminogen activator (tpa) catalyzes the zymogen plasminogen into plasmin, while tissue plasminogen activator inhibitor type-1 (tpai-1) is reduced. after hypoxia, injury, or sepsis, the damaged endothelium expresses large amounts of tf, while tfpi release is inhibited. also thrombomodulin receptor is internalized thus reducing apc formation, while decreased atiii synthesis and degradation by serine protease increases the activity of thrombin. finally tpai-1 synthesis and release are increased thus reducing the formation of plasmin like molecules [95] and thrombin-binding protein thrombomodulin (tm). heparin-like molecules are supported by glycosaminoglycans which cover the endothelial surface [90] . they act as cofactors for the antithrombinmediated inhibition of thrombin and activate factor x (stuart-power factor) [95] . tm is the major responsible for thrombin inactivation as when bound to thrombin, forms a potent protein c (pc) activator complex that equips the endothelium with anticoagulant properties. however, the inflammatory or septic stimuli decrease the anticoagulant properties of the endothelial cells by loss or internalization of tm [96] , while the contemporaneous stimulation of the thrombin receptor increases the inflammatory pathways. finally, the action of neutrophil proteases also contributes to reduced tm expression. moreover, the release of tissue plasminogen activator (tpa) is decreased, while its main physiological inhibitor, the plasminogen activator inhibitor-1 (pai-1), increases so that the profibrinolytic properties of the endothelium are diminished. tf is increasingly expressed on both monocyte and macrophage membranes as well as on other cell types, while the function of its main physiological inhibitor, the tissue factor pathway inhibitor (tfpi), is decreased by reduced synthesis of glycosaminoglycans on endothelial surface [97] . in practice, any single procoagulant or profibrinolytic has its physiological inhibitor. thus, tf is inhibited by tfpi [98] , the coagulation system is counteracted by the activated protein c system [99] , and tissue plasminogen activator (tpa) is coupled with tissue plasminogen activator inhibitor (tpai) [100] . the imbalance between these systems results in a net procoagulant state with increased fibrin deposition, microvascular thrombotic occlusion, and risk for tissue ischemia [101] . direct endothelial damage as occurs after trauma or ischemia-reperfusion injury contributes to activation of coagulation by decreased production of endothelial-derived substances as pgi and no. the anti-adhesive properties of such molecules reduce leukocytes and platelets aggregation and counteract the procoagulant effectors. localized coagulation has protective effects in discrete traumatic injuries, but its widespread activation is deleterious to the host with risk for multiple organ dysfunction and death [102, 103] . endothelial activation refers to the increased expression of adhesion molecules on endothelium surface. complementary molecules are also expressed on the outer membrane of neutrophil and monocyte cells [104] . the surface expression, modulation, and activity of these molecules are highly regulated by locally produced cytokines (endothelial and monocyte cells) as the chemokine il8 or paf, il1-β, and tnfα. the first step of the adhesion process consists of a "rolling" of leukocytes on the endothelial surface. this process involves selectins which are molecules expressed on leukocytes (l-selectin), endothelial cells (e-selectin), and platelets (p-selectin). selectins act as receptors that permit a loose binding with the endothelial surface thus allowing for leukocyte rolling in proximity of opportune signaling molecules expressed on the endothelium. the second step involves the linkage of integrins with receptors of the immunoglobulins superfamily expressed on the endothelium surface. integrins are a group of three heterodimer proteins expressed on the outer surface of activated leukocytes and collectively termed the cd11/cd18 complex. adhesion molecules include icams, e-selectins, pecam (platelet endothelial cell adhesion molecules), and vecam (vascular endothelial cell adhesion molecules). the final step consists of migration of activated leukocytes to the borders of endothelial cells and interaction with the endothelial-expressed adhesion molecules icam, pecam, e-selectin, and vecam [105] . adhesion can occur without intervention of adhesion molecules in the lung and liver. it has been suggested that actincontaining stress fibers increase at the leukocyte periphery, this, in turn, causing decreased pmns deformability and sequestration into the pulmonary and hepatic capillary beds [106] . the main effects of endothelial cells activation are summarized in fig. 2. 10. thrombin is the main effector of increased endothelial permeability thus reaffirming the bidirectional interplay between the coagulation and the inflammatory cascade ( fig. 2.11) . thrombin can cleave a set of protease-activated receptors (par1-4) expressed on the endothelium [90, 107] that induces rho-dependent cytoskeletal derangement in endothelial cells [108, 109] . par1 receptor activates rho kinase which inhibits the dephosphorylation of myosin light chains (mlc) [110] . phosphorylated mlc causes the actin-myosin interaction at the cell-to-cell contacts with contraction and rounding of endothelial cells and increased of vascular permeability. this facilitates the passage of protein molecules and leukocytes from the blood into the interstitial space (fig. 2.12) . furthermore, the formation of gaps in the endothelial barrier exposes the proteins of the external coagulation pathway to the abundant tf amount expressed by the basement membrane and vessel adventitia. clot formation is thus initiated, while collagen fibers from the extracellular matrix prompt vwf to polymerize and platelets to adhere. endothelial dysfunction refers to decreased no release and reduced expression or synthesis of the constitutive no synthase enzyme (enos). abnormality of endothelial relaxation properties lasts for many days after the acute insult (endothe-lial stunning) [111] . the impairment of no and pgi release by endothelial dysfunction have a profound impact upon cellular oxygenation with direct effects on tissues and organs well-being (fig. 2.10 ). according to the concept of intrinsic metabolic regulation, oxygen supply and demand are constantly matched. local vascular relaxation is therefore in dynamic equilibrium with the nervous-mediated vasoconstriction so that capillary blood flow is finely tuned to peripheral requirements. the endothelium plays a crucial role as a sensor of the local blood flow because of functional and structural coupling with the smooth muscle of arterioles and arteries. sensing is realized by depolarization/hyperpolarization of the endothelial cells, while communication involves electronic spread via endothelium/muscle cell-cell gap junctions [112] . therefore, the hyperpolarization of the capillary endo-thelium induces the upstream vasodilation of the feeding arterioles and arteries [113, 114] . when inflammation impairs this finely tuned cell-cell communication, a "malignant intravascular inflammation" ensues [115] . the abnormal vascular reactivity, increased fibrin deposition, and cells adhesion may well account for heterogeneity of intra-organ perfusion and impaired oxygen delivery. the relationship between organ perfusion heterogeneity, impaired organ oxygen supply, and development of metabolic acidosis has been demonstrated in an animal model of endotoxic shock [116] . in an in vivo model of septic shock, a 36% reduction of perfused capillary density with increased perfusion heterogeneity and mean intercapillaries distance contributed to functional shunting [117] with impaired oxygen extraction after endotoxin challenge or fecal peritonitis [118, 119] . the other fundamental player of the systemic immuneinflammatory activation after trauma and ischemiareperfusion or sepsis is the monocyte/macrophage cell. monocytes are the circulating form of resident macrophages and dendritic cells. resident macrophages can be found in the liver (kupffer cells), lung (alveolar macrophages), lymphoid tissue (spleen and lymph nodes), and kidney (intra-glomerular mesangial macrophages). all these cells are very active in the synthesis of cytokines and removal of particulate antigens. in addition, they play a central role in antigen presentation to the innate th1-type cells. apcs are fundamental for the eradication of pathogens, foreign antigens, or autologous cellular debris. their key functions include the recognition, uptake, and killing of invading organisms. tissue macrophages are initially activated after tissue injury by ischemic necrosis of mesenchymal cells in an il1-α-dependent process. after the necrotic insult il1-α is released from the dying cells with subsequent binding to the il-1 macrophage surface receptors. il1-α activates the assembly of inflammasome within the macrophage cytosol. inflammasomes are multiprotein cytoplasmic complex that cooperate with the toll-like receptors to respond to various insults by processing cytokines and promoting the inflammatory response. in the presence of pathogenic stimuli (e.g., alarmins), the inflammasome opens up so triggering the conversion of immature pro-inflammatory cytokines (e.g., il1-β) into mature forms and activating proapoptotic enzymes (e.g., caspases). thereafter, the extracellular il1-β secretion and upregulation of il1-β-induced chemokines, together with the increased expression of adhesion molecules, and cytokines secretion on the nearby endothelium [120, 121] allow for recruitment of circulating neutrophils and monocyte. no synthesis is activated with resulting vasodilation, opening of endothelial gaps, and loosening of endothelial barrier properties [122] (fig. 2.8 ). monocytes and macrophages are also pivotal cells for the interaction between myeloid and endothelial cells. after the expression of chemoattractant molecules (il8, monocyte chemoattractant protein-1 mcp-1, or macrophage inhibitory protein-1 mip-1), the endothelium attracts monocytes to the inflammatory focus where they migrate into the tissue and become macrophages. finally, the activated monocytes express large amounts of tf to propagate the procoagulant activity. after major uncomplicated trauma, the ability of apcs to express mhcii surface antigens is reduced [122] and returns to normal within a week [123] . after severe injuries, the apcs show a continuously decreased function. this deactivation leads to reduced expression of mhcii surface antigens and decreased ability to secrete cytokines which result into the increased susceptibility to infections [124] . many research findings have focused on the existence of premature programmed cell death (apoptosis) after injury, ischemia-reperfusion, and sepsis [125, 126] . although apoptosis is an adaptive mechanism in several tissue and organs, namely, the lymphoid tissue and gut, its role seems deleterious in acute inflammatory states. several studies suggest that the programmed cell death contributes to the derangement of cellular homeostasis in parenchymal (lung endothelial cells, kidney tubular cells, and skeletal muscle cells) and lymphoid organs with increased risk for sepsis and development of multiple organ dysfunction [87] . accelerated apoptosis induces lymphocytopenia [127] and decreased monocyte survival as reflected by loss of monocyte cd14 expression (co-receptor for the detection of bacterial lipopolysaccharide-lps) [82] . autoactivation of cytosolic and mitochondrial caspases are the two major pathways involved in apoptosis [128] (fig. 2.13) . briefly, the extrinsic pathways activate caspases via binding to members of the tnf-receptor superfamily, while the mitochondrial induced pathway requires the emission of cytochrome c, otherwise essential for mitochondrial survival, into the cytosol with subsequent caspases activation [129] . caspases are a family of proteolytic enzymes synthesized as inactive zymogens and activated by appropriate stimuli to express a death effector domain [130] . apoptosis is a series of coordinated processes [131] that lead to dna fragmentation, chromatin condensation, and blebs formation in the plasma membranes. most important this controlled form of cellular degradation carries out with minimal effects on surrounding tissues. therefore, apoptosis in a crucial physiological process during fetal development and throughout life as it maintains the normal development and regulation of cellular proliferation. for example, during organ development apoptosis eliminates those cells that are no longer necessary. it has been estimated that without apoptosis, about 2 tons of bone marrow and lymphoid tissue would accumulate in the body [132] ! conversely accelerated apoptosis of lymphocytes is as detrimental as delayed apoptosis in neutrophils. neutrophil cells are constitutively apoptotic as this ensures a tight conr e c e p to r the two main pathways of apoptosis are extrinsic and intrinsic pathway. each requires specific triggering signals and activates its own initiator caspase which in turn activate the executioner caspase-3. the execution pathway includes cell shrinkage, chromatin condensation, cytoplasmic blebbing, and formation of apoptotic bodies. finally phagocytosis of apoptotic bodies is performed by adjacent parenchymal cells or macrophages trol of the inflammatory response, but upon delayed apoptotic stimuli (cytokines), neutrophils become persistently activated and contributes to extensive organ damage by continuous release of toxic products (protease enzymes and reactive oxygen species) [133] . apoptosis causes a profound depression of immune functions with immunoparalysis and depletion of several immune cells including helper (cd4 + ) and suppressor (cd8 + ) t cells, b lymphocytes, and apcs (antigen presenting cells) [134] . postmortem studies have confirmed immune cells apoptosis in all age groups [134, 135] . it can be speculated that apoptosis of gut-associated lymphoid tissue (galt) [136] may be involved in bacterial translocation (see below). several findings support the current view of immunoparalysis as a dominant feature of patients surviving the early hyperinflammatory phase. although focal regions of inflammation ischemia and necrosis undoubtedly occur and contribute to the development of multiple organ dysfunction, death is the consequence of failure to control the primary infection or the development of new hospital-acquired infections often with opportunistic pathogens. this does not necessarily mean that infection is the main responsible for the patient's death and in fact many postmortem results are inconclusive. however, the severe decrease of innate immune function and the widespread hibernation of nonimmune cell type (cellular hibernation response) [137, 138] make apoptosis a primary mechanism for multiple organ dysfunction and ultimately death. interestingly cancer and sepsis show similar immune defects [139] , and increased survival after improving host immunity in cancer patients is encouraging to the field of sepsis. the most frequent manifestations of immunoparalysis include profound and persistent lymphopenia, loss of delayed type 4 hypersensitivity reaction [140] , reactivation of latent viruses infections (herpes simplex virus and cytomegalovirus) [141, 142] , and infection by low-virulent pathogens (e.g., stenotrophomonas spp., candida spp., acinetobacter spp.) [143, 144] . decreased pro-inflammatory cytokines production including ifϒ, tnf, and il-2, increased il-10 and other anti-inflammatory cytokines, decreased monocyte/macrophage and dendritic cells function with increased expression of apoptotic surface markers, and decreased cell survival are the hallmarks of immunoparalysis. dendritic cells and monocytes play a pivotal role in the development of decreased immune function. their reduced survival (apoptosis susceptibility) and function abnormalities (reduced hla-dr expression and increased production of il-10) induces t helper cell anergy and treg cell proliferation. the reduced capacity of monocytes to release pro-inflammatory cytokines (e.g., tnf, il-1, il-6, il-12), whereas the release of anti-inflammatory mediators (e.g., il-10, il-1ra) is not impaired or even enhanced [145] suggest for cellular reprogramming toward the anti-inflammatory pathway [146] . monocyte dysfunction is known as endotoxin tolerance that is reduced cellular response to an endotoxin challenge. the main consequence of endotoxin tolerance is the increased susceptibility to nosocomial infections and death. decreased immune t-cell function and t-cell exhaustion are the other leading causes of immunoparalysis. 1. the high antigenic load and the elevated pro-and antiinflammatory cytokine profile of the early phase of trauma and sepsis are ideal for the development t-cell exhaustion. phenotypic features of exhausted t cells are a decreased production of pro-inflammatory cytokines, mainly ifϒ and tnf, decreased cellular proliferation and cytotoxic function by excessive expression of pd-1 inducible co-stimulatory molecule and programmed cell death ligand pdl-1, and decreased expression of the il-7 receptor [148] . il-7 is a multipotential cytokine that acts to reverse immunosuppression by multiple mechanisms. it has shown to induce the two-to threefold increase of both naïve (th0-type) and cd4/cd8 cells in cancer [149] and hiv patients [150] so reversing their major pathological abnormality, i.e., profound lymphopenia. other effects include blockade of apoptosis [151] , reversal of t-cell exhaustion [152] , increased ifϒ [152] production leading to macrophage activation, and increased adhesion molecules expression on activated t lymphocytes [153] . the decrease of th1-type, th2-type [86, [154] [155] [156] , and th17-type cells [86, 157] with maintenance of the number of treg cells leads to the relative increase of regulatory functions and downregulation of effector t-cell response [86, 157] . the reduction of th17-type cells contributes to the increased susceptibility to fungal infection due to their important role in protecting against extracellular bacterial and fungal invasion [158] . the relative increase of treg cells is deleterious as being is associated with decreased effector t-cell proliferation and function. their increased resistance to apoptosis with respect to the other t-cell type is probably responsible for their increased proportion [159, 160] . treg cells can also suppress innate immune cells thus inhibiting myeloid-derived cell function [161] . therefore, increased treg cells impair the immune function by acting both on innate and adaptive immunity. neutrophils, nk cells, and ϒδ t cells are other effectors whose function is decreased or impaired during immunoparalysis. 2. briefly circulating neutrophils are markedly increased during the hyperinflammatory phase due to delayed apoptosis and release of immature cellular forms [162, 163] . loss of chemotactic activity is their most frequently encountered dysfunction [164, 165] . a subset of neutrophils with suppressive properties similar to the myeloid-derived suppressor cells (mdscs) contributes to the development of immunoparalysis by production of large amounts of il-10 [166] and interference in proliferation and function of effector t cells [167] . mdscs are a heterogeneous group of myeloid cells that expand during chronic inflammatory states and cancer as a result of altered hematopoiesis. mdscs possess strong immunosuppressive properties of both humoral and cellular type [168] . 3. nk cells are markedly reduced both in peripheral blood and tissues where they are most abundant [169] . impaired ifϒ by nk cells increases the host susceptibility to viral infections and reactivation of latent viruses, notably herpes simplex and cytomegalovirus [141, 142] . 4. ϒδ t cells are a subset of lymphocytes that possess functions common to the innate and adaptive immune systems. they are particularly abundant in the intestinal mucosa with innate-like defense mechanism (ifϒ and il-17 cytokines production). intestinal ϒδ t cells are a first-line defense against infections [170] , and their decreased number exposes to the risk of microbial invasion of the blood and peritoneal cavity. the main effects of chronic inflammation upon innate and acquired immune cells are shown in figs. 2.14 and 2.15. multiple organ dysfunction syndrome (mods) is defined as "the presence of altered organ function in an acutely ill patient such that homeostasis cannot be maintained without intervention" [2] . the mechanisms implicated in the development of mods include: anti-inflammatory cytokines secretion cytotoxic function cytokines secretion. ag-specific antibodies production. cd8 + t-cell b cell t reg cell lymphocytopenia results from massive loss of t helper and t suppressor lymphocytes. t regulatory cells are more resistant to apoptosis so that an immunosuppressive phenotype results. surviving t cells shift from a proinflammatory th-1 cell-type to an anti-inflammatory th-2 cell-type profile 3. immune system dysregulation 4. mitochondrial dysfunction although the above mechanisms can interact in a variety of ways and combinations, immune system dysregulation and mitochondrial dysfunction seem to play a prevalent role. as detailed above, the immune system dysregulation is the imbalance between pro-inflammatory and anti-inflammatory mechanisms. cars is a coordinated anti-inflammatory response to severe inflammatory stimuli that allows for the maintenance of immune system homeostasis. the "purpose" of cars is to limit the noxious effects of sirs while not interfering with pathogen elimination or healing of the injured tissue. however, cars may be dangerous when its effects lead to a condition known as "immunoparalysis" with an increased risk of nosocomial sepsis and septic shock. mitochondrial dysfunction with its hypoxic cellular implications is also important in the pathogenesis of mods. pmns production of reactive oxygen species (superoxide and peroxynitrite) and inflammatory cytokines induces oxidative stress that leads to uncoupling of oxidative phosphorylation. the derangement in cellular energy metabolism is known as cytophathic hypoxia, a functional concept that points out the imbalance between adequate oxygen delivery and poor oxygen utilization at the mitochondrial level. when cytophathic hypoxia develops, the result is cellular dysfunction and death. although "bad" from a clinical point of view, cytophathic hypoxia may be a cellular adaptive response [171] as it can be viewed as a cellular hibernation-like state. however, if this phenomenon occurs for too long, irreversible cellular damage may result [172] . bacterial translocation is another proposed mechanism for sustained inflammation and development of multiple organ dysfunctions. although the "sustained-hit model" [173] predicts that the persistent stimulation of natural and acquired immune system by noxious stimuli is responsible for the maintenance of the inflammatory status, recent acquisitions implicate the apoptotic loss of δϒ t cells in the intestinal mucosa [170] . there is good evidence that ischemia-reperfusion injury is the main mechanism for loss of gut barrier function [174] . as oxygenation of the villi is dependent on a counter-current mechanism such that o 2 tension at the tip of the villi is lower than that of arterial blood, any decrease in splanchnic blood flow may be associated with bacterial translocation. at this point endotoxin, bacteria, or gut ischemia can stimulate the gut-associated lymphoid tissue to generate an immune-inflammatory response that affects distant organs. otherwise stated the gut becomes a pro-inflammatory organ, releasing cytokines and other inflammatory mediators which give origin to sepsis and mods. according to the point of view of the intensive care medicine, the whole body is constituted by seven physiologically interdependent organ systems: nervous, respiratory, cardiovascular, hepatic biliary, renal, digestive, and coagulation system. although many of them can be involved, the respiratory and cardiovascular systems are most frequently damaged with acute lung injury (ali)/acute respiratory distress syndrome (ards) and sepsis-induced hypotension being the commonest clinical presentations. it is well established that the number or dysfunctional organs affects the prognosis. thus the probability to die with only one single dysfunctional organ is fairly less than half with respect to two or more organ dysfunction [175] . moreover, a multicenter study established that the mortality increases almost linearly from 21.2 to 76.2% when organ dysfunction increases from 1 to ≥4 organ systems [176] . mortality is also influenced by comorbidities notably chronic renal failure, diabetes, and cancer [176, 177] . cumulative comorbidities also increase the risk for the development of mods with the highest mortality rate [178] . by converse, survivors suffer from a persistent chronic illness that may last for months or even years. this illness is characterized by prolonged icu stay, slow recovery from organs dysfunction, recurrent infections, progressive or permanent cognitive deficit, and loss of overall sense of well-being and function [179, 180] . so the duration of healthy life expectancy is reduced although the patient's overall life span is intact. the common pathophysiological denominator of mods is the loss of cell membrane barrier function. this finding is strikingly evident from the autopsies of patients who die as a result of multiple organs dysfunction syndrome. despite the clinical evidence of acute myocardial dysfunction, progressive cholestatic jaundice, continuing impairment of renal function, and similar dysfunction of other organ systems, the histopathological findings are remarkably normal or limited to mild tissue edema. therefore, the preserved organ morphology is strikingly in contrast with the impairment of organ function. the loss of cellular and tissue (endothelial) barrier function seems the cornerstone for this sort of hibernation that prevents the multiplicity of organ expression, variability, and communication. the commonest clinical presentations of single organ system dysfunctions are shown below. recent acquisitions indicate that the brain is often involved in sepsis and mods as it is one target of sirs either by direct activation of resident pro-inflammatory cells (neuroglia) or by mediators generated elsewhere (complement, cytokines). although the cns is considered an immuneprotected organ due to the blood-brain barrier, the abundance of glial cells with their signaling receptors makes the brain one of the preferred target for organ system dysfunction. therefore, the local inflammatory reaction that follows traumatic or hypoxic (e.g., post-cardiac arrest) brain damage can spread outside the cns thus exacerbating sirs. conversely, the circulating cytokines and other inflammatory mediators may increase the permeability of the blood-brain barrier thus causing brain damage [181] . cytokines activate monocytes to transform into glial cells (equivalent to resident macrophages in extra-cerebral tissues) so that inflammation continues in the nervous tissue [182] . circulating cytokines also can stimulate the afferent fibers of the vagus nerve that activate the central nervous system. after stimulation of vagus nerve endings, cerebral endothelial cells are activated resulting in the breakdown of the blood-brain barrier [183, 184] . the activation of cerebral endothelium also induces microvascular dysfunction, loss of cerebral vascular tone (autoregulation), and coagulopathy (microthrombi deposition) [185] . receptors for the commonest cytokines have been found in the hippocampus, one of the most involved regions for mnemonic elaboration and neuroplasticity [181] . chronic inflammation of the hippocampus can lead to irreversible cognitive decline, especially in the elderly. histopathologic lesions of the brain in sepsis include cerebral edema, infarct and ischemic lesions, and microabscesses [185] . sepsisassociated encephalopathy has been reported in 23% of people in the icu [186] . however, the true incidence is difficult to estimate because of lack of clear definition and subjective nature of its assessment. moreover, sedative drugs may blunt the symptoms thus making the diagnosis difficult. symptoms include changes in consciousness, awareness, cognition, and behavior. symptoms seem correlated with the global severity of illness as assessed by the acute physiology and chronic health evaluation ii score (apache ii score). it is by far the commonest organ system involved. ali and ards are the two clinical manifestations of respiratory dysfunction. ali is defined as the presence of bilateral pulmonary infiltrates on chest radiograph and arterial hypoxemia (pao 2 /fio 2 < 300). if the pao 2 /fio 2 drops below 200, the condition is classified as ards [187] . ards can originate from two distinct pathways: 1. direct pulmonary damage as occurs after trauma, infection, aspiration, or inhalation injury 2. indirect damage as occurs with sepsis, pancreatitis, extrapulmonary trauma and burns, blood transfusions, and fat embolism [188, 189] respiratory dysfunction span from extremely severe forms requiring immediate tracheal intubation, mechanical ventilation, and paralysis to milder forms with mild hypoxia and tachypnea. the lung is the most frequent organ involved in sepsis and trauma probably because it is the single organ with the largest vascular bed and it harbors one of the largest populations of resident macrophages. ards is characterized by neutrophil infiltration, alveolar-capillary barrier disruption, pulmonary vascular leakage, and edema. in most severe forms the alveoli contain eosinophilic protein-rich exudate with presence of hyaline membrane [190] . electron microscopic examination reveals extensive damage to type i alveolar pneumocytes with cellular swelling and blebbing of the plasma membranes [190] . pulmonary edema and absorption atelectasis decrease the pulmonary volume with the occurrence of refractory hypoxia by increased pulmonary shunt. moreover, the widespread deposition of microthrombi and hypoxic pulmonary vasoconstriction impose a pressure overload upon the right ventricle with myocardial dysfunction and failure. the initial clinical presentation is often subtle with tachypnea, dyspnea, dry cough, and agitation. hypoxia with hypocapnia is the most prominent laboratory feature. the magnitude of hypoxia is often unexplained by the mild or absent roentgenographic signs. ct scan reveals diffuse but nonuniform ground-glass consolidations often with a non-conformal gravitational distribution [191] . in more severe forms, multiple patchy areas of lung consolidation coalesce in massive airspace consolidation [191] . characteristically the distribution is uniform and affects all lung zones extending to the extreme periphery of the lung fields. hypoxia and interstitial/alveolar edema stimulate the carotid body chemoreceptors and intraparenchymal lung mechanoreceptors to hyperventilate with decreased arterial carbon dioxide tension. in the latest phases, normocapnia ensues mainly as result of increased respiratory work due to alveolar collapse, decreased pulmonary volume, and respiratory compliance. the mechanical disadvantage of decreased pulmonary compliance causes the abnormal increase of trans-pulmonary pressure (the pressure across the alveoli) that is the driving force for pulmonary expansion. the shear stress upon the delicate alveolar and interstitial structure causes further inflammation as a vicious cycle ensue with endothelial activation and worsening of pulmonary function that influences each other. mortality from ards ranges from 15 to 80% [189, 192] with average values of about 30% according to most recent ventilatory strategies [193] . it is one of the commonest organ dysfunction of septic and/ or traumatic origin. some studies report an incidence of up to 66% [194, 195] with a mortality rate as high as 70% [195, 196] . the cardiovascular dysfunction has long been recognized as a consequence of the generalized inflammatory reaction [197] . its most prominent clinical features are (1) myocardial dysfunction with biventricular dilation and reduced ejection fraction [198] and (2) arterial hypotension despite fluid volume infusion and decreased response to vasoactive drugs. it generally accepted that the decreased cardiac function is the product of circulating myocardial depressant factors (endotoxin, il1β, tnfα, il6, paf, and others) and mitochondrial damage [199] [200] [201] . some experimental evidence indicates that endotoxin acts by inducing an inflammatory response through heart tissue macrophage, mast cells, and infiltrating blood leukocytes. the clinical picture includes the following: arterial hypotension, mental impairment, warm and dry skin, oliguria, elevated arterial blood lactates, and higher than normal mixed or central venous saturation. the hemodynamic pattern shows a high cardiac output and decreased peripheral vascular resistance [202, 203] . the picture of low cardiac output state (pale and wet skin, oliguria, and hypotension) is rarely seen and indicates a more severe form of cardiac depression [202, 203] . when examined in detail, the cardiac function appears depressed as, despite the high output state, the ejected volume is lower than normal (reduced stroke volume), the ventricle is dilated (increased diastolic volume), and its contractile state is impaired with reduced ejection fraction (increased end-systolic volume). these changes are often masked by the decreased peripheral vascular resistance. the reduced afterload allows for the maintenance of a high output state as the dysfunctional ventricle pump blood against a dilated arterial system. when the ventricular dysfunction is severe, the clinical picture resembles that of cardiogenic shock with low cardiac output, high arterial lactates, and intense peripheral vasoconstriction. several studies have shown that cardiac dysfunction is biventricular in origin and that is potentially reversible by 7-10 days [204] . the other essential feature of cardiovascular dysfunction is arterial hypotension by decreased peripheral arterial resistance. because a major fraction of peripheral vascular resistance is located in peripheral arteries, this segment is designated as the resistance vessels. resistance vessels control the blood flow to organs, and it is under the control of either intrinsic (e.g., metabolic) or extrinsic (e.g., neural and humoral) factors. intrinsic regulation plays a fundamental role in pressure-flow autoregulation (ability of an organ to keep constant the blood flow despite fluctuations in arterial pressure), reactive hyperemia (ability to increase the blood flow after relief of a vascular obstruction), and functional hyperemia (ability to cope the organ blood flow in accordance with local metabolic needs) [205, 206] . in sepsis, intrinsic regulation is compromised in all the vascular beds with the exception of the cerebral vasculature [207, 208] . this can explain the mismatch between whole oxygen supply and metabolic demands of septic shock. the increased cardiac output does not cope with the cellular energy requirements so that hypoxia and increased acid production develop. by converse, the extrinsic regulation of resistance vessels is responsible for maintenance of arterial pressure and blood flow to the whole organs. during sepsis and after trauma or ischemia-reperfusion injury, a marked hypo-responsiveness to vasopressor agents is a frequent finding with development of arterial hypotension and tissue hypoperfusion. it is generally accepted that the decreased responsiveness to vasoconstrictor stimuli is consequent to increased production of endothelial nitric oxide (no) by the induced nitric oxide synthase (inos) [209] . in normal arterioles, a small amount of no is produced by endothelial nos (enos) as maintenance of cardiovascular homeostasis [209] . however, cytokines and endotoxins induce inos to produce large quantities of no [209, 210] . although inos induction can be viewed as an adaptive response to noxious stimuli, the resulting hypotension by massive no production is deleterious for organs and tissues. also, there is some evidence that vascular smooth muscle can be another relevant source of no production. in smooth muscle, neural nos (nnos) is upregulated thus contributing to no generation in septic animals [211] . another factor implicated in arteriolar hypo-responsiveness may be the complex of morphological alterations of the endothelial lining of the arterioles. experimental studies show that the endotoxin challenge results into endothelium disruption with cell swelling, pseudopod formation, and frank denudation of the endothelial lining [212, 213] . finally, the loss of capillary to arteriolar signaling may induce the hypo-responsiveness of arterioles to vasopressor stimuli. in normal conditions, capillaries may regulate the arteriolar tone by the upstream spread of electronic signaling along the endothelium. in sepsis, the cell-to-cell communication is impaired so that the modulation of blood flow delivery to capillaries is severely compromised [214] . taken all together the above mechanisms may lead to the inappropriate redistribution of blood flow to tissues and organs [215, 216] . because the arterial control of blood flow is responsible for the convective flux of oxygen to capillaries, the alteration of arterial tone may cause regional hypoxia [215] and ultimately multiple organ dysfunction. treatment principles of hyperdynamic shock are briefly outlined in chap. 3. the coagulation system is designed: 1. to stop active blood loss 2. to seal the site of lesion, thus allowing for pathogens destruction and removal of debris 3. to initiate the healing process the most severe form of coagulation disorder is the disseminated intravascular coagulation (dic). this pathophysio-logic mechanism results from the imbalance between coagulation and fibrinolysis with upregulation of the procoagulant pathway, downregulation of anticoagulant pathway (namely, atiii, protein c, and s system), and decreased fibrinolysis. coagulation is regulated by three main anticoagulant mechanisms: antithrombin, the protein c system, and the tissue factor pathway inhibitor (tfpi). the loss of physiologic anticoagulation in sepsis results from the action of several humoral (il1-β, tnf-α, and il6, c-reactive protein) and cellular (vascular endothelial cells, monocytes, macrophages, and platelet) pro-inflammatory mediators. furthermore, monocytes and macrophages are the main source of tissue factor (tf) a potent thrombin generator via the vii (extrinsic) pathway. the widespread fibrin deposition causes the formation of microthrombi with diffuse microcirculatory flow disturbance. tf is a membrane-bound protein that possesses the most important procoagulant activity [217] . after its expression on the cellular membranes, it forms complexes with the circulating factor vii of the coagulation cascade (extrinsic pathway) thus activating the formation of thrombin [218] . tf is abundant in the subcutaneous tissue and virtually in all blood-tissue barrier [219] . as seen above, tf is also inducible on monocyte and macrophage surfaces. in meningococcal septicemia, large quantities of tf have been localized in microparticles (mps) shed from platelets and leukocyte aggregates [220] . after trauma or infection, large amount of tf can be expressed by the denuded endothelium, damaged subcutaneous, and muscular tissues with priming of the coagulation cascade. similarly, sepsis can induce disseminated intravascular coagulation by cytokines production and monocyte adhesion to the activated endothelium with further tf expression. after thrombin formation, the von willebrand factor (vwf) is released from the weibel-palade bodies of endothelial cells. vwf binds to platelet glycoprotein iib to stabilize the adhesion of platelets and entrapped white blood cells aggregates to the injured endothelium [221] . normally vwf multimers are cleaved by the protein complex adamts13, and this process is stimulated under conditions of shear stress. sepsis is associated with decreased circulating adamts13 levels resulting in increased levels of ultra-large vwf multimers [221] . after adhesion to the endothelium (white thrombus formation), platelets enhances the inflammatory and coagulation cascade by expression of tf on activated monocytes and endothelial cells [222] . clinical conditions associated with overt dic are summarized in table 2 .3. the role of dic in the pathogenesis of mods is widely supported by experimental and postmortem findings. firstly, the demonstration of intravascular thrombi at autopsy appears related to the clinical dysfunction of the organ [223] . secondly, experimental studies demonstrated the deposition of intravascular and extravascular fibrin in the kidneys, lung, liver, and brain [223] . finally, clinical studies demonstrated the prognostic importance of dic as being an independent predictor of death [224] . the clinical diagnosis of dic relies upon the simultaneous presence of: 1. consumption of coagulation factors (prolonged clotting times) 2. consumption of coagulation inhibitors (e.g., decreased atiii activity) 3. platelets consumption or count of less than 100,000/mm 3 4. presence of fibrin degradation products (increased d-dimer) 5. presence of a condition known to be associated with dic in the past decade, clinical simple scoring systems for dic were introduced (table 2. they have individual strengths (jaam has the highest sensitivity, while isth is the most specific) and collective weaknesses (poor diagnostic sensitivity for late-onset dic) [228] . fibrinogen levels are not sensitive indicators of dic as fibrinogen synthesis is increased during sirs as occurs in the acute phase protein reaction. finally, the diagnosis of dic can be difficult in the presence of hepatic dysfunction or portal vein thrombosis as many of diagnostic clues of dic are present in these pathologic conditions. coagulation dysfunction affects up to 30% of patients with sepsis [229] , while thrombocytopenia (<10 5 platelets/mm 3 ) occurs in up to 60% with an inverse relationship between platelet count and severity of disease [230, 231] . acute kidney injury (aki) is defined as an absolute increase in serum creatinine of 1.5-fold from baseline or a urine output <0.5 ml/kg/h for 6 h [232] . overall aki frequency was reported over 20% in a large cohort study of 325,395 patients admitted to icu with odds of death increasing in parallel with increased severity of acute kidney injury [233] . crude mortality has been found ranging between 28 and 82% [234, 235] . the severity of aki is classified using the risk, injury, failure, loss and end-stage renal disease (rifle) criteria listed in table 2 .5, [236] . two main pathways are responsible for aki: one is characterized by acute tubular necrosis, renal hypoperfusion, and ischemia-despite common belief, this form accounts for only 22% of sepsis-induced aki [237] -and the second form is specific to mods, and its predominant mechanism is apoptosis induced by inflammatory cytokines [238] . this pathogenic mechanism is more adherent to the hyperdynamic circulatory state that follows post-traumatic sirs and sepsis [239] . experimental studies suggest that the decrease in glomerular filtration rate in aki is the result of a disproportionate vasodilation of the efferent than the afferent glomerular arterioles [240] . in normal condition, the vascular tone of afferent and efferent vessels is regulated by humoral intra-glomerular factors (mainly angiotensin, thromboxane a2, and pgi2). the interplay between vasoconstrictor (angiotensin and thromboxane a2) and vasodilator (pgi2) factors causes the tone of efferent vessels to increase with respect to the afferent arterioles. this allows for the glomerular capillary pressure to ensure normal filtration. during sirs or sepsis, the dilation of the efferent arterioles decreases the glomerular pressure and hence the filtration rate [241] . this pathophysiologic mechanism can explain the positive effects of norepinephrine or vasopressin infusion in patients with sirs and oliguria. despite the common belief, norepinephrine and vasopressin have been reported to increase the urinary output in patients with hyperdynamic shock. their vasoconstrictor effect is probably helpful in restoring the tone of the efferent arteriole with resulting increase of capillary perfusion pressure. in addition to glomerular hemodynamic changes, circulating or locally (mesangial cells) produced tnfα induces tubular cell apoptosis with imbalance in water and electrolytes homeostasis [242] . aki is an important form of organ dysfunction because of its association with high mortality rates. in a multinational, multicenter study, the prevalence of aki has been estimated to be 5-6% of people with only 40% of them discharged alive [243] . septic shock is the most common cause of aki with a prevalence of 65% [244] . trauma patients who developed aki showed 29% of mortality rate as compared with 9% of the overall mortality rate [245] . the gut has long been recognized as the "motor" of mods due to its capability of amplifying the sirs response during shock and gut hypoperfusion. the incidence of gastrointestinal dysfunction is difficult to estimate due to the subjective nature of diagnosis and the lack of a clear diagnostic definition. symptoms and signs of intestinal dysfunction include hyporexia, anorexia, inability to tolerate enteral feeding, decreased intestinal motility, and diarrhea often of hemorrhagic type [246, 247] . after a catastrophic hypoperfusion of the gut as occurs with cardiac arrest or severe multiple jmhw japanese ministry of health and welfare [225] , isth international society of thrombosis and haemostasis [226] , jaam japanese association of acute medicine [227] a if hematologic malignancy, 0 pts for bleeding and platelets and add 3 pts to the total score loss of renal function >4 weeks end-stage renal disease (e) loss of renal function >3 months modified from [236] trauma, the normal intestinal flora is destroyed and substituted by an increasing number of pathogens [248] . this phenomenon is detrimental because the normal flora protects against the colonization of microbial pathogens thus preventing bacterial translocation (see above) [249] . however, the change of normal flora together with reduced intestinal motility and disruption of normal gut mucosal barrier increase the risk for bacterial translocation and development of mods. the loss of mucosal barrier function is the result of endotoxin, inflammatory cytokines, and decreased production of tight junction proteins [250] . hepatic dysfunction is schematically divided into two clinical forms: hypoxic hepatitis (hh) and jaundice-induced cholestasis (jic). hh can result from absolute (e.g., cardiogenic) or relative decrease in cardiac output with reduced oxygen supply to the liver. during septic shock, cardiac output and oxygen delivery are increased albeit not enough to cope with the high oxygen requirement of the liver and the inability of cells to extract oxygen [45] . moreover, the hepatic arterial buffer response and other vascular mechanisms of defense against portal blood flow reduction are impaired [251] . hh is also the product of oxidative stress after ischemia/reperfusion injury and the action of endotoxin and other inflammatory mediators which lead to activation of kupffer cells and activation and recruitment of leukocytes. the diagnosis of hh requires: 1. a clinical setting of impaired oxygen delivery (usually due to cardiac and/or respiratory failure) 2. a greater than 20-fold increase in serum aminotransferase activity 3. the exclusion of other causes of liver cell necrosis [252, 253] another landmark of hh is the acute drop of prothrombin levels which gives origin to a hemorrhagic syndrome [254] . these acute biochemical changes, especially those reflecting acute liver cells necrosis, decrease rapidly within 2 or 3 days and normalize within approximately 2 weeks. the delayed elevation of serum bilirubin is often seen but without apparent jaundice. jaundice-induced cholestasis is a more subtle form of hepatic jaundice usually associated with severe infections [255, 256] . the increase of bilirubin is a late event in these septic patients [257] , and histological studies reveal the intrahepatic origin of the cholestasis [258] . the reduction of bile salt excretion in the gut is responsible for the atrophy of the mucosal gut barrier [259] with loss of integrity and increased risk of bacterial translocation. moreover, the gut is deprived by the bacteriostatic action of bile salts with proliferation of pathogens and increased production of endotoxin [260, 261] . this creates a vicious circle that perpetuates the infection and hence further cholestatic liver damage. the production and transport of bile salts are a complex, highly coordinated, and energy-dependent mechanism. during sepsis many of these mechanisms are impaired because of a lack of energy due to hypoxia or hypoperfusion or both [262] . the major limiting factor for the bile flow at the canalicular level is the atpdependent bile salt export pump that is severely impaired in the septic liver [262] . the severity of hyperbilirubinemia appears directly related to the severity of impairment in the various steps of bile formation. no specific therapies can be recommended for acute liver dysfunction. the most recommended measures are merely supportive including the normalization of perfusion pressure and restoration of oxygen delivery [263] , the maintenance of normal oxygen tension, and serum glucose levels. the early use of enteral nutrition is also helpful in counteracting the atrophy of gut mucosal barrier and reducing the intestinal pathogens [264] . a high-impact trauma victim was admitted to ed with head trauma and severe hemorrhagic shock by ruptured spleen and multiple bone fractures. the patient was unconscious; his blood pressure was 85/55 mmhg with 135 bpm pulse rate. metabolic acidosis (ph 7.27, paco2 32 mmhg) and hyperlactacidemia (7.5 mmol/l) were also present. splenectomy and external fixation of bone fractures were quickly performed. during surgery, the blood pressure remained at about 100/60 with an average 120 bpm pulse rate. despite several liters of crystalloids and transfusion of blood products (4 prc and 4 ffp), an infusion of norepinephrine up to 0.15 μg/kg/min was started. at icu admission, the patient had stable vital signs (ap 115/50 mmhg) a mild tachicardia (hr 115 bpm), and compensated metabolic acidosis (ph 7.37, paco2 30 mmhg) with elevated arterial blood lactates (7 mmol/l). after 12 h from icu admission, the resuscitation phase appeared complete. at 24 h, fever of 38.8 °c, leukocytosis 21,000 cells/mm 3 , respiratory alkalosis, and tachycardia prompted the execution of a total body ct scan. a. sepsis b. sirs c. intra-abdominal abscess d. fever of cns origin a. il-4 and il-10 b. il-1 and tnf c. il-6 and il-7 d. the ct scan excluded intra-abdominal abscesses and revealed large bilateral psoas muscle and pelvic hematomas. microbiological cultures from urine, bronchial tube drainage fluids were negative. within 5 days fever and tachycardia disappeared so that the definitive fixation of bone fractures was scheduled for the following week. at day 10, a new appearance of fever and leukocytosis occurred. a. infection b. sirs c. drugs-induced fever d. fever of unknown origin the chest roentgenogram showed a right perihilar infiltrate and purulent bronchial secretions. broad spectrum antibiotics were started, but 24 h thereafter the patient develops hypotension, oliguria, metabolic acidosis, altered mental status, and decreased platelet count. a a multi-sensitive klebsiella pneumoniae was isolated from blood cultures, and antimicrobial therapy allows for rapid control of the septic source and resolution of the clinical picture. the patient was discharged from intensive care after 15 days and returned at home within 60 days from the hospital admission. please see chap. 58 for the correct answer. sir isaac newton, sepsis, sirs and cars of critical care medicine consensus conference: definitions for sepsis and organ failure and guidelines for the use of innovative therapies in sepsis the systemic immune response to trauma: an overview of pathophysiology and treatment 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production is not a generalized phenomenon: a review pamps and alarmins: all we need to know about danger how tissue injury alarms the immune system and causes a systemic inflammatory response circulating mitochondrial dampss cause inflammatory response to injury bacterial endotoxin stimulates macrophages to release hmgb1 partly through cd14-and tnf-dependent mechanisms the impact of endogenous triggers on traumaassociated inflammation trauma alarmins as activators of damage-induced inflammation early complementopathy after multiple injuries in humans molecular mechanisms of inflammatory and tissue injury after major trauma-is complement the "bad guy the role of complement in trauma and fracture healing alarmin(s) news about danger: workshop on innate danger signals and hmgb1 proinflammatory cytokines (tumor necrosis factor and interleukin 1) stimulate release of high mobility group protein-1 by pituicytes release of chromatin protein hmgb1 by necrotic cells triggers inflammation the nuclear protein hmgb1 is secreted by monocytes via a non-classical, vesicle-mediated secretory pathway high mobility group 1 protein (hmg-1) stimulates proinflammatory cytokine synthesis in human monocytes regulation of monocyte migration by amphoterin (hmgb1) inflammatory mediators in relation to the development of multiple organ failure in patients after severe blunt trauma the complement system in trauma-related and ischemic tissue damage: a brief review interleukin-6 potentiates neutrophil priming with plateletactivating factor trauma primes cells trauma: the role of the innate immune system neutrophil priming and activation in the pathogenesis of postinjury multiple organ failure neutrophil recruitment under shear flow: it's all about endothelial rings and gaps inflammatory serum markers in patients with multiple trauma. can they predict outcome systemic inflammation after trauma differentiation of effector c4 t cell populations postinjury neutrophil priming and activation: an 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inflammation complement activation in injured patients occurs immediately and is dependent on the severity of the trauma the role of the complement system in traumatic brain injury femur fracture induces site-specific changes in t-cells immunity reperfusion injury of ischemic skeletal muscle is mediated by natural antibody and complement innate autoimmunity natural antibodies, autoantibodies and complement activation in tissue injury role of biological modifiers regulating the immune response after trauma the role of neuroinflammation in traumatic brain injury evidence for a role of kallikrein-kinin system in patients with shock after blunt trauma molecular and functional interactions among monocytes, platelets, and endothelial cells and their relevance for cardiovascular diseases platelets: bridging hemostasis, inflammation and immunity coagulation abnormalities in acute lung injury and sepsis priming for enhanced alveolar fibrin deposition after hemorrhagic shock: role for tumor necrosis factor new treatment strategies for disseminated intravascular coagulation based on current understanding of the pathophysiology systemic inflammation and disseminated intravascular coagulation in early stage of ali and ards: role of neutrophil and endothelial activation extracellular proteolysis in brain injury and inflammation: role for plasminogen activators and matrix metalloproteinases the acute phase response function of c-reactive protein high concentrations of lipopolysaccharide-binding protein in serum of patients with severe sepsis or septic shock inhibit the lipopolysaccharide response in human monocytes neutrophil-mediated tissue injury and its modulation role of nitric oxide in inflammation the expanding universe of t-cell subsets: th1 and th2 and more signalling events involved in interferon-gamma-inducible macrophage nitric oxide generation th1 and th2 subsets: paradigms lost? marked elevation of human circulating cd4 + cd25 + regulatory t cells in sepsis induced immunoparalysis human cd4 + cd25 + regulatory t lymphocytes inhibit lipopolysaccharide-induced monocyte survival through a fas/fas ligand-dependent mechanism immunity to fungal infections th17 cytokines and the gut mucosal barrier giammarellos-bourboulis ejg. deficient candida-specific t-helper 17 response during sepsis longitudinal study of cytokine and immune transcription factor mrna expression in septic shock immunoparalysis after multiple trauma il-10-induced micro-rna-187 negatively regulates tnf-alpha, il-6 and il-12p40 production in tlr4-stimulated monocytes persisting low monocyte human leukocyte antigen-dr expression predicts mortality in septic shock inflammation endothelium and coagulation in sepsis plasma levels of the three endothelialspecific proteins von willebrand factor, tissue factor pathway inhibitor, and thrombomodulin do not predict the development of acute respiratory distress syndrome elevated circulating e-selectin, intercellular adhesion molecule 1, and von willebrand factor in patients with severe infection a single endotoxin injection in the rabbit causes prolonged blood vessel dysfunction and a procoagulant state vascular endothelial cell dysfunction in septic shock anticoagulant properties of the vascular endothelium initiation and regulation of tissue factor-dependent blood coagulation reversible regulation of tissue factor-induced coagulation by glycosyl phosphatidylinositol-anchored tissue factor pathway inhibitor the tissue factor-factor viia complex: procoagulant activity, regulation, and multitasking protein c anticoagulant pathway and its role in controlling microvascular thrombosis and inflammation plasminogen activator inhibitor 1: physiological and pathophysiological roles organ distribution of fibrin in disseminated intravascular coagulation recombinant e. coli-derived tissue factor pathway inhibitor reduces coagulopathic and lethal effects in the baboon gram-negative model of septic shock delayed treatment with recombinant human tissue factor pathway inhibitor improves survival in rabbit gram-negative peritonitis traffic signals on endothelium for lymphocyte recirculation and leukocyte emigration mechanisms of endothelial cell injury in acute inflammation leukocyte trafficking in alveoli and airway passage endothelial injury in sepsis mechanistic coupling of protease signaling and initiation of coagulation by tissue factor activation of endothelial cell protease activated receptor 1 by the protein c pathway regulation of myosin phosphatase by rho and rho-associated kinase (rho-kinase) stunning" following a brief exposure to endotoxin: a mechanism to link infection and infarction? incidence of myoendothelial gap junctions in the proximal and distal mesenteric arteries of the rat is suggestive of a role in endothelium-derived hyperpolarizing factormediated response endothelial cell pathway for conduction of hyperpolarization and vasodilation along hamster feed artery microvascular recruitment in hamster striated muscle: role for conducted vasodilation the splanchnic circulation: no longer a silent partner pathological supply dependence of systemic and intestinal o2 uptake and endotoxemia microvascular perfusion is impaired in a rat model of normotensive sepsis heterogeneity of gut capillary transit times and impaired gut oxygen extraction in endotoxemic pigs effect of a maldistribution of microvascular blood flow on capillary o2 extraction in sepsis cutting edge: critical role of mesothelial cells in necrosis-induced inflammation through the recognition of il-1 alpha released from dying cells interleukin-1 in the pathogenesis and treatment of inflammatory diseases interleukin-10 down regulates mhc class ii alphabeta peptide complexes at the plasma membrane of monocytes by affecting arrival and recycling immediate il-10 expression following major orthopaedic trauma. relationship to anti-inflammatory response and subsequent development of sepsis the significance and challenges of monocyte impairment: for the patient and the surgeon depletion of dendritic cells, but not macrophages, in patients with sepsis leukocyte apoptosis and its significance in sepsis and septic shock apoptosis and surgical trauma: dysregulated expression of death and survival factors on peripheral lymphocytes cell death: critical control points how cells die: apoptosis pathways human caspases: activation, specificity, and regulation cell death during sepsis: integration of disintegration in the inflammatory response to overwhelming infection the sirens' song dysregulated expression of neutrophil apoptosis in the systemic inflammatory response syndrome apoptotic cell death in patients with sepsis, shock and multiple organ dysfunction prolonged lymphopenia, lymphoid depletion and hypoprolactinemia in children with nosocomial sepsis and multiple organ failure rapid onset of intestinal epithelial and lymphocyte apoptotic cell death in patient with trauma and shock mechanisms of organ dysfunction in critical illness: report from a round table conference held in brussels mechanisms of sepsis-induced organ dysfunction sepsis-induced immunosuppression: from cellular dysfunctions to immunotherapy delayed hypersensitivity: indicator of acquired failure of host defenses in sepsis and trauma herpes simplex virus lung infection in patients undergoing prolonged mechanical ventilation cytomegalovirus reactivation in critically immunocompetent patients the late phase of sepsis is characterized by an increased microbiological burden and death rate predictors of 30-day mortality and hospital costs in patients with ventilator-associated pneumonia attributed to potentially antibiotic-resistant gram-negative bacteria bench to bedside review: endotoxin tolerance as a model of leukocyte reprogramming in sepsis endotoxin tolerance: new mechanisms, molecules and clinical significance lipopolysaccharide structure-function relationship in activation versus reprogramming of mouse peritoneal macrophages immunosuppression in patients who die of sepsis and multiple organ failure interleukin-7 and immune reconstitution in cancer patients: a new paradigm for dramatically increasing overall survival effects of recombinant human interleukin-7 on t-cell recovery and thymic output in hiv-infected patients receiving antiretroviral therapy: result of a phase i/iia randomized, placebo-controlled, multicenter study overexpression of bcl-2 in transgenic mice decreases apoptosis and improves survival in sepsis il-7 restores lymphocyte function in septic patients il-7 promotes t cell viability, trafficking, and functionality and improves survival in sepsis lymphocyte subset responses to trauma and sepsis cd4+ t-lymphocyte alterations in trauma patients early circulating lymphocyte apoptosis in human septic shock is associated with poor outcome early assessment of leukocyte alterations at diagnosis of septic shock icu-acquired immunosuppression and the risk for secondary fungal infections regulatory t cell populations in sepsis and trauma increased proportion of cd4(+) cd25(+)foxp3(+) regulatory t cells during the early-stage sepsis in icu patients innate immune functions of immature neutrophils in patients with sepsis and severe systemic inflammatory response syndrome neutrophil paralysis in sepsis the function of neutrophils in sepsis expression and function of the chemokine receptor cxcr1 and cxcr2 in sepsis neutrophils are significant producers of il-10 during sepsis a subset of neutrophils in human systemic inflammation inhibits t cell responses through mac-1 cd11+/gr-1+ myeloid suppressor cells cause t cell dysfunction after traumatic stress adib-conquy m. captain study group. toll-like receptors expression and interferon-gamma production by nk cells in human sepsis association of ϒδ tcells with disease severity and mortality in septic patients multiorgan failure is an adaptive, endocrine-mediated, metabolic response to overwhelming systemic inflammation the multiple organ dysfunction syndrome evolving concepts in the pathogenesis of postinjury multiple organ failure current status of bacterial translocation as a cause of surgical sepsis the impact of acute organ dysfunction on patients' mortality with severe sepsis epidemiology of severe sepsis in the united states: analysis of incidence, outcome and associated costs of care the epidemiology of sepsis in patients with malignancy the role of infections and comorbidity. factors that influence disparities in sepsis the lingering consequences of sepsis: the hidden public health disaster? hmgb1 mediates cognitive impairment in sepsis survivors cytokines and the nervous system ii: actions and mechanisms of action resolving postoperative neuroinflammation and cognitive decline understanding brain dysfunction in sepsis sepsis-associated encephalopathy: review of neuropsychiatric manifestations and cognitive outcome the neuropathology of septic shock impact of encephalopathy on mortality in the sepsis syndrome. the veterans administration systemic sepsis cooperative study groups acute respiratory distress syndrome: the berlin definition pulmonary and extrapulmonary forms of acute respiratory distress syndrome epidemiology of ards and ali pulmonary edema (including respiratory distress syndrome) chapter 14: pulmonary edema is the mortality higher in the pulmonary vs the extrapulmonary ards? a meta-analysis ventilation with lower tidal volumes as compared with traditional tidal volumes for acute lung injury and the acute respiratory distress syndrome the epidemiology of acute organ system dysfunction from severe sepsis outside the intensive care unit epidemiology of severe sepsis in japanese intensive care units: a prospective multicenter study cub-réa study group. incidence and impact of organ dysfunctions associated with sepsis myocardial dysfunction in septic shock: part 1. clinical manifestation of cardiovascular dysfunction profound but reversible myocardial depression in patients with septic shock prevention of endotoxin-induced sarcoplasmic reticulum calcium leak improves mitochondrial and myocardial dysfunction tumor necrosis factor-alpha decreases sarcoplasmic reticulum ca2+-atpase expressions via the promoter methylation in cardiomyocytes endotoxin and cytokines induce direct cardiodepressive effects in mammalian cardiomyocytes via induction of nitric oxide synthase patterns of septic shock in man: a detailed study of 56 patients bacteremia due to gram-negative bacilli other than the salmonella: a clinical and therapeutic study right ventricular dysfunction and dilatation, similar to left ventricular changes, characterize the cardiac depression of septic shock in humans microcirculation of the intestinal mucosa tissue perfusion and organ function: ischemia/reperfusion injury sepsis-induced vasoparalysis does not involve the cerebral vasculature: indirect evidence from autoregulation and carbon dioxide reactivity studies cerebral hemodynamics vascular reactivity and metabolism during canine endotoxin shock control of vascular reactivity inducible nitric oxide synthase and vascular reactivity in rat thoracic aorta: effect of aminoguanidine endotoxin induced skeletal muscle contractile dysfunction: contribution of nitric oxide synthases leukocyte activation does not mediate myocardial leukocyte retention during endotoxemia in rabbits endothelial structural integrity is maintained during endotoxic shock in an interleukin-1 type 1 receptor knockout mouse capillary and arteriolar responses to local vasodilators are impaired in a rat model of sepsis vascular reactivity and tissue oxygenation cardiac output and redistribution of organ blood flow in hypermetabolic sepsis the role of the tissue factor pathway in initiation of coagulation structural biology of tissue factor, the initiator of thrombogenesis in vivo cell biology of tissue factor, the principal initiator of blood coagulation cellular origin and procoagulant properties of microparticles in meningococcal sepsis inflammation-associated adamts13 deficiency promotes formation of ultra-large von willebrand factor tissue factor expression by monocytes: regulation and pathophysiological roles microvascular thrombosis and multiple organ dysfunction syndrome treatment effects of drotrecogin alfa (activated) in patients with severe sepsis with or without overt disseminated intravascular coagulation criteria for diagnosis of dic based on the analysis of clinical and laboratory findings in 345 dic patients collected by the research committee on dic in japan towards definition, clinical and laboratory criteria, and a scoring system for disseminated intravascular coagulation a multicenter, prospective validation of disseminated intravascular coagulation diagnostic criteria for critically ill patients: comparing current criteria prospective evaluation of three different diagnostic criteria for disseminated intravascular coagulation the hematologic system as a marker of organ dysfunction in sepsis incidence, risk factors, and outcome of severe sepsis and septic shock in adults: a multicenter prospective study in intensive care unit. french icu group for severe sepsis coagulation system and platelets are fully activated in uncomplicated sepsis epidemiology of sepsis acute kidney injury incidence and outcomes of acute kidney injury in intensive care units: a veterans administration study incidence classification and outcomes of acute kidney injury developing a consensus classification system for acute renal failure acute dialysis quality initiative workgroup: acute renal failuredefinition, outcome measures, animal models, fluid therapy and information technology needs the histopathology of septic acute kidney injury: a systematic review glucocorticoids potently block tumor necrosis factor-alpha and lipopolysaccharide-induced apoptotic cell death in bovine glomerular endothelial cells upstream of caspase 3 activation renal blood flow distribution in experimental septic acute renal failure pathophysiology of septic acute kidney injury: what do we really know? angiotensin ii in experimental hyperdynamic sepsis production of tumor necrosis factor by rat mesangial cells in response to bacterial lipopolysaccharide beginning and ending supportive therapy for the kidney (best kidney). acute renal failure in critically ill patients: a multinational multicenter study cooperative antimicrobial therapy of sepstic shock (catss) database research group. acute kidney injury in septic shock: clinical outcomes and impact of duration of hypotension prior to initiation of antimicrobial therapy incidence and outcome of early acute kidney injury in critically ill trauma patients altered gastrointestinal motility in critically ill patients: current understanding of pathophysiology, clinical impact, and diagnostic approach gastrointestinal dysmotility: clinical consequences and management of the critically ill patient dramatic changes of the gut flora immediately after severe and sudden insults gut flora in health and disease epithelial barrier leak in gastrointestinal disease and multiorgan failure mechanism and role of intrinsic regulation of hepatic arterial blood flow: hepatic arterial buffer response hypoxic hepatitis: underlying conditions and risk factors for mortality in cirtically ill patients hypoxic hepatitis: clinical and hemodynamic studies in 142 consecutive cases hypoxic hepatitis hepatic dysfunction during bacterial sepsis liver function in septic shock the use of maximum sofa score to quantify organ dysfunction/failure in intensive care. results of a prospective, multicenter study. working group on sepsis relate problems of the esicm sepsis and cholestasis pathophysiology of increased intestinal permeability in obstructive jaundice the value of bile replacement during external biliary drainage: an analysis of intestinal permeability, integrity, and microflora effect of internal biliary drainage on plasma levels of endotoxin, cytokines, and c-reactive protein in patients with obstructive jaundice sepsis and cholestasis surviving sepsis campaign: international guidelines for management of severe sepsis and septic shock guidelines for the provision and assessment of nutrition support therapy in the adult critically ill patient: society of critical care medicine (sccm) and american society for parenteral and enteral nutrition key: cord-015082-l629n8is authors: nan title: poster sessions 323-461 date: 2002-08-29 journal: intensive care med doi: 10.1007/s00134-002-1455-7 sha: doc_id: 15082 cord_uid: l629n8is nan total protein concentration in bal increased significantly and led to peak at 1689±323 mg/ml 1hour after intubations . mucin concentration was highest at 1hour after ventilation (155.75±31.6mg/ml). bal sp-a concentration ratio increased about 20 times after 1hour ventilation. compare to 100mg/ml total protein, the ratio was 2.16±1.14 in 1hour later, and 4.31±2.8 in 3hours after ventilation.the change of bal wbc level led to peak in after 1hour ventilation, but blood wbc level led to peak in 3hours later. for elastase level both peak were 3hours later in bal and blood.in the caller components of bal, the neutrophyl cells were dominant in 1hour after intubation, but 3hours after ventilation, mast cells with phagocyted mucine and dusts were dominant. just introduction. coronary disease is prevalent in diabetic patients resulting in a frequency of invasive cardiac procedures four times that of non-diabetics. after cardiac surgery diabetics have twice the mortality and morbidity in early and late phases after operation. the reasons for this increased risk are poorly understood. diabetics exhibit complex abnormalities of lung structure and of the control of the cardiorespiratory system. these include pulmonary micro-vascular disease, autonomic neuropathy associated with an increased cardiovascular instability, an increased incidence of central and obstructive sleep apnoea and a reduced response to hypercapnia. this study was undertaken to determine whether at risk diabetic patients could be identified pre-operatively. methods. 14 patients awaiting urgent cardiac surgical re-vascularisation were studied with measurement of: spirometry; percentage increase in transfer factor from sitting to lying position (tf) as an indicator of micro-vascular lung disease; overnight oximetry on air; and 24hour holter monitoring at present arf is one of the most spread and serious complication of postoperation period. practically the experience of carring nimv on patients with arf on early stadies of mosf is absent. until now, the criteria of uneffectiveness of nimv and indications for cessation of mask ventilation and moving of patients to mechanical ventilation are not determined. methods. there were included 48 patients with ali in the examination. the cause of this condition was the mosf, developed in postoperative period. diagnosis of ali/ards was stated on the criteria adopted the american european consensus conference on ards (1). presence of organs failure was determined on multiple organ dysfunction score (table 1) . nimv was carried out by seances from 5 to 8 hours. average duration of nimv consisted 39.5±5.3 hours. results. improvement of gases change was determined on 28 patients (58%) out of 48. though 20 patients with mosf were reintubated, out of which 13 patients (27%) died lately as the result of mosf progressing. the condition of gases changing functions before intubation is one of the determining factors of prognosis. the patients reintubated under satisfactory indices of gase blood composition and early symptoms of mosf survived. patients, who were reintubated on decreased indices of arterial oxygenation under mosf progressing died in 100% cases ( nimv is effective method in complex therapy of arf, developing in postoperative period after cardiac surgery, that leads to significant improvement of lungs biomechanics and gases change function. progressing of mosf and storage disturbance of lung oxygenation is absolute indication for intubation and applications of special regimes of mechanical ventilation. references. 1. bernard gr, artigas a, brigham kl. am j respir crit care med -1994: 149:818 introduction. several bioimpedance cardiac output systems have been developed in the past in order to measure cardiac output in a wide variety of clinical situations. however, open thorax surgery negatively influences the accuracy of the measurement of thoracic electrical bioimpedance cardiac output (teb-co) (1). the purpose of the present study was to evaluate the performance of a new bioimpedance cardiograph hl-4 (vrije universiteit medical centre amsterdam and hemologic amersfoort, the netherlands), using a new algorithm and a new electrode configuration, during open and closed chest in cabg patients, comparing teb-co with transcardiopulmonary thermodilution (tcpco). methods. after hospital ethics committee approval and written informed consent, fourteen patients with preserved lv-function at cineangiography or echocardiography, scheduled for coronary artery bypass grafting were included. for the teb system two current injecting electrodes were placed on the forehead and the left thigh respectively and two voltage sensing electrodes were used: one above the left clavicle at the base of the neck and the other at the level of the xyphoid in the left midaxillary line. for tcpco, the picco-system (pulsion, munich, germany) was used. hemodynamic measurements were recorded at three time points: t1 before the operation, t2 after weaning from bypass before sternal closure and t3 after sternal closure. teb-co and tcpco data were compared with pearson's r correlation coeficient. p<0.05 was considered significant. bland-altman analysis (2) with bias and precision was carried out at each of the three time points. results. ten males and 4 females with age 64±9 yr, body weight 78 ± 13 kg and height 172±8 cm were included. a total of 34 matched data pairs were available for analysis. table 1 shows the results of correlation, bias and precision of the measurements at the three different time points. teb consistently underestimated tcpco. at all time points, there was a good correlation between both techniques. introduction. isoflurane sedation of icu patients has previously been shown to be useful but has not come into wide clinical use for a number of reasons.a new device(the anesthetic conserving device,"acd") enables easy and safe administration of isoflurane in the icu setting.we conducted a randomised, controlled study to evaluate efficacy of sedation and environmental safety during administration of isoflurane with the acd. the acd is a modified heat and moisture exchanger connected to the breathing circuit at the endotracheal tube.isoflurane is administered via a syringe pump to a vaporiser rod in the acd.due to the physical properties of the acd most of the exhaled isoflurane is returned to the patient.5 mechanically ventilated patients were randomised to receive isoflurane via the acd.4 control patients received midazolam intravenously. all patients received morphine analgesia. quality of sedation was assessed hourly in all patients."adequate sedation" was pre-defined as a set interval on the bloomsbury sedation scale. additionally, the patient's nurse determined if sedation over the previous hour in general had been adequate or not. time from discontinuation of the sedative drug until the patient followed verbal command and to extubation was compared between groups. in the isoflurane group a gas evacuation system was used during isoflurane administration. athmospheric concentration of isoflurane was measured at 0.5 m from the acd. results. in the isoflurane group patients were adequately sedated by the bloomsbury scale for 58 ± 9 % of the study period, compared to 50 ± 28 % in the control group.nurse satisfaction in the isoflurane group was 88 % of time and 57% of time in the control group.mean time to extubation after cessation of sedative administration was 11 min in the isoflurane group and 1864 min in the control group, mean time to patient cooperation was 55 min in the isoflurane group, and 1866 min in the control group. no significant hemodynamic changes were noted at initiation of the sedation in either of the groups. no serious complications related to sedation were noted in either group.opioid requirements in the isoflurane group were lower, with a mean rate of 1.6 0.9 mg/hr, compared with a mean rate of 4.2 2.4 mg/hr in the control group.mean isoflurane infusion rate was 2.5 ml/hr, with mean end-tidal isoflurane concentrations of 0.37% (0.19-0.82%).environmental levels of isoflurane were generally low,with a mean of 0.43 ± 0.27 ppm, well below the recommended long-term exposure limit of 2 ppm. brief peaks (<2min) between 2 and 15 ppm were noted during endotracheal suctioning, etc on an average of 0.1 times/hour of exposure. conclusion. isoflurane administered via the acd for sedation of icu patients is environmentally safe, requires small volumes of isoflurane and may provide better quality of sedation than midazolam. it appears to be more titratable with a shorter time from adequate sedation to extubation and ability to cooperate. references. millane ta, bennett ed,grounds rm,anaesthesia 1992;47:768-774.spencer em,willatts sm,intensive care brudney c. s. 1 , gosling p. 2 , manji m. 3 1 anaesthesia, 2 biochemistry, 3 anaesthesia, university of birmingham, birmingham, united kingdom increased capillary permeability has been implicated in the pathogenesis of ards and organ failures. surgery and ischaemia-reperfusion injury are both associated with stimulation of the acute inflammatory response, an early feature of which is an increase in systemic capillary permeability. the kidneys amplify small changes in systemic capillary permeability (1).the aim of this study was to explore any association between acr during and after cardiopulmonary bypass (cpb) and subsequent pulmonary and renal function. methods. forty patients (9 female) mean (range) age 67.8 (50-85) yrs undergoing coronary artery bypass grafting were enrolled. patients with severely impaired left ventricular function (<35% ef) were excluded. ten ml of urine was collected at intervals from the start of surgery until 48 hours post cpb. microalbuminuria was measured by automated immunoturbidimetry and expressed as the albumin creatinine ratio (acr: ref. range <2.3 mg/mmol). acr was compared with po2/fio2 ratio, hours on ippv, renal function and duration of inotropic support, using spearman's rank correlation procedure. results. two patients were excluded (death at 6 hours and acute renal failure post cpb). the median (range) duration of ippv was 15 (4-72) hours. 22 patients required inotropic support for median (range) 12 (2-96) hours. median (range) acr increased during surgery and was maximal 10 minutes post cpb. (table) two hour acr was inversely correlated with the mean po2/fio2 ratio up to 12 hours (rs = -0.46 p = 0.0044). two and 12 hour acrs were both positively associated with duration of ippv (rs = 0.46 p = 0.0071 and 0.62 p <0.0001 respectively). acr at 4 and 12 hours were associated with serum creatinine 24 hours post cpb, (rs = 0.35 p = 0.045, rs = 0.50 p = 0.003 respectively). acr at 2, 4 and 12 hours post cpb were associated with serum creatinine 48 hours post cpb (rs = 0.45 p = 0.033, rs = 0.59 p = 0.003 and rs = 0.49 p = 0.016 respectively). there was no significant association between duration of inotropic support and acr at any time point up to 48 hours. conclusion. cpb leads to a perioperative microvascular insult, causing increased capillary permeability which influences later pulmonary and renal function. these rapid changes in microvascular permeability can be monitored as the acr, and in the patient group studied, the magnitude of the acr as early as 2 hrs post cpb is associated with later organ function. acr may provide a tool allowing early identification of patients at risk of developing organ dysfunction, who may benefit from early intervention aimed at modifying the inflammatory response. acute lung injury (ali) is a major complication of gram-negative bacterial sepsis. to date, bacterial lipopolysaccharide has been held responsible for triggering ali (1). whether additional bacterial toxins play a role in the development of acute pulmonary inflammation during gram-negative sepsis remains an unresolved issue. flagellin, a principal component of bacterial flagella, has been recently shown to elicit immune responses via activation of the toll-like receptor 5 (2). we have newly found that flagellin induces an expression of icam-1 and a massive production of il-8 by human lung epithelial cells. in mice, flagellin produces a severe acute lung inflammation with local release of pro-inflammatory cytokines, accumulation of inflammatory cells and increased pulmonary permeability that was more pronounced than following endotoxin (3). the purpose of the present investigation was to evaluate the influence of flagellin on lung fluid filtration in rats. wistar rats (250-300 g) were exposed either to intravenous injection of flagellin 0.1-2 mg/kg or corresponding volume of normal saline (controls). after 8-20 h, the rats were anesthetized and the lungs were isolated. the isolated lungs were ventilated under a normoxic condition and perfused with homologous blood (37ºc) at a constant flow for 4 h or until development of irreversible edema. airway pressure, pulmonary arterial pressure, pulmonary vascular resistance, and changes in the lung weight were assessed. the increments in outflow pressure of 0.77 kpa for 6 min were used to determine the fluid filtration rate and filtration coefficient in the lungs every 30 min (4). flagellin induced a dose-and time-dependent increment in the lung fluid filtration rate. in parallel, flagellin markedly increased airway pressure, pulmonary arterial pressure, pulmonary vascular resistance, and filtration coefficient. in contrast to the control lungs, all the lung preparations from flagellin-treated animals developed irreversible edema within the first two hours of perfusion. in isolated blood-perfused rat lungs, flagellin enhances fluid filtration, most likely, through elevation both of pulmonary microvascular permeability and hydrostatic pressure. the present study provides further evidence that flagellin may contribute to the development of sepsis-associated ali. . whether protein c conversely affects eosinophil function has not yet been reported. we investigated the effects of protein c and activated protein c on chemotaxis of eosinophils. possible involvement of endothelial protein c receptor (epcr) in the regulation was studied by using specific epcr antibodies. for preparation of eosinophils we used macs cd+16 microbeads according to the manufactor's protocol. chemotaxis assays were performed using a 48-well boyden microchemotaxis chamber in which a 5-micrometer pore sized cellulose nitrate filter separates the upper and the lower chamber. eosinophils were pretreated by various protein c preparations with or without epcr antibodies, followed by washing and assessment of their migratory responses toward eotaxin. protein c and activated protein c exerted no significant chemotactic effect on eosinophils. however, eosinophils pretreated with protein c or activated protein c showed a sigificantly reduced response to the specific chemoattractant, eotaxin. moreover, this effects of protein c and activated protein c were inhibited using an antibody against epcr. conclusion. protein c as well as activated protein c inhibit the chemotactic effect of eotaxin on eosinophils via mechanisms involving epcr. this result indicates that protein c as well as activated protein c may decrease the number of eosinophils in tissue and thereby inhibiting inflammation and coagulation. deleterious effect of severe sepsis may be related to an oxidative stress, particularly related to peroxynitrite. selenium (se) toxicity is supposed to be related to oxidative stress through reaction with thiols. we perform a study to compare these toxicities. methods. 100 wistar rats were studied. after 8 day quarantine lipopolysaccharide (lps) or se was administered intraperitoneally in 3 ml saline water. lps and se were administered in groups of 10 rats with increasing doses from 28 to 34 mg/kg for lps, and from 0.35 to 4.5 mg/kg for se. mortality was observed at 48 hours. animals were sacrificed under halothane. blood samples were taken in 2 surviving rats of each group. nitric oxide (no) and nitrotyrosine (nit), a marker of oxidative stress especially related to peroxynitrite, were measured by elisa techniques, and plasma se concentration using atomic flame absorption. results. septic rats were rapidly sick. they rolled up into a ball. their fur was dull, and stood on end. they were asthenic and had diarrhea. at autopsy, intestinal abnormalities, and in some rats echymotics dots and hemolytic plasma were observed. rats were dehydrated. se rats developed an encephalopathy the first day and later recovered. se rats were lively, and seemed to required higher level of halothane for induction. (1)however the mechanisms responsible for this alteration remains under investigation. depressed micochondial respiration has also been found in different tissues during sepsis. (2) the objective of this work was to study diaphragmatic function in rats after peritoneal sepsis and to correlate these findings with diaphragmatic mitocondrial respiration. cecal ligation and perforation was done under general anesthesia in wistar rats (septic group, n=7) . after 48 hours the animals were monitored for arterial blood gases, systemic hemodynamia and body temperature. then, they were sacrified and the diaphragm force-frequency curves were obtained in vitro before and after fatigue. contraction time and relaxation time were also measured. mitochondrias were isolated from the diaphragm and oxygen consumption and other respiratory indexes were studied in septic animals. the results were compared to sham operated animals (control group, n=7). the septic group showed significantly lower values of aortic blood flow, arterial oxygen partial pressure, body temperature and arterial bicarbonate (p<0.05) when compared to the control group. the forces measured at the different frequencies of stimulation were lower in the septic diaphragms both before and after fatigue when compared to controls (p<0.05). mitochondrial respiration evaluated by oxygen consumption and rcr indexes was found decreased in the septic animals (p<0.05). diaphragmatic contractile failure along with hemodynamic, respiratory and metabolic dysfunctions was found in peritoneal sepsis in rats. diaphragmatic dysfunction could be explained by mitochondrial damage during sepsis. we speculate that mitochondrial injury and dysfunction could be related to oxidative stress in this animal model. introduction. protein c is activated by thrombin bound to thrombomodulin and this effect is enhanced in the presence of the endothelial protein c receptor (epcr). in vivo and in vitro studies have revealed that components of this pathway may also inhibit inflammatory responses. protein c was able to inhibit leukocyte adhesion to vascular endothelial cells and to reduce neutrophil accumulation in rat lungs [1] . protein c inhibits proinflammatory cytokine release in monocytes [2] that were shown to express epcr [3] . soluble epcr binds to proteinase-3 and cd11b/cd18 of activated neutrophils [4] , which were previously shown to synthesize thrombomodulin but not to promote thrombin-dependent protein c activation [5] . if protein c directly affects neutrophil functions has not jet been sufficiently demonstrated. we investigated the in vitro effects of protein c and activated protein c on chemotaxis of isolated human neutrophils and explored wether epcr may be involved. neutrophils were obtained from forearm venous blood by standard methods. leukocyte migration toward gradients of soluble attractants into cellulose nitrate micropore filters was measured using a 48-well microchemotaxis chamber. cells were either directly exposed to gradients of protein c or were pretreated with protein c followed by washing; then chemotaxis toward typical attractants was tested. neither protein c nor activated protein c induce chemotaxis of neutrophils. both inhibit neutrophil chemotaxis toward interleukin-8, fmlp and c5a and there is no significant difference in the effects of these two substances. a blocking antibody against the epcr is able to diminish the effects of protein c and activated protein c. conclusion. protein c as well as activated protein c is able to inhibit neutrophil chemotaxis. this indicates that an activation of protein c is not necessary for effects on neutrophils to occur or that neutrophils are able to activate protein c followed by migration. the reduction of the protein c effects by an antibody against the endothelial protein c receptor suggests that neutrophils express epcr capable to signal anti-migratory stimuli. during sepsis increased vascular permeability results in fluid extravasation and edema. lymphatics contribute in draining interstitial fluid from the abdomen to central circulation, but several factors (outflow venous pressure, pattern of mechanical ventilation) can act upon flow in the thoracic duct (1, 2). we have tested if lymph flow is affected by endotoxin infusion under different ventilatory conditions. methods. 9 anesthetized pigs (29.4±3 kg) were studied. septic damage was induced by continuous infusion of endotoxin (lipopolysaccharide e.coli, lps). abdominal lymph flow was continuously recorded by an ultrasound flow probe positioned on the thoracic duct at the diaphragm level; hemodynamics, respiratory system data, bga and intra-abdominal pressure (iap) were registered. during the first 2.5 hours of lps infusion animals were ventilated in volume controlled mode tv 10-11 ml/kg, rr 20 bpm, peep 5, fio2 0.5; during the next 2 hours animals were divided in group 1 (control, peep 5), 2 (peep 15) and 3 (spontaneous breathing, cpap peep 5). during lps infusion lymph flow significantly increased from 2.3 to 4.6 ml/min (p<0.05), cardiac output and compliance decreased from 3.4 to 2.8 l/min * and 32 to 21 ml/cmh2o * respectively, while mean pulmonary artery pressure and iap increased from 20 to 47 mmhg * and 13 to 20 cmh2o * (* p<0.05). in all the pigs a positive correlation was found between iap and lymph flow (mean pearson´s coefficient 0.59). no correlation was found between lymph flow and central venous pressure and airway pressure (mean pearson´s coefficient 0.20 and 0.13). in group 1 and 2 lymph flow changes averaged -9% and +19% (versus value before randomization). cpap increased lymph flow by 55%. lymph flow from the abdomen increases during lps infusion: role of lymphatics in draining abdominal fluid could thus be significant during sepsis (~ 280 ml/h are drained). these preliminary results suggest that spontaneous breathing could improve lymphatic flow from the abdomen. despite the following rise in intra-thoracic pressure, increase of peep is not associated with lymph flow reduction. animals in peep 15 group have however shown different patterns of response, and more data are needed to clarify this aspect. introduction. : ischemia/reperfusion or sepsis is initially responsible of an acute activation of pro-inflammatory cytokines (e.g. tumour necrosis factor (tnf-)). it is followed by a rise of anti-inflammatory cytokines (e.g. interleukin-10 (il-10)). in human umbilical vein endothelial cell (huvec) tnf-induces a mitochondrial release of reactive oxygen species (ros) in a dosedependent manner. the signalisation pathway which links tnf-at mitochondria involves ceramide pathway (1).the goal of our study is to evaluate the action of il-10 on the oxidative stress induced by tnf-in huvec and to define the mechanism of this interaction. huvec were grown on plastic cover slides. at confluence they were placed in a perfusion chamber under a microscope equipped with a digital camera connected to acquisition software. cells were perfused with krebs solution containing two fluorescent probes: dichlorodihydrofluorescein diacetate (dcfh) to study the release of reactive oxygen species (ros) and propidium iodide (pi) to study cell mortality. three cell groups were studied: a reference group, a tnf-group where, after one hour stabilisation, tnf-was added (1 ng/ml) in perfusion medium during one hour, · a group tnf-+ il-10 where il-10 was added to perfusion medium 30 minutes before tnf-. variations in fluorescence were recorded each 10 minutes for dcfh and each one hour for pi. for a non lethal concentration (pi remaining unchanged), il-10 reduces significantly the ros production induced by tnf-(anova for repeated measures). interleukin-10 has an inhibitory effect on the release of ros induced by tnfin huvec. this effect could be the result of an interaction with acid sphingomyelinase. (1) am. j. cell. molecular biol. 24: 762-8, 2001 . the immunosuppresive drug cyclosporine a (csa) is an inhibitor of mitochondrial permeability transition (mpt) which could afford protection against cell death [1] .to test whether csa protects against endotoxin-induced myocardial apoptosis [2], we produced 123i-annexin v [3], a marker of apoptotic cells, and measured its myocardial uptake during endotoxaemia in csa-treated rats. the specificity of the signal has been previously verified with caspase inhibitors and 123i-human serum albumin. methods. 1) 123i-annexin v was produced with a radiochemical purity higher than 97% as confirmed by hplc. 2) young male sprague-dawley rats were either given iv : saline (0.5ml) : control group, n=10, or lipopolysaccharide (lps) from e coli (30mg/kg) ± csa (10mg/kg): lps group, n=10 and lps+csa group, n=8. 6h later, all animals were given 123i-annexin v (18mbq, 10mg protein). after 12h, hearts were harvested and divided into apex, septum, right and left ventricle (rv, lv) for determination of 123i-annexin v myocardial uptake with a lkb gamma counter. results were expressed as a mean percentage ± sd of the injected dose per gram of tissue (%id/g). statistical analysis was performed by mann-withney test; a p value <0.05 was considered as significant (*). 123i-annexin v myocardial uptake is significantly increased in the lps group compared to control group; there is no significant difference between the septic groups . control lps lps+csa mean + -sd 0.05 + -0.01 0.18 + -0.06 * 0.19 + -0.05 ns mortality 0% 25% 0% 123i-annexin v myocardial uptake conclusion. our results confirm that endotoxaemia is associated with significant myocardial apoptosis but fail to demonstrate that csa can reduce the cell death signal detected by 123i-annexin v . in spite of its action on mpt and its myocardial dysfunction reducing effect in septic rats [4] , csa provides no myocardial protection in this model . a reducing effect of csa on endotoxin-induced mortality is not excluded but remains to be demonstrated. further investigations are needed to clarify the effect of csa on the inflammatory responses due to endotoxaemia. sepsis induced alterations in hemostasis with dysbalances in fibrinolysis may lead to capillary obstruction due to fibrin deposition. the aim was therefore to investigate regional net fluxes of the fibrinolytic enzyme tissue-type plasminogen activator, tpa, and its main inhibitor plasminogen activator inhibitor type-1, pai-1, in response to endotoxemia. methods. anesthetized pigs (n=8) were instrumented for registration of cardiac output (co, thermodilution) and portal (qpv), hepatic (qha) and renal (qra) blood flows (ultrasound flowmetry, transonic). blood samples were collected from the aorta and pulmonary artery as well as the portal, hepatic and renal veins. after baseline registrations, all animals were subjected to an e. coli endotoxin infusion for 90 min, followed by a volume/norepinephrine resuscitation for 210 min targeting baseline co levels. plasma concentrations of both total and active tpa and pai-1 were determined as described [1, 2] and net organ fluxes (ng/min) were calculated based on in-/outflowing plasma concentrations and local plasma flow [1]. results. endotoxemia induced a low co state and a decrease in qpv. total liver blood flow was preserved due to a concomitant increase in qha. during resuscitation co and qpv were restored to baseline values. systemic plasma levels of total tpa increased over time during endotoxemia, peaking at 90 min, whereupon a decline occurred. however, plasma levels of total tpa had not returned to baseline values at the end of the registration period (300 min). changes in systemic levels of active tpa mirrored changes in total tpa. a marked (8-fold) increase in mesenteric net release of total tpa was observed. this response was paralleled by a pronounced increase in hepatic uptake of tpa. pai-1 described a different response to endotoxemia. by the end of the experiment plasma levels of both active and total pai-1 increased. in contrast, no significant net fluxes of pai-1 were observed across any of the investigated vascular beds except for the hepatic vascular bed, where a net release of both total and active pai-1 occurred at approximately 150 min. hepatic pai-1 release rates then increased progressively. conclusion. endotoxemia induced a marked increase in mesenteric release of tpa which however was not entirely responsible for the increase in systemic plasma level of tpa. the results indicate that this profribrinolytic response at later stages are counteracted by increased plasma levels of pai-1 and this increase is mainly derived from the hepatic vascular bed. thus, patients with altered regional endothelial functions or liver capacity prior to a septic challenge can be expected to demonstrate varying susceptibility to thrombotic events. antithrombin has been shown to reduce mesenteric venular leukocyte interactions and intestine injury in a leukocyte-dependent model of endotoxemia (1). however, endothelial damage during early endotoxemia has been shown to be leukocyte-independent (2). the role of antithrombin in this setting is still unknown. therefore, it was the aim of the study to investigate the effects of antithrombin on leukocyte-independent endothelial damage. in male wistar rats, microvascular permeability (mp) and leukocyte-endothelialinteraction (leukocyte rolling, lr) were determined in mesenteric postcapillary venules using intravital microscopy at baseline, 60 and 120 min after start of a continuous infusion of endotoxin (etx; 2mg/kg/hr, e.coli o26:b6) (group a, n=8). therefore animals were laparotomized and the mesentery was exposed beneath an in-vivo videomicroscope. mp was measured using fluorescein isothiocyanate (fitc) labelled albumin. leukocyte-endothelial interaction was blocked in all groups by fucoidin (25 mg/kg b.w.), a l-selectin-binding carbohydrate, 10 min before laparotomy. animals in group b (n=8) received antithrombin (kybernin®, aventis-behring, germany; 500 ie/kg b.w.) prior to baseline measurement and additionally to the procedure described above. animals in group c (n=8) received equivalent volumes of nacl 0.9 % instead of antithrombin and endotoxin. statistical analysis was performed using two-way repeated measures anova followed by the scheffé test. a p-value <0.05 was considered significant. in groups a-c, fucoidin prevented lr during the entire experiment. however, in all groups mp increased significantly, starting at 60 min. animals in group a were characterized by a stronger increase in mp and showed significantly higher values in mp in comparison to groups b and c at 120 min. there were no significant differences in mp between groups b and c. leukocyte-independent endothelial damage during early endotoxemia is attenuated by antithrombin. endothelial damage during early endotoxemia has been shown to be leukocyte-independent (1). paf (platelet-activating factor)-and serotonin-receptor antagonism has been shown to reduce leukocyte-independent macromolecular leakage significantly (2, 3). nevertheless, the exact mechanisms involved in leukocyte-independent endothelial dysfunction are unknown. therefore, it was the aim of the study to investigate the effects of nitric oxide (no) on leukocyte-independent endothelial damage during endotoxemia methods. in male wistar rats, microvascular permeability (mp) and leukocyte rolling (lr) were determined in mesenteric postcapillary venules using intravital microscopy at baseline, 60 and 120 min after start of the experiment. in all groups, leukocyte-endothelial interaction was blocked by fucoidin. rats were randomized into 7 groups, 11 animals each. the experiments were divided into two parts. part i (no-inhibitor): in group a, the mesentery was superfused with a l-name superfusion (100 mmol/l) combined with a continuous infusion of endotoxin (etx; 2mg/kg/hr) after baseline measurement. group b received a l-name superfusion of the mesentery combined with a continuous infusion of saline 0.9 %. groups c and d were treated like groups a and b but without l-name. part ii (no-donator): group x received sin-1 (initial bolus of 1 mg/kg b.w. followed by 0.5 mg/kg b.w. after 60 min-measurement) followed by a continuous infusion of endotoxin (etx; 2mg/kg/hr). group y was treated similar to group c and group z was treated similar to group d. statistical analysis was performed using two-way repeated measures anova followed by the scheffé test. a p-value < 0.05 was considered significant. fucoidin prevented leukocyte-endothelial-interaction in all groups. part i: pe increased in all groups, being significant in group d at 120 min (p<0.05 vs. baseline) and being significant in groups a-c starting at 60 min. animals in group d were characterized by a slighter increase in mp and showed significantly lower values in mp in comparison to groups a and b at 60 min, and to groups a-c at 120 min. there were no significant differences in mp between groups a-c at 120 min. part ii: pe increased in all groups being significant in group z at 120 min (p<0.05 vs. baseline) and being significant in groups x and y starting at 60 min. animals in group y were characterized by a stronger increase in mp and showed significantly higher values in mp in comparison to groups x and z at 120 min. there were no significant differences in mp between groups x and z. leukocyte-independent endothelial damage during early endotoxemia is a nitricoxide mediated event. overproduction of nitric oxide (no) is thought to be a principal cause of the hypotension of septic shock. two nitric oxide synthase (nos) enzymes have been described in blood vessels: endothelial nos (enos) and inducible nos (inos). constitutive activity of enos in the endothelium is a major determinant of blood vessel tone in health; however, in experimental sepsis it appears endothelial enos expression is reduced while smooth muscle inos expression is increased (1). in contrast, another model of human sepsis found an increase in enos but not inos in the vessel wall (2). to resolve this discrepancy, we studied enos and inos protein concentrations in arterial smooth muscle (asm) from patients with clinical sepsis. asm was isolated from mesenteric vessels from patients undergoing bowel resection for perforated viscus (who in the perioperative period met the accp/sccm criteria for septic shock), and from controls with bowel cancer. after mechanical removal of endothelium and adventitia, the tissue was homogenised in protease inhibitor and frozen until sufficient samples had been accumulated. western blotting was performed under reducing conditions, with membranes incubated in 1:2000 (inos) or 1:500 (enos) primary antibody followed by 1:2000 peroxidase labelled secondary antibody. protein bands were quantified by computer analysis of the chemiluminescence detection film, then normalised to the protein concentration of the sample prior to dilution. . enos protein was increased in arterial smooth muscle from patients with septic shock (control 39.3 14.7 units/mg, septic 96.5 27.1 units/mg; n=13 controls and 11 septics; p = 0.04, student's t test). in contrast, there was no increase in concentration of inos; indeed inos protein was only detectable in asm from 2 control and 3 septic patients. we suggest that overexpression of enos, rather than inos, in the arterial smooth muscle of patients with septic shock may be responsible for the hypotension observed in these patients. introduction. data published in the literature concerning the effect of sepsis on intestinal motility found a reduction as well as a stimulation of intestinal motility . the settings used are mostly in vivo settings, and therefore not usable to investigate intestinal motility independent from circulatory changes. the aim of our study was to evaluate the direct effect of endotoxinemia on guinea-pig small bowel motility in vitro, independent from circulatory changes, and in a second step to evaluate the effect of vasoactive drugs on motility of these septic animals. two groups of guinea-pigs received 1 mg/kg e. coli lps intraperitoneally 4 or 20 hours before the experiments started. in the following hours the animals developed severe symptoms of sepsis. a control group did not receive lps before the experiments started. the small bowel of sacrificed guinea-pigs was excised, cleaned and kept in tyrode's solution. after a resting period segments of 8 cm length were set up in parallel organ bathes containing oxygenated tyrode's solution. peristaltic contractions were elicited by perfusion of the segments with tyrode's solution at a rate of 0.5 ml/min, against an aboral resistance of 400 pascal. the intraluminal pressure increased gradually until it reached a pressure threshold (pt) which triggered peristaltic contractions. these contractions were recorded via a pressure transducer at the aboral end of the segments. increasing concentrations of epinephrine, norepinephrine, dopamine, dobutamine, clonidine and dexmedetomidine were cumulatively added to the organ bath at 15 min intervals. each drug was tested on 8 different segments. statistics was performed using ncss for windows, one-way and two-way anova for repeated measures were used, p values <0.05 were considered statistically significant. in the control group all tested vasoactive drugs had a dose-and substance-dependent inhibitory effect on peristalsis. higher concentrations of all tested substances led to a complete block of peristalsis. 4 hours after lps application a pronounced reduction of the inhibitory effects of clonidine, epinephrine, norepinephrine and dopamine were found. the reduced inhibitory effect of dexmedetomidine was not significant. 20 hours after lps application the inhibitory effect was reduced again, but for most substances this reduction was not statistically significant. dobutamine was the only tested substance with a more pronounced effect after 4 hours than after 20 hours. endotoxinemia per se did not affect small bowel motility in vitro. a possible explanation for the controversy to in vivo data demonstrating an inhibitory effect on peristalsis might be that intestinal ischemia is a common event during sepsis, and ischemia in turn might cause paralysis. a described reduced sensitivity of alpha-adrenoceptors during sepsis, or a central effect of lps additionally inhibiting peristalsis (1), might also be responsible for our findings. high cytokine levels in patients admitted to the emergency department are associated with an increased incidence of sepsis/septic shock. patients with cardiogenic shock (cs) who often develop sepsis during icu-stay,have not been particularly studied. we studied whether plasma levels of cytokines are better predictors of sepsis/septic shock than routinely determined laboratory parameters. il-1,il-6 and il-10 plasma levels were determined in 51 pts with cs(cardiac index <2.2 l/min/m²,pcwp >15, mean arterial pressure <60mmhg or need for vasopressor therapy and signs of organ hypoperfusion) on admission to the icu (median 16hrs after shock onset). 36 patients who were not surgically treated during icu stay were eligible for the study and evaluated for development of sepsis or septic shock within 1 week after onset of cs. c-reactive protein (crp) levels and white blood cell (wbc)-counts were routinely evaluated once daily in all patients until discharge. data are given as median and interquartile range. all pts with cs were free of demonstrable infection at time of blood sampling. nevertheless 60% had a crp-level >5mg/dl at time of enrollment.11 pts (31%) developed septic shock within 1 week after onset of cs. pneumonia (54%, n=6) and catheter related infections (27%, n=3) were the leading causes of sepsis. sepsis after cs was not associated with a higher mortality rate (82% vs. 80%, p=ns) and sirs that was encountered in 69% of cs pts at the time of blood sampling did not predispose for development of sepsis (64 vs. 72%, p=ns).crp levels,and wbc-counts as well as il-1, il-6 and il-10 plasma levels on admission to the icu did not differ significantly between cs-pts who developed sepsis and cs-patients without sepsis ( in pts who survived for more than 24hrs (n=28) the absolute crp levels 24hrs after admission (crp24hrs) and the increase in crp levels over 24hrs following icu admission (dcrp) were significantly higher in pts who developed sepsis as compared to pts without sepsis. (crp24hrs: 23.2mg/dl [11-35.6] vs. 9.4 [5.9-13.1], p=0.002; dcrp: 9.7mg/dl [7.5-11.8] vs. 4.1[0.5-5.6], p=0.011). a dcrp >6.5mg/dl in 24hrs was more sensitive than an absolute crp level >20 mg/dl 24hrs after icu-admission for predicting sepsis (82 vs. 73%), but both parameters had equal specificity (88%). conclusion. although many pts with cs exhibit elevated crp levels the increase in crp over 24hrs (dcrp24hrs)is a valuable parameter to identify pts at risk for sepsis. single-point determination of cytokines on admission to the icu is not superior to follow-up determinations of crp for predicting sepsis. mitochondrial dysfunction may be implicated in sepsis-induced multi-organ failure. glycolytically-generated atp may thus be an important alternative energy source if aerobic respiration is compromised. little is known about glycolysis during sepsis, though both up-and down-regulation are reported 1,2 . we therefore examined changes in glycolytic activity in a longterm sepsis model. an instrumented, fluid-resuscitated, faecal peritonitis rat model was used. this has a 72-hour mortality rate of approx. 50%. septic (n=57) and sham (n=35) rats were sacrificed at various time points (0, 24, 48, 72h) and liver samples harvested and assayed for maximal activity of the rate-limiting glycolytic enzymes, hexokinase (hk), phosphofructokinase (pfk) pyruvate kinase (pk). we demonstrate an initial rise (albeit non-significant) then significant downregulation in two rate-limiting glycolytic enzymes during sepsis. the lack of difference at 72 h may reflect prior demise of the severely ill animals. whether the degree of glycolytic down-regulation is related to subsequent death requires further study. we presume the interesting finding of upregulation seen in the sham animals to be a response to surgery and/or fluid loading. recent studies have shown that low-dose vasopressin infusion or terlipressin bolus (tp, its long acting analogue; o'brien, 2002) restores blood pressure and reduces norepinephrine (ne) requirements in septic shock. however they have no effect upon blood pressure in non-septic patients. exact mechanisms underlying this hyperreactive effect in sepsis patients remain unknown. we chose to investigate this using our established in vivo and in vitro models of endotoxic shock in rats. in vivo -spontaneously breathing anaesthetised male wistar rats was given either saline (sham) or endotoxin (lps) (klebsiella 40mg kg -1 ) over 30 mins and then fluid resuscitated with colloid 25mls kg -1 hr -1 for 210 mins. at 120 mins either a bolus of tp (1.5mg kg -1 ) or a bolus and infusion of ne (0.5mg kg-1 and 1mg kg hr -1 ) was administered. measurement of flow and pressure (mean arterial pressure -map) were made from appropriately sited probes and transducers. in vitro -rings of rat mesenteric artery (rma) were harvested, cleaned and incubated for 20 h with or without 1mg ml -1 lps (s. typhosa). they were then mounted in organ baths for measurement of isometric tension. cumulative concentration-response curves to phenylephrine (pe: 10 -9 to 10 -5 m) or vasopressin (vp: 10 -12 to 10 -6 m) were then constructed. statistical analysis was by anova. results. in vivo -while ne had a significantly greater effect upon map in shams compared with lps rats (p=0.042), tp caused a greater increase in lps animals than shams. a bolus of tp lasted approximately 75 mins. in vitro -lps significantly depressed contractile responses to pe compared to control tissues (max contraction controls -1.35±0.14g, lps -0.48±0.11g, p<0.001, anova). however there was virtually no contractile response to vp even in control tissues after 20 h incubation. the cytokine cascade activated in response to injury consists of a complex biochemical network with diverse effects on the injured host. leukocyte activation after trauma is essential for inflammation. it is a multistep process in which chemokine -interleukin (il)-8 has pivotal role. in two-hit hypothesis, sepsis represent a second insult to a previously injured and primed host, converting a low-grade or regulated host response into an accelerated or dysregulated host response, triggering new or progressive organ dysfunction (1). aim of this study was to assess pro-inflammatory response to trauma with or without sepsis as a second insult. twenty five patients with severe trauma (explosive and sclopetarious) who developed sepsis and 10 patients with same kind of severe trauma without sepsis were enrolled in this study. in the trauma+sepsis group 21 patients developed multiple organ dysfunction syndrome (mods) and 14 died. in trauma group 4 developed mods and 2 died. blood was drown on the first, third and fifth day of trauma. concentrations of il-8, il-12, tumor necrosis factor (tnf)alpha and interferon (ifn)-gamma were determined in plasma using elisa assays. when compared trauma+sepsis group with trauma group we found statistically highly significant difference (p<0.01) in il-8 and ifn-gamma and statistically significant difference (p<0.05) in tnf-alpha concentrations; mean values of il-8 were 230-fold higher, ifn-gamma 360-fold higher and tnf-alpha 17-fold higher in patients with trauma with sepsis. il-12 was not statistically different (p>0.05) between two groups. when compared mods group with group without mods, we found statistically highly significant difference (p<0.01) in il-8 and tnfalpha concentrations; mean values of il-8 were 60-fold higher and tnf-alpha 43.5-fold higher in patients with mods; il-12 and ifn-gamma were not statistically different (p>0.05) between two groups. when compared non-survivors with survivors, we found statistically highly significant difference (p<0.01) in il-8 and tnf-alpha and statistically significant difference (p<0.05) in il-12 concentrations; mean values of il-8 were 2.3-fold higher in non-survivors, mean values of tnfalpha were 2.2-fold higher in survivors, il-12 was also higher in survivors. ifn-gamma was not statistically different (p>0.05) between two groups. there is augmented pro-inflammatory response after trauma with secondary sepsis. high concentrations of il-8 and tnf-alpha indicated higher severity (mods). but, fatal outcome was predicted with high concentrations of il-8 only; survivors had higher concentrations of tnf-alpha and il-12. therefore, pro-inflammatory response was partly beneficial and partly detrimental to the host. in patients with shock hypoxia is considered to be the most important cause of organ failure and death. the goal of treatment therefore is to restore tissue oxygen delivery (tdo2). due to impaired oxygen extraction in distributive (septic shock) the relation between tdo2 and tissue oxygenation is less conclusive. direct measurement of tissue oxygen pressure (pto2) could be of great importance in gaining a better insight in tissue oxygenation in these patients. previously published data concerning pto2 in patients with sepsis/septic shock are contradictory (1,2). furthermore the techniques used were not easily applicable at the bedside. in a prospective observational study we performed bedside pto2 measurements in 8 patients with sepsis/septic shock to gain insight in pto2 values and their dynamic changes related to the course of the illness, as well as investigating the practical applicability of tissue oxygen measurement in the icu setting. pto2 was measured continuously during the course of the illness using polarographic clark-type o2 electrodes (licox catheter measurement system, gms), which were placed subcutaneous in the upper arm. disease progression over time was expressed as the daily calculated sequential organ failure assessment (sofa) score. results. five men and 3 women with septic shock n=6 or sepsis n=2 were included. the median (range) age was 39 years (21-85), median apache-score on the day of admission was 15 (9-29), median duration of pto2 measurement per patient was 5,0 days (3-7). in none of the patients technical problems were encountered during the pto2 measurements. the first day of measurement the median pto2 of the eight patients was 49 (18-92) mmhg. in the six surviving patients the sofa score decreased over time and this was associated with a concomitant decrease in pto2 to a median of 30 (16-69) mm hg. in the 2 nonsurvivors an increasing sofa score was associated with an increase in the mean pto2 to 57 mmhg on the day of death. in seven patients linear regression analysis showed a positive correlation between the daily sofa scores and the daily mean pto2: r= 0.71, 0.75, 0.88, 0.93, 0.85, 0.87, 0.61. in one patient no correlation was found. conclusion. bedside pto2 measurements in the icu using the licox measurement system are easily performed. pto2 in septic patients is variable but changes with the clinical course reflected by the sofa score: clinical improvement was associated with a decrease in pto2 while deterioration was associated with an increase of pto2. these findings suggest that in patients with septic shock decreased oxygen utilisation may play a more important role than tissue hypoxia as such. to precise the diagnostic value of macrophage migration inhibitory factor (mif) as a marker of severity in patients with sepsis and to determine relations between mif and interleukin 6 (il-6), we conduced a prospective, observational, cohort study, in two general intensive care units. we analyzed 20 patients with septic shock, 17 patients with sepsis, and 10 healthy volunteers. the median mif serum level was significantly higher in septic shock patients (2.01 ng/ml, range 0.77-12.0) than in sepsis patients (1.10 ng/ml, range 0.31-5.11) or in healthy volunteers (0.45 ng/ml, range 0.28-1.54). there was a direct correlation between mif and il-6 concentrations (r=0.693, p<0.01). the area under the curve (auc) of the receiver-operating characteristic (roc) for prediction of septic shock was 0.81 (p<0.01) for mif and 0.85 (p<0.01) for il-6. the auc under the roc curve for prediction mortality was 0.714 (p<0.05) for mif and 0.707 (p<0.05) for il-6. in this trial we found significant elevated serum levels of mif in patients with septic shock and sepsis. moreover, mif levels were discriminative for septic shock and mortality, and had a direct correlation with levels of il-6 with a similar diagnostic accuracy. in conclusion, mif appear to be a promissory marker of severity in sepsis. high density lipoprotein (hdl) modulates the inflammatory response to injury and infection via several pathways. hdl also directly binds and neutralises lps. administration of reconstituted hdl reduces cytokine release and attenuates shock in experimental endotoxaemia (1). the hdl associated enzymes paraoxonase (pon) and lecithin cholesterol acyl transferase (lcat), destroy oxidised lipids that induce inflammatory changes in vascular endothelium (2). incorporation of serum amyloid a (saa), an acute phase protein, into the hdl particle during the inflammatory response, may displace these protective enzymes producing a particle with proinflammatory properties (3). alterations in hdl composition may, therefore, be implicated in dysregulation of the inflammatory response and could influence outcome from septic shock. methods. patients with septic shock, not given tpn or propofol, were recruited. apache ii scores and icu mortality were recorded. plasma and serum samples were taken within 48 hours of the onset of shock. hdl cholesterol was measured by microenzymatic colorimetric assay. apolipoprotein ai (apo ai) was quantified by liquid phase radioimmunoassay. pon activity was determined by measuring the rate of paraoxon hydrolysis and described as percent of a control serum pool. lcat activity was quantified by measuring the esterification rate of 14 c labelled cholesterol. saa was measured by elisa. results were compared with those of a pool of healthy volunteers and between survivors and nonsurvivors. (mann whitney u test). results. 20 patients were recruited. there were 13 survivors (s) and 7 nonsurvivors (ns). pon activity was significantly higher in s than ns: 47.6 (11.5-135.6) vs. 22.6 (5.7-39.43), p<0.02. saa concentration was significantly higher in s than ns: 1915 (90.9-14300) severe trauma and sepsis are the major sources of morbidity and mortality despite the rapid development of intensive therapy. studies have indicated that there are marked alterations in immune response in patients exposed to major trauma or prolonged surgical procedures, including altered pro-and anti-inflammatory mediator/cytokine release (1). traumatic injury results in profound immunosuppression which predisposes the patients to sepsis and/or multiple organ dysfunction syndrome (mods). aim of this study was to assess the prognostic value of anti-inflammatory cytokines: interleukin (il)-1 receptor antagonist (il-1ra) , il-4, il-10 and transforming growth factor (tgf)-beta 1 regarding severity and outcome in patients with trauma and sepsis, trauma only and sepsis only. twenty five patients with severe trauma (explosive and sclopetarious) who developed sepsis, 10 patients with same kind of severe trauma without sepsis and 5 patients with severe sepsis were enrolled in this study. twenty nine patients developed mods (of all 40 patients), 19 died. blood was drown on the first, third and fifth day of trauma or sepsis. concentrations of il-1ra, il-4, il-10 and tgf-beta 1 were determined in plasma using elisa assays. when compared mods group (regardless of initiating insult -trauma or sepsis) with group without mods, we found statistically highly significant difference (p<0.01) in il-1ra and il-10 concentrations; mean values of il-1ra were 6-fold higher and il-10 70-fold higher in patients with mods; il-4 and tgf-beta 1 were not statistically different (p>0.05) between two groups. when compared non-survivors with survivors, we found statistically highly significant difference (p<0.01) in il-1ra and il-10 concentrations; mean values of il-1ra were 2.7-fold higher and il-10 1.4-fold higher in non-survivors; il-4 and tgf-beta 1 were not statistically different (p>0.05) between two groups. when compared trauma+sepsis group with trauma group, we found statistically highly significant difference (p<0.01) in il-1ra and il-10 concentrations, they were higher in trauma+sepsis group (il-1ra 3.7-fold, il-10 42-fold). il-4 and tgf-beta 1 were not statistically different (p>0.05) between two groups. when compared trauma+sepsis group with sepsis group and trauma group with sepsis group, we found no statistically significant difference in either one of 4 anti-inflammatory cytokines. our study shows that il-1ra and il-10 are excellent predictors of severity and outcome of critical illness; higher concentrations were found in group with more severe clinical status (mods) and in non-survivors. il-4 and tgf-beta 1 had no significance as predictors of severity and outcome what so ever. fifty-eight patients admitted to two medical intensive care units for reasons other than acute coronary syndrome were consecutively included and analyzed according to their troponin status. thirty-day mortality, left ventricular ejection fraction, the presence or absence of underlying coronary artery disease, and a panel of inflammatory cytokines were compared between troponin-positive and troponin-negative patients. thirty-two of 58 critically ill patients (55%) without evidence for an acute coronary syndrome were troponin-positive. positive troponin levels were associated with higher mortality (22.4% vs. 5.2%, p < 0.018) and lower left ventricular ejection fraction (p = 0.0006). troponinpositive patients had significantly higher median levels of tumor necrosis factor a, its soluble receptor and interleukin-6. a subgroup of ten aplastic patients was troponin-negative at study entrance. three became troponin-positive during leukocyte recovery and subsequently died, whereas all the others stayed troponin-negative and survived. conclusion. elevated troponin is a mortality risk factor for medical intensive care patients admitted for reasons other than acute coronary syndromes. it is associated with decreased left ventricular function, and this may be mediated by tumor necrosis factor a and mediators produced by neutrophilic granulocytes. it is very interesting to notice the high correlation among protein c and atiii activity levels and sofa scores (p< .01) and the dramatic decrease of the protein c system is already firmly present 48 hours before negative outcome (not survivors). we also register the significant alterations of c1 inhibitor specially in the group (ns) patients with severe candidemia and this marker assumes a significant role with an interesting clinical future . tumour necrosis factor-a (tnf) is an important pro-inflammatory mediator and high levels of this cytokine have been associated with a poor outcome from sepsis. recently, genetic polymorphisms of the tnf locus and its promoter region have been associated with the incidence and outcome of severe sepsis 1;2 , although the results have been conflicting 3 . we chose to investigate the association between a known functional single nucleotide polymorphism (snp) in the tnf gene promoter (-308 g/a, guanine to adenine substitution) and outcome in severe sepsis and septic shock. caucasian adult patients with a diagnosis of severe sepsis or septic shock on 6 icus in the uk and from an icu in sydney, australia were recruited. whole blood was collected in edta, dna extracted and amplified by pcr using specific primers and digested with the restriction endonuclease nco1. this enzyme cuts the wild type (allele g) but this cutting site is abolished by the polymorphism (allele a). the restriction fragments were then size separated, visualised and scored on agarose gels. fisher's exact test was used for statistical analysis. shedding of membrane bound tumour necrosis factor receptors to produce soluble molecules (stnfrsf1a and 1b) is an important inflammatory control mechanism 1 . we and others have previously demonstrated that increased levels of stnfrsf1a and stnfrsf1b are associated with decreased survival from sepsis. furthermore, there appears to be an association between polymorphisms of the tnfrsf1b locus and plasma levels of stnfrsf1b 2 . we have therefore investigated whether polymorphisms of the tnfrsf1b gene and its promoter region might influence outcome from severe sepsis and septic shock. 170 caucasian adult patients with a diagnosis of severe sepsis or septic shock from 6 icus in the uk and from an icu in sydney, australia were recruited. we analysed 3 polymorphisms of the tnfrsf1b gene. a single nucleotide polymorphism in exon 6 (snp 196 t/g) was studied by pcr-rflp, a microsatellite in intron 4 (ms4) using an abi373a sequencer and a 15 base pair insertion/deletion in the promoter region (indel) by polyacrylamide gel electrophoresis. analyses of associations between genotype and allele frequencies and outcome were by fisher's exact test. . icu mortality was 28%. overall genotype and allele frequencies for each of the polymorphisms were similar to published population frequencies. there were no statistically significant differences in allele frequencies in any of the three polymorphisms between survivors and non-survivors (snp196 p=0.19, ms4 p=0.52, indel, p=0.42). the mortality was lower in patients homozygous for the 15 base pair repeat in the microsatellite polymorphism in intron 4 (the genotype associated with high levels of stnfrsf1b) (mortality 20% v 32.1%) and was higher in those with the snp196 (t/g or g/g) (mortality 34.8% v 23.2%). these differences, however, did not reach conventional levels of significance (p=0.14 and 0.12 respectively). larger studies will be required to confirm or refute associations between tnfrsf1b gene polymorphisms, particularly ms4 15 homozygosity, and outcome from sepsis. when associated with end organ dysfunction, sirs is a major cause of morbidity and mortality in the intensive care unit (icu) population (1). lps concentrations in the gastro-intestinal tracts of these patients are elevated as a consequence of bacterial overgrowth. lps processing in the mesenteric circulation may influence the systemic inflammatory response (2). tlr4 is an integral part of the lps receptor complex. a tlr4 polymorphism (asp299gly) is associated with hypo-responsiveness to lps in human bronchial epithelial cells. we examined the association of this polymorphism with clinical outcome in icu patients with severe sirs. methods. adult icu patients with evidence of severe sirs were studied. patient demographics, apache ii data, length of stay and outcome data were collected. genotype was determined using pcr amplification. statistical analysis was performed using spss 11.0. results. 67 patients have been genotyped of whom 2 are still in icu. of the remaining patients, 20/65 (31%) died in icu and 9 died in hospital after discharge from icu, giving an overall hospital mortality rate of 29/65 (45%). mean (sd) apache ii score was 21 (5).the tlr4 genotype frequencies were asp/asp 88.1% (59/67), asp/gly 10.4% (7/67) and gly/gly 1.5% (1/67). the allele frequencies were asp 93% and gly 7%, similar to previously reported frequencies in caucasians. preliminary analysis revealed no significant differences between apache ii scores in patients with the asp/asp genotype (mean 20.7, sd 5.6) and those with asp/gly or gly/gly genotypes (mean 19.0, sd 2.8) (p=0.40, student's t-test). 5/29 (17%) of patients who died during the hospital episode carried the gly polymorphism, compared to 3/36 (5%) of those who survived the hospital episode (p=0.45, fisher's exact test, or 2.3, 95% ci 0.5-10.5). conclusion. no associations with severity of illness on admission to icu, icu length of stay or hospital outcome were detected with the present sample size. recruitment is ongoing, to attain sufficient power. we aim to study genes coding for components of the lps receptor complex, which are biologically relevant to innate immunity and the development of sirs. detailed, prospective study of the role of polymorphisms in innate immunity has the potential to improve our understanding of the pathogenesis of sirs, and to influence risk stratification and management of this severe complication of life-threatening infection. the present study was designed to evaluate the effect of low dose albumin infusion vs. control on the local inflammatory response following abdominal surgery. albumin loss during surgery is a well described phenomenon. in previous experiments a loss of plasma proteins, resp. albumin was observed during abdominal surgery. intravital microscopy for five hours was used to evaluate the effect of low dose albumin on the mesenteric microcirculation. urethan-anesthetized sprague-dawley rats underwent median laparatomy and placement of a doppler flow probe around the abdominal aorta. an ileal loop was prepared for eventration onto a microscopic stage using a plastic foil technique and the mesentery was immersed with krebs-henseleit buffer(5%co2 in n2). low dose albumin (0.001 g/(kg bw*h)) was given vs. control (nacl 0.9%) during the experiment. heart rate, map, aortic blood flow were registered on a beat-to-beat basis. abg's were drawn hourly for analysis of metabolic(be), respiratory (po2, pco2) and hct values. rolling and adherent leukocytes significantly increased in the control group until the end of the experiment, whereas they constantly remained on a low level in the albumin group. velocity and shear rate in the mesenteric microcirculation were significantly higher in the albumin group which was supported by increased abdominal flow and stroke volume vs. control. low dose albumin infusion significantly reduces the inflammatory response on the mesenteric microcirculation following abdominal surgery. beneficial effects on systemic hemodynamics, mesenteric microcirculation and attenuation of leukocyte rolling and adhesion in mesenteric venules could only be observed in the albumin group, whereas the inflammation progressed in the control group. iasonidou c. 1 , pertsas e. 1 , koletsos k. 1 , kapravelos n. 1 , tsagalof s. 1 , riggos d. 1 1 icu, g.papanikolaou, thessaloniki, greece optimizing patient's hemodynamics in the icu can be challenging.the pa catheter has been used to determine preload, afterload and myocardial performance. however,insertion of a catheter is not a risk-free procedure and the values obtained can in some circumstances be misleading.the use of tee in icu has been increasing.previous studies have examined the correlation between the pulmonary vein (pv) velocities and mitral valve (mv) velocities and pcwp. the purpose of our study was to evaluate the relationship between these variables during different loading conditions as assessed by tee in icu patients. (11) patients,with a mean age of 65± 10 years, requiring mechanical ventilation were prospectively studied. in all patients a pa catheter was inserted and baseline measurements were obtained.the pv velocities and mv velocities were evaluated during three different loading conditions:1) in a control situation 2) in a state of decreased preload by intravenous administration of nitroglycerin 3)in a state of increased preload by administration of fluids.in all patients we used the following indices from the pv velocity : s (systolic),d (diastolic), decelaration time(dt) of d wave, apv (atrial reversal) and from mv velocity: e,a wave and deceleration time of e wave. the decrease in preload resulted in a trend toward a lower amplidute of d wave peak velocity as compared with the control state and a significant prolongation of the deceleration time (p<0,05).there was a decrease in height of the systolic (s) wave (p<0,001)and the apv (p<0,05). the mv curve demonstrated a significant decrease in e velocity (p<0,05) and prolongation of deceleration time (p<0,05).the increase in preload resulted in a significant increase in systolic and diastolic wave in pv (p<0.01) with a shortening of the dt of d wave.the apv became significantly higher (p<0,05).the mv curves demonstrated a significant increase in e wave (p<0,05) with a decrease in dt.there was a good correlation between d wave and pcwp (r:0,6),apv wave and pcwp (r:0,7) and e wave and pcwp (r:0,6).a direct correlation was present between changes in e and d waves (r:0,68) and changes in dt of e and dt of d velocities (r:0.78). this study provides evidence that tee gives information additive to the pa catheter in the assessment of preload in an icu population. examination of pv velocities and mv velocities and their changes during different loading conditions provide additional information regarding diastolic function. this may prove useful in minimizing the use of invasive methods for hemodynamic monitoring in icu patients. further investigation is required to correlate these doppler measures with the invasive hemodynamic measurements. methods. 26 patients with spe (19 women and 7 men), with a mean age of 59 years (sd of 17; range: 21 to 83 years), were studied prospectively. coloured doppler-echocardiography was performed in all cases at admission, confirming that diagnosis by perfusion gammagraphy and / or helycoidal ct scan. emboli were observed in six patients (23%): 3 in right atrial chamber, 2 in pulmonary artery and 1 in output tract of right ventricle. in 17 patients (65%) right ventricular dilatation with a mean value of 38.8 mm (sd 5), and tricuspid insufficiency in 20 (74%) with mean estimated systolic pulmonary arterial pressure of 60 mmhg (sd 16). pulmonary acceleration time was measured in 13 patients and found shortened in all of them: 52 milliseconds (sd 16), and septal abnormal movement was detected in 10 patients (39%). 25 out of 26 patients had more than one sign of severe pulmonary embolism (spe), 10 had two sign and the other 15 had three or more signs. echocardiography is a simple technique, which allows the diagnosis of spe by the detection of emboli in the right heart cavity and / or the objectivation of indirect signs of functional alteration of right ventricle. coagulopathy and systemic inflammatory response have been previously reported in patients after cpr (1). the coagulopathy includes activation of coagulation and inhibition of fibrinolysis, alterations similar to those reported in sepsis where profound depletion of anticoagulation proteins have been evidenced, and had significant therapeutic consequences (2). however, anticoagulation proteins: protein c and s (pc -ps), as well as antithrombin (at) levels were not reported after cpr. consequently, serial measurements of markers of coagulation (thrombin-at [tat], d-dimers), fibrinolysis (plasminogen-activator inhibitor 1: pai-1), inflammation (il-6) and endothelial injury (soluble thrombomodulin: stm) were performed in 50 patients (age: 61±13 years; saps: 64±13) after successful cpr.analyses on biomarker levels by anova were performed. the aim was to evaluate the effect of thrombolytic therapy in massive and submassive pulmonary embolism methods. 22 patients (16 women and 6 men), with a mean age of 59 years (sd 18), range: 21 to 83, studied prospectively. diagnosis at admission was confirmed with spiral ct scan and/or ventilation-perfusion (v/p) gammagraphy. a study protocol was performed in all patients consisting of: complete analysis, electrocardiography, thorax radiography and echocardiography.one hundred milligrams of rt-pa was infused in 2 hours due to severity of the clinical presentation: haemodynamic instability (4 cases) and/or severe hypoxemia or echocardiographic abnormalities (8 patients). clinical improvement was seen in the entire group. studied variables pre and postthrombolysis are shown in the table. mean arterial pressure (map); right ventricular diameter (rvd), systolic pulmonary arterial pressure (spap), acceleration pulmonary time (apt), oxygen saturation (os), fibrinogen, hematocrit and heart rate (hr). postthrombolytic changes in electrocardiogram were objectivated, showing that some abnormalities had disappeared, such as: right bundle heart block in 8 out of 19 patients (42%), s1q3t3 pattern in 5 out of 20 (25%), t wave alterations in 6 out of 21 (29%), and the pulmonary p in 2 out of 4 (50%). minor hemorrhagic complications were observed in 7 cases; only one needed transfusion. one patient had hematuria, one other hemarthrosis, and another one suffered pericardial blood effusion (after coronary by-pass graft). we have previously shown that the measurement of gut intraluminal redox potential (eh) during progressive bleeding and reperfusion is useful to monitor changes in oxygen transport 1 . eh could provide with a different type of information from other parameters of tissue oxygenation, such as lactate and intramucosal ph. our goal was to show the rate of decrease of eh after the occlusion of superior mesenteric artery blood flow (qintestinal). eight anesthetized and mechanically ventilated sheep were studied. eh was measured as a voltage difference using a milivoltmeter with a platinum electrode, against a reference electrode. qintestinal was measured with an electromagnetic flowmeter. after basal measurements, superior mesenteric artery was occluded and eh was continuously registered during 30 minutes. data (mean ± sd) were analyzed with repeated measures of anova followed by dunnett's test. response time was defined as a decline greater than three sd from basal values. assessment of heart rate variability (hrv) has been used in risk stratification after acute myocardial infarction, in congestive heart failure, and in the early diagnosis of diabetic neuropathy. patients with end-stage renal disease (esrd) constitute a population of increased cardiovascular morbidity and mortality. hemodialysis patients often show signs of autonomic neuropathy. data on hrv are usually derived from 24-hour holter recordings. however, short term rr interval variation as measured on standard 12-lead ecg holds important prognostic implications in subjects with dilated cardiomyopathy. the purpose of this study was to look at short term rr variation in esrd patients, and its modification after dialysis. methods. 47 (32 male, 15 female) patients were included in the study.all of them were in three times a week hospital hemodialysis. twenty control subjects, of similar age and gender distribution, with normal renal function and blood pressure, were recruited among ward stuff. the rr intervals were measured from a continuous 2-min recording of lead ii. rr variation was calculated as the standard deviation of the rr intervals (rrsd), and the coefficient of variance of the rr (rrcv), i.e. standard deviation divided by the mean rr and expressed as percentage. ecgs were also analysed for left ventricular hypertrophy (lvh). . rrsd and rrcv were significantly decreased in dialysis patients compared to controls. rrsd was 11.28±8.13 vs 39.832.12 p=0.000, and rrcv was 1.56±1.13 vs 4.63±2.75, p=0.000. rrsd and rrcv were not affected by dialysis, but were significantly decreased in those with ecg evidence of lvh, compared to those without. rrsd was 6.87±6.1 vs 10.9±8.8 (p=0.043), and rrcv was 0.95±0.76 vs 1.36±1.26, (p=0.033). rrcv was associated with mg and k postdialysis. rrsd and rrcv were inversely correlated with cornell voltage, an ecg index of lvh. hemodialysis patients present with low short term rr variation in comparison with control subjects. electrocardiographically detected lvh among esrd patients is also associated with depressed rr variability. increased intracranial pressure,as that was seen in patients with large cerebral tumors, reduces frequency of the pulse.prolapsus of the brain masses in tentorial incissura of foramen magnum and consequently bradycardia,respiratory arrest,coma and death might occur in these patients. the aim of this study is to examine the ecg changes in patients with brain tumors in regard to the kind of tumors and localisation . the study group was consisted of 31 patients (15 male and 16 female) of average age 57,9 years (range 30 to 83). there were 10 patients with temporal lobe tumor (4 left and 6 right), 9 patients with frontal lobe tumor (5 left and 4 right), 9 with parietal lobe tumor (3 left and 6 right) and 3 with occipital lobe tumor (2 left and 1 right). ecg changes were evaluated during the first 72 hours from receiving in the icu. large cerebral tumors confirmed with ct, and definitive diagnosis was made pathohystologicaly. the most common ecg abnormalities associated with central lesions that we found were: prolongation of the q-t interval in 64.7% patients with right and 50% with left cerebral hemisphere tumor; elevated, peaked, or notched t waves in 52.9% patients with right and 42.8% with left cerebral hemisphere tumor; and increased p-wave amplitude in 23.5% with right and 28.5% patients with left cerebral hemisphere tumor. the most frequent ecg changes that we registered among rhythm and conduction disturbances were: narrow-qrs tachycardia with regular rhythm; sinus tachycardia in 35% with right and 28% patients with left cerebral hemisphere tumor, sinus bradycardia in 11.7% with right and 14% with left cerebral hemisphere tumor, and incomplete/complete right bundle branch block (bbb) in 28% patients with left cerebral hemisphere tumor. we did not find any specific differences according to the pathohystologicaly type of the tumors. conclusion. ecg abnormalities associated with central lesions in the patients with brain tumors are not depend from the kind of tumors and side of the brain where tumor is located. in the patients with brain tumors on left side of the brain prevails incomplete and complete right bundle branch block (bbb). prolonged mechanical ventilation support (mv) is associated with increased morbidity and less cost -effective admissions in the postoperative period (po) of heart surgery (hs). study conducted to identify variables associated with prolonged mv in patients that underwent hs. methods. cohort study; 293 consecutive patients enrolled from 1/12 to 11/17 of 2001. inclusion criteria: patients submitted to hs and admitted to intensive care unit (icu) in use of mv. exclusion criteria: non-cardiac surgery, admission to icu in spontaneous ventilation or death during the first 12 hours of the po. variables that could be associated with prolonged mv were pre-selected for analysis and grouped according to the period it represented. preoperative period: in-hospital stay duration, age, body mass index, gender, severity of left ventricular dysfunction (lvd), pulmonary hypertension, chronic obstructive pulmonary disease, redo, urgency for the procedure. peroperative period: surgery, extracorporeal circulation (ecc) and aortic clamping duration, fluid and blood input/output differences, type of surgical procedure, combined procedures, need for post-ecc intraaortic balloon counterpulsation (iab). admission to the icu: oxygen alveolar/arterial difference, blood oxygen partial pressure/oxygen inspired fraction ratio (p/f). first 24 hours of po: dobutamine or norepinephrine (nor) use, blood drainage volume, lowest blood lactate measurement and prognostic scores sofa, tiss and mods. for dichotomyc variables, mann-whitney test was applied; for continuous variables we used kendall's tau non-parametric correlation test; cuzick tendency test was used to evaluate association with lvd. results. median mv duration: 8 hours; mean duration time 37.8 hours (1 to 742). increased mv duration was associated with emergency surgery (p=0.0117), coronary artery bypass graft surgery (p=0.0307), need of iab counterpulsation after ecc (p=0.0005), use of nor (p<0.0001). increases in mv duration were associated with increasing values of some variables (positive correlation): age (p<0.0001), surgery duration (p=0.0058), blood input/output difference (p<0.0001) and sofa, tiss and mods scores (p<0.0001). negative correlation was presented for fluids input/output difference (p=0.0142) and p/f (p<0.0001). increased linear tendency for mv duration correlates with worsening of lvd (p=0.012). conclusion. more sophisticated statistical analysis should be applied in order to determine the cause-effect correlation for these variables. interventional studies will be conducted in the following months. cerebral vasospasm is a, potentially, life threatening phenomenon after aneurysmal subarachnoid hemorrhage (sah). as part of "triple h" therapy, fenylephrine and norepinephrine in combination with dobutamine and dopamine, are used most frequently to elevate systemic arterial blood pressure (abp) in order to preserve optimal cerebral blood flow. in obtaining increased arterial blood pressure during episodes of vasospasm we altered medical therapy from fenylephrine to norepinephrine but experienced an increasing incidence of paralytic ileus. in a retrospective cohort study we evaluated clinical outcome and the incidence of paralytic ileus. methods. in 1991-1999, a consecutive series of 146 patients had surgery (aneurysmal clip ligation) within 72 hours after sah. 80 patients with clinical vasospasm, were subdivided into two groups with respect to medication used. group a (n=48) was treated with a combination of dobutamine, dopamine and fenylephrine (mean increase syst. abp 40.0±20.5 mm hg). in group b (n=32) norepinephrine was used instead of fenylephrine (mean increase in syst. abp 57.8 ± 27.3 mm hg). we compared basic variables of the two treatment groups, and investigated the clinical outcome using the glasgow outcome scale (gos), one year after initial sah. complications were registered and compared between these treatment groups. results. the two treatment groups were evenly matched concerning age (p=0.28), wfnsscore at admission (p=0.89), amount of subarachnoid blood on ct-scan (fisher) (p=0.75), and observed prognostic variables as hypertension (p=0.50) and smoking (p=0.65). the clinical outcome, was not influenced by the kind of medication used (p=0.55). the incidence of paralytic ileus differed between the two groups (group a: 2/48 vs group b: 12/32, p<0,001). paralytic ileus occurred mainly in patients treated with norepinephrine (12/32=37.5%, odds ratio=13.8). no relationship was found in height of systolic abp or dosage of norepinephrine administered to these patients. we observed a significant difference in duration of administration of norepinephrine in patients, who did develop a paralytic ileus (see -12.14 / -3.42 p=0.002 table: norepinephrine use in cerebral vasospasm; patients with or without paralyticileus. conclusion. -the use of norepinephrine in patients with cerebral vasospasm after sah did not influence clinical outcome, although higher blood pressure levels were reached. -norepinephrine administered during longer periods than 12 days, increases the risk of paralytic ileus.-fenylephrine is recommended in the treatment of cerebral vasospasm after sah. amigues l. 1 , klouche k. 1 , massanet p. 1 , canaud b. 1 , béraud j. j. 1 1 intensive care unit, lapeyronie university hospital, montpellier, france introduction. slow discontinuous ultrafiltration (sduf) is nown recognized as an effective complementary treatment for congestive refractory end stage cardiac failure(escf). however, an organic kidney disease is often associated with heart failure and may worsen the prognosis. this study was undertaken to compare the effects of sduf in escf patients with and without previous kidney disease. methods. 39 patients fullfilling escf criteria with fluid overload and oliguria (grade iv nyha) were treated by sduf. sduf was performed with a double head pump monitor (bsm 22, hospal), blood flow rate: 150-200 ml/mn, ultrafltration rate: 0.2 to 1 l/h. vascular access was provided by femoral silicone twin catheters. a renal replacement therapy was institued when indicated. age, sex, cardiopathy, and nephropathy were collected in each patient. patient follow up before and after sduf: systolic arterial pressure, heart rate, diuresis, total fluid volume removed, cardiothoracic index, creatinemia, blood urea nitrogen, natremia, natriuresis, mortality and average survival. datas were compared between two groups: group 1without and group 2 with nephropathy. mean age was 65±9 yo and sex ratio was 1/4,3. myocardiopathy origin was: ischemia (11), hypertension (6), valvulopathy (4), primary non-obstructive cardiopathy (7), multifactorial (11). oliguric renal failure: fonctional in 20 patients (group 1) and associated with mild chronic nephropathy in 19 patients prior to heart attack (group 2). no significant differences in clinical and biological datas were observed between the two groups except for blood urea nitrogen: 26,2±15 in group 1 vs 39,2±16,8 mmol/l in group 2. during scud, hemodynamic stability was observed in both groups; diuresis and natriuresis significantly increased and remained stable at the end of the treatment despite significant decreased diuretic doses. mean sessions of sduf was 3,8±2,4 in group 1and 5,6±5,6 in group 2 (ns). renal replacement therapy was institued in both groups but the number of sessions was significantly higher in group 2: 3,8±4,7 vs 0,95±1,3. mortality during hospitalisation was 25% in group 1 and 21%in group 2. from the surviving patients, 4/15 patients in group 1 and 6/15 patients in group 2 underwent a chronic hemodialysis treatment. average survival was higher but not significant in group 2 (21,3±36 vs 7,7±9 months). our study sugests that sduf remains a long term beneficent treatment for patients with both escf and renal failure. paradoxically, prognosis is slightly better than in patients with isolated refractory congestive heart failure. organic renal failure could artificially worsens cardiac function by increasing diuretic resistance which may be improved by sduf. (1). this study aims to evaluate the influence of factors affecting renal blood flow including map, cvp, pulmonary artery wedge pressure (pawp), cardiac index (ci), systemic vascular resistance (svr) and pulmonary vascular resistance (pvr) on immediate graft function. methods. 15 consecutive patients undergoing live-related kidney transplant were included. prior to anaesthesia, a 7.5f continuous cardiac output pulmonary artery catheter was placed via the right internal jugular vein. a continuous cardiac output monitor (baxter edwards px 1800) was used for haemodynamic monitoring. baseline values of the map, cvp, pawp, ci, svr and pvr were computed. data was collected at 30-minute intervals thereafter and immediately before and after release of the vascular clamps. the ischemia time, intravenous fluids, dopamine use and blood transfusion were noted. if warranted by low preoperative haemoglobin or increased surgical blood loss, blood was administered as 200 ml packed cell units using leukocyte filters. the outcomes chosen for graft function were the urine output on table (uo-ot), 24-hour urine output (uo-24), fall of serum creatinine from the preoperative value on day 1 (creat-1) and day 7 (creat-7). using spss statistical software, multiple linear regression analysis was done to find the variables significantly affecting the outcome. results. the only variable found to have a statistically significant influence on uo-ot was the map. no variable had any effect on the uo-24. blood transfusion had a negative influence on the fall of creatinine on day 1 and day 7 ( since clinically adopted during cardiac surgery, cardiopulmonary bypass (cpb) has been implicated in complement activation and postoperative acute phase reaction. corticosteroids are usually employed as an attempt to dampen these phenomena and related postoperative morbidity. methods. after informed consent previously approved by the local ethical committee, we included 62 adult patients submitted to cardiac surgery under cpb at a non-emergency setting. preoperative risk stratification employed euroscore (es) and cleveland clinic score (ccs). methylprednisolone (mp) -15mg/kg, was added to cpb priming solution for group 1 (n=31) but not for group 2 (n=31). blood samples were collected from all patients at anaesthesia induction (t0), 3(t3), 6(t6) and 24-hour postoperative (t24) for measurement of total c3 and c-reactive protein (crp) levels, by nephelometric technique. postoperative multiple organ score (mods) were daily registered. results. the groups were considered comparable concerning to preoperative risk stratification, length of cpb and postoperative organ dysfunction at 72h postoperative (mod72, as well. starting from similar levels of c3 and crp, we did not observe significant differences between groups 1 and 2 concerning to postoperative levels of c3. nevertheless, patients treated with mp (group 1) exhibited higher crp levels at 24h postoperative, as shown bellow: 11.24 ± 6.34 6.57 ± 6.20 0.001 perioperative administration of mp failed to show evidences of beneficial effect over postoperative organ failure and complement activation. the acute phase response, expressed as crp systemic levels, instead of softened, was significantly enhanced among patients to whose cpb priming solutions was added mp. these results support a larger randomised trial to reassess routine use of corticosteroids during cpb. objective: to evaluate the influence of enteral application of an immunoglobulin enriched bovine milk preparation on endotoxin plasma levels, endotoxin neutralizing capacity of plasma (enc) and the acute phase response (il-6, crp) during and after cardiac surgery in a pilot study. design, patients and methods. 60 patients who were treated by coronary bypass operation were enrolled in a controlled randomized study. the patients were treated by enteral application of 40g of a bovine colostrum milk preparation per day for 2 days preoperatively. endotoxin and enc were sequentially determined intra-and postoperatively by a chromogenic modification of the limulus amebocyte lysate test. interleukin-6, crp, transferrin, alpha-2macroglobulin, albumin, apo-a, apo-b, igg, iga, igm were determined by "elisa" and nephelometrically. the clinical course was followed up by daily evaluation of the apache-ii-score. main results: demographic data were comparable in both groups. no differences of the apache-ii-score (6.5 1.9 verum group, and 6.8 1,8 control group, on admission) were observed. endotoxin plasma levels and enc showed high levels at the end of the procedure and seemed to have a trigger function for the acute phase response but were not significantly reduced throughout the observation period in patients receiving the milk preparation as calculated by comparing the area under the curve. plasma levels of endotoxin binding proteins did not differ significantly. plasma levels of il-6 increased to maximal median values of 655 pg/ml in the verum and 786 pg/ml in the control group 2 and 6 h after surgery. a tendency to lowered il-6 levels was observed throughout the whole observation period for the verum group. crp-levels showed their maximum values 48h after the procedure and were significantly reduced in patients of the verum group (p = 0.034). conclusion. this study revealed that endotoxemia occurs early during an elective surgical intervention, which is followed by a subsequent increase in mediators of the acute phase reaction. the prophylactic enteral application of a bovine milk preparation for two days in cardiac patients did reduce postoperative crp-plasma levels but contrary to a former prospective double blind study in abdominal surgery did not reduce perioperative endotoxemia. one reason could be the too low application of the bovine colostrum milk preparation. to compare a possible effect of improved therapeutical approaches in head trauma, epidemiological data should be compared at certain time points. due to the legal obligation to document all in-patient treatments in germany and to forward these data in an anonymous form to the office for statistical affairs (statistisches bundesamt) it is possible to provide a nationwide epidemiological analysis of head trauma and to compare the yearly obtained data. the incidence, the mortality, and the duration of hospital stay for the treatment of all hospitalized patients suffering from head trauma were calculated and compared to the data from 1996 while considering the data obtained from the office for statistical affairs in bonn and wiesbaden. the data were investigated while separating them according to the international classification of diseases 9 (icd-9; no. 800-804 and 850-854). to further elucidate the causes of altered mortality and duration of hospital stay the number of cts and mris in german hospitals in 1996 and 1999 were compared. in addition, data indicating the number of patients admitted to neurological rehabilitation centers were analyzed. the incidence of head trauma did not change between 1996 and 1999 and was calculated to be at 340/100.000. the mortality, however, decreased from 11.6/100.000 in 1996 to 9.4/100.000 in 1999 (9412 vs. 7705 patients). in addition, the duration of hospital stay declined in all icd-9 encoded subgroups including mild brain trauma. this could be due to the increased number of ct devices and mris in german hospitals (ct: 732 vs 904/ mri: 227 vs 350) while comparing 1996 and 1999. the number of patients transferred from hospitals to neurological rehabilitation centers increased from 35800 in 1996 to 61088 in 1999 (+ 71%). it could be speculated that both improved knowledge on the field of brain trauma therapy and a higher number of technical devices allowing rapid diagnosis of brain injury or potential intracranial complications following head trauma accounted for the reduction in mortality due to brain trauma in germany from 1996 through 1999. the decline of the duration of hospital stay especially in patients with more severe head injury could also be due to a more rapid transfer of patients with head trauma from hospitals to rehabilitation centers. therapeutic hypothermia may improve outcome in patients with severe head injury, but clinical studies have produced conflicting results. we hypothesised that severe side effects of artificial cooling might have masked positive effects in earlier studies, and treated a large group of patients with severe head injury with hypothermia, using a strict protocol to prevent the occurrence of cooling-induced side effects. methods. 136 consecutive patients admitted to our hospital with severe head injury (glasgow coma scale (gcs) <8) in whom icp remained above 20 mmhg in spite of therapy according to a step-up protocol described previously [1] were included in our study. those who responded to the last step of our protocol (barbiturate coma) constituted the control group (n=72). those who did not respond to barbiturate coma (n=64) were treated with moderate hypothermia (32oc-34oc). average apache ii scores were higher, and average gcs at admission slightly lower in the hypothermia group, indicating greater severity of illness and more severe neurologic injury. predicted mortality was 86% for the hypothermia group vs. 80% in controls. actual mortality rates were significantly lower: 62% vs. 72%, p<0.02. the difference in overall mortality between hypothermic patients and controls was statistically significant (p<0.05). the number of patients with good neurologic outcome was also higher in the hypothermia group: 15.7% vs. 9.7% for hypothermic patients vs. controls, respectively (p<0.02). these differences were explained almost entirely by the subgroup of patients with gcs of 5 or 6 at admission (mortality 52% vs. 76%, p<0.01; good neurologic outcome 29% vs. 8%, p<0.01). artificial cooling can significantly improve survival and neurologic outcome in patients with severe head injury, when used in a protocol with great attention for the prevention of side effects. these effects are especially clear in patients with gcs of 5 or 6 at admission. because there is likely to have been bias against the hypothermia group in this study, the positive effects of hypothermia might even have been underestimated. introduction. s 100 b, a glial calcium-binding protein, is a serum marker of cerebral damage. posttraumatically, however, s 100 b in increased in all patients suffereing from hemorrhagic-traumatic shock, regardless of whether trauma is cerebral or extra-cerebral. the aim of this experimental study was to determine whether the posttraumatic s 100 b increase is caused by extra-cerebral trauma or by hemorrhagic shock and whether it is influenced by the severity of shock. hemorrhagic shock was achieved by bleeding anesthesized rats to a mean arterial pressure (map) of 30-35 mm hg through a femoral catheter and maintaining this map until incipient decompensation. subsequently, map was either increased immediately to 40-45 mm hg (moderate shock) or maintained at 30-35 mm hg until 40% of shed blood had been returned (severe shock), and then increased to 40-45 mm hg. resuscitation was provided after 40-45 mm hg map had been maintained for 40 min. trauma was achieved by midline laparotomy. hemorrhagic-traumatic shock caused an early s 100 b increase at the onset of decompensation. s 100 b in serum was highest at the end of the 40 min. period during which map was maintained at 40-45 mm hg and was significantly higher at all time points after severe shock than after moderate shock. in contrast, trauma (laparotomy)without hemorrhagic shock did not cause any increase of s 100 b in serum. the posttraumatic s 100 b increase in serum appears to be caused by hemorrhagic shock rather than by extra-cerebral trauma. regardless of whether the source of s 100 b is cerebral, indicating cerebral damage linked to shock, or extra-cerebral, the main determinant in the clinical setting remains the severity of shock. romera m. a. 1 , chamorro c. 1 , silva j. a. 1 , pardo c. 1 , marquez j. 1 , ortega a. 1 1 intensive care unit, clínica puerta de hierro, madrid, spain in patients with non-traumatic subarachnoid hemorrhage (sah), the development of myocardial abnormalities has been widely described. however, the true incidence of myocardial injury in this group of patients is unknown yet. we analyze the incidence of myocardial injury, in this population, using cardiac troponin i (tn i) assay and also we assess if the increase in tn i concentration has prognostic value. prospective study, including all patients with non-traumatic sah admitted to our intensive care unit (icu), from december 1999 to december 2001. serum tn i concentration was measured, at least once, within the first 72 hours after onset of symptoms. inmunoassay based on the "sandwich" principle was employed. the chi-squared test and fisher exact test were used for statistical analysis. of the 96 patients admitted, 14 were excluded ( admission later than 72 hours, absence of tni determination, previous cardiopathy or renal failure ). eighty-two patients were included in the study (50 women ). mean age 50±15 years. the tni concentration was increased in 24/82 patients (29% ). sixteen (19.5% ) patients died in icu. twelve of the 24 (50% ) with a high tni concentration and 4/58 (7% ) with a normal tni concentration died [ relative risk (rr) 7.25 (2.6 to 20.2; 95 % confidence interval (ci); p<0.001 ]. thirty-seven ( 45 % ) patients had a hunt-hess (hh ) grade greater or equal to iii. poor grades of sah ( hh>or = iii ) were associated with a higher incidence of raised tni concentration ; ci 95 %); p<0.001 ]. among this group of poor grade patients, elevated tn i levels were associated with a higher mortality [12/21 (57%) with a raised tni compared with 3/16 (19% ) with a normal tni concentration; rr 3 (1.03-9; ci 95% ); p<0.05]. however, mortality in every case was related to neurological problems. seven patients (8.5%) suffered from pulmonary edema and all had elevated tni levels. echocardiography was performed in all 7 patients, being abnormal in 5 of them. conclusion. in our series, the incidence of myocardial injury in sah was 29 %. this cardiac injury was more frequent among patients with severe grades of sah. elevations in tn i levels had prognostic value, being associated with a higher mortality. therefore, we should closely monitor those patients with sah who develop an increase in the tni levels. renaud e. 1 , matéo . j. 2 , benlolo . s. 2 , payen . d. 2 1 dept of anesthesiology and critical care, lariboisiere hospital, 2 department anesthesiology intensive care, lariboisiere, paris, france respiratory failure is one of the major complication of acute stroke (1). we have investigated the impact of the location stroke on respiratory failure incidence, cause of intubation and outcome. we reviewed 40 consecutive patients with acute stroke admitted to icu from 1998 to 2001. following data were collected, glasgow coma score (gcs), cause of icu admission, presence of acute respiratory failure (arf), reason for intubation, presence of aspiration, length of mechanical ventilation (lomv), severity of hypoxia, length of stay in icu (los) and mortality. continuous data were compared by paired t-test and nominal data by chi-test. explicative variables for arf were assessed by univariate analysis. . 24 patients had a middle cerebral artery (mca) stroke and 16 had brainstem stroke (bs). age (mca 50±13sd yrs vs bs 52±13sd for), gsg score (mca 8±3sd vs bs 10±3sd for), length of stay in icu (12±15sd days for mca vs 15±13sd) were not significantly different. 68% bs and 33% mca patients were admitted in icu for respiratory failure (p=0.01). admission to icu with loss of consciousness was significantly higher in mca (19/24, 80%) than in bs (0/16) (p=0.001). indication for intubation was always for aspiration pneumonia that was the leading cause of arf (0.0007) associated with swallowing paralysis in bs (p=0.001) and to unconsciousness in mca (p=0.03). there was no difference for the lomv, the severity of hypoxia between the 2 groups. arf, intubation or reason for intubation were not associated with mortality in the 2 groups (p=0.1). the major cause of death was the presence of cerebral herniation in the 2 groups (p=0.004). pulmonary complication due to aspiration more predominant in bs than mca stroke, represents the major cause of intubation and arf for bs patients. in the contrary, loss of consciousness in mca stroke group predominates for icu admission. outcome in all patients (mca and bs) was not influenced by presence of respiratory failure or reason for intubation. the major cause of death for stroke's patients is the neurologic state, and especially the presence of herniation. stroke code (sc) is a guidelines of united actuatio between out of hospital enmergency services from barcelone and the most four important hospitals of the city; which aim is to optimize the sequence time for stroke treatment; this allows to increase the number of candidates for reperfusion therapy. the present study aim is to evalute differents times sequences in the acute strokes in which trombolysis has been practised according to the acute stroke code first priority; and secundary to describe findings in the ct scan of these patients pro-inflammatory cytokines, such as tnf and il-1 are released in the brain within hours after closed head injury (chi). they were shown to have deleterious effects, mainly when active in the early post-injury period. a variety of anti-inflammatory and anti-apoptotic modalities have been shown to ameliorate the outcome of chi. erythropoietin (epo) is a kidneyderived cytokine regulating haematopoiesis both by acting as a growth factor and by inhibiting apoptotic cell death. recently it has been shown to be produced in cultured neurons, brain astrocytes and neurons under hypoxic/ischemic conditions and in response to oxidative stress. other studies have shown that the erythropoietin receptor (epor) is present under normal conditions on neuronal and brain capillary endothelial cells. epo has been found to have newly discovered neuroprotective properties in different models. these models include neuronal cultures against glutamate toxicity, global glutamate toxicity and rodent models of cerebral ischemia. in addition it induces brain endothelial cell proliferation and stimulates neovascularization in vivo. the present study was designed to test the protective effects of epo in rats udergoing controlled chi. methods. chi in rats was induced using a weight-drop device. clinical status was evaluated by the neurological severity score (nss), which tests 10 tasks including reflexes, behavior and motor activity. a point is awarded for failing to perform a task so a higher score corresponds to a more severe trauma. study animals were treated with 2 doses of i.p. 5000 units/dose (1 ml) of rhu-epo, 1 h and 24 h after chi (treatment group) or with 1 ml of vehicle injected i.p. at the same time points (control group). nss was evaluated by an observer blinded to the different groups at 1, 3 and 7 days post chi. nss scores were compared using a two tailed student t-test. control and study rats were subjected to chi of similar severity, (1h nss 10.4+0.78 and 10.22+0.46 respectively, p=0.86) and followed at 1d, 3d and 7d following chi. clinical recovery was facilitated in the treatment group starting at 24 h after chi and reached statistical significance at 7 days post chi. the treatment group's 7 d nss was 5.0 (n=8) vs. 6.625 in control animals (n=9) p=0.037. the present findings point to a neuroprotective role of epo in traumatic brain injury. brain tissue of treated and control animals is currently being analyzed for parenchymal cytokine levels. we have examined the role of post trauma treatment with epo of rats undergoing chi. as has been shown in other models of brain injury (stroke, ischemia, glutamate toxicity) epo seems to have a neuroprotective effect in head trauma. the exact mechanism of this protection has yet to be elucidated. this is the first time, to our knowledge, that epo has been studied in an animal model of traumatic head injury. (1998) (1999) (2000) is 4.7%. it is difficult to know how to apply these figures to individual patients. we have used the anaerobic threshold in a prospective observational study to try and identify patients with an increased risk of mortality. forty-five patients scheduled for elective aaa repair had their anaerobic thresholds measured pre-operatively. the anaerobic threshold is the patient's oxygen consumption in ml/kg/min when anaerobic metabolism occurs(reference 2). it is calculated by using a bicycle ergometer and a metabolic cart. clinical presentation and evolution of valvular heart disease (vhd) patients have great significance in determining the best moment for surgical correction but lacks correlation with surgical outcome in most cases. this study tries to determine the preoperative variables associated with mortality in the course of surgical treatment of vhd. cohort study conducted from january 2001 to february 2002. inclusion criteria: patients submitted to vhd surgery during the period of study. exclusion criteria: vhd surgery combined with non -vhd procedures. data were analyzed with chi -squared, fisher and mann -whitney tests. one hundred five patients met the inclusion criteria. the preoperative variables associated with surgical mortality were: systemic arterial hypertension (p=0.016), peripheral vascular disease (p=0.031), redo (p=0.031), age (p=0.051), blood creatinine level (p=0.042), left ventricular dysfunction (p=0.02). conclusion. based on these data, efforts will be held in order to develop a prognostic score index for mortality in vhd surgical patients. in our setting, the diffusion of institutional education in basic cardiopulmonary resuscitation (bcpr) is low. the number of patients admitted to our units after resuscitation following cardiac arrest is rising due to the population demand on the out-of-hospital emergency services, 061. the patients with neurological sequelae secondary to incorrect bcpr in the first 10 minutes are common. through the association of ex-patients of the intensive care medicine department, and with the psycho-social support of voluntary helpers on patient discharge, relatives are offered bcpr as part of the quality care programme. every three months, professionals from the department organise this course for 20 relatives in the form of a 5 hour module. the concepts of the prevention of ischaemic heart disease are presented together with the content of the national plan for bcpr. practical sessions are undertaken in small groups of 6 to 8 persons, using dummies. a total of 294 relatives in 13 courses have received this training over the past 5 years. the mean age of the students was 38.3 (sd 15) years (11-75), 72% women, 32% with middle and higher education, 20% housewives, and 15% manual labourers. the evaluation of the scores obtained in the 16 item test before and after the course is shown in the tables below. multiorgan system failure (mosf) is an infrequent but very serious complications after cardiac surgery, with high rates of mortality. this study was undertaken to determine the frequency, prognosis and risk factors for mosf this study was performed in a twelve-bed cardiac surgery intensive care unit over a 24-month period. all adult consecutive patients undergoing coronary, valvular and combined (valvular and coronary) surgery were prospectively studied (n = 2102). all patients were assessed by the "modified" parsonnet score results. mosf developed in 138 (6.57%) patients, of whom 51 (36.9%) died. this was the main cause of overall hospital mortality (51/92, 55.4%). in a logistic-regression anlysis, the development of sepsis, postoperative low cardiac output syndrome, mechanical ventilation more than 72 hours, a "modified" parsonnet score more than 25 and and preoperative ventilatory support were independantly associated with the development of mosf. an organ system failure index (osfi) of 3 or more was most significantly associated with icu mortality (p<0.001). conclusion. in our series mosf was a leading cause of mortality after open-heart surgery. the development of mosf with an osfi of 3 or more was the main predictor of postoperative mortality. we studied 211 patients who underwent cabg surgery. fifteen patients (13 male and 2 female) were younger than 45 years and 196 patients (147 male and 49 female) were older than 65 years. perioperative death occured in one patient from the <45 years group and in 10 patients from the >65 years group (p=ns). categorical data were compared using the chi-square test and numerical data were analysed using the student t-test. differences were considered significant at p<0.05. in a n investigation conducted in ours icu, 38% of patients hospitalised after elective cardiac surgery presented a pain score > 30 ( min score 0 ; max 100) . these results were considered inadequate. a quality improvement initiative was undertake. the aim of the present study was to test if pain evaluation and treatement improved following pain guidelines implementation in a surgical icu. the design consisted in observing de pain evaluation both before and 1 month after implementation of guidelines. these guideline are divided in two item : first introduction of a regular pain evaluation using a visual analogue scale (vas) and second in a proportional vasderived analgesic prescription protocol. recommendation were given during repetitive meetings, feedback sessions and regular poster information on the icu walls. pocket guideline and vas tool was distributed. pain intensity evaluation of the nursing team was checked by an independent observer and compared with the nurses-charted vas. improvement of pain control was tested based on the following criteria: utilisation of the algorithm at least twice per working shift; corresponding analgesic drug to observed vas; and follow up check of vas after analgesic administration. the independent observer measure vas at 8 a.m. and at 4 p.m. postoperative day 2 and 3. proportion of algorithm adherence before and after introduction of the recommendation were tested using fisher's exact test. variance of median vas was tested using mann-whitney test. these preliminary results indicate that the implementation of an algorithm on pain intensity evaluation and treatment increases the number of pain evaluation and re-evaluation after drugs administration. although the administration of analgesic drugs increased, the number of patients with insufficient pain treatment stays still high. the prognosis of liver transplant has improved the last few years due to advance in surgical techniques and immunosuppressive regimes, but early complications show a high prevalence affecting morbi-mortality in these patients a 42 beds icu in a teaching 3rd level hospital. prospective observational study on all patients with the mentioned condition treated in our centre from october 2000 to october 2001. follow up during icu stay. we have collected data from 46 patients (47 grafts) with a mean age of 52.4±9.6 years, 28.3% women, mean apache ii on admission 17±4.1, median child score 7,5 (7-8) and mean sofa score 5.8±1.8. surgical data were as follows: fluids balance 478±2570, hours of graft ischemia 7.6±2.1, reperfusion syndrome in 21% and fibrinolysis in 4.3%. at admission mean core temperature was 35.3±1 ºc. median icu stay 3.5 (3-4, max. 17) days and median hours under mechanical ventilation 10 (7.5-16, max 67). the prescribed immunosuppression was cyclosporine in 50% and tacrolimus in 50% of patients results. icu mortality was 6.5% (3 patients). complications were present in 78% (45.7% of them more than two episodes). 2 patients had to be reoperated, one because early graft dysfunction treated with mars and retransplantation (death because a new graft dysfunction), and the other because abdominal haemorrhage. one patient developed an early rejection. metabolic complic 60.9% (high insulin requirements 41.3%) -renal failure 45.7% (renal replacement 6,5%)-cardiac complic 30.4% (chf 8.7%, hbp 10.9%) -respiratory complic 19.6% (10.9% sdra) -bleeding 13% -neurological complic 10.9% (myelinolysis 1 patient) -infection 2.2%. patients who died had higher apache ii, child and sofa scores, lower serum albumin levels, longer graft ischemia, higher percentage of fibrinolysis and reperfusion syndrome during surgery and higher percentage of acute renal failure an need for renal replacement (not statistical analysis due to the low mortality rate we report the effects of substitution with a virus-inactivated protein c (pc) concentrate in disseminated intravascular coagulation (dic) in preterm infants and children with sepsis (meningococcal in the children and aldolecent; staphylococcal and enterobacter in the preterms) associated with purpura fulminans. this was a prospective open-label study. a total of 16 patients,9 paediatric and adolescent patients age 0.2 to 18.25 years with dic associated with severe acquired pc deficiency (range 0.02 to 0.48 iu/ml; median, 0.22 iu/ml) in meningococcal septic shock and purpura fulminans; and 7 preterm infants with severe acquired pc deficiency (range 0.01 to 0.1iu/ml; median, 0.02 iu/ml) due to staphylococcal and enterobacter sepsis were studied. replacement therapy was initiated with a virus-inactivated pc concentrate with an initial intravenous bolus of 80 to 120 iu/kg followed by 50 iu/kg up to six times per day as an adjunctive therapeutic regimen to otherwise optimal intensive care treatment. after initial pc administration, plasma pc levels rose to normal ranges and were maintained under pc replacement therapy. improving or even normalising global hemostatic parameters were assessed in all patients. markedly elevated plasminogen activator inhibitor type 1 (pai-1) levels prior to treatment, reflecting a reduced fibrinolytic potential, decreased rapidly under pc substitution. concomitantly improving signs of purpura fulminans reflected by decreasing size of skin lesions, demonstrated a restoring microcirculation. seven of the nine paediatric and all of the neonatal patients survived. one patient (paediatric) required limb amputation; two patients died because of multiorgan failure. both presented with a severely low plasma pc activity of 0.02 iu/ml on admission to the hospital. no adverse effects were observed with the pc concentrate administration. ait can be concluded that the administration of pc concentrate had a marked benefit on the deranged coagulation status of patients with purpura fulminans and septicaemia. normalisation or even partial correction of haemostasis as well as improvement of microcirculation accompanied by improving signs of purpura fulminans were demonstrated in all patients the main purpose of this study is to report medical and surgical complications of spine surgery in a third level universitary pediatric hospital with a reference spine surgical program. methods. study design is a retrospective clinical series of 73 pediatric spinal surgeries.all spine surgeries performed on children under 18 years of age between january 2000 and january 2002 were included. patient were grouped in four diagnostic categories (idiopathic, neuromuscular, congenital scoliosis and miscellaneous) and procedure performed (posterior (p) fusion, anterior/posterior (ap) fusion, anterior (a) fusion, (iw) instrumental withdrawal). next data were recorded from clinical chart:age, gender, needs of transfusion products, volume demands during first postoperative day,days on mechanical ventilation,medical and surgical complications. results. study sample included 73 patients, 47 female and 26 male. age ranged between 4 and 18 years with average of 12.7 years. characteristics were: idiopathic 29, neuromuscular 26,congenital scoliosis 7,miscellaneous 11. procedures performed were:p fusion 53,ap fusion 17,a fusion 1,iw 2. .average lenght of stay in pediatric intensive care unit were 1.3 days (range 0-25).average days on ventilatory support 0.46( range 0-2.5). no patient required intubation after weaning.major complications were: deep wound infection(2), respiratory distress(1), large intraoperative blood loss (1),and paraplegia (1).no deaths were observed.minor complications were: atelectasis(6), pleural effusion(4), pneumonia(2), pneumotorax(1), superficial wound infection(4), urinary tract infection(3) and electrolitical disturbances(17). postoperative transfusion needs were 8.4 ml/kg (95% confidence interval (ci) 4.1-12.8) for ap fusion, 6.8 ml/kg (95% ci 4.8-8.7)for p fusion; a fusion and iw doesn't need postoperative blood replacement. total blood transfusion was 40.7 ml/kg (95% ci 29.7-51.8)for ap fusion, 23.2 ml/kg (95% ci19.3-27.1)for p fusion; 6.9ml/kg for a fusion and 7.6 ml/kg for iw.volume demands(no blood products)during first postoperative day were 81.4 ml/kg (95% ci 50.5-112.2)for ap fusion, 59.1 ml/kg (95% ci 48-69.7) for p fusion;38ml/kg for a fusion and 44.8 ml/kg for iw. conclusion. spine surgery has few major complications rate in a reference spine surgery pediatric hospital. minor respiratory complications affect 15% of our patients without repercussion in outcome. total blood loss is greater in ap fusion than in other procedures, but postoperative blood replacement in picu didn't differ between procedures. background elevated intra-abdominal pressure (iap) adversely affects pulmonary, cardiovascular, renal, splanchnic and central nervous system physiology, and it determines the common clinical picture called "the abdominal compartment syndrome". nevertheless the direct monitoring of iap is not always practicable, because it requires an abdominal drainage. a lot of authors demonstrated in the adults that the bladder pressure is a reliable index of iap, but there are not studies on pediatric population. the aim of this study is to evaluate the level of significance of this index in a pediatric population. population: we enlisted a group of pediatric patients, sedated and paralysed (3 oltx, 3 abdominal surgery, 1 cardiac surgery), age 20.8 ± 16,3 (range 4-48) months. methods. the bladder pressure was measured with the patient in supine position, with a trasduction circuit connected to the bladder catheter and to the abdominal drainage (3 jpratt, 3 pig tail, 1 catheter for peritoneal dialysis). to obtain a good transduction of pressure, a volume of saline was pushed into the bladder. the volume of saline was variable according to the weigth and age: we obtained a scheme (table 1 ) from our empirical evaluation of the pediatric bladder compliance and urodinamic data. table 2 , there aren't significative differences between the level of pressure measured in the bladder and in the peritoneal cavity ( p= 0.50). mean:0,00 ds:1,15 from 1998 to 2001, 7 pediatric patients (age range 0.58 to 10 years, mean 3.91 years) were treated using nppv during distinct episodes of acute respiratory failure (arf) of neuromuscular origin. in all patients immediate intubation for an acute, life-threatening presentation was avoided and respiratory status improvement was achieved. few data are available up to now about nppv application and indications in the acute setting in infants affected by neuromuscular disorders (nmd). a prospective observational study was carried out on 7 non-consecutive neuromuscular patients admitted to picu because of arf and managed with nppv in the acute phase;remarkably, 4 out of 7 were <15 months aged. all the patients were treated by a flowtriggered intensive care mechanical ventilator (siemens servo 300 ventilator, siemens-elema, sweden) through a tight fitting face mask. nppv was administered for at least 24 hours postadmission. a pressure-control mode was adopted for better compensation of leaks around the mask. flow-sensitive trigger permitted a better synchronization of patient's spontaneous breathing, limiting the need for deeper patient sedation (low-dose midazolam drip). initially, a relatively low ventilator frequency delivery was set (8-10 b/min). peak inspiratory pressure was tritrated upward to obtain an exhaled tidal volume of 5-6 ml/kg maintaining a paco2 value <65 mmhg and a ph >7.25; peep value was adjusted to maintain an oxygen saturation >91-92% with a required fio2 <0.6. results. all patients were referred to picu on spontaneous breathing: those admitted with et tube already positioned were not considered eligible for this study. an oxygenation improvement was obtained in all patients within 3 hours from the onset of nppv . the pao2/fio2 increased from 86.29 ±16.13 to 235.6±32.32 (p<0.05) and 292.6 ± 25.75 (p<0.001) on selected time points (3 and 12 hours after nppv introduction, respectively); conformly, alveolar-to-arterial oxygenation difference (a-ado2) decreased from a 462.3 ± 77.77 to 195.7 ± 59.67 (p<0.05) and 101.8 ± 23.92 (p<0.001) respectively. conclusion. nppv resulted a safe and effective therapeutic approach in both hypoxemic and hypercapneic arf episodes in this children group affected by nmd. even in cases of emergency presentation or when resuscitation is needed, it is of importance to identify nmd children with residual ventilator-free breathing ability thus performing a nppv trial. life-threatening respiratory distress and young age should not preclude nppv application in a picu setting. pulse oximeters are widely used in paediatric intensive care but they have some severe limitations. the technique relies on the presence of adequate peripheral arterial pulsations, which are detected as photoplethysmographic signals (ppg). when peripheral perfusion is poor as in states of hypovolaemia, hypothermia and vasoconstriction oxygenation readings become extremely unreliable. hence, pulse oximetry becomes unreliable in a significant group of children just at the time when accurate readings are most needed. to overcome this limitation, the oesophagus has been investigated as a potential measurement site on the hypothesis that perfusion may well be better preserved at this central site. studies on adult patients have shown that measurable ppg signals at red and infrared wavelengths can be detected within the whole depth of the oesophagus. a new system to investigate the quality of oesophageal ppg signals is being constructed with the aim of developing a neonatal and paediatric oesophageal pulse oximeter. a reflectance optical sensor has been constructed comprising miniature infrared and red emitters and a photodetector. the sensor was design to fit into a conventional disposable transparent stomach tube, 12 french gauge. the oesophageal ppg sensor within the stomach tube was inserted through the nose into the oesophagus of a 2 kg, 17 day old neonate. the stomach tube was advanced into the oesophagus under direct vision until the probe was 25 cm from the nose. ppg traces from the oesophagus were recorded for approximately 5 minutes at this depth on a laptop computer. measurements were repeated at 20 and 15 cm from the nose. measurable ppg traces of good quality were obtained in the oesophagus at all three depths. the ppg signals in the mid to lower region of the oesophagus on average had larger amplitudes at both red and infrared wavelengths than the ppgs recorded in the upper oesophagus. artefacts on both wavelengths due to oscillations as a result of high frequency ventilation. filtering successfully eliminates the artefact. the new oesophageal reflectance optical sensor has allowed ppg measurements to be made within the whole length of the neonatal oesophagus. the red and infrared wavelengths used are suitable for pulse oximetry. these results are the first to demonstrate that pulse oximetry may be feasible in the neonatal or paediatric oesophagus. further studies are required to develop a neonatal/paediatric pulse oximeter. we used protein c (ceprotein; baxter -immuno) in two patients with moderate or severe, therapy-resistant vod. . patient 1 (e.g, swiss, 1,75 y) suffered from an acute myelogenous leukemia (m 7 with t(1;22) of early infancy after complete first remission by conventional chemotherapy an allogeneic stem cell transplantation with a matched unrelated donor was performed. conditioning comprised busulfan, vp16 and cyclophosphamide. patient 2 (m.k, iranian, 11 y) suffered from beta-thalassemia major with secondary moderate hemosiderosis, as well as chronic persisting hepatitis c infection with liver fibrosis. he received a matched related bone marrow transplantation, using i.v. busulfan , reduced cyclophosphamide dose and fludarabine. both patients received low dose heparin (100 iu/kg) and antithrombin iii substitution. in addition, pat. 2 got prophylactical defibrotide (20 mg/kg) and n-acetylcystein. two weeks after transplant both patients developed vod (severe (pat 1); moderate to severe (pat 2))with weight gain, hepatomegaly, massive ascites and severe thrombocytopenia. maximal bilirubin was 15 mg/dl (pat 1) and 2 mg/dl)(pat 2). therapy with defibrotide (60 mg/kg) was started immediately. in pat.1 the pulmonary situation deteriorated rapidly with massive aszites and oxygen need and a reversed portal venous flow. defibrotide was stopped after 5 days. thrombolytic therapy using rtpa and a continuous pc substitution (pc level 16%; bolus 100 iu/kg, followed by 100 iu/kg every 4h)) were started. lysis therapy had to be abandoned due to respiratory tract bleeding global coagulation (pt 34 %, aptt 250 sec) and pc level normalized within hours after pc substitution. a normal centripetal portal flow could be achieved by high dose defibrotide (120 mg/kg) and continued pc substitution after several weeks. pat. 2 showed only a temporary improvement under defibrotide treatment. due to clinical deterioration (hepatic pain, increased ascites) and low pc level (38%) a continuous pc substitution (50 iu/kg every 6 h) was initiated. there was a prompt recovery after adding pc with a dramatic reduction of ascites, weight and abdominal pain within 2-3 days after start of pc infusion. elevated bilirubin levels returned to normal in both patients. in our 2 patients neither prophylactic administration of at iii nor of defibrotide were able to prevent moderate to severe vod.our data indicate that pc substitution may be a useful adjunctive treatment in severe vod. until controlled studied will be initiated we recommend a stratified treatment in vod, starting with defibrotide, and adding pc in unresponsive cases. (1) showed recently that restrictive strategy of red-cell transfusion could be at least as effective as and possibly superior to a liberal transfusion strategy in critically ill patients. the aim of this study was to assess the impact of local transfusion guidelines emphasizing restrictive strategy on patients undergoing heart surgery and the prognostic value of transfusion following those restrictive criteria. methods. two groups of 100 heart surgery patients were compared before and after the introduction of local transfusion guidelines. these guidelines involved general information on blood transfusion risks and obligation for the physician to respect predetermined transfusion criteria (hb <7g/dl or >7 g/dl associated to systolic arterial pressure <90 mmhg or age over 70yrs or hr >100/min or ci <2.2 l/mim/m² or other associated disease 5.5 5.2 6 6.4* 5.6 5.6 8.5 9.9* <0.05 mortality (%) 4 6* 6 19* <0.05 conclusion. introduction of local restrictive transfusion guidelines was associated to a significant reduction in red cell transfusion during the postoperative period of heart surgery. the global morbidity and mortality rates in the whole group of patients were not affected. however patients who required blood transfusion following the restrictive strategy had a worse outcome. transfusion was probably more the consequence than the cause of this worse prognosis. if transfusion was the cause of the worse prognosis, then morbidity and mortality rates would have been higher among patients requiring transfusion during liberal period than in the whole group of patient. (1) hebert pc and col. a multicenter, randomized, controlled clinical trial of transfusion requirements in critical care. n engl j med 1999 ;340 :409-17. risk factors and outcome in european cardiac surgery higgins tl. quantifying risk and assessing outcome in cardiac surgery 3 1 intensive care medicine, hospital del mar, 2 servicio de microbiología to analyze if morbid obesity (mo) is associated with critical pathology in relation to patients undergoing vertical banded gastroplasty (vbg). all critical patients (cp) suffering mo, with a mean body mass index = 50,61, receiving programmed surgical treatment, and admitted in the icu during the next period: 1st oct. 96 to 25th feb. 02, were prospectively included. · surgery procedures. *restrictive: vbg according to masson's technic. *derivative: vbg + gip according to salmon's technic. vbg + gip according to capella's technic. ·type of study: descriptive. -vbg in association to other surgical procedures: 16 cp (16 cholecystectomies, 1 right inguinal herniorhaphy, and other 3 procedures). · mortality: 2 cp: -septic shock -multiorganic disfunction · readmission: 2 cp (subphrenic abcess and ards). complications 1. hypoxemia: 69 cp (60% of the total) 1.1. 1.1. not secondary to hypoventilation: 52 cp (72.5%). 1.2. 1.2. associated to hypoventilation: 52 cp (72.5%) 2. need for noninvasive mechanical ventilation: 25 cp (22%) 3. high blood pressure: 61 cp (52.0%). 4. disturbances of cardiac rythm and conduction: 8 cp (7%). 5. metabolic acidosis: 13 cp (11%). 6. other complications: 16 cp (14%). conclusion. 1-the mo patient undergoing vbg, with or without gip, rather than a patient bound to the reanimation or recovery room, is indeed a patient who requires admission in the icu for, at least, 24-36 hours. 2-hypertension of difficult management and hypoxemia not due to hypoventilation nor shunt are the most frequent complications. 3-an important percentage of cp requires also mechanical ventilation. 4-complications related to surgery are exceptional. karlicek a. 1 , haveman j. w. 1 , verhoeven e. 1 , van den dungen j. j. a. m. 1 , tielliu . i. f. n. 1 , hulsebos r. g. 1 , nijsten m. w. n. 1 1 surgery, groningen university hospital, groningen, netherlands introduction. the mortality in acute abdominal aortic aneurysms remains high. recent series still report a hospital mortality rate of more than 50% (1,2). despite the large number of published studies on hospital outcome, long-term outcome after icu admission has hardly been studied. here we present hospital survival and long-term outcome in 308 patients with an acute abdominal aortic aneurysm. the records of all patients operated for aneurysm surgery between 1990 and 2001 were retrospectively reviewed. in 308 patients surgery was performed for an acute abdominal aortic aneurysm. all operation reports were analysed. for complete follow-up the general practitioner was contacted if necessary. after arrival in the emergency department and confirmation of the diagnosis by physical examination and/or ultrasound all patients were immediately brought to the operation room. in our hospital even patients with cardiac arrest on arrival in the operation room are treated without delay, and were thus included in our study. all surviving patients were admitted at the intensive care. in case of postoperative haemodynamic instability, multiple organ failure, sepsis or diarrhea a sigmoidoscopy was performed to assess the presence of ischemia or infarction. three hundred and eight patients were operated for an acute abdominal aortic aneurysm, 266 men and 42 women. operative mortality was 13% (39/308). calculated from the moment of icu admission, 30 day survival was 78%. cumulative survival rates calculated with the kaplan meier method at 1, 3, 5 and 10 years were 65%, 55%, 46% and 28% respectively. in 23 patients in whom sigmoid resection was performed, 30 day survival was 35% compared to 83% in the other patients. mortality in ruptured abdominal aortic aneurysm remains high, 30 day survival was 78% in our group. sigmoid resection was associated with lower survival but sigmoidoscopy should be augmented to exclude sigmoid necrosis. outcome in these patients is not invariably poor. long term follow-up shows that also after discharge from the hospital these patients have a high mortality. carvalho a. g. r. 1 , gomes r. v. 1 , santos jr. b. 1 , barbosa o. n. 1 , weksler a. 1 , pontes a. p. 1 , camara a. c. 1 1 surgical intensive care unit, instituto nacional de cardiologia laranjeiras, rio de janeiro, brazil introduction. prognostic markers developed in europe and north america cannot be applied in latin america where life expectancy is 30 % lower according to world health organization. the objective of this study is to analyze patients profile submitted to heart surgery (hs), type of surgery distribution and the impact of variables, previously reported in medical literature, in the mortality and duration of intensive care unit (icu) stay in a public tertiary hospital. cohort study of patients submitted to hs from january 1998 to april 2001. patients profile, type of surgery distribution and many variables were analyzed. variables that were studied: age, gender, body mass index (bmi), body area (ba), preoperative in-hospital stay (preop), extracorporeal circulation (ecc) duration, ventricular function (vf), surgical indication, combined procedures (comb), urgency for the surgery, presence of diabetes mellitus (dm), systemic arterial hypertension (ah) and cigarette smoke (cs). the profile and patients variables were analyzed and compared in two different groups. group a (ga): patients discharged from icu or in-icu stay lower than or equal to 3 days (median in-icu stay in this study). group b (gb): death during icu admission or in-icu stay longer than 3 days. t-student, mann-whitney, chi-squared and fisher tests were used in the statistical analysis. conclusion. there is a rather singular distribution of surgeries in this group. many of the previously described variables showed correlation with mortality or longer admission in the icu. prospective studies will be held in order to adjust these variables and determine new ones more relevant to underdeveloped countries. pierce c. m. 1 , fortune p. 2 , petros a. j. 1 1 picu, great ormond street hospital, london, united kingdom, 2 picu, royal children's hospital, melbourne, australia there are anecdotal reports of sildenafil, a type 4 phospodiesterase inhibitor, being used to reduce pulmonary artery pressure in children with mainly cardiac induced pulmonary hypertension (pht). we have given oral sildenafil to 10 children on our paediatric intensive care units with pht from various causes. diaphragmatic hernia (3), avsd (2) vsd (1) pda (1), pphn (2) pulmonary hypoplasia (1). the median age of the group was 1m (iqr 1-6m). 7 were receiving inhaled nitric oxide during sildenafil. median dose was 0.5mg/kg (iqr 0.3-0.53mg/kg 8hrly) and duration was 7 days (iqr 3-29). pulmonary artery pressures were directly measured in 3 of the 4 cardiac children and deduced from doppler echocardiographic measurements of the tr jet in 5 children.results. pap decreased significantly (p<0.05) following oral doses of sildenafil (n=7). mean pulmonary/system (p/s) ratios decreased from 0.99 to 0.75 (n=8) within 3 hours of the oral dose. systemic pressure was unaltered in all children. in one child with pulmonary hypoplasia the p/s ratio was unaffected. oral sildenafil can significantly reduce raised pap in children when there is a reversible etiology. this may be particularly useful in children and neonate with pphn. central venous catheters (cvc) are an important means of securing intravascular access in pediatric intensive care unit patients. one of the major morbidity's in use of cvc is catheter-related infection (cri). the incidence of bacteremia with cvc use is approximately 5.3/1000 catheter days and mortality as high as 25%. one approach to reduce the incidence of cri has been to decrease catheter bacterial colonization (cbc). reduction in cbc is achieved by coating or impregnating antimicrobial substances into the catheter material. use of minocycline/rifampin treated catheters has been shown to reduce the rate of cri in critically ill patients. the concern in pediatric population is the use of minocycline. tetracycline and its derivatives (minocycline), when used in young children, carry the risk of dental and sceletal abnormalities. the problem of potential eluting of minocycline from minocycline-impregnated catheters may pose a risk for young children. our study examined whether detectable levels of minocycline and rifampin were present in the serum of the pediatric intensive care unit patients with indwelling minocycline/rifampin impregnated cvc. methods. patients admitted to pccu age 8-18 years and in need of cvc were eligible for study. six patients were enrolled. each patient had two samples of blood 2 and 2.5 ml withdrawn for rifampin and minocycline assays respectively. collection times were at the time of catheter insertion and 24h thereafter for seven days or until catheter removal, whichever came first. rifampin serum samples were processed prospectively soon after colection by standard hplc. minocycline serum samples were stored frozen in -80 centigrade and assayed in one batch using reverse phase hplc. results. demographic data are in table. ranges with mean values in ( ). none of the minocycline samples had detectable level of antibiotic. the limit of sensitivity for minocycline was 0.4 mg/l. therapeutic levels are 1.4-1.8 mcg/ml. one patient had 4 consecutive samples 3 to 6 with low therapeutic levels of rifampin (4-5 mcg/ml). therapeutic levels of rifampin are 4-32 mcg/ml. rifampin sensitivity was 1 mcg/ml. rifampin has distinct peak time and no interfering substances were identified.sex (3), the emergence of this consequences required fast and corrective treatment. an inotropic agents commonly used in vlbw infants such as dopamine and norepinephrine in some cases do not produce elevation in blood pressure despite using of very high dose. in this study i would like to exam the influence of hydrocortisone administration in vlbw infants with hypotension unresponsive to stndard catecholamine treatment. i have reviewed the cardiovascular response to hydrocortisone therapy in 16 preterm infants. mean gestation age was 27.1 (25-29) weeks, postnatal age 4.8 (1-9)days, mean birth weight 1070g (710-1420). eight of them suffered from respiratory distress syndrome and eight from sepsis. the first line of hypotension therapy was always volume administration (normal saline or albumine) and catecholamine infusion. hydrocortisone at the dose 2mg/kg was administered when dopamine at the dose 10mcg/kg/min (12 patients or norepinephrine 0.5mcg/kg/min (4 patients)failed to normalized arterial blood pressure. pneumococcal meningitis is an important cause of morbility and mortality in children. we describe the epidemiological characteristics and clinical features of pneumococcal meinigitis in children admitted to a children's hospital in barcelona. medical records of 32 children with a diagnosis of pneumococcal meningitis based on identification of s. pneumoniae in the blood or cerebrospinal fluid between january, 1993, to april, 2002, were retrospectively reviewed. results. 32 cases of pneumococcal meningitis were diagnosed in 29 patients. median age was 24 months (range 1.8 m-9.8 y). 24 children were younger than 5 years old (75%). male-female ratio was 1.2:1. none of the children had a previous immunological deficit. thirteen patients (41%) were pre-treated with antibiotics. the most frequent signs on admission were fever (100%), vomiting (72%), headache and irritability (34%), othalgia (22%) and shock (16%). neurological findings were lowered level of consciousness in 18 patients (52%), signs of meningismus in 24 patients (75%) and arreactive mydriasis in 3 patients (9%). the mean leukocyte counts in blood were 19164/mm3 and the mean c-reactive protein was 141mg/l. cerebral spine fluid indices on admission were: white blood cell=2301 (50-9800) /mm3; protein=148 (43-396) mg/dl; and glucose= 38 (0-262) mg/dl. main serogroups were: 18 (16%), 6 (13%), 4 (9%), 19 (6%), 23 (6%), 7 (3%), 14 (3%) and 9 (3%). overall, 50% of the pneumococcal isolates were penicilin-nonsusceptible, 28% cefotaxim-nonsusceptible and 3% were vancomycinnonsusceptible. an initial abnormal cranial computed tomography was found in 6 patients. the median duration of parenteral antibiotic therapy was 10 days. all patients were empirically treated initially with cefotaxime (associated to vancomycin in 22 of them). twenty-six patients (81%) received dexamethasone. the administration of mannitol was necessary in 6 patients (19%) and anticonvulsants were administrated in 8 patients (25%). only 8 patients (25%) needed inotropic support (no longer than 48 hours). mechanical ventilation was required in 15 patients (47%) during a mean of 1.5 days (range 0-10). acute complications were: metabolic acidosis (3/32), disseminated intravascular coagulopathy (6/32), seizures (7/32), siadh (3/32) and diabetes insipidus (2/32). twelve patients (37%) suffered deafness, three patients (3%) hemiparesia and four (12%) were exitus. the mean hospital stay was 18.5days and mean intensive care stay was 3.8days. there is an increased prevalence of pneumococco with decreased susceptibility to penicillin and to cefotaxime. deafness is one of the most common and serious sequelae of pneumococcal meningitis. corticotherapy has reduced the incidence of hearing loss. the new, antipneumococcal conjugated vaccine will confer effective prevention from the age of two months and will reduce the incidence of this meningitis. aims : to analize sedation/anesthesy methods used in our hospital for painfull or unconfortable procedures in children in relation to : 1)patient confort, 2)sedation complications, 3)and efficacy of the procedure a prospective study was conducted from january to march 2002 in 154 disconfortable procedures in 154 children. mean age was 40m ; their asa score was 4 in 7%, 3 in 16%, 2 in 57%, and 1 in 20%. more frequent procedures were : 54 lumbar punctures (lp), 23 thoracentesis or drainages, 21 central catheters insertion, endoscopys 18. we identified 3 different groups in relation to methods of sedation/analgesy : 1-procedures done in the emergency department with local anesthesia; 2-procedures done with administration of intravenous midazolan+ketamine; 3-procedures done with anesthesic support. we used the ramsay scale to classify the degree of anesthesia and the serna behavioral scale to classify the reaction to the procedure.results. group1(n=36):42%patients fighted against the procedure (serna scale 1) and in 25% of the patients, complications of the procedure were found to be related to inadequate sedation. group2 (n=67): in 10%, sedation was considered inadequate -serna level 1 (n=1) and 2 (n=5)-and in 1 case there were complications of the procedure related to unsufficient sedation; there were 7 (10%)cases of minor complications sedation-related; group3(n=51): patient confort and adequacy of the sedation were found in 100%, with 2 (4%) complications of the anesthesic method.conclusion. sedation/anesthesia were needed for the confort of the patients; only minor complications of sedation/anesthesy were found ; efficacy of the procedure was best achieved with the anesthesic method. introduction. the goals of emergency airway management are to antecipate and recognize respiratory problems and support therapy. the endotracheal intubation ( et) is not a routine procedure and it requests planning and personnel qualified to reduce the complications associated to this technique 1 . the purpose of this study is to evaluate early complications associated with endotracheal intubation methods. data were collected prospectively from february 1995 to january 2000 in tertiary teaching hospital. the variables were obtained in four age groups: group 1 (>1 month); group 2 (between 1 month to 11 months); group 3 (between 12 months to 144 months)and group 4 (>144 months). the data were collected as demographic data, reason for endotracheal (et) intubation, sedation administered, local of et, physician responsible for et, complications associated with airway management. the major complications were defined as technical problems that resulted increased morbidity. minor complications were incidents that should be avoided. the complications were compared between emergency or elective et intubation. statistical analysis by chi-square, fisher exact test . we evaluated 612 (45% female and 55% male) no consecutive patients. indication for intubation were: respiratory failure (31%), coma or depressed sensorium (7.5%), post-operative (45%) and shock (17%). sedation and/or analgesic were used in 54% of patients and 3.5% did not receive a sedative or analgesic for et intubation. a total 125 et emergency intubation ( we report an outbreak due to rsv in a 20 bedded picu with an annual admission rate of approximately 1000 patients, cardiac and medical patients accounting each for 40% of the population and 20% surgical.methods. an outbreak is defined as an event in which minimally 2 patients develop bronchiolitis due to rsv following transmission via hands of carers within a limited period of 1 week.nasopharyngeal aspirates were obtained from children with symptoms of lower airway infection, all samples were tested for rsv using the enzyme immuno assay, followed by tissue culture when the assay was negative. rsv positive children were isolated in cubicles and strict standards of hygiene were implemented. introduction.the objective of the study was to investigate the validity of outcome prediction after severe head injury using serum levels of protein s-100 b and of neuron specific enolase nse. methods.fifteen patients with severe head injury were included in this prospective study (9 men and 6 women) mean age 38 yrs (18-69). none of the above patients had spinal cord injury or any other neurological disease. venous blood samples were taken on admission and consecutively the 1, 2, 3, 4, and 5 day. immunoluminometric assay was used for the specimens. we tried to correlate the s-100 b and nse serum concentrations with the ct scan intacerebral pathology as well with the age, gender and outcome. results. all patients had elevated s-100 b and nse serum concentrations, with a gradual reduction towards the 5th day of icu stay. the mean values of day 1, for s-100 b were 2.4 ìg/l and for nse were 32.5 ìg/l. of day 5, they were for s-100 b 0.57ìg/l and for nse 14.9 ìg/l. patients who died had the first day mean values of s-100 b 4.2ìg/l and nse 47.2 ìg/l, whereas the survivors had mean values of s-100 b1.32 ìg/l and of nse 27.4 ìg/l (p <0.05). there was no strong correlation between the ct scan findings, the initial serum s-100 b and nse values and the gcs, on admission. conclusion. the protein s-100 b and nse are biochemical markers that seems to be elevated during the first days of injury, in patients with severe head trauma and could be used as markers of he severity of the injury. if protein s-100 b and nse could be used as a prognostic factor of the patient outcome, needs more investigation. our study is continued. estimates such as 28-day survival may be grossly misleading for assessment of intensive care utility. late mortality and morbidity may severely affect overall outcome. we studied 100-day survival rate in addition to survivors´ general health evaluation and prevalence of signs indicating post-traumatic stress disorder (ptsd). the setting is a university general intensive care unit. during the study period all adult patients who had been intubated and mechanically ventilated for at least 48 hours were included (patients who died before 48 hours are excluded). three to six months after their critical illness, survival data were retrieved from hospital and national registers. all patients surviving at this time were sent a health survey questionnaire (sf-36) and the post-traumatic stress syndrome 10-questions inventory (ptss-10). results. 153 patients fulfilled the inclusion criteria. the mean age was 69 years, 40% were women. health questionnaires were returned by 36 (58 %) of the 62 survivors at follow-up time. 28-day survival rate was 58 %, at 100 days survival rate had fallen to 46%. among the 36 responding survivors the frequency of a response pattern compatible with ptsd was 28%. survivors without signs of ptsd had sf-36 mean scores more than 1 standard deviation (sd) below the swedish norm in the domains of physical functioning, role-physical and social functioning. survivors with signs of ptsd scored below non-ptsd survivors in every domain, and were more than 3 sd below the swedish norm in the domains of social functioning, roleemotional and mental health. in total, there were only five persons (14 % of respondents) who scored at or above the swedish norm for both the physical and the mental health summary scales. assuming the same outcome in non-respondents this figure would correspond to about 6% of all the 153 included patients. conclusion. in this cohort of severely ill patients 28-day mortality was in the expected range but much mortality (another 12 %) occurs in the following 10 weeks, indicating a number of patients who have been subjected to long-lasting care with very meagre benefits. at 3-6 months following onset of their disease, survivors show considerably reduced subjective rating of their general health and life quality. as much as 28 % of the survivors show signs compatible with ptsd. it could be estimated that about 6 % of all patients included will both survive and within 3 -6 months reach a level of general health comparable to that of the general population. the aim of this study is to probe that critically ill patients gender is not associated with differences in severity of illness and related mortality. we had tested the premise in front of a controversial evidence offered by several years of our icu activity. observational study. retrospective analysis using data prospectively collected in a medical-surgical icu of 15 beds, in a teaching reference hospital, from november 1997 to july 2001. we analyzed 3152 consecutive admissions considering reason for admission, age, icu length of stay, severity of illness (mpm0, mpm24, saps ii and spanish version of apache iii) and related risk of death. cases were analyzed according to gender and age decades. therapeutic effort was analyzed according nems system. standarized mortality ratio (smr) and its 95% ci was determined. one thousand and twelve cases out of 3152 were women. mean age (sd) was 60 (17) years. significative differences were founded in mpm0 prognostic values (0.243±0.249 for men and 0.267±0.264 for women, p 0.003). the rest of epidemiological data do not offer significant differences. smr for men was 0.96, and for women 1.21, but 95% ci overlapped 0.86-1.07 vs. 1.04 -1.41, p ns. the same differences were found when different age intervals were analyzed. only 3 admission diagnostic (ischemic cardiopathy, post cardiopulmonar arrest and multiple trauma with no head trauma) showed greater mortality rates in women, but these differences disappeared when age intervals were considered. in spite of certain confusing data about greater mortality ratios in women admitted to our icu, accurate analysis does not show significant differences in severity of illness, associated prognosis and mortality, and therapeutic effort between male and female. bacterial infection is one of the most frequent and most feared complications in patients with a hematologic malignancy (phm). in a retrospective study, we found that bacteremia precipitating icu admission in phm was associated with a better outcome [1]. however, it remained unclear whether this finding could be extrapolated to all bacterial infections. the aim of this prospective study was to evaluate whether bacterial pneumonia (bp) and bacterial sepsis or other bacterial infections (bs) had a better outcome compared to non-bacterial or noninfectious complications (nbc) in critically ill phm.methods. 106 consecutive phm admitted to the icu over a 2 year period were categorized into bp (n=32), bs (n=24) or nbc (n=50) according to strict diagnostic criteria by an independent panel of physicians who were blinded for the outcome. the impact of bp and bs on the inhospital mortality was assessed by logistic regression after adjustment for severity of critical and underlying hematologic illness, duration of hospitalization before icu admission and other potentially important prognostic factors. two models were tested, the first using a classical severity of illness score (apache iii) and the second using a score system especially designed for cancer patients (groeger score) [2] . bacterial infection is one of the more favourable complications precipitating icu admission in phm and is associated with comparable mortality rates as in general icu patients. therefore, reluctance to admit phm to the icu for advance support is unjustified, especially when a bacterial infection is suspected to be the cause of deterioration. (1,2) the cleveland clinic score is the only one, to compile intraoperative data until the timepoint of icu admission.(3,4) we wanted to find out, whether the combination of pre and postoperative score, in alliance with additional parameters, improves the predictability of outcome. from 1995 from until 1999 adult cardiothoracic patients were examined. logistic regression was used for analyzing those variables, dealing with mortality. the selection of significant factors is based on a stepwise forward procedure(p<0,05). the accuracy of multivariate analysis is shown as roc(receiver-operator characteristic) curve. . 21 variables, pre as well as intraoperative parameters proved to be statistically significant in the analysis, 9 in the multivariate analysis: both scores, operation and aox time, preop at iii, assessment of intraop course, hb at icu admission, blood loss 24h<1155ml and rethoracotomy for bleeding. the pre and the postoperative cleveland clinic risk score were both statistically significant in the uni and multivariate analysis, but their combination improved roc. additional parameters had only little further impact. pre and postoperative cleveland clinic score are reliable in predicting the risk of cardiothoracic patients. adding further intra and postoperative data, risk stratification becomes more precise. the appearance of unexpected intraoperative difficulties was highly significant for adverse outcome. the collection of data should be continued on the icu and therapy should be reevaluated and modified any time. objective: to describe the frequency, etiologies, forms of presentation, and foci of bacteremia identified in patients admitted to the icu. prospective epidemiological surveillance study carried out from april 1998 to march 2001. bacteremia was defined as the isolation of a pathogenic microorganism in one or more blood samples. bacteremias were classified into contaminating or true according to clinical manifestations. a descriptive analysis of variables including mean values, ranges, and standard deviations is presented. a total of 332 episodes of bacteremia were identified, 228 of which were true bacteremias (27.8 episodes per 100 patients). the characteristics of patients with true bacteremia were as follows: mean (sd) age 64 (16.3) years; male sex 58.8%; mean apache ii score on admission 14.9 (5.8); and mean length of previous hospitalization 22 (27) days. in 153 (67.1%) cases, bacteremias were acquired in the icu and in 49 (21.5%) episodes were polymicrobial. a total of 289 pathogens were cultured. these included gram-positive cocci in 173 (59.9%) cases, gram-negative bacteria in 94 (32.5%), and fungi in 17 (5.9%). initial presentation included severe sepsis in 36 (15.8%) cases and septic shock in 51 (22.4%). the most frequent origin of intra-icu true bacteremias was unknown in 42.5% of cases (primary bacteremia) followed by catheterrelated bacteremia. crude mortality was 45.2% and bacteremia-related mortality 21.5%. primary bacteremia and catheter-related bacteremia were the most common. a total of 21.5% bacteremias were polymicrobial. gram-positive cocci were the predominant causative pathogens. gyurov e. g. 1 , milanov m. s. 1 , milanov s. g. 1 , neichev p. g. 1 1 general icu, emergency medicine hospital "pirogov", sofia, bulgaria intensive care units are unique because they house seriously ill patients in confined environments where antibiotic use is extremely common. since our last publication (1) there is a substantial rise in emergence of nosocomial infection namely gram-positive as well as changes of pattern of emergence. to study the frequency of emergence of nosocomial infection (nci) in intensive care unit (icu) we studied retrospectively data from case records and flow sheets of 1451 postoperative patients in our icu during 1999-2000 and compared data with last period. results. of 1451 patients in our icu during two years, we include those 613 (42.2%) who stayed for more than 72 hours. according to results from cultures we divided them to three groups. group one included 355 (57.9%) patients without bacterial growth. group 2 included patients with proved nosocomial infections /nci/. we obtained samples: 898 from urinary catheters (376 positive-41.9%), 552 from tracheal tube (457 positive-82.8%), 597 from blood (282 positive-47.2%), 64 intradermal segments from central venous lines (34 positive-53.1%), and 17 from sputum (15 positive-88.2%). the most common place for emergence of nci in our icu is respiratory tract. on 5-th icu day the tract became infected in almost 56% of the patients. the major role among pathogens played acinetobacter spp. (27.4%), citrobacter spp. (20.3%), p.aeruginosa (12% and serratia spp. (10%). the second place for emergence of nci is "reserved" for blood-stream infections. almost the half of the cultures (47.2%) showed bacterial growth. the isolated pathogens were the same: acinetobacter spp (19%), serratia spp. (16%), but there was substantial rise in frequency of emergence of s. epidermidis during the last years (see figure) . its frequency almost equalized that of acinetobacter spp. the other two main sources for nci were urine catheters and cv catheters. they remained on 3-rd and 4-th place. group 3 included patients with endogenous surgical wound infections. in this group we obtained samples from surgical wounds and drainages. in 1999 25.6% of cultures showed bacterial growth. during next 2000 this figure rose nearly twice (48.3%). the leading role played the same acinetobacter spp., citrobacter spp., p. aeruginosa, enterococcus spp. and e. coli. the role of s. epidermidis increased greatly during this period de waele j. j. h. c. 1 , hoste e. 1 , blot s. 1 , colardyn f. 1 1 icu, ghent university hospital, gent, belgium introduction. intra abdominal infections frequently complicate the postoperative course of patients with acute necrotizing pancreatitis. the objective of this study was to analyze the incidence of pancreatic surinfection after surgery for acute necrotizing pancreatitis, describe its characteristics and identify associated risk factors. we retrospectively (1995) (1996) (1997) (1998) (1999) (2000) (2001) analyzed 46 patients treated surgically for acute pancreatitis. surgical treatment consisted of debridement and postoperative continuous lavage. we recorded demographic characteristics, incidence of organ failure, data on surgical and infectious complications, data on surgical and medical treatment and disease severity by ranson and apache ii score. surinfection of the pancreatic necrosis was present in 30 out of 46 patients (65%). the surinfection was polymicrobial in 17 patients. most of the organisms were gram-negative (54%), the others were gram-positives (28%) or fungi (17%). patients with surinfected necrosis were younger (50 y vs. 64, p<0.01), had surgical complications more often (70% vs. 26.7%, p=0.01), needed retroperitoneal lavage for a longer time (28 days vs. 9, p<0.01), and had a longer hospital stay (84 days vs. 29, p<0.001) than patients without surinfection. multivariate analysis demonstrated that age (or 0.90; 95% ci: 0.83-0.97, p<0.01) and the occurrence of a surgical complication or 10; 95% ci 1.53-66.6, p<0.01) were independently associated with pancreatic surinfection. the mortality in patients with infected necrosis was higher (72% vs. 40%, p=0.04), although in multivariate analysis no association was found. pancreatic surinfection is high after debridement and retroperitoneal lavage, with mainly gram negative bacteria involved. surgical complications and younger age are significant risk factors for surinfection. the aim of this report is to describe the current status of sap in spanish icu's methods. sap cases are identified in accordance with generally accepted criteria in each icu, such as ranson, imrie, pcr and ct-dynamic criteria. sap was selected from the data base of the national study of spanish nosocomial infection monitoring (envin). this study covered the period from 1997 to 2000. envin is an observational, prospective and multicentre study. sap patients hospitalized during more than 24 hours in all the participating icu's have been included in the study. these patients were monitored until their discharge from the icu or up to a maximum of 30-60 days. secondary infections have also been monitored. severity is measured by means of apache ii. infections, mortality, epidemiological data and antibiotics used as a means of prevention are described. the statistical analysis used the chi x2 test for the association of qualitative variables, the student t for the comparison of averages and the 5% statistical significance level results. 199 patients (1.18%) of the 16,927 patients monitored by envin were found to have sap. the average apache ii was 14.9 and the average stay was 12.2 days. the base illness was medical (94.5%) and surgical (5%). 30.7% of the patients underwent emergency surgery. ni accumulated incidence was 48.7% and density incidence was 40/1000 hospitalization days. crude mortality was 31% and ni-related mortality was 45%. 97 infections were detected: 27 of abdominal origin (27.8%), 17 ventilator-associated pneumonias (18.5%), 12 secondary bacteremias related to abdominal infection (12.4%), 12 catheter-associated urinary tract infections 12.4%; primary bacteremias (10.3%); 3 central venous catheter-associated bacteremia (1.5%). a total of 66 pathogens were isolated. bgn 58.5%, cgp 24.5% (mrsa 4.2%), fungii 17% (principally candidas), enterococci 15% and anaerobes 3.7%. 85.4% of the sap patients received antibiotic treatment. the antibiotic most frequently used in prophylaxis was imipenem-cilastatine (70%) and piperaciline-tazobactam (10%). the antibiotics most frequently used in absolute indication were imipenem 70%, piperaciline/tazobactam 19.4%, metronidazol 14%, vancomicina 13.5%, ciprofloxacino13%, amikacina in 13% and fluconazol 7.6% conclusion. sap cases in spanish icu's account for little more than 1% of all hospital cases, but they result in high levels of severity, morbidity and mortality. crude mortality and sap septic complication-related mortality in spanish icu's are much higher than the average indicated in the literature (29.7% and 14.9%). imipenem is the antibiotic most frequently used in prophlaxis. the irruption of candidas has been detected. fluid /blood warmers help to prevent hypothermia by raising the temperature of intravenously administered fluids & blood. the hl-90 hotline fluid warmer is the model used in our hospital. it consists of disposable tubing set with a central channel through which the fluid is infused and outer tubing through which heated water circulates. the water is contained in a reservoir, which is heated by an electric element. the manufacturer's instructions recommend changing the water in the reservoir every 30 days. this water is a potential source of infection and we therefore sampled the water in the reservoir for microbiological contamination. this study was conducted at royal london hospital during the month of december 2001.there are 10 fluid warmers, all hotline in our operating theatres. samples of water were taken from each of the reservoirs at the end of the working day. using aseptic techniques 20 ml of water were added to a labeled blood culture bottle. each sample was cultured for 48 hours. after one week we repeated procedure results. after 48 hours of incubation, pseudomonas sp. grown in 7 out of 10 culture bottles. the results from the second sets also grew pseudomonas sp. in the same 7 out of 10 water reservoirs.conclusion. the water in the reservoir is heated to 41-42 degree celsius. this temperature does not inhibit the growth of pseudomonas. each time the disposable tube is disconnected from the reservoir approximately 5 ml of water is spilled potentially spreading microorganisms. in addition there are case reports of cracks/splits in the inner tubing of the disposable tubing potentially exposing infused fluid or blood to heated water from the reservoir (1). methods. 161 (17.26%) of 933 consecutive cardiac surgery patients operated at onassis cardiac surgery center, from january 1st to june 30th 2001, developed t>38.5 c and leucocytosis, without evidence of specific site of infection. those patients were examined for possible catheter related infection, by removing central and arterial catheters and sending them along with blood specimens for culture. infections within the first postoperative 48h were defined as early, whereas those developed after the first 48h were defined as late. we examined the relation between the incidence of catheter related infection and the type of microorganism isolated, the type of operation performed, the icu stay and the hospital mortality. . 610 coronary artery bypass grafting(cabg), 202 valve or ascending aorta replacement(vr), 86 combined(cabg+vr), 13 acute dissecting aneurysm(ada) and 22 other operations were carried out. positive blood or catheter cultures were found in 36 patients (3.85%). staphylococcus epidermidis was cultured from all patients with early(n=6) and 83% of those with late(n=30) infection, while candida was found in 30% of those with late infection. icu stay and hospital mortality was ten times higher in patients with positive blood or catheter cultures compared to the general icu population (21.4 vs 2.1 days and 27.7% vs 2.36%, respectively). finally, mortality was higher in patients with late compared to those with early infection(30% vs 17%). (pts) who had suffered traumatic brain injury (tbi) as well as the immune response of these pts. pts with moderate to severe tbi (gcs =<11) and age >17 were enrolled under the presupposition they remained on mechanical ventilation (mv) >4 days. a total of 35 tbi pts were followed-up; infected pts were identified and associated factors were studied. in addition, serum immunoglobulin (sig) levels and soluble interleukin-2 receptors (sil-2r) were measured in infected pts. c. albicans species. candidemia in icu patients is associated with a high mortality rate. c. albicans was the most common yeast isolated from blood. non-c. albicans species have a frequent occurence among candidemic icu patients. the moderate susceptibility of azoles against non-c. albicans species indicates the usefulness of susceptibility testing for antifungal treatment. prospective, cohort, observational, and multicenter study. urine cultures were performed once a week to all patients admitted to the icu. samples were processed at the different clinical microbiology laboratories of the participating hospital using specific culture medium (sabouraud) and the bactec technique and the a20c (biomerieux) system for the identification of species. candiduria was defined as < 104 cfu of candida spp. in the urine. frequencies are expressed as cumulative incidence (%) and incidence density (episodes per 1000 days of urinary catheter). . results. a total of 1765 patients admitted >7 days to the 70 participating icus between may 1998 to january 1999 were included in the study. of these patients, 1730 (98%) had a urinary catheter inserted, with 40,273 urinary catheter days. one or more candida spp. in the urine were detected in 389 patients. the rate of candiduria was 22 per 100 patients/icu and the incidence density 9.5 per 1000 days of urinary catheter. in 23 cases, candida spp. in association with different bacteria (5.9%) were found, mostly gram-negative pathogens (13 cases), in particular p. aeruginosa (n=5) and e. coli (n=3), and gram-positive pathogens (10 cases) especially enterococcos (n=7). in respect to candida spp., c. albicans predominated (68.4%) followed by c. glabrata (8.2%), c. tropicalis (3.6%), c. parapsilosis (2.3%), and c. krusei (1.3%), independently of the week in which isolation of pathogens was made. conclusions: candiduria was diagnosed in 22% of critically ill patients admitted for more than 7 days in the icu. candida albicans was the pathogen most frequently recovered (68.4%), although c. non-albicans was isolated in one out of each three cases. a retrospective study was done over the last year that included 19 neonatal patients admitted at the intensive care unit. all patients had congenital anomalies (6 patients with gastroschisis (31.5%), 3 patients with esophageal atresia (15.7%), and others with intestinal obstruction, duodenal atresia and malrotation). 90.9% of the patients were on total parenteral nutrition and mechanical ventilation. the average stay in the icu was 9.6 days. candida albicans was checked for in swabs of wound, in blood-culture, stool-culture, urine-culture, tracheal aspirate, gastric asp results candida albicans was identified in 11 patients (57.8%). it usually appeared 3-4 days after the introduction of the antibiotic therapy. it was most commonly found in gastric aspirate (63.6%), stool-culture (36.3%) etc. it would first appear in gastrointestinal tract (stool-culture and gastric aspirate after 8 days). in respiratory and urinary tract candida was identified after 12 days, and in blood-culture after 18 days. 54.5% of the patients received cephtriaxon or ampicillin, and 36.3% amikacin or gentamycin and metronidazol. morbidity in patients with yeast infection is very high. the most common causative agent is candida, and the predilection organ is digestive tract. risk factors are: prematurity, mechanical ventilation, total parenteral nutrition, longer hospital stay and widespectrum antibiotics. due to unspecific clinical picture early diagnosis is usually made according to the results of taken cultures. there are still many dilemmas regarding systemic antimycotic profilaxys. key: cord-022592-g7rmzsv5 authors: wynn, james l.; wong, hector r. title: pathophysiology of neonatal sepsis date: 2016-07-06 journal: fetal and neonatal physiology doi: 10.1016/b978-0-323-35214-7.00152-9 sha: doc_id: 22592 cord_uid: g7rmzsv5 nan a successful immune response is critically necessary to eradicate infectious challenges and prevent dissemination of the infection in the host. however, if inflammation is not limited and becomes generalized, it can result in the constellation of signs and symptoms of a systemic inflammatory response syndrome (sirs). if the infection is not contained, the spread of the pathogen from its local origin through the blood may result in systemic endothelial activation and precipitate sepsis, severe sepsis, and septic shock. progression of sepsis to shock may lead to multiple organ dysfunction syndrome (mods) and ultimately death. host immunity is divided into innate and adaptive immune systems for purposes of discussion and teaching but there is a great deal of interaction between the two systems. innate immunity is rapid, largely nonspecific, and composed of barriers, phagocytic cells, the complement system, and other soluble components of inflammation. after breech of a barrier, cellular elements of the innate immune response are the first line of defense against the development and progression of infection. adaptive immunity, which is antigen specific, is long lived, and often takes several days to develop, provides immunologic specificity and memory. these systems work together to protect the host from pathogenic challenge but may also precipitate host injury through aberrant responses. the outcome of infection is dependent on at least four major factors: (1) the pathogen, (2) the pathogen load, (3) the site of infection, and (4) the host response. less is known about the host response in neonates compared with adults for a number of reasons, the principal one being a highly variable definition of disease. our understanding of the pathophysiology of sepsis is largely from investigations in adult populations, including both humans and animals. there is clear evidence from both preclinical models of sepsis and humans that neonates manifest different host immune responses as compared with adults. [1] [2] [3] [4] even in comparison with children, neonates manifest a unique host immune response to septic shock. 5 thus neonatal-specific clinical investigations, particularly in very preterm infants, are required to improve both survival and long-term outcomes for these populations. a better understanding of the pathophysiology will uncover new opportunities for interventional studies ultimately aimed at improving outcomes. to this end, in this chapter we explore the pathophysiology of sepsis in the neonate, with special attention paid to the immunobiology of sepsis. adult and pediatric intensivists currently use generally accepted definitions for sepsis for goal-based therapeutic interventions. [6] [7] [8] [9] these definitions are critical to facilitate epidemiologic studies, to accurately determine disease prevalence, to select patients for clinical trials, and ultimately to improve the delivery of care. the generally accepted pediatric definition for sepsis, established in 2005, was intended for all children (<18 years old), including term neonates (≥37 weeks' completed gestation). 6 preterm neonates (<37 weeks' completed gestation) were specifically excluded from the pediatric generally accepted definitions, and neonatal-perinatal subspecialists were not represented among the pediatric consensus experts. to investigate whether the pediatric generally accepted definitions for sirs and sepsis applied to term infants, hofer and colleagues 10 retrospectively examined 476 term neonates and found that the generally accepted definitions applied to only 53% of cases of culturepositive early-onset sepsis. neonatal sepsis has been inconsistently defined on the basis of a variety of clinical and laboratory criteria, which makes the study of this condition very difficult. 11 diagnostic challenges and uncertain disease epidemiology necessarily result from a variable definition of disease. the lack of a generally accepted definition for neonatal sepsis remains a significant hindrance towards improving outcomes and accurately describing disease pathophysiology. thus working definitions for the sepsis continuum, specific for preterm and term neonates, are needed to provide a uniform basis for clinicians and researchers to study and diagnose severe sepsis. the addition of immune biomarker-based staging of disease to clinical sign staging is highly likely to increase the accuracy of patient classification for future multicenter clinical trials that will test novel interventions. sepsis or serious infection within the first 4 weeks of life kills more than 1 million newborns globally every year. 12, 13 the incidence of neonatal sepsis is variable (from less than 1% to more than 35% of live births) on the basis of gestational age and time of onset (early-onset sepsis [<72 hours after birth] or late-onset sepsis [≥72 hours after birth]). [14] [15] [16] [17] [18] [19] [20] preterm neonates have the greatest sepsis incidence and mortality rates among all agegroups [21] [22] [23] [24] [25] [26] (figure 152-1) . risk factors for developing sepsis in neonates, particularly the very premature, have been well described. 14, 15, [27] [28] [29] [30] [31] [32] [33] prematurity, low birth weight (especially infants weighing less than 1,000 g), male sex, a maternal vaginal culture positive for group b streptococcus (gbs), prolonged rupture of membranes, maternal intrapartum fever, and chorioamnionitis are strongly associated with an increased risk for early-onset sepsis. 30 chorioamnionitis is associated with the greatest risk for subsequent clinical or culture-proven sepsis. 29 recent studies demonstrate the risk for sepsis in newborn infants born to women with clinical chorioamnionitis is strongly dependent on gestational age, with minimal risk in neonates aged 35 weeks or older and greater risk with increasing degrees of prematurity. [34] [35] [36] [37] [38] [39] [40] [41] the risk for neonatal sepsis conferred by maternal gbs colonization 29 is significantly reduced with adequate intrapartum antibiotic prophylaxis. 42 section xxvi pathophysiology of neonatal diseases ated with sepsis-like syndromes (e.g., echovirus, enterovirus, parechovirus, coxsackie virus, adenovirus, parainfluenza virus, rhinovirus, and coronavirus). 50, [63] [64] [65] physical barriers, including skin and mucosal surfaces, are the first point of contact between the host and potential pathogens. thus a successful immune defense in addition to epithelial barrier function is critical to prevent the development of local infection. multiple immune elements are present to prevent attachment and propagation of pathogens while simultaneously permitting the presence of commensal organisms required for homeostasis. vernix enhances skin barrier function in late-preterm and term neonates. vernix is a complex material comprising water (80.5%), lipids (10.3%), and proteins (9.1%) produced by fetal sebaceous glands during the last trimester 66 and is largely absent in preterm neonates born before 28 weeks' gestation. vernix provides a barrier to water loss, improves temperature control, and serves as a shield containing antioxidants and innate immune factors such as antimicrobial proteins and peptides (apps). 67 the apps on the surface of the newborn's skin (and replete in the amniotic fluid [68] [69] [70] ) are capable of killing/inactivating common neonatal pathogens, including gbs, e. coli, and candida species. 71 erythema toxicum is an immune-mediated manifestation that results from bacterial colonization of the skin occurring shortly after birth. 72, 73 this common cutaneous immune response is less common in preterm infants than in term infants, highlighting the impact of developmental age on host immune capabilities. 74 in contrast to the moist mucosal surfaces of the respiratory and gastrointestinal (gi) tracts, the skin is arid, which further reduces the chances for microbial invasion. the outermost layer of the skin, the stratum corneum, prevents microbial invasion, maintains temperature, and reduces the risk for dehydration through prevention of transcutaneous water loss. 75 the immature and incompletely developed stratum corneum of preterm newborns takes at least 1 to 2 weeks after birth to become fully functional 76 and may take up to 8 weeks in the extremely preterm neonate, significantly increasing the risk for barrier dysfunction. 77 disruption of the cutaneous barrier by trauma (e.g., placement of an intravenous catheter or heel stick) or chemical burn allows microorganisms to enter the subcutaneous tissue, increasing the likelihood of their establishing a local infection ( figure 152 -2). the likelihood of a microbial breach of the cutaneous barrier rises in the presence of intravenous catheters, which are essential for critical care. emollients, aimed at enhancing the barrier function of preterm newborn skin, increase the risk for nosocomial infection and their use is not recommended. 78 mucosal barriers contain multiple components that serve to prevent infection, including acidic ph, mucus, cilia, proteolytic enzymes, apps, opsonins such as surfactant proteins, sentinel immune cells such as macrophages, dendritic cells, polymorphonuclear neutrophils (pmns), and t cells, as well as commensal organisms. 79 like the skin, the gi mucosa is quickly colonized after birth and contains a significant repository of microorganisms. [80] [81] [82] gi barrier integrity, paramount for prevention of spread of microorganisms out of the intestinal compartment, is dependent on the interaction between commensal organisms and host epithelium. interleukin (il)-17, produced by type 3 intestinal innate lymphoid cells in the presence of the microbiota, drives granulocytosis and may protect the neonatal host from infectious challenge. 83 a loss of intestinal barrier integrity likely plays a role in the development of necrotizing enterocolitis (nec) and late-onset sepsis. 84, 85 prolonged antibiotic treatment, hypoxia, and remote infection are factors known to despite the efficacy of this intervention, the incidence of invasive gbs disease in african american neonates is still more than twice that in white babies, 43 and the incidence of escherichia coli sepsis may be rising in very-low-birth-weight (vlbw) neonates. 44 vaginal delivery in the presence of maternal active primary herpes simplex virus significantly increases the risk for a neonatal herpes simplex virus infection, which has a fulminant course and high mortality. [45] [46] [47] preexisting maternal immunodeficiency or sepsis also increases the risk for sepsis in the neonate. 48 in addition, care practices after birth, such as intubation, mechanical ventilation, and placement of central venous lines, increase the risk for the development of sepsis. 49 a number of pathogens have been associated with sepsis in the neonatal period. the predominant cause is bacterial; however, certain viral infections are associated with a fulminant course and significant mortality. [50] [51] [52] in a large (n = 104,676), multicenter study of vlbw infants (<1500 g), gram-positive organisms accounted for 34% of pathogens causing early-onset sepsis and 61% of those causing late-onset sepsis. in contrast, gram-negative organisms were responsible for 58% of early-onset sepsis and 26% of late-onset sepsis. 53 candida species accounted for 3% of cases of early-onset sepsis and 11% of cases of late-onset sepsis. infection by gram-negative organisms, particularly pseudomonas species, carries a higher risk for fulminant course and death than infection by other pathogen groups. 14, 15, 49, [54] [55] [56] gram-positive causes of sepsis are dominated by gbs and coagulase-negative staphylococci (cons). 15, 57 although the high mortality rate for gbs has been well described (especially among infants born prematurely), mortality rates associated with cons are significantly lower. 15, 16 fungi may also be associated with fulminant neonatal sepsis and predominantly affect vlbw infants. 15, 58, 59 independent predictors of in-hospital neonatal mortality after late-onset sepsis were pseudomonas infection (adjusted odds ratio [or], 14.31; 95% confidence interval [ci], 3.87% to 53.0%) and fungemia (or, 5.69; 95% ci, 2.48% to 13.01%). 60 the limited sensitivity of current methods to identify causative organisms is partially due to an inability to take a large sample of blood from newborn infants with suspected sepsis. 61 blood culture-negative ("clinical") sepsis is estimated to occur at a nearly 10-fold greater rate than blood culture-positive sepsis. 62 in some of these infants, sepsis may also be due to novel viral pathogens associy e a r s 3 0 -3 9 y e a r s 4 0 -4 9 y e a r s 5 0 -5 9 y e a r s 6 0 -6 9 y e a r s 7 0 -7 9 y e a r s 8 0 -8 9 y e a r s the production of mucus and mucociliary clearance of pathogens and debris. 95 premature neonates have relatively more goblet cells than do maturer neonates, leading to a decrease in mucociliary clearance. respiratory mucosal function can be impaired by surfactant and saliva deficiency, altered mucus production, and mechanical ventilation. ventilation is associated with decreased mucociliary clearance, airway irritation, and parenchymal lung injury (see figure 152 -2). intubation is also associated with the progressive accumulation of colonizing bacteria and bacterial endotoxin in respiratory fluids, with concomitant mobilization of endotoxin-modulating apps to the airway. 96 neonates with surfactant deficiency lack apps such as surfactant proteins a and d, which are also absent in commercially available surfactant preparations. 97 there is an age-dependent maturation in the ability of respiratory epithelium to elaborate apps (cathelicidin and β-defensins), such that the respiratory epithelium of preterm newborns mounts a deficient app response. 98 disrupt or injure the neonatal intestinal barrier (see figure 152 -2). [86] [87] [88] under these circumstances, the gut may become the motor of systemic inflammation. 89 mechanistically, paneth cells and intestinal lymphoid cells may release excessive amounts of il-17, which, in turn, plays a critical role in the development of sirs. 90 many interventions aimed at reducing the frequency of sepsis in neonates via enhancement of mucosal barrier integrity have been evaluated. neither probiotics nor glutamine supplementation has reduced the incidence of neonatal sepsis. 91 in contrast, human milk feeding is associated with a reduction in the risk for sepsis 92 and nec 93, 94 and is strongly encouraged, especially in preterm infants. respiratory mucosa is defended in utero by amniotic fluid and pulmonary apps, surfactant proteins a and d, alveolar macrophages, and pmns, among other immune elements. the surface and submucosal gland epithelium of the conducting airways is a constitutive primary participant in innate immunity through c the cell surface and in endosomes, whereas rlrs and nlrs detect pathogens only intracellularly. the discovery that tlr4 was integral for a robust lipopolysaccharide (lps)-mediated inflammatory response after gram-negative sepsis may be why tlrs have been more thoroughly investigated in the setting of sepsis than other prrs. 100 each of the 10 known tlrs in humans, present on and within multiple cell types, recognizes extracellular and intracellular pathogens via specific pamps. 101, 102 multiple tlrs may be activated in concert by intact or partial microorganisms and in turn activate multiple second-messenger pathways simultaneously. 102, 103 lps is the prototypic mediator of systemic inflammation and generates many of the clinical findings of sepsis and septic shock, including mods and death. 104 lps signals through tlr4 in conjunction with the adaptor proteins cd14 and myeloid differentiation factor 2. 105 in adults a reduction in mortality and improvement in hemodynamics were demonstrated when the level of serum lps was reduced. 106 the level of lps is elevated in blood from infected neonates and those with nec even in the absence of gram-negative bacteremia. 84 high levels of circulating endotoxin found during sepsis and nec are associated with multiorgan failure, thrombocytopenia, neutropenia, and death. 84 administration of anti-lps antibodies to a small number of these deficiencies as well as those related to cellular function in combination with invasive procedures lead to a reduction in respiratory barrier function that increases the risk for sepsis. once the local barrier function has been compromised, pathogen recognition by local immune sentinel cells is the first step towards the development of an immune response (figure 152-3) . elegant sensing mechanisms have evolved to facilitate detection of potentially pathogenic microorganisms. multiple classes of pathogen recognition receptors (prrs) have been discovered that serve as detectors of pathogen-associated molecular patterns (pamps), including cell wall and membrane components, flagellum, nucleic acids, and carbohydrates. 99 a litany of prr classes have been discovered, including the toll-like receptors (tlrs), nod-like receptors (nlrs), retinoic acid-inducible protein i like receptors (rlrs), peptidoglycan recognition proteins, β 2 -integrins, and c-type lectin receptors. the tlrs, β 2integrins, and c-type lectin receptors detect pathogens both on neonates with sepsis (n = 16) with serum endotoxin present reduced the time to recovery but not mortality as compared with the values in placebo-treated neonates. 107 reduction of serum lps levels by exchange transfusion in infected neonates (n = 10) was associated with improved survival. 108 bacterial cell wall components (such as lipoteichoic acid) signal primarily through tlr1, tlr2, and tlr6, flagellin signals through tlr5, and cpg double-stranded dna signals through tlr9. common viral pamps such as double-stranded rna or single-stranded rna signal through tlr3, and tlr7 and tlr8, respectively. agonist-tlr binding results in a signaling cascade of intracellular second-messenger proteins ultimately leading to production of cytokines and chemokines, as well as activation of other antimicrobial effector mechanisms. 101 signaling through tlrs typically leads to the production of nuclear factor κb (nf-κb)-dependent inflammatory cytokines and chemokines, whereas signaling through toll/il-1 receptor-domain-containing adapter inducing interferon (ifn)-β (trif) induces production of type i ifns, as well as nf-κb-related inflammatory cytokines. in neonates of all gestational ages, up-regulation of tlr2 and tlr4 messenger rna (mrna) occurs during gram-positive and gram-negative infection, respectively. 109 dysregulation or overexpression of tlr4 is involved in the development of nec in experimental animal models, 110 implicating the importance of tlrs in the initial immune response to pathogens and their role in neonatal sepsis. other important intracellular prrs include nlrs and rlrs. for nlrs, multiple cytosolic proteins are able to act as pamp sensors (e.g., nlrp1, nlrp3, and nlrc4) and coalesce with adaptor proteins and procaspase 1 to form a multimeric protein complex termed the inflammasome. 111 the formation of the inflammasome results in the conversion of procaspase to active caspase 1, which cleaves the inactive precursor proteins il-1β and il-18 to their active forms. 111 rlrs are cytoplasmic rna helicases that, like tlr3, sense double-stranded rna of viral origin and induce type i ifn production and nf-κb activation. 102 to date, the impact of rlr and nlr signaling has not been specifically examined in neonates with sepsis. in addition to its roles in leukocyte function (adhesion, phagocytosis, migration, and activation) and complement binding, complement receptor 3 (cr3, also known as mac-1 and cd11b-cd18) functions as a pathogen sensor on the surface of phagocytes. cr3 binds lps, as well as a broad range of other microbial products, in cooperation with or independently of cd14, leading to up-regulation of inducible nitric oxide (no) synthase and no production. 112 diminished expression of l-selectin and cr3 on stimulated neonatal pmns impairs activation and accumulation at sites of inflammation. 99, 113, 114 decreased expression of l-selectin and cr3 persists for at least the first month of life in term infants, possibly contributing to an increased risk for infection. 115 the expression of cr3 (cd11b) may be reduced further in preterm neonates as compared with term neonates. 116 in umbilical cord blood from neonates of less than 30 weeks' gestation, pmn cr3 content was similar to levels found in patients with type 1 leukocyte adhesion deficiency (failure to express cd18). 113, 114 thus decreased leukocyte cr3 surface expression increases the likelihood of suboptimal pathogen detection and cellular activation, particularly in the preterm neonate. c-type lectin receptors are prrs that recognize bacterial, viral, fungal, and parasitic carbohydrate moieties. c-type lectin receptors may be expressed on the cell surface (e.g., macrophage mannose receptor, mincle receptor, dectin 1, and dectin 2) or secreted as soluble proteins (e.g., mannose-binding lectin [mbl], (which is also named mannan-binding protein or mannanbinding lectin) as one of the acute-phase reactants. once bound to its carbohydrate ligand, mbl initiates activation of complement via the lectin pathway to promote opsonization and phagocytic clearance of pathogens. plasma mbl concentrations are low at birth (especially in preterm infants) but rise steadily throughout infancy and childhood. 117 low levels of mbl are associated with the increased incidence of sepsis in neonates. [118] [119] [120] in addition to decreased concentrations at birth, certain genetic polymorphisms of mbl (namely, mbl2), have been associated with an increased risk for infection in some, 121 but not all, studies. 122-124 m-ficolin activates the complement system in a manner similar to mbl and its level is elevated in neonates with sepsis. 125 prr stimulation results in rapid inflammatory mediator transcription and translation directed at cellular activation and clearance of pathogenic organisms 126 (see figure 152 -3). during sepsis and septic shock, multiple proinflammatory cytokines have been identified, including il-1β, il-6, il-8 (cxcl8), il-12, il-18, ifn-γ, and tumor necrosis factor (tnf)-α. 127 compared with adults with sepsis, neonates with sepsis produce less il-1β, tnf-α, ifnγ, and il-12. [128] [129] [130] [131] [132] [133] the decreased cytokine production is due in part to decreased production of important intracellular mediators of tlr signaling, including myeloid differentiation factor 88, ifn regulatory factor 5, and p38, which exhibit gestational age-specific decrements. 134 recent studies have demonstrated impaired inflammasome activation and mature il-1β production by neonatal mononuclear cells. 121, 135 in a comprehensive study (>140 analytes) of serum from neonates evaluated for late-onset sepsis, il-18 emerged as a predictive biomarker to differentiate infected neonates from uninfected neonates. 136 il-18 reduces pmn apoptosis, 137 drives ifn-γ production, 138 and induces production of tnf-α, il-1β, and cxcl8. 139 il-18 primes pmns for degranulation with production of reactive oxygen intermediates on subsequent stimulation. 140 dysregulation of many of these functions linked to il-18 are seen in sepsis and septic shock. increased il-18 levels have been demonstrated in premature neonates with brain injury 141 and also in an experimental model of nec, [142] [143] [144] highlighting activation pathways common with those in ischemia and inflammation. excessive levels of il-1β, tnf-α, il-6, cxcl8, il-10, and il-18, such as those seen with advanced-stage nec, severe sepsis, or septic shock, correlate with poor survival. 84, [145] [146] [147] [148] altered cytokine levels (increased il-10 and il-6 levels and decreased ccl5 levels) may identify those neonates at highest risk for the development of sepsisassociated disseminated intravascular coagulation (dic). 149 proinflammatory cytokine production leads to activation of endothelial cells, including increased expression of cell adhesion molecules that facilitate leukocyte recruitment and diapedesis ( figure 152-4) . up-regulation of cell adhesion molecules (soluble intercellular adhesion molecule, vascular cell adhesion molecule, l-selectin, p-selectin, e-selectins, and cd11b-cd18) during sepsis facilitates rolling and extravascular migration of leukocytes. [150] [151] [152] [153] decreased production of l-selectin and expression of c3 in pmns and monocytes derived from neonates may impair accumulation at sites of inflammation. 113, 114 chemokine gradients produced by endothelial cells and local macrophages are necessary for effective and specific leukocyte attraction and accumulation (see figure 152 -4). without adequate leukocyte recruitment, there is increased risk for propagation from a localized to a systemic infection. although poor cellular chemotaxis in the neonate has been observed, it is not likely a result of reduced serum concentrations of chemokines as baseline levels are similar in preterm and term neonates as compared with adults. 154 suboptimal cellular chemotaxis may be related to other mechanisms, such as poor complement receptor translocation. 162 the role of hmgb-1 and rage signaling in human neonates with sepsis has not been well characterized but has been shown to be involved in the pathophysiology of nec in an experimental model. 163 significantly lower soluble rage levels were found in human fetuses that mounted robust inflammatory responses and hmgb-1 levels correlated significantly with the levels of il-6 and s100β calcium-binding protein in the fetal circulation. 164 other specific damage-associated molecular patterns, including heat shock proteins and uric acid, may also stimulate tlrs, regulate pmn function, and serve as immune adjuvants. heat shock protein production in infected neonates has not been evaluated but polymorphisms in heat shock proteins increase the risk for acute renal failure in preterm neonates. 165 the levels of heat shock proteins are significantly elevated in infected adults and children. 166 elevated heat shock protein 60 and heat shock protein 70 level measured within 24 hours of pediatric intensive care unit admission were associated with septic shock and there was a strong trend towards increased mortality. 167, 168 uric acid can increase cytokine production, pmn recruitment, and dendritic cell stimulation 169 and may also serve as an antioxidant. 170 the level of uric acid is reduced in the serum of neonates with sepsis as compared with control neonates. 171 in addition to facilitating leukocyte attraction, proinflammatory stimuli result in production of vasoactive substances that decrease or increase vascular tone and alter vascular permeability (see figure 152 -4). these include platelet-activating factor, thromboxane, leukotrienes, no, histamine, bradykinin, up-regulation after stimulation, 99 deficiencies in another downstream signaling process, 155 or inhibition by bacterial products. 156 the levels of a wide variety of chemokines are increased during sepsis, including cxcl10 (ip-10), ccl5 (rantes), ccl2 (monocyte chemoattractant protein 1), ccl3 (macrophage inflammatory protein 1α), and cxcl8. 157 the levels of other chemoattractive molecules also increase with sepsis, including complement proteins c3a and c5a, apps, including cathelicidins and defensins, and components of invading bacteria themselves. 127, 136 the importance of chemoattractive substances in the pathogenesis of severe sepsis is highlighted by studies showing that cxcl8 can be used as a stratifying factor for survival in children, 158 and c5a is implicated in sepsis-associated organ dysfunction in adults. 104 chemokine investigations in infected neonates revealed that cxcl10 is a sensitive early marker of infection, 157 and low ccl5 levels may predict development of dic. 149 damage-associated molecular patterns (or alarmins), such as intracellular proteins or mediators released by dying or damaged cells, may also active prrs. for example, the damage-associated molecular pattern high-mobility group box 1 (hmgb-1) is involved in the progression of sepsis to septic shock in adults. 104, 159 macrophages or endothelial cells stimulated with lps or tnf-α produce hmgb-1, which signals through tlr2, tlr4, and receptor for advanced glycation end products (rage). 160 hmgb-1 results in cytokine production, activation of coagulation, and pmn recruitment. 159,161 hmgb-1 mediates disruption of epithelial junctions within the gut via the induction of reactive nitrogen intermediates, leading to increased bacterial a twin study which assessed the frequency of infections among monoygotic and dizygotic prematurely born twins concluded that 49.0% (p = .002) of the variance in susceptibility to lateonset sepsis was due to genetic factors alone. 215 the impact of genetics in the host response is also underscored by the increased risk for death from infection seen with african american race or male sex among low-birth-weight infants. 216 an ethnically unique single nucleotide polymorphism in the tlr4 promoter region was significantly associated with gram-negative bacterial infections in preterm infants. 217 several recent studies in newborn infants have demonstrated an association between small variations in dna, specifically single nucleotide polymorphisms, and infection development and outcomes. 122, [218] [219] [220] [221] [222] because tlrs play an essential role in recognition and response to pathogens, alterations in their expression, structure, signaling pathways, and function can have consequences for host defense. polymorphisms or mutations in tlrs are associated with increased risk for infection in adults [223] [224] [225] [226] and children 210, 227, 228 but are less well characterized in neonates. after confounders had been controlled for, the presence of a tlr4 single nucleotide polymorphism was associated with a three-fold increase in the risk for gram-negative infections in vlbw infants. 222 polymorphisms in the tlr2, tlr5, il10, and pla2g2a (which encodes pla 2 ) genes were associated with the development of neonatal sepsis. 218 modifications in expression or function of costimulatory molecules necessary for tlr activation are also associated with an increased risk for infection. for example, the levels of lpsbinding protein (lbp; which binds intravascular lps) and the lps coreceptor cd14 are both increased during neonatal sepsis. 211, 229, 230 furthermore, genetic variations in these proteins have been associated with increased risk for sepsis in adults. [231] [232] [233] genetic polymorphisms in myeloid differentiation factor 2, a small protein involved in lps signaling through tlr4, increase the risk for organ dysfunction and sepsis in adults 234 but the significance in neonates is unknown. polymorphisms in post-tlr activation intracellular signaling molecules, including myeloid differentiation factor 88, 235 il-1 receptor-associated kinase 4, 236 and nf-κb essential modulator, 237 are associated with invasive bacterial infection in older populations. additional genetic polymorphisms in intracellular second-messenger inflammatory signaling systems with impact on neonatal sepsis risk and progression are likely to be uncovered with the implementation of biobanking and mining of stored samples. mutations have been identified in nlrs that are involved in the pathogenesis of crohn's disease (nod2) 238 and neonatalonset multisystem inflammatory disease (nlrp3). 239 rlr mutations have been identified but have unknown clinical significance. 240 no mutations in specific domains of nlrs have been found in neonates with sepsis or nec. 220, [231] [232] [233] [241] [242] [243] [244] [245] [246] [247] [248] [249] [250] [251] [252] the importance of nlrs in listeria monocytogenes infections in neonates is unknown. complement is an important component of early innate immunity that facilitates killing of bacteria through opsonization and direct microbicidal activity. complement components also possess chemotactic or anaphylactic activity that increases leukocyte aggregation and local vascular permeability. furthermore, complement reciprocally activates a number of other important processes, such as coagulation, proinflammatory and prostaglandins. 172, 173 these substances are produced predominantly by host endothelium and mast cells. activated pmns produce phospholipase a 2 (pla 2 ), the level of which is increased in the serum of neonates with sepsis 174 and leads to generation of vasoactive substances, including prostaglandins and leukotriene. thromboxane produced by activated platelets and endothelin 1 produced by activated endothelium 175 are potent vasoconstrictors that participate in the development of pulmonary hypertension. [176] [177] [178] [179] overproduction of cytokines and vasoactive substances is associated with circulatory alterations and organ failure seen in severe sepsis and septic shock ( figure 152-5) . 6, [180] [181] [182] [183] if the pathogen is not contained locally and inflammatory homeostasis is not restored, sirs may develop, and lead to mods and death (see figure 152 -5). 184 the traditional paradigm for understanding the host response to sepsis consists of an intense proinflammatory response, or sirs, temporally followed by a compensatory antiinflammatory response syndrome. this paradigm has been challenged by the failure of multiple antiinflammatory strategies to improve sepsis outcomes in adults. 185 new data in adults and children demonstrate simultaneous proinflammatory/antiinflammatory responses where the magnitude of either response may determine outcome. 186, 187 near simultaneous increases in antiinflammatory cytokine production (transforming growth factor β, il-4, il-10, il-11, and il-13) occur in neonates during infection, countering the actions of proinflammatory cytokines. 127, 188, 189 these mediators blunt the activation and recruitment of phagocytic cells, reduce fever, modify coagulation factor expression, and decrease production of reactive oxygen and nitrogen intermediates, no, and other vasoactive mediators. [190] [191] [192] [193] [194] [195] soluble cytokine and receptor antagonists produced during sepsis also modulate proinflammatory mediator action. elevation of the levels of tnf receptor 2 (which regulates the concentration of tnf-α), soluble il-6 receptor, soluble il-2, and il-1 receptor antagonist have been documented in neonatal sepsis with resolution after effective treatment. 189, 196, 197 the role of these regulatory cytokine inhibitors in the immune response to neonatal sepsis and septic shock has been incompletely characterized. soluble rage competes with cell-bound rage for the binding of hmgb-1 and other rage ligands. 198 soluble rage has antiinflammatory effects and its level is elevated in adults during sepsis. 199 furthermore, soluble rage improved survival and reduced inflammation when given to infected adult rodents. 200 serum soluble triggering receptor expressed on myeloid cells 1 may reduce inflammatory signaling for triggering receptor expressed on myeloid cells 1, and predict mortality in preterm neonates. 201 micrornas may regulate inflammation at the level of gene expression via several putative mechanisms. 202 several pilot studies in rodents and humans have demonstrated regulatory functions for microrna in neonates. [203] [204] [205] [206] [207] [208] the impact of regulatory micrornas and their effects on the host inflammatory response in neonates with sepsis are unclear. endogenous cortisol is induced by proinflammatory cytokines and attenuates the intensity of sirs associated with severe sepsis and septic shock. 209 the use of cortisol in adults with sepsis has been controversial. 210, 211 cortisol production in newborn infants is significantly increased early in shock. 212 however, very preterm neonates may have relative adrenal insufficiency that may contribute to hemodynamic instability and hypotension. cortisol replacement may be critical in these infants and deserves further study. 213 it is important to note, however, that in children with septic shock, adjunctive corticosteroid therapy is associated with repression of gene programs corresponding to the adaptive immune system. 214 complement-mediated activation of leukocytes during sepsis occurs via up-regulated cell surface receptors (complement receptor 1 [cd35] and cr3). 257, 258 c3b and c5a facilitate opsonization (primarily c3b), redistribute blood flow, and increase inflammation, platelet aggregation, and release of reactive oxygen intermediates (primarily c5a). 259, 260 c5a-mediated local leukocyte activation also results in increased cytokine production with subsequent up-regulation of adhesion molecules on vascular endothelium and increased cell recruitment to the site of infection. 261 data in adults link elevated c5a levels with multiple facets of sepsis-associated disease, such as dic, cardiac dysfunction, increased proinflammatory cytokine levels, sirs, apoptosis of adrenal medullary cells leading to adrenal cytokine production, and leukocyte activation. 104 contrary to its name, the alternative pathway is the primary mechanism of amplification of complement activation after c3 convertase assembly (which cleaves c3 to c3a and c3b). dysregulation of complement activation may contribute to adverse effects in individuals with severe sepsis or septic shock. neonates, particularly the very premature, exhibit decreased basal levels of complement proteins and function for both the alternative pathway and the classical pathway. 253, 254 moreover, as compared with adults, neonates exhibit gestational age-related degrees of depressed complement-mediated opsonic capabilities. 255 as such, complement-mediated opsonization is poor in premature neonates and limited in term neonates. 255 nization and pathogen clearance may help explain the lack of efficacy of intravenous immunoglobulin to prevent sepsis or death from sepsis in neonates. [279] [280] [281] [282] [283] [284] [285] apps are the most phylogenetically ancient means of innate immune defense against microbial invasion. present in nearly every organism, including bacteria, plants, insects, nonmammalian vertebrates, and mammals, these small, often cationic peptides are capable of killing microbes of multiple types, including viruses, bacteria, parasites, and fungi, largely by disruption of the pathogen membrane. 286 constitutive expression of apps occurs in humans on barrier areas with consistent microbial exposure such as skin and mucosa. after microbial stimulation, both release of preformed apps and inducible expression are thought to contribute to early host defense. 287 importantly, there is no evidence for the development of microbial resistance to apps that target fundamental components of the microbial cell wall. some apps can bind and neutralize microbial components such as endotoxin, precluding engagement with tlrs and other prrs, and diminish inflammation. many apps can potentially reduce the intensity of the inflammatory response associated with the presence of bacterial toxins. [288] [289] [290] because endotoxemia is an important contributor to neonatal mods and death with sepsis and nec, 84 lps-binding/blocking strategies, including use of synthetic apps, may have a significant positive impact on outcomes. 288, 291 bactericidal/permeability-increasing protein (bpi) is a 55-kda protein present in the respiratory tract, pmn primary granules, and plasma. bpi exerts selective cytotoxic, antiendotoxic, and opsonic activity against gram-negative bacteria. 288 plasma bpi concentrations were higher in critically ill children with sepsis syndrome or organ system failure than in critically ill children without sepsis syndrome or organ system failure, and bpi levels positively correlated with the pediatric risk for death score. 265 pmns from term neonates are deficient in bpi, potentially contributing to the increased risk for infection. 292 whereas term neonates demonstrate up-regulation of plasma bpi during infection, premature neonates showed a decreased ability to mobilize bpi on stimulation, 293 which may contribute to their risk for infection with gram-negative bacteria. polymorphisms in bpi increase the risk for gram-negative sepsis in children, but the impact of these polymorphisms in neonates is unknown. 294 compared with pmns from adults, pmns from term neonates produce similar quantities of defensins but reduced quantities of bpi and elastase. 292, 295, 296 recombinant bpi (rbpi 21 ) treatment was associated with improved functional outcome, reduced amputation, but no difference in mortality in a multicenter study of children with severe systemic meningococcal disease. 297 lactoferrin is the major whey protein in mammalian milk (in particularly high concentrations in colostrum) and is important in innate immune host defenses. lactoferrin is present in tears and saliva and has antimicrobial activity both via binding iron and by direct membrane disruption activity via a portion of its amino-terminal lactoferricin. 298 lactoferrin is also an alarmin (e.g., hmgb-1 or il-33), capable of activating leukocytes, binding endotoxin, and modifying the host response by acting as a transcription factor that regulates mrna decay. 299, 300 bovine lactoferrin has been shown to reduce the incidence of bacterial and fungal sepsis 301, 302 and nec in preterm infants. 303 lysozyme is present in tears, tracheal aspirates, skin, and pmn primary and secondary granules and contributes to degradation of peptidoglycan in bacterial cell walls. secretory pla 2 can destroy gram-positive bacteria through hydrolysis of their membrane lipids. 174 pmn elastase is a serine protease released by activated pmns with microbicidal function and is believed to play a role in the inflammatory damage seen with pmn recruitment, particularly in the lung. 116, 136 cathelicidin and the insufficiency, and pmn dysfunction. 104 septic shock in adult humans was associated with extensive complement activation, c-reactive protein-dependent loss of c5a receptor on neutrophils, and the appearance of circulating c5a receptor in serum, which correlated with a poor outcome. 262 deficiencies in c5a receptor found in term neonates as compared with adults may limit the ability to respond to c5a and therefore increase the likelihood of infection. 240 the expression of c5a receptor on preterm pmns is unknown. the extent to which c5a or other complement proteins play a role in the development of disease in septic neonates remains to be determined. complement regulatory proteins modify the effects of complement and prevent potential damage due to overactivation. in particular, cd59 blocks c9 polymerization and target lysis, cd55 destabilizes cd35 and c3 and c5 convertases, and cd35 accelerates the deactivation of c3b. 263 dysregulation of complement activation can lead to a vicious activation cycle that results in excessive cellular stimulation, cytokine production, endothelial cell activation, and local tissue damage promulgating sirs and septic shock (see figure 152 -5). 264 in addition to the initial inflammatory response including complement activation, molecular detection of pamps promotes il-1β and il-6 production, which in turn increases the production of multiple other innate proteins that possess valuable immune function and serve to reduce pathogen load. 265 acutephase reactant proteins, produced predominantly in the liver, include c-reactive protein (opsonin), serum amyloid a (cellular recruitment), lactoferrin (reduces the level of available iron/ antimicrobial peptide lactoferricin), procalcitonin (unknown function), haptoglobin, fibronectin (opsonic function), pentraxin 3 (binds c1q and activates the classical complement pathway), mbl, and lps-binding protein. 127, 211, 229, [265] [266] [267] [268] [269] [270] acutephase reactant proteins have been studied in neonates with sepsis primarily to assess them for diagnostic utility rather than immunologic function. in particular, elevated plasma concentrations of c-reactive protein and lps-binding protein are often associated with early-onset sepsis. 229, 271 the levels of il-10 and c-reactive protein were significantly higher in preterm infants who did not survive sepsis, pneumonia, or nec. 272 a lack of sustained increase in the production of c-reactive protein and serum amyloid a during sepsis has also been associated with a fulminant course. 273 the fetus receives antibodies from the mother via active placental transfer, with a significant increase beginning around 20 weeks' gestation. as a result of a shorter period of gestation, preterm neonates have lower igg subclass levels as compared with term neonates, particularly igg1 and igg2 subclasses. 274 preterm neonates (24 to 32 weeks' gestation) with low igg levels (serum total igg levels below 400 mg/dl at birth) were at increased risk for development of late-onset sepsis but not death compared to those with levels above 400 mg/dl. however, igg titers and opsonic activity to cons were not predictive of late-onset cons sepsis. 275 reliance on other means of innate immune defense likely provides the premature neonate with alternative microbial control mechanisms. despite the presence of maternally derived immunoglobulin and acute-phase reactant proteins, neonates exhibit impaired opsonizing activity compared with adults, which likely increases the risk for progression of infection. 276 complement plays a critical role in immunoglobulin-mediated opsonization and effector cell phagocytosis. 277 although immunoglobulin has many putative beneficial immunologic functions, most of these have not been demonstrated or examined in preterm infants. 278 the dependence on complement for effective immunoglobulin-based opso-thrombocytopenia in neonates, 322 which is attributed to reduced megakaryopoiesis in the setting of consumption with clot formation. 323 decreased platelet function in preterm neonates with sepsis further increases the risk for bleeding. 324 in extremely low-birth-weight infants, platelets are hyporeactive for the first few days after birth, complicating the ability of the immune system to contain a microbiologic threat and increasing the risk for hemorrhage. 325 clotting can lead to propagation of inflammation via thrombin-induced production of platelet-activating factor. pmns activated by platelet-activating factor or platelet tlr4 may then contribute to further endothelial injury and dysfunction, leading to the development of a vicious clottinginflammation-clotting cycle. activated platelets may be consumed in clot formation and/or may also be removed from the circulation by the liver, 326 potentially resulting in thrombocytopenia, particularly during gram-negative and fungal infections. 196, 322, 327 systemic activation of coagulation is associated with consumption of clotting factors and increased risk for bleeding, prolonged proinflammatory responses, and dic. 128, 149, 328 this finding is consistent with the elevated serum levels of il-6 52 and high frequency of dic seen with disseminated herpes simplex virus infection. 329 in adult mice, protease-activated receptor 1 plays a major role in orchestrating the interplay between coagulation and inflammation. 330 protease-activated receptor 1 may modify the endothelial response during neonatal sepsis and thus is a target for therapeutic intervention. recent studies have shown the critical importance of vascular endothelial activation in the early recognition and containment of microbial invasion. in transgenic mice, it was shown that pulmonary endothelial cells sense blood-borne bacteria and their products, 156 whereas alveolar macrophages patrol the air spaces. 331 these data illustrate the role of endothelium and help to explain in part the occurrence of acute respiratory distress syndrome (ards) and persistent pulmonary hypertension of the newborn associated with severe sepsis in the absence of a primary pulmonary infectious focus. expression of tlrs allows endothelium to become activated in the presence of microbial components, leading to production of cytokines, chemokines, and adhesion molecules (e.g., vascular cell adhesion molecule, intercellular cell adhesion molecule, l-selectin, p-selectin, and e-selectin). these substances are all necessary to attract immune cells (primarily pmns) to the site of infection and to facilitate pathogen containment. [150] [151] [152] [153] 156 vasoactive substances released from activated leukocytes, platelets, and endothelial cells include platelet-activating factor, thromboxanes, leukotrienes, no, histamine, bradykinin, and prostaglandins. 172, 173 the balance of no and endothelin 1, a vasoconstrictor, may be disrupted with endothelial damage, favoring the constrictive effects of endothelin 1 and leading to ischemia and injury. 175 this phenomenon may explain in part why no inhibitors increased mortality in adults with septic shock. 301 stimulated endothelium can be a doubleedged sword, however, because excessive activation can lead to systemic overproduction of cytokines and vasoactive substances (including no). endothelial cell apoptosis, detachment from the lamina, and alterations in vascular tone combine to promote capillary leak, leading to hypovolemia, shock, and organ failure 156,302,303 (see figure 152 -5). release of myeloperoxidase from pmns may also injure surrounding endothelium. 332 activated or damaged endothelium establishes a prothrombotic environment that can result in local microvascular occlusion 314 or progress to dic. 333 the glucocorticoid receptor is the target for cortisol, the primary endogenous glucocorticoid in humans, produced in the zona fasciculata of the adrenal glands. endothelial glucocorticoid defensins are other apps that possess antimicrobial properties. 304 cathelicidin is present in the amniotic fluid, vernix, skin, saliva, respiratory tract, and leukocytes. α-defensins are cysteine-rich 4-kda peptides found in amniotic fluid, vernix, spleen, cornea, thymus, paneth cells, and leukocytes. β-defensins are found in the skin, gi tract, urinary system, reproductive organs (placenta, uterus, testes, kidney), respiratory tract, breast milk, mammary gland, and thymus. in addition to microbicidal action, apps have a wide range of immunomodulatory effects on multiple cell types from both the innate immune system and the adaptive immune system. 287, 305, 306 these immunomodulatory effects include altered cytokine and chemokine production, improved cellular chemotaxis and recruitment, improved cell function (maturation, activation, phagocytosis, reactive oxygen intermediate production), enhancement of wound healing (neovascularization, mitogenesis), and decreased apoptosis. the cytosolic granules of pmn are rich in apps, including α-defensins, lactoferrin, lysozyme, cathelicidin, soluble pla 2 , and bpi. gestational age-related decreases in the umbilical cord blood concentration of several apps (cathelicidin, bpi, calprotectin, soluble pla 2 , α-defensins) in comparison with maternal serum levels have been drescibed. 307 plasma app deficiencies may contribute to the increased risk for infection associated with prematurity, and their absence may increase the risk for endotoxemia. compared with term neonates, preterm neonates showed lower human β-defensin 2 levels in umbilical cord blood. 308 up-regulation of apps (defensins) occurs in blood of infected adults 309 and children (defensins, lactoferrin). 310 the effect of sepsis on the production of plasma apps in neonates has not been investigated in detail. the development of a procoagulant state in the surrounding microvasculature allows the trapping of invading pathogens and prevents further dissemination (see figure 152 -5). in general, the intrinsic pathway amplifies coagulation after initiation by the extrinsic pathway. 311 reduced levels of vitamin k-dependent factors (factors ii, vii, ix, and x), reduced thrombin generation, reduced consumption of platelets with formation of microthrombi, and reduced levels of counterregulatory elements (inhibitors) increase the risk for bleeding in infants and children. 312 during sepsis, a microvascular procoagulant state develops via stimulation of phagocytes, platelets, and endothelium, resulting in expression of tissue factor. 313, 314 tissue factormediated activation of the coagulation cascade results in activation of thrombin-antithrombin complex, plasminogen activator inhibitor type 1, and plasmin-α 2 -antiplasmin complex, 315 as well as inactivation of protein s and depletion of the anticoagulant proteins antithrombin iii and protein c. [316] [317] [318] decreased activated protein c levels were associated with increased risk for death from sepsis in preterm neonates. 319 a randomized controlled trial of activated protein c revealed no change in mortality among pediatric patients with sepsis, but term infants younger than 60 days old experienced increased bleeding. 320 the coagulation cascade is intimately tied to inflammation and complement activation. 104 cytokine production increases expression of endothelial tissue plasminogen activator inhibitor type 1. plasminogen activator inhibitor type 1 inhibits fibrinolysis by inhibiting the conversion of plasminogen to plasmin, which in turn is important for the breakdown of fibrin. deposition of fibrin in small vessels leads to inadequate tissue perfusion and organ failure. 321 increased plasminogen activator inhibitor type excessive local inflammation and tissue damage. 350 high early levels of circulating free pmn-derived dna produced by nets are associated with mods and death. 351 nets contain destructive proteases capable of killing bacteria even after the pmn has died. 352 formation of nets is reduced in pmns from preterm neonates and nearly absent in term neonates 353 but may occur with sustained cellular stimulation. 354 net formation may result in collateral damage to surrounding tissues when the target microbe is too large to be effectively phagocytosed (e.g., fungal hyphae). 355 the contribution of net production to detrimental outcomes in infected neonates is unknown but excessive net formation with collateral tissue injury may contribute to the poor outcomes seen in preterm neonates with fungal infections. 356 rapid depletion of bone marrow pmn reserves during infection, particularly in neonates, 357 can lead to neutropenia, with consequent impaired antimicrobial defenses and significantly increased risk for death. 358 in a multivariate analysis, neutropenia and metabolic acidosis were associated with fatal neonatal sepsis. 359 neutropenia is particularly common in gram-negative sepsis in neonates. 360 release of immature pmn forms (bands), which exhibit greater dysfunction than mature pmns, 361 may further predispose to adverse outcomes. murine neonates with experimental sepsis exhibit delayed emergency myelopoiesis (a process by which the host repopulates peripheral myeloid cells lost early during sepsis), that is independent of trif and myeloid differentiation factor 88. 362 interventions aimed at addressing reduced pmn numbers in neonates have included provision of mature pmns 363 and prophylaxis or treatment with colonystimulating factors (granulocyte colony-stimulating factor and granulocyte-macrophage colony-stimulating factor). despite strong biologic plausibility, these interventions have been unsuccessful at reducing the neonatal infectious burden. [364] [365] [366] in a metaanalysis, treatment with colony-stimulating factor therapy (granulocyte colony-stimulating factor, granulocyte-macrophage colony-stimulating factor) in a subgroup (n = 97) of neutropenic neonates (absolute neutrophil count less than 1700/µl) with culture-positive sepsis (largely gram-negative and gbs) significantly reduced the risk for death (relative risk, 0.34; 95% ci, 0.12 to 0.92). 365 therefore, stimulation of granulopoiesis may be beneficial under these specific circumstances, although further studies focused on this subpopulation and outcomes are needed. irreversible aggregation and accumulation of newborn pmns in the vascular space after stimulation leads to decreased diapedesis, rapid depletion of bone marrow reserves, vascular crowding, 367 and increased likelihood of microvascular occlusion. 368 neonatal pmn deformation compared with adult pmn deformation is reduced at the baseline, which increases the risk for occlusion. 367 furthermore, low blood pressure/flow states seen during septic shock further exacerbate existing microvascular ischemia. 295 in combination, these deficiencies increase the propensity for systemic spread of infection, and set the stage for microvascular occlusion. many other cells besides pmns are involved in the development of an immune response to infection. monocytes, macrophages, and dendritic cells amplify cellular recruitment through production of inflammatory mediators, activation of endothelium, phagocytosis and killing of pathogens, and antigen presentation to t and b cells of the adaptive immune system. the primary functions of monocytes are the synthesis of crucial inflammatory proteins 369 and antigen presentation to naïve cd4 + t cells. 370 the patterns of cytokine production can promote the differentiation of naïve cd4 + t cells into distinct subtypes of t cells that serve important roles in the clearance of pathogens. for example, t-helper 1 (t h 1) cells are produced from naïve cd4 + t cells after exposure to ifn-γ and il-12, and support cellmediated immunity against intracellular pathogens through production of ifn-γ, tnf-α, and lymphotoxin. t-helper 2(t h 2) cells receptor is a critical negative regulator of inducible no synthase expression and nf-κb activation, 334 demonstrating a protective role of the endothelium during sepsis. studies have revealed a potential role of plasma angiopoietin during pediatric septic shock. 335 the level of angiopoietin 1, which protects against vascular leak, was reduced, whereas the level of angiopoietin 2, which promotes vascular permeability, was elevated, highlighting a novel potential therapeutic opportunity to reduce endorgan injury. the roles for endothelial glucocorticoid receptor and angiopoietin 1 in neonatal sepsis are unknown. the role of endothelium activation during sepsis and septic shock in neonates, particularly in premature neonates, has been less well investigated. toxins from gbs have been shown to damage pulmonary endothelium 336 and likely participate in pulmonary complications associated with gbs pneumonia such as ards and the development of persistent pulmonary hypertension of the newborn. 337 the levels of the adhesion molecules e-selectin and p-selectin, expressed and secreted by activated endothelium, are increased in the serum of neonates with sepsis 136 and likely reflect significant endothelial activation. endothelial tlr4 activation impaired intestinal perfusion in an experimental model of nec, via endothelial no synthasenitrite-no signaling. 338 the pmn is the primary effector of innate immune cellular defense. endothelial cells produce activating cytokines and chemokine gradients that recruit circulating pmns to the site of infection. expression of cell adhesion molecules by pmns and endothelium allows cells to roll and extravasate into surrounding tissues. activated pmns phagocytose and kill pathogens via oxygen-dependent and oxygen-independent mechanisms. il-1β is produced by activated pmns largely via an nlrp3-asccaspase 1-dependent* mechanism that amplifies the recruitment of additional pmns from the bone marrow to the site of infection. 339 activated pmns may release reactive nitrogen species, reactive oxygen species, and proteolytic enzymes via activation of membrane-associated nadph oxidase. these reactive intermediates and enzymes can lead to destruction of nonphagocytosed bacteria but can also cause local tissue destruction, including neonatal endothelial and lung injury, as well as surfactant inactivation, 116, 340 and thus play a role in progression from sepsis to mods. neonatal pmns exhibit quantitative and qualitative deficits as compared with adult pmns. 295, 341 respiratory burst activity is suppressed in pmns during neonatal sepsis and may contribute to poor microbicidal activity. [342] [343] [344] compared with adult pmns, neonatal pmns exhibit delayed apoptosis, 345, 346 as well as sustained capacity for activation (cd11b up-regulation) and cytotoxic function (reactive oxygen intermediate production) that contributes to tissue damage. 347 reduced apoptosis with prolonged survival of pmns may result in improved bacterial clearance but may also paradoxically increase the risk for sustained pmn-mediated tissue damage. increased serum pmn elastase, urokinase plasminogen activator, and urokinase plasminogen activator receptor levels are found at the time of presentation in infected neonates. 136 with pmn death, dna (chromatin), histones, and apps are expelled into the environment and serve to trap bacteria (neutrophil extracellular traps [nets]). 348 the formation of nets can occur after activation of platelet tlr4 349 and may lead to *asc, apoptosis-associated speck-like protein containing a carboxyterminal card. eosinophils phagocytose antigen-antibody complexes and release cytokines, chemokines, cytotoxic molecules, apps, and other substances (prostaglandins, thromboxanes, leukotrienes) when stimulated. 387 eosinophilia is commonly observed in neonates with sepsis due to candida sp. 388 and bacteria, 389 and is seen in infants with nec. 387 in infants of less than 26 weeks' gestation, eosinophilia (absolute eosinophil count more than 1000/mm 3 ) may predict bacterial sepsis. 389 eosinophilia in premature infants is not associated with production of ige. 390 studies have demonstrated an integral role for eosinophils in adult intestinal integrity and revealed a novel innate bactericidal nonphagocytic function via extracellular catapulting of mitochondrial dna nets with associated bound toxic proteins. 391 the precise role of eosinophils in the neonatal immune response to sepsis and in maintenance of intestinal integrity has yet to be determined. mast cells play a role in the response to pathogen invasion as a part of the innate cellular immune system via production of histamines (which promote vasodilation and up-regulation of p-selectin), cytokines, pmn recruitment, bacterial phagocytosis, and antigen presentation. 392, 393 mast cell involvement was demonstrated in infants with erythema toxicum, where mast cell recruitment, degranulation, and expression of apps occurs. 394 adult rodents deficient in mast cells exhibit impaired pmn influx, 395 impaired clearance of enteric organisms, and decreased sepsis survival. 396 mast cell production of histamine likely contributes to the vasodilation associated with sepsis and septic shock. like eosinophils and pmns, mast cells are capable of killing bacteria via generation of extracellular traps in adults. 397 this means of immune protection has not been investigated in neonates. mast cells may also alter adaptive immune function by patterning the t h 2 immunophenotype seen in the neonate and therefore contribute to the increased risk for infection. immature dendritic cells exposed to histamine during maturation (with lps) exhibit altered t-cell polarizing activity with predominance towards the t h 2 phenotype via increased production of il-10 and decreased production of il-2. 398 furthermore, mast cells from neonates were shown to secrete significantly more histamine after stimulation as compared with adults, 399 which may contribute to the development of shock. 400 the role of natural killer (nk) cells in neonatal bacterial sepsis is incompletely defined. nk cell numbers increase with increasing gestational age, 401 furthermore, a reduced percentage of nk cells present at birth may be a risk factor for late-onset sepsis in preterm infants. 402 it is noteworthy that the numbers of circulating nk cells are not significantly different in neonates with or without infection 370, 403 ; however, the numbers of circulating nk cells are decreased in newborn infants with shock. 404 the mechanisms used by nk cells to destroy bacteria include secretion of apps (defensins), direct contact and lysis, antibody-dependent cellular cytotoxicity, and ifn-γ production. 405 in neonates with bacterial sepsis, nk cells are activated, as evidenced by up-regulation of cd69. 2, 406 despite activation, nk cell cytotoxicity is deficient in infants with sepsis and recurrent infections. 370, 405 although neonatal macrophages exhibit impaired baseline activation in response to ifn-γ, 341 nk cell-mediated production of ifn-γ can enhance their phagocytic capability. further studies are necessary to more clearly define the role of nk cells in neonatal bacterial sepsis. cd71 + ter119 + (erythroid) cells may contribute to the increased susceptibility of the neonate to infection by reducing the inflammatory response associated with bacterial colonization of the gut. for example, ex vivo tnf-α production by stimulated adult effector cells was reduced in the presence of murine neonatal splenic cd71 + erythroid cells via an arginase 2-dependnent mechanism. 407 the cd71 + erythroid population represents a large portion of murine fetal liver, neonatal spleen/bone marrow, and adult bone marrow. [408] [409] [410] furthermore, the murine neonatal spleen contains large numbers of colony-forming progenitor cells for 2 to 3 weeks after birth. 411 of note and in stark contrast arise in the presence of il-2 and il-4, produce il-4, il-5, and il-13, down-regulate t h 1 responses, and support humoral immunity, as well as defense against extracellular parasites. a third subset of t h cells, t-helper 17 cells, are generated in the presence of transforming growth factor β, il-6, il-21, and il-23. these cells produce il-17 and il-22, which are important for defense against extracellular bacteria and fungi. neonatal mononuclear cells exhibit a bias away from t h 1 cell-polarizing activity because of increased il-6 and low tnf-α production. 371 this may be beneficial because of mobilization of antiinfective proteins/ peptides that serve to protect the newborn during microbial colonization 266 and development of immune tolerance. 341 the adverse consequence is a reduced ability to respond to infection with microorganisms; particularly intracellular pathogens such as listeria sp. 372 and mycobacteria. 373 preterm infants (<30 weeks) may have greater attenuation of tnf-α and il-6 secretion compared with term infants and adults. 277 there is decreased monocytic recruitment to areas of inflammation during sepsis because of decreased chemotactic ability. 374 although the levels of peripheral monocytes decrease early during sepsis (between 60 nd 120 hours), secondary to extravasation and differentiation into macrophages, sepsis-related elevation of macrophage colony-stimulating factor 375 results in a late increase in the number of peripheral monocytes (>120 hours). 376 in addition to altered cytokine production and suboptimal recruitment, monocyte phagocytic function is reduced during sepsis. 377 antigen presentation to naïve cd4 + t cells is an important immune function performed by monocytes. the decreased antigen-presenting function in monocytes from newborn infants is in part due to decreased mhc class 2 molecule expression 378 and decreased expression of costimulatory molecules, including cd86 and cd40. 379 monocytes leave the bloodstream, enter the tissues, and differentiate into macrophages and dendritic cells. monocytes and macrophages are closely related to pmns (common myeloid progenitor) and can kill pathogens by similar means. circulating monocytes differentiate into macrophages after exposure to maturing cytokines, and exit the bloodstream into tissues. important substances produced by stimulated monocytes/macrophages include complement components, cytokines (both proinflammatory and antiinflammatory), coagulation factors, and extracellular matrix proteins. 369 located just below epithelial borders, macrophages encounter pathogens immediately after entry. macrophages are avidly phagocytic and generate apps to reduce bacterial burden, such as lactoferrin, defensins, transferrin, and lysozyme. in addition, macrophages play an important role in the amplification of the immune response through the production of cytokines and chemokines, as well as in antigen presentation to naïve cd4 + t cells. macrophages are poorly responsive to several tlr agonists. 380 dendritic cells are antigen-presenting cells that function as a liaison between the innate immune system and the adaptive immune system through induction of antigen-specific t cellmediated immunity. dendritic cells from newborn infants exhibit a reduced antigen-presenting function when compared with adult cells 379 and require increased stimulation for activation. 381 evaluations of neonatal dendritic cell function suggest a tendency towards poor up-regulation of costimulatory molecules (cd80/cd86) and activation markers (cd83), poor stimulation of t-cell proliferation, and a tendency towards the induction of immune tolerance. 382 although preterm and term infants and adults have similar numbers of "plasmacytoid" dendritic cells in their blood, the capacity to produce ifn-α on tlr9 challenge was significantly decreased in preterm neonates and may increase susceptibility to viral infections. 383 dendritic cells in umbilical cord blood can effectively induce cytotoxic lymphocyte responses. 384 depletion of dendritic cells has been reported in adult animals 385 and adult patients 386 with sepsis; their role in the immune response to neonatal sepsis is not well characterized. pmn and γδ t cells. 432 these data show that neonatal t cells are activated and are capable of playing a role in the host response to bacterial sepsis in vivo. neonatal lymphocyte function is skewed towards t h 2 responses, setting the stage for immune tolerance (t h 2) rather than immune priming for infection (t h 1). 417 newborn infants must overcome that immune modulation in order to mount effective responses to specific infectious challenges and respond to vaccination. examples of the impact of this immunopolarization include decreased ifn-γ production by cd4 + and cd8 + t cells as compared with production in children and adults. 433, 434 the likely significance of decreased ifn-γ production is a reduction in activation of other immune cells, such as macrophages. reports of lymphocyte function in infected newborns are very limited. expansion of lymphocytes after antigenic stimulation is important for development of sustained immunity. decreased lymphocyte proliferative responses have been shown during the first 8 weeks of life in vlbw neonates, 421 and may predispose the premature neonate to development of late-onset sepsis. for example, t-lymphocyte function was depressed in infected newborn infants, and especially in those with multiorgan failure, versus healthy term or growing preterm infants. 435 similarly, production of lymphocyte-associated cytokines after stimulation of umbilical cord blood peripheral blood mono nuclear cells with gbs was significantly deficient in preterm and term infants compared with adults. 277 cytomegalovirus infection in utero leads to the expansion and differentiation of mature cytomegalovirusspecific cd8 + t cells, which have characteristics similar to adult cd8 + t cells. 436 these cells showed potent perforin-dependent cytolytic activity and produce antiviral cytokines, highlighting the potential for adult-like immunocompetence of neonatal t cells under specific circumstances. an important location for effective lymphocytic function during systemic bacterial infection is the spleen. the marginal zone of the spleen facilitates the clearance of bacteria, particularly encapsulated organisms, from the bloodstream. these functions are accomplished via the interaction of multiple leukocytes, including macrophages, dendritic cells, b cells, and t cells, within follicles of the spleen. the neonatal splenic marginal zone is immature, owing to a lack of antecedent antigen exposure and is virtually devoid of cd21 + b cells. 412 as a result of this functional asplenia, there is decreased clearance of pathogens from the blood and potential for a more fulminant course with bacteremia. 437, 438 b cells are critically important in the adult host response to sepsis. data suggest antibody-independent and antibodydependent roles for b cells in the outcome of sepsis. 416 studies deciphering the role of b cells in neonatal sepsis are very limited, and thus the role b cells play in the neonatal host response is unclear. after gbs meningitis, the level of igm was increased, suggesting b cells from neonates can respond to pathogenic challenge. 439 premature neonates with perinatal infection or nosocomial infection may show signs of humoral immunoparalysis, manifested by decreased igm/igg production ex vivo as compared with production in their healthy age-matched counterparts. 440 sepsis in early life did not reduce serum antibody titers in preterm infants after heptavalent pneumococcal conjugate vaccine exposure but was associated with a reduced opsonization titer to a single serotype, suggesting the capacity to respond to vaccination or other immune challenge may be altered. 441 in the setting of reduced classic adaptive immune function seen in early life as compared with the function in adults, innate lymphoid populations (which lack b cell receptor and t cell receptor) may play a significant role in protecting the neonate from infectious challenge. [442] [443] [444] [445] [446] [447] [448] examples of innate lymphoid cells include γδ t cells, intestinal lymphoid cells, invariant nk t cells, mucosa-associated invariant t cells, and b1 cells. to the lymphoid and reticuloendothelial system roles of the spleen in the healthy adult, the spleen is normally a major site of erythropoiesis during fetal and neonatal life, to support rapid fetal and postnatal growth in the setting of significantly reduced erythroid reservoirs as compared with the adult reservoirs. 410, 412, 413 a lack of effect on neonatal murine survival to polymicrobial sepsis after adoptive transfer or diminution of cd71 + erythroid splenocytes suggests that the impact of these cells on neonatal infection risk and progression may be limited. 414 the contribution of the adaptive immune system in the neonatal host response to sepsis is uncertain. the 5-to 7-day interval required for development of an adaptive immune responsenamely, the selection and amplification of specific clones of lymphocytes (b cells and t cells) that results in immunologic memory-argues against a central role for adaptive immunity in the protective response to early neonatal bacterial sepsis. as a result, the neonate is thought to largely depend on innate immunity for protection from infection during the first days of life. in adults, absence or dysfunction of the adaptive immune system has a profound impact on survival in preclinical models. 415 b cells (and in particular b-cell cytokine production) and not t cells were shown to be important in the early host response to experimental sepsis. 416 studies using neonatal mice lacking an adaptive immune system showed no difference in polymicrobial sepsis survival as compared with survival of wild-type mice with an intact adaptive immune system. 3 furthermore, there are many quantitative and qualitative differences in lymphocytes from neonates compared with lymphocytes from adults, 417 each with a respective proposed clinical impact. 87 as these findings illustrate, the contribution of adaptive immunity for protection and response against sepsis, and in particular which components are protective, is unclear in the most immature and requires further investigation. peripheral blood examination has yielded inconsistent changes in the percentage, number, and type of circulating lymphocytes during sepsis. 403, [418] [419] [420] [421] [422] [423] moreover, changes related to the timing of sepsis onset (early-onset or late-onset sepsis) and prematurity have been incompletely characterized. t regulatory cells are abundant and potent at birth, facilitating inhibition of t h 1 cell immunity, 424 and perhaps mediating a state of immunologic tolerance. 425 although the numbers of splenic t regulatory cells are increased in murine neonates and adults with sepsis, depletion of t regulatory cells had no effect on survival of murine adults. 2, 426 alterations in the number or function of t regulatory cells in human neonatal sepsis have not been reported. examination of peripheral blood to identify markers of sepsis has yielded a number of lymphocyte cell-surface molecules whose levels increase during sepsis. activation of neonatal t cells is evidenced by increased cd45ro expression (present on t cells after antigenic stimulation) at the time of sepsis diagnosis, 419, 427, 428 and with congenital infection, 429 although changes in number may take several days to occur after stimulation. 430 other markers of lymphocyte activation may be found at different time points during the course of infection. for example, expression of the activation marker cd69 is increased on t cells (cd4 + ) early in the infectious process, whereas cd25 and cd45ro expression persists for several days. 406 increased expression of cd4 + t-cell carcinoembryonic antigen-related cell adhesion molecule 1 (cd66a) in preterm infants with lateonset sepsis may contribute to sepsis-associated immune suppression. 431 hla-dr expression is increased on multiple cell types during neonatal sepsis. 406 in contrast to adults, a large portion of neonatal t cells produce cxcl8, which activates of genes for innate immune and metabolic pathways with decreased levels of adaptive immune transcripts. 467 using a proteomics approach, ng and colleagues 468 identified proapolipoprotein cii and a desarginine variant of serum amyloid a as promising biomarkers for late-onset sepsis and nec in preterm infants. 468 it is very likely that implementation of unbiased "omic" approaches will reveal critical age-appropriate pathways and opportunities for therapeutic interventions aimed at improving neonatal sepsis outcomes. sepsis that leads to shock and organ failure carries the worst prognosis. sirs contributes to the development of organ failure in neonates (see figure 152 -5). 52, 148, 181, 469 persistent decreases in capillary perfusion are associated with mods and death in adults. 470 lethargy, shock, and birth weight less than 1500 g were independent predictors of sepsis-related death. 471 in neonates, impairment of the cardiovascular system, manifested by poor perfusion or hypotension, is invariably associated with septic shock. sustained poor organ perfusion in neonatal sepsis and septic shock due to cardiovascular dysfunction is associated with mods affecting the kidney, 472,473 liver, 474 gut, 475 and central nervous system 476 (see figure 152 -5) . the mechanism of organ failure may be decreased oxygen utilization associated with mitochondrial dysfunction rather than poor oxygen delivery to tissue. 477, 478 on the basis of available evidence, it has been speculated that the prolonged sirs associated with severe sepsis and shock leads to organ failure via a cessation of energy-consuming processes. 479, 480 development of severe nec is also associated with severe sepsis, shock, mods, and death. 84, 481 the need for intubation or initiation of vasoactive medications, and hypoglycemia, thrombocytopenia, increased prothrombin time, or excessive bleeding as presenting laboratory signs of sepsis are risk factors for sepsis-related death. 359, 475, 482 independent predictors of in-hospital late-onset sepsis death during the birth hospitalization were the presence of congenital anomalies (or, 4.12; 95% ci, 1.60 to 10.60), neuromuscular comorbidities (or, 3.34; 95% ci, 1.66 to 6.73), and secondary pulmonary hypertension with/without cor pulmonale (or, 23.48; 95% ci, 5.96 to 92.49), 60 underscoring the impact of organ-level comorbidities that increase neonatal sepsis mortality. the most common organ dysfunction associated with sepsis is cardiovascular dysfunction. cardiovascular dysfunction associated with sepsis may lead to shock that is a composite of hypovolemic, cardiogenic, and distributive shock. distributive shock is related to endothelial no production that leads to excessive vasodilation. cardiogenic shock may be related to mitochondrial death (induced by reactive nitrogen and reactive oxygen intermediates) with subsequent myocardial dysfunction. abnormalities in peripheral vasoregulation and myocardial dysfunction may play a larger role in hemodynamic derangements in pediatric patients, especially infants and neonates. in children, a non-hyperdynamic state with reduced cardiac output and increased systemic vascular resistance is most commonly observed in the setting of sepsis. [483] [484] [485] [486] [487] the hemodynamic presentation in neonates is much more variable 484 and is complicated by an unclear association between a normal blood pressure and adequate systemic blood flow. 488, 489 microcirculatory flow is impaired in term neonates even with mild to moderate severity of infection. 490 preterm neonates with sepsis have relatively high left and right cardiac outputs and low systemic vascular resistances. however, a decrease in right or left ventricular output of more than 50% has been associated with increased mortality in neonatal sepsis. 491 an elevated left ventricular output mechanistic investigations that fully explore the role of these newly discovered populations in the neonatal host response to sepsis are likely to uncover novel therapeutic opportunities. systems biology and the use of "omic" approaches have the potential to produce significant insights into the pathogenesis of sepsis. genomic and proteomic approaches have yielded important data associated with septic shock in older populations. [449] [450] [451] [452] [453] [454] [455] [456] [457] the use of these modern techniques in the study of neonatal inflammation and response to pathogen challenge has only just begun. 136, 198, 458, 459 the ability to profile genome-wide expression has significantly enhanced our understanding of the complexity of the host immune response to sepsis in children. 5, 449, 450, 453, 460 for example, genome-wide expression profiling revealed zinc homeostasis as an important feature of pediatric sepsis. 450, 453, 461 prophylactic zinc supplementation reduced bacterial load and mortality in a murine model of peritoneal sepsis. 462 however, oral zinc supplementation did not alter mortality in neonates with probable sepsis. 463 in a study of pediatric patients who met the criteria for septic shock, 6 a unique whole-blood transcriptomic response was found in neonates as compared with infants, toddlers, and school-age children. neonates manifested the largest number of uniquely regulated genes, representing both innate and adaptive immune system pathways, and showed a predominance of down-regulated transcripts representing the adaptive immune system. 5 the number of up-regulated genes increased in proportion with developmental age. investigation of the murine circulating leukocyte transcriptome revealed significant differences in the host immune response to sepsis across the age spectrum (neonate, young adult, elderly), despite similar increases in mortality among the neonates and elderly mice as compared with young adult mice. 1 these data underscore the impact of developmental age on the host immune response and suggest that therapeutics, which may have efficacy in older populations, may be ineffective in or possibly detrimental to neonates. because the transition to extrauterine life is associated with dramatic changes in physiology, the whole-blood transcriptome is likely to be quite different between both uninfected infants and in the host response to sepsis by timing after birth. indeed, an unsupervised analysis of the whole-blood genome-wide transcriptome on prospectively collected peripheral blood samples from infants evaluated for sepsis revealed the major node of separation between groups (infected or uninfected) was the timing of evaluation relative to birth (early, lass than 3 days or late, more than 3 days). 464 principal component analyses revealed significant differences between patients with early or late sepsis despite the presence of similar key immunologic pathway aberrations in both groups. this study highlights both the uninfected state and the host responses to sepsis are significantly affected by timing relative to birth. a study of vlbw infants with blood culture-proven late-onset sepsis (59% cons) identified a 554-gene signature associated with sepsis, with increased expression of the tnf-α network, including matrix metalloproteinase 8 and cd177 among the most commonly up-regulated genes. 465 elevated matrix metalloproteinase 8 mrna expression and activity in septic shock correlated with decreased survival and increased organ failure in pediatric patients. matrix metalloproteinase 8 is a direct activator of nf-κb. 466 inhibition (genetic or pharmacologic) of matrix metalloproteinase 8 leads to improved survival and a blunted inflammatory profile in a murine model of sepsis. most recently, a 52-gene network was uncovered and validated that accurately identified infected infants, who exhibited increased expression phosphorylation, ubiquitination, sumoylation) may occur after sepsis. 126, 517 these dna alterations may modify transcription factor access of gene-specific promoter regions, ultimately leading to short-term and long-term changes in gene expression and immune function. the dna methylation pattern in the promoter region of the calca gene varied in different types of bacterial sepsis in preterm infants, suggesting its potential use as an epigenetic biomarker. 518 trained immunity, the term coined to describe an adaptive innate immune response, may also be a positive or negative consequence of sepsis in early life. 519 mechanisms that underlie trained immunity are beginning to emerge, and include dna methylation and modification of energy utilization pathways. 520, 521 nonspecific vaccine benefits and resistance to subsequent pathogen challenge after innate immune priming or previous infection are likely manifestations of trained immunity in neonates. 3, 513, 522 the cell types, extent, and duration of trained immunity-based modifications in neonates with sepsis have not been studied. myeloid suppressor cells manifest immunosuppressive activity with sepsis 523 and were recently described in neonates. myeloid suppressor cells are present at high frequency at birth and decline in number with postnatal age. they inhibit t-cell proliferative responses and ifn-γ production. 524 reactivation of viral infection that may contribute to morbidity and mortality has been demonstrated in infected adults. 525 the impact of this phenomenon in neonates is unknown. acute hypoxic respiratory failure, ards, and acute lung injury are common pulmonary complications associated with severe sepsis. destruction of the alveolar capillary membrane leads to refractory hypoxemia. after direct or indirect insults to the lung, alveolar macrophages produce chemokines that mitigate pmn influx to lung parenchyma. activated pmns release reactive oxygen and reactive nitrogen intermediates that damage endothelial and epithelial barriers, leading to leakage of protein-rich edema fluid into the air spaces. other pulmonary complications with severe sepsis may include secondary surfactant deficiency, 526 primary or secondary pneumonia, 527 and reactive pulmonary hypertension. 528, 529 infants with sepsis and persistent pulmonary hypertension of the newborn may require inhaled no in addition to optimized ventilation strategies such as highfrequency oscillatory ventilation. 530 if oxygenation or tissue perfusion remains severely compromised despite optimal medical management, extracorporeal membrane oxygenation should be considered in neonates weighing more than 2 kg without contraindications. 531,532 the detrimental neurodevelopmental long-term impact of sepsis has been demonstrated in multiple studies and has been reviewed in detail. [533] [534] [535] [536] [537] central nervous system injury is predominantly white-matter injury (loss of pre-oligodendrocytes), manifested by focal cystic periventricular leukomalacia, diffuse necrosis, or a combination of these entities. 538, 539 central nervous system injury is, in part, mediated by inflammation with or without direct pathogen invasion. 141, 540, 541 the impact of sepsis on central nervous system injury is intensified with lower gestational ages, highlighting the detrimental effects of sepsis on the developing brain. 539 the importance of evaluating the preterm infant for disseminated infection that may include meningitis cannot be overemphasized. a complete evaluation, including cultures of blood, urine and cerebrospinal fluid, is uncommon, 356 although one third of the cases of culture-positive meningitis in vlbw infants are associated with negative concurrent blood cultures. 542 clinically apparent seizures may occur in 25% of vlbw preterm infants with meningitis. 542 low-voltage background pattern, sleep-wake cycling, and seizure activity on but normal ejection fraction in preterm neonates with septic shock suggests that septic shock in preterm neonates is predominantly due to vasoregulatory failure. neonatal sepsis may or may not be associated with left ventricular diastolic dysfunction; however, cardiac injury as manifested by elevated levels of cardiac troponin t may complicate the clinical picture. 492, 493 abnormal peripheral vasoregulation with or without myocardial dysfunction is the primary mechanism for the hypotension accompanying septic shock in the neonate. 494 therefore, infected neonates may present with hypotension and adequate perfusion (warm shock) or inadequate perfusion (cold shock). myocardial dysfunction can lead to ventricular wall stretch that in turn elevates b-type natriuretic peptide levels. b-type natriuretic peptide levels are elevated in children with septic shock, 495 and increased levels have utility as prognostic indicators of death. 496 plasma no level is elevated in neonates with sepsis and shock compared wit those with shock alone. 182 elevated serum lactate level (>3 mmol/l) distinguished nonsurvivors from survivors in a pediatric study that included neonates. 497 after severe sepsis or septic shock, there is an increased risk for subsequent infection and death in children and adults. this phenomenon is termed immunoparalysis and is associated with reduced mhc class 2 expression and tnf-α production by mononuclear cells after endotoxin stimulation. in addition to altered monocytic responses, there is significant loss of lymphoid cd4 + t and b cells via caspase-dependent apoptotic pathways. 415, 498 whether by clonal selection, apoptosis, or elevated endogenous glucocorticoid levels, [499] [500] [501] lymphocyte loss may lead to a state of immune compromise after the acute phase of sepsis. 412, 499, [501] [502] [503] [504] [505] new data suggests that il-7 may play an important role in promoting t-cell activation and prevention of apoptosis. 506 the importance of immunoparalysis has been convincingly demonstrated in infected adults 507-510 and children. 511 however, the clinical impact in the preterm neonate in whom adaptive immune function is less well developed is uncertain. 512, 513 in examinations of peripheral blood and postmortem spleens from infected adults, there is significant loss of b and cd4 + t lymphocytes and dendritic cells, 386, 498 resulting in decreased antigen presentation, antibody production, and macrophage activation. 514 circulating peripheral absolute lymphocyte counts can drop significantly in adults with sepsis but this phenomena is also seen in critically ill adults who are not infected. 477 sustained lymphopenia significantly increases the risk for secondary infection, mods, and death in children. 505 extensive loss of lymphocytes (both b and t lymphocytes) has been described in postmortem specimens from the thymus and spleen in infected preterm and term infants. 412, 499, [501] [502] [503] [504] the number and the size of the follicles in the spleen decreased significantly and the total number of cells decreased by more than three times; similar changes were found in lymph nodes. 478 however, these histopathologic splenic findings are in contradiction to earlier reports where no differences were described in infected and uninfected infants. 412 splenomegaly may occur in infants with late-onset sepsis and may be due to splenic congestion in the absence of hyperplasia of white pulp. 502 the mechanisms responsible for immune alterations after sepsis are beginning to emerge. the intensity of the inflammatory response may be modified by neural-based mechanisms. 515 t cell-secreted acetylcholine acts on macrophages to reduce production of tnf, il-1, il-18, hmgb-1, and other cytokines. 516 the role of vagal tone in the neonatal host response to sepsis is unclear. discovery and characterization of the impact of epigeneticmediated immune system functional alterations after sepsis is an area of intense research. dna methylation and posttranslational modification of histone proteins (methylation, acetylation, prevented hmgb-1 level elevation, and was associated with longer survival times. 559 increased plasma nitrite and nitrate concentrations are associated with the development of multiple organ failure in pediatric patients with sepsis 560,561 but have not been investigated in neonates. the incidence of neonatal sepsis remains high and outcomes remain poor despite considerable technologic advances in the field of neonatology. much remains to be learned about the impact of developmental age on the host response to sepsis and what facets are critically important. important considerations for future investigations include the development and implementation of a generally accepted definition for neonatal sepsis, the use of homogeneous systems (only neonatal components) for human ex vivo studies, and transgenic approaches in preclinical models, alongside observational studies in humans to ensure meaningful findings. complete reference list is available at www.expertconsult.com. the amplitude-integrated electroencephalogram may be helpful to predict neurologic outcome in infants with sepsis or meningitis. 543 significantly lower resistance, vasodilatation, and higher blood flow were noted in all the cerebral arteries of infants with sepsis. increase in cerebral blood flow velocity was correlated with elevated il-6 concentrations. 544 alterations in blood flow in preterm infants, in addition to factors associated with sepsis, such as respiratory distress, hypercarbia, hypotension, and patent ductus arteriosus, contribute to the risk for intracerebral hemorrhage. endocrine abnormalities may include altered thyroid function 545 and adrenal insufficiency associated with refractory hypotension. 546 inadequate adrenocortical responses are associated with increased mortality. 547, 548 cortisol production in the neonate is significantly increased early in septic shock. 212 however, very preterm neonates can have relative adrenal insufficiency that may contribute to hemodynamic instability and hypotension. hydrocortisone has not been evaluated in large prospective randomized clinical trials for the treatment of septic shock in the neonate but it has been shown to increase blood pressure, decrease heart rate, and decrease vasoactive medication requirements in preterm and term neonates in addition to its cytokine-suppressing effects. [549] [550] [551] if hydrocortisone treatment is considered, the obtaining of a pretreatment serum cortisol level is prudent in order to differentiate contributing causes of hypotension. sepsis was the most common cause (78%) of acute kidney injury in term neonates and was associated with high mortality (37%) (n = 49). 552 the frequency of acute renal failure (defined as a blood urea nitrogen level greater than 20mg/dl) in infected neonates was 26% and oliguria occurred in 15% of acute kidney failure cases. 553 acute kidney injury in preterm neonates is associated with high mortality. 554 hepatic injury and dysfunction are frequent associations with severe sepsis. the mechanisms include reduced hepatic perfusion associated with septic shock and mitochondrial energy failure. reductions in coagulation and complement factors, acute-phase reactant proteins, and increases in the levels of transaminases and bilirubin are commonly seen, especially in association with decreased perfusion states. energy expenditure and oxygen consumption are increased during sepsis, 555 and decreased mitochondrial oxidative function precipitated by hypoxia and the presence of reactive oxygen intermediates may lead to impaired growth, caloric deficiency, and energy failure. 556, 557 sepsis that leads to mods carries a dismal prognosis. inadequate cardiac output and microcirculatory failure, which may be combined with formation of microthrombi and dic, can lead to poor perfusion to the kidney, 472,473 liver, 474 gut, 475 and central nervous system. 52, 148, 181, 469, 476 recent studies suggest that the mechanism of organ failure in sepsis may relate to decreased oxygen utilization associated with mitochondrial dysfunction rather than poor oxygen delivery to tissues. 479, 480 mitochondrial dysfunction can initiate activation of cell death pathways, including apoptosis, pyroptosis, necrosis, and netosis (i.e. cell death mediated by nets). damage-associated molecular patterns (including nucleosomes and microparticles) created by activation of these cell death programs further amplify the host inflammatory response. free radicals play an important role in the inflammatory process of sepsis. 558 in a sepsis model in neonatal piglets, edaravone, a novel free radical scavenger, increased mean arterial pressure and cardiac output, lowered heart rate, reduced hydroperoxide, nitrite, and nitrate levels, delayed the tnf-α surge, 37 . jackson gl, rawiki p, sendelbach d, et al: hospital course and short-term outcomes of term and late preterm neonates following exposure to prolonged rupture of membranes and/or chorioamnionitis. pediatr infect dis j 31 (1) protective immunity and defects in the neonatal and elderly immune response to sepsis increased mortality and altered immunity in neonatal sepsis produced by generalized peritonitis defective innate immunity predisposes murine neonates to poor sepsis outcome but is reversed by tlr agonists the host response to sepsis and 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combined proinflammatory signals differential responses of cord and adult blood-derived dendritic cells to dying cells preterm neonates display altered plasmacytoid dendritic cell function and morphology efficient priming of antigen-specific cytotoxic t lymphocytes by human cord blood dendritic cells cd11c+ dendritic cells are required for survival in murine polymicrobial sepsis depletion of dendritic cells, but not macrophages, in patients with sepsis evaluation of eosinophilia in hospitalized preterm infants perinatal hematological profile of newborn infants with candida antenatal infections eosinophilia in newborn infants eosinophilia in premature neonates. phase 2 of a biphasic granulopoietic response catapult-like release of mitochondrial dna by eosinophils contributes to antibacterial defense mast cells in innate immunity mast cells and neutrophils release il-17 through extracellular trap formation in psoriasis urticaria neonatorum: accumulation of tryptase-expressing mast cells 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markers to detect neonatal infection immunosuppressive cd71+ erythroid cells compromise neonatal host defence against infection transferrin receptor 1 is differentially required in lymphocyte development transferrin therapy ameliorates disease in beta-thalassemic mice ineffective erythropoiesis in stat5a −/− 5b −/− mice due to decreased survival of early erythroblasts roles of spleen and liver in development of the murine hematopoietic system immaturity of the human splenic marginal zone in infancy. possible contribution to the deficient infant immune response fetal and neonatal development of human spleen: an immunohistological study neonatal cd71 + erythroid cells do not modify murine sepsis mortality caspase inhibitors improve survival in sepsis: a critical role of the lymphocyte b cells enhance early innate immune responses during bacterial sepsis neonatal adaptive immunity comes of age lymphocyte subpopulations in full-term septic neonates neonates with culture proven sepsis have lower amounts and percentage of cd45ra+ t cells pre-inflammatory mediators and lymphocyte subpopulations in preterm neonates with sepsis analysis of lymphocyte proliferative response subpopulations in very low birth weight infants and during the first 8 weeks of life hla-dr+, t lymphocytes and cd25+ cells in eutrophic full-term neonates with staphylococcal septicemia peripheral blood cells with expression of the interleukin-2 receptor (cd25+) in severely infected fullterm neonates cord blood cd4 + cd25 + -derived t regulatory cell lines express foxp3 protein and manifest potent suppressor function regulatory t cells mediate maternal tolerance to the fetus increased natural cd4+cd25+ regulatory t cells and their suppressor activity do not contribute to mortality in murine polymicrobial sepsis transient increase in cd45ro expression on t lymphocytes in infected newborns surface activation markers of t lymphocytes: role in the detection of infection in neonates can expression of cd45ro, a t-cell surface molecule, be used to detect congenital infection? increased cd4 + t cell co-inhibitory immune receptor ceacam1 in neonatal sepsis and soluble-ceacam1 in meningococcal sepsis: a role in sepsis-associated immune suppression? interleukin-8 (cxcl8) production is a signatory t cell effector function of human newborn infants increase of cord blood cytokineproducing t cells in intrauterine infection age-related changes in intracellular th1/th2 cytokine production, immunoproliferative t lymphocyte response and natural killer cell activity in newborns, children and adults identification of t-lymphocyte function in healthy vs. septic preterms and its relation to candidal infections in the hospital setting mature cd8 + t lymphocyte response to viral infection during fetal life asplenic-hyposplenic overwhelming sepsis: postsplenectomy sepsis revisited hyposplenism: a comprehensive review. part i: basic concepts and causes demonstration of circulating group b streptococcal immune complexes in neonates with meningitis in vitro immunoglobulin response of fetal b-cells is influenced by perinatal infections and antibiotic treatment: a study in preterm infants blood stream infection is associated with altered heptavalent pneumococcal conjugate vaccine immune responses in very low birth weight infants shades of grey-the blurring view of innate and adaptive immunity a critical role of t-cell receptor gamma/delta cells in antibacterial protection in mice early in life in vivo and in vitro activation and expansion of gammadelta t cells during listeria monocytogenes infection in humans gamma delta t cells respond directly to pathogen-associated molecular patterns influence of perinatal risk factors on cd3+/tcr αβ and cd3+/tcr γδ lymphocytes in cord blood of preterm neonates tcr usage and functional capabilities of human gamma delta t cells at birth il-23 activates innate lymphoid cells to promote neonatal intestinal pathology identification of pediatric septic shock subclasses based on genome-wide expression profiling genomic expression profiling across the pediatric systemic inflammatory response syndrome, sepsis, and septic shock spectrum divergence of canonical danger signals: the genome-level expression patterns of human mononuclear cells subjected to heat shock or lipopolysaccharide validating the genomic signature of pediatric septic shock genome-level longitudinal expression of signaling pathways and gene networks in pediatric septic shock gene-expression profiling of grampositive and gram-negative sepsis in critically ill patients the use of gene-expression profiling to identify candidate genes in human sepsis gene-expression profiling of peripheral blood mononuclear cells in sepsis recent progress of proteomics in critical illness proteomic profiling of the amniotic fluid to detect inflammation, infection, and neonatal sepsis optimized dna extraction from neonatal dried blood spots: application in methylome profiling validation of a gene expressionbased subclassification strategy for pediatric septic shock genome-level expression profiles in pediatric septic shock indicate a role for altered zinc homeostasis in poor outcome prophylactic zinc supplementation reduces bacterial load and improves survival in a murine model of sepsis oral zinc supplementation for reducing mortality in probable neonatal sepsis: a double blind randomized placebo controlled trial post-natal age is a critical determinant of the neonatal host response to sepsis genome-wide expression profiles in very low birth weight infants with neonatal sepsis a novel role for matrix metalloproteinase-8 in sepsis identification of a human neonatal immune-metabolic network associated with bacterial infection host-response biomarkers for diagnosis of late-onset septicemia and necrotizing enterocolitis in preterm infants fetal thymic involution: a sonographic marker of the fetal inflammatory response syndrome persistent microcirculatory alterations are associated with organ failure and death in patients with septic shock risk factors and predictors of mortality in culture proven neonatal sepsis anatomical study of the kidneys of newborn infants dying after a septic state renal failure, comorbidity and mortality in preterm infants liver perfusion in sepsis, septic shock, and multiorgan failure neonatal sepsis in karachi: factors determining outcome and mortality association of septic shock caused by early-onset group b streptococcal sepsis and periventricular leukomalacia in the preterm infant accelerated lymphocyte death in sepsis occurs by both the death receptor and mitochondrial pathways pathology of lymphoid organs in low birth weight infants subjected to antigen-related diseases: a morphological and morphometric study advances in pathogenesis and management of sepsis molecular biology of inflammation and sepsis: a primer multisystem organ failure and capillary leak syndrome in severe necrotizing enterocolitis of very low birth weight infants clinical and laboratory factors that predict death in very low birth weight infants presenting with late-onset sepsis heart failure in pediatric septic shock: utilizing inotropic support circulatory shock in children: an overview hemodynamic support in fluidrefractory pediatric septic shock clinical practice parameters for hemodynamic support of pediatric and neonatal patients in septic shock distinct hemodynamic patterns of septic shock at presentation to pediatric intensive care which inotrope for which baby? relationship between blood pressure and cardiac output in preterm infants requiring mechanical ventilation microcirculatory changes in term newborns with suspected infection: an observational prospective study hemodynamics in preterm infants with late-onset sepsis echocardiogram done early in neonatal sepsis: what does it add? el-arman mm: myocardial dysfunction in neonatal sepsis: a tissue doppler imaging study diagnosis and treatment of neonatal hypotension outside the transitional period elevation of brain natriuretic peptide levels in children with septic shock brain natriuretic peptide for prediction of mortality in patients with sepsis: a systematic review and meta-analysis predictors of mortality and multiple organ failure in children with sepsis sepsis-induced apoptosis causes progressive profound depletion of b and cd4+ t lymphocytes in humans acute thymus involution in infancy and childhood: a reliable marker for duration of acute illness thymus size and its relationship to perinatal events the grade of acute thymus involution in neonates correlates with the duration of acute illness and with the percentage of lymphocytes in peripheral blood smear. pathological study clinicopathological differences between early-onset and late-onset sepsis and pneumonia in very low birth weight infants spleen depletion in neonatal sepsis and chorioamnionitis acute thymic involution in fetuses and neonates with chorioamnionitis prolonged lymphopenia, lymphoid depletion, and hypoprolactinemia in children with nosocomial sepsis and multiple organ failure immunosuppression in sepsis: a novel understanding of the disorder and a new therapeutic approach prevention of lymphocyte apoptosisa potential treatment of sepsis? mutations in genes required for t-cell development: il7r, cd45, il2rg, jak3, rag1, rag2, artemis, and ada and severe combined immunodeficiency: huge review early circulating lymphocyte apoptosis in human septic shock is associated with poor outcome immunosuppression in patients who die of sepsis and multiple organ failure immunoparalysis and adverse outcomes from critical illness very low birth weight neonates who survive early-onset sepsis do not have an increased risk of developing late-onset sepsis early sepsis does not increase the risk of late sepsis in very low birth weight neonates the pathophysiology and treatment of sepsis nicotinic acetylcholine receptor α7 subunit is an essential regulator of inflammation neural reflexes in inflammation and immunity epigenetic regulation of immune cell functions during post-septic immunosuppression dna methylation pattern of calca in preterm neonates with bacterial sepsis as a putative epigenetic biomarker a prime time for trained immunity: innate immune memory in newborns and infants epigenetic programming of monocyteto-macrophage differentiation and trained innate immunity mtor-and hif-1alpha-mediated aerobic glycolysis as metabolic basis for trained immunity nonspecific effects of neonatal and infant vaccination: public-health, immunological and conceptual challenges myd88-dependent expansion of an immature gr-1 + cd11b + population induces t cell suppression and th2 polarization in sepsis myeloid derived suppressor cells are present at high frequency in neonates and suppress in vitro t cell responses reactivation of multiple viruses in patients with sepsis surfactant-replacement therapy for respiratory distress in the preterm and term neonate diminished inducible nitric oxide synthase expression in fulminant early-onset neonatal pneumonia the paradox of adult respiratory distress syndrome in neonates bench-to-bedside review: developmental influences on the mechanisms, treatment and outcomes of cardiovascular dysfunction in neonatal versus adult sepsis inhaled nitric oxide and persistent pulmonary hypertension of the newborn the diseases treated with ecmo: focus on pphn extracorporeal membrane oxygenation and sepsis neurodevelopment of extremely preterm infants who had necrotizing enterocolitis with or without late bacteremia neonatal infection and 5-year neurodevelopmental outcome of very preterm infants impact of sepsis on neurodevelopmental outcome in a swiss national cohort of extremely premature infants infection-induced inflammation and cerebral injury in preterm infants postnatal sepsis, necrotizing entercolitis, and the critical role of systemic inflammation in white matter injury in premature infants postnatal infection is associated with widespread abnormalities of brain development in premature newborns adverse neurodevelopment in preterm infants with postnatal sepsis or necrotizing enterocolitis is mediated by white matter abnormalities on magnetic resonance imaging at term association between high cytokine levels with white matter injury in preterm infants with sepsis the relationship of csf and plasma cytokine levels to cerebral white matter injury in the premature newborn to tap or not to tap: high likelihood of meningitis without sepsis among very low birth weight infants the prognostic value of amplitude integrated eeg in neonatal sepsis and/or meningitis cerebral blood flow velocity in earlyonset neonatal sepsis and its clinical significance serum cortisol and thyroid hormone levels in neonates with sepsis refractory hypotension in preterm infants with adrenocortical insufficiency hydrocortisone administration for the treatment of refractory hypotension in critically ill newborns circulating adrenocorticotropic hormone (acth) and cortisol concentrations in normal, appropriate-forgestational-age newborns versus those with sepsis and respiratory distress: cortisol response to low-dose and standard-dose acth tests hemodynamic changes after low-dosage hydrocortisone administration in vasopressor-treated preterm and term neonates cardiovascular effects of hydrocortisone in preterm infants with pressor-resistant hypotension dose-related inhibition of proinflammatory cytokine release from neutrophils of the newborn by dexamethasone, betamethasone, and hydrocortisone the main etiologies of acute kidney injury in the newborns hospitalized in the neonatal intensive care unit acute renal failure in neonatal sepsis acute kidney injury in preterm infants admitted to a neonatal intensive care unit effect of sepsis syndrome on neonatal oxygen consumption and energy expenditure nitric oxide inhibits neonatal hepatocyte oxidative metabolism impaired energy metabolism during neonatal sepsis: the effects of glutamine bench-to-bedside review: neonatal sepsis -redox processes in pathogenesis edaravone, a novel free radical scavenger, reduces high-mobility group box 1 and prolongs survival in a neonatal sepsis model inflammatory cytokine and nitric oxide responses in pediatric sepsis and organ failure plasma nitrite and nitrate concentrations and multiple organ failure in pediatric sepsis key: cord-020643-0yzkqykg authors: müller-werdan, u.; buerke, m.; christoph, a.; flieger, r.r.; loppnow, h.; prondzinsky, r.; reith, s.; schmidt, h.; werdan, k. title: schock date: 2006 journal: klinische kardiologie doi: 10.1007/3-540-29425-2_6 sha: doc_id: 20643 cord_uid: 0yzkqykg nan pathophysiologie des schocks -347 therapie der metabolischen azidose -375 6.4.6 volumentherapie -375 6.4.7 inotrope und vasoaktive substanzen -379 6.4.8 optimierung des intravasalvolumens -383 6.4.9 neue therapieansätze zur schockbehandlung -383 6.4.10 gerinnungsaktive substanzen -383 6.4.11 hypothermie/cooling -384 6.4.12 pharmakotherapie des kritisch kranken -384 6.5 multiorgandysfunktionssyndrom (mods) -384 6.5.1 hypothesen zur entstehung -384 6.5.2 schweregradeinteilung durch score-systeme -386 6.5.3 organdysfunktion der lunge -389 6.5.4 organdysfunktion der niere (akutes nierenversagen) -390 6.5.5 organdysfunktion des gehirns -393 6.5.6 organdysfunktion des peripheren und autonomen nervensystems sowie der skelettmuskulatur -393 6.5.7 organdysfunktion des herzens und des kreislaufs -394 6.5.8 organdysfunktion des gastrointestinaltrakts -394 6.5.9 dysfunktion des gerinnungssystems -395 6.5.10 dysfunktion des stoffwechsels und künstliche ernährung -397 6.5.11 dysfunktion des endokrinium -397 6.5.12 dysfunktion des immunsystems -398 6.6 spezifi sche schockformen -398 6.6.1 kardiogener schock -398 6.6.2 septischer schock -409 6.6.3 hypovolämischer schock -418 6.6.4 traumatischer schock -419 6.6.5 anaphylaktischer schock -419 die reduktion der effektiven gewebeperfusion kann dabei hervorgerufen werden durch eine abnahme der herzleistung, durch störungen der makro-und mikrozirkulation mit verminderter perfusion, durch eine kritische abnahme des blutvolumens, eine beeinträchtigung der o 2 -aufnahme in der lunge und der o 2 -abgabe im gewebe. zur beeinträchtigung essenzieller zellfunktionen kommt es im gefolge einer exogenen intoxikation, einer endogenen überschwemmung mit mediatoren (7 abschn. 6.2.6) oder endokrin/metabolischer krisen. klar zu trennen vom schockbegriff sind die synkope und der kollaps. eine passagere kritische herabsetzung der gehirndurchblutung mit gleichzeitiger bewusstlosigkeit wird als synkope bezeichnet (task force on syncope, european society of cardiology 2001). nicht immer ist damit eine allgemeine kreislaufi nsuffi zienz vergesellschaftet. bei einem kollaps über-wiegen vagale reaktionen, das ereignis ist von kurzer dauer, organschäden infolge o 2 -mangels treten nicht auf. von den zahlreichen schockklassifi kationsversuchen (adams et al. 2001; kumar u. parrillo 2001) erscheint die schockeinteilung nach hämodynamischen gesichtspunkten für den kliniker am hilfreichsten (. tabellen 6.1, 6.2 und . abb. 6.1): hypovolämischer schock infolge eines im verhältnis zur gefäßkapazität verminderten zirkulierenden blutvolumens, charakterisiert durch erniedrigte diastolische füllungsdrücke und -volumina des herzens; kardiogener schock infolge eines pumpversagens, hervorgerufen durch eine eingeschränkte myokardkontraktilität, einen verlust an funktionsfähiger myokardmasse oder eine intrinsischstrukturelle bzw. -mechanische behinderung der herzfunktion, charakterisiert durch erhöhte diastolische füllungsdrücke und -volumina des herzens; extrakardial-obstruktiver schock infolge einer flussobstruktion im herz-kreislauf-system, charakterisiert entweder durch eine behinderung der diastolischen füllung oder eine exzessive nachlasterhöhung; verteilungsschock infolge eines verlustes der vasomotorenkontrolle mit dilatation von arteriolen und venolen, charakterisiertnach adäquater volumensubstitution -durch einen erhöhten herzindex und einen erniedrigten systemischen gefäßwiderstand. f f f f abb. 6.1. zusammenspiel von schockursachen bei verschiedenen schockformen. im falle des kardiogenen, hypovolämischen und obstruktiven schocks resultiert die hypotonie im wesentlichen aus der abnahme des herzindex, mit konsekutivem anstieg des systemischen gefäßwiderstands. im falle des distributiven -insbesondere des septischen -schocks ist die hypotonie v. a. auf die erniedrigung des systemischen gefäßwiderstands zurückzuführen mit sekundärem anstieg des . herzindex. bei zahlreichen schockformen wird das hämodynamische profi l durch komponenten der hypovolämie, der myokarddepression (ischämisch und infolge anderer ursachen) und der gefäßdysfunktion (mit einfl uss auf die nachlast) mitbeeinfl usst. bei einem volumenverlust von 15-35% (stadium ii) beginnen die kompensationsmechanismen zu versagen, es treten eine milde bis mäßige hypotonie, eine abnahme des herzindex und eine orthostase auf. der systemische gefäßwiderstand nimmt beträchtlich zu, und das serumlaktat kann ansteigen. traumapatienten, die 35-40% ihres blutvolumens verloren haben (stadium iii), weisen eine ausgeprägte tachykardie, hypotension und bewusstseinsstörungen auf. ein hypovolämischer schock manifestiert sich bei einem verlust von über 40% (stadium iv) des zirkulierenden volumens, es resultiert ein abfall des herzindex und der gewebeperfusion auf weniger als die hälfte; die auftretende laktatazidose kündigt eine ungünstige prognose mit in der regel irreversiblem hämorrhagischem schock an. besteht ein blutverlust von 40% oder mehr über einen zeitraum von 2 h oder sogar länger, so muss bereits mit der erfolglosigkeit der dann eingeleiteten maßnahmen gerechnet werden. prognostisch ungünstige faktoren beim hypovolämischen schock sind die geschwindigkeit des auftretens des volumenverlustes (je rascher, desto ungünstiger) und eine vorbestehende einschränkung der herzfunktionsreserve (z. b. bei zustand nach herzinfarkt). povolämischen schock die eingeleitete therapie trotz adäquater volumensubstitution nicht mehr erfolgreich ist. blutverluste, plasmaverluste und exogene flüssigkeits-/wasserverluste zählen zu den häufi gsten ursachen dessen, was sich in komplizierter verkettung der sekundären reaktionen und regulationen klinisch-symptomatologisch als »hypovolämischer schock« manifestiert. der kardiogene schock ist die häufi gste todesursache von infarktpatienten in der krankenhausphase (hollenberg 2003; prondzinsky et al. 2004) . klinisch fi ndet sich ein zentralisierter kreislaufschock mit low-output-syndrom (hypotonie, oligurie, blasse, kühle haut) und den symptomen der akuten links-und rechtsherzinsuffi zienz mit 3. herzton, halsvenenund lungenstauung bis hin zum lungenödem. hämodynamisch (. tabelle 6.2) imponieren die zunahme der vorlast (zunahme von ventrikelvolumina, linksventrikulärem füllungsdruck, pulmonalkapillardruck und zentralem venendruck) und eine abnahme von herzindex, schlagvolumenindex und schlagarbeitsindex, verbunden mit einem anstieg des systemischen gefäßwiderstandes. als kriterien des infarktbedingten kardiogenen schocks (hochman et al. 1999; prondzinsky et al. 2004) myokardfunktionsverlust. die häufi gste ursache eines kardiogenen schocks (. tabelle 6.4) ist ein mindestens 40%iger myokardfunktionsverlust im rahmen einer khk: beim akuten, großen myokardinfarkt -häufi g bei proximalem verschluss des r. interventricularis anterior (riva) der linken koronararterie oder der lad (»left anterior descendens«)aber auch bei rezidivinfarkten und in der stunned-myocardium-phase. akute mitralinsuffi zienz. eine weitere ursache eines kardiogenen schocks ist die akute mitralinsuffi zienz infolge einer sehnenfadenruptur oder einer papillarmuskeldysfunktion/ruptur bei myokardischämie, akutem herzinfarkt (häufi g infolge eines lad-verschlusses), endokarditis, stumpfem thoraxtrauma, mitralklappenprolaps oder klappenprothesendysfunktion. die ischämische papillarmuskelruptur tritt häufi g 3-7 tage nach akutem herzinfarkt auf; ein neu aufgetretenes apikales holosystolikum kann der ruptur vorausgehen. akute aorteninsuffi zienz. als schockursache ist die akute aorteninsuffi zienz meistens auf eine fl oride endokarditis zurückzuführen, weiterhin auf eine prothesendysfunktion oder eine aortendissektion. > tabelle 6.4. ursachen des infarktbedingten kardiogenen schocks; daten von 1422 patienten des shock trial und shock registry a (hochman et al. 2000; mod. nach prondzinsky et al. 2004) . häufi gkeit (%) ventrikelruptur. die infarktbedingte ventrikelseptumruptur tritt ebenfalls wenige tage nach dem infarkt, häufi g bei lad-verschluss, auf. bei einem therapiefraktären hinterwandinfarkt ist auch an das vorliegen eines begleitenden (selten eines isolierten) rechtsherzinfarkts zu denken, der in 10-20% der fälle zum kardiogenen schock führt (zehender et al. 1993 ). weitere ursachen. weitere auslöser des auftretens eines kardiogenen schocks sind akute myokarditis, terminalstadien von kardiomyopathien, dekompensiertes hochdruckherz und vitien, maligne brady-und tachykarde rhythmusstörungen sowie traumatisch bedingte myokardkontusionen und intoxikationen mit negativ-inotropen pharmaka. die klinische symptomatik und die hämodynamischen befunde sind geprägt durch die kombination krankheitsspezifischer befunde mit einem low-output-syndrom. auch bei dieser schockform ist der zeitverlauf prognosebestimmend. bei einer akuten perikardtamponade durch ischämische ruptur der freien ventrikelwand (in der regel 5-7 tage nach herzinfarkt) oder infolge einer blutung bei thoraxtrauma bzw. nach thrombolysetherapie reichen bereits 150 ml blut im perikardbeutel zur schockentwicklung aus; bei einem chronischen malignen oder entzündlichen perikarderguss können dagegen 1-2 l ergussfl üssigkeit ohne schockentwicklung toleriert werden. bei einer akuten lungenembolie ohne vorbestehende kardiopulmonale erkrankung führt der verschluss von 2 oder mehr lappenarterien und damit von 50-60% des lungengefäßbettes zum schock; im falle chronischer, rezidivierender lungenembolien mit adaptativer rechtsherzhypertrophie werden teilweise wesentlich höhere anteile an lungenstrombettverschlüssen ohne schocksymptomatik toleriert. der gemeinsame nenner des verteilungsschocks (. tabelle 6.1) ist der verlust an peripherem gefäßwiderstand. neben den allgemeinen schockzeichen mit arterieller hypotonie, tachykardie, tachypnoe, oligurie und bewusstseinsstörung imponieren warme, relativ gut durchblutete extremitäten und ein erniedrigter diastolischer blutdruck. anamnese und charakteristische befunde können häufi g eine rasche ätiologische einordnung ermöglichen: eine urtikaria spricht für einen anaphylaktischen, eine rückenmarksverletzung für einen neurogenen und zeichen der infektion für einen septischen schock, der weitaus häufi gsten form des verteilungsschocks. hämodynamisch imponiert beim verteilungsschock eine ausgeprägte erniedrigung des systemischen gefäßwiderstandes, wobei jedoch in den einzelnen organgefäßbetten der widerstand erniedrigt, unverändert und auch erhöht sein kann. initial (vor volumentherapie) können herzindex und ventrikuläre füllungsdrücke erniedrigt sein, danach sind sie in der regel erhöht (. tabelle 6.2). der septische schock, die häufi gste todesursache auf der intensivstation, ist folge einer toxin-/mediatorkaskade, die sowohl durch gramnegative als auch durch grampositive bakterien ausgelöst werden kann, ebenso polymikrobiell und in seltenen fällen auch durch pilze oder viren (7 abschn. 6.2.6). 64% der patienten mit einer sepsis entwickeln innerhalb von 24 h (median) eine schwere sepsis, und 23% der patienten mit schwerer sepsis innerhalb der nächsten 28 tage (median) einen septischen schock (moerer et al. 2005) . die letalität der sepsis (ca. 10-15%) steigt bei der schweren sepsis auf ca. 30% und beim septischen schock auf 50-60% an. circa 50% aller sepsistodesfälle sind auf ein intraktables mods zurückzuführen, 40% auf ein therapierefraktäres kreislaufversagen, und bei 10% ist eine nicht beherrschbare septische kardiomyopathie (7 abschn. 6.6.2) die ursache. an der verbesserung der sepsisdefi nitionen und -klassifizierung (. tabelle 6.3) wird permanent gearbeitet: die piro-klassifi zierung ist ausdruck des bemühens um eine bessere charakterisierung der sepsis ähnlich der tumorklassifi zierung (levy et al. 2003) . wesentlich aussagekräftiger als die qualitative sepsisdiagnose (. tabelle 6.3) sind quantitative parameter zur beschreibung des schweregrads der sepsis (z. b. der sepsis-score nach elebute u. stoner, . abb. 6.22), des septischen multiorganversagens (z. b. der apache-ii-schweregrad-score, . abb. 6.15), des ausmaßes der septischen gefäßschädigung (abfall des systemischen gefäßwiderstands) und der septischen myokarddepression (abnahme des linksventrikulären schlagarbeitsindex). sie erlauben nicht nur eine abschätzung der prognose der sepsispatienten, sondern auch des ansprechens auf die therapie (müller. . tabelle 6.5 zeigt das charakteristische profi l von sepsispatienten im vergleich zu patienten mit fieber, aber lediglich lokaler infektion und im vergleich zu patienten mit kardialem pumpversagen. die sepsispatienten sind durch einen hohen sepsis-score, eine hyperdyname kreislaufsituation mit hohem herzindex bei erniedrigtem systemischem gefäßwiderstand und schlagarbeitsindex und durch störungen der o 2 -verwertung gekennzeichnet. die inzidenz der schweren sepsis liegt bei 6/1000 krankenhaus-und 24-136/1000 intensivstationaufnahmen, die inzidenz des septischen schocks bei 1,4/1000 krankenhausaufnahmen und 61-87/1000 intensivaufnahmen (moerer et al. 2004) . im verständnis der sepsis und des septischen schocks ist allerdings in den letzten jahren ein erheblicher wandel eingetreten (. abb. 6.2). stand früher die bakterielle oder pilzinfektion mit einschwemmung von keimen in die blutbahn ganz im vordergrund der betrachtungsweise, so sind es heute vielmehr die durch die mikrobiellen toxine via aktivierung von mediatorzellen induzierten zytokin-und mediatorkaskaden, die für das mods und den septischen schock verantwortlich gemacht werden (7 abschn. 6.2.6). diese einschätzung wird unterstützt durch die tatsache, dass nur bei jedem 2. oder 3. patienten mit sepsis eine positive kultur gefunden wird und diese auch nicht prognosebestimmend ist. auch dass nichtinfektiöse noxen (trauma, pankreatitis, herzchirurgische operationen mit der herz-lungen-maschine) zu einem ganz ähnlichen klinischen bild wie bei bakteri-ell ausgelöster sepsis und septischem schock führen können, spricht für eine mehr oder weniger gemeinsame zytokin-/mediatorendstrecke als verantwortliche schädigungskaskade sowohl bei infektiösen als auch bei nichtinfektiösen (sirs, . tabelle 6.3) systemisch-entzündlichen schockformen. der anaphylaktische schock ist ein akut eintretender, »warmer« schockzustand, der durch anaphylaktische und anaphylaktoide reaktionen ausgelöst wird: der blutdruckabfall infolge vasodilatation mit relativer hypovolämie kann einhergehen mit larynxödem, bronchospasmus, angioödem, urtikaria, erythemen und myokarddepression. tabelle 6.5. herz-kreislauf-veränderungen und o 2 -metabolismus bei patienten mit sepsis, fieber und kardialem pumpversagen. (nach werdan et al. 1991; boekstegers et al. 1994) . sepsis fieber kardiales pumpversagen (n=14) (n=11) (n=7) sepsis-score (elebute) 22 (4) 13 (4) 13 (3) multiorganversagen-score (apache ii) 32 (6) 26 (4) 29 (3) hämodynamik herzindex (l/min/m 2 ) 4,8 (1,4) 3,0 (0,5) 2,3 (0,2) linksventrikulärer schlagarbeitsindex (g · m/m 2 ) 37,5 (12,5) 36,6 (6,7) 21,4 (4,3) systemischer gefäßwiderstand (dyn · s · cm -5 ) 597 (149) klassische anaphylaktische reaktionen sind ige-vermittelte allergische ereignisse in reaktion auf ein antigen, entsprechend einer typ-i-reaktion nach gell u. coombs, die perakut und generalisiert systemisch ablaufen. antibiotika, insekten-und schlangengifte, impfstoffe, seren, jodhaltige kontrastmittel und nahrungsmittel gehören zu den typischen auslösenden allergenen. ige-spezifi sche effektorzellen der immunantwort sind im wesentlichen mastzellen und basophile, die nach stimulation eine vielzahl proinfl ammatorischer mediatoren freisetzen und damit das klinische erscheinungsbild der anaphylaxie hervorrufen. davon abzugrenzen sind ige-unabhängige unverträglichkeitsreaktionen ohne vorausgehende sensibilisierung mit einem sehr ähnlichen oder identischen klinischen erscheinungsbild: bei anaphylaktoiden reaktionen (typischerweise ausgelöst z. b. durch röntgenkontrastmittel, salizylate und opiate) kommt es durch chemische, physikalische oder osmotische stimuli zur mediatorfreisetzung aus mastzellen und basophilen. eine idiopathische anaphylaxie kann typischerweise bei jungen erwachsenen auftreten, häufi g nachts oder postprandial; auslösefaktoren und effektorzellen sind unbekannt. die anaphylaxis factitia wird dem münchhausen-syndrom zugerechnet. daneben wurden auch immunreaktionen vom typ iii nach gell u. coombs bei anaphylaktischen reaktionen beschrieben, bei denen komplexe aus igg und spezifi schem antigenkomplement komplement aktivieren und über die anaphylatoxine c3a und c5a die mediatorfreisetzung aus mastzellen und basophilen stimulieren. charakteristischerweise tritt diese reaktion bei patienten mit hereditärem iga-mangel, dem häufi gsten angeborenen defekt des immunsystems (1:500-700), im rahmen von bluttransfusionen auf: präformierte antikörper gegen iga im serum dieser patienten können die mediatorkaskaden auslösen. der begriff »anaphylaktoide reaktion« kann auch als überbegriff verwandt werden für akute unverträglichkeitsreaktionen mit den symptomen einer anaphylaxie, ohne damit eine aussage zum pathomechanismus zu implizieren walther u. böttiger 2004) . genaue zahlen zur inzidenz anaphylaktischer und anaphylaktoider reaktionen sind aufgrund deren unvorhersehbarkeit und unberechenbarkeit nicht bekannt walther u. böttiger 2004) . entsprechend einer retrospektiven epidemiologischen studie muss mit etwa 21 anaphylaxiefällen pro 100.000 einwohner gerechnet werden, wobei zu 36% nahrungsmittel, zu 17% medikamente, diagnostika oder immuntherapeutika, zu 17% insektenstiche und zu 15% unklare ursachen zur anaphylaxie führten (yocum et al. 1999 ). antibiotika, jodhaltige kontrastmittel. etwa bei 1 von 2700 hospitalisierten patienten kommt es nach einer älteren studie zur medikamenteninduzierten anaphylaxie. nach schätzungen ist bei verabreichung von penicillin bei einem von 10.000 patienten mit einer anaphylaxie zu rechnen, die in 9% der fälle tödlich verläuft, so dass jährlich in den usa mit mehreren hundert todesfällen zu rechnen ist. auch die einnahme von cephalosporinen, neueren β-laktamantibiotika und fluoro-chinolonen kann zur anaphylaxie führen. es wird geschätzt, dass etwa 3-7% der patienten mit penicillinallergie kreuzreaktionen gegen ein cephalosporin aufweisen. etwa 200-800 todesfälle gehen in den usa jährlich zu lasten jodhaltiger kontrastmittel. tierische gifte. typische verursacher der anaphylaxie sind insektengifte, übertragen durch stiche der tiere der ordnung hymenoptera (u. a. bienen, wespen, hornissen), und schlangengifte (z. b. klapperschlangen, mokkassinschlangen), die neben toxischen auch schwere allergische reaktionen hervorrufen können. etwa 0,5-5% der bevölkerung haben schon eine schwere allergische reaktion auf einen insektenstich durchgemacht, und 1% dieser reaktionen kann in eine lebensbedrohliche anaphylaxie münden. nahrungsmittel. in der praxis ist bei akuten reaktionen nach mahlzeiten die abgrenzung allergischer reaktionen zu nahrungsmittelintoleranzen und bakterientoxinerkrankungen schwierig. nahrungsmittelallergien bestehen bei etwa 1-6% der kinder und sind im erwachsenenalter seltener anzutreffen. schwere anaphylaktische reaktionen auf nahrungsmittel sind eher selten, sind jedoch gehäuft für erdnüsse, sojabohnen, eiweiß und schalentiere beschrieben worden. latex, exotische früchte. zunehmend ist die inzidenz latexallergischer anaphylaxien. diese können durch die benutzung von latexhandschuhen ausgelöst werden, es sind aber auch bei einführen von kathetern aus latex und bei kondomen anaphylaxien beschrieben worden. auch wenn keine exakten epidemiologischen daten vorliegen, so ist davon auszugehen, dass etwa 7-18% der ärzte und des pfl egepersonals auf latex allergisch reagieren. gefährdet sind neben arbeitern aus der latex-verarbeitenden industrie v. a. patienten, die sich mehreren operativen eingriffen unterzogen haben, ganz besonders spina-bifi da-patienten, die zu >50% sensibilisiert sind. ein hohes allergiesierungspotenzial haben auch schleimhautkontakte, z. b. bei urogenitalen katheterisierungen oder bariumkontrasteinläufen. inzwischen stehen kommerzielle tests zur verfügung, die präoperativ innerhalb weniger stunden eine aussage darüber zulassen, ob bei einem patienten latex-spezifi sche ige-antikörper im serum vorhanden sind. überzufällig häufi g ist die latexallergie vergesellschaftet mit nahrungsmittelallergien, häufi g gegen exotische früchte wie avocado, banane, kiwi, passionsfrucht oder auch kastanien. das risiko unerwünschter und multipler arzneimittelwirkungen erhöht. allergien. häufi g erlebt man anaphylaktische reaktionen bei allergikern und häufi ger nach intravenöser als nach oraler allergenzufuhr, weshalb der anaphylaktische schock nicht selten iatrogen verursacht wird. narkosemittel, muskelrelaxanzien. anaphylaktoide narkosezwischenfälle bis hin zum herzstillstand treten nach einer umfangreichen französischen studie mit einer inzidenz von 1:4500-1:6000 allgemeinanästhesien auf. in ca. 6% der fälle kam es trotz adäquater therapie zum tod des patienten. auslösende agenzien waren in 60-70% der fälle muskelrelaxanzien, in ca. 18% latexprodukte und in ca. 5% kolloidale volumenersatzmittel. aufgrund struktureller besonderheiten (quartäre ammoniumgruppe) können alle muskelrelaxanzien unverträglichkeitsreaktionen hervorrufen. aber auch opioide, lokalanästhetika und narkotika wurden als potenziell anaphylaktoid wirkende agenzien herausgestellt. kumulativer mediatoreneffekt. der kumulative effekt der freigesetzten mediatoren besteht im wesentlichen in einer erhöhten gefäßpermeabilität, einer ausgeprägten vasodilatation und einem bronchospasmus. autoptisch wurde bei tödlich verlaufenden anaphylaxien ein ödem der lungen mit oftmals fl üssigkeitsgefüllten alveolen, ein ödem der oberen atemwege, einschließlich des larynx und der epiglottis, der haut und der viszeralen organe gefunden. im zusammenhang mit ödemen der oberen atemwege kommt es oft zu einer pulmonalen überblähung oder auch zu einer ausgeprägten bronchokonstriktion. der neurogene schock wird durch einen verlust der peripheren vasomotorenkontrolle infolge einer dysfunktion oder einer verletzung des nervensystems hervorgerufen. das klassische beispiel dafür ist der meist mit einer rückenmarkverletzung assoziierte spinale schock (hund u. abel 2002) . dem verlust des venentonus mit erhöhter venöser kapazität scheint hier die entscheidende bedeutung zuzukommen, wobei auch der arteriolentonus reduziert sein kann. in letzterem falle kommt es nach flüssigkeitssubstitution zu einer steigerung des herzindex. eine -allerdings selbstlimitierende und vorübergehende -neurale fehlregulation fi ndet sich auch als ursache der vasovagalen synkope und bei der spinalanästhesie. die addison-krise kündigt sich mit unspezifi schen symptomen an wie anorexie, übelkeit, erbrechen, diarrhöen, abdominellen schmerzen, myalgien, gelenkschmerzen, kopfschmerzen, schwäche, verwirrtheit und agitation oder delir. bei exsikkose kann es zu hohem fieber kommen. das initiale hämodynamische profi l ähnelt dem des hypovolämischen schocks; nach adäquater volumensubstitution demaskiert sich ein hyperdynamer, vasopressorrefraktärer schock (claussen et al. 1992; parkar u. taylor 2001) , der einem septischen schock ähneln kann. überlagern sich beide schockformen, so sind das erkennen der nebenniereninsuffi zienz (kortisolbestimmung, acth-test) und die eingeleitete kortikoidbehandlung oft lebensrettend. zur addison-krise (parkar u. taylor 2001) kann es einerseits bei patienten mit chronischer, substituierter nebennierenrindeninsuffi zienz bei besonderen krankheits-oder operationsbelastungen ohne ausreichende steigerung der kortikoidmedikation kommen; andererseits kann auch eine beidseitige nebennierenblutung im gefolge einer sepsis (meningokokken, gramnegative bakterien), einer hiv-infektion, einer pilzinfektion, einer malignen infi ltration, eines anaphylaktischen schocks (lefevre et al. 1996) oder auch eine einblutung unter antikoagulation zur akuten nebenniereninsuffi zienz führen (rao et al. 1989 ). seltene schock-und synkopenformen sind in der . übersicht 6.1 aufgeführt. mit dem begriff »vasalperipherer schock« werden jene akuten zustände von kreislaufi nsuffi zienz bezeichnet, de-. der verlust von myokardmasse beim akuten herzinfarkt mit minderung der kontraktilität führt zur abfl achung der frank-starling-kurve und zur zunahme des vorhofdrucks (. abb. 6.4a, a-b). durch das positiv-inotrop wirkende dobutamin (ohne wesentlichen einfl uss auf den venösen rückfl uss) kann die kontraktionskraft des herzens gesteigert und damit der herzauswurf erhöht werden (. abb. 6.4a, b-c). im gegensatz zum dobutamin würden die katecholamine noradrenalin und dopamin nicht nur positiv-inotrop wirken, sondern auch die venöse kapazität reduzieren und damit den mittleren zirkulationsdruck erhöhen. hypovolämie vermindert das zirkulierende blutvolumen und damit auch den mittleren zirkulationsdruck pcm (. abb. 6.4b, a-b). die resultierende abnahme des venösen rückstroms und damit des herzauswurfs kann durch volumensubstitution ausgeglichen werden (. abb. 6.4b, b-c). das hämodynamische bild des hämorrhagischen hypovolämischen schocks ist nicht nur durch den blutvolumenverlust geprägt, sondern auch durch eine initiale kompensatorische vasokonstriktion, die im weiteren verlauf von einer therapeutisch schwer beeinfl ussbaren vasodilatation gefolgt wird. für letztere wird zum einen eine hochregulation der spinalen stickoxidsynthase verantwortlich gemacht (lu et al. 1999 ; 7 abschn. 6.2.6), zum anderen eine aktivierung der poly(adp-ribose)-synthetase (pars), einem schlüsselenzym der apoptose (7 abschn. 6.2.7). der septische schock (thiemermann 2000a) vor adäquater volumensubstitution reicht die abnahme des rv allerdings nicht aus, um die reduktion des pcm auszugleichen: der herzauswurf bleibt vermindert (. abb. 6.4c, a-b). durch eine adäquate volumensubstitution lässt sich der pcm normalisieren; aufgrund des weiterhin erniedrigten venösen widerstands (rv) führt dies, bei fehlen einer myokarddepression, dann sogar zu einem supranormalen venösen rückstrom und einem supranormalen herzzeitvolumen (. abb. 6.4c, b-c). die graphik verdeutlicht, dass die rv-abnahme bis zu einem gewissen grad eine meist vorhandene mäßige myokarddepression maskieren kann. erst bei einer ausgeprägten kontraktilitätsminderung wird diese als einschränkung des herzauswurfs manifest (. abb. 6.4c, c-d), was bei ca. 20% aller sepsispatienten zutrifft (kumar u. parrillo 2001) . die mikrozirkulation und deren erkrankungsbedingte störungen können heutzutage beim patienten mit der orthogonalen polarisations-spektral-bildgebung (»orthogonal polarization spectral imaging«, ops; groner et al. 1999 ) sichtbar gemacht werden (. abb. 6.5). eine regelrechte durchblutung auf mikrozirkulationsebene (gefäße bis 100-150 µm durchmesser) ist voraussetzung für eine adäquate gewebeperfusion; ein normales herzzeitvolumen und ein normaler blutdruck sind dafür noch nicht ausreichend. die regionale durchblutung auf mikrozirkulationsebene wird durch lokale intrinsische (autoregulation) und durch extrinsische faktoren (autonomes nervensystem und humorale faktoren) geregelt (holtz 2000) . der blutfl uss zu den einzelnen organen wird durch den tonus der präkapillären arteriolen und prä-und postkapillären sphinktergefäße sowie durch lokale veränderungen der metabolischen aktivität gesteuert. mit der ops-methode (groner et al. 1999 ) lässt sich bei schwerer sepsis zeigen, dass die sublinguale mikrozirkulation in ihrer gefäßdichte reduziert -4,5(4,2-5,2)/mm bei sepsis vs. 5,4(5,4-6,3)/mm bei gesunden -und der anteil der perfundierten kleinen gefäße (<20 µm) vermindert ist -48(33-61)% vs. 90(89-92)%. diese veränderungen sind bei versterbenden ausgeprägter als bei überlebenden. die topische applikation von azetylcholin kann diese veränderungen wieder rückgängig machen (de backer et al. 2002; ince 2002) . die durchblutung der einzelnen organe kann über einen weiten blutdruckbereich organspezifi sch konstant gehalten werden (. tabelle 6.6), für den menschen wird diese autoregulation für einen bereich von 60-100 mmhg angegeben (bond 1993 bei hypovolämie und anderen hypodynamen schockformen wird der blutfl uss in gehirn und herz autoregulatorisch aufrechterhalten, in allen anderen organgefäßbetten ist er jedoch teilweise erheblich eingeschränkt, damit der systemische blutdruck möglichst stabil gehalten werden kann (. tabelle 6.6). für diese vasokonstriktion sind v. a. ein erhöhter sympathikotonus und die freisetzung von katecholaminen aus der nebenniere verantwortlich. diese adaptativen mechanismen reichen aus, um bei gering bis mäßig eingeschränktem herzzeitvolumen lebenswichtige organe adäquat zu perfundieren; bei ausgeprägter hypotonie kommt es jedoch zur organischämie und zum organversagen. selbst nach wiederherstellung stabiler herz-kreislauf-verhältnisse können die störungen auf mikrozirkulationsebene über tage persistieren, v. a. im gehirn, in den nieren, in der leber und in anderen splanchnikusorganen (wang et al. 1990 ). die irreversible phase eines schweren hämorrhagischen schocks ist aufgrund experimenteller befunde durch die vasodilatation präkapillärer sphinkter charakterisiert. die splanchnikusperfusion beim kritisch kranken variiert sehr stark in abhängigkeit von der grunderkrankung, kompensatorischen mechanismen und therapeutischen interventionen (jakob u. takala 2000) : im falle einer systemischen hypoperfusion oder hypoxämie wird die splanchnikusperfusion reduziert, wobei der leberblutfl uss auch unter diesen bedingungen aufgrund intrinsischer kontrollmechanismen relativ hoch gehalten wird (jakob et al. 2002) . bei sepsis und septischem schock fi nden sich bereits bei noch relativ adäquaten blutdruckwerten störungen der organdurchblutung, die eine primäre schädigung auf mikrozirkulationsebene nahe legen (thiemermann 2000a sie beruht auf einer sepsisinduzierten aktiven vasodilatation und dem verlust der extrinsischen vasomotorenkontrolle (bond 1993) , wodurch die durchblutung ausschließlich vom herzzeitvolumen abhängig wird. eine ausnahme von dieser regel stellt die hirndurchblutung dar, die in der sepsis weiterhin die fähigkeit zur autoregulation beibehält: bei patienten mit sepsis ist die hirndurchblutung bereits vor der ausbildung des schockzustandes um ein drittel reduziert, wobei diese durchblutungseinschränkung jedoch nicht als ursache der septischen enzephalopathie angesehen wird. im koronargefäßsystem fällt dagegen der widerstand noch stärker ab als in den anderen organen und demzufolge ist die koronarperfusion bei patienten mit septischem schock sogar häufi g erhöht (dhainaut et al. 1993 schock. die systemische und regionale minderperfusion spielt als verursacher der ischämisch bedingten zellschädigung bei den meisten schockformen eine prägende rolle. sind die neurohumoralen adaptationsmechanismen (. abb. 6.9) nicht mehr in der lage, die schockinduzierte organischämie und hypoxie zu kompensieren, so sind die hemmung des aeroben zellstoffwechsels, die abhängigkeit der energieproduktion von der allein nicht ausreichenden anaeroben glykolyse, die daraus resultierende zelluläre energieverarmung mit laktatanstieg und azidose die folge. diese schwerwiegende ischämisch bedingte zellschädigung ist jedoch keine simple hypoxiefolge auf die mitochon-driale atp-produktion allein: die mitochondrien zeigen auch noch bei sehr niedrigem o 2 -partialdruck (michaelis-konstante <1 µm) eine adäquate atp-produktion. demzufolge garantiert diese hohe o 2 -affi nität auch eine regelrechte funktion der atmungskette, selbst in der hypoxie, sieht man von extrembedingungen einmal ab. der so gestörte substrat-und energiestoffwechsel verursacht zahlreiche zellschäden, die sich als destruktion von mitochondrien, als beeinträchtigung der strukturellen und funktionellen zellmembranintegrität, in form zytotoxischer effekte und schließlich als zelltod manifestieren. leber und niere scheinen auf einen abfall des zell-atp besonders empfi ndlich zu reagieren. sepsis und septischer schock. die bedeutung eines energiemangels in der sepsis ist noch nicht eindeutig geklärt. zumindest lässt sich nachweisen, dass der atp-gehalt im skelettmuskel bei nichtüberlebenden stärker vermindert ist als bei überlebenden (brealey et al. 2002) . zytokine (zell et al. 1997) , exzessive produktion von stickoxid und reduzierte zelluläre glutathionreserven (brealey et al. 2002) scheinen ursächlich für die der verminderten energieproduktion in der sepsis zugrunde liegenden hemmung der atmungskette zu sein, wie für den skelettmuskel (brealey et al. 2002) und den herzmuskel (gellerich et al. 1999) gezeigt. bei der diskussion dieser ergebnisse darf allerdings nicht vergessen werden, dass nicht nur ischämie und hypoxie zur beeinträchtigung der mitochondrialen funktion und atp-produktion führen können, sondern auch mediatoren, insbesondere zytokine (. übersicht 6.3; . tabelle 6.7; zell et al. 1997 endotoxin ist zweifellos ganz entscheidend für die schlechte prognose bei gramnegativem schock mitverantwortlich. darüber sollte jedoch nicht vergessen werden, dass es in niedrigen konzentrationen über eine nur moderate mediatorfreisetzung bei der infektabwehr durchaus günstig wirken kann (mäßiges fieber, steigerung der immunabwehr, abtötung der keime). die durch endotoxin induzierte hypotension scheint mit der zeit eine abschwächung zu erfahren (mailman et al. 1999) . auch bei der entstehung und perpetuierung der herzinsuffi zienz scheint endotoxin eine rolle zu spielen (rauchhaus u. müller-werden 2001) . aus kardiologischer sicht bietet das endotoxin aber auch überraschungen: neben der deletären direkten und indirekten kardiodepressiven wirkung (7 abschn. 6.2.8) lassen sich auch protektive effekte auf das herz nachweisen (mcdonough et al. 1995; song et al. 1994; yao et al. 1993 ). grampositive erreger verursachen mindestens ebenso viele sepsiserkrankungen wie gramnegative keime. da grampositive bakterien keine lipopolysaccharide synthetisieren und in ihre zellwand integrieren, scheidet endotoxin als induktor einer grampositiven sepsis aus. hierfür kommen einerseits peptidoglykane (. abb. 6.6), die wie endotoxin an cd14 binden können, und die lipoteichonsäuren der zellmembran in frage und andererseits toxine, die entweder als superantigene, als porenbildner oder als adp-ribosylierende toxine ihre zytotoxische wirkung entfalten. peptidoglykane und lipoteichonsäuren wirken pyrogen, aktivieren komplement und b-lymphozyten. dabei werden die makrophagen aktiviert und sezernieren zytokine, teils in größeren mengen als nach endotoxinstimulation. toxine mit superantigeneigenschaften, wie z. b. das toxinschocksyndromtoxin 1 bestimmter staphylokokken, besitzen wie konventionelle antigene die fähigkeit, t-lymphozyten zu aktivieren. unter umgehung bestimmter kontrollmechanismen stimulieren sie jedoch nicht nur 0,01% der zellen, wie konventionelle antigene, sondern 2-10-25% aller t-lymphozyten gleichzeitig. die folge ist eine überaktivierung der t-zellen mit massiver zytokinfreisetzung, insbesondere tnf-α, und evtl. letalem schockverlauf. die bedeutung der superantigene bei der pathogenese der sepsis kann gegenwärtig noch nicht ausreichend abgeschätzt werden (visvanathan u. zabriskie 2000) . endotoxin und toxine grampositiver keime können sich in ihrer wirkung verstärken und über die toll-like-rezeptoren miteinander kommunizieren (li et al. 2003 das pseudomonas aeruginosa exotoxin a ist ein adp-ribosylierendes toxin, das hochselektiv den ribosomalen elongationsfaktor 2 der proteinsynthese adp-ribosyliert und damit inaktiviert. das toxin stellt einen wesentlichen virulenzfaktor der pseudomonassepsis dar. an kardiomyoyzten führt exotoxin a über eine partielle proteinsynthesehemmung zu einer störung der neusynthese von β-adrenozeptoren und damit zur kontraktilitätsabschwächung auf katecholamine (müllerwerdan et al. 1997) . das peptidoglykan ist hauptbestandteil der zellwand grampositiver bakterien. es wird ebenso wie endotoxin von cd14-und toll-like rezeptoren gebunden; die bindung an letztere bewirkt eine zellaktivierung. weitere bakerielle toxine wie die lipoteichonsäure, lipoarabinomannan von mykobakterien oder nannuronan führen ebenfalls über einen cd14-und toll-like rezeptor-abhängigen weg zur aktivierung von leukozyten. bestimmte bakterielle dns-sequenzen sind in der lage, antigenpräsentierende zellen (monozyten) zu aktivieren. diese dns-abschnitte sind reich an nichtmethylierten cpg-motiven. oligodeoxynukleotide mit cpg-motiven werden von toll-like-rezeptoren 9 erkannt; diese dns-abschnitte werden in die zellen aufgenommen und führen nach einer azidifi zierung in den endosomen zu einer aktivierung sowohl von mitogen aktivierten proteinkinasen als auch dem transkriptionsfaktor nf-κb. das verständnis um die bedeutung von toxinen und mediatoren im schockgeschehen fußte zunächst auf erkenntnissen, die bei der aufklärung der pathogenese des septischen schocks gewonnen wurden (. abb. 6.6). heutzutage kann jedoch davon ausgegangen werden, dass die mediatorkaskade die gemeinsame endstrecke nicht nur des septischen schocks darstellt (. abb. 6.6), sondern dass auch ein hypovolämischer, traumatischer, anaphylaktischer und sogar der kardiogene schock komponenten dieser kaskade als auslösende oder unterhaltende ursachen beinhalten. dieses konzept spiegelt sich auch in der aktuellen terminologie von sepsis und sirs (»systemic infl ammatory response syndrome«, . tabelle 6.3) wider: ausgehend von infektiösen oder nichtinfektiösen stimuli (. abb. 6.6) kommt es zur aktivierung von mediatorzellen, die primäre mediatoren wie den tumornekrosefaktor α freisetzen. diese primären mediatoren beeinfl ussen weitere zielzellen, die einerseits geschädigt werden können, andererseits (wie z. b. die neutrophilen granulozyten) fi nale mediatoren auf diesen stimulus (wie z. b. reaktive o 2 -verbindungen oder stickoxid) freisetzen. die mediatorfreisetzung soll zwar eigentlich zur bekämpfung der bakterien, zur neutralisierung von bakterientoxinen und zur schadensbegrenzung nichtinfektiöser insulte dienen, die aggressiven verbindungen schädigen dabei jedoch auch das herz-kreislauf-system und weitere vitale or zytokine, insbesondere tnf-α und il-1, stehen am anfang des mediatornetzwerks von schock und sepsis. bakterielle toxine, aber auch sirs-stimuli, können monozyten und makrophagen zur bildung und freisetzung insbesondere von tnf-α aktivieren (. abb. 6.6; loppnow 2001). als resultat der direkten und indirekten zytokinwirkungen stehen beim septischen schock die häufi g irreversible herz-kreislauf-schädigung und das mods. die bedeutung der zytokine beschränkt sich jedoch nicht nur auf den septischen schock: auch beim hämorrhagischen schock fi nden sich erhöhte tnf-serumspiegel; beim kardiogenen schock sind die interleukin-6-spiegel vergleichbar hoch wie beim septischen schock , und das akute lungenversagen bei hypovolämischem und traumatischem schock lässt sich durch den einsatz von anti-tnf-α-antikörpern bessern. erhöhte zytokinserum-und -plasmaspiegel fi nden sich darüber hinaus auch bei zahlreichen infektiösen und nichtinfektiösen herz-kreislauf-erkrankungen (. tabelle 6.7). derzeit muss allerdings noch häufi g die frage offenbleiben, ob zytokine dabei kausale bedeutung haben, ein sekundäres phänomen von krankheitswert oder nur ein unwichtiges epiphänomen darstellen (ausführliche diskussion in müllerwerdan et al. 1996; rauchhaus u. müller-werdan 2001) . das verstehen der zytokinwirkmechanismen (loppnow 2001; rauchhaus u. müller-werdan 2001 ) -gezeigt am beispiel des tnf-α -bietet die möglichkeit einer -bisher leider weder beim septischen schock noch bei herzinsuffi zienz erfolgreichen -kausalen therapie, z. b. mit anti-tnf-α-antikörpern. an der zielzelle bindet tnf-α (als trimer) an einen der beiden tnf-rezeptoren (tnfr1 mit einem mg von 55.000; tnfr2 mit einem mg von 75.000) und löst damit verschiedene zelluläre signale aus (7 abschn. 6.2.8). unterschiedliche stimuli (fieber, endotoxin, tnf-α selbst und andere zytokine) können zur proteolytischen spaltung der tnf-rezeptoren und ihrer freisetzung als lösliche tnf-rezeptoren ins plasma führen. deren bedeutung ist jedoch bisher noch unklar: in niedrigen konzentrationen scheinen sie ein reservoir für das gebundene tnf-α darzustellen, in höheren konzentrationen das tnf-α und damit seine zytotoxische wirkung zu neutralisieren. zytokine sind nicht nur schädlich; es ist vielmehr die massiv gesteigerte produktion und damit die dysbalance von proinfl ammatorischen mediatoren wie tnf, il-1, il-2, il-8, ifn und dem plättchenaktivierenden faktor (paf) mit den antiinfl ammatorischen substanzen und mechanismen (il-10, interleukin-1-rezeptorantagonist, lösliche tnf-rezeptoren, glukokortikoide und auch fieber), die letztlich bei schock und sepsis die ungünstige prognose determiniert. so zeigt eine hohe interleukin-6/interleukin-10-relation bei kritisch kranken eine ungünstige prognose an (taniguchi et al. 1999) . die tatsache, dass niedrige zytokinkonzentrationen (z. b. tnfα) bei sepsis durchaus zur abwehrreaktion in protektiver weise beitragen können, belastet das kausale therapiekonzept der unterbrechung dieser zytokin-/mediatorkaskade. > > zytokine und gerinnung. bei schwerer sepsis und bei septischem schock kommt es regelhaft auf endothelzellebene zu störungen der gerinnung und fibrinolyse im sinne einer disseminierten intravasalen gerinnung (dic, verbrauchskoagulopathie), allerdings individuell in jeweils sehr unterschiedlichem ausmaß: gerinnungsvorgänge sind gesteigert, fibrinolyseaktivitäten vermindert, und das verarmen an gerinnungsfaktoren wie des aktivierten protein c trägt zur proinfl ammation bei (dempfl e 2003). zytokin-genpolymorphismen. die prognose eines schockpatienten hängt wahrscheinlich nicht nur von seinem phänotyp, sondern auch von seinem genotyp ab, wie bisher v. a. für patienten mit sepsis und septischem schock gezeigt worden ist. männliche sepsispatienten, die homozygot sind für das allel tnfb2 des tnf-polymorphismus (ncoi-restriktionsfragmentlängenpolymorphismus), produzieren höhere tnf-plasmaspiegel und haben eine ungünstigere prognose als die sepsispatienten mit dem genotyp tnfb1/b1 bzw. tnfb1/b2. verknüpft mit dem tnf-polymorphismus und ohne eigenständige prognostische aussagekraft sind polymorphismen der hitzeschockproteine hsp70-hom c/t und hsp70-2 g/a bei patienten mit schwerer sepsis. während sich für die genotypverteilung des interleukin 1β keine unterschiede zwischen sepsispatienten und gesunden zeigen, fi ndet sich das allel a2 des interleukin-1-rezeptorantagonisten (il-1ra) bei patienten mit schwerer sepsis häufi ger als bei gesunden. dies könnte auf eine erhöhte sepsisempfi ndlichkeit bei dieser allelkonstellation hinweisen. nicht nur genpolymorphismen proinfl ammatorischer zytokine, sondern auch die antiinfl ammatorischer scheinen relevanz zu besitzen: sepsispatienten mit spezifi schen interleukin-10-genpolymorphismen sezernieren geringere mengen dieses antiinfl ammatorischen zytokins und haben eine erhöhte sterblichkeit (lowe et al. 2003 ). reaktive o 2 -verbindungen stellen im schockgeschehen eine wichtige gruppe terminaler mediatoren dar (de vega et al. 2002; motoyama et al. 2003) . dabei handelt es sich um moleküle mit einem ungepaarten elektron in der äußeren hülle. ursprungsort von freien radikalen sind zum einen aktivierten mediatorzellen, wie neutrophile granulozyten, makrophagen und endothelzellen, sowie andererseits (in der reperfusionsphase der schockbehandlung) das hypoxanthin als abbauprodukt des atp. das superoxidanion (o 2 -), das hydroxylradikal (oh · ), das wasserstoffperoxidmolekül (h 2 o 2 ), die hypochlorige säure (hocl) neutrophiler granulozyten und das peroxinitritradikal (onoo -) sind dabei die wichtigsten verbindungen (. abb. 6.7). peroxinitrit -gebildet aus dem stickoxid-und dem superoxidradikal -ist ein potentes und hochreaktives oxidans mit ausgeprägter zytotoxischer wirkung, das besonders zur endothelschädigung führen kann, die thrombozytenaggregation steigert und die ansprechbarkeit der koronargefäße auf vasodilatatoren vermindert (lamy et al. 2001) . treten die so entstandenen sauerstoffradikale bzw. reaktiven verbindungen in kontakt mit einer zielzelle, so können sie auf verschiedene zelluläre strukturen wirken: auf die zellmembran, das zytosol, den zellkern und die mitochondri-> 6.2 · pathophysiologie des schocks en. schließlich wird durch diesen oxidativen stress eine reihe von genen aktiviert (»stress-response genes«), die zur bildung von antioxidanzien, o 2 -radikalabbauenden enzymen, zytokinen und transkriptionsfaktoren führen. sie dienen einerseits der reparatur von zellschäden durch die sauerstoffradikale und führen, falls dies nicht möglich ist, zum gerichteten zelltod, der apoptose. ein beispiel des schädigungsmusters reaktiver sauerstoffverbindungen bei ischämie und reperfusion im herzen gibt . abb. 6.7. das zunächst als »endothelial-derived relaxing factor« klassifi zierte stickoxid (no) wird enzymatisch aus arginin gebildet. die cnos-isoformen sind ständig vorhanden. sie können sofort aktiviert werden und produzieren geringe mengen an no, das dann zahlreiche physiologische funktionen erfüllt. die cnos-isoenzyme können weiterhin in die neuronale form (nnos oder nos i, lokalisiert im zytosol von zentralen und peripheren neuronen) und in die endotheliale form (enos oder nos iii, überwiegend membrangebunden) eingeteilt werden. die enos-isoform besitzt sowohl parakrine als auch autokrine aktivität; sie wird überwiegend durch den scherstress der gefäße aktiviert, das gebildete no diffundiert in die glatten gefäßmuskelzellen und führt zur erschlaffung. die enos ist nötig, um die gefäße in einem partiell relaxierten zustand zu halten, der durch noradrenalin und endothelin antagonisiert wird. sie reguliert damit den gefäßtonus, die organperfusion und den blutdruck. weiterhin hemmt das von der enos produzierte no die adhäsion von thrombozyten und neutrophilen an endothelzellen und möglicherweise auch die proliferation von gefäßmuskelzellen. die induzierbare form der stickoxidsynthase (inos oder nos ii) wurde zuerst in makrophagen gefunden, sie kommt jedoch in vielen zelltypen vor. ihre induktion durch zahlreiche triggersubstanzen (. tabelle 6.8) dauert einige stunden, das enzymmolekül ist relativ langlebig (halbwertszeit mehrere stunden), und es kann durch die inos-aktivität in kurzer zeit lokal sehr viel stickoxid gebildet werden. demzufolge besteht die möglichkeit, dass das zur verfügung stehende enzymsubstrat arginin die produktion von stickoxid durch inos limitiert, was im falle der wesentlich geringeren no-produktionsraten durch die cnos-aktivität nicht der fall ist. reaktive stickstoff-sauerstoff-verbindungen (rnos) -insbesondere das aus dem stickoxid-und dem superoxidradikal entstehende peroxinitritradikal -wirken, überschießend produziert, im zellstoffwechsel hochtoxisch (7 abschn. 6.2.6; lamy et al. 2001) . ein dritter, für das schockgeschehen möglicherweise sehr bedeutungsvoller bildungsweg ist die nichtenzymatische reduktion von nitrit zu no in minderdurchblutetem gewebe. im ischämischen, azidotischen myokard fi ndet sich nitrit in mikromolaren konzentrationen. mittels elektronenspinresonanzspektren ließ sich nachweisen, dass in diesem hochreduktiven stoffwechselmilieu nitrit in großen mengen nichtenzymatisch zu stickoxid reduziert wird (zweier et al. 1995) . das entstandene stickoxid kann entweder am ort der bildung reagieren (autokrine wirkung) oder in benachbarte zellen diffundieren (parakrine wirkung). ein großteil der stickoxidwirkungen (vasodilatation, kardiodepression) beruht auf der aktivierung der löslichen guanylatzyklase durch stickoxid, mit bildung des zyklischen guanosinmonophosphats (cgmp), des gegenspielers des zyklischen adenosinmonophosphats (camp). bildung und wirkung des stickoxids können auf verschiedenen stufen blockiert werden: die induktion der inos kann durch glukokortikoide unterdrückt werden; die stickoxidsynthasen lassen sich durch argininanaloga wie l-nmma (ng-monomethyl-l-arginin; thiemermann 2000b) hemmen, und die aktivität der guanylatzyklase kann durch methylenblau blockiert werden. in der schockauslösung und der schocktherapie ist stickoxid ein entscheidender mediator (. tabelle 6.8; thiemermann 2000a). die hemmung der zellulären o 2 -verwertungs-> störung in den organen durch no-inhibition einer reihe mitochondrialer enzyme (aconitase, nadh-ubichinon-reduktase, succinat-ubichinon-oxidoreduktase; brealey et al. 2002; thiemermann 2000a ) könnte dabei eine ursache für die erhöht gefundenen o 2 -partialdrücke im skelettmuskel von sepsispatienten sein (. tabelle 6.5). natriuretische peptide wie das atriale natriuretische peptid (anp) und das b-typ-natriuretische peptid (»brain natriuretic peptide«, bnp) sind indikatoren einer mit einer ungünstigen prognose gekoppelten herzfunktionseinschränkung. darüber hinaus können zytokine der interleukin(il)-6-familie die sekretion von anp und bnp induzieren. im septischen schock steigen plasma-anp (82,7±9,9 vs. 14,9±1,2 pg/ml) und plasma-bnp (12,4±3,6 vs. 5,5±0,7 pg/ml) an. der anp-anstieg korreliert dabei mehr mit dem il-6-anstieg und weniger mit der kardiovaskuären dysfunktion, während der bnp-anstieg die pumpfunktionseinschränkung des herzens (abnahme des herzindex) widerspiegelt (witthaut et al. 2003) . in die schockpathogenese sind noch zahlreiche weitere hormone und mediatoren -endothelin, vasopressin (7 abschn. 6.4.7), adrenomedullin, adhäsionsmoleküle, hitzeschockproteine, arachidonsäurederivate u. a. -involviert (thiemermann 2000a; müller-werdan et al. 1996 burchardi et al. 2000) . schockart (dhainaut et al. 1993 ). apoptose -der gerichtete zelltod -fi ndet sich bei patienten mit sepsis v. a. in lymphozyten und intestinalen epithelzellen, nicht dagegen in nennenswertem maße im herzen und anderen parenchymatösen organen. bei nichtseptischen schockzuständen ist die apoptose ein noch selteneres phänomen wie bei der sepsis (hotchkiss et al. 1999; moldawer 1999 ). zahlreiche der im schock und in der sepsis gebildeten mediatoren (übersicht und literaturangaben in werdan 2000; krishnagopalan et al. 2002; müller-werdan et al. 1996; können eine myokarddepression hervorrufen und damit zum bild der akuten septischen kardiomyopathie beitragen. welche von den zahlreichen beschriebenen wirkungen auf die herzmuskelzelle jedoch tatsächlich klinische relevanz besitzen, ist bisher nur mit einschränkung zu beantworten. die z. z. am meisten favorisierte »negativ-inotrope kaskade« ist die endotoxin-tnf-α/il-1-no-cgmp-kaskade (. abb. 6.8). zirkulierendes endotoxin stimuliert mediatorzellen zur systemischen und myokardialen freisetzung von tumornekrosefaktor α (tnf-α) und interleukin 1 (il-1). tnfα und (wohl von geringerer bedeutung) il-1 induzieren dann die bildung der induzierbaren stickoxidsynthase (inos) im herzen. das von ihr gebildete stickoxid (no) stimuliert die lösliche guanylatzyklase des kardiomyozyten, und der daraus resultierende anstieg des cgmp führt schließlich über eine hemmung des ca 2+ -einstroms in die zelle oder über eine desensibilisierung der myofi lamente gegenüber ca 2+ zur kardiodepression (kumar u. parrillo 2001; . alle einzelschritte sind weitgehend belegt. die gabe von endotoxin führt bei probanden zur hyperdynamischen > herz-kreislauf-situation mit blutdrucksenkung, vasodilatation und steigerung des herzindex bei gleichzeitiger abnahme des schlagarbeitsindex als klinischem inotropieparameter (suffredini et al. 1989) . auch der anstieg des zirkulierenden tnf-α nach endotoxinapplikation ist bei probanden gezeigt worden (michie et al. 1988) , und erhöhte tnfα-plasmaspiegel fi nden sich auch im septischen schock. bei therapeutisch mit tnf behandelten tumorpatienten kann es zur ausgeprägten myokarddepression kommen. im tierexperiment führt die gabe von tnf-α zu einer bis mehrere tage anhaltenden myokarddepression (müllerwerdan et al. 1996) . der nachweis der direkten depressorischen wirkung des tnf-α auf die funktion des kardiomyozyten lässt sich im zellkulturexperiment führen: tnf-α hemmt in klinisch relevanten konzentrationen nach mehrstündiger einwirkungsdauer die »positiv-inotrope« wirkung des β-adrenozeptoragonisten isoproterenol und auch die hoher ca 2+ -konzentrationen. die depressorische tnf-α-wirkung kann durch anti-tnf-α-antikörper neutralisiert werden. auch die induktion einer induzierbaren stickoxidsynthase in kardiomyozyten durch tnf-α und il-1 ist experimentell gut belegt, ebenso wie bei patienten mit septischem schock eine erhöhte aktivität der inos und ein cgmp-anstieg im herzen. es spricht also vieles dafür, dass die diskutierte »negativinotrope« kaskade für die myokarddepression in der sepsis und im septischen schock wesentlich mitverantwortlich ist. dennoch scheint dies nicht der einzige kardiodepressive mechanismus zu sein: einerseits kann tnf-α im experiment auch stickoxidunabhängig negativ-inotrop wirken, andererseits wird durch tnf-α nicht nur die positiv-inotrope wirkung von β-adrenozeptoragonisten, sondern auch die von α-adrenozeptoragonisten gehemmt. letzteres ist über eine alleinige induktion der stickoxidsynthase nicht zu erklären; so hemmt endotoxin, das in kardiomyozyten die stickoxidsynthase stimuliert, zwar auch die positiv-inotrope wirkung von β-adrenozeptoragonisten, nicht aber die von α-adrenozeptoragonisten (müllerwerdan et al. 1996) . aufbauend auf experimentellen befunden dieser art kann davon ausgegangen werden, dass neben der stickoxid-cgmp-kaskade auch noch andere mechanismen für die negativ-inotrope wirkung von tnf-α verantwortlich sein müssen (. abb. 6.8). die belegte hemmung des phosphoinositolstoffwechsels durch dieses zytokin erklärt die hemmung der positiv-inotropen wirkung von α-adrenozeptoragonisten, und die ebenfalls dokumentierte abschwächung des ca 2+ -transients durch tnf schwächt alle inotropen effekte ab. ebenfalls belegt ist die hemmung des ca 2+ -einwärtsstroms durch einen kardiodepressiven peptidfaktor, der sich in hämofi ltraten von patienten mit kardiogenem und septischem schock in konzentrationen nachweisen lässt, die diese hemmwirkung in humanen herzmuskelzellen hervorrufen ). die gewebeschäden sind nicht allein durch den mangel an sauerstoff während der ischämiephase, sondern auch durch die reperfusion und reoxygenation der ischämischen organe bedingt (abella u. becker 2002) . bei der enzymatischen reaktion der in der ischämiephase akkumulierten atp-abbauprodukte hypoxanthin und xanthin mit sauerstoff kommt es zur bildung freier sauerstoffradikale (wasserstoffperoxid, superoxidradikal, hydroxylradikal), die durch lipidperoxidation die aktivierung des prostaglandinsystems und eine denaturierung von zellmembranproteinen bewirken. katalysiert wird diese radikalproduktion durch das enzym xanthinoxidase, das in der ischämie durch umwandlung aus der xanthindehydrogenase (nicht zur radikalbildung befähigt) entsteht. dieser umwandlungsprozess läuft in den geweben bei ischämie unterschiedlich schnell ab (darm: 10 s; herzmuskel: 8 min; leber, milz, niere, lunge: 40 min), was die unterschiedliche empfi ndlichkeit der einzelnen gewebe auf reperfusionsschäden mit erklären soll freie sauerstoffradikale rufen darüber hinaus eine verstärkte akkumulation polymorphkerniger leukozyten hervor, begünstigen das leukozytensticking sowie über den »respiratory burst« die bildung weiterer sauerstoffradikale aus polymorphkernigen leukozyten (. abb. 6.7). demzufolge kommt der leukozyten-endothel-interaktion -leukozyteneinwanderung, -rollen, feste adhäsion, gefäßtransmigration -via adhäsionsmoleküle -selektine, integrine, immunglobulinsuperfamilie, cadherine -bei dem ischämiereperfusionsschaden eine große bedeutung zu (finney et al. 2002) . exzessive stickoxid-und damit peroxinitritbildung, komplementaktivierung (ciurana u. hack 2002) und apoptoseinduktion (abella u. becker 2002) tragen wahrscheinlich ebenfalls entscheidend zum komplexen ischämiereperfusionsschädigungsmuster bei. ein weiterer kandidat ist das kernenzym poly(adp-ribose)-polymerase (parp; synonym: par-synthetase, pars; liaudet et al. 2001; szabo u. liaudet 2002; . abb. 6.7): proinfl ammatorische zytokine im kreislaufschock produzieren reaktive sauerstoffverbindungen via stimulation der xanthinoxidase, de-novo-expression der induzierbaren stickoxidsynthase, induktion einer mitochondrialen dysfunktion und rekrutierung von neutrophilen granuloyzten mit expression der nadph-oxidase. als folge davon werden die sauerstoffradikale peroxinitrit, wasserstoffperoxid und hydroxylradi-kale gebildet, durch interaktion von superoxidanionen, stickoxid und der eisenkatalysierten oxidation von superoxidanionen. der so entstehende oxidative zellstress induziert die bildung des transkriptionsfaktors ap-1 und löst dns-einzelstrangbrüche aus. die dns-schäden aktivieren die parp. die parp-aktivierung verbraucht atp sowie redoxäquivalente (nadph), induziert damit eine endothelzellschädigung sowie den zelltod und potenziert die aktivierung der transkriptionsfaktoren ap-1 und »nuclear factor kappa b (nfκb)«. letzteres hat die verstärkte transkription und translation der ap-1-und nfκb-abhängigen gene für inos, icam, mip-1α, tnf-α und komplement-c3 zur folge. das aus c3 gebildete c5 und die gesteigerte endotheliale expression des adhäsionsmoleküls icam-1 führt zu einer verstärkten einwanderung von aktivierten leukoyzten in den infl ammatorischen fokus und bewirkt damit eine intensivierung der bildung reaktiver sauerstoffverbindungen. der teufelskreis wird mit der triggerung weiterer dns-strangbrüche durch den oxidativen stress geschlossen. da natürliche schutzstoffe gegen die effekte der sauerstoffradikale (scavenger-substanzen, superoxiddismutase (elsakka et al. 2001) , glutathion, glutathionperoxidase, katalase) im intrazellulärraum nur in geringer konzentration vorliegen, können sie den als folge der reperfusion und reoxygenation eintretenden reperfusionsschaden, der durch endothelläsion, intrazelluläres ödem und letztlich deletären einstrom von kalziumionen in die zellen charakterisiert ist, nicht verhindern. der reperfusionsschaden wird als wichtiger pathogenetischer faktor bei der entwicklung des mods im schock gesehen. noch ausstehend ist der nachweis der klinischen wirksamkeit einer antioxidanzientherapie (mullan u. mccloskey 2002) , der in zahlreichen studien für folgende substanzen nicht erbracht werden konnte: superoxiddismutase, katalase, glutathionperoxidase, ascorbinsäure, glutathion, harnsäure, α-tocopherol, karotinoide, der xanthinoxidasehemmer allopurinol, lazaroide, pyruvat (fink 2003) , der hemmung der komplementaktivierung (ciurana u. hack 2002) , von parp-inhibitoren (szabó u. liaudet 2002; liaudet et al. 2001 ) oder der inhibition der map-kinasen (tulleken et al. 2001 ). hrv-td zeitdomäne der herzfrequenzvariabilität; hrv-fd frequenzdomäne der herzfrequenzvariabilität zusätzlich zu den intrinischen anpassungsmechanismen der neurohumoralen und auto-/parakrinen regelkreise im schock können bakterientoxine (endotoxin) und möglicherweise auch mediatoren eine fehlstellung der stellglieder induzieren (schmidt et al. 2001; schmidt u. müller-werdan 2003) : sowohl die baro-als auch die chemorefl exsensitiät und ebenso die herzfrequenzvariabilität zeigen bei kritisch kranken im schockbedingten mods ausgeprägte, prognosebestimmende einschränkungen (. tabelle 6.9; 7 abschn. 6.5.1). auf welcher ebene toxine und mediatoren diese autonome dysfunktion bewirken -zentralnervensystem, autonomes nervensystem, zielzelle (. abb. 6.10) -bleibt noch zu klären; experimentell ist bei spontan schlagenden neonatalen kardiomyozyten (»schrittmacherzellen«) durch züchtung der zellen in endotoxin eine einschränkung der schlagfrequenzvariabilität zu erzielen, ohne dass dabei das autonome nervensystem involviert ist (schmidt et al. 2001 ). ) schock bedeutet für den patienten immer eine lebensbedrohliche notfallsituation. er erfordert ein sofortiges handeln, beginnend in der prähospitalphase (christ u. lackner 2004) , fortgesetzt in der notaufnahme (laggner 2004 ) und schließlich auf der intensivstation. bei der erstversorgung müssen basisdiagnostik und basistherapie zum frühestmöglichen zeitpunkt begonnen werden. diese basisversorgung dient primär der kreislaufstabilisierung und gilt für alle schockformen. sie mündet nach feststellung der schockform in eine schockformspezifi sche behandlung. je kürzer die schockdauer ist, je schneller die stabilisierung des kreislaufs erreicht wird und je früher die schockformspezifi sche behandlung begonnen werden kann, um so günstiger die prognose. patienten, bei denen ein kreislaufschock vermutet wird, sollten auf der intensivstation weiter betreut und behandelt werden. die in der . übersicht 6.4 aufgeführten technischen untersuchungen schließen sich an. patienten mit unklarer schockätiologie oder mit einer sich nach volumensubstitution nicht rasch stabilisierenden kreislaufl abilität sollten baldmöglichst einem invasiven hämodynamischen monitoring unterzogen werden (7 abschn. 6.3.4). extremitäten. blutungen und thromboembolische komplikationen können bei diesen patienten auf eine verbrauchskoagulopathie hinweisen. kranke mit einem septischen fieber und neutropenie weisen häufi g keinen fokus auf; der septische schock kann sowohl mit hohem fieber als auch mit hypothermie einhergehen. das klinische bild des schockpatienten ändert sich mit der behandlung (laggner 2004) : der unbehandelte patient mit septischem schock in der notaufnahme ist ein patient mit zentralisiertem, »kühlem« schock, der behandelte patient mit septischem schock auf der intensivstation zeigt das »klassische« bild des »warmen«, hyperzirkulatorischen schocks (. tabelle 6.10). das vorgehen bei klinischem schockverdacht im rahmen der erstversorgung zeigt die . übersicht 6.4. empfehlungen zur klinisch-hämodynamischen basisdiagnostik bei kardiogenem schock sind der . übersicht 6.5 zu entnehmen. die höhe der troponin-und ck-mb-anstiege und das flächenintegral korrelieren zwar mit der infarktausdehnung und der ungünstigen prognose, ein cut-off-wert für den infarktbedingten kardiogenen schock kann allerdings nicht angegeben werden. ein nicht unerheblicher anteil (>50%) der patienten mit schwerer sepsis und septischem schock zeigt erhöhte troponinserumspiegel und weist damit auf eine myokardschädigung in der sepsis hin. patienten mit erhöhten troponinwerten haben eine ungünstigere prognose (spies et al. 1998 bei schwerer sepsis und septischem schock scheint das serielle pro-nt-bnp-monitorig innerhalb der ersten 7 tage nach diagnosestellung prognostische bedeutung zu haben: während bei den später versterbenden die pro-nt-bnp-spiegel stark erhöht bleiben oder sogar weiter ansteigen, sinken sie bei den überlebenden signifi kant ab. gerinnungsparameter. da patienten mit akuten kardialen erkrankungen in der regel antithrombotische, antithrombozytäre und fi brinolytische substanzen singulär oder in kombination erhalten, ist die standardmäßige bestimmung von parametern der blutgerinnung obligat: thrombozyten, aptt, prothrombinzeit, fibrinogenkonzentration, d-dimere (bei verdacht auf beinvenenthrombose, lungenembolie). sobald die umstände dies gestatten, sollten vorrangig ein ekg und eine thoraxröntgenaufnahme angefertigt werden. bildgebende (janssens 2001; und invasive verfahren (s. unten), evtl. kombiniert mit interventionellen maßnahmen, ergänzen die diagnostik und bieten erste therapeutische ansätze. das monitoring und entsprechende therapeutische konsequenzen sind in den . übersichten 6.4 und 6.6 wiedergegeben (müller arterielle druckmessung. bei allen patienten mit kreislaufschock sollte eine kontinuierliche, blutige arterielle druckmessung erfolgen, da die manuelle messung mit der manschette oder mit nichtinvasiven oszillometrischen techniken aufgrund der zentralisierung und vasokonstriktion unzuverlässig. ist. in der regel wird dafür die a. radialis kanüliert; bei schwerster zentralisation sollte allerdings die a. femoralis bevorzugt werden. zentraler venendruck. die messung des zentralen venendrucks ist bei kritisch kranken, insbesondere schockpatienten, für das hämodynamische monitoring normalerweise nicht genügend, eine abschätzung der linksventrikulären vorlast kann damit nicht ausreichend sicher durchgeführt werden, ebenso wenig wie mit der klinischen einschätzung allein. für die therapieoptimierung des patienten im kardiogenen shock ist die kenntnis des herzzeitvolumen (hzv) zwingend erforderlich. das hzv ist die regelgröße des herz-kreislauf-systems und wird im wesentlichen durch vorlast, nachlast und kontraktilität sowie die herzfrequenz bestimmt. klassische klinische zeichen wie blutdruck, urinausscheidung, halsvenenfüllung, hautperfusion und hautturgor erlauben keine zuverlässige einschätzung der hämodynamik beim schwerkranken intensivpatienten (eisenberg et al. 1984) . pulmonalarterienkatheter. der pulmonalarterienkatheter (pak) ist weiterhin ein grundpfeiler der erweiterten hämodynamischen überwachung bei patienten mit kardiogenem schock (flieger et al. 2004; janssens 2001 bruch et al. (2003) f für das prognoserelevante monitoring weniger geeignet (müller-werdan u. werdan 2005) als anfangs gedacht. der grad der regionalen gewebeoxygenierung im schock ist spektroskopisch durch messung der hämoglobin-o 2 -sättigung und des cytochrom-aa 3 -redoxstatus möglich (beilman et al. 1999) . während die kontinuierliche messung der co 2 -atemgaskonzentration (kapnographie) v. a. in der notfallmedizin und in der kardiopulmonalen reanimation zur beurteilung von kreislauf-und respiratorischer funktion zum einsatz kommt, ist die sublinguale kapnometrie (messung des sublingualen co 2 -partialdrucks) ein neues, nichtinvasives verfahren zur beurteilung des schweregrades und der prognose von schockzuständen (weil et al. 1999 ). bei patienten mit akutem koronarsyndrom wurden klinische modelle entwickelt, die zuverlässig das auftreten eines kardiogenen schocks im weiteren verlauf vorhersagen können. für den patienten mit st-strecken-elevations-myokardinfarkt (stemi) erfolgte dies auf der grundlage der gusto-i-und -iii-studiendaten (. tabelle 6.11). dieser risiko-score, der innerhalb weniger minuten in der notaufnahme erhoben werden kann, erlaubt auf schnelle und unkomplizierte weise, eine frühzeitige abschätzung des risikos einer schockentstehung im weiteren klinischen verlauf (hasdai et al. 2000b das grundsätzliche ziel der symptomatischen schocktherapie ist die wiederherstellung einer suffi zienten durchblutung der vitalorgane und gewebe, ehe sich ein zellschaden ausbilden kann. dies erfordert einen ausreichenden herzindex und blutdruck. kurze perioden einer ausgeprägten minderperfusion werden besser toleriert als gravierende blutdruckabfälle. die aufrechterhaltung eines mittleren blutdrucks von mehr als 60-65 mmhg hat initial priorität vor dem anheben des herzindex auf werte von >2,1 l/min/m 2 im falle des kardiogenen und obstruktiven bzw. >4,0-4,5 l/min/m 2 beim septischen und beim volumensubstituierten hämorrhagischen schock. die durchblutung sollte zumindest so gesteigert werden, dass der arterielle laktatspiegel unter 2,2 meq/l verbleibt, eine weitgehende garantie dafür, dass der stoffwechsel weiter ae nicht länger als 30 s sollte die intubation als sicherste beatmungsmethode in anspruch nehmen. die intubation (klasse-i-empfehlung) sollte nur durch sehr gut trainierte personen praktiziert werden. im zweifelsfall ist (auch aus juristischen gründen) die leichter zu beherrschende maskenatmung (beatmung immer mit 100% sauerstoff) anzuwenden. larynxmaske oder kombi-tubus gelten als klasse-ii-option. der (peripher-)venöse zugang ist ebenfalls eine klasse-i-intervention. die anfl utzeit der medikamente beträgt 1-2 min. aufgrund vielfältiger komplikationsmöglichkeiten wird die anlage eines zentralvenösen zugangs nur in ausnahmefällen als indiziert angesehen. alternativ besteht insbesondere bei kindern, aber auch bei erwachsenen die möglichkeit, medikamente über eine intraossäre kanüle zu applizieren; dies wird unterhalb der tuberositas tibialis in der markhöhle platziert und ermöglicht neben der gabe von medikamenten auch die infusion von flüssigkeit. f f f f > adrenalin, atropin und lidocain können in notfällen auch endobronchial über den tubus gegeben werden (7 abschn. 6.4.8). hierbei muss dann die dosis mindestens verdoppelt (2-bis 2,5fach) in 10 ml 0,9%igem nacl appliziert werden. adrenalin ist das einzige routinemäßig empfohlene medikament, alle anderen, z. b. antiarrhythmika, natriumbikarbonat, kalzium-und magnesiumzubereitungen, sind speziellen reanimationssituationen vorbehalten. adrenalin. die standarddosis ist 1 mg i.v. alle 3-5 min bis zur kreislaufstabilisierung, gefolgt von jeweils 20 ml i.v.-spülvolumen. dosen über 1 mg haben bestenfalls den primären, nicht jedoch den langfristigen reanimationserfolg verbessern können (gueugniaud et al. 1998 ). höhere adrenalindosen korrelieren sogar mit einem ungünstigen neurologischen defi zit (behringer et al. 1998) . dennoch sehen die empfehlungen im verlauf der reanimation eine eskalation der adrenalindosis auf bis zu 0,2 mg/kgkg alle 3-5 min im sinne einer klasse-ii-empfehlung vor. vasopressin. ermutigend, aber noch nicht standardempfehlung, ist die wirkung von vasopressin (40 ie i.v.), das in vergleichsstudien mit adrenalin zumindest gleichwertig, bei nachweis einer asystolie sogar vorteilhaft war (wenzel et al. 2004) . im falle eines adrenalinrefraktären schocks infolge kammerfl immerns bei erwachsenen gilt vasopressin als klasse-iia-empfehlung. atropin. in einer dosierung von 1 mg als bolus i.v. alle 3-5 min bis zu einer gesamtdosis von 0,04 mg/kgkg ist atropin indiziert bei asystolie und pulsloser elektrischer aktivität. die dosis von 1 mg atropin sollte nicht unterschritten werden, da es bei niedrigeren dosen zu paradoxen effekten (bradykardien) kommen kann. bei symptomatischen bradykardien tritt die atropingabe hinter die transvenöse oder behelfsweise transkutane stimulation zurück. amiodaron. amiodaron wird empfohlen bei persistenz von kammertachykardie oder kammerfl immern nach defi brillation und adrenalingabe (dorian et al. 2002) . initial wird eine rasche 300-mg-infusion in 20-30 ml kochsalz-oder glukoselösung gegeben, ggf. gefolgt von einer infusion mit 1 mg/ min für 6 h und anschließend 0,5 mg/min bis zu einem täglichen maximum von 2 g. die gabe von lidocain ist generell nicht als routinemaßnahme zu sehen, auch nicht bei patienten mit akutem myokardinfarkt! in einer dosis von 1,0-1,5 mg/kgkg i.v. kann lidocain bei kammerfl immern/pulsloser kammertachykardie nach erfolgloser defi brillation und adrenalingabe verabreicht werden (wirksamkeit als unsicher eingestuft), ggf. gefolgt von weiteren 0,5-0,75 mg/kgkg innerhalb von 3-5 min (maximaldosis 3 mg/kgkg oder 200-300 mg/h). lidocain stellt z. z. allerdings bei dieser indikation im vergleich zu amiodaron nur ein antiarrhythmikum der zweiten wahl dar. > ! > 6.4 · therapieprinzipien bei schock zur rezidivprophylaxe nach erfolgreicher defi brillation oder kardioversion kann lidocain in einer dosierung von 1,0-1,5 mg/kgkg gegeben werden, gefolgt von 20 ml nacl 0,9%, mit einer anschließenden dauerinfusion von 1-2 mg/ kgkg/h. vor einer defi brillation angewandt, erhöht lidocain eher die defi brillationsschwelle. monomorphe kammertachykardien nicht primär ischämischer genese können mit lidocain nur zu etwa 10% unterbrochen werden. dagegen ist ajmalin (1,0 mg/kgkg i.v.) mit einer 60%igen erfolgsrate wesentlich effektiver. natriumbikarbonat. da eine bedeutsame arterielle azidose während der kardiopulmonalen reanimationssituation in der regel durch eine unzureichende ventilation begründet ist und nahco 3 selbst zu einem paradoxen intrazellulären co 2 -anstieg führen kann, ist die gabe von natriumbikarbonat üblicherweise nicht indiziert. in speziellen situationen kann die gabe von bikarbonat allerdings hilfreich sein: bei vorbestehender metabolischer azidose, bei hyperkaliämie und bei intoxikationen mit trizyklischen antidepressiva oder phenobarbital. nach protrahiertem herz-kreislauf-stillstand oder langdauernden wiederbelebungsbemühungen -nach ineffektiver defi brillation, herzmassage, intubation, beatmung und vasopressorentherapiekann bikarbonat (1 meq/kgkg als initiale dosis) von nutzen sein. andere puffersubstanzen haben bisher keinen eingang in die offi zielle empfehlung gefunden. elektrolyte: kalzium, magnesium. eine kalziumgabe als routinemaßnahme kann nicht empfohlen werden. nur im falle einer hypokalzämie, einer hyperkaliämie oder einer intoxikation mit kalziumantagonisten ist die gabe von kalzium bedingt indiziert (klasse-iib-empfehlung): 10%ige kalziumchloridlösung in einer dosis von 2-4 mg/kgkg, wiederholung in 10-minütigen intervallen möglich. die applikation von magnesium (intravenöse gabe von 1-2 g magnesiumsulfat in 1-2 min) ist indiziert bei bestätigter hypomagnesiämie und refraktärem oder rezidivierendem kammerfl immern. ebenfalls indiziert ist die intravenöse gabe von 1-2 g magnesiumsulfat bei torsade-de-pointes-tachykardien. enttäuschend war bisher der einsatz der transkutanen antibradykarden stimulation in einer studie mit 1056 patienten mit herz-kreislauf-stillstand. weder bei asystolie noch überbrückend nach defi brillation mit nachfolgender asystolie konnte dadurch der ausgang der prähospitalen kardiopulmonalen reanimation entscheidend effi zienter gestaltet werden (literatur in . in mehreren studien konnte gezeigt werden, dass nach kardiopulmonaler reanimation durch die induktion einer milden therapeutischen hypothermie mit temperaturen zwischen 32 und 34°c eine verbesserung des überlebens und der neurologischen funktion erreicht werden kann (bernard et al. 2002; group thacas 2003) ; aufgrund der guten ergebnisse ist das > »cooling« nach cpr bei herzstillstand infolge kammerfl immerns als klasse-i-maßnahme in die ilcor-empfehlungen eingegangen (nolan et al. 2003) . nach eventueller durchführung von reanimationsmaßnahmen erfolgt die stabilisierung des herz-kreislauf-systems und der lungenfunktion. zur herz-kreislauf-stabilisierung dienen kristalloide und kolloide plasmaersatzlösungen sowie der einsatz von katecholaminen (7 abschn. 6.4.6 und 6.4.7). dazu dienen auch anxiolyse und analgesie, relaxierung zur einsparung von sedativa und die beseitigung von fieber (hyperthermie steigert den o 2 -verbrauch um 7%/°c). ebenfalls beachtet werden müssen die negativen auswirkungen der peep-beatmung auf die herzfunktion (zunahme der rechtsventrikulären nachlast, zunahme des rechtsventrikulären durchmessers und abnahme der linksventrikulären diastolischen dehnbarkeit, direkte myokarddepressive wirkung), ebenso bei kontrollierter mechanischer (»controlled mechanical ventilation«, cmv) oder intermittierender ventilation (»intermittent mandatory ventilation«, imv). die langwierige und schwierige weaning-off-phase gerade bei herzkranken und schockpatienten kann durch einhalten standardisierter protokolle erleichtert werden (lehmann et al. 2003; seige et al. 2001) . die myokarddepression kann weiterhin verstärkt werden durch anästhetika und barbiturate. während benzodiazepine und opioide für sich keine relevante myokarddepression hervorrufen (mit ausnahme von meperiden), können sie in kombination einen additiven negativ-inotropen effekt induzieren. > > evidenzbasierte empfehlungen zur beatmung des schockpatienten, speziell des patienten mit kardiogenem schock, gibt es z. z. noch nicht (kontoyannis et al. 1999; lesage et al. 2004 ). bei überwiegen der vorteile der maschinellen beatmung bei patienten mit kardiogenem schock (reduktion der kardialen vor-und nachlast, abnahme der pulmonalen stauung, reduktion der atemarbeit) erscheint eine eher großzügige indikationsstellung bei diesen patienten gerechtfertigt. ein engmaschiges kardiopulmonales monitoring und ein rasches anpassen der beatmung an änderungen der herzfunktion ist angebracht (lehmann et al. 2001; seige et al. 2001 ). antiarrhythmische therapie. ein spezifi sches muster an rhythmusstörungen ist für den schockpatienten nicht dokumentiert. dennoch muss häufi g mit potenziell malignen rhythmusstörungen gerechnet und entsprechend behandelt werden. die im kardiogenen schock meist vorhandene hochgradige pumpfunktionseinschränkung reduziert die frequenztoleranz des patienten beträchtlich und birgt so die gefahr einer weiteren verschlechterung der schockzustandes, falls die hämodynamisch relevante rhythmusstörung nicht rasch beseitigt wird. abhängig von der klinischen situation (z. b. kardiogener schock bei herzinfarkt) kann eine schmerzbehandlung und eine analgosedierung erforderlich werden (ruß et al. 2005) . intravenöse morphinbolusgaben (2-5 mg i.v. alle 5-30 min bis zu einer gesamtdosis von 2-3 mg/kgkg) sind, z. b. in den ersten stunden eines herzinfarktes, das mittel der wahl. auf eine mögliche blutdrucksenkung infolge einer direkten und indirekten hemmung des sympathikus muss geachtet werden, v. a. bei relativer hypovolämie. im kreislaufschock kann die morphinclearance durch die minderperfusion der leber eingeschränkt sein. morphin hemmt nicht nur den schmerz, sondern trägt auch zur senkung des o 2 -verbrauchs bei. der wichtigste parameter der metabolischen azidose ist der laktatanstieg. die anhebung des extrazellulären ph-werts mit bikarbonat führt nicht immer zum gewünschten erfolg des intrazellulären ph-anstiegs, der zudem eine linksverschiebung der hämoglobindissoziationskurve und damit eine erschwerte o 2 -abgabe an das gewebe bewirkt. die therapie der laktatazidose kann aber zu einer besserung der systemischen und hepatischen zirkulation führen, die ihrerseits zum abbau verbleibenden laktats beiträgt. der therapeutische einsatz von bikarbonat sollte auf ph-werte von 7,10-7,15 beschränkt bleiben. klinisch wurde die hämodynamik von schockpatienten durch bikarbonat auch bei ausgeprägter azidose nicht verbessert (cooper et al. 1990 ); eine isolierte stimulation der pyruvatdehydrogenase mit dichloracetat senkte bei laktatazidose die hohe letalität von 70% nicht (stacpoole et al. 1992 ). ob kollodiale oder kristalloide lösungen zur volumensubstitution besser geeignet sind, wird seit langem kontrovers diskutiert; die art der lösung scheint allerdings für den therapieerfolg nur eine untergeordnete rolle zu spielen. unterschiede bezüglich morbidität und letalität konnten für verschiedene flüssigkeitsregimes bisher nicht eindeutig gezeigt werden (kreimeier u. prückner 1998; ; task force of the american college of critical care medicine, society of critical care medicine 1999). kristalloide lösungen sind kostengünstig, leicht zu lagern, steigern ausreichend die diurese und können zusätzlich extravasale flüssigkeitsverluste bei dehydratationszuständen ersetzen. nachteilig sind das auftreten ausgeprägter peripherer ödeme und die relativ kurze hämodynamische wirksamkeit (. tabelle 6.13). am häufi gsten kommen physiologische (0,9%ige) kochsalzlösung und vollelektrolyte (z. b. ringer-laktatlösung) zum einsatz, die sich beide gleichermaßen im intravasalraum und im interstitium verteilen; nach 1 h fi nden sich aber nur noch weniger als 25% des infundierten volumens in der zirkulation. beide lösungen senken den kolloidosmotischen druck. im vergleich zu kolloidalen lösungen muss etwa das 2-bis 4fache des intravasalen flüssigkeitsdefi zits an kristalloider flüssigkeit zur erzielung einer vorübergehenden normovolämie infundiert werden. 5%ige glukoselösung fi ndet sich 1 h nach infusion nur noch zu 8% im intravasalraum; sie erhöht neben dem volumen des extrazellulärraums unerwünschterweise auch das des intrazellulärraums (infolge des wassereinstroms in die zellen zum ausgleich des infusionsbedingten osmotischen gradienten). sie sollte deshalb zur volumentherapie nicht verwendet werden. zu den kollodialen lösungen zählen albumin, hydroxyäthylstärke, dextran und gelatine (. tabelle 6.13). diese verbleiben zunächst vorwiegend im intravasalraum und stellen dort den plasmaonkotischen druck wieder her. sie führen demzufolge in geringerem ausmaß zu peripheren ödemen, und es genügen kleinere volumina zur substitution als beim einsatz von kristalloiden lösungen. nachteile sind neben den hohen kosten die bekannten, substanzeigenen nebenwirkungen (s. unten). dass kolloidale lösungen v. a. bei der sepsis mit ihrer erhöhten kapillarpermeabilität das auftreten eines lungenödems fördern, ist viel diskutiert, aber bisher nicht gesichert worden. von den kolloidalen substanzen wird in deutschland am häufi gsten hydroxyäthylstärke eingesetzt. für die initiale volumentherapie mit kolloidalen lösungen erscheinen 6%ige lösungen von mittelmolekularer hydroxyäthylstärke (hes 200.000/0,5 oder 200.000/0,62) sowie die 6%ige dextran-60-lösung (macrodex) am besten geeignet. nicht nur wegen der limitierten tagesdosen können die kolloidalen lösungen mit kristalloiden (z. b. im verhältnis 1:1) kombiniert werden. die derzeitige datenlage spricht dafür, dass das kolloid der zukunft eine stärkepräparation der dritten genera-> 6.4 · therapieprinzipien bei schock tion (hes 130/04; . tabelle 6.13) sein wird (boldt 2003; dieterich 2001) . albumin. albumin fi ndet als 5%ige (kolloidosmotischer druck ca. 20 mmhg) und als 20-bis 25%ige lösung (kolloidosmotischer druck ca. 80-100 mmhg) verwendung und verbleibt relativ lange im intravasalraum (>90% nach 2 h). bei hypovolämie ist initial eher die 5%ige lösung angebracht; bei ödematösen patienten kann die verwendung der hyperonkotischen albuminlösung eine erwünschte flüssigkeitsverschiebung aus dem interstitium in den intravasalraum bewirken. beim direkten studienvergleich der volumensubstitution kritisch kranker erbrachten die gabe von albumin im vergleich zu kochsalzlösung keine unterschiede hinsichtlich letalität und morbidität (safe study investigators 2004). kristalloide und synthetische kolloidale lösungen haben das humanalbumin bei der volumentherapie weitgehend ersetzt; geblieben ist beim erwachsenen als indikation eine hypalbuminämie <2,5 g/dl bzw. ein gesamteiweiß <4,0-4,5 g/dl. es ist jedoch nicht sinnvoll, eine hypalbuminämie als folge eines kapillären lecks (wie bei der sepsis) mit albumin vollständig auszugleichen, da dieses mit einer halbwertszeit von 1-6 h aus dem intravasal-in den extravasalraum abwandert. der absolutwert des kolloidosmotischen drucks hat sich als parameter zur albuminsubstitution nicht durchgesetzt. unerwünschte, in der regel milde nebenwirkungen (fieber, schüttelfrost, urtikaria) treten mit einer häufi gkeit von ca. 0,5% auf; die blutgerinnung wird nicht beeinträchtigt. bei der infusion großer albuminmengen kann es zu einer senkung des ionisierten plasmakalziums kommen. > dextrane. dextrane sind hochmolekulare lineare polysaccharide mit vereinzelten seitenketten, gelöst in physiologischer kochsalzlösung. sie werden entsprechend ihrer molekularmasse entweder direkt (mg <50.000) oder nach enzymdegradation, bevorzugt renal, eliminiert. durch den hohen kolloidosmotischen druck füllt die 10%ige dextran-40-lösung den intravasalraum durch einen ausgeprägten einstrom aus dem interstitium auf, was im schock mit gestörter mikrozirkulation erwünscht, bei einem depletierten extrazellulärraum jedoch eher unerwünscht ist. dextran 40 reduziert die geldrollenbildung der erythrozyten, und es soll auch die gewebeoxygenierung verbessern. hydroxyäthylstärkepräparationen. im deutschsprachigen raum ist hydroxyäthylstärke (hes) sicherlich die am häufi gsten eingesetzte substanz zur therapie der hypovolämie. hes-lösungen werden aus kartoffel-oder maisstärke produziert innerhalb der letzten jahre ist es zur entwicklung deutlich verbesserter hes-präparationen (1. generation: 450/07; 2. generation: 70/05, 200/05, 200/0,62; 3. generation: 130/04; . tabelle 6.13) hinsichtlich intravasalem hes-abbau und nebenwirkungsprofi l -res-speicherung oder nephrotoxität (s. unten) -gekommen (boldt 2003 in einem cross-over-vergleich bei kritisch kranken erzielte die gabe von 100 ml albumin 25% eine zunahme des plasmavolumens 45 min nach infusionsende um 465 ml, nach 1 l ringer-laktatlösung waren es dagegen kurzfristig maximal 194 ml. wesentliche verbesserungen der hämodynamik und des o 2 -transports korrelieren eindeutig mit der plasmaexpansion. sie sind nach gabe kristalloider lösungen entweder gar nicht oder wesentlich schwächer als nach infusion kolloidaler lösungen nachweisbar. die wirksamkeit der einzelnen kolloidalen lösungen untereinander scheint vergleichbar (. tabelle 6.13), die wirkdauer der hes-lösungen dagegen länger als die der 5%igen albuminlösung. lungenfunktion. bei der diskussion um die ideale volumenersatzlösung spielt die potenzielle gefahr der auslösung eines lungenödems eine entscheidende rolle. verfechter des einsatzes kolloidaler lösungen führen an, dass kristalloide flüssigkeiten den kolloidosmotischen druck (kod) nachhaltig erniedrigen und damit diese gefahr hervorrufen. befürworter des einsatzes kristalloider lösungen fürchten dagegen bei der anwendung kolloidaler lösungen einen verstärkten abstrom kolloidosmotisch wirksamer moleküle durch die geschädigte alveolokapilläre membran ins interstitium, verbunden mit einem anstieg des extravaskulären kod und damit der gefahr der ausbildung oder verstärkung eines lungenödems. eindeutige vorteile hinsichtlich der organperfusion sind weder für kristalloide noch für kolloidale lösungen bei der volumensubstitution kritisch kranker überzeugend belegt (müller-werdan u. . als zielkriterien einer adäquaten flüssigkeitssubstitution des schocks dienen zunächst klinische parameter wie herzfrequenz, diurese und blutdruck, die sich in den physiologischen bereichen bewegen sollten. bei ausbleiben einer raschen hämodynamischen stabilisierung und bei patienten mit bereits vorbestehender eingeschränkter herzfunktion empfi ehlt sich das invasive monitoring mittels pulmonalarterienkatheter (7 abschn. 6.3.4). während der volumensubstitution muss mit dilutionsbedingtem abfallen von hämatokrit und hb (1-3 g/dl) gerechnet werden, die in der regel auch akzeptiert werden können: bei einem hämatokritwert von 30% liegt eine maximale o 2 -transportkapazität ohne gefahr einer gewebehypoxie vor. während der primären volumentherapie -in der initialen phase der hypovolämie -ist aus diesem grund ein hb-wert von 10-11,5 g/dl bzw. ein hämatokrit von 30-35% als ausreichend anzusehen. nachfolgend -bei normovolämie, erzielt durch volumengabe -ist eine substitution von erythrozytenkonzentraten erst ab hb-werten <8-10,0 g/dl indiziert > kritisch kranke. bei kritisch kranken ist eine mäßige anämie infolge eines okkulten blutverlustes und einer supprimierten erythropoese nicht selten. übereinstimmung besteht darüber, dass patienten mit akuter anämie und einem hämoglobinwert von ≤7 g/dl und darunter entsprechend den allgemeinen empfehlungen (simon et al. 1998 ) mit erythrozytentransfusionen substituiert werden sollten. dagegen konnte bei kritisch kranken mit hb-werten von 7-10 g/dl durch erythrozytentransfusionen die prognose nicht verbessert werden (hebert et al. 1999) . mögliche günstige effekte der transfusion könnten durch ungünstige reaktionen -wie rheologische störungen gealterter transfundierter erythrozyten, störungen der immunfunktion infolge der transfusion nicht leukozytengefi lterter erythrozyten (blumberg u. heal 1998 ) -wieder zunichte gemacht werden. sepsispatienten. die gabe eines erythrozytenkonzentrates erbrachte bei anämischen (hb <10 g/dl) sepsispatienten keine verbesserung der o 2 -utilisation, weder global (vo 2 ) noch regional (magentonometrie). beobachtet wurde eine 10%ige zunahme des linskventrikulären schlagarbeitsindex, aber auch eine ungünstige, 15%ige zunahme des pumonalvaskulären widerstands (fernandes et al. 2001) . bei patienten mit schwerer sepsis und septischem schock mit einem hämatokrit <30% und gleichzeitig einer auf <70% erniedrigten zentralvenösen sauerstoffsättigung (s cv o 2 ) wird die gabe von erythrozytenkonzentraten zur anhebung des hämatokrits auf ≥30% empfohlen, zumindest in der frühphase der sepsis innerhalb der ersten 6 h auf der notaufnahme (rivers et al. 2001) . ansonsten gilt die empfehlung, erythrozytenkonzentrate bei einem hämoglobinwert <7,0 g/dl (<70 g/l) zu geben und das hb auf einen wert von 7,0-9,0 g/dl anzuheben (dellinger et al. 2004 ). kritisch kranke. die verminderte erythropoetinbildung beim anämischen kritisch kranken und der hohe anteil von knapp 40% transfundierten intensivpatienten bilden die rationale für die erprobung einer erythropetingabe (epo) bei intensivpatienten: im rahmen einer studie mit 1300 intensivpatienten wurden wöchentlich -bis zu 4-mal -40.000 einheiten rhuepo bzw. placebo gegeben. die epo-gabe reduzierte den prozentsatz transfusionspfl ichtiger patienten von 60,4 auf 50,5% und die zahl der transfusionen um 19%, wobei der hb-anstieg mit 1,32 g/dl stärker ausfi el als in der placebogruppe (0,94 g/dl). die letalität der epo-gruppe war mit 14% nicht unterschiedlich im vergleich zu der der placebogruppe (corwin et al. 2002) . der nutzen dieser epo-gabe bei intensivpatienten wird z. z. kritisch gesehen (eckardt 2003) . sepsispatienten. bei patienten mit schwerer sepsis wird erythropoetin zur behandlung einer anämie nicht empfohlen (dellinger et al. 2004 ). mit ausnahme des hypovolämisch-traumatischen schocks mit versorgungsengpässen gibt es bei allen anderen schock-formen z. z. für den einsatz hyperton-onkotischer lösungen keine indikation (meier-hellman u. burgard 2004). im wesentlichen sind es die katecholamine dobutamin, dopamin, noradrenalin und adrenalin (. tabelle 6.14) und die phosphodiesterasehemmstoffe amrinon, milrinon und enoximon, (. tabelle 6.15), deren positiv-inotrope wirkung ausgenutzt werden kann . sie werden eingesetzt, um die herz-kreislauf-schädigung zu kompensieren und damit die durchblutung und die o 2 -versorgung der vitalorgane sicherzustellen. bei den klinischen symptomen eines kardiogenen schocks ist dobutamin (positiv-inotrope wirkung, mittel der 1. wahl bei > akutem myokardinfarkt; 2-20 µg/kgkg/min) und ggf. zusätzlich noradrenalin (positiv-inotrope und vasokonstriktorische wirkung; 0,05-0,3 µg/kgkg/min) indiziert; burchardi et al. 2000; . es handelt sich dabei um eine symptomatische, nicht um eine kausale therapie. bei vergleichbarer positiv-inotroper wirkung beeinfl ussen katecholamine herzfrequenz, blutdruck und gefäßwiderstand sowie den linksventrikulär-enddiastolischen füllungsdruck am gesunden herzen in unterschiedlicher weise (. tabelle 6.14; burchardi et al. 2000; : adrenalin wirkt am stärksten, noradrenalin am wenigsten positiv-chronotrop. vor allem noradrenalin erhöht den gefäßwiderstand und damit den blutdruck; in höheren konzentrationen tun dies allerdings auch dopamin und adrenalin. bei sepsis und schock können desensibilisierungsprozesse und toxin-und mediatorschädigungen das ansprechen auf katecholamine ganz erheblich beeinträchtigen (7 abschn. 6.4.8). . dosisbereich ( (hochman et al. 2000) . fragt man sich dann aber, wie denn die wirksamkeit der katecholamintherapie gesichert sei, so wird man enttäuscht: weder für den kardiogenen noch für den septischen schock gibt es hinsichtlich einer möglichen letalitätssenkung evidenzbasierte daten! lediglich für die behandlung der akuten herzinsuffi zienz fi ndet sich in der literatur eine metaanalyse, die sich mit der wirksamkeit von intravenös applizierbaren, über den adrenergen signalweg wirksamen inotropen substanzen -vorwiegend dobutamin, dopamin, dopexamin und phosphodiesterasehemmer -beschäftigt (thackray et al. 2002) . das ergebnis der kleinen studien mit geringer statistischer aussagekraft (21 studien mit insgesamt 632 patienten) ist enttäuschend: nicht eine letalitätssenkung, sondern eine im trend höhere sterblichkeit wurde bei dem einsatz der inotropen substanzen gefunden (odds-ratio 1,50; 95%iges konfidenzintervall 0,51-3,92). wegen der potenziell unerwünschten und gefährlichen nebenwirkungen der katecholamintherapie -wie die infl ammationsinduktion sowohl systemisch als auch im herzen selbst schwertz et al. 2004 ) -sollte man beim einsatz von katecholaminen im schock eher zurückhaltend sein. während dobutamin den linksventrikulär-enddiastolischen druck entweder unbeeinfl usst lässt oder ihn sogar geringfügig senkt, wird er durch dopamin meistens etwas gesteigert. die ursache dafür dürfte ein erhöhter venöser rückstrom durch eine α-adrenozeptorvermittelte venokonstriktion sein. bei der volumensubstitution des septischen schocks mit dobutamin können deshalb größere flüssigkeitsmengen erforderlich werden als im falle des dopamins müller-werdan u. werdan 2004 ; task force of the american college of critical care medicine, society of critical care medicine 1999). dobutamin ist das katecholamin der wahl zur therapie der eingeschränkten pumpfunktion der septischen kardiomyopathie! dagegen ist nach der empfehlung einer deutschen expertenkommission dopamin kein katecholamin der ersten wahl bei der therapie des septischen schocks in kontrollierten studien konnte durch dopamin in »nierendosis« (0,5-2-3 µg/kgkg/min) keinerlei nephroprotektiver effekt und auch keine prognoseverbesserung dokumentiert werden. der routinemäßige einsatz von niedrigdosiertem dopamin zur nephroprotektion bei schock-, mods-und sepsispatienten kann demzufolge nicht empfohlen werden dellinger et al. 2004; . bei patienten mit septischem schock zeigen katecholamine (dobutamin) eine geringere positiv-inotrope wirkung als bei patienten mit sepsis ohne schock (silverman et al. 1993) . diese katecholamintoleranz ist zumindest partiell auf eine dysregulation des β-adrenozeptor-adenylatzyklase-systems zurückzuführen (down-regulation der β 1 -adrenozeptoren; hochregulation der inhibitorischen g-proteine), hervorgerufen durch endogene und pharmakologisch applizierte katecholamine, sowie durch zytokine u. a. sepsismediatoren (. abb. 6.13a,b; müllerwerdan et al. 1996; reithmann et al. 1993; silverman et al. 1993) . sie lässt sich zumindest partiell durch eine steigerung der katecholamindosis ausgleichen. da diese β 1 -adrenozeptor-adenylatzyklase-desensibilisierung jedoch alle am myokardialen β 1 -adrenozeptor angreifenden katecholamine in gleichem maße betrifft, resultieren daraus keine differenzialtherapeutischen konsequenzen. über den erfolgreichen einsatz von β-rezeptorenblockern zur verbesserung der katecholaminansprechbarkeit wurde bisher beim patienten mit schock noch nicht berichtet. auch die gefäße zeigen im septischen schock eine katecholamintoleranz mit einer abgeschwächten bis fehlenden vasokonstriktion auf α-adrenozeptoragonisten. in diesem falle scheint jedoch nicht die im tierexperiment bei sepsis und endotoxinämie gefundene abnahme der zahl der gefäß-α-adrenozeptoren die entscheidende rolle zu spielen, sondern vielmehr das vermehrt gebildete stickoxid. durch hemmstoffe der stickoxidsynthase lässt sich im sepsisund endotoxin-tiermodell die stark abgeschwächte vasokonstriktorische katecholaminwirkung wieder restaurieren (. abb. 6.13a,b). die zahl der gefäß-β 2 -adrenozeptoren ist im tierexperiment bei sepsis und endotoxinämie als nicht verändert beschrieben. insbesondere bei kardiogenem schock nach herzoperationen scheinen sich phosphodiesterasehemmstoffe (pde-hemmstoffe; . tabelle 6.15) als katecholaminbegleittherapie zu bewähren . bei akuter herzinsuffi zienz konnte der pde-hemmer milrinon in einer randomisierten studie lediglich bei patienten mit nichtischämischer herzinsuffi zienz, nicht jedoch im gesamtkollektiv eine letalitätssenkung erzielen, wohingegen bei patienten mit akuter ischämischer herzinsuffi zienz sogar eine letalitätserhöhung erwartet werden muss (cuffe et al. 2002) . digitalis spielt bei sinusrhythmus in der therapie des kardiogenen schocks als inotropikum keine wesentliche rolle; da-> gegen ist es bei tachykardem vorhoffl immern und vorhoffl attern zur frequenznormalisierung bei schockpatienten das antiarrhythmikum der wahl. als neues therapieprinzip bei akuter herzinsuffi zienz scheint der kalziumsensitzer levosimendan erfolgreich (follath et al. 2002; kivikko et al. 2003; moiseyev et al. 2002) . levosimendan (in deutschland nicht zugelassen) sensibilisiert kalziumabhängig troponin c für kalzium und verbessert damit die kontraktilität des herzmuskels, wobei die diastolische relaxation entweder unbeeinfl usst bleibt oder sogar verbessert wird. durch öffnung atp-sensitiver kaliumkanäle wirkt levosimendan darüber hinaus vasodilatierend. bei akuter herzinsuffi zienz (herzindex 1,9 l/min/m 2 ) war levosimendan (bolus von 24 µg/kgkg über 10 min, danach 0,1 µg/ kgkg/min für 24 h) hämodynamisch effektiver als dobutamin (180-tage-letalität in der levosimendangruppe: 26%, in der dobutamingruppe 38%; follath et al. 2002) . ähnlich günstige effekte von levosimendan wurden für patienten mit akutem herzinfarkt und linksherzinsuffi zienz beschrieben (moiseyev et al. 2002) . zur senkung der vor-und nachlast dient bei akutem herzinfarkt in 1. linie nitroglyzerin und in 2. linie nitroprussidnatrium. nitroglyzerin wird in einer infusionsdosierung von 0,3-0,5-4 µg/kgkg/min gegeben; die nitroprussidnatriuminfusi-on wird mit 0,3 µg/kgkg/min begonnen und bis zum maximal erwünschten effekt alle 2 min bis zu einer dosis von 1-6 µg/ kgkg/min gesteigert; sie kann mit positiv-inotropen pharmaka kombiniert werden (alpert u. becker 1993) . nesiritide, das endogene b-typ-natriuretische peptid (in deutschland nicht zugelassen), senkt bei dekompensierter herzinsuffi zienz den pulmonarkapillardruck rascher und effektiver als nitroglyzerin. im vergleich zur behandlung mit dobutamin scheint nesiritide (i.v.-bolus 0,3 µg/kgkg, anschließend infusion mit 0,015 µg/kgkg/min bzw. i.v.-bolus 0,6 µg/kgkg, anschließend infusion mit 0,030 µg/kgkg/min) rascher eine rekompensation zu bewirken und möglicherweise sogar die prognose zu verbessern (silver et al. 2002 klinzing et al. (2003) infundierten vasopressin in einer dosis von 0,06-1,8 u/min, um die gabe von noradrenalin vollständig beenden zu können. ihre ergebnisse zeigen eine deutliche umverteilung des intestinalen blutfl usses zu lasten der magenschleimhaut. im vergleich zu anderen arbeiten wurde allerdings in dieser studie vasopressin nicht im sinne einer substitution bei vasopressinmangel, sondern in einer sehr hohen dosierung eingesetzt. dieselbe arbeitsgruppe hat auch die effekte einer niedrig dosierten (0,04 u/kgkg/h) vasopressininfusion auf die globale und intestinale hämodynamik untersucht. in dieser untersuchung wurden 12 patienten im septischen schock (noradrenalinbedarf: 0,13-1,4 µg/kgkg/min) mit arginin-vasopressin behandelt. ein adäquate splanchnikusperfusion blieb trotz eines signifi kantem abfalles des herzindexes erhalten. allerdings kam es auch bei dieser niedrigen dosierung zu einer redistribution des blutfl usses zu lasten der gastralen mukosa. auch eine niedrigdosierte vasopressintherapie kann mit einer bedeutsamen gastrointestinalen minderperfusion einhergehen. es gibt hinweise darauf, dass eine restriktive flüssigkeitstherapie kombiniert mit der infusion von vasopressin bei der behandlung einer unkontrollierbaren intraabdominellen blutung zu einer verbesserung der überlebensrate führt. wer dieses -im tierexperiment überaus erfolgreiche -konzept in die praxis einführen will, muss jedoch wissen, dass es zur zeit noch keine klinischen beweise seiner wirksamkeit gibt. beim septischen schock besteht ein vasopressinmangel. in mehreren untersuchungen konnte gezeigt werden, dass vasopressin sowohl in niedriger als auch in hoher dosierung bei septischen patienten zu einer deutlichen stabilisierung der hämodynamik führt. signifi kante nebenwirkungen traten dabei nicht auf. allerdings gibt es andere untersuchungen, in denen störungen der intestinalen und der hepatischen perfusion und generell der mikrozirkulation nach vaso-> pressininfusion nachgewiesen wurden. solange es so deutlich widersprüchliche erkenntnisse gibt, kann unseres erachtens die gabe von vasopressin im septischen schock nicht generell empfohlen werden, auch wenn die surviving sepsis campaign (. tabelle 6.20; dellinger et al. 2004 ) bei therapierefraktärem septischem schock die zusätzliche gabe von vasopressin (0,01-0,04 u/ml) für vertretbar ansieht. der kardiogene schock führt häufi g zu lungenstauung/lungenödem und zum prärenalen nierenversagen und damit zur oligurie. neben der kreislaufstabilisierung können diuretika und hämofi ltration erforderlich werden (7 abschn. 6.5.4). das im schock via induktion der inos überschießend produzierte stickoxid (no) zeigt neben seinen erwünschten wirkungen im rahmen der infektabwehr unerwünschte deletäre effekte (7 abschn. 6.2.6). insofern erscheint die blockade der überschießenden no-produktion mittels inos-hemmer (argininanaloga wie l-name) sinnvoll. klinische ergebnisse zu diesem therapieprinzip liegen für den septischen und den kardiogenen schock vor. gesicherte relevanz besitzen diese substanzen bei kardiogenem und bei septischem schock. kardiogener schock. der einsatz von thrombolytika und weiteren gerinnungsaktiven substanzen im kardiogenen schock wird in 7 abschn. 6.6.1 besprochen. gp iib/iiia-rezeptor-antagonisten scheinen neben ihrer thrombozytenaggregationshemmenden auch antiinfl ammatorische eigenschaften zu besitzen (bonz et al. 2002; köster et al. 2003; lincoff et al. 2001) , deren therapeutischer stellenwert aber noch nicht ausreichend abgeschätzt werden kann. nach größeren notfalloperationen muss in ca. 8% der fälle mit dem auftreten eines einorganversagens, in 4% mit einem zweiorganversagen, in 2% mit einem dreiorganversagen und in 1% mit einem vierorganversagen gerechnet werden; lungen-, nieren-und leberversagen sind dabei mit 7-9% etwa gleich häufi g. die prognose der patienten mit mods ist um so ungünstiger, je mehr organe geschädigt sind und je länger das organversagen anhält. die sterblichkeit nach 1-, 3-und 7-tägiger dauer eines einorganversagens liegt bei 20, 30 bzw. 40%, im falle eines zweiorganversagens bei 50, 60 bzw. 70%, und bei einem dreiorganversagen bei 80, 90 bzw. fast 100%. die aufgeführten zahlen gehen auf untersuchungen zurück, die vor 25 jahren durchgeführt worden sind. bei der ungünstigen prognose des mods hat sich seit 1973 mit einer berichteten letalität von 94% bis 1994 mit 60% zwar eine verbesserung gezeigt, die sterblichkeit ist jedoch weiterhin sehr hoch und weitgehend unabhängig von der art des geschädigten vitalorgans. ein lebensalter >65 jahre erhöht die letalität auf das doppelte. die mediatoren-/zytokin-hypothese (. abb. 6.2, 6.3 und 6.6) postuliert eine exzessive oder prolongierte produktion von zytokinen durch entzündungszellen, z. b. granulozyten und makrophagen, sowie durch dazu fähige körperzellen, z. b. kardiomyozyten. zu diesen zytokinen gehören interleukin 1, tumornekrosefaktor α, interleukin-6, das auch antiinfl ammatorische teilwirkungen hat, interleukin-8 u. a. diese zytokine induzieren die produktion fi naler mediatoren wie stickoxid, arachidonsäuremetaboliten, bradykinin und histamin, die neutrophile granulozyten und endothelzellen aktivieren und damit die gewebeschädigung induzieren. die induktion dieser überschießenden infl ammation kann sowohl durch infektiöse stimuli (infektion, sepsis) als auch durch nichtinfektiöse stimuli (sirs; 7 abschn. 6.1) induziert werden. die mikrozirkulations-und ischämie-/reperfusionshypothese propagiert die ischämie und/oder die gefäßendothelschädigung als ursache des mods, mit inadäquater o 2 -versorgung von geweben und zellen (alleinige ischämie), mit einer ischämie-/reperfusionsschädigung und der generie-! rung von toxischen o 2 -radikalen und/oder gewebeschädigung infolge der endothel-leukozyten-interaktion. diese hypothese favorisiert auch die annahme mehrerer toxischer stimuli in sequenz als auslöser des mods (»two-hit model of mods«). diese hypothese geht von der darmwandtranslokation von bakterien und endotoxinen als mods-ursache aus. bei patienten mit septischem und nichtseptischem mods fi ndet sich eine erhebliche autonome dysfunktion (»uncoupling of biological oscillators«; godin u. buchman 1996), die sich z. b. als einschränkung der herzfrequenzvariabilität messen lässt (. abb. 6.14a-d). im gegensatz zur autonomen dysfunktion herzkranker mit gesteigerter sympathikusaktivierung scheint diejenige des mods-patienten von einer abschwächung sowohl der sympathikus-als auch der parasympathikusaktivität geprägt zu sein (heinroth et al. 1999 ob es allerdings tatsächlich nur eine einheitliche ursache eines einheitlichen mods gibt, muss z. z. noch offen bleiben! viel wahrscheinlicher ist eine multifaktorielle mods-genese unter beteiligung der aufgeführten mechanismen. die komplexität des mods macht es verständlicherweise schwierig, den prognosebestimmenden schweregrad dieses krankheitssyndroms zu beschreiben. score-systeme stel-> len eine möglichkeit dazu dar (graf et al. 2003) . zwei versuche dieser art sollen hier vorgestellt werden: der schweregrad-der-erkrankung-score apache ii (»acute physiology and chronic health evaluation«) und der sepsisbezogene organversagen-score (sofa-score). in verschiedenen bereichen der medizin werden score-systeme seit längerem zur quantitativen erfassung von befunden eingesetzt. so dient z. b. die glasgow-coma-scale (. tabelle 6.17) zur abschätzung des schweregrades einer bewusstseinstrübung. bei patienten einer intensivstation können score-systeme eingesetzt werden, um objektive, quantifi zierbare parameter zu gewinnen zur (graf et al. 2003; scores (wie der schweregrad-der-erkrankung-score apa-che ii oder der sepsis-score nach elebute u. stoner) kön-f f f f nen am patientenbett innerhalb von 5-10 min mit einem auf einem mikrocomputer installierten programm einfach bestimmt und dokumentiert werden (müller. fortentwicklungen mit noch besserer prognostischer aussagekraft wie der mpm-score, der apache-iii-score und das supportsystem bedürfen noch der praktikabilitätsumsetzung und haben noch keine sehr weite verbreitung (graf et al. 2003; janssens et al. 2002) . bei einem schweregrad-der-erkrankung-score wie dem apa-che-ii-score (knaus et al. 1985; . abb. 6.15) korreliert die score-höhe mit der letalität, und zwar sowohl des gesamtkollektivs kritisch kranker als auch bestimmter subkollektive (z. b. patienten mit sepsis). ein mögliches problem bei der anwendung von score-systemen stellt jedoch die abhängigkeit der score-werte von der grundkrankheit dar. zusätzlich zu den bereits bestehenden organversagen-scores wurde von der europäischen intensivmedizinischen gesellschaft der sofa-score (»sepsis-related organ failure assessment score«) entwickelt (. tabelle 6.17). er erfasst die wichtigsten organdysfunktionen mit jeweils einem einzelnen pa-tabelle 6.17. schweregradklassifi zierung des mods-sofa-score. (nach vincent et al. 1996) . sepsisbezogener organversagen-score (sofa-score) punktzahl augen öffnen: 4 (spontan), 3 (bei aufforderung), 2 (bei schmerz), 1 (nicht); beste motorische antwort: 6 (gezielt nach aufforderung), 5 (gezielt nach schmerz), 4 (ungezielt nach schmerz), 3 (beugemechanismen), 2 (streckmechanismen), 1 (keine); verbale antwort: 5 (orientiert), 4 (verwirrt), 3 (inadäquat), 2 (unverständlich), 1 (keine) rameter und teilt den schweregrad der organdysfunktion entsprechend der abweichung dieses parameters von der norm ein. der sofa-score wird zunehmend auf bei nichtsepsispatienten zur abschätzung des schweregrades des mutiorganversagens eingesetzt, auch bei kardiologischen patienten. auch die prognose von patienten mit herzerkrankungen auf einer coronary care unit (ccu) kann mit score-systemen prognostiziert werden (7 abschn. 6.5.2): die aussagekraft des initial erhobenen saps-ii-score (»simplifi ed acute and physiology score«) ist bei ccu-patienten mindestens so prägnant wie bei patienten auf einer intensivstation (. abb. 6.16a,b; schuster et al. 1997) . dies unterstreicht eindrücklich, dass nicht nur die schwere der herzerkrankung, sondern v. a. das daraus resultierende multiorgandysfunktionssyndrom über das überleben des herzpatienten entscheidet. bei patienten nach herzoperationen erlaubt der apache-ii-score die identifi zierung der patienten mit eskalierendem systemischen infl ammationsreaktionssyndrom nach kardiopulmonalem bypass (eskalierendes cpb-sirs; . tabelle 6.4). es ist charakterisiert durch eine -im vergleich zum unkomplizierten postoperativen verlauf -überschießende systemische entzündungsreaktion infolge des operationstraumas und des einsatzes des kardiopulmonalen bypass, mit hohen tumornekrosefaktor-α(tnf-α)-und tnf-rezeptor-, leukozytenelastase-und neopterinplasmaspiegeln als ausdruck einer leukozyten/makrophagen-aktivierung, einem sepsisähnlichen multiorgandysfunktionssyndrom mit myokarddepression und hoher letalität (müller. dieses nur bei wenigen patienten (<10%) auftretende eskalierende cpb-sirs kann bereits am ersten postoperativen tag anhand eines apache-ii-score≥24 identifi ziert werden (. abb. 6.17). aufgrund der ergebnisse von beobachtungsstudien in den zeiträumen 1988 zeiträumen -1990 zeiträumen und 1996 pathophysiologie nach kreislaufschock kann es in abhängigkeit von der schockform und der zugrunde liegenden erkrankung zu 2 formen der lungenschädigung mit daraus resultierender insuffi zienz kommen: ein kardiogen bedingter kreislaufschock induziert primär eine erhöhung des lungenkapillardrucks, während die übrigen schockformen eine gesteigerte permeabilität der lungenkapillaren bedingen (engelmann 2000a ). das lungenödem bei erhöhtem kapillardruck hat als häufi gste ursache einen gesteigerten hydrostatischen druck in den lungenkapillaren, der wiederum in erster linie folge eines linksherzversagens ist. die filtration von flüssigkeit und protein wird als folge der erhöhten kapillarpermeabilität so gesteigert, dass der abtransport über die lymphwege bei weitem nicht ausreicht, um dies zu kompensieren. aufgrund der gestörten barrierefunktion kann auch kein nennenswerter osmotischer druckgradient aufgebaut werden, so dass sich diese ödemform mit rascher progredienz entwickelt. die störung der barrierefunktion des endothels wird nach gegenwärtiger auffassung durch mediatoren ausgelöst. die uniformität der pulmonalen reaktion bei sehr unterschiedlichen klinischen ausgangskonstellationen wird dabei > auf das limitierte repertoire des organismus an effektorsystemen zurückgeführt. es wird postuliert, dass alle formen des kreislaufschocks, die zu einem ards führen, letztlich ein ischämie/reperfusionssyndrom darstellen, wobei endotoxineinschwemmung, gewebehypoxie, makrophagen-und leukozyteneinwanderung, -adhäsion (mittels adhäsionsmolekülen) und -aktivierung mit freisetzung von zahlreichen entzündungsmediatoren als dominierende faktoren angesehen werden. bei 50% aller schockpatienten fi ndet sich eine akute respiratorische insuffi zienz. die frühesten veränderungen der lungenfunktion im rahmen eines kreislaufschocks basieren auf reaktionen des zentralen atemantriebs oder der atemmuskulatur. sowohl der gesteigerte atemantrieb infolge der stimulation der pulmonalen j-rezeptoren und der chemorezeptoren des karotissinus als auch die minderperfusion des medullären atemzentrums führen zur steigerung des atemminutenvolumens (tachypnoe, hyperpnoe), zur hypokapnie und zur initialen respiratorischen alkalose. ventilations-perfusions-störungen infolge des gesteigerten atemminutenvolumens bei gleichzeitiger reduktion des herzzeitvolumens können die folge sein. der lungenwiderstand ist initial unverändert oder nur minimal erhöht, falls nicht bereits schockbedingt eine arterielle hypoxämie besteht. die erhöhte atemarbeit bei gleichzeitiger minderperfusion der atemmuskulatur und des zwerchfells kann zur frühzeitigen respiratorischen insuffi zienz führen. falls der schockzustand nicht rasch behoben werden kann, sind kardiales lungenödem bzw. ein ards zu befürchten. zur klinik des lungenödems mit erhöhtem kapillardruck (kardiales lungenödem) 7 kap. 4.2. das lungenödem bei erhöhter kapillarpermeabilität bedingt ein klinisches syndrom, für das der begriff des »adult/acute respiratory distress syndrome« geprägt worden ist. die letalität des ards beträgt nach wie vor ca. 50-60%, wobei fortschritte auf dem gebiet der beatmungstechniken (engelmann 2000a, b) ten patienten mit ards nach unterschiedlichen auslösern zu einer vermehrten mikrobiellen belastung der lunge, für die im wesentlichen eine retrograde keimaszension aus dem gesamtintestinaltrakt mit mikroaspirationen in verbindung mit gestörten host-defense-mechanismen der lunge verantwortlich gemacht wird. in abhängigkeit von der dauer des ards kommt es zunehmend zu nosokomialen pneumonien, die bei über 10 tägiger beatmung mehr als 70% der ards-patienten betreffen können. die übergänge zwischen pneumonie und ards sind fl ießend und entziehen sich häufi g einer exakten klinischen defi nition (american thoracic society et al. 1999 bei zentraler atemdepression und bei pulmonal bedingter respiratorischer insuffi zienz mit erniedrigtem po 2 und erhöhtem pco 2 ist eine endotracheale intubation und eine assistier-> > te oder kontrollierte beatmung angezeigt (artigas et al. 1998; lehmann et al. 2003; seige et al. 2001) . für den beatmungspfl ichtigen patienten mit kardiogenem schock wurde in einer studie eine letalität von 51% angegeben und als wesentliche prognostische risikofaktoren ein apache-ii-sore >29, ein serumkreatinin >180 µmol/l und eine linksventrikuläre auswurffraktion <40% gefunden (lesage et al. 2004 vorbedingung für prophylaxe und therapie (kierdorf 2001 der routinemäßige einsatz von niedrigdosiertem dopamin zur nephroprotektion bei schock-, mods-und sepsispatienten kann nicht empfohlen werden (7 abschn. 6.4.7). diese substanz ist für die verbesserung der nierendurchblutung besonders attraktiv. ein signifi kanter überlebensvorteil durch die gabe von diuretika bei intensivpatienten mit akutem nierenversagen konnte bisher nicht gezeigt werden. eine retrospektive untersuchung erbrachte sogar im zusammenhang mit einer diuretikagabe eine letalitätssteigerung und eine erhöhung der inzidenz einer terminalen niereninsuffi zienz (mehta et al. 2002 bei freisetzung von chromoproteinen (hämolyse, myolyse), bei paraproteinämie und aminoglykosidinduziertem anv wirkt eine alkalisierung mit bikarbonat protektiv. sie wurden bislang unter der vorstellung, eine zelluläre schädigung zu verhindern oder abzuschwächen, bei folgenden formen des anv zur nephroprotektion eingesetzt: bei ischämieinduziertem anv nach nierentransplantation und bei nephrotoxininduziertem anv (kontrastmittel, aminoglykoside). zur differenzialtherapie stehen verschiedene verfahren zur verfügung: hämofi ltration, hämodiafi ltration, high-fl ux-dialyse, ultrafi ltration, plasmapherese, hämoperfusion (kierdorf 2001). bei der hämofi ltration liegt die standardhämofi ltrationsrate im anv bei 1-2 l/h. nicht bestätigt hat sich die hoffnung, damit relevante mengen deletärer mediatoren eliminieren zu können: zwar können immunmodulatorische substanzen wie zytokine und kardiotoxische faktoren hämofi ltrationsmembranen passieren, ein wesentlicher abfall der plasmakonzentrationen dieser substanzen scheint jedoch dadurch nicht zustande zu kommen, obwohl eine vorübergehende kreislaufstabilisierung -messbar als anstieg des systemischen gefäßwiderstandes -beschrieben ist (hoffmann et al. 1996 (hoffmann et al. , 1999 . auch die »high-volume-hämofi ltration« mit durchsatzraten von 6 l/h kann diesbezüglich trotz einsparung von vasopressoren kaum überzeugendere ergebnisse liefern (cole et al. 2001) . eine effi ziente endotoxinelimination bei patienten mit gramnegativer sepsis und mods sowie mit peritonitis ist mit dem matisse-adsorber möglich (reinhart et al. 2004 ). an makroporöse trägerkügelchen gebundenes immobilisiertes humanes serumalbumin bindet während eines hämoadsorptionzyklus endotoxin und bewirkt auf diese weise einen deutlichen abfall der serumendotoxinkonzentration. in einer phase-ii-studie mit 145 patienten mit vermuteter gramnegativer sepsis, davon 104 mit peritonitis, führte die tägliche endotoxinadsorption in den ersten 4 tagen nach diagnosestellung im trend zu einer vorübergehenden besserung des mods (stärkerer abfall des apache-ii-und des sofa-score während der ersten 4 tage) und zu deutlicheren senkung des serumendotoxins; die letalität -nicht primäres zielkriterium (!) -war mit und ohne endotoxinabsorptionstherapie nicht unterschiedlich (reinhart et al. 2004) . für aphereseverfahren (formica et al. 2003; kellum 2003; samtleben et al. 1998; shehata et al. 2002; stegmayr 2001; stegmayr et al. 2003) und die gekoppelte plasmafi ltrationadsorption sind bei patienten mit septischem mods günstige effekte bisher nur in fallberichten und in relativ kleinen kontrollierten studien beschrieben. die schwere leberinsuffi zienz ist charakterisiert durch eine, möglicherweise stickoxidbedingte, hyperdyname systemische kreislaufsituation mit erhöhtem herzzeitvolumen und erniedrigtem systemischem gefäßwiderstand. reaktiv dazu kommt es zur konstriktion der nierenarterien und damit zur na + -retention und aszitesbildung. diese hochgradige renale vasokonstriktion bei leberschädigung wird durch eine aktivierung des renin-angiotensin-aldosteron-systems und eine noradrenalinfreisetzung bewirkt und als hepatorenales syndrom bezeichnet. es ist charakterisiert durch eine oligurie (<300 ml/24 h), einen anstieg des serumkreatinins trotz adäquatem blutdruck und eine erniedrigte urin-na + -konzentration von <10 mmol/ l. im initialstadium ist es reversibel, im weiteren verlauf treten jedoch tubulusschäden auf. eine gesicherte therapie ist nicht bekannt, ggf. muss hämodialysiert werden. obwohl neurone sehr ischämieempfi ndlich sind, kommt es durch die protektive autoregulation der hirndurchblutung erst in relativ späten schockstadien zu ausgeprägteren zerebralen durchblutungsstörungen. ohne vorbestehende zerebrovaskuläre insuffi zienz fi nden sich erst bei einem abfall des arteriellen mitteldrucks auf 50-60 mmhg irreversible störungen der ischämieempfi ndlichsten kortex-und rückenmarksareale. bereits vorher können jedoch reversible bewusstseinsstörungen in form von konfusionen bis zur bewusstlosigkeit auftreten, je nach grad der durchblutungsstörung, ggf. verstärkt durch begleitende störungen des säure-basen-und des elektrolythaushalts. das elektroenzephalogramm zeigt dabei unspezifi sche veränderungen (zauner et al. 2000) . septische enzephalopathie. die septische enzephalopathie ist charakterisiert als eine reversible dysfunktion des zentralnervensystems ohne erkennbare strukturelle schäden, als deren ursachen toxinwirkungen oder ein geändertes neurotransmittermuster diskutiert werden (papadopoulos et al. 2000) . sie kann sich bereits bei blutdruckwerten >60 mmhg manifestieren und geht mit einer erhöhten letalität einher (briegel 2003; eggers et al. 2003; green et al. 2004) . sie äußert sich als irritabilität, agitation, desorientiertheit, konfusion, stupor und koma. eine spezifi sche therapie der septischen enzephalopathie ist z. z. noch nicht etabliert. bei erfolgreicher behandlung der sepsis kann in der regel mit einer raschen besserung der septischen enzephalopathie gerechnet werden. über 50% aller patienten mit länger bestehender sepsis und multiorganversagen scheinen eine axonale sensorische und motorische neuropathie zu entwickeln, die charakterisiert ist durch abgeschwächte oder fehlende sehnenrefl exe und schlaffheit der extremitätenmuskulatur (hund 2001) . die hirnnerven sind intakt; im liquor fi ndet sich bei manchen patienten eine erhöhung des proteingehalts, die zellzahl ist normal. eine spezifi sche therapie ist nicht bekannt. eine retrospektive analyse (33 patienten mit multiorganversagen) erbrachte erste hinweise auf eine mögliche günstige wirkung einer frühzeitigen immunglobulin-gma-gabe bei der polyneuropathie kritisch kranker mit gramnegativer sepsis (mohr et al. 1997) ; als gesicherte therapie kann dieses konzept z. z. noch nicht empfohlen werden. septisches und nichtseptisches mods sind durch eine autonome dysfunktion geprägt, die sich als einschränkung der herzfrequenzvariabilität sowie der baro-und chemorefl exsensitivität bemerkbar macht, im ausmaß mit dem schweregrad des mods korreliert und auch bei intensivpatienten verlässlich und praktikabel messbar ist (schmidt et al. 2001; schmidt u. müller-werdan 2003) . eine gezielte therapie der autonomen dysfunktion des mods-patienten existiert bisher nicht. generalisierte muskelschwäche und abgeschwächte oder fehlende muskeleigenrefl exe kennzeichnen auch das klinische bild der myopathie des kritisch kranken (hund 2001) . diese ist durch eine typ-i-und v. a. durch eine typ-ii-faseratrophie gekennzeichnet und geht nur selten mit einer erhöhung der serumkreatinkinasewerte einher. differenzialdiagnostische schwierigkeiten kann die abgrenzung zur polyneuropathie des kritisch kranken (muskelbiopsie), zur myopathie durch glukokortikoide und muskelrelaxanzien sowie zur -sehr seltenen -septisch-metastatischen pyomyositis bereiten. das klinische bild von polyneuropathie und myopathie ist durch eine schwäche der extremitäten, hyporefl exie, verzögerte respiratorentwöhnung und durch eine komplikationsreiche, verlängerte motorische rehabilitiation mit erhöhter letalität gekennzeichnet. muskelrelaxanzien und kortikosteroide können zusätzliche schädigende effekte am neuromuskulären system hervorrufen, die sich als transiente neuromuskuläre blockade, axonale motorische neuropathie oder myopathie der dicken filamente zeigen. wegen der eingeschränkten klinisch-neurologischen beurteilung eines intensivpatienten kommt den elektrophysiologischen untersuchungen (elektromyo-und elektroneurographie), der messung der kreatinkinase im serum, der muskel-und nervenbiopsie, der liquorpunktion und gelegentlich der kernspintomographie eine große diagnostische und differenzialdiagnostische bedeutung zu. nach schwerem krankheitsverlauf fi nden sich bei mehr als 90% der patienten noch nach jahren muskelschwäche und neurologische defi zite (fletcher et al. 2003) . die schockverursachte sympathikusaktivierung manifestiert sich am herzen als tachykardie, selten bei hämorrhagischem schock als eine vagusvermittelte bradykardie, und in form von tachykarden supraventrikulären und ventrikulären rhythmusstörungen. bei koronarkranken patienten kann sie über die steigerung des myokardialen o 2 -verbrauchs (bei gleichzeitiger hypotonie) ein defi zit der koronarperfusion hervorrufen und damit eine myokardischämie provozieren. eine sympathikusvermittelte gefäßkonstriktion wird an den koronararterien durch die gefäßautoregulation (7 abschn. 6.2.3 und . tabelle 6.8) weitgehend verhindert. die sympathikusaktivierung ist auch für die bei manchen schockformen (septisch, hypovolämisch, traumatisch) beschriebene steigerung der myokardkontraktilität verantwortlich, die jedoch durch zirkulierende kardiodepressive schockfaktoren auch vermindert werden kann. die akute septische kardiomyopathie wird in den 7 abschn. 6.2.8 und 6.6.2 dargestellt. zur komplexen organdysfunktion des kreislaufs bei den verschiedenen schockformen 7 abschn. 6.2. im schock reagieren die splanchnikusgefäße auf die sympathikusaktivierung mit einer raschen und intensiven vasokonstriktion. demzufolge ist der darmtrakt sehr ischämiegefährdet; typische schädigungsmuster sind in . tabelle 6.16 aufgeführt. aufgepfropft auf die darmischämie können die in der reperfusionsphase gebildeten o 2 -radikale eine schädigung der darmbarriere bewirken (bahrami et al. 1998; stallmach u. zeitz 1998) . alle symptome treten nur inkonstant auf. übelkeit, erbrechen und schmerzen im rechten oberbauch gehen dem fieber häufi g voraus; die allerdings nur selten palpable raumforderung im rechten oberbauch sichert weitgehend die diagnose. laborparameter helfen diagnostisch häufi g nicht weiter; wegweisend ist die sonographie. die therapie besteht in der gallenblasenentfernung (letalität ca. 13%) oder in der perkutanen, transhepatische gallenblasenpunktion mit und ohne drainageeinlage (cholezystotomie). wenige stunden nach einem akuten stressereignis können erosionen und ulzera der schleimhaut vorwiegend im magenkorpus, weniger im antrum oder duodenum, auftreten. die pathophysiologie dieser läsionen ist komplex und unterscheidet sich von der der durch säure und helicobacter pylori hervorgerufenen ulzera. durch eine mukosale minderversorgung mit oxygeniertem blut als folge von minderperfusion oder hypoxie wird ein schleimhautschädigender circulus vitiosus, bestehend aus venöser stase, sludge, vasospasmus, gewebehypoxie, mediatorenfreisetzung, radikalbildung und autokongestion der gefäße in gang gesetzt. die magensäuresekretion ist bei den meisten kritisch kranken reduziert und nur bei patienten mit sepsis, verbrennungen, erhöhtem intrakraniellem druck oder schädel-hirn-trauma erhöht (ruß et al. 2005 ). eine gastrointestinale blutung des kritisch kranken kann 2 schweregraden zugeordnet werden (ruß et al. 2005) : zum einen die offenkundige blutung, nachgewiesen durch hämatemesis oder aspiration aus einer gastralen sonde, und zum anderen die klinisch relevante blutung, bestehend aus einer offenkundigen blutung mit arterieller hypotension und transfusionsbedarf von 2 erythrozytenkonzentraten. die rate der offenkundigen blutung liegt bei beatmeten intensivpatienten bei 9% und im gesamtkollektiv der intensivpatienten bei 4,4%. die häufi gkeit klinisch relevanter gastrointestinaler blutungen bei kritisch kranken auf der intensivstation ist in den letzten 20 jahren von bis zu 20% ohne prophylaxe auf z. z. ungefähr 1,5% gesunken. dafür sind neben der ulkusprophylaxe überwiegend die verbesserten intensivmedizinischen therapiemöglichkeiten verantwortlich. für das auftreten klinisch relevanter stressulkusblutungen im gesamtkollektiv der intensivpatienten konnten in einer großen prospektiven studie nur 2 der vermuteten risikofaktoren statistisch signifi kant bestätigt werden: in der vorbeugung einer stressulkusblutung des kritisch kranken sind h 2 -rezeptorantagonisten, sucralfat und antazida hocheffektiv gegenüber placebo (ruß et al. 2005) . bei einem ph-wert <4 ist der magensaft von 90% der intensivpatienten steril. dies gilt nur noch für 15% bei einem intragastralen ph-wert >4 unter säurehemmender medikation, bedingt durch eine ph-wert-abhängige kolonisation des magensaftes mit gramnegativen keimen. diese erreger können u. a. durch regurgitation und/oder weitere kolonisation des oropharyngealen raumes und anschließender mikroaspiration in das bronchialsystem gelangen und nosokomiale, besonders beatmungsassoziierte pneumonien hervorrufen. der vergleich von ranitidin (3-mal 50 mg i.v./tag) mit sucralfat (4-mal 1 g intragastral/tag) zeigte in der prophylaxe einer klinisch relevanten blutung bei für mehr als 48 h beatmeten intensivpatienten eine signifi kant niedrigere blutungsrate für ranitidin (1,7 vs. 3,8%). im gegensatz zu früheren untersuchungen und metaanalysen konnte kein signifi kanter unterschied der sterblichkeit (23,5 vs. 22,8%) und nur ein trend in der reduzierung der gesamtrate beatmungsassozierter pneumonien (19,1 vs. 16,2%) unter sucralfat nachgewiesen werden. die für h 2 -rezeptorantagonisten, antazida und sucralfat beschriebene wirksamkeit bei der stressulkusprophylaxe kritisch kranker gilt auch für die protonenpumpeninhibitoren, wobei für diese substanzgruppe ähnlich umfangreiche untersuchungen und analysen bisher nicht vorliegen (weitere dosierungsempfehlungen s. ruß et al.2005 ). die leber reagiert empfi ndlich auf hypotonie und minderperfusion. dennoch ist die »schockleber« -massive ischämische nekrosen mit sehr hohem transaminasenanstieg -ohne vorbestehende lebererkrankung ein seltenes ereignis (böker u. manns 1998). viel häufi ger dagegen ist eine zentrilobuläre schädigung mit einem milden anstieg der transaminasen, der ldh, des bilirubins und der alkalischen phosphatase. der transaminasenanstieg erreicht am 1.-3. tag sein maximum, er normalisiert sich über die folgenden 3-10 tage. trotz der hepatischen synthese von akute-phase-proteinen ist die proteinsynthese der leber im kreislaufschock eingeschränkt; dies trifft v. a. für präalbumin, albumin und gerinnungsfaktoren zu. auch nach erfolgreicher schockbehandlung kann die biliäre stase mit erhöhten werten für bilirubin und alkalische phosphatase lange persistieren. die septische hepatopathie imponiert klinisch durch den ikterus, funktionell ist die eingeschränkte syntheseleistung das entscheidende. eine spezifi sche therapie ist nicht bekannt. therapierefraktärer aszites, hepatorenales syndrom, hyponatriämie und hepatopulmonales syndrom können bei patienten mit chronischen lebererkrankungen und mods/ sepsis den sepsisverlauf komplizieren (ruß et al. 2005) . pathophysiologie mods, sirs und sepsis verursachen eine aktivierung des gerinnungssystems sowie eine initiale aktivierung und anschließende hemmung der fibrinolyse (huhle 2003; dempfl e 2003; riess 1998 )! diese veränderungen führen schließlich zur disseminierten intravasalen gerinnung (dic) -ausdruck einer mikroangiopathischen hämolyse, verbrauchsthrombozytopenie, verbrauch an gerinnungsfaktoren und mikrothromben. an der pathophysiologischen bedeutung der thrombozyten bei verschiedenen klinischen und experimentellen schockzuständen ist nicht mehr zu zweifeln. bei einer reihe von noxen, die direkt (z. b. biogene amine, thrombin usw.) oder indirekt (z. b. kollagen) nach freisetzung aus subendothelialen gefäßstrukturen auf die thrombozyten einwirken, kommt es zu deren aggregation. diese aggregation ist zunächst reversibel. ihr kann unter geeigneten bedingungen die spontane desaggregation folgen. untersuchungen an patienten mit quantifi ziertem schweregrad einer sepsis (elebute-score) und eines multiorganversagens (apache-ii-score) sprechen dafür, dass die sepsis einen hyperaggregiblen, aber noch reversiblen zustand des thrombozyten induziert (erhöhte fibrinogenrezeptoraktivität, gemessen als libs1-expression), während der zunehmende schweregrad des multiorganversagens zur irreversiblen thrombozytendegranulierung führt, erkenntlich an der verstärkten oberfl ächenexpression der thrombozytenadhäsionsmoleküle gmp-140 und thrombospondin mit zunehmendem schweregrad des multiorganversagens (gawaz et al. 1995) . diese normalerweise in den plättchengranula gespeicherten adhäsionsmoleküle verstärken durch ihre oberfl ächenaggregation die plättchenmikroaggregation und führen somit zur irreversiblen thrombozytendegranulation im sinne der dic. die dic ist klinisch durch die kombination von blutungen (petechien, purpura, ekchymosen, verstärkte blutungen nach gefäßpunktionen) und thrombosen (gangrän, akrale zyanose, hautnekrosen, tiefe venenthrombosen) charakterisiert (dempfl e 2003). sie fi ndet sich bei unterschiedlichen erkrankungen, wobei sepsis und septischer schock im rahmen des mods im vordergrund stehen. die dic bei gramnegativem schock (wo sie am häufi gsten gesehen wird) wird mit dem endotoxin als auslöser in verbindung gebracht (7 abschn. 6.2.5); die fulminant verlaufende dic bei meningokokkensepsis (waterhouse-friderichsen-syndrom), die mit sehr hohen endotoxinspiegeln einhergeht, liefert dafür ein gewichtiges argument. allerdings fi ndet sich die dic auch bei grampositiver sepsis, wie z. b. der pneumokokkensepsis. beim hämorrhagischen schock ist das auftreten einer dic ein seltenes ereignis; hier steht die verdünnungsthrombozytopenie nach adäquater volumensubstitution im vordergrund. abfall von thrombozytenzahl, fibrinogen und antithrombin, anstieg von ptt, prothrombinzeit, thrombinzeit, d-dimeren, fibrinmonomeren, thrombin-antithrombin-komplex, fibrinopeptid a und prothrombinfragmenten (f 1 + f 2 ). im sofa-score (. tabelle 6.17) dient die thrombozytopenie als maß für den schweregrad der gerinnungsstörung in der sepsis. kontrollierte studien zur prophylaxe der dic bei mods, schock und sepsis gibt es bisher nicht. die gesicherten therapieempfehlungen zur therapie der klinisch manifesten dic bei mods, schock und sepsis sind in der . übersicht 6.9 zusammengefasst (riess 1998; . gerinnungshemmende medikamente werden bei sepsis nicht nur bei klinisch manifester dic, sondern häufi g auch unabhängig davon eingesetzt (kujath et al. 2003; werdan et al. 2005) . dieses vorgehen beruht zum einen auf der vorstellung einer »prophylaxe« der ingangsetzung der pathogenetisch gefährlichen gerinnungskaskade. zum anderen wird zunehmend evident, wie eng gerinnungsaktivierung und proinfl ammation in der sepsis verknüpft sind; darauf beruht das konzept, mit der bremsung der gerinnungskaskade -v. a. der thrombinaktivierung -auch das überschießende proinfl ammatorische potenzial des körpers zu dämpfen. lange zeit war heparin ein nachdrücklich propagierter ansatz bei sepsis, wahrscheinlich aber ein wenig oder sogar nicht wirksamer und dazu noch ein potenziell gefährlicher. in den letzten jahren konzentriert sich das interesse v. a. auf den ersatz endogener inhibitoren einer unerwünschten generalisierung der gerinnungsabläufe, die in der sepsis erniedrigte serumspiegel zeigen und meist mit einer ungünstigen prognose verknüpft sind: antithrombin, protein c, protein s und gewebsthromboplastininhibitor (tissue-factor-pathway-inhibitor, tfpi). und schließlich darf nicht vergessen werden, dass in der sepsis thrombozytopenie und thrombozytendysfunktion ebenfalls gerinnungsprobleme machen können. wenig bekannt ist, dass thrombozyten auch mikrobiozide peptide sezernieren und über kontaktabhängige mechanismen zur bakterienabtötung beitragen können (. übersicht 6.10). in der prowess-studie (»effi cacy and safety of recombinant human activated protein c for severe sepsis) wurden patienten in der frühphase einer schweren sepsis (sepsis mit organversagen, . tabelle 6.4) eingeschlossen und entweder mit aktiviertem protein c (drotrecogin alfa aktiviert, xigris) oder mit placebo behandelt (bernard et al. 2001) . nach einschluss von 1690 patienten wurde die studie wegen der überlegenheit des verumpräparates vorzeitig abgebrochen: die 28-tage-letalität der patienten mit standardtherapie lag bei 30,8%, die der patienten mit aktiviertem protein c bei 24,7% (p =0,005). dies entspricht einer letalitätssenkung absolut von 6,1% und relativ von 19,4% (nnt 16). der günstige effekt war dabei unabhängig von der höhe des initialen plasma-protein-c-spiegels. nicht alle sepsispatienten scheinen in gleicher weise zu profi tieren: pneumonie, patientenalter >50 jahre, apache-ii-score ≥25, mehrorganversagen und schock sind faktoren, bei denen eine besonders erfolgreiche behandlung erwartet werden kann; die prognostisch günstige urosepsis und die nichtabominelle chirurgische sepsis scheinen dagegen eher weniger anzusprechen. demzufolge ist die indikation in den usa auf den erwachsenen patienten mit schwerer sepsis und einem hohem letalitätsrisiko (wie einem apache-ii-score ≥25) und in europa auf den erwachsenen sepsispatienten mit mindestens 2 organversagen ausgerichtet. der gestörte stoffwechsel bei patienten mit mods, schock und sepsis und die erforderlichen ernährungsmaßnahmen besitzen für die prognose des kritisch kranken eine wesentlichere bedeutung als lange angenommen . bei patienten mit mods ist -wie bei allen kritisch kranken -eine möglichst frühzeitige enterale ernährung anzustreben (ruß et al. 2005) . trotz des ausgeprägten katabolen zustands kritisch kranker und insbesondere der schock-, mods-und sepsispatienten kann der gesteigerte abbau nicht einfach durch ein mehr an zugeführten kalorien ausgeglichen werden, da diese patienten, insbesondere mit septischem schock, gar nicht in der lage sind, ein übermaß an kalorienangebot zu verwerten. in der an die akute katabole krankheitsphase anschließenden anabolen erholungsperiode muss dann durch eine überdurchschnittliche kalorienzufuhr dem gesteigerten bedarf rechnung getragen werden (ruß et al. 2005) . immunonutrition. der einsatz der immunonutrition -die enterale zufuhr von mehrfach ungesättigten fettsäuren, nukleotiden und arginin sowie die parenterale/enterale gabe von glutamin -zur verbesserung des immunstatus kritisch kranker scheint bei intensivmedizinischen risikopatienten -ards (pacht et al. 2003) , mods-und sepsisgefährdete patienten in der perioperativen phase -günstige effekte zu haben; bei manifester sepsis sprechen studiendaten eher für eine ungünstige wirkung (bertolini et al. 2003 , heyland u. samis 2003 suchner 2002) . welche »drüsen« sind mods-anfällig? derzeit richtet sich das »endokrinologische augenmerk« bei mods, schock und sepsis auf den insulin-glukose-stoffwechsel einerseits (7 abschn. 6.5.10) und die hypothalamus-hypophysen-nebennieren-achse (»hypothalamo-pituitary-adrenal axis«, hpa-achse) andererseits (beishuizen u. thijs 2003; werdan et al. 2005) . endotoxin kann die hpa-achse auf unterschiedlichen ebenen beeinfl ussen (beishuizen u. tijs 2003 über verschiedenene mechanismen kann es im septischen schock zu einer relativen nebennierenrindeninsuffi zienz kommen . bei diesen patienten wird in unerwünschter weise die antiinfl ammatorische wirkung des endogenes cortisols abgeschwächt. die folge ist ein vermindertes ansprechens des kreislaufs auf vasopressorische katecholamine und wahrscheinlich auch eine erhöhte sterblichkeit (annane u. cavaillon 2003) . im prolongiert verlaufenden septischen schock geht die negative feedbackkontrolle der hypothalamus-hypophysen-nebennierenrinden-achse (hpa-achse) verloren, mit einem verlust des pulsatilen sekretionsverhaltens des cortisols. ursache dafür sind eine tnf-bedingte sekretionshemmung von corticotropin-releasing-hormon und acth sowie eine abschwächung der acth-stimulierbarkeit der cortisolproduktion der nebenniererenrinde. infolge der down-regulation zellulärer glukokortikoidrezeptoren mit abnahme von rezeptorzahl und affi nität bildet sich im septischen schock eine glukokortikoidresistenz aus. wahrscheinliche ursache dafür ist eine hohe lokale konzentration proinfl ammatorischer zytokine, die über die bildung von transkriptionsfaktoren wie activator-protein 1 (ap1) und »nuclear factor kappa b« (nfκb) zu einer komplexbildung mit aktiviertem glukokortikoid-rezeptorelementen führt. da die glukokortikoid-rezeptorzahl in den gefäßen mit dem mittleren arteriellen blutdruck korreliert, trägt die glukokortikoid-down-regulation entscheidend zur hämodynamischen instabilität im septischen schock bei (zonghai et al. 2003) . bei zwei dritteln aller patienten mit septischem schock lässt sich anhand des acth-tests die diagnose einer relativen nebennierenrindeninsuffi zienz stellen. bei diesen patienten können stress-hydrocortison-dosen von 200-300 mg/tag die erworbene glukokortikoidresistenz überwinden und die proinfl ammation im septischen schock dämpfen. für den behandelnden intensivmediziner ist es beeindruckend, wie unter dieser hydrocortisongabe die katecholaminansprechbarkeit der gefäße wiederhergestellt und damit innerhalb weniger tage eine drastische einsparung von katecholaminen möglich wird (annane et al. 2002) . die in einer relativ kleinen studie mit 300 patienten gefundene letalitätssenkung durch hydrocortison bei patienten mit septischem schock und relativer nebennierenrindeninsuffi zienz (annane et al. 2002) wird z. z. in der europäischen corticus-studie mit geplanten 800 patienten überprüft. nicht nur bei septischem schock, sondern auch bei eskalierendem sirs nach herzchirurgie scheinen stressdosen von hydrocortison die überschießende infl ammationsreaktion zu dämpfen und die prognose zu verbessern (kilger et al. 2003 ). die hohe infektanfälligkeit kritisch kranker und insbesondere der patienten im schock ist folge einer komplexen schädigung des humoralen und zellulären immunsystems bei schock und sirs (. tabelle 6.4, . abb. 6.4a-c und 6.6). von den möglichkeiten einer infektions-/sepsisprophylaxe bei intensivmedizinischen risikopatienten sind neben allge-> meinen hygienemaßnahmen und der erzielung einer normoglykämie mittels hochdosierter insulingabe (7 abschn. 6.5.10) v. a. die möglichkeiten einer prophylaxe mit immunglobulinen zu nennen (werdan 2001b; werdan et al. 2005) . durch prophylaktische gabe von immunglobulinen (ig) kann das auftreten von infektionen bei verschiedenen intensivpatientenkollektiven gesenkt werden, v. a. bei »infektions-hochrisiko-patienten« und bei »infektions-hochrisiko-operationen« (werdan 2001b) : anerge patienten (ca. 8%) nach herzchirurgischen operationen mit der herz-lungen-maschine: die prophylaktische gabe von iviggma senkt die im vergleich zu normergen patienten höhere inzidenz postoperativer infektionen auf werte unter diejenige normerger patienten (kress et al. 1999 einheiten, schweres abdominelles oder retroperitoneales trauma mit transfusionsbedarf von mehr als 10 einheiten und intubationspfl ichtigkeit länger als 24 h. bei patienten mit schwerem trauma (injury severity score 16-50) senkt die prophylaktische gabe von ivigg (0,25 g/kgkg an den tagen 1,2,3,6) zwar die häufi gkeit von infektionen, insbesondere pneumonien (um mehr als 50%), die infektionsverursachte morbidität und letalität wird allerdings nicht vermindert (douzinas et al. 2000) . spezifi sche schockformen k. werdan, r. prondzinsky, m. buerke, u. müller-werdan unabhängig von der art des kardiogenen schocks (. tabellen 6.1 und 6.2) besteht die initialbetreuung des patienten mit kardiogenem schock (. übersicht 6.11) zunächst in der hämodynamischen stabilisierung und ausreichenden oxygenierung, dem adäquaten hämodynamischen monitoring und der möglichst raschen klärung der schockursache. anschließend lassen sich, soweit möglich, kausale behandlungsmaßnahmen einleiten, die im infarktbedingten myogenen pumpversagen ischämischer genese in der möglichst raschen wiedereröffnung des verschlossenen infarktgefäßes besteht (. abb. 6.18). die schockbehandlung ist ein wettlauf mit der zeit! jede »schockminute« erhöht das risiko des auftretens des prognosebestimmenden multiorgandysfunktionssyndroms (7 abschn. 6.5)! die behandlung des kardiogenen schocks kann in symptomatische (medikamentöse therapie, mechanische unterstützungssysteme) und in kausale (interventionelle oder chirurgische koronarreperfusion, verschluss von shunts oder rupturstellen) maßnahmen eingeteilt werden. ziel der medikamentösen, symptomatischen (!) behandlung (bengur u. meliones 1998; goldberg et al. 1999; hollenberg 2003; hollenberg et al. 1999) ist die steigerung der myokardkontraktilität und die situationsbezogene optimierung von vor-und nachlast (volumenentzug, vasodilatatoren, blutdruckstabilisierung), um so -gesteuert mittels invasivem monitoring -einen adäquaten herzindex und blutdruck und damit eine ausreichende perfusion der vitalorgane aufrechtzuerhalten (7 abschn. 6.4.1, cotter et al. 2003a ). im wesentlichen sind es die katecholamine dobutamin, dopamin, noradrenalin und adrenalin und die phosphodies-! terasehemmstoffe amrinon, milrinon und enoximon, deren positiv-inotrope wirkung ausgenutzt werden kann (7 abschn. 6.4.7; . tabellen 6.14 und 6.15). digitalis. digitalis spielt bei sinusrhythmus in der therapie des kardiogenen schocks als inotropikum keine wesentliche rolle; dagegen ist es bei tachykardem vorhoffl immern und vorhoffl attern zur frequenznormalisierung das antiarrhythmikum der wahl. levosimendan. als neues therapieprinzip bei akuter herzinsuffi zienz scheint der kalziumsensitzer levosimendan erfolgreich (7 abschn. 6.4.7). der kardiogene schock führt häufi g zu lungenstauung/lungenödem und zum prärenalen nierenversagen und damit zur oligurie. neben der kreislaufstabilisierung können diuretika und hämofi ltration erforderlich werden (7 abschn. 6.5.4). zur senkung der vor-und nachlast dient bei akutem herzinfarkt in erster linie nitroglyzerin und in zweiter linie nitroprussidnatrium (7 abschn. 6.4.7). bei dekompensierter herzinsuffi zienz ist nesiritide, das endogene b-typ-natriuretische peptid (in deutschland nicht zugelassen) ein neuer therapieansatz (7 abschn. 6.4.7). zum experimentellen einsatz von stickoxidsynthetaseinhibitoren (l-name/l-nmma), inhibitoren der komplementkaskade und von pyruvat 7 abschn. 6.4.9. antiarrhythmische therapie (7 kap. 3) die konservativ nicht beherrschbare schocksymptomatik kann ein chirurgisches eingreifen erforderlich machen. die akutherztransplantation dieser patienten wird zwar weiterhin die ausnahme bleiben, mit den ventrikulären assist-systemen (wie z. b. dem novacor) stehen dem chirurgen jedoch über wochen einsetzbare bridging-maßnahmen zur verfügung (martin et al. 2003) , die auch dem patienten mit kardiogenem schock zugute kommen könnten. bei kardiogenem schock infolge eines myokardinfarkts ist aufgrund der guten erfolge der akut-pci eine aortokoronare bypass-notfalloperation nur noch selten nötig. als mögliche indikationen gelten hierfür: fortbestehende ischämie bei koronarographisch nachgewiesener mehrgefäßerkrankung und/ oder lca(linke koronararterie)-hauptstammstenose; nach pci-komplikationen bzw. nicht erfolgreicher pci. die operation sollte dann aber notfallmäßig erfolgen, da die präoperative ischämiedauer der entscheidende prädiktor der perioperativen letalität ist. auch bei der perkutanen koronarintervention (pci) muss mit einem akuten koronarverschluss und mit einer letalität von 0,4-1,0% rechnen, vorwiegend als folge eines sich entwickelnden kardiogenen schocks. vor allem bei der hochrisiko-pci (hochgradig eingeschränkte linksventrikuläre pumpfunktion, dreigefäßkrankheit, das zu dehnende gefäß als das . übersicht 6.11. basisbehandlungskonzept des kardiogenen schocks katecholamine, (phosphodiesterasehemmer) ggf. ca ++ -sensitizer vom typ des levosimendan vor-und nachlastsenkung (cave: hypovolämie): vorlast: nitrate (0,3-0,5-4 µg/kgkg/min bzw. 0,3-6,0 mg/h) vorlast + nachlast: nitroprussidnatrium (0,1 einzige noch perfundierte koronargefäß) kann das risiko der koronardehnung durch prophylaktisch oder im stand-by genutzte mechanische unterstützungssysteme (. übersicht 6.12) reduziert werden (ferrari u. figulla 2005) . in der wartephase auf die herztransplantation können auch hier ventrikuläre assist-systeme als bridging-verfahren eingesetzt werden. bezüglich der pci bei myokardinfarkt s. unten. die mechanischen unterstützungssysteme im kardiogenen schock reichen von der kardiopulmonalen reanimation (7 abschn. 6.4.2) über die wiederherstellung einer suffi zienten koronarperfusion bis hin zu mehr oder weniger komplexen kreislaufunterstützungssystemen. die jährlich 270.000 patienten mit einem akuten herzinfarkt in deutschland, von denen ca. 5-10% einen kardiogenen schock entwickeln, unterstreichen das quantitative ausmaß, das solche verfahren bereits haben oder haben könnten. mechanische kreislaufunterstützungssysteme fi nden beim akuten herzstillstand (ferrari et al. 1998) , im kardiogenen schock und bei der hochrisiko-ptca anwendung (martin et al. 2003) . beim herzstillstand wird ein komplettes kreislaufunterstützungssystem benötigt, das in 15 min unter fortlaufender kardiopulmonaler reanimation möglichst unter röntgenkontrolle installiert werden kann (ferrari et al. 1998) . im kardiogenen schock ist eine partielle kreislaufunterstützung mittels der intraaortalen ballonpulsation (iabp) für einen zeitraum bis zu etwa 10 tagen möglich. bei der hochrisiko-pci kommt die mechanische kreislaufunterstützung mittels iabp oder kardiopulmonalem bypass je nach befundkonstellation auf der stand-by-basis im bedarfsfall oder elektiv vor beginn der pci zum einsatz. da 90% der akuten reokklusionen innerhalb der ersten 6 h nach pci auftreten, sollten diese verfahren zumindest für diese zeit sofort verfügbar sein (ferrari u. figulla 2005) . methodik, komplikationen, kontraindikationen. ein ballonkatheter (8 f) von ca. 70 cm länge (z. b. ballonlänge 26 cm, insuffl ationsvolumen 40 ml) wird perkutan transfemoral über eine schleuse (z. b. 9 f) eingeführt und so platziert, dass der ballon in der thorakalen aorta distal des abgangs der linken a. subclavia zum liegen kommt. mit einem ekg-gesteuerten augmentierungsmodus von 1:1-1:4 wird der ballon in der diastole aufgeblasen (füllung mit heliumgas) und am beginn der systole abrupt desuffl iert (. abb. 6.19a-c) . die komplika-tionsrate (zumeist ischämie der unteren extremität, arterielle thrombose, infektion, persistierende blutung inguinal; bei längerer anwendung gelegentlich beeinträchtigte leber-und nierenfunktion) liegt aktuell bei 2,7%, bei einer mittleren liegezeit von 3 tagen; eine vorzeitige beendigung der iabp-therapie ist bei etwa 2% der patienten erforderlich (gregg et al. 2003) . kontraindikationen sind schwere iliofemorale gefäßveränderungen, aortendissektion, höhergradige aortenklappeninsuffi zienz und eine blutungsdiathese. wirkprinzip und erzielbare wirkungen. durch das streng auf die diastole beschränkte aufblasen des ballons nimmt der aortendruck phasisch zu (diastolische augmentierung; »kamelhöckerform« der blutdruckkurve; . abb. 6.19a) und bewirkt damit eine zunahme der koronarperfusion. die abrupte desuffl ierung des ballons am beginn der systole führt über eine sogwirkung zur absenkung des bei pumpversagen erhöhten diastolischen ventrikeldrucks (vorlastsenkung) sowie des systolischen aortendrucks (nachlastsenkung) und damit zu einer, allerdings nicht sehr ausgeprägten, senkung des myokardialen o 2 -verbrauchs. im transösophagealen echokardiogramm zeigte sich eine verbesserung der linksventrikulären pumpfunktion, und bei der dopplersonographischen koronarfl ussmessung steigerte sich die maximale sowie die integrale flussgeschwindigkeit um etwa 115% bzw. um ca. 80%. hypotensive patienten mit einem systolischen blutdruck ≤90 mmhg zeigen dabei den größten anstieg der koronaren flussgeschwindigkeit. der fluss im poststenotischen anteil (cohen et al. 2003) . der vorteilhafte effekt der intraaortalen gegenpulsation im zusammenhang mit perkutaner revaskularisation (brodie et al. 1999; lindholm et al. 2003) wie auch unter thrombolytischer therapie (barron et al. 2001; kovack et al. 1997 stomel et al. 1994) ist bei patienten mit infarktbedingtem kardiogenem schock gut belegt. amerikanische kliniken mit hoher iabp-implantations-frequenz (>35/jahr) erzielen bei schockpatienten eine um 30% niedrigere letalität (barron et al. 2001) . bei akutem koronarverschluss nach pci und bei pci-hochrisikopatienten mit mehrgefäßerkrankung und erheblich eingeschränkter pumpfunktion (im mittel 10% aller patienten) lässt sich die iabp mit gutem primärergebnis und ausreichend sicher einsetzen. bei patienten mit kardiogenem schock kann der iabp-einsatz bereits in der notaufnahme sicher durchgeführt werden (bur et al. 2002) . ein früher einsatz der iabp bei nichtherzchirurgischen krankheitsbildern scheint nach den daten der benchmark-registry mit weltweit über 19.636 patienten die prognose zu senken (cohen et al. 2003) . limitationen der iabp-anwendung sind die notwendigkeit eines mindestherzindex von 1,3 l/min/m 2 und eines weitgehend stabilen rhythmus sowie der eher bescheidene erfolg mit einer einsparung des myokardialen o 2 -verbrauchs um nur 10-20%. bei herzchirurgischen patienten mit »low-output-syndrom« kommt die iabp häufi g zum einsatz. ein prädiktiver score erlaubt die identifi zierung derjenigen patienten, bei denen die iabp-behandlung des low-output-syndroms nicht ausreicht und ein wechsel auf ein herzunterstützungssystem notwendig wird (hausmann et al. 2002) . die katheter können entweder perkutan oder nach chirurgischer gefäßeröffnung in die femoralgefäße eingeführt werden. im falle der »supported angioplasty« bevorzugen wir als anästhesieverfahren die elektive intubationsnarkose. das system kann von einem erfahrenen team innerhalb von 15 min installiert werden. indikationen. indikationen sind die überbrückung akut vital bedrohlicher mechanischer infarktkomplikationen wie der ventrikelruptur, der herzstillstand und die hochrisiko-pci. in den usa erleiden jährlich 70.000-100.000 patienten einen herzstillstand im krankenhaus; diese patienten könnten somit möglicherweise ohne zeitlichen verzug wiederbelebt werden. bei erfolgloser konventioneller reanimation wären diese patienten kandidaten für eine notfall-pcps (ferrari et al. 1998) . die zu erwartenden ergebnisse wären günstig, wenn man die resultate einer multizentrischen studie mit 218 pcps-reanimierten patienten zugrunde legt (34% der mit pcps reanimierten patienten konnten aus dem krankenhaus entlassen werden, im vergleich zu 14% der konventionell reanimierten eines historischen kontrollkollektivs). 7 von 9 patienten mit fulminanter myokarditis und kardiogenem schock überlebten bei vorübergehendem einsatz des perkutanen kardiopulmonalen bypass-systems für 3-10 tage ohne nachfolgende transplantation (kato et al. 1999) . natürlich müssen indikationsrichtlinien für die anwendung des perkutanen kardiopulmonalen bypass-systems vorliegen, soll dieses verfahren mit einer gewissen aussicht auf erfolg eingesetzt werden und nicht nur den irreversiblen schockzustand und damit das leiden des patienten verlängern. als richtlinien können gelten (ferrari u. figulla 2005): keine spontanzirkulation trotz optimaler reanimation über 5 min bei patienten unter 60 jahren mit dokumentiertem plötzlichem herzstillstand, sofort eingeleiteter reanimation und ohne neurologische defi zite; der einsatz der perkutanen kreislaufunterstützungssysteme als überbrückung bis zur diagnostik, bis zur koronarrevaskularisation (ptca), bis zur implantation eines biventrikulären unterstützungssystems (»bridging to bridging«). die bisher beschriebenen mechanischen unterstützungssysteme sind für den einsatz von stunden bis tage gedacht; sie sind geeignet, eine akute schockphase zu überwinden. jüngst konnte auch mittels eines perkutan einsetzbaren mechanischen unterstützungssystems (tandem-heart) ein günstiger verlauf für patienten im protrahierten kardiogenen schock aufgezeigt werden (thiele et al. 2001) , ebenso wie für das impella-system (ferrari u. figulla 2005) . komplexere ventrikuläre unterstützungssysteme, wie z. b. das linksventrikuläre system novacor oder biventrikuläre künstliche herzen, dienen für terminal herzinsuffi ziente patienten zur überbrückung der wartephase auf ein spenderherz (martin et al. 2003) . bei der konventionellen herzdruckmassage (hdm) wird ein herzzeitvolumen von nur 25-30% des normalwertes erreicht. diastolische drücke von 40 mmhg und systolische drücke von 60-80 mmhg werden selten überschritten. unter den bedingungen der manuellen herzdruckmassage reichen zerebraler und myokardialer blutfl uss nicht aus, um auf dauer einen hypoxischen schaden abzuwenden. um höhere blutfl üsse zu erreichen, werden alternative bzw. zusätzliche methoden zur hdm erprobt, wie die sequenzielle thorakoabdominelle kompression, die cpr mittels pneumatischer weste, die aktive kompressions-dekompressions-reanimation und die sog. »inspiratory threshold valve«. während die pneumatische weste v. a. den kompressionsvorgang automatisiert, haben die anderen genannten verfahren eine steigerung des venösen rückfl usses zum ziel. nach den aktuellen empfehlungen können mechanische hilfsmittel zur cpr verwendet werden. die herzdruckmassage mit interponierter abdomineller kompression (iac, »interposed abdominal compression cardiopulmonary resuscitation«) stellt ein vielversprechendes verfahren dar. bei der herzdruckmassage wird von einem zusätzlichen helfer mit der handfl äche eine kompression von 100±20 mmhg auf die nabelgegend ausgeübt, und zwar alternierend mit der kardiopulmonalen kompression im verhältnis 1:1. dadurch kommt es zu deutlich höheren blutfl üssen als mit konventioneller hdm. die aktive kompressions-dekompressions-pumpe (acd-cpr, »active compression-decompression cardiopulmonary resuscitation«) ist ein vielversprechendes und leicht anwendbares konzept (. abb. 6.21a,b). ein griff wird mittels eines saugnapfes (z. b. cardiopump) auf der brust des patienten fixiert. so kann sowohl druck auf den thorax ausgeübt als auch durch zug an dem griff ein unterdruck erzeugt werden (mit förderung des venösen rückfl usses und einem höheren aortalen blutfl uss bei der nächsten kompression). in mehreren randomisierten prähospitalstudien zeigte der einsatz der cardiopump leider keinen überlebensvorteil. bei patienten, die außerhalb des krankenhauses mit acd-cpr reanimiert worden waren, ließ sich jedoch eine verbesserte langzeitüberlebensrate nachweisen (plaisance et al. 1999 ). > 6.6 · spezifi sche schockformen während in den vergangenen jahren die dokumentierte infarktsterblichkeit seit der einführung der intensivüberwachung, der thrombolyse, effektiver antithrombotischer therapien sowie der perkutanen koronarintervention (pci) von ca. 30% um 1960 auf zwischenzeitlich 6%-7% deutlich gesenkt werden konnte, ist die historisch zwischen 70 und 80% angesiedelte sterblichkeit bei kardiogenem schock infolge eines akuten myokardinfarktes trotz moderner therapiemaßnahmen nicht in diesem ausmaß verbessert worden. unverändert stellt der kardiogene schock die hauptursache der infarktsterblichkeit im krankenhaus dar. ein kardiogener schock entwickelt sich bei 5-10% aller patienten mit akutem herzinfarkt (hochman et al. 1999 , jacobs et al. 2000 menon et al. 2000c) , und zwar am häufi gsten dann, wenn mindestens 35% der linksventrikulären muskelmasse akut oder sukzessive infarziert (menon et al. 2000a; prondzinsky et al. 2004) abb. 6.21a,b. aktive kompressions-dekompressions-pumpe (acd-cpr). a das system besteht aus einem saugnapf (radius 6,5 cm), einem gleitkolben und einem horizontalen griff (gesamthöhe 13,5 cm). b in die oberfl äche des griffes ist eine messskala (kg oder pounds) eingearbeitet. die aktive kompression/dekompression wird mit der mitten auf dem sternum platzierten pumpe mit 80-100 kompressionen (unabhängig vom atemzyklus) durchgeführt. der kompressionsdruck beträgt dabei (abhängig von der steifheit des brustkorbs) 29,5-50,0 kg, entsprechend einer kompressionstiefe von 3,8-5,1 cm. die gleichlange dekompressionsphase mit einem sog von im mittel 9,1 kg dient dazu, den brustkorb ohne kontaktverlust maximal zu expandieren. (nach cohen et al. 1994) . primäre pumpstörung mit schock bei ausgedehntem infarkt die daten der prähospitalen thrombolysestudie (captim; bonnefoy et al. 2002) legen nahe, dass der frühzeitige prähospitale einsatz von thrombolytika tendenziell zur verringerung nachfolgend auftretender schocksituationen führt. diese prospektiv erhobenen daten, die einen günstigen trend zu gunsten der thrombolyse mit einer annähernden halbierung der schockrate im weiteren hospitalverlauf aufwiesen, werden durch eine reihe retrospektiver daten positiv gestützt. verschiedene kleinere studien wie die daten des shock-trial-registry weisen auf einen deutlichen behandlungsvorteil für patienten mit kardiogenem schock in folge linksventrikulären pumpversagens mittels thrombolyse und begleitender intraaortaler gegenpulsation hin, insbesondere bei nachfolgender revaskularisation mittels pci oder aortokoronarer bypass-op. die autoren schlussfolgern, dass für krankenhäuser ohne möglichkeiten zur unmittelbaren koronarintervention eine strategie mit früher systemischer thrombolyse und gleichzeitiger intraaortaler gegenpulsation eine frühestmögliche vorbehandlung darstellt, bei nachfolgendem transfer des patienten in ein entsprechendes herzzentrum mit möglichkeiten zur pci oder aortokoronaren bypass-op. diese daten stehen interessanterweise in einem deutlichen kontrast zu den daten der danami-studie (andersen et al. 2003) , in der bei herzinfarktpatienten auch bis zu 2 h zeitverzögert interventioneller koronartherapie ein eindeutiger behandlungsvorteil zu gunsten der mechanischen koronarrevaskularisiation gegenüber der thrombolyse aufgezeigt werden konnte. hierbei ist jedoch einschränkend zu bemerken, dass die gesamtsterblichkeit in beiden gruppen überdurchschnittlich niedrig war und sich diese daten primär nicht auf schockpatienten übertragen lassen dürften. darüber zeigte sich in der isolierten betrachtung des kombinierten endpunktes nach 30 tagen (tod, reinfarkt, schlaganfall) nur für den einzelnen endpunkt »reinfarkt« ein signifi kanter vorteil für die patienten mit primärer pci. erst die wiederherstellung eines effektiven blutfl usses durch die sofortige wiedereröffnung des verschlossenen koronargefäßes mittels akut-pci (meyer et al. 1982 ) erbrachte nahezu eine halbierung der sterblichkeit. die shock-studie (hochman et al. 1999; ryan 1999) überprüfte die wirksamkeit der akutrevaskularisation (überwiegend akut-pci) bei 302 patienten mit kardiogenem schock nach herzinfarkt im sinne der evidenzbasierten medizin (. tabelle 6.19). primäres zielkriterium war dabei die senkung der 30-tage-letalität, sekundäres zielkriterium u. a. die senkung der 6-monate-letalität. die 17%ige senkung der 30-tage-letalität im gesamtkollektiv war nicht signifi kant unterschiedlich, wohl aber die der 6-monate-letalität, und -in einer nachuntersuchung -die 12-monate-sterblichkeit. besonders die patienten <75 jahren profi tierten von der akutrevaskularisation, ebenso die patienten mit einem zeitintervall <6 h von schmerzbeginn bis randomisierung und die mit einem zweitinfarkt, weiterhin männer mehr als frauen. zu ähnlichen aussagen kommt das alkk-schockregister in deutschland (zeymer et al. 2004 während im shock-trial (hochman et al. 1999 ) der eindeutige behandlungsvorteil auf die altersgruppe der unter-75-jährigen begrenzt war, konnten weitere arbeiten den behandlungsvorteil der pci auch für patienten im höheren alter aufzeigen (antoniucci et al. 2003; dzavik et al. 2003) . das alter stellt zwar bei kardiogenem schock einen eigenständigen risikoprädiktor dar; bei frühzeitiger und technisch erfolgreicher intervention ist allerdings der relative nutzen vergleichbar. die möglichkeit zur stent-implantation und der einsatz von gp iib/iiia-hemmern hat den effi zienzgrad und die sicherheit der akut-ptca bei kardiogenem schock noch weiter gesteigert. die effektivität des glykoproteinrezeptorantagonisten abciximab konnte sowohl bei perkutanen koronarhochri 6.6 · spezifi sche schockformen sikointerventionen als auch insbesondere bei infarktbedingtem kardiogenen schocks aufgezeigt werden (zahn et al. 2003; zeymer et al. 2003) . so wird in einer pci-serie (antoniucci et al. 1998 ) mit 66 schockpatienten über die implantation eines stents bei 47% berichtet; die primäre erfolgsrate lag dabei bei 94%, ein optimales angiographisches ergebnis wurde zu 85% erzielt, die 6-monate-überlebensrate lag bei 71%; und 80% der patienten hatten nach 6 monaten einen nyha-herzinsuffi zienzschweregrad von nur i-ii. von besonderem interesse war, dass gestentete koronarien eine geringere 6-monate-restenose-/reokklusionsrate (20%) hatten als dilatierte, aber nichtgestentete herzkranzgefäße (67%). ebenso ließ sich auch ein langfristiger überlebensvorteil bei erfolgter stent-implantation und verabreichung von abciximab gegenüber einer strategie mit alleiniger ballonangioplastie, alleiniger stent-implantation oder ptca sowie abciximabgabe ohne stent-implantation belegen. der initial signifi kant verbesserte timi-iii-fluss ließ sich auch nach 2,5 jahren als signifi kanter überlebensvorteil für die patienten, die mit stent und abciximab behandelt wurden, nachweisen (antoniucci et al. 2002; chan et al. 2002; giri et al. 2002) . die akut-pci kann bei kardiogenem schock infolge herzinfarkt als das verfahren der wahl angesehen werden (hamm et al. 2004) der überzeugende erfolg der pci im vergleich zur medikamentösen therapie mit i.v.-thrombolyse dürfte darauf zurückzuführen sein, dass mittels pci in einem sehr viel höheren prozentsatz ein timi-grad-3-koronarfl uss (komplette reperfusion) wiederhergestellt werden kann, als dies mit der thrombolyse möglich ist. wegen der günstigen ergebnisse der akut-pci bei infarktbedingtem schock scheint die empfehlung gerechtfertigt, infarktpatienten mit manifestem oder beginnendem schock (katecholaminpfl ichtigkeit) möglichst rasch in ein zentrum mit der möglichkeit zur akut-pci durch ein erfahrenes herzkatheterteam zu verlegen (hamm et al. 2004) . dabei kann durchaus vorab eine i.v.-thrombolyse durchgeführt werden, sie stellt für die akut-pci keine kontraindi-> kation dar. falls verfügbar, sollte der transport mit liegender intraaortaler gegenpulsation (iabp) erfolgen, ein beträchtlicher nutzen der iabp bei den so behandelten patienten (n =46) ist in einer retrospektiven analyse aufgezeigt: 1-jahres-letalität mit iabp-behandlung 33%, 1-jahres-letalität ohne iabp-behandlung 68% (p =0,019; kovack et al. 1997 ). die akute mitralinsuffi zienz (bei 1% aller infarktpatienten; 0,4-5% aller infarkttodesfälle; 2-7 tage, im mittel 4 tage nach hinterwandinfarkt) kann entweder als folge einer papillarmuskeldysfunktion oder einer papillarmuskelruptur entstehen; meistens ist die papillarmuskeldysfunktion von transienter natur und führt nur zu einer milden mitralinsuffi zienz, während der papillarmuskelabriss in der regel ein refraktäres lungenödem und eine schwere beeinträchtigung des kreislaufs hervorruft. die mitralinsuffi zienz infolge papillarmuskelabriss fi ndet sich typischerweise bei älteren patienten mit erstmaligem, oftmals eher kleinem infarkt und nur leicht eingeschränkter auswurffraktion; eine insuffi zienz infolge dysfunktion des halteapparates -ohne papillarmuskelabriss -tritt eher bei jüngeren patienten mit koronarer mehrgefäßerkrankung und eingeschränkter auswurffraktion auf (calvo et al. 1997) . frauen und ältere infarktpatienten sind von der akuten mitralinsuffi zienz, die gemeinsam mit der ruptur des ventrikelseptums etwa 8% aller schockinfarktpatienten ausmacht, häufi ger betroffen; möglicherweise führt die i.v.-thrombolyse zu einem früheren auftreten dieser komplikation (hochman et al. 2000) . bei der mehrzahl der patienten ist die rechte kranzarterie oder der r. circumfl exus das infarktgefäß. bei bis zu 50% aller patienten mit akutem myokardinfarkt lässt sich auskultatorisch oder echokardiographisch eine mitralinsuffi zienz nachweisen. eine höhergradige mitralinsuffi zienz fi ndet sich hingegen nur bei 3% aller infarktpatienten, wobei die hälfte dieser patienten ein lungenödem oder einen manifesten kardiogenen schock entwickelt (tcheng et al. 1992) . das systolikum -in typischer weise holosystolisch über dem apex zu auskultieren und in die axilla ausstrahlend -ist tabelle 6.19. shock-studie: akutrevaskularisation bei patienten mit kardiogenem schock nach herzinfarkt. (nach hochman et al. 1999 (zotz et al. 1993) . das hämodynamische monitoring zeigt eine abnahme des herzzeitvolumens und das auftreten einer v-welle in der pulmonalkapillare (. tabelle 6.2). die letalität innerhalb der ersten 24 h kann bis zu 50% betragen. nach initialer stabilisierung mit vasopressoren, nachlastsenkung (typischerweise mit dem gut steuerbaren nitroprussidnatrium) und implantation der intraaortalen ballongegenpulsation muss bei der papillarmuskelruptur zum frühestmöglichen zeitpunkt ein klappenersatz (ggf. mit anlegen eines aortokoronaren bypasses) durchgeführt werden. die ergebnisse der frühoperation sind mit einer letalität von 44% deutlich besser als die der spätoperation (letalität 78%; thompson et al. 2000) . falls eine papillarmuskelruptur sicher auszuschließen ist und »nur« eine papillarmuskelischämie vorliegt, kann eine pci des den betroffenen papillarmuskel versorgenden koronargefäßes in betracht gezogen werden. eine ventrikelseptumruptur (birnbaum et al. 2002; menon et al. 2000b ) tritt bei 1-3% aller infarkte innerhalb der 1. woche auf und ist etwa gleich auf alle infarktlokalisationen verteilt. die ruptur kann mit neuerlichen herzschmerzen einhergehen. typisch dafür ist das laute holosystolikum mit punctum maximum am linken sternalrand, gelegentlich vergesellschaftet mit einem schwirren. die diagnose lässt sich echokardiographisch stellen (ballal et al. 1993) , der o 2 -sättigungssprung zwischen rechtem vorhof und rechtem ventrikel lässt eine quantitative shunt-abschätzung zu. neben der shunt-größe ist prognostisch auch die lokalisation wichtig: die hämodynamische beeinträchtigung durch den shunt wird v. a. durch die gestörte compliance des rechten ventrikels geprägt, wenn dieser, wie bei einem hinterwandinfarkt, ebenfalls infarziert ist (held et al. 1988) . die hälfte der patienten entwickelt einen kardiogenen schock. die prognose wird in erster linie vom ausmaß der links-und rechtsventrikulären infarzierung und weniger von der größe des defektes bestimmt (cummings et al. 1997) . bei hämodynamischer instabilität besteht nach ansicht der meisten autoren eine dringliche bzw. notfallmäßige operationsindikation, da sich unter konservativer therapie bei diesen patienten häufi g ein multiorgandysfunktionssyndrom entwickelt (lemery et al. 1992; cox et al. 1996; killen et al. 1997) . therapeutisch bietet nur die rasche operative deckung des defektes nach vorheriger implantation der intraaortalen ballongegenpulsation (thiele et al. 2003 ) und medikamentöeser stabilisierung (nachlastsenkung mittels nitroprussidnatrium zur shunt-reduktion, ggf. in kombination mit katecholaminen) aussicht auf erfolg. aber selbst bei frühzeitiger operation muss mit einer sterblichkeit von 73% im vergleich zur sterblichkeit von 95% bei abwartender haltung gerechnet werden (hochman et al. 1997) . der perkutane translu-minale verschluss einer septumruptur gilt vorerst als experimentelle methode. die ruptur der freien ventrikelwand (figueras et al. 2001; slater et al. 2000) geht mit einer letalität von 90% einher. sie tritt am häufi gsten zwischen dem 2. und 8. tag nach infarkt auf, aber jede 3. ruptur fi ndet bereits innerhalb der ersten 24 h statt. sie betrifft häufi ger frauen, den erstinfarkt, den vorderwandinfarkt, den älteren patienten und diejenigen mit hochdruck. sie wird gehäuft gesehen bei vorausgegangener medikation mit kortikoiden und antiphlogistika. der frühzeitige einsatz von thrombolytika scheint die inzidenz zu senken, der späte einsatz sie zu erhöhen. die ruptur der freien wand führt zum hämoperikard und zur raschen perikardtamponade. oft kommt es zu neuerlichen starken herzschmerzen mit sofortiger elektromechanischer entkopplung. am häufi gsten führt der einseitige einriss zum tode; subakute einrisse können jedoch ebenfalls als akute tamponade, als großer perikarderguss oder als chronisches pseudoaneurysma imponieren. der akute schock nach infarkt muss immer an die diagnose einer ventrikelruptur denken lassen. eine sofortige perikardpunktion kann zur vorübergehenden stabilisierung führen, die zur zügigsten diagnostik (echokardiogramm: perikarderguss; swan-ganz-katheter: tamponadedruckprofi l) genutzt werden muss. sehr selten wird eine ventrikelruptur überlebt, nur dann, wenn der patient schnellstens (richtwert: 1 h) an die herz-lungen-maschine angeschlossen wird und der einriss erfolgreich übernäht werden kann. das geronnene blut im perikardbeutel verstopft sehr schnell eine perikarddrainage, was für die reanimation am offenen thorax spricht. zahlreiche erkrankungen sehr unterschiedlicher ätiologie können in einen kardiogenen oder extrakardial-obstruktiven schock münden (. tabelle 6.1). nach entsprechender diagnostik wird sich die therapie zunächst darauf konzentrieren, einerseits eine besserung der pumpfunktion des herzens zu erzielen, andererseits versuchen, kardiale und extrakardiale obstruktionen zu beseitigen (z. b. punktion eines tamponierenden perikardergusses, thrombolyse bei massiver lungenembolie) oder schockauslösende rhythmusstörungen zu beherrschen (z. b. kardiopulmonale reanimation). man wird dann bestrebt sein, die therapie in eine kausale münden zu lassen, soweit dies möglich ist. bei aktueller inoperabilität eines patienten mit dekompensierter aortenklappenstenose und schock kann auf der intensivstation bei sorgfältigem invasivem hämodynamischem monitoring eine nachlastsenkung mit nitroprussidnatrium zur entlastung des ventrikels versucht werden (khot et al. 2003) , und auch die aortenklappenvalvuloplastie kann im einzelfall zur rekompensation beitragen und damit die voraussetzungen für den späteren klappenersatz schaffen (melzer et al. 2001 ). > 6.6 · spezifi sche schockformen intoxikationen mit kardiodepressiven und vasotoxischen substanzen (schmidt et al. 1998) . trotz der potenziell akut lebensbedrohlichen situation ist die langzeitprognose dieser patienten nach erfolgreicher akutbehandlung in der regel sehr günstig. dies rechtfertigt auch den einsatz eines kardiopulmonalen bypasses für einige stunden, falls die konventionellen detoxikationsmaßnahmen und die symptomatische therapie zu keiner herz-kreislauf-stabilisierung führen. herzverletzungen (redling et al. 1998 ). offene, aber auch stumpfe thoraxtraumen mit schockentwicklung sollten immer an eine herzverletzung denken lassen; nach rascher diagnosestellung ist hier die sofortige herzoperation häufi g lebensrettend. perioperativer herzinfarkt mit schockentwicklung. wegen der häufi g nur kurz zurückliegenden operation muss bei auftreten eines herzinfarktes häufi g auf die i.v.-thrombolyse verzichtet werden, obwohl wegen des meist kurzen zeitintervalls zwischen infarktbeginn und infarktdiagnostik die erfolgschancen der thrombolyse günstig wären. in solchen fällen sollte -wann immer möglich -die akut-ptca auch des beatmeten patienten in erwägung gezogen werden. zwischen 1980 zwischen und 1987 f − − − f − − in deutschland werden z. z. jährlich 65.000 operationen mit der herz-lungen-maschine durchgeführt mit einer durchschnittlichen letalität von 3,3%. wesentliche todesursachen sind dabei septischer schock und mods. gründe für das erhöhte sepsisrisiko sind die kardiale vorschädigung der patienten, die folgen der thorakotomie mit einer erhöhten inzidenz an pneumonien sowie bakteriämie, endotoxinämie und vorwiegend pulmonalen mikrozirkulationsstörungen als folge der extrakorporalen zirkulation. ein weiterer wesentlicher aspekt ist die kontaktaktivierung des blutes in der herz-lungen-maschine: sie löst eine ausgeprägte systemische entzündungsreaktion (»post pump infl ammatory response«) aus. als folge davon kommt es zur aktivierung von blutzellen mit freisetzung von aggressiven mediatoren und proinfl ammatorischen zytokinen, die z. t. in höheren serumkonzentrationen gefunden werden als in der sepsis. diese entzündungsreaktion wird für das auftreten des mods nach herzchirurgischen operationen mitverantwortlich gemacht (prondzinsky et al. 2005 ). die möglichkeiten einer positiv-inotropen intervention sind bei dem muskelschwachen rechten ventrikel relativ begrenzt (maisch u. christ 2004) . insbesondere digitalis sollte vorsichtig dosiert werden, um intoxikationen zu vermeiden. die senkung der zur dekompensation führenden pulmonalen hypertonie steht im vordergrund. rechtsherzdekompensation bei ards und sepsis: 7 abschn. 6.6.2 in der therapie des rechtsherzversagens und der pulmonalen hypertonie nach operativen, insbesondere herzchirurgischen eingriffen, bewirkt inhalativ appliziertes stickoxid (no, 0,5-40 ppm) eine selektive pulmonale vasodilatation ohne systemvaskuläre nebenwirkungen rinne u. zwissler 2004) . die pulmonale vasodilatation geschieht ohne beeinträchtigung der hypoxischen pulmonalen vasokonstriktion, so dass es weder zu einer erhöhung des intrapulmonalen shunts noch zu einer reduktion der arteriellen oxygenierung kommt. die wirkung von inhalativ appliziertem no kann jedoch variieren. prinzipiell denkbare effekte auf koronarien und myokard scheinen nicht sehr ausgeprägt bis fehlend zu sein (cheifetz et al. 1996) . bedenken gegen den einsatz von inhalativem no bei akuten koronaren syndromen bestehen nicht (lindenfeld 1998) . zunehmend setzt sich zur intraoperativen anwendung neben inhalativem no das inhalativ applizierbare, stabile prostaglandin-i 2 (pgi 2 )-analogon iloprost (ilomedin) aufgrund seiner längeren hwz und seiner technisch einfacheren anwendbarkeit durch (rinne u. zwissler 2004 (bowers et al. 1998; jacobs et al. 2003; pfi sterer 2003; seyfarth u. schömig 2004) . hinweisend dafür sind st-strecken-hebungen in den rechtspräkordialen ableitungen v 3r und besonders v 4r . der rechtsherzinfarkt -charakterisiert durch einen schockzustand (hypotonie, niedriges herzzeitvolumen) ohne lungenstauung -lässt sich echokardiographisch als großer, hypokinetischer rechter ventrikel und kleiner linker ventrikel kenntlich machen. wegweisend für die diagnose sind neben klinik und echokardiographie auch der niedrige pulmonalkapillardruck trotz schockzustand. der rechtsherzinfarkt disponiert zu bradykarden rhythmusstörungen in form höhergradiger av-blockierungen. vorgehen der wahl bei rechtsherzinfarkt mit oder ohne begleitenden schockzustand ist die akut-ptca (bowers et al. 1998) : in einer untersuchung an 53 patienten mit rechtsventrikulärem infarkt konnte durch die akut-ptca bei 41 patienten (77%) eine wiedereröffnung der verschlossenen rechten koronararterie erzielt werden, mit einer prompten und beeindruckenden erholung der rechtsventrikulären pumpfunktion innerhalb einer stunde und einer niedrigen hypotonie-(12%) und letalitätsrate (2%). bei den patienten, bei denen die wiedereröffnung nicht gelang, persistierte die rechtsventrikuläre funktionsstörung; die hypotonierate lag bei 83% und die letalität bei 58% (bowers et al. 1998 trotz des gewachsenen verständnisses des krankheitsablaufs und neuer therapieansätze konnte die hohe letalität des septischen schocks von 40-60% bisher noch nicht wesentlich gesenkt werden. die sepsishäufi gkeit ist zunehmend, in den vereinigten staaten erkranken jährlich etwa 750.000 patienten an einer sepsis, und 200.000 versterben daran. wesentliches zur klassifi kation und ätiologie (. abb. 6.22, 7 abschn. 6.1.5), zur pathophysiologie (7 abschn. 6.2.3), zur basisdiagnostik (7 abschn. 6.3.2) und basistherapie (7 abschn. 6.4) und zum mods (7 abschn. 6.5). die typischen herz-kreislauf-veränderungen bei gramnegativem und grampositivem septischen schock sind: blutdruckabfall infolge der ausgeprägten vasodilatation, erniedrigung des systemischen gefäßwiderstandes bis auf 30% der norm, kompensatorische zunahme des herzzeitvolumens, des schlagvolumenindex und des schlagarbeitsindex. fehlt eine relative kardiale vorschädigung, so liegen die gemessenen herzfunktionsparameter dabei auch beträchtlich höher als die gesunder probanden mit einem normalen systemischen gefäßwiderstand um 1100 dyn · s · cm -5 (. abb. 6.23). die beiden toxinschocksyndrome werden durch das toxicshock-syndromtoxin 1 (tsst-1) von staphylococcus aureus bzw. durch das streptococcus-pyogenes-exotoxin (spe) von streptokokken produziert. beide toxine haben superantigeneigenschaften (7 abschn. 6.2.5), was den dramatischen klinischen verlauf über eine massive zytokinfreisetzung erklären könnte. bei verdacht auf eine der genannten erkrankungen müssen die erkrankten auf die intensivstation gebracht werden, wo schnellstmöglich mit der antibiotika-und der symptomatischen schocktherapie und engmaschigen überwachung begonnen werden muss, da sich sehr rasch ein schock entwickeln kann. die toxische vasodilatation mit massivem blutdruckabfall könnte ein gesundes herz durch einen drastischen anstieg des herzzeitvolumens kompensieren (. abb. 6.23). eine so weitgehende kompensation der erniedrigten nachlast, d. h. ein wesentlicher anstieg des herzzeitvolumens wie in . abb. 6.23 beschrieben, wird im septischen schock nur selten beobachtet, v. a. nicht bei protrahierten verläufen: die pumpfunktionsparameter des herzens sind dabei zwar im vergleich zu gesunden probanden mit normalem systemischem gefäßwi-> derstand meist nicht erniedrigt oder sogar leicht erhöht, berücksichtigt man jedoch die inverse korrelation mit dem systemischen gefäßwiderstand (. abb. 6.23), so wird die eingeschränkte pumpleistung des herzens bei vielen patienten bereits in der hyperdynamen phase des septischen schocks und sogar bereits bei septischen patienten mit noch normalem blutdruck (raper et al. 1989 ) rasch evident. die erklärung für die nur inadäquate steigerung der herzförderleistung ist eine potenziell reversible, multifaktorielle herzschädigung in der sepsis, die durch klinische befunde belegt werden kann. als charakteristika der akuten septischen kardiomyopathie (müllerwerdan et al. 1996; (redl et al. 1993) . insgesamt ist jedoch die diskussion um das »ideale« katecholaminregime bei primär rechtsventrikulärer septischer herzschädigung noch in vollem gange; die möglichkeit zur detaillierten quantifizierung des therapieerfolgs wird im einzelfall sicherlich hilfreich sein. der septische schock stellt ein bereits weit fortgeschrittenes krankheitsstadium dar! er ist in der regel hyperdynam (herzindex >5,5 l/min/m 2 ; systemischer gefäßwiderstand 600 dyn·s·cm -5 ) und nur in der spätphase oder bei nicht ausreichender volumensubstitution hypodynam (<2,5; 1200). die z. z. noch weitgehend symptomatische behandlung beinhaltet zunächst die rasche und adäquate volumensubstitution, ggf. gefolgt und unterstützt durch den einsatz von katecholaminen. die mikrobielle diagnostik (blutkulturen und organbezogener keimnachweis), die fokuselimination und der beginn einer antibiotischen therapie leiten bereits zu den kausalen maßnahmen über. sepsis und septischer schock können ein facettenreiches mods induzieren, das die prognose des patienten entscheidend prägt. das konzept der therapie des septischen schocks besitzt mehrere eckpfeiler (czock u. keller 2002; dellinger 2003; dellinger et al. 2004; freeman et al. 2001; werdan et al. 2005 bei der wahl des richtigen präparates zur kalkulierten und gezielten antiinfektiösen therapie sind die empfehlungen der paul-ehrlich-gesellschaft sehr hilfreich (bodmann u. vogel 2001) , wobei die zunehmende resistenzentwicklung ein ernsthaftes problem darstellt. mit einer innerhalb der ersten 6 h auf der notaufnahme begonnenen, an hämodynamischen zielkriterien orientierten volumen-und katecholamintherapie (. abb. 6.25) lässt sich bei patienten mit schwerer sepsis und septischem schock die 28-tage-sterblichkeit im vergleich zur »konventionellen intensivtherapie« von 49 auf 33% senken (rivers et al. 2001) . neu an diesem im vergleich zu früheren »sauerstoff-zielorientierten«, bisher erfolglos gewesenen ansätzen ist der sehr frühe beginn der behandlung. dennoch darf dieses »erfolgsrezept« nicht kritiklos von der notaufnahme auf die intensivstation übertragen werden: der noch nicht anbehandelte »notfallsepsispatient« unterscheidet sich hämodynamisch beträchtlich von dem klassischen »intensivstationsepsispatienten« (dan-nino et al. 2002; . einen die früh-und die stabile sepsisphase integrierenden vorschlag zur herz-kreislauf-therapie gibt . abb. 6.26. sepsispatienten sind durch ein absolutes und ein relatives intravasales volumendefi zit charakterisiert. die sofortige und adäquate flüssigkeitstherapie ist der entscheidende erste schritt zur behandlung des septischen schocks (. tabelle 6.20; müller-werdan u. . bei einem hämatokrit <30% und gleichzeitig einer auf <70% erniedrigten zentralvenösen sauerstoffsättigung (s cv o 2 ) wird die gabe von erythrozytenkonzentraten zur anhebung des hämatokrits auf ≥30% empfohlen (. abb. 6.25), zumindest in der frühphase der sepsis innerhalb der ersten 6 h auf der notaufnahme (rivers et al. 2001) . ansonsten gilt die empfehlung, erythrozytenkonzentrate bei einem hämoglobinwert <7,0 g/ dl zu geben und das hb auf einen wert von 7,0-9,0 g/dl anzuheben. eythropoetin wird nicht zur behandlung der anämie im rahmen einer schweren sepsis empfohlen (. tabelle 6.20; dellinger et al. 2004 ). berücksichtigt man weiterhin die auswirkungen der katecholaminbehandlung auf die leber-splanchnikus-perfusion, den magenmukosa-ph und den laktatspiegel, so schneiden dobutamin und noradrenalin hinsichtlich unerwünschter wirkungen günstiger als dopamin und wesentlich günstiger als adrenalin ab , so dass auch aus diesen gründen der einsatz von noradrenalin und dobutamin empfohlen werden kann. phosphodiesterasehemmstoffe und dopexamin sind im septischen schock eher pharmaka der 2. wahl. digitalis ist bei tachykardem vorhoffl immern und bei vorhoffl attern indiziert, bei sinusrhythmus ist seine bedeutung als positiv-inotrope substanz eher gering einzustufen. selten kommen vasopressin sowie -noch wesentlich seltener -angiotensin, vasodilatatoren, n-acetylcystein, glukagon und kalzium als positiv-inotrope und direkt sowie indirekt vasoaktiv wirkende pharmaka bei patienten mit septischem schock zum einsatz dellinger 2003; dellinger et al. 2004; . eine prognosebesserung ist nicht belegt. zu ernährung und stoffwechsel . tabelle 6.20 und 7 abschn. 6.5.5. die gabe von hydrocortison hat eingang in die offi ziellen therapieempfehlungen gefunden (. tabelle 6.20). hydrocortison bessert den vaskulär bedingten schock, mit einem anstieg des erniedrigten systemischen gefäßwiderstandes und einer verkürzung der behandlungsdauer mit vasopressorischen katecholaminen; die myokarddepression der septischen kardiomyopathie lässt sich durch hydrocortison allerdings nicht bessern: der erniedrigte linksventrikuläre schlagarbeitsindex steigt nicht an (briegel et al. 1999) . die hinweise auf eine letalitätssenkende wirkung (annane et al. 2002) werden z. z. in der corticus-studie überprüft (7 abschn. 6.5.11). auch die gabe von aktiviertem protein c (drotrecogin alfa aktiviert; xigris) ist bestandteil der offi ziellen therapieempfehlung bei schwerer sepsis in der frühphase (. tabelle 6.20; 7 abschn. 6.5.6). wie in einer cochrane-analyse beschrieben haben i.v.-immunglobuline -das iviggma-präparat pentaglobin -je 0,25 g/ kgkg an 3 aufeinanderfolgenden tagen -mehr noch als ivigg-präparate eine letalitätssenkung in mehreren kleinen studien gezeigt (septischer schock mit endotoxinämie, abdominelle sepsis; zitiert in werdan 2001b; werdan et al. 2005) ; die datenlage reichte der survival sepsis campaign jedoch nicht aus, um die gabe von iviggma in ihre therapieempfehlungen aufzunehmen (dellinger et al. 2004) . derzeit nicht zu empfehlen ist der einsatz von antiendotoxintherapien, von steroiden in hoher dosierung und prostaglandin e, von anti-tnf-α-antikörpern, interleukin-1-rezeptorantagonisten, paf-antagonisten, n-acetylcystein und antioxidanzien. die peripheren venen sind kollabiert und der jugularvenendruck ist niedrig, es sei denn, der hypovolämische schock ist assoziiert mit einer einfl ussstauung, z. b. durch ein myokardiales pumpversagen, eine perikardtamponade oder einen pneumothorax. die akren sind kühl, und v. a. bei älteren patienten kann es zu einem absinken der körpertemperatur kommen. die kapilläre füllung ist vermindert. weitere klinische zeichen des hypovolämischen schocks sind periphere zyanose, verminderter hautturgor und trockene schleimhäute. die patienten klagen über durst, schwitzen und kurzatmigkeit, sie sind ängstlich. im schweren schock werden die patienten zunehmend apathisch oder verwirrt. der blutdruck kann nichtinvasiv nicht mehr messbar sein, oder es kann ein großer gradient zwischen blutig und unblutig gemessenem blutdruck nachweisbar sein. die hypotension kann im sitzen oder orthostatisch verstärkt sein. die atemabhängigen schwankungen des systolischen arteriellen blutdrucks sind intensiviert. die typischerweise zu beobachtende tachykardie kann durch die vorausgegangene einnahme von β-rezeptorenblockern ausbleiben. gelegentlich kommt es zu einer bradykardie. der zur abschätzung des volumen-/blutverlustes berechenbare schockindex -quotient aus puls und systolischem blutdruck; normal 0,5 (blutverlust <10%); drohender schock 1 (blutverlust <20-30%); manifester schock 1,5 (blutverlust >30-50%) -ist keine große hilfe und kann zu fehleinschätzungen führen (große variabilität; interferenz mit vormedikation (β-blocker) und begleiterkrankungen (arterielle hypertonie). sind trauma und äußerliche blutung die ursache des schocks, so haben die kontrolle des blutverlustes, die bereitstellung von gekreuzten blutkonserven und die infusion von flüssigkeit und blutprodukten eine höhere priorität als weitergehende diagnostische maßnahmen (groneveld 2001). nach stumpfem abdominellem trauma sollte eine diagnostische peritoneallavage durchgeführt werden. eine computertomographie des abdomens gehört zur weiterführenden diagnostik; diese ist zwar zeitaufwendiger und weniger sensitiv als die peritoneallavage und nur bei relativ stabilen patienten durchführbar, sie ist aber diagnostisch spezifi scher. ein rupturiertes bauchaortenaneurysma kann sonographisch oder, falls der zustand des patienten es erlaubt, angiographisch erfasst werden. ein stumpfes thoraxtrauma kann kompliziert sein durch eine aortenruptur, einen spannungspneumothorax, hämatothorax, hämoperikard oder eine tamponade. eine thoraxröntgenaufnahme kann hier diagnostisch weiterführen. bei gastrointestinalen blutungen sollten weiterführende endoskopisch-diagnostische maßnahmen auch erst nach der initialen flüssigkeitstherapie erfolgen. eine magensonde sollte gelegt werden, um die aspiration von mageninhalt bei erbrechen zu verhindern und um eine magenblutung erkennen zu können. > > laboruntersuchungen umfassen neben der blutgruppenanalyse und bereitstellung gekreuzter blutkonserven die bestimmung des blutbildes, der elektrolyte, der laktatkonzentration, des serumkreatinins, der blutgase einschließlich des ph-werts und der glukose. unmittelbar nach einer blutung können der hämoglobingehalt und hämatokrit des blutes noch normal sein. ein akuter hypovolämischer schock kann mit einer leichten leukopenie und anschließenden leukozytose einhergehen. die überwachung der patienten erfordert neben einer kontrolle des arteriellen blutdrucks und der urinausscheidung ggf. die messung des zentralen venendrucks und ein hämodynamisches monitoring mittels pulmonalarterienkatheter zur überwachung der flüssigkeitstherapie. die messung des pulmonalkapillären verschlussdrucks ist besonders wichtig bei funktions-und compliance-störungen des linken ventrikels, etwa bei einer vorbestehenden herzerkrankung. umgekehrt kann bei schwerer pulmonaler hypertonie und rechtsherzinsuffi zienz der druck im linken herzen anhand des zentralen venendrucks unterschätzt werden. durch kontrolle der kardialen füllungsdrücke kann die entstehung eines lungenödems bei der flüssigkeitssubstitution vermieden werden. therapieziel ist neben der substitution des verlorengegangenen volumens die optimierung der o 2 -zufuhr an die peripheren gewebe (christ u. lackner 2004; groeneveld 2001) . der therapieplan beim hypovolämischen schock erfolgt nach den aufgeführten gesichtspunkten: lagerung und schmerzstillung, volumenersatz, kausaltherapie, zusätzliche (fakultative) maßnahmen, prophylaxe. aufgrund der mikrozirkulationsstörung und der sekundären volumenverluste sind oft weit höhere volumina erforderlich, um die makro-und mikroperfusion sicherzustellen, als aufgrund des initialen volumenverlustes bzw. schockereignisses zu erwarten wäre. seit jahrzehnten ist gegenstand der diskussion, ob die volumenersatztherapie mit kristalloiden oder kolloiden lösungen erfolgen soll (7 abschn. 6.4.6). in der regel werden bei der behandlung des hypovolämischen schocks kolloide und kristalloide in einem fi xen verhältnis (z. b. 1 teil kolloid: 2, evtl. 3 teile kristalloid) verabreicht. hyperton-onkotische lösungen zeigen beim hypovolämischen schock keine medizinischen vorteile (meier-hellman u. burgard 2004). vasoaktive pharmaka werden im hypovolämischen schock dann eingesetzt, wenn die therapieziele mit adäquater volumensubstitution nicht zu erreichen waren, oder als überbrückende maßnahme bis zur einleitung der volumentherapie. in abhängigkeit vom klinischen bild sind begleitende therapiemaßnahmen wie sedierung, analgesie und frühzeitige beatmung anzustreben. zur beherrschung schwerster hypovolämischer schockzustände kann der einsatz von antischockhosen (mast, »medical/military anti-shock trousers«) hilfreich sein. dabei werden durch getrennte pneumatische kammern unterschenkel, oberschenkel und abdomen mit zentripedal abfallenden drücken komprimiert. nach wie vor ist trauma die häufi gste todesursache bei personen im alter von 2-40 jahren. verkehrsunfälle sind die häufi gste ursache des stumpfen traumas. statistisch sind in der frühphase nach einem trauma verbluten oder schwere schädelverletzungen die häufi gsten todesursachen; einige tage nach dem ereignis kommt es zu einer häufung von todesfällen durch schwere schädel-hirn-traumen; im abstand von mehreren wochen zum trauma sind sepsis und multiorganversagen die häufi gsten todesursachen. die traumatische verletzung ist ein komplexes, multifaktorielles geschehen, das ein weites spektrum an verschiedenen autonomen endokrinen und zellulären reaktionen auslöst. schmerz, angst, blutverlust, gewebeverletzung, hypoxie und bakterielle kontamination wirken als neuroendokrine stimuli; zusätzliche einfl ussfaktoren sind der individuelle bewusstseinszustand und die gabe von medikamenten, v. a. anästhetika. meist handelt es sich beim traumatischen schock jedoch um die pathogenetische konstellation eines hypovolämischen schocks, der durch wiederholte blutverluste aggraviert verlaufen kann (shapiro 2001 ). beim hämorrhagischen schock fi nden sich die klinischen zeichen der hypovolämie (7 abschn. 6.6.3) und das bild der äußeren oder inneren blutung oder andere zeichen der gewalteinwirkung (shapiro 2001) . der traumatische schock ist nicht immer ein blutungsschock. viele weitere ursachen sind zu nennen und bestimmen das klinische bild, z. b. spinaler schock (hund u. abel 2002) , hirnödem, kardiale kontusion, herztamponade, spannungspneumothorax, hypothermie, flüssigkeitsverluste durch verbrennungen und crush-verletzungen mit einer massiven freisetzung von myoglobin. bei allen patienten, die ein trauma erlitten haben, ist nach zeichen des schocks zu fahnden. der genaue unfallhergang ist ebenso zu erfragen wie eventuelle internistische vorerkrankungen. gerade bei älteren patienten können auch scheinbar geringfügige gewalteinwirkungen zu schweren verletzungen führen, etwa aufgrund einer osteoporose. der vollständig entkleidete traumapatient sollte nach überprüfung der vitalfunktionen mit größter sorgfalt körperlich untersucht werden. zur weiterführenden diagnostik 7 abschn. 6.3.3. da das ausmaß der verletzungen oftmals nicht anatomisch genau feststellbar ist, kommen zur objektivierung des schweregrades des traumas score-systeme zur anwendung, häufi g wird der »revised trauma score« genutzt. parallel zur überprüfung der vitalfunktionen (atemwege, atmung, herz-kreislauf-funktion) sollte sofort eine rasche volumensubstitution erfolgen. bleibt der patient trotz volumengabe hypotensiv, so können zusätzlich erythrozytenkonzent-> rate transfundiert werden. die zentrale frage bei der weiteren behandlung des traumapatienten ist, ob eine sofortige operative therapie erforderlich ist. ein fehlendes ansprechen der schocksymptomatik auf die flüssigkeitsverabreichung ist verdächtig auf eine andauernde blutung. aktive externe blutungen können oft schon durch direkten druck unterbrochen werden. sind äußere blutungen kontrolliert, so muss nach versteckten blutungsursachen gefahndet und diese ggf. operativ angegangen werden. gerade in notaufnahmen muss darauf geachtet werden, dass die patienten im traumatischen schock nicht außerdem noch in eine hypothermie geraten, die den patienten zusätzlich gefährdet und die gerinnung beeinträchtigen kann. eine massive transfusion von erythrozytenkonzentraten kann zu einer schweren koagulopathie führen durch den prozentual zu geringen anteil an gerinnungsfaktoren im vergleich zu den blutzellen, durch hypothermie und durch die sekundäre gerinnungsstörung im schock. die koagulopathie wird durch gabe von ffp (»fresh frozen plasma«) und thrombozytenkonzentraten behandelt. eine klinische unterscheidung zwischen anaphylaktischer und anaphylaktoider reaktion gelingt nicht. das klinische erscheinungsbild (haupt 2001; walther u. böttiger 2004) variiert interindividuell stark, auch in abhängigkeit vom antigeneintrittsort, der absorptionsrate und dem ausmaß der sensibilisierung. initial können daher gastrointestinale symptome, übelkeit, erbrechen, durchfälle, kolikartige beschwerden, unwillkürlicher abgang von stuhl und harn, selten darmblutungen, hauterscheinungen oder beschwerden von seiten des respirationstraktes im vordergrund stehen. das zeitliche intervall bis zum auftreten von beschwerden kann minuten bis mehrere stunden betragen; ganz überwiegend treten die symptome innerhalb der ersten stunde nach antigenexposition auf. die sich meist rapide entwickelnde systemische reaktion geht sehr oft (mehr als 90%) einher mit hauterscheinungen wie pruritus, flush, erythem, urtikaria und in schweren fällen angioödem. häufi g sind juckreiz und schwellungen der nasen-, augen-und mundschleimhaut sowie ödeme der lippen, der augenlider und der zunge. häufi g und bedrohlich sind atemwegsobstruktionen, extrathorakal durch ödeme im larynx-und pharynxbereich, intrathorakal durch bronchialobstruktion. oft kommt es zum lungenödem. in schweren fällen einer anaphylaxie, etwa bei intravenöser antigenexposition, kann es ohne hauterscheinungen und atembeschwerden unmittelbar zum schock kommen. die hämodynamischen veränderungen des anaphylaktischen schocks sind aus kasuistischen beschreibungen bekannt (hanashiro u weil 1967) . im vordergrund stehen hypovolämie aufgrund von flüssigkeitsverschiebungen ins interstitium bei erhöhter gefäßpermeabilität und peripherer vasodilatation, tachykardie und erniedrigte kardiale füllungsdrücke (carlson et al. 1981; silverman et al. 1984) . initial im verlauf einer anaphylaxie wurden erhöhte herzzeitvolumi-! 6.6 · spezifi sche schockformen na beobachtet, die einerseits durch die erhöhten katecholamin-und histaminspiegel zustande kommen, andererseits die folge des erniedrigten systemischen gefäßwiderstands sein könnten (moss et al. 1981; hamberger et al. 1980) . im weiteren verlauf sinkt das herzzeitvolumen ab bei einem erniedrigten venösen rückstrom. niedrige pulmonalkapilläre verschlussdrücke, wie sie im anaphylaktischen schock gefunden werden, sprechen gegen eine kardiale verursachung des häufi g gefundenen lungenödems; vielmehr ist dies am ehesten die folge der erhöhten gefäßpermeabilität (carlson et al. 1981) . jedoch wurden im anaphylaktischen schock ekg-veränderungen gefunden im sinne von ischämiezeichen und arrhythmien (booth u. patterson 1970; sullivan 1982; austin et al. 1984) . auch über eine reversible myokarddepression wurde bereits berichtet (raper u. fisher 1988) , wenn auch ältere studien nur eine geringe beeinträchtigung der herzfunktion fanden. für die notfalltherapie (haupt 2001; waltheru. böttiger 2004) spielt die unterscheidung zwischen anaphylaktischer und anaphylaktoider reaktion keine rolle. im folgenden ist daher nur von der anaphylaxie als überbegriff die rede. bei verdacht auf eine anaphylaxie ist sofortiges handeln erforderlich (. tabelle 6.21), so dass eine rasche evaluation der situation unter berücksichtigung möglicher differenzialdiagnosen erfolgen muss (haupt 2001) . schwere anaphylaktische reaktionen können auch unter adäquater therapie progredient verlaufen, oder es kann nach einer vorübergehenden besserung zu einem erneuten einbruch der symptomatik kommen. daher ist oftmals eine intensivmedizinische überwachung dieser patienten angezeigt. grundpfeiler der sofortbehandlung bei hypotension und hypoxie sind: ausschalten des mutmaßlichen auslösers, offenhalten der atemwege, 100%ige o 2 -zufuhr, intravaskuläre volumenexpansion und katecholamine. differenzierte empfehlungen zur schweregradabhängigen (stadium 0-iv) akuttherapie anaphylaktoider reaktionen unter berücksichtigung der führenden klinischen symptomatik (kutan, pulmonal oder kardiovaskulär) wurden in einer interdisziplinären konsensuskonferenz erarbeitet (ahnefeld et al. 1994) . hinsichtlich der differenzialtherapie mit volumenersatz, katecholaminen, histaminantagonisten, glukokortikoiden und theophyllin wird auf die detaillierte abhandlung der konsensuskonferenz verwiesen. allgemeine therapiemaßnahmen und -prinzipien entfernung des auslösenden agens. das auslösende agens muss an der eintrittspforte entfernt (z. b. insektenstachel) oder die weitere systemische absorption vermindert (z. b. anlegen eines tourniquets bei eintrittspforte an einer extremität) bzw. die antigenzufuhr gestoppt werden. in bestimmten situationen (z. b. insektenstich) kann die subkutane injektion von adrenalin (0,1-0,2 mg) -möglichst in nähe der einstichstelle -sinnvoll sein. freie atemwege. freie atemwege müssen sichergestellt sein -schon ab stadium i (leichte allgemeinreaktion) o 2 -zufuhr über eine maske, bei bedrohlicher hypotension und/oder dyspnoe endotracheale intubation und 100%ige o 2 -beat-mung. ein larynxödem kann die intubation erschweren oder sogar unmöglich machen: in solchen fällen kann die koniotomie lebensrettend sein. entwickelt sich eine obstruktion der oberen atemwege, so wird eine intubation des patienten erforderlich, die dann meist schwierig ist. eine kontrollierte mechanische beatmung mit positivem endexspiratorischem druck wird häufi g notwendig, wenn sich hypoxie und lungenödem entwickelt haben. lagerung. die flachlagerung des patienten, möglichst trendelenburg-lagerung (ausnahme: lungenödem) wird empfohlen. volumen. schon ab stadium i (leichte allgemeinreaktion) sollte über einen zuverlässigen, möglichst großlumigen venösen zugang, möglichst rasch volumen (elektrolyt-und kolloidale lösungen) substituiert werden. die kausale therapie der relativen hypovolämie ist die adäquate volumensubstitution. schwere anaphylaktoide reaktionen erfordern nicht selten die zufuhr größerer flüssigkeitsmengen innerhalb kurzer zeit (2-3 l in 20-30 min). dies ist nur über einen großlumigen zugang möglich. auch nach primärer kreislaufstabilisierung können im verlauf der nächsten stunden infusionen von mehreren litern erforderlich werden. gelingt die zufuhr ausreichender volumina in kürzester zeit, sind häufi g keine weiteren therapeutischen maßnahmen erforderlich. dies gilt offenbar v. a. für anaphylaktoide reaktionen in der perioperativen phase, die sich primär oder ausschließlich am kardiovaskulären system manifestieren. bei kardial grenzwertig kompensierten patienten sollte die zufuhr großer volumina unter erhöhter vorsicht erfolgen, um eine akute kardiale dekompensation zu vermeiden. im stadium iii (bedrohliche allgemeinreaktion: anaphylaktischer schock) ist die alleinige gabe von elektrolytlösungen unzureichend. höhermolekulare lösungen sind zu bevorzugen. albumin bietet dabei gegenüber den künstlichen plasmaersatzmitteln keine vorteile. hydroxyäthylstärke (hes) mit einem mittleren molekulargewicht (hes 200.000) kann als volumenmittel der wahl zur soforttherapie anaphylaktoider reaktionen angesehen werden. begrenzt wird der einsatz durch die maximal zu verabreichende menge von etwa 20-30 ml/kgkg/tag (ca. 1,5 l beim erwachsenen). eine darüber hinaus erforderliche volumenzufuhr sollte bevorzugt mit elektrolytlösungen erfolgen. adrenalin. die pharmakologische behandlung der anaphylaxie beruht in erster linie auf dem einsatz von adrenalin, womit sowohl die hypotension als auch die bronchokonstriktion wirksam bekämpft werden kann. adrenalin kann intravenös, intramuskulär (sofortige selbsttherapie von patienten mit bekannter allergie nach allergenexposition im stadium ii -ausgeprägte allgemeinreaktion -mit kommerziell erhältlichen fertigspritzen, fastjekt), sublingual, endotracheal oder als dosieraerosol verabreicht werden. eine eindeutige indikation zur parenteralen verabreichung von adrenalin besteht im stadium iii (bedrohliche allgemeinreaktion: schock), jedoch kann der einsatz bei zunehmender hypotension trotz adäquater volumengabe schon im späten stadium ii erwogen werden. die intravenöse verabrei-tabelle 6.21. differenzialtherapie anaphylaktischer/anaphylaktoider reaktionen. (mod. nach ahnefeld et al. 1994; . kutane um eine ausreichend genaue dosierung zu erreichen, wird hierbei 1 mg (1 ml) adrenalin in einer 10-ml-spritze mit 9 ml nacl 0,9% aufgezogen. eine maximaldosis von 1 mg adrenalin (med nach dab) sollte in der regel nicht überschritten werden. steht kein intravenöser zugang zur verfügung, kann adrenalin endobronchial appliziert werden. in diesem fall sollte adrenalin etwa 2-bis 3-mal höher als bei intravenöser gabe dosiert werden (ca. 0,3 mg) und mit nacl 0,9% oder aqua bidest. auf ein volumen von etwa 5 ml verdünnt werden; evtl. erforderliche wiederholungsgaben sollten möglichst intravenös erfolgen. die wirkung von adrenalin hält bei endobronchialer gabe länger an als bei intravenöser verabreichung. eine pulmonale symptomatik (bronchospasmus) im stadium ii oder iii kann durch inhalative applikation von adrenalin oder -bei nichtverfügbarkeit -mit den zur asthmatherapie verwandten β 2 -sympathomimetika wirksam behandelt werden; die dosierung richtet sich nach den nebenwirkungen. die maximaldosis ist erreicht, wenn tachykardie und etwas später tremor auftreten. die therapie kardiovaskulärer reaktionen mittels adrenalininhalationen ist jedoch nicht gesichert. man sollte jedoch an diese möglichkeit denken, wenn keine parenteral applizierbaren katecholamine zur verfügung stehen. besondere aufmerksamkeit und vorsicht erfordert der einsatz von adrenalin bei patienten mit khk oder arrhythmien. in diesen fällen kann adrenalin zu einer akuten koronarinsuffi zienz bis hin zum myokardinfarkt bzw. kammerfl immern führen. andererseits muss gerade bei patienten mit khk der perfusionsdruck ausreichend hoch gehalten werden. dies gelingt im stadium iii häufi g nur durch gleichzeitige gabe von volumen und einem vasokonstriktor in ausreichender dosierung. stationäre aufnahme. alle patienten mit einer anaphylaktischen reaktion müssen stationär aufgenommen und kontinuierlich überwacht werden, auch dann, wenn die symptome rasch auf eine adäquate 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myocardial depression in normotensive sepsis the effects of coronary artery disease on cardiac function in nonhypotensive sepsis toll receptors and sepsis right ventricular function in early septic shock states open randomized phase ii trial of an extracorporeal endotoxin absorber in suspected gram-negative sepsis desensitization of rat cardiomyocyte adenyl cyclase stimulation by plasma of noradrenalinetreated patients with septic shock hämostasestörungen im umfeld von sepsis und sirs intraoperatives anästhesiologisches management bei patienten mit pulmonaler hypertonie early goal-directed therapy in the treatment of severe sepsis and septic shock allgemeine intensivtherapie early revascularization in cardiogenic shock -a positive view of a negative trial a comparison of albumin and saline for fl uid resuscitation in the intensive care unit help apheresis in the treatment of sepsis impact of thrombolysis, intra-aortic balloon pump counterpulsation, and their combination in cardiogenic shock complicating acute myocardial infarction: a report from the shock trial registry. should we emergently revascularize occluded coronaries for cardiogenic shock? warum sollte der intensivmediziner der autonomen dysfunktion seiner patienten beachtung schenken? lebensbedrohliche akute intoxikationen durch kardio-und vasotoxisch wirkende medikamente und drogen autonomic function in the icu patient pulmonary and left ventricular decompression by artifi cial pulmonary valve incompetence during percutaneous cardiopulmonary bypass support in cardiac arrest the ability of the simplifi ed acute physiology score (saps ii) to predict outcome in coronary care patients katecholamine im kardiogenen schock: hilfreich, nutzlos oder gefährlich critical care medicine -principles of diagnosis and management in the adult, 2 nd edn mortality in emergency department sepsis (meds) score: a prospectively derived and validated clinical prediction rule a review of randomized controlled trials using 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patients with severe sepsis and multiorgan dysfunction syndrome plasma exchange as rescue therapy in multiple organ failure ncluding acute renal failure treatment strategies for acute myocardial infarction complicated by cardiogenic shock in a community hospital enterale immunonutrition: wann, für wen, welche zukunftsperspektiven gibt es? the cardiovascular response of normal humans to the administration of endotoxin cardiac disorders in penicillin-induced anaphylaxis low-dose vasopressin in the treatment of septic shock in sheep myocardial ischemia-reperfusion injury: role of the peroxynitrite-poly(adp-ribose) polymerase pathway change in the ratio of interleukin-6 to interleukin-10 predicts a poor outcome in patients with systemic infl ammatory response syndrome task force of the american college of critical care medicine, society of critical care medicine (1999) practice parameters for hemodynamic support of sepsis in adult patients in sepsis outcome of patients sustaining acute 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out-of-hospital cardiopulmonary resuscitation hrsg) expertenforum: hämodynamisch aktive substanzen in der intensivmedizin mechanische und elektrische therapie kardialer arrhythmien assessment of ivig for prophylaxis and therapy of sepsis akute septische kardiomyopathie: bestandteil des multiorganversagens in der sepsis? plasma atrial natriuretic peptide and brain natriuretic peptide are increased in septic shock: impact of interleukin-6 and sepsis-associated left ventricular dysfunction cardioprotective effects of monophosphoryl lipid a, a novel endotoxin analogue in the dog epidemiology of anaphylaxis in olmsted county: a population -based study effectiveness of the glycoprotein iib/iiia antagonist abciximab during percutaneous coronary interventions (pci) in clinical practice at a single high-volume center metabolic encephalopathy in critically ill patients suffering from septic or nonseptic multiple organ failure right ventricuar infarction as an independent predictor of prognosis after acute inferior myocardial infarction tnf-a and il-1a inhibit both pyruvate dehydrogenase activity and mitochondrial function in cardiomyocytes: evidence for primary impairment of mitochondrial function prospective evaluation of early abciximab and primary percutaneous intervention for patients with st elevation myocardial infarction complicated by cardiogenic shock: results of the reo-shock trial predictors of in-hospital mortality in 1333 patients with acute myocardial infarction complicated by cardiogenic shock treated with primary percutaneous coronary intervention (pci) -results of the primary pci registry of the arbeitsgemeinschaft leitende kardiologische krankenhausärzte (alkk) study on glucocorticoid receptors during intestinal ischemia shock and septic shock diagnosis of papillary muscle rupture after acute myocardial infarction by transthoracic and transesophageal echocardiography enzyme-independent formation of nitric oxide in biological tissues transfundieren sie thrombozytenkonzentrate bei einem thrombozytenabfall auf <5000/mm 3 (5×10 9 /l), unabhängig vom vorliegen einer blutung, weiterhin bei thrombozytenzahlen von 5000-30.000/mm 3 (5-30×10 9 /l) und vorhandenem signifi kanten blutungsrisiko. höhere thrombozytenzahlen wie ≥50.000/mm 3 (50×10 9 /l) sind für chirurgische und invasive prozeduren erforderlich (empfehlungsgrad e) key: cord-015024-2xzc0uc5 authors: nan title: esicm 2010 wednesday sessions 13 october 2010 date: 2010-08-31 journal: intensive care med doi: 10.1007/s00134-010-2001-7 sha: doc_id: 15024 cord_uid: 2xzc0uc5 nan power spectrums for vt and eadi are shown in fig. 1 (ps and nava) for a typical patient. the enlarged section highlights how changes in eadi are highly synchronized with nava ventilation, but less so for ps. table 1 ) and complications of mechanical ventilation ( table 2 ) did not differ significantly between the two studied groups. introduction. high tidal volumes in mechanically ventilated patients with ards lead to baro/bio-trauma and increase mortality. also, it was recently shown that ventilation with high tidal volumes is a risk factor for ''acquired ards'' in a medical population. objective. we evaluated the impact of high tidal volumes after cardiac surgery. method. we analysed the prospectively recorded data of 3,434 consecutive patients who underwent cardiac surgery from 2002 to 2005. we predefined 3 groups of patients based on the tidal volume delivered immediately after surgery: (1) low: 7-9.9, (2) ''traditional'': 10-12.9, (3) high: above 13 ml/kg of predicted body weight (pbw). we assessed the risk factors for organ dysfunction (prolonged mechanical ventilation, hypoxemia, hemodynamic failure and renal failure) by univariate and multivariate analysis, including the initial tidal volume in the models. mean tidal volume/actual weight and tidal volume/pbw was 9.2 ± 1.3 and 11.1 ± 1.5 in men (p \ 0.0001), 9.1 ± 1.4 and 12.5 ± 2.2 in women (p \ 0.0001). 411 patients (12%) were ventilated with low tidal volumes, 2,194 (63.9%) with ''traditional'' tv and 829 (24.1%) with high tv. the mean body mass index in the 3 groups was 23.8 ± 4.0, 27.0 ± 4.1 and 31.5 ± 5.4 respectively (p \ 0.0001). with increasing bmi, the tidal volume/ actual weight decreased while the tidal volume/pbw increased (figure) . the percentage of women was 12.2, 20.8 and 52.2% respectively for low, ''traditional'' and high tv (p \ 0.0001). high tidal volumes were associated with prolonged intubation ([48 h) (1.5 vs. conclusion. traditional and very high tidal volumes are associated with prolonged mechanical ventilation and organ dysfunction after cardiac surgery and use of high tidal volumes is an independent risk factor. ''prophylactic'' protective ventilatory strategy should be provided in this population with inflammatory state at risk to develop ventilator induced pulmonary edema. women and patients with high bmi are more at risk to be ventilated with injurious tidal volumes. introduction. evidence shows that clinicians' non-technical skills (behavioural and cognitive skills) have a significant impact on teamworking, patient safety, efficiency of care provided and potentially patient outcomes (1) . such skills are key for cardiac arrest teams (cats), which are multi-professional (anaesthetists, physicians and nurses) and normally function under high pressure. to date, most tools to assess nontechnical skills in healthcare have focused on surgery (2) and anaesthesia (3) . no validated, robust tools are currently available for assessing non-technical skills in cats. objectives. to develop and validate an observational skill-based clinical assessment tool for resuscitation (oscar). this should be psychometrically robust for use in both training and assessment contexts. methods. oscar was based on a well-validated tool for surgery (otas) (4) and was developed in phases. six behaviours were included in the assessment: communication, cooperation, coordination, monitoring, leadership and decision-making. observable behavioural exemplars were derived for each one of these behaviours across the three cat subteams-anaesthetists, physicians and nurses (phase 1). quantitative expert consensus methodology was employed to assess content and face validity and observability of the exemplars (phase 2). two clinician observers used oscar to blindly rate eight cats performance in a series of simulated cardiac arrests. psychometric analyses of these ratings were used to determine observable behaviour applicability, internal consistency, and inter-rater reliability (phase 3). . 15 of 18 oscar behaviours demonstrated high internal consistency (cronbach a = 0.736-0.970). psychometric analyses dictated removal of three behavioural exemplars (two in anaesthetic group; one in physician group) to significantly improve internal consistency. inter-rater reliability was also high (inter-observer pearson r = 0.661-0.911, all p \ 0.005). inter-observer reliability analyses revealed a learning curve between the two observers, with significant reduction in scoring discrepancies from the first to the eighth observed resuscitations. conclusions. oscar is a psychometrically robust (reliable, content-and face-valid) tool for the assessment of teamworking skills in cardiac arrest events. the tool is feasible to use and can be employed for both training and assessment purposes. introduction. different educational methodologies are used to teach basic skills in emergency medicine. high-fidelity patient simulation offers an ideal venue for presentation of critical events that can be managed by medical students without risk to a patient. therefore full scale simulation training could be superior to paper case based seminary rounds to achieve these specific educational objectives. objectives. the aim was to compare simulation to a standard education measured by multiple choice questionnaire. after written informed consent and approval of the institutional research ethics board 160 fifth year medical students were included in the survey. they took part in the compulsory emergency medicine curriculum of charité universitätsmedizin berlin. the students completed a basic multiple question tests on day 1 including 25 questions concerning the topic of ''acute coronary syndrome'' (acs). on day 2 for the topic ''acs'' half the group was assigned a 45 min session simulation training while half the group was assigned a 45 min session paper case training. on day 3 groups were reversed and the topic ''aic'' was taught in either simulation training or paper case seminary round. the test of day 1 was repeated after each training sessions. results of the tests were evaluated using spss(tm) 18. the mann whitney u test was used to show any significant differences in reaching educational objectives in the test (a \ 0.05 was considered significant). there was an even distribution of men and women among the two groups. the test results showed no significant difference between the two groups on day 1. on day two for the topic ''acs'' the group with simulation training achieved significantly better test results. for the topic ''acs'' on day 3 there was no difference while students received further training in acs not using a high fidelity simulator. the results were not linked to specific teachers. introduction. rapid sequence induction (rsi) involves loss of spontaneous breathing and mandates airway control. steps to reduce adverse incidents include adherence to minimum monitoring standards, appropriate drug selection, access to difficult airway equipment and presence of skilled anaesthetists. there is substantial evidence that appropriate monitoring reduces risk by detecting the consequences of errors, and by giving early warning of patient deterioration. objectives. to assess conduct of emergency anaesthesia (monitoring and drugs) for critically ill patients not in an operating theatre (or) administered by intensive care doctors. methods. prospective analysis of rsi for critically ill patients in a uk nhs acute hospital over 1 month. or based practice was excluded. reason for anaesthesia, location, drugs administered, monitoring modalities, adverse events and access to airway equipment were recorded. results. data from 26 patient episodes were collected: predominantly in the emergency department (38%) and intensive care unit (27%) for respiratory failure (46%), reduced consciousness (19%) and to facilitate investigations (19%) . the most common induction agent was propofol (52%); thiopentone (26%) and etomidate (11%) were less frequently used. suxamethonium (73%) was preferred for initial neuromuscular blockade. during induction most doctors used pulse oximetry, electrocardiography and blood pressure monitoring. only 24% used capnography. no doctor used minimum monitoring to association of anaesthetists of great britain and ireland (aagbi) standards. 1 rescue airway equipment immediately available is shown in fig. 1 . complications occurred in 12 cases (fig. 2) . patients that had a hypotensive episode during induction all had thiopentone or propofol used as induction agents. 15% of patients had a period of desaturation, and 23% required more than one attempt for successful intubation. in cases with complications, rescue airway equipment was unavailable in[60 and 75% did not achieve uk minimum monitoring standards. conclusions. shortcomings during emergency anaesthesia were recorded including monitoring, access to rescue airways and physiological disturbance. procedural guidelines and training are to be developed for emergency anaesthesia; access to capnography and alternative airway equipment will be assured. these issues are unlikely to be unique to our trust and assessment of practice is recommended. introduction. critical care echocardiography (cce) is performed and interpreted by the intensivist at the bedside to establish diagnoses and guide the management of patients with circulatory or respiratory failure in the icu. competence in basic and advanced cce has been recently defined [1] , but no curriculum to reach the required cognitive and technical skills has yet been elaborated. objectives. to assess the efficacy of a limited, tailored training program for noncardiologist residents without experience in ultrasound to reach competence in basic cce. methods. six noncardiologist residents (anaesthesiology: n = 5, pneumology: n = 1) without previous experience in ultrasound participated to the study during two 3-month periods. the curriculum consisted in 4 h of didactics, 2 h of interactive clinical cases and 6 h of tutored hands-on. color doppler mapping was excluded from the training. after completion of the training program, all eligible patients underwent subsequently a transthoracic echocardiography (tte) performed in random order by a recently trained resident and an experienced intensivist with expertise in cce who was used as a reference. in each patient, the resident and the experienced intensivist answered binary ''rule in, rule out'' clinical questions covered by basic cce [1] : global left ventricular (lv) size and systolic function (eye-ball evaluation of ejection fraction), homogeneous or heterogeneous lv contraction pattern, global right ventricular (rv) size and systolic function, identification of pericardial fluid and tamponade, and assessment of both the size and respiratory variations of the ivc. in case of undetermined interpretation, the corresponding clinical question was considered not addressed. the agreement between responses to clinical questions provided by the two investigators who independently interpreted the tte study at bedside was used as an indicator of effectiveness of the tested curriculum. proportion of graduates to work within a ''critical care'' setting. the level of support available to trainees may vary with local resources but risk management and national guidelines stipulate that close supervision is provided to junior doctors in high stake decisions and procedures until deemed competent at the relevant tasks 1 . furthermore, substantial ongoing reduction in working hours places further limitations on training; both majors can impact adversely on junior doctors service output and experience. a modified delphi method was used 2 years ago to design a task focused single-day course on the theoretical basis of critical care and provide lab-based training in delphi identified high risk procedures and interventions 2 . objectives. assess the impact of the course on the following: trainee confidence and the start of the ''novice'' critical care post trainee performance in comparison to peers perceived educational benefit from their training post compared to peers methods. 30 junior doctors attending the course were enrolled in the study and matched for graduation year and medical school to 30 junior doctors who did not attend similar training prior to commencing their post. data was collected through anonymous standardized forms on the day of the course, first day of the job, end of week 1, week 6 and 4 months into the post. trainee confidence and self perceived competence were assessed on a ten point scale. in addition, trainees were requested to maintain a log of 3 interventions: 1-ultrasound guided central venous catheter insertion, 2-arterial catheter insertion, 3-ventilation problem solving. candidates attending the course demonstrated greater confidence at multiple points within their post as well as higher performance, satisfaction and educational value scores. conclusions. critical care trainees benefit from a task focused orientation to the fundamentals of critical care before commencing first post in this setting. . 100 enrolled 50 patients in each group. no differences in age and gender. incidence of vap-study group 8.8% compared to control group 28.8% p value 0.004. vap per 1,000 hospital days: control-29.5% compared to study-10.2% p value 0.06; average days in icu control-14.97 compared to study-11.8, p value 0.151; average ventilated days, control-12.68 compared to 8.9, p value 0.044; average antibiotic use in days control-9.7 compared to study-7.2, p value 0.413. introduction. nosocomial infections are the most common in-hospital complications with high morbidity and mortality. educating healthcare professionals is an important prevention measure. objective. to analyze the impact of a nurse consultant team on nosocomial infections prevention in the icu, the improvement in prevention knowledge of the nurse staff, and its impact in the application of the prevention measures in the daily practice. methodology: the nurse referent team was constituted by 8 nurses. the study subjects were all the staff icu nurses and all the patients admitted during pre and post-intervention phases. the study was conducted in our medical-surgical icu (16 beds) in 3 phases: 1 pre-interventional (15/01/09-15/02/09) observational. record of the accomplishment of 12 cdc recommended variables about mechanical ventilation associated pneumonia (vap) and catheter related bloodstream infection (cr-bsi) prevention measures. 2 interventional (1/03/09-1/10/09) eight educational meetings with the nurses staff groups to teach the most important aspects of the nosocomial infections prevention. before and after lectures every nurses answered an anonymous questionnaire about their knowledge in those subjects. a poster with the most important reminders was place in every icu patient room. 3 post-interventional (15/01/10-15/02/10) observational. new record of the same 12 cdc pre-interventional variables. we compared the accomplishment of these variables before and after the interventional phase as well as the number of correct questionnaire answers. statistics were made with spss software. results. during the interventional phase 70% of the staff nurses attended the educational meetings. the number of correct answers increased significantly after the conference (77.1 vs. 55.2% p \ 0.0001). regarding to the daily practice, we observed a significant increase in the accomplishment in most of the variables (see table below), while in 3 of them no improve was observed and in 2 the improvement was not statistically significant. during the study period we observed a decrease in the incidence of vap (6.7-14.9 episodes/1,000 mv days) and cr-bsi (2. introduction. glucose variability has been found to be associated with mortality in critically ill patients, independent of mean glucose concentration [1] . objectives. the aim of this analysis was to assess the impact of real time continuous glucose monitoring (cgm) on glucose variability in critically ill patients receiving intensive insulin therapy (iit). methods. this is the post-hoc analysis of a prospective, randomized, controlled trial [2] . data of 124 patients admitted to the icu either receiving iit according to a real time cgm system (guardian ò , medtronic, northridge, ca, usa) (n = 63) or according to an algorithm (n = 61) with selective arterial blood glucose measurements (simultaneously blinded cgm) for 72 h were analysed. insulin infusion rates were guided according to the same algorithm in both groups. mean glucose and standard deviation, as a marker of glucose variability, were calculated for the first 24 h (glumean1, glusd1) and for the whole study period (glu sd ). statistical comparison of parameters between study groups and between icu survivors (n = 94) and non-survivors (n = 30) was performed using student's t test. results. the variability of sensor glucose during the entire study period was comparable between the real time cgm group and controls (21.51 ± 1.10 vs. 23 introduction. in the gastrointestinal tract, the gut flora which comprises several hundred grams of bacteria is crucially involved in host homeostasis through their metabolic, trophic, and protective activities. however, the immediate changes in the gut flora in critical illness following severe insults are unknown. objectives. to investigate the changes in the gut flora at an early phase of severe insult in critically ill patients. methods. fifteen patients who experienced a sudden and severe insult including trauma, out-of hospital cardiac arrest, and cerebral vascular disease were studied, along with 12 healthy volunteers as the control group. two fecal samples were acquired from the subjects by swabs of the rectum within 6 h after admission to the emergency room (day 0). samples were serially collected from patients on day 1, 3, 5, 7, 10, and 14. samples were collected from control subjects. results. total bacterial counts, especially various obligate anaerobes and total lactobacillus, significantly decreased in comparison to those of the control subjects on day 0. in addition, on day 0, the total organic acid levels of the patients were significantly lower than those of the control subjects; particularly acetic acid, propionic acid, and butyric acid. the levels of these acids remained low throughout the 14 days period of study. the total bacterial counts did not recover to normal levels during the 14 day study period. obligate anaerobe counts of the patients did not improve until day 14. total lactobacillus counts were low on day 0 and increased gradually thereafter, but did not attain the levels found in controls. the counts of pathogens (enterococcus and pseudomonas) increased during the study period. conclusions. gut flora in critically ill patients can change drastically immediately after a severe insult, and may not recover for up to 14 days. at the same time, the number of harmful bacteria can increase. total bacteria 10.1 (9.8-10.4) 7.5 (6.7-8.0) .001 obligate anaerobes clostridium cocades group 9.4 (9.0-9.7) 6.1 (5.4-7.0) .001 clostridium leptam subgroup 9.2 (9.0-9.7) 6.5 (6.1-7.3) .001 bacteroides fragilis group 9.4 (9.0-9.8) 6 .8 (5.9-7.6) .001 bifidobacterium 9.2 (8.9-9.4) 6.0 (\5.0-6.9) .001 atopobium cluster 9.0 (8.5-9.5) 6.0 (\5.1-7.1) .001 results. mean serum 25(oh)d level was 18.9 ± 10.9 ng/ml. by current definitions the majority of patients (62.5%) were vitamin d deficient (\20 ng/ml) and 24.9% were vitamin d insufficient (c20 and \30 ng/dl). normal 25(oh)d levels ([30 ng/ml) were present in 12.6%. table 1 provides information on clinical and laboratory findings in the three 25(oh)d groups. both lower 25(oh)d tertiles were associated with increased hospital mortality after adjustment for age, sex and saps ii. for 330 patients both 25(oh)d and pth levels were available. adjusting the cox regression analysis also for pth and dialysis status increased the hr for hospital mortality to 2.3 (1.1-4.7) and 2.1 (0.99 and 4.4) for the two lower 25(oh)d tertiles. in addition tertiles of pth and serum calcium levels suggested higher mortality rates for patients in the highest pth (p = 0.09) and those in the lowest calcium tertile (p = 0.12). our results demonstrate that independent of baseline saps ii, age and sex, critically ill patients with low 25(oh)d levels seem to be at increased risk for hospital mortality. whether a rapid correction of vitamin d status may be beneficial in the icu setting remains to be further explored in randomized controlled trials. • the autonomic storm after brain death must be early diagnosed and treated with a standardized protocol including hormone therapy introduction. the use of filling pressures of the right atrium and left atrium is normal in the monitoring of critically ill patients undergoing mechanical ventilation. this monitoring is done through an invasive catheter placed in the superior vena cava and pulmonary artery, which is not free of complications. the ability to make measurements of these parameters in a non invasive way, makes the echocardiography an useful and essential tool when monitoring critically ill patients objectives. we focus the study on validate the reliability of noninvasive measurements by echocardiography and invasive measurement catheters of filling pressures methods. we conducted a prospective observational study relating the filling pressures, between central venous pressure (cvp) with the diameter of the inferior vena cava and left atrial pressures with the values of the ratio e/e 0 . the filling pressure variables were only discriminated as high or low. low values were accepted when invasive measurement of cvp was \6 and \4 mmhg in the lap; and by echocardiography when the diameter of the ivc was\12 mm and the ratio e/e 0 \8. high values were accepted when the measurement of cvp was higher than 10 and 8 mmhg in lap and in echocardiography when the diameter of the ivc [16 mm and the ratio e/e 0 [15. we collected data from 38 patients in the immediate postoperative period, under mechanical ventilation (vt 8-9 ml/kg, fio 2 50%, peep 3), sinus rhythm, good cardiac function and without postoperative drug support. all of them had a central venous line and right atrium catheter as habitual monitoring of postoperative cardiac patients. we performed an echocardiography when the patient presented hypotension, with low values of cvp and lap, and we repeated the measurements after the infusion of the habitual fluid protocol (500 ml hes 6% in 30-45 min). the data we record were: diameter of ivc and ratio e/e 0 by echo and cvp and lap values by invasive catheters. 23rd esicm annual congress -barcelona, spain -9-13 october 2010 s333 introduction. an attenuated cardio-hemodynamic response to dobutamine is associated with a poor outcome in established human sepsis [1, 2] . establishing a sensitive method to identify early cardiac dysfunction in both experimental and human sepsis would be a useful tool to explore timesensitive mechanisms further. objectives. to assess myocardial responsiveness to dobutamine in early sepsis. methods. all procedures were in accordance with uk home office laboratory animal legislation. under isoflurane anaesthesia, male adult wistar rats underwent left common carotid and right internal jugular venous cannulation for blood sampling/continuous bp monitoring and fluid administration respectively. rats received either 1.8 ml caecal slurry (sepsis; n = 8) or 1.8 ml saline (sham; n = 8) ip, before fluid resuscitation (0.9% saline 10 ml/kg/h) and conscious monitoring was commenced. after 4 h, rats were re-anaesthetized with isoflurane and transthoracic echocardiography was performed. stroke volume was optimised with saline boluses prior to an incremental dobutamine infusion (1.25-20 mcg/kg/ min). data are presented as mean (sd); analyzed with 2-way anova and post-hoc tukey test. results. figure 1 summarizes hemodynamic changes after sepsis, fluid resuscitation and dobutamine infusion. baseline parameters were similar after echocardiography-guided fluid resuscitation, with contractility and stroke volume restored in septic rats to sham values. septic rats demonstrated an enhanced chronotropic response to dobutamine compared to sham (p \ 0.002). both peak velocity and cardiac output were attenuated by c25% in sepsis (p \ 0.0001). in sepsis, baseline map was higher but neither sham nor septic maps were affected by dobutamine infusion. conclusions. dobutamine stress echocardiography is a sensitive, reproducible, dynamic physiological probe that reveals early cardiac dysfunction in septic rats with apparently similar baseline cardiovascular physiology. introduction. the evaluation of right ventricular (rv) function is clinically useful in patients with acute respiratory distress syndrome (ards) because the presence of rv failure has large prognosis implications. the purpose of the current study was to compare right ventricular myocardial strain imaging parameters with conventional echocardiographic indices evaluating right ventricular function during ards. objectives. we hypothesized that peak systolic strain would be more sensitive than conventional echocardiographic parameters in detecting subclinical right ventricular systolic dysfunction in patients with ards. methods. in total, 10 patients with ards and with normal right ventricle function assessed by two dimensional echocardiography and 10 age matched subjects under mechanical ventilation without heart or pulmonary disease were included in the present study. conventional echocardiography parameters for rv function assessment like rv fractional area change (rvfa) or the tricuspid annular plane systolic excursion (tapse) were measured and compared to tissue doppler imaging parameters with strain value obtained from the right ventricle free wall. . strain values were reduced in the rv free wall of the patients with ards compared with the control group (23.45% ± 3.8 vs. 31.8% ± 2.8 p = 0.005) moreover no significant difference was observed in conventional two dimensional parameters evaluating rv systolic function between these two groups of patients. in patients with ards a significant relationship was shown between peak systolic strain at basal free wall and arterial carbon dioxide tension (rho = -0.79 p = 0.017) and with the end inspiratory pressure (rho = -0.67 p = 0.04). conclusions. during the ards, doppler tissue imaging parameters can determine rv dysfunction that is complementary to conventional echocardiographic indices and is correlated with respiratory parameters. on doppler tissue imaging, patients with ards exhibit abnormal rv systolic function even in patients with normal rv function assessed with conventional echocardiographic parameters. objectives. studying the effect of olv on rv outflow impedance during inspiration and expiration using transesophageal echo-doppler in a trial to differentiate the rv consequence of increasing lung volume from those secondary to increasing airway pressure during mechanical ventilation. methods. thirty stable patients on mechanical ventilation because of different causes were enrolled prospectively in this single center, cross sectional clinical study. each patient was firstly subjected to conventional ventilation (cv) with volume controlled ventilation, followed by open lung concept (olc) ventilation by switching to pressure controlled mode, then recruitment maneuver applied until pao2/fio2 [375 torr. hemodynamic (mean arterial pressure ''map'', central venous pressure ''cvp'' and heart rate ''hr'') and respiratory (total and intrinsic peep, peak, plateau and mean airway pressure and total and dynamic lung compliance) measurements were recorded before, 20 min after a steady state of cv and 20 min after a steady state of olc ventilation. also, transesophageal echo doppler was performed at end of inspiration and end of expiration to calculate the mean acceleration (ac mean ), as a marker of the rv outflow impedance, 20 min after a steady state of cv and 20 min after a steady state of olc ventilation. results. during inspiration, ac mean was significantly lower during cv compared to olc ventilation (p value .001). inspiration didn't cause a significant decrease in acmean compared with expiration during olv (p value.001) but did do so during cv. in comparison to baseline and cv, olc ventilation was associated with a statistically significant higher cvp (p value .001 for both), higher total quasi-static lung compliance (p value .001 for both) and dynamic lung compliance (p value 0.001 for both). moreover, pao 2 /fio 2 ratio of olv was significantly higher than in baseline and cv (p value .001 for both). conclusions. olc ventilation does not change rv afterload during inspiration and expiration as rv afterload appears primarily mediated through the tidal volume. moreover, olc ventilation provide a more stable hemodynamic condition and better oxygenation and lung dynamics. introduction. among indices provided by the analysis of aortic blood flow through esophageal doppler, mean acceleration (acc) is supposed to reflect the left ventricular (lv) systolic function, but this has been poorly validated. in particular, acc could be influenced by loading conditions of the lv. objectives. to test whether acc actually behaves as an indicator of lv systolic function by testing if 1. it increased with inotropic stimulation, 2. it was not altered by fluid loading, 3 . it correlated with the echographic lv ejection fraction (lvef) and it reliably tracked the changes in lvef during therapeutic intervention. in 37 patients with cute circulatory failure (sapsii 59 ± 18, age 64 ± 10 years, 26 receiving norepinephrine), we administered either a volume expansion (500 ml saline over 20 min in 24 patients) or dobutamine (5 lg/kg/min in 13 patients). we simultaneously measured acc (cardioq, deltex medical) and lvef at baseline and after therapeutic intervention. results. volume expansion significantly altered neither lvef (from 56 ± 12 to 58 ± 12%) nor acc (from 9.5 ± 3.2 to 9.9 ± 3.1 cm/s 2 ) while dobutamine infusion significantly increased lvef by 27 ± 15% and acc by 44 ± 27%. considering the 74 acc/lvef pairs of measurements, an acc \8.2 cm/s 2 predicted a lvef b 45% with a sensitivity of 77% (95% ci [56-91%]) and a specificity of 68% (95% ci [52-80%]). the changes in lvef and in acc during fluid and dobutamine administration were significantly correlated (r = 0.64, p \ 0.05). conclusions. acc fulfilled the criteria required from a clinical indicator of lv global systolic function. a given value of acc allowed detecting a low lvef with a modest accuracy. by contrast, the treatment-induced relative changes in acc were reliable for tracking the treatment-induced relative changes in lvef. objectives. to compare the relationship between systolic or diastolic dysfunction at icu admission and the incidence of cardiologic complications and mortality at sixth months. methods. prospective study of forty consecutive patients diagnosed of acute myocardial infarction (ami) (23 nstemi, 17 stemi) who were admitted in the icu of university hospital puerto real (cadiz, spain) from 1st may 2009 to 30th september 2009. studied variables: age, gender, type of ami (nstemi, stemi), left ventricular ejection fraction (lvef) by biplanar simpson's rule, diastolic function (ratio e/e 0 of the mitral annulus included), incidence of cardiac complications (acute pulmonary oedema, atrial fibrillation with hemodynamic instability and cardiogenic shock) and mortality at sixth month. echocardiographic studies were performed with a ge vivid 7 pro(r) by an intensivist who had performed up to 500 doppler studies in critical patients. all studies were remeasured by a second observer in an echocardiographic workstation with no statistical difference in measured velocities. patients were classified according to their lvef in (a) preserved ([55%), (b) mildly depressed (45-55%), (c) moderately depressed (30-45%) and (d) severely depressed (\30%); and according to their e/e 0 ratio in (a) normal e/e 0 ratio (\10) and (b) elevated e/e 0 ratio (c 10). the results were statistically analysed with chi-square test and odds ratio calculus. results. diastolic dysfunction measured with e/e 0 ratio was associated with high incidence of cardiac complications (chi 2 test cl 95% p \ 0.001, or 24). systolic dysfunction measured by lvef was also associated with more complications but with less strength of statistical association (chi 2 test cl 95% p \ 0.05, or 8. 25 ). there were no significative statistical difference between lvef and e/e 0 ratio in mortality at sixth month. conclusions. in our study, diastolic and systolic dysfunctions in patients with ami at icu admission were associated with high incidence of cardiac complications, with more strength of statistical association in patients with diastolic dysfunction. the small sample volume didn't allow us obtaining significative statistical differences in mortality at sixth months. a new method has been developed to assess global end-diastolic volume (gedv) and extravascular lung water (evlw) from a transpulmonary thermodilution curve. our goal was to compare this new method to the established method currently in clinical use, over a wide range up to extreme pathophysiological conditions. objectives and methods. 11 anesthetized and mechanically ventilated pigs (90-110 kg) were instrumented with a central venous catheter and a right (5f pulsiocath, pulsion, munich, germany) and a left (5f volumeview, edwards lifesciences, irvine, ca) thermodilution femoral arterial catheter. the right femoral catheter was connected to a picco2 monitor (pulsion) and used to measure cop, gedvp and evlwp using the old method based on the equation: gedv = cop 9 (mtt -dst). the left femoral catheter was connected to the new ev1000 monitor (edwards) and used to measure coe, gedve and evlwe using the new method based on the equation: gedve = f (s2/s1) 9 coe 9 mtt, where s1 and s2 are respectively the maximum up-and down-slopes of the dilution curve, respectively. 137 measurements were done during inotropic stimulation (dobu), during hemmorhage (hypo), during fluid overload (hyper), and after inducing oleic acid-acute lung injury (ali). overall, cop and coe ranged from 3.1 to 15.4 and from 3.4 to 15.1 l/min, respectively. cop and coe were closely correlated (r 2 = 0.99), mean bias (± sd) was 0.18 ± 0.29 l/min and %error was 7%. gedvp and gedve ranged from 701 to 1,629 and from 774 to 1,645 ml. gedvp and gedve were closely correlated (r 2 = 0.79), mean bias was -11 ± 78 ml and %error was 14%. evlwp and evlwe ranged from 507 to 2,379 and from 495 to 2,222 ml. evlwp and evlwe were closely correlated (r 2 = 0.97), mean bias was -5 ± 72 ml and %error was 15%. parameters over the study period are presented in the table (*p \ 0.05 intervention vs. base or hyper). introduction. fluid resuscitation is a major therapy in icu. various mechanisms are involved in the regulation of the microcirculation and the macrocirculation. objectives. the goal of this study is to assess the sublingual microcirculatory changes in response to fluid challenge in preload-responsive and non preload-responsive patients. after approval by our local institutional review board, 18 patients in surgical icu have been included in an observational study. each patient was monitored by an arterial catheter and an oesophageal doppler. the decision of fluid infusion was taken by the physician in charge of the patient. preload-responsive patients were defined by variations in cardiac index (ci) c 15%. sublingual microcirculation videos were obtained using the orthogonal polarized spectral (ops) imaging technology. functional capillary density (fcd, cm cm -2 ) and microcirculatory flow index (mfi) were collected. the macrocirculatory and microcirculatory measurements were obtained before, during and after the infusion of 500 ml of saline. five sublingual sites were recorded before and after the fluid resuscitation. the ventilator settings and sedative and vasoactive drugs infusion rates were kept constant throughout the procedure. results. patients were admitted in icu for acute brain trauma (n = 5), hemorrhagic shock (n = 4), septic shock (n = 4), acute brain hemorrhage (n = 3) and acute pancreatitis (n = 2). the average age of the patient was 44 ± 17. the mean values of ci and mean arterial pressure (map) before the fluid therapy were respectively 2.7 ± 1.2 l/min/m 2 and 80 ± 15 mmhg. nine patients responded to fluid infusion (ci c 15%.). about the microcirculation, there was no significant difference between responders (r) and non-responders (nr) concerning the variations of mfi (0. introduction. passive leg raising (plr) was shown to discriminate hemodynamically unstable patients who will benefit from subsequent fluid administration or not. 1 concerned by the possibility of harmful hypotension starting the plr maneuver from a 45°semirecumbent position, in a previous study, we found that raising patients' legs from a supine position, we were not able to predict fluid responsiveness in a heterogeneous cohort of medical intensive care unit (icu) patients. 2 objectives. to investigate whether starting plr maneuver from a 45°semirecumbent position would better predict volume responsiveness without harmful hypotension in spontaneously breathing critically ill medical icu patients. methods. fluid responsiveness was tested in 27 consecutive patients (14 sepsis, 4 respiratory failure, 5 heart failure, 4 others) with a mean arterial pressure (map) \60 mmhg and/or a cardiac index (ci) \2.4 l/min/m 2 . heart rate (hr), mean arterial pressure (map), global end-diastolic volume index (gedvi), cardiac index (ci) and stroke volume index (svi) were recorded using the picco method. patients were stable in a semirecumbent (45°) position when first measurements were taken (baseline 1). for the plr maneuver, patient's bed was tilt to have the lower limbs raised to a 45°angle while the patient's trunk was then in a supine position. changes after 2 min were recorded. the patient was then brought into a supine position, and heamodynamic measurements were recorded when stable (baseline 2). thereafter, 500 ml of 0.9% nacl were administered over 15 min. positive predictive values (ppv) and negative predictive values (npv) of the plr maneuver were calculated using a cut-off value of 15 % increase for ci and svi and 10 % increase for map. results. patients' median age was 60 (29-82) years and their saps score 59 (16-90). all patients received vasopressors and/or inotropes. baseline hemodynamics and changes after plr and fluid challenge are shown in table 1 . results are given as median (range); n/a = not available, *p \ 0.05 versus baseline. ppv and npv for ci were 50 and 86%, for svi 20 and 77% and for map 12 and 80%, respectively. conclusions. in our hands, plr was not useful identifying fluid responders in this heterogenous population of severely ill medical icu patients, the starting semirecumbent position being associated with a potentially harmful decrease in map. however, it was helpful to detect patients who will not benefit (or even suffer harm) from further fluid administration. recently, some studies suggested that an impaired diastolic function is a predictive factor of mortality in patient with shock. it is not already known whether fluid infusion could improve diastolic function. objectives. the aim of the study was to determine the impact of rapid fluid infusion on diastolic function. after acceptance by the local ethic committee, 94 icu patients were prospectively included. volume expansion (ve) by 500 ml of saline was performed by the intensivist in charge. transthoracic doppler echocardiography was performed before and after fluid infusion. stroke volume (sv), early diastolic transmitral velocity (e), early diastolic mitral annular velocity (ea) and e/ea ratio (reflect of lv filling pressure) were studied. patients were divided in 2 groups according to their sv' increase: responders (r) (those who increased their sv by at least 15 %) and non-responders (nr). wilcoxon rank sum test was performed to compare data before and after ve. data are presented in median (iqr) results. fifty-three (56 %) patients were r and 41 (44%) were nr. in the overall population, ea increased significantly with ve [from 12.8 (4.5) to 13.9 (5.3) cm/s, p = 0.002]. in the r group ea increased significantly [from 12.6 (4.1) to 14.6 (4.9) cm/s, p = 0.01] and e/ea did not change significantly [from 6.1 (2.1) to 6.3 (2. 3), p = 0.4]. however in the nr group, ea did not change significantly [from 12 (5) to 12.7 (6.6) cm/s, p = 0.47] while e/ea increased significantly [from 6.7 (2.8) to 8.5 (3.9) cm/s, p = 0.008]. conclusions. according to these results, adequate fluid infusion seemed to enhance lv relaxation without increasing lv filling pressure while inadequate fluid infusion did not affect relaxation but increased lv filling pressure. objectives. the aim of our study is to compare the rapid variation of co measured by vigileo-flotrac ò with doppler-echocardiography which is considered as a reference method. during the 36 first hours of hospitalisation, we studied 16 mechanically ventilated patients receiving norepinephrine who underwent arterial pressure monitoring via a radial artery catheter. the flotrac ò pressure sensor and the vigileo ò monitor were connected to the arterial line. at each fluid expansion or norepinephrine dose modification a transthoracic doppler-echocardiography was performed and co was calculated. variations for co measured by each method were compared. results are presented as median (iqr). linear regression and the bland-altman method were used for statistical analysis. methods. for the in vitro experiments blood of 20 healthy donors was incubated (in the ratio 5:1) with one of the following solutions: ringer solution, ringer-lactate solution, modified gelatin (gelofusin); hydroxyethyl starch (hes) 130/0.42. after incubation, the following parameters of erythrocyte aggregation were measured: t1 and t2-characteristic times of spontaneous erythrocyte aggregation; b-hydrodynamic strength of aggregates; i 2.5 -index of strength of the largest aggregates at shear rate 2.5 s -1 . rbc deformability at various shear stresses was determined by ektacytometry. in vivo study on 60 patients with trauma treated randomly with either only crystalloids (group 1; n = 20), or crystalloids + hes 130/0.42 (group 2; n = 20) or crystalloids + gelofusin (group 3; n = 20) over 7 days, the same parameters as in vitro study were determined at day 1-7. twenty healthy men and women were included as controls. for statistical analysis the statistical package spss version 15.0 was used. statistical significance was considered at p \ 0.05. in vitro study in the final analysis effects of different colloids on rbc aggregation and deformability were considered as increasing impact (:), decreasing impact (;) and no impact (-) ( table 1) . in vivo study significant microrheological disturbances were detected at day 1 after admission. deformability index was lower in patients compared with controls (0.39 ± 0.012 vs. 0.44 ± 0.016; p = 0.017). simultaneously, the patients showed erythrocytes hyperaggregation compared with control (;t1, ;t2; :i 2.5 , :b). in the first group (crystalloids) described violations persisted throughout the study time. in group 2 (crystalloids + hes), the deformability was higher than in the 1st group, from 3 days till the end of the study, attaining the normal range, and also higher than in the third group (crystalloids + gelofusin). in the third group, deformability index was not significantly different from group 1. according aggregatometrical data in the first group hyperaggregation syndrome remained the entire period of observation. hes adding (group 2) decelerated aggregate formation (:t1, :t2; ;i 2.5 ). in contrast, modified gelatin adding enhanced erythrocyte aggregation (;t1 :i 2.5 , :b). conclusions. crystalloid solutions are not able to improve microrheological parameters. hes 130/0.42 increases rbc's deformability and reduced rbc's aggregability. gelofusin increases erythrocyte aggregation and no effect on deformability. introduction. trauma patients often require norepinephrine (ne) infusion and fluid challenge to keep normal blood pressure values. the reliability of dynamic predictors of fluid responsiveness during vasopressors therapy is under debate. we investigated the impact of norepinephrine (ne) infusion changes on pulse pressure variation (ppv) assessed with the mostcare system (vytech health, laboratoires pharmaceutiques vygon, ecouen, france) in intensive care unit patients. this device is a pulse contour method that provides cardiac output and fluid responsiveness variables and does not need any kind of calibration or preloaded data. methods. 15 trauma patients (8 female, 7 male, mean age 53 ± 22) admitted to a 13-bed university hospital medico-surgical icu were prospectively enrolled. inclusion criteria were: mechanically ventilated patients (tidal volume [8 ml/kg and constant respiratory rate); invasive arterial blood pressure monitoring; ne infusion. ppv values were recorded continuously during three different haemodynamic states: at baseline (t1), 10 min after a 0.05 lg/kg/min ne increase (t2), 10 min after a further 0.05 lg/kg/min ne increase (t3), 10 min following the reduction of ne to t2 dosage (t2 0 ) and 10 min after setting ne to baseline value (t1 0 ). during the study neither fluid challenge nor other vasoactive/inotropic drug changes were done. anova test was applied. results. see data in table 1 . at t1 ne mean dosage was 0.12 lg/kg/min (range 0.05-0.2 lg/kg/min). the mean ppv was: at t1 10.69 ± 6.9%, at t2 8.78 ± 5.9%, at t3 7.77 ± 6.1%, at t2 0 8.73 ± 5.5%, at t1 0 12.07 ± 8.2% (p \ 0.05). conclusions. our findings demonstrated that ppv was significantly affected by changes in ne: the higher the ne dosage the lower the ppv. changes in arterial tone due to ne infusion can impair ppv reliability in assessing fluid responsiveness in trauma patients. introduction. in mechanically ventilated patients respiratory variation in the arterial pulse pressure (dpp) is a reliable predictor of fluid responsiveness 1 . respiratory variation of pulse oximetry plethysmographic waveforms correlate to dpp 2 and can be calculated automatically in real time (heart-lung index [hli ò ] from hamilton medical). this prospective study evaluates the relationship between dpp and hli ò to predict fluid responsiveness. mechanically ventilated patients were investigated; all connected to an hamilton g5 ventilator and ventilated in adaptive support ventilation (asv), paralyzed and none had severe cardiac dysrhythmia. 15 were eligible for fluid expansion. dpp, hli ò (obtained from a finger probe pulse oxymeter integrated to the ventilator) and cardiac index (ci from transthoracic echo-doppler), were obtained before and after fluid expansion (8 ml/kg of hea over 20 min). ci-responders were defined by 15% increase from baseline. results. 10 out of the 15 patients were ci-responders and had significantly higher hli ò before volume expansion (21% ± 6 vs. 9% ± 4, p \ 0.02). before fluid expansion hli ò was correlated with dpp (r 2 = 0.68, p \ 0.01, fig. 1 ). hli and dpp were significantly correlated with change in ic induced by fluid expansion (r 2 = 0.53 and r 2 = 0.73, respectively). objectives. the primary end point of this study was to evaluate the rvd of the ivc in icu patients with spontaneous breathing. methods. icu patient with spontaneous breathing and signs of hypoperfusion (oliguria, mottles, serum lactate level [2 mmol/l) were eligible after the approval of the local ethics committee. we excluded patients with acute heart failure with pulmonary edema, moribund and arrhythmic patients. the trans thoracic echocardiographic (tte) evaluation was done by 4 confirmed intensivists (level [2 in echocardiography). the aortic diameter measured at the lv outflow chamber and the tvi were measured. the vena cava inferior diameters at inspiration and at expiration were measured on the sub costal view. the rvd of the ivc was defined as the (maximal ivc diameter -minimal ivc diameter)/maximal ivc diameter. these measures were realized at t0, before fluid challenge, and after a fluid challenge of 500 ml of hes 6% (130.9/1) over 30 min (t30). patients with an increase of tvi of more than 15% were considered as responders to the fluid challenge. the measures of tvi and of the rvd of the ivc were validated by an experimented intensivist and echographist (level 3) after blinding the patient' name and of the times of measurement. roc curves were constructed, and the cut off was determined as the closest point of the roc curve to the ideal point (sensibility = specificity = 1). the values are expressed as median and extremes. objectives. our objective was to test whether non invasive assessment by trans thoracic echocardiography of sub aortic velocity time index (vti) variation after a low volume of fluid infusion (100 ml of hydroxy ethyl starch, hes) can predict fluid responsiveness. methods. sub aortic vti was measured by transthoracic echocardiography before fluid infusion (baseline) in 39 sedated patients with acute circulatory failure and low tidal volume mechanical ventilation in whom volume expansion was planned. then, vti was recorded after 100 ml of fluid infusion over 1 min, and after an additional infusion of 400 ml of hes over 1 min. we measured the variation of vti after 100 ml of fluid (dvti 100 ) for each patient. receiver operating characteristic (roc) curves were generated for dvti 100 in all patients. when available, roc curves were also generated for pulse pressure variation (ppv) and central venous pressure (cvp). , volumes (gedvi) and variabilities (svv, ppv) have been suggested to predict volume responsiveness (vr). the final classification of a patient as ''volume responsive'' is usually made by a volume challenge (vc) with an infusion of a pre-defined amount of fluid over a certain time. among many variations of vcs, the infusion of 7 ml crystalloid over 30 min is one of the most established. despite superior predictive capabilities of svv, ppv and gedi compared to cvp and pawp in a number of studies, they fail to predict vr in a substantial number of patients. furthermore, the use of these parameters is limited due to femoral access of the cvc (gedi; cvp) or the absence of controlled ventilation and/or sinus rhythm (svv, ppv). repeated ''exploratory'' vcs with 7 ml/kg might result in volume overload in some patients. objectives. therefore, we investigated the usefulness of a ''small vc'' with 3.5 ml/kg crystalloid over 15 min compared to a standard vc with 7 ml/kg over 30 min. in 48 patients equipped with picco hemodynamic monitoring we performed a 30 min vc with 7 ml/kg of crystalloid. during the vc transpulmonary thermodilution (td) was performed at 0, 15 and 30 min to obtain td-derived ci (ci td ). additionally pulse contour ci (ci pc ) was recorded in intervals of 5 min. introduction. the prevalence of obesity, defined as a body mass index (bmi) c30 kg/ m 2 , reaches epidemic proportions. it is not only a risk factor for health problems, but also exacerbates illness progression. consequently, the number of obese patients on the intensive care unit (icu) has increased enormously. caring for obese patients can be quite challenging due to the weight and size of this person. the extent of and specific problems associated to the care of obese icu patients are unknown. the aim of this study is to identify and quantify problems nurses face in caring for obese patients on the icu. this study was performed on the icu at the radboud university nijmegen medical centre and contained two parts. in the first part a selection was made of 44 obese patients admitted between 2004 and 2009; these patients were matched with 44 normal weight patients (bmi 18.5-24.9 kg/m 2 ). patients were matched on gender, age, length of icu stay and apache-ii score. all patient files were screened for the presence and intensity of problems in caring for these patients. in the second part 25 nurses were asked in a survey to share their experiences in caring for obese patients. they were asked about the nature, frequency and intensity of the problems they faced. in total, 94 problems were identified in the 88 screened patient files. seventy-two problems (76.6%) occurred in care for obese patients and 23 (23.4%) in care for normal weight patients. in both groups, most of the problems were related to activities of daily living (adl) such as (re)positioning in bed, transfers and personal care. surprisingly, the intensity of the problems was similar in both groups. most of the problems were moderate (hardly to solve by one person) or severe (only to solve with two persons or special equipment). moderate problems occurred in 32.3% of normal weight patients and in 33.3% of obese patients; severe problems 29.0 and 31.9%, respectively. this result was also confirmed by the survey. the nurses qualified most of the problems they were asked about as moderate or severe, and the frequency of the experienced problems was much higher. from the files it appeared that in 31.8% of the obese patients nurses had adl problems. strikingly, in the survey nurses reported that they frequently (72.0%) or even always (28.0%) experienced adl problems in obese patients. nurses reported and experienced more problems in daily care for obese icu patients compared to normal weight icu patients. although the intensity of the problems with obese patients did not differ from normal weight patients, the frequency in which they occur was much higher. differences between reported problems and the survey suggest an underestimation of problems that can be solved by performing a prospective study. nevertheless, based on these results, and taking into account that obesity will increase in the future, we recommend anticipating to the needs of the nurses whenever possible. introduction. worldwide the number of obese patients (bmi [ 40) is increasing rapidly (1); this also includes patients admitted to the intensive care units (icu). this raises special demands on the staff, the surroundings and the equipment (2) . often the obese patient is not mobilised according to the clinical standard this causes complications to breathing, circulation and skin etc. furthermore the length of stay in the icu increases and the mortality rises. objectives. the aim of this study therefore was to make clinical guidelines and recommendations for mobilisation of the obese icu patient based on evidence. this will increase the knowledge and importance of mobilisation between staff and on longer term improves the daily average number of mobilisations performed with these patients. a secondary aim is that increased knowledge on this topic will improve the interdisciplinary work between the different professions based on the same overall aim. a systematic review of the literature concerning mobilisation of the obese icu patients was made in the year 2009-2010. the study is still work in progress analysing the literature to make guidelines and recommendations based on evidence. furthermore evidencebased education of special trained staff in mobilisation has been conducted in january/ february to improve their knowledge of the impacts mobilisation has on the respiration, circulation and skincare etc. the education was planned to aiming at a interdisciplinary audience. results. the preliminary results shows that it is more difficult to care for and mobilise the obese icu patient, because there is lack of space, non-availability of the correct equipment, too few available staff members and a significant negative attitudes among the staff towards the obese patient. recommendations are made within airway, breathing, circulation, nutrition, pain, equipment and patient experience according to the procedure of mobilisation of the obese icu patient. the recommendation was implemented in the already performed education and resulted in a changed attitude among the participant and improved the status of mobilisation in the daily prioritization. this knowledge was obtained in the evaluationinterview conducted approximately one month after the seminar. conclusions. according to the literature mobilisation of the obese icu patient needs special attention towards a safe clinical practise based on evidence with focus on both the patient and the staff. special attention towards this group of patient is created by performing evidence based research resulting in clinical guidelines that has to be implemented through theoretical and practical education on an interdisciplinary level. nurses are constantly exposed to the pain and suffering of those in their care 3 . the primary aim of this study was to investigate the risk of secondary traumatic stress/compassion fatigue (sts/cf-the trauma suffered by the helping professional) and burnout (bo-emotional exhaustion, depersonalization, and reduced sense of personal accomplishment), and the potential for compassion satisfaction (cs-the fulfillment from helping others and positive collegial relationships) among nurses working in icu. an additional goal was to test the relationship of these three constructs to each other. (1) . the use of closed suction circuits has been suggested beneficial as a prophylactic measure (2) . objectives. the aim of this study was to compare the incidence of vap and the occurrence of desaturation during suction using either oss or css. we also investigated contamination of the closed suction circuit and the occurrence of adverse events. methods. css were a new product in our clinic. all staff underwent a user course supervised by the manufacturer of the closed circuit. after this, data were collected during four periods in 2009, 2 month css followed by 2 months oss which was repeated twice. during the summer period css were used without any data collection and then followed by two periods of css and oss. all mechanical ventilated patients were consecutively included. a culture of deep endotracheal aspirate and a blind microbiology brush was taken in association with the intubation, after 72 h and every monday. after changing css and in case of extubation, the tip of the catheter was sent for culture. demographic data were retrieved from the hospital database. data were analyzed with descriptive methods. results. the incidences of vap were higher in the css group (table 1) . both suction systems showed almost no desaturation during and after suctioning. positive cultures were obtained in 55% of all the retrieved css catheters. the microbiological flora resembled the species found in the airway cultures. there were no inter patient contamination and neither did the bronchoscopy frequency differ between oss and css patients. in the css group six adverse events were seen; three tube occlusion and three incidences with secretion clogging. conclusions. the use of a css did not prevent vap, in our study. there were no benefit with css other than maybe to protect the staff and our finding of positive culture in 55% of the cases is in line with earlier studies. objectives. the aim of this study was to determine which intensive care patients the nurses defined as 'difficult' and their experiences in coping with such patients. the study was carried out as a qualitative design with 18 voluntary nurses employed in five intensive care units of a research and training hospital. the data were collected using demographic characteristics form and a semi-structured interview form. interviews with nurses were made individually and face to face. the data were evaluated by using colaizzi's phenomenological data analysis method. as a result of data analysis into two categories and two themes were identified. the categories were (1) difficult patient definition of the nurses, (2) the effect of difficult patients on their care, and (3) how the nurses are affected and cope with difficult patients. the nurses listed their reasons for defining some persons as difficult as difficult physical care of the patients, and the difficulty in communicating due to dementia, agitation, alzheimer's disease or the patient's personal characteristics. the nurses said that they found taking care of patients they found difficult physically and psychologically demanding. they used methods such as finding out the patient's problem and taking appropriate measures, increasing communication with the patient and providing explanations, trying to obtain spiritual satisfaction and transferring the patient's care to another nurse when communication problems were impossible to overcome. intensive care nurses have difficulty in caring for and communicating with some intensive care patients due to the characteristics of the disease, physical/psychological factors and personal characteristics. we found that nurses continued the care of these ''difficult'' patients by focusing on solving their problems, transferring the care to another nurse when necessary or by trying to obtain spiritual satisfaction. methods. teams of three delirium experts visited ten icu's in the the netherlands in which the cam-icu was incorporated in daily practice, twice. these teams consisted of two consultants in either psychiatry, clinical geriatrics or neurology, and either a research-physician (mmjve) or a research-nurse (mvdb). based on cognitive testing, inspection of the files and dsm-iv criteria for delirium, the teams classified patients as awake and not delirious, or delirious or comatose. this classification served as gold standard to which the cam-icu as performed by the bed-side nurses was compared. a simple 2 9 2 table was used to calculate the sensitivity and specificity. results. 15 delirium experts performed 333 assessments. 181 (54%) of these patients were assessable for delirium, 126 (38%) patients were excluded because the level of consciousness was too low, and 27 (8%) patients were non-assessable due to other reasons. overall, we found a sensitivity of 47% (95% ci 35-58%) and a specificity of 98% (95% ci 92-99%). the strengths of this study include the large numbers, the multicentre design, the extensive evaluations by teams of various delirium experts and the independent assessments of delirium experts and bed-side nurses. a limitations is the time interval between the expert assessment and the administration of the cam-icu (mean 125 min; standard deviation 118 min). there were striking differences in implementation strategies of the cam-icu between the centres. tables 1, 2 . 23rd esicm annual congress -barcelona, spain -9-13 october 2010 s341 introduction. presence of expiratory ineffective efforts in mechanically ventilated patients is a common problem associated with increased duration of mechanical ventilation, length of stay and also a higher cost and mortality. nowadays, identification and categorization of expiratory asynchronies can only be done at the bedside with the continuous observation of the ventilator interface. nurses must be skilled to understand non appropriate situations of anomalous patient-ventilator interactions. objectives. we tested the hypothesis that after specific training nurses would acquire enough skills to detect expiratory efforts as intensive care expert physicians would do. training phase: nurses were provided with selected bibliography on patient ventilator interaction and afterwards trained by intensivists with expertise on mechanical ventilation (2 h/day during 20 days) on airway pressure, flow and volume waveforms identification and eye interpretation of early and late ineffective expiratory efforts during expiration. validation phase: airflow and airway pressure waveforms were obtained from 8 different icu mechanically ventilated patients using and acquisition and processing biomedical signal software (better care ò ). one thousand and seven breaths were randomly selected from a total of 2,608,800 breaths. subsequently, selected breaths were blindly analyzed by 2 trained nurses and 3 intensivists to identify ineffective expiratory efforts. introduction. several publications indicate that manual hyperinflation is a widely used measure in the icu, but more important is the fact that there is no uniformity in the implementation of this measure. this is also on my ward. in literature there are a number of reasons given to start manual hyperinflation: abolish mucus retension, improve oxygenation and removal of atelectasis. the positive effects are improved compliance, improved oxygenation and a decrease in the number of vap's (ventilator associated pneumonia). the negative effects are a decrease in cardiac output due to high peak pressures, an increased risk of baro-/volutrauma and the risk of giving too much tidal volumes. the risk of barotrauma increase with pressures above 40 cmh 2 o. other side effects include the development of a pneumothorax and increased icp (intra cranial pressure). objectives. creating more awareness of the procedure with lower peak pressures as a result. methods. through literature review, clinical courses and the introduction of a pressure gauge achieve greater uniformity and awareness of the procedure. we used a flow analyzer of imt medical, a laptop with flowlab software version 4.1.1 and an artificial lung to demonstrate how much pressure and volume is generated during manual hyperinflation. conclusions. compliance with bts guidelines could be improved. unsurprisingly co-morbidities were frequent, but did not seem to affect outcome. use of a pneumonia severity assessment tool was sub-optimal, however mean curb-65 score didn't correlate with that recommended to prompt critical care assessment. apart from functional status, we are unable currently to identify any factors in this age group which can be used to guide critical care admission decision making. conclusions. in our study the incidence of complicated pneumonia was 316/10,000 patients admitted in picu. in necrotizing pneumonias the blood cultures were more positive than in non-necrotizing patients. although the surgical approach in necrotizing pneumonia is controversial, it resulted in a insignificantly lower mortality rate, comparing with non-necrotizing pneumonias. background. community-acquired pneumonia (cap) of mixed etiology has frequently been described in the literature, but its clinical significance remains unknown. the aim of this study was to describe the prevalence, clinical characteristics, and outcome of severe cap of mixed etiology in icu patients. a 5-year prospective study was conducted on consecutive patients with severe cap admitted to icu in whom an extensive microbiological investigation was performed. results. 316 patients were included. a single pathogen was detected in 128 (40.5%) cases, while two or more pathogens in 38 (12.0%) cases. the most frequent pathogens' combinations were those of two bacteria (28.9%) and bacterium plus virus (21.1%). compared with patients with monomicrobial pneumonia, patients with mixed pneumonia were older, had higher severity score (psi) and were more likely to have previous chronic pulmonary disease (see table 1 below). moreover, mixed cap patients showed similar clinical and analytical data at admission but increases in the frequency of respiratory distress and in length of stay and a trend to higher orotracheal intubation and mortality rates. a mixed etiology was detected in 12% of cases with cap requiring icu hospitalization and was associated with older age and increased severity. despite similar radiological features (n of involved lobes, pleural effusion) at admission, cap with mixed etiology showed a trend to worse clinical course and outcomes than monomicrobial pneumonia. objectives. to assess the incidence and aetiology of pneumonia in a mixed medicalsurgical icu, in order to develop local epidemiologically guided protocols to reduce antibiotic resistance selection in patients with pneumonia. methods. retrospective observational study on prospectively collected data in a mixed medical-surgical icu of a secondary care italian hospital. at our institution, epidemiological data on infections and data on antibiotic use are recorded since 1992; in 2005 a new electronic recording of icu infections was introduced. type of infection, germ characteristics, clinician diagnosis and antibiotic use were prospectively collected in an electronic database and retrospectively reviewed. antibiotic exposure index was calculated as each antibiotic total amount administered divided by its defined daily dose times total days of admission. between 2005 and 2009 a total of 690 patients were admitted to our icu. pneumonia was the commonest infectious disease at admission (148 cases, 21% of patients), and the commonest infectious complication during icu stay (38 new occurrences, 21% of total pneumonia patients). table 1 shows major epidemiological findings in the study population. the incidence of acquired pneumonia was remarkable: 9.7 cases every 1,000 days of mechanical ventilation. the most frequent isolated organisms were s. aureus (18 patients) and p. aeruginosa (12 patients). methicillin-resistant s. aureus (mrsa) accounted for 61% of pneumonia caused by s. aureus, and its prevalence matched closely the exposure index to vancomycin. such a high incidence of mrsa is consistent with other records in mediterranean countries. carbapenem-resistant p. aeruginosa was somewhat less of a problem (33% of pneumonia by p. aeruginosa), and was not apparently associated with antibiotic exposure, at least within the unit. conclusions. in our retrospective observational study we found a high incidence of pneumonia at our institution, as well as a high percentage of mrsa, the latter with strong relationship with exposure to vancomycin. new protocols for infection containment and antibiotic usage are urgently needed. introduction. community-acquired pneumonia (cap) carries a high morbidity and mortality. a major problem is the insufficient monitoring of cap by standard chest radiography, as the evaluation depends highly on the observer and the extent of pulmonary infiltration cannot be assessed properly (1). objectives. the aim of our study was to compare the process of inflammation in cap measured by alveolar nitric oxide (no)-analysis (2) in exhaled breath and the extent of the inflammatory infiltration by electrical impedance tomography (eit) (3) in spontaneously breathing patients. after approval of the local ethic committee and obtained written informed consent 24 patients with cap were included in the study. all patients showed an acute pulmonary infiltration in chest x-ray, pulmonary symptoms (coughing, shortness of breath), positive findings in auscultation, leukocytosis, elevated crp and a pneumonia severity index c4. no analyses (analyser cld88sp, eco medics, dürnten, switzerland) were performed at t0 (up to 24 h after admission), t1 (2 days after admission) and t2 (4 days after admission. eit measurements (eit evaluation kit, dräger medical, lübeck, germany) were performed at t0 and t2 and inhomogeneity of ventilation was assessed by offline analysis. all measurements were made at beside in sitting position. data were compared by t test and regression analysis. results. there was no significant correlation between the alveolar no concentration and the extent of inhomogeneity of the local infiltration measured by eit. also during the study the time course of the inhomogeneity index was not correlated with change in exhaled no. the right/left distribution of the pulmonary infiltration in the chest x-ray and the eit measurement showed a positive correlation (p \ 0.0001; r 2 = 0.69). conclusions. pulmonary regional infiltration in cap measured by eit can not predict the actual alveolar process of inflammation in the lung. nevertheless the monitoring devices give additional information to better evaluate the time course of inflammation and the dimension of the respiratory dysfunction in diseased lung. organizing pneumonia (op) presenting as acute respiratory failure (arf) is a relatively rare disease, and was only previously specifically reported in 2 small series [1, 2] , with mortality up to 90%. these studies were performed before the publication of international consensus classification of idiopathic interstitial pneumonias in 2001 [4] . objectives. to compare clinical features and prognosis of patients with op with those of patients presenting diffuse alveolar damage (dad), during arf. design: retrospective monocentric study in a university hospital conducted during an 11 yr-period. to determine predictors of niv failure in patients who were intubated for respiratory failure and extubated directly to niv. methods. this is a retrospective analysis of prospectively collected data from january 2008 to dec 2009. patients with respiratory failure were mechanically ventilated in a university hospital's medical intensive care unit (icu) and subsequently extubated to niv. physiological and biochemical parameters, using arterial blood gas measurements, were collected at the end of the spontaneous breathing trial and 1 h after the application of niv. failure of niv was defined as respiratory failure requiring re-intubation within 48 h. out of 175 patients, 78.9% were successfully extubated to niv. success rates were 70.6% in patients with chronic obstructive pulmonary disease (copd) and 80.9% in other patients (p = 0.24). patients who failed niv were more tachypnoeic, acidaemic and hypercapnic pre-niv, and more tachycardic, hypotensive, acidaemic, hypercapnic and hypoxaemic post-niv (p all.05). on logistic regression analysis, three physiologic parameters predicted niv failure: pre-niv respiratory rate (or 1.57, 95% ci 1.07-2.31 per 5 breaths increase), post-niv heart rate (or 1.28, 95% ci 1.08-1.52 per 5 beats increase) and post-niv systolic blood pressure (or 1.15, 95% ci 1.02-1.29 per 5 mmhg decrease). conclusions. physiologic parameters, including the respiratory rate pre-niv, and heart rate and systolic blood pressure post-niv, independently predict niv failure post-extubation. these parameters should be taken into account in the decision to extubate directly to niv. introduction. discontinuation of mechanical ventilation in critically ill patients is a challenging task and involves a careful weighting of the benefits of early extubation and the risks of premature spontaneous breathing trial (sbt). only a few studies have explored indices derived from both heart rate and breathing pattern variability analysis for the estimation of weaning readiness. objectives. to investigate heart rate (hr) and respiratory rate (rr) complexity in patients with weaning failure or success, using both linear and nonlinear techniques from signal processing theory. methods. forty-two surgical patients were enrolled in the study. there were 24 who passed and 18 who failed a weaning trial. signals were analyzed for 10 min during two phases: despite of passing the protocol the decision to extubate was postponed in some patients. to gain insight on the physicians reasons for continuing mechanical ventilation after passing the wean screen protocol. a wean screen protocol was introduced at a mixed medical (neuro-)surgical icu of a teaching hospital in december 2009 to april 2010. ventilation practitioners assessed ventilated patients and recorded the physicians reasons for continuing mechanical ventilation despite of passing the wean screen protocol. . 223 patients were ventilated in this period. 135 daily screens were performed, 111 screens were successful. only 24 passed wean screens resulted in extubation. the rate of extubation was 22%. 78% screens did not lead to liberation from mechanical ventilation. the extubation rate does not correspond with the findings of the abc trial with an extubation rate of 54%. table 1 shows the physicians' reasons to continuing mechanical ventilation. it should be noted that all patients with an unsafe airway were patients with a glasgow coma scale (gcs) of b 8 [intracerebral haemorrhage (68%), cerebral infections (13%), post-cpr encephalopathy (13%) and severe brain injury (6%)]. we accomplished a reduction in the use of sedatives (-12% midazolam and -35% propofol) and morphine (-27%) ( table 1 ). the amount of time spend on ventilators decreased, albeit not significantly (p = 0.902). this was probably due to the vap-ventilatorbundle (introduced last year), the heterogeneity of our cohort and the already short mv-duration. . non-invasive ventilation (niv) has been utilized in selected patients with hypoxemic arf to avert endotracheal intubation, which is related to life-threatening complications. niv has been also proposed to facilitate weaning and extubation in patients with hypercapnic arf. so far, no controlled randomized study has investigated the potential role of niv in weaning patients with hypoxemic arf. objectives. we designed this pilot study to assess safety and feasibility of niv to wean hypoxemic arf patients. twenty mechanically ventilated patients with hypoxemic arf were randomized to receive early extubation followed by niv application via helmet (helmet group) or conventional weaning through the endotracheal tube (tube group). primary outcomes were the duration of invasive mechanical ventilation and the adherence to the study protocol. secondary outcomes were protocol failure (i.e. need for re-intubation), icu and hospital mortality, rate of tracheotomy, duration of continuous intravenous sedation, weaning time, and septic complications. table 1 . weaning through helmet by niv application following early extubation was safe and feasible. overall the adherence to the study design was 90%. in addition, in the helmet group, there was a significant reduction in the rate of tracheotomy and a trend toward a lower rate of protocol failure, and fewer days on invasive ventilation. there was no difference with respect to days of continuous sedation, icu and hospital mortality, weaning time and septic complications. (1) . delirium is a common occurence on the icu and is associated with increased length of stay (los) and poor outcomes (2) . objectives. we developed a combined daily sedation hold, delirium management, and weaning (sdw) protocol and implemented this to reduce icu los and improve outcomes. methods. a sdw protocol was implemented in 2009. we prospectively audited all patients from january to march 2010. delirium was measured using the icdsc. data was analysed using graphpad statistical software. results. 109 consecutive patients were analysed. the incidence of delirium was 35% (38 pts). of these, 63% (24) had risk factors for delirium. there was no difference in onset of delirium between sexes, age, type of admission, or severity of illness. however, in patients with delirium, duration of mechanical ventilation (mv) and icu los were significantly longer and there was a trend towards increased hospital los ( conclusions. measuring the linear dependence of variables through time by k 2 and ø may be used to determine non-linear behavior between the variables of the emmv. non-linear behavior during weaning perhaps indicates the dependency of, either the resistance or compliance of the respiratory system, on the ventilatory support (i.e. pi). accordingly, k 2 and ø, estimated at the frequency interval form 6 to 1 (h) -1 , can provide information concerning to the dynamics of the respiratory system that can be used as a complement to determine the suitability of the mv withdrawal. objectives. to study the potential superiority of aprv on cmv in a subgroup of patients with severe ards. methods. retrospective observational study on 39 patients severe ards who were admitted between july 2008 and january 2010 to mafraq hospital icu in uae. the diagnosis of ards was based on presence of bilateral infiltrates in cxr and p/f ratio of less than 200 in absence of evidence of elevated left atrial pressure. all patients were managed according to ardsnet guidelines using low tidal volume cmv and iv steroids. criteria for transition to airway pressure release ventilation (aprv) included failure to wean down fio2 below 60% after 24 h, hemodynamic instability due to high peep, and failure to maintain plateau airway pressure below 30 cmh2o. initial settings of aprv were ph 26, pl 2, th 5, and tl 0.5 with titration of fio 2 as required keeping pao 2 more than 60 mmhg. we compared the outcome of cmv and aprv groups with special concern to the duration of mechanical ventilation, requirement for tracheostomy, and survival to icu discharge. twenty four male and 15 females were included in the study with a mean age of 42 years (± 24). fourteen out of them fulfilled the criteria and were shifted to aprv within 24 h of initiating mechanical ventilation. ten out of 14 (70%) patients in the aprv arm survived to icu discharge versus 16 out of 24 (67%) patients in cmv group (p 0.45). survivors in aprv group spent significantly shorter periods of mechanical ventilation compared to survivors in cmv group (9.6 vs. 12.1 days p 0.03). while 8 out of 16 (50%) survivors in cmv required tracheostomy for prolonged intubation or recurrent lavage, only 2 out of 10 (20%) survivors in aprv group required tracheostomy tube placement (p 0.02). we concluded that aprv can be effectively used as rescue measure of ventilation in patients with severe ards. although our study does not show any mortality benefit of using aprv over cmv, there was a shorter ventilation days and icu stay using aprv. we strongly recommend further studies to investigate the probability of using aprv as initial mode of ventilation in this subset of patients. weaning from mechanical ventilation is a common daily procedure when caring for critically ill patients, and a lifesaving practice on which nurses are taking an increasing role with the introduction of nurse-led protocols. the literature supports that nurse-led protocols facilitate weaning and increase nurses' input in decision-making. on the other hand, decision-making is a complex function affected by the nature of the task, the decision environment and the characteristics of the decision maker. although the cognitive process of clinical decision-making has been investigated with many different methodologies, little is known about the decision environment and its impact on decisions' during the weaning process. objectives. this paper aims to address one of the factors of the clinical environment and its impact on the decisions when discontinuing mechanical ventilation. methods. this paper is part of a large comparative ethnographic study looking at nurses' input during the weaning process of mechanically ventilated patients. participant observation of critical care nurses took place in an 12-bedded icu in greece and an 18-bedded icu in scotland for 6 months each to examine nurses' involvement in the decisions made. in-depth semi-structured interviews with the nurses followed focusing on how nurses perceived their participation in the decisions made. data from field notes and interview transcripts were analysed thematically using the qualitative data analysis software nvivo, version 8. inter-personal and inter-professional relationships were considered revealing influences of nurses' input in decision-making. clinicians' personality played a significant role in their involvement in decisions, whereas trust and appreciation, the sense of support and the sense of accountability were also considerable dynamics of inter-professional relationships and predisposed decision-making. clinical decision-making is a multi-dynamic process specifically in complex clinical long-term situations such as weaning. aspects of the decision environment, such as the interprofessional relationships should be acknowledged when introducing methods to enhance nurses' role in teamwork and collaborative decision-making in order to improve the weaning process of ventilated patients and their outcome. objectives. the objective of our study was to analyze the temporal trends and outcomes of two cohorts of patients ventilated with psv and pav+. a cohort of 16 consecutive patients who were ventilated with pav+ and another cohort of 15 consecutive patients who were ventilated with psv were compared. all patients had the same inclusion criteria (gas exchange, ventilatory mechanics, peep level, resolution/stabilization of the cause leading to invasive mv and appropriate level of consciousness). both modes were adjusted to predefined clinical criteria (psv to reach a respiratory rate about 25 bpm and pav+ to reach a physiological inspiratory effort introduction. presence of expiratory asynchronies (ea) (ineffective efforts, cough and continued contraction of inspiratory muscles) is a common problem associated with increased duration of mechanical ventilation, longer stay, higher costs and increased mortality. because of the lack of systems that automatically detect and report ea, their identification is currently done by examining ventilator interface at the bedside or by applying dedicated algorithms in investigational conditions. validate the accuracy of linear mathematical algorithms to automatically detect ea built in a new computerized system that grabs and process data from different bedside icu monitors and mechanical ventilators. observational and prospective study in a general icu of 16 beds. two beds were equipped with a software (better care ò ), a technological platform responsible for data acquisition and synchronization, processing, storing-as non static and processable dicom objects-and also for integrating all this data with health information systems. by using the better care ò platform, a total of 2,600,000 breaths from 8 consecutive adult patients were collected with at least 24 h of mechanical ventilation. algorithm #1: the ea algorithm consisted in a mathematical analysis of the airflow and airway pressure waveform variations during expiration not followed by a mechanical breath. algorithm #2: designed to select 1,000 breaths out of the total number. this algorithm sorted and classified the breaths by the percentage of deviation from the expected expiratory curve. the result was 1,000 breaths covering most of the shapes the expiratory curve could have. five expert attendant physicians independently analyzed the 1,000 selected breaths and classified them as ea or not. the ea algorithm processed the same 1,000 selected breaths and assigned a percentage to each one, according to the variation in the shape and direction of the expiratory airflow and airway pressure curves. the expert criterion against the ea algorithm scores was used to construct a logistic regression model. we calculated sensitivity, specificity, positive predictive value and negative predictive value. the predictive performance of ea algorithm was evaluated using roc curves. optimal sensitivity and specificity were achieved by setting the cut-off point at a ea algorithm score of 42%. a variation in the shape and direction of the expiratory airflow and airway pressure curves [42% compared to the theoretical curve identified an ea with a sensitivity of 91.5%, specificity of 91.7%, a positive predictive power of 80.3% and a negative predictive power of 96.7%. introduction. near-infrared spectroscopy (nirs) in combination with a vascular occlusion test (vot) has been proposed to assess and identify metabolic and microcirculatory alterations during sepsis and shock in critically ill patients. however, to automatize repeated measurements at the bedside, this technique can potentially cause discomfort to the patient. vascular arterial occlusion performed in the finger may be a more attractive method to execute repeated measurements at the bedside because of more tolerability from the patient. we have previously showed in healthy volunteers that nirs can be used on finger to assess the sto2 response to vot and that 5 min was an adequate occlusion time to provide the best curve fit for nirs dynamic variables 1 . objectives. we aimed to investigate whether sto2 response to vot obtained from the finger could predict conventional sto2 response measurements obtained from the thenar of critically ill patients. parameters of sto2 response were measured with an inspectra spectrometer model 650 (hutchinson technology inc.) equipped with a 15-mm or a 5-mm probe. the 15mm probe was placed over the thenar eminence and the 5-mm probe was place over the ventral face of the middle finger. we performed in each patient a series of two vascular occlusion tests (vot): one on the finger (5 min) followed by one on the arm (3 min). the measurements were obtained within 24 h of intensive care admission and every 24 h thereafter until day 3. vot-derived sto2 traces were analyzed for baseline, ischemic (rdecsto2, %/min) and reperfusion (rincsto2, %/s) parameters. we performed 63 paired of nirs measurements in 25 critically ill patients (age 62 ± 15; 16 m/9f). although sto2 did not differ significantly between thenar and finger (76% ± 11 vs. 79% ± 12; p = 0.31), rincsto2 and rdecsto2 were statistically lower in the finger (1.5%/s ± 0.7 vs. 2.5%/s ± 0.8, p = 0.001; 3.1%/min ± 1.0 vs. 11%/min ± 3.3; p = 0.001). we performed bivariate linear model with correlated errors in which sto2 outcomes on thenar and on finger were treated as responses. the correlation was significant for sto2 and rincsto2, but not for rdecsto2 (table 1) . furthermore, mixed model analysis showed that thenar-sto2 as dependent variable could be significantly predicted by finger-sto2 parameters with estimation coefficient (± se) of 0.7 ± 0.06 (p = 0.0001), 0.9 ± 0.09 (p = 0.0001) and 0.98 ± 0.3 (p = 0.0061) for sto2, rincsto2 and rdecsto2, respectively. correlation of sto2 response: finger vs. thenar a prospective randomized clinical trial performed in icu's of an university and teaching hospital during a 2.5 year period, involving 120 septic and non-septic patients, randomized (after stratification) to hemodynamic monitoring, by picco tm or pac with both techniques allowing cardiac output and central/mixed venous o 2 saturation monitoring. methods. hemodynamic management was guided by extravascular lung water index (evlwi) and global end-diastolic volume index (gedvi) in the picco tm group and by the pulmonary capillary wedge pressure (pcwp) in the pac group for 3 consecutive days. primary outcome measures were ventilator-free days (vfd), for which the study was powered, and lengths of stay in icu and hospital. secondary measures were the course of cardiorespiratory parameters, fluid and vasopressor requirements, lactate levels, organ functions and mortality. in the study period, 72 septic and 48 non-septic patients were included. 60 patients received a picco tm and 60 a pac catheter. monitoring arms were comparable at baseline, although sepsis differed from non-sepsis in hemodynamics and severity of lung injury. premorbidity was greater in non-septic patients. the fluid infusions and balances did not differ between monitoring arms, except at t = 24 h when the picco tm group had a more positive balance (p = 0.044). cardiac index and central venous o 2 saturation increased more in the course of time in the picco tm than in the pac group. the decrease in norepinephrine requirements strongly tended to favor the picco tm group (p = 0.06). the course of lactate levels and organ failure did not differ between monitoring arms. vfd did not differ among monitoring arms. picco tm monitoring was associated with relatively fewer mechanical ventilation and icu days in sepsis but more in non-sepsis (after day 28). the changes in respiratory parameters, sofa and number of catheter-related complications did not differ among the arms of the study. overall, 19 patients (32%) died in the picco tm group before day 28 and 21 (35%) in the pac group (p = 0.85). conclusion. hemodynamic management guided by picco tm monitoring is safe and results in better tissue oxygenation than guidance by pac, without inducing pulmonary overhydration, in septic and non-septic, critically ill patients. this was associated with fewer mechanical ventilation and icu days in patients with sepsis but more days in patients with non-sepsis (after day 28), partly attributable to greater cardiovascular premorbidity in the latter. the major primary and secondary endpoints, vfd and mortality, were not affected. introduction. non-invasive evaluation of endothelial function may be easily accomplished by ultrasound assessment of flow-mediated vasodilation (fmd) of the brachial artery, but this technique has not been fully explored in septic patients. objectives. this prospective study aims to investigate the role o fmd analysis on intra hospital prognosis of patients with severe sepsis and septic shock. adult patients admitted to the intensive care unit with a diagnosis of severe sepsis or septic shock (\24 h of duration) were consecutively included. fmd of the brachial artery was measured upon admission and after 24 and 72 h using a high-frequency linear transducer (7.5-10 mhz) according to internationally accepted protocols. a group of apparently health subjects paired for gender and age was used as controls for fmd analysis. patients were followed up to discharge or death. we studied 42 adult patients 9 mean age 51 ± 19 years, 26 females, 79% on vasopressors with sepsis predominantly of abdominal or respiratory etiology (75%). apache ii risk score was 23 ± 7 and intra hospital mortality rate was 33%. fmd was similar in patients with or without use of vasopressors at baseline (p = 0.56). fmd in septic patients was significantly lower than in health controls (1.5 ± 7 vs. 6 ± 4%; p \ 0.001). we observed that survivors depicted a gradual improvement on endothelial function, so that 72 h after sepsis onset fmd was significantly lower in nonsurvivors (-3.3 ± 10 vs. 5.2 ± 4%; p \ 0.05; time-group interaction p value = 0.03). conclusions. brachial fmd is altered in septic patients with hemodynamic instability and its improvement may be an early marker of favorable prognosis. introduction. change in pulse pressure variation (dpp) and respiratory variation of the pulse oxymetry plethysmogram (pop) may predict the hemodynamic effect of peep in mechanically ventilated patients [1, 2] . reported comparisons [3, 4] between pop variations (popv) and co or dpp are based on selection of 3-5 consecutive breaths (dpp3b) during a ''stable'' period of pop. recently, a fully automatic ventilation mode (intellivent ò , hamilton medical, switzerland) that incorporates an automatic and continuous popv calculation (hli ò ) using a dedicated algorithm has been developed. the present study was designed to compare dpp3b, dpp calculated with the algorithm as hli ò (dppalg) and hli ò. . . 11 sedated icu patients ventilated with hamilton medical s1 ventilator (with integrated pulse oxymetry (po)) were included (age = 66 ± 13 years, saps ii = 54 ± 24, no arrhythmia, norepinephrine: 1.0 ± 1.2 mg/h in 6 patients, map = 74 ± 10 mmhg, vt = 8.4 ± 0.6 ml/kg). waveforms of po from a finger sensor and of blood pressure from a radial catheter were recorded for 1-2 h. from the waveforms, breath by breath (using respiratory flow signal), without pre-selection of stable periods and using known formula [1] dpp3b (averaging 3 breaths without any filtering), dppalg and hli ò were automatically obtained (matlab ò ). dpp3b was compared to dppalg (2600 pairs) using mann-whitney t test. 18287 pairs of hli ò and dppalg values (see fig. 1 below) were compared using linear regression and bland-altman method. a dppalg threshold value of 13% was used to generate hli ò roc curves. results. dpp3b and dppalg were significantly correlated (r 2 = 0.99, p \ 0.001), but standard deviation of dpp3b were higher than the standard deviation of dppalg (1.2 ± 0.4 vs. 0.2 ± 0.1%, p \ 0.001). dppalg and hli ò were correlated (r 2 = 0.73, p \ 0.001), mean difference was 2 ± 4%. hli ò above 13% predicted dppalg above 13% with a sensitivity of 95% and specificity of 82% (roc: 0.96). conclusions. dpp3b should be interpreted with caution due to the high variance of this index. in real conditions and during long time monitoring dppalg and hli ò are in acceptable agreement and hli ò may help estimating continuously the hemodynamic effects of ventilation. introduction. transthoracic echocardiography (tte) is supposed not to be useful in ventilated patients (pt). echocardiography is usually performed transesophageally in ventilated pt and is thought to be independent of the examiner's skills. we want to demonstrate that tte in ventilated pt could be learned even by medical students with reasonable results and that tte could add useful informations for interpretation of the hemodynamic status. objectives. in a prospective observational study 42 consecutive patients (pt) were enrolled in a 9-bed medical intensive care unit of a university hospital. inclusion criteria was septic shock according to actual guidelines. transthoracic echocardiography (acuson cv70, siemens, germany) was performed by a medical student in each subject on day 1, day 7 and survival was reported on day 28. tte-examination was reduced to an apical 4-chamber view for interpretation of left ventricular global function and calculation of left ventricular ejection fraction (ef) with the simpson method and to a subcostal view in order to examine the diameter of the inferior caval vein (ivc) and to rule out pericardial effusion. each examination was digitally recorded and was interpreted by an experienced cardiologist. every single pt was mechanically ventilated. cardiac output (co) was measured with the transpulmonary thermodilutional technique (picco-catheter, pulsion, germany). the insertion of the picco-catheter took place due to an individual physician's decision. crp was measured as an parameter of inflammation. results. 42 pt, mean age 71 years ± 7.22, 27 male (64%), 23 pt with known coronary artery disease (55%), 2 pt with known dilated cardiomyopathy (5%). mean apache ii-score 35.9 ± 6.8. 22 pt died within 28 days (52%). picco-catheter was inserted in 21 pt (50%). tte could be successfully performed in 38 pt (91%). the following values are expressed as mean values ± sd, student's t test, p \ 0.05 denotes statistic significance. ef on day 1 43.6% ± 16.2, ef on day 7 42.9% ± 15.7, p = 0.04. ivc on day 1 20.3 mm ± 4.5, ivc on day 7 20.2 mm ± 4.5, p = ns. co on day 1 5.5 l/min ± 3.4, co on day 7 4.0 l/min ± 3.4, p = 0.05. crp on day 1 19.82 mg/dl ± 10.4, crp on day 7 9.91 mg/dl±, p\ 0.0001. pericardial effusion in no pt. in older pt coronary artery disease is common and ef is at the start of septic shock severely diminished. ef decreased slightly in the early course of septic shock, may be as an expression of septic cardiomyopathy. the ivc diameter did not change and may not be useful as a predictor of preload in ventilated pt. co decreased over time as the hyperdynamic circulation in septic shock is getting normalised. tte adds useful hemodynamic information and should be performed in each ventilated pt. tte could be performed in almost each ventilated pt and is easily learned even by medical students. (1, 2) , which can often be caused by anaemia. in current guidelines the transfusion trigger is haemoglobin (hb) \ 7 g/dl, but there is no recommendation for scvo 2 (3). objectives. the aim of this retrospective study was to evaluate the change in scvo 2 before and after transfusion and to reveal whether co 2 -gap reflects it. methods. over a 6 month period hb, scvo 2 , co 2 -gap and o 2 -extraction ratio (o 2 er) were recorded before and after transfusion. data are presented as median [interquartile range], for statistical analysis wilcoxon, mann-whitney tests and pearson correlation were used as appropriate. results. out of 128 transfusion events the scvo 2 was measured in 50 cases. after transfusion hb increased significantly: 7.7 [7.1-8.2]-9.0 [7.9-9.7] g/dl, p \ 0.001. the median scvo 2 was 71 %, therefore two groups were created: ''low'' (scvo 2 \ 71 %, n = 27); ''high'' (scvo 2 c71%; n = 23). hb increased significantly in both groups (p \ 0.001), but scvo 2 conclusions. in the high-group the low hb levels did not cause oxygen debt, as after transfusion hb increased significantly but scvo 2 did not, and o 2 er and co 2 -gap were within the normal range. our results give further support that not only the hb level should serve as a transfusion trigger, but measures of oxygen debt such as scvo 2 and co 2 -gap should also be considered, hence unnecessary transfusions could be avoided. introduction. intellivent ò is a fully closed loop ventilation designed to keep the patient within target ranges of etco2 and spo2. the system includes an automatic adjustment of peep and fio2 following the ardsnetwork tables [1] . if required peep is changed by 1 cmh 2 o every 6 min with a maximal possible value set by the user or depending on an automatic and continuous calculation of the respiratory variations of the plethysmogram from an integrated pulse oxymeter (hli ò ), i.e. the higher the hli ò the lower the maximal peep allowed by the system. the present study was designed to estimate whether changes in peep are reflected in hli ò changes. in 17 sedated icu patients ventilated for 120 min in fully closed loop ventilation with intellivent ò (hamilton medical s1 ventilator), 34 episodes of significant changes in peep (c 4 cmh 2 o) were selected and hli ò values within 5 min before and after peep changes were collected. statistics were done using sigmastats with p \ 0.05 as significant. results. changes in peep and in hli ò are shown in the table 1 15 ± 3 cmh2o 9 ± 4 cmh2o 17 ± 12% 9 ± 6% p \ 0.001 the correlation between change in peep and change in hli ò is shown on the fig. 1 . conclusions. based on these preliminary data changes in peep are reflected hli ò changes and may help estimating continuously the hemodynamic effects of ventilation. objectives. we have tested a 3 axis accelerometer sensor for detection of regional left ventricular ischemia. in 12 pigs a 3-axis accelerometer was sutured to the left ventricular (lv) apical region in left descending coronary artery (lad) supply area accelerometer x-axis measured longitudinal-, y-axis circumferential-and z-axis radial epicardial motions. epicardial displacements were calculated from the acceleration signals and systolic displacements within 150 ms after peak r on ecg was measured. lad was occluded for 60 s to induce regional lv dysfunction. myocardial circumferential strain (shortening) measured by echocardiography in the lv apical anterior region was used to confirm ischemia. the ecg st-segment in lead ii was also monitored. data are presented as mean ± se. early systolic displacement at baseline was 11 ± 4 mm, 12 ± 2 mm and 3 ± 2 in circumferential, longitudinal and radial directions, respectively. lad occlusion induced akinesia in circumferential (0 ± 3 mm, p \ 0.001) and radial (0 ± 1 mm, p = 0.01) directions, whereas longitudinal displacement changed less to 10 ± 2 mm (p = 0.280). ischemia was confirmed by echocardiography strain, showing lengthening in systole (p \ 0.001). no significant changes were observer in the ecg st-segment during coronary occlusion (p = 0.341). introduction. there is increasing evidence to suggest perioperative complications are predictive of long term survival and that reducing them may improve survival rates 1 . goal directed therapy has been shown to reduce mortality and morbidity perioperatively, with those unable to increase oxygen delivery perioperatively having demonstrably worse outcomes. the advent of non invasive tissue oxygenation monitors using near infrared spectroscopy has allowed further study of oxygen flux during goal directed therapy. objectives. to observe changes in tissue oxygenation during an 8 h oxygen delivery targeted post surgical optimisation program and provide long term mortality followup of a surgical cohort of high risk patients. methods. 40 patients undergoing high risk surgery and postoperative optimisation (targeting of oxygen delivery index of [600 ml/min/m 2 ) on the tensive care unit at a london teaching hospital were enrolled. each patient underwent a protocolised haemodynamic optimisation protocol as per our standard unit policy for 8 h with consecutive recordings of tissue oxygenation at the thenar eminence using an inspectra 325 monitor. additional variables relating to global and tissue perfusion were measured concurrently. patients were followed up for survival status at 3.5 years using routinely available information held within our hospital records. in hospital mortality was 17.5% (n = 7), whilst at 3.5 years this had increased to 50% (n = 20). there was no significant difference between apii scores 11 (4) versus 9.5 (4), age 61.95 ± 16.23 versus 65.8 ± 16.35 or operation type for survivors and non-survivors at 3.5 years respectively. significant differences between groups were found however for admission and mid optimisation protocol (4 h) hr and sto2 (see table 1 there were no significant differences in measured variables for 30 day mortality. conclusions. there appears to be a statistical and clinical difference in hr and tissue oxygenation between the long term survivors of high risk surgery who undergo monitored postoperative goal directed optimisation. introduction. bronchoscopic bronchoalveolar lavage (b-bal) is today the gold standard for sampling of inflammatory markers in the distal airways. nonbronchoscopic bronchoalveolar lavage (n-bal) by ordinary suction catheter has been investigated as a more easily accessible method for alveolar sampling in the setting of acute respiratory distress syndrome (ards). the results, however, were disappointing, probably due to more proximal sampling by the n-bal. to investigate wether n-bal by a catheter with physical properties similar to those of the bronchoscope is comparable to b-bal. methods. b-bal and n-bal by cook's airway exchange catheter was performed with 3 9 30 ml normal saline on opposite sides 15 min apart at nine different occasions on 5 anesthetized and intubated pigs. the volume of the recovered lavage was noted, after which the fluid was analyzed for albumin, total cell count, viability and differential cell count. statistical analysis was performed using wilcoxon's rank-sum test. results. n-bal yielded significantly higher albumin content than b-bal (20.1 ± 8.7 vs. 11.7 ± 3.4 mg/l, p = 0.027). in all other measurements there were no significant differences between n-bal and b-bal (recovered volume 52. objectives. we hypothesized that collagen synthesis and degradation are disturbed in acute respiratory failure. in the finnali-study we defined acute respiratory failure as need of noninvasive and/or invasive ventilatory support for more than 6 h (1). after informed consent we collected blood samples for serum procollagen propeptides i and iii (pinp, piiinp) and ictp levels at study admission, day 2, 7 and 21. patients with all four blood samples were included in this substudy. multiple organ dysfunction (mod) was defined as two or more individual organ sofa scores of 3-4 at any day during the first week. results. the study population comprised 68 of 958 finnali patients (1). the mean (sd, range) age was 60 years (15, 20-86) and the majority were male 74%. on admission the mean sapsii score was 45 (15, 2-83). 16 patients (24%) developed mod during the first 7 days. over time piiinp/pinp-ratio first increased and then decreased to baseline by day 21 while pinp/ictp-ratio decreased and then decreased to baseline by day 21(p \ 0.001 and p = 0.006, respectively) ( fig. 1 ). there were no statistical differences in the ratios between patients with or without mod. conclusions. we found that in patients with acute respiratory failure the balance of collagen synthesis was towards degradation of type i collagen and production of collagen type iii. 3 ± 43 ng/ml in the ards group, and significantly higher than the 53.8 ± 37.0 ng/ml in the ali (not ards) group. the difference in hmgb1 values in the early stage between the group that died up to the by 30th day and the surviving group was not significant, but the hmgb1 values were significantly higher in the group that died until the 60th day and 90th day than in the survival group. it was concluded that differences in hmgb1 values in the early stage after the onset of ali (not ards)/ards are useful as outcome determining factors after 30 days of onset. an inverse correlation was observed between the hmgb1 values and lung oxygenation, suggesting the possibility that hmbg1 is involved in the development of respiratory failure. s. shibata 1 , g. takahashi 2 , n. shioya 3 , s. endo 3 1 akita city hospital, anesthesiology, akita, japan, 2 iwate medical university, emergency medicine, morioka, japan, 3 iwate medical university, critical care medicine, morioka, japan sivelestat sodium hydrate (sivelestat) is a selective polymorphonuclear leukocyte elastase (pmn-e) inhibitor and has also been shown to be effective for pulmonary disorders associated with sirs in clinical patients. blood levels of inflammatory cytokines have been shown to be decreased in patients treated with sivelestat. however, since patients with sirs have already received other drugs, it remains indefinite whether or not sivelestat might suppress the production of cytokines. moreover, it is difficult to clarify any cells releasing cytokines. in the experiment using cells isolated from the blood, intercellular mutual actions and cytokine networks were blocked and the experiment failed to faithfully reproduce the in-vivo condition. objectives. the possibility of sivelestat suppressing the production of cytokines from granulocytes and monocytes was assessed by intracellular cytokine staining using the whole blood culture method and flow cytometry to faithfully reproduce the in-vivo condition. methods. blood samples were collected from 9 healthy volunteers. a vehicle (control group), lipopolysaccharide (lps; lps group), or lps + sivelestat (sivelestat group) was added to the whole blood, followed by the addition of a protein transport inhibitor in each group. after incubation, they were subjected to staining of the cytokines retained in the cells by the addition of an anti-interleukin 8 (il-8) or anti-tumor necrosis factor a (tnf-a) antibody and analysis by flow cytometry. the data were analyzed by the kolmogorov-smirnov test. values obtained [d/s(n)] result from the comparison of the fluorescence histograms of each sample with a control one. addition of sivelestat at low concentrations (1 and 10 lg/ml) significantly (p \ 0.01) suppressed the production of il-8 from granulocytes induced by a low concentration (1 ng/ml) of lps. on the other hand, the granulocytic production of tnf-a induced by a high concentration of lps (10 ng/ml) was significantly (p \ 0.01) suppressed by treatment with sivelestat at high concentrations (10 and 100 lg/ml). with regard to the monocytic production of tnf-a and il-8 induced by lps, there was no significant suppression of either tnf-a or il-8 production by sivelestat. conclusions. sivelestat, a neutrophil elastase inhibitor, suppressed granulocytic production of il-8 and tnf-a, suggesting the potential usefulness of sivelestat for the treatment of various morbid conditions involving il-8 and tnf-a in their onset. introduction. coagulation, fibrinolysis and extravascular fibrin deposition are the hallmarks of the pathogenesis of acute lung injury (ali). pai-1 has a central role in antagonizing fibrinolysis by decreasing the plasminogen turnover to plasmin. pai-1 has been suggested as a clinical severity marker of ali. in previous studies it was associated with higher mortality and morbidity in the critically ill. upar is a cell surface receptor activating the serine protease upa. increased expression of upar is found in various stages, including inflammation, tissue remodelling and malignancies, indicating poor prognosis. pai-1 antagonizes the proteolytic activities of upa and plasmin. objectives. we sought to evaluate the prognostic value of supar and pai-1 for 90-day mortality of patients with acute respiratory failure (arf). the finnali-study patients needed invasive or non-invasive ventilation for more than 6 h (1). blood samples were collected from patients at baseline and on day 2 after baseline. healthy volunteers were also analyzed. sera were frozen at -80°c until analyses. concentrations of supar and pai-1 in blood serum were measured by enzyme linked immunosorbent assay (elisa). data are presented as median (iqr). the prognostic value of supar and pai-1 for 90-day mortality was determined with roc analysis. in the critically ill, supar and pai-1 were 12.7 (10.7-14.7) ng/ml and 2.33 introduction. acute lung injury is a common disease in intensive care, associated to various septic or inflammatory diseases. inflammation is part of the defense mechanisms of innate immunity, occurring after tissue injury. objectives. the aim of the project was to decipher the transcriptional changes occurring after the onset of an inflammatory injury by intravenous injection of oleic acid. experimental study of the lung transcriptome after oleic acid injection. thirtysix c57bl/6 j mice, aged of 7 weeks, were sacrificed at 1h, 1h30, 3h, 4h, 18h and 24h after physiological serum or oleic acid injection (20 ll) in the caudal vein. left and right lung were separated for mrna extraction and pathological examination. labelled cdna were hybridized on cdna nylon microarray (tagc, marseilles, france) and raw data were extracted from scanned images with bzscan2 software. raw data were normalized with the quantile method, and supervised analysis was conducted with significance analysis of microarray algorithm within the r statistical suite and bioconductor libraries. after the administration of oleic acid, the mice were tachypneic and prostrated. all survived during the 24 first hours. the pathological analysis of lung tissue revealed an early inflitration of the lung tissue by polynuclear cells, as well as a pulmonary edema. these alterations were not observed after 4h. the time course analysis of transcriptional lung data identified a thousand genes which expression is modulated after injury. hierarchical clustering identified 3 major groups of genes. the first one (365 genes) is composed of genes transiently up-regulated between 1h and 3h after oleic acid injection. th second group (366 genes) is composed of genes expressed between 1h30 and 4h. the third group (366 genes) is composed of genes expressed at the later time points (18h-24h). the functional annotation linked these signatures with keywords related to pro-inflammatory response, vascular endothelium modification and lipid metabolism, respectively. rt-pcr analysis of pro-(tnf, il6) and anti-inflammatory (il4, il10) markers related the pro-inflammatory phase to the earlier time points (1h-1h30) and the anti-inflammatory phase to the late points (after 4h). conclusions. oleic acid injection in mice induced a transient acute lung injury. this is confirmed by clinical, pathological and transcriptional modifications. the modulation of gene expression after the oleic acid injection revealed an early pro-inflammatory response, followed by an anti-inflammatory response and lipid metabolism modificiations. this model could now be used to describe the specific modulation occuring during pulmonary infection and critical injuries like acute respiratory distress syndrome. introduction. ventilator associated lung injury (vali) is influenced by tidal volumes, airway pressure and cyclic opening of alveoli during mechanical ventilation. preserved spontaneous breathing during partial ventilatory support may be protective, but it is not known whether the transpulmonary pressure generated by spontaneous breathing has the same effect on vali as if generated by the ventilator. to determine whether hemodynamics, respirtory function and vali are influenced by the amount of support provided by pressure support ventilation. after approval from the institutional animal care committee, acute lung injury was induced in 50 anesthetized sd rats by acid aspiration. ten animals each were then ventilated with positive end-expiratory pressure 5 cmh2o in pressure control (pc), pressureregulated assist control (ac) or pressure support mode with 100% (ps100), 60% (ps60) or 20% (ps20) pressure support of initial distending pressure needed to maintain tidal volume. pc animals were paralyzed. after 4 h animals were killed and vali determined. results. there were no differences in baseline characteristics. acute lung injury was characterized by a decrease of the p/f ratio from 447 ± 75 to 235 ± 90 mmhg and of the dynamic compliance from 0.5 ± 0.2 to 0.28 ± 0.1 ml/cmh 2 o. conclusions. compared to controlled ventilation, preserved spontaneous breathing activity improved hemodynamic stability, respiratory function and lung edema clearance. the reduction in pressure support did not lead to reduced tidal volume, but transpulmonary pressure was preserved by muscular activity of the chest wall. no difference was observed between full or 60% of pressure support, but further reduction in pressure support resulted in increased wet-dry ratio. objectives. we studied the effects of metabolic acidosis on enzymatic and non-enzymatic no-production in hypoxic and hyperoxic lung regions in a pig model. eighteen healthy anesthetized pigs were separately ventilated with hypoxic gas to the left lower lobe (lll) and hyperoxic gas to the rest of the lung. six pigs received hcl infusion (hcl group), six pigs received n w -nitro-l-arginine methyl ester (l-name) and hcl (l-name + hcl group) and six pigs received buffered ringer's solution (control group). no concentration in exhaled air (eno), no synthase (nos) activity in lung tissue, and regional pulmonary blood flow were measured. results. metabolic acidosis, induced by infusion of hcl, decreased the relative perfusion to the hypoxic lll (q lll /q t ) from 7 (±3) to 3 (±1)% in the hcl group (p \ 0.01), and from 4 (±1) to 1 (±1)% in the l-name + hcl group (p \ 0.05), without any measurable significant changes in eno from hypoxic or hyperoxic lung regions there were no significant differences between the hcl and control groups for ca 2+ -dependent or ca 2+ -independent nos activity in hypoxic or hyperoxic lung regions. metabolic acidosis augmented the hypoxic pulmonary vasoconstriction, without any changes in pulmonary enzymatic or non-enzymatic no-production. when acidosis was induced during ongoing nos-blockade, the perfusion of hypoxic lung regions was almost abolished, indicating acidosis-induced pulmonary vasoconstriction was not no dependent. assessing and monitoring biomarkers in acute lung injury (ali) may improve knowledge of its pathogenesis, early recognition, and management and predict remote organ injury and multiple organ failure. objectives. early consents for research are difficult to obtain in patients with or at risk of ali because of the emotional burden of the severity and sudden onset of the disease. however, study samples may be obtained from left-over clinical blood draws, which are readily available if processed adequately. the aim of this study was to compare fresh and ''waste'' blood samples prospectively in a series of consecutive critically ill patients. the hypothesis is that ''waste'' blood samples if appropriately processed provides accurate and reliable results comparable to the gold-standard, which is immediate collection and processing of fresh blood samples. prospective study comparing biomarkers of epithelial injury (srage) and inflammation (20 different cytokines/chemokines) in critically ill patients measured on fresh blood or waste blood, kept at 4 degrees celsius for 24 h. an automated system performed a daily screening of adults in the icu with an increased risk for ali (lung injury prediction score, lips) within 12 h of admission and/or on recognition of the diagnosis of ali, using the american-european consensus conference criteria. risks factors for ali include pneumonia, sepsis, pancreatitis, shock, aspiration, high risk surgery and high risk trauma. irb approved the protocol and written consent was obtained from patients or their surrogates. statistical measurements were performed using the bland-altman analysis for correlation between fresh and waste blood sample data. between may and december 2009, 30 patients were enrolled. one patient was excluded due to lack of sample. samples were obtained either at one time point (n = 25) or two, on consecutive days (n = 13). female/male patient ratio was 12/17. seven of the 29 patients had ali. twenty two patients had risk for ali with a median lips score of 4 (iqr 3.5-4.5). sepsis was the most common risk factor, present in 23 patients. in-hospital mortality was 28% (8/29). the bland-altman plot (mean bias ± se, limits of agreement) showed good correlation for il-1ra (-9 ± 14.3 pg/ml, -38.3 to 20.4 pg/ml), il-6 (0.9 ± 2.9 pg/ml, -5 to 6.8 pg/ml), il-8 (-0.3 ± 0.3 pg/ml, -0.9 to 0.3 pg/ml), il-12 (p40) (-0.2 ± 0.3 pg/ml, -0.7 to 0.4 pg/ml), mcp-1(-0.2 ± 2 pg/ml, -4.2 to 3.9 pg/ml) and srage (-24 ± 53 pg/ ml, -130 to 83 pg/ml) between fresh blood and ''waste'' blood samples. in patients with ali, properly stored blood, drawn for clinical purposes, can be processed within 24 h for research purposes. however, the stability of each biomarker of interest needs to be individually validated before using stored blood introduction. pulmonary surfactant inactivation following acute lung injury might promote alveolar derecruitment and reduce the airspace available for ventilation, making the lung more prone to ventilation-induced lung injury (vili). our aim was to test the potential for a protective effect of exogenous surfactant treatment in a model of acid aspiration and vili. methods. 17 male c57/bl6 mice were anesthetized, mechanically ventilated (vt 15 ml/ kg; rr 130/min; peep 2 ± 0.2 cmh 2 o; fio 2 0.5) and immediately subjected to intrabronchial (right) instillation of 1.5 ml/kg hcl 0.1 m. mechanical ventilation went on for 420 min. 180 min after the acid instillation, 9 mice were treated with exogenous surfactant (80 mg of phospholipids/ml) given as bolus of 1 ml/kg in the right bronchus (surf group). we measured oxygenation, lung compliance (measured every 60 min throughout the experiment), macrophage inflammatory protein (mip) 2 in broncho-alveolar lavage (bal) fluid. . pao 2 at the end of the experiment was significantly higher in the surf than in control group (383 ± 163 vs. 198 ± 104 mmhg, p \ 0.05). although surfactant bolus caused a reduction in lung compliance measured 10 and 60 min after treatment, in the surf group compliance restored to 98 ± 7% of the post injury level, while it decreased in control group to 88 ± 8% (p \ 0.05). there were no differences between groups in the dosage of mip-2 in bal neither in right or left lung. conclusions. exogenous surfactant treatment improved lung function in a murine model of two hit lung injury. grant acknowledgment. introduction. ventilator induced lung injury significantly contributes to the mortality in patients with acute respiratory distress syndrome, the most severe form of acute lung injury. understanding the molecular basis for response to cyclic stretch and its derangement during high volume ventilation is of high priority. objectives. to identify specific molecular regulators involved in the development of ventilator induced lung injury. we undertook a comparative examination of cis-regulatory sequences involved in the coordinated expression of cyclic stretch responsive genes using microarray analysis. analysis of stretched vs. non-stretched cells identified significant enrichment for genes containing binding sites for the transcription factor atf3 (activating transcription factor 3). to determine the role of atf3 in vivo, we compared the response of atf3 gene deficient mice to wild type litter mates in an in vivo model of ventilator induced lung injury. results. atf3 deficiency results in increased sensitivity to mechanical ventilation alone or in conjunction with inhaled lipopolysaccharide (10 mg/kg) as determined by assessment of lung and bronchoalveolar lavage cell infiltration and pro-inflammatory mediator release, pulmonary edema and indices of tissue injury. the expression of genes containing an atf3 cis-regulatory region was significantly altered in gene deficient animals. atf3 protein expression and nuclear translocation is increased after mechanical ventilation. conclusions. atf3 deficiency confers increased sensitivity to mechanical ventilation alone or in combination with inhaled endotoxin. in our model, atf3 acts to ''counterbalance'' cyclic stretch and high volume-induced inflammation, limiting its potential to cause additional lung injury and consequently protecting animals from injurious cyclic stretch. objectives. our aim was to evaluate the role of the alveolar macrophages in a murine model of ali, by selective depletion of this type of cells from the air space achieved by clodronate administration. mice were treated (it) with 100 ll of clodronate (clo)-or pbs (pbs)-liposomes. after 24 h mice were anesthetized and ventilated (vt 8-10 ml/kg, rr 140 min -1 , fio 2 0.21); in order to induce lung injury 2 ml/kg of hcl (0.1 m) or air bolus (sham group) was instilled in the right bronchus. mice were ventilated for 10 min, and extubated after awakening. 24 h after injury, animals were sacrificed and broncho-alveolar lavage (bal) and blood gas analysis (fio 2 = 0.21) were performed. . 24 h after lung injury animals with alveolar macrophages depletion, showed a better oxygenation versus pbs-treated group. however, recruitment of neutrophils in bal was not statistically different between clo_hcl and pbs_hcl group. results. high levels of oc were found in patients treated by 150 mg of ot bid. oc levels ranged from 2,507 to 4,551 ng/ml in these patients. concentrations of oc were five-to tenfold higher than concentrations reported in healthy volunteers. lesser levels were found in patients treated by 75 mg of ot bid. nevertheless, the patient with the moderate renal failure seemed to accumulate oc (levels ranged from 906 to 1470 ng/ml) whereas concentrations reported in the patient with a normal renal clearance were below (152-349 ng/ml). conclusions. ecmo seemed not to have any influence on oc concentrations while renal insufficiency seemed to be the parameter leading to oc accumulation. as ic50 was very low and reached even with usual dosage, increasing ot dose to 150 mg bid appeared to be unnecessary. objectives. aim of our study was to evaluate the effect of nursing care on patients undergoing venous-venous ecmo for acute respiratory distress syndrome (ards). methods. we recorded physiological and ecmo parameters (heart rate, arterial blood pressure, mixed venous saturation (svo 2 ), arterial oxygen saturation (spo 2 ), body temperature and extracorporeal blood flow (bf)) before and during daily nursing in 5 patients undergoing vv-ecmo for several days (each patient was followed on average for 4.6 days, 23 cases in total). arterial blood gases were also collected before and after nursing care. daily nursing was performed following defined steps (sponge bath, oral hygiene, change position of endotracheal tube, elevation with scooping stretcher for sheets replacement and back hygiene, dressing replacement) in agreement with a standard protocol in use in our department. (expressed as mean ± standard deviation). all patients were affected by ards h1n1-related. patients were sedated with propofol (182 ± 68 mg/h) or midazolam (7.9 ± 1.8 mg/h) plus an opioid drug (fentanyl 175 ± 45 mcg/h or remifentanil 0.2 ± 0.01 mcg/kg/min or sufentanil 0.32 ± 0.09 mcg/kg/min). ramsey score before nursing was 5.4 ± 0.9. in 11 cases patients were paralysed. in table 1 we summarized the adverse events observed during nursing care, divided into hypertensive or tachycardic episodes, blood oxygen desaturation, reduction in svo 2 or reduction in bf. forty-nine sedative bolus were administered during nursing (mean request for each patient: 2.1 ± 1.9), always after an episode of hypertension or tachycardia (most frequently during elevation with scooping stretcher and changing position of endotracheal tube). although in 13 cases preventive bolus of sedation were administered before nursing, in 8 of those cases (61%), additional bolus were required. we found an inverse correlation between bf and the increase in heart rate, drop in arterial saturation and svo2. despite active warming, we observed a drop of 0.31 ± 0.15°c (p \ 0.01) in body temperature. nursing care may have a significant impact on physiologic parameters of patients during vv-ecmo. tachycardia, hypertension and reduction in oxygenation were commonly recorded and were not prevented by pre-nursing bolus of sedation but were attenuated in patients with higher bf. introduction. prone position has been used in cases of ards with refractory hypoxemia but some physiological effects are still unknown. prone position could increase intraabdominal pressure (iap) and could lead to acute renal failure (arf). acute kidney injury in icu is associated with increased mortality. objectives. the aim of this study was to determine whether prone position could increase intraabdominal pressure and possibly promote arf. we studied all adult ards patients who were ventilated using the protective strategy defined by ards network criteria and who needed prone position to improve oxygenation. we collected respiratory data (ventilator parameters and gas exchange) and hemodynamic variables (heart rate, systolic, diastolic and mean arterial pressure). iap was measured using the abdo-pressure tm bladder transducer following world society of acute compartment syndrome recommendations. abdominal perfusion pressure was calculated as mean arterial pressure minus iap. main renal parameters were: filtration gradient (fg), creatinine clearance, fractional excretion of sodium (fena) and urea (feurea). patients were classified according to rifle score after each manoeuvre. all data were recorded in prone and in supine position at least once per day. results. the study included 18 patients (14 male) admitted to a medical-surgical icu over a one-year period. their mean age was 47.3 ± 17.8 and length of icu stay was 23 ± 18 days. all 18 patients had primary ards and 9 had received nephrotoxics. icu mortality reached 50%. we recorded at least 3 manoeuvres per patient (a, b, c). prone positioning improved pafio 2 ratio from 101.5 ± 5 to 150 ± 72 (p = 0.027). iap showed a small increase from 6.1 ± 2.7 to 8.9 ± 2.9 mmhg (a; p = 0.004), from 8.4 ± 3.4 to 11.2 ± 3.7 mmhg (b; p = 0.038) and from 7.6 ± 1.3 to 9.9 ± 0.8 mmhg (c; p = 0.001). there were no statistically significant changes in hemodynamic parameters or abdominal perfusion pressure. renal function parameters (fg, creatinine clearance, fena and feurea) showed no modification after each prone positioning. in contrast, when patients were classified according to rifle score, we observed a trend towards worsening, though this was not statistically significant. conclusions. prone positioning improved arterial oxygenation in primary ards patients and was associated with an increase in iap. however, creatinine clearance and glomerular filtration remained unchanged. percutaneous extracorporeal life support system (p-ecls) including ecmo becomes widely used in medical and surgical emergent situation, such as refractory cardiogenic shock, cardiac arrest and acute respiratory failure. patients requires highly specialized intensive care and monitoring system. we reviewed our ecls experience and tried to analyze the clinical outcomes, factors for survival and frequently faced problems during management for improving weaning and survival rate (medical vs. surgical patients). introduction. in spite of the huge efforts spent over the last years, conventional treatment of acute hypoxemic respiratory failure (ahrf) is often inadequate and alternative procedures must be instituted. icus skillful in extracorporeal membrane oxygenation (ecmo), as recently shown [1] , may improve survival of these patients. since 1989 we developed a treatment algorithm for ahrf which encomprises: (1) low flow venous-venous ecmo (lf-ecmo) consisting in a relatively low initial blood flow (bf, 2-2.5 l/min) to maximize extracorporeal co 2 removal while providing partial oxygenation (if needed, bf can be increased up to 4.5-5 l/min to keep arterial po2 above 45 mmhg); (2) femoral-femoral percutaneous cannulation with 21-25 fr cannulas to allow free movements of the neck and increase patient's tolerance; (3) early institution of spontaneous assisted ventilation (sb) and weaning from sedation and mechanical ventilation (mv) while on ecmo. objectives. to review our last 10 years lf-ecmo activity. methods. study period was january 2000-2010. lf-ecmo entry criteria were: potentially reversible acute hypoxemic respiratory failure, lis c3, no evidence of intracranial bleeding and no absolute contra-indications to heparinization. ecmo was performed with different type of heparin coated hollow-fiber artificial lungs. . we treated 31 patients (mean ± sd, 37 ± 18.5 years old, 68% males, bmi 27 ± 6, sofa 7.5 ± 3, oi 38 ± 14). 48% of these patients were placed on ecmo at other hospitals and transported to our icu by a dedicated ecmo team. ventilation days before ecmo were 12 ± 16 (range 1-69). before ecmo vt/kg was 6 ± 1.5 and rr was 33 ± 12: after ecmo beginning vt/kg was unchanged while rr decreased to 11 ± 4 (p\ 0.01). ecmo was set at bf 2.5 ± 1 l/min, gf 3.3 ± 1.5 l/min, fio2 0.98 ± 0. introduction. ventilating patients with acute lung injury (ali) in supine position potentially leads to an impaired pulmonary gas exchange. prone position (pp) is an attractive means to improve ventilation-perfusion (v/q) ratio [1, 2] but has several contraindications and showed no improvement in survival so far [3] . another therapeutical option is an upright position, which is easy to perform and has theoretical advantages over pp: the upward shift of the abdominal compartment is less pronounced, thus increasing thoracoabdominal compliance [2] . 4 however, to date regimes of an upright position did not tilt patients more than 45° [4] . 4 objectives. we hypothesised that a 60°standing position (sp) during mechanical ventilation may improve respiratory function. furthermore, we aimed to determine the feasibility of a sp for 2 h during mechanical ventilation. we studied 30 adult patients, receiving mechanical ventilation for more than 48 h in the intensive care unit of an university hospital. after recording baseline data, patients were placed in a 60°sp with the body entirely straight. further data sets were recorded during 2 h in sp, and after patients position was readjusted to supine position. functional residual capacity (frc) increased immediately after reaching sp (p \ 0.001) and remained elevated after repositioning to supine position. pao 2 /fio 2 ratio and compliance decreased initially during sp, but increased (p \ 0.05) after patients were retransferred to supine position. haemodynamic variables remained stable under a moderate increase of doses of catecholamines during the study period. conclusions. changes in respiratory function during sp are probably explained by a downward shift of the diaphragm due to gravitational forces 2 leading to an increased frc but not altering v/q ratio as demonstrated by the pao 2 /fio 2 ratio. after reaching the initial supine position the opening of the lung proved by the elevated frc is the predominant effect now associated with an increase in oxygenation as reflected by the pao 2 /fio 2 ratio due to an optimised v/q ratio. our results are confirmed in a subgroup analysis for 9 patients meeting ali criteria. ventilating patients in sp may be a new therapeutical approach to improve respiratory function in patients with ali. (1) . there are several clinical trials investigating the efficacy of the free radical scavenger n-acetylcysteine (nac) in ards, but its advantage remains uncertain. objectives. critically appraise and summarize all randomized clinical trials involving intravenous nac administration in adult patients suffering from ards. we included trials involving participants with ards according to the american-european consensus conference criteria (2) regardless of the underlying cause, and where one of the groups was treated with intravenous n-acetylcysteine in bolus intravenous doses or as continuous infusion, or combination of the two, and the other group was given placebo or standard treatment. conclusions. the main finding of this meta-analysis is that intravenous nac is ineffective in reducing mortality, length of stay or duration of mechanical ventilation in ards. we also found that late administration of nac may be associated with adverse outcome. the mechanism of this potentially deleterious effect remains unclear, but dosing and timing of nac appear to be critical issues. objective. to evaluate if extubation during ecls is harmful or beneficial. a 28-year-old woman was admitted to our intensive care unit (icu) after removal of a left ventricular assist device. this device was implanted as bridge to recovery for postpartum cardiomyopathy and ventricular function seemed to have recovered sufficiently. however, shortly after icu admittance she developed massive left and right ventricular failure. therefore a centrally cannulated veno-arterial ecls (maquet permanent life support) was implanted as a bridge to transplant. four days later she was extubated while on full ecls support, in order to reduce the risk of ventilator associated pneumonia. while on ecls, the patient was mobilized, practiced with an ergometer and chatted with her family. three days later the patient underwent cardiac transplantation. the postoperative period was characterized by temporary pulmonary failure, due to the combination of lung edema and atelectasis. eventually she made a full recovery. discussion. ecls provides a valuable means as bridge to transplantation, bridge to bridge or bridge to recovery. with the increasing use of ecls for circulatory failure, debate about the necessity of mechanical ventilation during this treatment ensues. ecls is usually applied under deep sedation and controlled mechanical ventilation. discontinuation of sedation possibly prevents intensive care acquired weakness. extubation during ecls may provide better pulmonary perfusion due to negative intra-thoracic pressure. furthermore, the awake and extubated patient is able to mobilize and exercise which may reduce the risk of atelectasis and ventilator associated pneumonia. our patient however developed pulmonary edema and atelectasis after discontinuation of ecls. the edema was probably a consequence of reperfusion injury, due to severely decreased pulmonary flow while on ecls. an absent ventilatory drive while on ecls may have led to hypoventilation while the patient was extubated, resulting in atelectasis. an extensive medline search resulted in one other case report describing an extubated patient on ecls. 1 intermittent non-invasive positive pressure ventilation was used to prevent atelectasis, but the patient developed pneumonia after 30 days of ecls. our patient was successfully extubated while on ecls. however, we conclude that there is insufficient evidence to recommend or oppose extubation of patients on ecls for circulatory failure. severe ards and refractory hypoxemia were defined with a pao2/ fraction of inspired oxygen (fio2) ratio of b100, or uncompensated hypercapnea with a ph of \7.20 despite receiving optimal conventional treatment. the ecmo can be used as a rescue treatment in these case. objectives. evaluation of severe ards treated with extracorporeal oxygenation (ecmo). all these ards were due to bacterial pneumonia or h1n1 influenza. over the last year (december 2009-january 2010), the recourse to extracorporeal oxygenation (ecmo) was used in ten patients with severe ards and severe hypoxemia. two groups were defined: bacterial pneumonia with ards (bp group, n = 5), and h1n1 influenza with ards (h1n1 group, n = 5). all ecmos were implanted at the bedside to facilitate intra-hospital or inter-hospital transfer, because of severe hypoxemia or hemodynamic instability making impossible patient mobilization before ecmo. results. data sets of 44 patients of 47 consecutive patients treated with ecmo were complete and included into analyses. we had no clinical or radiological evidence for thrombosis or clotting within ecmo-circuit with a target-ptt of 35 s. one patient with systemic aspergillosis died because of intracranial hemorrhage. one ecmo circuit had to be replaced due to insufficient oxygenator function after 10 days. further data are presented in tables 1 and 2 . conclusions. in this retrospective analysis of 44 patients who underwent ecmotreatment, ac with low-dose heparin (target-ptt of 35 s) was safe and without any observation of macroscopic thrombosis or clotting within the circuit. transfusion requirements and intracranial hemorrhage were low as compared with previous reports [1, 2] . therefore our data suggest that it is possible and safe using ecmo-therapy with low-dose heparin. introduction. in response to h1n1 pandemy, italy and lombardy created a national and a regional icu network, respectively, for treatment of ards patients. our hospital policlinico san matteo of pavia participated with a team for inter-hospital ecmo implantation and subsequent patient transport. objectives. description of the pavia ecmo team and activity analysis. methods. our team is composed by a cardiac surgeon, two intensivists, a perfusionist, an icu nurse, two emergency rescue technicians and a driver. all necessary aids for implantation and intensive care are ranged in three trolleys and three transport bags. equipments are firmly mounted on a two-level steel bridge connected to a spinal board. a portable ultrasonograph is also available. the ecmo team was alerted by the national call center. each mission used two ambulances, and in one case the ambulances were embarked on a hercules c130 j. from 31 october to 31 december 2009, four patients were implanted and transported, three suffering from h1n1 influenza (including a 190-kg body weight patient) and one from acute mitral valve rupture. all patients, already mechanically ventilated with maximal support, had veno-venous ecmo implanted by femoro-femoral percutaneous cannulation. the median mission duration was of 7.25 h (range 6-17 h). all patients were transported to our icu, where the median ecmo duration was of 8 days (range 6-10 days). no major managing issue occurred during the ecmo missions, and patient hospital survival was of 100%. a multispecialist team with good knowledge of ecmo can provide an effective support in severe respiratory failure, with ecmo implantation in peripheral hospitals and subsequent patient transport, thus realizing a fast and safe continuum between phone call activation and admittance to the reference center. introduction. when patients with sever respiratory failure are treated with v-v ecmo the right heart sometimes fails. this is a serious complication with a high mortality. in our unit these patients have been converted to v-a ecmo, although it is not fully agreed upon in the ecmo community due to previously depressing results. objectives. to evaluate the results of conversion to v-a from v-v ecmo in case of right heart failure. retrospective analyses of all patients with severe respiratory failure, treated between 1987 and 2009 at the karolinska ecmo centre. patients who were converted to v-a ecmo due to right ventricular failure were evaluated. a total of 214 patients (78 adults, 57 peadiatric, 79 neonatal) were treated on v-v ecmo for severe respiratory failure. of them 50 (33 adults, 9 peadiatric, 8 neonatal) needed conversion to v-a ecmo due to right ventricular heart failure demonstrated clinically by multiorgan failure and verified by echo cardiography. the survival after conversion to v-a ecmo was 10/33 (30%) in the adult age group, 7/9 (78%) in the peadiatric age group and 2/8 (25%) among the neonates. conclusions. given the high risk of fatality if not treated, conversion to v-a from v-v ecmo should be considered when the right ventricle fails. patients on v-v ecmo with right ventricle heart failure have very bad prognosis. it is concluded from the present results that conversion to v-a ecmo can save some of these patients. cardiac surgery and regional hemodynamics: objectives. to test whether tapse and right ventricular systolic (sm) and diastolic (em and am) tissue doppler imaging velocities are related with pulmonary artery systolic pressure (pasp) and length of the weaning process in mechanically ventilated patients with acute heart failure (ahf). methods. rv fractional area change (rvfac), left ventricular ejection fraction (lvef), pasp, tapse, sm, em, am rv tdi velocities, early diastolic mitral e wave and e 0 maximal tdi velocities of the mitral annulus at the lateral wall were obtained at admission by doppler echocardiography in a cohort of 32 patients with ahf, presented with pulmonary oedema, who required positive-pressure ventilation for more than 48 h in the intensive care unit (icu). echo-derived measures were compared between patients with and without pulmonary hypertension, whereas their association with duration of mechanical ventilation and length of the weaning process was tested with multivariate linear and logistic regression analysis. and increased e/e 0 ratio (10.8 ± 1.21 vs. 7.6 ± 0.54, p \ 0.001) compared with subjects with normal pasp (n = 10). these variables were negatively associated with duration of mechanical ventilation (r 2 = 0.55, beta slope = -0.89 for tapse, r 2 = 0.52, beta = -0.57 for sm, r 2 = 0.45, beta = -0.27 for em/am, p \ 0.001) and were proven to successfully discriminate patients with (n = 12) and without (n = 20) prolonged weaning ([7 days of weaning after the first spontaneous breathing trial failure, p \ 0.001 for all comparisons). conclusions. we suggest that in critically ill patients with ahf presented with pulmonary oedema, low tapse and rv tdi velocities upon admission are associated with pulmonary hypertension and prolonged length of the weaning process. objectives. the aim of the study was to study changes in cerebral blood flow (cbf), as determined by tcd, during the early postoperative course of cvs and to correlate such changes with post-operative nc. we studied 72 patients undergoing extracorporeal circulation cvs (coronary by-pass, valve replacement or both) between march 2007 and march 2008. cbf was assessed by measuring bilateral mca flow velocities by tcd before and 10, 24 and 48 h after cvs. changes c20% between consecutive tcd results were considered significant. demographic and clinical variables, co morbidities, euroscore, sofa, type and duration of surgery and type and severity of nc were also recorded. patients were assigned to 3 groups according to cbf changes from baseline: a) changes b20%; b) cbf increases c20%, c) cbf decreases c20%. nc were classified as major (stroke, tia and coma) and minor (delirium, encephalopathy, transient cognitive impairment). we used descriptive statistics and inference by v 2 , anova and pearson's correlation. of the 72 patients, 20 were excluded (2 early post-operative death and 18 due to technical difficulties or incomplete tcd recordings). of 52 evaluable patients, 17 (33%) had no cbf changes (group a), 12 (22%) had increases c20% (group b) and 23 (45%) had decreases c20% after cvs (group c). a positive correlation was found between cbf changes and duration of circulatory arrest (p \ 0.02), maximum sofa score (p \ 0.001), respiratory dysfunction (p \ 0.016) and duration of mechanical ventilation (p \ 0.001). neurological complications occurred in 16 patients (31%), of which 4 (25%) were major and 12 were minor (75% introduction. the sole monitoring of macrohemodynamic variables is not always sufficient in the early detection of tissue hypoperfusion, especially in cardiac surgical patients that frequently present with microcirculatory derangements. near infrared spectroscopy (nirs) is an easily applicable non invasive technique that has been used to provide an estimate of tissue oxygenation at the bed side. objective. the aim of our study was to evaluate the effect on outcome of guiding hemodynamic therapy and specifically inotrope titration in cardiac surgical patients postoperatively with nirs. methods. patients operated on with cardiopulmonary bypass were assigned, after stratified randomization (gender, euroscore-cutoff of 6), to an intervention (ig) and a control group (cg). postoperatively, following cardiac intensive care (cicu) admission, after initial resuscitation according to cicu protocol, sto 2 (%) was measured in patients of the ig in 3 muscle sites: thenar, masseter and deltoid. if it was less than 80% in 2/3 sites, dobutamine was administered in incremental doses (2.5 lg/kg/min), with the sto 2 (%) measured every half hour. the interventional period began upon cicu admission and lasted for 6 h, after which both groups were treated according to cicu protocol. primary outcome measured was the oxygen consumption rate at the end of the 6 h intervention period as assessed with nirs vascular occlusion technique. . 18 patients were included in the study (8 in the intervention group and 10 in the control group). the 2 groups did not differ statistically significantly regarding age, euroscore, and macrohemodynamic variables postoperatively (with the exception of cvp). microcirculatory parameters upon admission to the cicu also did not differ, excluding masseter sto 2 (%). the oxygen consumption rate and the reperfusion rate increased in the 6 h study period in both groups, without differing statistically significantly between the groups at any time point (cg oxygen consumption rate 20.1 ± 12.4 upon cicu admission and 32.8 ± 15.5 6 h later, ig 15.4 ± 6.6 and 19.7 ± 7.7 respectively) (cg reperfusion rate 297 ± 210 upon cicu admission and 592 ± 254 6 h later and ig 201 ± 119 and 304 ± 163 respectively). as far as outcome parameters were concerned, the 2 groups did not differ statistically significantly in the total hours and total dose of vasopressors ± inotropes received, in the hours of mechanical ventilation, in the duration of cicu or hospital stay, and in sofa scores the days following the operation. conclusion. nirs guided titration of inotropes did not lead to a greater improvement in the microcirculation 6 h postoperatively, or to a better outcome. the limited power of the study prevents definite conclusions on the role of nirs in hemodynamic therapy in cardiac surgery patients. objectives. to estimate the prevalence of pulmonary embolism among mv patients in icu and its association to deep vein thrombosis (dvt). in a monocentric prospective observational study, we included all the patients requiring mechanical ventilation with no previously diagnosed pe, who underwent a thoracoabdominal ct contrast scanner for any medical reason. we used a modified protocol for pe diagnosis with a 64-multidetector row ct scan read by two independent radiologists. the association with a dvt was explored by performing venous compression ultrasound of four limbs. objectives. the aim of this animal study was to evaluate the effect of intraabdominal hypertension on left ventricular diastolic function. after approval by an institutional animal care committee, 7 rabbits were anesthetised before mechanical ventilation. an intraperitoneal infusion of 1.5% glycine solution was used to increase intraabdominal pressure to 20 mmhg. the right common carotid artery was catheterised in the neck in order to introduce a millar mikro-tip catheter (millar instruments inc., houston, usa) into the left ventricle. heart rate, arterial pressure, central venous pressure, oesophageal pressure and intraabdominal pressure were measured. the s time constant of relaxation which is considered as best index of relaxation was calculated using the derivative method (1). all haemodynamic measurements were registered at baseline and after inducing intraabdominal hypertension. data are presented as mean (iqr) and were compared using a wilcoxon rank sum test. results. heart rate (from 202 ± 100 to 166 ± 100 beat/min, p = 0.6), mean arterial pressure (from 60 ± 29 to 50 ± 34 mmhg, p = 0.20) and dp/dt max (from 3,590 ± 255 to 2,111 ± 197 mmhg/s, p = 0.14) were not significantly modified by intraabdominal hypertension. however, the s time constant of relaxation increased significantly (from 16 ± 3 to 43 ± 18 ms; p = 0.05). conclusions. in this animal model, intraabdominal hypertension impairs left ventricular relaxation. these changes in the condition of the microcirculation have been related to the degree of organ dysfunction and thus patient outcome ie hospital length of stay. near infrared spectroscopy (nirs) is an easily applicable non invasive technique that has been used to provide an estimate of tissue oxygenation at the bed side. objectives. the aim of our observational study was to examine whether impaired tissue oxygenation as assessed with nirs immediately postoperatively correlates with hospital length of stay. patients undergoing a planned cardiac surgical procedure on cpb were included in the study. patients' thenar tissue oxygenation (sto 2 %) was assessed with nirs postoperatively in the cardiac intensive care unit (cicu). results. 22 patients undergoing cardiac surgery on cpb (13 male/9 female) (age: 64 ± 14 years, euroscore: 5.7 ± 3; mean ± sd) were enrolled in the study. patients length of stay was 8.5 (3-26); median(range). the haemodynamic parameters of our patients upon admission to the cicu were: map 84 ± 12 mmhg, cvp 9 ± 4 mmhg, pcwp 12 ± 4 mmhg, mpap 24 ± 5 mmhg, ci 2.6 ± 1.1 l/min/m2, svr 1407 ± 501 dyne x s/ cm5, pvr 239 ± 106 dyne x s/cm5, hr 97 ± 15 bpm, hb 11.7 ± 1.7 g/dl, lactate 2.8 ± 1.8 mg/dl; (all variables expressed as mean ± sd). upon admission to the cicu all patients were mechanical ventilated, under vasopressor ± inotrope support and their central temperature was 36.9 ± 0.68; mean ± sd. the thenar sto 2 % was 80 ± 11; mean ± sd. thenar sto 2 % correlated statistically significantly with hospital length of stay (r = 0.53, p = 0.11). discussion. tissue oxygenation as assessed with nirs reflects the balance between regional oxygen delivery in relation to oxygen utilization. an elevated sto 2 in the presence of normal macrohemodynamics may reflect impaired oxygen consumption and thus an impaired microcirculation. conclusion. patients with impaired tissue oxygenation immediately postoperatively have a longer hospital length of stay. further studies are needed to confirm these results and to investigate the potential benefit from incorporating this information regarding tissue oxygenation in the treatment algorithm. objectives. the goal of this study was to compare two different sedative agents for implantation of crt-ds related to incidence of adverse events and patient's satisfaction. methods. the study included forty-two, asa iii-iv patients, undergoing transvenous implantation of crt-ds under local infiltrative anesthesia with 20 to 30 ml of 1% lidocaine. intraoperative sedation was established with intermittent boluses of midazolam (1-5 mg) to achieve desirable level of sedation. before the induction of ventricular fibrillation in order to test the defibrillator function of the crt-d device, patients received an additional bolus of either propofol (1.5-2.5 mg kg -1 , p group, n = 20) or etomidate (0.1-0.25 mg . kg -1 , e group, n = 22) targeting bis values in the range 50-60. the incidence of apnea, hypotension, nausea, myoclonus, pain at injection site, allergic reactions as well as patient's satisfaction with anesthesia described as feel of well being were registered and compared between groups. results. in 23 subjects (56%) no complications were recorded. myoclonus was registered in 5 patients from e group (23%) and in none from p group (p \ 0.05). no patients receiving etomidate reported pain at injection site compared to 4 patients (20%) receiving propofol (p \ 0.05). there was no significant difference in incidence of apnea between two groups (15 vs. 9%, p = 0.30). two patients in p group (10%) and 1 in e group (4.5%) became hypotensive after delivering the hypnotic agent (p = 0.36). also, there was no statistically significant difference between groups considering the frequency of nausea (0% vs.9%, p = 0.26). all the patients whom propofol had been delivered (100%) reported feel of well being and only four of them filed the same after etomidate (18%) (p \ 0.01). no allergic reactions and major adverse events were registered. conclusions. implantation of crt-ds and its testing can be successfully performed with administration of both propofol and etomidate as a safe procedure with low per operative morbidity and shorter complication rates. still, treating with propofol tends to be more satisfactory for the patients. introduction. ultra-short-acting b 1 selective adrenergic antagonists are now widely used to control tachycardia and tachyarrhythmia perioperatively. among them, landiolol, a new ultra-short-acting b 1 -blocker, has been reported to exert a more potent negative chronotropic effect with little effect on blood pressure than esmolol (1). however, detailed mechanisms underlying different cardiovascular actions are still unknown. objectives. in this study we evaluated direct effects of landiolol on cardiac performance and single cell electrophysiology in comparison to those of esmolol. methods. the present study composed of two parts. the first part of the study used isolated guinea-pig hearts which were perfused in the langendorff mode at constant flow with oxygenated tyrode solution at 37°c. the coronary perfusion pressure (cpp) was continuously monitored throughout the experiment, and intrinsic heart rate (hr) and isovolumetric left ventricular contraction were measured with a thin saline-filled balloon inserted into the left ventricle. the second part of the study was to measure action potentials and ionic currents in ventricular myocytes isolated enzymatically from guinea-pig hearts. comparison of data was conducted by repeated-measure anova with post hoc test (bonferroni's correction). conclusions. esmolol had a more potent negative inotropic effect than landiolol. this effect is, at least in part, derived from shortening of apd. in addition, increase of the coronary resistance would facilitate the negative chronotropic action of esmolol in vivo. conclusions. nma moderates hpv in the conscious spontaneously breathing beagle, but not to the same degree as acz. as compared to acz, the additional methyl-group in nma may impair its capability in vivo to act on a non-ca acz-sensitive cellular receptor or channel or that both, ca-dependent and ca-independent actions of acz yield a greater effect. introduction. tee with bubble test is considered as the ''gold standard'' method to detect a pfo with right to left shunt. tcd is a non-invasive method which has been shown to be as accurate as tee for pfo detection. we conducted a multicenter trial to estimate the prevalence of pfo, the influence of the size of the heart chambers on the prevalence of pfo and the accuracy of tcd as a non invasive method for pfo detection in mechanically ventilated icu patients. one hundred icu patients (74 m and 26 f) under mechanical ventilation who needed a tee study for hemodynamic assessment were included in the study. in each patient, the presence of a pfo was detected by tee and tcd. three bubble tests with agitated haemacel ò were performed by each method, with tee probe at 608 and 908 rotation and with tcd the gate of pulse wave doppler (pwd) at the m 1 segment of the middle cerebral artery (mca). patients without temporal acoustic window to perform tcd were excluded from the study. the size of pfo was classified as grade i, ii and iii according to the number of microbubbles passing from the right to the left atrium and the number of hits (high intensity transient signals) detected with pwd in the mca (grade i: \6 microbubbles or hits, grage ii: [6 and \25 and grade iii: more than 25 microbubbles or hits). for each patient included in the study we measured and correlated the presence of pfo with the tidal volume (v t ), the plateau pressure (p plat ), the compliance of the respiratory system (c rs ) and the size of the right (rv) and left (lv) ventricle. results. mean p a o 2 /fio 2 was 198 (min 50, max 350), mean c rs was 39 ml/cmh 2 o (min 11, max 72), mean v t was 574 ml (min 260, max 980) and mean p plat was 22 cmh 2 o (min 14, max 37). the prevalence of pfo detected with tee was 28% and with tcd 48%. there was no pfo detected with tee and missed by tcd. tcd was more sensitive than tee in detecting pfo of grade i (7 with tee, 17 with tcd) and ii (6 with tee, 16 with tcd), while for grade iii the two techniques had equal sensitivity (15 with tee, 15 with tcd). no correlation was found between p plat , c rs , v t and the presence of pfo. on the contrary, a strong correlation was found between rv dilatation and the presence of pfo (p \ 0.001). conclusions. the prevalence of pfo detected by tcd is very high in mechanically ventilated icu patients and this may have important clinical implications. tcd is more sensitive than tee in detecting a small pfo. the presence of rv dilatation increases the prevalence of pfo. objectives. the aim of our study was to identify in mechanically ventilated patients for ali/ards the prevalence of pfo and to evaluate the factors that may influence the prevalence of pfo. methods. two groups of mv patients, one with ali/ards and one without respiratory failure (rf), were enrolled in the study. all patients underwent a tee study for hemodynamic assessment. in each patient three consecutive bubble tests with agitated haemacel ò were performed at 60 0 and 90 0 rotation of the tee probe. the bubble test was performed through a central line in the inferior or superior vena cava (ivc, svc). a pfo was diagnosed by the presence of microbubbles in the left atrium within five cardiac cycles following the injection. furthermore, in ali/ards patients in whom a pfo was not detected at baseline mv, three consecutive bubble tests during recruitment maneuver at 45 cmh 2 o for 20 s were performed. the compliance of the respiratory system (c rs ), blood gas exchange and the ventilatory settings (p plat , v t ) were recorded in both groups. o, respectively. the presence of rv dilatation was a strong predictor for the fo opening (p \ 0.01); on the contrary, no statistical significant difference was found between the site of injection (svc vs. ivc), the c rs , v t , and p plat and the presence or absence of a pfo. a high prevalence of pfo was found in ali/ards patients. rv dilatation seems to be the reason of this high prevalence. rv dilation may be due to the lower c rs and higher p plat of the ards patients. introduction. the clinical evaluation of arterial tone is mainly based on the calculation of total systemic vascular resistance (tsvr). however, given the pulsatile nature of arterial flow, this parameter provides an inadequate assessment of vascular tone. another approach proposed would take account of changes in pulse pressure and blood flow, relationship known as arterial elastance (ea). so, for a given stroke volume, the blood pressure generated in the circulatory system will depend on ea (1). to assess the ability of the dynamic arterial elastance (ea dyn ), defined as the relationship between pulse pressure variation (ppv) and stroke volume variation (vvs), to predict the hemodynamic response in mean arterial pressure (map) to a increase in stroke volume (sv) in hypotensive preload-dependent patients with acute circulatory failure. we performed a prospective clinical study in a 17-bed multidisciplinary intensive care unit, including 21 patients with controlled mechanical ventilation and monitored with the vigileo ò monitor, for whom the decision to give fluids was taken due to the presence of circulatory, including arterial hypotension (map b 65 mmhg or systolic arterial pressure \90 mmhg), and preserved preload-responsiveness condition, defined as svv c10%. dynamic arterial elastance (vpp/vvs ratio), arterial pulse pressure to sv ratio, map/sv ratio, tsvr and map were compared to predict a map increase c15% after volume expansion (map-responders). results. at baseline, only ea dyn was significantly different between map-responders and nonresponders. ve-induced increase in map was strongly correlated with baseline ea dyn (r 2 = 0.82, p \ 0.0001) and changes in ea dyn after ve (r 2 = 0.77; p \ 0.0001). the only predictor of map increase was ea dyn (auc 0.98 ± 0.03; 95% c.i.: 0.8-1). a baseline ea dyn value [0.89 predicted an increase c15% in map after fluid administration with a sensitivity of 92.9% (95% c.i.: 66.1-99.8%) and a specificity of 100% (95% c.i.: 59-100%). conclusions. dynamic assessment of arterial elastance by pvv to svv ratio during controlled mechanical ventilation could be used to predict mean arterial pressure increase after volume loading in hypotensive preload-dependent patients. severe sepsis is one of the major reasons for intensive care unit (icu) admission and leading causes of mortality. some of these score systems have been customized for patients such as apache ii, apache iii, sasp ii and mods. this study is to assess the validity of mortality prediction systems in severe septic patients. objectives. the aim of this study was to compare and evaluate four severity scoring systems in intensive care unit (icu), including apache ii, apache iii, sasp ii and mods in severe septic patient. methods. fifty-six severe septic patients were divided into two groups. one was survival group and the other was non-survival group. besides general data, the continuous surveillance of apache ii, apache iii, sasp ii and mods were recorded by 1st, 3rd and 7th day. results. compared with survival group, mods was significant difference in non-survival group only in 1st day (7.9 ± 3.1 vs. 10.1 ± 2.9, p \ 0.05) but apache ii, apache iii and sasp ii were significant difference through 1st, 3rd and 7th day(p \ 0.05). in seven-day comparison, p value of apache iii in non-survival group was the minimum (p = 0.000) and p value of mods was the maximum (p = 0.008). in optimal survival evaluation, it seemed that apache iii was the best (apache iii [ apache ii = saspii [ mods). conclusions. in order to evaluate the critical condition and prognosis of severe septic patients, apache iii was the best and apache ii and sasp ii were followed and mods was the worst. objectives. to assess compliance with the cem standards for management of severe sepsis across three ed sites in the west midlands. methods. data was collected retrospectively over 3 months. patients presenting to the ed within this period were assessed for likelihood of severe sepsis by the diagnostic code given to each patient upon leaving the ed. data was analysed using a scanned copy of the ed clerking. patients' notes were assessed for sirs criteria and signs of new infection. if these criteria were met, and organ dysfunction was present, they were included in the audit. results. 255 patients with severe sepsis were identified. of these 17% were documented as septic by ed staff. the cem standards of care were received in 41% of patients with a documented diagnosis of severe sepsis in the ed, and 23% of patients overall. 89% of patients received the 'treatment' aspects of care: oxygen, iv antibiotics (with blood culture) and iv fluids. 71% of severely septic patients had no documented consideration of icu referral. conclusions. early recognition of severe sepsis in the ed led to greater performance in meeting the cem standards. although 100% of patients received observations and 89% received the treatment interventions, we performed poorly in meeting the remaining cem standards. the trust has developed a severe sepsis proforma which incorporates the cem standards to accurately record the completion of each intervention. a sepsis course for staff has been launched trust wide, and a formal referral process to icu for all severely septic patients is being implemented. objectives. to observe association of body temperature (bt) and antipyretic use with mortality in the critically-ill. a prospective multi-national, multi-center observational study. consecutive patients whose icu stay were expected to be more than 48 h were recruited from 15 centers in japan and 10 centers in korea. patient's bt was prospectively recorded every 4 h until patient's death, discharge from the icu or up to 28 days. information including patient's clinical characteristics at admission, presence of infection, and use of steroids, extracorporeal circuit, and antipyretics were recorded. (1). while blood culture results take time, treatment for bloodstream infection should be provided swiftly, usually before results are available (2) . prior treatment with antimicrobials increases the chances of false negative results. haste, poor technique and alteration in commensal flora may increase the chances of falsely identifying pathogens. objectives. we have investigated the utility of blood culture tests in our general critical care unit over 1 year in terms of results yielded and actions prompted. methods. the indication for blood culture was clinician's discretion. all critical care sourced blood cultures for the period oct 2008 to sept 2009 were reviewed from the microbiology laboratory database. blood culture specimens were collected in bact/alert ò bottles (biomerieux, durham, nc, usa 27704). notes review was made of the positive blood culture episodes to determine actions after the results were known. consideration was given to the source of the blood sample: clean stab versus from an intravascular device. categoric data was analysed using the chi-squared test and p value of 0.05 was accepted as significant. objectives. we hypothesized that in the emergency department of our hospital many patients with sepsis are not recognized as such. methods. in a retrospective design, patients of an age of 18 years and older who were admitted to the emergency department during a period of 3 months between january-april 2008 and diagnosed as having an infection were included. the diagnose infection was made on admission by the emergency department nurse. the included patients were either classified as having sepsis or not having sepsis, according to the sirs criteria. conclusions. h1n1 infection was associated with significant morbidity and mortality. it occurred mainly in young pts with co-morbidities and was associated with severe hypoxemia, a trigger for prolonged mechanical ventilation and frequent use of lung rescue therapies. a significant delay in hospital admission and start of antiviral therapy should also be noted. admission to administration time difference between cycles was 1.26 h, with a mean reduction of 1.04 h between clinician assessment and prescription time in cycle two. we identified delays against the standard after both cycles of the audit. we demonstrated that the method of prescription should be taken into consideration when prescribing antibiotics in patients with suspected sepsis. there are a multitude of factors that could contribute to a reduction in the clinician assessment to prescription time, which may be investigated in further audits. conclusions. despite high levels of resistance among psa and ab from these icus, cfr for most carbapenem dosing regimens were above the reported susceptibility. doripenem provided greater cfr than meropenem, which was superior to imipenem against these isolates. while higher doses combined with prolonged infusions significantly improved cfr against psa, alternative therapeutic strategies will be required to address these highly resistant ab. grant acknowledgment. the passport study is supported by a grant from janssen-ortho-mcneil. introduction. drug interactions are common, and the effects of these interactions can range from innocuous to deadly. critically ill patients often receive a variety of potent drugs, including antimicrobials, making this population extremely susceptible to drug-drug interactions. therefore, physicians must be familiar not only with the antimicrobial drugs capable of producing adverse drug events, but also their potential drug-drug interactions. there are scarce data about the incidence of these types of drug interactions and the how frequently it might cause adverse events. objectives. the purpose of this study is to evaluate the incidence of potential drug interactions involving antimicrobials and the possibility to cause adverse events. the clinical pharmacist has prospectively analyzed icu prescriptions between january 2009 and december 2009 with the purpose to identify potential drug-drug interactions involving antimicrobials. the screening was done with the relief from a software (epocrates rx ò drug reference). the interactions detected were classified in eight groups according to the affected system (neurological, cardiovascular, gastrointestinal, renal, endocrine, hematological, musculoskeletal and others) and through the type of interaction (pharmacokinetic, pharmacodynamic and others). we have identified the most common potential effects, the medications involved and have observed the incidence of adverse drug events. results. the icu admitted 347 patients during the study period. we have analyzed 661 physician orders with 8209 prescribed items. we have identified 871 antimicrobial drug interactions (71 different interactions) which compound 20% of the total drug interactions (n = 4349). the cardiovascular system and the pharmacokinetic interaction were the most potentially affected (38%; 42%). the most common medications involved were: fluconazole (24%), clarithromycin (23%), levofloxacin (12%); linezolid (11%). the clinical pharmacist has made an intervention regarding medication safety in 3% (n = 27) and the acceptance rate by the medical icu staff was 74%. we have not been able to identify any adverse drug event caused by drug interaction even with our active search and the spontaneous reports. however, sub notification must be taken into consideration. conclusions. clinicians should be aware of potential drug-drug interactions when making therapy selections for critically ill patients. antimicrobial drugs are susceptible to interact with other drugs, which may increase the risk of adverse drug events. the clinical pharmacist interventions may improve clinical outcomes by optimizing medication use, monitoring potentially preventable adverse drug events and promoting information about this important issue to the icu multi-professional team. introduction. cefazolin is one of the most frequently administered antimicrobial agent for prophylaxis in ''clean'' surgery. its broad spectrum against gram + micro-organisms and its pharmacological characteristics make it an easy-to-use choice to prevent infections caused by staphylococcus aureus and coagulase-negative stapylococci. objectives. the aim of this study is the evaluation of the plasma concentrations of cefazolin administered as a prophylactic antimicrobial agent during cardiac surgery with cpb. adequate cefazolin plasma levels can maintain a tissue concentration high enough to prevent the risk of developing post-operative infections. after obtaining ethical committee approval and personal written consent, two groups of patients were enrolled in this prospective study. the first group, 12 patients, received cefazolin, 2 g, 30-60 min before skin incision and 1 g adjunctive dose after 4 h. then, three 2 g doses were administered every 8 h. in the second group of 12 patients the adjunctive 1 g cefazolin dose was given at the beginning of the cpb. blood samples were collected immediately before the first dose and every hour for the whole time of surgery, and, only in the second group, after surgery, at 12th, 18th and 24th hour. plasma cefazolin concentration was determined with a biological radial diffusion assay. results. plasma cefazolin was constantly higher than the mic90 of the most involved micro-organisms (according to clsi). in the first group, cefazolin concentration suddenly decreased after starting cpb. the 1 g adjunctive dose immediately restored it. the earlier administration of this dose in the second group prevented this sudden fall. plasma cefazolin was maintained at effective inhibitory levels for the whole time of surgery in all patients ([32 mcg/ml). during the postoperative period cefazolin decreased slowly, but inhibitory plasma levels were always maintained. the rate of cefazolin clearance was found equal to the creatinine clearance in all patients. perioperative plasma cefazolin concentration conclusions. the administration of cefazolin 2 g every 8 h can guarantee effective inhibitory plasma concentrations during surgery and during the first 24 h after surgery. cpb causes a sudden fall in cefazolin plasma levels. this can be avoided administering an adjunctive 1 g dose immediately before starting cpb. objectives. vancomycin dose regimen was adjusted based on trough plasma levels in burn patients that were distributed according to the extension total burn surface area (tbsa); also pharmacokinetics changes were compared. methods. twenty seven adult burn patients of both sexes, requiring antimicrobial therapy with vancomycin for the control of sepsis were investigated. pharmacotherapeutic follow up was performed in a serial of periods (112 observations) for all patients investigated by collection of blood samples, 1 ml each from the venous catheter as follows: 1st blood sample collection, 2 h after the beginning of drug 1 h infusion and a 2nd sample blood collection at the trough, immediately before the next dose. if necessary, additional sample blood collections were performed based on the laboratorial data for patients any time, for dose adjustment purpose and optimization of drug therapy. vancomycin plasma concentrations were determined by highperformance liquid chromatography. plasma curve decay was plotted, and pharmacokinetics was analyzed by one-compartment open model against the reference data reported. results. burn patients receiving the empiric dose regimen showed trough plasma level lower than the minimum effective concentration, consequently dose adjustment was required. vancomycin adjusted dose regimen showed statistical significance differences according to tbsa (p \ 0.05) as follows for daily dose normalized to body weight and expressed by mean ± sd: 25.5 ± 11.9 mg/kg/day were required for patients with tbsa below 20%, 32.0 ± 6.4 mg/kg/day for tbsa 20-40% and 34.8 ± 12.2 mg/kg/day were required for tbsa above 40%. relevant changes on pharmacokinetics were observed by drug plasma clearance increased according the increase of tbsa (p \ 0.05), while the apparent volume of distribution and also the biological half-life remained unchanged. additionally, a weak correlation was observed between vancomycin plasma clearance and creatinine clearance (r 2 = 0.36; p = 0.0004), probably due to the contribution of the extra-renal clearance on total drug elimination. on the basis of data obtained in the present study and to prevent therapeutic failure and also to reduce the risk of bacterial resistance, dose adjustment in burn patients is recommendable based on vancomycin plasma monitoring and also on the extension of total burn surface area. introduction. the importance of early antibiotic therapy has been recently demonstrated. regarding a rapidly increasing number of obese patients, appropriate drug dosage in these patients is an important challenge of critical care since it has been shown that not only early start of antibiotics but also correct target concentrations decrease mortality. vancomycin is administered according to body weight (bw). nevertheless, little is known about the percentage of obese patients achieving pre-defined target serum levels within 24 h after initiation of vancomycin therapy compared to patients with normal bw. objectives. therefore, it was the aim of our study to analyze the appropriateness of serum vancomycin levels in patients with a bw between 40 and 150 kg. vancomycin is almost entirely excreted by the glomerulus and may be responsible for nephrotoxicity [1] . however, there is a lack of definitive evidence linking concentrations to either outcome or toxicity [2] . few reports exist comparing intermittent dosing and continuous infusion. ingram [3] suggested that whilst associated with a slower deterioration in renal function, there was no difference in the prevalence of nephrotoxicity. similarly, hutschala [4] demonstrated worsening creatinine in 149 patients following cardiac surgery with both intermittent and continuous infusion but infusion tended to be less nephrotoxic despite receiving higher doses. we wish also to report our experiences with vancomycin infusion in critically ill cardiac patients. methods. we examined retrospective data from 2,512 patients treated with vancomycin. we perform adjusted and un-adjusted analysis using sofa on the day of starting vancomycin and total dose received. to assess the differences in either an initial pulmonary or non pulmonary presentation. methods. prospective, observational, multi-center study conducted in intensive care (icu). we reviewed demographic and clinical data for all pandemic h1n1 influenza a infections reported in the esicm h1n1 registry. results. 512 patients were screened from the registry. 330 patients with completed data entry for pulmonary and non pulmonary with outcomes were identified and analysed. all patients had either suspected, probable or confirmed pandemic h1n1 influenza a infection and were being cared for in an icu. 53% of the patients were male with a median age of 43 (iqr 32-55) years. the admission mean saps3 score was 54 ± 16 and the apache ii score was 21 ± 9. 33% of the patients subsequently received non invasive ventilation and 72% received invasive mechanical ventilation. the icu mortality rate was 30%. the hospital mortality was 32%. 77% of patients presented with a pulmonary presentation. 73% of these were admitted with ards and/or bacterial pneumonia and 22% with an acute bronchospastic exacerbation. 23% of patients were admitted to the icu with a non pulmonary presentation. the main reasons for admission in these patients were: cardiovascular instability (33%), altered level of consciousness (20%), renal failure (11%) and acute coronary syndromes (5%). patients with a pulmonary presentation were older, had a increased history of asthma or copd and were more likely to be ventilated. they had a higher mortality rate in the icu. non pulmonary presentations were more likely to suffer from chronic renal impairment. a total of 512 episodes of pandemic influenza a (h1n1)v infections in critical care setting were analyzed: 169 with bacterial pneumonia (94 males and 73 females) and 343 with wheezing or viral pneumonia (182 males and 161 females). the mean age was 45 (± 17) years in patients with bacterial pneumonia and 40 (± 17) in patients viral pneumonia. the mean apache ii score was 25 (± 9) and 20 (± 9), with a corresponding probability of death of 37 (± 25)% and 24 (± 21)%. comorbidities were common, but without significant differences between the two groups (only exceptions pregnancy-more prevalent in patients without bacterial pneumonia-and dialysis dependence-more prevalent in patients with bacterial pneumonia). at icu admission shock and acute renal failure were more common in patients with bacterial pneumonia. in patients without pneumonia; severe hypoxia and ards did not presented significant differences between groups. aims. evaluation if an isocaloric beginning of artificial nutrition in critically ill medical patients is associated with increased nutritional related side effects compared to a hypocaloric start. methods. 110 critically ill medical patients with an expected need for artificial nutrition of [5 days were included into this prospective, randomized clinical study. artificial nutrition was started either isocalorically right from the beginning (group a; n = 55) or hypocalorically (50% of the energy demands) followed by a stepwise increase over the next 2 days (day 2: 75%); day 3: 100%) (group b; n = 55). nutrition related side-effects were defined as the occurrence of hyperglycemia, hyperlactatemia, hypertriacylglycerolemia, upper digestive intolerance, cholestasis, or diarrhea as well as disturbances of serum electrolytes and were assessed on a daily basis. patients were randomized to receive either an artificial nutrition started isocalorically (group a) or hypocalorically followed by a stepwise increase (group b). of the 150 patients, 100 patients completed the study (group a: n = 55; group b: n = 55). the calculated, cumulative energy requirements of patients of group a and b were 9,058 ± 1,236 and 9,125 ± 1,569 kcal, respectively (p = ns). patients of group a received 76 ± 17% and patients of group b 69 ± 12% of the calculated energy requirements (p \ 0.01). the incidence of nutritional related side effects was not different comparing both groups, except for hypophosphatemia, which was more pronounced in group a. additionally, exogenous phosphate needs were higher in patients of group a. the number of interruptions of the artificial nutrition did not differ between groups. conclusions. an isocaloric start of artificial nutrition provided more energy during the first 5 days of their icu stay than a hypocaloric beginning. there was no difference in the number of interruptions and in the incidence of nutritional related side effects, except hypophosphatemia suggesting the presence of refeeding syndrome. in studies carried on to demonstrate positive effects of glutamine (gln) that has innumerable biological features, the main point of discussion isn't whether gln has positive effects in sepsis but rather the effect difference between different administration routes. only enteral (en.) or parenteral (pn.) administration was analyzed in this respect and no studies on combined administration were performed. the primary endpoint in this study was to analyze the effects of administration of en. and pn. gln together or separately on intestinal mucosa + immune system in the experimental sepsis model. for this purpose villus atrophy, bacterial growth in blood and tissue, levels of blood gln, tnfa and il10 were examined. the secondary endpoint was to evaluate the different administration models in terms of cost. wistar, adult female rats were used. they were fed standard. sepsis was developed in 4 groups (all rats) by injection of intraperitoneal(ip.) 1 ml (5 9 10 7 cfu/ml) e. coli. grup c (n = 12):en./pn. isotonic saline (1 ml/day; 2 ml/d); grup e (en., n = 10):en. gln (0.5 g kg -1 day -1 ) + pn. saline (2 ml/d); grup p (pn., n = 11):pn. gln (0.4 g kg -1 day -1 gln) + en. saline (1 ml/d); group ep (en. ± pn., n = 11):pn. gln (0.3 g kg -1 day -1 ) ala-gln = 0.2 g kg -1 day -1 gln) + en. gln (0.3 g kg -1 day -1 ); were administered. feeding of rats began 4 h (h) after administration of ip e. coli. blood gln (with spectrophotometer), tnfa and il10 concentrations(with elisa) were examined at the start (baseline levels) and at 24-96 h after the experiment started. samples of tissue from mesenteric lymph node, liver, lung, blood and small intestine were collected. ala-gln = 0.27 g kg. . rates of reproduction of the strain administered were found lower for group ep than group c (p \ 0.05). rates of villus atrophy in ileum of group ep, p and e were lower than group c (p \ 0.05).plasma gln levels were found lower in groups ep and p at 24 h, and higher at 96 h than other groups (p \ 0.05). when plasma gln levels at 96 h were compared with their baseline levels, significant increases were detected in groups ep and p and significant decreases were detected in groups c and e (p \ 0.01). serum tnfa and il10 levels were found lower for groups ep and p at 24 and 96 h when compared between groups (p \ 0.05). when serum tnfa and il10 levels at 96 h were compared with their baseline levels, more distinctive increases were detected in groups c and e than other groups (p \ 0.01). significant positive correlation was determined between tnfa and il10 levels at 24 h (p \ 0.01) and 96 h (p \ 0.01). cost of simultaneous administration of en. and pn. gln was higher than en. administration but close to pn. administration at these doses. methods. medline and embase were searched. hand citation review of retrieved guidelines and systematic reviews was undertaken and academic and industry experts were contacted. only methodologically sound randomised controlled trials (rcts) were eligible for inclusion in the primary analysis. the primary analysis was conducted on clinically meaningful patient oriented outcomes, which included mortality, functional status and quality of life. secondary analyses considered vomiting/regurgitation, pneumonia, bacteremia, sepsis and multiple organ dysfunction syndrome. meta-analysis was conducted using the peto analytic method, which is known to minimize bias in the presence of sparse events. the impact of heterogeneity was assessed using the i 2 metric. results. 4,217 unique abstracts were identified, resulting in the retrieval of 701 papers for detailed eligibility review. four rcts were identified to be on topic however one rct reported excessive loss to follow-up such that an intention to treat analysis could not be conducted. analysis based on the three methodologically sound rcts demonstrated the provision of early en was associated with a significant reduction in mortality (or = 0.20, 95% confidence interval 0.04 to 0.91, i 2 = 0). no other outcomes could be pooled. sensitivity analysis including all four on-topic rcts (or = 0.26, p = 0.04, i 2 = 0), and a simulation analysis conducted using a different analytical method. (or exact = 0.14, 95% ci 0.01 to 0.97), confirmed the presence of a mortality reduction. conclusions. although the detection of a statistically significant reduction in mortality is promising, overall trial size was small. the results of this meta-analysis should be confirmed by the conduct of a large multi-center trial. reference(s). results. the mean ibp was 10.7 ± 4.1 and mean igp was 11.6 ± 4.1. correlation between the ibp and igp was significant however moderate (r2 = 0.51). analysis according to bland and altman showed a bias and precision of 0.8 and 2.7 mmhg respectively, however the limits of agreement (la) were large and ranged from -4.5 to 6.1 mmhg. the median grv proto was 80 ml (0-1,050) and median grv classic was also 80 ml (0-1,250). correlation between the 2 methods was excellent (r2 = 0.89). analysis according to bland and altman showed a bias and precision of -0.8 and 52.3 ml respectively and the limits of agreement (la) ranged from -103 to 102 mmhg. the median drainage time and return times were 5 min (0.5-15) and 2.5 min (0-21) for grv proto compared to 2 min (0.1-10) and 1 min (0-8) for grv classic. a preliminary cost effectiveness analysis shows that the price of measuring grv with the classic method ranges from 3.84€ to 24.18€ per day, depending on the grv size. price of measuring grv with the gastro pv system is independent of grv size and is estimated at 9.49€ per day. the gastro pv system if priced at 8.5€ could become cost effective at grv of 100 cc and more. conclusions. the interim results of an ongoing multicentre pilot study show that the gastro pv is a good alternative to the standard method for measuring grv. because the nurse can perform other tasks during drainage and return of the grv, and the fact that the system remains closed during measurement, this could be a major step forward in standardisation of grv measurement. furthermore it allows screening for intra-abdominal hypertension via igp estimation. acknowledgment. the gastro pv devices were provided by holtech medical, free of charge. introduction. the importance of early enteral feeding of the critically ill patient has been well documented. it is the more physiological approach, which is associated with lower rates of infectious complications. early enteral nutrition within 24 h is recommended by the espen guidelines on enteral nutrition. a recent meta-analysis revealed that mortality and the incidence of pneumonia were significantly reduced in patients with enteral nutrition within 24 h. parenteral nutrition may be associated with higher mortality. objectives. evaluation of a new technique for the placement of postpyloric feeding tubes by intensive care physicians. methods. prospective cohort study in 27 critically ill patients subjected to transnasal endoscopy and intubation of the pylorus. attending intensive care physicians were trained in the handling of the new endoscope for transnasal gastroenteroscopy for 2 days. a jejunal feeding tube was advanced via the instrument channel and the correct position assessed by contrast radiography. primary outcome measure was successful postpyloric placement of the tube. secondary outcome measures were time needed for the placement, complications like bleeding and formation of loops and the score of the placement difficulty graded from 1 (easy) to 4 (difficult). data are given as mean values and standard deviation. out of 34 attempted jejunal tube placements, 28 tubes (82%) were placed correctly in the jejunum. the duration of the procedure was 28 ± 12 min. the difficulty of the tube placement was judged as follows: grade 1: 17 patients, grade 2: 8 patients, grade 3: 7 patients, grade 4: 2 patients. in 3 cases, the tube position was incorrect, and in another 3 cases, the procedure had to be aborted. only in one patient, bleeding occurred that required no further treatment. conclusions. fast and reliable transnasal insertion of postpyloric feeding tubes can be accomplished by trained intensive care physicians at the bedside using the presented procedure. this new technique may facilitate early initiation of enteral feeding in intensive care patients. grant acknowledgment. the authors acknowledge the support of pentax, hamburg, germany, who provided the endoscope used in the study and of fresenius kabi, bad homburg, germany who provided the feeding tubes. a well-nourished condition before prolonged endotoxemia results in a better ability to adapt to endotoxin-induced metabolic deterioration of arginine-nitric oxide metabolism than does reduced caloric intake before endotoxemia (1). the role of individual organs in the arginine-citrulline metabolism during malnutrition and sepsis is unknown and may be key to direct future interventions. to study the effects of reduced caloric intake and endotoxemia on the citrulline-arginine metabolism in the gut-liver-kidney axis. organ arginine-nitric oxide metabolism was measured by using a primedconstant stable-isotope infusion of [15n2]arginine and [13c-2h2]citrulline during 4 conditions; a 7-day reduced caloric intake feed regimen (starv; n = 9), normal control feed regimen (co; n = 9), endotoxemia alone (ce) and reduced caloric intake and endotoxemia (re) in. catheters for blood sampling were placed in the abdominal aorta, which, in combination with the catheters in the portal, hepatic and renal veins, served for metabolic measurements across the portal-drained viscera, liver and the kidneys, respectively. results. interestingly, re animals had similar citrulline appearance from the gut (331 ± 36 nmol/kg/min) compared to control and animals during ce, but higher in endotoxemia alone (102 ± 14, p \ 0.01). this was related to a significantly higher no production from the gut in the re group (1,644 ± 153 vs. 720 ± 41, p \ 0.01). in the kidney arginine appearance from citrulline decreased significantly during re compared the control animals (3 ± 7 vs. 224 ± 30 nmol/kg/min, p \ 0.01). in contrast, the liver disposed more arginine in the re group compared to the other conditions, while no production was not higher. conclusions. despite reduced caloric intake prior to endotoxemia, the gut remains capable of increasing release of citrulline, although the capability of the kidney for the de novo production of arginine is severely compromised. metabolic control of the citrullinearginine metabolism in the gut-liver-kidney axis should focus on increasing de novo arginine production from citrulline. objectives. the aim of this study was to measure duodeno-caecal transit times of enteral feed in this patient group using a scintigraphic technique. a prospective observational study was performed in 16 mechanically ventilated critically ill patients (12 m, age 49 ± 17 yr, bmi 25 ± 5 kg/m 2 , icu admission day 5 ± 3, apache ii on study 20 ± 7; mean ± sd) and 6 healthy subjects (3 m, age 24 ± 9 year, bmi 24 ± 45 kg/m 2 ). after a 6 h fast a 100 ml enteral feed (ensure 1 kcal/min), labelled with 20 mbq 99m tc-sulphur colloid, was infused into the distal duodenum over 6 min. dynamic anterior scintigraphic images were recorded in 3 min frames for 240 min and the time of first appearance of activity in the caecum was recorded by two blinded operators (kj, ar). data were assessed using mann whitney u test and are presented as median (iqr). introduction. erythromycin, a macrolide antibiotic is widely used as a prokinetic agent in intensive care unit (icu) despite the lack of data supporting its prolonged effectiveness in enteral nutrition (en) intolerant critically ill patients. to evaluate impact on clinical outcome of erythromycin prescription as prokinetic agent in icu. all patients consecutively admitted from january 2008 through december 2009 mechanically ventilated for more than 5 days and receiving en were included in an observational cohort study. en intolerance was defined clinically as a 8-hourly gastric residual volume (grv) c150 ml or vomiting. successful en was defined as a grv\150 ml with a feeding rate c40 ml/h. erythromycin prescription was left to practician appreciation. objectives. this study aims at evaluating the relationship between diarrhoea and en in icu patients. methods. during 1 month, the days with and without diarrhoea (c 3 liquid stools/day) and the characteristics of nutritional support of all patients staying in our icu were recorded. patients staying \24 h or presenting an intestinal stomy were excluded. we compared, between days with and without diarrhoea, total energy coverage and energy coverage by en as % of needs, en energy intake and en volume for each patient. needs were estimated as 25-30 kcal/kg body weight for women and men, respectively. the relationship between antibiotics, laxative treatment and diarrhoea was also analysed. results are presented as mean ± sd. comparisons were made by mann-whitney test. the risk of diarrhoea with en was calculated by odds ratio and confidence intervals (ci). the study included 1133 days of hospitalisation of 171 patients (60 ± 16 years, bmi 26 ± 5 kg/m 2 , sofa score at admission 6 ± 6). en was present in 85 days of diarrhoea and 581 days without diarrhoea. determining the small bowel function is of great concern in icu patients, because a malfunctioning small bowel may predispose to malnutrition and may increase the risk of sirs. a recently developed test, the citrulline generation test (cgt), measures the enterocytes' capability to convert glutamine into citrulline. the production of citrulline exclusively takes place in functioning enterocytes, therefore this conversion represents small bowel function. objectives. we aimed to define the cgt reference values in 16 'stable' icu-patients to assess small bowel function. secondly, we wanted to compare four different cgt methods; enteral and iv administration of dipeptiven and measurement of citrulline in both arterial and venous samples. we performed the cgt on 16 stable icu-patients, defined as having respiratory failure but not dependent on vasopressors. they had a normal renal function and were able to tolerate enteral nutrition. a 5 h fast was followed by administration of 20 g of glutamine-alanine (dipeptiven ò ) either intravenously or enterally, randomly determined. the next day the same test was performed by using the other route. after each administration of dipeptiven, citrulline levels, both arterial and venous, were measured at fixed time points using reverse-phase high performance liquid chromatography (hplc). results. nine females and 7 males were admitted to the icu with either a medical (11) or a surgical (5) diagnosis. they had a mean (± sd) age and bmi of 60.9 ± 10 years and 26.7 ± 7, kg/m 2 respectively. their median apache ii score was 24.5 (iqr = 19.5-26.8). on the day the cgt was performed their median sofa score was 4.0 (iqr = 3.0-4.8 early post-pyloric feeding has been shown to improve clinical outcomes [1] . commonly used methods for placing a nasojejunal tube (njt) are blind, endoscopic or fluoroscopic placement. the later two methods are relatively invasive, expensive and can cause delay to feeding, whereas blind placement is often unsuccessful. electromagnetic sensor guided njt insertion is a bedside technique able to confirm successful placement without the need for abdominal x-ray. the system incorporates a liquid crystal display and a receiver unit. the receiver is placed over the patient's xiphoid process and picks up the signal from an electromagnetic transmitter located at the tip of the feeding tube. the screen provides a visual aid to enable the operators to trace the route of the tube tip and identify its' location according to anatomical markers. objectives. we were interested to determine the suitability of electromagnetic sensor guided njt insertion especially in relation to success rate and procedure time. methods. fifty patients were referred for electromagnetic njt insertion on units at the leeds teaching hospitals. insertion time was measured from oesophageal visualisation until post-pyloric placement. various positional manoeuvres were employed along with administration of sedatives, prokinetics and air insufflation when applicable. all insertions were carried out by experienced investigators. all njt insertions were confirmed by abdominal x-ray. data collection included patient demographics, hospitalisation and procedural information. results. forty male and 10 female patients, mean age 40 (range 1-81 years), bmi mean 25 (14-33), had attempted electromagnetic njt placement. patients had been hospitalised for a median of 12 days (1-180). indication for njt insertion was either large aspirate and/or reflux (86%). seventy six percent of patients had an artificial airway and 50% of patients were receiving sedation. forty six percent of patients received metoclopramide and 58% air insufflation. thirty six percent of patients were moved into either left or right lateral position. successful post-pyloric placement was achieved in 90% of patients confirmed by additional abdominal x-ray. procedural time varied from 2 to 180 min (mean 23). two of the placement failures were due to patient intolerance. conclusions. bedside electromagnetic guided njt placement technique is an acceptable method of placing post pyloric feeding tubes with a high success rate. gastrointestinal failure (gif) score has been suggested (1). the gif score defines gi failure as the occurrence of feeding intolerance (fi) and intra-abdominal hypertension (iah) simultaneously. to compare the outcome of patients with primary vs. secondary gif. methods. all consecutive, mechanically ventilated (mv) patients treated for at least 24 h during january 2008 to december 2009 in two icus were studied. gif was defined as gif score equal or above 3 points according to the gif score (1). 3 points = fi and iah simultaneously; 4 points = abdominal compartment syndrome (acs). fi was defined as the need to stop enteral feeding for any clinical reason (vomiting, high gastric residuals, bowel distension etc). iah was defined as mean intra-abdominal pressure (iap) c12 mmhg on any day. acs was defined as iap [ 20 mmhg with the new onset organ failure. when gif developed in a patient with primary pathology in abdomino-pelvic region it was classified as primary gif, when occurred without previous pathology in abdomino-pelvic region it was taken as secondary. objectives. in this study the biochemical quality and prion safety of the pharmaceutically licensed plasma octaplaslg ò was evaluated. the prion reduction factor achieved by western blot was confirmed by animal studies. eighteen consecutive batches of octaplaslg ò (octapharma ppgmbh, vienna, austria) were tested on global coagulation parameters, fibrinogen levels, activities of coagulation factors and protease inhibitors, activation markers, as well as von willebrand factor multimers. in parallel studies, plasma pool was spiked with exogenous spike material, derived from brains of hamsters infected with hamster-adapted scrapie 263 k, and a down-scale of the octaplaslg ò manufacturing process was performed. the prp sc reduction factor for the resin was investigated in both western blot and hamster bioassay studies. a reduction factor of c3.1 log10 prp sc was found for this process step by western blotting. the outcome of the hamster bioassay confirmed that the high level of removal prp sc seen during octaplaslg ò manufacturing was equivalent to a removal of infectivity (3.0 log10). in octa-plaslg ò , a parallel reduction of the s/d virus inactivation step led to significantly higher activities of plasmin inhibitor. our studies demonstrated that the same amounts of prp sc and prion infectivity bind rapidly and with a very high affinity to the chromatography resin. octaplaslg ò has the same clinical safety and efficacy profile compared to that demonstrated by octaplas ò over the last 18 years, except for the increased safety margin in terms of prion disease transmission and the possible effect of a significantly increased plasmin inhibitor activity. uniplas ò is a second generation solvent/detergent (s/d) treated, coagulation active plasma for infusion produced with an implemented prion removal step. it was developed as an alternative to the blood group specific s/d plasma products, octaplaslg ò and octaplas ò , in order to obtain an universally applicable (i.e. blood group independent) plasma that can be used without taking into account the blood group of the recipient. due to an initially controlled, optimal mixing of plasma of different blood groups prior to s/d treatment, in uniplas ò , the blood group specific antibodies (anti-a and anti-b of both igm and igg type) are neutralised and/or removed by free a and/or b substances and red blood cells (rbcs) to a clinical acceptable level with very limited or no complement activation. objectives. in this study an extensive biochemical characterisation of the first uniplas ò validation batches was performed. methods. three batches of uniplas ò were produced by octapharmappgmbh (vienna) under production conditions in [2008] [2009] . uniplas ò batches were tested on all important coagulation factors, protease inhibitors, activation markers, adamts13 and factor h levels, as well as von willebrand factor multimers. in addition, anti-a and anti-b titres of igm-and igg-type were investigated. finally, complement activation products, as well as key components of the complement system, were measured. results. in uniplas ò batches, all coagulation factor activities were higher than 0.7 iu/ml and all protease inhibitor activities, including protein s and plasmin inhibitor, were higher than 0.5 iu/ml. uniplas ò contained standardised levels of adamts13 and factor h, within the normal ranges for single-donor freshfrozen plasma. there was no activation of fvii obtained during manufacturing, thrombin-antithrombin (tat)-complex, prothrombin fragments (f1 + 2) and d-dimer levels were within the normal ranges. anti-a and anti-b titres were within the uniplas ò specification, i.e. anti-a igm and anti-b igm\1:8 as well as anti-a igg and anti-b igg \1:32, respectively. uniplas ò did not contain an increased amount of immune complexes and the manufacturing of uniplas ò associated with more complement activation than the one seen for octaplaslg ò . conclusions. the present study confirmed that uniplas ò displays the same high quality and clinical efficacy as the s/d treated blood group specific plasma octaplaslg ò , but with the additional advantage in being a blood group independent universally applicable plasma. most pts received more fluids than calculated by parkland formula (7 ± 7.5 ml/kg 9 %tbsa). interestingly, nonsurvivors received less (3.9 ± 4.1 vs. 8.3 ± 8.2 ml/kg 9 %tbsa). gastric decompression, ascites drainage and the implementation of a stool protocol with rectal enemas (18 interventions in 12 pts) was able to remove 2.2 ± 1.3 l of body fluids and this was related to a significant decrease in iap and cvp and an improvement in oxygenation and urine output ( conclusions. pris is a difficult condition to diagnose and routine monitoring of the adverse effects of high-dose propofol remains sub-optimal. hypothermia has been reported to alter propofol pharmacokinetics 3 and we propose that active cooling may increase the risk of developing pris. this may be particularly relevant in patients with tbi who are on high doses of propofol to control icp in addition to concomitantly administered catecholamines to maintain cerebral perfusion pressure. we recommend that further research is required in this area in view of the increasing use of induced hypothermia in icu. objectives. to compare differences in fluid resuscitation based on direct or indirect admissions to the london burns unit. methods. 67 admissions to the burns unit with [20% burned surface area (%bsa) were identified over 5 years. 16 were excluded from analysis due to palliation or death within the first 8 h. 12 sets of notes were randomly selected for analysis of fluid balance in the first 24 h period of fluid resuscitation after the burn injury. results. mean (sd) time from burn injury to arrival at the burns unit was lower for patients transferred direct to the burns hospital rather than via another hospital (117.9 ± 75.27 vs. 372.5 ± 255.7 min p = 0.032). mean (sd) error in burn size estimation was lower for patients initially treated by burns specialists versus non-burns specialists (4 ± 3.6 vs. 10.6 ± 4.3%, p = 0.016). all patients were resuscitated according to the parkland formula calculated at one of 2, 3 or 4 ml/kg/%bsa. the mean (sd) actual fluid volume differed from the target by 30.06% (±40.43%); the lower the calculated fluid target, the greater the error between actual and planned resuscitation volumes; there was no difference in accuracy of fluid resuscitation at 24 h between patients initially managed by burns specialists versus non-burns specialists (44.14 ± 49.04 vs. 10.34 ± 7.70% respectively, p = 0.163). conclusions. burned patients transferred directly to specialist burns care receive a faster and more accurate assessment of their burn injury. despite this, we found no difference in fluid targeting errors at 24 h, though this may reflect corrective fluid management on arrival at the specialist centre. echocardiography is an useful and minimally invasive tool that allows to know the heart filling pressures, also it has proven highly accurate in predicting the response to volume in critically ill patients. we try to determinate the response to fluid infusion by static variables as cvp or lap, comparing with the variation of ivc. methods. an observational prospective study with 18 patients undergoing coronary cardiac surgery (7 patients were excluded by a no presenting a good echo views), in the postoperative period under mechanical ventilation (vt 8 ml/kg, fio2 50%, peep 3). we performed an echocardiography if the patient presented hypotension, just before the habitual fluid load protocol were started (500 ml hes 6% in 30-45 min). we collected data before and after the infusion, and determine the responsiveness to volume if the cardiac output increased more than 10%. data in the report included invasive cvp and lap, and echo measures, ratio e/e', diameter and variations of inferior vena cava (ivc) and variations of stroke volume by echocardiography (ølvot x vti lvot) and with vigileoò system. . the correlation between low values of cvp/lap and volume response was poor, the relationship between cvp below 6 mmhg with increased cardiac output had a correlation (pearson correlation -0.06) with a significance (2-tailed) 0.981, and the relationship between lap \4 mmhg and an increase in cardiac output had a correlation (pearson correlation 0.229) with a significance of (2-tailed) 0.362. the measurement of the variation of the inferior vena cava, led us to calculated a cutoff point more sensitive to determine which patients were responders to volume. through the roc curves (sensitivity/specificity), with the area under the curve of 82.5% (se = 11.2%) and with a confidence interval of 95% (p significance of 0.002), resulted in a 12% variations of ivc with a sensibility of 100% and specificity of 42% (younden's index of 42.9%). the same calculation, based on kraemer's quality indices (qi) gave us a 13% of variation in ivc, with a w = 0.2 specificity rather than sensitivity (qi 0.58), and with a w = 0.8 sensitivity rather than specificity (qi 0.61) objectives. to ascertain whether postoperative hypothermia is linked to high or low risk surgical patients. we conducted a prospective systematic analysis looking at the incidence of postoperative hypothermia in adults who underwent general anaesthesia. children age \18, pregnant women and patients undergoing regional anaesthesia were excluded from the survey. to identify the current level of doctors' knowledge on perioperative fluid management. methods. the survey was conducted at george eliot hospital, nuneaton, uk in may 2009. questionnaires consisting of ten multiple-choice questions on basic sciences and clinical scenarios were devised by a consultant anaesthetist. these were personally distributed to doctors of all grades working in anaesthetics and the surgical specialties. doctors were asked to complete the questionnaire within 5 min. of the 66 questionnaires distributed, 52 were completed. results. the mean questionnaire score varied between specialties from 87% in the anaesthetics department to 64% for doctors in surgical specialties. the mean score of registrars and fy1 doctors in surgical specialties was found to be 72 and 65% respectively. the overall mean score was 74%. of all doctors surveyed, the daily maintenance water requirement was known by only 52%, 62% knew the daily maintenance sodium requirement and 75% knew that of potassium. the electrolyte contents of 0.9% sodium chloride and hartmann's solution was answered correctly by 65% and 63% respectively. there is a significant deficiency in doctors' knowledge on perioperative fluid management. more emphasis on optimal perioperative fluid management is required in undergraduate and postgraduate training. increased awareness of the british consensus guidelines on intravenous fluid therapy for adult surgical patients would aid training. based on this survey, a regional online survey of junior doctors is planned to further identify gaps in perioperative fluid management training. optimal fluid management could also help to reduce prolonged hospital stay which can result from fluid-related complications. objectives. to evaluate dynamic echocardiographic parameters as predictors of volume responsiveness in surgical patients. methods. 25 patients were included in the study after laparotomy surgery performed on the same day (4 breathing spontaneously and 21 mechanically ventilated in volume controlled mode with tidal volume of 10 ml/kg). a fluid challenge was performed in spontaneously breathing patients by passive leg raising and infusing saline (7 ml/kg). echocardiographic analysis of respiratory changes of inferior vena cava diameter (ddivc) and aortic blood flow (dabf) was performed in all patients. a threshold of 18% for ddivc was used for classifying patients as volume responders or non-responders. age, sex, gender, bmi, cvp, iap, map, left ventricular ejection fraction, left ventricular systolic and diastolic area, and stroke volume in all patients, as well as itbvi, ci, ppv and svv in 16 patients were measured. a positive correlation with ddivc was established for itbvi (r = 0.49, p = 0.05), iap (r = 0.47, p = 0.01) and ef (r = 0.38, p = 0.05). a positive correlation with dabf was not established for any variable measured. 16 patients (64%) were classified as volume responders and 9 (36%) as non-responders. responders had overall higher iap than non-responders (13.45 ± 4.99 mmhg vs. 9.82 ± 3.33 mmhg respectively, p = 0.04). respiratory changes of ivc diameter showed positive correlation with itbvi. so, conclusions about itbvi could be indirectly made from ddivc values in patients who are not being invasively monitored. ppv and svv did not show positive correlation with itbvi. surprisingly, we confirmed a positive correlation between ddivc and iap. we detected 9 patients with high iap, while all the volume responders had overall higher iap. although further investigations are needed to establish how longer duration of high iap may influence ddivc, it seems that ddivc is a good parameter of volume responsiveness during first 24 h after laparotomy surgery. unlike from other studies, we could not establish a positive correlation between dabf and any variable measured. these studies were performed in hypovolemic septic patients, so this could be the reason for such different results. more studies are needed in a larger set of patients undergoing laparotomy surgery to evaluate dabf. introduction. fluid optimization after major cardiac surgery was shown to improve patients postoperative outcome significantly. several hemodynamic parameters were proposed for the guidance of therapy but never compared in a head to head trial. objectives. in this prospective randomized trial patients scheduled for elective cardiac surgery underwent early goal directed fluid therapy guided either by stroke volume variation (svv) or by oxygen delivery index (do2i). we hypothesized that while svv is easier to obtain it will not be inferior to do2i in outcome parameters. methods. following ethics committee approval and signing of a written informed consent, 84 patients were randomized in two groups to undergo either fluid optimization guided by do2i or svv in the first 8 postoperative hours in the icu following elective cardiac surgery (cabg). following a standardized egt protocol the parameters were collected by using hemodynamic monitoring based on a pulse contour analysis and a transpulmonary lithium dilution (lidco plus, lidco,uk). we compared amount and type of volume infused, need and amount of inotropic or vasopressor substances, time spent on ventilator, los in the icu and postoperative complications. statistics were evaluated by using a t test for unpaired samples. table 1 . compared to the do2i group fluid optimization using svv showed reduced ventilator times (p = 0.043) and less complications (p = 0.004) in the first 80 days after surgery. no differences between the groups were detected concerning the type and amount of volume infused, need for inotropes or vasopressors or the los in hospital conclusions. while svv is less invasive, cheaper and easier to be obtained than do2 outcome was at least not inferior and even showed improvements in postoperative cardiac surgery patients. 23rd esicm annual congress -barcelona, spain -9-13 october 2010 s377 introduction. over the years, there have been concerns over incompatibility of transfused blood with various intravenous fluids during blood transfusion, especially related to increased levels of haemolysis. it is often impractical, particularly in an emergency situation, to flush through a giving set with a so-called ''safe'' fluid prior to and after delivering blood. we wanted to investigate whether this is actually necessary and whether the usual fluids used in the perioperative period really do cause any demonstrable alteration in the composition of transfused blood. objectives. the purpose of this study was to expose packed red cells to a variety of different intravenous fluids commonly used during the perioperative period and to measure a number of parameters in the blood following their contact with each different fluid, including a blood film to examine for clumping of cells or haemolysis. a unit of a positive blood was passed through blood giving sets which were primed with various intravenous fluids. after adequate mixing of blood with fluids, samples were collected for full blood count, urea and electrolytes and blood films. one millilitre of mixed blood was taken in each bottle at a time. the intravenous fluids used in this study were normal saline, hartmann's solution, 5% dextrose, 10% dextrose, starch and gelatin. there was no significant rise in blood parameters suggestive of haemolysis. the potassium and ldh levels were not significantly different with various fluids. the haemoglobin and haematocrit levels were also comparable to one another. there was no demonstrable changes in blood parameters suggestive of haemolysis, nor were there any change in electrolyte values. this suggests that all of the fluids investigated during this study would be suitable to be used via the same giving set before and after the transfusion of pack red cells. objectives. to assess the compliance with the national guidelines in avoiding inadvertent peri-operative hypothermia in an acute district general hospital in england. we prospectively studied our local practice on maintaining normothermia in 165 consecutive adult surgical patients {61 men, mean age 57.4 years, 112 patients with asa grade 2 (67.7%), 60 emergency surgical patients (36.4%), 49 patients with significant cardiac disease 29.7%}. we used a questionnaire that was filled pre-operatively by anesthetic nurses, intra-operatively by anesthesiologists, and post-operatively by recovery nurses. patients were recruited from the following surgical subspecialties: general surgery (34%), gynecology (24%), trauma (22%), breast surgery (9%) and orthopedics (7%). day surgery patients were excluded. peri-operative hypothermia was defined as temperature \36°c as per the nice guidelines. results. less than half of our patients (47.9%, n = 79) had their temperature measured preoperatively, on whom incidence of hypothermia was 25.3% (n = 20). only one of these patients was warmed prior to induction. patients requiring emergency surgery and those with asa grade 2 had increased incidence of preoperative hypothermia (30.8% and 28.6% respectively, p \ 0.05). based on nice guidelines, 164 patients needed intraoperative forced air warming but only 64 (38.8%) patients received it. intraoperative temperature measurement was made on 111 patients, of whom 28.8% (n = 32) were hypothermic. incidence of intraoperative hypothermia was high in surgical procedures lasting longer than 30 min (p \ 0.05) but was not affected by the use of regional anesthetic techniques. 161 patients had their temperature measured on arrival to recovery of whom 46 (26.6%) were hypothermic. 38 patients (23.6%) had their temperature measured every 15 min (nice recommendation) and the mean time interval for temperature measurement in recovery was 26 min. 10 patients were still hypothermic on leaving recovery. conclusions. majority of our surgical patients did not receive adequate perioperative care on maintaining normothermia. consequently, the incidence of hypothermia was significant pre-, intra-and post-operatively. we are currently analyzing the data to investigate the effect of hypothermia on duration of recovery stay, length of hospitalization and mortality in our patients. we completed a double-blind randomized trial in patients undergoing cardiac surgery in which we compared fluid resuscitation with a hydroxyethyl starch (hes, 10% 250 mw pentastarch) and saline. use of hes resulted in markedly less use of catecholamines the morning after surgery. an underlying design principle was that assessment of cardiac index (ci) is essential for a proper fluid protocol. in this analysis we examine that supposition. all subjects had pulmonary artery catheters. patients were consented preoperatively, but randomized post operatively to receive up to 4 blinded 250 ml boluses for predefined hemodynamic targets; ci \2.2 l/min/m 2 , blood pressure (bp) set by admitting team, cvp \3 mmhg, or urine output \20 ml/h. hemodynamic measurements were made before and after each bolus. after the 4 study boluses, only saline was used. results. 237 patients received fluids, 119 hes and 118 saline. there were 727 study boluses, 348 hes and 379 saline. of these, 99 boluses (14%) could not be assessed for this hemodynamic analysis (but were still used for the primary outcome) because of protocol violation or missing data. of the rest, 235 (37%) of boluses were given for a low ci; in 33 bp and 11 cvp were also low so that ci was the only trigger in 30%. a low bp was a trigger in 290 (46%). low cvp was the trigger in 95 (15%). only 50 hes and 65 saline patients required the maximum allowed 4 blinded boluses. at the 4th bolus, low ci was the trigger for 8 (16%) of hes but 20 (31%) of saline patients. there were 511 that could be evaluated for hemodynamic response based on four possible outcomes of cvp and ci. objectives. the aim of our study was to evaluate the predictive value of cvp with regard to gedi, and to correlate these parameters to cardiac index (ci). conclusions. volume depletion according to gedi was found in more than half the patients. the predictive values of cvp with regard to volume depletion were low gedi and its changes significantly correlated to ci and its changes, which was not observed for cvp. therefore, gedi appears to be more appropriate for volume management during mayor liver resections. introduction. regional anticoagulation with citrate is an effective and established anticoagulation strategy during crrt in critically ill patients, especially in surgical patients with a high risk of bleeding and in case of a heparin-induced thrombocytopenia (1). however, citrate crrt could be associated with major metabolic derangements such as metabolic alkalosis, hypocalcemia, hypernatremia and citrate toxicity. objectives. the aim of our study was to investigate efficacy, safety and metabolic stability during citrate crrt in critically ill patients with acute kidney injury. methods. the retrospective study was performed in a mixed surgical and trauma icu in a university hospital. patient charts were reviewed for demographic data, the period and dosage of citrate crrt and metabolic parameters. reasons of admission, comorbidities and severity of illness were also evaluated. citrate crrt was performed using commercially available equipment and fluid solutions (multifiltrate ò with integrated ci-ca ò -system; fresenius medical care; germany). to maintain stable metabolic and haemodynamic conditions we used an internal standard protocol for citrate crrt. statistical analysis was performed using descriptive methods (mean, median and standard deviation) and a mann-whitney u test where appropriate. p \ 0.05 was regarded as statistically significant. conclusions. although minor metabolic imbalances were observed, none led to a termination of citrate crrt and all of them could be managed by adjustments of blood flow and dialysate rates according to a preset protocol. our findings suggest citrate crrt to be a safe and effective strategy for crrt even in patients with hepatic dysfunction. nevertheless, metabolic parameters need to be monitored regularly to avoid severe metabolic derangements. introduction. the liver is central to ammonia metabolism, being the main site of urea cycle enzyme pathways. in acute liver failure (alf) and decompensated chronic liver disease (cld) ammonia dysmetabolism results in hyperammonaemia, thought to be of central importance in the pathogenesis of hepatic encephalopathy and, in alf cerebral oedema [1] . continuous renal replacement therapy (crrt), commonly used in critically ill patients may be an effective method of clearing ammonia. little is known of the efficacy such techniques have on ammonia clearance. objectives. to quantify the clearance of ammonia using an aquarius haemofilter (ahf) using different renal replacement doses and techniques. methods. patients with a circulating ammonia level[100 lmol/l due to commence crrt were enrolled. the ahf was programmed to run in either pre-or post-dilution modes at a blood flow rate of 200 ms/min using a 1.2 or 1.9 m 2 filter depending on the crrt ultrafiltration (uf) dose, which included 35, 60 or 90 ml/kg/h (adjusted for ideal body weight). 2 ml of blood and effluent fluid were collected, on ice into lithium/heparin and serum separation tubes, from pre and post filter access points and effluent tubing to calculate urea and ammonia clearance using the cordoba formula [2] . delta whole body ammonia clearance was determined by measuring arterial ammonia at 0 and 60 min. ammonia measurements were performed using a pocketchem ò blood ammonia bedside testing machine. results. 20 patients (9 alf and 11 cld) were recruited (mean age 45 years, sd (14), with mean arterial ammonia 123 lmol/l, sd (49). 60 min whole-body ammonia clearance was -12 lmol/l, p = 0.016, paired t test). ammonia and urea clearance were correlated (r = 0.748, p = 0.020); uf rate correlated negatively with filtrate ammonia (r = -0.496, p = 0.007) and positively with ammonia clearance (r = 0.515, p = 0.005). filter ammonia clearance was not dependent on filter size for the standard blood flow rate. pre or post dilution modes did not affect ammonia clearance (p = 0.100, student's t test). a constant filter size and blood flow rate achieved ammonia clearance of 39 ml/min/m 2 for 35 ml/kg/h, 60 ml/min/ m 2 for 60 ml/kg/h and 52 ml/min/m 2 for 90 ml/kg/h (p = 0.008, one way anova). conclusions. 60 ml/kg/h based on ideal body weight appears to be the optimum dose of crrt for ammonia clearance when using a blood flow rate 200 ml/min and a 1.2 m 2 filter. filter and delta whole body ammonia clearance may be increased further using the combination of a higher dose (90 ml/kg/h) with a larger filter size and higher blood flow rates. introduction. malnutrition is common in intensive care following the catabolic state induced by critical illness. patients who progress from enteral nutrition back to oral feeding are usually in an energy deficit. espen guidelines recommend increasing calorie delivery during the recovery period to cover this anabolic phase. oral nutritional supplements (ons) are widely used to facilitate calorie delivery within the hospital setting however the effectiveness of this strategy is dependent on patient compliance with the products. compliance among the elderly ward-based population has been considered (1) however that of intensive care patients has not been reported. to evaluate compliance to ons in a mixed medical and surgical adult intensive care unit (icu) in a district general hospital. prospective observational study was conducted over a 2 month period with data compiled from fluid chart analysis and discussions with nursing staff. all adult icu patients prescribed, or offered without prescription, an ons were included until the point they were discharged to the ward. the supplements studied, resource ò energy, 2.0 fibre, fruit and dessert (nestlé nutrition), were selected based on their availability within the trust. patients were offered a choice of flavour. results. data was collected and analysed for 51 patient days. a total of 122 supplements were prescribed. of the prescribed supplements, 42.6% were offered to patients and 26.6% consumed. 20% were offered the same at nursing discretion based on clinical need and 39.8% were consumed. resource ò energy was the most frequently prescribed and offered product (76.2 and 59.2% respectively). most common flavours selected by patients were strawberry and vanilla. resource 2.0 fibre was better tolerated (91.7%) than resource energy, resource fruit and resource dessert (73.9, 70.8 and 50.0% respectively). across all products the best tolerated flavours were apricot, chocolate and coffee (100%). the highest calorie supplement, resource ò 2.0 fibre, resulted in the best compliance in both tested flavours. compliance with ons demonstrated here is higher than previous studies (1) partly attributable to one-on-one nursing of icu patients enabling active encouragement with feeding. nursing staff discretion had better uptake than routine prescription of ons. however, difficulties with ons still remain. interestingly in our study the highest calorie density supplement was tolerated the best and thus giving the most benefit to the patient. despite the difficulties associated with ons uptake we would recommend its regular use on icu with a drive towards the highest calorie supplements being offered. introduction. cirrhosis is a chronic disease and the patient's quality of life is affected in a negative way due to the problems like ascites, jaundice, nutrition deficiency, fatigue, activity intolerance, itching, pain, insomnia, anxiety, hopelessness, work loss and depression. objectives. the aim of this study is to examine the changes in patient's lives that diagnosed with cirrhosis of the liver disease owing to the symptoms they experienced. methods. this research is a qualitative study that has been carried out with 13 inpatients diagnosed with liver cirrhosis in the gastroenterology clinic of a teaching and research hospital. average age of patients was 54 (ranging 39-70). descriptive characteristics form and semi-structured interview form were used in the data collection. interviews with patients have been performed individually and face to face. the data were evaluated by using colaizzi's phenomenological data analysis method. as a result of the data analysis, three categories and six themes were identified. categories include: (i) problems of symptoms related to the physical limitations (ii) psychosocial issues. patients suffer mostly from fatigue and malaise (12 patients), while those in the later stages suffer from, additionally, physical ailments caused by acid. inability to sleep due to anxiety and increase in tendency to sleep in advanced stages have been identified after being diagnosed. the majority of patients were identified to have undergone an anxiety besides having a fatal disease due to concern for the future, being forced to quit the job and being affected by the experiences of the patients in advanced stages. it also has been discovered that the patients had experienced social isolation because of fatigue and weakness in particular. as the result of this study it has been determined that patients with cirrhosis have mainly problems of fatigue, weakness, sleep disorders, anxiety and associated problems. 23rd esicm annual congress -barcelona, spain -9-13 october 2010 s379 [1] . while in patients with acute liver failure, elevation of arterial ammonia levels has been linked to cerebral complications and increased mortality, the role of arterial ammonia in hh patients is unknown. our study aims at evaluating arterial ammonia levels in patients with hh. furthermore, we wanted to elucidate the potential consequences of high ammonia levels in these patients. arterial ammonia levels were measured and documented in 72 hh patients without liver cirrhosis who were admitted to the medical icu. icu mortality and overall 28day-survival were documented. cox regression was performed to describe the impact of ammonia levels on mortality. mann-whitney test was used for comparison of metric variables. results. overall median arterial peak ammonia level in our patients was 65 lmol/l (41.9-100.6 lmol/l), whereas median arterial peak ammonia value was significantly higher in icu non-survivors compared to survivors (77 (53-119.2) vs. 52.9 (36.4-71.1); p \ 0.01). saps ii and sofa score were significantly higher in icu non-survivors (p \ 0.01 and p \ 0.05, respectively). cox regression revealed that arterial peak ammonia levels were significantly associated with higher 28-day-mortality (p \ 0.01), even after adjustment for saps ii. median arterial peak ammonia levels in patients with verified brain edema were significantly higher than in patients without (130.7 lmol/l (92-349.7 lmol/l) vs. 70.6 lmol/l (43.2-81.7 lmol/l); p \ 0.05) after exclusion of patients following cardiopulmonary resuscitation with consecutive hypoxic brain damage. our results suggest that increased levels of ammonia are associated with high mortality and can lead to brain edema in patients with hh. 53% of patients had a diagnosis of sepsis and 21% of patients were admitted under the neurosurgical team, the latter of which may have contributed to the relatively low anticoagulant use of 54%. systemic heparinisation was the sole anticoagulant used, but compliance with local protocols was poor with 48% of appts below the therapeutic range and 46% of infusions commenced at the wrong rate. 68% of filter changes were due to clotting and mean filter life was 32 h. despite this, dose delivery was acceptable, with 84% of prescribed dose delivered. conclusions. as previously reported 1 , our demographic data confirm the relatively poor outcome of patients needing crrt. we have identified areas where care for these patients could be optimised and endeavour to do this locally via improved protocol design and an ongoing educational programme. many of the components of crrt could be incorporated into care bundles, but certain aspects of treatment remain controversial 3 which may be a barrier to their adoption. given the high numbers of neurosurgical patients in our unit, consideration should be given to the use of regional anticoagulation such as citrate. introduction and objectives. accurate prognostic indicators of patient survival in an intensive care unit (icu) help guide clinical decision making. factors known to portend poor prognosis in acutely ill cirrhotics in icu include the need for mechanical ventilation, development of shock, renal failure and sequential increase in the number of failing organs. while serum lactate is now an established marker of survival and/or the need for transplantation in fulminant liver failure, its impact on critically ill cirrhotics is less well known. methods. we retrospectively studied 133 consecutive acutely ill cirrhotics admitted to the icu between 2005 and 08 at the royal free hospital, a tertiary referral centre in liver diseases and transplantation. data were collected on demographic variables, aetiology of liver disease, liverspecific prognostic scores [child-turcotte-pugh (ctp), model for end-stage liver disease (meld), united kingdom model for end-stage liver disease (ukeld)], and acute illness scores [acute physiological score and chronic health evaluation (apache ii), sequential organ failure assessment score (sofa) ]. in addition, serum lactate levels at 0, 24 and 48 h were also recorded. multivariable logistic regression analysis was performed, and the discrimination ability of each of the above-mentioned scoring models in predicting icu and hospital survival of these patients was evaluated using the area under the receiver operating characteristic (roc) curve. conclusions. one third of lt recipients present a documented bacterial infection within 1 year after surgery. we found a high prevalence of ciprofloxacin resistance and a low incidence of s.aureus witch was often resistant to methicillin. non fermentative gram negative bacilli represent 18% of the pathogens and should be taken in account for treatment of the most severe patients. extracorporeal liver support therapy is in its infancy but is valued as a detoxification treatment option for patients with cirrhosis who have rapid worsening of their liver function. we report the use of prometheus ò , a new extracorporeal liver support system allowing the removal of protein bound and water soluble toxins by fractionated plasma separation and absorption (fpsa) in a patient with wilson's disease (wd) who developed rapid worsening of their liver function. a 26-year-old female patient, diagnosed with wd since the age of 17, was initially treated in an irregular pattern with penicillamine. therapy was discontinued. now, 2 years later, she developed acute decompensated liver failure with hepatic encephalopathy with a meld 29. liver transplantation (lt) was the treatment option for this patient. but, in this case, the rapid and adverse evolution of the liver failure with renal failure and the unknown waiting time for a emergency liver donor in our country led us to use the extracorporeal liver support therapy. after 5 h 30 min of therapy we reduced the amount of bilirrubin for less than a half, we increase the urinary output and next day the patient went to liver transplant, stable, with a renal function improved. conclusions. acute liver failure due to wd is most of the time fatal without emergency lt. this case report highlights discontinuation of chelants treatment in a patient with wd. as the patient progressed to decompensated liver cirrhosis with encephalopathy, lt was the only treatment option but while we don't get a donnor, we can use, for a short period of time, an extracorporeal liver support therapy as a very useful bridge. results from two studies presented at the recent easl 2010 congress have shown that treatment with extracorporeal devices may not confer a survival advantage for severe liver failure patients, despite positive dialysis effects. however, results among a small sub-group of patients show promise like severely ill patients with hepatorenal syndrome type 1 or a meld score over 30. (1) . metoclopramide is used to stimulate the upper gi tract and seems to have no effect on colonic motility. objectives. the aim of this in vitro study was to compare the prokinetic potency of those substances. a tissue bath with guinea pig colonic segments fixed on a polyacrylic tray allows the evaluation of the transit time (tt), the time necessary for a wooden pellet to perambulate. a decrease of the tt reflects stimulation, and an increase inhibition of peristalsis. after stable peristalsis activity the effect of increasing concentrations of prucalopride, neostigmine or metoclopramide on tt were evaluated. dose response curves were constructed, two way anova (sigma stat) was used for statistics, p values b 0.05 were considered to be significant. effect of prucalopride and neostigmine on motility results. prucalopride stimulates normal peristalsis in vitro only in the highest tested concentration of 3 lm (p \ 0.05). neostigmine's prokinetic effect was limited to a small concentrations range (0.1 lm, p \ 0.05), the concentration of 0.03 lm had a moderate, but not statistically significant prokinetic effect and the highest tested concentration (1 lm) lead to a complete block of peristalsis (fig. 1) . metoclopramide, as expected, was devoid of any effect on colonic motility. conclusions. this experimental setting is a reliable method to evaluate the effect of different substances on colonic motility in vitro. prucalopride's prokinetic activity is concentrations dependent and limited. neostigmine is well known to improve colonic motility, but it seems imperative that the drug's effective dose range be use-higher concentrations have inhibitory effect on peristalsis. objectives. robotic radical prostatectomy involves extreme changes in patient position and often associated with a longer operative time than other commonly performed laparoscopic procedures. this review discusses the anesthetic considerations in robotic radical prostatectomy while analyzing potential risk factors related to pulmonary complications. we retrospectively reviewed the medical records of all the patients who had undergone robotic radical prostatectomy at our institution. among the total patients of 80, aged 49 to 82 years, 58 patients were capable of spontaneous respiration at the end of surgery (group i) whereas 22 patients needed assist ventilation (group ii). the demographic characteristics, coexisting diseases, anesthesia and operation time, anesthetic agents, the amounts of blood loss, infused fluid and transfused blood products were compared between the groups. results. the mean age of the patients was 67.2 ± 7.3 years. the mean operation times were 445.3 ± 210.3 min (range, 235-1250 min). age, body mass index (bmi) and asa status did not differ significantly between the two groups, whereas operation time, the amount of blood loss and the incidence of transfusion were significantly higher in the group ii. although 3 patients with subcutaneous emphysema and atelectasis needed prolonged ventilator care for 48 h, the incidence of atelectasis and subcutaneous emphysema was similar between the groups. conclusions. prolonged laparoscopic surgery in a steep trendelenburg position has a high possibility of postoperative respiratory insufficiency and the possible contributing factor is a long operation time. objectives. we examined the frequency of postoperative cough reflex and its effect on postoperative clinical outcome retrospectively. we examined the patients who admitted into the icu after the esophagectomy with lymphadenectomy during the period from september, 1999, to february, 2010. in addition to usual criteria for extubation we removed their tracheal tube if the cough reflex was identified when one milliliter of half saline was distilled into their trachea. if the cough reflex was absent until 7 days after the operation the patient underwent tracheostomy and after that they weaned from the ventilator. results. there were 125 patients (f/m 24/101), and their mean age was 58.5 ± 7.8. cough reflex were confirmed by seventh postoperative day in 95 patients (76%) but residual 30 patients underwent tracheostomy because of absence of cough reflex ( introduction. the technique of laparoscopic cholecystectomy carried with carbon dioxide pneumoperitoneum may lead to adverse events in mechanical, hemodynamic and respiratory systems as a consequence of physiopathological changes such as increased intraabdominal pressure. _ it may cause hypoxemia, hypercapnia, hemodynamic instability and impairment of oxygenation. decreased functional residual capacity, ventilation/perfusion imbalance and sympathetic stimulation effects of co2 that is absorbed from peritoneum are basic problems. in perioperative period, application of mechanical ventilation and anesthesia should be reviewed because of these physiopathological mechanisms. in this study, we aimed to investigate the effects of 5 cmh 2 o peep application on etco2, minute ventilation and arterial oxygenation during laparoscopic cholecystectomy operations. for this reason, the study included total 40 patients and they were randomly divided into two groups. same anesthetic protocol was applied in both groups. for general anesthesia induction; 1 mg/kg dose of fentanyl, 2 mg/kg dose of propofol were administered. following this procedure endotracheal intubation was applied with 0.15 mg/kg dose of cisatracurium. patients received %50 o2-%50 n2o (mixture with equal amounts) with 0.8-1.2 mac end-tidal sevoflurane for anesthesia maintenance. before co 2 insufflation, respiratory parameters were recorded on the respiratory apparatus adjusting etco2 32-36 mmhg, respiration rate 12/min., inspiration/expiration rate 1:2, vt: 8-10 ml/kg. patients were ventilated by volume controlled mechanical ventilation. heart beats, mean arterial blood pressure and peripheric o2 saturation (spo2), etco2, minute ventilation(v) and peak airway pressure(p _ ip) values of all patients were recorded just before insufflation (t0). after recording, 5 cmh 2 o peep was applied to the first group (group 1). peep wasn't applied to the 2nd group (group 2). these parameters were repeated in 5 periods such as 5 (t1) and 30 (t2) minutes after insufflation, preexsufflation (t3) and postexsufflation (t4) in both groups. before insufflation, respiration rate (12/min) and etco2 (32-36 mmhg) values were adjusted as planned in both groups and minute ventilation was also adjusted. at the same time, total insufflated amount of co2 for distending abdomen was recorded. arterial blood gas analyses were made just before induction (while patients were breathing normal room air, t0), 30 min after induction (t2) and just before the end of the operation (t4). in our study, we found that minute ventilation to stabilize etco2 32-36 mmhg was significantly increased in group 2 in which peep was not applied (p \ 0.01). none enhancement was needed in minute ventilation in group 1 and arterial oxygenation was significantly increased in group 1 (p \ 0.01). aside from the cholesterol lowering effects of statins, as a class of drugs they have been shown to exert anti-inflammatory effects and have the potential to be therapeutic in neuroinflammatory disorders 3 . we tested the hypothesis that atorvastatin improves memory retrieval post unilateral nephrectomy in a murine model. methods. c57/bl6 mice were randomly allocated into 4 groups (n = 8-10/group): control plus placebo, control plus atorvastatin, nephrectomy plus placebo and nephrectomy plus atorvastatin. animals were given either a placebo (0.4 ml normal saline) or 250 lg in 0.4 ml normal saline of atorvastatin by gavage once a day for 5 days. on day 4 all animals underwent fear conditioning training using a conditional stimulus of a 70 db tone and an unconditional stimulus of a 0.70 ma electric shock. on day 5 the surgical animals underwent unilateral nephrectomy, whilst the control animals received no surgery. at post-surgical day 3 all animals were tested for hippocampal dependent memory retrieval using the fear conditioning paradigm, with freezing response to the 70 db tone as a marker of memory retrieval. all animals were then terminated. results. surgery evoked a reduction in hippocampal dependent memory retrieval in the nephrectomy plus placebo group as measured by % freezing time (mean ± sd: 40 ± 16) when compared to the control plus placebo group (70 ± 19; p \ 0.01); a situation mimicking pocd. this change was obviated in the nephrectomy plus atorvastatin group (59 ± 16; p [ 0.05 vs. control plus placebo). conclusions. our data suggested that atorvastatin has the potential to improve postoperative cognitive performance in a murine model of pocd. the proven safety of the drug along with its already widespread use and cost effectiveness would permit rapid instigation of a human randomized controlled trial to explore efficacy in the clinical setting. a. puxty 1 , r. docking 1 1 glasgow royal infirmary, department of anasethetics, glasgow, uk hypotension in the post-operative period is common but guidelines recommend its prevention/treatment [1] . epidurals are common practice following major surgery in many institutions and can prevent pulmonary complications [2] but have also been associated with falls in blood pressure when compared to other analgesic techniques [3] . fluids therapy is a common intervention for hypotension but fluid overload has been associated with worse outcomes in surgical patients [4] . we decided to audit the incidence and management of hypotension in the surgical high dependency unit of a large tertiary referral hospital. to determine the incidence and management of hypotension in the surgical high dependency unit in pancreatic, upper gi and lower gi patients. we prospectively looked at 48 patients who underwent major upper gi, lower gi or pancreatic surgery involving epidural analgesia. the first 24 h of care from onset of anaesthesia was closely looked at with regards to fluid management, epidural management and actions taken on episodes of hypotension or severe hypotension (defined as systolic blood pressure of \90 and \80 respectively). each episode of hypotension was looked at to determine the actions taken at that point. of the 48 patients looked at, 17 were major pancreatic, 17 lower gi and 14 upper gi patients. 40 (83%) had at least one episode of hypotension, with 11 (23%) having at least one episode of severe hypotension. mean fluid in during the first 24 h was 6523 ml, with a mean fluid balance of 4133 ml. there was no difference between the doses of epidural local anaesthetic in 24 h between the hypotensive and non hypotensive groups (p = 0.275). management of hypotensive episodes was variable, but the most common intervention at episode one was fluid bolus (80%) and discontinuation of epidural was most common at episode two (39%). use of vasopressors for hypotension was very low with only two infusions being started altogether. conclusions. hypotension is very common in our high dependency unit. fluid balance in our patients was far more positive that we had expected. management of hypotension was variable. we plan to institute a protocol for hypotension and fluid administration to determine if improvements can be made. objectives. to identify predictive factors associated with the need for relaparotomy in patients with ssp. adult ssp patients undergoing laparotomy between 2004 and 2009 included within a single-center peritonitis registry (perit) were collected. patients subjected to relaparotomy were studied. we excluded patients with severe peritonitis secondary to appendicitis. apache ii and sofa score at icu admission after the initial laparotomy were recorded. variables with a p value.1 in a bivariate analysis were included in a multivariate logistic regression for further analysis of predictors for need for re-laparotomy. results. two-hundred forty-seven patients were obtained from perit registry. a total of 212 patients with spp were included in the analysis. eighty seven patients (41%) required relaparotomy. median number of re-laparotomies was 3. most spp were associated to colon (n = 94, 44.3%), small intestine (n = 83, 39.2%) and biliary tract (n = 33, 15.6%) perforations. cultures were positive in 74.5% of first laparotomy: gram negative bacteria were isolated in 53.3%, gram positive bacteria in 16.5% and fungi in 4.7%. hospital mortality was 17% (n = 36). multivariate analysis is described in the table 1 . conclusions. in obese patients scheduled for surgery, the previous use of cpap has not shown an improvement in blood gas parameters. the use of cpap in the hours before and immediately after surgery has not been associated with better postoperative oxygenation. combined icu-surgery dpt. action in these cases seem to contribute to better patient outcomes. objectives. we set out to quantify the intensive care workload and changes to that workload over the first 4 years following the transfer of a specialist bariatric service to our hospital. a prospectively collected bariatric surgical database was cross-referenced to the itu database (ward-watcher) to identify admissions to the 10-bedded critical care unit of all patients who had undergone any bariatric procedure. for each patient identified; demographics, reason for admission, level of support, length of stay and outcome were recorded. data were grouped into 12-month periods for trend analysis. research in emergency situations and especially in resuscitation field raises important ethical and regulatory issues. the globalization of the resuscitation science through multicentric trials for example highlights the need for a more consistent approach to regulatory aspects to enable the science to grow while protecting human rights. objectives. the purpose of this analysis is to compare the different regulations approaches in emergency research in north america (canada, usa) and in europe (european directive, france). conclusions. this analysis emphasizes the lack of international standardization of regulatory measures and ethical decisions. however some countries like the us seem to advance in the democratic process by mandating additional regulatory measures (community consultation, public disclosure to the communities) prior to initiation of clinical investigation; nonetheless, there is little evidence of their effectiveness. many challenges are raised. firstly, the variability in regulations, and consequently in local board's assessments, is problematic, pleading for international regulations. secondly, the current heterogeneous ethical review process and demanding unsubstantiated regulatory measures poses a risk to all when it is not evidence based and it is applied inconsistently between countries, within a country and worse at the level of each individual hospital review board. it puts the investigator at risk for unnecessary criticism and the community at risk as it is unknown if we truly consult or inform our target communities about waiver of consent research through our current ethical and regulatory processes. globalization and evaluation of the ethical and regulatory processes are urgently needed; regulatory community has to work towards a standardized evidence-based process upon which to base regulatory decisions. introduction. in research outside the intensive care field it is known that a high score for the psychological factor ''perceived hopelessness'' experienced by healthy individuals increases risk of death several fold. objectives. the aim of this study was to examine if the score of the psychological factor ''perceived hopelessness'' may predict long term mortality (mean or high perceived hopelessness score) when assessed post icu care in former icu patients. methods. prospective, multicenter study in three mixed icu's in sweden. questionnaires, including the 2-item hopelessness scale, demographic data and previous illnesses, were sent 6 months after discharge to all former adult icu patients who thereafter were followed for another 3 years. a reference group of 6093 individuals from the uptake area of the hospitals served as controls. results. 980 (59%) patients returned the questionnaires. the icu patients reported significantly higher mean scores in perceived hopelessness score compared with the general population, 2.5 (sd 2.2) compared with 2.1 (sd 2.1) (p \ 0.001), and 40% (n = 392) of the icu patients perceived a mean or high hopelessness score compared with 36% of the general population (p \ 0.001). the icu patients who died during the follow-up period reported a significantly higher perceived hopelessness score (n = 123) 3.7 (sd 2.5) (p \ 0.001) as compared with those who survived up to 3 years after discharge (n = 857) 2.4 (sd 2.1). in a logistic regression model the long term mortality for the icu group was found to be affected by: pre-existing disease [odds ratio (or):2.2], age (or: 1.04) and perceived hopelessness score (or: 1.18). the new and interesting finding of this study is that icu patients score higher on ''perceived hopelessness'' than a control population and this increase is predictive for the post icu mortality. furthermore, the size of this effect is significant and only exceeded by pre-existing disease and age. we performed a retrospective observational study to evaluate what proportion of met calls was associated with lomt issues. to estimate the proportion of met reviews involving patients with a not-forresuscitation (nfr) order and the timing of met calls in relation to admission and death or discharge from hospital. to compare the patient characteristics and outcome for met calls associated or not associated with lomt issues. we obtained hospital research ethics committee approval. we performed a retrospective observational study involving five-year (august 2005-april 2010) in a single tertiary australian hospital. we obtained information on demographics, on the met review and hospital outcome. lomt included nfr orders, not for met orders and palliative care plans. results. we analysed 4829 met reviews in 3629 patients. table 1 and fig. 1 summarize major findings for overall population and the two subgroups of patients with or without lomt. patients with lomt care plan were older, more likely to have medical diagnoses, were reviewed later during their hospital stay and closer to their hospital discharge or death. fewer lomt patients were admitted to icu. hospital length of stay was shorter, mortality in lomt care patients was double that of non-lomt patients. however, more 50% of patients with lomt were discharged alive from the hospital. conclusions. more than one third of met activations deal with lomt issues. although the mortality of these patients is high, a large proportion survives to hospital discharge. evaluation of the patient experience in intensive care (icu) frequently depends on reports from surrogates such as relatives. there is a concern regarding the validity of the surrogate opinion which might not represent the values of the incapacitated patient and treatment decisions therefore maybe biased [1] . others have found that there is a strong preference within a population for utilizing relatives as surrogate decision-makers in the event of admission to icu and this attitude is not influenced by ethnicity, religion or education level [2] . objectives. the objective was to measure the ability of the relative to answer on behalf of the patient. a further wish was to determine the validity of their surrogate responses. a retrospective study, which surveyed relatives of patients who had died within a critical care service during a 2-year period (2005, 2006) . the 31 item questionnaire allowed for the collection of quantitative and qualitative data with respect for each item to overcome the limitations of the quantitative format which may not be sensitive to all the issues which can surround the provision of end-of-life care [3] . for 3 items, relatives were asked specifically to grade their capacity to represent the patient. results. quantitative data from the 3 items designed to test the relatives' perception of their ability to act as surrogates indicates that relatives considered they could respond to these items for 46% (average) of instances. when the relative did answer on the patient's behalf, the level of concordance between the surrogate (relative) and the patient's perceived opinion was 60% suggesting that when the relative is willing to act as surrogate the response is likely to have validity. (table 1) . results from the qualitative data indicates that the low (46%) level of willingness to answer these questionnaire items reflected a reluctance to answer on behalf of a sedated or ventilated patient, rather than an inherent inability to represent the patient. conclusions. the response rate to the 3 items vindicates concerns regarding the ability of relatives to represent the patient in icu settings and supports a need for further study. where the relative is willing to act as surrogate, concordance does exist. qualitative data clarified quantitative results and was instrumental in promoting a better understanding of the concerns of relatives who have a family member admitted to icu. . the majority of patients that died in icu were provided some kind of therapy restriction. an important conflict strains between clinical practise, bioethical principle and jurisdiction laws; the solution of this conflict is more and more urgent. therapy restriction has also important economical aspects since the number and cost of available treatments constantly increase. our survey studied therapy restriction procedures in hungary for the first time. in 2007 we performed a survey with questionnaire among intensive care physicians. questionnaires were sent out electronically to 743 registered members of the hungarian society of anaesthesiology and intensive care. respecting anonymity we have statistically evaluated 103 replies (14%) with t test and anova. we grouped intensive care physicians based on gender, years spent in work, religion and type of department they were working, and we compared data from these groups. intensive care physicians generally make their decisions alone, based on the patient's long-term life prospects and physical status (3.75/5 points). they are slightly influenced by the opinion of the patient (2.57), the relatives (2.14) and other medical personnel (2.37). if the physician sees any chance of recovery but the patient or relative requests treatment restriction then 67.3% of physicians that completed the forms would continue therapy against the will of the patient or relative. only 27.7% would accept the patient's/relative's opinion and autonomy in such a case and would stop therapy. in fact 28.3% of physicians would make their decisions without considering or even against the opinion of patient if they think therapy is useless. if there is no chance of recovery despite medical treatment 3% of physicians stop the treatment, 8.1% would continue it without informing the patient or the relatives, 17.2% informs the relatives but continues useless treatment irrespective of the will of the patient or relative. having analyzed the groups we found two significant differences. in case of useless treatment physicians working in university hospitals more often choose treatment restriction without informing relatives (p \ 0.001) then those working in non-university hospitals. physicians who declare themselves as atheist rather choose the continuation of treatment without informing relatives (p = 0.007). conclusions. the hungarian practise of end of life decisions among intensive care specialists is paternalistic, physicians make their decisions alone, do not consider the requests of the patient or relatives. our goal is to strengthen patient autonomy and to support their opinion by training icu physicians. on the other hand it is inevitable to define what useless medical treatment exactly is and to introduce this category in medical ethics and also in jurisdiction practise. objectives. to determine the frequency and processes of eol care at our centre. between october 2007 and december 2009, 150/388 (39%) patients staying in the icu for more than 1 day, underwent some form of eol care in the icu. icu staff notified investigators whenever an eol decision was made. we recorded demographic details, documentation of the eol care process in the case notes, and interviewed icu staff to determine the eol care processes involved. results. 97 patients (65%) were male, 53 (35%) were females. mean age was 42.4 ± 2 years. icu stay was 6.7 ± 6.3 days, admission apache ii score was 18.1 ± 7.8 which increased to 24.9 ± 9.4 on the day of eol care decision. 33% patients had metastatic cancer. reasons for initiating eol care were refractory acute illness in 53%, advanced cancer in 51%, brain death in 11%, and lack of finances in 11%. eol discussions were initiated by the family in 11%, and by the icu medical team in 89% patients. 16 families wanted to take the patient home to die. the icu consultant was involved in all discussions with the family, the primary consultants in 91% and primary team residents in 79%. nurses were involved in only 3 patients. agreement on eolc was reached after 1 discussion in 70%, 2 discussions in 22%, and 3 discussions in 8% of cases. documentation of the eol care process was not done in 51% cases. withholding of life support (wh) was practised in 110/150 patients (73%) and withdrawal of life support (wd) in 27%. intubation was withheld in 14.5% patients, cardiopulmonary resuscitation in 50%, inotropes in 76% and dialysis in 8%. regarding wd, only 3/40 patients were extubated and the ventilator withdrawn in another 7/40 patients. inotropes were withdrawn in 20 patients (50%). reduction of fio2 0.21 without discontinuing mechanical ventilation was the commonest mode of wd, in 35 patients (88%). all patients received morphine infusions during lols/wols. family members were present by the bedside in 61% cases. conclusions. wh is preferred over wd. documentation of the eol process does not occur in a significant proportion of cases. nurses are rarely involved in the eol care decision making process. legal issues may be barriers to good eolc in our icu, and perhaps in india. objectives. to know the point of view of the staff is essential to understand their beliefs, attitudes and decisions. brazilian private general icu with 23 beds. the following items were analyzed: profile of the interviewed; their opinion about end of life questions: fear of death, fear of experience pain before death, the best place to die, advanced directives, decision-making process, therapeutic withhold of mechanical ventilation, nutrition, fluid management, antibiotics, vasoactives drugs, sedation and analgesia in patients which death is imminent and irreversible. results. about 84.41% of our icu team answered the research (n = 65). the mean age is 32.61 years (sd 6.74), 58.50% of female, 58.50% married, 50.85% protestants and 38.50% catholics and icu professional experience of 6.75 years (sd 4.65). using a visual analog scale (0, no fear to 10, the worst fear possible) the team pointed 5.96 as their fear of death; the fear of suffering pain before death was 8.17. for 44.60% of the responders, the best way to die would be with their lovely ones, no matter if at home or at hospital. only 9.20% would prefer to die an icu. the majority of the team (88.25%) would share the eol decision-making process with the family instead only by the medical staff (52.30%). about 73.10% would leave an advanced directive with their therapeutic preferences like do not resuscitation orders. the icu team agreed on the withdrawal of vasoactives drugs (41.50%), antibiotics (57.00%), nutrition (18.50%) and mechanical ventilation (1.53%) in patients out of treatment. our results showed the staff vision about their own death and their opinion about the end-of-life care issues. in developing country as brazil there is a still gap between everyday practice and the current legislation. fortunately, the debate about eol issues has increased in last years. the end-of-life discussions and decisions should begin by respect to points of view of all involved: patients, family, medical staffs with a legal support of the society's beliefs and expectations. prospective observational study conducted in 8 greek multidisciplinary icus. we studied all consecutive icu patients who died, excluding those who stayed in the icu \48 h or were diagnosed with brain death. 306 patients comprised the study population [mean age 64 ± 17 (sd) years, mean apache ii score on admission 21 ± 7]. results. of patients studied, 41% received full support including unsuccessful cardiopulmonary resuscitation (cpr). 48% died after withholding of cpr, 8% after withholding of other treatment modalities besides cpr, and 3% after withdrawal of treatment. patients in whom therapy was limited had a longer hospital (p = 0.01) and icu (p \ 0.01) stay, a lower admission gcs score (p \ 0.01), a higher apache ii score 24 h prior to death (p \ 0.01), and were more likely to be admitted with a neurological diagnosis (p \ 0.01). patients who received full support were more likely to be admitted with either a cardiovascular (p = 0.02) or trauma diagnosis (p = 0.05), and to be surgical rather than medical (p = 0.05). the most important factors affecting the physician's decision to provide full support were reversibility of illness and prognostic uncertainty; the physician's religious beliefs and legal concerns had minimal impact. the main factors guiding the decision to limit therapy were unresponsiveness to treatment already provided, prognosis of underlying chronic disease, and prognosis of acute disorder; old age was not a determinant, while economic cost and lack of icu beds seem to play no role. relatives' participation in decision-making occurred in 20% of cases and was more frequent when a decision to provide full support was made (p \ 0.01). the principal reason for not discussing end-of-life dilemmas with relatives was the fact that the family was thought not to understand (60%) advance directives were rare (1%). icus. however, in a large majority of cases, it involves the withholding of cpr only. withholding of other therapies besides cpr and withdrawal of support are infrequent. physician has a dominant role in decision-making. objectives. the primary objective of this study is to determine the prevalence of inappropriate or non-beneficial care in icu patients as perceived by their icu healthcare providers, as well as the reasons for this perception. second, we want to determine which factors are associated with the perception of inappropriate care. a descriptive survey design is used. a single-day cross-sectional evaluation of perceptions of inappropriate care among 2,327 icu healthcare providers in 85 icu centres in 10 european countries will take place on may 11th 2010. questionnaires will be administered to icu healthcare providers (nurses, head nurses, junior and senior icu physicians) providing bedside care to adult icu patients on that particular day. in this study, inappropriate care is defined as a patient care situation that is similar to one or more of seven scenarios. these scenarios were created based on the literature and a multidisciplinary conference attended by experts in intensive care, geriatrics, and palliative care. . the cross-sectional study will take place on may 11th 2010. preliminary results will be given at the esicm conference. we have designed a one-day cross-sectional study to record inappropriate or non-beneficial care in european icu's. results will be available for the esicm conference. grant since the introduction of the mental capacity act in the uk in 2007, the impact within research in the intensive care environment has not been elucidated. since many of the patients are incapacitated and therefore unable to consent, it is now stipulated by the ethics committee that the researcher must make reasonable attempts to identify a consultee, failing this, nominate a person unrelated to the research project to be consulted. in order to comply with the mental capacity act, retrospective consent must be obtained, once the patient regains capacity. objectives. the aim of the study was to highlight the difficulties in obtaining retrospective consent, evaluate the methods used and demonstrate the adaptations made to increase retrospective consents. methods. this explorative analysis investigated the process of obtaining consent in 246 patients enrolled in an observational study on critically ill patients. consent was obtained on admission if the patient had capacity. assent from the patient's next of kin or a legal professional representative was obtained before enrolment in patients who lacked capacity. after discharge from icu, a member of the research team re-visited these patients to explain their involvement in the research, its purpose, procedures, implications and any further participation required by the subject. at this point, the patient could consent or withdraw from the study. if the patient decided to withdraw from the study, all data collected and samples stored were destroyed. the researcher visited the patient for a minimum of two visits; firstly to explain the study; secondly to establish if the patient has retained the information and to gain retrospective consent. results. 246 patients were recruited within the time period of which 44 (17.7%) died. in 65 (26.2%), consent was obtained on admission as the patients had capacity, 36 (14.6%) were discharged prior to obtaining retrospective consent, 10 (4%) lacked capacity on the researcher's visits, and 1 patient (0.4%) withdrew from the study. 90 patients (36.3%) were successfully consented retrospectively. overall, the researchers performed 221 visits to obtain from the 137 patients for whom retrospective consent was required. conclusions. the process of recruiting patients who lack capacity within the intensive care unit is challenging and time consuming. stipulations set by the ethics committee to seek retrospective consent once the patient has regained capacity, has a major impact on research staff time and finances. detailed recommendations as well as guidelines how to assess capacity in the post-icu patient and how the assessment of capacity has to be applied to intensive care research are needed to fully comply with ethical and legal requirements. objectives. we wanted to know if patients expressed to surrogate decision makers, after icu discharge, specific resuscitation directives, and we have investigated any factors related to the patients and their illness or care process that might be associated with this. we reviewed patients admitted in the icu between december 2008 and may 2009. a random sample of 60 survivor patients has been defined. seven patients were excluded (3 for language barrier, 2 died, 2 were no more reachable). fifty three patients took part in semistructured interview at 6-12 month post icu discharge. the questionnaire discussed in detail the aspects of advance directives. patients had also completed a quality of life questionnaire (euroqol 5d), and we calculated the eq-5d visual analog scale. we reviewed medical records in icu data base: age, gender, length of stay, saps ii, bmi, length of ventilator support and central venous catheterization as well as prescription of transfusion, hemodialysis or adrenergic agonist. multivariate logistic regression was practiced to investigate any factor associated to expression of specific resuscitation directives after icu discharge. after icu discharge, 60% of interviewed survivors expressed specific resuscitation directives to an appropriate identified surrogate (written ''living will'' or oral statement). eq-5d visual analog scale was 58 ± 19. on multivariate regression analysis, only one studied variable was significantly associated to the post-icu expression of specific resuscitation directives: age (odds ratio = 0.94, z = -2.1, p = 0.035). conclusions. after icu discharge, a majority of our patients expressed to surrogate decision makers specific resuscitation directives, especially the younger patients. our findings suggest that surviving to icu is an opportunity to specify oral or written directive, and both may help to illuminate future decision making from the patient's perspective. objectives. to explore the issues around eolc provision for cancer patients in a critical care unit through family, professional and patient experiences. to explore how a diagnosis of cancer impacts upon eolc provision for critically ill patients. a heideggerian phenomenological interview approach was undertaken, in order to gain personal experiences. families of those patients who died after decisions to forgo lifesustaining treatment (dflsts) were interviewed. patients who were seriously critically ill (apache ii [ 25 or had received cpr) who experienced critical care were also interviewed, since patients' views about eolc provision are very rarely explored. doctors and nurses also contribute their vision for, and experiences of, eolc in a cancer critical care unit. thirty seven participants were interviewed. tensions between treating families versus treating patients impacted on timeliness of eolc. achieving a good death was possible through caring activities that made best use of technology to prevent prolonged dying. decision-making and eolc could be difficult to separate out which, in turn, affects prospects for eolc. three main themes included: dual prognostication; the meaning of decision-making; and care practices at eol: choreographing a good death. these themes outlined the essence of moving along a continuum toward patients' deaths and the impact that had on opportunities for care and a good death. conclusions. cancer affected the trajectory in unexpected ways. the trajectory could be very quick, especially in unexpected death and some newly diagnosed cancers. even in the face of a life-limiting and serious disease like cancer, death could be unexpected. the rapidity of trajectory related to cancer diagnosis, prognosis, withdrawal and patient demise significantly impacted on the potential for, and timing of, eolc. a sentiment of moving on from historical practices around critical care for cancer patients, and related poor prognoses, was overwhelmingly agreed on but important caveats in cancer prognostication remains. conclusions. these data suggest that oscillation settings of 5 and 10 hz provided more optimal pef/pif ratio ([1.1). our data also suggests that airway clearance using hfcwo may facilitate improved gas exchange in mechanically ventilated patients. further study is required to confirm these results grant acknowledgment. partial funding support in the form of devices was provided by hill-rom inc. a. esquinas 1 , m. folgado 2 , j. serrano 3 1 hospital morales meseguer, intensive care unit, murcia, spain, 2 hospital virgen de la concha, zamora, spain, 3 hospital reina sofia córdoba, intensive care unit, cordoba, spain objectives: we hypothesized that the use of intrapulmonary percussive ventilation (ipv) could effect hypercapnia/acidosis and airway secretions control during treatment with noninvasive mechanical ventilation (nippv) in exacerbations of copd associated with bronchial secretions. prospective multicenter study. the study was performed in the medical icu of 3 spanish university hospitals members of the spanish ipv working group. we enrolled copd exacerbation patients with secretions and the need for nippv in icu. criteria of exacerbations of copd are: a respiratory frequency c25/min, a pao 2 [ 45 mmhg and ph b 7.35. we define two ipv strategies as complementary treatment during nippv to evaluate the effects of ipv. strategy group i: nippv at first line and combination of ipv in early periods without nippv in spontaneous breathing and ph c7.35. strategy group ii: first line of ipv with mouthpiece/face mask and oxygenation previous to the application of nimv with ph \ 7.35. in both groups daily sessions ipv were applied by for 30 min/3 day by mouthpiece or face mask during stay in icu. nippv was applied with bipap ventilator (respironics) and face mask with bipap mode. cardiopulmonary monitoring, clinical and arterial blood gases were evaluated. therapy was considered as successful when patients did not need nippv support and clinical and arterial blood gases returned to baseline. results. 65 patients with copd exacerbation were admitted in icu for nimv, age 70 ± 12 years, male (90%) 15 were excluded for severe hypoxemia (pao 2 :fio 2 b 200) associated with pneumonia (5/15) and cardiac insufficiency (10/15). fifty patients were enrolled in the study. -up tilt-table rehabilitation better than sitting in a chair for ventilated adults in intensive care in terms of improving lung function? j. manners 1 , a. thomas 1 , s. boot 1 , g. mandersloot 1 1 barts and the london school of medicine and dentistry, london, uk physiotherapy intervention is a fundamental part of the patient stay in an intensive care unit (icu) and treatment is often aimed at maintaining/improving respiratory function. physiotherapists use the upright posture to elicit these improvements and sitting in a chair and standing with a tilt-table are commonly used interventions. to date there are no published reports comparing the efficacy of these interventions in ventilated subjects. • to compare the effects of these two positioning techniques employed with icu patients. • to measure changes in respiratory rate, tidal volume and minute volume during these positioning interventions. • to measure functional residual capacity during positioning interventions. • to measure the change in metabolic demand during positioning interventions. methods. convenience sampling of ventilated subjects meeting the inclusion criteria was employed. subjects acted as their own controls undergoing sitting in a trauma chair and standing on a tilt table at 70 degrees in random order on the same day. respiratory rate (rr), tidal volume (v t ), ventilation (v e ) and oxygen consumption (vo 2 ) were measured at minute intervals during baseline and intervention for 10 min. functional residual capacity (frc) was measured once at rest and following each intervention. measurements were recorded using the ''e-covx'' module for the ''ge carestation ventilator''. results. 10 subjects were recruited. no adverse events occurred during interventions. significant increases from baseline rr (p \ 0.0001), v e (p \ 0.0001) and vo 2 (p = 0.009) occurred during the tilt table intervention. there was an increase in frc during tilting of 0.6 l which failed to reach significance. significant increases from baseline rr (p \ 0.0001), vo 2 (p = 0.024) and a decrease in v t (p = 0.015) occurred with the chair intervention. conclusions. these interventions are safe in a critical care population. increased muscular activity associated with upright interventions elicited expected elevations in vo 2. the tilt-table produced an increase in v e driven by an increased rr at the expense of v t . v e was not elevated during chair sitting despite an increased vo 2 and was accompanied by an unexpected decrease in v t. introduction. uk guidelines about rehabilitation after critical illness highlight the need for outcome measures to determine patient progress and efficacy of treatment [1] . there is no consensus about the most appropriate measures of patient function. the austoms [2] tool was designed by therapists in australia to measure activity and function across nine scales assessing structural and functional difficulties and ability to perform activities. scales are split into four domains (impairment, activity limitation, participation restriction and distress/wellbeing) and scored from 0 to 5 with 0.5 intervals allowed. acceptable inter-rater variation is defined as an absolute difference of 0.5. austoms has not been appraised in patients recovering from critical illness. objectives. to prospectively determine the inter-rater reliability of the austoms physiotherapy scales in adult patients who had undergone cardiothoracic surgery and required critical care admission for over 5 days. methods. the therapy (physiotherapy and occupational therapy) team underwent a 1 h teaching session using the austoms handbook prior to commencing the trial. austoms was then used over eight consecutive weeks during the weekly therapy goal setting meeting. each week a patient was selected to be scored using the most appropriate functional scales. the clinical history was presented to the team by the therapist leading the patient's care. therapists were then asked to independently score patients across the four domains for each scale. reasons underlying differences in scores were explored by group discussion. the difference between the 10th and 90th centiles of the initial scores was calculated for each domain as a measure of inter-rater variability. results. 6-8 therapists were present at each meeting. respiratory function and musculoskeletal movement related function were the most common scales used. the mean difference between 10th and 90th centiles was greater than 0.35 (± 0.21) for all domains. none of the scales/domains showed consistent inter-rater reliability over the 8 week period. overall the activity limitation domains of each scale showed the least inter-rater variance of scores. clinical experience of therapist did not appear to influence scores. conclusions. the austoms outcome measure showed poor inter-rater reliability when evaluated over an 8 week period on our intensive care unit. further work is ongoing to evaluate the ability of austoms to reveal changes over time when scored by therapists. introduction. uk guidelines on the rehabilitation of patients after critical illness highlight the importance of establishing and reviewing individualised rehabilitation goals for all patients that are at risk of developing physical and non-physical morbidity [1] . our institution's practice is to create objective goals that are smart-specific, measurable, achievable, realistic and timed [2] . objectives. the aim of this audit was to prospectively collect data regarding the setting of rehabilitation goals in a group of patients admitted to a cardiothoracic intensive care unit. methods. all consecutive patients admitted under the intensive care team in november 2009 were included. data regarding the timings of initial physiotherapy assessment, goal setting, and concomitant sedation were collected using a structured questionnaire completed by the treating physiotherapist. results. 30 patients were admitted under the critical care team. 29 patients were assessed by a physiotherapist within 24 h of admission. of these 29, 21 had smart goals set within a median of 7 days of initial assessment (range 1-29 days). there was a correlation between level of consciousness and the number of days taken to set goals. patients who were fully conscious or drowsy on initial contact (n = 11) had a smart goal set in a median of 3 days. by contrast patients who were sedated/paralysed on initial assessment (n = 18) had goals set in a median of 12 days. initial goal setting did not include other therapists or the family. goals fell in to 5 categories, range of movement, hoisting out to chair for periods of time, sitting on the edge of the bed, transferring out to the chair by standing and mobility goals-i.e. walking set distances. the maximal interval between reviews of the patients' goals was 7 days. most patients had smart goals defined and regularly reviewed. however, despite physiotherapy assessment within 24 h of admission, there was often a delay in setting these objective goals. the need for continuous sedation acted as a barrier to explicit setting of goals. the results emphasised the need to improve patient and family/carer involvement with initial goal setting in order to be compliant with uk standards. objectives. investigation of ems effects on muscle strength and exploration of issues in relation to handgrip dynamometry in icu patients. one hundred seventy two consecutive patients with apache ii score c13, were randomly assigned to the ems (n = 86, age: 62 ± 18 years, apache ii: 18 ± 5) or the control (n = 86, age: 59 ± 18 years, apache ii: 18 ± 5) group. ems sessions applied daily in muscles of both lower extremities. the strength evaluation of various muscle groups of the upper and lower extremities was made clinically upon awakening with the mrc scale, ranging from 0 to 5 (normal strength) for each group. the same scale was also employed in the diagnosis of cipnm (mrc \ 48/60). a subgroup of these patients also performed handgrip dynamometry. results. fifty seven patients (ems: 28, control: 29) were finally evaluated. ems patients scored higher than controls (p b 0.05) in wrist flexion, knee extension, ankle dorsiflexion and right side hip extension, while they tended to perform higher in all other muscle groups (p: 0.20-0.10) ( table 1) . grant acknowledgment. this project has been co-financed by e.u. and the greek ministry of development. background. secretion removal is major aim of respiratory physiotherapy in intensive care. manual hyperinflation provides a tidal volume to the lungs that is greater than baseline. it is effective in secretion clearance and is frequently used [1, 2] . there is a limited evidence that addressed the effects of combining rib-cage compression and suctioning on oxygenation, ventilation, and airway-secretion removal in mechanically ventilated patients [3] . objectives. the aim of this study was to investigate the effects of manual hyperinflation administered in combination with expiratory rib-cage compression on lung compliance, gas exchange, and secretion clearance in mechanically ventilated patients. methods. twenty-two intubated, mechanically ventilated, and hemodynamically stable patients were studied. the patients received manual hyperinflation, with or without expiratory rib-cage compression, with a minimum 3-h interval between the two interventions. manual hyperinflation with or without expiratory rib-cage compression was performed for 5 min before endotracheal suctioning. respiratory mechanics and hemodynamic variables were measured 5 min before (baseline) and then 5 and 20 min after the interventions. arterial blood gases were determined 5 min before (baseline) and 20 min after the interventions. secretion clearance was measured as sputum weight. the two measurements were obtained on the same day. results. no significant differences were observed in gas exchange and secretion clearance between the two interventions (p [ 0.05). in each case, static lung compliance and tidal volume improved significantly at 5 min post-intervention (p \ 0.01), whereas at 20 min postintervention, only static lung compliance had improved significantly above baseline (p \ 0.01). our results suggest that the addition of expiratory rib-cage compression to manual hyperinflation does not improve lung compliance, gas exchange, or secretion clearance in mechanically ventilated critically ill patients. recently, there has been an interest in mobilization of acutely ill patients who are in an intensive care unit (icu). in the literature, the major safety issues while mobilizing critically ill patients has been outlined. cardiac reserve [(cr) (% of age predicted maximal heart rate)] and respiratory reserve [(rr), ratio of partial pressure of oxygen in arterial blood to the inspired fraction of oxygen (pao 2 /fio 2 )] are the important factors that can affect the ability to tolerate the mobilization. patient who has rr more than 300 and cr lower than 50% is considered to have sufficient reserve to tolerate mobilization [1, 2] . objectives. the aim of this study was to compare the effects of mobilization on respiratory and hemodynamic parameters in patients with sufficient and insufficient respiratory and/or cardiac reserve. mobilization events are divided into two groups (sufficient, insufficient) according to the pre-mobilization cr (sufficient, \50%; unsufficient, [50%) and rr (sufficient, [300; insufficient, \300). heart rate (hr), systolic/diastolic/mean arterial blood pressure (sbp, dbp, mabp), respiratory rate (rsr) and percutaneous oxygen saturation (spo 2 ) were recorded from the monitor. respiratory and hemodynamic parameters were collected just prior to the mobilization, just after the completion of the mobilization when the patient had been returned the supine position and 5 min of the recovery period and compared between the groups. a total of 87 abdominal surgery patients (47 male, 40 female) received 113 mobilization treatments in icu. the mean age was 60.1 years, mean body mass index (bmi) was 24.9 kg/m 2 , mean apache ii score was 20.5 and mean icu stay was 7.3 days. mobilization events included 88 (78%) sitting on the edge of the bed, 12 (11%) standing, 13 (11%) walking to chair and sitting in the chair. 85% (96) of mobilization events had insufficient rr and 15% (17) of mobilization events had sufficient rr. 74.3% (84) of mobilization events had insufficient cr and 25.7% (29) of mobilization events had sufficient cr. all respiratory and hemodynamic parameters were found similar in sufficient rr and insufficient rr group at all stages of the mobilizations (p [ 0.05). spo 2 was higher, while hr and rsr was lower at all stages in sufficient cr group compared to insufficient cr group (p \ 0.05). resting hr and cr may affect the safety of mobilization, for this reason it is important to consider respiratory and hemodynamic parameters prior to and while mobilizing the icu patients. introduction. obesity is a chronic disease and a major health problem. obesity in critically ill patients is associated with a prolonged duration of mechanical ventilation and intensive care unit (icu) length of stay [1] . objectives. the aim of this study was to investigate the effects of mobilization on respiratory and hemodynamic parameters in the critically ill obese patients. [ 30.00 kg/m 2 )] were included as soon as their cardiorespiratory stability allowed mobilization protocol. mobilization was defined as sitting in the bed, sitting on the edge of the bed, standing, walking to chair and sitting in the chair. heart rate (hr), systolic/diastolic/mean arterial blood pressure (sbp/dbp/mabp), respiratory rate (rr) and percutaneous oxygen saturation (spo 2 ) were recorded from the monitor. respiratory and hemodynamic parameters were collected just prior to the mobilization (supine position), just after the completion of the mobilization when the patient had been returned the supine position and 5 min of the recovery period. all parameters were compared with initial values. the ratio of partial pressure of oxygen in arterial blood to the inspired fraction of oxygen (pao 2 /fio 2 ) was calculated from the arterial blood gas samples before and after the mobilization. introduction. the use of respiratory therapy for patients with a variety of lung disease is a standard in medical care [1] , including in the intensive care unit (icu) setting [2] . in this context, it is widely accepted the routine use of physical therapy in several situations in the intensive care, such as the care of critically ill patients not requiring ventilatory support, assistance during the postoperative recovery and the assistance to critically ill patients requiring ventilatory support [3] . at present definitive recommendations cannot be made regarding the use of respiratory physiotherapy for decreasing relevant clinical outcomes in critical ill patients requiring mechanical ventilation. objectives. this study aimed to determine the impact of providing chest physiotherapy on the duration of mechanical ventilation, intensive care length of stay, intensive care and hospital mortality in mechanically ventilated patients. single-centre, randomized, controlled trial in a university hospital general intensive care unit (icu). were included in the study 139 patients aged more than 18 years, admitted to the icu needing mechanical ventilation for longer than 48 h. physiotherapists provide group intervention (p) with the intensity and frequency of therapy they felt appropriate based on their assessment of the likely treatment benefit. control patients (group c) only received suctioning, decubitus care and general mobilization. results. primary outcomes were icu and hospital mortality regardless of the cause of death. secondary outcomes were length of icu and hospital stay, length of mechanical ventilation, weaning and extubation failure. patients in the p group more frequently achieved parameters to start weaning, but there were no significant differences between p and c groups on weaning and extubation failure, length of mechanical ventilation and length of icu stay. there was fewer hospital, but not icu, mortality in the p group. conclusions. we demonstrated that respiratory physiotherapy decrease hospital mortality and suggest that this effect was, in part, secondary to the effect of the intervention on weaning from mechanical ventilation. introduction. critical illness can cause diverse cerebral dysfunctions ranging from unconsciousness to minor cognitive impairments (mci). severe cerebral dysfunction, as delirium, is known to affect outcome after critical illness but it is uncertain whether minor impairments affect mortality or morbidity [1] . objectives. the primary aim of this study was to estimate the incidence of mci in a group of general icu survivors immediately after icu stay and three and 12 months after discharge. secondary we wanted to explore if type of cerebral dysfunction after icu discharge affected mortality and morbidity. methods. patients admitted to our general icu were included prospectively. we included 80 patients. 11/80 (14%) were delerious and 14/80 (18%) were not delerious but had mmse \ 24 after icu stay. of the 55 patients with mmse c24, 28 were possible to classify as having mci or not. 18/28 (63%, 95% ci: 45-83%) were found to have a mci after icu discharge. on 3 and 12 months these numbers were respectively: 11% (95% ci: 0-20%) and 10% (95% ci: -1 to 21%) there was an increased risk of both death and being institutionalised at both 3 and 12 months regarding delirious patients and patients with mmse \ 24 compared to patients with mmse [ 24. no such differences were found regarding patients with or without mci. (tables 1 and 2) . conclusions. the incidence of mci after critical illness is high on discharge but drops on 3 and 12 months after. severe cognitive impairments affect mortality and morbidity, but minor cognitive impairments do not. objectives. this study analyzes mid-term survival and risk factors associated with survival of patients undergoing cardiac surgery in son dureta hospital. methods. 1938 patients were consecutively operated from november 2002 to december 2007. patients who were discharged alive from hospital were followed until december 2008. we did kaplan-meier survival analysis and logistic regression study of variables associated with mid term mortality. results. in-hospital mortality was 1.96% (95% ci: 1.36-2.60%). information was available on 1,844 (97%) of 1,900 patients who survived until hospital discharge. at the end of the follow-up period, observed mortality was 6.5% (ci 95%: 5.4-7.7%). survival probability at 1, 3 and 5 years of follow-up was 98, 94 and 90%, respectively. the mean time of follow-up was 3.2 years (range 0.01-6.06). patients c70 years showed a lower survival rate than patients \70 years of age (log rank \ 0.0001). age c70 years, history of severe ventricular dysfunction (ef \30%), diabetes mellitus, preoperative anemia and hospital stay were independently associated with mid-term mortality. conclusions. mid-term survival of patients alive after hospital discharge was very satisfactory. mid-term mortality varied according to age and several preoperative chronic diseases. a closed-ended questionnaire was developed by the nurse congress commission of the société de réanimation de langue française (srlf). an invitation to complete it online was sent by email to caregivers registered on the srlf push-list. results were analyzed by icu or by respondent. results. 731 caregivers working in 263 icus completed the questionnaire (65% were nurses, 24% were doctors, 9% were nurse's aides, 86% worked in adult icus and 14% in pediatric icus). 7% of adult icus (n = 222) had unrestricted policy but 58% had a visiting time of less than 4 h per day. at the opposite, 59% of pediatric icus (n = 41) had unrestricted policies. 63% of the respondents working in icus with a visiting time \4 h per day considered very useful or essential to enlarge visiting periods but 29% of them considered this enlargement as unhelpful. at the opposite, 9% of the respondents working in icus with unrestricted policy found very useful or essential to reduce visiting periods. 81% of caregivers working in icus with unrestricted policy but only 35% of caregivers working in other icus thought that an unrestricted policy was able to improve often or systematically the relations with families. moreover, only 8% of caregivers working in icus with unrestricted policy but 46% of caregivers working in other icus thought that an unrestricted policy disturbs the organization of care. 96% of respondents found very useful or essential to give information in a dedicated room whereas it was often or systematically done in only 52% of icus. identically, 97% of respondents found very useful or essential to give information to proxies with the patient's nurse whereas it was often or systematically done in only 54% of icus. some cares were often or systematically programmed for family participation in 56% of pediatric icus but in only 0.5% of adult icus. indeed, proxies often or systematically participated in nursing in 66% of pediatric icus but never in adult icus. at the opposite, proxies often or systematically participated in tracheal aspirations in only 5% of pediatric icus and in 0.1% of adult icus. conclusions. more than half of respondent's adult icus are closed but caregivers working in icus with unrestricted policy perceive it favorably. some improvements are also expected by caregivers on the use of dedicated rooms for information and on the participation of nurses in meetings with families. finally, participation of families to care is not a practice of french adult icu caregivers. methods. included: patients with dysfunction of two or more organs in the first 24 h, admitted and discharged from icu during 2008. excluded: neurocritical and politrauma patients. contact 1 year following discharge; questions were asked concerning symptoms related to a period in intensive care that presented following discharge and which were not present prior to admission. in the case that the patient was not contacted, the next of kin was asked. results. 154 patients included. general characteristics during admission to icu: 63% male; age 60.2 ± 17.5 years; sofa* 7 ± 2.8; apache** ii 16.8 ± 6.4; apache** iv 57 ± 18.8; length of stay in icu: 5.1 ± 28.4 days; 42.9% were on invasive mechanical ventilation and 17.5% on non-invasive mechanical ventilation. data collection was carried out over 18 ± 4.4 months, on average 19 months (range: 12-26 months). 18.2% (28 patients) had died at the time of contact. the person contacted was the patient in 33.3% of the cases, the spouse in 13.7% and immediate family (patient's parent/child/sibling) in 43.1% of the cases. 46.6% had difficulty sleeping following discharge from icu with an average time since discharge of 10.5 ± 8.9 months; 51.1% suffered feelings of sadness and difficulty in finding enjoyment which had persisted for 8.7 ± 8.1 months; 30.3% had experienced difficulty in concentrating over an average of 9.3 ± 8.8 months; 28.2% had suffered some form of memory loss after discharge over an average period of 8.9 ± 8.6 months; 76.5% presented with asthenia over an average of 10.8 ± 8.3 months; 64.7% had arthromyalgia over a period of 12.6 ± 8.4 months; 35.3% had experienced changes in appetite over an average of 6.6 ± 7 months; 43.3% had changes in intestinal habit over an average of 7.78 ± 9 months; of which 64.3% had diarrhoea, 30.4% constipation, and 5.4% both symptoms; 18.8% presented with headache over a period of 12.9 ± 9.2 months; 15.2% had tremors, that had not previously been present, over an average of 12 ± 7.3 months; 12.8% had experienced reduced vision, over an average period of 16.5 ± 7.1 months; 10.7% presented with speech/ language problems, over an average period of 14.6 ± 8.6 months; 14.5% exhibited newly presenting changes in micturition, over 16 ± 7.7 months. another less frequently occurring symptom was loss of hearing (1.3%). conclusions. severely ill patients that are admitted to icu frequently present with ''residual'' symptomatology following discharge, most notably arthromyalgia and asthenia. many of these conditions persist for months. intensive care unit (icu) readmission rates range from 3 to 12%, in spite of initial recovery from critical illness. previous researches report that the revised acute physiology and chronic healthy evaluation (apache ii) score at either admission or discharge is an important predictor for readmission after icu discharge. however, there are a few papers concerning the association of discharge apache ii score with readmission after discharge from surgical intensive care unit. objective. we compared the ability of the discharge apache ii score with that of the admission apache ii score in predicting readmission, especially early readmission within 48 h, after discharge from icu. conclusion. this study showed that both discharge apache ii score and admission apache ii score are useful predictors for readmission after icu discharge, but discharge apache ii score is only independent factor in predicting early readmission within 48 h after icu discharge. introduction. health related quality of life (hrqol) is decreased in former icu patients. in research outside the intensive care field it is well known that the psychosocial factors, coping strategies and perceived hopelessness affect hrqol. however, the influence of coping and hopelessness on hrqol after intensive care is unknown. objective. the aim of this study was to examine how coping strategies and perceived hopelessness among former icu patients compares to corresponding in a reference group. we also evaluated the effect of coping and hopelessness and icu related factors on hrqol. methods. prospective, multicenter study in three mixed icu's in sweden. patient demographics, length of stay, apache ii score, reason for admission and time on ventilator were collected for all adult patients. questionnaires, including the coping instrument pearling-schooler mastery scale (pms), the 2-item hopelessness scale, sf-36, demographic data and previous illnesses were sent 6 months after discharge from hospital to the patients. the reference group (n = 6,093) was a random selection of persons from the same catchment area as the study patients. . 780 (59%) icu-patients, 18-74 years, returned the questionnaires. the patients reported significantly lower mean scores in coping 13.6 (sd 4.6, p \ 0.001) and higher perceived hopelessness 2. conclusions. this study indicates that coping strategies and perceived hopelessness are important for the hrqol of previous icu patients. however, the magnitude of these effects are smaller than that of pre-existing diseases. introduction. mortality on a medical intensive care unit (icu) is estimated to occur in about 25% of patients. its association with age, severity of illness and comorbidities is well established. for other diseases like coronary artery disease it has been shown that pre-existing depression is a risk factor for worse outcome. the role of depression regarding the outcome of icu patients has not been investigated so far. we studied the association between pre-existing depression and mortality in medical icu patients and present preliminary data of this ongoing study. objectives. assessment of a possible association between mortality of icu patients and prevalent depressive mood at time of icu admission. the primary endpoint was 28-day mortality. methods. prospective cohort study. all patients admitted to a medical 8-bed icu in a university hospital, older than 18 years, were eligible. postoperative patients and patients who had an expected length of stay below 48 h (survey) were excluded. patients whose cognitive function allowed appropriate comprehension and response answered the hospital anxiety and depression scale (had). prevalent depressive mood at admission was defined by a score c8 in the depression dimension. all other patients were assessed by observer rating by next-ofkin. in this case the hammond scale, a validated instrument for observer rating of depressive mood (cut-off c4), and a modified version of the had for observer rating (cut-off c10) were used. in addition apache ii, saps ii, sofa, age, sex, comorbidities, reason for admission, length of icu stay and ventilator days were recorded. . by now 173 patients had complete follow up data. of these patients 53 (31%) were classified to have depressive mood at icu admission. in total 46 patients had died by day 28 (27%). the 28-day mortality was 40% (21/53) in patients with depressive mood and 21% (25/120) in patients without (p = 0.01). patients with and without depressive mood did not differ with respect to age, sex, apache ii, saps ii or sofa score at admission. multiple logistic regression analysis with 28-day mortality as the dependent variable revealed that prevalent depressive mood at the time of icu admission was an independent risk factor for mortality (table 1) . conclusions. pre-existing depressive mood is an independent risk factor for mortality in medical icu patients. introduction. some classical post-icu discharge predictors of death are described, such as age, severity of disease and level of nursing care [1] . besides these factors, some laboratorial data at icu discharge are potential predictors of post-icu death. objectives. the aim of this study was to investigate whether standard base excess (sbe), ph, lactate, hemoglobin level, creatinine, platelets, leukocytes and albumin at the icu discharge as well as the 25% decrease on c-reactive protein concentrations (crp [ 25%) from the day pre-icu discharge to the day of icu discharge may be useful predictors of in-hospital outcome. patients discharged from the icu after at least 72 h of stay were retrieved from our prospective collected data base. a multivariate analysis was performed using a backward-lr binary logistic model taking in-hospital death as a dependent variable and the cited data as independent variables. results. 396 patients were retrieved. the average age was 49 ± 19 years old, mean apache ii score was 17 ± 8, and the main causes of admission were septic syndromes and respiratory failure. the in-hospital mortality after icu discharge was 18%. the icu length of stay was 9 ± 9 days. at the time of icu discharge ph was 7.41 ± 0.04, sbe was -0.3 ± 4.0 mmol/l, lactate was 1.9 ± 1.0 mmol/l, hemoglobin 9.6 ± 2.0, creatinine was 1.3 ± 1.46 g/dl, albumin was 2.68 ± 0.64 g/dl, platelets was 280,645 ± 166,100/mm 3 , leukocytes was 10,795 ± 8,769 cells/mm 3 and the number of patients who lowered crp at least 25% were 159 (40% conclusions. this study demonstrated that sbe, lactate, hemoglobin and albumin concentrations on the day of icu discharge are independent predictors of in-hospital mortality. moreover, the reduction on crp levels above 25% in the last 24 h of icu stay is a strong predictor of better in-hospital clinical outcome. we suggest that these variables together with the clinical judgment may be taken into account on the icu discharge decision process. readmissions to the intensive care unit (icu) are usually associated with increased morbidity and mortality, and they may evidence the quality of patients' care. the risk for icu readmission varies across studies, and is generally analyzed just before icu discharge, leading to deviation of icu team and patients' daily goals. early prediction may improve the care for patients in risk for icu readmission, and help developing mechanisms for its prevention. objectives. to analyse risk factors for readmission in intensive care unit looking at the first 24 h data after unit admission. methods. the first intensive care unit admission of patients was analyzed from january to december 2009 in a medical-surgical unit. readmission to the unit was considered those during the same hospital stay or within 3 months after intensive care unit discharge. deaths during the first admission were excluded. demographic data, acute illness and comorbidity prognostic scores, and use of mechanical ventilation were submitted to uni and multivariate analysis for readmission. numeric variables were expressed as median or percentage. conclusions. age, medical admission, sofa score and respiratory-and/or sepsisrelated admission are early associated with increased icu readmission risk. objectives. the aim of this study was to examine patient perceived hrqol in former icu patients that die in the period from 6 month up to 3 years after discharge from intensive care unit and the hospital. methods. prospective, multicenter study in three mixed icu's in sweden. questionnaires, including hrqol (sf-36), demographic data and previous illnesses, were sent out six, 12, 24 and 36 months after discharge to all former adult icu patients. data for this study were only collected among those dying before the 36 months post-icu follow-up. of the 980 patients who returned the questionnaires 123 (12.6%) died, 31 (25.2%) between 6 and 12 months, 55 (44.7%) between 12 and 24 months, and 37 (30.1%) between 24 and 36 months. the most frequent admission diagnoses were respiratory problems n = 36 (29.3%) and gastrointestinal diseases n = 33 (26.8%). examining hrqol in the former icu patients the following observations were made: (see fig. 1 ). a pronounced and quantitatively large decrease in hrqol is seen for the surviving patients with pre-existing disease as compare to the previously healthy survivors. although already at a very low value further decreases in hrqol for the patients dying before 3 years post icu is significantly less as compared to the icu patients with pre-existing disease that survives. the decrease is mainly in physical function, role physical function and role emotional function (marked in the figure). conclusions. yes, health related quality of life is extensively affected, mainly in the dimensions physical function, role physical function and role emotional function. importantly, in these two affected physical dimensions a shorter time to death increases such a decrease. the finding further stresses the importance of pre-existing diseases for the final hrqol outcome of former icu patients. introduction. despite initial recovery from critical illness requiring icu admission, many patients remain at risk of subsequent deterioration and death [1] . recent studies have shown readmission rates ranging between 4 and 7% [2] ; this population had mortality rates six times higher and were eleven times more likely to die in hospital [1] . 1. to calculate the readmission rate in our mixed icu unit over a 6 months period 2. to identify risk factors associated with readmission into the icu 3. to study the outcomes of these readmissions methods. a retrospective observational study, data was collected from an icu computer database (metavision) and analysed manually results. the total number of admissions in this period was 365, average patient age was 57 ± 17 with 52.4% being males. readmissions constituted 10.5% of the total admissions with 23.8% of those readmitted within 24 h of their initial discharge. 40% of the initial discharges from the unit were made out of hours i.e. unplanned, presumably due to heavy demand on beds. readmissions were particularly associated with patients discharged to surgical wards 23.8% and the hepatobiliary hdu 28.6%, the latter might reflect the proportion of that particular patients population received. 52.4% of the readmissions required to stay 72 h or less in icu. the overall mortality of the patients requiring more than one admission in this 6 months period was 57.1%. there is an urgent need for expanding icu services in our hospital, i.e. extra beds, staff, outreach teams, etc in addition to investing in nursing capacity building especially in surgical wards. we agree with others studies that compared with the general population, icu survivors report lower hrqol. moreover, a relationship between several factors like sepsis, renal failure, sofa (first and second day score), critical illness polyneuropathy, mechanical ventilation, sedation time, previous psychiatric history and blood products transfusions were found in our study population. conclusions. according to our data, subclavian vein was the most common insertion site used, especially as 2nd and 3rd placement and was related with the lower incidence of becteremia episodes. although the risk of placing a cvc for inflection complications is against the risk for mechanical complications, we have to improve our cvc policy, preferring the subclavian or the jugular site of insertion, in order to minimize the infection risk for a nontunneled cvc. objectives. objectives for this study were to determinate the frequency and the risk factors associated with bos. secondarily, we searched several variables as civil status, age, sex, work seniority as potential risk factors. inclusion criteria were to work in critical care unit (ccu) the hospital clínico universidad de chile (hcuch). this unit included 2 subunits: intensive care unit (icu), middle care unit (mcu). the mbi ò instrument was applied between april to july of 2009. all staff of ccu were asked to response the instrument. as previously reported, bos was defined with high ee, high dp and low pa. risk of bos was anything of the three dimensions positive for bos. we gave information on specifics objectives and the schedule of a future intervention programme. for analysis, comparisons were made based on student t test, chi-square test with yates corrections or fisher exact test as corresponded. for all tests we used confidence interval 95% with p \ 0.05. a total of 111 mbi ò tests that included all sub-units in ccu. this is a 56% of all personal working in the ccu. bos was found in 8.26% of cases. women (75%), unmarried (75%), with an average of age 31.9 years old. (23-47 years old) and with a work seniority younger than 5 years (50%). ee is high (31.5%), for nurse and paramedical personals. dp was 22.5 and 31.5% to middle level, for nurse and medical doctor, and low pa in 34% for paramedical personal, with longer work seniority (more than 10 years). risk factors were female gender, unmarried status, childless, middle aged (23-47 years old) and recent start in the job (stay younger than 5 years). introduction. burnout is a prolonged response to chronic emotional and interpersonal stressors on the job, and is defined by three dimensions: exhaustion, cynism (depersonalization), and inefficacy. icu physicians are exposed to several stress factors and are particularly predisposed to this syndrome. to describe the prevalence of burnout syndrome among intensivists and its relation to their quality of life. methods. an epidemiological cross-sectional survey conducted to evaluate all adult icu physicians in salvador, ba (brazil), from october to december 2006. the quality of life and burnout syndrome were evaluated respectively by the whoqol-bref instrument and the maslach burnout inventory (mbi). burnout was classified into low, moderate and high levels for the three studied dimensions, according the mbi classification, and it was defined by the presence of a high level in at least one dimension. the quality of life was evaluated in four domains: physical, psychological, social relationships and environment, graduated from 0 to 100, with higher scores denoting higher quality-of-life. [1] ) has been successfully used to measure nursing workload on an intensive care unit over a 24-h period. in contrast to intensive care, the nursing care workload on mc is not evenly spread over a twenty four period, but tends to vary between shifts. objectives. the aims of this pilot study were (1) to assess the fitness of nas as an accurate reflection of nursing workload on an mc unit. (2) to determine the nursing work load, per patient, per 8 h shift. prior to the commencement of the study all thirty one nurses taking part received instruction in the content and registration of nas. at the end of each 8 h shift, each nurse retrospectively scored their patient(s) using nas. this consists of a check list containing twenty three items giving a possible score between 0 and 177, where 177 equates to 1.77 full time equivalent (fte) intensive care nurse. the nas were entered in to a database and the average scores, per patient, per shift were calculated. three hundred patients were retrospectively scored over a 2-month period in october and november 2009. not all patients were scored on all three shifts as some patients had been transferred out of the unit before shift end. in addition any incorrectly completed forms were discarded and excluded from the study. methods. this multicenter pilot study included 14 doctors working at (pediatric) intensive care units (icu). subjects were randomly assigned to two groups: one was first tested during day, then during night, while the other was tested in reverse order. the d2 test of attention [3] was used to assess attentional performance. total performance (tn-f) score, standardized for age and level of education, was used to express attentional performance. subjective, 1-to-10 scores were gathered in two questionnaires. results. figure 1 displays standardized total performance scores of 14 doctors. measured attentional performance showed high intra-and interpersonal variability and did not differ between both shifts (p [ 0.9). in contrast, doctors expected alertness to be decreased (7.5 ± 1.7 and 6.1 ± 1.7 (mean ± sd) on subjective 1-to-10 scale during day and night shifts, respectively; p \ 0.02) and the chance of making errors to increase (from 3.3 ± 1.7 to 6.2 ± 1.3 (mean ± sd); p \ 0.01) during night shifts. conclusions. physicians working at icu are aware of the risk of making errors during night shifts. however, we showed that doctors perform equally during night and daytime when confronted with a short-time challenging task. consequently, a discrepancy between measured attentional performance and expected alertness was observed. these results suggest nocturnal alertness might be comparable to daytime during short-lasting tasks that elicit a high level of stress and motivation (e.g. testing, medical emergency). further research is needed to elucidate if longlasting (routine) tasks reflect decreased sustained attention and contribute to medical errors. we studied 297 physicians, the majority of whom were male (70%). mean age and time since graduation were 34.2 and 9 years, respectively. high levels of emotional exhaustion, depersonalization, and reduced personal accomplishment were found in 47.5, 24.6, and 28.3%, respectively. prevalence of burnout syndrome, defined as a high score in at least one dimension, was 63.3%, while prevalence was 7.4% for all three dimensions. in conclusion, burnout syndrome was common in this sample of icu physicians. aims. our goal was to assess the physician's opinion about potential competencies of a triage nurse. a representative cross sectional study design was applied with self-fill-in questionnaire about physician's attitude related to skills of triage nurses. the questionnaires were distributed between september and november 2007 in 24 (out of 31) eds. in this survey 159 physicians' questionnaires were processed. chi-square and student-t test was used for comparison of variables. p values less than 0.05 were considered statistically significant. results. 79.2% of physician would support the special training of triage nurse. 55.3% of physician suggests that the nurses use the patient's physical examination regularly in eds. the full time (ft) emergency physician significantly would reduce the basic competencies of nursing (e.g. dressing, feeding of patient, p = 0.008, and p \ 0.001, respectively) than parttime (pt) emergency physicians. significantly greater part of the ft physician would widen the competency of triage nurses in the field of physical examination of nervous system (p \ 0.001) and cardiovascular system (p = 0.005) than the pt physician. conclusion. hungarian emergency physician would widen the competency of triage nurse, but only half of physician would like to that nurses apply physical patient examination in practice. the full time physician would give more competencies for triage nurse than part time ones, but the final field of competency will be depended on other factors. healthcare-associated infections (hcai) are estimated to affect 1.4 million people worldwide, causing longer hospital stay, increasing hospital costs and excess mortality [1] . hand hygiene represents the single most effective way to prevent healthcareassociated infections. compliance with hand hygiene amongst healthcare workers (hcw) has been demonstrated to be quite low at 40% [2] . to quantify the degree of compliance to hand hygiene norms in the icu and to assess the short term success of strategies to improve hand hygiene compliance. setting. 34 bedded medical-surgical icu in a tertiary care centre. design. prospective observational. method. unobtrusive observer (single person). observed over sessions of 1 h. the compliance was calculated as :number of times the staff performed hand hygiene/number of hand hygiene opportunities. the number of hand hygiene opportunities was based on the who tools [3] : before touching a patient, before clean/aseptic procedures, after body fluid exposure risk, after touching a patient and after touching patient surroundings. introduction. icu delirium represents a form of brain dysfunction that in many cohorts has been diagnosed in 60-85% of patients receiving mechanical ventilation. delirium is a common but complex clinical syndrome characterized by disturbed consciousness, cognitive function or perception, which has an acute onset and fluctuating course and is associated with poor outcomes. and yet, it can be diagnosed and treated. in the uk, reporting of delirium is generally considered to be poor. in light of updated nice guidelines on delirium due out this year, specialist clinical assessment will soon become gold standard as a means of diagnosing and reducing the prevalence of this condition in the icu setting. nice recommends that cam-icu (confusion assessment method) be used by healthcare professionals who are trained and competent in the diagnosis of delirium. on our 16-bed unit, we are currently implementing cam-icu assessments to be performed twice daily (at the commencement of each nursing shift) as well as rass (richmond agitation and sedation scale) scoring on an hourly basis for all patients. objectives. to implement training of all our icu nursing staff in the use of cam-icu and rass scoring. to periodically validate and reinforce earlier training, so as to improve assessment and reporting of delirium. methods. our 'delirium group' comprising both nursing and medical staff, taught cam-icu and rass to 48 staff members using multimedia presentations in small groups and/or individual teaching sessions over 6 weeks. scoringofcam-icu andrasswassubsequentlyauditedon3occasions post training. discrepancies were discussed and post-audit retraining provided where necessary. results. the following audit and validation data were generated on our unit as documented in table 1 . no statistical analysis was undertaken. we anticipate focusing on the challenges encountered and strategies used in managing this change in our icu practice. methods. the factors causing resistance to change based on multisource data. qualitative technical methods were used: brainstorming and focal groups. the data collection elaboration was created by the collaboration of icu nurse, quality department nurse and external reviewers. finally, the main factors were classified in different categories. each category was scored by 1 to 5 according to gravity and prevention possibility. finally, priority was given to more serious and easier prevention problems. results. the most serious problems for icu professional was the historical factors. the easier solution problems were ''the lack of information'' and all evaluators were agree with it. we arranged the factors in order to the next classification (tables 1 and 2 ). discussion. all investigators were agreed with the low importance of problems with payments and low prevention probability of low organisational flexibility, so they were agreed on not to work about them. the icu professionals were more pessimistic and have lower confidence in prevention possibilities but they showed more confidence about the capacity to learn new skills. they weren't worried about resistance to do experimental things. probably, historic factors play an important role in this pessimistic attitude. on the other side, quality and safety experts have more experience in prevention programs and they put all their trust in its. after doing the analysis, we chose the ''lack of information problem'' to plan prevention activities. we consider it is a serious and real problem but at the same time, easy of prevent. conclusions. the implementation of the patient safety program in the icu means a real cultural change. the priority analysis could help to plan strategies in order to avoid the program failure. objectives. we concerned about whether medical personnel could recognize management of the cuff of artificial airway or not. we asked to doctors and nurses working in intensive care unit of konyang university hospital, daejeon, republic of korea. we asked 12 questions with 18 contents of questionnaire that was composed of methods of set initially, maintenance and appropriate pressure of cuff. results. 96 of medical personnel replied to us. most of them had worked in intensive care unit, so they had placed of artificial airway. 86.4% of them used manometer to adjust the cuff. we could find that nurses had more cognition compared to doctors for it (94 vs. 76%). only 28.5% of doctors described pressure of the cuff in medical record. 89 of medical personnel replied that they knew the appropriate range of cuff pressure. 37% (32/89) of them replied that the range of cuff pressure was kept with 25-30 mmhg and 34% (30/89) was 20-25 mmhg. 70% of nurses in the icu knew that range of cuff pressure was 15-25 mmhg. most of them knew complications of high and low pressure of the cuff. 69.8% of medical personnel monitored the cuff balloon during receiving mechanical ventilation and they used manometer to adjust it. 78% of nurses knew that the cuff should be adjusted continuously, but 58% of doctors did. interval measuring the cuff pressure was 38% of once a day, 38% of three times a day, 17% of more than four times a day conclusions. most of the medical personnel knew to keep appropriate cuff balloon to prevent various complications of artificial airway. they had insufficient cognition about maintaining the cuff balloon and appropriate level of cuff pressure. that was more prominent in doctors than nurses interhospital transfer is occasionally required as a consequence of limited therapeutic options or because of a need for a higher intensity of medical care that cannot be given in rural intensive care units. along with the potential benefit for the to be transferred patient, transport may also lead to hemodynamic and pulmonary deterioration. in order to minimize additional risk of interhospital transport of critically ill patients, a mobile intensive care unit with a specialized retrieval team was established in our university hospital-based intensive care unit. from march 2009, transport of the critically ill patients in our adherence region are performed by micu. objectives. in this prospective audit adverse events and patient stability during micu transfers were assessed and compared to our previous data on transfers performed by standard ambulance [1] . results. 74 interhospital transfers over a 10-month period were evaluated. systolic blood pressure, glucose and haemoglobin were significantly different at arrival compared to departure, although never significant values for major deterioration were reached. an increase of total number of variables beyond threshold at arrival was found in 38% of patients, 32 percent exhibited a decrease of one or more variables beyond threshold and thirty percent showed an equal number of trespassed thresholds. there was no correlation between the patients status at arrival and the duration of transfer or severity of disease. icu mortality was 28%. compared to standard ambulance transfers of icu patients performed in 2005, there were far less adverse events: 12.5 vs. 34%, which in the current study were merely caused by technical (and not medical) problems. although mean apache ii score was significantly higher, patients transferred by micu showed less deterioration in pulmonary parameters during transfer than patients transferred by standard ambulance. conclusion. transfer by micu imposes less risk to critically ill patients compared to transfer performed by standard ambulance and has therefore resulted in an improvement of quality of interhospital transport of icu patients. introduction. previous studies in adult intensive care units (icus) reported rates of pre-mortem to post-mortem discrepancies ranging between 7 and 32% depending on the population studied. and, most of them were retrospective studies, which included small number of patients. to compare clinical and pathological diagnoses and to determine the types of errors in a large and multidisciplinary icu-patient population. we conducted a prospective study of all consecutive autopsies performed on patients who died in the icu of the hospital universitario de getafe, madrid, spain, between january 1982 and december 2007. the diagnostic errors were classified in two categories: class i errors that were major misdiagnoses with direct impact on therapy, and class ii diagnostic errors which comprised major unexpected findings that probably would not have changed therapy. conclusions. this study found significant discrepancies between clinical diagnoses before death and post-mortem findings. this reinforces the importance of the post-mortem examination in detecting otherwise unexpected diagnoses and improving the quality of care of critically ill patients. introduction. unplanned extubation is associated to a high risk of reintubation end correlates with increased risk of nosocomial pneumonia. on the other hand, reintubation significantly increases morbidity and mortality in critical ill patients, increasing the incidence of ventilator associated pneumonia (vap) rate and makes the airway management risky. objectives. the aim of our study was to test the rate of unplanned extubation as well as the reintubation rate in our icu, in order to evaluate the efficiency of our airway and weaning time protocols. methods. during a nearly 4 year's period, 216 patients admitted to the icu, mean age: 61.8 years, mean apache ii score: 18.5, mean los: 13.5 days, with predicted and actual mortality: 31.1 and 22.47% respectively. from these, 191 were intubated and included retrospectively in our study. 109 patients were extubated, while the others either underwent bedside percutaneous tracheostomy or died. we concerned that the number of days of mechanical ventilation were about equal to the number of days of intubation. reintubation was defined as the need to reintubate during the first 48 h after extubation. we recorded four episodes of unplanned extubation. three of them caused by malfunction of the tube due to secretions and airway obstruction and one of them was undesired extubation caused by the patient himself. the total number of days of intubation was 2,111, mean ± sd: 11.06 ± 11.87, min: 1, max: 51 days. therefore the rate of unplanned extubation was 1.897%, while the standard limit is below 15 %. the total number of reintubations was 19, while the total number of scheduled extubations was 109. therefore, the reintubation rate was 17.43%, while the standard limit is below 12%. conclusions.the recorded rate of unplanned extubation was low in our icu patients, below the acceptable limit, assuming that our sedation and airway management policy is effective. on the other hand, the recorded rate of reintubation was high in our study, above the acceptable limit. although a low rate of reintubation might indicate excessively long mechanical ventilation times, this did not recorded to our study. nevertheless, our data suggest that we have to improve further our weaning time protocols, making the extubation procedure safer, and avoiding risk factors for vap. . pvs such as inappropriate enrollment of patients with a contraindication to the study treatment may lead to excess harm in the active intervention group [1] and failure to deliver the study intervention according to the study protocol may underestimate true treatment efficacy [2] . full reporting of pvs may aid in the interpretation of rct results however there are no published reviews on this topic [3] . objectives. to determine reporting rates for key types of pvs and to investigate study characteristics that may be related to reporting. publications were excluded because they were subgroup or economic analyses of a previously published rct [6] , not a rct [5] , not published in the target journal [4] , systematic reviews [2] , or other reason [5] . median trial size was 700 participants (range: 20 to 162,387). 13/80 (16%) of rcts were single centre, 58/80 (72%) were industry funded and 21/80 (26%) reported negative findings. overall 71/80 (89%) of rcts reported some form of pv, these included: 51/80 (63%) patient compliance; 40/80 (50%) discontinuation of study intervention due to safety; 32/80 (40%) study intervention-related researcher error; 16/80 (20%) inappropriate enrollment and; 8/80 (10%) technical errors in randomisation. multi-centre rcts may be more likely to report study intervention-related researcher errors (44% of multi-centre trials vs. 15% of single centre trials, p = 0.06). academic trials were less likely to report discontinuation of study intervention due to patient safety (38% of academic trials vs. 80% of industry trials, p = 0.002) and were less likely to report technical errors in randomization (5% of academic trials vs. 23% of industry trials, p = 0.03). conclusions. multi-centre trials are accepted to be organizationally complex. on-site education may be required to reduce errors in study intervention delivery attributable to the research team. it is possible the apparent excess harm attributable to industry trials is a reporting artifact however, if it is real, it must be addressed. additional research is required to investigate patient safety-related pvs and technical randomization errors, which may be lower in academic trials. to determine the occurrence of harmless incidents and ae related to physician's competences in icus, disclosing their potential risk factors. conclusions. this prospective study was essential to identify the proportion of our icu admissions affected by md-inc and md-ae, disclosing their nature. our md-ae rates, affecting more than 40% of admissions, were higher than those described in prior general studies, including not only icus. among the detected md-ae, hypoglycemic episodes not related to insulin administration predominated, indicating important deficiencies regarding nutritional support. severity on admission and length of stay were important risk factors for the occurrence of at least one md-ae. a systematic measurement and analysis of unintended events (ue) have been recommended for patient safety and improvement of quality of care in critically ill patients. however, a spontaneous reporting system may be inefficacious in intensive care unit (icu) because of a poor data collection, particularly by physicians staff. objectives. the aim of this study was to evaluate the reliability of a staff spontaneous event report by comparison with events collected by an external observer in a surgical intensive care unit (icu). to facilitate the reporting and the analysis, we identified a series of events with a serial number and a colour code related to their for each of the following 5 macro-phases: icu bed booking, admission procedures, patient stay, discharge and emergency procedures. a specific structured form including ue's code and colour, date and hour of the event and type of patient has been prepared and proposed to staff 1-week for each month after a proper phase of education. the report was voluntary and anonymous and the data collected during the morning shift from september to december 2009 have been compared to those collected from an external observer. in the studied period, healthcare staff reported 53 ues: 51% collected by nurses, 47% occurred during the morning shifts and 67% were classified as moderate or severe. the rate of ue in the morning shift was 12 ues per 100 patient days. the external observer identified 55 events in 8 morning shifts with an incidence of 76 ues per 100 patient days. the violation of isolation rules for patient with multi-drug resistant bacteria infection both by icu staff and surgical consultant was the ue observed more frequently by the staff (26%) and by the external observer (35%). conclusions. the above data indicated that: 1. in our icu the incidence of ue is very high, particularly for compliance to isolation of infected patients and 2. the spontaneous reporting system under-estimated largely the real incidence of ues. introduction. importance of renal assessing in intensive care unit (icu) patients is unquestionable for a correct drug dosing, fluid requirements or decisions for renal replacement therapies. serum creatinine (sc) is a very common biochemical parameter in clinical practice for assessment of renal function. many equations have been designed to estimate creatinine clearance based on sc, but their capacities for providing a correct estimate of glomerular filtration rate (gfr) are suboptimal. this is even worse in critically ill patients due to malnutrition and/or immobilization. in clinical practice, despite its limitations, 24 h-urine creatinine clearance (crcl 24h ) is used as a reference method to determine gfr. data show that cystatin-c could be promising as an endogenous filtration marker in icu settings. objectives. to assess in a medical icu population whether the arnal-dade formula of cystatin-c clearance (cc) developed from serum cystatin-c (scc) shows better predictive performance of gfr than sc-based formulae, as regards to patients' renal function: crcl 24h c60 ml/min 1.73 m 2 or crcl 24h \60 ml/min 1.73 m 2 . results. all formulae showed notable bias from the reference method. interestingly, all equations based on sc-values clearly overestimated crcl 24h (cg: 19.6%; mdrd: 1.1%; fv-mdrd: 27.5%), whereas cc showed underestimation of these crcl 24h (cc: -17.5%). in the crcl 24h c60 ml/min 1.73 m 2 group (n c60 = 146; 89 patients), cc showed the best correlation indexes (cc-crcl 24h ; r 2 = 0.296, r = 0.544), the second most biased (-16.7%) and the worst precision (24.9%). in this group, mdrd was the least biased (-2.4%) and the most precise (21.3%). in the crcl 24h \60 ml/min 1.73 m 2 group (n \ 60 = 95; 54 patients), cc was the worst correlated with crcl 24h (r 2 = 0.214, r = 0.462), in contrast to mdrd (r 2 = 0.299, r = 0.547). in terms of precision, mdrd showed again better results than cc: 33.5% vs. 38.9%, respectively. conclusions. in our icu population, cc did not demonstrate a clear improvement on the remainder sc-based formulae in either of the two groups according to crcl 24h . however, in a patient with high mdrd values and suspicion of low gfr, cc could be useful as guidance before obtaining the definitive confirmation by crcl 24h . introduction. there are well established and robust techniques for measuring and categorizing renal function in people with chronic kidney disease (ckd). a number of rapid bedside estimates of renal function have been devised incorporating routine daily measurements, such as serum creatinine, in combination with demographic data (e.g. cockroft-gault, the mdrd series). the addition of serum cystatin c measurements to some equations may also improve accuracy of estimation. the current and accepted categorical classification of acute kidney injury (aki: akin/rifle) has been useful epidemiologically but does not provide a continuously variable measure of severity of aki which would be valuable for both clinical management and research. objectives. previously published abstracts have suggested a role for egfr in describing renal function in the critically ill but a more comprehensive analysis was needed. methods. 37 (21 male) (mean age 72 range 38-90) critically ill patients with aki were recruited. a 4 h creatinine clearance ( 4 crcl) (previously validated as a measure of renal function in critically ill patients) was measured and simultaneous blood sampling was done for creatinine, urea, albumin and cystatin c. various equations used to estimate gfr were compared to 4 crcl with regression and bland-altman analysis. all patients had a 4 crcl of\60 ml min per 1.73 m 2 introduction. epithelial-mesenchymal transition (emt), a key process in tissue development and repair, has also been identified as a major mechanism in fibrogenesis. the cytokine tgfb has been shown to induce transformation of epithelial cells into matrixforming and smooth muscle actin (sma)-expressing myofibroblast (mf) via emt. the other prerequisite is an injury-induced loss of intercellular contact, including adherens junctions (ajs). the classical experimental method to induce aj disruption is the uncoupling of e-cadherin-mediated contacts by low calcium medium (lcm). this concept has been termed as the two-hit model of emt (1). b-catenin, a scaffold protein of the aj, released by cell contact injury, can act as a transcription factor and has been shown to facilitate emt. however, the mechanism whereby cell contact injury promotes emt is not understood. our recent studies have shown that smad3, one of the main signal transducers of the tgfb pathway is a strong inhibitor of epithelial sma expression, by interfering with myocardinrelated transcription factor (mrtf) [2] . the latter is the main driver of the sma promoter, through it association with serum response factor (srf). intriguingly, b-catenin can bind to smad3. to clarify the mechanisms whereby aj injury promotes sma expression. methods. ajs were manipulated in kidney tubular cells, either by sirna-mediated downregulation of e-cadherin, b-catenin or through chemical uncoupling of ajs by lcm. protein expression was detected by western blotting and immunofluorescence microscopy, proteinprotein interactions were monitored by co-immunoprecipitation, and the activity of the sma promoter was determined by luciferase reporter assays. knockdown of e-cadherin promoted b-catenin translocation to the nucleus and induced a threefold rise in the tgfb-triggered sma expression. conversely, silencing of b-catenin strongly suppressed the two-hit (tgfb + lcm)-induced activation of the sma promoter, and inhibited sma protein and mrna expression by 50%. the same stimuli induced strong association of b-catenin with smad3. transfection of cells with a b-catenin expression vector dose-dependently prevented the inhibitory action of smad3 on the mrtfinduced activation of sma promoter. moreover the active (myogenic) mrtf-srf complex was restored, as b-catenin preempted smad3's inhibitory effect on the complex. these studies define a novel mechanism whereby epithelial injury activates the myogenic program, a central process in organ fibrosis. our results imply that b-catenin, liberated from the injured ajs, facilitates the activation of the myogenic program by preventing or mitigating the inhibitory action of smad3 on mrtf. these hitherto unknown interactions among smad3, b-catenin and mrtf represent novel targets to lessen fibrogenesis. introduction. in intensive care unit (icu) patients, kidney function is monitored by the creatinine clearance (crcl). it can be measured by two methods. urinary crcl (ucrcl) is directly measured, using the urinary and serum creatinine. but commonly crcl is estimated from serum creatinine (scr) alone, as estimated glomerular filtration rate (egfr); using equations validated in chronic kidney diseases. there is paucity of literature on validation and comparison of these methods in icu (hoste) . objectives. we compared 4-h timed ucrcl and egfr in the newly admitted critically ill. we also sought to ascertain the incidence of high crcl and the agreement between 2 methods in this subgroup. conclusion. the use of rifle criteria gives a high incidence of aki in the icu setting. in this unselected population of critically ill pts, cysc seems to be superior to cre in predicting pts who will develop aki and will need rrt during their hospitalization in the icu. early identification of high risk patients may allow potentially beneficial therapies to be initiated early in the disease process, before irreversible injury occurs. introduction. the contrast-induced nephropathy (cin) is consider to be the most frecuence reason of acute renal failure in hospitalized patients. they are defined by a fixed increase (0.5 mg/dl) o a 25% rise serum creatinine level after to be exposed 24 h to the contrast. the main complications are kidney and cardiac problems and this will lead to longer hospitalization and increased mortality. objectives. to compare cin occurrence after a injecting a iso-osmolar contrast (ioc, idixanol) or a low-osmolar contrast (loc, iohexol) to a group of patients submitted to coronary angiography, with o without percutaneous coronary intervention (pci). to establish unrelated cin markers and to evaluate the efficiency of the kidney protection protocol used in our hospital. conclusions. the loc was associated to a greater number of cin than ioc. patients who developed cin were significantly longer hospitalized. the use of point giving system that includes cin's predictors like dm, hematocrit \39%, ami, and treatment with diuretics helps us to classify cin risk and use a correct kidney protection protocol. introduction. the incidence of acute renal failure in the intensive care unit (icu) is around 50% of cases and is related to increase in mortality in patients who required dialysis as far as 80%. early detection of acute kidney injury (aki), after damage is not on set could be crucial to develop therapeutic strategies to modify the course of injury. blood and urinary concentrations of ngal are early biomarkers of aki 1 ; to date, little information exists regarding ngal usefulness in critically ill patients. objectives. to analyze: 1. the capacity of urine ngal (ungal) to predict akievaluated by rifle score-in critically ill patients and, 2. the ungal values in patients with sirs, sepsis or septic shock. methods. ngal was measured in urine sample by an automatic analyzer device (architect ci4100 ò ; abbott diagnostics) at admission and 48 h later in patients admitted to a general icu. patients were classified both by rifle score at admission and 24 and 48 h later and by ungal concentrations at admission. to the later classification, the cut-point for aki prediction was obtained by roc curve analysis. ungal values at admission were compared in patients with sirs, severe sepsis or septic shock. clinicians were blinded to ngal results. the study included 76 consecutively-admitted patients (29 female) with mean age 59.6 ± 17.6 years, and length of icu stay of 10.4 ± 10 days. fifty-four sirs, 6 severe sepsis and 16 septic shock. thirteen patients developed rifle f score, 9 of them at icu admission; extracorporeal renal therapies were required in 6 cases. when patients were classified according to their rifle score at 24 h of admission, ungal values at admission were: 26 (10-81) ng/ml in 48 patients with rifle 0, 163 (49-998) ng/ml in 5 with rifle r, 357 (151-596) ng/ml in 8 with rifle i and 384 (31-539) ng/ml in 10 with rifle f (p = 0.02). five patients were excluded, three died before 24 h with ungal 816 (17-1,029) ng/ml and two were discharged before 24 h with ungal 182 (13-352) ng/ml. the area under roc curve of ungal at admission for aki prediction was 0.84 (95% confidence interval 0.74-0.92, p \ 0.0001), with an optimal cutoff value of 114 ng/ml with 69% sensitivity and 84% specificity. forty-seven patients have ungal b 114 ng/ml. ungal concentrations at admission were 30 (10-132) ng/ml in 54 patients with sirs, 36 (10-235) ng/ml in 6 patients with severe sepsis and 400 (126-592) ng/ml in 16 patients with septic shock (p = 0.001). conclusions. urine ngal concentrations measured at icu admission appeared as a useful predictor of aki in critically ill patients; in addition, ungal concentrations showed an increasing pattern from sirs to severe sepsis and septic shock. 23rd esicm annual congress -barcelona, spain -9-13 october 2010 s403 introduction. two previous studies using the rifle criteria in intensive care patients have found the incidence of acute kidney injury (aki) to be 36 and 67%. however, these studies used calculated basal value of creatinine in a considerable proportion of their patients, which is a possible source of error. objectives. the aim of this study was to investigate the incidence and severity of acute kidney injury in intensive care patients using true baseline creatinine values. objectives. the aim of this study was to define the status of hcy and b vitamins at admission and 7 days of icu stay in critically ill patients, and to evaluate its relationship between them. a prospective study was done on critically ill consecutive patients with inclusion criteria: c18 years old, sirs and apache ii [15. hcy, b12 and folic plasma levels were measured by enzymoimmunoassay and enzymatic method. for b1, b2 and b6 in erythrocyte. permission was obtained from an institutional ethical committee and written informed consent was asked. results. at 0 and 7 days of icu stay 50 and 53% of patients were b 1 deficient, respectively. 12 and 20% were b 12 deficient on both times, respectively. folic levels show significant differences between 0 and 7 days of icu stay. we found association between b 2 vitamin and hcy at admission and 7 days. no differences were found between 0 and 7 days hcy values. introduction. cytochrome p450 3a (cyp3a), the most abundantly expressed cytochrome p450 enzymes in liver, are responsible for the metabolism of over 50% of drugs used across several therapeutic classes. in adults, cyp3a is represented primarily by the major isoform, cyp3a4, and a polymorphically expressed isoform, cyp3a5. individuals with at least one wild-type cyp3a5*1 allele synthesise functionally active enzyme while homozygotes for the *3 allele are functional non-expressers of the enzyme. the presence of functional cyp3a5 increases the hepatic metabolism of cyp3a substrates such as tacrolimus. ckd is known to reduce the hepatic metabolism of drugs via the cyp3a enzyme system and we have shown, recently, that aki has a similar effect and that the length of time with aki is the most important variable. we hypothesise that expression of functional cyp3a5 may reduce the impact of aki on hepatic drug metabolism as has been shown to be the case for drug interactions with the imidazole antifungals. methods. 72 (45 male) (mean age 72 range 23-90) critically ill patients with no aki and varying degrees of severity of aki were recruited. midazolam concentration was measured 4 h after intravenous administration as a probe-drug for hepatic cyp3a 4/5 enzyme activity (t 4 [midazolam] ). this is a validated method for testing cyp3a activity in critically ill patients. patients were excluded if they were on any known cyp3a4/5 inhibitors. results. two patients with severe aki had unexpectedly high t 4 [midazolam] . figure 1 demonstrates the following: without a cyp3a5*1 allele, the rate of midazolam metabolism increased with duration of aki (r 2 = 0.24; p \ 0.0001) (solid line). patients who had at least one *1 allele (dashed line) were protected from the inhibitory effect that aki has on hepatic drug metabolism (significant difference between the correlation lines p = 0.011). if the two major outliers are removed (dotted grey line) from the *3/*3 group (r 2 = 0.23; p \ 0.001), the correlation lines remain statistically different (p = 0.048). conclusions. the presence of an allele which codes for functional cyp3a5 protects critically ill patients from the inhibitory effect of aki on the hepatic metabolism of midazolam. thyroxine replacement therapy has become commonplace in the management of organ donors to reverse hemodynamic instability and homeostasis, yet the pharmacokinetics of thyroxine are unknown in this patient population [1, 2] . since t3 is only available in oral form, we studied the pharmacokinetics of oral versus intravenous t4 to determine if oral administration is suitable. objectives. (1) to study the pharmacokinetics of oral versus iv t4 therapy; (2) to determine if oral thryoxine therapy is suitable. with ethics approval and signed consent from the substitute decision maker, 34 patients who were determined to be neurologically dead and consented for organ donation, were randomized to receive either an oral or intravenous dose of t4 (2 mcg/kg). all patients received an oral and iv preparation; one of which was a placebo. this study was also double blinded and randomization occurred in blocks of 4-6. free serum levels of t4 and t3 were measured hourly until the time of organ procurement. the area under the curves (auc) were determined and compared using. results. there were 15 patients (13 males) in the oral versus 17 patients (8 males) in the iv group, with an average age of 61 ± 14 vs. 53 ± 17, respectively. there was no significant difference at baseline or 6 h between groups for hemodynamic variables, free t4, free t3 or tsh levels. the only exception was map where it was higher at baseline in the oral group and there was a significant increase at 6 h in the iv but not the oral group (77-96 vs. 91-89 in the oral). the auc for t4 was greater for the iv group (301 pmol/l/12 h) compared to the oral group (274 pmol/l/12 h). there was no statistically significant difference in any of the levels from 0 to 12 h between the oral and iv groups. oral bioavailability of t4 was 91%. conclusions. administration of iv t4 resulted in a slightly greater auc compared to oral administration. however, oral bioavailability of t4 in our population was very high, at 91%. t3 is currently the recommended thyroid replacement in neurologically dead organ donors. however, intravenous t3 is unavailable in many jurisdictions. iv t4 has been used as a substitute. our study shows that in this select population, oral bioavailability is high suggesting that oral t4 may be a reasonable alternative. further work is needed to determine whether there was a difference in the number and rate of organ retrieval in the oral versus intravenous groups. introduction. specific characteristics of metabolic derangements occurring in critical illness is domination of developing catabolic state particularly in acute necrotizing pancreatitis. as a result, we faced such a problem as developing a clinically apparent protein-calorie deficiency which is resistant to standard nutritional support. the treatment of acute necrotizing pancreatitis in chronic abuse patients is difficult to handle for the clinician and should include sufficient energoplastic supply. objectives. in our research we aimed to assess the efficacy of adding of ornithineaspartate complex in carbohydrate metabolism in chronic abuse patients with acute necrotizing pancreatitis. methods. 56 comparable chronic abuse patients with acute necrotizing pancreatitis (control group n = 28, mean age 45.1 ± 8.5; ornithine group n = 28, mean age 44.0 ± 8.1) received early parenteral nutrition from the moment of admission to hospital with universal system ''three-in-one''. ornithine group also received ornithine-aspartate complex by parenteral administration (40 g/day). on the second day the patients were admitted parenteral nutrition and tube feeding 24 h/day. the volume of parenteral nutrition was gradually decreasing. biochemical and metabolic endpoints were measured at baseline and on 6th day (nitrogen balance, amino acids spectrum, plasma whole protein, transferring concentrations, glucose and insulin levels) at the clinical laboratory in all patients metabolic disturbances with protein status and carbohydrate metabolism shifts were revealed. dynamic of the whole protein, albumin/protein ratio and nitrous balance in both group showed similar tendency of metabolic improvement. dynamic of essential and nonessential amino acids concentration remained normal showing adequate energoplastic supply in both groups. glutamine concentration in ornithine group remained stable and even increased by the 6th day of nutritional support, while in control group glutamine concentration was decreasing, and by the 6th day of nutritional support it was below normal values. in ornithine group higher levels of endogenous insulin at normal values of glucose and faster fisher index improvement were detected. conclusions. administration ornithine-aspartate complex in therapy of acute necrotizing pancreatitis in chronic abuse patients, probably, may influence on disease outcome. in ornithine group duration of delirium tremens causes was 4 ± 1 days versus control group (6 ± 1 days). restoration of metabolic activities confirms adequate nutritional support in both groups but ornithine-aspartate complex adding provides faster improvement of protein and carbohydrate metabolism. objectives. this study was designed to evaluate the nutrition indexes including serum prealbumin level as prognostic indicators of patient recovery in critically ill patients with comparing severity scoring systems. we selected 59 patients over 16 years old, supplied with total parenteral nutrition (tpn) for more than 7 days in surgical intensive care unit, ajou university hospital, suwon, korea. the serum prealbumin, albumin levels and total lymphocyte count were measured at the first, 3rd, 5, 7, 10, 14th days of nutrition support care by tpn. we checked apache (acute physiology and chronic health evaluation) ii score, saps (simplified acute physiology score), mods (multiple organ dysfunction score) and sofa (sequential organ failure assessment) score of patients. results. there were 43 male patients and 16 female patients with mean age 54.3 years. the mean day of sicu staying was 13.2 days. we compared two groups; survivor group (n = 49) and non-survivor group (n = 10). there were significant statistical differences in icu staying days (p = 0.021), apache ii score (p \ 0.001), saps (p \ 0.001), mods (p = 0.007) and sofa score (p = 0.005) between two groups. however, serum prealbumin level (p = 0.370), albumin level (p = 0.444) and total lymphocyte level (p = 0.584) did not showed significant difference between two groups. receiver operating characteristic curve showed low accuracy of serum prealbumin level as a prognostic factor (area = 0.559). prealbumin level showed correlation with albumin (r = 0.561), however did not show correlation with apache ii (r = -0.151), saps (r = -0.056), sofa (r = -0.056) and mods (r = -0.076). conclusions. nutrition indexes including prealbumin did not correlated with clinical outcome of critically ill patients. introduction. physical function is impaired following critical illness [1] . anaemia is a common complication of critical illness and has the potential to influence physical function [2] . it is not known whether anaemia affects the physical components of quality of life, the ability to carry out the activities of daily living (aodl) or the actual physical function of patients during recovery from critical illness. to determine the physical quality of life, ability to perform activities of daily living and actual physical function in a cohort of icu survivors dichotomised on the presence of anaemia at 3 months following icu discharge. one other organ failure were recruited from a general icu population. patients with a preexisting haematological condition were excluded. baseline and characteristics of icu stay were recorded. the patients were assessed with the sf-36 quality of life questionnaire (pcs), the frenchay activities index (fai) of aodl recalled for pre-morbid status and at 3 and 6 months, and the 6 min walk test (6 mwt) for actual physical function at 3 and 6 months following discharge from icu. organotopic measures of haemaglobin, creatinine, serum c-reactive protein and albumin concentration were also recorded. the results were dichotomised on the presence of anaemia at 3 months for statistical analysis. baseline characteristics were compared with student's t test. a 2 way anova was performed on the pcs and fai score as well as comparisons with t test between each time-point. the distance walked as part of the 6 mwt was compared with mann-whitney u test. patients who remained anaemic at 3 months were older, had a longer icu stay and had a greater requirement for inotropes during their icu stay. the pcs score of quality of life and the fai score was significantly impaired in both groups during follow up, but there was no effect of anaemia. the results of the t tests showed that there was a significant difference between the groups at 6 months for pcs but not for fai scores. the distances walked were severely impaired compared to the normal population (170 and 228 m at 3 and 6 months for anaemic group and 240 and 358 m for non-anaemic) in both groups was not significantly different between the two groups. the non-anaemic group did increase the distance walked significantly from 3 to 6 months. there was no difference between albumin, crp and creatinine concentrations between the groups. methods. this experiment was divided into two procedures. the first procedure is to choose two kinds of cell strains, including jurkat cell strain (comes from leukemia) and ccrf-cem cell strain (comes from acute lymphocyte leukemia).we cultivate this two kinds of cell strains to mature stage, then inoculate every kind of cell strain into four culture dishes, two culture dishes was stimulated by 1 lg/ml lipopolysaccharide(study group), and the other two culture dishes serve as blank control(not stimulated by 1 lg/ml lipopolysaccharide). eight hours later, we extracted the microrna in each culture dish. the second procedure is to use the technique of gene microarray to analysis the difference expressions of microrna. in the context of a high altitude expedition human subjects can safely be submitted to prolonged hypoxia and the resulting changes in mitochondrial function can be explored in a controlled fashion. the effect of hypoxia on immune cells-key players in the pathophysiology of sepsis-is of particular interest. to measure mitochondrial function of monocytes during prolonged hypobaric hypoxia. methods. serial blood samples were collected and oxygen saturation was measured in twelve climbers before and throughout a high altitude climbing expedition to pik lenin (7,134 m). measurements were performed at 440 m (baseline) and at the altitudes of 3,600 m (day 1), 4500 m (day 4) and 5,400 m (day 9) above sea level. pure monocytes were isolated by the use of an antibody-antigen mediated immunomagnetic cell isolation procedure and lysed for determination of activities of mitochondrial enzymes cytochrome c oxidase and citrate synthase. repeated measurements anova followed by least significant difference (lsd) post hoc test were used to compare results on different altitudes. mean oxygen saturation was 91 ± 3% on 3,600 m, and decreased to 86 ± 3% on 4,500 m and 81 ± 5% on 5,400 m (p = 0.001). we observed an increase in citrate synthase activity on all altitudes compared to baseline levels (p = 0.003). compared to the baseline, prolonged hypobaric hypoxia induced an increase in the mitochondrial respiratory chain enzyme cytochrome c oxidase enzymatic activity only at 4,500 m (p = 0.003). normalization of cytochrome c oxidase enzymatic activity by citrate synthase activity (relative enzymatic activity) yielded a decrease in relative cytochrome c oxidase enzymatic activity during hypoxia on 3,600 and 5,400 m (fig. 1) . expressing cytochrome c oxidase enzymatic activities as a ratio to citrate synthase is intended to act as a safeguard for potential differences in mitochondrial enrichment. conclusions. the data demonstrates that prolonged hypobaric hypoxia leads to a decrease in relative cytochrome c oxidase activity. this is due to an increase in citrate synthase activity as a marker enzyme for the mitochondrial matrix representing mass and/or number of mitochondria which is not counterbalanced by a corresponding increase of cytochrome c oxidase activity. results. glycocalyx degradation was increased in the lps-treated animals (0.3 lm, p \ 0.05) compared to controls. intracellular tissue no concentrations were two-to threefold higher in the lps-treated mice compared to controls (liver, kidney, heart, gut). the number of infiltrating mpo-positive cells increased significantly during endotoxemia. levels of both plasma arg and cit were significantly lower in lps-challenged mice than in controls, whereas plasma ornithine levels were significantly higher. conclusions. in this new developed murine sepsis model, the prolonged infusion of lps resulted in increased glycocalyx degradation and associate endothelial leakage. the enhanced no levels correlated with decreased plasma levels of arg and cit. our murine model with prolonged infusion appears applicable as a model for the human clinical situation, enabling adequate investigation of the influences of the arg-no metabolism on endothelial dysfunction in sepsis. critical illness polyneuromyopathy is a muscular weakness occurring in intensive care unit. one of the major risk factor is sepsis. an early decrease in membrane excitability was described [1] but corresponding mechanisms are imperfectly known. tnfa is released in the first time of sepsis and could be involved in the physiopathology. objectives. the aim of our study was to investigate tnfa effects on muscular voltage gated sodium channels (nav) in an in vitro model. early effects of tnfa on nav were analysed by macro-patch clamp on muscular fibers isolated from rat peroneus longus. measurements were performed on control fibers and after addition of tnfa at concentrations ranging from 2.5 to 100 ng.ml -1 . the effects of chelerythrine, a specific inhibitor of protein-kinase c (pkc), were also tested. experimentations were realised in a laboratory with permission of experimental research on animals and under the supervision of an authorized person (no 29-028). tnfa produced a concentration-dependant inhibition of nav currents (fig. 1) . maximal inhibition (75% of control current) was observed with concentrations from 50 ng ml -1 and above. this decrease was fast: 40% of maximum inhibition was observed in less than 5 min. moreover, chelerythrine inhibited tnfa action on nav. conclusions. in our experimental model, tnfa induce a rapid and concentration dependant decrease of muscular nav currents like observed in chronic sepsis [2] . as this effect is too quick to be a transcriptional one, and as it is blocked by chelerythrine, it can be assumed that tnfa action is mediated by a nav phosphorylation secondary to pkc activation. in conclusion we evidenced that tnfa reduce muscle excitability in the early stages of sepsis. further studies are needed to obtain a precise description of tnfa mechanisms. may also contribute to cell signaling and regulation of the immune response. nad(p)h oxidase in leukocytes and the vascular wall is a major regulated source of o 2 . we hypothesized that mice deficient in the p47phox (ko) component of nad(p)h oxidase would have less pulmonary inflammation than wild type (wt). we treated wt or ko mice with iv saline or lps and assessed lung injury by: 1. wet-dry-weight ratio; 2. leak of evans blue (eb) labeled albumin; and 3. histological score for edema. we used myeloperoxidase activity to indicate neutrophil (pmn) accumulation in lungs, and measured accumulation of macrophages and neutrophils in bronchial alveolar lavage (bal). apoptosis was assessed by tunnel staining. we also expression of icam-1, an adhesion molecule, and nitric oxide synthase (nos) enzymes, enos and inos (western and northern analysis) as well as nitrotyrosine formation. results. lung injury was increased in both groups. surprisingly there was greater eb leak in ko than wt at 2 h and a greater edema score at 4 and 8 h. pmn and macrophage accumulation in bal were the same in both groups at 2 h but greater in ko mice at 8 h. myeloperoxidase activity was similar at 2 h post lps in ko and wt indicating that similar accumulation of pmn in the lungs. apoptosis was increased in both groups at 4 h, but resolved in wt at 24 h and persisted in ko. nitrotyrosine was increased in both groups but appeared higher in ko. expression of enos and inos increased in both groups but was greater in ko than wt. conclusions. in contrast to our prediction, lung injury was greater in p47phox ko mice which indicates that this complex is not essential for lung injury. however, the injury was more severe and prolonged in ko mice indicating that o 2 may regulate the inflammatory response. introduction. septic shock remains the main cause of mortality in the icu, thus a persistent challenge. recently, dna and mrna analysis by microchip and gene expression by real time pcr highlighted 2 proteins s100a8, s100a9 and their complex, known as the calgranulins, as potential key prognostic markers for this disease: those two proteins, whose expression seems to be restrained to phagocytes cells are newly recognized components in sepsis-induced inflammation. moreover, they were shown to be at significantly higher concentrations in the plasma of septic shock patients that were going to die. in the contrary, those who were to survive saw their plasmatic concentration decrease, all severity scores in between the 2 population being the same. objectives. the aim of this study was to determine the repartition of these proteins in immune cells, their intracellular variation, at baseline and after cell activation and finally to understand the relation between their intracellular and extracellular expression. we used an in vitro model close to the immuno-inflammatory aggression that is septic shock. we stimulated in vitro for 1, 3 and 6 h whole blood from healthy volunteers using agonists found in the inflammatory storm that is septic shock (lps, fmlp, gmcsf, ifng). we also induced death cell, either using an apoptotic agonist, or by necrosis technics. we then analysed the intracellular variation of the calgranulins using flow cytometry technics. the extracellular quantification was made using elisa methods. all the statistic analysis were made using a mann-whitney test. we showed in this work for the first time that the intracellular repartition of the calgranulins is different depending on the type of cell: the complexe is the main form in the monocyte cytoplasma, whereas s100a9 is the main intracellular form of the pmn. this repartition remains after cell activation. we also checked the absence of calgranulins in lymphocytes. after cell activation we showed that intracellular s100a8, s100a9 and s100a8a9 increased, but at different levels depending on the cell and the agonist used. extracellular s100a8 also raised after cell stimulation, but the concentration found were very low compare to those found in the plasma of septic shock patients. conclusions. together, these results suggest a different regulation depending on the form of the protein and of the cell and thus of proper distinct function of each monomer and of the complex. in the limits of our model the increased concentrations found in the plasma of patients with a septic shock can't be explained by immune cell activation. objectives. although there is no specific antidote for these potent toxins, drugs like penicillin g and silibinin have been used with conflicting evidence. we successfully managed two patients with mushroom poisoning by using silibinin and nac. methods. two members of a family, a mother 38 years old, and her son 15 years old were admitted to our icu 36 h after the ingestion of wild mushrooms. they presented with abdominal cramps, vomiting, profuse diarrhea ([20/day), myalgias, confusion and agitation. the clinical examination showed severe dehydration, tachycardia, oliguria with grade i-ii hepatic encephalopathy. laboratory exams revealed elevation of liver enzymes sgpt: 1106/880 u/l, sgot: 1884/930 u/l. coagulation parameters were as following: prothrombin time 15.5 00 /15.1 00 , factor v \55%/60%, factor vii \50/55%. high ammonia levels were noted, reaching 360 and 400 ng/dl, respectively. metabolic acidosis was also present with mild renal dysfunction. the ultrasound performed in both patients showed hepatosplenomegaly. aggressive fluid and electrolyte replacement started upon admission. silibinin was given at a dose of 20 mg/kg/day intravenously, in four divided doses, for three consecutive days, while nac was given as a continuous infusion at a dose of 150 mg/kg for the first hour, 50 mg/kg for the next 4 h, and thereafter 100 mg/kg/day for the following four days. hepatic encephalopathy, mild jaundice and renal dysfunction resolved within 48 h, and liver function tests returned to normal within 4 days. the patients recovered fully and were discharged to a medical ward. recent experimental and clinical studies have shown a strong protective and antioxidant effect against hepatic cell injury in amanita toxicity by the administration of nac and silibinin, either as monotherapy or as a combination therapy. although further clinical research is required to confirm their efficacy in reducing mortality and transplantation rate, nac has been used in our icu in hepatic dysfunction of different etiologies with promising results. we have recently shown that in patients with lactic acidosis due to metformin intoxication (serum drug level = 61 ± 25 lg/ml; therapeutic level is b4 lg/ml) systemic oxygen consumption (vo 2 ) can be abnormally low despite a preserved global oxygen delivery (do 2 ) (1). the study, however, suffered from being retrospective. objectives. to prospectively clarify whether metformin primarily impairs vo 2 . methods. eight sedated, paralyzed and mechanically ventilated pigs received a continuous i.v. infusion of metformin, at a rate of 1.6 g/h. the amount of metformin administered to each animal ranged from 4 and 8 g. the experiment always finished 9 h after the initiation of drug infusion. use of sedative and neuromuscular blocking drugs, as well as ventilatory setting, were always kept constant. serum metformin concentration was measured at the end of the experiment, using high performance liquid chromatography (hplc). arterial ph, lactatemia, vo 2 (indirect calorimetry) and do 2 (computed from cardiac output measured by pulmonary artery thermodilution) were recorded hourly. data are presented as mean ± sd. statistical testing was performed using the one-way repeated measure anova and the linear regression analysis. metformin infusion produced toxic serum drug levels (77 ± 30 lg/ml; n = 5). arterial ph drop from 7.63 ± 0.05 (prior to infusion) to 6.96 ± 0.26 (end of the experiment) (n = 8; p \ 0.001) and lactatemia rose from 1 ± 0 to 17 ± 7 mmol/l (n = 8, p \ 0.001). vo 2 progressively decreased (from 117 ± 37 to 66 ± 30 ml/min; n = 8, p \ 0.001) while do 2 did not significantly change over time (from 274 ± 74 to 239 ± 66 ml/min; n = 7, p = 0.39). the decrease in vo 2 was proportional to the dose of metformin administered (r 2 0.62; n = 8, p = 0.02) and to the serum drug level reached by the end of the experiment (r 2 0.69; n = 5, p = 0.08). conclusions. lactic acidosis develops during metformin intoxication in the presence of a diminished vo 2 but in the absence of any clear evidence of inadequate do 2 . this finding suggests that impaired oxygen utilization, rather than availability, may have a role in the pathogenesis of metformin-induced lactic acidosis. : 180 min) . death was consequent to multiorgan failure, anoxic encephalopathy or capillary leak syndrome if ecls was performed under cardiac massage. four patients presented with documented brain death, allowing organ donation in 2 cases. among these patients, the heart of one flecainide-poisoned patient was successfully transplanted, after normalization of ecg and myocardial function as well as toxicant elimination under ecls. prognostic factors in ecls-treated poisoned patients were as follows: qrs enlargement on admission (p = 0.009), saps ii score on admission (p = 0.005), ecls performance under massage (p = 0.008), arterial ph (p \ 0.001), lactate concentration (10.7 [6.6-19.6] versus 15.0 mmol/l [6.2-29.5], p = 0.003), as well as red cell (p = 0.008), fresh plasma (p = 0.003), and platelet (p = 0.03) transfusions within the first 24 h. conclusions. to our knowledge, this is the larger series of ecls-treated poisoned patients ever reported. ecls appears to be an efficient salvage technique in case of refractory toxic cardiac failure or arrest, with a 28% survival rate. our series clearly demonstrate that toxic refractory cardiac failure remains the best indication with a 46% survival rate. objectives. aim of the study was to investigate the incidence of infections in patients treated with hypothermia while receiving sdd. in this retrospective case control study 35 patients treated with prolonged hypothermia (cases) were identified and 169 patients with severe brain injury were included (controls). propensity score matching was performed to correct for differences in baseline characteristics and clinical parameters. primary outcome was the incidence of infection. the secondary endpoints were the micro-organisms isolated from surveillance cultures and during infection. the demographic and clinical data indicated that the cases and controls were well matched. the length of stay in the icu and duration of mechanical ventilation were comparable between the groups. the overall risk of infection during icu stay was 20% in the hypothermia groups versus 34.4% in the normothermia group (p = 0.388). pneumonia was diagnosed in 11.4% of patients in both groups (p = 1.000). the incidence of meningitis, wound infection, bacteremia, and urinary tract infection was low and comparable between the groups. staphylococcus aureus was most frequently identified as the causative infectious microorganism in both the hypothermia (14.3%) and normothermia (36.3%) group (p = 0.375), followed by coagulase negative staphylococci (14.3% in the hypothermia and 27.3% in the normothermia patients, p = 0.625) gram-negative bacteria were isolated from the surveillance cultures in 51.4% of patients treated with hypothermia and 31.4% of patients in the control group (p = 0.143). colonization of the rectum with gram-negative bacteria was significantly more frequent in patients treated with hypothermia compared with normothermia (48.6 vs. 20.0% respectively, p = 0.041). in contrast, colonization of the upper gastrointestinal tract and sputum was comparable between the groups with an incidence of 14.3% in the hypothermia patients versus 11.4% in the normothermia patients (p = 1.0). use of sdd mitigates the increased risk of infection in patients treated with hypothermia. based on the surveillance cultures, it seems that oropharyngeal decontamination is the most effective part of the sdd regimen in the prevention of pneumonia. introduction. prognostic scores specific for critical patients were developed in order to predict mortality based on physiologic and laboratorial variables. on the other hand, specific scores for burn patients are calculated taking into consideration inhalation injury, age and total burned surface area (tbsa), among others. however, scores utilized in general icu have not been evaluated in burn patients. objectives. therefore, the aim of the present work was to validate apache ii, saps 3 as well as initial sofa in a population of patients with massive burn. these scores were compared to some specific burn patient scores, including absi (abbreviated burn severity index) and estimates of the probability of death. retrospective study employing data collected prospectively from may 2005 to february 2010 (58 months) at an icu specialized in burn patients at a teaching hospital which is considered a reference centre in trauma care. all patients admitted during this period were included. one hundred and fifty-four consecutive patients were studied (male: 73%; female: 27%), with averaged age of 37.7 ± 15.9 years and a hospital stay of 33.8 ± 28.4 days. mortality rate of our sample was 43.5%. incidence of inhalation injury was 45% and total burn surface area (tbsa) was the following: 35.5% of patients had 20% or less; 36.1% had 20-40% of tbsa whereas 28.4% showed 40% or more. area under curve of receiver operating characteristic (roc) of evaluated indexes is displayed on table 1 . computerized head tomography is routinely performed as a diagnostic tool after the occurrence of neurologic deterioration in the icu adult patients. however, the ct findings in this setting are rarely reported. we hypothesized that the analysis of a series of cranial cts would help to understand the neurologic conditions of the critically ill patients and improve their management. objectives. to analyze, over a three-month period, the head ct scans performed in the adult icu in the albert einstein hospital in são paulo, brazil. methods. all cranial cts performed in the icu patients during the studied period were analyzed by two radiologists from the albert einstein hospital staff from may 1st to august 31st, 2009, according to a pre-established protocol: 1. presence of acute cerebral ischemia; 2. presence of previous cerebral ischemia; 3 . presence of acute cerebral hemorrhage; 4. presence of cerebral edema; 5. cerebral aneurisms; 6. cerebral tumors and 7. normal cerebral tomography. we studied 69 ct scans from 34 (49.2%) males and 35 (50.8%) females, mean age 64.43 ± 21.7 years. the head ct findings were the following: (1) presence of acute cerebral ischemia = 10 (14.4%); (2) presence of previous cerebral ischemia = 17 (24.6%); (3) presence of acute cerebral hemorrhage = 27 (39.1%); (4) presence of cerebral edema = 7 (10.1%); (5) cerebral aneurisms = 2 (2.8%); (6) cerebral tumors = 2 (2.8%) and (7) c16 years c 18, abc (assessment blood consumption) cp: c2 and ets (emergency transfusion score) cp: c3, c4 years c6. these scales handle the following combinations of variables for calculation: age, sex, type of admission, mechanism, blood pressure, focussed assessment for the sonography of trauma, hemoglobin, orthopedic or pelvic trauma, heart rate. mt was defined as the transfusion of 10 units or more of packed red blood cells in the first 24 h. we study the sensitivity (s), specificity (sp), positive and negative predictive value (ppv, npv), likelihood ratios positive and negative (lhr+ , lhr-) and area under the receiver operating characteristic curve (auroc) of different scales for the predictive power of tm validated in the literature. 568 patients were available for analysis (77.6% men, iss 30 ± 13, blunt trauma 93.8% objectives. we measured patient-reported outcome following surgical management with dc using a quality of life instrument. methods. survivors discharged between 3 and 18 months after severe tat were contacted after obtaining approval by our institutional irb. we excluded patients with neurotrauma. we applied self-response version euroqol questionnaire (eq-5d) and visual analog scale (eq-vas: 0 (worst health)-100 (best health). euroqol it is based on a descriptive system that defines health in terms of 5 dimensions: mobility, self-care, usualactivities, pain/discomfort and anxiety/depression. each dimension has 3 levels of response: no problems (level 1), some problems (level 2) severe problems (level 3). results. thirty four patients were contacted. mean ± sd age was 31.8 ± 11.6 yrs, male were 88.2% and penetrating trauma occurred in 79.4%. mean ± sd in severity scores were: ati 24.4 ± 9.6, iss 28.1 ± 8.5 and apache ii 20 ± 6. the median time from discharge was 12 months (iqr 6-15 months). the eq-5d dimensions in which the largest proportion of patients reported severe problems were usual-activities (work, study) and pain/discomfort 14.7% and 5.9% respectively as shown in the conclusions. survivors of severe trauma and dc, reported acceptable quality of life with minimal limitations with social functioning. a prospective study should assess quality of life in these patients from hospital discharge and systematically over time. introduction. brain tumors surgery is one of the main causes of admittance to the nicu. it is important to know the risk factors associated to hospital mortality of patients admitted to nicu due to this reason. to identify perioperative factors associated to higher hospital mortality in a series of patients admitted to nicu immediately after a bt elective resection. methods. data of 101 patients operated for bt elective resection and consecutively admitted to nicu at imss umae 1 bajío were prospectively obtained. nicu bt database includes 269 perioperative items. we divided the series in two groups: surviving and deceased patients. then, we analyzed the perioperative behavior differences between both groups. either student's t test or chi-square test was used, as it corresponded, for the analysis of differences observed between both groups. values of p lower than 0.05 were considered significant. results. the hospital mortality observed in this series of 101 patients was 8.9% (9/101). data of the nine variables showing significant differences between surviving and deceased patients groups are shown in table 1 . even if hypoxic brain injury has been reported as the strongest factor affecting the poor outcome of near-drowning patients, little has been known about prognostic factors affecting the outcomes of those patients receiving mechanical ventilation. to define prognostic factors affecting the outcomes of patients mechanically ventilated after near-drowning. , white blood cell counts (or, 1.00; 95% ci, 1.00-1.00; p = 0.001), serum creatinine (or, 0.19; 95% ci, 0.06-0.67; p = 0.010), and serum lactic acid (or, 0.95; 95% ci, 0.93-0.98, p = 0.002) were associated with favorable outcomes, respectively. however, only higher body temperature as a clinical parameter and the level of serum lactic acid as a laboratory parameter were significant predictors of favorable outcomes in multivariate analyses; the or were 2.87 (95% ci, 1.01-8.14; p = 0.048) and 0.96 (95% ci, 0.93-0.99; p = 0.011), respectively. conclusions. initial body temperature and the level of serum lactic acid were two most important clinical and laboratory prognostic factor in nearly drowned patients. the outcomes were not affected by the degree of initial hypoxemia. to determine the use of automated external defibrillators (aed) and manual defibrillators deployed in the various hospital wards (unmonitored areas) in a university hospital. a prospective study was performed according to utstein style of all cardiac arrests occurred in the hospital during the first 6 months after the implantation of a new protocol of care for hospital cardiac arrest. because of this plan automated external defibrillators were located for hospital wards and common service areas (radiology areas, outpatients, …) where one would expect a lower incidence of cardiac arrests, according to the risk map elaborated previously. in areas of greatest risk manual defibrillators previously existed. all resuscitation attempts in these areas were analyzed, excluding the emergency department because of a separate protocol against the rest of the hospital. special attention was given to the use of aeds by wards staff before the arrival of resuscitation team. also a comprehensive volunteer training program was designed, but it began after the analyzed period was finished. results. during the first 6 months we collected a total of 20 pcr in hospital wards and public areas, with a median age of 71 years and predominantly male (12 patients). the most common origin was respiratory (13 patients) followed by cardiac (4 patients). the most frequent rhythm detected was non-shockable (17 patients), only in 2 was shockable and unknown in 1. before the resuscitation team arrival only two patients had been manually defibrillated and were never used the new aeds. conclusions. the aeds provided in the hospital were completely useless in the first months after placement, probably due to the lack of a comprehensive training plan associated to the population goal. methods. descriptive longitudinal study. 208 patients were studied by encephalic death, as potential donors of organs, alerted to the network of regional transplant 5 (cdtot), by units of intensive care, for 36 months, in barranquilla's city. it was applied qualifying each of the variables in agreement to the vital opposing signs and biochemical tests brought in this moment. . 78.3% of the subjects were male; the average of age was 37.34 years (±sd: 16.24). the values of blood sugar, sodium, osmolaridad, tonicidad, po2, fc, pam, and glasgow, determined a score of 22, qualification that there had patient with encephalic death with the scale mbcm, as a test of certainty of the scale to diagnose encephalic death in total absence of reflections of stem. conclusions. there is recommended the application of mbcm's scale to every neurological patient by diagnosis of encephalic death in proof of certainty, in absence of others. by the high specificity of the already demonstrated scale there is recommended that scores lower than 19 they should restate the qualification. a score of 22 is an encephalic death in absence of reflections of stem. grant acknowledgment. clínica general del norte-cdtot introduction. prospective analysis of tracheostomies performed in patients admitted to a neurotrauma icu, the reasons for its implementation, and intraoperative complications in the first week. methods. all patients admitted to the icu of neurotrauma, which underwent a tracheostomy after admission. data were collected: affiliation, cause of admission, average stay, cause for realization of tracheostomy, tracheostomy time delay from its indication, place of performance of the procedure (icu or operating room), perioperative complications (event at transfer to operating room or during surgery: hypoxia, hypotension, arrhythmia, bleeding, premature extubation, false cannulation, cardiac arrest, pneumothorax or death), and postoperative complications in the first week (bleeding, difficulty in changing cannula, stomal infection, pneumothorax, death). introduction. the s-100b protein is a brain-specific protein release from astroglial cells into the circulation after traumatic brain injury (tbi). researches indicate that the s-100b serum level could be a useful indicator of tbi severity, however there is not evidence enough about the role of s-100b in nonsevere head trauma. the hypothesis that s-100b is a useful screening tool to detect brain injury in patients with a normal level of consciousness after a head trauma was tested. a total of 115 patients with the diagnosis of mild tbi without decrease of consciousness (according to the gcs) with at least one neurological symptom or finding like amnesia, headache, dizziness, convulsion and vomits, were prospectively included. we recorded the clinical data on admission and a blood sample before 6 h after tbi, for s-100b inmunoluminescence analysis. a routine cranial computed tomography scan (ct) was obtained within 24 h after the injury (categorized in normal or pathological). the diagnostic properties of s-100b serum levels.105 lg/l, for prediction of intracranial lesions revealed by ct were tested with receiver operating characteristic (roc) analysis. seventy of the patients (60.9%) were men, with a mean (sd) age of 49.07 (20.65) years (range, 14-92 years). a total of 11 patients (9.6%) had intracranial lesions. serum s-100b levels were significantly higher in patients with intracranial lesions than in the remaining patients. the average value of the protein in patients without intracranial lesion was 0.315 lg/l with a ci 95% (0.265-0.365 lg/l), and in those with pathological findings in ct was 0.601 lg/l with a ci 95% (0.403-0.798 lg/l). significant differences were found between levels of s100b protein and the presence of pathological findings in the ct (p = 0.001) (fig. 1) . the roc curve analysis showed that s100b protein is a useful tool to discriminate the presence of intracranial injury in ct (auc, 0.76, 95% ci, 0.685-0.907, p \ 0.001). s100b analyses with a cut-off level of 0.105 lg/l showed a sensitivity 100% but a specificity 19.2%. we evaluated different cut off values and in our series, the best cut off of the s100b protein is at 0.253 lg/l with a sensitivity of 100% and specificity 58%. (fig. 2) conclusion. determination of serum protein s-100b is a useful biochemical indicator of brain damage in head trauma. our results show that an increase in the cut-off point of s-100b to 0.253 lg/l increases its accuracy in the prediction of the existence of macroscopical lesions. key words. protein s-100b, brain injury, minor head trauma, cranial computed tomography. critically ill patients with systemic inflammatory response syndrome frequently suffer muscle weakness due to critical illness myopathy (cim) and polyneuropathy (cip). several in vitro studies have shown that the cause of muscle weakness is a loss of membrane excitability accompanied by membrane depolarization [1] . objectives. we investigated membrane polarization and excitability parameters in muscle and motor nerve in vivo within the first week after intensive care unit (icu) admission. methods. the study was approved by our local ethics committee. patients with sofa scores c8 on 3 consecutive days underwent nerve conduction studies including direct muscle stimulation to categorize patients as icu-control, cim-(dmcmap \3 mv) and/or cippatients (reduced snap amplitude) within the first 8 days after icu admission. to assess excitability parameters we recorded stimulus-response behaviour, threshold electrotonus, current-threshold relationship and recovery cycle from abductor pollicis brevis muscle following stimulation of the median nerve [3] . data are shown as median and 25%/75% percentile. conclusions. we describe for the first time that critically ill patients in general show muscle-and nerve membrane depolarization, whereas patients later suffering from muscle weakness due to cim or cim/cip feature additionally reduced membrane excitability. this suggests that membrane depolarization in critically ill patients is caused by energy failure leading to dysfunction of the na-k pump, the motor of membrane repolarisation-whereas reduced membrane excitability in cim or cim/cip needs an additional dysfunction of voltage gated sodium channels for example occurring in the presence of endotoxins [2] . in intensive care patients with central nervous system (cns) disease, the systemic inflammatory response syndrome (sirs) criteria are often unreliable as a basis for identifying the inflammatory process. even with the presence of some infection they could be signs of the diencephalons-catabolic syndrome. diencephalons-catabolic syndrome like sirs constitutes of hyperthermia over 38°c, tachypnea of over 20 per minute, tachycardia, and arterial hypertension. thus, sirs symptoms may occur after antibacterial treatment even if there is no infection or inflammation. we suggest a more precise method which could help to avoid the excessive antibacterial therapy and to control it in patients with cns disease-a procalcitonin test. objectives. reduce the use of wide specter antibiotics makes the control over antibacterial therapy in patients with cns diseases more precise; reduce the number of complications related to unnecessarily long antibacterial treatment. after obtaining the informed concern, in our investigation we included 30 patients with different neurological disorders, who had recently transferred neurosurgical operations. all of them demonstrated sirs symptoms on different postoperative terms. when sirs symptoms occurred, we checked the level of procalcitonin in the patient's serum by a semi quantitative method on a disposable brahms pct-q system. the procalcitonin level was determined against a color scale. procalcitonin level over 0.5 ng/ml (16 patients) considered a sign of infection and in such cases we prescribed antibacterial treatment 2,000 mg of selenase for 5-7 days. if the test result was negative (14 patients) we repeated it in 24 h and in cases with the same results, no antibacterial treatment was administered even if there were sirs symptoms. if pct-q test was negative patients were sedated (fentanyl 0.8-1.5 lg/kg/h and clonidine 0.2-0.5 lg/kg/h) to achieve autonomic stability and attenuate clinical manifestation of sirs. we had not observed any cases of sepsis in both groups of patients. by mince of pct, we had managed to reduce the quantity of wide specter antibiotics, used in neurosurgical patients for 34.7%. conclusions. procalcitonin test in neurosurgical clinic let us determine the necessity of antibacterial treatment reduce the use of wide specter antibiotics, medical costs and prevent the forming of polyresistant infection. l. combe 1 , r. appleton 1 , c. gilhooly 1 , j. kinsella 1 1 university of glasgow, department of anaesthesia and critical care, glasgow, uk intensive care unit-acquired weakness (icuaw) is increasingly recognised as a common complication of critical illness with potentially prolonged debilitating sequelae. the estimated incidence is 46% in patients with sepsis, multi-organ failure or prolonged mechanical ventilation [1] and suggested risk factors include: the systemic inflammatory response syndrome (sirs), sepsis, higher severity of illness, hyperglycaemia, renal replacement therapy and parenteral nutrition. objectives. the aims of this study were to determine the incidence, risk factors and outcomes for patients diagnosed with icuaw in glasgow royal infirmary's (gri) icu. the study was undertaken in two parts, firstly as a case-control study [matched for age (within 5 years), sex and admission apache ii score (within 5 points)] and secondly by comparing identified cases of icuaw to a 3-month cross-sectional sample (1/10/09-31/12/09, 112 patients) of gri's icu patients. data for both parts of the study was obtained from two electronic databases, wardwatcher and carevue. carevue was searched to identify patients with icuaw and wardwatcher was used to identify the controls. data collected included: patient and illness characteristics, severity of illness scoring, organ support and treatments provided, laboratory results and outcomes. minitab software was used for statistical analysis. conclusions. the incidence of icuaw was very low, we hypothesise this to be explained by the absence of systematic evaluation of patients for icuaw. the risk factors and outcomes for icuaw were consistent with some of the published literature. prospective study is now planned to systematically evaluate this condition. with increasing age, comorbidity, and socioeconomic deprivation being associated with higher risk pregnancies, there comes a potential higher risk of complications. neurological and neurosurgical complications, which can be particularly devastating during the peripartum period, include those due to medical conditions of pregnancy (hypertensive disease, sepsis, thromboembolic disease, hypoxic-ischaemic brain injury), iatrogenic complications secondary to anaesthetic or obstetric interventions, incidental illness or injury (pharmacological alterations, trauma, tumour), and deliberate self-harm and violence. objectives. to ascertain the frequency of neurocritical care admissions in the west of scotland, the nature of the admission diagnoses, the impact they have on our service (length of stay), and maternal and foetal outcome. methods. using the scottish intensive care society audit group wardwatcher patient database, female patients aged 14-50 years old who were admitted to the neurocritical care unit were identified (january 2008-december 2009). we manually reviewed the electronic admission note for each of these women in order to gain diagnoses; a targeted case note analysis ensued. within the 24 month study period there were a total of 915 admissions to neurocritical care, of whom 164 fulfilled the age and gender criteria; 6 admissions (0.66% of total) were for neurological complications in the peripartum period. the age range was 18 to 45 years (median 31 years). three women (50%) were intrapartum (20-36 weeks gestation) at the time of their admission, and three were postpartum (1 day-7 months). half of admissions were due to incidental illness or injury, a third to pregnancyrelated medical complications, and one case was iatrogenic in nature. length of stay in icu was 1 to 12 days (median 4.3 days). one patient sustained a residual facial nerve weakness and deafness. conclusions. this survey provided insight into the incidence and nature of pregnancyrelated pathology requiring acute referral to a regional neurosciences centre. as highlighted in other surveys, there may be many more peripartum patients with neurological complications who are cared for in general critical care units, and do not require admission to a tertiary referral centre [1] . further work is underway to ascertain the true numbers of neurological complications of pregnancy countrywide. our approach represents a paradigm for the continuing audit of pregnancy-related critical care resource use in scotland. introduction. hypertonic saline has an osmotic effect on the brain because of its high tonicity and ability to effectively remain outside the blood-brain barrier. there may be a minimal benefit in restoring cerebral blood flow, which is thought to be mitigated through local effects of hypertonic saline on cerebral microvasculature. most comparisons with mannitol suggest almost equal efficacy in reducing icp but not compared their effects on eeg. objectives. we aimed to compare the effects of 20% mannitol, 3% or 7% hypertonic saline on hemodynamic parameters, intracranial pressure and electroencephalography in experimental head trauma. bilateral craniotomy were carried out in the parietal region and head trauma was applied for all rabbits. the rabbits were randomly divided into four groups. in group i rabbits were only observed. in group ii: 20% mannitol, in group iii: 3% hypertonic saline and in group iv: 7% hypertonic saline was administered intravenously to achieve similar osmolar load. electroencephalography, mean arterial pressure, heart rate, intracranial pressure were recorded before trauma and 5 and 60 min after trauma. results. increased intracranial pressure was significantly decreased by mannitol, 3 and 7% hypertonic saline solutions at the end of study (p \ 0.05). but intracranial pressure values of mannitol and 7% hypertonic saline groups were lower than the other groups (p \ 0.05). the electroencephalography scores decreased after trauma in all groups (p \ 0.05). at end of the study, 3 and 7% hypertonic saline groups had similar electroencephalography scores with pretrauma scores (p [ 0.05). the mean arterial pressure and heart rates increased after trauma in all groups (p \ 0.05). mean arterial pressure values were found lower only in mannitol group at end of the study (p \ 0.05). our study showed that when used in intracranial hypertension treatment, 7% hypertonic saline solution is as effective as mannitol, and preserves hemodynamic parameters, and normalizes traumatic electroencephalography abnormalities better than mannitol. objectives. to identify the causes of new onset seizures in patient admitted in medical icu. methods. all the patient admitted in icu and who had new onset seizures were evaluated. the patients were evaluated for metabolic profile. imaging (ct/mri) was done whenever needed. patients with preexisting seizure history were excluded from study. . 44 (29 males, 15 females) patients, who had first seizure during hospitalization in icu were included. 43 patients had generalised and one had focal seizures. 28 patients had metabolic abnormalities. 13 (46.42%) had evidence of hepatic encephalopathy. 3 (11.36%) had only hepatic encephalopathy while rest had associated uremia, hyponatraemia, hypophosphatemia and hypomagnesemia. out of 10 patients, who had renal failure, 4 had evidence of uremia while rest had associated hyponatraemia or hypophosphatemia. only one patient had evidence of hypocalcemia. imaging was done in 16 patients. 14 (30%) had abnormal ct scan results. 4 (9.09%) had intracranial hemorrhage, 4 (9.09%) had infarct, 2 (4.55%) had brain metastasis, 2 had evidence of hydrocephalus and one each had evidence of extradural hemorrhage and tuberculoma. csf analysis was done in 7 (15.9%) patients. 3 (6.8%) had evidence of tuberculosis and 1 (2.27%) had evidence of pyogenic infection. to study the role of various investigations and ct in evaluating these patients. all patients admitted with new onset seizures within 72 h prior to presentation were included. all the patients were questioned and an attempt was made to assign an electroclinical syndrome to seizure. patients were evaluated for metabolic profile, neuroimaging. csf examination was done in those who had persistently altered mental status, infectious symptoms and fever. results. 110 patients were admitted (1.03% of total patients who came to emergency) with history of new onset seizures. 71.8% patients were diagnosed to have acute symptomatic seizures and were placed in ilae category 4.13 and three patients were placed in ilae category of remote symptomatic seizures. the cause of seizures was established in 82 (74.5%) patients and remained unestablished in 28 (25.5%) patients. 14 (12.7%) patients were diagnosed to have neurocysticercosis. other important causes were acute infarct, uremia, hyponatremia, hypernatremia, viral encephalitis, post partum eclampsia, pyogenic and tubercular meningitis. alcohol withdrawal seizures were seen in 3.6% patients. metabolic derangements were seen in 19 (17.3%) patients. computed tomography was done in 88 patients and 50% had abnormal findings. mri was done in 13 patients and 12 had abnormalities. conclusions. neurocysticercosis was found to be most common cause of seizure activity in our part of country. though metabolic derangement can cause significant proportion of new onset seizure patients routine imaging of brain should be performed in patients with new onset seizures. work environment and organisational issues: 1335-1347 1335 subjective and objective research into the working conditions and their effect on the health and safety of people working in icu, focusing mainly on the natural factors of temperature, humidity, ventilation, lighting and noise (part 1) n. karachalios 1 , e.c. katsilaki 1 , d. sfyras 1 1 general hospital of lamia, icu, lamia, greece the aim of the project is the subjective and objective investigation of the conditions of work and the relation repercussions on the health and safety of people working in the icu, focusing mainly on the natural factors that are likely to cause the sick building syndrome. for this purpose a protocol of research in two phases has been planned. the first included objective measurements, with the use of suitable equipment, of the natural factors of temperature, humidity, ventilation, lighting and noise. the second phase included the subjective estimation of the working people about their own health and conditions of their work, in the particular area of the hospital with the use of substantiated anonymous questionnaire. after the subjective and objective study and analysis of 32 questionnaires and 55 measurements of natural factors, we found that the medium temperature of the icu was 24°c. the mean relative humidity of the icu was 38% (highest 38.3% and lowest 37.8%). the mean ventilation rate of the icu was 17 m 3 /h (highest 28.8 and lowest \7.2 m 3 /h). the mean sound pressure was 64.8 db (highest 75 and lowest 63.5 db). the average lighting was 331.5 lux (160 lux lowest and 598 lux highest). the objective data seem to keep pace with the subjective opinions of the working people, as they were impressed in the questionnaires of subjective estimate. the objective data were compared with the subjective. the results of the research were also compared with data from the existing bibliography and current legislation, leading to a line of conclusions. (1) insufficient and bad quality ventilation. (2) the existing temperature of the environment contributes to the appearance of sick building syndrome. (3) the working environment is noisy. (4) the environment of work has problematic or insufficient lighting. (5) the icu under study is a building area which can be characterized as ''sick'' if immediate action is not taken. background. up to 75% of critical care nurses test positive for (symptoms of) post traumatic stress disorder (ptsd) [1, 2, 3] . it is assumed that these symptoms are caused by professional involvement in life-threatening events [3] . in a sample of intensive care nurses, we investigated which work related incidents were perceived as most distressing. method. in interviews, 12 nurses (75% female) were asked to memorize and tell about their most traumatic work related event. all interviews were recorded. after verbatim transcription, the 'most critical events' were extracted and categorized bij two independent psychologists. . none of the nurses reported major life-threatening events such as trauma-related injuries, massive bleeding or seeing patients die as their 'most critical incident'. conclusion. not the major life-threatening events but relatively 'normal work related events' under unusual circumstances are mentioned as most critical by nurses. in contrast to major life-threatening events, these 'normal events' are usually underestimated by colleagues, and thus potentially compromise peer-support. a care bundle refers to evidence based interventions and information grouped together to improve outcomes and consistency of provided care [1, 2] . at the icus charge nurses and intensivists as shift leaders are responsible for daily management of unit activities. several immediately made decisions by shift leaders are made under time pressure and high information load with inadequate information. though we have evidence of structure and process based factors such as material and human resources, admission and discharge decisions or bed utilization, the support for information transfer and integration is poor in organizational decision-making concerning these factors. objectives. to identify immediate information needs of charge nurses and intensivists during the management of daily activities at the icu and evaluate how necessary this information is for their decision-making. from september 2009 to november 2009, all charge nurses (n = 515) and intensivists (n = 223) of 17 university affiliated icus providing comprehensive care in finland were surveyed with an on-line questionnaire using 122 statements. the questionnaire was developed based on our previous observation study and statements of our survey regarded information needs related to the icu care activities. a rating scale from 0 to 10 (completely unnecessary-absolutely necessary) was used to assess the necessity of the information. for each statement, a response with mean 7 or over was regarded as necessary information for immediate decisions. results. the response rate was 47.97% (charge nurses 50.1%, intensivists 43.1%). the working experience varied from 0 to 35 years (mean 13.6, sd 7.9). over 50% of respondents worked as a shift leader once a week or more often. 72 statements of 122 were valued as a necessary (mean [7 or more) for immediate decision-making. absolutely necessary information (mean [9 or more) for immediate decision-making were assessed related to the 11 statements. these statements concerned isolations, mechanical ventilation, admissions and discharges, special treatments, patient's condition, and scheduled dates or times for surgery or other procedures. conclusions. both icu charge nurses and intensivists identified several information needs that are crucial for immediate decision-making during the whole icu care process. information needs of the shift leaders differed and they were strongly connected to the needs of one's professional requirements. an integrated overview and summarization of immediately needed information-a care bundle for organizational decision-making-at the icus is highly needed for icu shift leaders. the common interests of both professionals, charge nurses and intensivists, should be emphasized when new technology-based systems are developed. background. the nursing shortage is an international problem that is expected to worsen in the coming years. studies show that one of the main reasons nurses leave the profession is their dissatisfaction with their work environment. structural empowerment and nurse-physician collaboration are two elements of the nurses' work environment that are potentially related to one another according to kanter's theory (1977) . in addition, a nurse's clinical specialization has been found to influence perceptions related to these two concepts. to examine the level of perceived structural empowerment, the perceptions of nurse-physician collaboration and the relationship between these two variables, among intensive care unit (icu) nurses and general ward nurses in israel, and to compare the groups. a descriptive, correlational, comparative study design was used on a sample of 84 icu nurses and 88 nurses from internal medicine and general surgery wards in a large university hospital in israel (response rate 79%). a three section, self administered questionnaire was used to measure the study variables: the condition of work effectiveness scale-ii (cweq-ii), the collaboration with medical staff scale (cmss) and demographic-professional background. results: perceived structural empowerment was found to be moderate (m = 19.41, sd = 3.66, range = 6-30). nurses tended to agree that there was nurse-physician collaboration (m = 2.69, sd = 0.55, range = 1-4, 1 = strongly disagree, 4 = strongly agree). a correlation was found between structural empowerment and the nurse-physician collaboration (r = .52, p \ 0.01). a significant difference was found between icu nurses and general ward nurses on their perceptions of nurse-physician collaboration (t (170) = -3.39, p \ 0.01; general wards: m = 2.83, icu: m = 2.55). no significant differences were found between nurse specialization on perceived level of structural empowerment. conclusion. nurses in this study tended to agree that there was nurse-physician collaboration on their unit/ward. nurses who perceived themselves as having a higher level of structural empowerment, felt that there was a higher level of nurse-physician collaboration. general ward nurses had more positive perceptions about nurse-physician collaboration on their ward as compared to icu nurses. no difference was found between the two groups on the level of structural empowerment. recommendation. the findings of this study can be used as the basis for the design of interventions, aimed at enhancing structural empowerment and nurse-physician collaboration, in order to improve nurses' work environment, as one of strategy to decrease the nursing shortage. further study of additional hospitals in the country is also recommended. teams have expanded and in some hospitals 24 h cover has been instituted. researchers are questioning the validity of outreach services and its impact on patient outcomes. as cco has been viewed as the panacea to all problems, data collection and analysis is fundamental in proving its financial and clinical benefits. objectives. this comparative study aims to evaluate retrospective data from 1 month in 2006 and 1 month in 2010. data does not encapsulate patient outcomes; it will compare frequency of referrals and interventions. this data provides an indication to the extent cco has participated in the care of the acutely ill over a given time period. methods. data was collected from the 4d medicus database collating intervention data. analysis occurred using 13 key interventions using excel software conclusions. whilst the validity of services has been questioned, the data itself indicates that more patients are referred and frequency of interventions has increased. various system changes occurred during this time period such as a change of mews trigger scores, the advent of 24 h cco and courses such as alert and survive sepsis were introduced into the basic training of staff. it must be noted that the intention in the uk for cco was a service that empowered staff through education to undertake this care themselves; therefore the increase in interventions could indicate that the educational approach hasn't made progress. although the study compares 13 interventions, an increase in the type of interventions was also noted such as ward based cco supervised cpap and establishing a picc line service. therefore this highlights the changing application of interventions. further analysis is required to look at the appropriate skills required for the delivery of safe care to the acutely ill in the ward environment. whilst ward staff are increasingly under resourced, both in skills and manpower, cco do provide the skills, knowledge and time to meet the shortfall in safe timely care. introduction. working as a critical care nurse involves situations where teamwork is essential and rapid, effective communication is of importance [1] . the education to become a specialist icu nurse gives skills and knowledge to manage patients who are critically ill with rapidly changing conditions [2] . experimental research is one way of contributing to the acquisition of such knowledge. to describe how icu nurses may contribute and perform in the experimental research process, an environment usually unfamiliar to them. we describe our experiences with regard to clinical contribution and our subjective evaluation of involvement in animal experimental research. method. three icu nurses in a swedish hospital were asked to participate in a research project investigating myocardial metabolism in porcine models of shock. the tasks were anaesthesia and pain management, assisting with catheter insertion and haemodynamic monitoring the pigs during the process results. although the situation was new, the nursing role and function in the team were at once similar and different to the daily work situation in the icu. one major skill learnt was the rigour of experimental measurements and sources of error, which is sometimes neglected in clinical care. being able to observe changes due to shock in a controlled setting, we improved our ability to critically 'think ahead' in anticipation of clinical deterioration [3] . our first-hand experiences at the animal experimental laboratory allayed many anxieties and misconceptions with this type of research. conclusions. the critical care environment demands skills such as the ability to accurately define and change priorities rapidly, good communication and teamwork [4] . we believe that the experimental research setting is one way of enhancing this ability. in these units patients condition may change rapidly and they may need close inspection as well as emergency response. early warning scoring (ews) system may make early recognition of and response to bad condition possible by observation based on systematic parameters. ews was developed as a simple scoring system to be used at ward level utilizing routine observations taken by nursing staff. ews is based on five physiological parameters; systolic blood pressure, pulse rate, respiratory rate, temperature and avpu score (alert; reacts to voice; reacts to pain; unresponsive). objectives. the aim of this study was to evaluate ews among patients admitted to pacu. methods. ews parameters were recorded four times from 167 patients after their admission to pacu. the first record was taken during the first admission to pacu (ews 1), the second (ews 2) after 15 min, the third after 30 (ews 3) and the fourth record after 60 min. the correlation between variables like differences of four ews, patients age, the asa score, duration of operation were statistically examined. early treatment and recognition of sepsis is a stated aim of the surviving sepsis campaign [1] but in busy clinical environments the delivery of antibiotics and fluids can often be delayed. we describe the implementation of an audit proforma, based on the survivesepsis.org [2] resuscitation bundle, as a tool to deliver six aspects of management within 1 h of recognition sepsis. 1. improve the early recognition and treatment of sepsis in acute medical patients. 2. provide a sustainable change in the management of septic patients 3. improve mortality and length of hospital stay methods. the proforma consist of six treatment management steps, based on the survivesepsis.org ''septic six'': 1 oxygen, 2 blood cultures, 3 antibiotics, 4 lactate, 5 iv fluids, 6 strict fluid management. it is triggered by patients satisfying two or more of the systemic inflammatory response syndrome criteria. all management steps should be implemented within 1 h of the trigger time stated on the form. the forms are collected and analysed every month and the results are displayed for staff working on the medical admissions unit and accident and emergency. a total of 239 forms have been collected, 30% diagnosed with severe sepsis. the progress on all six parameters is shown below. over the initial seven month period we have demonstrated a sustained improvement in the rapid delivery of all six of the management parameters. introduction. the early goal-directed resuscitation has been shown to improve survival in patients presenting with septic shock. a recent systematic review demonstrated the inability of central venous pressure (cvp) to predict the hemodynamic response to fluids infusion, and it should not be used to make clinical decisions regarding fluid management in critical patients. the clinical implication of this fact in septic shock is not well-known. objectives. the aim of this study is to determine if the resuscitation with fluids guided by cvp has clinical implications in patients with septic shock. post-hoc analysis of a patients' cohort with septic shock admitted in the medical intensive care unit since june 2007 to june 2009. all of them were treated on basis of a bundle for severe sepsis management. chi-square analysis was used to compare categorical data. continuous data were compared using student's t test. we used multiple logistic regression model to assess the association between the independent variable and mortality, after adjustment for possible confusing factors (we considered variable to be confounding if the estimate of the coefficient changed by more than 10%). eighty-five patients were studied. 66% were male. their average age was 57 ± 17 and 40% had previous chronic diseases. severity scores: apache ii 24 ± 8, sofa 10 ± 4 and 76% of patients had multiorganic dysfunction. infectious focus was respiratory in 48%. cvp mean was 12 ± 4 mmhg, scvo2 73 ± 8% and the mean amount of fluids provided was 2840 ± 844 cc. 71% of patients needed mechanical ventilation. hospital-stay middle was 28 days (1-160) and 10 days in icu conclusions. in our patients' cohort with septic shock treated under the basis of the early goal-directed resuscitation, the volume of fluids infused was associated independently with mortality. a lower fluid administration in the resuscitation probably could be caused by the early reach of a high central venous pressure. blinding of study interventions is necessary to prevent bias in randomized controlled trials (rct). since normal saline and 5% albumin are packaged in bags and bottles, respectively and they have different color and texture, a blinding procedure is necessary to ensure the fluids appear identical for comparative rcts. objectives. to describe the blinding procedure and evaluate sterility and stability involved in the transfer and storage of study fluids in the precise pilot rct. a standard operating procedure for concealment, meeting pharmacy guidelines and good manufacturing practices was developed by the manufacturing pharmacist at the coordinating centre and used by all participating sites. fluids were transferred with aseptic technique into identical 500 ml bottles under a sterile hood by the pharmacy or transfusion medicine technician then covered with an opaque wrapping. average time to transfer of study fluids from their original packaging was recorded to understand labor involved with creating each study fluid package. yellow intravenous tubing was manufactured to also conceal the fluid color. six blinded bottles of normal saline and 5% albumin from the participating centers were stored at room temperature for at least 3 months. cultures of the fluids using blood culture media and/or endotoxin levels (measured by commercial assay) were obtained to document sterility of the study fluids. protein electrophoresis was used to assess albumin stability. results. transfer of the study fluids was the responsibility of the research pharmacist/ technician and blood bank at 3 and 3 sites, respectively. average time to transfer 20 containers of normal saline and 5% albumin into bottles was 65 ± 29 and 77 ± 19 min, respectively. sterility (culture negative and/or endotoxin undetectable) of study fluids was confirmed from all bottles of normal saline and albumin that underwent testing. protein electrophoresis of albumin samples showed a single band suggesting no degradation of albumin during transfer and storage. conclusions. the standardized blinding procedure developed for transfer of study fluids in this pilot rct confirmed sterility and stability of our study fluids for 3 months. these data are important when considering the length of allowable storage time for these study fluids. due to the resources and time involved with the transfer of these fluids for individual sites, this transfer method needs to be incorporated into budgeting and may not be feasible in the context of a large rct. grant acknowledgment. the precise pilot rct was funded by a grant from canadian blood services. covidien, singapore, singapore, 18 yong loo lin school of medicine, national university of singapore, biostatistics unit, singapore, singapore introduction. the surviving sepsis campaign recommends a 6-h resuscitation bundle and a 24-h management bundle to improve outcomes in severe sepsis. compliance with and relevance of these recommendations to asian intensive care units (icus) are unknown. objectives. the primary objective of the present study was to assess the compliance of asian icus and hospitals to these bundles. the secondary objectives were to evaluate the impact of compliance on mortality, and the organisational characteristics of asian hospitals which are associated with higher compliance. methods. this was a prospective observational study of patients with severe sepsis who were admitted to the participating icus in july 2009. we recorded the organisational characteristics of participating centres, the patients' baseline characteristics, and the achievement of targets within the resuscitation and management bundles. results. sixteen countries and 150 icus participated, enrolling 1285 patients. hospital mortality was 44.9%. achievement rates for the bundle targets were: lactate measurement, 39.8%; blood cultures, 62.5%; broad-spectrum antibiotics, 64.0%; fluids ± vasopressors, 81.4%; central venous pressure, 39.7%; central or mixed venous oxygen saturation, 10.8%; low-dose steroids, 55.8%; drotrecogin alfa, 3.2%; glucose control, 27.1%; lung-protective ventilation, 11.7%. compliance rates for the entire resuscitation and management bundles were 7.6 and 3.5% respectively. on logistic regression analysis, achievement of the targets for blood cultures, antibiotics, and central venous pressure independently predicted decreased mortality. high-income countries, university hospitals, icus with an accredited fellowship programme, and surgical icus were more likely to be compliant to the resuscitation bundle. conclusions. compliance to the resuscitation and management bundles is generally poor across asia. given the resource limitations in asia, the most appropriate strategy to improve outcomes in severe sepsis may be to concentrate on ensuring early administration of antibiotics after blood cultures, and appropriate fluid therapy. cerebral oxygen desaturation predicts cognitive decline and longer hospital stay after cardiac surgery monitoring brain oxygen saturation during coronary bypass surgery: a randomized, prospective study the work is supported by departmental sources. 1032 clinical features and prognosis of organizing pneumonia pre-senting as acute respiratory failure in icu reference(s). 1. webster nr. ventilation in the prone position prone position in acute respiratory distress syndrome effect of prone positioning on the survival of patients with acute respiratory failure acute effects of upright position on gas exchange in patients with acute respiratory distress syndrome this study was funded by arjo international ag, florenzstrasse 1d metabolic acidosis and fatal myocardial failure after propofol infusion in children: five case reports longterm propofol infusion and cardiac failure in adult head-injured patients mild hypothermia alters propofol pharmacokinetics and increases the duration of action of atracurium intermittent haemodialysis versus crrt for arf in the intensive care unit dialysis dose in acute kidney injury: no time for therapeutic nihilism cirrhotics admitted to icu, and when added to the liver-specific scores of meld or ukeld, improves their respective predictive value intensive care, london, uk, 3 royal free hospital epidural anesthesia, hypotension and changes in intravascular volume intraoperative fluid restriction improves outcome after major elective gastrointestinal surgery surrogate designation: can we trust our relatives? does chest physical therapy work? physiotherapy in intensive care: towards an evidence-based practice fisioterapia no paciente sob ventilação mecânica this research was supported by grants from the following brazilian funding agencies/programs: cnpq, capes, fapesc and unesc readmission to surgical intensive care increases severity-adjusted patient mortality physiological scoring systems and audit predicting death and readmission after intensive care discharge a case-control study of patients readmitted to the intensive care unit severity of illness and risk of readmission to intensive care: a meta-analysis a comparison of admission and worst 24-h acute physiology and chronic health evaluation ii scores in predicting hospital mortality: a retrospective cohort study learning from the past to inform the future-a survey of consultant nurses in emergency care assessing emergency nursing competence post-traumatic stress among swedish ambulance personel levels of mental health problems among uk emergency ambulance workers partial and full ptsd in brazilian ambulance workers: prevalence and impact on health and on quality of life ambulance personnel and critical incidents impact of accident and emergency work on mental health and emotional well being 3 artemis health institute, director, critical care, pulmonology and sleep medicine, gurgaon, india, 4 artemis health institute, nursing, gurgaon, india reference(s) the australian incident monitoring study in intensive care: aims-icu. the development and evaluation of an incident reporting system in intensive care adverse events in critical ill patients ministry of health and social policy 1254 communication: a key factor in the patient safety? anemia of the critically ill: acute anemia of chronic disease impact of allogenic packed red blood cell transfusion on nosocomial infection rates in the critically ill patient high dose recombinant human erythropoietin stimulates reticulocyte production in patients with multiple organ dysfunction syndrome: the journal of trauma: injury, infection and critical ca to the staff of the critical care department, faculty of medicine injury severity and quality of life: whose perspective is important? quality of life and persisting symptoms in intensive care unit survivors: implications for care after discharge variations in health-related quality of life in critical patients funded in part by fogarty international center nih grant no. 1 d43 tw007560-01 and clinical research institute-fundacion valle del lili glasgow coma score, use of mechanical ventilation and vasoactive agents, and the occurrence of severe sepsis (according to bone's criteria-1992). the causes of admission were divided as: ischemic stroke, hemorrhagic stroke, subarachnoid hemorrhage, status epilepticus, traumatic brain injury, elective neurosurgeries, and miscellanea. the foci of infection, microbiological data and bacteremia were analyzed from septic patients. numeric data were expressed as median and interquartiles, while categorical data were calculated as percentage. univariate and multivariate (logistic regression) analysis was carried out to point factors associated with hospital mortality. results. we included 661 patients, with median age 65 years (iq range 50-78) and 54% were male 30%) patients, while it occurred during icu stay on 110 (70%) patients. hospital mortality was associated with age, the admission cause (higher for hemorrhagic stroke, traumatic brain injury and status epilepticus), apache ii score, glasgow coma score and severe sepsis on the univariate analysis cnpq 1317 perioperative factors associated to higher mortality in patients admitted to the neurological intensive care unit (nicu) immediately after brain tumor (bt) resection saldívar 2 1 umae 1 (high-specialty medical unit no 1) el bajío, imss and nicu, hraeb (high-specialty regional hospital of el bajío) anaesthesiology and intensiv care medizin anaesthesiology and intensive care unit charité universitätsmedizin-berlin, department for anesthesiology and intensive care medicine after approval of the local ethics committee, the pdr icg was measured within 24 h post injury (day 0) using the non-invasive limon system (pulsion medical systems of pdr icg to supranormal values higher sofa scores were indirectly associated with lower pdr icg values, particularly for sofa scores[8. when patients were grouped by icu length of stay (\11, c11 days, corresponding to the mean icu los of the german trauma registry), logistic regression analysis identified pdr icg consumables were provided by pulsion medical systems influence of apoe polymorphism on cognitive and behavioural outcome in moderate and severe traumatic brain injury genetic variation of the apoe promoter and outcome after head injury effects of apolipoprotein e genotype on outcome after ischaemic stroke, intracerebral haemorrhage and subarachnoid haemorrhage the association between apoe « 4, age and outcome after head injury: a prospective cohort study decreased cerebrospinal fluid apolipoprotein e after subarachnoid hemorrhage correlation with injury severity and clinical outcome « association of ventilation rates and co2 concentrations with health and other responses in commercial and industrial buildings « sensitivity to noise, personality hardiness, and noise-induced stress in critical care nurses recommended lighting level for offices » the chartered institution of « sick building syndrome, sensation of dryness and thermal comfort in relation to room temperature in an office building: need for individual control of temperature silent misery: most severe critical incidents post traumatic stress disorder in the emergency room: exploration of a cognitive model trauma exposure and post-traumatic stress disorder in intensive care unit personnel increased prevalence of post-traumatic stress disorder symptoms in critical care nurses drivers of quality in health services: different worldviews of clinicians and policy managers revealed systems thinking, system dynamics the fifth discipline: the art and practice of the learning organisation the development of system dynamics as a methodology for system description and qualitative analysis finnish funding agency for technology and innovation nursing activities score tradução para o português e validação de um instrumento de medida de carga de trabalho de enfermagem em unidads de terapia intensiva: nursing activities score (nas) nursing activities score in the intensive care unit: analysis of the related factors 1341 the self-perceived health between medical-surgical and crit-ical care nurses in hungary deutsch 1 , i. boncz 1 , a. sebestyen 3 , a. olah 1 1 university of pecs faculty of health sciences a longitudinal study design was used to explore the self perceived health of inhospital nurses in acute care settings (surgery, casualty, internal medicine, intensive, coronary care, emergency room) in two hungarian factors predicting team climate, and its relationship with quality of care in general practice nurse working conditions, organizational climate, and intent to leave in icus: an instrumental variable approach critical care nurses' work environments: a baseline status report quality of practice in an intensive care unit (icu): a mini-ethnographic case study vasps/intv 2010). medicinska fakulteten, lunds universitet critical thinking and clinical decision making in critical care nursing assessing and developing critical-thinking skills in the intensive care unit gulhane military medical academy, haydarpasa training hospital, istanbul, turkey, 3 gulhane military medical academy technology as a catalyst to transforming nursing care devices and desire: gender, technology and american nursing surviving sepsis campaign: international guidelines for management of severe sepsis and septic shock division of pulmonary and critical care medicine, seoul, republic of korea, 3 peking union medical college hospital, department of critical care medicine mai hospital, intensive care department, hanoi, viet nam, 10 king saud bin abdulaziz university for health sciences, king abdulaziz medical city, intensive care department 12 dr soetomo general hospital, department of intensive care republic of china, 14 ripas hospital, intensive care unit surviving sepsis campaign: international guidelines for management of severe sepsis and septic shock the surviving sepsis campaign: results of an international guideline-based performance improvement program targeting severe sepsis great differences in compli-ance with surviving sepsis campaign bundles surviving sepsis campaign: international guidelines for management of severe sepsis and septic shock delayed diagnosis is associated with increased morbidity, mortality and cost in the icu. as the mortality rate of severe sepsis remains unacceptably high, a group of international expert developed guidelines in 2004, termed the surviving sepsis campaign (ssc). the ssc group has introduced the ''sepsis care bundles surviving sepsis campaign guidelines for severe sepsis and septic shock implementation of a bundle of quality indicators for the early management of severe sepsis and septic shock is associated with decreased mortality improving outcomes for severe sepsis and septic shock: tools for early identification of at-risk patients and treatment protocol implementation observational, prospective follow-up. patients who were admitted into the intensive care unit in university hospital complex a coruña (chuac) during the months of hospital mortality was 30 surviving sepsis campaign: international guidelines for management of severe sepsis and septic shock early goal-directed therapy in the treatment of severe sepsis and septic shock associated with decreased mortality translating research to clinical practice: a 1-year experience with implementing early goal-directed therapy for septic shock in the emergency department improvement in process of care and outcome after a multicenter severe sepsis educational program in spain duration of hypotension before initiation of effective antimicrobial therapy is the critical determinant of survival in human septic shock delta co2 (pvco2-paco2) as a prognostic factor in septic shock septic shock using the new device inspectra 650: relation to macro-and microhemodynamic and outcome c. luengo 1,2 , f. vallée 1 , c. damoisel 1 , m. resche-rigon among the techniques assessing microperfusion, near infrared spectroscopy (nirs) gained interest. more than baseline sto 2 values, the reperfusion slope after a vascular occlusion test (vot) nirs parameters, especially the reperfusion slope scvo 2 or svo 2 ); metabolic (ph, base excess and lactate) parameters were collected. microperfusion data consisted in: nirs (baseline sto 2 , occlusion and reperfusion slopes (%/s), automated software); skin laser doppler microflow (baseline flow (tpu), peak flow (tpu) and slope during reperfusion (tpu/s), measured during and after a 3 min vot. survivors (s) and non-survivors ] differed between s and ns at day 1. macro-hemodynamic and metabolic data did not differ between s and ns plan quadriennal ea3509 svo 2 does not predict fluid responsiveness in critically ill septic patients supported by msm 0021620819 research grant: replacement of and support to some vital organs years) were studied. apache ii and sofa score at study entry were 21 (range: 5-32) and 8 (range: 1-16) respectively. the septic syndrome was due to sepsis (n = 12), severe sepsis (n = 3) or septic shock (n = 39). sites of infection included the lung reference(s). 1. ungerstedt u: microdialysis: principles and applications for studies in animals and man the pathophysiology and treatment of sepsis management of sepsis surviving sepsis campaign guidelines for management of severe sepsis and septic shock relation between muscle na + k + atpase activity and raised lactate concentrations in septic shock: a prospective study long-term continuous glucose monitoring with microdialysis in ambulatory insulin-dependent diabetic patients whether it is worth to correct acidemia by infusion of alkaline solutions is a matter of discussion. there are a number of evidences against the use of alkalinization therapy with respect to the benefits of reversing ph and the side effects of sodium bicarbonate infusion [1]. nonetheless, as recently shown by means of an on line survey, 67% of critical care physicians administer base to patients with lactic acidosis mmol/l), animals were randomized to 60 min of: a) sustained lactic acid infusion, a + b) sustained infusion + sodium bicarbonate, o) transient infusion, b) transient infusion + sodium bicarbonate. in the transient infusion (group o and b), at randomization lactic acid was replaced with normal saline. acid-base status and lactate levels were measured over time. in a number of animals phosphofructokinase (pfk) enzyme's activity was also measured. results. following lactic acid infusion blood lactate rose unnecessary use of alkali perturb acid-base status and lactate metabolism potentially overcoming metabolic adaptive strategies. reference(s). 1. boyd jh, walley kr. is there a role for sodium bicarbonatein treting lactic acidosis from shock? use of base in the treatment of acute severe organic acidosis by nephrologists and critical care physicians: results of an online survey strong ions gap (sig) quantifies unmeasured blood anions and it is calculated by the difference between strong cations and strong anions (all of them, dissociated in blood plasma) retrospective, observational study of all patients with septic shock as defined by the american-european consensus, admitted to the icu from arterial blood gases, albumin, lactate and electrolytes were obtained at admittance and 24 h later; apache and sofa score, central venous saturation and lactate comparison of acid base models for prediction of hospital mortality following trauma forty-five sepsis patients [median age, 62 (iqr, 47-37) years; admission saps ii, 45 (33-53) pts; severest multiple organ dysfunction syndrome score interaction of vasopressin infusion, corticosteroid treatment, and mortality of septic shock comparing two different arginine vasopressin doses in advanced vasodilatory shock: a randomized, controlled, open-label trial lambert 1 1 university of leicester, division of anaesthesia, leicester, uk blood samples were taken: at induction of anaesthesia, at 3 and 18-24 h post-cpb. neutrophils were isolated, mrna extracted, dna cleaned and reverse transcribed supported by a grant from the association of anaesthetists of great britain and ireland, and the british journal of anaesthesia/royal college of anaesthetists secretoneurin (sn), a neuropeptide, is specifically expressed in endocrine elevated nucleosome levels in systemic inflammation and sepsis extracellular histones are major mediators of death in sepsis 23rd esicm asymmetric and symmetric dimethylarginines (adma, sdma) are protein-breakdown markers; both compete with arginine for cellular transport and are excreted in urine. moreover adma, sdma, their ratio (marker of adma catabolism), arginine, interleukin-6(il-6), tumor-necrosis-factor-a (tnf-a), c-reactive-protein(crp) on day 1, 3, 6, 9, 12 and at discharge in 72 consecutive severely-septic patients were measured sdma were higher than normal, adma/sdma ratio was halved, arginine was low. adma was related to total sofa and arginine, inversely related to il-6 and crp; sdma was related to saps ii, sofa, blood urea, creatinine, arginine. adma/sdma ratio was inversely in 14 non-survivors, creatinine, il-6, tnf-a, crp and adma were stable, sdma increased, adma/sdma ratio remained low figure: time course of adma and sdma blood levels (mean ± standard error) during icu stay and the last icu day protein-hmgb-1 levels as predictors of outcome in patients with sepsis and septic shock hmgb1 as a predictor of organ dysfunction and outcome in patients with severe sepsis early low dcs counts may be correlated to disease severity and could predict fatal outcome. however, little is known about dc number in other shock than septic. objectives. to evaluate and compare the circulating dcs number in patients with severe sepsis, septic or cardiogenic shock. methods. in a prospective multicentric study (3 icu), consecutive immunocompetent patients with severe sepsis (ss), septic shock, cardiogenic shock were included. peripheral blood dc counts, measured by flow cytometry, were evaluated and compared between the three populations at admission and 24 h later. correlation to disease severity evaluated by clinical scores and day 28 mortality was studied. results. 100 patients were included (age 61 ± 14 years, 56 male, sofa d0 10.5 ± 3.9, saps ii 61 ± 18): 49 septic shock, 16 severe sepsis and 22 cardiogenic shock. mortality at d28 was respectively 43, 14 and 59%. 13 patients presented a sepsis associated to cardiogenic shock. at baseline and at day 1, a dramatic diminution in the numbers of total dcs either myeloid (mdcs) or plasmacytoid (pdcs), was observed in sepsis (severe sepsis or septic shock) compared to cardiogenic shock patients. no difference was seen between severe sepsis and septic shock patients (fig. 1). we did not observe any correlation between the number of total dcs at admission or at day 1 and severity of illness scores dc reduced number is a valuable marker of severe sepsis in shock and is not affected by hemodynamic changes. it could not be used as a prognostic marker in severe septic patients. 1382 preliminary results from a prospective study assessing the relationship between standard laboratory coagulation and global tests of clot-formation using thromboelastography in patients with fulminant hepatic failure v the routine use of international normalized ratio (inr) to establish the coagulation status in patients with fulminant hepatic failure (fhf) may be misleading. anecdotally, fhf patients, despite a significantly deranged inr, may display a normal or even hypercoagulable state, as recently shown, albeit in an extracorporeal setting, with frequently clotted circuits, despite raised pt we prospectively studied coagulation, demographic, survival and outcome measures of fhf patients (defined by de-novo liver failure, coagulopathy-inr [1.5, and encephalopathy) admitted to the royal free hospital liver and/or intensive care unit(s) (icu), a tertiary referral centre in liver diseases and transplantation we present the standard clotting tests and teg results from 10 (of a required 20) patients currently enrolled, demonstrating variable degrees of encephalopathy and coagulopathy effect of norepinephrine on cardiac output and preload in septic shock patients apparent heterogeneity in splanchnic vascular response to norepinephrine during sepsis aggressive use of high-dose norepinephrine in the treatment of septic shock norepinephrine requirement is not an independent variable to predict outcome in severe septic shock patients aim. the aim of this study was to measure the level of ptsd among hungarian ambulance workers, and explore factors which can influence it.sample and methods. 411 hungarian ambulance workers were involved to this crosssectional study (147 ambulance drivers, 173 ambulance nurses, and 91 ambulance team leaders: medical doctors and ambulance officers). self filling questionnaire were used for data collection, including briere's trauma symptom checklist, and socio-demographic questions. chi square test, independent t test and variance analysis were used for comparison of variables.results. the average ptsd-points of ambulance workers was 20. there was significant association between level of ptsd and gender: women's average 23, men's average 18 ptsd-points (p = 0.049). there were no correlations between level of ptsd and type of settlement, location of ambulance station and level of education. those who would need psychological support (p = 0.01), and those who had psychologically traumatic experiences in the last 2 years have significantly higher ptsd-points (p = 0.01).conclusions. hungarian ambulance workers are exposed with many effects which can lead ptsd. professional psychological support is needed in order to cope with ptsd successfully.the results were presented and discussed in our weekly meeting on patient safety and healthcare for all icu personnel. by the end of this year all the recommendations will be implemented in our icu.conclusions. we improved the safety and quality of in hospital transportation of icu patients by performing a prospective risk analysis. bow-tie is a good instrument to identify health care risks. to determine the incidence of phrenic neuropathy associated with the catheterization of internal jugular and subclavian veins, without ultrasound support, in patients admitted to an icu. a prospective study was performed by following patients admitted in the icu between october 2008 and may 2009. a normal neurography of both right and left phrenic nerves at the moment of their admission was the main inclusion criteria. after this baseline study, a new neurography was repeated weekly (chen and resman method, sinergy medelec), during their stay and at the moment of being discharge from icu. simultaneously, all vascular subclavian and internal jugular vein catheterization were registered. a final neurography and a fluoroscopy study were performed after being discharged from hospital. results. 40 patients were included and two hundred and ten neurographies of both right and left phrenic nerves were performed. 14 patients did not receive any vascular punctures in the cervical region during the follow up period, acting as control group. 26 patients underwent a total of 48 vascular catheterization, 43 in subclavian vein (89.6%) and 5 in internal jugular vein (10.4%). a phrenic neuropathy was diagnosed in 4 patients. this represented an incidence of 15% (4/26) of phrenic neuropathy per patient and 8% (4/48) related to subclavian and internal jugular vein catheterization. in relation to patients without phrenic nerve injury who underwent subclavian and internal jugular vein catheterization, patients affected of phrenic neuropathy had longer mechanical ventilation time (33 ± 36 days vs. 19 ± 13, p = 0.3) and longer average stay time in icu (49.7 ± 35 days vs. 25 ± 14, p = 0.08), although these differences have not statistical significance. we did not find significantly differences related to age (53 ± 9 vs. 55 ± 18, p = 0.2) and apache ii index (16 ± 3.5 vs. 15.8 ± 5.8, p = 0.4) between both groups (wilcoxon two-sample test). we performed a control neurography of 2 case patients after being discharged from hospital. we checked the cmap phrenic nerve reappearance after 3 weeks and 6 months of being diagnosed its neuropathy, respectively. conclusions. we found an incidence of phrenic neuropathy of 15% per patient and 8% related to subclavian and internal jugular vein catheterization, during the follow-up period. the time of reappearance of phrenic cmaps after being detected its neuropathy points to a neuroapraxia or partial axonotmesis as pathogenic type of injury.discussion. phrenic neuropathy has to be considered in cases of difficult weaning of unclear etiology. the catheterization of subclavian and internal jugular veins should be recommended employing ultrasound support. p. merino 1 , m.c. martin-delgado 2 , j. alvarez 3 , i. gutiérrez-cía 4 , á . alonso-ovies 3 , syrec 1 hospital can misses, icu, ibiza, spain, 2 isde, á rea de salud, madrid, spain, 3 hospital de fuenlabrada, icu, madrid, spain, 4 hospital clínico universitario, icu, zaragoza, spain introduction. syrec project aims to improve icu patient safety. the project includes an epidemiological study. we present the main results.objectives. to estimate the near miss (nm) and adverse events (ae) rate in spanish intensive care units (icus). we study the incidence and nature. finally, we classify and analyze its severity.methods. multicenter prospective observational cohort study. inclusion criteria: patients admitted to the 79 participant icus during the 24-h observation period. during this period, nm and ae detected and reported inside and outside icu were included. only outside icus were considered when its were the reason for admission. we evaluate the kind of incident, severity and preventability. data collection studied under the distribution of frequencies.results. 1,017 patients were included. 1,424 incidents were reported in 591 patients, 943 were nm and 481 ae. risk: the median risk of nm was 73% versus ae 40%. 22.1 incidents per patient admitted. incidence rate: the incident rate median was 5.89 per 100 patients per hour icu stay, the nm of 3.47 per 100 patients per hour icu stay and that of ae, 2.04 per 100 patients time of stay in icu. the 66% of the incidents reported have been nm and 34% ae. this incidents causing temporary damage in the 29.50% of occasions and in the 4.28% permanent damage, compromised the patient's life or contributed to death. classification of incidents (table 1) . conclusions. our study shows a high individual risk. our icus services present a highrisk environment. therefore we have to go into the developement of epidemiological studies depth, in order to create further strategies supporting patient safety. restore cardiovascular performance in severe lactic acidotic rats a. kimmoun 1 , n. sennoun 1 , n. ducrocq 1 , b. levy 1,2 1 inserm u961, groupe choc, vandoeuvre-lès-nancy, france, 2 chu nancy brabois, intensive care unit, vandoeuvre-lès-nancy, france introduction. lactic acidosis during shock is responsible for myocardial failure, vascular hyporesponsiveness and a decrease in sensitivity to vasopressor agents. sodium bicarbonate is a proposed treatment to correct acidosis, although with deleterious cardiovascular effects. indeed, hypocalcemia and hypercapnia, both powerful myocardial depressants, are the main side effects of the administration of this therapy [1] . objectives. already studied in experimental models of isolated lactic acidosis, the cardiovascular effects of sodium bicarbonate administration have never been explored after correction for hypocalcemia and hypercapnia. methods. we therefore compared, in a rat model of severe lactic acidosis (ph \ 7.2, hyperlactatemia[3 mmol/l) induced by a state of controlled hemorrhagic shock, the cardiovascular effects of: (1) standard resuscitation plus administration of sodium bicarbonate with correction for calcemia and paco 2 (''adapt'' group, n = 5); (2) standard resuscitation plus administration of sodium bicarbonate without correction for paco 2 and calcium (''nonadpat'' group, n = 5);(3) standard resuscitation; (''stand'' group, n = 5); (4) standard resuscitation plus calcium administration (''calc'' group, n = 5). evaluation at steady and shock state, 60 min and 120 min was focused in vivo on arterial gas and myocardial contractility (emax) by conductance catheter. ex vivo vasoreactivity was tested on mesenteric arteries (300 lm) by myography. sodium intakes were equivalent between groups. results. our model displayed a profound acidosis from 7.31 to 7.18 ± 0.01 (p = 0.02) and hyperlactatemia from 2.8 ± 0.4 to 9.2 ± 0.6 mmol/l (p \ 0.01). emax decreased from 2.5 ± 0.33 to 0.54 ± 0.06 mmhg/ll p = 0.01. in the adapt group, at 60 min, ph was normalized at 7.36 ± 0.01 (p = 0.04). furthermore, emax was enhanced at 332 ± 43% (p \ 0.001) (stand: 145 ± 26%, nonadapt: 94 ± 18%, calc: 234 ± 52%). the cumulative dose of infused norepinephrine was significantly lower in the adapt group 145 ± 26 lg/kg compared to other groups (stand: 344 ± 10 lg/kg, nonadapt: 438 ± 10 lg/kg, calc: 423 ± 103 lg/kg, p = 0.02). ex vivo mesenteric vasoreactivity in the adapt group was normalized (graph 1).mesenteric vasoreactivity to phenylephrine conclusions. in severe lactic acidosis, infusion of sodium bicarbonate after correction of its side effects improves myocardial function and vasoreactivity. [1] . the prevalence and significance of 25-hydroxyvitamin d deficiency in the intensive care unit have not been fully determined. a recent study of an unselected group of itu patients [2] has suggested low itu admission 25-hydroxyvitamin d levels are common. objectives/hypotheses to be tested. royal free hospital intensive care unit patients exhibit low circulating levels of 25-hydroxyvitamin d. circulating levels of 25-hydroxyvitamin d decrease further during the course of hospital admission. admission circulating levels of 25-hydroxyvitamin d affect itu morbidity and mortality methods. all itu admissions were assessed within 72 h of presentation and patients who were deemed to have the potential to require admission for at least 1 week were included. demographic and clinical data were obtained in a prospective manner. results were recorded from samples obtained at admission, 7 days and 28 days. standard itu nutrition protocols were used. no interventions were performed. results. clinical and outcome data were obtained for 40 patients. no significant differences between apache 2, saps 2 or apache 3 scores for survivor and non-survivor groups at either itu or hospital discharge were noted. 60 further patients await complete data analysis. 0% (0 of 39 for whom results were available) achieved an adequate ([75 nmol/l) circulating 25hydroxyvitamin d level. 3 patients (7.7%) demonstrated levels within the insufficient range (50-75 nmol/l). 2 patients (5.1%) did not have any detectable 25-hydroxyvitamin d. the remaining 34 patients (87.2%) were either in the deficient (41.0%, 25-50 nmol/l) or severely deficient (46.2%, 10-25 nmol/l) ranges. admission 25-hydroxyvitamin d levels in survivors and non-survivors were compared at itu and hospital discharge. no significant differences between the four groups (p [ 0.05, anova) were observed, indicating that in this data set, admission 25-hydroxyvitamin d levels do not appear to alter or determine clinical course. mean 25-hydroxyvitamin d levels were compared at admission, day 7 and at day 28. no significant differences between the three groups (p [ 0.05, anova) were identified. no significant differences between the mean 25-hydroxyvitamin d levels of the survivors and non-survivors at day 7 or day 28 were apparent (small numbers). admission [1, 2] and patients undergoing surgical procedures [3] . patients with neurological illness can receive significant quantities of ns, chosen primarily for its iso-osmolar properties. objectives. ns is commonly used as maintenance and resuscitation fluid by the anaesthetist, and as intravascular flushes by the radiologist during prolonged interventional neuroradiological (inr) procedures. this pilot feasibility study aimed to ascertain the effect of ns infusion on acid-base measurements in patients undergoing inr procedures under propofol-remifentanil anaesthesia. methods. we collated routine electrolyte, albumin and acid-base data of 5 patients who underwent coil/glue embolisations of intracranial aneurysms and vascular malformations, both before and after the procedure. base excess (be) was partitioned into the effects of sodium chloride difference (na-cl), albumin, lactate and unmeasured anions (uma), using the stewart-fencl-story approach [4] . all values are reported as medians (ranges objectives. to investigate the erythropoietic response to hight dose of a weekly schedule of recombinant human erythropoietin (rhuepo) in critically ill anaemic septic patients. a total of 60 patients admitted to the intensive care unite (icu) were enrolled in this study, patients were randomized to receive either rhuepo or not, 30 patient did to form the rhuepo group, 30 did not to form the control group.results. the epo treated group of patients showed significant increase in reticulocyte count compared with baseline p \ 0.001, as well as with the control group p \ 0.006. the epo treated group exhibited also a significant increases in hb concentration compared with baseline p \ 0.001 as well as the control group .0 3. all patients in the control group received rbc blood transfusion 100%, while only 83.33% of the epo group did. the epo treated group showed significant decreases in their apache ii score during the study period compared with baseline p \ 0.001 as well as with the control group p \ 0.05. the epo treated group showed no significant difference in their sofa score compared with baseline p \ 0.923, however the control group exhibited continuous and significant increase in their sofa score throughout the study period compared with their baseline p \ 0.003, there was no significant difference in the final outcome recovery, mortality or morbidity p .337, p \ 0.286 respectively.conclusions. the administration of rhuepo to critically ill anaemic septic patients is effective in raising their reticulocytic counts, hb concentrations and in reducing the total number of units of rbcs they require. in addition there was a trend toward better in hospital clinical course, increased recovery and decreased mortality in the rhuepo group.conclusions. anaemia is common following critical illness but does not appear to affect the physical aspects of recovery during medium term rehabilitation. this may be due to an overwhelming degree of symptom burden from other complications of critical illness impairing physical function to such a degree that the effects of anaemia are negligible in the medium term. although decreases in number and function has mainly been described in skeletal muscle, also other organs seem to be affected and it has been hypothesized that mitochondrial dysfunction might be involved in the development of organ failure. to study the effect of plasma of patients with septic shock on mitochondrial function in vitro to potentially later on identify a central factor affecting mitochondria in all tissues during sepsis and leading to multiple organ failure.methods. after sacrificing 6-8 week old sprague-dawley rats, mitochondria from soleus muscle were isolated through homogenization and a series of centrifugations. mitochondrial function was assessed by measuring of oxygen consumption, using an oxygraph containing a clarke-electrode, after addition of adp. before these measurements, mitochondria were incubated with plasma from septic patients or healthy volunteers, respectively, for 30 min. in our second series, the mitochondria were incubated with different concentrations of il-6, tnf-a or buffer. respiration rates were measured in the presence of adp (state 3; a measure for the oxidative capacity to produce atp) and without the presence of adp (state 4; a measure for the amount of uncoupling). respiratory control ratio (rcr; a measure for the respiratory efficiency of the mitochondria) was calculated by dividing state 3 by state 4 activity. all measurements were related to citrate synthase activity to compensate for the amount of mitochondria. statistical differences between the groups were analyzed using a student's t test.results. adp dependent (state 3) respiration was 98% higher and rcr 165% higher in the mitochondria incubated with plasma from the septic patients compared to those incubated with plasma from healthy volunteers (table) . there were no significant differences between the groups incubated with preservation buffer or the different cytokines (table) . introduction. microvascular fluid loss from the intravascular to the interstitial space generates tissue edema and is one of the major challenges in emergency and intensive care medicine. isolation of interstitial fluid (if) from skin makes it possible to study the microcirculation and proteins in this environment both during normal as well as pathophysiological conditions such as acute inflammation.objectives. by studying bio-markers from proteomic analysis by mass spectrometry in an inflammation model, we wanted to find proteomes that could be important in explaining inflammation. we have applied a recently described centrifugation method in a porcine model and compared it with implanted wicks. in nine anesthesized piglets we compared the methods and evaluated the if, by overhydrating the pigs with 3000 ml of acetated ringer's solution for 1 h, and thereafter continuously supplemented for 1 h according to fluid losses. if was isolated from implanted dry wicks, wet wicks and by centrifugation of excised skin. the methods were evaluated by the ability to reflect overhydration and to show the expected composition of plasma proteins in if by use of hplc. the if was also processed further with mass spectrometry to find possible tissue degradation or inflammation due to overhydration. statistics: by spss v 13.0 and graphpad instat (version 3.05). significance level: p = 0.05. colloid osmotic pressure in if was significantly lowered after overhydration for all the tree methods. wet wicks p = 0.05, dry wicks p = 0.001, skin samples p = 0.05. hplc of if collected with centrifugation after overhydration, identified peaks representing molecules smaller than albumin. mass spectrometry of the same if identified several proteins associated with inflammation: alpha-1-antichymotrypsin and lumican, the latter a protein identified as a modulator of inflammation. we have introduced a new centrifugation method for isolation of if from the skin of pigs. by further analysis of if isolated by centrifugation we were able to distinguish proteins found only in the if of the pigs overhydrated with ringer's acetate. these proteins could be associated with an inflammatory condition in the skin caused by massive overhydration, again causing tissue degradation. identification and validation of proteomic biomarkers can be a useful tool in future treatment of inflammation in general, and in sepsis in special. objectives. to define the pattern of change in metabolites by mrs in experimental sepsis. male sd rats (weight 325-375 g) underwent cecal ligation and puncture or sham procedure (n = 8 per group), and 24 h after surgery were euthanized. pulmonary tissue was extracted for magic angle mrs (hr-mas) and processing by the r metabonomic package. a supervised statistical analysis of main components (mc) was performed on the processed spectra.results. the mc analysis discriminated both group (septic and nonseptic) indicating a different metabolite profile. in addition, the analysis of mc loading revealed 4 displacement positions in the discrimination between groups with a variation in the signal intensity of 50%.conclusions. metabolomic analysis of pulmonary tissue by mrs is a potentially useful technique for the detection of biomarkers in sepsis.grant acknowledgment. introduction. cd14 + cd16 + neutrophils are a key subset of phagocytes associated with severe bacterial sepsis [1] . their characteristics, and potential neuro-immunomodulation, have not been explored in humans neutrophils exposed to septic plasma from icu patients. to assess the effect of adrenergic/cholinergic neurotransmitter molecules on human neutrophil adhesion and activation markers following exposure to human septic plasma. with irb approval, neutrophils were isolated from 5 healthy volunteers (ficoll density gradient separation) and incubated for 24 h with either plasma from healthy volunteers or septic patients plus pathophysiological concentrations of epinephrine (e), norepinephrine (ne) or acetylcholine (ach) and nicotine (nic) to assess potential parasympathetic-related neuro-immunomodulation. flow cytometry (dako cyan) measured expression on neutrophils of cd11, cd14, cd16 antibody markers and viability. median values are shown; analyzed by anova.results. neutrophils were unaffected by ne, e, ach or nic after incubation with plasma from healthy volunteers. after incubation with septic plasma, marked neutrophil activation occurred (p = 0.02). however, nic reduced cd14 + cd16 + activation (* fig. a ) by 95% (median (35-97%; 25th-75th centiles); p = 0.01). nic also attenuated cd11 expression, suggesting reduced neutrophil adhesion (* fig. b) . neutrophil viability was similar across drug and plasma treatments. conclusions. these preliminary data suggest that nicotine attenuates both the activation and adhesion of human neutrophils exposed to human septic plasma, but does not affect viability. objectives. the aim of this study was to evaluate the potential impact of lag between sepsis initiation and start of treatment on mitochondrial respiration. methods. 24 animals [40.1 ± 4.1 kg] were randomized (n = 6/group) to a control group (group i) and three groups resuscitated at 6 (group ii), 12 (group iii), and 24 (group iv) hours, respectively, after fecal peritonitis induction. fecal peritonitis was induced with instillation of 2.0 g/kg of autologous feces via intra-peritoneal drain. resuscitation was performed according to the ssc and esicm sepsis guidelines for 48 h. respiration of permeabilized skeletal muscle fibers and their isolated mitochondria was assessed at baseline and after 6, 12, 24, 48 and 72 h, when applicable, or before death occurred, if earlier. at the end of the experiment, also isolated brain, hepatic and myocardial mitochondrial respiration was measured using high resolution respirometry (oxygraph-2 k, oroboros instruments, innsbruck, austria). results. mortality (33%, each) and organ dysfunction was highest in groups iii and iv. in these two groups, different pattern of changes of skeletal muscle mitochondrial complex i-dependent respiratory control ratio (rcr) were observed (table 1) . no significant differences between groups were observed for complex i-and ii-dependent rcr values of hepatic, myocardial and brain mitochondrial respiration (fig. 1 ). there were no significant differences between the groups for any of the complexes in permeabilized skeletal muscle fibers mitochondrial respiration (data not shown). conclusions. despite the high mortality observed in groups resuscitated at later time points after induction of sepsis, end organ mitochondrial function assessed using physiological substrates was preserved. despite significant changes in skeletal muscle mitochondrial respiration efficiency in the two groups with the highest mortality, our findings do not support the view that mitochondrial dysfunction plays a major role in the pathogenesis of multiorgan dysfunction in experimental sepsis. grant acknowledgment. swiss national fund, nr: 3200-061988; stiftung für die forschung in anästhesiologie und intensivmedizin. adipose tissue is an endocrine organ which produces signalling proteins involved in inflammation and glucose homeostasis [1] . one of these proteins, adiponectin, promotes glucose utilisation and fatty acid oxidation and thus improves insulin sensitivity via its two receptors, adipor1 and adipor2 [2] . adiponectin expression has been shown to be reduced in type ii diabetes, obesity and endotoxaemia [2, 3] . adiponectin also exhibits antiinflammatory properties [4] . in this study, we have examined whether adiponectin and its receptor gene expression changes in murine adipocytes stimulated by lps. methods. 3t3--l1 adipocytes were grown in culture media (dmem with 10% fetal calf serum) until confluent. pre adipocytes were differentiated with the addition of 10 mg/ml insulin, 1 mm dexamethasone and 100 mm ibmx. media was changed every 48 h. cells were treated on day 12 with 100 ng/ml, 1 or 10 mcg/ml lps (escherichia coli, sigma-aldrich). cells were harvested at 4 and 24 h. mrna levels were determined by rt pcr in a 12.5 ll reaction volume consisting of 12.5 ng of reverse transcribed cdna mixed with optimal concentrations of primers and probe and qpcr tm core kit (eurogentec, uk) in 96-well plates on a mx3005p detector. results. cell response to lps was confirmed using il6 as a reference gene. expression of adiponectin mrna was significantly reduced in cells treated with 10 lg/ml lps harvested at 4 h (7.1 fold p = 0.002). there were no changes in cells treated with lower concentrations of lps. there were no changes at 24 h. r1 gene expression was significantly reduced following treatment with 100 ng/ml lps at 4 h (1.5 fold p = 0.04), but treatment with higher concentrations did not change expression. there were no changes at 24 h. r2 expression levels were significantly reduced at 4 h in the 1 and the 10 mcg/ml groups (2.6 fold p = 0.02 and 4 fold p = 0.01) respectively. there were no changes at 24 h. discussion. our results add to the evidence that changes occur in the adiponectin system during inflammation. in this model, we observed rapid reduction (at 4 h) in adiponectin at high dose lps, r1 at low dose lps and r2 at medium and high doses. there were no changes in expression levels at 24 h. this suggests that a rapid change in the adiponectin system may occur in response to lps but this change is not maintained at 24 h. in a previous study, our group has shown reduced adiponectin gene expression in adipose tissue depots in lps induced endotoxaemia [3] . it is interesting that different concentrations of lps induce different changes within the adiponectin system. further studies are needed to elucidate whether reductions in both adiponectin and its receptor may contribute to the inflammatory changes and hyperglycaemia commonly observed during sepsis including all co poisoned patients treated with hyperbaric oxygen. following parameters were seized: age, sex, date of admission, sofa, the source of the intoxication, the gravity co score, the initial clinical examination (realized by first aid), biology, the rate of hbco, the murray score and the rate of complication. results. 405 patients were included in the study. the sex ratio was 61%, the mean age was 42 ± 18 years and the global mortality was 5, 9%. among the 405 patients 29% were poisoned by smoke (s group), 60% by pure co (c group) and 11% by exhaust fumes. more than 30% of the exhaust fumes victims were suicide origin. this characteristic is associated with neurological impairment induce by ingested drugs. then, their neurological status is impossible to link to the co poisoning. we have therefore decided to exclude this group. the sofa score was higher in the s group compared with the c group (0.38-2.17; p \ 0.0001). a co score equal to 4 was present in 33 versus 1% respectively in s versus c group (p \ 0.001). in the under group of patients having a co score at 3, 0% (0/101) of co poisoned patients versus 18.4% (7/38) of smoke poisoned patients were ventilated (p \ 0.001). these patients were intubated either during transport or in the intensive care and none of them received hydroxycobalamine during the first aid (before intubation). the laboratory data showed in the s group a higher lactates level (2.2 vs. 3.9 mmol/l; p = 0.005) and lower initial pao 2 /f i o 2 ratio (365 vs. 306; p = 0.002). nine percent of the s group present a murray score at 4 versus 0% for c group (p \ 0.0001). pneumonia, shock and death were significantly more frequent in the s group (respectively 8.6 vs. 0.8%, p \ 0.0005; 11.2 vs. 0.8%, p \ 0.0001; and 17.9 vs. 1.2%, p \ 0.0001)conclusions. as expected the smoke poisoned group has a higher mortality than pure co group (mortality 17% vs. overall mortality 5.9%). at equivalent co gravity score, mortality and complications are always more frequent in the smoke poisoned group. the smoke poisoned group has a high risk of degradation. those patients require specific monitoring and support and probably early administration of hydroxycobalamine. hypothesis. at administration and maintaining higher plasma levels of at can reduce the need for inotropes in burn shock patients. we performed a retrospective cohort study of burn shock patients admitted to a single tertiary care center over 7 years period. patients were eligible for inclusion if they were received fluid resuscitation with ringer's solution and colloid according to clinical guidelines. data were abstracted including demographic, burn injury characteristics, resuscitation fluid volume, the type of colloid and the average of plasma at levels within 72 h after burn injury. administration of fresh frozen plasma and/or recombinant human at was defined as at administration. the decisions of at administration and inotropic support (dopamine or dobutamine) were made by the attending intensivists. primary outcome measure was the need for inotropes within 72 h after burn injury. cox regression model was used to estimate the risk reduction by at administration and average of at levels. [1] . argon, another member of the noble gas family has been reported previously to have a neuroprotective property [2] . the aim of this study was to investigate whether it attenuates neuronal injury in a rat model of neonatal asphyxia. methods. seven-day-old postnatal sd rats underwent right common carotid artery ligation and then recover with their dim for 1 h. thereafter, they were exposed to 8% o 2 balanced with nitrogen for 90 min. after 2 h, they were treated with 70% argon or 70% nitrogen (positive control group) for 90 min. the cohort pups without intervention served as naïve control. they were perfused 7 days later and their brains were sectioned and stained with 0.5% cresyl violet. microphotographs were taken from ca area of the hippocampus near -3.6 bregma relative to adult brain at 40 9 magnification. healthy cells were counted in a blind manner and their mean value was used for data analysis. results. the thickness of healthy layers in the right ca area of the positive control group was remarkably reduced compared with other groups (fig. 1 ). quantitative analysis revealed that argon treatment significantly increased healthy cell numbers in the right ca area of hippocampus from 36.53 ± 1.201 in the positive controls to 54.37 ± 1.162 (p \ 0.01) (fig. 2 ). grant acknowledgment. this study was supported by a grant from action medical research, uk. objectives. our objective was to study the mechanisms of death following high-dose citalopram administration in rats. experimental study in sprague dawley rats with intraperitoneal (ip) citalopram administration; determination of the median lethal dose (mld)using the dixon and bruce upand-down method; clinical descriptive study of citalopram-induced features and measurement of alterations in respiratory pattern (arterial blood gases and plethysmography) and biological parameters including blood lactate (scout ò , ekf diagnostic), plasma and platelet serotonin concentrations (high-liquid performance chromatography-fluorometry); determination of the preventive activity on seizures and death of diazepam, cyproheptadine, and propranolol pretreatments with the determination of their minimal effective dose; comparisons using anova for repeated measurements followed by bonferroni post-test.results. citalopram ip-mld was determined as 102 mg/kg in rats. seizures were significantly increased in rats receiving 80 and 120% of citalopram mld versus controls (p \ 0.01 and p \ 0.05, respectively), while death rate was only significantly increased in rats treated with 120% of citalopram mld (p \ 0.001). significant decrease in body temperature was observed after 90 min in rats treated with doses[60% mld in comparison to controls (p \ 0.05). occurrence of serotonin behavioural syndrome was comparable in all groups. citalopram administration did not result in significant hypoxemia, hypercapnia, and lactate elevation, thus not supporting the hypothesis of the occurence of any significant deleterious cardiovascular effect in citalopraminduced toxicity. however, a significant moderate increase in the inspiratory time (p \ 0.05) accompanied with an expiratory braking was observed. a significant decrease in platelet serotonin and increase in plasma serotonin concentrations were measured (p \ 0.05). pre-treatment with diazepam (1.77 mg/kg) and cyproheptadine (17.1 mg/kg) of rats receiving a lethal citalopram dose prevented seizures and death, while propranolol was ineffective.conclusions. citalopram respiratory toxicity remains mild, while deaths result from seizures probably related to serotonin toxicity. our observations may be helpful to better understand and manage human citalopram poisonings. objectives. to define the population pharmacokinetics (pk) of phenytoin in the critically ill, in addition to risk factors for sub-therapeutic dosing.methods. free and total ptn concentrations were measured in serum by means of high performance liquid chromatography following microfiltration, two to three times in the first 24 h after a loading dose. population pk modelling, including intra and interindividual variability, were determined using nonmem (r) . in the netherlands the use of diazepam is advised as first line treatment although evidence is not established and mainly provided through case-reports [1] . to compare the effect of diazepam on mortality in (hydroxy) chloroquine intoxication to standard therapy. we performed an extensive medline search (1950-april 2010) with a manual reference search of identified papers. (hydroxy) chloroquine intoxication studies and case reports in english, dutch or french were evaluated. patients older than 15 years with severe intoxications, based on measured concentrations or life-threatening symptoms, were included. pooled relative risk (rr) for mortality with corresponding 95% confidence interval (ci) were calculated by means of a fisher exact test. our results were compared with two retrospective and one prospective study.results. there were 32 case reports identified from which 25 case reports met our inclusion criteria. thirteen patients received diazepam of whom two died, compared to twelve patients who did not get diazepam of whom one died. statistical analysis demonstrated that treatment with diazepam was not associated with a lower mortality rate (rr: 1.1 ci 0.8-1.5; p = 1.0).although pooling of case reports is debatable, these results were comparable to the retrospective and prospective studies that didn't show any benefit from diazepam in chloroquine intoxication [2, 3, 4] . the positive effect of diazepam may have been underestimated, due to the fact that it has been given only as rescue therapy.conclusions. based on our analysis there is a lack of evidence concerning any antidotal effect of diazepam. good supportive treatment is pivotal. if the clinical manifestations of (hydroxy) chloroquine intoxications require sedation or treatment of seizures, diazepam is a good choice based on its pharmacological profile. a prospective study which compares diazepam to sedativa with similar pharmacokinetic and dynamic profile is required to prove that diazepam has any antidotal effect. introduction. brain is one of the first organs affected in sepsis and evaluation of brain function is difficult since patients are under sedation. it has been shown that mitochondrial dysfunction may play a significant role in the pathogenesis of septic encephalopathy. here we investigated inflammatory and metabolic parameters in a model of polymicrobial sepsis in mouse. methods. sepsis was induced by intraperitoneal injection of feces. animal received imipenem 6 h after the procedure. control animals received intraperitoneal saline and imipenem after 6 h. blood cytokines and serum lactate were measured. the animals were sacrificed by cervical dislocation. brain slices of 400 mcm were used to measure oxygen consumption and glucose uptake.results. interleukin 6, mip 1a and interleukin 1b significantly raised in the first 6 h after sepsis induction (p = 0.001; p = 0.0017; p = 0.0001 respectively). in 24 h only mip 1a was significant higher (p = 0.0001). lactate was elevated 6 and 24 h after sepsis induction (p \ 0.01 and p \ 0.001 respectively). oxygen consumption increased after 6 h of sepsis and drops under control values 24 h after the induction of sepsis. glucose uptake, measured by the nbdg fluorescence, was higher after 6 h (p = 0.0475) and 24 h after sepsis induction.conclusion. in a murine model of abdominal sepsis, inflammatory markers, lactate production, and brain glucose uptake increased and were parallel to alterations in the mitochondrial oxygen metabolism. introduction. the royal bournemouth hospital has one of the highest out-of-hospital cardiac arrest admission rates in the uk. in 2005, following ilcor/aha guidelines [1] , a cooling protocol was developed for patients with return of spontaneous circulation after advanced life support for ventricular fibrillation or pulseless ventricular tachycardia. in preparation for potential new ilcor/aha guidelines in 2010, the prospective database of outcomes for these patients was analysed.objectives. to evaluate the outcomes of therapeutic hypothermia for patients with return of spontaneous circulation following cardiac arrest. outcome data from our prospective registry of cooled patients are summarised.results. sixty-three patients were cooled in 4 years (median age 68 years; mode 76; range 19-88 years). 84% survived to itu discharge and 70% to hospital discharge. 98% of these were discharged home (14% to a rehabilitation hospital before home and one patient to a long term care facility). ninety-five percent of survivors were alive at 6 months and 80% alive at 1 year with seven status results still pending. median itu length of stay was 3.5 days (range 1-13). six patients required temporary percutaneous tracheostomies for airway protection and weaning from ventilation. median duration from itu to hospital discharge was 16 days (range 0-62).conclusions. this series is large by comparison to other uk centres. survival to hospital discharge, at 6 months and 1 year were better than other published results. although neurological outcomes were not formally assessed, we believe that the capacity to discharge home is a desirable patient outcome and represents the beneficial neurological effect of our cooling protocol. selection bias will have undoubtedly affected our results. however the age of our patients was higher than in published trials and in other reports is considered an adverse outcome predictor. our data would not support restricting induced hypothermia on the basis of age alone. we consider the itu and hospital lengths of stay required to discharge these patients to be long. these data were not reported in original trials. discharges may obviously be delayed for non-clinical reasons. this aside, neurological recovery progresses for months after cardiac arrest and discharge home may still prove possible if time is allowed. however, post-itu resource implications should be considered when introducing a cooling protocol. introduction. acute ischaemic stroke (ais) is the third largest cause of mortality and the leading cause of chronic disability in the industrialized world. in some parts of europe and the united states 26-43% of patients with ais may be admitted to a neurological intensive care unit (icu) for supportive therapy with 8-10% receiving mechanical ventilation [1, 2] . there are currently no agreed uk criteria for the admission of ais patients to critical care.objectives. to review the incidence and outcome of ais in our tertiary icu over the last five years. november 2005 and november 2009. ais was classified as thrombo-occlusive or embolic. subarachnoid haemorrhage and primary intracerebral haemorrhage were excluded. demographic and outcome data were recorded and compared against a mean value of all icu admissions.results. ais comprised 0.8% of icu admissions during the study period. demographic data is presented in table 1 as mean ± standard deviation or median (interquartile range) as appropriate. in 2008 4% (11/272) of hospital ais admissions were admitted to icu. 14 patients had surgical procedures including 8 decompressive craniectomies. 27% of survivors had a discharge gcs of 15/15. mortality for unselected medical admissions over the study period was 36%. there are differences of significance in the mortality according to the age, classified by age groups with an age cut off of 80 years (\80 years 38.6 vs. c80 years 40%, p \ 0.01). apart from the gcs, the rest of the variables analyzed in the ich score are not of significance; supra and infratentorial, presence of intraventricular blood neither on the divided volume over or under 30 cc although, in the latter, a p \ 0.1 can be observed and if we only analyze the supraventricular, it comes out as significant. other analyzed data are the time of the surgery, which is not significant, the need for mechanical ventilation, which is (42.9 vs. 15.8%, p \ 0.05), and the days of ventilation with a mortality clearly higher on those patients with\5 days of ventilation (83%) and on those of shorter stay (lesser then 5 days 79%).conclusions. let be remarked that the samples have been taken from patients admitted in the intensive care unit, losing a possible sample of less serious patients, and with a higher level of consciousness, what might explain why supra or infratentorial location and the volume don't come out as forecasting factors, since its likely that there are many small infratentorial outside the intensive care unit. we highlight also that the high mortality in the first few days can be caused by those patients who are admitted as donors, developing an encephalic death in the first days, conditioning also the data regarding the mortality on fewer days with ventilation. the finish up, we have to point out the fact that the presence of previous hypertension during the treatment might be a bad forecasting factor that should be deeper studied. to determine whether a delay exists between the time of diagnosis of intracranial haemorrhage and the time of reversal of anticoagulation, in patients presenting within our region. following approval by all audit and haematology departments a 6 month retrospective analysis was performed. we reviewed consecutive patients who received reversal of anticoagulation with pcc and vitamin k having presenting with intracranial haemorrhage whilst on warfarin. time of diagnosis was obtained from the time of scan and time of pcc issue was obtained from the blood bank database. case note analysis was performed to obtain further information.results. 47 patients were identified, 12 in the neurosurgical centre and 35 in peripheral hospitals. the median time from scan to issue of pcc was 84 min. 14 patients were reversed within 60 min and 14 patients waited longer than 120 min to have pcc issued. no adverse thromboembolic events were encountered.conclusions. avoidable delay exists between ich diagnosis and pcc issue. pcc could be stored in the emergency department and a stat dose administered immediately after diagnosis facilitating rapid correction of inr. repeat audit will be required to assess safety and efficacy. objectives. the aim of this study was to compare the functional ability and muscle strength between these two groups of patients. twenty-nine patients were evaluated (m:24, f:5) (age: 55 ± 17 years).the diagnosis of critical illness polyneuromyopathy was based on muscle strength measurement according to the medical research council (mrc) of muscle strength methodology. nine patients were diagnosed with critical illness polyneuromyopathy during their icu stay (mrc \48/60).the patients were evaluated with mrc and hand-grip dynamometry (hgd) every 10 days until their discharge from the hospital. the fim scale (functional independence measure) was used to evaluate the functional ability (18-126).the first evaluation was done at the discharge from the hospital and the second one 8 ± 1 months afterwards.results. the patients who developed critical illness polyneuromyopathy had statistically significantly lower mrc (39 ± 12 vs. 57 ± 2, p \ 0.001) and hgd at icu discharge (left 8 ± 5 kg vs. 23 ± 8, and right 9 ± 7 kg vs. 27 ± 9, p \ 0.001) compared to those who did not. the muscle strength as assessed with the mrc 10 days after icu discharge had statistically significantly lower (40 ± 11 vs. 56 ± 3, p \ 0.001), just as the second hgd evaluation (left 11 ± 7 kg vs. 25 ± 8 and right 9 ± 7 vs. 21 ± 8 kg, p \ 0.05).compared to those who did not develop critical illness polyneuromyopathy, the patients who did, had statistically lower fim values during their discharge from the hospital (49 ± 26 vs. 77 ± 30, p \ 0.05)and 8 months afterwards (90 ± 27 vs 125 ± 1, p \ 0.05).conclusions. the patients who developed critical illness polyneuromyopathy had significantly inferior muscle strength at their discharge from the icu. these patients also had lower functional ability. this functional ability remained defected even 8 months after their discharge from the hospital. these initial findings are suggestive that the appearance of critical illness polyneuromyopathy affects the patients mobility after their discharge either from the icu or from the hospital and persists for several months after icu discharge. further studies are needed to evaluate the effect of this impairment on the quality of life of these patients and also to evaluate therapeutic tools for critical illness polyneuromyopathy. introduction. this poster presents a qualitative system dynamics (sd) analysis of the factors which influence the care of acutely unwell ward patients in new zealand. this systems thinking approach is commonly used in organisational research and offers a way to make sense of complex relationships between variables. this approach has previously been used in health care to demonstrate differences in mental models between policy makers and clinicians (cavana et al., 1999) . since the factors which influence the care of acutely unwell ward patients are complex and multi faceted the qualitative sd method becomes an ideal analytic approach (e.g. see wolstenholme and coyle, 1983; senge, 1990; vennix, 1996; or maani and cavana, 2007) .objectives. the aim of this study was to examine the factors which influence the care of acutely unwell ward patients from an organisational perspective. key objectives were to determine the enablers and barriers to care from a nursing, medical and managerial (at ward and executive level) perspective.methods. using a multiple case study approach in four wards in two new zealand hospitals, focus groups and one to one interviews were conducted with key stakeholders identified as nurses, doctors and managers. initial coding of the data generated themes. these themes were then clustered to provide variables which were mapped to generate separate causal loop diagrams (clds) for each of the stakeholder groups to provide the basis for analysis. the clds were compared for characteristics and world views. preliminary results demonstrate a difference between clinical and managerial staff in characteristics and world view regarding the factors which affect the care of acutely unwell ward patients.conclusions. the qualitative sd approach has offered a novel and helpful way to make some sense of the complexity associated with caring for acutely unwell ward patients. organizational responses that may improve care delivery to these patients should be based on frank and open discussions between staff at all levels to ensure a shared mental model as the basis for change. objectives. the aim of the study is to explain the nursing in the technologicallyadvanced intensive care units. in this phenomenologically-designed study, a face-to-face in-depth interview was performed with 20 nurses, who were experienced for 1-22 years in the intensive care unit of cardiovascular surgery clinics. during the interviews, a semi-structured form was used. data were analysed using colaizzi's method of data analysis. the study was approved by the ethics committee of the institution.results. according to the nurses, nursing in technologically-advanced environment has three stages. these stages constituted three themes of the study: technology shock (first stage), understanding the technology-supported care (second stage), competency in technological environment (last stage). in the first stage, the nurses focus on themselves and technology; perceive the environment as frightening and complex. in the second stage, nurses gain control on technology, feel themselves safe and recognize their responsibility. in the last stage, the nurses experience anxiety related to their accountability. this anxiety may be motivating but also may be wearisome.conclusions. the nurses passes through three stages in a technologically-advanced environment. helping nurses to pass through these three stages appropriately will increase the contribution of technology to the patient care, more utilization of technology by nurses and more job satisfaction. unexpectedly, the compliance rate with the recommendations was significantly better over night. although the number of nurses is constant in the 24 h, the number of doctors is lower and less differentiated in the night shift. in an attempt to find an explanation for these findings we looked at the patient flow and time span until the first medical observation in the different time periods and we found that over night admissions (between 00:00 a.m. and 08:00 a.m.) corresponded only to 10% of all admissions and were seen sooner, which might explain our findings. a. objectives. the purpose of the study was to assess whether the completion of the sepsis resuscitation bundle within the first 6 h after icu admission, but beyond the specific time limit of the various bundle interventions, is related to an improvement in survival in patients with severe sepsis/septic shock. this was a single-center prospective observational study of patients admitted to the medical-surgical icu of an urban tertiary care teaching hospital with severe sepsis/septic shock. patients were recruited from june 2005 to november 2009. we assessed the compliance with the different tasks included in the 6-h resuscitation bundle. furthermore, we ascertained within the first 6 h after icu admission the compliance with those tasks not carried out within their specific time limits; we have called this variable ''bundle improvement at the icu''. results were stratified by the number of tasks of the bundle completed before admission at the icu, and the lag time between the beginning of severe sepsis and admission to the icu. these late completed tasks at the icu were related to hospital mortality by a cox regression model. objectives. the aims of this study were to assess the compliance rate with 6 h bundle as defined in the surviving the sepsis campaign guidelines 2008 in patients diagnosed with sepsis regardless of severity and whether compliance affects the rate of mortality and/or hospital stay. we conducted a prospective observational study. we randomly recruited 145 adult patients from acute admissions unit and intensive care in an acute district general hospital in england who met the diagnostic criteria for sepsis. for each patient, compliance with sepsis care bundle was obtained from medical notes. the following components of the 6 h sepsis bundle were assessed: obtaining blood cultures, initiating antibiotic therapy, measuring serum lactate and in the event of septic shock administration of fluid therapy. conclusions. long and unacceptable delays in admission to iccu were identified despite evidence of significant organ dysfunction in many of these patients. with all bundle elements being met for only 1 patient it is apparent that evidence based endpoints aimed at reducing mortality from severe sepsis are not being met despite all the bundle elements being practically deliverable. poor compliance with taking blood cultures prior to antibiotic administration and lack of scvo 2 measurement are areas requiring particular attention. further work is recommended to identify potential contributing factors to non-compliance. introduction. international guidelines recommend that cardiac output measurement is required in addition to arterial pressure monitoring in patients with persistent shock after initial therapy [1] . nevertheless, these recommendations are not supported by any comparison of arterial pressure and cardiac output for monitoring the effects of the most current treatments like fluid therapy. objectives. to evaluate in which extent monitoring the haemodynamic effects of a standardized fluid challenge with the sole arterial pressure could help for detecting the fluidinduced changes in cardiac index (ci). in 228 critically ill patients with acute circulatory failure deemed at receiving a 500-ml saline infusion over 20 min, we measured the systolic (sap), diastolic (dap), mean (map) and pulse (pp) arterial pressure and transpulmonary thermodilution ci before and after volume expansion.results. volume expansion significantly increased ci, sap, dap, map and pp by 24 ± 25%, 15 ± 19%, 9 ± 16%, 13 ± 17% and 21 ± 29%, respectively. the fluid-induced changes in pp, sap and map were significantly correlated with the fluid-induced changes in ci (r = 0.56, 0.55 and 0.37, respectively). the changes (in %) in pp were significantly related to the changes (in %) in stroke volume for all quartiles but with different coefficients of correlation: r = 0.39 for the 1st quartile (36-53 years), r = 0.44 for the 2nd quartile conclusions. pp and sap were the best arterial pressure values for detecting the fluidinduced changes in ci. using the sole pp for assessing fluid responsiveness led to a non negligible proportion of false negative cases. this supports the recommendation that when a precise monitoring of fluid resuscitation is required, like in refractory shock, a direct assessment of cardiac output is required. objectives. aim of our study is to show that it is possible to reduce high catecholamines in previous improper volume resuscitated patients by forced volume resuscitation combined with active dose reduction and generate the hypothesis of an avoidable catecholamine induced circulation injury. introduction. the sialic acid content of the red blood cell (rbc) membrane decreases early in sepsis [1] , and this alters the rbc shape and metabolism [2] . an increased ratio of the rbc proteins band 3/alpha spectrin was observed in a mouse model of septic shock, suggesting a possible alteration of the rbc membrane integral/peripheral proteins ratio [3] . as there are interspecies differences in membrane composition, these observations need confirmation in humans. we studied rbcs from patients with (n = 15) and without (n = 9) sepsis at icu admission and on day 3 in the septic patients. exclusion criteria were recent rbc transfusion, hematologic diseases, cirrhosis and diabetes mellitus. procedures included screening for rbc membrane protein alterations by cryohemolysis test and separation of the rbc membrane and skeletal proteins by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate [4] . comparison between groups was made by the student's t test or the mann-whitney test. a p value .05 was considered as statistically significant.results. the hemogram, including reticulocyte count was similar in septic and non-septic patients at icu admission. no significant difference was observed for cryohemolysis test results and the amount of the rbc proteins (table 1) . objectives. our purpose was to compare a new method (patrol fr 08-02110) with the reference method (randox tm ) during cbp. patients scheduled for coronary artery bypass (cb) and aortic valve replacement (avr) under cbp were enrolled after written informed consent in this protocol approved by local ethics committee. anesthesia protocol was standardized with systematic use of tranexamic acid. three blood samples were harvested: t1 = induction; t2 = 30 min. after cross aorta clamping; t3 = 24 h after induction. the patrol method was performed after serum exposition to a photosensibilizer agent then to a laser irradiation leading to the formation of free radicals. oxidation by those free radicals of a fluorometric sensor allowed an indirect measure of tas. this measurement in arbitrary unit (au) corresponded to area under curve compared to a control value from a pool serum. a value higher than 1 indicated a lower capacity for the given serum to neutralize free radicals whereas a lower value indicated a higher capacity. the same sample allowed tas determination (lmol/l) with randox tm method. results were expressed as absolute numbers, mean ± sd. tas were compared with anova test; p \ 0.05 was significant.results. the seven patients (5 male, 2 female; 71 ± 10 years old) enrolled underwent cardiac surgery (4 cb and 3 avr) without any problem. there was no variation in tas determination with the randox tm method: t1: 1.39 ± 0.13; t2: 1.41 ± 0.26; t3: 1.33 ± 0.15 lmol/l. conversely a two fold significant increase was measured during cpb with the patrol method: ti: 1.12 ± 0.31; t2: 2.21 ± 0.60*; t3: 0.99 ± 0.34 au. *p \ 0.001 versus t1.conclusions. oxidative stress due to overwhelming release of reactive nitrogen/oxygen species (rn/os) is held largely responsible for sepsis-induced organ failure and mortality [2] . up-front and/or ongoing distortion of the pro-oxidant/anti-oxidant balance is likely to play an important role in this situation and in ischemia-reperfusion. therefore the patrol test which appeared to be more sensible than the randox tm method could a good tool in these cases and for evaluation of new anti-oxidant treatments in critical care medicine. these results have to be confirmed in a larger population. introduction. sepsis is the leading cause of death in critically ill patients. despite attempts to improve standardized strategies in resuscitation and treatment of sepsis, the morbidity and mortality remain unacceptably high. early diagnosis and stratification of the severity of sepsis is the key to start timely the appropriate treatment. sepsis is the systemic inflammatory response syndrome to infection; it can lead to hypoperfusion and organ dysfunction and at the cellular level to aerobic mitochondrial dysfunction. lactate is the product of anaerobic metabolism and thus may serve as a prognostic factor in this subset of patients.objectives. the authors propose to test the association of the first serum lactate at hospital admission with shock and icu mortality in patients with community-acquired severe sepsis. during the study period 1,223 patients were admitted in the unit, of those 300 (25%) had severe community-acquired severe sepsis (cass). crude icu mortality rate among cass was 33%. considering the model previously described in methods and when the variables were adjusted only gender, age, saps ii, severity of sepsis and serum lactate were retained in the final model for icu mortality and saps ii nad serum lactate for shock (see table 1 ). a first blood lactate level was independently associated with shock and icu mortality in patients community-acquired severe sepsis admitted in intensive care. objectives. the objective of this study was to test whether svo 2 can predict fluid responsiveness in these patients. we studied 65 patients who were monitored with a pulmonary artery catheter for severe sepsis and septic shock. hemodynamic measurements were obtained before (baseline values) and after a fluid challenge with colloids or crystalloids. responders were defined as those with a[10% increase in cardiac index (ci). no additional interventions were performed during the test. student's t test and linear correlation were used for the statistical analysis.results. mean patient age was 70 ± 2 years and the mean sofa score 10 ± 2. mean arterial pressure was 71 ± 9 mmhg, cardiac index 3.0 ± 0.8 l/min/m 2 , pulmonary artery balloon-occluded pressure 13 ± 3 mmhg, and heart rate 104 ± 19 bpm. thirty-four patients (52%) responded to the fluid challenge. responders and non-responders had similar baseline svo 2 (67 ± 9 vs. 67 ± 8%, p = 0.98). baseline svo 2 was[70% in 13 responders (38%) and in 11 non-responders (35%). there was no correlation between changes in ci (%dci) and the baseline svo 2 (fig. 1) . sepsis is a disorder of microcirculation [1, 2] . although the pathogenesis of microvascular dysfunction in sepsis is extremely complex, neutrophil activation and their interaction with endothelial cells are considered central features of sepsis-induced microcirculatory alterations. to our knowledge, however, no study evaluated the microvascular pattern of septic patients with chemotherapy-induced severe leukocytes depletion.objectives. to assess early microcirculatory response to sepsis in patients with and without drug-induced neutropenia.methods. demographic and hemodynamic variables together with sublingual microcirculation recording (ops-sdf videomicroscopy) were collected in four groups of subjects: septic shock (ss, n = 9), septic shock in neutropenic patients (nss, n = 8), neutropenia without inflammation (neutr, n = 7) and healthy controls (crtl, n = 13). except for controls, all measurements were repeated after complete resolution of septic shock and/or neutropenia (tp2). collected video-files were processed using appropriate software tool and semi-quantitatively evaluated (functional capillary density, fcd (cm/cm 2 ); mean flow index, mfi [1] ) [3] . conclusions. microvascular derangements in sepsis did not differ between non-neutropenic and neutropenic patients. surprisingly, neutropenia per se without measurable systemic inflammation was also associated with alterations of the sublingual microcirculation. although we cannot exclude the role of residual neutrophils, our data could indicate that leukocytes are not the only and exclusive modulators of septic microvascular dysfunction. in addition, the role and mechanisms of microvascular changes associated with chemotherapyinduced neutropenia warrants further investigation. multiple organ failure is a leading cause of death in critically ill patients. improvements in outcome will most rely on our capacity to measure rapidly accessible biomarkers.objectives. to investigate if the time sequence of reactive oxygen metabolites (roms) production with sofa score could be prognostic for outcome. the study included 33 critically ill patients (from september to december 2009) who had roms measured (hydroperoxides) during icu stay, when the diagnostic criteria for sepsis (observed n = 14), severe sepsis (observed n = 11) and septic shock (observed n = 68) were present, 4-5 days and 3 weeks after the diagnosis (samples n = 93); on the same days, the sofa score was calculated. the plasma roms values were assayed by a diacron-italia kit, applied to an automatic instrument (olimpus au 640). statistical analysis was performed used mann-whitney test and the linear regression analysis. the roms values and sofa score were inversely correlated (r 2 = 0.64 for sepsis; r 2 = 0.42 for severe sepsis; r 2 = 0.37 for septic shock). the droms (the difference between the first and the last measurement of roms levels in each individual patient) was significantly different between survivors and non-survivors. clinical characteristics of the patients are presented in table 1 . values are presented as median and interquartile rangers. a p value .05 was considered as statistically significant.conclusions. the plasma roms values decreased when the critically conditions rapidly evolved towards organ failures with higher sofa. to explore: (a) stress neuropeptides (acth, cortisol, prolactin, neuropeptide y (npy) and substance p (sp)) in critically ill subjects and controls, (b) potential association between levels of stress neuropeptides, disease severity and pain. a prospective correlational study, with repeated measurements and cross-sectional comparisons. fifty-three critically ill patients with diverse primary diagnoses and 53-age and gender-matched healthy controls were studied for 8 days. serum neuropeptides were quantified by elisa (npy, sp) and chemiluminescence immunoassays (acth, cortisol, prolactin). pain levels were assessed by payen and puntillo scales. clinical severity was quantified by multiorgan failure scoring system (mof) and the multiple organ dysfunction score (mods). results. we observed: (a) statistically significant differences between critically ill and control subjects in regard with cortisol (p \ 0.0001), npy (p \ 0.0001) and sp (p \ 0.05) levels throughout the study. specifically, cortisol levels were higher and npy and sp levels were lower in patients compared to controls, (b) significant bivariate associations between stress neuropeptides (p \ 0.04), (c) statistically significant associations between acth and pain intensity levels assessed by payen (r = 0.391, p = 0.018) and puntillo (r = 0.509, p = 0.002) scales. there was also a constant but not statistically significant (p = 0.1) trend for lower sp levels in patients receiving opioids than in controls. moreover, npy levels were significantly lower in patients receiving analgesia (p = 0.042), (d) lower acth and cortisol levels in survivors (p \ 0.05) (e) at the day of least severity, a significant association between sp levels and mof was observed (r = 0.29, p \ 0.05).conclusions. (a) despite the fact that npy and sp are stress neuropeptides, their levels appear to be decreased in mods patients. it is worth-exploring whether critical illness may be a state of suppressed activity of some neuropeptides, (b) the observed association between stress neuropeptide levels and survival in critical illness needs to be explore further, (c) bedside measurement of selected neuropeptides in the future may provide an estimation of pain in uncommunicative patients.hence, the study of stress neuropeptides may provide new insight for the management of the critically ill. objectives. the objective of this study was to compare septic and non-septic inflammatory process in critically ill patients with respect to paraoxonase 1 activity, lipid profile and lipid peroxidation markers. methods. analyzed were serum paraoxonase 1 activity, lipid profile, oxidized low density lipoproteins and conjugated dienes in critically patients with sepsis n = 30), age/sex/ap-acheii matched critically ill controls with non-septic sirs (n = 15) and age/sex matched outpatient controls without inflammation (n = 30).results. the activity of pon 1 was lower in septic patients (96.7 ± 47.4 u/ml) as well as in patients with non-septic sirs (107.8 ± 39.8 u/ml) compared to healthy controls (158 ± 40.5 u/ml). the decrease in paraoxonase 1 activity, high density lipoprotein cholesterol and apolipoprotein a-1 concentrations was closely followed by the counter increase of serum amyloid a in both groups of patients. there was no difference in paraoxonase 1 activity between septic and non-septic critically ill patients. the concentration oxidized low density lipoproteins and conjugated dienes as markers of lipid peroxidation, were raised in both septic and non-septic sirs critically ill patients as compared with healthy controls. however there was no difference between both critically ill patient groups.conclusions. the decreased activity of paraoxonase 1 in negative correlation with lipid peroxidation markers offers a potentially useful nonspecific marker of inflammation in critically ill patients.grant acknowledgment. objectives. in the present study, we studied the short-term and direct effects of ivig with sepsis.methods. 16 patients was investigated. following the administration of 5 g of ivig for 1 h, we took blood samples immediately following ivig treatment and at 1 h after ivig treatment. blood samples taken at 1 h and just prior to ivig administration were used as controls. while there was no difference between 1 h before and just prior to ivig treatment, statistically significant decreases were observed in the levels of il-6 after the administration of ivig. no significant changes were observed in the levels of tumor necrosis factor-a and high mobility group box-1.changes in serum tnfa, il-6, hmgb1 we confirmed the results of previous animal studies. while we reported that the administration of ivig directly reduces the levels of il-6 in patients with sepsis, a further prospective study of the ant-cytokine effects following ivig treatment will be conducted in the near future. objectives. to investigate the levels of nucleosome in septic patients and to determine whether nucleosome could serve as a biomarker for sepsis. sixty-four consecutive patients who were newly admitted in surgical intensive care unit at two university hospitals were enrolled in this study. whole blood samples were drawn within 12 h of admission and on the third, fifth and seventh days. a last blood sample was drawn after recovery at icu discharge in survivors or at imminent death in the cases of non-survivors. plasma levels of nucleosome as well as cytokines il-6 and il-10 were detected by means of enzyme linked immunosorbent assay. . fifty patients were diagnosed as sepsis and the other fourteen patients were classified as controls. plasma levels of nucleosome were significantly higher in septic patients than in controls (two-way anova, p \ 0.05), while the levels of il-6 and il-10 were comparable between septic patients and controls. the septic patients presented the highest levels of nucleosome on the admission day, which was significantly different from the admission levels of nucleosome in controls (4.16 ± 3.48 vs. 1.61 ± 1.36, p \ 0.05). the plasma levels of nucleosome between survivors and nonsurvivors showed no statistical significance.conclusions. plasma levels of nucleosome may serve as a valuable biomarker for sepsis.introduction. high mobility group box protein 1 (hmgb-1) is a cytokine that can mediate inflammatory response in different conditions included rheumatoid arthritis, infections, sepsis and septic shock. hmgb-1 released by activated macrophages/monocytes acts as a late mediator of sepsis. studies have shown that serum hmgb-1 concentrations were elevated in patients with severe sepsis.objectives. in the present study, we evaluated the role of the hmgb-1 levels at the time of admission at the intensive care unit (icu) as predictor of outcome in patients with sepsis and septic shock.methods. forty-four patients admitted to the icu with sepsis and septic shock was recruited. serum samples were obtained at the time of admission for the determination of hmgb-1 levels. the results were correlated with the origin of sepsis, severity, organ dysfunction, requirements of mechanical ventilation and vasoactives, days at the icu, comorbidities and mortality at the icu and 28 days after admission. twenty-six patients were male (68.2 vs. 31.8%). septic shock was present in 28 patients (63.6%). the mortality rate at the icu was 31.8% (n = 14) and 38.6% (n = 17) at day 28th. hmgb-1 levels were 34.7 ng/ml ± 39.02 (1.34-230.2 ng/ml). hmgb-1 levels were significantly higher in non-survivors at the icu than in survivors (56.81 ng/ml ± 58.69 vs. 24.38 ± 19.18, p \ 0.05). higher levels of hmgb-1 in serum at the admission were correlated with a higher mortality rate in the icu (p \ 0.05) but not at day 28th (p = 0.07). these levels were not correlated with days at the icu, requirements of vasoactives, mechanical ventilation, and apache score.conclusions. the determination of hmgb-1 levels at admission at the icu in patients with sepsis and septic shock is a good predictor of worse outcome and lethality.introduction. recent experimental and clinical data (1, 2) support the hypothesis that costimulatory molecules, such as cd40, play an essential role in the innate immune response during sepsis. expression of cd40 on the surface of monocytes could represent an important pathway in the modulation of the production of several key inflammatory mediators.objectives. to investigate whether the expression of cd40 molecule on the surface of plasma monocytes differs among the various stages of sepsis. a total of 32 participants (12 icu patients with sepsis, 14 icu patients with septic shock and 6 healthy controls) were included in the study (male patients 73.1%, mean age 46.6 ± 16.8 years). inclusion criteria: icu patients on mechanical ventilation with first episode of sepsis or septic shock during current hospitalization. exclusion criteria: immunosuppression, neoplasia, autoimmune disease, cardiovascular disease. age, gender and comorbid conditions were recorded. a blood sample for quantification of cd40 expression was obtained at the time of enrollment (day 1), and on the fifth day after the onset of sepsis; measurement was made on the same day. cd40 expression on the surface of plasma monocytes (on days 1 and 5) was assessed by flow cytometric analysis. statistical analysis: kruskal-wallis test to identify difference of cd40 expression among the 3 groups was performed. post-hoc analysis was made by mann-whitney u test between independent groups, using bonferroni correction for multiple comparisons. roc curve analysis was used to determine the accuracy of cd40 in identifying patients with sepsis or septic shock. patients with sepsis had significantly higher levels of cd40 (day 1) compared with healthy controls subjects (0.86 ± 0.41 vs. 0.34 ± 0.07, p b 0.006). on the contrary, patients with septic shock did not show any significant difference compared with controls. a roc curve analysis for cd40 (day 1) (auc = 0.72, p b 0.05), revealed that a cut-off value of 0.53 could predict patients with sepsis with a sensitivity of 91% and a specificity of 57%.conclusions. upregulation of cd40 expression may reflect a protective phenomenon during sepsis. on the contrary, low cd40 expression could represent impaired immune function associated with more severe disease. in order to increase the cardiac output in the septic shock patients, according to surviving sepsis campaign team, norepinephrine (ne) or dopamine administration was recommended. the both agents increase the sympathetic tone which antagonize against parasympathetic activity used for gastrointestinal motility (involved gastric emptying). then, it is raised a question whether ne delayed the gastric emptying or not.objectives. this study was aimed to evaluate the gastric emptying in the septic shock patients with norepinephrine. a prospective observational study involved 54 adult septic shock patients, who received ne continuously in icu sardjito general hospital (yogyakarta, indonesia). patients with any head pathologies (trauma, surgical procedures for tumor or bleeding), any gastrointestinal or abdominal pathologies (diarrhea, trauma, surgical procedures for cancer, peritonitis, ileus etc.), and administrations of metochlopramide or alinamin were excluded. nutrition fluids (100 ml) was given passively via nasogastric tube, then after 60 min the tube was aspirated. the volumes of aspirates were recorded in % as a gastric residue. once measurement was done with time randomly for every patient. at the measurement time were recorded the dose of ne and the vital signs.results. the gastric residues were 41.76 ± 28.11% (17 patients), 45.46 ± 25.27% (13 patients) and 13.57 ± 8.52% (7 patients) for the doses of ne of 0.1, 0.2 and 0.3 lg/kg b.w./ min respectively. at the ne doses of 0.4, 0.5 and 0.6 lg/kg b.w./min, all of the gastric residues were zero (17 patients). the correlation between the ne doses and the gastric rescues was statistically significant (p: 0.01). the mean arterial pressures (map) were 84.16 ± 18.01 mmhg (ranges from 58 to 120 mmhg. there was no significantly correlation between map and the gastric residues.conclusions. the gastric emptying in the septic shock patients was not disturbed by administration of ne. introduction. anemia is a frequently encountered problem on the intensive care unit. several factor lead to anemia, among which are traumatic blood loss and the drawing of blood for routine laboratory tests. it's not known how this may affect innate immunity. hepcidin is a central regulator of iron homeostasis. it is induced in response to iron and inflammation and reduced in response to anemia and hypoxia. the suppression of hepcidin leads to the internalization and degradation of the iron exporter ferroportin on intestinal cells and macrophages, leading to the uptake of iron from the gut and the release of iron from the macrophages from the reticulo-endothelial system (res). these cells are central to the innate immune response and the altered iron status of these cells due to suppression of hepcidin may affect the inflammatory response of these cells. we tested the hypothesis that phlebotomy in human volunteers would lead to a suppression of the innate immune response. this abstract provides data of a pilot study carried out in 3 subjects. to investigate the effect of phlebotomy on the innate immune response of whole blood in human volunteers.methods. three volunteers were subjected to the letting of 400 ml of blood by phlebotomy. blood for the determination of hemoglobin and iron parameters, leucocyte count and differential, and hepcidin-25 was drawn at day 0, 4 and 7 after phlebotomy. further whole blood stimulation was carried out at each time point by adding 0.5 ml heparin anticoagulated whole blood to a prepared tube containing endotoxin, pam3cis or rpmi as a control. final concentrations of lps and p3c were 10 ng/ml and 1 lg/ml respectively. these tubes were incubated at 37°c for 24 h and centrifuged for 10 min at 4,000 g. the supernatant was frozen at -80 until the measurement of tnf-alfa and il-6 by elisa. cytokine production was corrected for the number of monocytes present. data are expressed as mean ± sem. hemoglobine decreased from 8.5 ± 0.3 mmol/l at baseline to 8.0 ± 06 at day 4. it returned to normal at day 7. there were no apparent changes in serum iron levels. there was however a clear decrease in serum ferritin levels from 162 ± 48 at baseline to 109 ? 46 at day 7. leucocyte count and differentiation did not show any significant changes. hepcidin was clearly suppressed from 4 to day 7 after phlebotomy (from 6 ± 1 to 2 ± 1). tnf-alfa production dropped from 81 to 47 ng/10 7 monocytes at day 7. il-6 production dropped from 327 to 195 ng/10 7 monocytes. hepcidin levels correlated well with cytokine production (r 2 0.53 for tnf-alfa, r 2 0.51 for il-6).conclusions. phlebotomy leads to suppression of the innate immune response in whole blood. this could be a result of the intracellular decrease of iron in immune cells due to the systemic suppression of hepcidin. these findings are relevant to critical care patients that are subject to the repeated drawing of blood while their immune system is often compromised. introduction. hypothermia and hyperthermia occur in many pathological states presenting to the emergency department. both these processes are known to significantly impair coagulation pathways but as yet there is little evidence to show what affect they have on the evolving clot structure. previous studies have attempted to determine the effect of temperature on whole blood coagulation using techniques such as thromboelastometry (teg) but its ability to provide meaningful outcomes in terms of clot quality and structure remains elusive. recent studies have highlighted the potential of a new technique, gel point (gp) and fractal dimension (d f ), as a functional biomarker in haemostasis. to explore both the changes in coagulation pathways and their associated effect on clot structure and quality based on the new biomarkers, gp and df. following full ethical approval, 30 healthy whole blood samples were obtained from individuals and tested at temperatures of 27°c (n -6), 31°c (n -6), 35°c (n -6), 37°c (n -6), 41°c (n -6). an oscillatory shear technique [1] using an ar-g2 instrument (ta instruments) was applied to each sample. the gp, which indicates the formation of the fibrin network, was obtained for each sample using the chambon-winter gel point criterion [2] . this method provides the basis from which d f can be determined [3] to interpret the structural properties of the clot network. the results were compared with the standard teg analysis. firstly, results showed a significant progressive change in the clot structure by this new biomarker across the whole temperature range (27-43°c). secondly, it also highlighted a significant and meaningful correlation between coagulation pathway change (time to gp, tgp) and the eventual clot outcome (fractal dimension). the tgp of the incipient clot was prolonged and the corresponding d f decreased with reduced temperature values. although, the changes in the coagulation pathway of the teg (r time) and the rheometer (tgp) correlated, the new biomarker, d f , provided additional structural data on the fibrin network formed and highlighted the relationship between coagulation pathway changes and the eventual fibrin clot structure.conclusions. in this study, we describe and quantify for the first time how temperature affects the coagulation pathways and how this impacts on the fibrin clot network, morphology and strength by using the new biomarkers, gp and d f . the potential of these new biomarkers in determining the effects of temperature change in critical illness and injury needs to be evaluated clinically. key: cord-014996-p6q0f37c authors: nan title: posters_monday_12 october 2009 date: 2009-08-06 journal: intensive care med doi: 10.1007/s00134-009-1593-2 sha: doc_id: 14996 cord_uid: p6q0f37c nan a huge variability in excess risk of death, ranging from 0 to 40%, from ventilator associated pneumonia (vap) have been reported in the literature [1] . this large between-study variation can be attributed to difference in definitions but also to incorrect estimation by standard statistical methods, i.e. inappropriate adjustment for informative censoring and time dependent confounders. the aim of this study was to take into account above statistical shortcomings and to assess the excess risk of vap by using an extension of a recently developed techniques from the field of causal inference [2] . materials and methods. data was retrieved from a large longitudinal, high quality multi-centric icu database from france (outcomerea). a random sample of consecutive patients ventilated [48 h from 17 icus over a 10 year period were included. vap was defined as clinical suspicion plus at least one positive proximal or distal sampling with quantitative count using classical thresholds. only the first vap episode was taken into account. we considered discharge from the icu as a competing risk and estimated the attributable 30-day icu mortality of vap by comparing the counterfactual cumulative incidence of death for the entire population under different hypothetical infection paths. baseline characteristics indicating underlying co-morbidity and longitudinal (daily measured) severity of illness indicators together with all other known confounders until vap developed were taken into account through the use of a marginal structural model [2] . results. a total of 4,333 icu patients were included. mean (sd) age and saps ii score were 63 (17) and 49 (18), respectively. seventeen and 21% were admitted after scheduled and emergency surgery respectively, 62% were medical patients. forty-two percent had an underlying chronic illness (knaus). nine percent received dialysis in the icu. a total of 647 (15%) patients developed vap within 30 days of admission (48 and 82% within 7 and 14 days, respectively). crude icu-mortality rates in patients with and without vap were 35 and 25%, respectively. when taking into account all the confounders, we found a 3.5% increase (95% ci 1. 7-5.3 , p = 0.0002) in the hazard of 30-day icu-death per additional day since the development of vap. provided vap could have been prevented in the whole population 30-day mortality would have decreased by 0.7%. conclusion. the excess risk of death from vap estimated by marginal structural models is lower than commonly reported in the literature. this indicates that underlying comorbidities and the evolution of severity of illness until vap are insufficiently taken into account by current standard statistical methods. reference (s) . 1. rello et al (2003) introduction. the intensive care society (uk) has recently published national guidelines regarding many aspects of percutaneous tracheostomy management [1] , however, these guidelines do not make any recommendations regarding antibiotic prophylaxis during the procedure. we recently audited uk practice and have established that only 9% of the units give prophylactic antibiotics for patients known to be colonised with methicillin-resistant staphylococcus aureus. the low rate of antibiotic use is surprising given that pneumonia and/ or bacteraemia following pt is frequently caused by organisms (non-mrsa) that colonise the patients' skin and/or airway [2, 3] . objective. to establish the incidence of mrsa positive sputum and/or blood cultures following pt in patients colonised with mrsa in their nose or throat. methods. we audited all the patients who had pt performed between 2005 and 2009 who were known to be colonised with mrsa in their nose or throat, (but who had negative sputum cultures) before pt was undertaken. we wanted to find how many of these patients developed mrsa positive sputum and/or blood cultures in the first week following their pt. results. from a total of 2004 patients admitted to critical care between 2004 and 2009 only seven mrsa colonised patients required pt. all of these 7 patients had mrsa colonised throat or nose with negative sputum and blood cultures prior to the pt. no patients were given prophylactic antibiotics during the pt as this was our standard practice. four (57%) developed mrsa positive sputum cultures in the first week following the procedure. one (14%) developed mrsa bacteraemia on day 4 following pt. over the same period there were no case of mrsa bacteraemia in the 118 mrsa colonised patients who did not undergo pt. microbiology bacterial biofilm has been observed in the surface of the endotracheal tube (ett) in mechanically ventilated patients and recent studies relate the presence of biofilm with the incidence of ventilator-associated pneumonia (vap). acinetobacter baumanii is a gramnegative opportunistic nosocomial pathogen involved in the production of vap, and capable of biofilm formation on abiotic surfaces. objective. to analyse the presence of biofilm in the ett by using scanning electron microscopy (sem) and to identify the microorganisms contributing to its formation in patients admitted in an icu endemic for a. baumanii. methods. from march to september 2008, 86 consecutive patients admitted to our unit and mechanically ventilated were included in the study. etts after extubation were (a) sent to microbiological culture, and (b) fixed with 4% paraformaldehyde-2% glutaraldehyde for one hour, dehydrated with increasing ethanol concentrations, and processed for sem. etts were observed under sem to assess the presence and extension of the biofilm, and to recognize bacterial or fungal forms. results. there were 57 males (66%) and 29 females (44%) with a median age of 60 years (range 16-86).the median apache ii score in the first 24 h was 20 (range 14-45). the median duration of intubation was 6 days (range . causes of intubation were coma in 41 patients (48%), respiratory failure in 34 (40%), and heart failure in 11 (13%). the microbiological isolations showed: a. baumanii (24%), staphylococcus non aureus (16%), pseudomonas spp. (16%), streptococcus viridans (13%), staphylococcus aureus (12%), enterococcus faecalis (9%), candida albicans (8%), and others (15%). under the sem, biofilm was identified in the 79% of all cases and was abundant in 29 (34%), regular in 25 (30%), and scarce in 13 (15%). morphological identification of microorganisms observed under sem showed: cocci in 11 (13%), bacilli in 15 (17%), and yeast in 5 (6% introduction. hospital-acquired infection is often linked to the standard of ward cleaning however the impact of increased quality of cleaning and deep cleans are unknown. objectives. this study aimed to determine the effect of enhanced cleaning on local contamination rates of hospital pathogens and whether this results in a reduction in patient colonisation. a cross-over one-year study was performed in the intensive care units (icu) of two teaching hospitals, which screened patients weekly for mrsa. in randomized two-month periods and in addition to conventional cleaning using detergent and mops, high contact areas were cleaned twice daily by a team of trained hygiene technicians using microfibre cloths. using contact plates, samples were taken at nine sites around the bed area and ward over 12 bed-days per week. hand hygiene was encouraged and compliance audited. results. only 2.5% of the planned 20, 736 local samples were missed and this was equal between study phases. average hand hygiene compliance was similar between enhanced and standard phases [hospital a 47.8% (261/546) vs. 56% (268/478) and hospital b 57.3% (259/ 452) vs. 52.7% (266/505)]. patient characteristics were similar during standard and enhanced cleaning periods. of the sites tested, samples taken from bed rails were most likely to be contaminated with mrsa (or = 4.16; 95% ci: 2.65-6.54) followed by nurses' hands (or = 1.89; 95% ci: 1.14-3.11). analysis of these site-samples also confirmed that enhanced cleaning significantly reduced environmental contamination (or = 0.45; 95% ci: 0.34-0.61; p \ 0.001). the effectiveness of enhanced cleaning in removing mrsa contamination did not vary with the sample site. a sub-group analysis of samples only taken from nurses' hands showed a non-significant reduction in mrsa hand contamination associated with enhanced cleaning although associated uncertainty was large (or = 0.56; 95% ci: 0.29-1.08; p = 0.077 conclusions. this is the first prospective controlled study examining the effectiveness of enhanced cleaning in preventing spread of multiresistant pathogens within icu. although both environmental and hand contamination were reduced, enhanced cleaning of high contact surfaces was not associated with a reduction in cross infection. conclusions. isd of secretions reduces the incidence of vap in patients receiving. css alone, or in combination with isd has no significant effect on incidence of vap. hence, isd may be recommended for vap prevention, considerations other than prevention of vap should determine the choice of the suction system in a mechanically ventilated patient. to show a mortality benefit, larger, multi-center trials may be required. decreasing incidence of ventilator-acquired pneumonia (vap) is increasingly regarded as a priority in icu quality programs. subglottic secretion suctioning (sss) has been associated with a decreased risk of vap. a previous metaanalysis concluded that sss reduces the risk of vap, but it included only five randomized controlled trials (rct), and sss is still underused, perhaps considering the available evidence is insufficient. we planned a systematic review and metaanalysis of sss for vap prevention. pubmed, embase and cdsr were searched for rct studying the influence of sss on vap incidence. additional outcomes were mortality (within icu or hospital), icu and hospital stay, mechanical ventilation duration and time from intubation to vap diagnosis. additional references and sources of information were searched, and authors were contacted as necessary. 11 rct were found, but one of them was excluded for not having enough data for analysis. 10 rct were analysed, including 2,019 patients. quality of the rct was only moderate. qualitative outcomes were homogeneous between studies, so were analysed by a fixed effects model; quantitative outcomes were very heterogeneous, and were analysed by a random effects model. compared to control, sss decreased vap incidence (rr 0.53; 95% ci 0.43-0.66), but not mortality (rr 0.97; 95% ic 0.80-1.16). sss delayed vap onset for 4.08 days (95% ci 2.63-5.52), shortened mechanical ventilation for 1.58 days (95% ci 0.71-2.45) and decreased icu length of stay for 1.97 days (ic 95% 0.02-3.91). in two rct, no differences were found in the hospital length of stay. conclusion. sss reduces vap incidence and delays vap onset, shortening mechanical ventilation and icu length of stay, but not decreases icu or hospital mortality. data from 10 rct support the use of sss as an adjunctive tool to prevent vap. introduction. in comparison to ventilator-associated pneumonia (vap), less data are available on ventilator-associated tracheobronchitis (vat). however, vat may be associated with considerable morbidity [1] . aim. to investigate prospectively the incidence and outcomes of vat. we studied prospectively all patients who received mechanical ventilation in the general intensive care unit of a tertiary hospital in greece between september-november 2008. vat diagnosis required temperature ([38°c) or leukocyte count [12.000 per ml or leukopenia \4.000 per ml) (at least one of these) plus new onset/change of purulent endotracheal secretions. vap diagnosis required the aforementioned criteria plus appearance of new and persistent pulmonary infiltrates on chest radiography. microbiological documentation was based on the growth of microrganisms in bronchial aspirations ([100.000 cfu) or bal ([10.000 cfu) . results. forty-six patients were included, median (iqr) age was 57 (49. years. eleven (24%) patients presented vat, 11 presented vap and 24 patients presented none of these two disorders (np). there were no significant differences between vat and vap cases in terms of baseline characteristics (diagnosis, respiratory compliance, apachee, murray score), occurrence of sepsis or ards and microbiology; pseudomonas aeruginosa, acinetobacter baumannii, staphylococcus aureus and klebsiella pneum. were the most common bacteria in both vat and vap. patients who presented vat or vap had significant longer hospitalization and mechanical ventilation duration (days) compared to , 24 (14-33) vs. 6 (4-7), (p = 0.01)] and [18 (9-25) , 19 (10-26), 5 (3-6) , (p = 0.01), respectively]. icu mortality was 36, 9, 8%, for patients with vap, vat and np, respectively (p = 0.1). conclusions. incidence and microbiological pattern was similar in vap and vap in these case series. both vat and vap were associated with longer hospitalizations and mechanical ventilation duration. further analysis with a larger cohort of patients is required to give conclusive remarks. reference (s) . 1. nseir s et al (2002) nosocomial tracheobronchitis in mechanically ventilated patients: incidence, aetiology and outcome. eur respir j 20:1483-1489. g. c. choutas 1 , v. g. nolas 1 , a. kalantzi 1 , a. moutzouri 1 , g. k. anthopoulos 1 1 intensive care unit, \ \251[ [ general air force hospital, athens, greece introduction. endotracheal suctioning is an essential part of care for patients receiving mechanical ventilation, to keep the airways free from bronchial secretions, assuring ventilation and oxygenation. there are two types of suction systems. in the open system, endotracheal suctioning requires disconnecting the patient from the ventilator and introducing a single-use sterile suctioning catheter into the endotracheal tube. closed systems are changed every 24 and 72 h. to determine whether ventilated patients treated with css in an intensive care unit (icu) differ as to airway bacterial colonization and colonization of the suction system based on cultivation of both bronchial secretion and suction catheter tip and if cultivation of suction catheter tip is adequate in place of bronchial aspirate cultivation. methods. 164 patient, incubated and ventilated in the icu ward were studied in a period of one year (2007 to 2008) , on admission to the icu a css (trach care mac) was connected. closed multi-use catheters were changed daily. two-pass endotracheal suctioning was performed as needed. ba cultures were obtained on admission and the next day. radiographs taken before, during, and after ba and css cultures were graded for pneumonia and a modified score for vap. of the 164 patients css samples 104 (63.4%) and ba samples 99 (60.4%) were sterile. airway colonization with gram-negative bacteria and fungi occurred in the majority of the patient 24.1% and gram-positive bacteria in 20%. cultivation of css revealed gramnegative bacteria and fungi occurrence 31.5% and gram-positive bacteria in 11.5%. with the current sample no significant difference was found between the positive results of trach care tip cultivation and ba cultivation p = 0.568. objectives. to reduce the use of sedatives and to decrease the amount of time spend on a ventilator by specific ramsay-instructions and checks for sedation-protocol-adherence. methods. in april 2008, after introductional lessons to doctors and nurses, we started and collected data for 8 months. a yellow reminder was attached to the medical-instructions-form and doctors were requested to fill in the ramsay-score on a daily basis. once in a week patients and records were screened to assess protocol-adherence. each nurse and each intensive care-unit received feedback on their compliance to the bundle-elements. the total amount of sedatives per month was divided by the number of ventilator days, resulting in an average dose midazolam/propofol per ventilator day. the median and interquartile range of ventilator days/patient was also calculated and all data were compared with the same period in 2007. we accomplished a reduction in the use of sedatives and costs. introduction. ventilator associated pneumonia (vap) often occurs in patients who are mechanically ventilated. the incidence rate varies between 23 and 34% for patients in the intensive care unit. it has been the second most common hospital-associated infection after that of the urinary tract. the diagnosis of vap is difficult because of different existing definitions. hypothesis. our hypothesis was, that lowering vap incidence rate, could be done by a bundle of five interventions. the purpose of introducing multiple ventilatory interventions as a bundle, was to lower vap incidence rate by 10%. methods. during the last 6 months of 2007 all patients who were ventilated [48 h, were investigated for vap. the diagnosis of vap was done according to the criteria supposed by the dutch working group on infection prevention. a new infiltrate on chest x-ray after 48 h ventilatory support in combination with fever, leucocytosis, increased need for oxygen and culture of blind bronchial secretion c10 5 cfu/ml. a ventilator bundle was introduced on all icu wards as inspired by the institute of healthcare improvement. five interventions were introduced: head of bed [30°, reduction of sedatives as low as possible according to prescribed ramsay score, assessment of readiness to extubate, cuff-pressure measurement 3 times a day with application of cuff pressures between 25 and 30 cm h 2 o, and oral care with chlorhexidine 0.12% 2 times a day. icu nurses were trained in the first 6 months of 2008. the last 6 months of 2008 were used to evaluate the bundle intervention in comparison with the last 6 months of 2007. results. 63 patients were included in 2007 and 72 in 2008. after introduction of the ventilator bundle, the incidence per 1,000 ventilator days decreased from 23.3% to 9.6% per 1,000 ventilator days. introduction. continuous positive airway pressure (cpap) may improve oxygenation in patients with mild to moderate acute hypoxemic respiratory failure (ahrf) and avert further deterioration and need for intubation. objectives. aim of our study was to assess the physiologic effects produced by the addition of periodic hyperinflations (sigh) to cpap in patients with ahrf. we studied 10 patients with non-cardiogenic ahrf. four trials of one hour each were performed at a constant fio 2 60% during (1) spontaneous breathing (sb) via a venturi mask, (2) cpap 1 , (3) cpap ? 1 sigh/min of 25 cm h 2 o for 8 s (cpap sigh ), (4) cpap 2 . cpap, via helmet, was maintained at 10 cm h 2 o troughout the whole study period. pao 2 /fio 2 ratio (p/f), paco 2 , ph, respiratory rate (rr), arterial blood pressure (abp), heart rate (hr), dyspnea and patient comfort (by means of 2 separate visual analog scales) were measured at the end of each trial. results. overall, p/f was significantly (p \ 0.05) improved by cpap (cpap 1 192 ± 53 mmhg, cpap sigh 227 ± 56 mmhg, cpap 2 214 ± 75 mmhg), as opposed to sb (113 ± 9 mmhg). overall, the sigh did not significantly improved p/f. in the six patients with bilateral infiltrates, however, the rate of improvement in p/f significantly (p \ 0.05) augmented with the introduction of a sigh as compared with those with monolateral infiltration (150 vs. 107% respectively, being 100% the increase from venturi to cpap 1 ). paco 2 , ph, rr, hr, abp, dyspnea and comfort were not significantly different between trials. conclusions. in patients with ahrf, cpap improves oxygenation without affecting hemodynamics. the addition of a sigh to cpap further improved oxygenation only in patients with bilateral pulmonary infiltrates. background. adaptive support ventilation (asv) is a novel electronic ventilator protocol that incorporates the recent and sophisticated measurement tools and algorithms. the target tidal volume and respiratory rate are continually adapted to patient's respiratory physics and varying medical conditions. in injured lung, the asv should actively adjust ventilatory parameters achieving minimal work of breathing to meet the lung protective strategies. but there were little literatures describing its efficacy when applied to korean population. methods. from may 2008 to january 2009, we observed initial mechanical ventilation parameters in 114 patients receiving asv due to various causes (48 lung injuries including 27 community acquired pneumonia, 9 hospital acquired pneumonia, 5 interstitial lung diseases, 4 pulmonary tuberculosis and 3 idiopathic cases; 66 without lung injury which comprise 33 trauma cases, 18 strokes, 9 suicidal attempts and 6 other cases). the mean age of studied population was 57.5 years (male:female = 42:15). the data were collected within the first 12 h of mechanical ventilation. conclusion. as expected, adaptive support ventilation delivered smaller tidal volume and higher respiratory rates for injured lungs. asv efficiently operated in korean ali patients without any serious drawbacks and favorably adjusted the tidal volume and respiratory rates combination in relation with rcexp to meet lung protective strategies. introduction. neurally adjusted ventilatory assist (nava) is a mode of mechanical ventilation that uses the electrical activity of the diaphragm to control the ventilator obtaining an improved patient-ventilator synchrony and an efficient unloading of the respiratory muscles. nava is characterised by a variability of the breathing pattern and the absence of a constant flow, which makes impossible the determination of reliable data of respiratory mechanics by using the rapid occlusion method. we have previously demonstrated that the least squares fitting method (lsf) could be used during pressure support ventilation (psv), provided that the level of ps is sufficiently high to unload the inspiratory muscle. hence we made the hypothesis that (1) reliable data of respiratory mechanics can be obtained by applying the lsf method during nava and (2) the lsf method should work better during nava because of the characteristics of the flow and pressure traces. methods. ten patients undergoing mechanical ventilation for acute respiratory failure were enrolled. they were ventilated using randomly either psv or nava with the the same peepe and tidal volume (v t ). data of resistance (r rs ), elastance (e rs ) and total positive end expiratory pressure (peep tot ) were obtained by fitting the equation paw = r rs 9 v 0 ? v t / c rs ? peep tot during inspiration, because of the possible presence of expiratory flow limitation. the coefficient of determination (cd) of the applied equation was used to compare data obtained during nava and psv, the higher being the cd, the better the quality of the data. moreover patients were sedated and ventilated in volume controlled ventilation (acv) with the aim of calculating data based on rapid occlusion method and compare them with those obtained with lsf method by using the bland-altman analysis. (1) data obtained with lsf were statistically more reliable during nava (mean cd: 0.98 ± 0.01) than during psv (mean cd: 0.90 ± 0.1; p \ 0.001). (2) the cd obtained at every level of nava was always higher then 0.96. on the contrary, the cd obtained at low level of psv was less than 0.90 due to the presence of inspiratory muscle activity. (3) the bland-altman analysis demonstrated lower bias and higher precision between traditional data and those calculated during nava (bias: 2.8; limit of agreements: -2.4/ 8.9) compared to psv (bias: 5.2; limit of agreements: -17/6.5). conclusion. the application of the lsf method during nava allows calculation of reliable data of rrs, ers and peeptot, which are independent of the level of nava applied. this is of clinical relevance since psv allows calculation of reliable data only at high level of pressure support. it appears that the influence of inspiratory muscle activity on respiratory mechanics is less relevant during nava ventilation, suggesting a more physiological ventilation during nava both in terms of timing and of delivering adequate level of assist throughout each breath. introduction. nava is a new spontaneous-assisted ventilatory mode based on the detection of diaphragmatic electrical activity (eadi) and its feedback to adjust ventilator settings. nava uses the eadi, an expression of the respiratory center's activity, to initiate pressurization, set the level of pressure support and cycle the ventilator into exhalation. therefore, nava should theoretically allow near-perfect synchronization between the patient and the ventilator. however there are few data documenting these effects in intensive care patients. to determine whether nava can improve patient-ventilator synchrony compared to standard pressure support (ps) in intubated intensive care patients. comparative study of patient-ventilator interaction during ps with clinician determined ventilator settings and nava with nava gain (proportionality factor between eadi and the amount of delivered inspiratory pressure) set as to obtain the same peak airway pressure as the total pressure obtained in ps. a 20 min continuous recording with each ventilatory mode was performed allowing determination of trigger delay (t d ), patient neural inspiratory time (t in ), duration of pressurization by the ventilator (t iv ), excess duration of pressurization (t i excess = t iv -t in /t in 9 100) and number of asynchrony events by minute: non-triggering breaths, auto-triggering, double triggering, premature and delayed cycling. results are given in mean ± sd. p is considered significant if .05. 12.9 ± 19.6 2.2 ± 0.6 70.8 ± 37.8 n asynchrony/minute 7.6 ± 6.4 4.1 ± 3.7* 47.5 ± 17.0 respiratory rate (min -1 ) 16.8 ± 2.6 20.4 ± 4.7 na * p \ 0.05 conclusion. compared to standard ps, nava improves patient ventilator interaction by reducing td and the overall incidence of asynchrony events. there is also a strong trend in reducing delayed cycling. this ongoing trial should provide evidence that nava can indeed improve patient-ventilator synchrony in intubated patients undergoing ps. introduction. high fio 2 and hyperoxia may induce pulmonary injury and may increase oxidative stress. guidelines suggest a target arterial oxygen tension of 8 kpa [1] . a canadian questionnaire study found considerable variation in the attitudes, beliefs and practices of intensivists in the management of oxygen therapy [2] . however, the actual response of intensivists to hyperoxia in patients has never been studied. in this retrospective database study we investigated adherence to guidelines concerning oxygen therapy in a dutch academic intensive care. all arterial blood gas (abg) data from mechanically ventilated patients from 2005 to 2009 were drawn from an electronic storage database (metavision) of a mixed 32-bed icu in a university hospital in amsterdam. mechanical ventilation settings at the time of the abg as well as the successive abg were retrieved. the statistical analysis was carried out with spss 16.01. results. 126.778 abg's from 6251 mechanically ventilated patients were retrieved including corresponding ventilator settings. in 28.222 (22%) of the abg's po 2 was[16 kpa. initial ventilator settings and adjustments based on abg's of this group are shown in table 1 [data represent median (10th/90th percentile)]. in 88% of the lowest fio 2 group, fio 2 was exactly 40%. in only 25% of cases with po 2 [ 16 kpa the fio 2 was decreased. hyperoxia was accepted with no adjustments in ventilator settings if fio 2 was 40% or lower. introduction. major patient ventilator asynchronies are frequent during non-invasive ventilation (niv) especially due to leaks.niv can be delivered using icu ventilators or specifically designed niv ventilators. although icu ventilators are traditionally used for invasive mechanical ventilation, specific niv modes have been recently implemented. the impact of these niv modes as well as niv ventilators on patient ventilator asynchrony is unknown. our objective was to compare the incidence of patient ventilator asynchrony between invasive or niv mode of icu ventilators and niv ventilators. patients and methods. icu patients with acute respiratory failure requiring niv were studied during three randomized consecutive 20 min-periods of niv: icu ventilator with and without niv mode and niv ventilator. we used two icu ventilators: evita xl (dräger) and engström (general electrics) and 1 niv ventilator: bipap vision (respironics). flow and airway pressure were continuously recorded to determine breathing pattern. to detect major patient ventilator asynchrony we used surface diaphragmatic and/or sternocleidomastoid electromyogram allowing to assess neural patient's inspiratory time and to define asynchronies: ineffective triggering, auto-triggering, double-triggering, prolonged and short cycles. asynchrony was quantified using an asynchrony index as previously described. results. these preliminary results concern ten patients (9 males and 1 female) with a mean age of 71 ± 8 and a saps 2 of 52 ± 21. reason for niv was acute exacerbation of copd (n = 4), acute pulmonary edema (n = 2) and post-extubation (n = 4). at time of study, ph was 7.37 ± 0.08, paco 2 50 ± 16 mmhg and pao 2 80 ± 28 mmhg. ventilatory settings were set by the clinician and kept constant during the three periods with a ps level of 9 ± 2 cm h 2 o and a pressurization ramp of 110 ± 32 ms, a peep level of 5 ± 1 cm h 2 o and a fio 2 of 36 ± 14%. using icu ventilators, inspiratory trigger was 1 l/min and cycling off was 40% when adjustable. median asynchrony index was 5.9% (3.0-12.1) using icu ventilators versus 4.2% (1.8-6.6) using niv mode and 0.6% (0.4-1.0) using bipap vision (p = 0.20 between invasive and niv mode, p \ 0.01 between niv mode and bipap vision). asynchrony index was greater than 10% in three patients using invasive mode, two patients using niv mode and no patient using bipap vision. auto-triggering was the main asynchrony. conclusion. niv ventilator (bipap vision) allowed a marked reduction in patient ventilator asynchrony during niv as compared to niv mode currently available on new generation of icu ventilators. grant acknowledgement. this study was supported by a research grant from respironics. a. armaganidis 1 , k. stavrakaki-kallergi 1 , c. sotiropoulou 1 , a. koutsoukou 1 , j. milic-emili 1 , c. roussos 1 1 athens university, athens, greece prevalence of expiratory flow limitation (efl) was estimated using the negative expiratory pressure method (nep) in 96 anesthetized, paralyzed mechanically ventilated patients in icu. patients were studied in supine position at zero positive end-expiratory pressure (peep). a nep device especially designed and in build in an evita 2-draeger respirator, allowed the application of a pressure equal to-5 cm h 2 o, starting at 8 ms after the onset of expiratory flow and sustained throughout the end of the expiratory time set on the ventilator. patients were categorized in two groups: 1. non efl (47 patients without flow limitation), in whom nep elicited an increase of expiratory flow over the entire expiratory flow-volume (v 0 -v) curve. 2. efl (49 patients with efl), in whom part or the expiratory v 0 -v curve during nep was superimposed on the baseline v 0 -v curve. half of our patients (51%) were flow-limited. no patient without pulmonary disease was found flow-limited, except of a percentage of morbidly obese patients (57%). efl was recorded in 65% of icu patients with pulmonary diseases: 65 % of ards patients, 75% of patients with respiratory infection, 75% of asthmatics and 85% of patients with copd. time constant (s) and inspiratory flow (v 0 insp) were found to predict the severity of flowlimitation expressed as efl % v t . objective. tidal volume (v t ) administered to ards patients can be adjusted depending on body weight. body weight may be estimated, measured or calculated for an ideal or a predicted value based on different formulas [1, 2] . besides, those formulas require the measurement of height and may differ depending on gender. we hypothesized that v t value (ml/kg of body weight) may be different and show intrameasure variability depending on the method used. methods. ards patients were included prospectively in the first 72 h after icu admission. the ventilatory parameters were selected by the attending physicians that were foreign to the study. all patients were ventilated by volume controlled-assisted mode. five independent observers estimated the weight (estw) of each patient. they also measured height with a metric tape for calculate the predicted body weight (pbw) [1] and the ideal body weight (ibw) [2] . after previous measurements, patients were weighed once with a calibrated scale (scaw). results were compared using analysis of variance. results. 18 patients were studied, 45% women (age 52.5 ± 15.9, saps ii 49.6 ± 17.6; apache ii 23.4 ± 9.1). ventilation parameters at inclusion: v t /scaw (ml/kg) mean ± sd 5.6 ± 0.86 6.7 ± 1.05 5.9 ± 0.87 5.8 ± 0.93 0.001** mean ± sd mean ± standard deviation, min minimum, max, maximum, pbw 50 (men) or 45.5 (female) ? 0.91 9 (height in centimetres -152.4), ibw 25 9 (height in meters) 2 , mean diff average of the intraindividual differences of calculated/estimated weight, range intraindividual difference in weight (estimated/calculated) expressed as minimum and maximum * estw versus pbw p \ 0.001, pbw versus ibw p \ 0.001, pbw versus scaw p = 0.005 ** v t /estw versus v t /pbw p \ 0.001, v t /pbw versus v t /ibw p \ 0.001, v t /pbw versus v t /scaw p \ 0.002 conclusions. our data show that there is no gold standard method for estimate or calculate body weight to adjust tidal volume in ards patients. recommendations based on pwb and ibw not guarantee that tidal volume administered is really those that we want to administrate. reference (s) . 1. ardsnet. nejm 2000; 342:1301 -8. 2. stewart te et al (1998 3 hôpital raymond poincaré ap-hp, service de réanimation médicale, garches, france, 4 hôpital saint-louis, paris diderot university, paris, france, 5 centre hospitalier d'etampes, etampes, france, 6 université versailles saint quentin en yvelines, versailles, france rationale-objectives. dyspnea is a major respiratory symptom, which can reveal a severe disease. additionally, it can also result from an inappropriate ventilator setting in mechanically ventilated patients. if these patients are nowadays more and more conscious, prevalence of dyspnea and its clinical, biological and radiological correlates has never been assessed in this population. prospective cohort study conducted in two medical intensive care units (icu) during 6 months. all patients intubated more than 48 h and conscious have been included. the first day when the patient regained consciousness, dyspnea, anxiety and pain were assessed using a visual analogic scale (vas). if dyspnea was found, patient was asked if he experienced ''air hunger'', and/or ''excessive respiratory effort'' and if dyspnea vas was improved after ventilator setting has been changed. demographic, clinical, biological and chest x-ray data and ventilator settings have been collected. results. 96 patients were included (age: 61 ± 18 years; simplified acute physiology score ii (saps ii): 43 (iqr 31-60). reasons for mechanical ventilation included acute respiratory failure (n = 46, 48%), neuromuscular diseases (n = 29, 30%), coma (n = 10, 10%), and exacerbation of chronic obstructive pulmonary disease (n = 7, 7%). dyspnea was present in 45 (47%) patients and was qualified as ''air hunger'' in 31 patients (32%), ''excessive effort'' in 23 (24%) and both in 4 (5%). age, saps ii, reason for mechanical ventilation, respiratory rate, clinical examination, x-ray chest, pao 2 /fio 2 ratio, paco 2 were not statistically different between patients with and without dyspnea. anxiety .0); p \ 0.0001), assist controlled ventilation [4.77 (1.60-14.3) ] and diastolic blood pressure ; p = 0.038) were independently associated with dyspnea in multivariate analysis. ''air hunger'' tended to be associated with controlled ventilation (p = 0.051) whereas ''inspiratory excessive effort'' was significantly associated with low inspiratory flow, severe hypoxemia (median pao 2 /fio 2 ratio: 166, p = 0.044) and marked hypercapnia (median paco 2 : 48 mmhg, p = 0.019). in 35% of breathless patients, of ventilator resetting decreased dyspnea. length of icu stay was greater in patients with dyspnea (p = 0.017) whereas extubation within three days and icu mortality did not differ between the two groups. conclusions. dyspnea is frequent in mechanically ventilated patients and is strongly associated with anxiety, more frequently when controlled ventilation is used and is often reduced after ventilator resetting. assessment of dyspnea in conscious mechanically ventilated patients should be routinely performed in order to improve patients' comfort. methods. an experimental study design was used with a group of 66 first year health care provider students. the students were divided into two groups related to familiarity of the location of exam. a part of students (n = 35) were examined in demonstration room (dr) and the other part of students (n = 31) in public place (pp) . every student received the same number of training hours (28 h) and the same training method in demonstration room. during this exam the students performed a 2 min long, single person cpr related to erc 2005 guideline. their performance was measured with calibrated ambu cpr software and the adapted point system of brendan b. spooner's scale. v 2 and t test were used for comparison. p values less than 0.05 were considered statistically significant. we did not find difference between dr and pp groups in the correct sequence of bls steps, hand position, adequate frequency and depth of chest compression. between groups of characteristics of ventilation were not significant differences observed. it is first critical point in bls process to assess the quality of patients' spontaneous breathing; therefore it is crucial that the duration of check breathing may be sufficient long. the duration of checking for breathing was significantly (p = 0.001) shorter in dr groups than pr groups. in the pp groups time interval between 30 chest-compression cycles were significantly (p = 0.016) longer-more than 2 s-than in dr group. conclusions. the altered location of bls final exam shortens duration of checking for breathing which determines bls providers' decision making on starting chest compressions. the students may be full of confident in the well-known place represented by the shorter time of checking for breathing. the changed place of exam extended time interval between chestcompression cycles, therefore weaken the continuity of chest compressions, and decrease the chance of return of spontaneous circulation. aims. this paper reports an evaluation of the student experience of using a clinical competence assessment tool (ccat) in postgraduate critical care nursing education. the focus is on the perceptions of students in relation to the validity, reliability and usability of the assessment tool. the domains of competence assessed are based on five domains outlined by an bord altranais (2000) . they are: professional/ethical practice, interpersonal relationships, practical and technical skills, utilising a holistic approach to care, clinical decision making and critical thinking skills and organisation and management of care. the assessment process encompasses three clinical assessments and clinical competence is measured using the developmental process of novice, advanced beginner, and competent as described by benner (1984) . students are asked to reflect on their own learning needs prior to each assessment. the assessment includes a discussion on the knowledge that underpins practice thereby showing the integration of theoretical and practical knowledge. questionnaire was administered to all students who recently completed a graduate diploma in nursing studies (critical care) at a specific third level institution. results. the evaluation of the ccat as a mode of competence assessment in postgraduate critical care nursing education was generally positive from the students' perspective. some students considered the holistic nature of the ccat document to be a limitation, suggesting that their level of competency could have been better addressed with a tool that was more oriented toward critical care rather than being so 'broad' in nature. overall respondents considered that the ccat helped them to identify learning needs and found the use of the tool to be a positive experience and easy to use although some respondents considered that the wording of some of the sub-domains and indicators was difficult to interpret. competence assessment is about ensuring the delivery of safe and competent patient care. in order to determine competence a valid and reliable tool is needed. this small scale study presents the views of post registration critical care nursing students on using a competence assessment tool. the findings of this study cannot be generalised, however they do provide insight for educators and students using competence assessment tools in programmes preparing registered nurses for specialist nursing practice. the use of a holistic assessment process needs further explanation. students need to be encouraged to move away from the reductionist approach, which is focussed on tasks and move towards a broader understanding of competent practice. reference (s) . 1. an bord altranais (2000) requirements and standards for nurses registration education programmes, 2nd edn. 2. benner p (1984) from novice to expert excellence and power in clinical nursing practice. addison-wesley, california. to assess the usefulness of a web-based interactive learning package designed to supplement an undergraduate acute care course (very basic) taught to final year medical students. a web-based interactive learning package was developed to supplement a highly rated traditionally taught 8-day acute/critical care course consisting of pre-course reading, lectures, skill stations and interactive tutorials [1] . the additional web-based package consisted of narrated lectures, interactive lessons, videos and animations to demonstrate practical procedures and clinical signs, self assessment quizzes and a question and answer forum. topics covered included arterial blood gas sampling and interpretation, acute metabolic disturbances, non-traumatic coma, acute respiratory failure and sepsis. both the package and the course are available to other medical schools free of charge. usefulness of the package was assessed by examining activity logs, a student questionnaire, formal focus group (conducted by an investigator not involved in course preparation or teaching), comparing the results of a post-course mcq based summative assessment with historical controls and comparing results of formative assessment included in the package with the summative assessment. results. over 50,000 student-activities were logged by 135 students during the two week course. 107 students completed the questionnaire. with regard to usability, [60% agreed or strongly agreed that interactive lessons and self assessment ran smoothly without faults, with a corresponding score of [80% for narrated lectures and ease of browsing. with regard to usefulness, c80% agreed or strongly agreed that the question and answer forum was useful in clarifying areas of doubt and narrated lectures improved understanding of the course material; [90% agreed or strongly agreed that the content as a whole was useful in preparing the respondent to work as a doctor, interactive lessons improved their understanding of how to apply their knowledge, and their understanding of arterial blood gas interpretation and self assessment exercises improved their understanding of the course material. participants in the focus group indicated that the resources provided in the website were useful for learning, specifically the animations, narrated lectures and the question and answer forum. suggestions for improvement included improving the quality of the video and animations, increasing the range of topics covered and ensuring consistency with the printed course manual. there was no correlation between formative and summative assessments but, compared to 36 historical controls, performance in the summative assessment improved (75 vs. 67%, p \ 0.001). conclusions. the package provided a useful supplement to a traditionally delivered acute care course. introduction. faculty development refers to that broad range of activities that institutions use to renew or assist faculty in their roles. it includes activities that improve an individual's knowledge, skills and attitudes in important areas in teaching, education, research, leadership, administration and career development. in this abstract we will introduce one of the most important methods of faculty development programs. a meeting by the authors ''organizing group'' was conducted to decide on a topic for our workshop and discussed the planning and designing process. we decided on conducting a workshop on clinical teaching methods. a scientific and organizing committee was established, and accordingly work loads and assignments were distributed among them. we gave this workshop a title of ''i am the best clinical educator…are you!? our target audience was acute care management providers, with a capacity of up to 45 participants. we gave a specific time and location of this event. venue was arranged. computers for group work, audiovisual and other logistics were provided. after summarizing the main points for the workshop the organizing committee distributed an invitation letters throughout the higher management and educational leaderships. an address remark was done through invitation from the organizing committee. hot and cold beverages and break lunch meals were provided. posters on the workshop were distributed through out the institution. folders with educational materials were provided for each candidate. pre-course registration was done. once the program for the workshop was finalized a reminder was sent out to the participants on the date and venue for the workshop. participants attended on time, folders, badges with usbs were handed out. a questionnaire was distributed to the audience to estimate their learning experiences and approaches towards teaching styles and methods which were used in their practice. certificates of attendance with cme credit hours were distributed. results. 45 candidates attended this faculty development workshop. 70% were nurses and 30% were physicians, during this workshop, three topics were distributed over three groups, one group on how to break bad news. second group about how to conduct microteaching and the third group about how to give feedback. each group was evaluated by three members of the organizing committee, each group was ranked accordingly. all were performed by role play. at the end of the workshop an evaluation form was filled 100% responders. a five performance scale was used. the strength of the workshop was innovativity and ranked as strongly agree. the only weakness was the place constraint. conclusion. we concluded that a well organized workshop using role play, interactive sessions are effective modality for faculty professional development programs among acute care providers with high satisfaction rate. only 35 of the respondents (17%) indicated they did understand the statistics they encountered in journal articles and 90% felt it was important to understand these concepts and that they would like to access more easily to biostatistics training. a patient is referred to a higher centre when services are needed to maintain continuity of care.there are guidelines for the safe inter and intrahospital transport of critically ill patients but no guidelines are available for the minimal mandatory content of interhospital referral notes of critically ill patients.this problem is manifold in developing countries. objective. to educate the critical care physicians regarding the deficits in the physicians referral notes with which critically ill patients are referred from one centre to another. it is a prospective observational study on 96 out of hospital referred patients transferred to our intensive care unit (icu) over a period of 1 year. after permission from the institutes ethical committee we reviewed the referral summaries of these patients at the time of icu admission regarding the information available of clinical details, course in the previous hospital and therapeutic interventions. patients with more than 24 h of hospitalization before transfer were included in the study. introduction. in japan, closed icus have been gradually increasing at university hospitals. a closed icu is necessary for a university hospital not only for the hospital activity but also the education of medical students and the training of fellows. they can learn how to manage the circulation and respiration status of severely ill patients in icu. it is indispensable for effective education to ensure sufficient proper icu staffs. but the present condition of our country is that there are not so many intensivists enough to perform both of clinical duties and education of students and fellows. each icu of university hospitals is endeavoring to increase the number of intensivists. one of the popular methods is the announcement on web site to promote interest of young fellows. regrettably, the homepage of the japanese society of intensive care medicine has no such specific pages. each icu of university hospitals has to create attractive its own pages in the homepages of the hospitals. [1] . most subjects are taught using lectures and group tutorials and the theory is applied in clinical areas to facilitate greater understanding of the newly acquired knowledge. there is no reported best practice mechanism for teaching medical ethics in a practical setting to medical students. objectives. the routine use of an ethical checklist has been proposed as a tool for the medical team to consider ethical issues on critical care [2] . its use as a tool for teaching medical ethics within critical care has not previously been reported. the aim was to use this checklist to facilitate learning providing clinical case material for discussion in daily tutorials. one medical student (sm) undertook a one week period of study to learn about ethics in critical care practice. the checklist was used to review patient notes, guide further discussion with patients, when observing the professional behaviours and communication of the multidisciplinary team, and as a guide for case based discussions. results. the complexity and severity of patient conditions in critical care makes it the ideal setting for learning about ethics. sm considered more ethical dilemmas in this practical attachment than in the previous 2 years of clinical placements. the checklist allowed identification of possible ethical issues relating to each patient and a deeper understanding of the patient's health care needs. it was used for daily tutorials to discuss the ethical principles and observed professional behaviours in a similar way to a discussion of clinical diagnosis and management of a patient case with a supervising doctor on a normal clinical attachment. complex issues such as capacity to consent, end of life treatments or resource allocation were seen in relation to ongoing care. on ward rounds it was observed that their conduct in an open environment could at times potentially compromise patient confidentiality. there was also a benefit from the consideration of ethics issues in a real time basis which allowed exploration and reflection on personal moral or spiritual beliefs and how they may differ from those of the patients and other medical professionals. conclusions. using an ethical checklist allowed application of theoretical lecture and workshop material to real life situations. by discussing the cases and observed behaviours with a senior critical care doctor it is possible for trainee staff to appreciate how difficult medical management decisions are made, and to improve the acquisition of the skills necessary to start to assess and discuss ethical issues surrounding a patient's care confidently. introduction. accurate data on patient's weight and height are important for management in intensive care units (icus). unfortunately, weight beds or bed scales are not available in a significant number of icus and these variables are often estimated by health care personnel. the accuracy of such estimations is poorly described. objective. to investigate the accuracy of visual estimation of weight and height in critically ill patients. methods. prospective study conducted in a 10-bed mixed medical and surgical icu. patients were consecutively weighed by an unblinded physician with a stretcher scale (t3 metric), and measured by a physical therapist using a measuring tape. the ideal weight was calculated using the ardsnet's formulas for predicted body weight. medical staff (ms), internal medicine resident (imr), nursing staff (ns), physiotherapist (pt) and nutritionist (nu) were asked to estimate patient actual weight, ideal weight and height. they were blind to the estimations during all the protocol. estimations in each healthcare group were computed as means, medians and percentage of error from actual and ideal weight and height, respectively. anova test was used to compare mean estimations between the groups. there were no significant differences between the groups in estimation of either weight (p = 0.73) or height (p = 0.68). conclusion. weight estimations from healthcare personnel are often inaccurate. there are no significant differences in accuracy between the estimations of weight and height in different healthcare groups. an effort should be made to weigh all critically ill patients. intraabdominal hypertension (iah) is often diagnosed in icu and it can lead to abdominal compartment syndrome, multiple organ failure and death [1] . in clinical setting biochemical signals based on which iah is considered severe or detrimental on visceral tissues are scarce. currently, the only clinically relevant signal is decreasing hourly diuresis. in an attempt to find an early sign of metabolically relevant signal on clinically marked iah we investigated abdominal wall metabolite concentrations. previously high lactate/pyruvate has been detected in dialysate from rectus abdominis muscle (ram) in animal models of iah [3] . in the present experiment we hypothesized that laparoscopic surgery which induces iah could lead to clinically significant increase of l/p ratio as a signal of anaerobic metabolism caused by iah and insufficient tissue perfusion. introduction. among the techniques proposed to assess microperfusion and oxygenation, nirs sounds to be convenient [1] . if baseline measurements do not provide useful information for outcome of micro-vascular impairment, functional evaluation using vascular occlusion test (vot) seems to be promising [1] . technological development of the nirs device (inspectra models 325 and 650, hutchinson technology, hutchinson, minn) proposes to use a new probe measuring hemoglobin saturation at less depth than previously (15 vs. 25 mm between fiberoptic) with more data output (1 value/2 s vs. 1 value/3.5 s) associated with an automated software to compute occlusion and reperfusion slopes. objective. to compare nirs results obtained, using the two different probes, at day 0 of septic shock (ss) in two groups of patients having similar clinical characteristics. methods. 43 patients (g1) and 30 patients (g2) were included within the first 24 h of ss. macrohemodynamic: heart rate (hr), mean arterial pressure (map), central venous pressure (cvp), cardiac output (co) and svo 2 (mixed venous o 2 saturation), ph, base excess, and lactate were collected as saps ii and sofa scores. baseline sto 2 at thenar eminence was continuously monitored and a 3 min upper arm(brachial artery) vot was performed. sto 2 occlusion and reperfusion slopes were calculated manually in g1 (probe 25 mm) using linear adjustment (r 2 c 0.90 to be valid) or calculated by the software in g2 (probe 15 mm) using the same method, p \ 0.05 was considered significant. results. median ± iqr. the two groups did not differ for macrohemodynamic nor for metabolic data (table 1) . nirs data surprisingly were largely significantly different between the two groups for both baseline and slopes ( background. hypovolemia and hypovolemic shock are life-threatening conditions that occur in numerous clinical scenarios. near-infrared spectroscopy (nirs) has been widely explored, successfully and unsuccessfully, in attempt to function as an early detector of hypovolemia by measuring tissue oxygen saturation (sto 2 ). in order to investigate the measurement site-and probe-dependence of nirs in response to hemodynamic changes, such as hypovolemia, we applied a simple cardiovascular challenge; a posture change from supine to upright, causing a decrease in stroke volume (as in hypovolemia) and a heart rate increase in combination with peripheral vasoconstriction to maintain adequate blood pressure. methods. multi-depth nirs was used in nine healthy volunteers to assess changes in peripheral vascular tone in the thenar and forearm in response to the hemodynamic changes associated with a posture change from supine to upright. a posture change from supine to upright resulted in a significant increase (***) in heart rate. thenar sto 2 did not respond to the hemodynamic changes following the posture change, whereas forearm sto 2 did. in the forearm, sto 2 was significantly lower (***) in the upright position with respect to the supine position. conclusion. the primary findings in this study were that (1) forearm sto 2 is a more sensitive parameter to hemodynamic changes than thenar sto 2 and (2) cerebral hyperperfusion syndrome, caused by inflow at normal blood pressure into maximally dilated fine vessels, is a recognized complication of carotid endarterectomy (cea) strict blood pressure control in the early postoperative period can minimize the risk of cerebral hyperperfusion. until yet, diagnosis of cerebral hyperperfusion mainly relies on intermittent postoperative examinations (spect; ct angiography). non-invasive absolute cerebral oxygen saturation (scto 2 by fore-sight technology) was validated to jugular bulb saturation (sjo 2 ) monitoring with a constant difference of 10% higher for scto 2 values. previously, sjo 2 monitoring after severe head injury indicated cerebral hyperemia. in this study, we evaluated scto 2 monitoring after carotid surgery as possible continuous on-line monitoring of cerebral hyperperfusion. fourteen pts scheduled for cea were monitored for 12 h postoperatively after cea. bilateral scto 2 monitoring was started before induction of anesthesia and maintained until 12 h postoperatively. intra-operative eeg monitoring guided the decision to intraluminal shunt insertion. strict blood pressure control was applied at maintaining normotensive levels throughout the clamping procedure. early postoperative care focussed on strict maintenance of normotensive blood pressure. in no pt, any change in eeg was observed after carotid clamping. in all pts, ipsilateral scto 2 significantly decreased after carotid clamping, without any scto 2 value below 55%. we observed no changes in contralateral scto 2 . mean clamping time was 28 min (19-37 min). in all pts, clamp release restored ipsilateral scto 2 to baseline values. in all pts, emergence from anesthesia was uneventful, without any new neurological deficit. in 6 of 14 pts, significant increases (scto 2 [ 85%) in ipsilateral scto 2 were observed in the postoperative period (m scto 2 87.5%), without any changes in contralateral scto 2 . this increase occurred at a mean of 3.4 h after carotid declamping with a mean duration 5.3 h. in these 14 pts, we could not make any significant correlation to arterial blood pressure, as none of these 6 pts needed more aggressive antihypertensive control. we noted that 4 of these 6 pts suffered from diabetes mellitus, while 5 of 6 pts revealed high ([90%) contralateral stenosis. further data will have to reveal the importance of these comorbide factors. non-invasive cerebral oximetry, enabling absolute cerebral oxygen saturation monitoring, could provide on-line estimation of cerebral perfusion state after cea. this could allow bedside detection (and eventual therapeutic interventions) of cerebral hyperperfusion after cea. introduction. analysis of microcirculatory alterations obtained by side-stream dark field (sdf) is time consuming. automated analysis with modern softwares could accelerate this process and help to quantify blood flow velocity. however, perfusion detection is based on the contrast between pixels and this may be influenced by image settings. objective. we aimed to compare data obtained with a new software to the traditional semi-quantitative analysis of sdf images. methods. we selected from our database six images of poor sublingual microcirculatory perfusion and six images of good microcirculatory perfusion registered by the sdf technique (microscan; microvision medical, amsterdam, the netherlands). the proportion of perfused vessels [ppv = (number of vessels with continuous flow/number of all vessels) 9 100] \ 70% was used to define microcirculatory perfusion. total vessel density (tvd) was determined automatically by the software ava 3.0 (microvision medical) and also by the semi-quantitative technique, considered as the gold-standard (number of capillary crossing three equidistant vertical and horizontal lines divided by the total length of these lines). ava software was also used as default definitions or set to optimize analyses according to manufacturer instructions. vessels falsely detected (false positive = fp) or missed (false negative = fn) by the software, in comparison to the semi-quantitative evaluation, were also counted. results. tvd was significantly higher by the ava software either on default or on optimized mode than by the semi-quantitative method, and these differences were present with good or poor perfusion images (table 1) . overall fp rate was 22%, and it was greater in poor perfusion images (40%). optimization of the ava set parameters attenuated fp rates both in poor and good perfusion images, at the expense of increasing fn rates (table) . due to intrinsic characteristics of the software, the mean total grid length was significantly lower in the ava than in the semi-quantitative analysis (3.8 vs. 5.6 introduction. perfusion index (pi) is the proportion of constant absorbed light compared to pulsatile absorbed light emitted from a pulse oxymeter. it ranges from a value below 1 up to 20 depedant of peripheral perfusion. it is measured primarily to evaluate the signal quality for the pulse oxymeter and is displayed by some pulse oxymeters to be acknowledged by the clinician. the pi changes with vasodilation and vasoconstriction. however, intubation is a stimulus able to increase endogenous catecholamines and thus leading to vasoconstriction possibly declining the perfusion index. therefore we found intubation with a double lume tube in a thoracic surgery setting as a suitable setting to evaluate changes in perfusion index as a reaction to intubation. after informed consent, we enrolled seven patients undergoing lung surgery requiring an double lume tube. they were monitored as it is standard of care in our institution with invasive blood pressure, ecg, and a pulse oxymeter displaying the pi. (radical 7, masimo, irvine, ca) the patients received the medication to induce anesthesia calculated adequately to their body weight. midazolam, propofol and fentanyl where used to anesthetize the patient, cisatracurium was used for muscle relaxation to facilitate intubation. pi, pulse and arterial saturation were recorded every minute from prior to induction until after successful intubation. a baseline value was recorded prior to induction and compared to the value 3 minutes after induction. then the pi measured next to intubation was compared to the pi after induction and analysed using students t test. introduction. anticoagulation strategies for albumin dialysis suppose a difficult compromise between risk for bleeding and a high tendency to clot in the circuit. even thought the sessions are short, a premature clotting is a serious event because the lost of blood (high priming volume) and a high cost of the systems. we intended to demonstrate that the classical approach based in heparin is not adequate in these patients and should be substituted for a different strategy (mixed low dose of heparin plus epoprostenol). methods. data of a prospective registry of all cases treated in our centre (a third level, teaching hospital) with albumin dialysis (mars system). initially we used non-fractionated heparin at 5-7 u/(kg h) in patients without coagulation problems, epoprostenol [4-6 ng/ (kg min)] in cases with risk or thrombocytopenia and no anticoagulation when high risk for bleeding or contraindication for anticoagulation. after an intermediate analysis of our registry we detected a high number of filters clotted when heparin was used and changed our approach to use as first indication a mixed protocol with non-fractionated heparin [3 u/(kg h) ] plus epoprostenol [4 ng/(kg min) ]. data are presented as percentages. analysis was performed with chi-square test. to detect variables related to coagulation a stepwise backward logistic regression analysis was performed. we registered 72 patients with a total of 216 sessions. selecting only the first session for each patient to validate the first choice for anticoagulation, we used heparin in 26 cases and detected the loss of 11 filters (42.3%) because clotting. after the change to mixed anticoagulation we used this as first indication in 17 patients and in only 4 (23.5%) the sessions were prematurely ended because clotting (p ns). the rest of patients received isolated epoprostenol in 19 cases (with 4-21%-cases of premature clotting) and no anticoagulant in five cases (with 3-60%-premature clotting). between the 26 cases with heparin as first choice, three episodes of mild and one episode of severe bleeding were detected while no patients in the mixed group presented bleeding complications (p ns). in a logistic regression analysis over all registered sessions using coagulation of filters as dependent variable and type of patient, anticoagulant, arterial pressure, inr, tpta, platelets, haematocrit or bilirubin as independent variables, none of these was included in the regression model. even though more studies are necessary to validate this conclusion, a mixed protocol based in low dose heparin plus epoprostenol could be adequate as first indication for non-complicated patients submitted to a mars treatment with lower risk for bleeding than the classical approach of isolated non-fractionated heparin. optimizing oxygen delivery in critically ill patients is vital for the promotion of aerobic cellular metabolism. current practice includes the measurement of variables such as partial pressure of arterial oxygenation (pao 2 ), cardiac index (ci) and percentage of oxygenated haemoglobin in arterial blood (sao 2 ). these parameters reflect global oxygen delivery. the real point of interest is the end point of the oxygen cascade; oxygen utilisation in tissue mitochondria. near infrared spectroscopy (nirs) has been developed in an attempt to measure tissue oxygen saturation (sto 2 ) in peripheral muscle microcirculation. manufacturers state normal values as 87 ± 2%. it uses four wavelengths near the infrared spectrum (680-800 nm) to measure sto 2 , a ratio of oxygenated haemoglobin to total haemoglobin. it is continuous and non-invasive. sto 2 has proven efficacious in predicting oxygen delivery in trauma patients and claims to have been successfully used to guide early resuscitation [1] . objectives. we were interested in assessing whether sto 2 had a role in measurement of oxygen delivery in the intensive care population, and how it compared to the parameters currently used to predict oxygen delivery. we had particular interest in the usefulness of nirs in septic patients, where the pathophysiology of tissue oxygen utilization is disrupted. patients from a general, adult intensive care unit were enrolled over an 8 month period. all patients had lidco monitoring. exclusion criteria were gtn, atrial fibrillation and patient refusal. 15 mm sto 2 probes were sited on the thenar eminence. serial recordings of sto 2 , cardiac index, hr, sao 2 , map, and pao 2 were recorded. sto 2 results were compared to more traditional parameters of oxygen delivery. sixteen patients were recruited, all met criteria for sirs and shock. four were excluded with incomplete data. results were analysed for individual patients and as a collective series. we found: • no statistical correlation between nirs and sao 2 or pao 2 . • a weak and clinically insignificant correlation between cardiac index and nirs (p \ 0.001). • supra normal nirs readings (normal [ 75%) were not infrequently gained in patients where all other parameters were indicating severe shock and poor oxygen delivery. conclusion. theoretically nirs has potential to be beneficial in measuring oxygen delivery. our results demonstrate that nirs is not accurate for our septic population. we found poor correlation with current methods used to predict oxygen delivery and it may well be more misleading than beneficial. more traditional methods of intensive care monitoring, although sometimes invasive, appear to provide a more accurate representation of a patient's oxygen delivery. background. urine output is a crucial parameter of renal function and fluid balance. conservative urine output monitoring harbors problems such as subjective reading, sampling time errors and nursing workload. an electronic urine collection device was introduced into the icu and connected to a computerized information system. this created a more reliable and accurate means for urine output monitoring and the ability to develop new calculated parameters. 1. to evaluate the effects of introducing an electronical urine collection device into a fully computerized icu. 2. to evaluate new parameters that were created by the combination of the device and a computerized data management system. patients included were all admitted to the icu at rambam medical center, haifa, israel, during the years 2007-2008. urine production and flow were monitored continuously by the urinfo2000 ò device (med-dynamix, israel), a novel electronic urinometer, connected to a patient data management system (imdsoft, israel). graphical analysis of urine production was done and derived parameters continuously calculated. comparison was done to the conventional mechanical urine collection system. variables studied were: measurement accuracy, sampling time accuracy, nursing workload before and after the implementation process. correlation between derived parameters and conventional renal function measurements such as plasma creatinine and creatinine clearance time. results. the conservative urine output measuring system demonstrated percentage error span in range of 68-100%, compared to a range of 25-32% percentage error in measurement after implementation of the computerized system. before implementation, sampling time error span was found to be 12-59 min, while no sampling time error was present after implementation due to the automated recording system. time consumed by the workload of the conservative urine output monitoring system was measured at 32-64 min per nursing shift (8 h). the computerized system eliminated this workload completely. derived parameters evaluated were continuous urine flow (in cc/min or cc/h), urine production acceleration rate (calculated via the slope of the ''up-rise'' in cc/min 2 ) and the peak urine production rate (cc/min). these parameters were able to demonstrate immediate changes in renal function, hours before conventional measurements and calculations would show them. conclusion. implementation of a computerized urine monitoring system can lead to improved accuracy in renal function monitoring and eliminate a significant amount nursing workload. use of derived calculated parameters may lead to earlier detection of renal malfunction and thus lead to earlier intervention. (40 g/1,000 ml), cica dialysate k2 tm (na 133 mmol/l, k 2.0 mmol/l, mg 0.75 mmol/l, cl 116.5 mmol/l, hco3 20 mmol/l, glucose anhydrous 1.0 g/l, ph * 7,4) and calcium chloride mayrhofer tm cacl 2 0,5 mol/l. the filter was an ultraflux av 1000s tm , the material of the bloodline tubing system was medical grade soft pvc. in three circuits used in two different patients we found an opaque white precipitation starting at the cacl 2 side port growing along the line with the direction of the bloodflow up to a maximal 2 mm wide and 80 mm long stripe. to identify the composition of the white stripes we included histological examination of hematoxylin-eosin stained sections and lyophilisation with wet chemical analysis. blood samples were simultaneously taken from the venous port of the cvvhd circuit and the arterial line of one patient. results. histology showed **an organic material in form of calcific deposit, covered with coagulated blood. chemical analysis identified this deposit as calcium phosphate. the results of the blood samples are shown in table 1 . calcium phosphate precipitates may have reached patient circulation and been deposited in the capillary bed of the lungs or other organs. no histological examinations of tissue were taken and adverse events could not be attributed to the described phenomenon. citrate anticoagulation was stopped and switched to combined heparin-epoprostenol sodium anticoagulation. conclusions. the combination of the fluids and materials used in this specific cvvhd circuit with citrate anticoagulation resulted in some patients in a detectable calcium phosphate formation in the circuit. physicians using the described setting should be aware of the phenomenon and stop citrate anticoagulation as soon as a deposit occurs. in vitro studies, using different compositions and concentrations of dialysate and substitution fluids and simulating different patient conditions (ph, ph, hb, alb,…) should clarify, which solutions could safely be used. in addition the material of the circuit should be investigated, since surface characteristics have been identified to influence the formation of a calcium phosphate layer [1] . reference (s) objective. the aim of this study was to assess, in a medical population of critically ill patients, whether intraabdominal pressure at admission was an independent predictor for mortality and to evaluate the effects of intraabdominal hypertension on organ functions. all patients admitted to the medical icu of the hgu gregorio marañón over a period of 45 days were studied prospectively. patients who fulfilled two or more risk factors for wsacs (diminished abdominal wall compliance, increased intra-luminal contents, increased abdominal contents and/or capillary leak /fluid resuscitation.) were included. iap was measured via a foley bladder catheter, according to the modified kron technique. data recorded on admission were the patient demographics with, acute physiology and chronic health evaluation ii score (apache ii), and type of admission; during intensive care stay, sepsis-related organ failure assessment score (sofa) and clinical concomitant factors and conditions. intraabdominal pressure were measured at least daily together with fluid balance. patients were followed throughout their hospital stay. forty-four patients were included in the study (age 58 -17, apache ii 20. . half were admitted for cardiopulmonary disease. twelve (27%) had pancreatic or gastrointestinal disease. twenty-two (50%) had severe sepsis or septic shock. the incidence of iah was 80%. mortality was 50%. the cause of the iah was capillary leak syndrome/fluid resuscitation in 51% of cases. there was no relationship between the presence of iah and the number of organ failure during admission. the only variables associated with mortality of the patients were sofa and apache ii. the presence of iah was not a factor associated with increased mortality, although these results may be confounded by sample size. conclusions. there is an unusually high incidence of iah in the population of critically ill medical patients with two or more medical risk factors for wsacs. however, unlike in other populations, our study does not demonstrate that the iap monitoring allow detecting a group at higher risk of developing multi-organ failure or death. background. drainage of ascitic fluid is a common practice in order to relief the respiratory discomfort of patients. the aim of the present study was to determine abdominal compliance after ascitic fluid removal by transcutaneous drainage. methods. twelve patients presenting with ascitic fluid were included. all patients had transcutaneous blind drainage with a wide catheter. the ascitic fluid removed was recorded, while the intraabdominal pressure (iap) was measured as proposed by wsacs. iap was measured before and 15 min after the puncture. abdominal compliance (cabd) was calculated. results. the pre-drainage iap was 10.33 mmhg (ranging from 8.82 to 13.24 mmhg, sd 2.02 mmhg), while the post-drainage was 5.90 mmhg (ranging from 5.15 to 8.82 mmhg, sd 1.32 mmhg). the mean volume of ascitic fluid removed was 1774 ml (ranging from 430 to 3,000 ml, sd 977 ml). cabd after drainage was 307 ml/mmhg (ranging from 67 to 333 ml/ mmhg, sd 144 ml/mmhg). a linear correlation was found between ascitic fluid removal and iap variations. conclusion. the drainage of ascitic fluid reduces iap, facilitating in this way respiration. moreover, iap variation seems be in linear relation with the volume of ascitic fluid removed. this linear relation between iap and volume may probably predict the cabd quite accurately and vice versa. however, larger studies are necessaries in order to safely draw predicting diap-dv (cabd) diagrams, and determine the optimal ascitic fluid removal in order to achieve best comforting of the patient and slower fluid reformation. introduction. use of stroke volume variation (svv) to guide fluid therapy in preload responsive state has been studied well in patients undergoing cardiac or neurosurgery during anaesthesia. use of this dynamic monitoring variable has not been studied much in septic shock. we undertook this prospective study to evaluate utility of svv to optimize preload in patients with septic shock and ards. setting. 40 bedded medical surgical icu of a 350 bedded tertiary care centre in pune, india. inclusion criteria: (1) patients with ards (po 2 /fio 2 b 200), svv readings were taken every 3 h with flotrac-vigileo system after confirming abolishment of spontaneous breaths by sedation or paralysis and increasing tidal volume transiently to 8 ml/kg. fluid boluses were given to keep svv \ 13% for 24 h after enrollment. attempts were made to reduce vasopressor doses keeping map c 70 mmhg. results. 20 patients with average age 57.9 ± 16.55 years and apache ii score 22.2 ± 5.14 were studied. each patient received an average 6.22 ± 2.54 l fluid in 24 h after enrollment to keep svv below 13%. svv at 24 h after enrollment was 10.1 ± 6.13% improvement in microcirculation was evident as plasma lactate reduced from 5.43 ± 3.64 (at 0 h) to 3.09 ± 2.78 mmol/l (at 24 h) there was no worsening in pulmonary edema as po 2 / fio 2 increased from 165.3 ± 106.1 (at 0 h) to 224 ± 100.66 (at 24 h) only 6 out of 20 patients needed renal replacement therapy. in 13 patients, vasopressors could be stopped completely in 49.7 ± 19.02 h. 12 of them survived till discharge from the icu and 1 died of ards. in 7 patients, vasopressors could not be weaned off completely and all of them succumbed. overall survival rate was 60%. conclusion. svv guided fluid therapy is a promising modality for pre load optimization in mechanically ventilated patients with septic shock and ards. introduction. cardiovascular function is an important determinant of outcome in sepsis, and heart rate (hr) has been associated with cardiovascular risk and mortality in large patient cohorts [1] . to investigate the association between hr and 28 or 90 day mortality in septic shock. methods. this study is a post hoc analysis of 358 septic shock patients who were included in the control group of a multicenter trial [2] . demographic and clinical data, average hr and catecholamine requirements during septic shock, occurrence of acute circulatory failure, 28 and 90 day mortality were documented. a binary logistic regression model adjusted for the simplified acute physiology score ii (excluding hr) was used to investigate the association between mean hr and acute circulatory failure or 28/90 day mortality. a multiple logistic regression model was applied to identify independent risk factors for developing hr critical for outcome. conclusions. hr is associated with 28 and 90 days mortality in septic shock. hr persistently exceeding 110 bpm during septic shock seems associated with a significant risk of death. introduction. different colloids can be used for treatment of hypovolaemia in septic pts. recently, small-volume resuscitation was introduced for initial therapy of severe hypovolaemia and shock. the concept of small-volume resuscitation encompasses the rapid infusion of a small dose of 7.2% nacl/colloid solution [1] . however, in septic pts hypovolaemia often associates with acute lung injury (ali). therefore in these pts great importance has influence of colloids on oxygen transport. objectives. the aim of the study was to evaluate and compare the effects of hhes and hes on oxygen transport in pts with sepsis and ss. methods. 34 hypovolaemic pts with sepsis and ss were enrolled in the study. 17 pts received 3-5 ml/kg (250 ml) hhes (7.2% nacl ? 6% hes) (fresenius kabi) within 45 min and 17 pts received hes 130/04 (voluven, fresenius kabi) 15 ml/kg. in all pts before and after infusion the parameters of oxygen transport was measured by pulmonary arterial catheter and transpulmonary thermodilution (pulsion medical system). after infusion of hhes oxygen delivery index (ido 2 ) increased because of increase of cardiac index (ci) despite of decrease of hemoglobin (hb) levels and absence of changes of arterial oxygen content. extravascular lung water (evlw) and shunt increased significantly immediately after hhes infusion, but this increase was not accompanied by deterioration of pao 2 /fio 2 . introduction. severe sepsis is characterised by a wide array of haemodynamic changes including increased capillary leak, vasodilatation, vascular hyporeactivity and myocardial depression. the resultant tissue hypoperfusion is an important catalyst of multi-organ failure [1] . to further develop our understanding of the underlying mechanisms, we have developed and characterised a fluid-resuscitated mouse model of intraperitoneal polymicrobial sepsis. objectives. to assess alterations in cardiac performance in mice at 0, 3, 6 and 24 h following faecal peritonitis. methods. sepsis was induced in 25 week old male mice (n = 25) by intraperitoneal (i/p) injection of dilute faecal slurry. sham animals (n = 10) received n-saline i/p. animals were fluid resuscitated at time 0 (30 ml/kg 0.9% saline), and at 6 and 18 h (50 ml/kg 0.8% saline-5% dextrose each time). under a minimum concentration of isoflurane to achieve light anaesthesia, peak velocity, stroke distance, heart rate and fractional shortening were measured in the short axis plane by echocardiography at the 0, 6 and 24 h timepoints. in separate sham and severe septic mice (n = 6 per group) the cardiac response to intravenous colloid boluses was assessed at 6 and 24 h. results. we clinically characterised septic animals into 'mild' and 'severe'. mice with severe sepsis showed a 65% drop in peak velocity and cardiac output at 6 h (vs. 30 and 14% falls in the mild septic and sham-operated animals, respectively, p \ 0.05). while mild septic animals showed recovery by 24 hr, cardiac output in severely ill mice remained significantly depressed (due to both low heart rate and stroke volume) compared to mild septic and sham animals [*p \ 0.05 ( fig. 1) ]. stepwise 0.5 ml boluses of intravenous fluid at 6 h in severe septic animals led to restoration of cardiac output to baseline (0 h) values. however, in the 24 h septic animals, fluid challenge produced an initial improvement in cardiac output followed by deterioration [ fig. 2 purpose. myocardial dysfunction has been well-documented in sepsis even in hyperdynamic state, and may develop and contribute to morbidity and mortality. nicaraven, a radical scavenger, has been shown to protect the coronary endothelial and myocardial function from ischemia and reperfusion injury due to hydroxyl radical scavenging activity. the purposes of present study were to determine the effects of nicaraven on cardiac function and cytokine production in lipopolysaccharide (lps) induced sepsis. methods. this protocol was approved by our institutional committee. following arterial and venous cannulation and tracheostomy, rats (330-350 g) were anesthetized with pentobarbital, and mechanically ventilated with a control mode (v t = 12 ml/kg, rr = 40 rpm). after baseline measurements, rats (n = 20) were administrated with lps (10 mg/kg, intravenously) and randomly assigned to following two groups: the nicaraven group treated with nicaraven [3 mg/(kg min), intravenously] and the control group treated with saline. the left ventricular pressure and volume were measured with the pressure and conductance catheter every one hour. cardiac function, including cardiac output (co), ejection fraction (ef), and maximal elastance of left ventricle (e max ) were analyzed with a computer soft. blood was collected, centrifuged (2,000 g, 15 min, 92) , and stored (-80°c) from rats every 2 h after operation to measure plasma concentration of tnf-a, il1-b and macrophage migration inhibitory factor (mif) using enzyme-linked immunosorbent assays kits. blood lactate concentration was also measured. data were analyzed by repeated measure anova. results. the co in the nicaraven group was kept significantly higher than the control group (p \ 0.05). the ef and e max in the nicaraven group were also kept significantly higher than the control group (p \ 0.05). arterial lactate, tnf-a, il1-b and mif were significantly lower in the nicaraven group versus the control group (p \ 0.05). conclusion. the current study indicates that the treatment with nicaraven improved cardiac dysfunction and reduced plasma concentration of cytokines, and improved lactic acidosis in septic model. methods. this protocol was approved by our institutional committee. following arterial and venous cannulation and tracheostomy, rats (330-350 g) were anesthetized with pentobarbital, and mechanically ventilated with a control mode (v t = 12 ml/kg, rr = 40 rpm). after baseline measurements, rats (n = 20) were administrated with lps (10 mg/kg, intravenously) and randomly assigned to following two groups: the oxytocin group treated with oxytocin (15 iu/kg iv and followed by the continuous infusion of 5 mg/(kg min), intravenously) and the control group treated with saline. the left ventricular pressure and volume were measured with the pressure and conductance catheter every 1 h. cardiac function, including cardiac output (co), left ventricular peak pressure (lvpp), and cardiac work (cw) were analyzed with a computer soft. blood was collected from rats every 2 h after operation to measure plasma concentration of blood lactate. data were analyzed by repeated measure anova. results. the co in the oxytocin group was kept higher than the control group but there is no significance (p \ 0.15). the lvpp and cw in the oxytocin group were kept significantly higher than the control group (p \ 0.05). arterial lactate was significantly lower in the oxytocin group versus the control group (p \ 0.05). conclusion. the present study indicates that the treatment with oxytocin improved cardiac dysfunction and reduced plasma concentration of lactate in septic model. introduction. conventional hemodynamic monitoring parameters like heart rate, mean arterial pressure (map), and central venous pressure may be misleading in assessment of circulating blood volume in severely septic patients. inadequate blood volume may compromise renal blood flow leading to acute kidney injury (aki). stroke volume variation (svv) is a sensitive indicator of relative preload responsiveness and has high sensitivity and specificity when compared to conventional indicators of volume status and their ability to determine fluid responsiveness. to assess the efficacy of svv guided fluid therapy in preventing aki in patients with severe sepsis on ventilatory support. mechanically ventilated patients with septic shock who had undergone resuscitation based on surviving sepsis campaign guidelines and still requiring vasopressor support were enrolled. patients with pre-existing renal failure were excluded. a total of 101 patients were randomized to receive fluid therapy according to conventional indices or svv, in the first 24 h after mechanical ventilation. svv was measured with flotrac vigelio after abolishing spontaneous ventilation by sedation and paralysis if required. fluid boluses were given to keep svv less than 13%. vasopressor therapy was optimised to maintain map [ 70 mm hg. patients were followed during their icu course with respect to development of aki, need for renal replacement therapy (rrt), length of icu stay and icu mortality. aki was diagnosed as per the rifle criteria. primary outcome measure was development of aki. results. patients in both groups were similar with respect to age (p = 0.6), sex (p = 0.7), and admission apache ii score (p = 0.6). incidence of aki was 29/48 (60.4%) and 21/53 (39.6%) in conventional and svv groups, respectively (p = 0.037). there was no statistically significant difference in terms of need for rrt, icu length of stay and icu mortality ( [1] . moreover, pnu-37883a (pnu), an inhibitor acting through the poreforming subunit of the channel, did not affect bp in our awake peritonitis rat model [2] . given that vasoconstrictors, including ne, inhibit k atp channel activity [3] , we speculate that the high sympathetic tone seen in sepsis [4] objectives. the goal of this study was to determine if hfav improves microcirculatory alterations in ss patients. methods. by using side dark field videomicroscopy (microscan ò , microvision medical) we evaluated sublingual microcirculation in 12 ss patients who according to our local protocol care [1] underwent a 12 h-hfav as rescue therapy for refractory septic shock. hemodynamic parameters and microcirculation were assessed at baseline, after 12 h of hfav, and 6 h after stopping hfav. microcirculation assessments were performed at 3 to 6 different sublingual areas (10-20 s/image). images were analyzed according to recent consensus [2] by semiquantitative scores of flow (mfi, mean flow index and ppv, proportion of perfused vessels), density (tvd, total vascular density; pvd, perfused vascular density), and heterogeneity (het mfi) of small vessels (\20 lm introduction. disturbances within the microcirculation represent an important factor in the pathogenesis of multiple organ dysfunction in sepsis and septic shock [1] . gender-specific effects may modulate the septic pathophysiology [2] . therefore, we studied sepsis-induced changes within the intestinal microcirculation in randomly cycling and ovariectomized female rats. objectives. we hypothesized that estradiol (e2) and dehydroepiandrosterone (dhea) may have a beneficial effect on the microcirculation during experimental sepsis and resubstituted these hormones in the ovariectomized animals. methods. fifty female rats were divided in to five groups of ten animals. group 1 received sham laparotomy without further treatment. in group 2-5 we induced experimental sepsis (colon ascendens stent peritonitis-casp model). animals of groups 3-5 were additionally ovariectomized 3 weeks before sepsis induction. in group 4 we administered 10 mg/kg estradiol immediately after and 12 h following casp surgery. the animals of group 5 received 50 mg/kg dhea immediately after sepsis induction. twenty-four hours after casp surgery intravital microscopy was performed to study leukocyte-endothelial interactions and functional capillary density. blood samples were taken for the measurement of estradiol, dhea and inflammatory cytokines. results. in ovariectomized rats subjected to casp the number of activated leukocytes was significantly increased in comparison to sham and not ovariectomized casp animals (p \ 0.05). in ovariectomized rats treated with e2 leukocyte adhesion was significantly reduced in comparison to untreated ovariectomized rats subjected to casp (p \ 0.05). the same observation was made in ovariectomized rats treated with dhea. in addition, in ovariectomized rats subjected to casp the functional capillary density was significantly decreased in comparison to sham and casp groups (p \ 0.05). in ovariectomized rats treated with e2 or dhea functional capillary density was completely restored. the results demonstrate the role of e2 and dhea in the sepsis-induced changes within the microcirculation. a rapid, non-genomic effect of both e2 and dhea is suggested [3] . dhea may play a role through conversion to e2 or through direct acting on the e2 receptor. further investigations should be done to elucidate the underlying mechanisms. both e2 and dhea appear to be a promising adjunct for the prevention and treatment of sepsis-induced multiorgan failure. liver is involved in the production of no. the aim of this experimental study was to evaluate the time course of hepatic no production at the onset of hypotension occurring during septic shock. methods. male wistar rats were anesthetized with isoflurane ò , and mechanically ventilated. a first group (sepsis group) underwent a cecal ligature and puncture (clp) peritonitis, the second one (control group) a laparotomy only. animals were euthanized at different times: 2 h after surgery, at shock onset, 2 and 3 h after shock. shock was defined by systolic blood pressure lower than 80 mmhg. each rat of sepsis group was matched with rat of control group. liver perfusion was measured using a direct laser doppler flowmetry probe. no generated in the liver was measured using a pulse voltametric method. results. 38 rats were studied (19 in each group). in sepsis group, shock occurred at 233 ± 51 min after clp. in sepsis group, a significant decrease of hepatic perfusion was identified 2 h after clp ( fig. 1 ) whereas hepatic no production was increased only at the time of shock onset (fig. 2 ). intra hepatic no production conclusion. this study shows a time shift between hepatic perfusion disturbance, hepatic no production and shock onset in a septic animal model. introduction. microvascular blood flow alteration is a key element of severe sepsis and septic shock [1] . one study show that microvascular alterations in septic patients could be improved with a nitric oxide donor nitroglycerin [2] . studies in human have shown that infusion of magnesium sulphate has endothelium dependent and independent vasodilation properties, increase of red blood cells deformability in specific conditions. we hypothesized that combination of nitroglycerin with magnesium sulphate and order of priority influence microvascular improvement in patients with severe sepsis and septic shock. methods. ten septic patients who had already been fluid resuscitated randomly assigned to one of two groups. one group received magnesium sulphate infusion 2 g/h with nitroglycerin 0.5 mg/h infusion added after 30 min. another group received nitroglycerin 0.5 mg/h infusion with additional magnesium sulphate 2 g/h infusion after 30 min. if required we added crystalloids and use norepinephrine. sublingual microcirculation was evaluated using side dark field videomicroscopy (microscanò, microvisionmedical) . each patient's microcirculation was evaluated by examining 5 different sublingual areas (10-20 s/image). in all patients measurements were obtained at baseline, at 30 and 60 min. images were analyzed by semiquantitative scores of flow (mfi, mean flow index; ppv, proportion of perfused vessels) and density (tvd, total vascular density; pvd, perfused vascular density). capillaries were defined as microvessels with a diameter \ 20 lm. data are presented as median values (percentiles 25; 75). . the median age of the patients was 60 (50; 71) years. in both groups we see tendency progressively increase of pvd, ppv and mfi after drug alone and combination after 60 min, but pvd has tendency to be higher [9.2(9; 9.8) n/mm 2 vs. 8.0 (7.8; 8.3) n/mm 2 , p = 0.05] after 60 min. in group, where magnesium sulphate infusion was given first. combination of magnesium sulphate with nitroglycerin, when magnesium sulphate is given first, has tendency to higher potential for improving of microcirculation in severe sepsis and septic shock patients, but further studies are needed to obtain more detailed results. severe sepsis remains one of the leading causes of death in critical care, with around 30% of patients dying within one month of diagnosis. rapid diagnosis and therapy of sepsis improves survival [1] . in november 2007 the whittington hospital introduced a hospital severe sepsis guideline, based on the first surviving sepsis campaign guideline [2] . the sepsis guideline was published on the hospital intranet and specified 20 actions to be completed within the first 24 h of the diagnosis of severe sepsis or septic shock [3] . objectives. to assess whether publication of a sepsis guideline on the hospital intranet, coupled with departmental educational campaigns, improved the management of severe sepsis. the whittington hospital is a university associated general hospital in london. we audited the early management of severe sepsis and septic shock before and after the introduction of the new hospital sepsis guideline. the 'before' phase comprised 22 patients with severe sepsis or septic shock admitted to critical care between november 2004 and november 2007. the 'after' phase comprised 30 patients with severe sepsis or septic shock admitted to critical care between january and november 2008, after introduction of the guideline. data was retrospectively collected from case notes and observation charts. the audit tool compared immediate, 1, 6 and 24 h actions following diagnosis against the hospital guideline. the main outcome measures were compliance and 28 day mortality. compliance was defined as the average of the percentage compliance with each of the 20 items specified in the guideline. results were compared by chi squared. compliance with the severe sepsis guidelines was only 64% after publication of the hospital sepsis guideline, compared with 68% before publication (p. 71)! there was similarly no significant difference in 28 day mortality (before 32%, after 30 %, p. 88). publication of a sepsis guideline on the hospital intranet, coupled with departmental teaching sessions, failed to improve compliance with surviving sepsis recommendations, perhaps because the guideline competes for attention with over 475 other guidelines on the intranet. next we will implement an interdepartmental educational programme to try and improve guideline compliance. as guidelines proliferate it is difficult to ensure they are followed, but failure to implement a published hospital guideline may represent a significant clinical and medicolegal risk. methods. the icu is an intensivist-led 10 bed intensive care in a 500 bed non-academic teaching hospital. hospital mortality from sepsis in icu-patients was 27.7% in 2007. patients are treated under modern icu conditioning, including continuous venovenous hemofiltration and a lung-protective ventilation strategy including prone position. an intensive insulin therapy protocol for glycemic control is used. in the period between march until june 2008, we prospectively screened all patients admitted to the icu for (severe) sepsis, without the knowledge of the nurses and most of the doctors. all severe septic patients were included in our surviving sepsis database. after 24 h, we examined how many targets of the resuscitation and management bundles were applicable and reached. in the period between march until june 2008, 154 patients were admitted to the icu. twenty-two of them were suffering from severe sepsis (14.3%), of which 18 had a septic shock. focus of the sepsis was abdominal in 10 patients (46%), pulmonary in five patients (23%), urogenital tract in five patients (23%), meningitis in one patient (4%) and catheterrelated in one patient (4%). table 1 shows us the applicability and achievement of the bundle elements. only in one of the 22 patients all targets were reached. however, mean individual bundle element performance was 65.3% (sd 39.7). all patients received fluid resuscitation when indicated, and all patients on mechanical ventilation were ventilated in a lung-protective manner with plateau pressures \30 cm h 2 o. only 33 percent of patients had glucose levels within the target range. scvo 2 was never measured, though it was indicated in 18 patients. one patient had an apache iiscore c25 and had no contraindications for administration of activated protein c. treatment was not considered for this patient by the attending physician. of these 22 patients suffering from severe sepsis, three died within 30 days after the diagnosis (13.6%). introduction. the surviving sepsis campaign (ssc) guidelines give a group of interventions (''sepsis bundles'') expected to improve the outcome of patients with severe sepsis [1] . objetives: the aim of this study was to evaluate the impact of the implementation of the ssc guidelines on the mortality in our intensive care unit (icu). methods. prospective, observational study. during one year period (january 2008-january 2009) the sepsis bundles were applied to each patient with severe sepsis-septic shock and they were followed up until discharge. we considered as ''time o'' (the time of delay of the implementation of the sepsis bundles) the time of admission of the patients in the icu. for each severe septic patient the following data was registered: time delay, apache ii and sofa scores at icu admission, diagnosis, the rate of compliance with the resucitation and management bundles, microbiological data, evolution of levels of serum lactate, empiric antibiotic therapy, length of stay and mortality in icu. the application of guidelines impact on mortality was compared with historical data years before implementation in our icu (46.3%) and spanish icu (48.2%) [2] . a total of 61 severe septic patients were included in the study. 7 (11.5%) patients had severe sepsis and 54 (88.5%) septic shock. the median age was 70 years. the mean apache ii was 21.7 (±8) and sofa was 8.5 (±4.3). the main sources of infection were abdomen (59%), lungs (21%), urinary tract (11.7%) and soft tissues (3.27%). the most common clinical diagnosis related to an episode of severe sepsis was peritonitis (59%). a microbiological diagnosis of the infection was reached in 75.4% and the infections were mostly caused by gram-bacilli. once the antibiogram was obtained, the initial treatment was considered appropriate in 96.7% patients. the rate of compliance with sepsis bundles was 100%. the length of icu stay was 15.3 days. mortality was 21.3%. the implementation of the sepsis bundles decreased icu mortality significantly (48.2% before implementation vs. 21.3% after implementation). non survivors were older (median age 77 ± 4.8), had higher apache ii (mean 29.3 ± 9) and sofa (mean 14 ± 3.6), 100% had septic shock, 61.5% had negative cultures and an increased on the levels of serum lactate in 24 h. age, apache ii and sofa scores and the increased on the levels of the serum lactate were useful tools to predict mortality. conclusion. implementation of the surviving sepsis campaign guidelines was associated with a reduction in icu mortality. introduction. the objective of this before-after study is to assess the impact of a protocol of care for severe sepsis in a french emergency setting. methods. two 4 months periods were surveyed before and after the initiation of a protocol of care for severe sepsis and septic shock. after the control period (p1: november 2004-february 2005), a procedure for early recognition, aggressive treatment and standardized antibiotherapy of severe sepsis was initiated. a campaign to raise medical physicians and nurses awareness concerning this new strategy of care was performed. the intervention period (p2: november 2006-february 2007) assessed the impact of these actions. . 82 patients with severe sepsis or septic shock were included during p1 (20% of patients with a suspected infection and 1.2% of all non trauma admissions) and 105 during p2 (22.8% of patients with a suspected infection and 1.3% of admissions). the age and the proportion of patients with co-morbidities were similar during the p1 and the p2 periods (82 years in median versus 82 years, and 49 vs. 50%, respectively). 39 and 41% of the patients lived in long term care facilities. severe sepsis and septic shock were correctly identified by the emergency team in 25/82 (30.4%) during p1 and in 57/105 (54.3%) in p2 (p = 0.002). the delay between the admission in the emergency department and the administration of antibiotics was in median equal to 4 h 28 min in p1 and 2 h 45 min in p2 (p = 0.0015). adequate iv fluid resuscitation was administered to 40% of patients in p1 and 63% of patients in p2 (p = 0.002). during p1, 20% of patients did not qualify for admission to the intensive care unit compared to 44.7% in p2. hospital mortality did not change from 29.3% (24/82) in p1 to 26.7% (28/105) in p2 (p = 0.81). conclusion. the introduction of a standardized treatment protocol in an emergency department allowed a better recognition of severe sepsis with earlier adapted treatment . the study was not powered to demonstrate a reduction of the mortality in this elderly population. multiple studies have shown that early detection and therapy is crucial for the prognosis of a severe septic patient. many hospitals have joined the surviving sepsis campaign and its fight for the decrease of mortality in severe sepsis and have implemented the severe sepsis bundles into their daily practice. other institutions such as ours had so far not taken this step, perhaps because the process is hard and time consuming. we have tried to find an easy way to audit the implementation of severe sepsis bundles and its change in time in an institution without a set system and database for the implementation of severe sepsis bundles to help us prove, that a systemic change in clinical practice is essential. we have decided to use the first step of the resuscitation bundle-the measurement of lactate and audit the lactate requests in blood samples with elevated inflammatory markers in our hospital laboratory information system. we retrospectively audited the number of lactate requests in blood samples with c-reactive protein (crp) c 200 mg/l and its evolvement in time between 2004 and 2007before and after the introduction of surviving sepsis guidelines and severe sepsis bundles in our regional hospital with 980 beds. we compared the total number of blood samples with elevated crp c over 200 mg/l with or without procalcitonin request in our institution with the number of blood samples with crp c 200 mg/l and lactate request (both arterial and venous) in the hospital laboratory information system. . the total number of lactate requests in samples with crp c 200 mg/l had increased in time, the incidence widely differed between departments. the main increase was in patiens from intensive care units, the number of lactate requests in samples from general wards, emergency department and intermediate (step down) units had also increased (18 lactate requests in 1667 samples with crp c 200 mg/l-1,1 % in 2004 and 104 lactate requests in 1857 samples-5.5% in 2007) but still remains insufficient. surprising was that procalcitonin was in non icu patiens with crp c 200 mg/l requested more often than lactate. although many lectures and seminars on severe sepsis bundles and the guidelines for the management of severe sepsis were organised in our intitution between the year 2004 and 2007, it was not sufficiently effective. conclusion. retrospective audit in the hospital laboratory information system of the number of lactate requests in samples with elevated inflammatory markers appears to be a fast and a very easy first step for auditing how the surviving sepsis guidelines and severe sepsis bundles are implemented in your institution. the results help to quantify the present state, its change in time and may serve as an impulse to make systemic changes in the system of early detection and therapy of septic patients. introduction. early goal-directed therapy (egdt) is the accepted gold standard for resuscitation in septic shock [1, 2] . international guidelines for the treatment of septic shock [2] set an initial 6 h limit to accomplish this goal. to test the hypothesis that egdt with fluids and vasopressors has better patient outcomes if each intervention is completed within 6 h. thirty septic shock patients from the spring of 2006 and 32 from spring 2008 were reviewed prospectively (n = 62). septic shock was defined as a lactic acid c 4mmol/l and/or hypotension unresponsive to fluids. apache ii and sofa scores were calculated. patients were subjected to the hospital septic shock protocol according to guidelines [2] . firstly, egdt compliance was met if the following 3 interventions were achieved within 6 h: lactate levels drawn, map c 65 mmhg and cvp c 8 mmhg; and secondly, if antibiotics were given\3 h, blood cultures were taken before antibiotics and if 20 ml/kg fluid bolus was administered prior to vasopressors. in 27 patients 5/6 interventions were performed in time (''egdt-compliant''). the other 35 were deemed ''egdt-noncompliant''. outcomes were mortality rate and discharge destination. fisher test was used in statistical analysis. . mr was 30% amongst the compliant and 43% amongst the noncompliant and admission to long-term care facilities (ltcf) was 52 and 40%, respectively. neither one of these differences was statistically significant. a power analysis revealed that 248 patients are required to attain statistical significance for mortality. discharge home was the same in both groups. there was no difference between groups in the number of new tracheostomies or new hemodialysis. conclusions. in a us community teaching hospital, compliance with guidelines in the treatment of septic shock had a trend towards lower mortality and higher discharge rates to ltcf but the difference was not statistically significant. larger numbers are needed for the benefits/effects of egdt-compliant therapy to reach statistical significance in the treatment of septic shock in this hospital setting. improve the survival rate of septic patients by means of education and implementation of a sepsis operative protocol including the activation of a specific consultation by an intensivist and an infectious disease specialist (i.e. sepsis team, st). aim of this study was to describe the first 30 months activity of st, with a focus on the patients not admitted in intensive care unit (noicu). methods. the sepsis operative protocol, introduced in clinical practice in june 2006, provides for specific instructions for the early identification and management of septic patients and for the early activation of the st for patients with severe sepsis or septic shock admitted in non-intensive departments. the st consultation ought to support the departmental health personnel in the management of septic patient and allows an early intensive care admission in case of shock or if mechanical ventilation is needed. to assess st activity, we evaluated in noicu patients the correct st activation rate, the number of st activations for each patient, the rate of central venous catheter insertion (cvc) and the 30 days mortality. results. from june 2006 to december 2008, the st was activated for 222 patients (7.4 patients per month) whose 109 (49%) were admitted to icu and 20 (9%) were considered too sick to benefit. in 60 (64%) of the remaining 93 patients, st was properly activated: 57 patients with severe sepsis and 3 with septic shock. thirteen patients (14%) had no sepsis and 20 (22%) had sepsis without organ dysfunction. 74% of st activations originated from medical departments (including emergency department) and 26% from surgical departments. the number of st activations for each single patient was 2 ± 1. the 30 days mortality was 0.05% in patients with sepsis, 23% in patients with severe sepsis and 33% in patients with septic shock. conclusion. the rate of correct activation of st and the number of activations for each patient were acceptable considering that more than 80% of the activations refers to septic patients and that a mean of 2 activations was sufficient for patient management. mortality rates observed are slightly lower than those reported by others, but further data are needed to evaluate the impact of st on patient outcome. a. estella 1 , l. pérez fontaiña 1 , j. i. sanchez angulo 1 , e. moreno 1 1 hospital of jerez, emergency and critical care unit, jerez, spain clinical evidence suggests that an early diagnosis and treatment of severe sepsis has been shown to improve outcome. frequently the initial management of septic patients occurs outside of the icu. objective. to describe clinical characteristics and outcome of septic shock patients admitted in icu and to compare mortality according origin prior admission in the icu (emergency department versus medical or surgical wards). consecutive patients with septic shock admitted in icu from july 2007 to november 2008 were registered. age, icu length of stay, source of infection, isolated bacteria, blood lactate concentration, apache ii score and mortality were collected. patients were classified according the origin prior admission in icu. . 97 consecutive septic patients were admitted in icu during the time of study, global mortality was 34%. 58 patients were admitted from medical or surgical wards and 39 patients from the emergency department. mean age was 64 years, 58 male and 39 female, icu length of stay was 8.4 ± 5.8 days, the mean apache ii score at admission in icu was 20.3 ± 7. abdominal infection, 44.3%, was the commonest source of infection followed by pulmonary and urinary infection, 21.6 and 20.6% respectively. 33 patients (34%) had a positive bacterial culture, the mean baseline lactate level was 3.8 ± 3.5 mmol/l p \ 0.05 (3.6 ± 4 mmol/l in the medical and surgical wards group versus 4.1 ± 2 mmol/l in the emergency department group).there were not differences in clinical characteristics according origin prior admission in the icu except for lactate level, and mortality, 56.9% in the medical and surgical wards group and 17.9% in the emergency department group (p \ 0.05). conclusion. there were not differences in clinical characteristics, icu length of stay, source of infection, isolated bacteria and apache ii score between groups. mortality was lower in the group of patients admitted in icu from the emergency department than the group admitted from medical and surgical wards. although very high circulating concentrations are detectable in plasma, it is not known which organs actually produce the cytokines. we hypothesized that key abdominal organs affected by sepsis such as the kidney and liver produce cytokines and tested this hypothesis by measuring cytokine flux. materials and methods. 12 pigs (15-26 kg) were randomised to control (n = 5) and endotoxin (n = 7) groups. hemodynamic measurements using picco and pulmonary arterial catheters and arterial blood gases were collected hourly. portal, hepatic and renal arterial blood flows were measured with transit time probes. arterial and venous cytokine concentrations (tnfa, il-1b, il-6 and il-10) were measured from samples taken from each respective organ. cytokine flux was calculated as: organ blood flow 9 (venous-arterial cytokine concentration difference). endotoxemic pigs had significant increases in heart rate (p \ 0.01) and mean pulmonary arterial pressure (p = 0.03) and decreases in cardiac output (p = 0.04). in contrast, these hemodynamic variables remained stable in the control animals. renal, hepatic and portal vein flows decreased significantly in all endotoxemic animals but remained stable in the control group. renal [ml/(kg min)]:control 5.5 ± 0.4, 5.4 ± 0.1, 5.4 ± 0.3, 5.3 ± 0.4 versus endotoxin 5.8 ± 0.6, 5.9 ± 0.9, 3.0 ± 1.0, 3.0 ± 0.4 for baseline, t = 2, 4, 6, respectively. portal [ml/(kg min)]: control 38.6 ± 3.0, 38.8 ± 5.8. 39.8 ± 5.0, 41 ± 6.7 versus endotoxin 42.7 ± 8.5, 15.1 ± 3.5, 12.4 ± 3.4, 12.4 ± 3.3 for baseline, t = 2, 4, 6, respectively. hepatic [ml/(kg min)]: control 5.1 ± 1.0, 5.3 ± 0.9, 5.5 ± 1.5, 5.2 ± 1.2 versus endotoxin 5.6 ± 0.9, 4.8 ± 1.4, 2.5 ± 0.7, 2.6 ± 0.1 for baseline, t = 2, 4, 6, respectively plasma cytokines tnfa was detectable in very low concentrations (\20 pg/ml) in 2 of the 7 endotoxemic animals, and none of the control animals. il-1b, il-6 and il-10 increased significantly with time peaking at t = 6, 4 and 2 respectively in the endotoxin group. in the control group only few animals showed a cytokine response, in numbers insufficient for statistical analysis. in the endotoxin group there was a negative cytokine flux in the renal circulation, maximal at t = 6 [-290.0 ± 429.6 for il-1b and -5210.7 ± 5865.6 for il-6 (pg/ml), respectively]. there was a positive cytokine flux for il-10 reaching its peak at t = 4 (9.1 ± 156.6 pg/ml). a similar pattern was seen in the hepatic ? portal circulation with maximal flux for il-1b and 6 at t = 6 (-291.0 ± 690 and -1346.4 ± 2687.4 pg/ml, respectively). for il-10 there was a positive flux peaking at -= 6 (1203.1 ± 1112.3 pg/ml). although there was a negative il-1b and il-6 cytokine flux in the renal, portal and hepatic circulations indicating net uptake, and vice versa for il-10, none of these values reached statistical significance. conclusions. these data do not support that cytokines are produced nor consumed in the kidney and liver during endotoxemia. discussion. non-survivors show more severity at the beginning and during their icu stay, more altered biological markers and a higher mean glycemia, but do not show significant difference either at initial glycemia, history of diabetes, hypoglycemia event or insulin treatment. elevated mean glycemia appears to be a factor independently associated with higher mortality. hyperglycemia prevalence in critically ill patients is very high and the controversy whether it is a mortality marker or a mediator still remains. our results would justify starting an intensive insulin protocol and its subsequent analysis. the interest of continuous scvo 2 was proven in the management of severe septic patients [1] , but the place of discontinuous scvo 2 remains unclear. objectives. to compare continuous scvo 2 to discontinuous scvo 2 concerning the number of therapeutic interventions in the management of severe sepsis (ss) and septic shock (ssc). methods. prospective randomized comparative study. inclusion criteria: age [ 16 years, ss or ssc [2] . two groups were defined: continuous scvo 2 (c group) monitored by a central venous oximetry catheter (edwards lifescience x3820hs, irvine, usa), and discontinuous scvo 2 (d group) measured on blood samples drawn every 4 h and at the request of the treating physician. the hemodynamic management of these patients was based on the algorithm established by rivers [1] . the primary endpoint was the number of therapeutic interventions (fluids, transfusions, inotropic drugs) triggered by a scvo 2 \ 70%. non parametric tests (chi-square and mann whitney) and repeated-measures anova were used in statistical analysis (p \ 0.05 was considered significant). results. 31 patients were included in a polyvalent intensive care unit (icu). the two groups were comparable concerning age, sex, weight, height, apache ii score, mods on admission and mechanical ventilation (mv). there were no statistical differences between the two groups concerning: mortality, duration of icu stay, duration of mv and the evolution of mods and plasma levels of lactate from day 1 to day 5. the therapeutic interventions data are shown in table 1 . introduction. the calcium activated potassium channel (bkca) exists in smooth muscle cells in most vascular beds and is believed to be important in sepsis induced hypotension and vascular hyporeactivity [1] and also in neutrophil killing and macrophage production of proinflamatory cytokines. however the latter two roles have been disputed [2] and we have found that bkca expression is not upregulated in aorta from septic mice using real time polymerase chain reaction. as its role in sepsis remains uncertain we sought to determine whether null mice for the bkca channel were (a) resistant to hypotension and (b) showed improved survival in a clinically relevant model of fecal peritonitis. methods. bkca null mice (based on balc) were obtained from jax ò mice. agematched litter mates homozygous for bkca were wild types (wt). mice (age 18-24 weeks) had tethered arterial and venous lines inserted under isoflurane anesthesia. the tether enabled mice to roam cages freely whilst continuous blood pressure (bp) traces were obtained. 24 h post surgery, echocardiogram and intraperitoneal injection of rat slurry was administered under anesthesia. fluid resuscitation of 0.3 ml/h voluven/5% dextrose (50:50) was given. at 6 and 24 h echo was recorded and mice culled with mesenteric arteries dissected for myography. data expressed as mean(sem) and statistical analysis anova. results. genotypic study and whole cell patch clamp recording in aortic smooth muscle cells confirmed bkca current was absent in null mice. fecal peritonitis induced equivalent hypotension in both wt (n = 5) and bkca null mice (n = 4) at 3-12 h (fig. 1a ). echocardiography at 6 h post slurry showed no difference in cardiac output between wt-23.1 (3.1) and bkca null mice-23.4 (2.6) ml/min and no difference or improvement cf time 0 (fig. 1b) . thus this fall in bp is due to reduction in total peripheral resistance not myocardial depression. in addition 3/4 of the bkca null mice died prior to 24 h as opposed to 1/5 wt. hence myography was only performed on wt mesenteric arteries which were hyporeactive to norepinephrine (p = 0.005, fig. 1c ). conclusion. there is no evidence from this transgenic mice study of fecal peritonitis that inhibition of the bkca channel would be beneficial for the treatment of hypotension in septic shock or would improve survival. reference(s). introduction. pro-and anti-inflammatory responses play a key role in the pathophysiology of sepsis [1] . phosphodiesterase (pde) inhibition could play an anti-inflammatory role in this setting [2] . previously, it was shown that among the three inhibitors of pde five currently available (sildenafil, vardenafil, tadalafil), only tadalafil could exhibit anti-inflammatory properties on endothelial cells (ec) stimulated by modified oxidized ldl or tnf alpha [3] . to assess the potential anti-inflammatory role of tadalafil in ec stimulated by lps. methods. thp-1 cells (2.10 6 /ml in rpmi) were incubated alone (control group) or in the presence of either tadalafil (3 lm; eli lilly, in, usa), lps from e.coli 0111:b4 (100 ng/ml; sigma-aldrich, inc.) or both. tnfa production, as a marker of inflammation, was measured in the supernatant (elisa assay; roche, mannheim, germany) after 16 h of incubation (3 independent experiments in quadruplet). comparisons were made by one-way anova, with bonferroni's post hoc test (mean ± sem). results. production of tnfa increased significantly after stimulation by lps alone compared to control (49.9 ± 7.7-fold over the control, p \ 0.05) or tadalafil (49.9 ± 7.7 vs. 1.0 ± 0.2-fold over control, p \ 0.05). levels of tnfa were significantly reduced in the lps ? tadalafil group, compared with the lps group (14.8 ± 2.1 vs. 49.9 ± 7.7-fold over the control, respectively; p \ 0.001) (graph 1). we hypothesized that daa provides varying protective effects in different organs as indicated by higher amounts of epcr in early murine sepsis. methods. sepsis was induced by cecal ligation and puncture (clp) in male nmri-mice (n = 21, body weight 30 ± 3 g). animals were randomly assigned to vehicle infusion (control), or clp sepsis with daa infusion [daa; 24 lg/(kg hr)]. a third group received only sham operation and vehicle infusion (sham). 48 h prior to clp all mice were given a permanent central i.v.-line and an arterial transmitter (pa-c10, st. paul, mn, usa) to measure heart rate (hr) and mean arterial pressure (map). clp was adjusted to survive 24 h. after 12 h hearts, livers and kidneys were fixed in formalin and embedded in paraffin. immunohistochemical analysis of the paraffin sections was performed using the avidinbiotin-peroxidase complex (abc) method. for analysis an anti-mouse epcr antibody (clone 1560, natutec, frankfurt, germany) was used (dilution 1:100) after heat pretreatment. anti-epcr positive cells were counted in 10 fields in light microscopy (original magnification: 62.5) of each tissue and the average was recorded. data are presented as mean ± sd. *p \ 0.05 was considered significant. results. there were no significant differences in hr between the groups (sham 650 ± 38 per min; daa 547 ± 175 per min; control 530 ± 143/min). map was significantly higher in sham group (137 ± 15 mmhg; p = 0.031) and non-significantly higher in daa group (111 ± 26 mmhg) when compared to control (97 ± 4 mmhg). anti-epcr positive cell count in heart tissue was significantly higher in sham-treated mice (6.3 ± 0.9 cells; p \ 0.001) and daa mice (6.0 ± 3.9 cells, p = 0.007) compared to controls (1.8 ± 1.3 cells). in kidney tissue epcr positive cells were significantly more in sham group (9.5 ± 4.0; p = 0.036) compared to control, but not in daa group (6.8 ± 2.5). liver samples showed no significant differences (sham 8.0 ± 2.3; daa 7.2 ± 3.8; control 6.3 ± 5.1). conclusion. our data showed higher amounts of epcr in murine sepsis undergoing daa therapy in heart and kidney tissues, but not in the liver when compared with control animals. this suggests that daa provides different effects in early experimental sepsis. background. caspofungin treatment is often initiated in hypovolemic shock patients, what could affect its pharmacokinetics and efficacy. the present study investigated the influence of hypovolemic shock and fluid loading on the plasma pharmacokinetic parameters and the pulmonary penetration of caspofungin in a pig model. after anesthesia and mechanical ventilation, 8 pigs (40 ± 3 kg) were bled to induce a 2-h deep shock and resuscitated for 2 h using normal saline based on hemodynamic goals. a 1-h perfusion of 70 mg caspofungin was started at the beginning of the resuscitation period. lungs were removed 6 h after the initiation of hemorrhage. sixteen animals were used as controls without hemorrhage. caspofungin concentrations were measured using high performance liquid chromatography method. in the shock group, the volume of removed blood was 39 ± 7 ml/kg and a volume of 90 ± 17 ml/kg of saline was infused through the resuscitation period. conclusion. hypovolemic shock followed by fluid loading in pig results in a significant decrease in plasma caspofungin exposition. it resulted in a decrease in the pulmonary concentration of caspofungin without affecting its diffusion to the lung. future investigations should focus on the interest for monitoring of plasma caspofungin concentrations in icu patients and on optimal dosing in these patients. objectives. the present study was designed to assess the effects of mps from septic origin on systemic hemodynamics as well as on the inflammatory, oxidative and nitrosative stresses. methods. forty healthy rats were randomly allocated to three groups: 10 animals inoculated with mps isolated from control rats (cmps), 15 animals inoculated with mps isolated from sham rats (shmps) and 15 animals inoculated with mps isolated from rats with peritonitis (smps). rats were anesthetized, mechanically ventilated and were infused with the same amount of cmps or shmps or smps. we measured heart rate (hr), mean arterial pressure (map), carotid artery blood flow (cbf) and portal vein blood flow (pbf). hemodynamic parameters were recorded during 7 h, and then animals were sacrificed. aorta and heart were harvested for further in vitro tissue analyzes. 1. the cellular origin (phenotype) but not the circulating concentration of mps was different in septic rats, characterized particularly by a significant increase in leukocyte derived mps. 2. smps but not cmps or shmps decreased mean arterial pressure without any effect on carotid artery and portal vein blood flows. all rats survived in the cmps and shmps groups whereas three rats died before the end of the experiment in the smps group. 3. rats inoculated with smps exhibited an increase in superoxide ion production and nf-kb activity, over-expression of inos with subsequent no overproduction and decrease in enos activation. pulse blood pressure recordings conclusions. rats with sepsis induced by peritonitis exhibited a specific phenotype of mps which could play a detrimental hemodynamic effect as a systemic vasodilatation. inoculation of smps in healthy rats decreased map likely by up-regulating nf-kb activity with subsequent inos, no and superoxide anion overproduction. these data confirm a proinflammatory detrimental role of mps in the vascular pathophysiology of septic shock. introduction. heat shock proteins (hsps) play an active part in modulating intracellular responses to stress. in the classical model for their activation de-repression of heat shock transcription factor 1(hsf1) occurs as a result of the titration of hsps away from hsf1 by misfolded proteins [1] . however, hsps may change in many diseases without any changes in the levels of denatured proteins [2] . objective. we propose that hsps are activated, in part, by a membrane dependent calcium channel receptor, possibly transient receptor potential vanilloid type-1 (trpv1). capsaicin, a known inducer of trpv1, and capsazepine, a selective antagonist, were used on different mammalian epithelial cell lines. cells were pre-treated with micromolar concentrations of capsaicin or heat shock (hs) followed by treatment with capsazepine. results. capsaicin or hs induced hsf1 activation and the consequent accumulation of hsp70, 90 and 25 chaperones. pre-treatment with capsazepine prior to hs or capsaicin abolished the heat shock response (hsr). capsazepine treatment prevented capsaicininduced stabilization of ikb and cell to cell adhesion and induced apoptosis. capsazepinemediated blockage of the heat shock response was reproduced with egta. moreover, treatment with trpv1 sirna resulted in a similar response to capsazepine. conclusion. hsr-sensing and signaling in mammalian cells depends, in part, on the transient entry of calcium by way of membrane dependent calcium channel receptor. these hsr modulators may hold promise in treating inflammation in the future. introduction. hydrogen sulphide gas, or its intravenous donor-sodium hydrogen sulphide (nahs), are promising therapeutic agents in ischaemia-reperfusion and haemorrhagic shock [1] . we studied nahs in a short-term endotoxaemia model as relatively little is known about its effects during sepsis. methods. under isoflurane anaesthesia, male wistar rats (approx 280 g weight) underwent left common carotid and right jugular venous cannulation for blood sampling/continuous bp monitoring and fluid administration, respectively. animals were kept normothermic on a heating mat. tissue oxygen tension (tpo 2 ) was monitored using oxylite probes (oxford optronix, oxford uk) placed in thigh muscle. after a 30-min stabilization period, fluidresuscitated rats [10 ml/(kg h)] were subjected to iv lps (20 mg/kg over 15 min). comparisons were made against animals receiving nahs (0.2 mg/kg bolus given immediately after lps, followed by a 2 mg/(kg h) infusion). echocardiography (vivid 7, ge healthcare, bedford) and blood gas analysis were sequentially performed. sham-operated, non-septic animals also received nahs (n = 5) or placebo (n = 7). at the doses given, nahs had no effect on either sham-operated animals (data not shown), nor on the endotoxic rats (table 1) . data shown as mean (±se). timepoints chosen reflect the biphasic response to endotoxin: 0 = baseline, 15 = initial hypotensive phase, 120 = maximal recovery, 240 = end of experiment. conclusion. nahs does not improve haemodynamics, tissue oxygenation nor shockrelated biochemical parameters in a severe model of fluid-resuscitated endotoxaemia. we will further investigate the effects of dose and time of therapeutic intervention in this model, in addition to testing it in a long-term septic model. intestinal endothelial and epithelial barrier dysfunction remain severe clinical problems as they may contribute to the development of sepsis and multiorgan failure. we have recently established an isolated rat small intestine model with access to vasculature, lumen and lymphatics for study of inflammatory changes in fluid balance [1] stable for 140 min, rendering it less suitable for examination of changes in gene and protein expression profile. the aim of this study was to assess the long term functional and metabolic stability of this model. adult female wistar rats were anaesthetized, small intestines cannulated and perfused vascularly (7.5 ml/min) and luminally (0.15 ml/min) and placed in a warm humidified chamber for up to 6 h. arterial, venous and luminal pressures as well as venous, luminal and lymphatic effluent flows and intestinal weight were recorded continuously. as measures of metabolic integrity, oxygen consumption, lactate/pyruvate ratio and galactose uptake from luminally administered lactose were analysed every 30 min. structural and barrier integrity were assessed as histostability score (mesenteric and antimesenteric fraction of fully epitheliated villi), wet/dry weight ratio and translocation of vascularly applied fitc albumin to lumen and lymphatics. data were compared using paired t tests. 34 ± 0.03/0.29 ± 0.04 ml/(min g) dry weight (**)) as well as galactose uptake (0.24 ± 0.07/0.25 ± 0.07 mg/(min g) dry weight (n.s.)) were very stable with time pointing towards high metabolic stability. during the whole experiment, luminal effluent flow was slightly lower than applied (0.13 ± 0.01 ml/min, 240 min) resulting in net liquid absorption over the whole time period (0.022 ± 0.018/ 0.017 ± 0.012 ml/min (n.s.)), and lymph production stayed in the physiologic range (0.014 ± 0.010/0.035 ± 0.039 ml/min (n.s.)). the organ weight did not change with time which, together with the balanced luminal fluid flow and end experimental wet/dry weight ratio of 4.38 ± 0.17 (compared to 3.97 ± 0.08 at the beginning of the experiment (**)), indicate absence of edema. minimal leakage of vascular fitc albumin to the lumen (0.4 ± 0.3%) and a histostability score of 0.97 ± 0.04 show integrity of the vascular-luminal barrier until the end of the experiment. the isolated small intestine model presented earlier [1] displays excellent long term physiologic, metabolic and histologic stability and opens up a wide field of applications including inflammatory gene transcription and protein expression. introduction. mitochondria play a major role during ischemia-reperfusion as well on cytotoxic pathways as protective such as ischemic preconditioning. the aim of this study is a better understanding of the mitochondrial pathophysiologic response to several oxygen regimens in an isolated mitochondria model. mitochondria were isolated from rat heart. enriched mitochondrial pellets were conditioned in presence of glutamate (5 mm) and malate (2 mm) inside the oxygraph chamber during 15 min. oxygen partial pressures were: 155 mmhg for control group; 0 to 4 mmhg for hypoxia group and 0 mmhg for anoxia group. then, after a 5 min oxygenation period, several measurements were realized: oxygen consumption (vo 2 ) were measured with or without adp (5 mm) (state 3 and 4 of mitochondrial respiration); calcium retention capacity (crc); mitochondrial membrane potentiel (dwm). to explore the involvement of reactive oxygen species (ros), mitochondrial vo 2 were measured in presence of a specific mitochondrial antioxidant drugs (xbj). all results were expressed in percent of variation in comparison to control group [median (minimum-maximum)]. the different groups were analyzed using a kruskal-wallis, a mann-whitney with a bonferroni correction or a sign test when necessary. after hypoxia and reoxygenation the mitochondrial function was altered. this impairment of mitochondrial function was not found after anoxia and reoxygenation. this difference in mitochondrial function between hypoxia and anoxia suggests the involvement of ros. this hypothesis was confirmed by the effect of the antioxidant xbj that reestablished after hypoxia the same level of vo 2 than after anoxia. [1] . superoxide dismutase (sod) catalyses the dismutation of superoxide oxygen free radicals to oxygen and hydrogen peroxide (h 2 o 2 ). the therapeutic potential of exogenous sod administration in ards is evidenced by demonstrations of efficacy in acute lung injury models [2] . anti-oxidant defenses, particularly the extracellular sod isoform, extracellular sod (ec-sod), are downregulated by endotoxin [3] . we proposed that ec-sod delivered via a novel viral vector would ameliorate lung injury caused by lipo-polysaccharide (lps) pulmonary instillation. methods. three groups with nine rats per group were randomised to receive either adenoassociated virus expressing ec-sod (aav-ec-sod), adeno-associated virus coding for no product (aav-null), or vehicle control, 5 days prior to planned lps instillation. a model of lipo-polysaccharide (lps) induced acute lung injury by pulmonary instillation was established in male sprague dawley rats. twenty-four hours following lps delivery, animals were anaesthetized and mechanically ventilated and their baseline compliance and oxygenation recorded. there was a statistically significant improvement in the oxygenation of animals recieving aav-ec sod as compared to aav-null or vehicle control (mean pao 2 = 14.348 vs. 9.756 and 8.679, respectively). there was a significant increase in amount of ec-sod as determined by real time pcr in the group who were administered aav-ec sod. no significant differences in static compliance or bronchoalveolar lavage cells counted were noted. conclusion. aav delivered ecsod is protective in a animal model of lps induced acute lung injury. the down regulation of the ec sod system seen in the systemic inflammatory response [3] and its subsequent replacement exogenously may explain our findings. further work will focus on other components of cellular anti-oxidant pathways and confirmation of down regulation of ec sod in our injury model. aims. the endothelial specific angiopoietin (ang)-tie2 ligand-receptor system has been identified as a non-redundant mediator of endothelial activation in experimental sepsis. binding of circulating ang-1 to the tie2 receptor physiologically protects the vasculature from leakage, whereas binding of ang-2 antagonizes tie2 signaling and disrupts endothelial barrier function. we tested whether administration of exogenous recombinant ang-1 improves survival and attenuates multi organ failure in a lethal murine sepsis model. to induce septic acute kidney injury and to evaluate survival time cecal ligation and puncture (clp) was performed in twenty sv129 mice. half of the mice received an intravenous application of recombinant human ang-1 (50 lg) immediately before clp and every 8 h thereafter. in the other half, saline was administered in the same fashion. for tissue assessment (western blot, immunohistological) clp was induced in 5 versus 5 (ang-1 vs. saline) additional mice; animals were sacrificed after 24 h. laparotomy served as sham control (n = 5). further, a panel of cytokines has been assessed with a cytometric bead array (cba) system after 24 h. 23.1 ± 3.6 mmol/l, p \ 0.01) were lower in ang-1 treated septic mice compared to controls. similar results were obtained at 16 h after clp. renal tissue revealed that saline treated mice exhibit a marked loss of expression of vascular endothelial (ve)-cadherin, a major component of endothelial adherens junctions. in contrast, loss of ve-cadherin expression was prevented by ang-1 (pre-) treatment (wb densitometry: ang-1: 0.82 ± 0.2; saline: 0.5 ± 0.1; p = 0.05). however, contrary to previous reports, intravenous injection of exogenous ang-1enhanced not only the expression of adhesion molecules (icam-1, vcam-1) in renal vasculature, but also circulating cytokine levels (tnfa, mcp-1, il-6, il-12). conclusions. our study demonstrates that administration of exogenous recombinant ang-1 improves survival time in a lethal experimental sepsis model. enhanced survival was accompanied by an improvement in microcirculatory function, probably via stabilization of adherens junctions. however, ang-1 injection deteriorated expression of vascular adhesion molecules and raised plasma cytokine levels. although ang-1 may have utility as an adjunctive agent for the treatment of septic multi-organ failure, additional dose-finding and efficacy studies are required. adaptive immune responses to infection. in contrast to neutrophils, macrophages or lymphocytes, there are virtually no data on the time course of circulating dcs in septic shock (ss). using a novel specific and sensitive assay, we analyzed the evolution of circulating myeloid (mdcs) and plasmacytoid (pdcs) dcs in ss. we enrolled immunocompetent adult patients with ss (n = 43), shock from other etiologies (nss, n = 29) and with sepsis without organ dysfunction (s, n = 15). 16 age-matched healthy controls (hc) served as reference for mdcs and pdcs. blood samples (200 ll) were drawn on the day of shock, then after 3 and 7 days. dcs were counted using the dc-labelling kit trucount ò assay (bd biosciences). cd11c? cd123-(mdc) and cd11c-cd123? (pdc) cells were selected by flow cytometry (facscanto tm , bd biosciences). hla-dr mean fluorescence index (mfi) was measured. age, sex ratio, saps ii, sofa score, nosocomial infection (ni) and mortality rates did not statistically differ between ss and nss pts. at day 1, mdcs and pdcs counts were significantly lower in ss and nss pts as compared to hc and s ( fig. 1 ). pts with ss had significantly lower mdcs and pdc counts than nss at days 1 and 3. hla-dr mfi of mdcs and pdcs was lower in ss pts compared to hc (p = 0.005 and 0.037, respectively). interestingly, 10 of the 43 ss pts developed ni after a median time of 9 (7.5-11) days in the icu. whereas mdcs increased in pts without ni, mdcs counts remained low at day 7 in pts who developed ni: mdcs counts and their relative variation between day 1 and 7 were significantly lower in pts who developed ni than in those who did not (p \ 0.05). logistic regression analysis indicate that a negative mdcs relative variation is associated with an increased risk of nosocomial infection with an or 22 (2.53-191) (p = 0.005). figure conclusion . ss is associated with quantitative and qualitative abnormalities of circulating mdcs and pdcs as early as day 1, independently of the haemodynamic injury. the persistence of low counts of mdcs after ss is associated with the advent of nosocomial infection during the icu stay, suggesting that dcs play a role in the development of sepsisinduced immunosupression. introduction. liver dysfunction is common in sepsis but its mechanisms are unclear. the aim of the study was to evaluate the effects of lps on cultured primary human hepatocyte respiration over time. methods. human hepatocytes were isolated and cultivated from human liver resection specimens. cultivated cells were exposed to lps (1 lg/ml) for 4, 8 and 16 h. after incubation, cells were trypsinized and respiration rates were measured using a high-resolution oxygraph (oxygraph-2 k, oroboros instruments, innsbruck, austria). glutamate ? malate (g ? m), succinate (s) or ascorbate/tmpd (a/t) were used as substrates to test the function of complex i, ii and iv, respectively. human hepatocyte mitochondrial function in the cells treated with lps for 8 h exhibited a significant reduction in the maximal complex ii-dependent mitochondrial respiration [control: 464 ± 142 vs. lps: 406 ± 163 pmol/(s 9 million cells) ( table 1) ]. after 4 and 16 h of lps incubation no significant reduction in cellular respiration was observed (4 and 16 h: n = 5 and 8 h: n = 4). statistics: paired t test, *p = 0.028 control vs. lps (8 h incubation). introduction. acute kidney injury (aki) in critically ill patients is a frequent clinical problem and a rising incidence has been reported over the past several years. recently two consensus definition for aki have been developed: rifle [1] in 2004 by the acute dialysis quality initiative workgroup (second conference) and akin [2] in 2007. insofar akin and rifle criteria have been applied in large retrospective studies, limited to the initial days of icu. nefroint is an italian initiative for an observational prospective multicenter study to evaluate epidemiology of aki in italian icus employing rifle and akin classifications. a pilot study has been performed in one of the centers enrolled. objectives. primary endpoints of nefroint are: application and comparison of rifle and akin criteria for aki definition in a prospective observational study; estimate, along such criteria, of aki incidence in critically ill patients; correlation of aki stages with prognosis. method. an observational prospective multicenter study has been designed, in italian adult icus (medical and surgical). all incident icu patients have been enrolled over a 6 month period. exclusion criteria was age \15 years, or icu stay \24 h. data collection about patients was performed on a web-based electronic case report form. data included icu admission diagnosis, daily urine output (3 h interval), daily laboratory data. sepsis events diagnosed on clinical and/or microbiological basis where as well marked for each patient. severity scores have been calculated at admission and daily. aki patients had higher severity of illness scores and higher serum creatinine values on admission. they also were older and more likely to have a respiratory diagnosis as reason for icu admission. conclusions. nefroint is an initiative aimed at comparing rifle and akin scores to promote a uniform use of a single definition of aki that will render subsequent studies comparable. early aki recognition could potentially allow implementation of timely corrective interventions, and hopefully prevent progression to more severe stages. aim. sepsis and septic shock remain the most important causes of acute kidney injury (aki) in critically ill patients and account for more than 50% of cases of acute renal failure (arf) in intensive care units (icu). its mortality varies with the severity of sepsis from 21% to 57%. the aim of this preliminary study was to investigate the differences in the course and prognosis of aki that was induced by community and hospital acquired sepsis. method. patients with sepsis induced aki were included in the study. rifle criteria were used to define aki. clinical and laboratory characteristics of the patients were compared with student t test and chi square tests. results. forty-one patients were included in the study and 24 of them had community acquired septic aki (akic). ninety percent of the patients received mechanical ventilation (mv). etiologies of sepsis were mostly community acquired pneumonia and ventilator associated pneumonia. age, gender, admission apache ii scores and sofa scores at the time of aki diagnosis were similar across the groups (p [ 0.05). hospital acquired septic aki (akih) developed later when compared to community acquired septic aki (10th and 3rd days of sepsis respectively, p 0.004). akih was significantly and more frequently associated with oliguria (73 vs. 35%, p 0.020), bacteremia (47 vs. 4%, p 0.001), nephrotoxic antibiotic usage (59 vs. 21%, p 0.013) and tend to progress more frequently to acute renal failure(63 vs. 18%, p 0.005) compared to akic. akic episodes were more frequently (74 vs. 41%, p 0.37) and rapidly (7 vs. 12 days, p 0.44) reversible. mean blood pressure and scvo 2 % were significantly lower and more vasopressor and steroid therapies were required during akih episodes compared to akic (p \ 0.05). while length of mv and mortality rates were similar, duration of hospitalization was significantly longer in the akih group (32 vs. 18 days, p 0.045). conclusion. these results suggest that, akih has worse clinic and prognosis than the akic so further and larger studies are necessary to investigate the preventive and therapeutic approaches. introduction. severity-of-illness or organ dysfunction scores are inaccurate to predict outcomes in patients with acute kidney injury (aki), even when specific aki scores are used. in recent years, the third versions of simplified acute and physiology score (saps 3) [1] and of mortality probability model (mpm0-iii) [2] scores were developed, and information on their use in patients with aki is scarce. objectives. to validate the use of saps 3 and mpm0-iii at the start of renal replacement therapy (rrt) in patients with aki. prospective cohort study conducted in the icus of three tertiary-care hospitals. data used to calculate the scores were collected at start of rrt. discrimination was assessed by area under receiver operating characteristic (aroc) curves and calibration by hosmer-lemeshow goodness-of-fit test. a total of 244 consecutive patients were included between january 2007 and july 2008. the mean age was 69.5 ± 16.6 years. the main contributing factors for aki were ischemia/shock (75%), sepsis (74%), contrast/nephrotoxins (33%), rhabdomyolysis (5%) and urinary tract obstruction (2%) (a patient could have more than one contributing factor). eightnine (36%) patients received rrt on the first day of rrt and 155 (65%) thereafter; continuous rrt was used as first indication in 206 (84%) patients. the icu and hospital mortality rates were 63 and 68%, respectively. the mean saps 3 score at the start of rrt was 70.0 ± 13.0 points. both the standard equation of saps 3 and mpm0-iii scores tended to underestimate mortality. discrimination was better for saps 3 [aroc = 0.82 (95% ci, 0.76-0.88)] than for mpm0-iii [aroc = 0.73 (95% ci, 0.66-0.80)], as was the calibration. however, mortality prediction and calibration improved when the customized equation of saps 3 for countries from central and south america was used. in multivariate analyses, both higher prognostic scores and length of icu stay prior to rrt were the main predictive factors for hospital mortality. conclusions. the saps 3 score at the start of rrt was accurate in our cohort of patients and seems a promising instrument for predicting hospital mortality critically ill patients with aki. objectives. the aim of this study was to investigate the effect of hes administration on kidney function compared with other colloids or crystalloids. methods. systematic review and meta-analysis of the effects of hes administration on kidney function. inclusion criteria for the study were prospective randomized trials comparing hes to control with reporting on variables of kidney function. aims. during the initiation phase of experimental acute kidney injury (aki), subtle but devastating changes, such as loss of brush borders, disruption of tubular cell polarity and cytoskeletal changes are detectable only to a certain extent by routine histologic methods. for this reason, subjective and moderate reproducible semi-quantitative scoring of tubular changes (e.g. vacuolization, detachment, cast formation, and necrosis) still remains the method of choice to quantify the extent of experimental aki. lectins are glycoproteins which are able to bind carbohydrate structures specifically. it has previously been shown that immunolabeling of the lectin phaseolus vulgaris erythroagglutinin (pha-e) is highly specific to the brush border of proximal tubular epithelial cells of rats, mice, and humans. the aim of this study was to (1) develop a simple and fast lectin (pha-e) based staining protocol (2) to objectively quantify, and (3) to analyze brush border loss in a murine model of septic aki. methods. septic aki in mice (n = 10) was induced by cecal ligation and puncture (clp). animals were sacrificed 24 h after clp.sham operated (n = 5) and healthy animals (n = 5) served as controls. in order to specifically stain the tubular brush border, binding of biotinylated lectin pha-e was visualized by the biotin-avidin-complex (abc) glucose-oxidase (go) method coupled to tetranitroblue tetrazolium (tnbt) in 1-lm paraffin sections of renal tissue. the mean brush border area of five randomly chosen, non-overlapping cortical highpower fields was analyzed by planimetric software. lectin pha-e staining was highly selective for brush border of proximal tubules (black colour). virtually no staining was present in glomeroli and medulla. the xx software reliably identified lectin-positive areas, as confirmed by image overlay controls. we found a significant difference between sepsis induced aki, sham operated animals, and healthy mice (clp: 0.091 ± 0.019; sham: 0.141 ± 0.048; healthy controls: 0.266 ± 0.14 pixel ratio; p \ 0.001). our findings with the pha-e staining protocol correlated significantly with the conventional semi-quantitative scoring system (r = 0.67, p \ 0.01). conclusion. the here presented lectin pha-e staining method followed by computerassisted planimetric quantification of brush border area is a highly reproducible and objective tool to analyze early histological changes during septic aki in mice. when an imbalance between oxygen supply and demand exist, anaerobic respiration commences and a metabolic acidosis develops. base excess and lactate have been used to identify a higher risk group of patients who should be admitted in icu prior to development of multiple organ failure. and at a time when appropiated therapy may previne the decline to death. acute kidney injury failure is a common complication in critically ill patients and it always difficult separate the acid base effects of critical illness per se from those of aki. the aim of this study was to examine wheter values of base excess or lactate taken on admission of patients with aki to a intensive care unit indicate prognosis and if wheter this can be used as screening tool for future intensive care admissions. we restropectively examinated data from 100 patients with aki. to define the unique acid base characteristics of aki patients, we used a control group. the matched group consisted of 80 icu patients wihtout aki matched for apache ii score. the base excess and lactate were collected at admission and then at 24 h. a total of 180 patients were enrolled at study over a 10 month-period. there were no difference with respect age, sex and apache score between groups. the icu survival rates were 72% to the aki group and 78% to control group. the value of base excess with the best predictive prognosis ability was -9 mmol/l to the aki group and-3.5 (p \ 0.001) to the matched group and the corresponding value for lactate was higher than 2.6 to both groups. the combination of these two markers on admission to the intensive care unit led to a sensitivity of 82% and specifity of 58% for mortality. conclusion. both base excess and lactate, or the combination of the two, can be used to predict 28 day mortality in patients admitted to the intensive care unit. in patients with aki a different cut off of base excess should be used.these variables could be utilized to identify patients who have a higher risk for mortality to whom resources could be better directed. nonthyroidal disease (ntd) is a common finding in patients who are critically ill or on dialysis or with cardiovascular disease. its presence has been associated with inflamatory conditions. the aim of this study was to analyse the posible association of ntd with the development of acute kidney injury (aki). secondary targets where to estimate the incidence of ntd in a polyvalent icu and observe the realationship between the levels of t3 and some inflamatory markers: c reactive protein (crp), albumin and cortisol. during 7 months in 2008, after approval of the local ethical committee, we prospectively determined the following parameters in every patient admited to the icu: t3, t4, tsh, serum creatinine (scr), crp, albumin and cortisol. after excluding patients who died or were discharged before 48 h, 107 patients were studied. the degree of aki was calculated using the rifle scale. at admission the values of the analysed parameters were (mean ± sd): t3 1.63 ± 0.8 pg/ml; t4 1.13 ± 0.4 ng/dl; tsh 1.14 ± 1.5liu/dl; scr [1] . its incidence (5-38%) is rising due to increasing numbers of ct scans and contrast studies conducted, and the higher prevalence of risk factors such as chronic renal impairment, diabetes mellitus and old age. although usually selflimiting, cin can be associated with a need for ongoing dialysis or increased mortality [2] . to highlight the problem of contrast induced nephropathy and the difficulties in interpreting the current evidence for possible prevention strategies. we present the case of a 75 year old man admitted to intensive care with acute pancreatitis. he underwent eight contrast-enhanced abdominal ct scans and received nacetylcysteine (nac) for all but one of these, after which he developed acute renal failure which did not recover. we also present a review of evidence for various proposed strategies. results. several studies have examined possible renal protective strategies around contrast administration. saline and bicarbonate have been shown to be beneficial when given pre-contrast [3, 4] . theophylline has been shown in meta-analysis to have a significant beneficial effect, but heterogeneity of methodology between studies makes it difficult to clarify the degree of benefit achieved [5] . nac has shown benefit in 8 of 22 trials. twelve meta-analyses showed inconsistent results, with 7 showing nac to be beneficial. none showed harm. we analysed the heterogeneity of methods, endpoints and patient groups that makes these studies difficult to compare. critically ill patients may be considered at even greater risk of cin. only one study has specifically looked at this group. strategies such as volume loading may be inappropriate in some patients and there may not be time for nac for 24 h pre-contrast. we were unable to find specific guidelines for the prevention of cin in critically ill patients. conclusion. the evidence for strategies to prevent cin specifically in critically ill patients is unclear. we review the current literature and propose renal protective strategies including hydration, nac and theophylline for this patient group based on the evidence available. objectives. the present study addresses the issue of how the different modes of rrt are currently used and performed. we conducted a prospective observational study in three portuguese intensive care units (icu). patient demographics, type of rrt used and outcomes were collected. we studied 84 patients who were treated with rrt for rf, with a median age of 58 years and a saps-ii score of 51.9 ± 15.9, a sofa score of 9.8 ± 4.1 at admission; 59 patients (70.2%) were treated with continuous replacement therapy (crrt), 14 patients (16.6%) with sustained low-efficiency dialysis (sled)and 11 patients (13.2%) were initially treated with crrt and latter with sled. using the rifle criteria for the stratification of acute renal dysfunction at the beginning of the rrt we observed: risk-2 (2.4%), injury-18 (21.4%), failure 52 (61.9%), loss-1 (1.2%), esrd-11 (13.1%). we used anticoagulation in almost all patients (83.3%). among patients who received anticoagulation, heparin was the most common choice (71.4%), followed by low molecular weight heparin (18.6%), and by sodium citrate ( introduction. in the intensive care unit (icu), severe sepsis and multiple organ failure are frequently associated with renal failure. continuous veno venous hemofiltration (cvvh), which is used as renal replacement therapy, also removes circulating inflammatory mediators. standard cvvh is currently prescribed with a substitution flow of 35 ml/(kg min). theoretically, when hemofiltration is performed with higher volumes, buffer balance will be restored more rapidly, while also more inflammatory mediators will be removed. this may result in faster stabilisation from septic shock. indeed, animal-and some human studies show promising results, but have several (methodical) limitations. to evaluate hemodynamic and metabolic changes during hv-cvvh in patients with septic shock in comparison to (standard) cvvh. we performed a retrospective, observational, single-center study. all patients admitted with septic shock who were treated with cvvh in the period 2005 until 2008 were included. cvvh was defined as a substitution-flow b3,000 ml/h, hv-cvvh as[3,000 ml/h. the decision to start with lv-cvvh or hv-cvvh was made by the attending icu-physician on an intention-to-treat basis. statistical analyses were performed with spss 13.0 introduction. haemostatic changes in critically ill patients are complex due to simultaneous pro-and anticoagulant processes. routine ptt and aptt assays monitoring clot formation poorly reflect hypo-or hypercoagulant state, especially during anticoagulation. endogenous thrombin potential (etp) comprises an in-vitro system for measuring thrombin generation beyond clot formation and may be more informative. objective. to assess whether etp has a role in monitoring systemic anticoagulation and predicting circuit clotting in critically ill patients receiving cvvh. methods. in a prospective study in an 18-bed general icu, we included 14 patients with acute renal failure (arf) requiring cvvh (postdilution, 2-4 l/u). patients received a bolus of 2,850 iu of nadroparin followed by 380 iu/h. samples of arterial and postfilter blood were taken at baseline and 1, 6, 12 and 24 h after start of cvvh to measure aptt, ptt, anti-xa and etp. we compared patients with early circuit clotting (circuit life £ lower quartile) and those with normal circuit life. median baseline arterial etp-area under the curve (auc) was 277 ma (iqr 175-385 ma) (normal values 346-520 ma). baseline etp-auc was positively related to antithrombin and inversely to ptt, aptt, anti-xa (p \ 0.01) and sofa score (p = 0.001). median circuit life was 24.5 h (iqr 12-37 h). at baseline, the four patients with early filter clotting (£12 h) had prolonged ptt and aptt, higher sofa score and a tendency to lower etp (table 1) . during cvvh and nadroparin infusion, arterial and postfilter ptt and aptt were prolonged (p \ 0.001), antixa lower (p = 0.003) and etp-maximal concentration (cmax) lower (p \ 0.004) when circuits clotted early. while arterial etp-auc tended to be lower (p = 0.08), postfilter etp-auc was not different between groups. in critically ill patients with arf requiring cvvh with nadroparin anticoagulation, baseline etp is lower than normal and inversely related to organ failure and (a)ptt, probably reflecting consumption of coagulation factors. within the cvvh circuit, etp-auc and anti-xa show opposing patterns. the concurrence of early filter clotting with prolonged (a)ptt, lower antixa, lower etp and higher sofa score emphasizes the role of severity of disease and associated coagulation activation and heparin resistance in circuit clotting. introduction. nadroparin is a low-molecular-weight heparin (lmwh) used to prevent clotting in the extracorporeal circuit during cvvh. in renal failure lmwh accumulates and is associated with more bleeding (ref) . whether nadroparin is removed by hemofiltration and whether the anticoagulant activity accumulates during continuous infusion is controversial. objective. to study the kinetics and removal of anti-xa activity during continuous infusion of nadroparin in patients requiring cvvh using a cellulose tri-acetate filter. methods. in a randomized crossover trial in an 18-bed general icu, patients with acute renal failure (arf) were randomized. in group 1, postdilution cvvh was initiated at filtrate flow of 4 l/h (blood flow (bf) 220 ml/min), which was converted to 2 l/h (bf150 ml/min) after 60 min; in group 2, 2 l/h was converted to 4 l/h. patients (\100 kg) received a bolus of 2,850 iu nadoparin followed by 380 iu/h. samples of arterial blood, postfilter blood and ultrafiltrate were taken at baseline, 1 h after the start and 15 min, 6, 12 and 24 h after the conversion to measure anti-xa activity. results. fourteen patients with arf were equally randomized. patients in group 1 had higher median sofa scores (14 vs. 9, p = 0.004), baseline coagulation markers were not significantly different. arterial and postfilter anti-xa values are presented in fig. 1 . during cvvh arterial anti-xa tended to decrease in time (p = 0.05). the median ratio of postfilter to arterial anti-xa was 1.7 (iqr 1.4 to 2.1). there were large differences between patients; differences between groups were not significant, except for postfilter anti-xa at 6 h, which was significantly higher in group 2 (4 l/h) (p = 0.02) . anti-xa activity was not detectable in the ultrafiltrate. conclusions. critically ill patients receiving nadroparin during cvvh showed no signs of accumulation of anticoagulant activity, although extracorporeal removal of anticoagulant activity could not be demonstrated. apparantly, nadroparin is cleared by these patients despite renal failure. the differences in anti-xa between patients may be related to severity of disease. introduction. unfractionated heparin (ufh) is used as the first-line agent for anticoagulation of the extracorporeal circuit during continuous renal replacement therapy (crrt) in 96% of icus in the uk (uk) [1] . its use is monitored with serial measurements of activated partial thromboplastin time (aptt) or its ratio (apttr) in 83% of icus [1] . there is, however, considerable variation in practice [1] . anticoagulation is useful for prolonging haemofilter life and facilitates the provision of continuous therapy, but must be balanced against the risk of haemorrhage, which has been correlated with increasing apttr [2] . most icus in the uk use an apttr target of 1.5-2.5 [1] , despite recent guidance that a target range of 1.0-1.4 provides adequate filter life with less risk of bleeding [3] . objectives. to investigate the adherence to our local target range for ufh therapy (apttr 1.8-2.2) and the occurrence of over-anticoagulation in our patients. ]. there were 241 apttrs (39%) which were above our target range, and 43 incidences (7%) where the apttr was greater than or equal to 5.0. the apttr was greater than 1.4 on 512 occasions (83%). conclusions. this study was conducted in an icu which delivers crrt at a higher than average frequency [4] , and which consistently has a standardized mortality rate below the national average. despite this, there was wide deviation from our target apttr range and a considerable incidence of significant over-anticoagulation, which may place our patients at risk of haemorrhage. the vast majority of apttrs were in excess of recent guidance [3] . regional citrate anticoagulation (rca) may provide longer filter life with a lower incidence of bleeding [3] . its use is increasing worldwide [5] , though it is not commonly used in the uk [1] . we are investigating the possibility of introducing rca in our icu. in the meantime, we will set a lower apttr target for our patients. we prospectively studied patients who received cvvh from july 2005 to december 2008. age, gender, admission diagnosis, and apache-ii were obtained and the patients were divided into three groups: low dose heparin group, low molecular weight heparin group (lmwh), and no anticoagulation group (normal saline washing) based on assessment of coagulation status. for each circuit, circuit life, bleeding, platelet count, pt, inr, aptt, creatinine and urea were collected before and after crrt. results. seventy-seven critically ill patients with acute renal failure were treated with crrt and 226 circuits were observed. among these circuits, 50 received unfractionated heparin (ufh) anticoagulation, 78 received lmwh anticoagulation and 98 received no anticoagulation. the mean circuit life (44.6 ± 23.9 h) in low dose ufh group, was significantly longer than in lmwh (31.5 ± 21.3 h) and in no anticoagulation group (31.9 ± 20.8 h). there was no significant difference in baseline patient pre-crrt hb, creatinine and urea among three groups. the inr and pt and aptt in baseline were significantly higher in no anticoagulation group compared to the other two groups (p \ 0.05). the platelet count was significantly lower in the no anticoagulation group compared to ufh group and lmwh group in baseline and during crrt. there was no significant difference in the filter pt, aptt, among the three groups during crrt. the clearance of creatinine and urea during crrt were no significant difference among the three groups. bleeding complication secondary to crrt were no significant difference among the three groups. objectives. the purpose of the study was to assess the duration of time spent off therapy during the first five days of crrt in post-traumatic arf, and to identify the reasons for this. ullevaal between 1 january 1997 and 31 december 2007, were retrospectively reviewed. the hospital is the regional trauma referral centre for approximately 1.93 million adult ([18 years) persons. according to the local treatment protocol, dialysis filters were routinely changed after 72 h due to time-out. individuals were identified and data collected using several institutional registries. patients were grouped according to presence of rhabdomyolysis based on peak serum creatine kinase levels exceeding 10,000 u/l or not. categorical data were compared employing two-sided pearson chi-square test, whereas continuous data were analyzed utilizing two-tailed mann-whitney u test. results. 39 patients were included during the study period. during the first five days of therapy there was a total of 162 dialysis days, and the total number of pauses was 131. the median duration of crrt was 20.0 h per day, giving a downtime of 4.0 h per day. the number of pauses per day was significantly larger in patients with rhabdomyolysis compared to patients without rhabdomyolysis (71 pauses in 66 dialysis days vs. 60 pauses in 96 dialysis days, p \ 0.01). this resulted in a shorter duration of crrt in rhabdomyolytic compared to non-rhabdomyolytic persons (18.5 vs. 22.0 h per day, p \ 0.01). overall the reasons for pauses during crrt were filter clotting (53%), therapeutic procedures (24%), catheter problems (11%), filter time-out (7%) and diagnostic examinations (5%). patients with rhabdomyolysis had more pauses due to therapeutic procedures (32 vs. 15%, p = 0.02), whereas non-rhabdomyolytic persons had more pauses due to catheter problems (17 vs. 6%, p = 0.04) and filter time-out (13 vs. 1%, p \ 0.01). the number of pauses per day stayed relatively stable during the first five days of crrt, but the reasons for pauses changed during the study period. conclusions. this study indicates that trauma patients with rhabdomyolysis had more frequent dialysis pauses during the first days of crrt than those without rhabdomyolysis, resulting in shorter duration of dialysis therapy. the reason for this was more frequent use of therapeutic procedures, i.e. surgery and radiological interventions, in rhabdomyolytic compared to non-rhabdomyolytic persons. grant acknowledgement. the author is supported by institutional grants. introduction. treatment of acute pancreatitis is aimed at correcting any underlying predisposing factor and at the pancreatic inflammation itself. hypertriglyceridemia is an uncommon cause of pancreatitis. a serum triglyceride level of more then 1,000 to 2,000 mg/ dl is an identifiable risk factor. interestingly, serum pancreatic enzyme levels may be normal or only minimally elevated in such cases. severe necrotizing pancreatitis is associated with a high rate of complications and significant mortality. the reduction of triglyceride level to below 1,000 mg/dl effectively prevents further episodes of pancreatitis. this study aimed to determine the effectiveness of plasma exchange (pe) in reducing triglyceride levels during an acute attack of hyperlipidemic pancreatitis (hlp). methods. prospective, observational study including six patients hospitalized with hyperlipemic pancreatitis treated with plasmapheresis between 2005 and 2008 in the medical icu of a teaching hospital in malaga. demographic data, apache ii score, organ support needed and prognosis were prospectively collected. a total of 6 hypertriglyceridemic patients with the complication of acute pancreatitis received one or two consecutive sessions. mean age was 38 ± 11 years and mean apache ii was 12 ± 4. icu mortality was 33%. we performed 9 sessions. the development of multiorgan failure in patients with hyperlipemic necrotizing pancreatitis was associated with grave prognosis (33%), needed mechanical ventilation, vasoactive agent and renal replacement therapy. however, we had a good outcome in the majority (67%) with a effective reduction of triglycerides after the session of plasmapheresis (pe). four of six patients (66%) recovered completely in a single session. two patients developed intraabdominal abscess, requiring more than one consecutive session and surgical debridement of infected necrosis and died due to both septic shock and multi-organ failure. the respective mean removal rates during a single pe for triglyceride were 76%. conclusions. the best treatment of hypertriglyceridemic pa is a drastic reduction of tg-s to normal. experiences with plasmapheresis are limited. we report six patients of hypertriglyceridemic necrotizing pancreatitis with mildly elevated amylase and lipase, treated successfully with plasmapheresis. in summary, pe treatment is an effective method to clear lipids and enzymes from plasma in a single session for most hlp patients. the presence of multisystem organ failure appears to be a more important indicator of outcome than does the presence of infection. results. sixteen (61%) patients were female and ten (39%) were male. the median age was 42 years old. the median apache ii score was 16.5. mechanical ventilation (57%), vasoactive agents (42%) and renal replacement therapy (34%) were the most common forms of organ support needed. 71 sessions of plasmapheresis were performed. 10 (38%) patients had been diagnosed with thrombotic thrombocytopenic purpura (ttp), six (23%) patients had hyperlipemic pancreatitis, five (19%) patients had pulmonary-renal syndrome (prs), three (11%) patients had guillain-barré syndrome (gbs) and two (7%) had myasthenia gravis. we obtained a decreased in the values of apache ii score following the plasmapheresis performed. there were six death (23% mortality) due of the severity of the disease. the number of complications were minimal and commonly described in the literature and there was a low mortality as a result. conclusion. results indicate that the performance of plasmapheresis was on a heterogeneous sample of patients with neuroimmunological diseases, rheumatology diseases and hyperlipemic pancreatitis. we conclude that plasmapheresis is a safe treatment which can be made by the staff trained in intensive care in any moment with a wide spectrum of clinical indications and with a minimum adverse effect. the aim of the study is to evaluate that early treatment of septic shock with cpfa may improve patient outcome. methods. twenty septic patients who were admitted to the icu have been enrolled in this study. cpfa treatment was performed immediately after septic shock was diagnosed (early group 6 h after diagnosis). every patient had 3-4 cpfa treatments for 8 h with q blood = 220 ml/h, q ultrafiltration = 30 ml/(kg h) and q plasma = 20% of q blood. we measured the plasma concentration of procalcytonin (pct), blood lactic acid levels, crp, serum creatynine, wbc and pao 2 /fio 2 ratio. the apache ii score, hemodynamic parameters, norepinephrine dosage were evaluated before cpfa (t 0 ), t 1 (after first cycle), t 2 (after second), t 3 (after third cycle) and t 72 (after 72 h). introduction. the development of electrolyte disturbances in intensive care patients could be prevented by the use of better adapted dialysis fluids. a common problem is hypophosphatemia which has been shown to occur in up to 80% of the patients. correction by intravenous phosphate supplementation is known to improve respiratory muscles, cardiac index, oxygen delivery to tissues and insulin resistance. lately it has been reported that phosphate can be added directly to the dialysis fluid. this facilitates phosphate handling, but there is a risk of precipitation with calcium. an additional problem is that the amount of phosphate required to correct total body deficit varies and repeated serum measurements are needed to establish phosphate insufficiency. the process is time consuming and leads to treatment delay and excessive cost. objectives. this study evaluated the possibility to achieve and maintain normal phosphate balance over time by using a new phosphate-containing dialysis fluid. objective. the purpose of this study was to evaluate the impact of different dialysate and replacement flows in the acid-base balance of the blood. furthermore we tried to assess the way partial pressure of oxygen (po 2 ) in the blood is affected by high flow crrt. methods. this was a prospective observational study. thirty consecutive critically ill patients that were admitted in our icu and required crrt during their course were enrolled in the study. for each patient, blood flow, dialysate and replacement flow as well as ultrafiltration adjustments were performed by the responsible intensivist. any time that the clinical condition required a modification in any of these parameters, and after a period of time of no less than 1 h, a simultaneous blood sample was drawn from both the arterial and the venous part of the circuit and the samples were analysed by a blood gas analyzer. arterial and venus samples were then compared for differences in ph, po 2 and pco 2 concentration. results. in total we performed 113 measurements in 30 patients. mean patient age was 62.1 years, mean apache ii score was 18, mean icu stay was 22 days and mean crrt days was 6 days. overall, ph in the venous line of the circuit was higher, pco 2 was lower and po 2 was lower as well compared to the respective values in the arterial line of the circuit, with no difference reaching a statistical significance. concerning the blood flow, we observed that when using high hemodiafiltration flows the difference in oxygen partial pressure between the arterial and the venous line of the circuit was greater, but again it did not reach statistical significance. conclusion. the use of crrt may influence the po 2 in the returning blood. although we did not reach statistical significance in our study, there was a definite trend towards lower po 2 in the venous line of the circuit when high flow crrt was applied. introduction. renal failure (rf) is a common complication in critically ill patient and is associated with high mortality and has a separate independent effect on risk of death. the continuous renal replacement therapy (crrt) is physiologically superior; however, there is lack of strong evidence to prove a clinical benefit. hybrid therapies (sled) that combine the benefits of intermittent haemodialysis and continuous therapies have emerged in the past few years. objectives. the aim of this study was to assess what type of renal replacement therapy (rrt) used and relate them to severity of the illness and outcome we conducted a prospective observational study in three portuguese intensive care units (icu). patient demographics, type of rrt used, saps ii and sofa score at admission and when we started the rrt and outcomes were collected. we studied 84 patients who were treated with rrt for rf, with a median age of 58 years and a median saps-ii score of 52; 59 patients (70.2%) were treated with continuous replacement therapy (crrt), 14 patients (16.6%) with sustained low-efficiency dialysis (sled) and 11 patients (13.2%) were initially treated with crrt and latter with sled. aim. tetanus is traditionally treated with very high doses of diazepam and morphine. it often required prolonged periods of paralysis and was associated with very high mortality and prolonged periods of ventilation. magnesium sulphate (mgso 4 ), due to its effects on neuromuscular and autonomic system should be effective in controlling muscle rigidity, spasm and autonomic instability in patients affected with tetanus. we introduced an icu protocol using mgso 4 as first line treatment. we wanted to evaluate our patient outcome following the introduction of our protocol. we retrospectively analysed the effects of introduction of mgso 4 in our intensive care for management of tetanus. aim. electrolyte disturbances were often seen in patients in intensive care unit (icu). hypomagnesemia is not enough described but can be contributed in icu mortality. the aim of this study was to define the prevalence of hypomagnesemia in critically ill patients and to evaluate its relationship with duration of mechanical ventilation day, length of icu stay and mortality. a prospective study was done on 60 patients with respiratory failure admitted to the icu between 01.01.2008 and 01.07.2008. total serum magnesium level, electrolyte levels, albumin, total protein, and lactate levels were evaluated at the admission. patients demographic features, accompanying neurological and cardiac diseases, apache ii score, duration of mechanical ventilation, and the length of icu stay and mortality were recorded. at admission 27% of patients had hypomagnesemia. a positive correlation was found between serum magnesium and calcium level (p = 0.03), but there was no relationship between other laboratory tests. also there was no relationship determined between hypomagnesemia and duration of mechanical ventilation, and the length of icu stay and mortality (p [ 0.05). conclusion. electrolyte levels are important in critically ill patients. however routine monitoring of serum magnesium level is not necessary. so we should increase the case number and also evaluate the serum magnesium level with urine magnesium level to see the effects of hypomagnesemia. method. medical records of copd patients who underwent invasive mechanical ventilation (imv) were reviewed. the patients' age, sex, body mass index (bmi), apache ii scores at admission, previous diagnosis of hypothyroidism or hyperthyroidism, history of thyroid replacement therapy or antithyroid medications, and the serum thyroid stimulating hormone (tsh), free triiodothyronine (ft3), and free thyroxine (ft4) at admission were recorded. the primary outcome measure was prolonged mv (pmv), which was defined as dependence on mv for [7 days. the outcome and the relation between the serum thyroid levels were evaluated. results. ninety-five copd patients were included, 80% were male, with a mean age of 65.25 ± 11.3 years. bmi's of the patients were 29.62 ± 4.7 and the mean value of apache ii score was 27.36 ± 10.36. only two patients (2%) had a history of hypothyroidism. two more patient were diagnosed hypothyroidism at admission and treated with thyroid medications. the patients treated with thyroid replacement therapy were liberated from mv successfully. 50 patients (52.6%) could not be weaned. serum ft3 level (1.52 ± 0.10) of the patients, who could not be weaned, was statistically lower than other group who could be liberated (p = 0.034).however there was no statistical difference between serum ft4 and tsh levels and two groups. hypothyroidism is an uncommon cause of ventilator dependent respiratory failure with an incidence of 3%, but it is treatable, so it should be considered in patients who can not be liberated.more prospective studies are also needed to evaluate the significance of hypothyroidism in patients with respiratory failure and failure to wean. smoke inhalation injury represents an important prognostic factor in patients admitted in the hospital after smoke exposition. objectives. we determined whether initial antithrombin (at) levels help in diagnosis and prognosis of sepsis after smoke inhalation. smoke inhalation was diagnosed according to classical clinical and laboratory findings in 34 patients admitted in the hospital with suspected inhalation after smoke exposition. at levels, coagulation parameters (fibrinogen levels, prothrombin time (pt), activated partial thromboplastin time (aptt) and liver function tests were determined on admission and correlated each other and with outcome of the patients. . initial at and fibrinogen levels were significantly lower in patients with severe smoke inhalation compared to control (p \ 0.05). initial at levels were lower in the ones who developed septic complications with disseminated intravascular coagulation (dic) compared to those without dic (p \ 0.05). initial at levels were significantly lower in patients who died as compared to survivors (p \ 0.05 introduction. x-ray finding of pleural effusion is fairly common in icus. this may vary from mild to massive effusions and of different etiologies. epidemiological and outcome data for this icu problem are scarce in literature. the objective of this study was to find how common this finding is in our icu, their respective etiologies and any bearing on icu mortality. a single centre, prospective, observational study conducted in two mixed medical and surgical icus in kolkata, india. over six month period (october 2008 to march 2009) all consecutive patient admissions to these two icus were screened for a x-ray evidence of pleural effusion, either on admission or during their icu stay. as per icu protocol apache ii scoring were done in all patients. those with effusions were grouped according to etiology. finally in icu mortality were observed for those with or without an effusion. a total of 617 icu admissions were studied. among these 212 patients were found to have x-ray evidence of pleural effusion. median apache ii score was 19 (iqr 18-20) among the study population with predominant (86.8%) medical admissions. incidence of bilateral effusions were a total of 87 (41%). the common causes of pleural effusion include chronic kidney disease (n 91-42%), heart failure (n 33-15%), pneumonia (n 28-13%), post operative (n 23-10%), chronic liver disease (n 13-6%) and rest others (e.g. trauma, pancreatitis, pte, malignancy). the overall icu mortality was 81 (38.2%) and 61 (15.1%) in groups with and without effusion respectively with a p value of .0001, showing number of deaths in pleural effusion group were significantly higher. our study showed x-ray finding of pleural effusion quite common in icu patient population even many a times being bilateral. in this small study the overall icu mortality were also higher in pleural effusion group, but a wider multicentric study is needed. introduction. acute lung injury (ali) is a clinical manifestation of respiratory failure caused by lung inflammation and the disruption of the alveolar-capillary barrier. to prevent alveolar edema, it is of critical importance to preserve the physical integrity of the alveolar epithelial monolayer which is regulated by the balance between centripetal forces arising from cytoskeletal tension and cell-cell and cell-matrix tethering forces [1] . intercellular junctions, such as tight junctions are closely related to actin cytoskeleton-related barrier regulation. proteins of the coagulation cascade such as thrombin (thr)-that stiffens [2] and contracts [3] alveolar epithelial cells (aec)-or activated protein c (apc)-an endothelial barrierprotective agent [4] -could modulate this balance of forces in the epithelial monolayer. to study the combined effects of thr and apc on the barrier integrity through the tight junction zo-1 of aec by western blotting and immunofluorescence. methods. aec (a549) were incubated for 3 h with apc (50 lg/ml) or vehicle (control). subsequently, thr (50 nm) or medium was added to the cell culture. for zo-1 western blotting, cell lysates were first ultracentrifuged (100,000g, 30 min, 4°c) to obtain membrane and cytosol fractions. then the samples were subjected to western blotting and the amount of zo-1 fractions was calculated by densitometry. for zo-1 immunofluorescence, aec were grown on glass coverslips and fixed in 3.7% formaldehyde solution. zo-1 antibody was used to localize the tight junction and the zo-1 integrated optical intensity was then measured. . treatment with apc did not induce significant changes in any zo-1 amount of fraction protein analyzed by western blot. thr induced a *fivefold increase (543 ± 90% of control values) in zo-1 membrane fraction while no changes were detected in zo-1 cytoplasm protein content (97 ± 13% of control values). by contrast, apc concentration of 50 lg/ml showed a clear tendency to reduce the effects induced by thr on zo-1 membrane fraction (209 ± 65% of control values). for zo-1 inmunofluorescence, apc and thr treatments resulted in different patterns of zo-1 in the cell-cell contacts. after thr challenge cells showed discontinuous staining of zo-1 compared to untreated cells indicating a disruption of alveolar monolayer. conclusions. the increase in zo-1 amount of membrane fraction after thr challenge lends support to a protective mechanism avoiding cell-cell contacts disruption. treatment with apc reduced the increased zo-1 amount of membrane protein induced by thr suggesting an improvement of the barrier integrity in this model. (2001) interleukin-18(il-18) is said to be involved in organ injury. we investigated the il-18 values of septic acute lung injury (ali) and acute respiratory distress syndrome (ards) patients. the subjects were 38 patients during the 3-year period from 2004 to 2007 from whom it was possible to collect a blood specimen within approximately 6 h of the onset of septic ali or ards. their mean age was 67 years, and their mean apache ii score was 29. their sofa score was 13, and their mean pao 2 /fio 2 (p/f) ratio was 170. the p/f ratio was 246 in the ali group and 135 in the ards group. there were 4 cases (10.5%) in the 28-day mortality group, and 6 cases (15.8%) in the 90-day mortality group. the value of il-18 in died group was significantly higher than in survived group (1,649 ± 1,056 vs. 4,523 ± 2,798 pg/ml; p \ 0.05), and in the ards group also significantly higher than in ali group (2,467 ± 1,880 vs. 1,314 ± 800 pg/ml; p \ 0.05). these results suggested that il-18 may play an major role in progression of ards in respiratory disorder as multiple organ failure (mof). [1] . it is well-known that the pathophysiological mechanisms and factors involved in the liberation of no and the activation of inflammatory responses differ between aud and non-aud patients. objectives. the main hypothesis of this study is that ards patients with aud and non-aud differ in their response to the application of evidence based algorithms with respect to no response (aud patients are more frequent non-responders). patients with ards (meeting aecc criteria) were included in this ethically approved study. patients with severe chronic lung fibrosis and/or bridging for lung transplant were not included. patients were allocated to aud and non-aud patients. the auddetection was performed by the published algorithm [2] . statistical analysis: wilcoxon-mann-whitney and chi-quadrat test was used. results. so far, 23 patients with ards were included. prevalence of aud was 52% in our ards patients. baseline characteristics are given in table 1 . frequencies of no nonresponse, extracorporeal lung support and mortality are given in table 1 . frequency of no non-response was in tendency different: 67% in aud patients versus 36% in non-aud. overall mortality was 50% in aud patients versus 45% in non-aud patients. introduction. acute lung injury (ali) is a critical illness characterized by increased vascular permeability and impaired gas exchange leading to death in some cases. inflammation plays a pivotal role in the induction and maintenance of ali and is therefore therapeutic target to treat ali. rho, a small gtpase, is involved in the regulation of inflammation through the activation of recruitment of neutrophils to the site of inflammation and through activation of transcription factors such as nf-kb. we hypothesized that a rho kinase (rock) inhibitor, y-27632 may be beneficial to dampen the inflammatory response in ali. male sd rats were intravenously pre-treated with either saline or rock inhibitor (y-27632, 5 mg/kg). ali was induced by intratracheal instillation of 1 mg/kg e. coli lipopolysaccharide (lps). control rats received saline intratracheally. 24 h after the induction of ali, lungs were harvested and analyzed for myeloperoxidase (mpo) activity and expression of the proteins ijb, inos and enos. bronchoalveolar lavage fluid (balf) was used to assess total protein concentration as a measure of vascular permeability. pre-treatment with the rock-inhibitor resulted in significantly decreased levels of lps-induced mpo expression and prevented the upregulation of both lps-induced inos and enos expression. furthermore, lps-induced degradation of ikb was attenuated by pretreatment with y-27632. finally, y-27632 improved vascular permeability by decreasing the lps-induced protein concentration in the balf. conclusion. inhibition of rho-kinase decreases lung inflammation and vascular permeability in acute lung injury and may therefore be a good approach to treat patients suffering from ali. we hypothesized that due to the cyclic changes of pulmonary air content there are po 2 oscillations also in the mixed venous blood (pvo 2 ), potentially influencing pao 2 oscillations. in each of three healthy pigs of 30 kg, anesthetized and ventilated with constant minute volume we studied three different tidal volume settings (5, 8 and 11 ml/kg) resulting in different respiratory rates. a calibrated oxygen probe (fiber optic, fluorescence-quenching probe, foxy-al300; ocean optics, dunedin, fl, usa) was inserted into the pulmonary artery through a 4 fr catheter. the catheter position was previously controlled by pressure tracing. pvo 2 was sampled with temperature compensation at 10 hz with a multi frequency phase fluorometer (mfpf 100, tau theta, fort collins, co, usa) after a generated timestamp to synchronize with the electric impedance tomography (eit) signal (goettingen goemf ii, viasys healthcare, the netherlands) sampled at 13 hz. eit and pvo 2 were simultaneously recorded for 3 min during each tidal volume setting and analysed with and without low pass filtering at the heart rate. we obtained pvo 2 oscillations with amplitudes between 3 to 10 mmhg with the main frequencies matching the respiratory rate. ventilation with tidal volumes of 11 ml/kg provided higher pvo 2 amplitudes than ventilation with 5 ml/kg. these results are preliminary and the source of the measured pvo 2 oscillations is not clear. alternate backflow from the superior and inferior vena cava due to changes in intrathoracic pressures during mechanical ventilation may be responsible for these oxygen partial pressure oscillations in the mixed venous blood. conclusion. mixed venous oxygen partial pressure oscillates in accordance to the respiratory rate. whether arterial po 2 oscillations are due to cyclic recruitment and derecruitment of the lung or to corresponding mixed venous oscillations remains to be evaluated. [2] and in neonates [3] . to our knowledge this is the first validation of the model using a large cohort of samples from intensive care patients. aim. to assess the ability of the severinghaus equations [1] to estimate values for po 2 and so 2 in critically ill adult patients. methods. 34,191 sequential blood gas samples were analysed to validate the severinghaus oxygen dissociation curve, of these 14,228 measurements had a so 2 b 96.5% and were included in subsequent analyses. bland-altman plots were used to examine the agreement between measured po 2 and that calculated from the severinghaus equations, and between measured and calculated so 2 , both with and without correction for ph. the differences between measured and estimated values were analysed using paired t tests with a p value \ 0.05 considered significant. results. the severinghaus oxygen dissociation model accurately reflects the relationship between po 2 and so 2 observed in clinical samples. there is reasonable agreement between the measured and calculated values for po 2 and so 2 , with the majority of values falling between the lines of 95% agreement. there was a statistically significant difference between observed and calculated values of po 2 even when adjustment for ph was made (p \ 0.0001), however the mean difference between the groups was not clinical significant (5.4 mmhg when ph adjusted). there was also a statistical difference between measured and calculated values of so 2 (p \ 0.001), again, however, this difference may not be considered clinically significant (1.6%). patient data and severinghaus oxygen dissociation conclusions. the severinghaus equations accurately reflect the oxygen dissociation curve in critically ill adult patients and whilst they provide values for po 2 introduction. zinc (zn) is an essential trace element, which plays a role in many biological functions including immune function. development of respiratory infections and changes in respiratory tract cells may be affected by low zn levels. in critically ill children mortality of septic shock and degree of organ dysfunction were associated to low blood zn levels 1,2 . our aim was to study serum zn in the beginning of acute respiratory failure (arf) and its association to development of organ failures and day 90 mortality. during an 8-week study period (from 16 april 2007 to 10 june 2007) 958 adult patients with arf were treated in intensive care units (= finnali-cohort). after consent blood sample for zn analysis was drawn at baseline). samples were taken in zn-free tubes, freezen and stored in -80°c for analysis. all samples were analyzed with an atomic absorption spectrophotometry in the oulu university hospital laboratory. the range of normal values is 11-22 lmol/l. organ failures were assessed by daily maximal sequential organ failure assessment (sofamax) score. results. 551 serum zn samples were obtained during 24 h after the baseline with median time of 6 h. only 21 zn values were within and two over the normal range. median (iqr) serum zn levels were 4.7 (2.9-7.0) and 5.0 (3.3-6.9) lmol/l for survivors (n = 393) and nonsurvivors (n = 158), respectively, with no significant difference (p = 0.38). in patients with or without infection (pneumonia, respiratory infection or sepsis) during 48 h prior to arf, zn levels were 4.9 (2.8-6.9) and 5.0 (3.0-7.1) lmol/l, respectively (p = 0.09). zn levels were significantly lower (p \ 0.001) in patients with cardiovascular sofa 3-4 than 0-2, 4.3 (2.8-6.1) and 5.9 (3.7-8.0) lmol/l, respectively. a significant correlation of zn level and daily sofamax (spearman's q -0.151, p \ 0.001) was found (fig. 1) . conclusions. low serum zn levels were detected in almost all patients with arf. no association to day 90 mortality was detected to support the earlier findings with pediatric critically ill patients. however, we found a significant correlation to organ failure development in adult patients with arf. mountaineering is closely related to a range of adverse influences. the overriding factor that affects a climber may be the hypobaric hypoxia, which is compensated by hyperventilation and other adaptive changes in the pulmonary and systemic circulation. west (1993) theoretically predicted hypoxemia combined with respiratory alkalosis [3] , and low oxygen saturation (59…88%) has been observed on peak broad, karakorum [4] . lack of adaptation is known as mountain sickness (occurrence 27…47% [1, 2] ), which may be alleviated by acetazolamide. the importance of understanding pathophysiology of mountaineering is dictated by the gradual expansion of western consumer-oriented society to higher altitudes. the goal of our study was to obtain precise information on changes in arterial blood gas composition, acid-base status, and degree of hemoglobin desaturation relative to altitude. materials and methods. experienced athletes-four males between 32 and 57 years and 1 female 38 years attempted to ascend mt. makalu (8,463 m) in april-may 2008. acetazolamide 0,125 bid was used from april 21 till may 21. femoral arterial blood rather than radial arterial blood was analyzed before reaching base camp ( background. ventilator associated lung injury is a complication of mechanically ventilated patients. knowledge about pathological pathways comes from animal studies, which are necessary to generate hypotheses to be tested in humans. various experimental methods of inducing acute lung injury (ali) have been used in animal models. the results of animal studies and human research appear to be conflicting; however, this may be a consequence from the different animal models used as such for comparison. we hypothesized that effects on gas exchange, respiratory mechanics, histo-pathologic lung damage and systemic inflammation are depending on the model of ali used. in five groups of pentothal anesthetized rats acute lung injury was induced by either lung lavage or hydrocloric acid aspiration. rats were then ventilated with lung protective settings in pressure controlled mode with positive endexpiratory pressure (peep) of 4 cm h 2 o or breathing spontaneously with continuous positive airway pressure (cpap) = 4 cm h 2 o for 4 h. blood pressures, cardiac output, pulmonary mechanics and gas exchange were measured. results. the tidal volume was 9.3 ± 2.5 ml/kg in ventilated and 6.6 ± 1.1 ml/kg in cpap groups. respiratory rate and minute ventilation were constant in ali animals and controls, but showed variability in spontaneous breathing animals. only half of the cpap animals with ali survived [ 2 h. no significant differences were found for pco 2 , cardiac output or blood pressure between models, but mean arterial pressure decreased in ali. in the lavage and aspiration model, pao 2 was lower after induction of ali (151 ± 65 and 162 ± 34 mmhg, respectively) than controls, and increased in lavage (394 ± 65 mmhg) but not the aspiration model (198 ± 100 mmhg) after 4 h (p \ 0.05). dynamic compliance of the respiratory system decreased permanently after induction of ali to 0.22 ± 0.03 ml/cm h 2 o (lavage) and 0.34 ± 0.11 ml/cm h 2 o (aspiration) as compared to controls, which maintained at 0.66 ± 0.16 ml/cm h 2 o after 4 h. the lungs from five additional anesthetized, unassisted breathing animals, taken directly after induction, showed significant atelectasis, neutrophil infiltration and interstitial and alveolar edema (diffuse alveolar damage (dad) score 5.5 ± 1.0), as compared to control animals without ali (dad 3.0 ± 2.7 in ventilated, 3.8 ± 2.9 in cpap, respectively). the dad was higher in aspiration (7.2 ± 1.6) than in lavage (4.3 ± 2.2) induced ali, with no significant differences between ventilated and cpap animals. no hyaline membranes were observed. conclusions. anesthesia induces significant alveolar inflammation, which is partially reversible by use of peep. the ali model of acid aspiration induces persistent changes in gas exchange, respiratory mechanics and alveolar damage, which are more severe and consistent than those induced by the lavage model. background. acute lung injury (ali) is characterized by exaggerated inflammation and a high metabolic demand. mechanical ventilation can contribute to ali, resulting in ventilator induced lung injury (vili). a suspended animation-like state induced by hydrogen sulfide (h 2 s) may reduce metabolism and co 2 production, allowing for a lower minute ventilation to maintain gas exchange, thereby decreasing vili. h 2 s may also limit lung injury via reduction of inflammation. the effect of h 2 s-induced suspended animation on myocardial function is unknown. methods. in rats, vili was induced using a peak inspiratory pressure (pip) of 25 mmhg and zero peep. controls were ventilated with a pip of 13 and peep of 5 mmhg. respiratory rate was adjusted to maintain normocapnia. suspended animation was induced by infusion of a h 2 s donor, controls received saline. blood gases were drawn, bronchoalveolar lavage fluid (balf) was collected, lungs were removed. aortic flow was measured. statistics include kruskal-wallis and mann-whitney u. introduction. alveolar oedema is a hallmark of ards and ali. fluid clearance and the influence of anaesthetics on oedema resolution are poorly understood on a molecular level in the injured lung. oedema resolution is mediated by osmotic water reabsorption, following active sodium reabsorption via the apically located epithelial sodium channel (enac), driven by sodium-potassium-adenosin-triphosphatase (na ? /k ? -atpase). objectives. our aim was to investigate the influence of 1 mac (=2.2 vol%) sevoflurane on mrna and protein levels of enac and na ? /k ? -atpase in injured alveolar epithelial cells (aec). methods. primary culture of aec was stimulated with lipopolysaccharide (lps, 20 lg/ ml) and exposed to normal air containing 5% co 2 with or without sevoflurane. mrna levels were measured at 8 h using the taq-man real-time pcr method. additionally, proteins for western blotting were analyzed at 4, 8 and 24 h (n = 6). in the presence of sevoflurane mrna level of the a 1 -subunit mrna of na ? /k ? -atpase in control cells was downregulated by 15% (p \ 0.05). a-subunit na ? /k ? -atpase protein expression, however, was not influenced by lps or sevoflurane at all time points. mrna of c-enac was decreased by 30% in the presence of sevoflurane and by 70% upon stimulation with lps. in the lps-sevoflurane group downregulation was even more pronounced with 79% (p \ 0.05) after 8 h, but not statistically different from the lps group. on the protein level of c-enac protein expression a first change was observed at 24 h with a downregulation of 27% upon lps exposure (p \ 0.05). sevoflurane did not have an effect of this transporter protein. previous studies have shown that halothane decreases na ? /k ? -atpaseand sodium channel activities in alveolar epithelial type ii cells [1] . despite this finding for halothane, we could not see similar effects for the volatile anaesthetic sevoflurane. our results suggest that neither the driving force of alveolar oedema resolution, the sodium potassium atpase, nor c-enac, which is considered the rate limiting step in sodium coupled water reabsorption are influenced by sevoflurane and lps in an in vitro model of ards. to further characterize the impact of sevoflurane on water transport, functional analysis of these two transporters have to be performed. grant acknowledgement. objectives. we evaluated the effects of two nebulised sfa perfluorohexyloctane (f6h8) and perfluorobutylpentane (f4h5) at different dosages (1 ml/kg vs. 0.1 ml/kg) on pulmonary mechanics and gas exchange in healthy lungs. design. after approval by the local animal care committee, prospective, randomized animal study. subjects. thirty-five new zealand white rabbits. interventions: tracheotomised and ventilated juvenile rabbits were nebulised intratracheally with either a high or a low dose of two different sfa (f6h8 low/high and f4h5 low/high ) or saline (nacl). ventilated healthy animals served as controls (sham). arterial blood gases, lung mechanics, heart rate and blood pressure were recorded prior to nebulisation and in 30 min intervals during the 6-h-study period. results. immediately after starting aerosol therapy p a o 2 /f i o 2 -ratio and dynamic lung compliance decreased in all groups, with the exception of the f4h5 low group which behaved like the sham group. although p a o 2 /f i o 2 -ratio showed a continuous improvement in the other groups over time respiratory mechanics still remained impaired. high dose groups with nebulisation of liquid perfluorohexyloctane (f6h8 high ), perfluorobutylpentane (f4h5 high ) or saline (nacl) showed no significant differences neither in oxygenation, blood pressure nor in pulmonary compliance and resistance. in contrast to f6h8 high , there were no residues of f4h5 high detectable in bronchoalveolar lavage. regarding f4h5 low we were not able to detect any adverse effects on gas exchange or pulmonary mechanics. additionally, wet-dry-ratio of apical lung tissue samples revealed no significant edema. conclusions. high dose aerosolized sfa (1 ml/kg), either f6h8 or f4h5, equals effects of high dose inhalation of saline. when comparing the low-dose sfa-groups, there is a convincing discrepancy in favour of f4h5. f6h8 low impairs pulmonary function, whereas a low dose application of f4h5 (low) shows no interference. this may be due to the faster evaporation of f4h5. a new sfa-based pulmonary drug delivery system for lipophilic or water-insoluble substances could be developed on the basis of a low-dose application of f4h5. objects. hypertonic exposure reduces cell volume and thereby creates a relative excess of plasma membrane (pm). as a result the lipid bilayer of the pm can simply unfold with a minimal increase in lateral tension when an externally imposed shape change demands it. to test this hypothesis, we determined the effects of osmotic pressure on the susceptibility of deformation injury and pm wound repair. we measured deformation injury and repair responses of a549. cell culture media were consisted of 1x hmem and mannitol (v/v 50/50) with osmolarity of 300 (iso), 440 (hyper) and 170 mosm (hypo). cells conditioned with media were either stretched or deliberately injured with a scalpel. the fraction of wounded and healed cells was measured using a dual label method. . (1) exposure to a hypertonic environment tends to lower the susceptibility of a549 to deformation injury (5.35 ± 1.15% for iso, 4.36 ± 1.16% for hyper), while exposure to a hypotonic environment uniformly increases it (6.23 ± 1.51% for hypo) in stretch injury. introduction. the migration of polymorphonuclear leukocytes (pmns) into the lung plays a critical role in the development of acute lung injury (ali). adenosine receptor a3 (a3ar) is one of four g protein-coupled adenosine receptors that has been demonstrated to modulate pmn trafficking in various models of inflammatory disorders including sepsis and asthma. however, the role of a3ar in ali has not been investigated systematically yet. the objective of this study was to determine the role of the a3ar in a murine model of lpsinduced lung injury and in an in vitro transmigration system with human cells. methods. the migration of pmns into the different compartments of the lung was determined by flow cytometry in adult male c57bl/6 mice (wildtype [wt] ) and homozygous a3 receptor knockout (a3ko) mice. we used chimeric mice that were generated by transferring bone marrow between wild-type and a3 ko mice to differentiate the role of a3 on hemopoietic and nonhemopoietic cells. furthermore, microvascular permeability was assessed by the extravasation of evans blue and the release of chemotactic cytokines into the alveolar airspace was determined by elisa. paraffin-embedded sections of the lung were stained for pmns after lps inhalation to illustrate their accumulation in the lung. in a human in vitro assay, we quantified neutrophil transmigration across an epithelial monolayer (a549 cell line). in all murine in vivo experiments and in the in vitro transmigration assay, we assessed the effectivity of the specific a3-agonist cl-ib-meca. all statistical analyses were performed by using anova. p \ 0.05 was considered statistically significant. results. inhalation of lps significantly increased the number of pmns in wt and a3ko mice in all lung compartments. no differences in pmn counts were observed between wt, a3ko, and chimeric mice. pretreatment with cl-ib-meca led to a significant decrease of pmns in all lung compartments of wt mice but not in a3ko mice. pharmacological activation of a3ar diminished the lps-induced microvascular permeability in wt mice but not in a3ko mice. upon lps-inhalation, a3ko mice exhibited significantly higher levels of the cytokines cxcl1 und cxcl2/3 in the alveolar airspace than wt mice. in wt mice, pretreatment with cl-ib-meca reduced levels of tnfa and il-6 significantly. transmigratory activity of human pmns across an epithelial monolayer was reduced when a3 was activated in pmns. in contrast, pretreatment of the epithelial cells did not inhibit migration of pmns. introduction. lung overdistention during mechanical ventilation causes an increase in pulmonary vascular permeability, which is characterized by interstitial and alveolar edema secondary to a diffuse endothelial and epithelial injury. thrombopoietin (tpo), a humoral growth factor that stimulates the proliferation of megakaryocytes, has also been identified as a pro-inflammatory mediator in various clinical conditions. the receptor of tpo, c-mpl, is constitutively expressed on endothelial cells and may modulate the permeability of the endothelium. we investigated the contribution of tpo in the development of acute alveolar edema formation by mechanical stretch. in an ex-vivo model of mechanical ventilation (mv), lungs of c57bl6 mice were ventilated for 2 h with high stress pressure cycled ventilation (end inspiratory pressure = 20 cm h 2 o, peep = 0 cm h 2 o, i:e ratio = 1:1) and perfused with 4% bovine serum albumin rpmi medium at a rate of 1 ml/min, in the presence or absence of tpo (1 mg/ml). following ventilation, lung elastance was measured and protein concentration was analyzed in the bronchoalveolar lavage. data are mean ± se. mechanical ventilation (mv) to treat patients with ards or acute lung injury (ali) has the end objective to increase the dynamic functional residual capacity (dfrc), thus increasing overall functional residual capacity (frc). simple methods to estimate dfrc at the end of expiration for a given positive end expiratory pressure (peep) would provide a valuable metric to track and modulate therapy. however, such methods do not exist and current methods are time-consuming and relatively invasive. methods. this study utilizes a constant stress strain ratio for an individual patient's volume responsiveness to peep to estimate dfrc at any peep. the estimation model identifies two population parameters from clinical data to estimate a patient-specific dfrc, b and mb, where b captures physiological parameters of frc, lung and respiratory elastance and varies depending on the peep level used, and mb is the gradient of b versus peep. dfrc was estimated at 5 different peep values (5, 7, 10, 12, 15 ) cm h 2 o, and compared to the measured dfrc for 12 ali/ards patients to validate the model. patients and methods. in a 4 years period 24 patients (12 males, 12 females) with haematological malignancies were admitted in icu. malignancy type, reason for admission, haematological profile, requirement for invasive ventilation, bronchial and blood cultures and survival rate were recorded. results. patients suffered from: hodgkin's lymphoma (4), non-hodgkin's lymphoma (12), chronical lymphocytic leukaemia (3), acute myelogenous leukaemia (3) and multiple myeloma (2) . admission to icu was precipitated by: emergency surgical procedure (3), respiratory failure (14), sepsis (5), pulmonary oedema (1) and coma (1) . pulmonary infiltrates was the main finding in chest x-ray. bronchial secretions cultures were positives in 12 patients while blood cultures were positives in 8 patients. apache ii score ranged from 10 to 31 (average 21.85) and the icu days ranged from 1 to 36 (average 11.29). all the patients required invasive ventilation. all the patients with sepsis and serious neutropenia were died, while the total mortality was 15/24 (62.5%). conclusion. the admission of patients suffering from haematological malignancies in icu is associated with high mortality. immunosupression that renders them susceptible to infections, thrombocytopenia, and invasive ventilation are factors that contribute to this. early recovery of bone marrow and non invasive ventilation could improve the outcome in these patients. in liver transplanted patients, immunosuppressive therapy can increase the risk of infections and post-operative arf. nimv has been proposed as an alternative technique to reduce complications related to endotracheal intubation. the aim of our study was to evaluate nimv in liver transplanted patients, developing arf in the post-operative period. materials and methods. in this study we evaluated 24 liver transplanted patients, developing postoperative arf. measurements of respiratory and haemodynamic parameters were performed at baseline, after 1 h and at the end of the treatment. we evaluated intubation rate, nimv tolerance, length of stay in the icu (los), icu and hospital mortality. results. 18 (75%) out of 24 patients were successfully treated with nimv, while 6 (25%) failed and were intubated. we observed no significant differences among groups in gas exchange, but rr was significantly reduced in the success group during treatment (p \ 0.01). in both groups we found no significant differences in pao 2 /fio 2 initial improvement, but the success group showed a significantly higher rate of pao 2 /fio 2 sustained improvement (p \ 0.01). no significant differences between the two groups were found in terms of hours and days of nimv. success and failure groups were significantly different in saps ii (p \ 0.02) los (p \ 0.02), icu and hospital mortality (6 vs. 50%, p \ 0.001, 17 vs. 50%, p \ 0.03). reasons for nimv failure were not related to respiratory causes, but acute systemic causes such as septic shock and mods. conclusions. nimv can represent a valid alternative to invasive mechanical ventilation for the treatment of postoperative arf in liver transplanted patients; in nimv success patients reduced los and mortality can be expected. the influence of body posture on expiratory flow-limitation (efl) was estimated in 44 flowlimited, mechanically ventilated patients using the negative expiratory pressure (nep) method. a device especially designed and in build in an evita 2-draeger respirator allowed the application of a pressure equal to-5 cm h 2 o, starting at 8 ms after the onset of expiratory flow and sustained throughout the end of expiration. patients were considered flowlimited, if despite the application of nep part or the expiratory flow-volume curve was superimposed on the baseline curve. patients were studied in supine and in semi-seated position (45) at baseline and then 30 min after administration of bronchodilators (5 mg of inhaled salbutamol) with a nebulizer connected to the inspiratory port of the ventilator. supine position was significantly related to the occurrence of efl (p = 0.001). efl was abolished in 9% of our patients when changing from supine to semi-seated position, while in general a significant improvement of efl was noticed (from 41 to 36% of v t , p = 0.001). significant improvement of efl was achieved as well (p = 0.002) after bronchodilative therapy. peepi was the only variable significantly related to efl improvement when changing body posture from supine to semi-seated, while for bronchodilative therapy, none of the variables studied was significantly related to efl improvement. l. c. woodson 1,2 1 university of texas medical branch, anesthesiology, galveston, usa, 2 shriners hospital for children, anesthesiology, galveston, usa aims. laryngeal injuries are common among burn patients and can result in long term functional deficits. we have included careful laryngeal examination with our initial fiberoptic bronchoscopic evaluation of burn patients. the goal has been to allow early identification of laryngeal injuries and to facilitate laryngology consultations. methods. digital video recordings were made of upper airway endoscopies performed during airway management on admission or at the time of anesthesia for initial wound excision. these recordings were used to identify laryngeal injuries and to facilitate laryngology consultations. a wide variety of laryngeal injuries were identified and the digital recordings (which can be communicated by email) greatly facilitated laryngology consultations. in many cases these recordings guided therapeutic interventions and were often sufficient to avoid a separate exam under anesthesia. diagnosis of thermal necrosis provided an indication for early tracheostomy. identification of the mechanism of mechanical airway obstruction (e.g. supraglottic edema, fibrinous exudates, granulomas, vocal fold dysmotility) resulting in failure of a trial of extubation frequently guided therapy. early identification of posterior glottic damage provided more timely corrective laryngological interventions. educational use of these videos helps increase awareness of risks of laryngeal injury in thermally injured patients. aims. the most used weaning predictor f/v t ratio, is not a consensual predictor. when it was reported on the first time, this ratio was considered highly sensitive and specific. but others papers seems to disagree with it, suggesting other cutoff values to determine weaning failure in specific populations, as the elders. advanced age is thought to be an import associated factor in the intensive care unit (icu), but its effect on the weaning process is unclear. no studies have found strong evidence that conventional weaning parameters are reliable for this population. the widest used weaning criteria, f/v t ratio, does not seems to keep the same performance in this kind of population. the main purposes of this study were to identify the possible differences of the f/v t ratio measured in a spontaneous breathing trial, between an adult and an elderly group. we designed a protocol to study the variation, sensibility and specificity of the frequency-to-tidal volume ratio between an adult group (ag; up to 65 years) and an elderly group (eg; older than 65 years) in a daily weaning screening trial. methods. the study cohort comprised 239 patients ready to undergo weaning trial. the parameters studied were: weaning success (48 h of spontaneous ventilation after extubation), respiratory rate (f), tidal volume (v t ), frequency/tidal volume ratio (f/v t ), gasometric and ventilatory parameters. the weaning method was spontaneous breathing trial (sbt). measurements were made in the beginning of sbt (t0) and 30 min after (t30). we analyze possible differences in the sensibility and specificity of the f/v t ratio between elderly and adults and compare with previous values already published. the chi-square test, anova and the t test were used in the statistical analysis. weaning success was 74.8% in eg and 78.1% in ag (p = 0.552). the baseline characteristics were similar. comparisons of ag and eg at t0 and t30 showed statistical differences in weaning criteria: f, v t and f/v t ratio. conclusion. weaning success in our study was low, but similar to the described in other trials. elderly patients showed higher f and lower v t . consequently, f/v t ratio was lower too. the area under the roc curve for f/v t ratio was smaller than already published. results. 562/645 (87%) were successfully extubated and 83 patients required re-intubation. the demographic data showed no differences in age, bmi, apache ii, icu admission diagnosis or sex distribution between groups (table 1) . patients who failed extubation had small but statistically significant differences in vital capacity (vc), peak negative inspiratory pressures (pnip), pao 2 /fio 2 ratios (pf) and were ventilated longer prior to extubation. paradoxically, patients failing extubation had positive end expiratory pressures that were statistically but not clinically significant higher. the ratio of respiratory rate to tidal volume (f/vt) was not significantly different. patients failing extubation were also more likely to have weaker cough, gag, level of consciousness as measured by glasgow coma scale and more secretions ( table 2 ). having no cuff leak did not predict failure of extubation. the most common reasons for reintubation were secretion retention and/or absence of cough (90%). pressure support ventilation (psv), a widely used assisted mode, has the purpose to avoid diaphragm disuse allowing the patient to generate spontaneous inspiratory efforts optimizing comfort and work of breathing. however, still little is known about the individual response of respiratory muscles under these conditions. we hypothesized that respiratory muscles of patients ventilated with clinic psv might result, at least sometimes, excessively unloaded. we performed an observational study in the intensive care unit on patients ventilated with psv set by the clinician in charge. twenty intubated, mechanically ventilated patients (71 ± 9 years old) during the weaning phase entered the study. the patients had no sedation at least for the last 24 h. respiratory timing, tidal volume (v t ), peak airway pressure (paw peak ), electrical activity of diaphragm expressed as percentage of its maximum (edi/ edi max ), inspiratory (ptpes) and diaphragm (ptpdi) muscle effort were measured during 30 min of clinic psv. results. we found that seven out of twenty patients generated a negative pes swing only during the psv inspiratory triggering phase (psv t ) in comparison with the remaining 13 patients in whom pes was negative throughout most of the mechanical breath (psv n ). in the psv t group, pes swing was either flat or positive after inspiratory triggering. therefore, in the psv t group both ptpes /min and ptpdi /min were fivefold lower than normal values. v t / predicted body weight (pbw) was significantly higher in the psv t versus psv n group (see table 1 ). during weaning with psv: (1) a significant number of patients (35%) showed a pes shape similar to that observed during pressure assist/control modes, and inspiratory muscle effort abundantly lower than normal, both indicating excessive inspiratory muscle unloading; (2) among the variables used to set psv, only a high v t /pbw (higher than 8 ml/kg) hallmarked excessive unloading; (3) due to the ample prevalence of the phenomenon, the question whether high levels of inspiratory muscle unloading can cause detraining and prolonged mechanical ventilation merits an answer from further research. introduction. the liberation from mechanical ventilation (mv) should be done as soon as possible in order to avoid complications and the risks associated with prolonged unnecessary mv, such as ventilator-associated pneumonia, ventilator induced lung injury, and increased icu and hospital stay. this procedure should be carried out properly and safely. objective. evaluate the extubation success rate, mv time and weaning time using a daily weaning screen followed by a spontaneous breathing trial (sbt). patients who were ventilated for more than 24 h were subject to this procedure, which was carried out by respiratory therapists. methods. in our icu, between february and august of 2008, all intubated patients who were ventilated for more than 24 h underwent a daily weaning screen, which contained variables such as hemodynamic, gas exchange, consciousness and resolving the need for mv. if these variables were stable, these patients were submitted to a sbt and were extubated if they did not show any signs of respiratory discomfort or hemodynamic changes for at least 30 min. conclusion. the use of a daily weaning screen followed by a sbt was associated with a high extubation success rate and a very short weaning duration with 11% of unsuccessfully extubations. c. chatt 1 , d. pandit 2 , g. raghuraman 2 1 birmingham heartlands hospital, critical care, birmingham, uk, 2 birmingham heartlands hospital, birmingham, uk aim. the aim of the study was to assess the impact of pmv on the course of weaning in mechanically ventilated patients. we wanted to assess the optimal pressure support at which pmv can be initiated which would enable prolonged use of pmv without affecting the duration of respiratory support. method. data on all patients who were mechanically ventilated for greater than eight days were obtained from icnarc database, who underwent tracheostomy as part of their weaning process. satisfactory level of pressure support was achieved (between 10 and 15 cm h 2 o) pmv was introduced into the patient's breathing circuit and spontaneous ventilation was attempted. we applied mann whitney u tests for parametric data, fisher exact tests for non-parametric data and anova was used to compare the three groups with different pressure supports at initiation of pmv. a p value \ 0.05 be statistically significant. results. 157 patients who were ventilated for greater than eight days identified. of these, 29 patients were excluded because they did not have a tracheostomy during their period of ventilation. of the remaining 128 patients, pmv was used in 92 patients (72%). there were no significant differences between the demographic data (sex, age) or the data on admission to intensive care (apache ii score, ratio of medical to surgical patients) and duration of mechanical ventilation between the two groups. however, there were significant differences in the mortality, total respiratory support days after tracheostomy and length of stay in intensive care and length of hospital stay between the two groups. in the group on pmv, no record of aspirations was found documented on the intensive care charts. in patients in whom pmv was used (n = 92), pmv was initiated at cpap (continuous positive airway pressure) in 40 patients (44%). 30 patients (33%) had pmv initiated at a pressure support of b10 cm h 2 o and in 21 patients (23%) pmv was initiated at a pressure support of [10 cm h 2 o. there was a significant difference in the duration of mechanical ventilation post tracheostomy (p = 0.04) and the length of hospital stay (p = 0.02) between the two groups, with the cpap group being ventilated for a shorter duration but with a longer stay in hospital the same difference was shown when comparing three groups of pressure support when pmv was commenced (cpap, pressure support b10 cm h 2 o and pressure support [10 cm h 2 o). however, in the pressure support [10 cm h 2 o group we observed that the duration of use of pmv was lower than in the other two groups despite longer duration of mechanical ventilation and total respiratory support days. although this was not statistically significant, it could be clinically significant. our study suggests that use of pmv at pressure support b10 cm h 2 o could increase the duration of its use without affecting the length of mechanical ventilation. we would therefore recommend weaning to a pressure support b10 cm h 2 o before pmv is commenced in the acute setting. no data are available concerning the oxygenation target to aim during the weaning phase from mechanical ventilation. also, in opposition to ards patients there is no clear recommendation for the upper limit of spo 2 to maintain during weaning. this study is part of a research project on peep and fio 2 settings automation during mechanical ventilation. methods. this observational study was designed to assess the spo 2 target aimed during the weaning phase of invasive mechanical ventilation (fio 2 b 0.60 and peep b 10 cm h 2 o). patients were recruited in icus from several countries (canada, france, italy, tunisia, argentina). the following data were prospectively collected by the respiratory therapists at each round during a 3 months period: spo 2 , fio 2 , peep level, ventilatory mode, anatomic site of the pulse oxymetry sensor, quality of the spo 2 signal. results. data from 3 centers (5 icus) from quebec city, canada, and 1 center from créteil, france are available. 687 patients were prospectively included. 12,907 observations were performed. the mean level of fio 2 was 0.43 ± 0.10 with fio 2 c 0.50 and 0.70 in 24.8% and 3.5% of observation times respectively. the mean level of peep was 5.6 ± 1.6 cm h 2 o and was below, equal or above 5 cm h 2 o in 3.1, 71.5 and 22.2% of the cases respectively. the most frequent ventilatory modes were pressure support (56%), simv (27%) and acv (12%). the pulse oxymetry sensor was applied on a finger of the hand in 94.5% of the cases and was deemed of good quality in 91% of the time. the mean spo 2 was 97.1 ± 2.7% for the whole population and was 97.3 ± 2.8% for patients with fio 2 c 0.70. spo 2 was higher than 95% in 66% the observations. desaturation with spo 2 below 92% were recorded in 2.6%. the spo 2 signal was deemed available by the bedside nurse in 94.4 ± 10.2% of the time. conclusion. this study demonstrates that spo 2 levels may be maintained at high levels unduly. this may have an impact on the weaning phase of mechanical ventilation. this study also shows that the spo 2 signal availability was high enough to be used in a closed-loop oxygenation system. introduction. automated weaning systems are viewed as a challenge to weaning decision-making autonomy by some clinicians. clinician perceptions of the utility of such systems may influence uptake in to practice. to assess the perceived utility of the automated weaning system, smartcare/ ps. a survey was generated based on comprehensive literature review and 1-year's experience using smartcare/ps. survey pilot testing was conducted with 5 senior clinicians experienced in smartcare/ps weaning in an independent icu. questions addressed perceived system usability and appropriateness of automated weaning, system benefits and disadvantages, as well as patient indications deemed suitable and unsuitable for smartcare/ps weaning. participants were also asked to indicate if they would continue using smartcare/ps on trial completion. the survey was administered to clinicians on completion of a randomized controlled trial conducted to compare smartcare/ps to non-protocolized weaning 1 . of 85 staff surveyed, 34 surveys were returned by 27 nurses and 7 doctors (response rate 40%). eight respondents had no experience with smartcare/ps despite the 1year trial duration, leaving 26 surveys with evaluable responses. the majority of respondents perceived smartcare/ps was easy to activate (19/26, 73%) and to use once activated (23/26, 88%). the system was observed to wean appropriately by 13/26 (50%) respondents; 13 experienced smartcare/ps to wean inappropriately. comments on inappropriate weaning identified clinically unacceptably increases of pressure support (ps) for patients with profound tachypnea and complicated lung pathology. smartcare/ps' ability to reduce the overall duration of weaning was questioned by all but 3/26 (12%) respondents. ps adjustment according to patient requirements was the most frequently perceived benefit (15/26, 58%). most respondents did not perceive any advantage of smartcare/ps for patient comfort 22/26 (86%), assessment frequency (15/26, 58%) and automated control of weaning (15/26, 58%). less control over weaning was the most regularly cited disadvantage of smartcare/ps (12/26, 46%). system issues such as program abortion without identifiable reason and mandatory peep reduction prior to a spontaneous breathing trial to assess readiness for separation were less frequently cited disadvantages [10/26 (38%) and 7/26 (27%) respondents respectively]. most respondents (18/26, 69%) felt smartcare/ps was best suited for weaning postoperative patients and should be avoided for patients with neurological dysfunction (18/26, 69%). only 4/26 (15%) respondents stated they would not continue to use smartcare/ps. clinicians demonstrated moderate acceptance of smartcare/ps. more work is needed to identify those patients more likely to benefit and confirm the overall utility of smartcare/ps as a weaning tool. introduction. physician approaches to ventilation withdrawn varies among physicians whereas the prompt recognition of respiratory failure reversal and usefulness of weaning protocols in reducing duration of mechanical ventilation (mv) have been largely demostrated as nursing staff attend patients 24 h a day its leadership in this process can be effective and safe. objective. to demonstrate that a nurse-directed protocol to withdraw mv could reduce a 25% its duration. prospective sequential study performed in two periods. during de first period (6 months) data concerning weaning definite criteria appearance, duration of mv, reintubation or need for nonninvasive ventilation (nimv) and demographic data were collected to all mechanically ventilated patients blinded by attending nurses and physician. after a three months phase of staff training there was a second 6 months period where weaning criteria were checked al each nurse working shift during the first 10 days of mv. when criteria were fullfilled a 120 min of spontaneous breathing trial was perfomed and tracheal tube removed if there were no intolerance criteria. same data as the first phase were collected. we used mann-whitney 0 s u test to compare mv duration, time to reach weaning criteria (trwc) and extubation delay (mv duration minus trwc). weaning failure was compared using x square. data are presented as median (25-75 percentile). results. 203 patients were screened (114 in the first period and 89 in the second) but only 164 patient reached weaning criteria in the first 10 days (96 in the first period 68 in the second), 63.5% men, aged 69 (55-76) years. aim of this study is present our experience about elective bedside pdt with the blue-rhino kit over an 1 year period, in order evaluate its efficacy in terms of intraoperative and postoperative complications. patients and methods. the study included a total of 73 consecutive icu patients requiring tracheostomy. all pdt were performed by icu staff physicians at patients' bedsides, using a blue rhino kit. the following data were recorded: age, sex, simplified acute physiology score (saps) ii, fraction of inspired oxygen (fio 2 ) before the tracheostomy, days on mechanical ventilation before the tracheostomy, bleeding, tracheal tear, subcutaneous emphysema, pneumothorax, wound infection, hypotension, lowering sao 2 during the procedure, inability to complete the procedure, and procedural mortality. distance follow-up included fiberoptic bronchoscopy to evaluate tracheal stenosis. results. there were a total of 2 (2.73%) complications (tracheo oesophagel fistula and bleeding). forty -one patients died in the icu (41%), although none of these deaths were related to technique complications. mean duration of the procedure was 7.5 ± 1.2 minutes. the pdt performed at bedside in the icu, using the blue rhino kit is a simple and safe procedure that offers many advantages in terms of safety and efficacy. objectives. questioning the need for several specialized physicians or extra assistance to perform a single percutaneous tracheostomy using fibrescopic tracheoscopy, we performed a prospective study into the complication rate of percutaneous tracheostomy without tracheoscopy on our mixed medical and surgical icu. , 99 consecutive patients were included after having received a percutaneous tracheostomy. indication for tracheostomy was always a long anticipated duration of mechanical ventilation. if no contra indications were present, percutaneous tracheostomy was performed. if contra indications against the use of percutaneous tracheostomy without tracheoscopic control were present, tracheoscopy was performed to ensure maximum patient safety. the mean age at the time of receiving a tracheostomy was 66.9 (17-83) years. the cohort consists of 69 male patients en 30 female patients. only ttwo percutaneous tracheostomy were performed under fibrescopic control due to contra-indications for an uncontrolled procedure. in 99 procedure, sixteen minor, and no major complications were encountered. this resulted in a 16.2% minor complication rate. conclusions. the number of complications in our group is approximately the same as those which are suggested in international literature where tracheoscopy was performed during percutaneous tracheostomy. none of the complications encountered could have been prevented by the use of tracheoscopy. therefore we postulate that in the hands of an experienced team and in adherence to strict guidelines, percutaneous tracheostomy can safely and successfully be performed without tracheoscopy. objective. to assess the risks and complications associated with the bedside pdt in our 9 years experience of over 1000 pdts in icu. pdt is a relatively newer technique and has been introduced as an alternative to open tracheostomy as a safer and convenient procedure. however, the risks and complications of the pdt have not been highlighted in the icu of a developing country. a retrospective analysis of the data gathered from patients undergoing pdt was done in a 40-bedded tertiary level multidisciplinary icu of a teaching hospital. the data was collected between april 2000 and march 2009. all intubated patients with indications for elective tracheostomy, as well as patients who required emergency tracheostomy were included in the study. demographic and other clinical details of the patients who underwent pdt were collected. griggs [1] technique was most commonly adopted while other adopted techniques were ciaglia [2] , white tusk/blue rhino tapered dilator and percutwist technique. a total of 1,010 pdts were done in 1,005 patients, over a period of 9 years. of the 1,005 patients 710 (71%) were males and 295 (29%) were females. the mean age of patients was 53.9 years. the average duration of intubation before pdt was 9.3 days. 970 (96%) pdt were done bedside in icu while 40 (4%) were done in wards, coronary care unit, high-dependancy unit and liver transplant unit. griggs technique was adopted in 841 (83.26%), ciaglia in 92 (9.12%), white tusk/blue rhino tapered dilator technique in 69 (6.83%) and percutwist technique in 8(0.80%) patients. long-term ventilation was the most common indication in 602 (59.60%) followed by airway protection in 547 (54.14%), facilitation of weaning in 75 (7.42%) while airway obstruction/difficult intubation was observed in 5 (0.50%) patients. pre-procedure coagulopathy was observed in 126 (12.47%) patients, 11 (1.08%) were morbidly obese while 12 (1.18%) required emergency tracheostomy. no complications were observed in 890 (88.11%) patients. procedural complications were seen in 120 (11.88%) patients. bleeding from the site was the leading complication affecting 60 (5.94%) patients. difficult tube placement was seen in 25 (2.54%) patients, premature extubation in 12 (1.18%), false passage in 4 (0.39%), guidewire dislodgement in 4 (0.39%), subcutaneous emphysema in 5 (0.50%), arrhythmia in 3 (0.29%) and bleeding requiring transfusion was seen in 6 (0.59%) patients. no procedure related mortality was observed. conclusion. on the basis of this large single centric study we found that pdt is a safe, reliable and convenient procedure which can be easily performed bedside by experienced intensivists. results. of the 16 recipients who underwent pdt, 11 were liver, 3 kidney and 2 heart transplant recipients. the respective mean values for age, weight and apache ii score were 42.7 ± 14.7 years, 61.5 ± 11.9 kg, and 28 ± 7. all pdts were performed at bedside by experienced staff anesthesiologists with direct bronchoscopic guidance. in all cases, the indication for pdt was prolonged mechanical ventilation due to acute respiratory failure. the mean time from transplant to pdt was 13 ± 22 months and the mean duration of endotracheal intubation before pdt was 3 ± 2 days. twelve patients had coagulopathies. the calculated lung compliance and pao 2 :fio 2 ratio improved after pdt (39.3 ± 27.6 vs. 54.4 ± 37.6 ml/cm h 2 o, p = 0.038 and 194 ± 55 vs. 248 ± 80, p = 0.005 respectively). transient hypoxemia (n = 2) and mild extratracheal bleeding (n = 2) were the only early complications. there were no procedural failures and no pdt-related late complications and deaths. conclusion. the results suggest that percutaneous dilational tracheotomy is an efficacious and safe technique for prolonged airway management with improved ventilatory mechanics in solid organ transplant recipients. a. vianna 1 , g. cabral 1 , r. azambuja 1 , g. carleti 1 , t. balbi 1 , g. pereira 1 1 clinica são vicente, rio de janeiro, brazil aims. we studied diferent aspects of tracheostomy procedures performed in intensive care units (icus) located in the municipality of rio de janeiro and compared them with the medical literature. a questionnaire was elaborated and sent through email to the coordinators of every icu in the city of rio de janeiro in the period of july to august 2008. the questionnaire was sent to the coordinators of the 87 icus located in rio, and was answered by 39 (44.82%) of them. among the studied icus, 11 (28.2%) are public, 25 (64.1%) are private, and 3 (7.7%) are part of university hospitals. 36 (92.3%) are medical/ surgical, 2 (5.1%) are medical, and 1 (2.6%) is a surgical unit. the average number of beds is 16 ± 7.3. the decision to perform the procedure is taken by the icu team in 38 (97.4%), by the patient's primary team in 11 (28.4%), and by both in 10 (25.6%). tracheostomy is performed by a surgeon in 36 (92.3%) units, by an intensivist in 2 (5.1%), and by both in 1 (2.6%). the procedure is performed at the bedside in 35 (89.7%) of the icus. the most frequent indications for tracheostomy are: prospect of prolonged mechanical ventilation, coma, and airway protection. 53.8% of are performed between the first and second week of mechanical ventilation, and 41% between the second and third week. control chest x-ray is performed in 87.1% of the units. surgical tracheostomy is available in all the studied units. only 7 (17.9%) units perform percutaneous tracheostomy. the reasons given for the preference for surgical tracheostomy were the lack of a qualified team for performing the percutaneous tracheostomy or material needed for this procedure. all icus that perform the percutaneous procedure use the ciaglia technique with bronchoscopic guidance. late followup is performed in 15 (38.5%) of the studied units. the study showed great differences between the tracheostomy protocols used in the hospitals of rio de janeiro and those found in the medical literature. in particular, the use of percutaneous tracheostomy is still infrequent in the icus of rio. on all the patients, aged at least 80 years or more, admitted to our postoperative icu since january 1994 through december 2006, we collected demographic profiles, operative data and short and long-term outcomes. spss 13.1 was used for statistical analysis and p \ 0.05 was considered the level of significance. a total of 428 patients (4.8%), 53.5% males and with a median (iqr) age of 83 (82-85) were admitted to our post-operative icu over the study period. iddm was recorded in the 19.6% of the population, copd in the 13.8%, hypertension in the 64.5%, chronic renal failure in the 21.7% and arteriopathy in the 37.9%. out the total population, 51.6% of patients, with a median (iqr) pre-operative crs of 7 (5-8) underwent a coronary-artery bypass grafting (cabg) surgery, whereas 20.7% of them, with a preoperative median (iqr) nyha of 3 (2.25-3) needed a valve replacement (vr) and 16.1% of them combined (cabg ? vr) operations; moreover, 11.6% of patients underwent other type of cardiac and aortic surgery. overall median (iqr) post-operative mechanical ventilation length was 15 (10.75-22) hrs. while no statistically significant difference was recorded in terms of mv duration among the four surgical groups. overall recorded mortality rate was 10%, with the lower 7.7% for cabg and the higher 12.6% for vr (p = 0.388). kaplan meier curves showed no differences in survival likelihood at 28th (log rank = 0.404, p = 0.817) 60th (log rank = 0.707, p = 0.702) and 90th (log rank = 0.742, p = 0.690) days after surgery among the different surgical groups. conclusions. the outcome after heart surgery in octogenarians is excellent; the operative risk is acceptable and the late survival rate is good. therefore, cardiac surgery should not be withheld on the basis of age alone. introduction. re-do cardiac operations have been reported to be increasing in incidence and are associated with a higher operative risk [1] . this study aimed to determine the impact on intensive care provision. methods. data from 15,346 procedures spanning twelve years (april 1996-march 2008) was examined. the 798 re-do operations were further analysed by gender, age, pathology (new, progressive, combined) , duration between procedure, theatre time, length of stay, complications and mortality. as the number of cardiac operations performed has increased over the twelve year period, the relative incidence of re-do procedures have remained stable at 5.2%. operative length at re-do was significantly longer (mean 316 min vs. 253) however anaesthetic time pre surgical incision was not significantly increased. subsequent length of stay on the intensive care or high dependency unit increased by 25% (mean 3.9 vs. 4.9 days), with higher complication rates affecting all systems (except post operative myocardial infarction). renal and pulmonary complications showed the most significant increases. renal related complications occurred in 14% and pulmonary in 20.1% of cases which represents an 80 and 25% increase on first operation rates. infection rates were also significantly increased at double that of the initial procedure. the total hospital stay was found to be 24% longer (by 11.5 vs. 14.2 days, respectively) while in hospital mortality increased from 2.4% at initial procedure to 7.6% at re-do. mortality rates were further elevated in the presence of renal failure post operatively, as re-do valve mortality increased from 6.3 to 20% and re-do cabg from 3.1 to 50% in this subgroup. conclusion. these results, combined with the stability of percentage re-do surgery over the twelve year period, enable specific planning and management of intensive care provisions. the knowledge of extended theatre times and subsequent stay in intensive care/high dependency units has a further impact on the throughput of routine cases. the data also highlights the increased costs associated with these patients, as they not only require longer hospital stays but also suffer increased complications requiring more investigations and interventions. specific costing therefore applies to this subgroup of intensive care patients. objectives. we aimed to assess hrqol at 30 days after surgery in relation to preoperative hrqol. we compared patients with decreased hrqol to patients with unchanged or increased hrqol to identify disparities between these two groups. a prospective cohort study including patient scheduled for cardiac, vascular, abdominal and orthopedic surgeries in a tertiary hospital was performed. patients filled-out a hrqol questionnaire (sf-12) the day before surgery and 30 days after. preoperative, intraoperative, postoperative data were collected. changes of pre-and postoperative physical component summary ( simultaneus kidney-pancreas transplantation is the best treatment option for type 1 diabetic patients with chronic kidney disease. currently, the medical and surgical complications have decreased significantly, although these represent a high risk of morbidity and mortality in the short term. objectives. this study sought to investigate the incidence of medical and surgical complications, the clinical characteristics and prognostic factors influencing graft and patient's survival in a recent cohort of pancreas-kidney recipients. patients and methods. the present study included 40 patients who received simultaneous pancreas-kidney transplantation in our center from january 2004 to february 2008. we studied demographic, clinical and immunological characteristics of patients, and surgical and medical complications during his admission to intensive care unit. results. the average age of recipients was 38.8 years and mean age of donors was 24.8 years. the median cold ischemia time was 10.7 h (95% confidence interval 5-14). the average stays on the waiting list was 98.5 days. 75% of patients were extubated within the first 10 h. 52% of patients required transfusion during their icu admission, amine infusion was started at 25% patients in the early hours. during follow-up, surgical reintervention in the immediate postoperative occurred in 30% of the patients. major surgery complications reported in the literature are graft thrombosis, although in our serie there have been only 2 kidney graft thrombosis and 2 pancreas graft thrombosis. only 5% of patients died within the first 3 months posttransplantation surgery. conclusion. surgical complications after pancreatic transplantation remain a significant concern. hence we our results add further evidence to support the notion that the double and simultaneous pancreas-kidney transplant is in fact the treatment of choice in selected patients with end-stage renal failure due to type 1 diabetes mellitus. a. shono 1 , t. mihara 1 , y. murakami 1 , j. ota 1 , f. kono 1 , y. saito 1 1 shimane universiy hospital, anesthesiology, izumo, japan pulmonary catheter is widely used for cardiac surgery. the complications of indwelling pulmonary catheter, such as perforation of pulmonary artery, pulmonary embolism, are well known. however, the thrombosis associated with introducer sheaths has received much less attention. we evaluated the incidence and risk factors for internal jugular vein thrombosis (ijvt) associated with introducer sheaths for pulmonary catheter after cardiac surgery. methods. the patients who underwent cardiac surgery and insertion of introducer sheaths (8.5f) at right internal jugular vein (ijv) were included. ultrasonographic evaluations of ijvt were performed prior to insertion and daily until introducer sheaths removal. we investigated demographic data, underlying disease, length of surgery, use of cpb and iabp, complications during cannulation and duration of catheterization. coagulation status (pt, aptt, platelet count, d-dimer) and cardiac index at before and after surgery were also recorded. the student's t test, v 2 test, and fisher's exact test were used for statistics and p value of .05 was considered significant. results. 53 patients were included in this study. mean age of patients was 70 ± 9 years (range 47-88), mean duration of catheterization was 4 ± 2 days. 19 (35.8%) patients developed ijvt which occurred only one day after insertion. the incidence of ijvt was related to presence of underlying disease (relative risk, 3.9; 95% confidence interval, 1.02 to 14.85) and was unrelated to emergency operation, the use of iabp and cpb, number of insertion attempts. there were significant differences between patients with or without ijvt in duration of catheterization (4.5 ± 2.2 vs. 3.4 ± 1.7 days, p = 0.044), cardiac index at 1 day after surgery (2.5 ± 0.4 l/(min m 2 ) vs. 2.9 ± 0.5 l/(min m 2 ), p = 0.002), the value of d-dimer at 1 day after surgery (5.3 ± 5.3 vs. 2.7 ± 3.1 lg/ml, p = 0.044). no clinical symptoms related to ijvt were found in observation period. our results demonstrated that ijvt associated with introducer sheaths was a frequent complication and cardiac index was significantly lower in patients with ijvt. though the incidence of ijvt was higher in patients with prolonged catheterization, it developed even on 1 day after surgery and was usually asymptomatic. this risk should be carefully considered when the insertion of pulmonary catheter is chosen for cardiac surgery. methodology. it is a prospective study of patients admitted to our icu after undergoing robotic radical prostatectomy (da vinci) in the time interval going from january 2007 up to april 2008. we analyzed clinical and demographic data, the length of stay in the icu and hospital, the need for blood transfusion, surgical times and the complications suffered during hospital stay. data are expressed as mean, median or percentage, using the student's t test and chi-square to compare averages and detect possible associations between variables. results. seventy three patients underwent surgery with a median of 60 years. mean surgical time was 240 min. in recent months this time is reduced to 180 min. the mean haemoglobin at admission (12.5 g/dl) was significantly higher than when dismissed (11.7 g/ dl), p \ 0.001. an average of two units of concentrated red blood cells was transfused in the surgery room in 13.7% of patients. only one patient required transfusion at the icu. cardiac or renal mild complications appeared in 5.5% of patients. this could not be associated with age. the median of mechanical ventilation length was 2 h. one patient required conversion to open surgery due to profuse bleeding. there was no hospital mortality and no need for reoperation. mean stay at icu was 1 day, significantly less than those patients who suffered complications (p \ 0.001). the median stay in ward was 4 days. conclusion. robotic radical prostatectomy (da vinci) has a very low associated morbidity, minimal blood transfusion requirement and short is the stay at the icu and hospital, in contrast to published data with open surgery. there was no hospital mortality. objectives. to evaluate short-and long-term outcome in patients undergoing coronary artery bypass grafting (cabg), who received an intra-aortic balloon pump (iabp) prior to surgery. methods. between january 1990 and june 2004, all patients (n = 154) who received an iabp prior to on-pump cabg in our center were included. patients received the intra-aortic balloonpump for vital indications (i.e. either unstable angina refractory to medical therapy or cardiogenic shock; group 1; n = 99) or for prophylactic reasons (group 2; n = 55). a cox proportional hazards model was used to identify predictors of long-term all-cause mortality. compared with the euroscore predictive model, observed 30-day mortality in group 1 (15.2%) was not significantly higher than predicted (10.3%). a dramatic decrease in 30-day mortality occurred in group 2 (median predicted mortality was 7.2% and observed was 0%, p \ 0.05; fig. 1 introduction. physiological abnormality is associated with adverse outcome and a high percentage of patients admitted to icu have abnormal physiology in the hours prior to admission [1] . track and trigger systems are used to enable timely intervention to a deteriorating patient. the mews system is used in our level 1 care general wards with more intensive monitoring in our level 2 care facilities. objectives. we hypothesised that the mews for patients admitted from the general wards were being inadequately documented and that this be may influencing outcome. we therefore also hypothesised that patients admitted from the level 1 care general surgical and medical wards in our hospital had a poorer outcome following icu admission than those admitted from sites of level 2 care, such as the high dependency unit (hdu), emergency department and theatre recovery. we undertook a prospective 10 week audit of all patients admitted to our district general icu. all patients' case notes and monitoring charts were reviewed by an icu consultant with additional data (e.g. apache ii scores) obtained from the ward watcher icu database. 30 day mortality data was obtained from the hospital's sci patient database. we then compared the data with that available for patients admitted over the previous year (2007) background and aim of study. aorto-femoral bypass (afb) is widely used for the patients with peripheral vascular disease (pvd). nevertheless, there is no consensus about the type of anesthesia for this difficult group of patients. we hypothesized that continuous spinal anesthesia (csa) will be more secure and suitable for the patients with pvd combined with pulmonary and cardio-vascular co-morbidities. the aim of our study was to compare alterations of the mean arterial pressure (map) and delivery of oxygen (do 2 ) during afb and in the early postoperative period under the influence of ga and csa. after approval of our hospital helsinki committee prospective randomized study was performed between 2004 and 2008: 34 male patients with pvd were included in our work. risk of anesthesia was equal to the third degree according to asa scale. in the first group of patients (n = 15) ga with mechanical ventilation was employed. in the second group of patients (n = 19) csa was used. both groups of patients were similar with respect to age and co-morbidities (copd 100% in both groups, ischemic heart disease and arterial hypertension). for ga we used propofol, midazolam, fentanyl, and isoflurane. bupivacaine was used for csa. in combination with mental sedation by intravenous midazolam. map was measured directly through radial artery catheter and do 2with the help of tetrapolar rheovasography. both parameters were measured during fixed 7 points: before the operation, at the end of induction of anesthesia, after cross clamping of the aorta, after release of aortic clamp, first hour after operation, 12 h after operation and 24 h after operation. mann-whitney test was used for statistical analysis of our results. in the patients with csa during the operation and in the early postoperative period, map was lower but statistically non significant. map was lower statistically significant only during the cross clamping of the aorta and in the first postoperative hour, most probably due to the influence of the sympathetic block. at the same time do 2 had almost no difference in both groups. only in induction stage it was lower in the ga group that most probably was connected with negative influence of propofol on cardiac output. conclusions. both methods of anesthesia ga and csa gave us opportunity to preserve stable map and do 2 during afb that we performed in this difficult pvd patients with copd and cardiovascular co morbidities. over the last few decades, several scoring system have been developed for use in critically ill patients, not only to assist therapeutic decision making but also to guide resource allocation and quality of care. to evaluate the tiss-28 in surgical intensive care unit (icu) patients and the possible relationship between tiss-28 and the type of surgery, severity of illness, and outcome in these patients. prospectively collected data from all patients admitted to a postoperative icu between 1st march 2004 and 30th june 2006 were analyzed retrospectively. a-priori subgroups were defined according to gender, age, saps ii score, sofa score, surgical procedures, and the occurrence of major morbidity or death in the icu or in-hospital. a total of 6,903 patients were admitted during the study period (63.5% male, mean age 62.3 years) constituting 29,140 observation days. the highest tiss-28 scores were observed on the day of admission. the highest tiss-28 was observed in patients who underwent cardiothoracic surgery, the lowest in neurosurgical patients. during the first week in the icu, tiss-28 was correlated to the severity of sepsis syndrome; however tiss-28 scores remained elevated only in patients with severe sepsis/septic shock. tiss-28 score was correlated to saps ii (r 2 = 0.42, p \ 0.001) and sofa score (r 2 = 0.48, p \ 0.001) throughout the icu stay and was consistently higher in non-survivors than survivors during the first 2 weeks in the icu. conclusions. the highest tiss-28 scores are observed on the day of admission to the icu with marked variations according to the type of surgery. tiss-28 correlates well with the severity of sepsis syndrome and outcome in these patients. our data could be helpful in icu planning, risk stratification, and resource allocation in the surgical icu setting. introduction. pain and opioids for treatment of pain can affect immune function in cancer patients, which may in turn influence metastatic capability of a primary tumour during and after surgical excision. it is also been shown morphine has a direct effect on cancer cells, but results of these studies have been conflicting. we therefore aimed to determine effect of morphine, commonly used in anaestehsia and intensive care, on in vitro breast cancer cell migration using two breast cancer cell lines. we used two cell lines: mcf7 is er positive breast cancer cell line while mda-mb-231 is er negative, less differentiated and more invasive. cells were incubated with or without morphine (concentrations 10-100 ng/ml) for 12, 24 and 36 h, corresponding to clinically relevant concentrations and exposure times. cell proliferation was determined using an mts (promega inc.). 24 h cell migration was determined using a 96-well flourescent kit (chemicon). results were compared using independent sample t test for differences between the groups. morphine had positive effect on cell proliferation, which was greater in mda-mb-231cells. proliferation of mda-mb-231 was increased the most at 12 h incubation and higher concentrations (100 and 75 ng/ml caused 41 and 26% increase in proliferation at 12 h incubation and up to 25% increase at 36 h incubation). proliferation of mcf7 cells was increased by 4% in 24 and 36 h incubation periods. morphine caused an increase in migration of both cell lines, which was again more evident with mda-mb-231 cells at higher concentrations of morphine (23, 10% and 44% increase with 100, 75 and 50 ng/ml respectively). our experiments have shown morphine has potential to directly stimulate breast cancer cell proliferation and migration in vitro, especially in less differentiated breast cancer cell line. further studies are needed to determine its effect on other metastatic mechanisms such as invasion and gene expression as well as the implication of these results for clinical practice. objectives. aim of this study was to evaluate the predictive value of nt-probnp levels and inflammatory markers (crp, il-6, tnf) on late mortality in patients who underwent thoracic surgery for lung cancer. methods. 21 patients median age (61 ± 11 years) without history of heart disease or renal failure. the blood tests for nt-probnp, crp, il-6 and tnf analyses were drawn one day preoperatively, 1 h, 48 h and 7 days postoperatively. patients' demographic data, laboratory results and mortality were collected and assessed. results. nt-probnp at 48 h was significant higher in non-survivors (1,330 ± 1,489 pg/ ml) compared to survivors (364 ± 231 pg/ml, p = 0,026). furthermore, nt-probnp at 48 h was associated with survival in the cox-regression analysis (p = 0.038, hr = 1,073, 95% ci: 1,004-1,147, units: 100 pg/ml). crp preoperatively was significant higher in non-survivors (64 ± 49 mg/dl) versus survivors (12 ± 18 mg/dl, p = 0,002). il-6 preoperatively was significant higher in non-survivors (61 ± 42 pg/dl) compared to survivors (20 ± 16 pg/dl, p = 0.011). conclusions. high nt-probnp levels at 48 h postoperatively, associated with increased mortality in patients undergoing thoracic surgery but there was no relation ship between crp, il-6 and mortality. introduction. the occurrence of post-operative delirium in elderly orthopaedic patients is associated with neurological complications and cognitive decline [1] . although the etiology of the decline is less understood, cerebral ischemic events may be involved [2] . high plasma concentration of n-methyl-d-aspartate (nmda) receptor antibodies (nr2ab) has been proven highly predictable for occurrence of the postoperative neurological events in cardiac surgery [3] . objectives. the aim of the present study was to investigate the predictive value of blood levels of nr2ab for postoperative delirium, cognitive dysfunction or any other neurological complications after hip and knee replacement surgery. methods. the study enrolled 47 consecutive patients, aged over 65, requiring acute or elective knee or hip replacement surgery. cognitive impairment was evaluated by minimental state evaluation (mmse) test administered before and after surgery. daily postoperative delirium was evaluated by confusion assessment method for intensive care unit (cam-icu). plasma levels of nr2ab were recorded before surgery, at the moment of hospital discharge (10-14 days postoperative) and 6 weeks after discharge. all other possible risk factors for postoperative delirium were also recorded. cognitive decline was present in 16 patients (34%) before surgery and in 35 patients (74%) at the moment of hospital discharge (p \ 0.0002). plasma levels of nr2ab were 1.30 ± 1.85 ng/ml preoperatively, 1.28 ± 1.01 ng/ml at the moment of hospital discharge and 1.61 ± 0.95 ng/ml 6 weeks postoperatively, with no significant differences. conclusions. the incidence of cognitive decline in elderly patients after othopaedic surgery was significantly higher when compared with the preoperative status but there was no correlation between the cognitive decline and the plasma levels of nr2ab. methods. patients older than 18 years, whose performed endoscopic, colonoscopic or both procedures, under sedation performed by the intensive care deparment of the hospital del tajo. data were collected for 6 months. demographic characteristics, medical history, asa (american society of anesthesiologists classification), drugs bolus and total dosages, respiratory and hemodynamic data, the length of procedure and recovery, and complications were collected. tolerance was assessed by endoscopist, with a 1 (very bad) to 5 (very good) scale. quantitative data are expressed with mean and standar desviation, and qualitative data with percentage. results. 93 procedures were included. table 1 shows main characteristics. tolerance and complications are referred in table 1 . the 91.4% of the procedures were appropriate (4 or 5) . the main complications were vomiting (5.4%) and hallucinations (12.9%). there were only 2 incidences of respiratory depression and 2 of hypotension. background. the authors hypothesized that the efficacy and quality of a remifentanil (r)-based regimen versus a piritramide (p)-based analgosedation in major post-surgical patients with renal and hepatic impairments still more potent even if prevention of narcotic induced hyperalgesia (nih) [1] was done. the nih was made by sulphate magnesium (m), ketamine(k) or clonidine(c). methods. 66 patients were randomly allocated to receive a blinded infusion of either r at a rate of 0.15 l/(kg min) (±0.10) (g1: n = 33) or p at 0.2 mg/(kg h) (±0.15) (g2: n = 33) coupled to an hypnotic sedation of propofol. r and p were titrated in icu after surgery, to achieve an optimal sedation as defined by a sedation conclusions. the remifentanil-based regimen allowed a more rapid emergence from sedation and facilitated earlier extubation diminishing total icu hospitalisation time and cost. even if we prevent the narcotic induced hyperalgesia by used of magnesium, ketamine or clonidine, needs of tramadol in rescue still lower in the remifentanil group due to its high power coupled with its high fexibility compared to piritramide. its reducing, by the way, risks of tramadol's metabolites accumulation in case of renal or leaver impairment. a ''fast track'' approach to cardiac surgery has significantly shortened the length of icu stay. however, quick awakening from anesthesia and subsequent extubation after discontinuation of sedative drug sometimes cause instability of hemodynamics, such as increase of bp or hr. dexmedetomidine (dex), a 2 agonist, is a sedative drug that can be continuously infused during weaning and extubation. the aim of this double-blind study was to evaluate the effect of dex on time to extubation and hemodynamics during weaning from mechanical ventilation after cardiac surgery. with irb approval and informed consent, the patients undergoing cardiac surgery were randomly divided into two groups, dex group [infusion of 0.5 lg/(kg h) of dex] and saline group. drug administration was started at sternal closure and continued 24 h. ramsay sedation score, times to extubation, systolic blood pressure, heart rate, respiratory rate, pulmonary artery pressure, central venous pressure, cardiac index were examined. we analyzed these parameters on icu admission, when the patients opened their eyes on order, at immediately before extubation, 30 min, 3 h, and 6 h after extubation. unpaired t test was used for statistics and p value less than 0.05 was considered significant. results. 24 patients were included in this study (n = 11 in the dex group, n = 13 in the saline group). there were no significant differences between two groups in age, length of surgery, length of anesthesia, and total dose of propofol and fentanyl. time to extubation was 625 ± 332 min in the dex group and 446 ± 309 min in the saline group (mean ± sd), which were also no significant differences. ramsay sedation score were maintained33 and no patients needed additional sedative drug during assisted ventilation in the dex group. althought mean systolic bp and mean pa, mean cvp, rr were similar in both groups during infusion. hr at eye opening, immediately before and after extubation were significantly lower in the dex group than in the saline group. conclusion. our results demonstrated that the infusion of 0.5 lg/(kg h) of dex decreased hr during weaning from mechanical ventilation. dex could not only provide adequate sedation but also suppress the stress response after cardiac surgery. dex is a useful sedative drug for preventing instability of hemodynamics on fast track approach. however, most anxiolytics impair intellectual function. dexmedetomidine (dex) is an alpha 2agonist that may offer sedation without overt cognitive decline. we performed a comparison between dex and propofol (pro), a drug often used for icu sedation. methods. prospective, randomized, double-blinded, cross-over study of 30 awake and intubated brain-injured (bi, n = 18) and non-bi (12) icu patients, each receiving pro and dex using a cross-over design with periods of baseline (analgesic use fentanyl only), drug a, interval washout (fentanyl only), drug b. sedation was titrated to a score of 0 or -1 (calm, cooperative) on the rass and hopkins nics scale. cognitive testing was performed at each study period using the validated 100-pt hopkins ace cognitive battery. objectives. we evaluated the effect on final outcome of a treatment regimen with lowdose haloperidol initiated when a positive cam-icu occurred as a quality improvement project. methods. the cam-icu was previously implemented in daily care in all patients who stayed[48 h in our 10 bed medical-surgical icu. in the first 9 months of the study (period 1), the cam-icu was used as an adjunct to daily care and the treatment of delirium was left to the physician on an intention to treat basis. subsequently, a 2 month study pause was defined. thereafter, in the second 9 months of the study (period 2), the cam-icu was judged to indicate the presence of delirium and haloperidol was directly started with a loading dose of 5 mg iv with subsequent daily dosing of 2.5-2.5-5 mg iv (age \ 70) or 1 oversedation is still common in many intensive care units (icu) despite the demonstrated benefits associated with sedation sparing strategies, including shorter duration of mechanical ventilation, and shorter length of stay in the icu and hospital. our aim was to observe whether a minimal sedation policy could be feasible in a multidisciplinary department of intensive care. prospective observational study over a two month period (december 1, 2008 to january 31, 2009) in a 35-bed medico-surgical department of intensive care of a university tertiary hospital. all adult patients who stayed in the icu for more than 12 h were included. data were collected on duration, type, dose, and indication for sedative and opiate analgesic agents. self extubation was used as a safety surrogate. disease severity was assessed by the apache ii score within the first 24 h of admission. statistical analysis was performed with spss software (spss incorporation, chicago, il, usa). a total of 335 patients (male 57%) with a median age of 61 years were included; 142 (42.4%) received some sedation, the majority [131 (92.3%)] during mechanical ventilation. midazolam (54%) and propofol (49%) were the most frequently used sedative agents. the most common indications for sedation were: early postoperative (50%), severe respiratory failure (19.7%), short term procedures (19.7%), and withdrawal syndrome (9.9%). the median percentage of time during which patients received mechanical ventilation without sedation was 87.5%, and was not related to severity as assessed by the apache ii score (rho 0.10-p = 0.2). in the group of patients who required sedation for longer than just short procedures or uncomplicated postoperative care ([2h), the median percentage of time during which patients received mechanical ventilation without sedation was 73.6%. analgesic opiates were often required (65%), predominantly by continuous infusion (56%). morphine was the most frequently used agent (54%). self-extubation occurred in 6 patients, but only 1 needed re-intubation. conclusion. in a mixed medical-surgical population of critically ill patients, a strategy of minimal or no sedation (''sedationless'') is feasible and without major adverse effects. we propose that comfort, hemodynamic instability, and mechanical ventilation should be abandoned as usual indications for sedation. grant acknowledgement. drs received grants from the doctoral fellowship program of capes/ brazilian ministry of education and from the federal university of rio de janeiro. r. russai 1 1 the royal free hospital, anesthetics, northwood, uk postoperative cognitive dysfunction (pocd) is reported to occur frequently after cardiac surgery, even in low-risk patients. predictors of neurocognitive deficits can suggest the potential etiology and outcome of patient that has developed pocd. there is a wide range of neurological manifestations from subtle cognitive impairment to deadly stroke. over a period of 20 weeks a population of 376 patients underwent cardiac surgery in our hospital. we have looked for any signs of pocd in correlation with the possible etiology. data have been collected prospectively focusing on past medical history (pmh), possible contributors, manifestation, complications and treatment. pocd has occurred in 25 patients (21 male, 4 female) with age range of 45 to 90 years (median age 73 years), of whom 28% has had pmh of neurological impairment (predementia; cerebrovascular disease). multifactorial etiology was found: respiratory failure 32%, morphine 26%, tramadol 15%, renal failure 12%, remifentanyl 9%. in 2 patients no related causes were recognised. all 25 patients showed confusion as leading manifestation, although in 20 patients confusion presented in combination with aggressive behaviour (11), cognitive dysfunction (6), paranoia (3). in 15 occasions pocd resulted in major complications such as difficulties in airway management (5), removal of cvp line (4), removal of arterial line (6). the majority of patients (22) required pharmacological treatment with single or multiple drugs therapy, the most common used was haloperidol (48% pts). the average length of stay in itu was 6.7 days, and the average length of hospital stay was 22.5 (6-102) days. conclusion. pocd is a common and potentially devastating complication with a complex and broad etiology, which may affect the rehabilitation process and the final outcome. early diagnose is essential for personalise treatments and therefore preserve in both life quality and life expectancy. introduction. sedation and analgesia is given to the icu patient for adaptation to the intensive care environment. however, side-effects of drugs used are increasingly acknowledged as negative factors increasing the risk of delirium, vasopressor therapy, prolonged ventilation and length of stay. objectives. the purpose of the proposed study was to study the practice of sedation in norwegian icu's and the challenges experienced by nurses and physicians. a national postal survey for clinicians in all norwegian icu's was conducted in september and october 2007. all intensive care units treating mechanically ventilated patients for more than 24 h (n = 54) were included. two respondents from each unit (1 intensive care nurse and 1 icu physician) were invited to answer the questionnaire. the survey was based on two previous danish surveys. questions on practice and perceived problems were scored on a numeric rating scale and a lickert scale, as well as a few questions with response categories based on theme. results. the response rate was 93% (n = 100). all 54 icus were represented with 53 nurses with formal education in intensive care and 47 physicians specialized in anesthesiology as respondents. written protocols are not routine in norwegian icus, but half of the departments titrated sedation according to a scoring system, most commonly maas. the most commonly used sedatives were propofol and midzolam, while fentanyl and morphine were the most used analgesics. the main indication for sedation was to achieve tolerance to ventilation and to treat other bothersome symptoms. side-effects were reported to be frequent with both sedatives and analgesics. the most frequent side effects (% reported as often present or always present) with sedative agents were circulatory instability (74%), delayed awakening (64%) and sleep disturbances (48%), while the most frequent side effects experienced with analgesics were gastrointestinal problems (75%), circulatory instability (53%) and delayed awakening (47%). the main indication for tracheostomy was reported as longterm ventilation and the wish to reduce sedation. discussion/conclusion. written protocols were not routinely used. side-effects of sedation are perceived as a problem by the majority of clinicians, leading to circulatory instability and delayed awakening. tracheostomy is used first of all to be able to reduce longterm ventilation and sedation. these results indicate a potential for new sedative agents and analgesics with fewer side-effects and more focus on the use of sedation protocols. [1] and has been associated with gaba agonist use [2] . delirious patients may not be overtly agitated, so signs of delirium must be actively sought. the confusion assessment method for the intensive care unit (cam-icu) is a validated and easy to use screening tool [3] . in a recent study on this 13 bed medical and surgical intensive care unit (icu), more patients who had received gaba agonists were delirious compared with those who were sedative free [4] . however, the percentage of patients having at least one episode of delirium was lower than expected (24%), perhaps because they were screened only once daily and in the daytime. we repeated this study with twice daily assessments (morning and after dark) and a larger number of patients in order to obtain a more accurate prevalence of delirium and confirm an association with gaba agonist use. methods. two doctors attached to the icu received a 30 min tutorial on using the richmond agitation and sedation score (rass) and the cam-icu assessment tools. the cam-icu was performed on all rousable patients (rass score [ -4) twice daily (morning and after dark). the following information was also noted: ( conclusion. this study shows that the prevalence of delirium on this unit is comparable with published research [1, 2] and higher (44.3 vs. 24%) [4] when patients were screened after dark as well as in the daytime. the study shows that any sedating drug was associated with significantly increased prevalence of delirium. the unexpected higher prevalence of delirium in the patients receiving non-gaba agonists versus gaba agonists cannot be explained by haloperidol use to treat delirium. results. there were 93 patients in the 11 icus. the mean age was 61.2 years old with a predominance of chinese (76.3%) and a slight male predominance of 57.0%. forty-six per cent of the patients were mechanically ventilated and 14.0% had tracheostomy done. there were an average number of 4 devices per patient. sedation was administered in 25.8% of the patients with no sedation scales used in a quarter of these patients. only 20.8% of the sedated patients were on sedation protocol. the majority of patients (58.3%) were monitored hourly and on propofol (50%) and midazolam (37.5%). up to 41.2% of sedated patients did not have daily interruption of sedation. there were no significant difference noted in the use of sedation between medical and surgical icus. slightly more than a third of patients were given analgesia (n = 33) with no analgesia scales used in a third of these patients. one third of them were administered with paracetamol and about a third with morphine. patients in surgical icus were more likely to receive analgesia compared to medical icus patients. most of these patients (81.8%) were monitored hourly. only 2 patients were on neuromuscular blockade. there was no usage of any formal delirium assessment tools at all with 38.8% of the patients being assessed for delirium based on clinical judgement of the caring team. only 24% of the patients had some form of sleep promotion in the icu. conclusions. this national multi-center study reveals several deficits in the adult icu with regards to sedation, analgesia and delirium assessment and management. several initiatives should be implemented to improve patients' safety and quality of care in the icu. methods. this study was conducted in 41 patients who visited emergency care center following caustic ingestion during a period ranging from january of 1998 and august of 2008, in whom a retrospective analysis of medical records was performed. findings for the esophageal lesion were classified according to the change of the esophageal wall and the infiltration of periesophageal soft tissue. also, clinical, laboratory, and endoscopic data from this patients were reviewed. the correlation between the degree of esophageal damage seen on ct scans and esophageal constriction seen on esophagography were then evaluated. a total of 41 cases of caustic ingestion were identified (age range, 20-82 years). the most common caustic agent ingested was acid (70%). the most frequent cause for ingestion was attempt of suicide (70%) as opposed to accidental (30%). the findings of thoracic ct in 41 patients were follows: first-degree esophageal injury in 4 (9.8%), seconddegree in 8 (19.5%), third-degree in 17 (41.6%), fourth-degree in 12 (29.3%). fourteen patients (34.1%) developed caustic esophageal stricture. the degree of esophageal damage got closer to grade iv, the more prevalent esophageal constriction became. this correlation was statistically significant (p \ 0.001). of the total 41 patients, 26 underwent endoscopy in the early stage after they visited emergency care center. an analysis of the correlation between the degree of esophageal damage seen on endoscopy and that seen on ct scans was performed. this revealed a significant correlation (p = 0.002, r = 0.585). conclusions. thoracic ct grading suggesting periesophageal soft tissue infiltration and fluid collection (grade iii to iv) rather than only edema (grade i) may be associated with stricture formation. early ct grading was very safe and useful for predicting the development of stricture induced by caustic ingestion. conclusion. in our area critical care transport teams provided safe transfers for critically ill patients.adequate preparation, strict adherence to checklists and adequate personal are the key of optimal solving of problems. introduction. although endovascular repair of traumatic aortic injury (ertai) has revolutionized the practice of vascular surgery, many questions still remain unanswered. endoleaks, coverage of the left subclavian artery, stent fold/collapse, access complications and durability are the most important complications associated with the procedure. we describe our experience with stent fold/collapse after endovascular repair of blunt aortic injury in otherwise healthy and young patients. from january 2005 to december 2008, 14 patients (mean age 39 years, mean apache score 11, mean length of stay 13 days) who underwent endovascular repair of a blunt aortic injury were admitted in our icu. every day clinical examination and invasive blood pressure monitoring were employed for all our patients. when persistent hypertension was detected, transthoracic (tte) and transoesophageal echocardiography (tee) were initially used, followed by spiral computed tomography (ct) and angiography as confirmatory methods. of the 14 patients, 4 (28%) developed a pressure gradient of [50 mmhg at the level of the stent that was initially investigated with continuous wave doppler at the descending thoracic aorta (suprasternal view). the complication presented with refractory hypertension (requiring more than two classes of antihypertensives in high doses) and difficult weaning. the cause of hypertension in 1 of those 4 patients was a stent collapse, while in the other 3 patients the stent appeared folded but not collapsed. endograft revision by open surgery was necessary in 1 of the 4 patients. conclusion. the absence of especially designed grafts for the treatment of blunt aortic injury and the subsequent use of oversized grafts are associated with severe complications. refractory hypertension after ertai can be a manifestation of poor stent alignment and/or stent collapse. echocardiographic monitoring proved to be a useful tool in the early diagnosis of this kind of stent-graft complication. as far as we know, it is the first time that echocardiography is described in the relative literature as an early diagnostic technique for this serious complicationction. facing an aging population, the number of interventions of the french emergency medical service (ems) among very elderly is increasing. a previous retrospective study showed that except from out-of-hospital cardiac arrest survival to discharge was remarkably high after ems intervention for life-threatening pathology [1] . the aim of the present study was to evaluate prospectively outcomes of very elderly patients managed by ems. methods. after irb approval, we conducted a prospective study over 1 year, including all patients aged 80 years or more managed by our physician staffed ems department. characteristics of patients including previous medical status (mccabe and knaus scoring systems), functional independence (katz adl scale), clinical conditions, the index de gravité simplifié ambulatoire (igsa) severity score were recorded. patients were followed until their hospital discharge. the 3-month mortality was recorded as well as the adl score. data are expressed as mean ± sd, median [iqr] or percentage of patients and compared using univariate and multivariate analysis. a p \ 0.05 was considered the threshold for significance (*). results. of the 523 patients included, 53 died on-scene, 440 were transferred to the hospital and 30 patients were left on scene because of significant improvement in medical status making hospitalization unnecessary, or on the contrary in near-death situations. mean age was 86 ± 5 years (214 men). their adl was 2 (0-9) and 63% of patients were living at home. main conditions were pneumonia (n = 80), acute coronary syndrome or chest pain (n = 76) and acute pulmonary oedema (n = 67). at 3 months, survival rate was 66% (n = 273). the proportion of patients living at home was 64% and adl among survivors was 2 (0-8) (vs. 1 (0-6) initially for this subpopulation, p = 0.01]. when compared with deceased patients, survivors were significantly younger (86 ± 5 vs. 87 ± 5 years*), had lower adl penetrating anterior chest wounds causing cardiac injury are typically fatal, with only 6% of patients surviving to reach hospital. while the majority of patients with thoracic trauma can be managed conservatively or with simple intercostals catheter, a small but significant number of blunt (10%) and penetrating (15-30%) injuries, require emergency resuscitative mediam sternotomy as a component of initial resuscitation. case report. a 35 years old men, fall from 20 meters high, while working in a truss. he was immobilized with semirigid cervical collar and backboard in the scene and transport to our trauma center, witch was a 1 h car-distance. anesthesiologist team was present since the initial management in emergengy room (er) and act according to advanced trauma life support principles. in primary survey, patient was paraplegic, had a gcs of 14/15, a normal respiratory rate, a slight hypotension and a slight tachycardia. when surgeon places a chest drainage, it drains immediately more than 1,500 ml blood, and the patient vital signs started to fade, to extreme bradicardia. the patient was then intubated with a rapid sequence intubation, with a single lumen endobronquial tube, and ventilated with protective lung ventilation. hypotension postinduction was promptly treated with vasoactive drugs (nor-adrenaline and dobutamine) and ongoing volume resuscitation. an emergency resuscitative mediam sternotomy incision was perfomed in the er and reveled a clavicule and sternum fracture and laceration in the braquiocephalic artery which has repaired. maintenance was performed with total intravenous anesthesia with fentanyl and nondepolarizing muscle relaxant. monitoring include standard monitoring plus direct arterial and central venous pressures. during the surgical procedure we treat massive blood loss, with multiple transfusions of 17 units of red blood cells (unmatched type-specific), seven units fresh plasma, and 2 pools of plaquets, fibrinogen and cryoprecipitate. at the end of the surgery, still ongoing blood loses, made us suspect of coagulopathy, and to use octaplex ò . it was also performed a nasal tamponade, to stop severe epistaxis and suture a major scalp wound with evidence of basal skull fracture. patient was transferred to an intensive care unit (icu) ventilated.we was extubated at the 18th day post-operative. after 27 days, he still remains in icu, because he is recovering from lumbar spine fixation for a total fracture-dislocation of d8-d9. discussion. although he remains paraplegic, we think emergency sternotomy have had a significantly impact in this life-threating situation. the use of cell-saver ò would have been beneficial, but it was unavailable in er. we included only patients attended in this unit by icu personnel. these patients belonged to the area assigned to cruces which has been reference centre of the northern area of spain until december 2008. we collected all the information needed from the clinical history and the treatment sheet, and used the spss 15.0 programme to perform the statistic analysis. results. we found 83 patients that meet the severely burned patients criteria and that were attended by the icu personnel in colaboration with the plastic surgey unit. their medium age was 49.88 ± 19.29 years, 77% of those patients were men, and the medium burned body surface was 32 ± 21.59%. these patients remained hospitalized in this unit during a medium time of 28.43 ± 21.7 days. during their stay, the 100% of them needed mechanical ventilation, 36% presented acute renal failure, 57% had a pao 2 /fio 2 less than 300, and 75.9% suffered some kind of infection. methods. prospective and observational study developed in a burn unit, in which were included all patients with total surface body area burn (tsba) [ 20% and/or inhalation syndrome who were admitted in our unit from march 2007 to december 2008. we used transpulmonar thermodilution by means of monitor picco ò in a total of 10 measurements per patient (admission and every 8 h). we collected measurements of cardiac index (ci), intrathoracic blood volume (itbv), extravascular lung water (evlw), inhalation syndrome or not (it was diagnosed by broncoschopy), percentage of tsba and abbreviated burn severity injury score (absi) in a total of 40 patients.the average change of measurements of ci, itbv and evlw was studied in the following determinations and their association with few factors with a general and lineal model of mixed effects longitudinal data unbalanced. results. the evolution of thermodilution measurements was the following (graphic 1) cardiac index: ci 1 = 2.83, ci 2 = 2.6, ci 3 = 2.86, ci 4 = 3.14, ci 5 = 3.41, ci 6 = 3.7, ci 7 = 3.95, ci 8 = 4.13, ci 9 = 4.37, ci 10 = 4.37. intrathoracic blood volume: itbv 1 = 753, itbv 2 = 733, itbv 3 = 783, itbv 4 = 742, itbv 5 = 757, itbv 6 = 813, itbv 7 = 890, itbv 8 = 896, itbv 9 = 923, itbv 10 = 908. extravascular lung water: evlw 1 = 6.78, evlw 2 = 7.3, evlw 3 = 7.82, evlw 4 = 8.74, evlw 5 = 8.71, evlw 6 = 8.65, evlw 7 = 9.31, evlw 8 = 9.13, evlw 9 = 8.71, evlw 10 = 8.37. in our serie, 75% of patients were male and the average age was 50.12 (20-86). nine out of all the patients (22.5%) suffered inhalation syndrome, the average absi was 8.69 (5) (6) (7) (8) (9) (10) (11) (12) (13) (14) (15) (16) and the average of tsba was 37% (20-95%). mortality in our serie was 25% (10 patients).ci and itbv measurements increased significantly while the reanimation advanced (ci p 0.0001) (vsit p 0.0001). in evlw we only find significantly differences in post hoc study between first measurements and fourth one (p 0.03), 5th (p 0.04), 6th (p 0.04), 7th (p 0.007), 8th (p 0.01) and 9th (p 0.04).in the evlw/itbv ratio (permeability index = pi) we did not observe significantly changes in the evolution.the inhalation factor did not change ci outcomes neither magnitude nor in the measurements evolution (p 0.48 and p 0.71 respectively), the same form, itbv (p 0.73 and p 0.28 respectively), but inhalation modified evlw (p 0.04) and the permeability index was in the signification stadistic limit (p 0.06).mortality was higher in patients who ci was lower and evlw was in higher values. conclusions. thermodilution in the reanimation period in critically ill burn patient shows significantly haemodynamic changes in the evolution that can help to adapt the treatment.the inhalation syndrome only modified the measurements of evlw significantly in this period but it influenced neither ci, itbv nor pi. introduction. one major issue in trauma management is to get every patient directly from the scene to the appropriate hospital for the injury he sustained. patterns of interfacility transfers have been thouroughly investigated in trauma system settings, but scarce data are available about transfers in non trauma system settings. objectives. this study aims to assess interfacility transfers that eventuate in the abscence of a formal trauma system and to estimate the potential benefits from the implementation of a more organized process. the 'report of the epidemiology and management of trauma in greece' is a one year project of trauma patient reporting throughout the country. it provided data concerning the patterns of interfacility transfers. in greece there is no formal trauma system employed and to our knowledge, all available data concerning the epidemiology of trauma in the country are either extrapolations of relevant data from other countries or based on police reports and individual hospital reports. in this study, we attempted to evaluate the paterns of interfacility transfers, information reviewed included patient and injury characteristics, need for an operation, intensive care unit (icu) admittance and mortality. trauma patients were devided in two groups, the transfer group was compared to the non-transfer group. analysis employed descriptive statistics and chi-square test. interfacility transfers were furthermore assessed according to each health care facility's availability of five requirements; computed tomography scanner, icu, neurosurgeon, orthopedic and vascular surgeon. results. data on 8,524 patients were analyzed; 86.3% were treated at the same facility, whereas 13.7% were transferred. in transferred group there were more male, the mean age was lower than that of the non transferred group and the injury severity score was higher. transferred patients were admitted to icu more often, had a higher mortality rate but were less operated on compared to non-transferred. the transfer rate from facilities with none of the five requirements was 34.3%, whereas the rate of those with at least one requirement was 12.4%. facilities with at least three requirements transferred 43.2% of their transfer volume to units of equal resources. conclusions. the assessment of interfacility transfers can reflect current trends in a nontrauma system setting and could indicate points for substantial improvement. results. 1,090 patients included, 2,172 injuries analyzed. average age was 36.5, 79.5% men. 43.3% were car accidents, 20% falls, 13.8% motorcycle, 6.6% run over and 2.2% bicycle. 49.9% had one injury, 30.8% two and 15.1% three. most frequent injury was tbi (33.7%), thoracic traumatism (20.2%) and ortophaedic (15.6%); severe tbi was 56.4%. ctrate according to marshall classification was 38.1% ii, 20.3% v and 12.6% iii. iss averaged 20, higher in dying patients than in the survivors (43 ± 21.6 vs. 20.1 ± 10.9; p \ 0.001). of the non mechanical-ventilated patients, 43.2% were so in the first 12 h following admittance. during this, 31.6% patients were given blood transfusions, platelets 6.3%, plasma 14.6% and prothrombinic complexes 2.2%. in the first 24 h 36.4% underwent surgery, most frequent was neurosurgery (52.8%). complications: nosocomial pneumonia (25.3%), catheter related bloodstream infection (6.9%), acute kidney injury (6.3%), ards (10.3%), cns infection (0.9%). 2.4% renal replacement therapy. invasive ventilation was used in 67.3% with 7.32 ± 11.7 days, non invasive ventilation in 0.8%. average stay in the icu was 4 days. 76.4% of the patients were transferred to a ward. 8.6% were transferred to another hospital. gos on discharge was higher than 3 on 67.3%. 15% died in icu, 7% brain death. tbi as a main injury showed a 32.4% mortality rate. depending on trauma type, mortality was higher in fall (40%) and run over (27.1%). if due to car accident (14.4%), motorcycle (27.1%) or bicycle (4.3%), mortality in icu was lower (p \ 0.001). prehospitalary variables related to mortality were age, gcs \ 7 and a motor component \ 5, pupil alteration, shock, respiratory failure, prehospitalisation intubation and iss (p \ 0.001). on arrival to hospital, the variables were: haemodynamic instability in the first 12 h, transfusions need and number, marshall iv-vi, mechanical ventilation (p \ 0.001) and initial fibrinogen (p \ 0.05). evolutive variables related to a higher mortality rate were days of stay, invasive ventilation, tracheostomy and the show up of complications (catheter related sepsis -p \ 0.01-, nosocomial pneumonia, acute kidney injury, ards, renal replacement therapy (p \ 0.001). in a logistic regression model, prehospitalisation variables having an influence on icu mortality rate were age (or 1.05; p \ 0.001), mydriasis (or 2.9; p \ 0.005), gcs-motor component (or 0.7; p \ 0.001), shock (or 3; p \ 0.001) and iss (or 1.1; p \ 0.001). conclusions. multiple trauma patients show a high need of resources, with many peaks of treatment involving a high monitorization and handling. many of the variables are related with a higher mortality rate in icu: iss, mydriasis, gcs motor component and shock. introduction. trauma systems are multifactorial modules that incorporate any aspect of traumatic injury from the very moment of the injury to the final outcome. a significant prerequisite for the development of a trauma system is the trauma registry. trauma registries are the actual core of any trauma system since they provide valuable information about the standard of care offered to the patients and are amenable to quality control and statistical evaluations, which in turn allow improvements and amendments in the definite care. contrary to what is common practice in the usa, trauma registries in european countries are in embryonic stage. in our country with no actual trauma system, the epidemiology of trauma and the reports on care outcomes are based on police reports and national emergency service reports. objectives. the purpose of this study was to assess the possibility of a national trauma registry in greece and to provide accurate data on the epidemiology of trauma in the country. methods. the project, entitled ''report of the epidemiology and management of trauma in greece'', was initiated in october 2005 and lasted for twelve months. all the national representatives of the hellenic society of trauma were invited to participate. the representatives are certified surgeons employed in hospitals receiving trauma. data presented here are those reported from two tertiary care facilities in athens and twenty eight other primary and secondary hospitals around the country. inclusion criteria were defined as trauma patients with documented need for admission in the hospital, patients that arrived dead or died in the emergency department of the receiving hospital and patients that required transfer to a higher level center. in total 8.862 trauma patients were included in the study in twelve months time. of them 68.7% (n = 6,084) were male, aged 41.8 ± 20.6 (mean ± sd) and 31.3% were female (n = 2778), aged 52.7 ± 24.1 (mean ± sd). as expected and reported in most trauma registries, males are leading in all subcategories of iss. the age group 0-54 years incorporates 68.4% of the total injuries, in accordance to the axiom that trauma is the disease of the young. conclusions. trauma registries are the cornerstone of any trauma system. even in a non-trauma system setting, registries are a valuable tool for quality control of the provided health care and for further development of the health care system. objectives. determine the impact of rurality in epidemiology, injury severity, health care facilities, length of stay (los), mortality, functional outcome and quality of life in trauma patients. retrospective study in trauma patients that were admitted in our emergency room(er) between 2001 and 2008. data was collected from the prospective trauma registry and follow-up registry 6 months after the accident. we classified patients according to statistical national institute classification: urban areas-areas with more than 500 inhab/km 2 or have a place with more than 5,000 inhabitants; semi-urban areas-areas with more than 100 inhab/km 2 and less than 500 inhab/km 2 or have a place with more than 2,000 inhabitants and less than 5,000 inhabitants; rural areas-areas that were not classified as semi-urban or urban areas.patients were divided in three groups according to residence area: r (rural), su (semi-urban), u (urban). we studied several variables in order to find a relation with rurality: sex, age, type of injury, los in hospital and intensive care (icu), anatomic severity (ais), politrauma severity (iss), physiologic severity (rts), surveillance probability (triss index), pre-hospital care, previous admission in other hospital, icu admission and mortality. we report two outcome measures: euroqol to evaluate quality of life and extended glasgow outcome scale for functional outcome. we used qui-square test, t test, mann-whitney test, kruskal-wallis test for statistic analysis. results. 1,311 patients were admitted in the er. 161 patients (12.3%) were excluded with missing data related to residence area. we studied 1150 patients, where 214 patients were from rural areas (18.6%), 219 from semi-urban areas (19%) and 717 from urban areas (62.3%). we find a statistical significant relation between rurality and pre-hospital care, previous admission in other hospital and icu admission. urban area patients had a higher incidence of pre-hospital care(r: 12.2%; su: 17.7%; u: 70.1%, p \ 0.01). semi-urban and rural patients were admitted more frequently in other hospitals before admission in er (r: 89.2%; su: 85.8%; u: 61.9%, p \ 0.01) and also had higher admissions in icu (r: 82.2%; su: 78.5%; u: 72.4%, p \ 0.006). there were no statistical differences in the other variables studied. conclusions. rural trauma patients are similar from those that live in urban areas concerning epidemiology, injury severity and outcome. despite lack of pre-hospital care and higher previous admission in other hospital in rural patients, mortality between groups did not differed in our trauma centre. introduction. metformin-associated lactic acidosis (malta) is a rare but severe complication (0.08/1,000 patients/year) of metformin treatment in type-ii diabetes. metformin impairs neoglucogenesis and liver lactate clearance in the presence of a disease that enhances its production. although frequently used, there is no recommendations regarding hemodialysis in this poisoning. to study the prognostic factors of malta and the interests of blood metformin measurement. . on admission, patients presented profound lactic acidosis with arterial ph 7.19 (6.84-7.31), serum bicarbonate 11.0 mmol/l (6.8-15.6) and plasma lactate 17.2 mmol/l (9.8-19.7). early symptoms associated coma (50%), asthenia (47%), vomiting (27%), abdominal pain (27%), and diarrhoea (20%). renal function was significantly altered [creatinine clearance: 37 ml/ min (14-54); p \ 0.001). all patients received massive alkalinization, 12/16 (75%) were hemodialyzed while 10/16 (63%) were mechanically ventilated and received catecholamines. six patients (38%) died in the icu. duration of icu stay was 3 days [3] [4] [5] [6] [7] [8] [9] [10] . there was no significant differences regarding malta severity and treatments between suicidal and accidental poisonings. neither lactic acidosis severity nor acute renal failure were predictive of death. there was no correlation between prognosis and the time-course of plasma metformin concentrations, with or without dialysis. toxicokinetics showed significant tissue distribution when the patient was early admitted or plateaued concentrations if he was later admitted and survived, even though his situation improved and his lactates decreased. metformin dialysance suggested an interest for extra-renal elimination enhancement although its impact on survival could not be analysed based on this limited study. our study showed that malta is severe with elevated mortality in icu whatever the poisoning is accidental or intentional. metformin toxicokinetics are useful case by case to better understand the patient's outcome. the most important guidelines [1] for trauma care recommend (us)-fast as the first step investigation to rule out major bleedings. however, us is less sensitive and accurate than multislice computed tomography (ct). the spreading concept that ''moderrn cts require little time'' often brings surgeons to ask for total body ct also in haemodynamically unstable patients. to understand how long it really takes to perform a total-body ct in patients suffering from major trauma (mt) we analysed the data of the ritg project, a pilot multicenter study to define the national standards for trauma care and establish a national trauma registry. methods. 3 italian level 1 trauma centers were involved into the first stage of the ritg project. data of all major trauma patients (iss [ 15) who were admitted to either one of the three hospitals during a 12 months period of time (01 july 2004-30 june 2005) were prospectively entered into the ritg database. time between hospital admission and the first scan were recorded for all patients. patients who, for any reason, were submitted to a ct with a delay of 120°or more were excluded. the time elapsed between the first scan and the patient's exit from the ct room was measured in a subset of 235 pts from a single center equipped with a new generation ct next to the emergency room. during the study period 753 mt patients were admitted to the three trauma center. 486 patients were submitted to an emergency total body ct scan within 120 0 . 12 patients died before arrival in the ed. 11 more died soon after admission and before the secondary survey. the interval times are shown in table 1 . seven patients died in the ct room. the average interval between hospital admission and the first available scan was 45°. however, even where a new generation ct next to the er was used, the average time needed to stabilize the patients, get a correct position on the ct and start the scanning process was 34°as an average. in the most severely injured patients, who are frequently artificially ventilated, the time required to stabilize the patient and perform a total body ct scan is longer than expected and to a certain extent, independent from the ct scanner itself unless the very last technology as the sliding ct scanners are employed [2] , thus ct should be considered with extreme caution in the unstable patients. in univariate analyses, survival to discharge was significantly lower with two of 11 acute conditions (acute coronary syndrome and acute inflammation), and with five of 10 chronic conditions (chronic heart failure, diabetes mellitus, kidney failure, hepatic cirrhosis and malignancy). recent surgery was strongly associated with higher odds of survival. the most consistent multivariate predictors of survival to discharge were liver cirrhosis (or 0.32; 95% confidence interval 0.15-0.70) and malignancy (or 0.40; 0.25-0.63). malignancy was not predictive for outcome after cpr attempts, whereas liver cirrhosis was predictive both in all dispatches and in dispatches involving cpr. recent surgery was strongly associated with higher multivariate odds of survival (or 6.1; 2.2-16.4) after cardiac arrest. in dispatches without cpr, absence of chronic conditions was associated with higher likelihood of survival (hr 6.9; 95% ci 5.2-9.3). increasing numbers of chronic conditions were significantly and continuously associated with lower survival (p for trend \ 0.0001). advanced age only weakly predicted survival, but age c75 years was, along with malignancy, the strongest predictor of not attempting cpr in patients with cardiac arrest. conclusions. comorbidities are important determinants of survival after in-hospital met dispatches, with and without cardiac arrest. survival odds are lowest with malignancy and liver cirrhosis. recent surgery increases odds of survival by exclusion of those most severely ill. advanced age at best weakly predicts worse survival, but strongly predicts not attempting cpr. design. retrospective, cohort study. setting. emergency department (ed) and intensive care unit in a university hospital. measurements and main results. the study subjects included of 314 consecutive severe trauma patients. a systematic review of the computer-based medical records of the patients was conducted to provide the base line characteristics and dic-related variables. the worst data of these variables were obtained at 4 time points within 24 h after arrival to the ed; time point 1, immediately after arrival at the ed to 4 h after the arrival; time point 2, 4 to 8 h after the arrival; time point 3, 8 to 16 h after the arrival; time point 4, 16 to 24 h after the arrival. one hundred and forty one patients (141/314, 44.9%) diagnosed as jaam dic showed significant differences in the prevalence of multiple organ dysfunction syndrome (mods) and the outcome in comparison to the non-dic patients. a stepwise logistic regression analysis showed that the maximum jaam dic scores during the study period independently predicted the patient death (odds ratio 1.338, 95% confidence interval 1.070-1.672). all of the patients who developed isth overt dic during the study period could be identified by the jaam dic criteria at early time points. the mortality rate and the incidence of mods of the patients with the isth overt dic were higher than those only met the jaam dic criteria. stepwise increases in the isth overt dic scores and the incidence of the isth overt dic were observed in accordance with the increases in the jaam dic scores. while the mortality rates were identical, there were marked differences in the incidence of mods and sequential organ failure assessment scores between the dic patients associated with trauma and sepsis. conclusions. the jaam scoring system has acceptable validity for the diagnosis of dic at an early phase of trauma. the jaam dic further exists in a dependent continuum to the isth overt dic. in addition to mods, other factors may affect the prognosis of the trauma patients associated with dic. introduction. in the uk, more than 300,000 units of fresh frozen plasma (ffp) are transfused every year. since 2004 there has been a reduction of 16% in its use, but it has been suggested that as many as 47% of transfusions in critical care may be inappropriate. there is significant morbidity associated with the transfusion of ffp. guidelines for the use of ffp do exist, but the indications for its use are limited. coagulation studies, such as prothrombin time (pt), abnormalities are often assumed to be a risk factor for bleeding prior to invasive procedures, but evidence suggests that ffp may not have a prophylactic role. in addition to this the pt or international normalised ratio (inr) were not intended to assess haemostasis in patients without a history of bleeding. review of the blood bank database between 1st january 2008 and 31st december 2008 revealed all prescriptions of ffp for patients on intensive care (itu). the case notes were analysed to find the indication and timings of administration and weight of the patient. the pathology database was examined to find the clotting studies immediately before transfusion. in 2008 98 patients received ffp; this was only 6.8% of the total admissions to the itu. there were 170 prescriptions and a total of 462 units transfused. the mean prescription of ffp was 2.7 units and overall each patient received a mean of 4.7 units. the mean dosage of ffp was 9.96 ml/kg. the pt pre-transfusion mean was 24.0 ± 13.6 s with a median of 19.7 s. the aptt pre-transfusion mean was 52.1 ± 20.0 s with a median of 46.1 s. only 6.5% of transfusions had not had a clotting screen done prior to administration of ffp. 32% of administrations were given prior to procedures being undertaken on the itu and a further 10% were given in preparation of the patient for the operating theatre. a significant number of patients are receiving ffp outside international guidelines. a third of transfusions were given for prophylactic correction of coagulopathy prior to an invasive procedure where there is least evidence for using ffp. most patients received a sub-therapeutic dose of ffp; there is ongoing debate on the correct dosage required to normalise coagulopathy, but it is likely to be greater than 15 ml/kg. [1] . little data exists on the demographics of mt and subsequent demand on a hospital's blood bank. we examined the mt requirements of a 1071 bedded tertiary referral hospital over a 20 month period. objectives. to establish the mt demographic, speciality distribution, prbc demand and associated mortality; within a tertiary referral hospital over a 20 month period. to assist with future mt logistics, planning, implementation and audit. methods. the hospital blood bank database was reviewed for cases of mt from jan 2007 to aug 2008. inclusion criteria were the administration of c 10 units of (prbc) within a 24 h period. cross referencing with the laboratory records and medical notes was undertaken to establish patient demographics, hospital specialty, diagnosis, outcome and number of prbc transfused. results. 59 patients received mt over a 20-month period; 34 male (57.6%) 25 female (42.3%). median age 71 years (range 16-95). median mt of prbc was 12 (range 10-50) units. 881 units of prbc were transfused in the treatment of mt during the study period, accounting for a hospital expenditure of over €175,000. the main specialties associated with mt were the: emergency department (17 patients, 28.8%), cardiothoracic surgery (13 patients, 22.0%), and 11 general surgery (11 patients, 18.6%). 25 of the 59 patients receiving massive transfusions (42.3%), did not survive to hospital discharge. 56% of those patients who died, did so in the first 48 h with a further 4% dying in the next 24 h. 20% of the further deaths occurred within and 20% after thirty days. conclusions. mt in our establishment is associated with a high mortality and predominantly early deaths. recipients were generally elderly with significant co-morbidity. provision of prbcs and blood components for massive transfusion recipients, is challenging for blood bank services [2] . the demand of mt, within our establishment, was predominantly within the acute specialties; emphasizing the need for close communication between them and the laboratory services. in light of this data we propose the implementation of a mt protocol together with continuous audit, to assess its effect on outcome. in 2007 the department of health updated the 'better blood transfusion' circular, a drive to decrease the use of blood products which have become increasingly scarce and expensive [1] . there is evidence that blood product transfusions in icu patients are associated with an increase in morbidity, length of stay and mortality. there has been concern over the increasing use blood products and despite guidelines [2] for their use, both national and local audits have demonstrated a high degree of inappropriate transfusion [3] . derriford hospital is a 1071-bedded tertiary referral centre. the blood bank database was examined for the use of blood products on icu from 1st january 1998 to 31st december 2008. there was a steady rise in icu admissions from 1,131 patients in 1998 to 1,443 patients in 2008. during this time there was a marked decline in both the transfusion of fresh frozen plasma (ffp) and cryoprecipitate. the decreased use of these blood products has occurred with only a very modest introduction of new pro-coagulant therapies, prothrombin complex concentrate (pcc) and recombinant factor viia (rfviia). usage of rfviia and pcc conclusions. our usage of blood products does not reflect the national trends of increasing use of cryoprecipitate and pcc with a small reduction in ffp transfusion, and is more in line with the hsc requirements for better use of blood products. this has been achieved with little use of the newer rfviia and pcc. the evidence for the use of all these blood products is not strong, particularly in critically ill patients. national guidelines exist for their use, but these are poorly adhered to. to test the hypothesis that transfusion of prbcs has a deleterious effect on clinically relevant outcomes in patients with septic shock receiving early goal directed therapy (egdt). retrospective cohort study of 93 patients who presented in an academic center in septic shock and received egdt. data was collected on patients identified via the surviving sepsis campaign chart review database and linked to the project impact database. pearson chi square and fisher's exact test were used to test for clinical significance. primary outcome was mortality and secondary outcomes included mechanical ventilation days, intensive care unit (icu) length of stay, and hospital length of stay. . 89/93 patients presented via the emergency department. 34/93 patients received at least one prbc transfusion during their hospital stay. the two groups were balanced with respect to age, gender, apache ii, and baseline lactate levels. the prbc group had a mortality of 41.2 vs. 33.9% in the no prbc transfusion group (p = ns). the prbc group also had more mechanical ventilation days (11.2 vs. 5.0 days, p = \ 0.05), longer hospital length of stay (25.9 vs. 12.5 days, p = \ 0.05), and longer icu length of stay (11.4 vs. 3.8 days, p = \ 0.05). conclusions. in this study, transfusion of prbc was associated with worsened clinical outcomes in patients with septic shock treated with egdt. this trial is limited by its small sample size and retrospective nature. however, the results are consistent with data from previous trials pointing to a deleterious effect associated with prbc transfusion. further studies are needed to determine the impact of transfusion of prbc within the context of early resuscitation of patients with septic shock, as the beneficial effects gained by an early and goal oriented approach to resuscitation may be lost by the negative effects associated with prbc transfusion. introduction. blood transfusion therapy (btt) is thought as one of transplantation of living cell, that means btt includes several risk such as infection and btt should be thought to derived from precious material by courtesy of donors. patients with traumatic cardiopulmonary arrest on arrival on the hospital (t-cpa) usually suffered from lethal hemorrhage and required rapid supplement of red blood cells for resuscitation of circulation and oxygen transport, that is to say btt. however, the prognosis of t-cpa patients is well known hopeless. the aim of this study is to evaluate the propriety of our strategy concerning btt for t-cpa patients. we retrospectively examined the medical records of t-cpa patients for the past 10 years. we do btt until 1998 (the first period) for t-cpa patients regardless of rosc without any restriction. after then (the second period), we do btt case by case but only after rosc in principle. the rate of rosc, admission to icu, survive to discharge were compared between these two period, and were compared within the first period between the patients group who underwent btt (btt group) and the group who did not underwent btt (non-btt group). in 477 blunt t-cpa and 29 penetrating t-cpa patients, 34 and 59% achieved rosc, 18 and 38% admitted to icu, and 3 and 14% were survive to discharge. in penetrating t-cpa in the first period, 12 units of packed red cells (prc) were used before rosc for 1 non-survivors. in the second period, no prc was used for non-survivor before rosc. in blunt t-cpa in the first period, prcs were used for 29 non-survivors before rosc. in the second period no prc was used for non-survivors before rosc. concerning the effect of btt on the prognosis of t-cpa in all cases, the rate of rosc and admission to icu were statistically higher in the first period than in the second period (p = 0.0003 and 0.032). however, there was no statistical difference in the rate of survive-to-discharge between these periods. there was a same tendency in witnessed cases. in cases with electrical rhythm on the scene, only the rate of rosc were higher in the first period (p = 0.007). restricted in the first period, only the rate of rosc was statistically higher in non-btt group than btt group in all cases, in witnessed cases, and in cases with electrical rhythm on the scene (p = 0.005, 0.008, and 0.003). however, there was no statistical difference in the rate of admission to icu and survive-to-discharge between these groups.. our retrospective serial study showed a possibility that btt before rosc for t-cpa improves the success rate of rosc but add no effect on the improvement of survival rate. btt is thought to be futile for t-cpa before rosc. management of refractory coagulopathy due to adult onset acquired autoimmune haemophilia. d. hendron 1 , g. allen 1 , m. brady 1 , g. benson 2 1 belfast city hospital, intensive care, belfast, uk, 2 belfast city hospital, department of haematology, belfast, uk we report a case of life-threatening haemorrhage occurring as a result of a rare acquired condition caused by the production of an antibody to clotting factor viii. this necessitated administration of recombinant activated factor viia (novoseven) to bypass this step of the clotting cascade. a 71-year-old man presented to intensive care following ogd for acute upper gastro-intestinal haemorrhage, with recent haemoptysis and haematuria. ogd had demonstrated a large clot obstructing the oesophagus and extending through stomach into duodenum. this could not be removed and no bleeding points were identified. a coagulopathy was detected which failed to correct with administration of appropriate amounts of fresh frozen plasma, cryoprecipitate and activated prothrombin complex concentrate (apcc), necessitating clotting factor studies. this demonstrated a factor viii level of 2% with a detectable antibody inhibitor. acquired haemophilia was diagnosed and activated factor viia was administered resulting in rapid correction of coagulation studies and arrest of haemorrhage. it was necessary to continue daily activated factor viia at a dose of 40 mg a day in addition to anti-inhibitor coagulant complex (feiba-vh)-an activated prothrombin complex with factor viii inhibitor bypassing activity. definitive treatment of the coagulopathy was chemotherapy with cyclophosphamide, vincristine and rituximab. this destroyed the factor viii inhibitor and returned his factor viii levels to almost 100%. laparotomy and gastrotomy were required to relieve the oesophageal obstruction from the accumulated clot. he was eventually discharged from hospital and remains well. acquired haemophilia is a rare haematological condition that presents with refractory haemorrhage and coagulopathy and these patients are likely to be referred to critical care services for ongoing support and management. it has an incidence of approximately 1.5 cases per million per year [1] . underlying medical conditions can be identified in up to 50% of patients and include autoimmune disease, solid tumours, lymphoproliferative malignancies and pregnancy [2] . international recommendations on the diagnosis and treatment of patients with this condition have recently been published and advise recombinant activated factor viia to control bleeding followed by a combination of corticosteroid and chemotherapy [2] . the paucity of cases presents an obstacle for randomised controlled trials and therefore these recommendations are based on anecdotal evidence and expert opinion. reference (s) objective. to analyze the application of blood transfusion in critically ill trauma patients after wenchuan earthquake. a retrospective study was made in icu of huaxi hospital on patients who had received transfusion at least once during 1 month after the earthquake. their primary diagnosis and clinical features and apacheii score were obtained at admission. non-active bleeding patients were classified into s group if operation was done during his icu stay, otherwise n group. the function of liver and kidney, and the state of circulation and oxgenation were compared between groups, as well as the hemoglobin level before each transfusion were investigated. a total of 75 patients (52.8%) had received transfusion at least once, among which 55 were non-active bleeding. the average frequency was 9.6 ± 5.7 and 2.0 ± 1.5, amount was 7302.6 ± 5402.8 ml and 921.8 ± 709.9 ml, the incidence of transfusion-related complication was 10.4%(19/182) and 13.0%(14/108) in active and non-active bleeding patients respectively. the apacheii score, mean arterial pressure, ast, serum creatinine, oxgenation index and hemoglobin level on day 1, 7, 28 after admission to icu showed no statistically significant difference between s and n group. the frequency and amount of transfusion were similar also, while the hb level before each transfusion was significantly lower in n group (87.4 ± 17.3 g/l)than in s group (75.6 ± 9.6 g/l) (p \ 0.05). the incidence of transfusion-related and infectious complications, time with ventilator and the 28-day mortality were similar. conclusion. transfusion strategy is more strict in icu doctors than surgeons, while the similar result on organ function, incidence of complications and outcome raises the need for a more wide-accepted transfusion trigger. keywords. earthquake trauma transfusion trigger. extracorporeal life support (ecls) represents an ultimate rescue technique in poisonings. the optimal anticoagulation protocol remains unclear. objectives. we aimed to investigate the coagulation status at ecls decision in order to validate the best heparin protocol to administer. [5 packs (2-12) ] and fresh plasma [4 packs (2-12) ] transfusions were required within the first 24 h. hemorrhages (9/57), thrombosis (3/57) or lower limb ischemia (4/57) seemed equivalent to previous series using more complicated anticoagulation protocols. conclusions. poisoned patients present at ecls time with important alteration in their clotting tests, associated with various degrees of hepatocellular failure, dic, defibrination, as well as dilution. a simple heparin protocol appears optimal to reduce complications in these critical situations. henna is the dried and powdered leaves of the henna plant. the plant is lawsonia alba and the powdered leaves are used to apply decorative designs over the skin. henna application is widely practiced in the arab and asian communities. they create fascinating designs over the skin, especially over the hands and feet. it is widely practiced during wedding ceremonies and at childbirth. g6pd deficiency is common in the community of the arab world. lawsone, the chemical compound in the henna leaves, is capable of inducing severe acute hemolysis in g6pd deficient cases. the compound is chemically related to naphtha. we report a case of acute sever hemolysis in a young girl who presented with dizziness and jaundice and diagnosed to have acute severe hemolysis. her symptoms had started while preparing for her wedding by henna application. the girl was g6pd deficient, and found to have severe hemolysis resulting from henna application on her skin. very few cases have been reported of similar nature. the matter is also of tremendous practical implication in areas of g6pd deficiency. the relevant literature is reviewed as well. background. lactate has prognostic use in critically ill medical and trauma patients, and is a core component in identification of early sepsis. elevated lactate levels in these patients prior to icu admission, e.g. in an a&e setting or pre hospital setting identify patients at risk of death and can trigger an earlier optimization of triage decisions and earlier targeted treatment. a range of poc methodologies for lactate measurement are available but there is little standardization between methodologies. stat sensor lactate is a new poc lactate meter based on a patented multiwell and multilayer electrochemical technology that incorporates control wells that measure and correct for common interfering substances. the electrochemistry technology is layered onto a gold platform providing a stable and robust surface for the electrochemical reaction kinetics. the aim of this study was to assess the performance and functionality of stat sensor lactate. whole blood venous samples were collected from 100 adult patients admitted to a&e. samples were tested for lactate using statsensor lactate (nova biomedical) and the omni b221 bga (roche) routinely used for lactate measurement. precision was assessed using donated whole blood and spiked with a concentrated lactate solution. results. within run precision was acceptable at all levels tested. for the lowest level sample (mean 1.5 mmol/l) %cv for the two meters tested was (6.7 and 7.0%) at the three other levels tested (mean 5.8, 10.4, 23 .9 mmol/l) % cv precision was \4%. lactate values during the method validation ranged from 0.6 to 11.1 mmol/l by the reference method (nova 0.8 to 10.6 mmol/l background and objectives. this research work's intention is to describe the epidemiology in patients suffering from anemia who were interned into emergency room (er). a preliminary work will be conducted in which three days in june will be randomly chosen. during these days, all patients satisfying certain criteria will be registered. the criteria fitted to this work are the following: be using the emergency channel of the hospital, score any diagnostic and be over 14 years old. paediatric, gynaecologic and traumatic cases fall out of this research. anemia was diagnosed according to who criteria. outcome. 861 patients were interned through the aforementioned er channel. 44.6% were subject of blood analysis using classification proceedings. from the latter, 17.7% were diagnosed with anemia. age, intake of clopidogrel and/or aspirin, admission and place of admission resulted statistically significant among anemia patients versus non-anemia. anemia was to be found in 8.5% of patients younger than 65 years old, 23% of the times in patients between 65 and 74 and 30.6% among patients above 74 years old. according to vcm, 41.7% were microcitic-anemia, 48% were normocític-anemia and 10.3% resulted macrocític ones. conclusion. anemia is among a large share of patients coming into the hospital through emergency proceedings. its likelihood increases accordingly to the risk group analysed and dominating among the elderly population and among patients suffering from renal disfunction and non aggregated. most common are normocitic and macrocitic anemia-types. early identification and valuation could bear prognostic consequences. a. s. omar 1 1 tawam hospital, critical care medicine, al ain, united arab emirates introduction. an elevated serum creatinin phosphokinae (cpk) and the presence of myoglobin in the urine characterize rhabdomyolysis. rhabdomyolysis had been described in various traumatic and non-traumatic conditions [1] , there are few reports of its association with anaphylaxis. in this paper, we report 2 cases of anaphylaxis both complicated with rhabdomyolysis. aim of the work. to discus the association between rhabdomyolysis and anaphylaxis and the value of early screening of cpk in such cases. setting. two patients were included in this review in multidisciplinary intensive care unit of tawam hospital/uae. the two patients survived, both developed rhabdomyolysis shortly after admission, evidenced by fivefold or greater increase in serum cpk [2] . both patients had transient hypotension through the presentation, but none of them had persistent shock requiring vasopressors or complicated with acute renal failure. conclusion. we observed rapid increase in serum cpk in our two cases suggesting the potential benefits of early assessment of cpk in such patients which may amplify early goal guided management and avoiding logistic organ dysfunction. keywords. rhabdomyolysis, anaphylaxis. the blood oxygen and carbon dioxide levels are a direct measure of the effectiveness of ventilatory support in patients on mechanical ventilation. head injury patients require strict control of the cerebral homeostatic state. these patients also need careful management of sedation, maintaining a fine balance between patient comfort, hemodynamic instability and ability to assess conscious levels. biphasic intermittent positive airway pressure (bipap) ventilation is thought to be better tolerated by the patient allowing for spontaneous breathing at any point, thus reducing the amount of sedatives and muscle relaxants used. but the effectiveness of this ventilatory mode in achieving stable blood oxygen and carbon dioxide levels in this group of patients is not known. we hypothesised that bipap is more labour intensive to adapt to the target blood gas parameters as the volume delivery is not constant and that the blood gases may be more unstable in the initial resuscitation phase of head injury patients without conferring much advantages in terms of usage of sedatives and muscle relaxants. retrospective data collected from case record review of 13 head injury patients with no primary respiratory insult, requiring mechanical ventilation with volume controlled synchronised intermittent mandatory ventilation (simv) was compared to the data from 13 similar patients treated with bipap ventilation. both the data groups specifically looked at two time periods, the first 24 h and 24-72 h after intensive care admission. blood gas parameters classified as hypocarbic, hypercarbic and/or hypoxic, use of muscle relaxants, number of episodes of raised intracranial pressure (icp) above 20 mmhg as recorded in the intensive care chart every hour, number of episodes of cerebral perfusion pressure (cpp) below 70 mmhg as recorded in the chart every hour was noted. need for muscle relaxant in the first 24 h of admission was noted. the outcome was recorded as either ''alive'' or ''dead'' at the end of itu stay. the data was checked for normality of distribution and compared using non parametric tests (spss 15 for windows). results. baseline characters were comparable between the groups. increased episodes of hypoxia (9.3 ± 12.5 vs. 6.9 ± 12% p = 0.53) and hypocarbia (32.9 ± 18.5% vs. 20.7 ± 20.1% p = 0.046) in bipap mode, compared to simv volume control mode. all measurements being percentages of total blood gases for that patient in the first 24 h. there was no difference in the usage of muscle relaxant (35.7 vs. 66.7% p = 0.214), raised icp, reduced cpp or mortality between the groups. conclusion. bipap mode of ventilation requires more intensive monitoring and changes in ventilatory settings before adapting to the target blood gas parameters in the first 24 h of admission. at the same time the quoted advantage of using less sedatives and muscle relaxants is not significant. acute post-traumatic brain swelling is one variety of the pathological forms, which needs emergent treatment following traumatic brain injuries. we investigated the effects of clinical effects of decompressive craniectomy (dc) in patients with acute post-traumatic brain swelling (bs). seventy-four patients of acute post-traumatic bs with midline shifting more than 5 mm were divided randomly into two groups: dc group (n = 37) and routine temporoparietal craniectomy group (control group, n = 37). the vital sign, the intracranial pressure (icp), the glasgow outcome scale (gos), the mortality rate and the complications were prospectively analysed. the mean icp values of patients in dc group at 24, 48, 72 and 96 h after injury were much lower than those of routine temporoparietal craniectomy group (15.19 ± 2.18, 16.53 ± 1.53, 15.98 ± 2.24 and 13.518 ± 2.33 mmhg vs. 19.95 ± 2.24, 18.32 ± 1.77, 21 .05 ± 2.23 and 17.68 ± 1.40 mmhg, respectively). the mortality rates at 1 month after treatment were 27% in the dc group and 57% in the control group (p \ 0.01). good neurological outcome (gos score of 4 to 5) rates 1 year after injury for the groups were 56.8 and 32.4%, respectively (p = 0.041). the incidences of delayed intracranial hematoma and subdural effusion were 22 and 5%, respectively (p \ 0.05). in conclusion, dc has superiority in lowering icp, reducing the mortality rate and improving neurological outcomes over routine temporoparietal craniectomy. however, it increases the incidence of delayed intracranial hematomas and subdural effusion, some of which need secondary surgical intervention. therefore, the effects of dc in patients with acute post-traumatic bs should be further evaluated. we analyze among others variables: age, injury severity score (iss), abbreviated injury score (ais); admission and discharge glasgow coma score (gcs), extended glasgow outcome score (gose), complications, icu and hospital mortality. differences between groups were tested with students t test and v 2 testing for statistical analysis. results. fourteen patients with intracranial hypertension were treated with decompressive craniectomy . compared with control group, patients with dc had a better gcs (9 ± 5 g1; 5 ± 4 g2 p = 0,004) and gose index not only at icu discharge (4 ± 3 g1; 2 ± 2 g2 p = 0,008) but also at hospital discharge (4 ± 3 g1; 2 ± 2 g2 p = 0.03). the mortality rate was lower in the craniectomy group (g1: 14%, g2; 73% p = 0.05). conclusions. in our center, the use of dc for treat patients with severe tbi and refractory cranial hypertension (gcs b 8 and pic c 25) improved outcome and mortality significantly compared with medical conventional approach. method. in this retrospective study we present 10 patients who underwent decompressive craniectomy following traumatic brain injury at king's college hospital between 2003 and 2008. results. 60% of these patients presented at a&e with a glasgow coma scale of 8 or below whereas the remaining 40% presented with gcs above 8 and deteriorated following admission. the patients underwent decompressive craniectomy to reduce raised icp resistant to medical treatment (barbiturate coma excluded). the procedure resulted in significant decrease in icp. 6 out of 10 patients had the operation within 24 h following their injury. we also found that dc in younger patients (\40 years) was correlated with lower icp following the operation compared to older patients ([40) . our study also showed that early dc (\24 h) is correlated with a shorter stay in itu. conclusions. the findings of the present study are limited by its retrospective nature and small sample size which does not permit any definitive conclusions from these results. however, they form the basis for further investigation. we present the study with a review of the recent literature. introduction. the objective is to study the correlation of secondary icp indices with ct findings and outcome in tbi. a cerebrovascular pressure reactivity index (prx) can be determined as the moving correlation coefficient between mean icp and mean arterial blood pressure . it is a surrogate marker of cerebrovascular reactivity. the rap coefficient was calculated as the running correlation coefficient (r) between slow changes in pulse amplitude (a) and mean icp (p). it is a surrogate marker of pressure-volume compensatory reserve. all components of the icp waveform that have a spectral representation within the frequency limits of 0.05 to 0.0055 hz can be classified as slow waves. methods. prospective observational study of 38 patients with tbi at the royal london hospital. all patients were managed according to the local guidelines for the management of tbi . secondary indices derived from the icp waveform were analyzed by icm ? software. an initial ct was performed in all patients before admission to icu. marshall classification has been shown to predict mortality in tbi. we found a strong association between all these secondary indices and the initial ct findings. all these markers of cerebral haemodynamics correlate significantly with outcome in headinjured patients. conclusions. surrogate markers of cerebrovascular reactivity and pressure-volume compensatory reserve correlates with ct findings and outcome in tbi. these secondary icp indices may be used in the management of tbi. introduction. following the introduction of national guidance [1] on the management of patients with head injury, the use computed tomography (ct) imaging of the head has increased markedly. the impact on anaesthetic and critical care services is unknown. 1. determine the impact of national guidelines on ct scanning in the head injured patient upon anaesthetic and critical care services in a university teaching hospital. 2. determine the incidence of acutely abnormal ct appearances in patients referred for ct scanning under the guidelines. 3. estimate in-hospital mortality in this population and its sub-groups. a case-note analysis was performed in october 2008 of 300 consecutive emergency department (ed) patients who were recorded as having a ct head. of the 300 cases, 10 did not actually have ct head. details of the 290 analysed subjects, indications for the scan and 30 day mortality rates are reported in table 1 . in patients with severe traumatic brain injury pro-and anti-inflammatory mediators are released into the systemic circulation. however, the relationship between the inflammatory response and the kind and duration of secondary insults remains unclear. objectives. the aim of this study was to investigate in severe traumatic brain injured patients the relationship between the systemic concentrations of pro-and anti-inflammatory mediators and the total duration of secondary insults occurring during the icu stay. methods. ten consecutive traumatic brain injury patients admitted to the icu were included. physiological variables were continuously recorded and analyzed minute-by-minute to identify the occurrence of secondary insults (intracranial hypertension, systemic hypotension, hypoxemia and hyperthermia) according to the edinburgh university secondary insult grading scale. serum samples were obtained at admission, 24, 48 and 72 h, in which pro-and anti-inflammatory mediators were analyzed by a bioplex assay. results. ten male patients were enrolled, mean age 35 ± 15, gcs 7 ± 1, apache ii 13 ± 4, iss 32 ± 9. patients were monitored for 3.5 days (median value, range 1-5; 46,442 total minutes recorded); intracranial hypertension occurred for 7,099 min (15.3% of total period recorded, range 0.3-99%), hypotension occurred for 6,176 min (14.8% of total period recorded, range 0.75-43%), hypoxemia occurred for 180 min (0.4% of total period recorded), not enough data were validated for fever. interleukin (il)-6, il-1beta, il-8, il-10 and il-1ra were in the detectable range. a significant correlation was found between the total duration of intracranial hypertension and the median value of il-6 (p \ 0.01, r 2 = 0.48), il-1beta (p \ 0.05, r 2 = 0.37), il-8 (p \ 0.05, r 2 = 0.34), il-10 (p \ 0.05, r 2 = 0.42), and il-1ra (p \ 0.05, r 2 = 0.34) measured during the period of observation. no correlation was found between these inflammatory mediators and the occurrence of hypotension or hypoxemia. no significant correlation was present between the baseline values of these inflammatory mediators and the severity indexes (gcs, iss and apache ii). conclusions. these results suggest that the duration of secondary insults such as intracranial hypertension was associated with a systemic inflammatory reaction, while the severity of injury on admission was not related to the initial concentrations of these inflammatory mediators. grant acknowledgement. aim. assessing behavioral responses to pain is difficult in severely brain-injured patients recovering from coma. we here propose a new scale developed for assessing pain in vegetative (vs) and minimally conscious (mcs) coma survivors: the coma pain scale (cps) and explore its concurrent validity, inter-rater agreement and sensitivity. methods. concurrent validity was assessed by analyzing behavioral responses of 48 postcoma patients to a noxious stimulation (pressure applied to the fingernail) (28 vs. and 20 mcs; age range 20 to 82 years; 31 non-traumatic and 17 of traumatic origin). patients' were assessed using the cps and four other 'pain scales' employed in non-communicative patients: the 'neonatal infant pain scale' (nips) and the 'faces, legs, activity, cry, consolability' (flacc) used in newborns; and the 'pain assessment in advanced dementia scale' (pa-inad) and the 'checklist of nonverbal pain indicators' (cnpi) used in dementia. for the establishment of inter-rater agreement, fifteen patients were concurrently assessed by two examiners. results. concurrent validity assessed by spearman rank order correlations between the cps and the four other validated pain scales was good. cohen's kappa analyses revealed a good to excellent inter-rater reliability for the cps total and subscore measures, indicating that the scale yields reproducible findings across examiners. finally, a significant difference between cps total scores was observed as a function of diagnosis (i.e., vs or mcs). conclusion. the cps constitutes a sensitive clinical tool for assessing pain in severely brain injured patients with disorders of consciousness. this scale constitutes the first step to a better management and understanding of pain in patients recovering from coma. methods. study group: 75 consecutive patients with cervical spinal cord injury admitted to icu. mean age: 35, 4 years. 51 patients asia a, 18 asia b, 6 asia c. the more frequent neurological level was c6 (50%). the requirement of mechanical ventilation was considered the key sign for establishing the diagnosis of severe respiratory failure. the blood gas values (po 2 , pco 2 , and pao 2 /fio 2 ) before and after connection to mechanical ventilation [mv(if needed)], were used to estimate the more probably mechanism of respiratory insufficiency. the increase of pco 2 levels was considerate as a sign of neuromuscular weakness; the low po 2 level before ventilation, and the persistence of pao 2 /fio 2 below normal values was considered a sign of v/q mismatch. for this purpose statistic analysis (mean values comparison using student t test) comparing blood gases before and after mechanical ventilation treatment was performed. results. 39 (52%) patients developed severe respiratory failure. mean delay between admission and mechanical ventilation was 108 h. previously to mechanical ventilation 22 patients developed pulmonary atelectasis, and four pneumonia. the incidence en respiratory failure was significantly higher in patients with neurological level above c6 (p \ 0.05). conclusions. the incidence of respiratory failure is related with the severity of neurological deficit (relationship between incidence of respiratory failure and neurological deficit level). in addition, our data support that, besides the neuromuscular weakness (moderate increase of co 2 levels), a significant v/q mismatch with shunting phenomena associated (significant hypoxemia no completely solved after mv) is involved in the respiratory failure of cervical spinal cord injured patients. 1. moderate and severe traumatic brain injury is more likely in middle aged men; more than one third present other major trauma and intensive first level medical treatment is required in most of them. 2. the most frequent complications found were infectious diseases like ventriculitis and vap. 3. independent mortality risk factors in moderate and severe trauma brain injury were age, high apache ii score, neuromuscular blocking drugs and icu los. 4. outcome was significantly improved after six months, and most of the patients only present mild disability and good recovery. nosocomial infections are leading causes of increased morbidity and mortality of severe brain injured patients [1] . the mechanism underlying the susceptibility to the infections is a subject of great scientific interest and still to be clarified [2] . it has been recently recognized that injury of brain induces a disturbance of balance between the central nervous and immune system [3] . objective. the aim of this study was to investigate changes in frequency of lymphocytes subpopulation in peripheral blood of patients with severe brain injury during the course of intensive care treatment. human peripheral blood samples were taken from the severe brain-injured patients at day 1, 4 and 7 and peripheral blood mononuclear cells (pbmc) were immediately isolated by gradient density centrifugation. the percentage of lymphocytes subpopulation were analyzed by simultaneous detection of surface antigens using fluorochrome conjugated monoclonal antibodies directed toward cd3, cd56, cd16, cd4, cd8, cd5, cd19. t lymphocytes were distinguished from the other lymphocyte subpopulation as cells labeled with anti-cd3 monoclonal antibody but negative for cd56 staining (cd3? cd56patients. eighty-seven patients with head injury, glasgow coma scale \ 9. measurements and main results. clinical and demographic data, and head ct scan were taken at admission. patients underwent advanced neuromonitoring and were treated according to brain trauma foundation guidelines. s100b concentration was quantified at admission and 24, 48 and 72 h post-tbi (days 0, 1, 2 and 3). outcome was assessed 12 months after discharge using glasgow outcome score. significant negative correlations were found between 1-year gos and s100b concentrations on days 1-3, but not on day 0 (day 0, p = 0.1; day 1, p = 0.003; day 2, p \ 0.0005; day 3, p \ 0.0001). patients who deceased showed higher s100b concentration than survivals for all the samples. good versus poor outcome (gos = 1-3) differed significantly on days 2 and 3. logistic regression analysis showed that samples 24, 48 and 72 h post-tbi sample predicted death outcome. roc curve analysis showed 72-h sample was the strongest predictor for decease. poor outcome was only predicted by the 72-h sample. conclusions. s100b levels 72 h post-tbi was the strongest predictor for poor and fatal 1-year outcome, whereas levels at admission do not. a temporal profile of s100b release from admission to 72 h post-tbi is strongly recommended for use in identifying the subset of patients liable of developing a worse outcome. according to our results, s100b protein might be an early, sensitive, accurate and useful biomarker for predicting long-term outcome in patients with acute severe tbi. grant acknowledgement. this research was made possible in part by the generous donation of protein s100b electrochemiluminescence assay kits by roche diagnostics, mannheim, germany. introduction. brain intercellular fluid glycerol concentration as measured by microdialysis catheters has been recognized as an index of glial and neuronal cellular destruction. we present a data analysis correlating glycerol levels with intracranial pressure (icp), cerebral perfusion pressure (cpp), brain tissue oxygen partial pressure (pbtio 2 ), lactate to pyruvate concentration ratio (l/p) and outcome. methods. data of 30 head injured patients is presented. all had simultaneous monitoring of icp, pbtio 2 and metabolic biochemistry by three brain intraparenchymal bolt catheters inserted via the same one burrhole (icp codman or camino, pbtio 2 licox and microdialysis-cma). there was not a clear straight correlation of raised glycerol levels with bad outcome. however, glycerol elevation seemed to be a predictor of intracranial hypertension together with l/p raise. in subarachnoid hemorrhage patients glycerol elevation was an early sign of secondary ischemic insult. conclusion. multimodal monitoring with intracranial catheters is a useful clinical tool for management of critical neurosurgical patients. metabolic biochemistry as measured by microdialysis, and specially l/p and glycerol levels, can early predict incoming intracranial hypertension as well as secondary ischemia. the pulsatility index (pi), a parameter derived from the blood velocities along the cardiac cycle, has been used as an indirect way to evaluate intracranial pressure. the aim of this research has been to evaluate the accuracy of transcranial doppler sonography (through pulsatility index) in the inference of intracranial pressure. methods. population of the study group (high-pi-group): 50 severe head injured patients (gcs at admission \ 9; mean age 37.7 years; patients with diffuse injury (traumatic coma data bank) type ii (43%) and iii (29%)) who presented episodes of increase of pulsatility index (pi [ 1.2) in the acute phase of head injury. control group (normal-pi-group): 50 severe head injured patients, with tcd recordings of normal pi (pi b 1.2). in all the patients the intracranial pressure (icp) was continuously monitored using a intraparenchymal device. all the tcd recordings are referred to the middle cerebral artery of the cerebral hemisphere were icp catheter was inserted. in the transcranial doppler recording, the pulsatility index was automatically calculated derived from the formulae: pulsatility index = (systolic velocity -diastolic velocity)/mean velocity. 100 transcranial doppler sonography recordings of with pulsatility index c 1.2 (high normal value of pulsatility index) were correlated with the simultaneous icp value. the incidence of intracranial hypertension (icp [ 20 mmhg) was analyzed in the high-pi-group, and compared with the incidence of intracranial hypertension in the normal-pi-group. methods. in a double-blind, randomized, placebo-controlled clinical trial, 80 patients scheduled for elective cabg was randomly assigned into two groups. after matching inclusion and exclusion criteria and induction of general anesthesia, one group received intrathecal sufentanil (s) and the other group received the same dose of sufentanil plus supplemental bupivacaine (sb). except for this, all the cases were similar regarding anesthesia and surgery. mean arterial blood pressures were measured before and after induction of anesthesia, during the bypass time and after weaning from bypass were checked. also, the need of the patients for administration of inotropic agents after weaning was compared. results. there was more stable mean arterial blood pressure and less inotropic need after weaning from cardiopulmonary bypass in the sb group. also, the sb patients had a more stable hemodynamic profile during the bypass period; especially after the initiation of the bypass. less inotropic agents were needed after weaning in the sb patients. there was no difference between the two groups regarding the extubation time. discussion. the administration of intrathecal sufentanil plus bupivacaine seems to keep the hemodynamic status of the patients more stable than intrathecal sufentanil alone. methods. this study was approved by the hospital 0 s ethics committee. prospective observational study including 20 consecutive patients. preoperatory and postoperatory data were collected. interventions included blood samples for nt-pro bnp taken prior to operation, and 18 and 36 h in postoperative. troponin-i was taken 12 and 36 h postoperatively. blood obtained was processed for nt-probnp with cobas h 232 system ò point of care (poc) by roche diagnostics, with range from 60 to 3,000 pg/ml. the serum nt-probnp level was also correlated with the logistic euroscore and ejection fraction (ef). serum ntpro-bnp and troponin i values were compared between patients with and without postoperative length of stay in the intensive cardiac unit (icu) [ 48 h. and hospital [7 days. all results are in median ± sd * p \ 0.05, **p \ 0.01 tables 1??? and 2??? conclusions. preoperative euroscore and nt-probnp levels were higher in patients with ef \ 40%. the troponin i after surgery increased more in patients whose length stay in icu was longer. after surgery nt-probnp levels increased significantly,and they differ significantly between patients with length stay in icu for more than 48 h and 7 days at hospital. our data collection confirmed that measurement of nt-probnp is useful and helpful during postoperative period and it also predicted a higher possibility for a long stay in icu and a later hospital discharge. however, owing to the small size sample, these results must be regarded as preliminary. conclusions. in spite of the limitations of our trial, percutaneous aortic valve implantation appears to be safety. a high rate of maccv events were observed, essentially due to a disruption of the a-v conduction, in most cases transitional. despite the definition of ''inoperability'' is difficult, less-invasive aortic valve procedures will undoubtedly find a place within current cardiac surgical practice. objective. to describe the evolution of cardiac transplant patients, presenting clinical low cardiac output in the immediate postoperative period, and after handling routine, they are treated with levosimendán (lv). descriptive, prospective and observational in a postoperative care unit for cardiac surgery from a terciary hospital. study period: january 2006-december 2008. lv was used when the patient had inotropic dependence over 72 h, to try to remove the amines or added to them in those cases that do not get these drugs with an adequate hemodynamics. bolus was used in 4 occasions and then infusion of 0.1-0.2 mcg/ (kg min). we analyzed demographic variables, hemodynamic response to the input of the drug if you can reduce or discontinue other medications, clinical tolerance and side effects, overall development, the icu and hospital stay. we studied 13 patients (6 women and 7 men). presented a mean age of 48.85. before surgery, all of whom were in nyha functional class iii-iv. three patients were transplanted in emergency. in this series, there is a case without pulmonary hypertension (pah) pre-transplant, 5 patients with mild htp and htp 7 moderate to severe, with a transpulmonary gradient(gtp) between 9 and 21 mmhg. the 3 patients with gtp [ 15 mmhg had a positive reversibility test with sildenafil. ischemia time of surgery was 195.46. in the immediate post, all the patients studied had low cardiac output syndrome by graft postoperative ventricular dysfunction, cardiac index measured by pulmonary artery catheter. in all patients echocardiography was performed to rule out a pericardial effusion with hemodynamic deterioration in cardiac cavities and showed ventricular dysfunction, right dominance in 10 patients. in all patients we observed a good tolerance to the drug. in 11 lv cases facilitated the withdrawal of the remaining. 2 patients were used lv only after the withdrawal of treatment with inotropic dependence on it. in the remaining cases to be associated with other drugs. only two cases could not withdraw inotropic treatment after the lv infusion. in five patients with pulmonary arterial hypertension and prevalence of right ventricular failure, to reduce poscarga also added pulmonary arterial vasodilators. patients have a stay in icu between 4 and 6 days. one patient mortality. 1. the primary graft failure is a severe potential complication of post-cardiac, which is associated with a worse prognosis. 2. lv shows good tolerance, without serious adverse effects attributable to the drug, and facilitated the removal of amines and clinical recovery. 3. it is necessary to expand the case to confirm the results, and to establish the most appropriate indications and patterns of use of this drug. post-infarction ventricular septal defect (infarctvsd) is a rare but serious complication of myocardial infarction, usually quickly followed by low cardiac output. repair of infarctvsd is still a challenging procedure with a high risk of mortality. improvement of surgical outcome depends on results of large studies in this setting. the aim of this retrospective study was the evaluation of preoperative and surgical parameters influencing the 30-day mortality following surgical repair. conclusions. in this large study, pre-operative left ventricular function and troponin level were found to be the best predictors identifying patients at high risk for 30-day mortality following surgical closure of infarctvsd. both parameters may be helpful in deciding on the time of the operation and preoperative preparation. in contrast to other findings, in our cohort the location of the vsd (anterior vs. posterior) did not affect mortality. this may be due to improvement of surgical technique and perioperative management over time. adequate fluid therapy is the first step of hemodynamic optimization after cardiac surgery [1] . cardiac surgery exposes patients to ischemia and reperfusion, which are well known risk factors for a systemic inflammatory response and increased capillary permeability in the lungs [2] . it is still unclear what type of fluid should be given in the presence of increased pulmonary vascular permeability at hypovolemic status. objectives. aim of this study was estimate the optimal type of fluid for intravascular volume deficit treating without evoking pulmonary oedema. a prospective clinical study at the intensive care unit was performed on 24 mechanically ventilated patients within 1 h after elective cardiac surgery involving cardiopulmonary bypass. patients, divided into four groups, were subjected to fluid challenge according to the global end-diastolic volume index (gedvi) measurements with normal saline 1,000 ml or the colloids 4% gelatin, 6% hes 130/0.4 or 5% albumin 250 ml in 20 min. hemodynamic and extravascular lung water index (evlwi), gedvi measurements were performed exactly before fluid challenge, afterwards and 30 min after challenge. results. the change in evlwi did not differ between saline and colloid fluid challenge. gedvi increased by 12% in saline group, by 15% in 4% gelatine, 16 in 6% hes 130/0.4 and 17 in 5% albumin. conclusions. all colloid fluid infusion leads to the greater increase in cardiac preload compare to normal saline (saline in four times larger volume). the change in evlwi did not differ between saline and colloid fluid groups and did not increase pulmonary oedema despite in the presence of increased pulmonary vascular permeability, when fluid overloading is prevented. introduction. the annual incidence of prosthetic valve thrombosis is up to 1-2% (patients-year) despite the anticoagulant therapy. conventionally, the treatment of choice for this event was the surgical valve replacement. however, fibrinolytic therapy has become a valid alternative for the treatment of this serious complication, especially in high-risk surgery patients. to analyze the clinical factors, diagnosis and treatment management of patients with prosthetic valve thrombosis admitted to the acute cardiac care unit. we designed an observational-descriptive study, including patients admitted between 1998 and 2008. clinical factors were analyzed: sex, age, prosthetic valve position, time from valve replacement, inr at admission, clinical features, diagnostic technique and treatment used. results. 14 patients were included. 35.7% were women, 64.3% men. mean age was 57.2 ± 4.15 years. the highest incidence was at the tricuspid prosthetic valve position (57.1%), followed by the mitral (35.7%) and the aortic position (7.1%). when a triple valve replacement was performed, the tricuspid position was the most often affected. mean time from the first valve replacement surgery was 5.9 ± 2.7 years. clinical features which led to the diagnostic were: acute heart failure (71.42%), peripheral embolization (14.28%), chest pain (7.14%) and syncope (7.14%). the diagnostic techniques used were transesophageal echocardiography (tee) and cinefluoroscopy in all the patients. inr at admission time was lower than adecuate anticoagulation recommendations in 71.42% of patients. the most widely used treatment was the systemic fibrinolytic therapy (78.57%), followed by surgery (14.28%) and conservative treatment with heparin alone (7.14%). the most widely used thrombolytic was rtpa in 90.9% of patients, with a mean dosage of 89.3 ± 15.4 mg. one patient was treated with 1.4 mil. ui of streptokinase. unfractionated heparin was added to all patients whom received fibrinolytic therapy, with a mean dose of 683 ± 331 ui/h. a 57.1% incidence of minor bleeding was found in the fibrinolytic group. there were no major complications due to fibrinolytic. total mortality rate was 14.28%. our experience, suggests that systemic fibrinolytic therapy is safe and effective in patients with prosthetic valve thrombosis. objective. to describe the outcomes of patients with acute, refractory, non-ischaemic and not postcardiotomy, cardiogenic shock treated with extracorporeal membrane oxygenation (ecmo) and to evaluate whether survivors and non-survivors differed with respect to clinical characteristics, pre-ecmo treatment and laboratory values. design. in this retrospective cohort study, information is collected from a database with additional review of medical records. patients. 20 consecutive adult patients, 14 males, mean age 39.7 ± 12.7 year, presenting to hospital with non-ischaemic acute severe, refractory cardiogenic shock, supported by central or peripheral venoarterial (va) ecmo. measurements and main results. characteristics of survivors and non-survivors were compared using chi square test. twelve patients (60%) were transported to our institution on ecmo. eleven patients (55%) were weaned from ecmo, seven (35%) bridged to ventricular assist devices. in two patients (10%) ecmo support was withdrawn. mean duration of ecmo support was 145.5 ± 69.7 h. overall survival was 70%, and did not differ between patients with myocarditis (n = 10), cardiomyopathy (n = 7) and acute on chronic non-ischaemic cardiogenic shock (n = 3). a larger proportion of the three patients with 2 or more complications died as compared to the seventeen patients with less than 2 complications (67% versus 24%, p = 0.133). pre-ecmo intra-aortic balloon counterpulsation (iabp) was used in 11 patients, 91% survived, as compared to 44% of those who did not receive iabp (p = 0.024). we have not identified any other significant differences between survivors and non-survivors. conclusion. the survival of patients on ecmo in this unique heterogeneous patient cohort is similar to the survival of ecmo support for fulminant myocarditis in the literature. we recommend to institute ecmo early in all medical patients with acute non-ischaemic cardiogenic shock, refractory to conventional therapy, or to refer these patients in time to an ecmo centre. introduction. human parvo b19 virus is associated with a broad spectrum of clinical manifestations, mostly in children or immune-compromised patients. in adults, severe myocarditis due to this viral agent is a rare disorder, presenting as acute congestive heart failure. we describe a patient with rapidly progressive heart failure, needing circulatory support by extracorporeal membrane oxygenation (ecmo). methods. case report. a 24-year-old previous healthy female was admitted to our icu with nausea, vomiting, bradycardia and hypotension with a blood pressure of 80/40 mmhg. two weeks before admission, patient had signs of erythema infectiosum. on physical examination the patient was pale, with venous congestion, third heart sound and hepatomegaly. the initial electrocardiogram showed a slow, regular, ventricular rhythm. admission chest x-ray showed normal heart size with bilateral pleural effusion. echocardiography revealed dilated ventricles (rv and lv) with depressed systolic function and a thrombus in the rv apex. patient was initially treated with intravenous medical therapy, but unfortunately developed progressive cardiogenic shock. troponin levels, serum transaminases and bun were extremely elevated. it was therefore decided to implant a percutaneous ecmo by femoro-femoral cannulation which permitted to stabilize hemodynamic conditions while peripheral organ functions returned to normal range. progressive cardiac recovery was observed after 7 days with a circulatory assistance with a mean flow rate of 4.1 l/min. as myocardial function improved, ecmo was gradually weaned and removed after 10 days of support. however, atrioventricular conduction did not recover, necessitating implantation of temporary vvi-pacemaker, which was later replaced by a permanent ddd pacemaker system. pathology of the endomyocardial biopsy showed extensive lymphocytary infiltration with destruction of myocytes. parvo b19 dna-pcr was positive in both the biopsy and serum. these findings suggest that this patient developed severe myocarditis induced by parvo b19 viral infection. to our knowledge, parvo b19 viral infection is an uncommon cause of severe myocarditis in adult patients. sparse literature is available describing the use of ecmo in these adult patients. conclusion. this case report shows that parvo b19 virus should be recognised as a potential infective agent in adult patients presenting with severe myocarditis. furthermore, ecmo can be safely used to stabilize hemodynamics and peripheral organ perfusion in expectation of myocardial recovery in these patients. copeptin is easier to measure than vasopressin, and could be used as a marker of vasopressin release [1] . the aim of the study was to compare plasma concentration of avp and cop during cardiac surgery, and specifically during post cardiac surgery vasodilatory syndrome (pcsvs). methods. two-month consecutive patients scheduled for cardiac surgery with cardiopulmonary bypass (cpb) were included in the study except patients suffering from chronic renal failure and under dialysis. blood samples were obtained from blood withdrawals routinely operated before cpb, during cpb and after surgery, at the postoperative hour 8 (h8). these samples were used for avp and cop measurements. pcsvs, assessed as hypotension unresponsive to volume replacement therapy and without cardiogenic shock features, was treated with norepinephrine (ne). patients treated with ne have been compared to the others. statistical test consisted of variance analysis, non parametric test (mann whitney or wilcoxon) and linear regression. a p value of less than 0.05 (p \ 0.05) was considered statistically significant. results. 64 patients have been included, out of which 10 have been treated with ne. correlation between avp and cop plasma concentrations is significant (r = 0.70, p \ 0.01). avp and copt concentrations increased significantly at h8 but the increase is less pronounced in ne-treated patients (fig. 1) . ne-treated patients had lower preoperative left ventricle ef (45.0 ± 14.9 vs. 55.5 ± 10.5%, p = 0.03), longer cpb (123. 1 ± 46.4 vs. 200 .0 ± 80.7 min, p \ 0.01) and clamping times (87.7 ± 41.5 vs. 157.1 ± 70.2 min, p \ 0.01), higher incidence of low output syndrome (8/ 10 vs. 4/54, p \ 0.05) longer extubation time (27.5 ± 14.8 vs. 8.0 ± 7.0 h, p \ 0.01) and higher plasma cop before (t0) and during cpb (fig. 1) . avp (ng/ml) et copeptin (cop, pmol/l), in patients. *p \ 0.05 ne versus others discussion. correlation between avp and cop is similar to that observed in other studies [1] . the correlation coefficient is rather weak that is possibly related to avp dosage limitations (binding of avp to blood platelets, lack of antibody-specificity). increased cop plasma concentrations before and during cpb is observed in sicker patients undergoing more complex surgery, which seems to expose them to relative postoperative vasopressin deficiency and pcsvs. background. waiting list for heart transplantation has been growing up. high doses of cathecolamines has been an exclusion criterion for heart donation and norepinephrine use is still controversial. to assess if norepinephrine used on heart donors modify receptors outcome. methods. historical cohorts study from april 1991 to march 2007. patients were divided in two groups: group 1: patients with local donors treated with norepinephrine (n = 58). group 2: patients with local donors managed with other cathecolamines (n = 39).cathecolamines were used at least for 1 h and doses were between 0.05 and 5 mcg/(kg min) if norepinephrine and between 1 and 20 mcg/(kg min) if dopamine or dobutamine. mortality risk factors published on the last international society for lung and heart transplantation guidelines were recorded. graft dysfunction risk factors were also collected. heart transplant outcome was measured by 30-day mortality, mortality rate at first, second, fifth and tenth years; and graft dysfunction incidence. chi-squared and t student test was used. multivariate logistic regression was used to evaluate norepinephrine impact on the outcome. mortality in group 1 was 36.2 and 41% in group 2. no differences in mortality or graft dysfunction incidence were found in multivariate analysis. conclusions. norepinephrine used for donors management compared with dopamine and dobutamine does not increase mortality or graft rejection incidence in heart transplantation. groups were not uniform so further studies may be made to determine this association. introduction. coronary artery bypass surgery on cardiopulmonary bypass is associated with significant morbidity and mortality. with present technology, all arteries on the heart can be bypassed off-pump. the benefit of this technique is higher for patients whom are at increased risk of complications from cardiopulmonary bypass, such as those who have heavy aortic calcification, carotid artery stenosis, prior stroke, and compromised pulmonary or renal function. to evaluate the short-term follow up results of off-pump coronary artery bypass (opcab) and postoperative management of these patients admitted to our coronary care unit. we designed an observational study that included patients who underwent opcab from july 1998 to december 2008. data were collected on preoperative age, sex, major cardiovascular risk factors, history of prior ami, number of affected vessels and ventricular function. after the surgery we evaluated: the extubation time, postoperative bleeding, troponin maximum level, need for blood transfusion, use of vasoactive drugs and intra-aortic balloon pump, development of renal failure, atrial fibrillation, neurological complications and reintervention. results. 69 patients were included. 79.9% were men and 20.3% women. mean age was 65.3 ± 8.7 years. 100% of patients had one or more cardiovascular risk factor: hypertension was present in 73.9%, smoking 55.1%; diabetes mellitus 37.7% and dyslipidemia in 62.3%. there was prior myocardial infarction in 53.6% of patients. prior coronary angiography showed 26.1% of patients with 2 vessels disease and 73.9% of 3 vessels disease. mean lvef was 53%. mean number of grafts was 1.7. mean extubation time was 9.6 h. mean postoperative bleeding was estimated in 410 cc. 37.7% of patients needed blood transfusion; 36.2% vasoactive drugs; and 2.9% needed an intra-aortic balloon pump. 39.1% of patients developed troponin t elevation with a mean level of 6.3 ng/ml. 23.3% of patients developed atrial fibrilation, and 10.1% renal dysfunction (two patients needed hemodialysis). there was no neurological complications. 1 patient needed a reintervention. mean of intensive care unit stay was 3.7 ± 5.3 days. total mortality rate was 2.76%. our experience shows that the off-pump coronary artery bypass graft surgery is a safe and effective technique for coronary revascularization, with low mortality and morbidity rates and reduced postoperative complications. objectives. to assess if deterioration of left atrial function in patients with severe sepsis and septic shock could predict mortality. we studied 30 patients with severe sepsis or septic shock with mean age of 49.8 ± 16.17. underlying echocardiographic parameters were measured on admission, 4th and 7th day, which comprised left ventricular ejection fraction (ef), and atrial function which is expressed as atrial ejection force (aef), with aef defined as the force that the atrium exerts to propel blood into the left ventricle (lv). all patients were subjected to bnp assay well. multivariate analyses adjusted for acute physiology and chronic health evaluation score ii (apache ii score) was used for mortality prediction. results. underlying source of sepsis was lung in 10 patient (33%), blood in seven patient (23.3%), abdomen in seven patients (23.7%), while three patient (10%) had urinary tract infection (uti) as a cause of sepsis. only one patient had cns infection. severe sepsis was admission diagnosis for 20 patients, 10 patients were labeled as septic shock. look for 28 days mortality. in-hospital mortality was 23.3% (7 patients) . admission ef showed significant difference between survivors and non-survivors 49.01 ± 6.51 versus 56.44 ± 6.93% (p \ 0.01), on the other hand admission aef showed insignificant changes between the same groups 10.9 ± 2.81 versus 9.41 ± 2.4 k/dynes p = 0.21, while bnp was significantly higher in the non-survivors 1,123 ± 236.08 versus 592.7 ± 347.1 pg/ml (p \ 0.001). multivariate logistic regression, the predictable variables for mortality was apache ii score, bnp then ef. conclusion. in septic patients, left atrial function unlike the ventricular function and bnp levels cannot be used as independent predictor of mortality. objectives. to analyse the relationship between plasma levels of nt-probnp and lcd diagnosed by echocardiograph during ss. methods. prospective observational cohort study. inclusion criteria: consecutive patients with ss [3] . non inclusion criteria: creatinine clearance \ 60 ml/min, 60 years \ age \ 18 years, cardiac surgery patients, pre existing coronary or cardiac insufficiency, neoplasia and systemic diseases. the evaluation of the left ventricular function was realised by a trans-thoracic or a trans-oesophageal echocardiograph on day 2. the lcd was defined by a left ventricular fraction of ejection \40% evaluated by teicholtz. the blood tests for nt-probnp analyses were drawn on days 0, 2, 4 and 7. serum nt-probnp measurements were made automatically by elecsys 2010 analyser with the truss nt-probnp (roche diagnostics, myelan, france) by the electrochemiluminescence immunoassay method (eclia). data are expressed as mean ± sd and percentages. statistical analysis was performed by repeatedmeasures anova and roc curves (p \ 0.05 indicated statistical significance). . 36 patients were included in a period of 30 months (medical patients n = 16, surgical patients n = 7 and trauma patients n = 13), age = 53 ± 10 years, bmi = 24 ± 2 kg/m 2 , apache ii = 15 ± 9, igs ii = 35 ± 16, duration of intensive care unit stay = 24 ± 32 days, mortality = 50%. lcd was observed in 11 patients. the statistical analysis showed a significant elevation of nt-probnp in patients with lcd (table 1) . on day 2, the area under roc curve was 0.79, and the cut off value of nt-probnp predictive of lcd was 2,600 pg/ml (sensibility = 72%, specificity = 74%). introduction. fluid responsiveness can be predicted by the respiratory variation of arterial pulse pressure (ppv) or of pulse contour-derived stroke volume (svv) as well as by the changes in pulse contour-derived cardiac index during a passive leg raising manoeuvre (plr) or a tele-expiratory occlusion (teo). we evaluated the ability of an infrared photoplethysmography arterial waveform (cnap device) to estimate ppv. we also tested the ability of this non invasive estimate of ppv to predict fluid responsiveness compared to the invasive measure of ppv, to svv and to the plr and teo tests. in 29 patients with septic shock (63 ± 13 years of age, 25 receiving norepinephrine, saps2 = 65 ± 22, lactate = 3.3 ± 3.1 mmol/l), we measured the response of cardiac index (pulse contour analysis, picco device) to fluid administration (500 ml saline over 20 min). before fluid administration, we recorded the ppv directly calculated from the non invasive arterial pressure signal (ppv ni ), the ppv directly calculated from the invasive arterial pressure signal (ppv i ), the ppv automatically provided by the picco device (ppv picco ), the svv automatically provided by the picco device, the changes in cardiac index induced by a plr test and the changes in cardiac index induced by a 15-s teo. results. five patients were excluded because the arterial curve could not be obtained by the cnap device due to excessive vasoconstriction. in the remaining 24 patients, fluid administration increased cardiac index by more than 15% (31 ± 19%) in 8 ''responders''. the fluid-induced changes in invasive (?20 ± 36%) and non invasive (?20 ± 29%) mean arterial pressure were correlated (r = 0.79, p \ 0.05). at bland-altman analysis, ppvni accurately reflected ppvi (bias 1%, limits of agreement ±8%). for predicting fluid responsiveness in the 24 patients, the receiver operating characteristics (roc) curves for ppv ni , ppv i , ppv picco , svv, plr and teo were 0.83 ± 0.10, 0.82 ± 0.10, 0.86 ± 0.09, 0.77 ± 0.11, 0.92 ± 0.07, 0.94 ± 0.06 (all non significantly different). when considering only the 15 patients ventilated with a tidal volume b7 ml/kg predicted body weight, 2 were falsely classified as non responders by ppv ni , ppv i and two others by ppv picco and svv, but all four were well classified by plr or teo. in septic shock patients, provided that vasoconstriction is not excessive, the non invasive assessment of arterial pulse pressure seems valuable for predicting fluid responsiveness. introduction. mechanical ventilated patients often require inotropic support. however, the role of mechanical ventilation (mv) in myocardial depression is not well understood. septic patients often have impaired cardiac function and are in need of mechanical ventilation. we hypothesized that mv enhances sepsis-induced myocardial depression. objectives. in this study we investigated the influence of mechanical ventilation on cardiac function in an acute sepsis model. sepsis was induced in male wistar rats using ip injection of lps. healthy and septic rats were randomized to one of three ventilation groups; (1) non-injurious ventilation with a tidal volume of 6 ml/kg and 5 cm h 2 o peep (low tidal volume, ltv), (2) injurious ventilation with a tidal volume of 19 ml/kg and 5 cm h 2 o peep (high tidal volume, htv) and (3) spontaneous breathing. arterial pressure was kept at least at 60 mm hg. cardiac output (co, thermodilution method), central venous pressure (cvp) and mean airway pressure were measured in vivo. after 4 h of ventilation, animals were sacrificed and cardiac function was measured ex vivo in a langendorff setup and expressed as developed pressure and ?dp/dt. cardiac wet to dry weight ratio was calculated. results. cardiac output in vivo was lower during htv ventilation than during ltv ventilation (p \ 0.001). cvp did not differ between ventilation strategies while mean airway pressure was higher in htv ventilation than in ltv ventilation (p \ 0.001). ex vivo, cardiac function of septic animals was depressed compared to healthy controls (p \ 0.001) in septic animals, cardiac function was better in htv ventilated animals than in non ventilated animals (p \ 0.05). ventilation lowered cardiac wet/dry ratio (p \ 0.05). developed pressure (p \ 0.01) and ?dp/dt (p \ 0.05) correlated inversely with cardiac wet/dry ratio. [1] . perfusion may be also evaluated by other parameters such as lactate or venous-arterial pco 2 gradient (delta pco 2 ). objectives. to evaluate if early normalization of scvo 2 after emergency intubation in septic patients persists over time and if it is associated with similar trends in lactate and delta pco 2 . methods. ten septic patients subjected to emergency intubation for respiratory or circulatory failure and in whom scvo 2 increased to [65% after the procedure. these patients were included in a large prospective study published elsewhere [1] . patients used a common intubation protocol and we evaluated several perfusion related parameters before, 15 min and 6 h after emergency intubation. statistical analysis included friedman and wilcoxon tests. results. evolution of perfusion parameters after intubation is presented in table 1 . five patients died during icu stay. as a whole, scvo 2 remained stable in 5 pts and decreased dramatically at 6 h by[20% in 3 non-survivor patients (lowest 29%). only 2 pts had a high lactate before intubation that did not normalize at 6 h (both non-survivors). delta pco 2 exhibited erratic changes over time with no correlation with scvo 2 changes and with mortality ( fig. 1 ). introduction. venous to arterial carbon dioxide difference (pv-aco 2 ) could reflect the sufficiency of blood flow in shock states. time evolution of pv-aco 2 during early phases of resuscitation in septic shock has not been widely characterized. we proposed to describe the association between time course of pv-aco 2 during the initial resuscitation and outcomes in septic shock. methods. patients with a new septic shock episode admitted to icu were included. general management was guided according surviving sepsis campaign recommendations. time 0 (t0) was set when a central venous catheter was inserted to guide reanimation. simultaneous measurements of lactate and arterial-venous gases were obtained at t0 and 6 h after (t6). pv-aco 2 was calculated as the difference between venous co 2 (blood samples drawn from a central catheter) and arterial co 2 . a value of pv-aco 2 [ 5 was considered as high. survival at 28 day was described for four groups: persisting high pv-aco 2 (high at t0 and t6), increasing pv-aco 2 (normal at t0, high at t6), decreasing pv-aco 2 (high at t0, normal at t6) and persistently low (normal at t0 and t6). survival probabilities were estimated using kaplan-meier method. log-rank test was use to estimate a two-tailed p value for the differences in survival among groups. results. sixty septic shock patients were analyzed. mortality rate was 36.7%. no demographic differences at baseline between survivor (s) and non-survivors (ns) were found. there were no differences in the amount of fluids administered at t0 and t6. no significant differences in scvo 2 at t6 for s introduction. septic shock (ss) has been defined as sepsis related hypotension despite adequate fluid resuscitation ? perfusion abnormalities such as lactic acidosis [1] . despite this, an operationally simplified definition overlooking perfusion parameters, has been utilized in several landmark studies during the last decades [2] [3] [4] [5] . more recently, a new consensus reemphasized the pivotal role of hypoperfusion in ss definition and added low svo 2 as a surrogate [6] . several problems emerge from these apparently interchangeable definitions, including pathophysiologic and epidemiologic (incidence, outcome) issues. objectives. our aim was to evaluate if applicating different commonly used ss definitions to vasopressor-requiring septic patients leads to distinct outcomes. methods. we applied the two most utilized ss definitions to hypotensive septic patients managed with a ne-based algorithm [7] for 10 years, generating two major subgroups for analysis (fig. 1) . statistical analysis included chi-square test. (fig. 1) . 102 pts of subgroup 2, exhibited persistent normal lactate levels with a mortality of 15.2% which was similar regardless of svo 2 [ or \70: p = 0.22. (fig. 2) . conclusions. commonly used ss definitions are not interchangeable and when applied to the same vasopressor requiring septic patients lead to statistically different mortalities. our data suggest that lactate and svo 2 cannot be used indistinctly to define shock condition. a reappraisal of clinical septic shock definition appears to be necessary. objectives. to assess intra-and inter-observer agreement of ecg interpretation in adults with septic shock (vasst, nejm 2008; 358:877) . methods. patients were randomised to receive a blinded infusion of low-dose vasopressin or norepinephrine in addition to open-label vasopressors. eight icus participated in this ecg sub-study; and 12-lead ecgs were recorded at baseline (prior to study drug infusion), and 6 h, 2 and 4 days after initiation of study drug. an intensivist (reader 1) and a cardiologist (reader 2), blinded to patient data and randomization group, interpreted all of the ecgs in duplicate, using a checklist. prior to ecg interpretation, a calibration exercise was performed to refine definitions and maximize inter-observer agreement; both readers reviewed 25 ecgs (from the current study) representing the spectrum of normal to abnormal. cohen 0 s kappa statistic was used to assess intra-and inter-rater reliability. methods. the model consists of eight elastic chambers including the heart and circulations. identification of the parameters is made only from measured pressures in the aorta and pulmonary artery, and the volume in the right ventricle. septic shock was induced in (n = 6) healthy pigs with endotoxin infusion over 30 min. right ventricular pressure-volume loops were recorded by conductance catheter and end-systolic ventricular elastance was assessed by varying right ventricular preload. consent was obtained from the university of liege medical ethics committee. errors for the identified model are within 8% when the model is identified from data, re-simulated and then compared to the clinically measured data. even with a limited amount of available experimental data to identify the parameters of the model, all simulated parameters trends match physiologically expected changes during endotoxic shock. in particular, a close match of the trends of the right ventricular end-systolic elastances are obtained, when compared to previously reported experimental results [1] , including capturing of the peak after 30 min and a decaying oscillation after 30 min. conclusions. pig-specific parameters for the cvs model were accurately identified using a significantly reduced data set. this research shows the ability of the model to adequately and realistically capture the impact of pressure-volume changes during endotoxic shock. in particular, the model is able to aggregate diverse measured data into a clear, clinically and physiologically relevant diagnostic picture as the condition develops. this research thus increases confidence in the clinical applicability and validity of this overall diagnostic monitoring approach. background. conflicting data exist concerning the effects on the microcirculation of increasing mean arterial pressure (map) with norepinephrine (ne) in septic shock. nearinfrared spectroscopy (nirs) has been proposed as a tool to quantify microvascular dysfunction in patients with sepsis. by inducing a vaso-occlusive test (vot), a variety of nirsderived variables can be measured to assess local metabolic demand and microvascular dysfunction. this trial was conducted to test the effects of increasing map by ne on microvascular reactivity in patients with septic shock. after local ethical committee approval and informed consent, we enrolled 10 patients in septic shock with an arterial pressure stabilized by ne. in addition to hemodynamic measurements, svo 2 and blood lactate level, we measured thenar muscle oxygen saturation (sto 2 ) and muscle tissue hemoglobin index (thi) by a tissue spectrometer (inspectra tm model 650, hutchinson technology inc, mn). serial vot (upper limb ischemia induced by a rapid pneumatic cuff inflation around the upper arm) were performed. we also recorded during the vot: basal sto 2 , thi, the slope of the decrease in sto 2 during the occlusion (desc slope; %/min) and the slope of the increase in sto 2 following the ischemic period (asc slope; %/s). muscle oxygen consumption (nirvo2i) was calculated as the product of the inverse of the slope value by the mean of thi over the first minute of arterial occlusion and is expressed in arbitrary units (u) (skarda shock 2007). all these data were obtained at 5 different times: baseline 1 and 2 with map of 65 mmhg, then at 75 mmhg and 85 mmhg of map by increasing the ne doses and finally to baseline 3. we report here data corresponding to the mean and sd of baseline 1 and 2 versus map 85 mmhg analyzed by repeated measures analysis of variance (at 5% level) with bonferroni adjustment to account for multiple comparisons. increasing ne dose induced an increase in cardiac output (from 6.08 ± 1.20 to 6.81 ± 1.62 l/min, p \ 0.05) without any changes in heart rate and an increase of svo 2 (from 70.9 ± 3.2 to 77.7 ± 5.8%, p \ 0.05 objectives. to investigate: 1. the effects of ''successful'' protocolised resuscitation (egdt) on microvessel perfusion (particularly density). 2. whether there is different effects of egdt on the microcirculation of septic compared to critically ill non-septic patients and 3. whether there is a difference in the behaviour of ''true'' capillaries (i.e 1-10 lm) compared to larger microvessels (11-20 lm) at baseline or after resuscitation. prospective observational study in the emergency and intensive care departments of an urban teaching hospital. subjects: 20 septic and 10 critically ill control patients requiring shock resuscitation (map less than 65 mmhg, ±cvp less than 8 mmhg, ±central venous saturations less than 75%). all patients had invasive monitoring and identical cardiovascular targets. patients with known cardiogenic shock or pre-stabilised trauma were excluded. we performed sidestream dark field (sdf) videomicroscopy of sublingual microcirculation at the point of egdt initiation and again on attainment of at least 2 out of 3 cardiovascular goals. three sites were imaged for 15 s and the clips were analysed randomly off-line to provide an average value for capillary density (total length and count per mm) and a semi-quantitative description of microvessel flow (continuous, intermittent or stopped) as previously described. vessels were grouped according to diameter as small (0-10 lm) and medium (11-20 lm). non parametric analysis was used for all within or between group comparisons, data is displayed as median values with [range]. *p \ 0.05 was considered significant. 2 (11) 6 (33) 1 (5) 12 (67) duration of occlusion (min), mean ± sd 5.4 ± 2.8 3.0 ± 0.0 6.7 ± 3.9 3.0 ± 0.0 minimal sto2 (%), mean ± sd 40 ± 0 5 2± 14 40 ± 0 5 4± 13 as expected, all septic shock patients, except one (for the vot fa40% ) and two (for the vot a40% ) had a recovery slope lower than normal when sto 2 decreased to 40% during arterial occlusion. by contrast, when occlusion lasted 3 min, many patients including patients who eventually died, were misclassified since their recovery slopes were in the normal range. these results could be due to the smaller decrease of sto 2 and in turn a less strong hyperemic response when ischemia lasted only 3 min. additionally, a significantly (p \ 0.03) shorter time to reach 40% was required when arm (compared to forearm) occlusion was performed. conclusion. when a vot is required for assessing microcirculatory disturbances in septic shock, we recommend performing it using an arm occlusion until sto 2 reach 40%. aims. to analyze the correlation between sto 2 (and its changes derived from a transient ischemic challenge) and global oxygen delivery (do 2 ) parameters measured invasively using a pulmonary artery catheter (pac). observational study, performed in a 26-bed medical-surgical icu, at a university hospital. we recruited adult patients with cardiovascular insufficiency that required a pac placement for hemodynamic monitoring and resuscitation. we collected demographic data, and hemodynamic and oxymetric data derived from the pac. simultaneously, we measured sto 2 and its changes derived from a vascular occlusion test (vot). results. twenty-two patients were studied. all the patients had a mean arterial pressure (map) above 65 mmhg. the do 2 index (ido 2 ) range in the studied population was 212-674 mlo 2 /(min m 2 ). the mean svo 2 value was 57 ± 9%, mean cardiac index (ci) 2.7 ± 1 l/ (min m 2 ), and blood lactate 4.5 ± 3.1 mmol/l. the correlations found between sto 2 and invasive oxygen delivery-related variables are shown in table 1 . the sto 2 -deoxygenation slope (deox) during the vot showed a significant correlation with svo 2 (r 0.6, p 0.02). we did not find any correlation between sto 2 and global flow measurements, such as cardiac index (ci), but we found a correlation between sto 2 and ido2. this correlation seems related to the arterial oxygen content, and not to global flow. normal sto 2 values could not rule out low ido2 and low ic states. therefore, sto 2 seems to be poorly sensitive to exclude hypoperfusion states. in clinical practice there remains issues over the appropriate prescribing of antibiotics in patients with unproven sepsis. the prescribing of antibiotics is not without risk and creates a selective pressure on existing bacterial flora resulting in the emergence of virulent and resistant organisms [1] . there is also a cost issue from the inappropriate prescription of antimicrobials [2] . the diagnosis of sirs can be made with confidence [3] , sepsis cannot and requires confirmation from microbial tests. empirical usage of antibiotic therapy is commonplace but not ideal. rapidly detectable, reliable markers of sepsis would help in directing antimicrobial therapy. objectives. the aims of this study are to determine the significance of % band forms in sirs patients suspected to have sepsis. can they be used as a diagnostic tool in conjunction with procalcitonin in order to direct antimicrobial therapy? methods. this is an observational study aiming to assess the ability of serum procalcitonin and percentage band forms in identifying nosocomial sepsis in patients with sirs. 32 patients were recruited over an 18 month period in a mixed medical-surgical university teaching icu. all patients had suspected sepsis arising 'de novo' and had not received prior antimicrobial therapy. patients had a septic screen performed along with baseline, 24 and 72 hpct and % band form count. introduction. pneumonia is the most frequent infectious complication after successfully resuscitated cardiac arrest (ca). however, diagnosis is difficult because of many clinical, biological and radiological confounding factors as well as the widespread use of therapeutic hypothermia. this could lead to a broad antibiotic prescription. to assess the utility of plasma procalcitonin (pct) measurements for diagnosis of early-onset pneumonia in successfully resuscitated ca. monocentric study (july 2006 -march 2008 with retrospective review of a prospectively acquired icu database focusing on all consecutive patients admitted for ca and surviving more than 24 h. patients with an infection prior to ca or with an extra-pulmonary infection developing within 5 days following admission were not studied. all files were reviewed to assess the diagnosis of early-onset pneumonia p(?), or not p(-) during the first 5 days of icu stay. p(?) was defined by the presence of a new pulmonary infiltrate on chest radiography, persistent for at least 48 h, associated with either positive quantitative culture of the endotracheal aspirates, either, in case of lack of bacteriological sample, conjunction of purulent sputum and hypoxemia (p/f \ 200). pct was measured at admission, days (d) 1, 2 and 3 (brahms kryptor ò ). among 245 patients admitted for ca, 132 were studied (48 death before 24 h, 11 evolutive infections and 54 incomplete samples). pneumonia was diagnosed in 86 patients (65%), and antibiotics were prescribed in 115 during the first 5 days of icu stay. characteristics of p(?) and p(-) patients were (median, iqr): age 60 (47-70) versus 60 (50-70) (p = 0.9), ''no flow'' 3 (0-10) versus 6 (1-10) min (p = 0.06), ''low flow '' 15 (5-20) versus 15 (6-23) min (p = 0.95), shockable rhythm 47 versus 37% (p = 0.16), cardiac etiology 68 versus 48% (p = 0.02), therapeutic hypothermia 99 versus 100% (p = 0.46), post-resuscitation shock 51 versus 48% (p = 0.72) and icu mortality 55 versus 54% (p = 0.9). using a threshold value of 0.5 ng/ml, negative predictive values were 39% at admission, 42% at d1, 52% at d2, whereas positive predictive values were 72, 68 and 70%, respectively. patients with post-resuscitation shock had higher pct levels than those that did not require vasopressors: 6.66 versus 0.95 ng/ml at d1 (p \ 0.001), 6.58 versus 0.82 at d2 (p \ 0.001) and 5.2 versus 0.95 at d3 (p = 0.03). conclusion. diagnosic value of pct is poor in survivors of ca and pct should not be recommended to assess early-onset pneumonia. post-resuscitation disease could play a major role in the lack of specificity and predictive values. in acute community respiratory infection, low levels of procalcitonin (pct) have been shown to allow a marked reduction of antibiotic use. the aim of the study was to look for the same efficacy in case of suspicion of infection during icu stay. method. from april 2008 to december 2008, 529 patients hospitalized in the five intensive care units (icu) of the university hospital of liège in belgium, were prospectively randomized to either a procalcitonin guided approach to antibiotic therapy (pct group, n = 268) or to a standard approach (ctrl group, n = 261) when they were suspected of developing an infection. for pct group guided therapy only, the use of antibiotics was more or less strongly discouraged (pct level .25 or.50 lg/ml, respectively) and more or less recommended (pct level [1 or[0.5 lg/ml, respectively) . number and duration of antibiotic treatments were recorded. diagnosis and treatment decisions were reviewed by infectious disease (id) specialists at the end of icu stay. results. there were no differences between groups in terms of age (66 vs. 65), saps ii score (40.3 ± 16.3 vs. 39.2 ± 16.7), type of patients (medical: 61 vs. 58%, scheduled surgery: 8 vs. 10%, emergency surgery: 21 vs. 21%, trauma: 10 vs. 11%), icu length of stay [8 (iqr 4-18) vs. 7 days (iqr 4-16)] for pct and ctrl group respectively. suspicion of infection was either evoked on admission (in 41 and 44%) or during icu stay (in 61 and 59%) in pct group and ctrl group respectively. at the time of suspicion, pct levels was.25 lg/ l in 26.8% of the infectious episodes in pct group and 24.1% in ctrl group. 79 episodes of suspected infection with pct level .25 lg/ml were recorded. clinicians decided not to treat 38% of these episodes (n = 30). the remaining 49 episodes were treated, of which 65 % were eventually considered as probable or confirmed infections by id specialist (n = 32). at the end of icu stay, id specialists classified 635 infectious episodes of both groups as confirmed (n = 262; 46.3%), probable (n = 140; 22%), possible (n = 70; 11%) or absent (n = 163; 25.7%). for confirmed episodes of infection, pct levels were .25 lg/ml in as much as 28.1% and above 1 lg/ml in 63.6%; for absence of infection, pct levels were .25 lg/ml in only 56.8% and above 1 lg/ml in 28.8%. the ability of pct to discriminate between confirmed and probable infections on the one hand and possible or absent infection on the other hand, was tested by the measurement of the surface under the roc curve, which was 0.67, which is too low to recognize pct as a valuable marker of infection. there were no difference in the number of treated patients (88 vs. 87%) nor in the number of antibiotic days (64 vs. 60%) between pct and ctrl group respectively. conclusions. procalcitonin level as an aid for the decision to treat infection in icu patients appeared not to be helpful. antibiotic consumption was not reduced using this tool in our study. introduction. respiratory infections, pneumonias in particular are a common cause of mortality in the intensive care unit (icu) patients worldwide. early identification and prompt management of these patients especially with associated sepsis is crucial in reducing the mortality. many clinical and laboratory markers have been studied extensively to predict the outcomes in them. there have been numerous studies on the clinical utility of serum procalcitonin (pct) in the past decade, in systemic inflammation, infection and sepsis. objective. to evaluate the role of serum procalcitonin, in predicting the outcomes of patients admitted in the icu with respiratory infections associated with sepsis. setting: 10 bedded icu of a tertiary referral hospital. study design: prospective observational study. subjects: 85 adult ([18 years) patients admitted in the icu with lower respiratory tract infections with associated sepsis during the period july 2007 to january 2009 were prospectively followed up. primary outcome measure: 28 day mortality. we measured pct levels using the brahms immunochromatographic technique(semiquantitative estimation) on the first day of admission into the icu . normal pct was taken as .5 ng/ml. patients were grouped into four groups-group a (pct \ 0.5 ng/ml), group b (pct [ 0.5 -2 ng/ml), group c (pct [ 2 -10 ng/ml),group d (pct [ 10 ng/ml). sepsis, severe sepsis, septic shock are defined according to the 1992 accp/sccm criteria. results. the overall mortality was 25.8% with mortality of 3.1, 7.6, 50, and 50% in groups a, b, c and d, respectively. there is a statistically significant difference (p \ 0.05) in the mortality rates of groups c and d as compared with group a and b, but no difference was observed in the mortality rates between groups a and b and groups c and d .also significant statistically are the apache ii scores, septic shock and multiorgan failure incidence in the groups c and d as compared to groups a and b. conclusions. serum procalcitonin level [2 ng/ml on the first day of admission in icu appears to be a good predictor of mortality in patients admitted with lower respiratory tract infections and associated sepsis. methods. in a retrospective study we assessed 458 acutely ill patients investigated for pct and treated by a physician blinded for pct value. for each patient we also calculated new simplified acute physiology score (saps ii). we evaluated many clinical and instrumental parameters and diagnosis was done upon our usually clinical practice results. the mean age of patients (pt) was 73.9 yeats, shock was found in 129 patients (28.4%),median value of saps ii score was 33 (iqr 40-26), and median estimated mortality from saps ii was 14% (iqr 24-27). bacterial infection was found in 77.3% (septic shock 16.7%, pneumonia 51.1%, cholecystitis 3.7%, pleural empyema 1.4%, other infections 27.1%) non infective disease in 22.7% (pulmonary embolism 7.7%, acute coronary syndrome 9.6% heart failure 18.3% other disease 64.4%. a pct value [ 0.25 ng/ml was considered positive: so pct was elevated in 59.6% of bacterial infection patients and in 30.8% of non infective disease patients. we also compared pct values with antibiotic therapy and considered appropriate the administration if pct [ 0.25 ng/ml: there was discrepancy in 39.1%. the review of these cases found medical decision wrong in 72 cases versus 248 (38.2%); 70 pt with pct \ 0.25 ng/ml had antibiotic therapy without bi and 2 cases with pct [ 0.25 ng/ml did not have antibiotic therapy but had a bacterial infection. subsequent to this review discrepancy felt to 23.1% (ci 95% 19. 3-27.3) and was found especially in pt with pct \ 0.25 ng/ml. at cut off point of 0.25 the sensitivity was 0.87 (ci 95%:0.82-0.90) specificity 0.33 (ci 95%:0.27-0.40) or 3.1 and at point 5.0 the sensitivity was 0.90 (ci 95%:0.92-0.95) specificity 0.26 (ci 95%:0.22-0.30) or 3.1, with high predictive positive value. all-causes mortality was 19.4%. mortality if pct \ 0.25 ng/ml was 9.8%, if pct 0.25-5.0 ng/ml was 24.5%; if pct 5.0-10.0 lg/ml was 38.2% and if pct [ 10 ng/ml was 31.0% without significant difference between bacterial infection and non infective disease group. comparing pct with saps ii score, area under roc-curve was not significantly different (pct 0.68-ci 95%: 0.60-0.74) (saps ii 0.75-ci 95%: 0.68-0.80). conclusions. pct in acutely ill patients is a useful marker to discriminate bacterial infections with high sensibility but low specificity and it may be useful to guide the therapy also with values higher than 0.25 ng/ml. our data suggest a real prognostic utility of pct in these patients, regardless of bacterial infections, but our efforts to elaborate a mathematical predictive model aren't still satisfying and further data are required in this setting. h. taniuchi 1 , t. ikeda 1 , k. ikeda 1 , s. suda 1 1 tokyo medical university, hachioji medical center, division of critical care medicine, tokyo, japan introduction. its apparent that detection of the causative bacteria is useful for the therapeutic strategy. however, conventional tests for the detection of the causative bacteria are not high sensibility. in order to diagnose sepsis or septic shock and start appropriate therapy rapidly, it's also important to know whether the infection is cause of gram negative bacteria, that is to say, whether the infection is cause of endotoxin. in this study, we investigate the severity level of sepsis and initiation criteria of direct hemoperfusion with polymixin b immobilized fiber column (pmx-dhp) treatment from the result of severity level by using endotoxin activity assay (eaa) and using measurement of procalcitonin (pct). subjects and methods. 26 patients who developed severe sepsis or septic shock and admitted to icu were included. on the day of icu admission, a general blood biochemistry, eaa and pct levels, and apache ii and sofa score were measured. patients were evaluated retrospectively the relationship between the severity of sepsis and each measurements and investigated the relationship between the measurements and pmx-dhp. serum eaa level was measured using smart line eaa luminometers. serum pct level was measured using immune luminometric assay. results. the average age of the patients is 72 ± 12, apacheii score was 23.5 ± 6.5, sofa score was 9.0 ± 3.1, the median pct was 26.0 ng/ml (range 0-200), eaa was 0.52 ± 0.26. the underlying diseases of the enrolled patients were the abdominal infection (12 patients), the urinary tract infection (3), pneumonia (3), the meningitis (2), the soft tissue infection (2) and other infection (5) . the causative bacteria were gram positive bacteria (7), gram negative bacteria (9), virus (1), and unknown (9). there was no statistical correlations between eaa or pct level and apacheiiscore. there was no statistical correlations between eaa level and sofa score. although there was no statistical correlation between pct level and sofa score, the pct level tended to rise as pct level rises. we investigated the relationship between eaa and pct levels. there was also no statistical correlations between eaa and pct. we investigated the relationship between the causative bacteria (gram positive bacteria, gram negative bacteria and the others) and eaa or pct level. there was no statistical correlations between the causative bacteria and eaa level nor pct, that was contrary to our expectation that eaa level should be high for gram negative bacterial infection. we further investigated the relationship between whether or not the pmx-dhp was implemented and eaa or pct level. there was no statistical relationships. conclusion. high levels of the eaa and pct would not indicate the severe infection with gram negative bacteria, and the initiation of pmx-dhp. further study is needed, in which more patients will be enrolled and evaluated. introduction. sepsis still the major cause of death in the late post traumatic period in patients with major burns. early diagnosis of sepsis is crucial for management and outcome of critically burn patients. attempted in this study to assess whether plasma procalcitonin (pct) level was related to diagnostic and prognostic of sepsis in burned patients. patients and methods. pct was measured over the entire course of stay in patients with predictive signs of sepsis according to american college of chest physician. the patients were assigned to two groups depending on the clinical course and outcome: a = no septic patients, b = septic patients. optimum sensitivity, predictive values, and area under the receiver operating characteristic (roc) curve were evaluated. results. over a 6 month period starting from 1 july 2008 to 31 december 2008, 157 patients were admitted. 62 were investigated. 40 in group a et 22 in group b. procalcitonin was significantly higher in septic group 7.26 ± 7 ng/ml compared to no septic group 0.25 ± 0.32 ng/ml. area under the curve was 0.94 on the day of sepsis diagnostic. pct cut-off value of 0.75 ng/ ml was associated with the optimal combination of sensitivity (85%), specificity (87%), positive predictive value (91%), and negative predictive value (73%). in survived septic patient the pct value was significantly lower than in deceased septic patients 3.5 ± 0.87 versus 10.18 ± 9.6 ng/ml. pct cut-off value for optimum prediction of outcome in septic patients was 3.66 ng/ml with sensitivity (91%), specificity (75%), positive predictive value (78%), and negative predictive value (90%). conclusion. procalcitonin appears to be a powerful marker of sepsis in burn patients. it is sensitive, specific, reliable and easy to measure. a high pct concentration ([3.66 ng/ml) would indicate poor outcome in septic patients. n. v. beloborodova 1 , a. s. khodakova 1 , a. y. olenin 1 , s. t. ovseenko 1 1 bakulev scientific center for cardiovascular surgery, moscow, russian federation objectives. accurate and timely diagnosis of sepsis remains challenging for clinicians. the diagnosis of sepsis is defined as typical symptoms of systemic inflammation (temperature, tachycardia, respiratory rate, leukocytosis) with clinical evidence of an infection site, but the criteria are met by a large number of intensive care unit (icu) patients. among studied biomarkers, serum procalcitonin (pct) has been described as one of the most promising predictors of bacterial sepsis, but in some clinical situations it is not enough. the search of reliable markers of sepsis is still in progress. in present study the significance of raised levels microbial phenylcarboxylic acids in serum of patients with sepsis are assessed. methods. the present study evaluated 22 serum samples of patients (pts) with documentary sepsis, according to well known consensus criteria. the comparison groups were: no. 1-16 clinically healthy volunteers, no. 2-36 pts. with acquired heart diseases, no. 3-19 pts with ventilator-associated pneumonia. blood concentrations of phenylcarboxylic acids were determined by gas chromatography-mass spectrometry (gc-ms). results are presented as median and range of 25th and 75th percentiles. the statistically significant differences between the various groups were calculated using mann-whitney test. results. increased levels of phenyllactic (pla), p-hydroxyphenylacetic (hpaa), p-hydroxyphenyllactic (hpla) acids were observed in group of pts with sepsis. the level of hpaa was increased up to two orders in comparison with groups no. 1 and 2 [14.1 (7.8-35.9) vs. 0.5 (0.4-0.6) and 0.7 (0.3-1.4) lm, p \ 0.0001). the levels of hpla and pla were increased up to one order [(7.6 [2.9-15.5] table 1 for illustration of importance of phenylcarboxylic acids blood level monitoring. introduction. acute kidney injury (aki) is a frequent complication of sepsis, and is associated with high mortality and morbidity rates. routinely used measures of renal function, such as levels of blood urea nitrogen (bun) and serum creatinine, increase only after substantial kidney injury occurs, resulting in delayed diagnosis of aki. therefore biomarkers, which enable early diagnosis, are needed. objectives. this clinical study was designed to investigate whether human interleukin-18 (il-18) and neutrophil gelatinase-associated lipocalin (ngal) are early predictive markers for sepsis-induced aki. urine and blood samples have been collected prospectively from icu patients, who met defined clinical criteria of severe sepsis. aki was defined by rifle criteria. urinary and serum levels of n-gal and il-18 have been quantified by elisa in patients with sepsis without aki (n = 11) and in patients with sepsis induced aki (n = 13). results. both, urinary il-18 and serum il-18 considerably increased (respectively, 7.22 and 19.54-fold over the baseline) two days before the patients reached rifle risk. urinary ngal raised significantly (1.72-fold over the baseline) one day before occurrence of aki, whereas serum ngal did not show any prior elevation. no increase in the levels of any of these markers could be found in patients who did not develop aki. conclusions. both urinary and serum il-18 seem to be sensitive early biomarkers for sepsis associated aki, while urinary ngal has less accuracy for aki prediction. objectives. to define a biomarker panel able to predict infection in case of severe acute dyspnea in emergency situations. we designed a prospective observational study of patients admitted in the emergency department (ed) and in medical polyvalent intensive care unit (icu) in a university hospital. inclusion criteria were acute dyspnea with spo 2 b 92% and/or respiratory rate (rr) c 25 b/min. patients with an immediate need of coronarography or with obvious spontaneous pneumothorax were excluded. five biomarkers were measured from blood sample at admission on ed or icu: nt b type natriuretic peptide (nt probnp), cardiac troponin i (ctni), ddimeres (dd), c-reactive protein (crp) and procalcitonin (pct). all clinical and biological data were recorded. an independent blinded data monitoring committee classified the patients according to all the available data including response to treatment and outcomes but blindly to biomarkers. the roles of biomarkers were assessed quantitatively and then using terciles of the distribution. the contribution of the biomarkers in the diagnosis was assessed using multiple logistic regression taking into account other clinical and biological explanatory variables. . 172 patients were enrolled consecutively. the final diagnosis was: severe sepsis (n = 50), acute heart failure (n = 42), pulmonary embolism (n = 11), copd (n = 21), other causes (n = 48). the 28 days mortality was 17%. there was no significant association between infection diagnosis and dd, ctni, nt probnp. interestingly, a crp value of less than 5 mg/l was not discriminant in predicting infection. adjusted on clinico-biological covariates selected, both pct with cutpoints of 0.1 and 0.4 ng/ ml (discrimination auc 0.850; p = 0.008) and crp with cutpoints of 30 and 100 mg/l (discrimination auc 0.857; p 0.001) were significantly associated with the diagnosis of sepsis. both biomarkers used simultaneously lead to a discrimination of the model (auc 0.879). conclusion. both crp and pct are able to predict the diagnosis of infection in case of severe acute dyspnea independently of clinico-biological variables. in this particular subpopulation, the best threshold for crp is higher than the standard one. an external validation is needed to prospectively validate the clinical utility of these findings. t. trefzer 1 , i. nachtigall 1 , a. weimann 2 , c. de grahl 1 , c. spies 1 1 charite universitaetsmedizin berlin, campus virchow, department of anesthesiology and operative intensive care medicine, berlin, germany, 2 charite universitaetsmedizin berlin, campus virchow, zentralinstitut für laboratoriumsmedizin und pathobiochemie, berlin, germany aims. infections are the most relevant icu-admission complication. crp and pct are labvalues used for diagnosis of infections. however, their use is often not evidence based. this study aimed to access whether the adherence rate increased after introducing an evidencebased standard operating procedure (sop). in 2008 an evidence-based sop was approved by experts of our department. in july 2008 it was made available to icu-physicians via intranet, which is accessible from every work station. altogether, we assessed sop-adherence rates of 602 patients: 188 in june 2008 (pre-sop), 204 patients in august 2008 (one month post-sop) and 194 in january 2009 (6 months post sop). every crp and pct measurement was assessed for adherence to the standard operating procedure (sop). at first, the three periods were assessed for significant differences concerning the adherence. according to the percentage of sop-conform measurements the patients were then divided into two groups: the sop-group (c70% of measurements sop conform) and the non-sop (nsop) group (\70% conform) in a second step, patients in the sop-and nsop-group were compared concerning icu scores (sofa, tiss, apacheii, saps) and outcome parameters (length of icu-stay, length of hospital stay, duration of mechanical ventilation, hospital mortality). statistics: p b 0.05 was considered as statistically significant; hospital mortality was assessed by a v 2 test, icu scores and outcome parameters were compared using the mann-whitney u test. all parameters with p \ 0.1 were included into a logistic regression analysis. no change was observed concerning the implementation of the sop pre and postintroduction: 33.5% in june 2008, 36.3% in august 2008 and 33.0% in january 2009. the non-conform pct-and crp-measurements resulted in additional costs of approximately 40.000 euros/year. the univariate analysis revealed significant differences in the sop-and nsop-group: the nsop-group had higher saps-, sofa-and tiss-scores, as well as increased length of icu-stay, length of hospital stay and duration of mechanical ventilation. logistic regression analysis revealed tiss score and length of hospital stay as an independent predictor for low sop adherence. conclusion. distribution of an evidence based sop without further education did not lead to a significant increase in adherence rates, but tiss score and length of hospital stay have shown to be independent predictors for low adherence to the sop. the significant higher tiss-scores in the nsop group might be a indicator for actionism of clinicians in the face of more severely ill patients. objetives. to asses the evolution of the risk-adjusted mortality rates of sepsis and septic shock in our icu in a ten years period. patients and method. analisys of prospectively recorded data of all pacients admitted with severe sepsis and septic shock in a 10 bed icu during a period of 10 years. patients were followed up until death or discharge from the hospital, excluding those with unknown outcome. mortality prediction was made using apache ii model with 95% confidence intervals. statistical analisys was made with spss 16.0 using anova test or t test to compare means and chi square test to compare categorical variables. results. from january 99 to december 2008 a total of 733 patients with sepsis were admitted, with an anual increase to reach 20% of all icu admissions. age and severity of illness increased anually as did sofa in the first 24 h (sofa1) thus rising up calculated risk of death. from 1999 to 2004 mortality rate was between 95% ics of calculated risk of death, falling below inferior ic from and after 2005. (fig. 1 ). hospital mortality versus risk of death per year mortality was 51.6% in the pre-2005 period and 44.4% from 2005 and on (p = 0.05) with non significant differences in apache ii, risk of death nor sofa1, but with significantly greater age in the post-2005 period (62.6 vs. 58.3 years p = 0.01). this non significant difference between the two periods of the study became significant when we analized the outcome in both sex. being significant in women (mortality 41.1% in pre-2005 period vs. 31.4% in post-2005 p = 0.02) but not in men (57.3 vs. 52.5% p = 0.2). overall sepsis moratlity is lower in female without significant differences in age, apache ii score nor risk of death (table 1) , being the only signifficant difference found in sofa1 (8.63 in male vs. 7.92 in female p = 0.03). introduction. low-grade systemic inflammation has been shown to play a key role in the pathophysiology of several chronic noncommunicable diseases [1, 2] and may be attenuated by anti-inflammatory treatments such as administration of statins [3] . so far, the association between acute systemic inflammation experienced during critical illness and long-term mortality after hospital discharge has not been investigated in intensive care unit (icu) patients. objectives. to assess the association between acute systemic inflammation, assessed by crp levels, and post-hospital mortality in non-surgical icu patients. methods. the study was performed as a prospective, observational follow-up study and included non-surgical critically ill patients with an icu length of stay [24 h. patients who died during the icu or hospital stay, were \18 years or pregnant, as well as patients discharged from the hospital with the plan to limit life support were excluded. demographics, chronic diseases, admission diagnosis, the simplified acute physiology score ii, length of icu stay, maximum crp levels during the icu stay (crpmax) and crp levels at icu discharge (crpdis) were documented. after a mean ± sd follow-up time of 2.04 ± 1.08 years, mortality and causes of death were determined. adjusted cox models were calculated to investigate the association of crpmax and crpdis with post-hospital mortality. a receiver operating characteristic analysis was used to identify optimal cut-off levels to predict post-hospital mortality. background. the prevalence of hiv infection is increasing worldwide as a public health problem. survival of hiv/aids patients has improved since highly active antiretroviral therapy, but sepsis has grown as an important cause of icu admission in this population. an international conference has set a system composed of specific risk factors, site and microbiology of severe infections and host response and organ dysfunctions (piro) to help identify patients at risk for sepsis. piro factors have not been classified for hiv/aids population yet. objectives. to identify predisposing factors, microbiology of infections, host clinical response and incidence of early organ dysfunctions of severe sepsis on hiv/aids patients, admitted to a specialized infectious diseases icu; to analyze long-term survival of hiv/aids critically ill patients. a prospective case-control study of septic and non-septic hiv/aids patients admitted between june 2006 and may 2008 was performed. demographic data, causes of admission, time since aids defining condition, cd4 cell count, and opportunistic infections were evaluated as predisposing factors to sepsis. microbiology and site of infections were registered. clinical response to severe infections was evaluated by ali/ards and shock incidence on day 1 of icu admission. organ dysfunctions (sofa score) were reported soon after icu admission. icu length of stay, hospital and 6-month mortality were compared between septic and non-septic groups. a multivariate regression analysis was done to identify risk factors for icu mortality. kaplan-meyer survival curve was built. . 108 icu admissions of 101 hiv-infected patients were studied. half (54) fulfilled criteria for severe sepsis diagnosis. septic group was younger (39.0 ± 11.8 vs. 43.4 ± 11.7 years, p \ 0.05) and had more female patients (35 vs. 17%, p \ 0.05). time since aids diagnosis, cd4 cell count and opportunistic infections prevalence were not different. sites of infection were predominantly pulmonary (48%) and catheter-related (19%). ninety percent of infections were nosocomial. forty-three percent of septic patients presented bacteremia. pseudomonas sp, s aureus and enterobacteriacae were commonly identified, but five patients had mycobacterium tuberculosis isolated (2 on blood cultures). multiple organ dysfunction syndrome was frequent, and incidence of cardiovascular, respiratory and hematological dysfunctions was significantly higher in septic group. longer length of icu stay (19.6 ± 20.5 vs. 9.4 ± 8.7 days, p \ 0.01) and icu mortality (61 vs. 18%, p \ 0.01) was observed for septic patients. severe sepsis also influenced long-term survival, as mortality continues significantly higher after 6 months (log rank 17.3, p \ 0.001). conclusions. piro system is applied to septic hiv/aids patients. shock, ali/ards and hematological dysfunctions are prominent for septic hiv/aids population. septic hiv/ aids patients are at severe risk of short and long-term mortality. international guidelines for management of severe sepsis and septic shock suggest the use of recombinant human activated protein c (rhapc) in adult patients with high risk of death (apache ii c 25 or multiple organ failure). the objective of this study is to analyse the characteristics and outcome of patients treated with rhapc in our medical intensive care unit. retrospective study of patients with severe sepsis/septic shock treated with rhapc between january 2005 to december 2008. all of them were c18 years, with apache ii c 25 and two or more organ dysfunction, and were treated on basis of a bundle for severe sepsis management: complete early goal-directed therapy, early administration of broadspectrum antibiotics; corticosteroids in vasopressors unresponsive patients and monitor for lactate clearance. chi-square analysis were used to compare categorical data. continuous data were compared using student's t test. prognostic factors of mortality were studied by means of multivariable logistic regression analysis. results. forty-one patients were studied. 80% were male. their mean age was 55 ± 18 years. 44% had comorbidities (26% immune pathology). severity scores. apache ii 29 ± 7, sofa 11 ± 3, 39% of patients had three o more organ dysfunction. 93% had septic shock. serum lactate level was 5.1 ± 2.6 mmol/l. the primary location of infections was: respiratory 71%, abdominal 15%, urinary 7%. 31.7% were positive blood culture. 83% of patients needed mechanical ventilation (18 ± 15 days). 26% of rhapc infusions were not completed, mainly for bleeding risk (36%) and death (27%). 2.4% of patients had bleeding event. at the end of the infusion 58% of patients remained with two or more organ dysfunction and 56% were vasopressors dependent. mean hospital stay was 31 days and 19 days in icu . 28 days mortality was 39%, icu mortality 41.5% and hospital mortality 48.8%. analyzed data included age, comorbidities, primary location of infections, severity scores and serum lactate level. univariable analysis showed that statistically significant factors related to mortality were: apache ii (31 ± 7 vs. 27 ± 6, p = 0.04), organ dysfunction number: 2 vs. [2 (25 vs. 58%, p = 0.02) and primary location of infections: pneumonia versus others (59 vs. 25%, p \ 0.05). a multivariable logistic regression analysis showed that age (or 1.10, 95% ci 1.01-1.12, p = 0.03), organ dysfunction number (or 14.75, 95% ci 1.81-117.38, p = 0.01) and serum lactate levels (or 1.50, 95% ci 1.01-2.21, p = 0.04) had statistically significant relationship to mortality. conclusion. in our study the patients with severe sepsis and septic shock remained with high vasopressors dependency and organ dysfunction at the end of the rhapc infusion. despite of rhapc therapy the mortality of patients was very high. the age and the severity at icu admission were independent prognostic factors of mortality. a higher incidence of severe sepsis in blacks compared to whites is well documented, however prior analyses do not discriminate whether this is due to a higher incidence of infections, a higher risk of developing organ dysfunction once infected, or both. objectives. we sought to understand whether higher severe sepsis incidence in blacks is due to higher infection susceptibility, higher risk of organ dysfunction once infected, or a combination of both. we analyzed 11,430,630 hospitalizations from 2005 hospital discharge records of 7 us states (25% of us population). we linked these records to us census data to generate age and sex-standardized incidence rates. we identified infections of bacterial and fungal etiology based on icd-9 cm criteria, including characterization by site and type of infection (gram negative vs. gram positive). we defined severe sepsis as documented infection plus acute organ dysfunction based on previous work by angus et al we estimated the risk of organ dysfunction among those hospitalized with infections using logistic regression, adjusting for age, sex and comorbidities (charlson score). fig. 1b ]. the combination of both events led to a 66% higher severe sepsis hospitalization rate for blacks (9.3 vs. 5.6 per 1,000 population, irr: 1.40-2.05). these differences persisted when stratified by sex, comorbidities, site and type of infection. infection incidence and severe sepsis risk conclusion. the higher incidence of severe sepsis among blacks is due to a higher hospitalization rate for infections, as well as a greater likelihood of organ dysfunction once infected. future interventions to reduce racial disparities in severe sepsis incidence should target both distinct events. grant acknowledgement. dr. mayr was supported by t32 hl007820-10. objective. to describe recent epidemiological data and mortality risk factors of patients admitted to icu for severe pneumococcal pneumonia (pp). multicentric retrospective study (january 2001-june 2008). prospective acquired data from patients admitted in 37 french medical icu for severe pp were considered. patients with concurrent meningitis, severe copd with known sp colonization, hiv or aspiration pneumonia were not included. pp was defined by the combination of a suggestive clinical context, the presence of a new pulmonary infiltrate on chest radiography and a s.pneumoniae positive bacteriological sample (pulmonary quantitative culture, pleural fluid, blood culture or urinary antigen assay). all files were reviewed and approved by two independent investigators (nm, am). . 224 patients were included. median age was 60 ± 16. hospital survivors were significantly younger (58 ± 16 vs. 66 ± 15, p = 0.001). sex ratio m/f was 151/73, but male sex was associated with higher risk of death (male: 59 vs. 83%, p = 0.001). active tabagism (39%) or alcohol abuse (29%) were more common than asplenia (1%). organ dysfunctions were mainly respiratory (82%), haemodynamic (70%) and renal failures (30%). low doses steroids were prescribed in 44% of patients with septic shock. icu mortality rate reached 25% (22% in the first 5 days); hospital mortality rate was 30%. univariate analysis demonstrated that age, male sex, cirrhosis and organ failure support were strong predictors for icu mortality. multivariate analysis only highlighted age [or 1.5 (1.01-1.08)], cirrhosis [5.27 (1.28-21.66) ] and renal replacement therapy [3.56 (1.56-8. 14)] as independent mortality predictors. activated protein c treatment was associated with decreased mortality [or 0.27 (0.08-0.77)]. bacteremia had no impact on outcome. conclusion. this is the most important cohort of pp requiring icu admission. despite adequate antibiotherapy, mortality is still preoccupant. determination of factors related to the bacteria (virulence) or to the host (genetic susceptibility) could allow a better understanding of this important health problem. introduction. to identify the risk factors of mortality for patients with severe community-acquired bacteremic pneumococcal pneumonia. retrospective study realised in the intensive care units of two hospital medical centers. the studied population was 70 patients with serious community-acquired bacteremic pneumococcal pneumonia. all the patients entered the intensive care units between january of 1997 and december of 2008. study variables were: age, sex, concomitant pathology, toxic habits, pre-vaccinal (1997-2001) and postvaccinal periods (2002) (2003) (2004) (2005) (2006) (2007) (2008) , serotype and sensitivity of streptococcus pneumoniae to penicillin, the initial use of the non-invasive mechanical ventilation, the development of empyema pleural, apache ii and sofa scores during the first 24 h after admission. results. the age average was of 55 years. forty one percent of our patients required mechanical ventilation, and 31% had acute renal failure that required hemofiltration. average values of apache ii and sofa were 19.8 and 7.6 respectively. in hospital mortality of the series was of 25%. in patients with severe community-acquired bacteremic pneumococcal pneumonia: (1) the presence of empyema pleural is an independent risk factor for mortality. introduction. procalcitonin (pct) is an interesting marker of pulmonary infection [1] . it is useful as an help for infection diagnosis but also for treatment follow-up [2] . besides, initiation of effective antimicrobial therapy is the strongest predictor of outcome in patients with septic shock [3] . the aim of the study was to analyse whether kinetics of pct decline may reflect sensitivity of identified infectious agents to initial antimicrobial therapy (at). patients with diagnosis of severe pneumonia following major cardio-thoracic or vascular surgery were retrospectively included in the study. severe pneumonia was suspected as a combination of several manifestations including fever or hypothermia, hyperleucocytosis or leucopenia, new radiological infiltrate, and/or a clinical pulmonary infection score [6, pct [ 1 ng/ml and pao 2 /fio 2 \ 200. initial antimicrobial treatment was chosen according to the guidelines in use in our institution for community-acquired or nosocomial infections. microorganism identification from endotracheal aspiration or bronchoalveolar lavage, and antibiotic susceptibility testing, allowed to classify patients according to appropriate (aat) versus inappropriate initial at(iat). pct was measured daily over 14 days and its kinetics compared between both groups. data are expressed as median (extremes) or mean ± sd (decrease rate). results. 28 patients aged 69 ± 9 (66-73), operated on vascular (n = 6), thoracic (n = 7), or cardiac surgery (n = 15) have been studied from october 2007 to july 2008. pneumonia occurred within the 1st to the 31st postoperative day (median 6.7 days), with a septic shock in 8 cases and 4 deaths at day 14. initial at was appropriate in 75% (21/28) patients. pct peak was not statistically different between aat versus iat patients (17.7 ± 42.2 ng/ml vs. 12.7 ± 26.6, respectively) but pct decrease was significantly steeper and constant in iat patients (fig. 1 ). pct decrease (%) from peak value over days discussion: the results suggest that absence of early decrease in pct within 2 days may reflect failure of the at. conversely, an average decrease in pct plasma concentration of 50% in 2 days seems to be a good marker of sensitivity of the causative infectious agent to the initial at. in case of unchanged pct within 2 days at change should be considered. icu mortality was 43% for pts with rb early vap while it was 22% (16 of 72 cases) in those with sb or negative cultures (p = 0.04). mortality was higher than the predicted according to apache ii score in pts with rb vap (43 vs. 30 ± 22%, p \ 0.01). however, it was lower than predicted in those with negative or sensible isolates (22 vs. 27 ± 24%, p = 0.08). conclusions. rb were the most common cause of early vap among our patients. the burden of illness, los in icu before intubation and previous use of antibiotics were associated with early vap due to rb. inappropriate empiric therapy and mortality were higher among patients with early vap due to rb. to evaluate the performance of the saps 3 piro model in patients with severe community acquired pneumonia (cap), over a period of 5 years (2003-2008) , in a general icu in a central hospital. material and methods. we analysed data prospectively registered in an informatic data base, which contains information referring to all patients admitted in this unit. analysed were 74 patients. discrimination was accessed by the area under the roc curve (aroc). calibration was evaluated by the by the hosmer-lemeshow ĉ test. conclusion. implementation of a sedation protocol requires constant follow up and regular adaptation to prove efficient over time. constant feed back information to both the medical and nursing staff is mandatory. treatment of hyperactive delirium. hal haloperidol, bzd benzodiazepine, pro propofol, aa atypical antipsychotic, nd no drugs, na not answered discussion. there tends to be a general pessimism regarding obese patients within the intensive care community. our data indicates that this opinion could be misplaced. reduced ventilator days may reflect a reluctance to invasively ventilate obese patients. the apache ii scoring does not take into account the bmi which would eliminate any severity scoring bias. high bmi alone should not be a consideration in the decision regarding suitability for admission to critical care. during the last decades, a growing medical knowledges have changed the clinical approach to elderly patient diseases. they receive major surgery or intensive treatment for acute medical illness but often the recover is condictioned by the previous chonical diseases. this determines a long period to stay in intensive care unit (icu) because the slow improvement and cause an occupation of bed places. in our hospital, after a period of training performed by an intensivist (bc) and an internist (ag), icu patients who need a non invasive ventilation (niv) or tracheostomized elderly patients who have difficult weaning were admitted in a dedicated area in a medical department (md). this study desribes the results of one year of observation. in the last year, forty nine patients (age 70.2 ± 12.3; m 26 f 23) were transferred from icu to md. twenty three patients were treated with niv (age 67.2 ± 12.5; m 12 f 11), fourteen tracheostomized patients (age 72.2 ± 10.8; m 9 f 5) receive positive pressure ventilation because the difficult weaning in icu while twelve don't need any respiratory support. at the admission was performed a multidisciplinary plan and many specialists were involved (dietist,physiotherapist, pneumologist) and in invasively ventilated patients (ivp) was done a program of weaning. we follow all the patients until the discharge at home where someone need oxigenotheraphy, niv or mechanical ventilation. for the invasive ventilated patients we try to identificate significative differences beetween patients discharged at home and patients who died in hospital. data are given as mean ± sd and statistical analisis t test was performed results. patients underwent niv stay in hospital for 21.3 ± 19.2 days (7.0 ± 4.9 days in icu-15.1 ± 9.2 in md) and ventilation was performed for the entire period in icu while for 8.4 ± 5.6 days in md. all the patients were discharged at home: twelve with niv, fourteen with oxygen. the lengh to stay in hospital for the ivp in wich weaning was failed in icu was 91.2 ± 25.4 days (36.0 ± 18.4 days in icu-55.1 ± 23.6 in md). in md they continue the invasive ventilation for 44.4 ± 34.7 days. seven were weaned from ventilation after 20.9 ± 14.2 days, one was discharged at home with the ventilator while six died in hospital. patients who died were older (79.2 ± 7.7 vs. 67.8 ± 11.2 years-p 0.05), have more chronical diseases (3.0 ± 0.7 vs. 2.2 ± 0.44-p 0.03), longer hospitalization (112.2 ± 12.0 vs. 81.5 ± 20.3 days-p 0.005), glascow coma scale (8.2 ± 3.8 vs. 15). elderly patients often require a long period of recovery from acute ilness. in selected patients md could be a useful place where continue the treatment started in icu. in our study ivp who died had more chronical diseases and a more significative cognitive compromission. aim. documenting the qualitative and quantitative properties of administered and lost fluids is a common critical care monitoring practice. these nurse-registered fluid balances (fb) are used to optimize patient care and in clinical decision-making. this ''good clinical practice'' has also found application in research: recent studies reporting superior outcomes expressly refer to (negative) fb. we prospectively assessed the accuracy (review of all fluid balance charts and correction of arithmetic errors) and consistency (gold standard: body weight changes [bwc] registered with standardized measurements of body weight on admission and discharge [precision ± 50 g]) of nurse-registered cumulative fb. total (tfb) and daily fb (dfb = total fb/los) were calculated. we analysed the unadjusted cumulative fb (unafb: without considering additional losses, i.e. perspiration/fever/liquid faeces) and the adjusted cumulative fb (adjfb: considering the above as proposed in the literature) in all patients (all) and in three subgroups (cardiaccerebral:card; septic:septic; others). exclusion criteria: lack of admission/discharge weight, incomplete fb data. we calculated 1l = 1 kg. among 385 patients admitted during the study period 147 were eligible and analyzed. fb were inaccurate in 49 cases (33%) (error range: -3.61 to ?2.02 l, mean arithmetic error ± sd: ?0.03 ± 0.81 l, mean absolute error: 0.45 ± 0.6 l). the body weights at admission and discharge were 78.58 ± 18.92 kg and 79.29 ± 18.77 kg, with a bwc of 0.75 ± 3.25 kg (0.30 ± 1.27 kg per day). unatfb were 2.01 ± 4.02 l, unadfb 0.66 ± 1.25 l. adjtfb was 0.57 ± 3.45 l, adjdfb 0.20 ± 1.23 l. correlation (r 2 ) and bland and altman was poor between bwc and unatfb (0.552 and -1.26 ± 5.41 kg) and slightly better between bwc and adjtfb (0.714 and ?0.18 ± 3.68kg). the sd of the difference between bwc and fb per day of the icu stay was always [1 kg. a multiple regression model including unatfb, duration of intubation, maximum temperature, estimation of liquid faeces, age and the calculated caloric deficit during the icu stay, only modestly improved correlation (r 2 0.773). compared to the two other groups, septic were significantly more severely ill, had a higher and longer fever, a longer los, larger bwc and cumulative fb, and presented larger differences between bwc and cumulative fb (poor correlation and bland and altman). though, consistency betwenn bwc and cumulative fb in card and other was still scarce. conversely, another multiple regression model (including only unatfb and the maximal temperature) in septic yielded an r 2 of 0.988. conclusion. fb are often inaccurate and they are not consistent with the gold standard of bwc. the correlation and the agreement with bwc of both adjtfb and unatfb are poor, with sd per icu day-stay[1 kg or l. multiple regression models including several variables slightly improve correlation, yet remaining disappointing. consequently, clinical decisions should rather be based on other methods than fb. a prolonged hdu los was associated with a high sofa score for respiratory, hepatic and coagulation variables, preoperative ecg alterations, an increased urea and bmi and important bleeding. sofa score should be use in the first 24 h to assess organ failure and a possible icu transfer for patients with an elevated score. evaluation of procalcitonin, neopterin, c-reactive protein, il-6 and il-8 as a diagnostic marker of infection in patients with febrile neutropenia financed by the following fellowships: rd 06/0020/ 1056 from retics, fiss pi070444 and fijc p-ef-07 reference(s). 1. bohoun c (2000) a brief history of procalcitonin biomarkers of sepsis: is procalcitonin ready for prime time? definitions for sepsis and organ failure and guidelines for the use of innovative therapies in sepsis procalcitonin assay in systemic inflammation, infection, and sepsis. clinical utility and limitations usefulness of procalcitonin for diagnosing complicating sepsis in patients with cardiogenic shock patients at our er, icu and wards of internal medicine and surgery meeting the ssc criteria were included. blood cultures were taken before administration of antibiotics, other cultures when appropriate. laboratory tests included wbc, crp, lactate, and pct. we categorized patients using bacteriological criteria group 2 bacteriological proven infection (negative blood cultures but any other culture(s) positive) using chi square test there was no difference in survival, pct and findings on chest x-ray between the groups. with the mann whitney u test we found no differences in wbc, crp and lactate between survivors en nonsurvivors in a cohort of patients meeting the ssc criteria only 30 % met bacteriological criteria for sepsis. wbc, crp, lactate and pct did not differ between patients with and without bacteriological proof of infection nor between survivors and non-survivors il1rn and tnfr2 in the severity and outcome of community-acquired pneumonia to investigate whether polymorphisms within genes encoding for inflammatory or anti-inflammatory molecules are associated with susceptibility design. prospective observational, cohort study a cohort of 1,162 spanish caucasians with cap and 1 subjects were genotyped for the following polymorphisms: tnfa -238 and -308, lta ? sequential kaplan-meier survival analysis of tnfrsf1b ?676 g/t polymorphism showed a protective role of the gt genotype. cox regression analysis adjusted for age, gender, hospital of origin and comorbidities showed that patients with gt genotypes had lower mortality rates compared with those patients with gg or tt genotypes (p = 0.02 our study does not support a role for the studied polymorphisms of the tnfa, lta, il6 and il1rn genes in the susceptibility or outcome of cap. a protective role of heterozygousity for the functionally relevant tnfrsf1b ? variability in genes involved in the inflammatory response in patients with community-acquired pneumonia to investigate whether polymorphisms within genes encoding for inflammatory or anti-inflammatory molecules are associated with susceptibility design. prospective observational, cohort study a cohort of 1,162 spanish caucasians with cap and 1,413 controlsinterventions: subjects were genotyped for the following polymorphisms: tnfa -238 and -308, lta ? sequential kaplan-meier survival analysis of tnfrsf1b ?676 tt versus gg/gt genotypes suggested a detrimental role of the tt genotype. longrank c2 tests at 20 and 28 days yielded p = 0.052 and 0.042, respectively; cox regression for 20-and 28-day survival, adjusted for age, gender ghent university hospital, intensive care we aimed to describe the incidence and characteristics of healthcare-associated pneumonia (hcap) diagnosed at the emergency department of our university hospital compared to cap, hcap occurs in more debilitated or at-risk patients, is more frequently caused by nosocomial pathogens, and has worse outcome. nursing-home pneumonia is included within the definition of hcap but could have characteristics different from the other categories of hcap ) with a diagnosis of 'pneumonia'. episodes were categorized in cap and hcap according to the definition of the american thoracic society/infectious diseases society of america. within hcap, distinction was made between nursing-home pneumonia (nhp) and non-nhp hcap. severity of the pneumonia was assessed using curb-65 during the study period, 287 episodes of pneumonia were diagnosed in 269 patients; 159 episodes (55%) were categorized as cap, and 128 (45%) as hcap. within hcap, 32 (12%), respectively 96 (33%) episodes were further classified as respectively nhp and non-nhp hcap. median age of the patients was 71 years (57-81) and 61% of patients were male median curb-65pneumonia severity score in patients with cap and hcap was 1 (0-2) and 2 (1-3) respectively (p = 0.001); in nhp and non-nhp hcap, median curb in bivariate logistic regression analysis, both increasing curb-65 (or 1.8, ci 1.2-2.8) and categorization as nhp (or 3.4, ci 1.2-9 half of the episodes of pneumonia diagnosed at our emergency department could be classified as hcap. severity of the pneumonia was higher in patients with hcap as compared to cap. categorization as nhp, but not as non-nhp hcap was independently associated with hospital mortality after adjustment for severity of the pneumonia medical intensive care unit community-acquired pneumonia (cap) is the leading cause of infectious death, severe sepsis and the seventh leading cause of overall death. severe cap (scap) is defined as need of aggresive intensive care unit (icu) management due to shock to describe the episodes of severe community-acquired pneumonia (scap) in a multicentric european study and to assess management practices and outcome of scap patients admitted to icu observational, prospective, multi-centre study conducted in 27 icus of 9 100 consecutive patients requiring invasive mechanical ventilation for an admission diagnosis of pneumonia or mv for [48 h were recruited in each icu. statistic analysis was performed using spss years p \ 0.01) and presented a higher saps ii score at admission (45.45 sd15.59 vs. 51.30 sd 17.29 p \ 0.01). patients were treated with monotherapy in 19.7% and combination therapy 80.3%. empirical antibiotic treatment was in accordance with idsa guidelines in 100 (45.9%) patients. combination was prescribed with macrolides in 46.0% and quinolones in 54.0%. in patients receiving combination therapy in accordance with idsa guidelines, a cox regression analysis adjusted by saps ii and age identified that macrolides use was associated with lower icu mortality when compared to the use of quinolones in patients with severe community acquired pneumonia who had therapy in accordance with the idsa guidelines, only combination therapy with macrolides was associated with better outcomes the eu-vap/cap project was endorsed by eccrn hospital de mataró, critical care, mataró, spain, 3 hospital universitario germans trias i pujol, microbiology, badalona, spain, 4 hospital de mataró, microbiology, mataró, spain, 5 hospital universitario germans trias i pujol this leads to improved survival in a sterile acute lung injury model in wild-type mice, compared to mmp-8 knockout mice. we recently showed that mmp-8 deficient mice had better survival in a cecal ligation and puncture (clp) sepsis model than wild-type mice. in humans, functional genetic variations of the mmp-8 gene exist, but their relation to outcomes of severe infections, such as cap, is unknown. objectives. we hypothesized that functional human single nucleotide polymorphisms (snps) leading to increased mmp-8 levels are associated with worse survival and higher incidence of severe sepsis in patients with cap. methods. we examined data from genims, a multicenter prospective cohort study of patients with cap and analyzed 3 potentially functional snps (rs3765620, rs1940475, rs11225395) in the mmp-8 gene in 1567 caucasians by polymerase chain reaction (pcr) the overall incidence of severe sepsis was 28.7% (n = 450), and 11.6% of patients (n = 181) died within 90 days. the rs1940475 genotype distribution was significantly associated with 90-day mortality by armitage's trend test failure plot for rs1940475 conclusions. the non-synonymous rs1940475 snp is associated with 90-day survival in patients with cap. our findings suggests a trend towards a transcriptome analysis of ventilator associated pneumonia in trauma patients the sepsichip project the diagnosis of acute infection in the critically ill remains a challenge. transcriptional profiles (tp) of circulating leukocyte can be used to monitor the host response to infection. ventilator-associated pneumonia (vap) is a frequent complication of major trauma, raising morbidity and mortality data files were analyzed with r and bioconductor. unsupervised analysis was conducted using the dbf-mcl algorithm. supervised analysis was conducted using the significance analysis of microarray algorithm, using siggenes library. all statistical analysis used corrections for multiple comparisons. results. vap occurred in 41 of the 165 trauma patients (24.8%). one hundred and fifteen samples were hybridized on husg9 k microarray (41 sirs and sepsis samples for the patients who developed a vap, and 33 sirs samples for those who did not). whereas clinical parameters (iss, chest trauma) discriminated trauma patients with or without vap, admission samples transcriptome analysis did not lead to the identification of prognostic markers. analysis of paired samples of the 41 patients who developed a vap identified a transcriptional signature. these genes were involved in transcriptional regulation, cell survival, hemostasis and endocrine regulation. conclusions. by comparing whole blood admission samples, we were not able to identify transcriptional prognosis markers in trauma patients who did or did not develop vap. however, using vap as a model of sepsis in trauma patients, we identified a set of genes which may serve to diagnose vap in trauma patients to investigate the correlation between transfusion practice and the development of ventilator associated pneumonia (vap) in patients with traumatic brain injury (tbi) we analyzed which tbi individuals developed vap in regard to the transfusion practice and if the number of transfused prbcs increases the risk of pneumonia development. we counted the total amount of prbcs units received by each patient during icu stay, as well as those given before vap development. patient's data included: demographics, apache ii, iss, gcs, vap characteristics, duration of mechanical ventilation (mv), length of stay (los) and outcome. cpis and mods were calculated on the day of vap detection.statistical evaluation was performed using univariate and multivariate logistic regression, students t-test and pearson's chi square test 7%) tbi patients who developed vap received on average 6 units of prbcs during icu stay, compared with non vap individuals who were transfused on average with two units of prbcs (p \ 0.002). vap patients received on average four units prbcs before vap development. after correcting for age, apache ii, gcs and iss, transfusion was independently associated with vap. the odds ratio for vap aspiration pneumonia (ap) in comatose patients (pts): clinical and microbiological findings ap is a common complication in comatose pts. we aimed to update data on their incidence standard guidelines were used for diagnosis and treatment of ap. daily chest x-ray were retrospectively reviewed. ap was diagnosed if pts met following criteria: persistent radiographic infiltrate within 4 days following ti, and at least two of the following: purulent sputum, fever/hypothermia duration of mv was 3 days (2-5) and length of stay in icu 4 days (3-6) on the day of ap diagnosis, main pts characteristics were: saps ii 5%) received empirical antimicrobial therapy. main empirical antibiotics were coamoxiclav (63%) and third generation cephalosporin (13%) 001) and mv duration (6 vs. 2 days, p \ 0.001), even considering only non-cardiac arrest pts. gcs 24 h after ti and ap were associated with a [3 days duration of mv in multivariate analysis ap was associated with higher overall 28-day mortality in univariate analysis (53 vs. 29%, p = 0.02) but no longer in multivariate analysis mortality in the icu was 44.6% with a corresponding hospital mortality of 55.4%. a microbiological documentation was obtained in 21.6% of the patients, with streptococus pneumonia being the most frequent (25% of the isolates). the cap was classified as localized in 68.9%, unilateral mean (±sd) saps 3 piro score was 42.12 ± 12.34 points, with a corresponding predicted mortality of 60.39 ± 21.48% (standardized mortality ratio 0.917). the aroc was 0.702 (0.585-0.819). the value of the hosmer-lemeshow ĉ test was 27 saps 3 piro presented a discrimination similar to the originally described. however, it significantly overestimated mortality sepsis mortality prediction based on predisposition, infection and response the piro-cap score was proposed earlier this year to stratify patients with severe community acquired pneumonia (cap) to evaluate piro-cap score in patients with severe cap, over a period of 5 years we analysed data prospectively registered in an informatic data base, which contains information referring to all patients admitted in this unit survival curves were built as proposed by the original authors. outcome was evaluated at icu discharge overall, it was a severe population: 90.5% of the patients presented at least one chronic disease, saps 3: 78.38 ± 15.07 points (predicted mortality 66.01% ± 21.84), length of stay in the icu 11.05 ± 10.13, icu mortality was 44.6%. a microbiological documentation was obtained in 21.6% of the patients, with s. pneumoniae being the most frequent (25% of the isolates). the cap was classified as localized in 68.9%, unilateral icu mortality was 12 piro-cap presented an excellent discrimination. however, mortality rates were greater than the ones described by the original authors in all groups (except group 4), with the system significantly under-predicting mortality. consequently, we recommend caution in their widespread use. cumulative survival reference(s). 1 clinical and biological assessments in icu patients delirium is a life-threatening, acute organ dysfunction with an incidence of 65% in uk mechanically haloperidol is recommended as treatment [2] despite limited evidence base. objective. a national postal survey of consultant members of the uk intensive care society (ics) was performed to determine the current management of delirium in the ) drug treatment of hypoactive and hyperactive delirium as described by two clinical vignettes and (3) level of agreement with five statements regarding delirium. results. six hundred and eighty one replies were received from 1,308 questionnaires senta response rate of 52%. twenty five percent of respondents routinely screen for delirium. only 17% use a validated screening tool, most (65%) of whom use the confusion assessment method, icu. hyperactive delirium is treated pharmacologically by 95%, the majority using haloperidol. hypoactive delirium is treated pharmacologically by 25%, with haloperidol again the most common treatment (80%) grant acknowledgement. intensive care foundation, intensive care society a practical algorithm to diagnose delirium in critical care-validity and reliability delirium occurs in up to 80% of critical care patients [1], but often remains undiagnosed because standardized delirium monitoring is often dismissed as being too time-consuming or too complicated [2]. the 'harvard flowsheet', derived from the 'confusion assessment method for intensive care unit' (cam-icu) [3], provides a practical, algorithm-type handling advice to assess the four dsm-iv delirium criteria in a standardized fashion in intubated patients. it mostly allows for truncation of assessments to save time after approval from our institution's ethics committee, patients of a 31-bed surgical icu-department were screened in five sessions for delirium (1) by a psychiatrist as the reference rater using the dsm-iv delirium criteria, and (2) by two 'harvard flowsheet'-investigators, each unaware of other's ratings. motoric delirium subtypes were classified according to the richmond agitation sedation scale (rass) [4], which was rated for the feature 3 ('altered level of consciousness') of the 'harvard flowsheet'. patients were deemed as having hypoactive delirium if they were dsm positive by the reference rater and had rass -3 to 0, or having hyperactive delirium if their rass was between ?1 and ?4. for interrater reliability the median time to complete the 'harvard flowsheet' in delirious patients was 50s (iqr, 40-120 s) vs. 45 s (40-75 s)] in non-delirious patients. conclusions. the 'harvard flowsheet' has high sensitivity, high specificity and very high interrater reliability. false-negative ratings can occur infrequently and likely reflect the fluctuating course of delirium with intermittent lucid states a decrease of the overall cost of sedation and of sedation/day of mv followed protocol implementation and has been pursued each year: sedation cost which was greater than 50 €/day in 2000/2001 has decreased to less than 12 €/day in 2006 acute renal failure in critically ill patients: a multinational, multicenter study to evaluate current transfusion practice and the association between the age of red blood cells (rbcs) and outcome in critically ill patients design. prospective, multicenter observational study patients: 757 critically ill adult patients receiving at least one unit of rbcs 5%)] revealed an unadjusted absolute reduction rate (arr) in mortality of 9% (95% ci 3-16%). after adjustment for disease severity, patient age, other product transfusions, number of transfusions, pre-transfusion haemoglobin concentration, pre-icu transfusions, and cardiac surgery the odds ratio (or) for hospital mortality in critically ill patients in australia and new zealand transfusion of rbcs is delivered within current international recommendations. however, within such a practice patients enrolled included 328 men and 173 women, with mean age of 45 ± 2 years. there were 152 (30%) liver transplantation, 271 (54.9%) renal transplantation and 54 (10.7%) pulmonary transplantation and 24 (4.7%) renal-pancreas transplantation. the 90 days mortality for liver, renal, pulmonary and renal-pancreas was: 18 (11.8%), 7 (2.6%), 13 (24.1%) and 1 (4.2%). the mean saps 3 score for liver, renal ic apache ii auc 0.786 95% 0.588-0.918, ic 95% 0.643-0.929. conclusions. in these study, no differences were observed comparing saps 3 and apache ii in the mortality prediction from liver, renal and pulmonary transplantation current opinion in critical care14 introduction of a rapid response team: why we are glad we met dew ma and members of the medical emergency response improvement team (merit) committee (2007) mature rapid response system and potentially avoidable cardiopulmonary arrest in hospital the effect of a rapid response team implementation in a private hospital adult patients often exhibit physiological deterioration hours before cardiopulmonary arrest to determine the effect of a rapid response team on the rate of in-hospital cardiac arrests, total and unplanned intensive care unit admissions, and icu and hospital mortality before and after implementation of a rapid response team standard criteria were used to activate the rrt and included acute changes in the patient's mental status, respiratory rate, heart rate, oxygenation, or blood pressure and hypoxia, chest pain, or worry from clinical staff. we measured: admitting diagnosis after rrt were a total the 276 activations. the most common reasons for rrt activation were ventilator dysfunction (39%), cardiac changes (22%) and acute neurological changes (16%). 22% were transferred to icu and the main reasons were cardiac changes (39%), ventilatory dysfunction (36%) and acute neurological changes (16%) the rrt implementation was associated with decreases in rates of inhospital cardiac arrest, but was not associated with reductions in hospital or icu mortality the 100.000 lives campaign: setting a goal and a deadline for improving health care quality gender difference in critical care response team activations impact on outcome saudi arabia, 2 king saud bin abdulaziz university for health sciences, riyadh, saudi arabia introduction. gender-related differences in outcome of ccrt intervention has been documented in the literature indicating that more men were admitted to the intensive care unit our center is the only center in the kingdome of saudi arabia which implements an intensivest-lead ccrt services 24 h/7. the team is leaded by in house board certified in critical care medicine. ccrt services started in chest pain unit-viable option when risk of cardiac etiology is modest. multitudinous patient population brought together for structured survey and care at appropriate level what alters physicians' decisions to admit to the coronary care unit? m. camara 1 , g. silva 1 , s. silva 1 , c. dias 1 , j. nóbrega 1 , e. maul 1 1 hospital central do funchal, funchal, portugal introduction. procalcitonin (pct) and c reactive protein (crp) are markers of sepsis and the levels correlate with the severity of illness.aims. to evaluate the relationship of procalcitonin (pct) and c-reactive protein (crp) kinetics within the first days of sepsis with the appropriateness of antibiotic therapy and the outcome. a prospective cohort study, over 3 months including 49 patients with documented sepsis in our 10-bed intensive care unit. crp and pct were simultaneously measured four times (m1-m4) during the first 8 days of antimicrobial treatment. the pct and crp time course were analysed according to the appropriateness of the empirical antibiotic therapy as well as according to the patient outcome.results. between 1 january and 31 march of 2009, 149 patients were admitted to the icu. 49 patients presented with sepsis on admission or during their stay. the most common infection site was the lung (83.7%) followed by primary bacteraemia (4.1%). gram-negative and gram-positive bacteria were isolated in the following proportion: 30.6 and 22.4%, respectively. enterobacter, acinetobacter and escherichia were the most frequently isolated (6.1% each). gram-positive sepsis was mainly caused by haemophylus influenzae (12.2%). sepsis was polimicrobial in 6.1% of cases. 24.5% of the patients were given inappropriate antibiotics. the proportion of gram-negative bacteria isolated was significantly higher in patients who did not receive appropriate antibiotics. the magnitude of the pct and crp elevation was not associated with the appropriateness of antibiotic therapy. logistical regression analysis showed that infection without agent was an independent predictor of inappropriateness of antibiotic therapy.age, saps ii, apache ii and sofa were not associated with an unsuccessful treatment. regarding the absolute value of crp and pct there was no significant difference between successful or unsuccessful. multivariate analysis showed that dpct 3 was not associated with antibiotic appropriateness and mortality.conclusions. although the sample is small, our study suggests that crp and pct kinetics are not associated with the appropriateness of antibiotic therapy and outcome. introduction. patients with hematological malignancy who need advanced life support in the icu because of a life-threatening complication may have a poor prognosis. that's why it is necessary to identify clinical, analytical and biological factors that can help doctors with the decision to admit these patients into the icu.objective. the aim of this study was to assess the utility of procalcitonin serum levels (pct) in predicting the outcome of patients with hematological malignancies admitted to the icu. a total of 53 patients with hematological malignancy were admitted to the icu from january 2007 until march 2009. epidemiological data were collected before admission, and patients were followed up clinically and analytically during icu stay. serum samples were collected from icu admission until a maximum period of 10 days. pct values were measured by an immunofluorescent assay based on trace (time-resolved amplified cryptate emission) technology (kryptor pct, brahms ag, hennigsdorf, germany). mean age: 49 (sd 14); 29 men/24 women. among the patients included, hematological diseases were: non-hodgkin lymphoma (10 patients), acute myeloblastic leukemia (24), acute lymphoblastic leukemia (3) , multiple myeloma (6), chronic lymphoproliferative disorder (2) , others (8). twenty patients (38%) had previously received hematopoietic stem cell transplantation. thirty patients (57%) presented neutropenia at the moment of icu admission. the main causes for icu admission were respiratory failure in 23 patients (43%) and septic shock in 17 (32%). pct levels were not significantly higher in those patients that required mechanical ventilation. pct levels were significantly higher (p = 0,023) in those patients admitted because of septic shock. pct levels were lower in days 1, 2 and 3 in the patients who survived with respect to those who died: day 1: 12.67 ng/ml (sd 28.89) versus 18.54 (sd 23); day 2: 8.40 (sd 18.91) versus 19.43 (sd 22.78) ; day 3: 4.50 (sd 9.86) versus 17.22 (sd 25.72 ). the differences were significant in days 2 (p = 0.015) and 3 (p = 0,024). there was a trend to have higher pct levels in those patients who had microbiologically documented infection respect to the rest; day 1: 7.69 ng/ml (sd 10.18) versus 20.75 (sd:26.71 ); day 2: 7.01 (sd 7.73) versus 17.83 (sd 23.29 ) and day 3: 4.35 (sd 5.48) versus 16.29 (sd 23.83) .conclusions. serum pct levels are higher in patients with septic shock. serum pct measurement might be useful for predicting mortality in patients with hematological malignancy who require advanced life support. introduction. sepsis is a major cause of mortality in the intensive care unit (icu). efforts have been made to reduce the time needed to diagnose sepsis in order to reduce mortality from sepsis-related multiple organ dysfunction. procalcitonin (pct) has been reported elevated levels at the onset of bacterial infections and seemingly correlated to severity of infection. several clinical trials have detected a high pct level in patients with evidence of systemic bacterial infections, whereas relatively low pct levels occur in patients with only localized bacterial infections.objective. the aim of the present study was to assessed the ability of pct through sensitivity, specificity, positive and negative predictive value (ppv, npv) in patients with suspected sepsis, septic shock, inflammatory systemic response syndrome (sirs) and compared it with variables like crp, mortality, band%, renal failure, active cancer and an isolated bacterial cultures. finally we wanted to evaluate if exists a no infectious correlation in patients who received blood transfusions. we conducted an observational study including all patients admitted to the multidisciplinary icu of the abc medical center (tertiary reference hospital) to whom requested pct at admission in the suspect of sepsis and we followed their outcomes. total populations was 293 patients (p). 51% were females and 49% were males. median age was 58 years. of the total of pct sample 40% were positive and 60% were negative. the sensitivity and specificity in septic patients were 42 and 61%. ppv and npv were 57 and 47%, respectively. we did not found any statistical difference between positive value of pct and sepsis, septic shock, sirs, mortality, crp, band%, acute renal failure, acute lung injury, ards (acute respiratory distress syndrome), blood transfusions and active cancer. the mortality in the populations was 18%.conclusions. the pct has a wide range of diagnostic in the septic patients. in our study the rate of false positive was 42% and limited the use for sepsis diagnosis. we suggest that the better utility is for outcome biomarkers more than diagnosis biomarkers of sepsis. y. jin 1 , c. guolong 1 , iit study group of zhejiang province in china 1 zhejiang hospital, hangzhou, china introduction. the use of intensive insulin therapy (iit) in severe sepsis and septic shock has been shown to decrease morbidity and mortality rates significantly when given to high risk surgical patients.objectives. the aim of this study was to assess the efficacy of iit in severe sepsis and septic shock patients in intensive care unit.methods. this is a muticentre, prospective, randomized and controlled study. we randomly assigned patients who admission to icu with severe sepsis or septic shock into three groups: a group (target range for blood glucose is 80-110 mg/dl); b group (target range for blood glucose is 120-150 mg/dl); c group (target range for blood glucose is 180-200 mg/dl as a control). primary end point (28-day mortality for any cause) and secondary end points (icu stay days, mv duration, apacheii scores and mods scores) were obtained serially for 28 days and compared between the three groups. of the 356 enrolled patients, 115 were randomly assigned to group a and 110 to group b and 131 to group c; there were no significant differences between the groups with respect to base-line characteristics. 28-day mortality was 20 percent in the group a and 21.8 percent in the group b assigned to iit, as compared with 38.9 percent in the group c assigned to conventional therapy (p = 0.008).during the interval from first hour to 28-day stay in icu, the patients assigned to group a and group b had a significantly lower apache ii scores(12. 1 ± 4.1 and 11.4 ± 4.4 vs. 15 .0 ± 4.5, p = 0.02) and mods scores(3.58 ± 1.73 and 3.25 ± 2.12 vs. 4.75 ± 3.06, p = 0.027) than those assigned to conventional therapy, there were no differences in icu stay days(13.90 ± 1.70, 9.65 ± 1.21, 14.40 ± 1.54, p = 0.7)and mv duration(9.0 ± 1.2,6.0 ± 0.8, 8.2 ± 0.9, p = 0.09) between the three groups. compared with the conventional therapy group, the group a had a higher rate of severe hypoglycemia [blood glucose level b 40 mg/dl (2.2 mmol/l); 8.5.0 vs. 0.8%; p \ 0.001]. intensive insulin therapy provides significant benefits with respect to outcome and scores in patients with severe sepsis and septic shock in icu, on the other hand, intensive insulin therapy brings a higher rate of severe hypoglycemia. to determine the prognosis factors in elderly patients (c65 years) with severe sepsis admitted to an intensive care unit (icu).method. an observational, prospective and multicenter study was realized. it includes all the patients of the database edusepsis study (adults with severe sepsis admitted to 59 spanish medical-surgical icus). the clinical and demographic characteristics of all patients including age, sex, origin of the infection, location of the patient at the moment of diagnosis of sepsis, apache ii modified score (apache ii score age excluded), number of organic failures, initial therapeutic strategy (measures of resucitación and measures of treatment), icu length of stay and hospital mortality were registered. the patient were classified in young cohort (\65 years) and elderly cohort (c65 years). elderly cohort patients were also classified in young-old patients (65-79 years) and very-old patients (c80 years). descriptive comparative study of both cohorts was realized and multivariate logistic regression for the two subgroups of elderly patients was performed to study the risk factors of hospital mortality. a total of 2,796 patients wer enrolled (62.2 ± 16.3 years, apache ii modified score of 17.5 ± 7.5, 3.3 ± 1.5 organic failures, hospital mortality 41.6%). the elderly cohort (n = 1489; 53.2%) presented a lower apache ii modified score (18.0 ± 7.9 vs. 17.1 ± 7.0, p 0.002), higher abdominal infection as origin of the sepsis (35.6 vs. 23.1%, p \ 0.001), higher nosocomial infection (60.0 vs. 57.7%, p 0.013) and a lower application of measures at initial treatment (13.2 vs. 16.2%, p 0.025) than the young cohort. there were not significant differences in the number of organic failures and days of stay in uci between both cohorts. the apache ii modified score (or 1.08; 95% ic 1.06-1.11; p \ 0.001), the nosocomial infection (or 1.54; 95% ic 1.17-2.04; p \ 0.001), the thrombocytopenia (or 1.53; 95% ic 1.11-2.11; p 0.009) and the acute renal failure (or 1.47; 95% ic 1.07-2.03; p 0.018) were associated independently to mortality in the subgroup of young-old patients. in the very-old patients only the apache ii modified score (or 1.12; 95 % ic 1.07-1.17; p \ 0.001) was independently associated with higher mortality and in this population subgroup the application of measures of initial resuscitation was a protective factor (or 0.21; 95 % ic 0.05-0.94; p 0.042).conclusions. the elderly patients (c65 years) admitted in the icu whith severe sepsis have higher mortality, more abdominal infections as origin of the sepsis and fewer application of measures of initial treatment than the young patients (\65 years). nevertheless, in the subgroup of very-old patients (c80 years) the aggressive initial treatment decreases the mortality. objectives. the aims of this study were to determine the crude and related to bacteremia mortality rates in icu patients with bacteremia who receive appropriate empirical antibiotic therapy and to describe the factors associated to mortality in this appropriated treated patients material and methods. during a twelve years and a half period, from 1996 to 2008, 413 icu-patients with clinically significant bacteremia were prospectively evaluated. for purposes of this investigation, appropriate empirical antimicrobial treatment of a bloodstream infection (aeat) was defined as the microbiological documentation of infection that was effectively treated based on its antibiotic susceptibility at the time the causative microorganism were suspected. clinical and microbiological variables were recorded. logistic regression analysis was performed to determine the risk factors associated to global and associated to infection mortality. results. among 413 icu-bacteremic patients, aeat was applied in 283 patients (68.5%). apache ii and sofa score were 21.06 ± 8.1and 8.4 ± 4.1, respectively and the incidence of septic shock was 40.3% in this aeat patients. global and associated to infection mortality rates were 53.4 and 24.7%, respectively in aeat patients. logistic regression analysis confirmed copd (or 7.07; 95% ci: 0.009-0.77) and age (or 1.06; 95% ci: 1.01-11) as factors independently associated to global mortality and diabetes mellitus (or 0.20; 95% ci: 0.05-0.77) presentation as septic shock (or 0.18; 95% ci: 0.38-0.93) and serum levels of albumin (or 0.28; 95% ci: 0.10-0.75) as a protective factors for global mortality whereas factors as nosocomial origin (or 0.08; 95% ci: 0.009-0.77) and again serum levels of albumin (or 0.06; 95% ci: 0.12-0.38) were considered protective for related mortality to bacteremia conclusions. mortality rates remains excessively high in aeat bacteremic-icu patients. different factors were identified as predictive factors for global and associated to mortality in aeat patients. only serum levels of albumin seems to be an independent protective factor for both global and associated to infection mortalities. introduction. severe sepsis is hallmarked by organ hypoperfusion or dysfunction. the transition from severe sepsis to septic shock carries with it an increase not only in morbidity but also in mortality [1, 2] . objectives. the aim of the study was to demonstrate the effect of shock at admission in sepsis comparing severe sepsis and septic shock admission diagnoses.methods. single center retrospective study in a 12 bed mixed icu of a tertiary university hospital. during a 2-years period of study 792 patients were unplanned admitted in the unit: the median was age of 55 (38-70), the males were 63.4% and the mean of sapsii was 44 ± 15. we randomly select two groups: severe sepsis (72 patients) or septic shock (126 patients) at admission. statistical analysis of variables: v 2 , mann-whitney test, unpaired t student test, cox regression. no statistical significant differences were found about age and sex between groups. about the origin of infection no statistical significant differences were found between groups, meanwhile the diagnosis respiratory infection appears to be more frequent in the severe sepsis group (58.3 vs. 44.4%, p 0.06). the proportion of post-operative admissions (in surgical related conditions) was not different between groups. the sapss ii and sofa at 24 h were higher in the septic shock group [49 (39-61) vs. 39 (29-45) , p \ 0.01]; 9 (6-11) vs. 4 (3) (4) (5) (6) (7) , p \ 0,01, respectively]. sofa at discharge appears to be higher in the shock septic group (excluding deaths) [3 (2-5) vs. 3 (2-4) (p 0.053)]. the mortality and length of stay (excluding deaths) was higher in the shock septic [33.3 vs. 12.5% (p \ 0.01); 10 (6-13) vs. 5 (3-11) (.01), respectively]. the ventilator associated pneumonia was not significantly different between groups. the probability of discharge, across an initial period of 15 days, was lower in the septic shock group [hazard ratio 0.47 (95% ci: 0.40-0.87)], mainly between the 5th and 10th days, as shown in the kaplan-meier plot (see graph 1) .admission diagnosis: probability of discharge conclusions. septic shock at admission patients had a poorer outcome. the difference in the probability of discharge between groups was higher when mechanical ventilation related events are likely to occur [2, 3] . we emphasize the importance of the institution of early goal-directed therapy in the wards and emergency departments prior to admission in an intensive care unit [1, 4] .introduction. it is not clear whether patients with community acquired severe sepsis (cass) or hospital acquired severe sepsis (hass) have a same presentation. objectives. to evaluate the characteristics of a severe sepsis (ss) population admitted through the er (cass) and those coming from the ward (hass). all patients were treated by the same team of intensivists and er doctors in a shock room, so we could minimise the differences due to management. methods. all adult patients admitted to the medical icu were eligible if they met the criteria for ss. we collected demographic characteristics, apache ii and sofa score, comorbidities and immuno-compromised conditions. scvo 2 or svo 2 (if possible), lactate concentrations. the milestones of the surviving sepsis campaign (ssc) were measured regularly during the first 6 h of treatment. the data collection went on in the icu stay too. treatment for septic shock was conformed to the recommendations of ssc. results. we enrolled 44 pts with ss, including 20 with cass and 24 with hass. there was no difference in demographic features and comorbidities, including immuno-compromised conditions, haematological malignancy and chronic respiratory diseases .there were no significant differences in hemodynamic variables or indices of tissue perfusion like scvo 2 (or svo 2 ) and blood lactate levels, or in amounts of fluids infused or needs of vasopressor agents. the need for mechanical ventilation (mv) after the first 6 has greater for hass than for cass patients, but during the icu stay the need for mv was the same for both groups; similarly, during the icu stay there was no difference in the need for extracorporeal renal support or need for adrenergic agents. at the beginning the scvo 2 was around 65% for the entire population. after the first 6 h both groups reached the target of 70%. at the admission 10% of patients had a scvo 2 less than 50% (12.5% for hass patients and 7.1% for cass, without any difference between groups) and 40% of patients had a scvo 2 higher than 70%. the mean svo 2 for both groups was higher than 65% already at the beginning of the observational period. conclusion. only a half of pts with ss or sho had fever. the presence of fever is often associated with a positive microbiological diagnosis, but better prognosis. while hypothermia was often viewed in severe ill pts and was associated with a worse prognosis. to investigate the possible differences in characteristics and outcome between early and late-onset severe sepsis in surgical intensive care unit (icu) patients. we conducted a retrospective analysis of prospectively collected data from all adult patients ([18 years) admitted to our 50-bed surgical icu between 1st march 2004 and 30th july 2006.results. of 5,925 patients admitted to our icu during the study period, 234 patients (3.9%) had severe sepsis; 74 (31.6%) had early-onset and 160 (68.4%) late-onset severe sepsis. respiratory infections (48.1 vs. 27.0%, p = 0.002) and infections of unknown origin (21.9 vs. 12.2%, p = 0.005) were more frequently recorded in patients with late-onset than those with early-onset severe sepsis, whereas abdominal infections were more frequent in early-onset than in late-onset severe sepsis (20.3 vs. 7.5%, p = 0.005). gram-positive infections were more frequent in late-onset than in early-onset severe sepsis (63.1 vs. 51.4%, p = 0.036). the time of onset of severe sepsis was not independently associated with an increased risk of in-hospital death (early vs. late: or 0.68 95% ci 0.36-1.29, p = 0.689).conclusions. respiratory infections and infections of unknown origin were more frequently recorded in patients with late-onset than in those with early-onset severe sepsis, whereas abdominal infections were more frequent in early-onset than in late-onset severe sepsis. the time of onset of severe sepsis has no impact on mortality. objectives. to describe the causes, microbia spectrum, and prognosis of pregnancyassociated sepsis treated in icu in france along the last 33 years. we conducted a retrospective study in a medico-surgical icu of 10 beds in a non-teaching hospital in france where a high risk maternity unit was opened in 1995. patients admitted between 1976 and 2008 for sepsis occurring during pregnancy or the post-partum period were included. the patients were excluded if the sepsis was due to a nosocomial icuacquired infection. charts were reviewed to collect data on sources of infection, microbia, maternal and fetal prognosis. data are shown as median (extremes) or percentage. data before and after 1995 were compared using non parametric tests.results. 66 patients were admitted for pregnancy-associated sepsis (10% of total pregnancy-related icu admissions). included patients had the following characteristics on admission: age: 31 (17-40) years, gravidity: 2 (0-8) pregnancies, parity: 1 (0-8) children, , . vasopressors, mechanical ventilation, and hemodialysis were required in respectively 25, 25, and 5% of cases.characteristics of infections are shown in table 1 . microbiological data about bacterial infections, and specially infections of pelvic origin (chorioamniotitis, endometritis, septic thrombophlebitis), are shown in table 2 .all urinary infections were due to e. coli. lung infections were most often documented clinically but not microbiologically.maternal mortality rate was 6% (2 deaths before and 2 deaths after 1995). for those infections that occurred in the pre-partum period, fetal mortality was 40%. after exclusion of fetal deaths that had occurred before icu admission, pregnancy was interrupted during icu stay in 18% of cases, resulting in fetal mortality of 9%.conclusions. despite of disappearance of post-abortum sepsis in france, sepsis remains a significant cause of icu admission during or after pregnancy, and a significant cause of maternal and fetal mortality.grant acknowledgement. none. introduction. the glasgow coma scale (gcs), universally used for assessing comatose states, has the advantage of ease of use making it accessible to all levels of clinical competence. there are, however, significant drawbacks. its relative subjectivity in the interpretation of verbal and eye responses and its mesencephalic limit in the rostro-caudal assessment of brain vitality. these two drawbacks limit its use particularly in icu intubated patients. we propose a new score, the sousse coma score (scs) that overcome the eye and verbal responses and explore the brain vitality up to brain death. the score ranges from 10 (normal consciousness) to 0 (brain death). the performance of this score was compared to a modified gcs (gcsm; gcs reduced to its only eye and motor components) and four score. our study interested 100 prospective and consecutive comatose patients who were admitted to a medical icu, intubated and under ventilatory support. the level of consciousness was assessed at admission by physicians of different levels of competence. the inter-observer reliability was assessed by measuring the spearman correlation between the responses of different observers to the scs, gcsm, four score and their respective components. the prognostic predictive value of the three studied scores was assessed by the analysis of possible correlation with mortality and the roc curves. inter-observer reliability was excellent (spearman rho [ 0.8) for the three studied scores, but with better performance for the scs (0.913, p \ 0.0001) compared to the gcs (0.863, p \ 0.0001) and four (0.893, p \ 0.0001). the level of overall inter-observer reliability for gcs and four was paradoxically higher than that of their respective motor components. this is probably the result of a summation effect of their respective components.regarding the relationship between mortality and the studied coma scores, there was for all three scores a threshold below which mortality was 100%; 5/10 for scs, 3/15 for the gcs and 5/16 for the four. beyond this threshold, only the scs provides a highly significant correlation with the risk of death (spearman = -0.907, p = 0.0001). a similar correlation was observed with the motor components of the gcs and four. a better correlation was found between mortality and the scs. the area under the roc curve, however, was poor for all three scores. in evaluating the minimally consciousness states and from eight value of scs, the gcs and four scores provided a wider and more subtle level of consciousness assessment. the scs provides a better inter-observer reliability and a better prediction of death while being easier to achieve. the apparently good inter-observer reliability of gcsm and four was simply the result of summation effect. however, scs was not very sensitive in detecting variations of the minimal consciousness states. the results of our study should be confirmed in larger multicenter studies for the external validation. introduction. the use of a sedation assessment scale and a sedation goal is recommended in critically ill adults [1] . several studies have found a reduction of icu length of stay (los), of mechanical ventilation (mv) duration, and of cost [2, 3] . but, durability and efficiency of such procedures over time have not been evaluated. a 15-bed medical icu in a university hospital. a sedation and analgesia protocol has been implemented since 02-2002. the sedation goal is prescribed each day by the intensivist; the ramsay sedation score is evaluated by the nurses every 4 h and doses of drugs are adapted accordingly. we conducted an annual survey from 2001 to 2008 to evaluate the quantity of sedative drugs utilized, the impact on icu los and on mv duration, and cost of sedation; both the medical and nurse staff were regularly informed and the protocol was modified if necessary. u. guenther 1 , j. weykam 1 , u. andorfer 1 , t. muders 1 , h. wrigge 1 , c. putensen 1 1 university of bonn, anaesthesiology and intensive care, bonn, germany background. acute brain dysfunction (delirium and coma) is reported to occur in up to 92% [1] , and to be associated with longer mechanical ventilation and stay in the icu, and increased 6-months mortality rates up to 34% [2] . such outcome data, to the best of our knowledge, are predominantly given on medical patients with delirium incidences and mortalities much higher than we expected in surgical patients. this study assessed incidence and impact of acute brain dysfunction on length of stay on the ventilator and in the icu, and mortality in cardiac surgery patients. after approval from our local ethics committee, every patient admitted to our cardiac surgery 11-beds icu from october through november 2007 was daily monitored for delirium with the ''confusion assessment method for the intensive care unit (cam-icu)'' [3] , level of consciousness was assessed with the richmond-agitation-sedation scale (rass) [4] . acute brain dysfunction was diagnosed if patients were comatose without sedative medication or delirious. patients were contacted 6 months later to obtain information about their further clinical course.results. 75 patients were eligible for analysis [male 42, female 33, age, mean (iqr), 69 (67-72) years]. 56 % had acute brain dysfunction while in icu, these had significantly higher apache-scores on admission, higher tiss-and saps-scores, were longer mechanically ventilated [11 (6-15) vs. 2 (1-2) days, p \ 0.0001, mann-whitney test) and had longer stay in icu [15 (10) (11) (12) (13) (14) (15) (16) (17) (18) (19) (20) (21) vs. 5 (4-7) days, p = 0.017, mann-whitney test). 8 patients were lost to follow-up; 6-months mortality in patients with acute brain dysfunction in icu was 30 vs. 8% (p = 0.045, log-rank test).discussion. incidence of acute brain dysfunction and mortality found here in cardiac surgery patients are lower compared to reports on medical patients. even though, duration of mechanical ventilation, length of stay in icu, and 6-months mortality were increased in patients with acute brain dysfunction in our icu.conclusion. these data emphasize the need for routinely monitoring of consciousness and delirium and to develop strategies to reduce incidence of acute brain dysfunction. introduction. the incidence of obesity is increasing globally, with 1 billion overweight people (body mass index (bmi) [ 25 kg/m 2 ), and 300 million of them obese [1] (bmi [ 30 kg/m 2 ). the number of obese individuals presenting to critical care is likely to increase, but data on effect of obesity on outcome is conflicting [2] . there is a general perception that obese patients are likely to have a higher incidence of adverse outcome in critical care. to investigate the effect of bmi on length of stay and mortality in patients admitted to critical care in city hospital, birmingham, uk.methods. an observational study was performed over a 3 month period from february-april 2008. pre-morbid data on bmi was collected from medical records and direct questioning of patients. apache ii score at 24 h, critical care length of stay (los) and survival data to hospital discharge were collected. the figures were compared against their predicted mortality. readmissions, patients with a los \ 12 h, patients \ 16 years old and those without complete apache ii data were excluded. a total of 162 patients were admitted. 17 met the exclusion criteria, bmi data was unavailable for 27 patients and apache data was unavailable for 4 patients. 114 patients were included in the final analysis. 45.6% were female and 54.4% were male.63% of patients were above their ideal body weight, and 27% were obese. the obese cohort had a mean apache ii score of 17.8 with a mean los of 6.75 days and a mean hospital mortality rate of 12.5 %. the corresponding figures for the non-obese group (bmi \ 30) was 17.11, 10.7 days and 33.7% (table 1) . obese patients had reduced hospital mortality in comparison to predicted rates from apache ii scores. statistically, there was no increase in mortality of obese patients. the los and ventilated days were also comparable to the nonobese patients. introduction. managing glucose levels in critically ill hospitalised patients has been shown to play a role in improving clinical outcomes. as a result glycaemic control protocols are widely used in critical care settings and require rapid and frequent testing of patient glucose levels. poc glucose meters have migrated from ambulatory testing into hospital.there is increasing recognition that the clinical accuracy of nearly all commonly used glucose meters are affected by components or substances often present in the blood matrix of critical care patients giving rise to an increase risk of adverse incident. the aim of this study was to challenge the accuracy of a glucose meter designed to correct for these interferences.methods. 100 paired random arterial whole blood samples were collected from icu patients admitted for [72 h, samples were tested for glucose using stat strip glucose (nova biomedical) and the omni b221 bga (roche) routinely used for blood gas analysis.statistical methods: spearman rank correlation/regression, bland-altman analysis. results were compared to iso 15197 standard for glucose measurements. omni bga: correlation coefficient r 2 = 0.99, slope = 0.96 with intercept -0.09.bland altman plot for absolute glucose concentration showed that mean bias compared to reference was -0.16 ± 0.27 mmol/l with limits of agreement -0.69-0.37 mmol/l. study aim. to assess predictors of t and its impact on icu-, hospital-length of stay and costs in a cardiac surgical patient cohort admitted to our eight bedded icu, since 1997 through june 2004.methods. all the pre-, intra-and post-operative variables were prospectively put into an electronic database. patients were divided into: (1) ntg group, not needing a tracheostomy;(2) tg group, undergoing a tracheostomy. p values \ 0.05 were considered significant. out of a total of 5,123 patients with a median (iqr) age of 67 years (59-73) (1) from post-operative icu to the cardiac surgical ward and (2) from the cardiac surgical ward to the rehabilitative one, increased significantly higher in the ntg group than in tg group (respectively: log-rank = 260.9, p = 0.0000 and log-rank = 77.1, p = 0.0000,). tg group showed a lower mortality (3.2 vs. 36.6%, p = 0.000) than ntg one.conclusion. this study allowed us (1) to define a predictive model for identifying patients that are likely to undergo a tracheostomy (2) introduction. patients admitted to itu increasingly have significant medical comorbidities that require chronic therapy for adequate control1. on admission to itu, acute medical problems take precedence and many long term medications, such as thyroxine, may be withheld or ceased. in chronic hypothroidism the patient is physiologically dependent upon ongoing administration of thyroxine. the optimal management of chronic medical conditions such as hypothyroidism within the itu may be essential to patient recovery and should be a quality assurance issue. to assess the prescription of thyroxine and the thyroid function in patients admitted to itu with previously diagnosed chronic hypothyroidism. a six year retrospective review of the electronic records of patients with hypothyroidism who were admitted to a 30 bed tertiary referral hospital itu from 2002 to 2008 was performed. patients were included if they were admitted to the itu for a period of more than 7 days and were on thyroid replacement therapy prior to itu admission. patient demographics, daily thyroid replacement dose/route, thyroid function tests (tsh and free t4) and rate and type of nutrition were obtained. patients were grouped by their worst recorded tsh according to predefined ranges . conclusions. patients did not receive their thyroid replacement for a significant proportion of their admission. this was predominantly due to either lack of prescription or lack of tolerance of enteral feed. the tsh was appropriate for the free t4 level. a significant proportion of patients (22%) did not have their tsh measured at all. of those that did, abnormal tests were inconsistently repeated or acted upon. having processes in place to ensure the appropriate prescription and adjustment of relevant chronic medications is essential in the provision of high quality care in itu.background. cell-free dna has been investigated as a diagnostic marker in many diseases, including acute conditions such as stroke, myocardial infarction, burns, sepsis etc. its serum and plasma levels have been shown to correlate with disease severity in all those. free circulating dna is released from dead cells (necrotic or apoptotic) and activated inflammatory cells. our hypothesis was that in acute pancreatitis free serum dna correlates with the extent of pancreatic necrosis and that it may be an early marker of severity. free dna was measured in sera from 30 patients with acute pancreatitis at admission, on the first, fourth and seventh day following admission. severetiy of illness was assessed with atlanta criteria. on the first day following admission patients who would develop severe pancreatitis had significantly higher serum dna levels than those with mild disease (median 0.271 vs. 0.059 ng/ml respectively; p \ 0.001). this parameter showed very good characteristics as a potential predictor (area under roc curve 0.97). free serum dna was in correlation with the extent of pancreatic necrosis.conclusions. free serum dna correlates with the extent of pancreatic necrosis and is a potential early marker of severe acute pancreatitis.keywords. acute pancreatitis, cell-free dna, prognostic marker, pancreatic necrosis. introduction. the performance of general prognostic models in patients with transplantation in need for intensive care unit (icu) admission is poor, showing a tendency towards significant underestimation of the risk of dying. the objective of our study is to evaluate the acute physiology and chronic health score ii (apache ii) and simplified acute physiology score (saps 3) and their 90 days mortality prediction after liver, renal and pulmonary transplantation [1] [2] [3] .methods. this is a prospective cohort study in a transplantation icu in porto alegre, brazil, during the period of may 2006-december 2007. clinical data of 501 pos transplantation patients admitted at icu were collected at admission and saps 3 and apache ii calculated with respective estimated mortality rates. the area under receiver operating characteristic curve (auroc) was obtained for both scores. objectives. to validate the saps 3 model, over a period of one year, in a general icu. material and methods. we analysed data prospectively registered in an informatic data base (icdoc), which contain information referring to all patients admitted in this unit.we studied all the patients admitted in the year 2008 (385 patients). excluded from the analysis were 25 readmissions and one patient still in the hospital. analysed 359 patients.results. discrimination was accessed by the area under the roc curve (aroc) and calibration by the hosmer-lemeshow ĉ test for the general equation and for regional equations (southern europe and mediterranean countries).the mean age of the patients was 63.9 ± 15.5 years. from the total of the patients, 163 were medical (45.4%), 143 were scheduled surgical (39.8%) and 53 were emergency surgical (14.8%). the mean icu and hospital mortality was 18.7 and 33.1%. the mean saps 3 score was 58.84 points (20-115). discrimination was good with an aroc of 0.800 (0.752-0.849).there was a statistical significant difference between the mortality predicted by the general equation and the observed mortality (ĉ = 16.4; p = 0.036); this discrepancy was not significant by using the regional equation (ĉ = 12.77 and p = 0.12). the saps 3 overestimated hospital mortality with the predicted mortality by the regional equation getting closer to the observed mortality [standardized mortality ratio (smr) = 0.93] than the predicted by the general equation (smr = 0.92).conclusion. the saps 3 model, particularly using the regional equation introduction. saps 3 has been previously validated in our icu and it has been routinely used in hospital mortality prediction. as we have shown before, saps 3 had a good accuracy regarding discrimination and calibration, with better predictions done by north american and western europe customized equations than the south american one [1] . in a larger sample we have been observed deterioration in calibration model, especially among groups of lower probability of death, regardless of the equation in use. therefore we tested saps 3 accuracy considering 30 days mortality in comparison to hospital mortality. we considered 576 consecutive admissions in a medical-surgical icu in a private tertiary hospital in sao paulo -brazil, in the period from january to november of 2008. probability of death was derived from given equations of the original study [2] . hospital and 30 days mortality were considered as end point. discrimination was performed by the area under the roc curve (auroc) and calibration by the hosmer-lemeshow (hl) statistic. observed to expected (o/e) mortality ratio was also calculated. to assess factors concerning prognosis of patients older than 64 years admitted to the icu: group a, 65 to 74 years old and group b, older than 74 years. both groups were compared for the apache ii, admission group, lenght of stay, mortality and usual intensive care procedures (arterial and venous catheters, mechanical ventilation). statistical anlysis: quantitaive variables were expressed as mean and standard deviation (sd). student t test was employed for these variables. categorical variables were compared by the chi-square. p \ 0.05 was considered statistically significant. a total of 804 patients were included in group a (mean age 69.96, sd 2.8) and 605 in group b (mean age 78.81, sd 3.58). apache ii score was 13.86 for group a and 15.24 for group b (p = 0,04); predicted mortality was 22.14 and 23.41% respectively (p = 0.33). ther were no differences for admission group or procedures among groups. mortality was significantly higher in group b (16.5 vs. 20.8%, p = 0.04). when mortality was analyzed for admission groups, it was higher just in cardiological group, wich included ischemic cardiopathy, cardiac failure and arrhythmia (5.1 vs. 9.7%, p \ 0.05). the investigation of the association between a differential access to intensive care services and patient or hospital outcomes is increasing markedly [1] [2] [3] [4] . objectives. the aim of this study was to compare demographic, clinical characteristics, and outcomes of patients admitted to tertiary-level intensive care units from a tertiary hospital ward (intrahospital transfer) to patients transferred from a secondary hospital ward (interhospital transfer). single centre retrospective study in a 12 bed mixed icu of a tertiary university hospital. during the study period (2007) (2008) conclusions. the interhospital transferred patients are younger, but at admission severity of the disease is comparable.these findings, within this case mix of patients, suggest there are not significant differences in mortality, length of stay, icu-nosocomial respiratory infection or physiological disability at discharge between intrahospital and interhospital transferred patients to our unit. in this study we did not find a different impact in outcome considering these differential sources of admission. we prospectively analysed data of all patients (pts) undergoing cardiac surgery between january 1998 and june 2005, and discharged from our icu by 24 h from surgery. on all patients the following was collected:(i) demographics, risk factors and gravity scores anamnestic illnesses (ii) intra-operative variables [i.e. type of operation, cardiopulmonary by-pass (cpb) and aortic cross clamp (acc) times] (iii) icu-related variables. one-way anova test was used for continuous variables whereas, differences in proportions were compared using chi-squared test.a binary logistic regression model was used to estimate the effect of each considered risk factor on discharging from cardiac surgical to rehabilitative ward, considered as a dycotomous outcome (yes = early b 7 days/no = late [ 7 days). statistic analyses were performed using spss software. p values less than 0.05 were considered significant. on all the patients, aged c80 years, admitted to our post-operative icu since january 1994 through december 2006, we collected demographic profiles, operative data and outcomes. a logistic regression model was set up to assess predictors of hospital outcome. a total of 428 patients (4.8%), 53.5% males and with a median (iqr) age of 83 (82-85) were admitted. the below table shows the outcome predictors (see table 1 ). objectives. the purpose of this study was to evaluate prospectively in our medium the capacity of apache iii score to stratify prognostically critically-ill-patients upon their admission to the icu, not only with regard to hospital mortality, but also to hospital length of stay. study aim. to assess if cardiopulmonary by-pass (cpb), aortic cross clamp (acc) time and duration of mechanical ventilation (mv) may impact on icu and hospital length of stay in a cardiac surgical patient cohort admitted to our 8 bedded icu, since 1997 through june 2004. all the patient pre-, intra-and post-operative variable were prospectively put into an electronic database. on all patients the following was collected:(i) demographics, risk factors and gravity scores anamnestic illnesses (ii) intra-operative variables [i.e. type of operation, (cpb) and (acc) times] (iii) icu-related variables (i.e. duration of mechanical ventilation, use and type of inotropes. statistic analyses were performed using spss software. p values \ 0.05 were considered statistically significant. a total of 5,123 patients with a median (iqr) age of 67 years (59-73) were admitted through the study period. 63.5% underwent a cabg operation, whereas 22.8% valve surgery and 13.6% aortic and lung surgery. a bivariate analysis was performed considering as independents variables respectively the natural logarytm (nl) of (1) cpb time, (2) acc time, (3) mv duration, whereas dependent variable was considered the nl of the total hospital stay. we showed that a linear correlation exists between total hospital stay (ln) and (1) conclusion. this audit allowed us to assess that the longer is the cpb and acc time and mv duration the longer is likely to be the total hospital length of stay of the patients undergoing heart surgery. introduction. high-dependency units (hdu) were designed as a bridge between the operating theatre and the surgical ward for postoperative patients demanding a higher than standard level of care. the aim of our study was to determine the risk factors as well as the predictive value of four severity scores for a prolonged hdu length of stay (los). three hundred fifty-eight consecutive adult patients were included in the study for a period of 6 months. asa, saps ii, possum and sofa scores were calculated for the first 24 h following admission. the demographic and the scores variables were subjected to a univariate and, consecutively, a multivariate logistic regression analysis. a receiver operating curve (roc) model was used to determine the predictive value of the scores for a prolonged los. the presence of a patient for three or more days in the hdu was defined as prolonged stay.results. the median los was 3 (1-18) days, 14 patients were transferred to the intensive care unit and the in-hospital mortality was 4.1% (15 patients). the univariate logistic regression revealed the following variables as significant for a prolonged los (p \ 0.05): asa, possum preoperative, possum postoperative, possum total, possum cardiac, possum ecg, possum type of surgery, possum blood loss, sofa, sofa respiratory, sofa cardiovascular, sofa liver, sofa coagulation, igs ii, igs respiratory, igs urinary output, igs urea, igs potassium, igs bicarbonate, and bmi. seven variables were identified as having a statistically significant association with the los (table 1) . according to the roc model, sofa score was the best predictor for a prolonged los, with an area under the roc (auroc) of 0.753.warning systems and rapid response team: 0375-0388 0375 s. saxena 1 , s. jafrey 2 , j. zwaal 2 1 kingston hospital, anaesthetics, kingston, uk, 2 kingston hospital, kingston, uk background. published data suggests that the patient group with the highest mortality in icus comprises those patients admitted from the hospital wards [1] . studies have shown that in-hospital cardiac arrests are commonly preceded by physiological abnormalities [2] . if admission to icu, is preceded by specific physiological derangement, then early identification of these high risk hospital in-patients may be possible. this may improve survival of patients. objectives. to determine 1. the effectiveness of new track and trigger pathway in identifying patients requiring icu admissions. 2. the impact of new system on outcome of icu admissions method.1. retrospective case notes survey of all icu admissions from the ward over a 4 month period. 2. the pathway is triggered when abnormalities are present in two or more of the following parameters: response to painful stimuli, respiratory rate, oxygen saturation, systolic blood pressure, and heart rate. 3. 3.triggering steps progress through involvement of junior medical staff and outreach teams at step 1, to more senior staff at step 2, to consultant involvement at step 3, depending on the level of deterioration of the patient. . 30 forms were collected over a period of 4 months. icu mortality: patients with 1 abnormality at any time prior to icu admission: 2/10 (20%) icu mortality: patients with c2 abnormalities any time prior to icu admission: 10/19 (52.6%) mortality of patients who were pathway followers: 2/5 (40%) mortality of patients who were pathway non-followers: 10/24 (41.6%) average length of stay in icu who were survivors from pathway followers: 25 days average length of stay in icu who were survivors from pathway non-followers: 3.6 days discussion.1. there was low sensitivity of pathway for identifying icu admissions. 2. poor documentation of triggering events 3. pathway followed inadequately in majority of patients due to combinations of delay in, or absence, of triggering when indicated 4. lack of consultant involvement at step 3 5. no patients with chronic kidney disease admitted to intensive care have poor outcomes [1, 2] . in 2001 68% of cardiac arrest calls in our hospital were from the renal unit (personal communication.) at this time critical care outreach teams were recommended as means of improving intensive care outcomes through earlier ward assessment of critically ill patients [3] [4] [5] . in 2006 modified early warning system (mews) charts to wards and a dedicated seven-day ward-based consultant led service (0800-1800) were introduced on our renal unit [6, 7] . aims and methods. the impact of these change interventions was analysed. primary outcomes were the incidence of cardiac arrests calls to the renal unit, admission apache ii scores and icu mortality. secondary outcomes were age, sex, intensive care and hospital length of stay, in-hospital mortality, cpr prior to icu admission and emergency admissions to the renal unit. cardiac arrests, mortality rates and emergency admissions were compared with the v 2 test; other outcomes via mann-whitney u test. a p value \ 0.05 was regarded as significant.results. the results are outlined in the table 1 [1] ; this group has a high mortality [2] . deterioration in vital signs often precedes referral to critical care and this is evidenced by a rise in the patient at risk score (pars). higher pars may be associated with worse patient outcomes [3] . most pars systems have a trigger value at which critical care input should be sought. we hypothesised that the duration of physiological deterioration prior to critical care admission would be associated with mortality and used the delay between pars trigger and admission as an estimate of this.methods. we collected data on over 200 consecutive patients that had deteriorated on the ward and required admission to general critical care (hdu and icu) at both acute hospitals in sheffield. patients admitted to specialist facilities such as cardiac and neurosurgical units were excluded. those already triggering at time of hospital admission were also excluded. to assess if any of the ccrt activation criteria was associated with higher incidence of icu or hospital mortality. our hospital is 900 bed tertiary care center. cohort analysis of prospectively collected data of each ccrt activation including demographic data of the patients and their outcome in terms of icu and hospital mortality and ccrt activation criteria.634 ccrt activation from 1st january 2008 to 30th september 2008.the activation criteria for ccrt includes: threatened airway,tachypnea defined as respiratory rate more than 30 or less than 8 breath/min, hypoxemia defined as oxygen saturation less than 90% on oxygen flow 6 l/min, arrhythmias defined as heart rate less than 40 or more than 130 beat per minute, hemodynamic instability if systolic blood pressure less than 90 mmhg or more than 200 mmhg, decrease level of consciousness defined as drop of gcs = 2 or more points from baseline, seizure and serious concern about the patient.we analyzed each factor separately as independent predictor of icu and hospital mortality. [1] . with this in mind efforts have been made to develop physiological early warning scoring systems which have been shown to predict subsequent outcome [2] . we have recently introduced an early warning system (ews) chart for all patients in our hospital and we wanted to assess its impact on our icu admissions. to assess the calculation of the ews, the scores of patients admitted to icu and the compliance with guidelines regarding further intervention for patients who were ultimately admitted to icu.methods. chart review of twenty five consecutive emergency icu admissions, examining the ews in the 24 h prior to admsission.results. ews charts were completed for 96% of emergency icu admissions; of these 88% of scores were calculated correctly. only 32% of ews had all parameters completed for all set of observations. the mean peak ews prior to icu admission was 5.9 with a range from 3 to 10. higher peak ews was strongly associated with increased icu mortality: a ews of 3-7 was associated with mortality of 10.5%, whereas a ews of 8-10 was associated with a mortality of 80% (see below). for each ews recorded specific action was required to be triggered according to the protocol. in 50% of cases appropriate action was taken, however, in 50% the required action was not taken and a number of patients were thought to have delayed referral to critical care as a result of this.conclusions. following this audit we have introduced a critical care outreach team and have embarked on an educational programme for staff with emphasis both on the complete and accurate recording of early warning scores and the necessity for appropriate action to be taken on the basis of these scores. aim. the quality of care prior to icu admission has been a focus of attention [1] . mews had been chosen by the trust as a trigger device to identify deteriorating (sick) patients in the general wards. this retrospective study looked at the clinical characteristics of unplanned admissions to our icu and assessed the mews as a predictive tool to trigger early intervention in such cases.methodology. all patients who were non-electively admitted to our icu from the medical wards were included in the study (january-march 2009 [1] score and standardised mortality ratio(smr) had occurred since the introduction of our nurse lead outreach serrvice and high dependency unit. this seemed counterintuitive so we decided to look at in futher detail and whether the phenomena of lead time bias occurred.objectives. primary end point was to assess if the apache ii scores and smr were different if assessed from the point of contact with outreach or hdu for patients admiitted to general intensive care. secondary endpoints looked at which physiological scores were most altered by these systems. a cohort prospective study was setup with ethics committee approval. all patients seen by outreach (group 1) or on hdu (group 2) prior to admission to general icu were included over a six month period. two sets of apache ii scores and mortality prediction were generated for each group, a 'pre' and 'post' score. the pre score was a 24 h scoring period started from up to 12 h prior to admission to icu on point of contact on hdu or by outreach. the post score was a period for scoring taken 24 h from the point of admission to icu, ie the conventional apache ii score .therefore each patient had two sets of scores for apache ii and predicted mortality. the apache ii and predicted mortality scores were then compared using a two tail paired t test, the individual physiology scores were compared via a wilcoxon rank sum score. in total 35 patients from hdu were included and 41 patients from outreach were included.the primary question was answered as a significant difference in apache ii and smr was found in both groups. introduction. unplanned admissions to intensive care units (icu) are associated with an increased mortality. in order to identify in-hospital patients at risk of deterioration, several scores based on physiological parameters have been published. however, routine application of these parameters is not common in all european hospitals yet. the goal of this prospective study was to evaluate the efficiency of the current practice of handing over ward patients at risk for decompensation by physicians and nurses. furthermore, factors associated with admission to the icu or alarming of the physician on duty should be identified. the study was conducted at the university hospital of regensburg, germany on wards with predominantly gastroenterological and general medical patients (38 beds). over a time period of 3 months, the daily routine report of patients at risk to the physician on call after hours was recorded. in addition, the nurse assessment of patients at risk and the documentation of the decompensation defined by calling the physician on duty during the night were registered.results. 526 patients were treated during the surveillance period. in total, 298 patients (134 women, 164 men) with a mean age of 59 ± 17 years were either judged by the attending physicians or the nurses at risk for deterioration. 260 patients suffered from decompensation during the night shift. of those, 86 patients were correctly identified by physicians and 69 patients by nurses, respectively. in 12 patients (2%), an icu admission was necessary.discussion. only a small portion of patients reported at risk experienced a severe decompensation at night, defined as icu admission. interestingly, those were only in part correctly identified by the physician and nurse reports. a further evaluation of the correlation of those reports with the previously published ''early warning score'', and physiological parameters associated with decompensation are currently being performed in order to estimate the value of standardized patient assessment, and will be presented at the meeting. introduction. the need to implement a patient follow-up program after icu discharge arises from several facts: (1) at icu discharge patients are now more fragile (aged, chronic comorbidities, complex). (2) the demand for intensive care exceeds its availability. as much as 25% of patients die after discharge from the icu, many of them in spite of a low predicted mortality, perhaps due to premature icu discharge. (3) compared to nursing care in the icu, the level of that received upon transfer to the floor, as measured by tiss-28, may be reduced up to more than 60%. we believe that, in order to change icu behavior towards focus on long term outcomes, we need to increase global awareness of disability post-icu discharges, and expand the involvement of the icu team in key decision management outside the icu. we propose an alternative model of care for the critically ill patient. this involves an expanded role for clinicians with expertise in critical illness at several points along the continuum of care.objectives. due to lack of adequate clinical resources to care for some recoverable patients when are discharged to hospital wards after a long time in icu, we have planned a follow-up program focused in detecting risk factors associated to bad prognosis and, decreasing adverse events in general hospital wards. qualitative, prospective and interventional study realised during seven months (from march to september 2008), in the medical uci of a teaching hospital in malaga. we determined demographic data, icu admission reason, comorbidity index (charlson scale), follow-up reason (polineuropathy, tracheostomy, analgesia), family support in ward, difference in nursing activities score between icu and ward (tiss-28), intervention done out of icu with patient; satisfaction of patient, icu readmissions, reason to end follow-up and mortality at day 60 after icu discharge. we enrolled 33 patients in this analysis. comorbidity was charlson scale 4 (very high) in 60.5% of patients, apache-ii mean score 18 points and mean expected mortality rate 27%. more than 72% of patients had five or more risk factors (age [ 74 years, icu stay [ 11 days, transfusions, inotropic drugs, mechanical ventilation, tracheostomy, kidney failure, parenteral nutrition, polineuropathy). nursing activities score in icu was 27.4 before discharge versus 10.2 in ward (62.8% decreasement). mean follow-up were 2.5 (range 1-5).in hospital mortality rate was 4.5%, rest of the patients were discharged at home. our study found the implementation of continue follow-up program from icu staff is associated with an important decreased of the mortality. encouraging clinical results and a non excesive workload for icu staff justify to continue this follow-up program in cases in which is going to be an important decrease in nursing care after icu discharge, and have bad prognosis risk factors. objectives. we examined the prevalence of adverse events (ae), suboptimal assessments of vital signs and whether there were advance directives prior to icu admission from the general wards among patients who died within 30 days of icu admission. the patients were those admitted to the general icu from the general wards at the university hospital, lund in 2008 and who died within 30 days after icu admission. there were 69 patients with a mean age of 73 years and a mean apache ii score of 26. we used the global trigger tool model for measuring ae (http://www.ihi.org). the frequency of vital functions assessments, and which parameters were controlled were studied in relation to patient status and the local routine for frequency of modified early warning scoring (mews).the patient records were also controlled for descisions to forgo treatment before admission to the icu. . 19 patients (27%) suffered from at least one ae prior to icu admission. 16 patients had an ae contributing to death, among those14 patients suffered from an ae that with a probability greater than 50% was deemed avoidable. seven of those patients suffered from a most likely avoidable ae contributing to death.vital signs were recorded inadequately in 43% of the patients in the 24 h before admission to the icu. the vital signs most often recorded were blood pressure, heart rate and oxygen saturation, whereas consciousness and breathing frequency were the least recorded parameters.descisions to forgo resuscitation, or some other limitations due to ethical considerations were found only in 25% of the patients before admission to the icu.conclusions. patients admitted to the icu who died within 30 days suffer from a considerable proportion of avoidable ae contributing to death. vital signs are not recorded in a satisfactory way during the 24 h before admission to the icu in this most severely ill population. there are very rarely documented descisions to forgo treatment in this group of patients before icu admission. thus, poor control of vital signs in the general wards leads to severe and avoidable ae contributing to death. the lack of descisions to forgo treatment before icu admission in this group of patients most probably contributes to prolonged dying and suffering, and unnecessary intensive care. results. a significant correlation (p \ 0.05) was detected between the type of prehospital care and the early outcome among the patients. the majority of the patients was transported by ambulance services (60.6%) from which half of the patients (29.4% of the total) were seen by a paramedic and the rest by a physician beforehand. a relevant proportion of the patients visited the cpu without having been seen by medical personnel (37.5%) before. a smaller group of patients was referred by an attending hospital physician to the cpu (1.8%). 68.2% of all patients were admitted to a cardiologic ward, 15.4% to icu and 12.1% underwent immediate cardiac catheterization. the rest was referred to other departments within the hospital, other hospitals or was discharged and no one died within 24 h after admission to the cpu. almost half of the patients (45.2%) who underwent immediate cardiac catheterization was transported by emergency physician car whereas half of the rest (25.4%) visited the cpu as out-patient (p = 0.00). this very similar ratio can be seen within the patient admitted to icu (27.3%). conclusion. the detection of the early symptoms of chest pain is an important prevention strategy for lay people because they can immediately turn to a chest pain unit (41.4%) where almost half of them might have life threatening situation (52.7%) requiring acute intervention (cardiac catherization or icu-treatment). the adequate in-time treatment can reduce the length of hospitalization and secure quicker recovery. key: cord-005497-w81ysjf9 authors: nan title: 40th international symposium on intensive care & emergency medicine: brussels, belgium. 24-27 march 2020 date: 2020-03-24 journal: crit care doi: 10.1186/s13054-020-2772-3 sha: doc_id: 5497 cord_uid: w81ysjf9 nan ventriculostomy-related infection (vri) is a serious complication in patients with hemorrhagic stroke. in such patients, diagnosis of vris is complicated by blood contamination of csf following ventricular hemorrhage. we aimed to evaluate the diagnostic potential of white blood cells count (wbc), c-reactive protein (crp), and procalcitonin (pct) to identify vris in patients with hemorrhagic stroke during the time of external ventricular drain (edv) in situ. this retrospective study was conducted at the neurosurgical-icu, university hospital of zurich. a total of 347 patients with hemorrhagic stroke and an external ventricular drain (evd) were admitted over a 6 years period at the icu. of those, 14 patients with vris ("vri"), defined by positive csf bacterial culture and increased wbc in csf (>250/ul), and 115 patients without vris and with serial csf sampling ("no-vri") were analyzed. patients with csfcontamination or suspected vri (negative csf cultures but antibiotic treatments) were excluded. wbc, crp, and pct were measured daily. csf was sampled routinely twice a week or by t>38°c. for the analysis, mean peak values of wbc, crp, pct during the time of evd in situ were compared between groups (t test). data are expressed as mean with ci 95%. results: between groups, wbc and crp were similar (wbc: 15.13 g/l and 14.55 g/ l, p=0.68 and crp: 115.93 mg/l and 129.44 mg/l, p=0.56 in the group vri and no-vri, respectively) ( figure 1, panel a and b ). in the group vri, pct was low and significantly lower than in the group no-vri (0.16 ug/l and 2.61ug/l, p=0.03 in the group vri and no-vri, respectively) (panel c). wbc in csf were similar between groups (710.14/ul and 675.16/ul p=0.93 in the group vri and no-vri, respectively). in this study, serum-inflammatory markers were not able to screen patients with vris. their routine measurement should be carefully evaluated. introduction: central nervous system (cns) infections constitute a potentially lifethreatening neurological emergency. patients admitted to the intensive care unit (icu) usually present with a severe disease and organ failure, leading to high mortality and morbidity. we have performed a retrospective analysis during a 5-year period of patients admitted to a polyvalent icu. clinical, demographic and outcome data were collected to evaluate its clinical impact on the outcome of patients with cns infections. we identified 30 patients with the diagnosis of meningitis, meningoencephalitis and ventriculitis, where the median age was 57,6 years (range 24-80). upon clinical presentation, their most frequent signs were fever (70%), meningeal signs (40%), seizures (30%), and a glasgow coma scale score <8 (66%). all needed ventilation support and 66% needed cardiovascular support. a definitive microbiological diagnosis was achieved on 22 patients and antibiotic therapy was adjusted on 18 of them. most common microorganisms were streptococcus pneumoniae (n=7), listeria (n=5) and pseudomonas aeruginosa (n=4) (figure 1 ). other gram negative microorganisms were detected and lead to more adverse outcomes. meningitis was the cause of admission on 26 patients and on a minority (n=4) meningitis was considered to be a secondary diagnosis on patients admitted for other causes (traumatic brain injury, subarachnoid or intraparenchymal hemorrhage, postoperatively of neurosurgical tumor). patients that eventually died had at least one risk factor (age>65, immunocompromised due to diabetes, corticotherapy, hiv or heart transplantation). patients admitted to the icu were not so aged, but had some comorbidities and risk factors leading to more uncommon microorganisms, increasing the risk of adverse outcomes. this lead to an increase of mortality: 23% in the icu and an overall of 43%. study of selenium levels in unresponsive wakefullness (uws) patients with systemic inflammatory response syndrome (sirs) e kondratyeva 1 , s kondratyev 2 , n dryagina 2 the objective of this study was to evaluate the pharmacokinetics (pk) of levetiracetam (lev) in critically ill patients with normal and augmented renal clearance (arc), and determine if the recommended dosage regimen provides concentrations in the therapeutic range (12-46 mg/l) [1] . a prospective observational study was conducted in a tertiary hospital. six blood samples were taken during a dose interval at steady state and lev was quantified by hplc. a population pk study was carried out. statistical analysis was conducted to evaluate the differences in pk between patients with and without arc. the suitability of drug concentrations was also assessed. results: seventeen patients were included, 13 with normal creatinine clearance (crcl) (80-129 ml/min) and 4 with crcl≥130 ml/min (arc). ten patients received 500mg q12h, one 1000mg q12h and two 1500mg q12h. the data were best fitted to a two-compartment model. figure 1 shows lev concentrations during the dosing interval. mean clearance (cl) was 4 l/h and mean volume of distribution of central compartment (v) was 44 l. interindividual variability was 38 and 61% for cl and v, respectively. no differences were identified between both groups (p>0.05) in pk parameters. no correlation was found between lev cl and crcl. trough levels were below the minimum concentration (c min ) 12 mg/l of the therapeutic range in all patients except 1. furthermore, between 3-5 h 50% of samples were below the c min . conclusions: administered doses were not able to maintain lev concentrations in the recommended therapeutic range. other dosage strategies, such the extension of infusion time with higher doses, could be evaluated in order to obtain a more favourable profile. no correlation between lev cl and crcl was found. the mechanical properties of muscles such as tone, elasticity, and stiffness are often affected in chronic critical ill (cci) patients. a hand-held device known as the myotonpro demonstrated acceptable relative and absolute reliability in a ward setting for patients with acute stroke [1] . the technology works on the principle of applying multiple short impulses over the muscle bulk via the testing probe. the aim of our study is to assess the feasibility of objective measurement of muscle tone in cci patients with neurological dynamics and serum biomarkers. the study included 23 cci patients with neurological disorders (stroke, traumatic brain injury, neurosurgical intervention for brain tumors) with more than a 3-weeks stay in icu. dynamic measurements of the muscle properties were taken on the deltoideus, brachioradialis, quadriceps femoris, gastrocnemius using the myo-tonpro. to identify the leading factor in impaired muscle tone also were measured neurological (s100, nse), inflammatory (il-6), bacterial load (pct) biomarkers using elecsys immunoassay and the serum level of microbial metabolites using gc-ms (thermo scientific). results: all patients were divided into groups depending on positive and negative clinical dynamics. significant differences were obtained in parameters characterizing changes in muscle tone of lower limbs -f gastrocnemius (tone) -15.5 vs 18.5 hz, r quadriceps femoris (the mechanical stress relaxation time) -16.5 vs 13.6 ms (p < 0.01, respectively). some significant correlations between five parameters of muscle tone biomarkers and microbial metabolites were revealed. the results of a quantitative measurement of muscle tone objectively reflect the dynamics of neurological status, which in the future may be promising technique for the personalized approach cci in patients. introduction: changes in hormonal status in patients with unresponsive wakefulness syndrome (uws) remains poorly understood. methods: 275 patients in uws were examined at the period from 2007 to 2017. 152 patients (115 men) with tbi and 123 patients (63 men) after hypoxia. acth, cortisol, tsh, free t3 and t4, sth, prolactin and natriuretic peptide were studied in the period from 2 to 4 months uws. in men, the level of total testosterone, lh and fsh was additionally studied. the obtained data was compared with the uws outcome in 6-12 months (crs-r scale assessment). none of the studied hormones of the hypothalamic-pituitary-adrenal axis were a reliable criterion for predicting the outcome of uws. most often and consistently was revealed a tendency of disrupt the rhythm of cortisol secretion, with higher rates in the evening hours. the average value of sth was higher in men with the consequences of head injury who had recovered consciousness than in those who remained in uws. significant decrease in testosterone levels, regardless of age, was found in patients with a consequence of tbi. mean levels of lh were higher in patients with tbi and hypoxia who remained unconscious than in patients who later restored consciousness. the average level of fsh was higher in patients who had recovered consciousness . the increase of natriuretic peptide level was observed both in patients who remained in chronic uws and in those who restored consciousness. no certain endocrine background, characterising this category of patients was found. violations of some hormones secretion rhythms, in particular, cortisol can be considered usual for uws patients, especially in patients with tbi. therapeutic hypothermia has not been used before our research in chronically critically ill (cci) patients. temperature decrease in neuronal cells is a strong signal that triggers endogenic cytoprotection programs using early response genes expression. our goal is to determine influences of craniocerebral hypothermia (cch) on level of consciousness in cci patients. we examined 98 patients with different types of brain injuries. 54 males and 44 females, mean age 45.56 ±16.03. patients were divided into 2 groups: main group -47 patients (vegetative state (vs) -28, minimally conscious state (mcs) -19), comparison group -51 patient (vs -32, mcs -19), groups were equal on main parameters (severity, functional state, comorbidity). patients from main group received courses of cch, duration -180 minutes, scalp temperature 5-8°с, cerebral cortex cooling up to 32-34 o c, session end was without slow reheating period, and session's amount was set -until signs of consciousness recovery. cortex temperature check done noninvasively by using detection of brain tissue emi in shf-range. consciousness recovery in vs and mcs patients controlled using crs-r scale. results: cch sessions significantly increased level of consciousness in vs and mcs patient groups. in vs patients vegetative state increased until minimally conscious state and mcs +, and in mcs group until lucid consciousness (p <0.05) (figure 1 ). craniocerebral hypothermia is used in chronically critically ill patients for the first time. our research results demonstrated effectiveness of cch as an additive treatment tool in such patients. this let us optimistically determine the perspective of inclusion of cch method in chronically critically ill patient's rehabilitation to increase level of consciousness. despite the clinical benefit of endovascular treatment (evt) for large vessel occlusion (lvo) in ischemic stroke, space-occupying brain edema (be) represents a common complication during the course of disease. routinely, ct imaging is used for monitoring of these patients, notably in the critical care setting, yet novel and easy bed-side techniques with the potential to reliably predict be without repetitive imaging would be valuable for a time and cost effective patient care. we assessed the significance of automated pupillometry for the identification of be patients after lvo-evt. we enrolled 40 patients admitted to our neurocritical-care unit who received evt after anterior circulation large vessel occlusion. we monitored parameters of pupillary reactivity [light-reflex latency (lat; s), constriction and re-dilation velocities (cv, dv; mm/s), and percentage change of apertures (per-change; %)] using a portable pupilometer (neuroptics®) up to every 60 minutes during the first 72 hours of icu stay. be was defined as midline-shift ≥5mm on followup imaging within 3-5 days after evt. we assessed differences in pupillary reactivity between patients with and without be (u-test) and evaluated prognostic performance of pupillometry for development of be (roc analysis). in 32 patients (19 women, 74.3±12.0 years) without be, 1,224 assessments were compared to 207 assessments in 6 patients (3 women, 71.7±11.8 years) with be. on day 1, day 2, and day 3 after evt, patients with be had significantly lower cvs and dvs, and smaller perchanges than patients without be, whereas lat did not differ between both groups. roc-analyses revealed a significant negative association of cv, dv, and per-change with development of be. conclusions: automated pupillometry seems to identify patients at risk for be after evt. a prospective study should validate whether automated pupillometry harbors the potential to reduce unnecessary follow-up ct imaging. the aim of this preliminary analysis is to detect differences between the qualitative and quantitative evaluation of the pupillary function carried out by doctors and nurses of an intensive care unit (icu) of a tertiary level hospital. secondary purpose is to investigate new indications for the use of pupillometry in a population admitted in icu methods: the study has been conducted (currently in progress) at the intensive care unit and ecmo referral center at careggi teaching hospital (florence; italy). the enrolled patients are adult subjects (> 18 years) with alteration of consciousness defined by a glasgow coma scale (gcs) < 9, following a primary brain injury and/or the use of sedative drugs. the studied parameters, obtained with neurolight pupillometer ® (id-med, marseille, france) are analyzed, integrated and visual/qualitative evaluation of the pupil function shows a lower reliability if compared to automated pupillometry. the estimated error in the proper determination of photomotor reflex is 34.9% (p< 0.01). no significant difference is reported between quantitative and qualitative pupillometry in the detection of anisocoria. our preliminary results are compatible with previously reported data [1] [2] [3] , even if there was no difference in anisocoria determination. interestingly, a longer latency period among patients treated with opioids has been observed. other results are still in progress. introduction: due to the dynamic of critical care disease, a rapid bedside, noninvasive and highly sensitive and specific method is required for diagnosis. in this study we set out our experience with trancranial color-coded duplex ultrasound (dxt) [1] . the dxt study identifies cerebral arteries as well as hemorrhagic phenomenon, hydrocephalus, mass-occupying lesions and midline shift. this is the main difference between dxt and conventional transcranial doppler (dtc) which is a blind study and do not provide any image. descriptive, cross-sectional and observational study from december 2018 to june 2019. 21 patients were included. inclusion criteria: neurocritical patients. exclusion criteria: no acoustic window, presence of ultrasound artifacts. data collection was performed. it was used a lowfrequency transducer from 1.5-3.5 mhz with trancranial duplex preset ( figure) . the patterns were defined as normal, vasospasm, high resistance, hypermedia and cerebral circulatory arrest, depending on the cerebral flow velocity, lindegaard ratio (lr) and pulsatility index (ip). results: 12 men (57.1%) and 9 women (42.9%). average age 55.6(20-79). patients diseases: subarachnoid hemorrhage 6, traumatic brain injury 5, av malformation 4, stroke 2, hemorrhagic cerebrovascular accident 2 and mass occupying lesions 2. normal pattern: 10 patients (rel. freq 0.47). vasospasm: 5 patients (rel. freq 0.23). high resistance: 4 patients (rel. freq 0.19). hyperemia: 1 patient (real. freq 0.04). cerebral circulatory arrest: 1 patient (rel. freq 0.04) conclusions: dxt should be part of the routine of neuromonitoring, it allows real time images especially useful in unstable conditions. although it will be needed a large amount of patients to be statistical significant, dxt is useful considering a non invasive study, bedside and it allows early identification of different clinic conditions. introduction: embolization of the draining vein during endovascular treatment of arteriovenous malformation (avm) may result in venous outflow obstruction and hemorrhage. anaesthesiologist can use deliberate hypotension to reduce blood flow through avm which may be somehow helpful to prevent this scenario. adenosine-induced cardiac arrest may facilitate the embolization too. the goal of our study was to improve the results of endovascular treatment of avm using adenosine-induced cardiac arrest. methods: after obtaining informed consent 13 patients (8 male, 5 female) were selected for adenosine-induced cardiac arrest during endovascular avm embolization. main age was 40,8±6 years old. 9 of them were evaluated as iii class asa, 4 as iv. endovascular treatment in all cases was performed under general anaesthesia. propofol, fentanyl, rocuronium were used to induce anaesthesia, then all the patients were intubated and ventilated with parameters to keep etco2 32-35 mm hg. sevoflurane 2,1-2,6 vol% (12 cases) or desflurane 6 vol% (1 case) were used to maintain anaesthesia. hemodynamic monitoring consisted of ecg, pulsoximetry, non-invasive blood pressure measurement. onyx or/and squid were used as embolic agents. ct was performed to every patient just after procedure as well as neurological examination. results: adenosine dosage was 0.875-1.26 mg/kg. time of consequent cardiac arrest was 10-40 sec. there were 10 cases we administered adenosine for 1 time, in one case we had to administer it twice, in one fig. 1 (abstract p014) . circle of willis and pulsed-wave doppler mode of middle cerebral artery -3 times and 4 times in one more case as well. hemodynamic parameters recovered without any particular treatment in all the patients. embolization has been performed in all the cases uneventfully. postoperative ct showed no hemorrhage. nobody from investigated group had neurological deterioration in postoperative period. our study shows that adenosine-indused cardiac arrest is not very difficult to perform method and it can be useful during avm embolization. a major risk factor for stroke is atrial fibrillation (af). to treat af anticoagulation is needed. there are now several anticoagulants available. however, a lack of head to head data as well as the absence of accurate techniques makes it difficult to compare them and measure determine there efficacy. stroke is known to produce an abnormal clot microstructure which is a common factor in many thrombotic diseases. this pilot study aims to use a functional biomarker of clot microstructure (d f ) and clotting time (tgp) to investigate the therapeutic effects of different anticoagulants in stroke and af. we recruited 114 patients (59 af and 55 stroke & af). two samples of blood were taken: before anticoagulation (baseline) and post anticoagulation (6-10weeks) . patients were either given warfarin (31%) or axipaban (69%). d f and tgp were measured and compared before and after anticoagulation. results: warfarin increased t gp (267±56 secs to 332±78 secs (p<0.05)), and decreased d f (1.71±0.05 to 1.65±0.06 (p<0.05)). apixaban increased tgp (235±66 sec to 410±105 sec (p<0.05)) but did not change df (1.72±0.04 & 1.72±0.05). interestingly we found that in the apixaban group tgp significantly correlated (p=0.05) with blood drug concentration levels. in this study we show that d f and tgp can quantify and differentiate between the therapeutic effects of two different oral anticoagulants. showing that warfarin prolongs clotting and weakens the ability of the blood to form stable clots. conversely apixaban prolongs clotting time but does not affect the bloods ability to form stable clots. this shows the utility of the d f and tgp biomarkers in comparing two different treatment options, something no other current marker has proven able to do. where d f and tgp may prove useful tools in a personalized approach to anticoagulation treatment and monitoring in an acute setting. hospital mortality compared to the model with the original hairscore. patients with poor-grade aneurysm subarachnoid hemorrhage (asah) world federation of neurological surgeons (wfns) grades iv and v, have commonly been considered to have a poor prognosis (70-100% mortality). though early intervention and aggressive treatment in neuroicu has improved outcome in the past years, it is controversial because most of the patients left hospital severely disabled. the objective of this study was to investigate the clinical and social outcomes in intracranial aneurysm patients with poor-grade asah underwent different intervention therapies. a single center observational registry of 25 poor-grade asah consecutive patients, defined as wfns grades iv and v, treated at tertiary chilean referral center from december 2013 to march 2019 were enrolled in this study. the clinical data including patient characteristics on admission and during treatment course, treatment modality, aneurysm size and location, radiologic features, signs of cerebral herniation (dilated pupils), and functional neurologic outcome were collected. clinical outcomes were assessed via gose and and sociooccupational outcome, both at discharge and at 6 months. figure 1 ). 20% mortality is less than previously reported, and survivors had a favorable recovery, confirmed with neuro psychological test. poor-grade asah patients in our study shows a more positive outcome than previously considered. prognosis of subarachnoid hemorrhage (sah) is scarce, indeed almost half patients die or become severely disable after sah. outcome is related to the severity of the initial bleeding and delayed cerebral infarction (dci). infection and more precisely pneumonia have been associated with poor outcome in sah. however, the interaction between the two pathologic events remains unclear. therefore, we hypothesized that dci may be associated to pneumonia in sah patients. thus the aim of our study was to analyze the association between delayed cerebral infarction and pneumonia in patients with sah. in this retrospective, observational, monocentric cohort study, patients included in the analysis were admitted in neurosurgical intensive care unit or surgical intensive care unit in the university hospital of brest (france) for non-traumatic sah. primary outcome was diagnosis of dci on ct scan or mri 3 months after sah. multivariate analysis was used to identify factors independently associated with dci. a total of 224 patients were included in the analysis (female male ratio 141/83, median age 57 [47-65] years). multivariate analysis was adjusted on sedation, intracranial surgery, fisher classification of sah severity, pneumonia occurrence and non-pneumonia infectious event occurrence ( figure 1 ). pneumonia occurred in 66 patients (29.5%) and other causes of infections in 45 patients (20.1%). dci was found in 108 patients (48.2%). factors independently associated with dci were pneumonia (or 3.10 [1.41-7.06]; p=0.006) and non-pneumonia infectious events (or 2.50 [1.20-5.39 ]; p=0.016). interestingly severity table 1 (abstract p023). correlation of safety and efficacy markers of thrombolysis and thrombolysis time with distance from stroke centre results expressed as odds ratio with 95% confidence interval of initial bleeding evaluated by fisher scale was not independently associated with dci. dci is independently associated with the occurrence of pneumonia or other cause of sepsis. those results may highlight the need for rigorous approach for prevention protocol, early diagnosis and treatment of hospital acquired infectious diseases in sah patients. introduction: traumatic brain injury (tbi) can have devastating neurological, psychological and social sequelae. increased psychiatric morbidity after tbi has been shown in both adult and the pediatric population. also, critical illness as such is a risk factor for psychiatric problems in youth. our aim was to assess risk factors for later being prescribed psychiatric medication in survivors of intensive care unit (icu)-treated pediatric tbi. we used the finnish intensive care consortium (ficc) database to identify patients 5-17 years of age, treated for tbi in four icu in finland during the years 2003-2013. we examined electronic health records and ct scans and collected data on drug prescription after discharge. we used multivariable logistic regression models to find statistically significant risk factors for psychiatric drug reimbursement. we identified 248 patients of which 46 patients received psychiatric drug prescription (19%) during follow up. the median time to prescription was 14 months after tbi (interquartile range [iqr] 5-31 months). 33 patients received antidepressants, 9 received stimulants and 18 received antipsychotics. increasing age showed a positive association with all drug prescriptions except for stimulants, where an inverse relationship was observed (table 1) . using multivariable analyses, we could not find any admission or treatment related factors that significantly associated with being prescribed psychiatric medications. teenage survivors with moderate disability (glasgow outcome scale [gos] 4) showed high numbers of psychotropic drug utilization (45% received any medication, 36% received antidepressants, 24% received antipsychotics). our data suggests, that the risk of psychotropic drug prescription after tbi depends on factors other than those related to injury severity or treatment measures. the incidence of drug prescription is especially high in patients with moderate disability. the effects of 1-adamantylethyloxy-3-morpholino-2-propanol hydrochloride on the formation of steroid neurotoxicity in rats with brain injury a. semenenko 1 , s. semenenko 2 , a. solomonchuk 3 , n. semenenko 3 depending on the nature of the brain injury and the severity of the victims, mortality in traumatic brain injury (tbi) ranges from 5 to 65% [1] . one of the targets for pathogenetic influence on the course of tbi is the use of pharmacological agents that are able to counteract the negative effects of excess concentrations of glucocorticoids on brain. the therapeutic effect of new pharmacological derivative 1adamantylethyloxy-3-morpholino-2-propanol hydrochloride (ademol) in rats with tbi was evaluated for 8 days. the pseudoperated animals and control group received 0.9% nacl solution and the comparison group received amantadine sulfate. cortisol levels were used to determine the efficacy of the test drugs in tbi. in rats treated with ademol, the level of cortisol in the blood ranged from 179 to 188 ng/ml (p5-p95) and was 2.58-fold lower (p<0.05) compared to control pathology group on the 8 day of therapy. instead, the effect of amantadine sulfate on the level of cortisol in the blood was significantly less than that of ademol. the concentration of cortisol in rats with amantadine sulfate in the blood ranged from 271-280 ng/ml (p5-p95), was 1.73 times lower (p<0.05), compared with the control pathology group, and by 49.2% (p<0.05) exceeded the corresponding value in animals treated with ademol. therapeutic treatment of rats with severe tbi with a solution of ademol, preferably better than rats in the group with 0.9% nacl and amantadine sulfate protect the brain from the formation of steroid neurotoxicity by cortisol (p<0.05). although cerebrovascular pressure reactivity (prx) well correlate to patient's outcome [1] , it requires continuous monitoring and mobile average calculation for its determination. we therefore hypothesized that a simplified model of variation between mean arterial pressure (map) and intracranial pressure icp over the first three days of admission would have been able to predict patient outcome: we call this new parameter cerebrovascular pressure correlation index (cpc). we performed a retrospective observational study of all adult patients with severe tbi admitted to icu from january 2017 to april 2018 inclusive. all consecutive patients with a clinical need for icp monitoring were included for analysis. both for icp and map data were mean value over 2-hours registration, for a total of 12 observations/day, cpc was therefore calculated as the pearson correlation coefficient between icp values (x axis) and map values (y axis), obtaining one single value every 24 hours. variables included in the model (i.e. cpc, cpp, icp, systemic glucose, arterial lactate, paco 2 , icp, and internal body temperature) were collected for the first 3 days since trauma. for the main outcome only the minimum value of cpc fit the regression analysis (p = 0.004). the correspondent roc curve showed an auc of 0.80. the associated youden criterion was ≤0.26 (sensitivity = 0.90; specificity = 0.68). of all the variables considered for the secondary outcome only cpcmin fit the regression model (p =0.03). table 1 reports the median and iqr range for sg and nsg of all the variables considered in the model. this observational study suggests that cpc could be a simplified model of variation between map and intracranial pressure icp over the first three days of admission predicting patient outcome. introduction: impaired cerebrovascular reactivity (car) after traumatic brain injury (tbi) is a marker for disease severity and poor outcome. it is unclear how dynamic changes in body temperature and fever impact car and outcome. we calculated the pressure reactivity index (prx) using the center-tbi high-resolution intensive care unit cohort, as a moving correlation coefficient between intracranial pressure (icp) and mean arterial pressure (map). minute and hourly values of prx and temperature were averaged in patients with simultaneous recording of icp and abp. demographic data was based the core registry (v2.0). linear mixed models were calculated based on minute-by-minute data using r with lme4 v1.1-21 and ggeffects v0.9.0. generalized estimating equation models were used to analyze changes during effervescence (increase of temperature of >1°c within 3 hours). we assessed high frequency physiological data during 567 days of 102 patients admitted to the icu with predominantly a closed injury type (n=94/102). median age was 46 years (iqr 29-62), baseline gcs was 6 (iqr 3-9), and 27% had at least one unreactive pupil. the main measurement site for temperature was the urinary bladder 55/ 102(54%). half of the patients (49/102) developed fever(>1h with mean t ≥ 38.3°c) with a total of 834h fever and a median of 9h fever(iqr 4-24) per patient. of 110 effervescence episodes 30(27%) reached the febrile threshold of 38.3°c which was associated with an increase in prx from 0.09 (±sd 0.25) at baseline (2h before) to 0.26 (±sd 0.3) during the febrile peak (p=0.014) (figure 1-a) . linear mixed models showed a quadratic relationship between prx and temperature (p<0.001) with an increase in predicted prx with febrile and hypothermic temperatures ( figure 1b ). the association of increasing body temperature with worsening of car supports prevention of fever in severe tbi. prospective studies are needed to further differentiate between mechanisms involved (i.e. inflammation) and central autonomic dysregulation. fig. 1 (abstract p030) . the patients with a good 6-month outcome (gose>3) after severe traumatic brain injury showed an increase in root mean square of successive differences between normal heartbeats (rmssd) (compared to baseline 30 -minutes before tracheal succtioning) acute kidney injury (aki) is relatively common in patients with severe traumatic brain injury (stbi) and it can contribute to morbidity and mortality [1] . nephrocheck is a point-of-care urine test that flags two biomarkers that indicate if a critically ill patient is at risk for aki. we investigated the incidence of subclinical aki in patients with stbi. we performed a prospective observational study of all adult patients with severe tbi admitted to icu from january 2017 to april 2018 inclusive. all consecutive patients with a clinical need for icp monitoring were included for analysis. urine samples of severe tbi patients was collected at icu admission from 33 patients to measure nephrocheck (nc) test [igfbp7] x was performed using the nephrocheck® astute1 40 ™ meter. serum creatinine was collected at admission, during the first three days, at icu dismission and 60-days follow up to assess renal recovery. the diagnosis of aki was based on kdigo criteria. hemodynamics, electrolytes, peep, p/f, kind of fluid administered, fluid balance, % fluid overload, length of stay, the sequential organ failure assessment score, injury severity scores and mortality were collected. a total of 15 patients (45%) presented a median nc higher values at icu admission. one patient with positive nc value experienced aki at 24 hrs. the positive nc group had more plasma transfusion (p-value 0.03) and a lower median hematocrit at 24 hrs (p-value 0.013), but similar hospital length of stay (p=0.17) and mortality rate (p=0.80) conclusions: nc at icu admission identifies subclinical aki in tbi patients and it maight be used to predictclinical aki. hemodilution (but not fluid overload) seems to be associated with development of subclinical aki. higher nc at icu admission is not associated with worst longterm outcome in tbi patients. severe traumatic brain injury (tbi) is considered a serious public health problem in europe. partly because of the heterogeneity of tbi, considerable uncertainty may exist in the expected outcome of patients. the international mission for prognosis and analysis of clinical trials in tbi (impact) and the corticosteroid randomization after significant head injury (crash) prediction models are considered the most widely validated prognostic models [1, 2] . however, studies using these prediction models for benchmarking of outcomes have been scarce. we aimed to compare actual outcomes in a tbi cohort of critically ill tbi patients with predicted outcomes in a quality of care initiative in an academic hospital. in this retrospective cohort study, we included consecutively admitted tbi patients to the icu adults of erasmus mc, university medical center, rotterdam, the netherlands between january 2018 and february 2019. we included 87 patients with tbi. 14-day mortality was 25%, sixmonth mortality was 36% and six-month unfavourable outcome was 50%. the impact core+ct+lab model predicted 34% 6-month mortality (vs 35% actual, p=0.89) and 51% unfavourable outcome (vs 50% actual, p=0.9). the 14-day mortality prediction by crash prognosis calculator was 43% versus actual 14-day mortality of only 25% (p=0.01), whereas 6-month unfavourable outcome prediction by crash was 67% (vs. 50% actual, p=0.02) ( figure 1 ). the impact model, although developed more than a decade ago, seemed appropriate for benchmarking purposes in this single center cohort in the netherlands, while crash predictions were less applicable to our setting. introduction: out of hospital cardiac arrest (ohca) continues to be associated with significant mortality and morbidity. centralisation of care has considerably improved patient survival but has resulted in increased morbidity in the form of neurological deficit. accurate neurological prognostication remains challenging incorporating repeated clinical examination and ancillary investigations [1, 2] . data was collected retrospectively and analysed for 96 patients admitted post ohca from october 2018 to october 2019. patient arrest demographics were collected in conjunction with extensive inpatient investigation findings including ct, traditional pupil assessment, pupillometry and eeg. results: 50% of patients survived to hospital discharge. patients presenting in a shockable rhythm continue to have higher survival rates ( table 1) . 53% of patients who received immediate cpr survived to hospital discharge in comparison to 41% of patients who did not receive immediate cpr. 73% of patients underwent non-contrast ct head. 74% of patients had traditional pupillary examination performed on arrival. pupillometry was introduced in december 2018; 31 out of a possible 85 patients had pupillometry during their inpatient stay. eeg was undertaken in 11% of cases. our data shows receiving immediate cpr and presenting with a shockable rhythm remain positive prognostic factors. ct head as a stand-alone prognostic modality is unreliable with 14% of patients who survived to discharge, with intact neurology, had an admission ct head reported as hypoxic brain injury. a new neuroprognostic strategy is required in our unit that adds further certainty to likely clinical outcome. this includes increased use of tests such as eeg and pupillometry and the introduction of biomarkers such as neuron specific enolase, somatosensory evoked potential testing and magnetic resonance imaging. introduction: post-resuscitation care of patients following an out-of-hospital cardiac arrest (oohca) is set out by the uk resuscitation council [1] . this is in line with the european resuscitation council guideline [2] . the aim of this audit was to review compliancy to this guideline at the intensive care unit at the bristol royal infirmary . a retrospective audit was performed over a six-month period in adults who were admitted to the intensive care unit at the bri following an oohca whom later died during that admission (41 patients). the focus was on whether the neuroprognostication and end-of-life (eol) care received was as per the standards set by the uk resuscitation council. the main neuroloical examinations documented were pupillary reflex (100%), corneal reflex (75%) and motor response to pain (100%). 61.5% of patients received an ssep analysis >72 hours post-rosc, 81.5% underwent an eeg and 66.7% had >2 serum neuron-specific enolase measurements recorded. all patients (100%) underwent a ct head during their admission. 5.6% of patients were referred to palliative care during their admission. 22% of patients were prescribed all eol medications. most common prescriptions included alfentanil (90.2%) and midazolam (58.5%). finally, 100% of appropriate patients were referred to be potential organ donors. the audit reflected our local practice and that some parameters were not being maintained as set by uk resuscitation guideline. multiple introduction: the prognostication of neurological outcome in comatose out-ofhospital cardiac arrest (ohca) patients is an integral part of post cardiac arrest care. biochemical biomarkers released from cerebral cells after hypoxic-ischemic injury represent potential tools to increase accuracy in predicting outcome after ohca. currently, only neuronspecific enolase (nse) is recommended in european prognostication guidelines. in this study, we present the release dynamics of gfap and uch-l1 after ohca and evaluate their prognostic performance for long-term neurological outcome in ohca patients. serum gfap and uch-l1 were collected at 24, 48 and 72 h after ohca. the primary outcome was neurological function at 6-month follow-up assessed by cerebral performance category scale (cpc), dichotomized into good (cpc 1-2) and poor (cpc [3] [4] [5] . outcome prognostic performance was investigated with receiver operating characteristics (roc) by calculating the area under the receiver operating curve (auroc) and compared to nse. results: 717 of 819 included patients had at least one serum gfap or uch-l1 value at 24, 48 or 72 h after ohca. gfap and uch-l1 levels were significantly elevated in patients with poor outcome. gfap and uch-l1 discriminated excellently between good and poor neurological outcome at all time-points (auroc gfap 0.88-0.89; uch-l1 0.86-0.87) and overall predictive performance measured by auroc of gfap and uch-l1 was superior to nse (auroc 0.76-0.85) ( figure 1 ). however, the roc at the highest specificities of uch-l1 and gfap overlap those of nse and comparing the sensitivities for uch-l1 and gfap with those of nse for the highest specificities (>95%) revealed higher sensitivities for nse than for uch-l1 and gfap at 48 and 72 h. gfap and uch-l1 predict poor neurological outcome in patients after ohca excellently and with a higher overall accuracy than nse, but both biomarkers perform inferior to nse at specificities over 95% at 48 and 72 h limiting their clinical use to guide decisions on prognosis. blood pressure after cardiac arrest and severity of hypoxicischemic encephalopathy c endisch 1 , s preuß 2 , c storm 3 introduction: blood pressure management in post cardiac arrest (ca) patients ensures sufficient cerebral perfusion to avoid secondary brain injury. in local chain-of-survival improvements affect p-ohca survival [1] [2] [3] [4] [5] . also initial rhythm in p-ohca is an important predictor of survival [1, 4] . little is known about the relationship between initial rhythm in p-ohca and long-term outcome [6] [7] [8] . our aim was to establish the relation between shockable rhythm and favorable long-term outcome in pohca. all children aged 1 day-18 years who experienced non-traumatic ohca between 2002-2017 and were admitted to the sophia children's hospital in rotterdam were included. long-term outcome was determined using a pediatric cerebral performance category score at the longest available follow-up interval. the primary outcome measure was survival with favorable neurologic outcome, defined as pcpc 1-2 or no difference between pre-and postarrest pcpc. the association between shockable rhythm and the primary outcome measure was calculated in a multivariable regression model, adjusted for the pre-defined variables. from the 329 patients included in the 16 year study period 126 (38%) patients survived to hospital discharge of which 99 patients (30%) had favorable neurologic outcome (median follow-up duration of 24 months). the rate of favorable neurologic outcome rose from 17% in 2002 to 52% in 2017 (p < 0.001 for trend) (fig. 1) the odds of favorable neurologic outcome at the longest follow-up duration were significantly higher after a shockable initial and unknown rhythm. secondly, trend analysis showed an increase in aed defibrillation and shorter cpr duration. this was followed, finally, by a rise in rosc, survival to hospital discharge and favorable neurologic outcome rate. low socioeconomic status is associated with worse outcome after cardiac arrest. this study aims to investigate if patients´socioeconomic status impacts the chance to receive early coronary angiography after cardiac arrest. in this nationwide retrospective cohort study, 4011 patients admitted alive after out-of-hospital cardiac arrest (ohca) and registered in the swedish registry for cardiopulmonary resuscitation were included. individual data on income and educational level, prehospital parameters, coronary angiography results and comorbidity were linked from other national registers. in the unadjusted model there was a strong correlation between income level and rate of early coronary angiography where 35% of patients in the highest income quartile received early angiography compared to 15% in the lowest income quartile. when adjusting for confounders (educational level, sex, age, comorbidity and hospital type) there were still higher chance of receiving early coronary angiography with increasing income, or 1.31 (ci 1.01-1.68) and 1.67 (ci 1.29-2.16) for the two highest income quartiles respectively compared to the lowest income quartile. when adding potential mediators to the model (initial rhythm, location, response time, bystander cardiopulmonary resuscitation and if the arrest was witnessed) no difference in early angiography related to income level where found. the main mediator was initial rhythm (figure 1 ). higher income is strongly related to the rate of early coronary angiography after ohca. this finding is consistent when adjusting for known confounders. however, the association between income and early angiography seems to be mediated by initial rhythm. patients with low income more often presents with non-shockable rhythms which lowers the likelihood to undergo early coronary angiography. a. the total amount of mortality as a stacked bar: in light-red the number of patients who deceased at scene, in green the number of patients deceased during admission, in red patients who died after discharge. the grey line is the total number of inclusions. b. the rate of bystander aed use, rate of initial shockable rhythm, rate of less than 15 minutes of cpr and rate of favorable neurologic outcome over time. p for trend significant for bystander aed use, less than 15 minutes of cpr and favorable neurologic outcome. trend analysis performed using binary logistic regression for dichotomous data (and a kruskal-wallis test for non-normally distributed continuous data) effect of simulation teaching of cardiopulmonary resuscitation for nursing v spatenkova 1 introduction: simulation teaching is a modern type of critical care (cc) education. the aim of this study was to assess the effect of simulation teaching of cc on a comparison of final examination in different model levels of cardiopulmonary resuscitation (cpr) after the first (cc1) and third, final cc3. the success rate of cpr was tested in prospective study (2017) (2018) on two groups with a total of 66 students in cc1 and cc3 at the faculty of health studies. three semester of undergraduate nursing simulation education (lectures and training) used the laerdal simman 3g. quality of cpr was evaluated according to 4 parameters: compression depth, compression rate, chest release and time of correct frequency. we tested if cpr quality differed between the two groups. for the compression depth and compression rate parameters, first the conformity of variance was verified and then two-sample t-test. as the chest release and time of correct frequency are recorded as percentages, the wilcoxon rank-sum test was conducted for these parameters. to ensure good resuscitation, all recorded parameters must be properly performed during resuscitation. thus, pivot tables were used to generate statistics and test if the number of correctly performed resuscitation parameters for cc1 and cc3 differ. the compression depth parameter was statistically significantly higher for the cc3 than for the cc1 (p=0.016). there were no differences in compression rate (p=0.210), chest release (p=0.514) and time of correct frequency (p=0.586). it was also tested how many of the parameters were performed correctly by students at cpr. the chi-square test shows the relative frequency of cpr success is higher for the cc3 group than for the cc1 group. at least 3 out of 4 parameters were correctly performed by 13% of cc1 students compared to 28% of cc3 students. the study showed a significant improvement of cpr in the final cc3 and supported the three semester simulation education. changes in blood gases during intraoperative cardiac arrest jj wang, r borgstedt, s rehberg, g jansen protestant hospital of the bethel foundation, anaesthesiology, intensive care and emergency medicine, transfusion medicine and pain therapy, bielefeld, germany critical care 2020, 24(suppl 1):p049 introduction: blood gas analysis (bga) is a common approach for monitoring the homeostasis during surgery. while it is well known that cardiac arrest (ca) leads to circulatory collapse and disturbances in homeostasis, little is known about changes of blood gas during peri-operative ca. we retrospectively analysed patients ≥18 years who suffered from peri-operative ca during non-cardiac surgery from 01/2014 to 12/ 2018. peri-operative ca was defined as need for cardiac compression during anaesthesia care. collected data included ph, paco2, pao2, return of spontaneous circulation (rosc) and 30-day mortality after ca. within the study period, we observed 56 peri-operative ca (m=35, f= 21; age 69±16) during 62742 anaesthesia procedures (rosc occurred in 38 patients (68%). 30 days after ca, the mortality was 45% (n=17), 23% (n=13) were discharged, and 14% (n=8) still in hospital. 87% (n= 49) of ca patients had an invasive blood pressure monitoring, 52% (n=29) had bga before and 66% (n=37) during peri-operative ca. prior to ca, the average values were: ph 7.3±0.1, paco2 39±8 and pao2 225±107. during ca, the average values were ph 7.2±0.2, paco2 50±17 and pao2 215±138. table 1 shows the distributions of blood gas before and during ca. there were no statistical differences between the groups (ph: p=0.4; paco2: p=0.19; pao2: p=0.21). hypercapnia and respiratory acidosis is common in peri-operative ca. these data suggests inadequate ventilation during peri-operative resuscitation. further studies should focus on its impact on the outcome. ]. comparing cases with and without rosc, there were significant more diagnostics done in the group without rosc but more therapeutic consequences seen in the rosc-group (table 1) . icu-ca is frequent. diagnostics to detect reversible causes of ca were used rarely in icu-ca (44%), even in patients without rosc. notably, diagnostics often had therapeutic consequences particularly in rosc. further studies are required to define standardized diagnostic algorithms during icu-ca. continuous monitoring of cardiac patients on general ward were improved short term survival of in-hospital cardiac arrest uj go 1 introduction: the importance of early detection in the in-hospital cardiac arrest (ihca) is emphasized. previous studies have reported that clinical outcomes are improved if ihca is witnessed, or if a patient admitted to a monitored location [1, 2] . this study aimed to evaluate the association between continuous monitoring and survival of ihca on general ward. a retrospective cohort study of ihca in patients admitted to ward at an academic tertiary care hospital between january 2009 and december 2018 was performed. the primary outcome was return of spontaneous circulation (rosc). the secondary outcomes were 72hour survival and survival to hospital discharge. (table 1) . cardiac patients with continuous monitoring on general ward showed improving rosc and 72-hour survival but not survival to hospital discharge in ihca. in-hospital cardiac arrest is associated with poor outcomes. although steroids are frequently used in patients with septic shock, it is unclear whether they are beneficial during cardiac arrest and after return of spontaneous circulation (rosc). of 369 cardiac arrest patients evaluated, 100 were enrolled. advanced life support was conducted according to the2015 resuscitation guidelines. forty-six patients were randomly assigned to receive methylprednisolone 40 mg during resuscitation, and 54 to receive saline (placebo). after resuscitation, steroid-treated patients received hydrocortisone 240 mg daily for up to 7 days, followed by tapering . there was no significant difference between the two groups in scvo 2 andall the secondary outcomes (p>0.05 for all comparisons). the present study found no significant physiologic benefit of corticosteroid administration during and after resuscitation in hospitalized patients with cardiac arrest. the experiences of ems providers taking part in a large randomized trial of airway management during out of hospital cardiac arrest, and the impact on their views and practice. results of a survey and telephone interviews m thomas 1 introduction: the aim is to explore ems experiences of participating in a large trial of airway management during out-of-hospital cardiac arrest (air-ways-2), specifically to explore: 1. any changes in views and practice as a result of trial participation. 2. experiences of trial training. 3 . experiences of enrolling critically unwell patients without consent. 4. barriers and facilitators for out-of-hospital trial participation. an online questionnaire was distributed to 1523 ems providers who participated in the trial. in-depth telephone interviews explored the responses to the online questionnaire. quantitative data were collated and presented using simple descriptive statistics. qualitative data collected during the online survey were analysed using content analysis. an interpretive phenomenological analysis approach was used for analysis of qualitative interview data results: responses to the online questionnaire were received from 33% of airways-2 study paramedics and 19 study paramedics were interviewed. paramedics described barriers and facilitators to trial participation and changes in their views and practice. the results are presented in five distinct themes: research process; changes in views and practice regardingairway management; engagement with research; professional identity; professional competence. conclusions: participation in the airways-2 trial was enjoyable and ems providers valued the training and study support. there was enhanced confidence in airway management as a result of taking part in the trial. study paramedics expressed preference for the method of airway management to which they had been randomized. there was support for the stepwise approach to airway management, but also concern regarding the potential to lose tracheal intubation from 'standard' paramedic practice. causes of medical care-associated cardiac arrest on the intensive care unit s entz 1 introduction: cardiac arrest on intensive care unit (icuca) following therapeutic interventions is of imminent importance, because the interventions are comparatively predictable and precautions can potentially be taken. this study investigates medical care associated complications that led to icuca. intensive care database was screened for patients ≥18 years who experienced icuca in a tertiary hospital with five icu (two medical, two surgical, one interdisciplinary, with a sum of 71 icu beds) in germany from 2016-2018. icuca was defined as receiving chest compression and/or defibrillation after admission on icu and classified as "medical care associated" if it was preceded by a therapeutic intervention (i.e. induced by medication, bedding procedures, iatrogenic injuries, procedure associated). subgroups included patients with recurrence of spontaneous circulation (rosc) vs. no-rosc and patients with vs. without vasopressor therapy before intervention. there were 125 icuca in 114 patients of totally 14,264 icu patients. medical care associated complications leading to icuca were detected in 28 cases (22%) [incidence 19.6/10,000(ci95 12.3 -26.9)]. icuca following therapeutic interventions occurred because of circulatory insufficiency [n=20(70%)], respiratory failure [n=5(17%)] and airway associated problems [n=3(10%)]. nine of the 28 patients (32%) with care-associated icuca died. table 1 demonstrates therapeutic interventions followed by icuca. care-associated complications were common reasons for icuca. most of events were induced by circulatory insufficiency due to induction of anaesthesia and bedding procedures. further investigations should focus on preventive strategies, such as vasopressor infusion before therapeutic interventions. in-hospital cardiac arrest (ihca) is a lethal event. however, ihca has received less attention than out-of-hospital cardiac arrest (ohca). there have been some studies on ihca; however, there is a lack of information on the evidence and clinical features of ihca compared with information for ohca. we therefore conducted this study to clarify important aspects of the epidemiology and prognosis of ihca in patients with code blue activation. we carried out a retrospective observational study of patients with code blue events in our hospital during the period from january 2010 to october 2019. we obtained information on the characteristics of patients including age and gender, ihca characteristics including the time of cardiac arrest, event being witnessed, presence of bystander cardiopulmonary resuscitation (cpr), initial shockable rhythm, vital signs 1 h or 6 h before cardiac arrest, survival to hospital discharge (shd), and the cardiac arrest survival postresuscitation in-hospital (caspri) score. the primary endpoint was shd. we performed univariate and multivariate logistic regression analyses. a total of 293 code blue events were activated during the study period. finally, 81 patients were included in this study. overall, the shd rate was 28.4%. the median time of cpr was 14 min (interquartile range, 6-28 min). the rate of initial shockable rhythm was 19.8%. there were significant differences in cpr duration, shockable rhythm, and caspri score between the shd group and non-shd group by univariate-logistic regression analysis. caspri score was found to be the most effective predictive factor for shd (or=0.98, p=0.006) by multivariate-logistic regression analysis. our results demonstrated that caspri score is associated with shd in cpa patients with in-hospital code blue events. caspri score in ihca patients would be a simple and useful adjunctive tool for management of post-cardiac arrest syndrome (pcas). peri-operative cardiac arrest in prematurityincidence and causes at a tertiary care hospital between 2008-2018 g jansen, j popp, e lang, r borgstedt, b schmidt, s rehberg protestand hospital of the bethel foundation, anaesthesiology, intensive care and emergency medicine, bielefeld, germany critical care 2020, 24(suppl 1):p058 the peri-operative care of premature pediatric patients requires special expertise and is therefore reserved for specialized centers. although premature birth is described as a risk factor for peri-operative complications and cardiac arrest (poca) there are no data on its incidence and causality in this particular population [1] . the present study investigates the incidence and causality of pediatric poca at a tertiary care hospital and level i perinatal center in germany. in the anesthesia database of the study center, all anaesthesiological procedures in patients <16 years of age were examined for poca in preterm infants (gestational age <40th week of gestational age) between 2008 and 2018. the peri-operative period was defined between the beginning of anesthesiological care up to 60 minutes after anesthesia and/or sedation. we defined cardiac arrest as the necessity of chest compressions. the perioperative phase and the cause of the poca, gestational age and birth weight were recorded. between 2008 and 2018, 308 (1.3%) of the 22,650 pediatric anesthesiological procedures were performed on 301 premature infants. in total, 10 poca occurred in 9 of these patients (f=6, m=3; average gestional age 208±27 days; average birth weight 1510±747g (incidence 3.2%, ci 96 1.3-5.2%). the time of occurrence and the causes of poca are shown in table 1 . poca in premature babies is rare and has an incidence of 1.3%, which is significantly higher than the non-premature babies. the main causes are problems or complications associated with the respiratory tract and its management, as well as massive hemorrhage. introduction: peri-operative cardiac arrest (poca) in children's anesthesia care is a dreaded event. depending on the country and population, studies describe incidences between 2.9-20.6 per 10,000 children's anesthetics. there are no data on the current incidence of pediatric poca in germany. the present study investigates the incidence of poca at a tertiary hospital and level i perinatal center in germany. in the anesthesia database of the study center, all anaesthesiological procedures in patients <16 years were examined for poca. the peri-operative period was defined between the beginning of anesthesia care up to 60 minutes after anesthesia or sedation. cardiac arrest was defined as the necessity of chest compressions. age, weight, asa status, cause of death and survival after 30 days were recorded. results: 18 poca (median weight was 2525g [q1 7151;q75(14748)]) were observed in 22,650 anaesthesiological procedures (incidence 7.9±4.2 per 10,000 [ci95 4.3-11.6]). table 1 shows the distribution of the individual age groups, incidences and mortalities of poca. peri-operative 30-day mortality was 3 per 10,000 [ci 95 [1] [2] [3] [4] [5] . three children died intraoperatively as a result of hemorrhagic shock, one on the picu as a result of malignant hyperthermia. 30 days after poca, 4 more children had died on the icu due to their underlying disease. poca is a rare event. risk factors are an age <28 days and an asa status ≥ iii. the main cause of peri-operative death in patients <16 years of age is massive hemorrhage, the 30-day mortality is determined by the underlying disease. in-hospital cardiac arrest -predicting adverse outcomes t partington, j borkowski, j gross northwick park hospital, anaesthesia/critical care, london, united kingdom critical care 2020, 24(suppl 1):p060 introduction: cardiac arrest occurs in 1.2 per 1000 hospital admissions in the uk. return of spontaneous circulation (rosc) is achieved in approximately half of resuscitation attempts, but rate of survival to hospital discharge is substantially lower [1] . in our centre, post-arrest care accounts for 6.4% of icu admissions. premorbid social function is purported to affect outcomes, but comorbidity scores are more often used for risk stratification. using a novel social function score alongside an existing comorbidity scale, we aimed to identify trends to inform management of patients at risk of deterioration. a six-month prospective observational study was conducted in a major uk hospital from october 2017 to april 2018. for all adult inpatient cardiac arrests, medical notes were reviewed and data collected on the following domains: patient demographics comorbidities and functional status admission details post-arrest events statistical analysis was performed using student's unpaired t-test. results: 54 cardiac arrests occurred. 85% were in medical patients, with the majority male (63%) and aged over 75 (63%). 89% were emergency admissions, with mean duration of hospital stay pre-arrest 9 days. in 17 cases (31%) sustained rosc was achieved. however, seven of these (41%) were not subsequently admitted to the icu. only six patients (11%) survived to hospital discharge. pre-admission function and comorbidity were worse in patients who did not survive to discharge ( fig. 1 ), but these were not statistically significant in view of small survivor group size. in an increasingly frail inpatient population, a substantial proportion of patients in whom circulation is restored after cardiac arrest are subsequently considered unsuitable for icu admission. given our understanding of inferior outcomes in patients with poor physiological reserve, we encourage early discussion regarding the appropriateness of cpr in selected patients, guided by social function and comorbidity. references: 1. national cardiac arrest audit 2017/18 introduction: there are studies that determine events related to poor outcome in cardiac arrest [1] . in our study, following parametres were determined ohca patients; age median years, asian/europe/syrian, bystander cpr, bystander aed, ems defibrillation, initial cardiac rhythm, prehospital rosc, corneal and pupillary light reflex and day survival. we determineted poor prognostic sign with post-cardiac arrest patients. in this study, we identified the causes of poor outcome in patients with ohca. this was a single-centre, retrospective study. we determined incidence and epidemiological factors including: demographics, initial cardiac rhythm. our study population were non-traumatic ohca. our icu, all ohca patient were evaluated wtih echo, and fluid, inotrope and vazopressor were added according to cardiac performance. results: during our study, 5970 patients who were admitted to intensive care unit between 2012-2019 were screened. 133 of these patients were out-of-hospital arrest and 41 of them were in-hospital arrest. development of cerebral oedema during treatment in hospital remains a poor prognostic sign. the evaluation of initial cardiac ritm is useful to predict neurological outcome in post-cardiac arrest patients. survival after ohca remains low. the evaluation of initial cardiac ritm is useful to predict mortality and neurological outcome in postcardiac arrest patients. basic life support (bls) education and training for school children is active in japan. however, the bls action by schoolchildren may be limited by school rules. this study aimed to analyse the time factors for basic life support performance and outcome in classmatewitnessed out-of-hospital cardiac arrest (ohca) and to investigate how schoolchildren act when they detect ohca. methods: nation-wide database for 1,068 school children cases with ohca and local extended database for 5,478 ems-unwitnessed ohca, both of which were prospectively collected during the period of 2011-2016, were retrospectively analysed. proportion of schoolchildren-detected ohca was low in classmate cases (16.8%, 179/889) in nationwide database and extremely low in all ems-unwitnessed ohcas (1.6%, 88/5,478) in local database. nationwide database analyses revealed that both emergency call and bystander cpr were delayed when a classmate witnessed the ohca case: median, 1 vs. 0 min and 3 vs. 2 min, respectively. classmate-witnessed cases were associated with higher incidences of shockable initial rhythm, aed use and traumatic causes. the rate of neurologically favourable outcome was 19.6% and 12.3%, respectively in classmate-witnessed and other cases: adjusted or; 99% ci, 1.24; 0.63-2.47. of 88 cases detected by schoolchildren in our prefecture, 8 (34%) cases had presumed cardiac aertiology and 12(13.8%) cases were caused by suicide attempts (hanging and fall). school children placed emergency 119 calls as the first action only in 32 (36.4%) cases. emergency calls were largely delayed when school children dialled other numbers or left the scene to seek adult help. school children were rarely involved in bystander cpr (21%) and aed placement (1%). school children are rarely involved in entire bls. emergency calls and bystander cpr are delayed when schoolchildren act to seek help. because schoolchildren detect suicide-related ohcas, psychological care to schoolchildren involved in bls may be necessary. prognostic value of neutrophil/lymphocyte and platelet/ lymphocyte predicting cardiopulmonary resuscitation with spontaneous circulation recovery c li the affiliated suzhou hospital of nanjing medical university, suzhou, china critical care 2020, 24(suppl 1):p063 to investigate the predictive value of peripheral blood neutrophil-tolymphocyte ratio (nlr) and platelet-to-lymphocyte ratio (plr) on inhospital mortality in patients with spontaneous circulation recovery after cardiac arrest. a retrospective analysis was made of 30 patients who recovered from cardiac arrest in our hospital from april 2012 to november 2018 and were admitted to the intensive care unit for more than 24 hours. they were divided into survival group and death group according to the outcome of discharge.the dynamic changes and differences of nlr and plr in 24 hours and 48-72 hours after admission to icu between the two groups were analyzed and compared. multivariate analysis and roc curve were used to explore the predictive value of nlr and plr for in-patient mortality. compared with the survival group, plr in the dead group was significantly lower within 24 hours of admission to the intensive care department (p < 0.05), while nlr in 48-72 hours was significantly higher (p < 0.05). the nlr of surviving group was significantly lower than that of 24 hours (p < 0.05), while the nlr and plr of death group were not significantly different (p < 0.05) from that of 24 hours (p < 0.05). multivariate logistic regression analysis and roc curve showed that nlr of 48-72 h in icu was an independent risk factor for predicting in-patient mortality, and had high sensitivity and specificity in predicting death outcomes. neutrophil to lymphocyte ratio, platelet to lymphocyte ratio can help to judge the outcome of patients with cardiac arrest and recovery of autonomic circulation after cardiopulmonary resuscitation. [2, 3] patients with sofa score > 12 (vs sofa score ≤ 12) had a higher free iron level (35.5 μmol/l vs 16 μmol/l, p = 0.0333) ( figure 1 ). we found a positive correlation between free iron level at h0 and changes of sofa score between h0 and h48 (r= 0.56 ic95[0.08;0.76]). out-of-hospital cardiac arrest is associated with a significant change of plasma free iron level. free iron level at admission is associated with short term outcome. further research is warranted to better determine the significance of such changes. the optimal level of arterial oxygen in the post-resuscitation period is unknown. recent studies show conflicting results in regard to hyperoxia and its association with survival after out-of-hospital cardiac arrest (ohca) [1] . the aim of this trial is to study the association between early hyperoxia after ohca with return of spontaneous circulation (rosc) and 30-day survival. observational study using data from three swedish national registers (i.e. intensive care, cardiac arrest and national patient registries after a successful resuscitation, a systemic inflammatory response occurs, and the c-reactive protein (crp) level represents the degree of inflammation [1] [2] [3] . this study examined the association between increased inflammation and early-onset pneumonia (eop) in patients treated with extracorporeal cardiopulmonary resuscitation (ecpr) after out-of-hospital cardiac arrest (ohca). this retrospective study included data of patients with ohca treated with ecpr admitted to st. luke's international hospital between april 2006 and april 2019. the exclusion criteria were as follows: age < 18 years, therapeutic hypothermia withdrawal due to death or circulatory failure, or sepsis as a suspected cause of cardiac arrest. patients were diagnosed with eop according to clinical signs and symptoms acquired after a hospitalization period of >48 h and within 7 days of admission. the crp levels were measured daily from admission to day 3. we studied 55 patients with a median age of 55 years (interquartile range: 42-65 years). furthermore, 52 (95%) patients were males, and the median time interval from collapse to adequate flow was 51 (42-63) min. all patients received prophylactic antibiotics, and 18 (33%) of them had favorable neurological outcomes (cpc, 1-2). eop occurred in 32 (58%) patients, with a significantly higher crp level on day 3 than that in those without eop (13. 9 categorizing reasons for death after ecpr is important for comparing outcomes to other studies, assessing benefits of interventions, and better define this heterogeneous patient collective. a categorizing for death after cardiac arrest in both in-hospital (ihca) and outof-hospital (ohca) arrests has been proposed in non-ecpr patients by witten et al. here, we adopt this categorization to ecpr patients. single-center, retrospective, cohort study of patients without rosc after ihca or ohca and ecpr between 2010 and 2017. patients with survival below 24 hours were excluded. patients were allocated to one of five predefined reasons for death. results: 231 va-ecmo patients were included (age 58.6±14.3, 29.9% female, 58% ecpr, 30 day survival 42.9%). reasons for death for patients with va-ecmo for shock (survival 53%) and ecpr (36%) were: neurological withdrawal of care (10% vs 25%), comorbid withdrawal of care (18% vs 4%), refractory hemodynamic shock (16% vs 33%), respiratory failure (3% vs 2%), and withdrawal due to presumed patient will (0% vs 1%) ( figure 1 ). the differences in reasons for death among the two groups were significant (p <0.001), driven by withdrawal due to neuroprognostication, comorbidity and hemodynamic instability. categorizing death after va-ecmo into five categories is feasible. there are significant difference between patients with va-ecmo for shock and ecpr. interestingly, only a quarter of patients after ecpr died due to brain damage. introduction: scarcity of potential dead brain donors and the persistent mismatch between supply and demand of organs for transplantation has led the transplant community to reconsider donation after circulatory death (dcd) as a strategy to increase the donor pool. normothermic regional perfusion (nrp) by extracorporeal membrane oxygenation (ecmo) may be the most effective method for preserving abdominal organs in dcd, especially in liver transplantation [1, 2] . a pitfall of this method is its complexity and the unavailability of this resource in some hospitals, especially in regional hospitals, where potential dcd donors may exist. aim of this study is to report the use of mobile ecmo team in controlled dcd. from june 2018 to november 2019 our group has worked as a mobile ecmo team for cdcd outside our center. portable equipment included cannulation material and the ecmo device. the transplant team consisted of 1 transplant coordinator (anesthesiologist-intensivist, ecmo operator and organ extraction supervisor), 1 cardiac surgeon (cannulation), 1 interventional radiologist (cannulation) and one cardiovascular perfusionist (ecmo operator). twenty-five cdcd donations were performed. characteristics of donors and organs retrieved are summarized in figure 1 . from 25 cdcd, 17 livers, 4 lungs, 45 kidneys were obtained. the evolution of grafts and receptors was favorable at day 30 post-transplant. mobile ecmo teams may enable cdcd in hospitals without these resources, thereby increasing the pool of donors and optimizing graft outcomes. what is the useful coagulation and fibrinolysis marker for predicting extracorporeal membrane oxygenation circuit exchange due to intra-circuit thrombus? y izutani, k hoshino, s morimoto, k muranishi, j maruyama, y irie, y kawano, h ishikura fukuoka university hospital, emergency and critical care center, fukuoka-shi, japan critical care 2020, 24(suppl 1):p071 a thrombus formation is one of the most frequent and adverse complications during extracorporeal membrane oxygenation (ecmo) support. previous studies have reported that increased d-dimer is a useful predictor of thrombus formation within the ecmo circuit. the purpose of this study was to identify coagulation/fibrinolysis markers for predicting the replacement of ecmo circuit due to intra-circuit thrombus during ecmo support. fourteen patients who underwent veno-venous ecmo for acute respiratory failure between january 2014 and december 2018 were enrolled. these patients received a total of 125 days of ecmo support. of these, 9 days (times) on which the ecmo circuits were replaced was regarded as the replacement group, while the remaining 116 days were considered as the non-replacement group. the several coagulation/fibrinolysis markers were routinely measured every day during ecmo support. we compared with the levels of these markers between two group to identify the most relevant marker for ecmo circuit replacement due to thrombus. the mean duration of ecmo support was 9±11 days, and the mean number of ecmo circuit replacement was 0.6±1.0 times per patient. ddimer, thrombin-antithrombin complex (tat), plasmin-α2 plasmin inhibitor complex (pic), and soluble fibrin (sf) were significantly higher in the replacement group rather than in the non-replacement group (p < 0.01, respectively). according to a multivariate analysis, sf was the only independent predictor of ecmo circuit replacement due to thrombus. the odds ratio (95% confidence intervals) for sf (10 μg/ml) was 1.2 (1.1-1.3). the area under the curve and optimal cut-off value were 0.94 and 85 ng/ml for sf, respectively (sensitivity, 100%; specificity, 85%). from these results, we concluded that sf may be the useful marker rather than d-dimer for predicting the replacement of ecmo circuit due to intra-circuit thrombosis. inhomogeneity of lung elastance in patients who underwent venovenous extra corporeal membrane oxygenation (v-v ecmo)-a computed tomography scan study rd di mussi 1 , ri iannuzziello 2 , fm murgolo 2 , fd de carlo 2 , e caricola 2 , na barrett 3 , lc camporota 3 , sg grasso 2 1 università degli studi di bari "aldo moro", department of emergencies and organ transplant, bari, italy; 2 università degli studi di bari "aldo moro", bari, italy; 3 department of adult critical care, guy´s and st thomas´nhs foundation trust, king´s health partners, london, uk critical care 2020, 24(suppl 1):p072 in patients with acute respiratory distress syndrome (ards), nonaerated, poorly aerated, and normally aerated regions coexist to variable degrees in lung parenchyma. the recruitment maneuvers aim to reopen collapsed lung tissue. in a theoretical point view, this strategy may also prevent the normal aerated lung tissue hyperinflation [1] . the objective of our study was to evaluate lung characteristics in terms of hounsfield units (hu), volume and elastance before and after a recruitment maneuver. in 37 patients with severe ards who underwent v-v ecmo, computed tomography scans (ct-scans) at 5 cmh 2 o of continuous positive airway pressure (cpap) and 45 cmh 2 o were performed. the same ct image was selected at the two different levels of pressure. the distribution of lung opacities, in terms of hu, was classified using the "ucla" colour coding table (osirix image processing software, geneva, switzerland). correspondent lung regions of about 1020 voxels were selected. the quantitative analysis, in terms of volume air (vair) was performed with maluna software (version 3.17; maluna, goettingen, germany). elastance was calculated as the pressure(cmh 2 o)/ vair (ml) ratio. results: see figure 1 . lung inhomogeneity occurs also after recruiting maneuvers. our data confirm that the elastance of recruited lung regions is higher than the elastance of the normal aerated lung regions at low positive end-expiratory pressure (peep) (baby lung). on the contrary the "baby lung" frequently develops hyperinflation. the unpredictable pattern of distribution of volume after recruitment maneuverers may explain the controversial role of peep during the ards treatment. . formal recommendations on target, timing, and rate of at supplementation are lacking. we conceived this study to evaluate the effect of prolonged at supplementation in adult patients requiring veno-venous ecmo for respiratory failure on heparin dose, adequacy of anticoagulation and safety methods: before ecmo start patients were randomized to either receive at supplementation to maintain a functional at level between 80 and 120% (at supplementation group) or not (control group) for the entire ecmo course. anticoagulation was provided with unfractionated heparin following a standardized protocol [1] . the primary outcome was the dose of heparin required to maintain the ratio of activated partial thromboplastin time between 1.5 and 2. secondary outcomes were the adequacy of anticoagulation measured with anti-factor xa and the incidence of hemorrhagic and thrombotic complications and amount of blood products fig. 1 b) . conclusions: this retrospective analysis was not able to show a survival benefit for additive pp to ecmo support in general. early initiation of pp could be an important factor for improving survival in this setting and should be considered in a randomized controlled trial for further evaluation. cause-specific mortality during extracorporeal membrane oxygenation, a single center review of medical records m panigada, d tubiolo, p properzi, g grasselli, a pesenti fondazione irccs ca´granda ospedale maggiore policlinico, intensive care unit, milano, italy critical care 2020, 24(suppl 1):p075 introduction: mortality during extracorporeal membrane oxygenation (ecmo) settles around 35% and the occurrence of bleeding during ecmo is associated with a high mortality rate. however, cause-specific mortality is rarely reported, probably due to the difficulty of its classification. the purpose of the study was to evaluate the agreement between two expert icu physician in the classification of the cause of death of patients supported with ecmo for either respiratory or cardiac support. methods: two intensive care unit (icu) expert staff physicians independently reviewed the entire medical records of all ecmo patients who died before icu discharge from january 2011 to september 2019 at fondazione irccs ca' granda, milan. they were asked to choose the cause of patient's death among six categories. in case of disagreement, a third expert adjudicated the case. the two reviewers were also asked whether, in their opinion, bleeding during the last 24 hours contributed to death. elso definition of major bleeding [1] during the last 24 hours was also recorded for each patient. results: two-hundred and two patients were supported with ecmo of whom 70 (34.6%) died. most of these patients (n=53, 75.7%) died during ecmo. interrater agreement for cause-specific mortality between the two expert physicians was substantial (k 0.71, se 0.07, p<0.01) of the 14 discordant cases 6 were categorized as refractory respiratory failure and 4 as multiorgan failure and septic shock respectively. the distribution of cause-specific mortality is shown in figure 1 . major bleeding (elso) was present in 27 (38.6%) patients, only in 9 (33.3%) of them bleeding contributed to death according to the reviewers. patients treated with early pp while ecmo showed a superior survival to patients treated with late pp or without pp while ecmo. optimal cut off value for duration of ecmo initiation to first pp was calculated using roc-analysis (auc = 0.789) and the youden-index. highest sensitivity and specificity for beneficial survival were achieved for a beginning of pp in <0.71 days. (log rank=0.018). pp: prone positioning p076 non-invasive mechanical ventilation in veno-venous extracorporeal membrane oxygenation j rilinger, v zotzmann, x bemtgen, pm biever, d duerschmied, c bode, dl staudacher, t wengenmayer heart center freiburg university, department of cardiology and angiology i, freiburg, germany critical care 2020, 24(suppl 1):p076 introduction: veno-venous extracorporeal membrane oxygenation (ecmo) support can be combined with a variety of different non-invasive ways to deliver oxygen to the patient's lung. several positive effects might be linked to this so called "awake ecmo". so far there is little evidence about indications and outcome of this approach. we report retrospective registry data on all ards patients treated with ecmo support at a university hospital between 10/2010 and 04/ 2019. in a systematic review of medical records, we distinguished between patients with invasive mechanical ventilation (imv) from the initiation of ecmo therapy (imv group) and patients that received any kind of non-invasive oxygen supply (non-imv group). a total of 276 patients could be analysed. 16 (5.8%) patients received non-imv ecmo support. patients receiving non-imv ecmo therapy showed severe underlying pulmonary disease and immunosuppression (fig. 1) . these patients had higher rates of lung fibrosis, long-term oxygen therapy, pulmonary hypertension, renal insufficiency and immunosuppression (p<0.05). 12 of 16 patients (75%) required imv during the hospital stay in average 5.3±5.0 [0.8-17.1] days after ecmo initiation. reasons were hypoxia despite of ecmo, insufficient ecmo-flow, insufficient protective reflexes or patient agitation. patients with initially non-imv ecmo support showed a numerical but not significant lower icu and hospital survival (25.0% vs. 45.4%, p= 0.111). non-imv ecmo support was applied in patients with severe underlying pulmonary disease and/or immunosuppression. in a high proportion of patients the ventilation regime had to be switched from non-invasive to invasive. survival in this very selected cohort was low. in this retrospective analysis no evident benefit for a noninvasive ventilation strategy could be found. the high proportion of patients who switched from non-imv to imv therapy underlines the need for rigorous patient selection. intra-hospital transportation on extracorporeal membrane oxygenation (ecmo) -a single centre experience in ireland. z siddique, s o´brien, e carton, i conrick-martin mater misericordiae university hospital, department of critical care medicine, dublin, ireland critical care 2020, 24(suppl 1):p077 the objective of this study is to evaluate intra-hospital transportation of patients on extracorporeal membrane oxygenation (ecmo). it is a retrospective analysis of prospectively collected database, performed as part of ongoing quality improvement initiatives. the setting of this study is an 18-bed, combined surgical and medical adult intensive care unit (icu) located in a 570-bed hospital that serves as the national referral centre for cardiothoracic surgery, heart & lung transplantation and ecmo in ireland. we reviewed 33 months of data (from 2017 to 2019) regarding patients admitted to our critical care unit who required intra-hospital transfer for diagnostic and/or therapeutic interventions. we also compared the data to available local guidelines. results: 23 patients were transported on ecmo on a total of 28 occasions; the most common indication being ct brain (table 1) . ecmo cannulation sites were peripheral in 20 patients, 3 patients were centrally cannulated. median time from start of the transfer until the patient was returned to icu was 50 minutes (range: 35-195). the ecmo console was placed on a dedicated ecmo trolley apart from two occasions where it was placed on the patient's bed. number of staff required for transport was between 4 to 10; with an icu consultant as team leader. ecmo specialist nurses were always present on the transport team. 27 transfers were during normal working hours with 1 happening on a weekend. a total of 12 complications occurred during the transports, of underlying pulmonary disease or status of immunosuppression in ecmo patients without invasive mechanical ventilation which 1 was significant and 11 were not. the significant complication encountered was ventricular tachycardia in a v-a ecmo patient which required electrical defibrillation. no adverse events related to transport were seen following return to icu. in this single-centre study, we have demonstrated safe intra-hospital transport of ecmo patients. the use of local guidelines, appropriate personnel and performance during normal working hours is recommended. a novel approach for flow simulation in ecmo rotary blood pumps a supady 1 , c benk 2 , j cornelis 3 , c bode 1 , d duerschmied 1 1 heart center freiburg university, cardiology and angiogiology i, freiburg, germany; 2 heart center freiburg university, department of cardiovascular surgery, freiburg, germany; 3 fifty2 technology gmbh, 79108 freiburg, germany critical care 2020, 24(suppl 1):p078 introduction: extracorporeal membrane oxygenation (ecmo) is used increasingly in critically ill patients suffering from acute respiratory failure, cardiogenic shock or cardiac arrest. however, this therapy can have deleterious side effects such as bleeding or clotting complications and hemolysis. these complications are particularly caused by physical stress acting upon the blood components while passing through the ecmo system, especially within the rotary pump. we here present a novel approach to simulate blood flows through rotary blood pumps used in current ecmo systems in order to better understand the genesis of these complications. geometries of the xenios dp3 (xenios ag, heilbronn, germany) rotary pump were reconstructed by ct-scans and manual measurements using computer-aided design (cad). the computational fluid dynamics (cfd) simulation was performed using the software preon-lab (fifty2 technology gmbh, freiburg, germany), which implements a mesh-free lagrangian method requiring minimal preprocessing of the cad data. the geometries are introduced to the simulation model as tessellated surfaces. five operating points have been specified by the rotation of the centrifugal fan and the corresponding inflow and outflow of blood. the blood is approximatively modelled as a newtonian fluid with a density of 1040 kg/m 3 . preonlab allows detailed assessment of the blood flow while passing through the rotary pump including analysis of local flow rates, pressure gradients and shear stress acting upon the blood. dead zones in the fluid flow can be detected which gives reference points for optimizations of the pump design. for the first time, we demonstrate a novel approach for flow simulation in an ecmo rotary pump ( figure 1 ). this approach may help better understand hemodynamics within the extracorporeal system to define optimal operating points or re-design components aiming to limit hemolysis, coagulation disorders and bleeding in seriously ill patients. one-year experience of bedside percutaneous va-ecmo decannulation in a territory ecmo center in hong kong km fong, sy au, pw leung, kc shek, hj yuen, sk yung, hl wu, so so, wy ng, kh leung queen elizabeth hospital, intensive care unit, hong kong critical care 2020, 24(suppl 1):p079 when veno-arterial extra-corporeal membrane oxygenation (va-ecmo) support can be terminated, arteriotomy wounds of the patients of are traditionally closed by open repair in the operation theaters. lots of manpower are involved and timeslots in operating theaters are scarce. transport of the critically-ill is risky. successful va-ecmo decannulation using percutaneous device called proglide has been reported and our group had adopted and modified this approach [1] . methods: this is a retrospective study analyzing the one-year experience of bedside va-ecmo decannulation. our institution is a 23-bed tertiary ecmo referral center in hong kong. our first bedside decannulation was performed in november 2018, and since then, this practice had replaced the traditional open repair, unless contraindicated. data from november 2018 to october 2019 were analyzed. in the study period, 39 patients received va-ecmo. 28 survived to decannulation and 25 received bedside percutaneous decannulation. their median age was 59 (52-67). the default arterial catheter size was 17fr, with 15 fr in 3 cases and 19fr in one. five (20%) failed percutaneous closure and they were subsequently surgically repaired without extra corporeal life support (ecls) continues to be associated with high mortality rates. our ability to predict outcome prior to initiation ecls remains limited. here we take a single cell rnaseq approach in an effort to identify novel immune cell types that are associated with-and may contribute to-survival on ecls. whole genome transcriptomic profiles were generated from~40,000 peripheral blood monocytes obtained from 38 patients at the time of cannulation for veno-arterial ecls (va-ecls). within each subpopulation, differential gene expression analysis was performed to identify new markers associated with survival. findings were validated in a additional cohorts by flow cytometry. surviving patients had significantly higher proportions of cd8 + nkt cells (cd3 + /cd8 + /cd19 -/cd56 + ) that were cd52 + (p = 0.001, fdr < 0.05) ( figure 1 ). to validate this observation, we performed fc analysis of a second cohort of 20 patients. for each patient, we quantified the proportion of cd8 + nkt cells that were cd52 + . using the median proportion as the cutoff, we again found that a high proportion of cd52 + cells among cd8 + nkt cells was predictive of 48 hour survival (p=0.024). we noted that while high levels of cd52+ cells among the cd8+ nkt cells was protective in this cohort of va-ecls patients, this relationship did not hold for patients with sepsis. as only a few the va-ecls patients were septic, we analyzed a third cohort of septic ecls patients. we observed that high levels of cd52+ cells among the cd8+ nkt populations was not protective in this population. the proportion of cd8+ nkt cells that are positive for cd52 is predictive of survival among patients undergoing va-ecls for noninfection related indications. introduction: the use of calcium sensitizers has grown enormously in the last decade, probably due to their interesting pharmacodynamic properties. levosimendan (ls) is frequently administered in patients under mechanical circulatory support. we performed a retrospective evaluation of patients treated with ls prior to weaning from mechanical support. this evaluation was combined with a review of the literature. a query of our icu patient data management system revealed 22 patients receiving ls prior to or during vad/ecls support. outcome data were obtained from the patients medical records. of our 22 patients, 78% was successfully weaned off ecls. fourteen patients (63 %) died before being discharged of whom 5 while on ecls support. of the weaned patients, 9 died afterwards. 4 of the converted patients needed subsequent veno-venous ecls support for right ventricular support after the implantation. survival to discharge ratio for the whole group was 31 %. more detailed demographic results can be found in table 1 . a pubmed search using the terms "(ecmo or ecls) and ls and weaning" resulted in 7 publications which dealt specifically with weaning of ecls support. several weaning approaches are available, however poor outcome has remains a problem. some recent studies show a possible beneficial effect of ls infusion prior to weaning from ecls. however most of these studies are retrospective or observational at best. because ls is primarily reserved for the most severe cases, outcome interpretation is difficult. overall weaning success ranges from 82%-92% and variation is very dependant of inclusion criteria. the calcium sensitizer ls can be used when weaning off patients from ecls, certainly given its low incidence of complications. future, large randomized trials are however needed in order to confirm this strategy. cardiogenic shock is well described in newly diagnosed pheochromocytoma, and crisis may be precipitated by hemorrhage into tumour. v-a ecmo represents a rescue therapy in a subset of these patients refractory to medical management, facilitating cardiac recovery and subsequent definitive surgery. consent to publish: written informed consent for publication was obtained from the 2 patients. during a spontaneous breathing trial respiratory mechanics can worsen, and respiratory muscle effort can increase, leading to respiratory muscle fatigue, pump failure, hypercapnia and an unsuccessful weaning from mechanical ventilation. this case report discusses the possibility of applying extracorporeal co 2 removal (ecco 2 r) to reduce respiratory muscle effort in a liver transplant recipient who already failed three weaning attempts from mechanical ventilation. the ecco 2 r membrane lung was integrated into a conventional renal replacement therapy circuit and blood flow was increased from 150 to 300 ml/min. measurements of respiratory mechanics (including esophageal pressure, as shown in fig. 1 ) were used to assess the reduction of respiratory effort before and during the application of ecco 2 r. was delivered through a 13fr-double-lumen-cannula; 350 ml/min blood-flow with 10lt oxygen sweep-gas-flow and aptt 1.5-2 baseline were maintained (iv-heparin). in all cases respiratory and metabolic parameters improved without complications ( figure 1 ). ecco2r-crrt facilitated extubation (4 out 9 imv pts). in 4 out of 5 pts at risk of niv failure, it avoided imv. treatment mean duration was 73±31 hours, mean lenght of icu stay was 6±4 days. all patients survived to the treatment, nevertheless 2 patients died due to irreversible multiple mof. in our aecopd series prismalung®-prismaflex® facilitated weaning from imv and avoided intubation in patients at risk of niv failure without complications. these positive results may be related to minimal invasiveness of the low-flow device used and may constitute the rationale for a larger randomized controlled trial. consent: written informed consent for data publication has been obtained. extracorporeal the primary outcome findings from the supernova trial [1] demonstrated that the use of extracorporeal carbon dioxide reamoval (ecco 2 r) allows a reduction in tidal volume (tv) to ultraprotective levels (≈4 ml/kg predicted body weight or pbw) during mechanical ventilation in ards patients without significant increases in the arterial partial pressure of carbon dioxide (paco 2 ). unfortunately, it was not feasible to directly measure ecco 2 r rates during the trial. we used a mathematical model of whole-body oxygen (o 2 ) and carbon dioxide (co 2 ) transport and biochemistry [2] to calculate ecco 2 r rates that permit a fit to the data reported for hemolung (alung technologies) and ila (novalung)/cardiohelp (getinge) devices in the supernova trial [3] . the mathematical model was calibrated under baseline conditions where patients were mechanically ventilated at a tv of 6 ml/kg pbw in the absence of an ecco 2 r device; the o 2 consumption rate, co 2 production rate and pulmonary shunt fraction were adjusted to match the measured baseline arterial partial pressure of o 2 and paco 2 . assuming all baseline parameters were fixed, tv was then reduced to 4.1 ml/kg pbw and the mathematical model predicted the ecco 2 r rate to the change in the paco 2 level. model predictions for the devices are shown in table 1 . these predictions suggest that ecco 2 r rates for ila/cardiohelp devices were approximately twice those for hemolung devices during the supernova trial. these results may be useful to evaluate the expected performance of novel ecco 2 r devices. efficiency and safety of a system crrt plus ecco2r to allow ultraprotective ventilation protocol in patients with acute renal failure f maldarelli 1 despite renal function replacement techniques (crrt), a patient who develops acute renal failure(aki) in intensive care unit (icu) has a mortality rate of 5-80%. this risk is partly due to the adverse effect of aki on other organs than the kidney. respiratory complications are frequently associated with the development of aki. new machines combining crrt with a carbon dioxide removal membrane (ecco2r) allows the setting up of an ultra-protective ventilation (4 ml/kg of predicted boby weight (pbw)) to reduce any lung damage from mechanical ventilation (mv). the reduction in tidal volume (vt) is associated with a decrease in lung damage partly triggered by aki. we evaluated the efficacy of a combined system crrt+ecco 2 r to reduce the vt to ultraprotective values in patients with acute respiratory failure and aki. ards is a syndrome with high morbidity and mortality. an emerging treatment option is ecco2r, but the benefit its remains unclear. we assess different degrees of ecco2r and varying dead space (ds) on ventilator settings in order to minimize mechanical power. we calculated mechanical power as (1) power=rr*{δ〖vt〗^2*[1/2*el+rr*(1+i:e)/(60*i:e)*r]+ δvt*peep} (el: system elastance, r: airway resistance, peep: positive end expiratory pressure, i:e: inspiratory to expiratory ratio). we calculated the combination of respiratory rate (rr) and tidal volume (vt) ("optimal rr" and *optimal vt*) leading to minimal applied power for a stable carbon dioxide elimination of 300 ml/min (vco2) for two scenarios: 1) variation of physiological ds from 10 to 40 % of vt at a fixed rate of eccor2. 2) variation of ecco2r of either 80, 120, 160 or 200 ml/min at a fixed physiological ds of 20%. the alveolar ventilation (va) necessary to eliminate the vco2 was calculated as (2) va= (-vco2*σ_co2*r*t*(1+k_c ))/(vco2/q-p_vco2*σ_co2*r*t*((1+k_c ))/760) σco2: co2 solubility in blood, r: gas constant, t: temperature. pvco2: venous partial pressure, kc: function of ph (12.5 for a ph of 7.2), q: blood flow [5 l/min]). increasing ds from 10 to 40% increases the minimal mechanical power from 5.9 to 10.8 j/min, primarily caused by an increase of optimal vt (495 -672 ml). optimal rr was only slightly increased (6.4 -7.5 /min, figure 1 panel a). for varying ecco2r removal, necessary ventilation ranges from 1.6 to 3.6 l/min. this predicts a minimal power between 5.6 and 10.4 j/min with an unchanged optimal vt (540 -543 ml) and an increasing optimal rr (5.4 to 12.3 /min ( figure 1 panel b)). in order to minimize mechanical power, increasing shunt or co2 production should be met with increases in rr while increases in ds should be met with increases in vt. our results indicate that during ecco2r, mechanical power and thus risk for lung injury can be minimized with higher vt compared to conservative ventilation strategies. validity of empirical estimates of physiological dead space in acute respiratory distress syndrome jd dianti, eg goligher, as slutsky university of toronto, interdepartmental division of critical care medicine, toronto, canada critical care 2020, 24(suppl 1):p091 increased physiological dead space fraction (v d /v t ) is a hallmark of the acute respiratory distress syndrome (ards) and has been shown to predict ards mortality. v d /v t is also important in estimating the reduction in tidal volume (v t ) and driving pressure (δp) with extracorporeal co 2 removal (ecco 2 r). v d /v t can be measured with volumetric capnography but empirical formulae using the patient's age, weight, height, gender and paco 2 have been proposed to estimate v d /v t based on estimates of co 2 production (v co2 ). the accuracy of this approach in critically ill patients, however, is not clear. secondary analysis of a previously published trial [1] in which v d /v t and v co2 were measured in ards patients. estimated dead space fraction (v d,est /v t ) was calculated using standard formulae. agreement between methods was evaluated by bland-altman analysis. the predicted change in δp with ecco 2 r was evaluated using both measured and estimated alveolar dead space fraction (v dalv /v t ). results: vd,est/vt was higher than measured vd/vt, with a low correlation between the 2 (r2= 0.21). vco2 was underestimated by the predicted approach (table 1) , accounting for 57% of the error in estimating vd/vt. the expected reduction in δp with ecco2r using vdalv/ vt was in reasonable agreement with the expected reduction using introduction: acute respiratory distress syndrome (ards) is a common condition in critically ill patient. however neuromuscular blockers (nmb) result controvertial in early treatment of ards [1] . we ought to search systematically and realize a meta-analysis on the matter. an electronic search of randomized clinical trials in adult patient treated with early neuromuscular blockers compared without neuromuscular blockers in ards. the primary objective of the analysis was the mortality at 21 to 28 days. secondary endpoints included mechanical ventilation free days, icu acquired weakness and barotrauma. the search obtained 6 studies for the analysis [1] [2] [3] [4] [5] [6] (figure 1 ). the early use of neuromuscular blockers in ards showed no increase in mortality, but the results should be taken with caution. there was no differences in mechanical ventilation free days. barotrauma is less with the use of nmb. ultrasound is fairly sensitive in the detection of lung infiltrates in patients with hematologic malignancies. in patients with pneumonia requiring intensive care (icu) admission, we hypothesise that abnormal right ventricular (rv) function is associated with an increased 90-day mortality. rv dysfunction in critically ill patients has a well-known association with adverse outcomes [1] . however, its impact on mortality in patients with pneumonia has not been directly studied. patients admitted to the queen elizabeth hospital birmingham icu between april 2016 and july 2019 with a diagnosis of pneumonia who had a formal cardiologist tte were included. abnormal rv function was defined by either depressed function, dilated size or moderate to severe risk of pulmonary hypertension (phtn). abnormal lv function was defined by an lv ejection fraction £ 45% or grade ii or more diastolic dysfunction. patients with a clinical suspicion of pulmonary embolism were excluded. the primary outcome was 90-day mortality. continuous data is presented as median (iqr). categorical data is presented as % and analysed using a chi-squared test. results: 942 patients were admitted to icu with pneumonia, of which 347 (37%) had a tte. patients were 59% male, had a median age of 67 (46-88) and 90-day mortality of 31%. abnormal rv function was present in 30% (n=103), with 15% depressed, 15% dilated and 14% with moderate to severe risk of phtn. rv dysfunction was associated with an increased 90-day mortality compared to normal rv patients (62% vs. 18%, p<0.0001). lv function was abnormal in 25% (n=88) and was not associated with a higher 90-day mortality compared to normal lv patients (38% vs 29%, p = 0.20). rv dysfunction was associated with a higher 90-day mortality than lv dysfunction (62% vs 38%, p = 0.001). conclusions: this is one of the first studies to demonstrate that abnormal rv function is associated with an increased mortality in icu patients with pneumonia. interestingly, abnormal lv function was not associated with an increased mortality. rakuno gakuen university, anesthesiology, hokkaido, japan critical care 2020, 24(suppl 1):p097 we previously reported a simple correction method of estimating pleural pressure (ppl) by using central venous pressure (cvp) and that it can be used to estimate ppl and transpulmonary pressure in pediatric patients with respiratory failure. however, it remains unknown that this method can be applied to patients with various levels of chest wall elastance and/or intravascular volume. the objective of this study is to investigate whether our method is accurate in various conditions of chest wall elastance and intravascular volume. the study was approved by the animal care and use committee of rakuno gakuen university. ten anesthetized and paralyzed pigs (43.2 ± 1.8kg) were mechanically ventilated and subjected to lung injury by saline lung lavage. each pig was subjected to 3 different intravascular volume and 2 different intraabdominal pressures; in each condition, the accuracy of our method was tested. specifically, airway flow, airway pressure (paw), esophageal pressure (pes), and cvp were recorded in each condition, then changes in pes (δpes) and δppl calculated using a corrected δcvp (cδcvp-derived δppl) were compared. cδcvp-derived δppl was calculated as κ × δcvp, where κ was the ratio of the δpaw to δcvp during the occlusion test. means and standard deviations of the two variables that reflect δppl (δpes and cδcvp-derived δppl) in all pigs with all conditions were 6.1 ± 4.1 and 6.4 ± 5.3 cmh 2 o. the bland-altman analysis for the agreement between δpes and δcvp showed a bias of -0. 3 the activity and functionality of the diaphragm are difficult to measure in patients ventilated in intensive care. ultrasound can be a useful tool for monitoring diaphragm muscle activity during different ventilation modes. few data currently exist on diaphragm muscle activity in critically ventilated patients [1] . our goal is to evaluate the respiratory muscular work of the diaphragm with different settings of the respirator by means of an ultrasound scan. the ultrasound assessments of the diaphragm were performed with a 10mhz linear probe at the apposition zone. we measured the thickening of the diaphragm with the respiratory acts, through the thickening fraction (thickening fraction, tf), defined as:tf = (tdimax -tdimin / tdi min)% tdimax: diaphragm thickness at the end of inspiration (maximum thickness) tdimin: diaphragm thickness at the end of expiration (minimum thickness). ventilatory support was divided into 4 classes: 1 -spontaneous breathing (sb) or continous positive airway pressure (cpap); 2 -pressure support ventilation (psv) with low pressure support (5-12cmh2o); 3-psv with high pressure support (> 12 cmh2o); 4 -controlled mechanical ventilation (cmv). a total of 223 assessments were performed in 70 patients. the evaluations were all possible at the right hemidiaphragm, while on the left they were not possible in 7% of the cases. the median tf (iq range) of the 4 ventilation classes was respectively: 42% (25-62%) in sb / cpap; 26% (17-31%) in low-psv; 17% (9-22%) in high psv; and 5% (2-13%) in cmv. the kruskal-wallis test confirms a significant difference between the groups (p <0.0001). the ultrasound of the diaphragm can be a valid tool for monitoring respiratory muscle activity during mechanical ventilation. introduction: extubation failure is defined as reintubation after 48 hours of extubation in mechanically ventilated critically ill patients. it is associated with morbidity and mortality. the aim of our study was to assess reintubation rates in a busy district general hospital and evaluate the impact of high flow nasal oxygen therapy (hfno) on reintubation rates. we performed a retrospective observational study looking at patients admitted to our 7 bedded level 3 critical care unit (370 patients a year) for a period of 5 years between 1 st november 2014 and 31 st october 2019. we included patients over 16 years of age who were mechanically ventilated and length of stay was greater than 48 hours. exclusions were age < 16 years, tracheostomy and patients requiring ventilation for < 48 hours. data was collected from ward watcher, a sicsag database and electronic patient records. our study failed to show any impact of hfno on reducing extubation failure. further work is needed to develop a standardized approach to weaning and to consider routine application of noninvasive ventilation to reduce reintubation rates [1] . fig. 1 (abstract p097) . the bland-altman analysis for the agreement between δpes and cδcvp-derived δppl in various conditions. low: low intravascular volume, normal: normal intravascular volume, high: high intravascular volume, abd-: without an abdominal compression band, abd+: with an abdominal compression band oral endotracheal intubation is common to critically ill patients in intensive care unit. oral care for an intubated patient is important to maintain the moisture of oral mucosa. also, the securement method of oral endotracheal tube developed from cloth tape to commercial tube holder. training powerpoint and video for microteaching was prepared to train up 30 icu nurses to perform the new practice. demonstration and re-demonstration was arranged to assess skills of every nurse. afterwards, each nurse answered a quiz to evaluate the understanding of oetth and its special techniques in application. questionnaire was designed to collect the feedback from all nurses too. the result showed there was 21 nurses (72%) out of 30 nurses achieved full marks in the post-quiz which demonstrated their full understanding of the use of oral ett holder and its nursing care. about the feedback from nurse, 72% of nurses claimed that they were confident in using the new oetth in clinical setting after training. 96% of nurses agreed in time-saving of nursing care routine with the use of an oetth. however, only 56% of nurses agreed that the oetth is effective in prevention of oral mucosa injuries and another 24% of nursing staff disagreed on its function in improving the patient's oral care. in conclusion, some of the nurses did not agree the prevention of oral mucosa injuries by the new securement method with oetth while some nurses welcomed the new oetth as more easy and effective in oral care to intubated patients. execution of percutaneous dilatational tracheostomy using the standard laryngeal mask airway for ventilation: a prospective survey study g gagliardi 1 , v gagliardi 2 , c chiani 3 , g laccania 4 , f michielan 3 1 aulss 5 -veneto, anesthesia and intensive care, adria, italy; 2 aulss 5 -veneto, university of padua, adria, italy; 3 aulss 5 -veneto, anaesthesia and intensive care, adria, italy; 4 aulss 6 -veneto, anaesthesia and intensive care, padua, italy critical care 2020, 24(suppl 1):p101 we fulfilled a survey study dealing with bronchoscope-guided percutaneous dilatational tracheostomies (pdt), using the classic laryngeal mask airway (lma) for the airway management [1] . the aim was to verify the safety and the effectiveness of the aforementioned procedure methods: we performed an observational prospective survey study enrolling 150 patients hospitalized in the intensive care unit. before performing the tracheostomy, the endotracheal tube has been replaced by the laryngeal mask airway. arterial blood gases, ventilation pressures and tidal volumes have been monitored, registered and compared. the median peak inspiratory pressure has been detected stable in all patients. furthermore, during the ventilation with the laryngeal mask, the tidal inspiratory and expiratory volume difference observed between before and after the bronchoscope positioning, has shown a statistically significant variation. finally, in all cases etco 2 , spo 2. , pao 2, and blood ph values persisted within the normal range. the standard lma provides for a reliable airway management and allows an effective ventilation while performing the pdt. once positioned in the supraglottic zone, the lma does not need to be moved throughout all the pdt performance, avoiding risks of displacement, glottic harm and airway device damage, and permitting an easy handling of the bronchoscope, which gives an appropriated visualization of the trachea and a more efficient aspiration. in consequence to the large internal diameter of the lma tube, ppeak has continued to be stable in all patients, providing for minor resistance and inspiratory work. eventually, no late complications, such as tracheal stenosis and infections, have occurred. tracheostomies are the most common surgical procedure performed on critically ill patients. randomized control trials comparing tracheostomy timing in intensive care patients have been equivocal. in order to perform non-urgent tracheostomy in our icu, consent is required from the patient or a formal guardian appointed ad hoc by the courts. since tracheostomies are practically the only elective surgery performed in the critically ill, icu requested guardianship almost always indicates a clinical decision to perform tracheostomy. as appointing a guardian and arranging a tracheostomy takes about a week, the decision to appoint a guardian offers a unique "intention to treat" opportunity to evaluate outcomes in patients for whom tracheostomy is planned. we performed a retrospective analysis over 3 years on patients for whom guardianship was sought excluding those requiring urgent tracheostomy and those with a do-not-resuscitate order. patients were divided according to outcome (tracheostomy, extubation or death prior to tracheostomy) and compared. guardianship was sought for 233 ventilated patients. a decision to withhold tracheostomy was made for 13 patients, who were excluded, leaving 220 patients for analysis. tracheostomy was performed for 131/220 (60%) patients, 62/220 (28%) were extubated and 27/220 (12%) died while waiting for tracheostomy (from nonairway related reasons). tracheostomy was performed on mean ventilation day 16±1. comparing extubated patients to those who had tracheostomy (table) shows similar demographics, but significantly lower mortality and hospital length of stay. a significant proportion of patients initially planned for tracheostomy were successfully extubated. despite demographic similarities, mortality in this group was significantly lower than for patients undergoing tracheostomy. for a selected subgroup of possibly difficult to characterize patients, delaying tracheostomy may be beneficial. figure 1 ). ptis were analysed by speciality and by outcome. complications occurred in 6 cases (incidence 6.5%). there were 3 cases of subcutaenous emphysema, 1 pneumothorax (occuring d6 post procedure) and 1 case each of stoma and suture site infection. there was 1 unplanned cannula change within 7 days of insertion. 24% of cases had cuff inflated on discharge from icu. handover of care was suboptimal; follow up care plans were documented in 18% of cases. a supervising consultant was present for all ptis. there was a trend of increased insertion by consultant and increased reliance on theatre, with corresponding decrease in the number inserted by trainees. pti in our training icu appears safe with low incidence of complications and good senior support for tracheostomy insertion. emphasis must continue on training junior intensivists in pti. transition of care beyond icu requires further work where currently there is suboptimal handover of care and safety netting for non-icu colleagues. supplemental oxygen administration is ubiquitous in the critical care environment, yet evidence is mounting for the deleterious effects of hyperoxia [1] . concerns over the adverse effects from hypoxaemia often exceed those of hyperoxaemia in developing world settings, and inconsistent availability of blood gas monitoring may limit judicious oxygen titration. the aim of this project was to audit oxygen delivery practice and introduce qi measures to avoid excess oxygen delivery in a tertiary icu in lusaka, zambia. a prospective snapshot of ventilatory parameters were recorded for critically ill patients over a 5-week period, including positive end expiratory pressure (peep), fio 2 , and time-course spo 2 . systematic education was provided through group and one to one tutorials to empower nursing and medical staff to titrate oxygen safely and appropriately. repeat data collection was then performed over 4 weeks. initially 18/30 patients (60%) were over-oxygenated, as defined by fio2 >0.5 and spo 2 consistently >95%. 12/18 patients with an fio 2 of >0.5 had peep ≤ 5cm (67%). no patient had a pao 2 recorded in the past 24 hours. education was provided as well as implementation of unit protocols above all patient beds documenting a stepwise approach to titration peep and fio 2 . post intervention fewer patients were over-oxygenated: 7/21 (33%) had fio2 >0.5 and spo 2 consistently >95%, and 7/18 with an fio 2 >0.5 (39%) had a peep ≤ 5cm. in addition, 7/21 (33.3%) had a pao 2 recorded within 24 hours. this qi project has shown that nurse engagement and systematic education to titrate fio2 and peep can be achieved in a resource poor setting and may decrease the incidence of hyperoxia in critically ill patients. availability of blood gas monitoring and knowledge of interpretation was a major barrier to oxygen titration tracheal intubation (ti) in adult burn patients might be unnecessary in 30 to 40% of cases [1, 2] . in pediatric burn patients, there is little data on both the rate of ti and the rate of early extubation [3] . it has been common practice for a child with a facial burn and/or a suspected airway injury to be intubated early due to the risk of losing airway patency. however this risk should be mitigated against the potential risks of ti and mechanical ventilation in children. therefore the aim of this study was to describe the airway status of child burn victims taken in charge of in our pediatric burn intensive care unit. focused on patients arriving with ti, we investigated the rate of early extubation. in addition we compared non intubated patients with those with prolonged ti. this retrospective study described a cohort of 1520 patients hospitalized between 2010 and 2018. data was retrospectively recorded from the patient's paper clinical chart. the mean age of our patients was 2.8 ±3.1 years [mean±sd] with an average burn area of 14±11%. 86% had scald burns and 45% had facial burns. 4% of the children were admitted in the burn icu with ti. for 36% of them, tracheal tube was removed within the first 48 hours after admission. the probability of prolonged ti increased independently with the burned skin area (bsa) (p <0.0001), the presence of facial burns (p = 0.001), and in case of flame burns (p = 0.007) ( figure 1 ). among patients with more than 70% bsa, 85% were intubated more than 48h. among patients with less than 20% bsa, 0.5% were intubated more than 48h. according to our retrospective data, it seems appropriate to intubate children with 70% and more bsa, while for patient with less than 70% bsa, it might be relevant to seek guidance from physician of the nearest burn center. under 20% bsa, ti seems rarely required. an analysis of the predictive applicability of initial blood gas parameters for the need for intubation and the presence of inhalation injury in patients with suspected inhalation injury c pirrone 1 , m chotalia 2 , t mangham 1 , r mullhi 1 , k england 1 , t introduction: we hypothesise that initial blood gas parameters have a good predictive applicability in detecting the need for intubation and the presence of inhalation injury in patients with suspected inhalation injury. to the best of our knowledge, this has not been directly studied in the literature. patients with suspected inhalation injury admitted to the icu at queen elizabeth hospital, birmingham between april 2016 and may 2019 were included. the initial blood gas parameters analysed were pao 2 (kpa), paco 2 (kpa), ph, carbon monoxide level (cohb; %) and pao 2 /fio 2 (pf) ratio. receiver operator characteristics (roc) for these parameters were plotted against the need for intubation for more than 48 hours and the presence of inhalation injury as detected by bronchoscopy and laryngoscopy. area under the curve (auc) for each parameter was calculated. results: 85 patients were admitted with suspected inhalation injury to the icu. 68% were intubated for more than 48 hours. of patients who were intubated, 69% had inhalation injury as indicated by bronchoscopy or laryngoscopy. table 1 outlines the auc for initial blood gas parameters in detecting the need for intubation for more than 48 hours and the presence of inhalation injury. ph was the parameter with the most prominent auc, with reverse correlation indicating fair accuracy. no clear inflection point was identified, although all patients with ph < 7.25 required intubation and had inhalation injury. paco 2 had a fair predictive applicability in detecting the need for intubation. pf ratio, pao 2 and cohb had poor accuracy. conclusions: initial blood gas parameters had a broadly poor predictive applicability for the need for intubation and the presence of inhalation injury in patients with suspected inhalation injury. severe acidosis (ph < 7.25) was the most useful blood gas parameter. clinicians should be cautious in using blood gas parameters alone to inform intubation decisions. lung cancer surgery is associated with a high rate of pulmonary complications including ards and mandates lung protective ventilation strategies [1, 2] . such strategies include non-intubated video assisted thoracic surgery (nivats) with spontaneous breathing [3] . currently neither data on respirator settings nor on gas exchange have been reported for applying the latter. this data constitutes a prerequisite for meaningful evaluating the respiratory consequences of non-intubated spontaneous breathing during lung cancer surgery. the aim of this case series was for the first time providing such data from lung cancer surgery including pneumonectomy. during a 12 month period 32 patients without contraindications [3] scheduled for video assisted thoracic surgery (vats) for non-anatomical and anatomical lung resection including one pneumonectomy (px) were offered non-intubated spontaneous breathing. all patients gave informed written consent to the procedure as well as for analysis and publication of data. anaesthetic management included target controlled infusion of propofol and remifentanil, laryngeal mask airway, and pressure support ventilation. we present early data that early trials of cuff deflation within 48 hours of tracheostomy insertion can be achieved using a standardized protocol. its impact on length of stay, duration of ventilation and patient-centered outcomes needs to be investigated in larger multi-centre trials. preventing underinflation of the endotracheal tube cuff with a portable elastomeric device. a randomized controlled study je dauvergne 1 , al geffray 2 , k asehnoune 2 , b rozec 1 , k lakhal 1 1 hopital laënnec -chu de nantes, service d´anesthésie-réanimation, nantes, france; 2 hotel-dieu -chu de nantes, service d´anesthésieréanimation, nantes, france critical care 2020, 24(suppl 1):p112 the management of the endotracheal tube cuff pressure (p cuff ) is routine practice for critical care nursing staff. underinflation could lead to ventilator-associated pneumonia [1] whereas overinflation exposes to tracheal damage [2] . multi-daily check and adjustment is recommended to ensure that p cuff lies between 20 and 30 cmh 2 o [3] . to automate this task some devices exist but may be inconvenient, bulky and/or ineffective. their use is not supported by guidelines. a portable elastomeric device could be appealing for p cuff automated regulation. this prospective randomized controlled study tested whether the tracoe smart cuff manager tm reduced the rate of patients undergoing ≥1 episode of underinflation (p cuff <20 cmh 2 o), as compared with routine manual p cuff adjustment. monocentric, randomized controlled study. patients with acute brain injury and receiving mechanical ventilation were prospectively allocated to one of the two arms: manual reading and adjustment of p cuff at least every 8h (routine care) or adjunction of the smart cuff manager tm (intervention). this study was approuved by an institutional review board. among 60 randomized patients (routine care in 32, smart cuff manager tm in 28), 506 measurements were performed in 48h. with routine care, a higher rate of patients experienced at least one episode of underinflation (62.5 vs. 17.8%;p<0.001). episodes of underinflation episodes (15% vs. 2%;p<0.001) and manual adjustments (77% vs. 56%;p<0.001) were more frequent with routine care. for overinflation, there was no between-arms difference (p>0.99). the adjunction of continuous p cuff control with the tracoe smart cuff manager tm reduced the incidence of p cuff underinflation as compared with manual intermittent adjustments. overinflation was not promoted by this device. direct laryngoscopy as a technique for tracheal intubation is a potentially lifesaving procedure that healthcare professionals in a variety of fields are taught. however, this skill is challenging to acquire and difficult to maintain. poorly performed intubation technique can lead to potentially serious complications [1] . the intersurgical iview video laryngoscope is a new intubation tool which may have advantages over direct laryngoscopes, such as the macintosh, in the hands of novice personnel. a prospective randomized counterbalanced trial of 30 medical students, who did not have previous airway management experience, was conducted. each student received brief didactic teaching,following this, participants were directly supervised performing laryngoscopy and intubation using the macintosh and iview devices in an alternating pattern. students were permitted up to three attempts to successfully intubate under four conditions, three laryngoscopy conditions using alaerdal intubation trainer and one using a laerdal simman manikin. there was no significant difference in the success rate of intubation or time to intubation between the two devices. the iview outperformed the macintosh in time to intubation in the normal airway in the final scenario, once students gained experience with both devices. no significant difference was found in the number of optimisation manoeuvres, or intubation attempts between groups. areas where the iview outperformed the macintosh included severity of dental trauma and participants' perception regarding ease of use ofthe device. the iview may prove to be a useful teaching tool for novice personnel who are acquiring the skills of tracheal intubation. patients with a primary pulmonary pathology were more likely to respond to aprv. this association has not been described before and warrants further multi-centre exploration in a larger patient group. introduction: airway suctioning is common during mechanical ventilation, using either an open endotraqueal suctioning or closed endotracheal suctioning (ces). closed circuits were developed to prevent arterial desaturation and atelectasis associated to ventilator disconnection. however, ces may cause substantial loss of lung volume. the purpose of this study was to investigate the effects of a compensation method to prevent the loss in aeration during ces. the suctioning technique was performed for 15 seconds, negative pressures limited at 150 mmhg. closed suction catheters with 14fr (halyard health, georgia, eua) were used. electrical impedance tomography (eit) monitoring and arterial blood gas were collected. a nihonkoden mechanical ventilator (nkv550, california, eua) was applied, having a newly developed algorithm for suctioning which overcomes any pressure loss during suctioning (inlinesuction-app). when activated, the app delivers pcv ventilation, adding 2 cmh 2 o of end-expiratory pressure above peep, and delivering driving pressures of 15 cmh 2 o. results: pigs (30±5.4kg) with injured lungs and mechanically ventilated. we tested the aspiration procedures using low peep=5cmh 2 o, or high peep=±12.3cmh 2 o with v t2 o), whereas maintenance of compliance was observed when the app was on (from12.2±1.4 ml/cmh 2 o to 12.5±4.5 ml/cmh 2 o. blood gas in a representative animal showed a drop in pao 2 when app was off (from 247, to 149 mmhg after 2 min, and to 176 mmhg after 10 min) ( figure 1 ). with app on the pao 2 changed from 259 (pre-suction), to 223 (2 min), to 253 mmhg (10 min). the new nksoftware, delivering pcv ventilation during suctioning, could prevent atelectasis and functional loss associated to the procedure. tyrosine kinase inhibitor: an effective tool against lung cancer involvement responsible for acute respiratory failure in icu y tandjaoui-lambiotte 1 patients with advanced-stage non-small-cell lung cancer have high mortality rates in the intensive care unit (icu). in the last two decades, targeted therapies have changed the prognostic of patients with lung cancer outside the icu. the fast efficacy of targeted therapies led some intensivists to use them as rescue therapy for icu patients. we performed a national multicentric retrospective study with the participation of the grrroh (groupe de recherche en réanimation respiratoire en onco-hématologie). all patients with non-small-cell lung cancer admitted to the icu for acute respiratory failure between 2009 and 2019 were included in the study if a tyrosine kinase inhibitor was initiated during icu stay. cases were identified using hospital-pharmacies records. the primary outcome was overall survival 90 days after icu admission. results: thirty patients (age: 60+/-14 years old) admitted to a total of 14 icus throughout france were included. seventeen patients (59%) were nonsmoker. adenocarcinoma was the most frequent histological type (n=21, 70%). most patients had metastatic cancer (n=21, 70%). epithelial growth factor receptor mutation was the most common oncologic driver identified (n=16, 53%). during the icu stay, 17 (57%) patients required invasive mechanical ventilation, 13 (43%) catecholamine infusion, 3 (10%) renal replacement therapy and one (3%) extracorporeal membrane oxygenation. eighteen patients (60%) were discharged alive from icu and 11 (37%) were still alive after 90 days (see figure) . moreover, 6 patients (20%) were alive one year after icu discharge. despite a small sample size this study showed that, in the context of lung cancer involvement responsible for acute respiratory failure, the use of tyrosine kinase inhibitor should not be refrained in patients with severe condition in icu. the burned patient is one of the most complex patients whith a very high mortality. those patients with inhalation injury have a worst prognosis, typically associated with respiratory complications. the aim of our study is to evaluate the mortality of burn patientes with inalation injury in a critical burn unit. a prospective, observational and descriptive study was conducted over a period of 3 years. inhalation injury was defined with these criteria (≥ 2): history of injury in an enclosed space, facial burns with singed nasal hair, carbonaceus sputum and stridor. if they were intubated it was diagnosed by bronchoscopy. demographic data, tbsa, absi, baux score, apache ii, sofa, mechanical ventilation (mv), complications, length of stay, hospital course and mortality data were collected. results: 362 burns patients were admitted. 24% (84 patients) had inhalation injury. mortality among patients with inhalation injury was 28,6% (24 patients). most patients were men and those who died were older and with higher severity scores (fig. 1) . we found no significant differences between groups in the need for mv (95% vs. 85%) or in the percentage of tracheostomy performed (33.3 vs. 28.3). however, patients who died had more respiratory complications like ards, and also shock, renal failure and need of renal replancement therapies although infectious complications were similar in both groups. there was no statistically significant difference in volume used during initial resuscitation in the different groups. patients with inhalation injury who died had higher severity scores at the begining. although there were no differences in the need for mv patients who died had more respiratory complications as well as shock, renal failure and need of rrt, but no infectious complications.the volume used during inicial resuscitation, that was always related to the prognosis, was similar in both groups. further studies are needed to see if this greater initial severity corresponds to the degree of inhalation. aerogen, medical affairs, galway, ireland; 2 aerogen, science, galway, ireland critical care 2020, 24(suppl 1):p120 patients with acute exacerbations such as asthma are prescribed aerosol therapy from presentation in the emergency department to progression through to the intensive care unit. however, the variability in dose delivery to the lung across the possible patient interventions is not well characterized. here, we assess the predicted lung dose of a bronchodilator in a simulated spontaneously breathing adult patient via both facemask and nasal cannula, and via tracheostomy during mechanical ventilation. a standard dose of 2.5 mg in 2.5 ml salbutamol was aerosolized using the aerogen solo nebulizer (aerogen, ireland). for facemask testing, the nebulizer was used in combination with the aerogen ultra with 2lpm supplemental oxygen flow. for nasal cannula testing, the nebulizer was used in combination with the airvo 2 system (fisher and paykel, nz) system at both 10 and 50lpm gas flow rate. tracheostomy-mediated ventilation was assessed in combination with a hme, with the nebulizer placed between the hme and the tracheostomy tube. international standard iso27427 adult breath settings (vt 500ml, bpm 15, i:e 1:1) were used across all tests, and generated using a breathing simulator (asl5000, ingmar medical, usa) or mechanical ventilator (servo-u, maquet, sweden). the dose delivered to the lung was assessed using a capture filter at the level of the trachea, with drug mass determined using uv spectrophotometry at 276nm and interpolation on a standard curve. the results of testing are illustrated in figure 1 . the bronchodilator dose delivered to the simulated patient was seen to be relatively consistent between progressive interventions, except during high flow therapy, with the more clinically relevant 50lpm gas flow rate having a profound effect on the dose. these results may go some way towards explaining how different patient interventions can affect aerosol dose. the the mechanical ventilation (mv) have been identified as an independent factor indicating a worse prognosis for lung cancer patients [1] . this study was conducted in order to assess the results of noninvasive mechanical ventilation (niv) and/or invasive mechanical ventilation (imv) modalities in lung cancer patients admitted to the icu with acute respiratory failure (arf). in this study, lung cancer patients with respiratory failure who were admitted to the icu between january 2017 and december 2018 were evaluated retrospectively. results: 93 patients were included in the study. the mortality rate was 18.3%. 83 patients had niv. imv was applied to 10 patients. in the first 24 hours, 39 of the 83 patients who were initially treated with niv were administered imv. the duration of hospital stay, diagnosis of pneumonia and mortality rate were found to be significantly lower in patients treated with niv alone (p≤0.001, p=0.004, p=0.025), but glaskow coma score (gcs) was significantly higher in this group (p≤0.001). the mortality rate was similar between the patients who were initially treated with imv and those who were treated with imv in the first 24 hours. charlson comorbidity index (cci) and mv duration were significantly higher in patients who died (p=0.01, p= 0.021), but gcs was significantly lower in this group (p=0.032). in the linear regression model for the likelihood of mortality, ccl≥9 and unsuccessful niv increased the mortality rate by 3.4 (1.1-10.5) and 5.2 times (12-23.6) respectively (p=0.036, p=0.032). niv has been an effective modality for respiratory support in most lung cancer patients presenting with arf. however, failed niv seems to be a factor for increased mortality. therefore, the choice of respiratory support modality to be applied in this patient group should be decided by considering the gcs, cci and etiology of arf. the interaction between ventilator settings and the occurrence of acute kidney injury is not fully elucidated. this study aimed at investigating the effect of stepwise increase in peep level on the risk of acute kidney injury as evaluated with the renal resistivity index (rri).the primary outcome is to investigate whether increased levels of peep could lead to increase rri and whether rri could predict the occurrence of aki. methods: patients mechanically ventilated for at least 48 hours and without aki at admission were included in the study. rri was calculated at icu admission. posterolateral approach was used for kidney ultrasound. the peak systolic velocity (v max ) and the minimal diastolic velocity (v min ) were determined by pulse wave doppler, and the rri was calculated as (v max -v min )/v max . the exam was performed modifying the peep levels: 5, 10 and 15 cm h 2 o in random order for 15 minutes. occurrence of aki was defined within 7 days according to kdigo criteria. sixty-four patients were enrolled in the study and incidence of aki was 14/64 (22%). demographical and clinical characteristics are reported in table 1 . increase in peep showed a significant increase in rri from peep 5 to peep 10 (p<0.001) and from peep 10 to peep 15 (p=0.001) ( figure 1 ). the area under the roc curve of rri to predict aki was 0.845 at peep 5, 0.898 at peep 10 and 0.894 at peep 15 (all p<0.001). the youden index analysis showed an rri>0.70 as the best cut off for aki with a sensibility of 65% and a specificity of 96%. patients with rri>0.70 were 11/64 (17%), 13/64 (20%) and 22/64 (34%) at peep 5,peep 10 and peep 15 respectively. patients ventilated with a peep value associated with rri>0.70 had higher incidence of aki (11/14 vs 6/50, p<0.001). the application of peep can increase intrarenal vascular resistance,which is associated occurrence of aki; peep level should therefore be balanced taking into account the rri. the rri seems able to predict occurrence of aki in mechanically ventilated patients. alveolar and respiratory mechanics modifications produced by different concentrations of oxygen in healthy rats subjected to mechanical ventilation with protective ventilatory strategy d dominguez garcia 1 , r hernandez bisshopp 1 , jl martin barrasa 2 , d viera camacho 1 , a rodriguez gil 1 , j arias marzan 1 , s garcia hernandez 3 high oxygen can damage tissues [1] . in this study, we analyze the histological and pulmonary mechanics modifications that can occur when identifying different inspiratory oxygen fractions (fio 2 ) in lungs of healthy rats during protective mechanical ventilation. we use sprague-dawley rat. 4 groups were designed, each with 6 animals, the tidal volume (6 ml/kg), peep (3 cmh 2 o) and respiratory rate (90 rpm) were kept constant, changing the fio 2 between the groups. four groups were established: fio 2 0.21, 0.4, 0.6 and 1. after 4 hours, the lungs were removed for histological study and obtaining the wet/dry index. the histological modifications studied were: alveolar septa (as), alveolar hemorrhages (ah), intraalvelolar fibrin (if) and inflammatory infiltrates (ii). each parameter was rated from 0 to 3 [2] . peak pressure (pp) and pulmonary compliance were monitored every 60 minutes. different statistical tests will be used to analyze the data. results: references to the damage produced in the as, ah, if, ii and the global histological pattern were identified in the groups with the highest fio 2 and there was more damage (p <0.00001) ( figure 1 ). the wet/dry index rose significantly as the oxygen concentration increased (p = 0.001). in the groups to which a fio 2 of 0.6 and 1 was administered, the pp selected specific values with respect to the baseline intake from the first 60 minutes, an aspect that was not appreciated in the other groups (p <0.0001). regarding pulmonary compliance, it will be seen that, in the fio 2 0.6 and 1 groups, it decreased from the first 60 minutes, finding differences with respect to the other groups (p <0.0001). conclusions: mechanical ventilation applied for 4 hours in healthy animals produces disorders that are more pronounced as oxygen concentration increase. fio 2 greater than or equal to 0.6 should be avoided without clinical justification. introduction: patients requiring prolonged acute mechanical ventilation (pamv, defined as 4+ days on mv) are sicker and incur disproportionate morbidity and costs relative to patients on short-term mv (stmv, <4 days of mv). we quantified specific clinical outcomes among patients requiring pamv vs. stmv in a contemporary database. we conducted a multicenter retrospective cohort study within~700 hospitals in the premier database, 2014-2018. using icd-9-cm and icd-10 codes we identified pamv and stmv patients, and compared their baseline characteristics and hospital events. because of the large sample size, we omitted hypothesis testing. a total of 691,961 patients met the enrollment criteria, of whom 266,374 (38.5%) received pamv. at baseline, patients on pamv were similar to stmv with regard to age (years: 62.0 ± 15.8 pamv vs. 61.7 ± 17.2 stmv), gender (males: 55.6% pamv vs. 53.9% stmv), and race (white: 69.1% pamv vs. 72.4% stmv). pamv group had a higher comorbidity burden than stmv (mean charlson score 3.5 + 2.7 vs. 3.1 + 2.7). the prevalence of each of the indicators of acute illness severityvasopressors (50.3% vs. 36.9%), dialysis (19.4% vs. 10.3%), severe sepsis (20.3% vs. 10.3%), and septic shock (33.5% vs. 15.9%)was higher in pamv than stmv, as were hospital mortality and combined mortality or discharge to hospice (figure 1 ), extubation failure (12.3% vs. 6.1%), tracheostomy (21.6% vs. 4.5%), development of c. difficile (4.5% vs. 1.7%), and incidence density of ventilator-associated pneumonia (2.4/1,000 patient-days vs. 0.6/1,000 patient-days). conclusions: over 1/3 of all hospitalized patients on mv require it for 4 days or longer. pamv patients exhibit a higher burden of both chronic and acute illness than those on stmv. commensurately, all clinical outcomes examined are substantially worse in association with pamv than stmv. identifying the readiness of patients recovering from critical illness for liberation from invasive mechanical ventilation (imv) is not always straightforward [1] . the scottish intensive care society (sics) trainee audit 2018 conducted a scotland-wide study to understand current practices relating to liberation from imv. data were prospectively collected on patient demographics, indication for intubation, spontaneous breathing trial (sbt) practices, physiological markers, icu outcome and icu los. all patients >18 years ventilated with imv for > 24hrs from the 1 st nov. 2018 -30 th nov. 2018 were eligible for inclusion. exclusion criteria included extubation for end-of-life, death whilst intubated and presence of tracheostomy. logistic regression was performed to detect factors associated with extubation failure (ef). results were analysed via excel 2010 and stata v.14.1. patient benefit and privacy panel approval was granted. total population of 172 patients were included: 108 (63%) male and median apache2 score 19 (iqr 13-23). ef at first attempt occurred on 27 occasions (15.7%), median icu los of 10 days (iqr 7-12), mortality rate 22.2%. the cohort successfully extubated first time had a median icu length of stay of 5 days (iqr 3-9) and mortality rate of 1.4%. methods of sbt and extubation outcomes detailed in table 1 . no sbt prior to extubation had higher odds of ef (or 2.52, ci 1.09-5.84, p=0.03); patient ventilation for < 3 days had a three times higher odds of ef (or 3.31, ci 1.09-10.1, p=0.03). these were independently associated with ef on multivariate analysis conclusions: we found a reintubation rate of 15.7% in scottish icus. type of sbt most commonly used is divergent from the methods advocated in the literature. the lack of sbt and early extubation attempt was associated with failure, which in turn was associated with longer icu los and higher mortality. in patients undergoing prolonged invasive ventilation we hypothesise that abnormal right ventricular (rv) and left ventricular (lv) function are associated with increased 90-day mortality. whether changes in lv or rv function could aid in the prognostication of these patients has not been directly studied. patients admitted to the queen elizabeth hospital birmingham icu between april 2016 and july 2019 who were intubated and ventilated for more than 7 days and had a formal transthoracic echocardiogram (tte) whilst in icu were included. abnormal rv function was defined by the presence of depressed function, dilated size or moderate to severe risk of pulmonary hypertension. abnormal lv function was defined by the presence of lv depression (lv ejection fraction £ 45% or grade ii or more diastolic dysfunction) or a hyperdynamic lv (formally mentioned in tte report). patients who had a neurological cause for prolonged ventilation were excluded. the primary outcome was 90-day mortality. categorical data is presented as % and analysed using a chi-squared test. continuous data is presented as median (iqr). results: 871 patients required prolonged ventilation, of which 350 (40%) had a tte. patients were aged 62 (49-75), were 61% male and had a 36% 90-day mortality. the median ventilator days were 13 (6-20) and 77% required a tracheostomy. abnormal rv function was present in 26% (n=90) and was associated with an increased 90-day mortality compared to normal rv function (68% vs. 25%, rr 2.71 [2.10-3.50], p< 0.0001). lv function was abnormal in 27% (n=95) and was associated with an increased 90-day mortality compared to normal lv function (54% vs 28%, rr 1.91 [1.47 -2.49], p < 0.0001). abnormal rv function had a trend towards an increased mortality compared to abnormal lv function (68% vs 54%, rr 1.26 [1.00 -1.60], p = 0.07). in this study, abnormal rv and lv function were present in a quarter of patients undergoing prolonged ventilation and were associated with an increased mortality. introduction: tidal volume delivered by mechanical ventilation (mv) in sedated patients is distributed preferentially to ventral alveoli, causing overdistention and associated collapse in dorsal alveoli, driving volutrauma, atelectrauma and ventilator-induced lung injury [1] . temporary transvenous diaphragm neurostimulation (ttdn) stimulates diaphragm contraction [2] . when used in synchrony with mv, ttdn encourages increased dorsal ventilation due to the change in pressure gradients with diaphragm contraction, mimicking a more normal physiological pattern. this may improve gas exchange and reduce injury. a pilot study was conducted using 50 kg pigs undergoing mv in a mock icu. deeply sedated subjects were provided lung-protective volume-control ventilation at 8 ml/kg. ttdn diaphragm contractions were delivered in synchrony with inspiration on every second breath, reducing the ventilator pressure-time-product by 15-20% during mv+ttdn breaths. tidal volume distribution was recorded in each condition using electrical impedance tomography, and compared to never-ventilated, spontaneously breathing subjects (nv). results: dorsal ventilation changed from 49% during mv breaths to 54% during mv+ttdn breaths, compared to 60% in the nv group (p=0.035). ventral ventilation changed from 51% during mv breaths to 46% during mv+ttdn breaths, compared to 40% in the nv group (p= 0.042, figure 1 ). conclusions: ttdn diaphragm contraction used as an adjunct to mv yields a more physiological pattern of volume distribution. this translates into less overdistension in the ventral areas and less atelectrauma in the dorsal areas and reduces ventilator-induced lung injury. this technology introduction: by measuring the pes and its derivatives, we can measure the relationship that exist between the diaphragmatic excursion and the oscillation of the esophageal pressure curve: pswing (ps) so we infer that, just as with the pes, the variations of it might be related to a weaning failure [1, 2] . however, no nominal value exists in the bibliography to predict the test result. patients who meet with the inclusion criteria start the weaning process through a test of 30 minutes of spontaneous ventilation, t-tube (tt). and also the respiratory rate (rr) and the tidal volume (tv). from this analysis, an average ps (aps) is determined for each moment of the test (aps1, initial and aps2, final.).a quotient was obtained in relation to these variables using the value previously obtained (quotient dtv/dps x100. a total of 13 patients were included (n=13).regarding the evolution during tt, 9 (n=9) (69%) were successful, while 4 (n=4) (30.76%) failed when analyzing a rate that relates the variables tv and ps, a quotient was obtained in relation to these variables using the value previously obtained (quotient dtv/dps) for patients who were successful and who failed, (dtv/dps)/100 successful patients presented a value of 18.75 while those of the failure group presented a value of 45.83, (or 1,2 -3 p=0.082) ( table 1) . when presenting the relationship between tv and ps through the quotient (dvt/dps)/100, it is observed a tendency to have a higher quotient among patients who failed versus those who did not fail. the process of weaning from mechanical ventilation imposes an additional workload on the cardiovascular system, which may result in impaired myocardial function, increase in left ventricular filling pressure and respiratory distress. among surgical patients, those undergoing heart surgery are particularly susceptible to cardiac dysfunction induced by weaning because of inadequate cardiovascular reserve. the aim of our study was to depict the pathophysiological changes assessed by echocardiography during the steps of weaning and to identify possible predictors of weaning failure (wf). we enrolled 34 consecutive patients undergoing isolated coronary artery bypass grafting in our institution. data were obtained by intraoperative transesophageal echocardiography before sternotomy (t0) and by transthoracic echocardiography at the beginning of weaning (t1) and at the time of extubation (t2). wf was defined as deferral of planned extubation or respiratory failure needing reintubation or non-invasive mechanical ventilation within 48 hours. results: wf occurred in 7 patients (20.6%) and involved manifestations of respiratory distress in 5 (14.7%). we found a significant association between left ventricle outflow tract-velocity time integral (lvot-vti) and ventricular-arterial coupling measured at t1 and wf, with lvot-vti emerging as the best predictor of wf with an area under roc curve of 0.8669 ( figure 1 ); an optimal cutoff value of 15 cm provided 100% sensitivity and 71% specificity. significant increase in e/e' measured at t2 (13.44 vs 9.96, p 0.02) suggested a cardiac etiology of respiratory distress in patients who failed the weaning trial. our study showed that serial assessment of hemodynamic parameters by means of echocardiography is feasible in cardiac surgical patients and can provide insight into pathophysiological changes during weaning. although these preliminary data need to be confirmed in a larger population sample, lvot-vti emerged as a promising predictor of subsequent wf. compliance with guidelines for respiratory therapy in preclinical emergency medicine g jansen, n kappelhoff, s rehberg protestand hospital of the bethel foundation, anaesthesiology, intensive care and emergency medicine, bielefeld, germany critical care 2020, 24(suppl 1):p131 introduction: current guidelines on pre-hospital emergency ventilation are based on the guidelines for lung protective ventilation in the intensive care unit. the present survey was designed to determine the accordance of actual pre-hospital emergency ventilation by german emergency physicians (gep) with these recommendations. recommendations include a respiratory rate (rr) between 10-16/min, a tidal volume (vt) between 6-8 ml/kg, a maximum pressure (pmax) <30 mbar and a positive end-expiratory pressure (peep) of 5 mbar. an anonymous web-based questionnaire encompassing 7 questions was sent to gep from september to december of 2018. gep were asked to specify their level of education, their preferred ventilation settings and the usually chosen parameters employed to guide mechanical ventilation. statistical analysis was performed using the ch²-test with a significance level ≤0.05. 60% of the questionnaires were completed (159/261). 25% of the participants were trainees (tr), 75% consultants (co). as target parameters for guidance of ventilation, 87% of the tr and 91% of the co use capnometry. the vt controlled 62% of the tr and 54% of the co on the basis of body weight. 81% of the tr and 81% of the co reported to control oxygenation using spo2. table 1 shows our analysis of the given answers. there were no statistically significant differences between the groups. deviations from the guidelines of pre-hospital emergency ventilation settings are common and mainly concern the use of a guidelinecompliant peep. in addition, recommended target parameters for guidance of ventilation were not applied in a significant proportion of gep. prospective observational study including ltx recipients admitted to our icu from february2017 to january2019, who underwent a spontaneous breathing trial (sbt) using a t-piece for 30 minutes. clinical variables and arterial blood gas samples were recorded before starting sbt and after 20 minutes on the t-piece. diaphragmatic excursion (de) and thickening fraction (dtf) were also assessed using ultrasound(us) after 20 minutes on the tpiece. us-dd was defined as de<10 mm or dtf<0.3 of at least one hemidiaphragm. patients who successfully completed a sbt, defined according to clinical criteria,were extubated. extubation failure was defined as the need for reintubation within 48h. results are expressed as medians (iqr) or frequencies (%). 193 ltx recipients were admitted to the icu, 79 of whom underwent an sbt. 51 were male, and the median age was 58y. main indications for ltx were interstitial lung disease (43.0%), copd and cystic fibrosis. 59 were bilateral ltx, and 13 and 7 were left and right unilateral ltx respectively. 69 patients were extubated after sbt and 6 required reintubation within 48h. 53 presented us-dd, though there were no differences between patients who succeeded and those needing reintubation. in contrast, patients who succeeded showed higher pao2/fio2 after 20 minutes on the t-piece (table 1) . similarly, higher reductions in deltapao2/fio2 after 20 minutes on the t-piece were observed in patients who failed. oxygenation after sbt performed using a t-piece may predict extubation failure in ltx recipients with successful sbt. us-dd was not associated with the need of reintubation. descriptive study about the relationship between self-extubation episodes and patient-ventilator interaction s nogales 1 , introduction: to evaluate the relationship between self-extubation and patientventilator interaction, among other physiological variables, in order to predict and to prevent these events. self-extubation (se) are quality indicators in patients under invasive mechanical ventilations (imv) and are related with mortality [1] . planned secondary analysis of a prospective data base of clinical and physiologic signals of patients receiving imv. we included se episodes (2012-2018) with continuous record of ventilator and monitor signals (bclink bettercare®). we analysed demographic data, physiological parameters (peripheral oxygen saturation spo2, heart rate hr, respiratory rate rr and media arterial pressure map) and patientventilator interaction (asynchrony index ai, ineffective efforts during expiration iee and double cycling dc). we studied a period of 12 hours prior to the se episode. we used the wilcoxon non-parametric test and for a proper analysis a linear mixed effects model. we included 21 episodes of se, mean age 62±13years, 76%men, apache ii at admission 17±10, 4,6±3,8days under imv until the episode, reintubation rate 47.6%, icu stay 20,9±17,6days, icu mortality 14%. at the time of the se, 65% were under sedation, 65% with physical restraint. the 67% were in weaning. we observed a trend to increase in spo2, rr, hr, map and asynchronies in the 2-hour period prior to se episode. we compared these variables from this period with a 2-hour period before and we observed a statistically the data presented in this study show that our results are in accordance with the literature with favorable mortality and early postoperative complication rates and support that this procedure is an excellent alternative for surgery in the elderly patients. it is reported that patients with pulmonary hypertension (ph; systolic pulmonary arterial pressure (spap)≥35 mmhg)) have frequent cardiac complications after transcatheter aortic valve implantation (tavi). ph often gets worse in some patients despite the normal cardiac function after tavi. no studies have ever examined prognosis after tavi in patients with or without worsening of ph. therefore, we retrospectively examined the frequency of mid-to long-term heart failure and cardiac death in patients with and without deterioration of ph after tavi. among 113 patients who underwent tavi at our hospital between february 2014 and march 2016, we analysed 27 patients with ph (spap≥35 mmhg) before surgery. spap was measured in transthoracic echocardiography before and within 1 week after tavi. patients were divided into two groups according to whether spap worsened/ did not change or improved after tavi. we examined the frequency of admission due to heart failure or cardiac death (death caused by heart failure, angina, or myocardial infarction) during the period of 3 years after tavi. ph worsened or did not change after tavi in 9 patients, while it improved in 18 patients. the left ventricular ejection fraction measured within 1 week after tavi showed no difference between the two groups (56.6±11.9% vs 58.4±10.0%, p=0.71). the worsened/ no change group was higher in frequency of admission due to heart failure (logrank; p<0.05) and cardiac death (logrank; p<0.04). despite successful treatment for as by tavi, the frequency of heart failure and cardiac death was higher in patients who did not show improvement of ph after tavi, even in the absence of cardiac function decrease. vigorous intervention for ph worsening after tavi may be helpful to improve prognosis. the there are several different anti platelet drugs that can be used to treat acute cardiac events. currently there are no effective markers that can assess how these drugs modify coagulation profile and quality. a new functional biomarker that measures fractal dimension (df ) and clot formation time (tgp) has been developed [1] . df quantifies clot microstructure whereas tgp is a real-time measure of clotting time. we aimed to validate df and tgp in st elevation myocardial infarction (stemi) and assess the effect of two p2y12 inhibitors which have different pharmacological mechanisms: clopidogrel and ticagrelor. we prospectively recruited 72 stemi patients in the emergency setting. venous blood samples were collected 12 hours after admission, following treatment with either ticagrelor or clopidogrel, in accordance with the local guidelines at the time. the blood samples were tested using the df and tgp biomarker, platelet aggregometry, clot contraction and standard markers of coagulation. results: 36 patients received clopidogrel and 36 received ticagrelor. the df for clopidogrel was higher than ticagrelor (1.75±0.05 vs 1.73±0.06, p=0.18 which corresponds to a decrease in clot mass of 20% figure 1 ) and the tgp was reduced (205±91sec vs 257±89 sec, p=0.06 a 20% reduction in time). the results of the study suggest that clopidogrel is less powerful in its effects on clotting characteristics compared to ticagrelor. blood from patients receiving clopidogrel formed quicker and denser clots. this would suggest the risk of secondary events or stent occlusion is lower in those patients on ticagrelor, highlighting that df and tgp may be important in identifying patients at risk of future thrombotic events, the study is ongoing and will investigate the long term outcome in these patients. introduction: new onset atrial fibrillation (noaf) during critical illness frequently resolves prior to discharge. however long-term risks of noaf (i.e. heart failure, ischemic stroke and death)remains high [1] . previous studies noted that nearly half of noaf cases did not have diagnosis recorded [2] . addressing this may reduce post critical illness mortality by increasing af surveillance post intensive care (icu) discharge. retrospective data was collected from an electronic health record for icu admissions over a 10 month period from a biomarker is defined as a measurable indicator of some biological state or condition. combined with a good clinical evaluation, they can enable an early and safe diagnostic, thus a faster management for the patient. cardiac biomarker testing is not indicated in routine in the emergency department (ed) because of low utility and high possibility of false-positive results. however, current rates of testing are unknown. the aim of our study was to evaluate the importance of measuring cardiac biomarkers especially troponins, d-dimer, and btype natriuretic peptide in our daily practice, and to identify the latest recommendations for a better use of these biomarkers in the diagnostic and therapeutic approaches. we conducted a prospective observational study, over a 13 months periods performed in the ed of the university hospital center ibn rochd, casablanca, morocco, including all patients admitted during our study period and having a blood test for at least one biological marker. the dataset was analyzed by spss statistics 21.0. a total of 182 patients was enrolled. troponins were tested in 85.3% patients (high sensitive in 49.5% and troponin i tni in 35.8%), ddimer in 30.9%, bnp 19% and nt pro bnp in 9.5% of cases. the diagnostic impact was significant in 94.4% of cases for troponins, 84.6% of cases for d-dimer and 87.5% for bnp. the therapeutic impact was considered important in 80.6% cases for troponins, 69.2% for ddimer and 87.5% for bnp. cardiac biomarkers have an important role in the ed, not only do they confirm the diagnosis (including the role of troponins in acs) but also eliminate others (with a strong negative predictive value of d-dimer for thromboembolic disease) and prove the cardiopulmonary origin of acute dyspnea (the significant place of bnp in confirming the diagnosis of acute heart failure). a multicenter study on the comparison of inter-rater reliability of a new and the original heart score among emergency physicians from three italian emergency departments the heart (based on history,ecg,age,risk factors,troponin) score is a valid tool to stratify the acs in chest pain. but some reports suggest that its reliability could be low for heterogeneity in the assignment due to the subjective interpretation of the history. we used the chest pain score for the "history". in this study we compare the reliability of the new heartcps and original heart. this is a multicenter retrospective study conducted in 3 italian ed between july and october 2019 using clinical scenarios. ten physicians were included after a course on heart and heartcps score. we used 53 scenarios which included clinical and demographic data. each participant independently assigned scores to the scenarios using the heart and heartcps. we tested the interrater agreement using the kappa-statistic (k), the confidence intervals are bias corrected ; we used stata/se 14.2 statistical software . a p-value of < 0.05 defines statistical significance. the overall inter-rater reliability was good for heart and heartcps: kappa =0.63 (ci 95%;0.57-0.72)and 0,65(ci95%;0.63 -0.67); with good agreement among all the class of risk for heartcps but moderate in the medium class for heart . we found significant differences of inter-rater reliability among the senior and junior physicians who used the heartcps:k=0.56(ci95%;0.52-0.57)and 0.75(ci95%;0.70-0.79). heartcps score increased its history inter-rater reliability specially among the junior physicians from k=0.35 (ci 95%; 0.27-0.43) to k=0.69(ci 95%;0.62-0.71).the junior physicians seem to be more reliable than senior with the heartcps:k=0.75 (0.71-0.79) vs k=0.56 (ci95%;0.52-0.57). the heartcps showed inter-rater reliability better than original heart among the medium class of risk and the junior group. it could be proposed to young doctors to stratify the acs risk of chest pain. limit: we used scenarios rather than real patients. a hybrid approach as treatment for coronary artery disease: endo-cabg or pci first, does it matter? introduction: the aim of this study is to discuss the short-term results of a hybrid approach combining minimally invasive endoscopic cabg (endo-cabg) with a percutaneous coronary intervention (pci). to bypass the disadvantages and potential complications of conventional cabg via median sternotomy, we developed the endocabg technique to treat patients with single-and multi-vessel coronary artery disease (cad). this procedure is performed with three 5-mm thoracic ports and a mini-thoracotomy utility port (3 cm) through the intercostal space. this technique can be combined with pci: the hybrid approach. the sequence of the 2 procedures (endocabg followed by pci or vice versa) may result in different outcomes. from 02/2016 to 12/2017 data from 81 consecutive patients scheduled for a hybrid technique at jessa, belgium, were prospectively entered into a customized database. this database was retrospectively reviewed. subgroup analysis was performed to compare outcomes of patients who first received endocabg with patients who first received pci. a p-value < 0.05 is considered significant, a p-value < 0.1 is considered as a trend toward significance. four patients underwent revision surgery and 2 patients died within the first 30 days. in 79 patients the left anterior descendens artery (lad) was grafted with the left internal mammary artery (lima), the right coronary artery (rca) was the most stented vessel using pci. patients first treated with pci received more units of fresh frozen plasma after endocabg compared to those who were first treated with endocabg (p=0.03). there was also a trend toward significant more transfusion of packed cells in this small subgroup (p=0.07). the hybrid approach is a feasible technique as a treatment option for patients with multi-vessel cad. if cabg follows the pci, patients are more likely to receive transfusion. a possible explanation could be the need for dual antiplatelet therapy prior to surgery in this group, but this needs further investigation. prognostic difference between troponin elevation meeting the mi criteria and troponin elevation due to myocardial injury in septic troponin t (ctnt) elevation in critically ill patients is common and is associated with poor outcome. using common assays, 40-50% of patients in the icu will have elevated troponin level. our aim was to determine whether there is any prognostic difference between troponin elevation meeting the mi criteria (rise and fall more than 20% together with echo and ecg new abnormalities) and troponin elevation due to myocardial injury in septic patients. we enrolled 101 patients with sepsis and mean sofa score 5,2 respectively in which ctnt level was measured more than once and analyzed there ecg and echo findings. patients were classified into three groups:definite mi (rise and fall ctnt ≥ 20% and contemporaneous changes on ecg and/or echo),possible mi (rise and fall ctnt ≥ 20% and no other findings),myocardial injury (ctnt rise less than 20%) results: data from 101 patients were analyzed (49% female; mean age 61.9 (sd 16.9)). a total of 101 patients had at least one elevated ctnt more than 0.03 mkg/l. in 71 (70%) of patients ctnt level rised more than 20% from the first elevated measurement.64 (63%) of patients met mi criteria considering new ecg and echo findings. the overall mortality rate in all patients was 53.9%.the mortality rate didn't differ significantly in three groups: in the definite mi group 62.4%, in the suspected mi group 52%, in the non mi ctnt elevation group 56,4%, p=0,6. coronary angiography was performed in 46 (73%) of patients from the definite mi group,pci was performed in 18 (39%) of patients. the mortality rate in the invasive group was not significantly lower comparing to the nonivasive group 29% vs 37,8%, p=0,06. bleeding complications were significantly more frequent in the definite mi group 13% vs 7% and 8% respectively conclusions: ctnt level elevation is associated with poor outcome regardless coronary or non coronary injury. myocardial revascularization may be beneficial in patients with sepsis and definite mi, but it is also associated with increased bleeding risk. diagnostic interest of "marburg heart score" in patient consulting the emergencies department for acute chest pain chest pain is a common reason for emergency department visits, although this primarily refers to acute coronary syndrome (acs), this symptom may be frequently related to other non-ischemic etiologies. the aim was to validate the marburg heart score as a tool to exclude coronary artery disease in emergency department patients with nontraumatic acute chest pain. methods: a prospective, observational, descriptive and analytic cohort study conducted in the emergency department, from february 1st to march 31st, 2019, collecting patients consulting for nontraumatic acute chest pain, the "marburg heart" score was calculated for all these patients. telephone contact was made after 6 weeks to look for an ischemic cardiovascular event. we included 171 patients. the mean age was 57 +/-13 years, the sex ratio was 0.86. the majority of the patients (78.9%) consulted directly to the emergency department, 21.1% were referred by a primary care physician. the median time to consultation after the onset of chest pain was 24 hours. high blood pressure was the most common risk factor (43.9%), followed by smoking (31%), diabetes (24.8%) and dyslipidemia (23.4%). thirty-five patients (20.5%) had already coronary heart disease, ecg was pathological in 19.3% of patients, 8 patients had an acs with st segment elevation. at six weeks, 20.6% of the patients had an acute coronary event. according to the patients' answers on the 5 questions of the marburg heart score. the area under the roc curve of this score was 0.78 with a negative predictive value of 87.2%; the "marburg heart score" is a simple, valid and reproducible clinical score with a discriminatory power to rule out the diagnosis of coronary artery disease from the first contact with the patient presenting for chest pain in emergencies. the abdominal aortic aneurysm (aaa) surgery is a complex procedure in elderly patients with high cardiovascular risk. anesthesiological techniques should play special attention to the volume status during cross-clamping as well as to the blood loss. goal directed fluid therapies (gdt) in aaa surgery in elderly patients decrease the perioperative morbidity and mortality [1] . aim of this study is to investigate administration of fluid-based on either a gdt approach or a control method (fluid administered based on static preload parameters and traditional hemodynamic) in all phases of aaa surgery and especially in the phase of clamping and de-clamping. a total of 30 patients asa iii, randomly scheduled for elective, open aaa surgery were included in this clinical trial. they were randomly assigned to two groups i -gdt with targeting stroke volume variation (svv) and ii -control group where fluids were administered at the discretion of the attending anaesthesiologist. in both these groups hemodynamic parameters, central venous pressure (cvp), temperature, blood loss and diuresis were registered during the operation and 48 hours postoperatively. each group was assessed for postoperative complications. gdt group received less fluids and had a higher cardiac index (ci) (3.9± 0.6 vs. 2.9± 0.8 l/minute per m 2 , p < 0.01) and stroke volume index (55.1 ± 5.4 vs. 35.1 ± 5.8 ml/m 2 , p < 0.01) than the control group. there were significantly fewer complications in the intervention than control group (3 vs. 9, p = 0.02). gdt fluid administration enables less use of fluids, improved hemodynamic and fewer postoperative complications in elderly patients undergoing aaa surgery. ultrasonography is a valid diagnostic tool, used to measure changes of muscle mass. the aim of this study was to investigate the clinical value of ultrasound-assessed muscle mass, in patients undergoing cardiothoracic surgery that present muscle weakness postoperatively. for this study, 221 consecutive patients were enrolled, following their admission in the cardiac surgery intensive care unit (icu) within 24 hours of cardiac surgery. ultrasound scans, for the assessment of quadriceps muscle thickness, were performed every 48 hours for 7 days. muscle strength was also evaluated in parallel, using the medical research council (mrc) scale. of the 221 patients enrolled, ultrasound scans and muscle strength assessment were performed in 165 patients. the muscle thickness of rectus femoris (rf), was slightly decreased by 2.18% ([95%ci: -0.21; 0.15], n=9; p=0.729) and the combined muscle thickness of the vastus intermedius (vi) and rf decreased by 3.5% ([95% ci: -0.4; 0.22], n=9; p=0.530). patients whose combined vi and rf muscle thickness was below the recorded median values (2.5cm) on day 1 (n=78), stayed longer in the icu (47 ± 74 vs 28 ± 45 hours, p = 0.015). patients with mrc score ≤ 48 on day 3 (n=7), required prolonged mechanical ventilation support compared to patients with mrc score ≥ 49 (n=33), (44 ± 14 vs 19 ± 9 hours, p = 0.006). the use of muscle ultrasound seems to be a valuable tool in assessing skeletal muscle mass in critically ill patients after cardiothoracic surgery. moreover, the results of this pilot study showed that muscle wasting of patients after cardiothoracic surgery is of clinical importance, affecting their stay in icu. prediction of cardiac risk after major abdominal surgery s musaeva, i tarovatov, a vorona, i zabolotskikh, n doinov kuban state medical university, anesthesiology and intensive care, krasnodar, russia critical care 2020, 24(suppl 1):p147 the aim is to assess the incidence of cardiovascular incidents in major abdominal surgery [1] using the revised lee index. a study was conducted of 144 elderly patients who underwent major abdominal surgery in the krasnodar regional clinical hospital no. 2 under combined anesthesia. in the preoperative period, the risk of cardiovascular incidents was assessed using the revised lee index and the functional status was assessed by met. depending on the lee index, 3 groups were identified: group 1 (n = 69) -low risk (index value -1), group 2 (n = 52) -intermediate risk (index value -2); group 3 (n = 23) -high risk (index value> 3). we estimated the incidence of critical incidents in groups: hypo-, hypertension, arrhythmias, and bradycardia. in the general population, cardiac risk was 2.2 ± 0.7 points; functional status -7.7 ± 1 met. the greatest number of critical incidents was recorded in patients with high risk (58.4%), the smallest -in patients with low risk (9.1%), in patients with intermediate risk -26.5% (n <0, 05 between groups according to chi-square criterion). in the structure of critical incidents, hypotension was most often encounteredin 62 (43%) patients, while some patients revealed several incidents from the circulatory system (n = 116). overall, the lee scale showed good prognostic ability (auroc = 0.81) in predicting hemodynamic incidents. the revised lee index is a useful tool to help assess the risk of cardiovascular incidents and determine patient management tactics in the perioperative period. postoperative cognitive dysfunction (pocd) remains an unresolved problem due to lack of consensus on its etiology and pathogenesis. some believe that pocd is the result of the direct toxic effect of general anesthetics on the nervous system. others claim that surgical trauma activates proinflammatory factors that induce neuroinflammation. wistar rats were allocated into 2 groups: 1-minor surgery (n=20), 2major surgery group (n=20). after 5 days of handling and habituation rats undergone surgery under isoflurane general anesthesia (2 vol.%). group 1 rats underwent laparotomy with gentle gut massage followed by wound closure. rats in group 2 undergone left side nephrectomy. starting from the 4th postoperative day spatial memory in rats was studied in morris water maze which is a cylinder metal pool with a diameter of 1.5 and a height of 0.5 m filled with water (temp.26±1 o c) up to half. it has a platform with a diameter of 12 cm and a height of 1 cm below the water level. testing was preceded by a training stage, which included 8 sessions daily for 4 days. thus, rats developed spatial memory to the location of the platform. on the 5th day of the study test stage was conducted to assess spatial memory: rats were launched from 3 points into maze without platform and data were recorded for 60 seconds at each session. time spent on the target quadrant (ttq) and the number of target area crossings (tac) were registered. a second test was conducted 14 days after the first test to evaluate long-term spatial memory. the duration of surgery and anesthesia did not differ significantly between groups. there was a significant difference between groups in average ttq and tac in test 1 (table 1 ). in test 2 minor surgery group showed better results but they were less significant. major surgery is associated with a more pronounced deterioration of spatial memory in rats in early postoperative period compared to minor surgery. cardiac inflammatory markers in icu patients with myocardiac ischemia after non cardiac surgery (a pilot study) p manthou 1 , g lioliousis 2 , p vasileiou 3 , g fildissis 1 1 national kapodistrian university of athens, athens, greece; 2 national kapodistrian university of athens, general thoracic hospital´´sotiria´´, athens, greece; 3 national kapodistrian university of athens, university of athens, athens, greece critical care 2020, 24(suppl 1):p149 patients with known coronary artery disease have higher perioperative risk for myocardial ischemia [1, 2] . mortality is frequent following cardiac ischemia in the intensive care unit (icu) after non-cardiac surgery. the first group includes patients admitted to the intensive care unit for post-operative follow-up without myocardiac ischemia in the first 24 hours. the second group includes patients with myocardiac ischemia postoperatively and needs intensive care monitoring. cardiac risk assessment was made with the lee index,hemorrhagic risk assessment with the has-bled bleeding score and thrombotic risk assessment with cha2ds2-vasc score. postoperatively, pathological test values such as bnp, troponin, crp, calcitonin were estimated. the sequential organ failure assessment (sofa) systeme was used to assess sepsis. the nursing activity score (nas) scale was used to measure the workload of various nursing activities in the icu. according to the pilot study, the sample consists of 35 patients. 31.4% had myocardial ischemia. the lee index was significantly higher in patients with myocardial ischemia. the duration of hospitalization, the high dose of vasoconstrictive drugs, the length of stay in the icu, the duration of mechanical stay and the nursing workload were higher in patients with myocardial ischemia. ck-mb and troponin levels differed significantly between the two groups. creatinine, bilirubin and bnp during the 24 hours were significantly higher. patients with myocardial ischemia had significantly higher mortality. cardiac risk assessment, has-bled score and cha2ds2-vasc score in combination with cardiac enzymes such as troponin could predict myocardiac ischemia in severely ill icu patients. introduction: according to the literature an airway complication followed thyroid gland surgery are: difficult trachea intubation, tracheomalacia, postextubation stridor and bleeding [1, 2] . most common cause of death was problem with respiration and airway obstruction [3] . subsequent hypoxia could require emergency airway and even tracheostomy [3] . aim of our study was to determine the most common of airway complications and their association with type of surgery in our region. the retrospective cohort study included 400 pts., (369 women, 31 men) was performed in odessa regional hospital, oncology centre odessa. there were three types of patients: with euthyroid goiter -170 (43%), polynodos goiter -125(31%) and thyroid cancer -105 (26%) ( table 1) . airway complications were diagnosed after trachea extubation based on indirect laryngoscope, presence of stridor, desaturation. the pearson's criteria was calculated. the ratio of airway complications after thyroid surgery was 9.7% (39 pts). the main reasons of airway complications in thyroid surgery included: laryngeal edema -22 pts (5.5%); recurrent laryngeal nerve injury -12 pts (3.0%) and postoperative bleeding 5 pts (1.2%). thyroid gland cancer and polynodosal goiter associated with laryngeal edema and recurrent laryngeal nerve injury (pearsen criteria were 0.271 -moderate and 0.203 consequentially). it's may require more attention from the anesthetists after extubation and readiness for an urgent airway. serum iron level and development of multiple organ dysfunction syndrome in patients in the perioperative period s tachyla mogilev regional hospital, department of anesthesiology and intensive care, mogilev, belarus critical care 2020, 24(suppl 1):p151 recently there has been attention of researchers to the problem of perioperative anemia. it was found that it increases the risk of death and postoperative complications. threatening complication is multiple organ dysfunction syndrome (mods). the objective was to determine the level of serum iron in the perioperative period in patients with endoprosthetics of large joints, and with the presence of mods in abdominal surgery. a prospective cohort study was conducted in 77 patients, including 18 men and 59 women, age 61.9 ± 15.1 years. two groups were identified: 1st (control) -patients after endoprosthetics of large joints (n = 40), 2nd (main) -patients in abdominal surgery with the presence of mods (n = 37). the presence of mods was established based on the criteria for the 2016 sccm / accp conference. serum iron was monitored using an au 680 analyzer (usa). the study identified several stages: 1st -before surgery, 2nd -1st day after surgery, 3rd -3rd day, 4th -7th day, 5th -10th day. when studying the indicators of serum iron, its significant decrease (p <0.05) in the postoperative period was established. in the 1st group: 1st stage -15.2 (10-19.4) mmol / l, 2nd stage -5.2 (3.9-7.6) mmol / l, 3rd stage -6.6 (5-8.7) μmol / l, stage 4 -9.7 (8.6-12.1) μmol / l, stage 5 -9.4 (7.8-11 9) μmol / l. in the 2nd group: 1st stage -11.9 (10-15) mmol / l, 2nd stage -3.7 (1.7-4.1) mmol / l, 3rd stage -3, 6 (2.4-4.5) μmol / l, stage 4 -6.5 (4.4-8.2) μmol / l, stage 5 -7.6 (6.5-9 4) μmol / l. moreover, in both groups, iron increased at the 4th stage against the 2nd stage (p <0.05). when comparing the level of iron between the groups, significant differences were found (p < 0.05) at the 2nd, 3rd and 4th stages. in patients in the postoperative period, a decrease in serum iron is observed, the level of which rises by the 7th day, but does not reach the initial values. this decrease is more pronounced in patients with the presence of mods after abdominal surgery. kidney and pancreatic graft thrombosis happened in 5.6% and 18.9%, respectively, and bleeding in 21.1%. forty-one (45.6%) developed at least one infection during hospital stay. infection during icu was found in 13.3% and main pathogens were gram negative bacilli sensible to beta-lactam. after icu, the incidence of multi-drug resistant pathogen was 13.5%, predominantly gram negative bacilli. fungal infection was lower 4%. all-cause hospital mortality rate was 5.6%. infectious complications are the main cause of morbidity and mortality following spk transplantation. the administration of broadspectrum prophylactic antibiotics are leading to the appearance of multi-drug resistant pathogens. knowing local microbiological flora may be helpful, allowing more adequate antibiotic prophylaxis. introduction: cardiopulmonary bypass (cpb) is associated with thrombotic complications. occurrence of thrombosis after cpb is 12% which takes the third place between cpb-associated complications. our study determined preoperative predictors of thrombosis in children with congenital heart defects. 138 patients with congenital heart diseases in age up to 11 months 29 days (median age -4,7 months, youngest age -2 days after birth, oldest -11 months 29 days), underwent cardiac surgery with cpb, were enrolled in this study. all patients were divided into two groups: 1 st -without thrombosis, 2 nd -with thrombosis. protein c, ddimer, von willebrand factor and plasminogen plasma levels were assessed directly before surgery. thrombotic cases were proven by performing doppler ultrasound or mri. thrombotic complications were diagnosed in 30 children (21%). between all thrombotic complications ischemic strokes were diagnosed in 73% (22 cases), arterial thrombosis in 17 % (5 cases), intracardiac thrombus in 7% (2 cases) and mechanical mitral prosthetic valve thrombosis 3%(1). receiver operating characteristic (roc) curves are created for the listed indicators. area under the curve (auc) for protein c 0,64 (sensitivity(sn)-65%, specificity(sp) -50%), d-dimer is 0,65 (sn -65%, sp 50%), for plasminogen activity -0,62 (sn 60%, sp 40%) and for von willebrand factor level -0,64 (sn 80%, sp 55%). an roc curve was created for all three indicators, the auc was 0.7 (sn -80%, sp -40%). these parameters can be recommended as predictors of thrombosis in children after cardiac surgery. cpb is related with a large number of life-threatening complications. in our work, preoperative predictors of thrombosis were identified. based on this data, it is possible to create thrombosis risk scale change the tactics of the anaesthetic approach, the prevention of thrombosis in the postoperative period. further studies are needed to identify other possible predictors of thrombosis. introduction: abdominal ischemia occurs in 9% of patients submitted to aortic aneurysm repair. its early diagnosis requires an elevated index of suspiction, particularly in more severe patients. we hypothesized that earlier increase and higher levels of c-reactive protein (crp) may help to predict intra-abdominal ischemia. we performed a retrospective study of patients admitted to the intensive care department (icd) after abdominal aorta aneurism surgery. we included all patients admitted during a two-year period, that survived for more than 48 hours. primary outcome was splanchnic ischemia assessed by abdominal ct-scan. we also evaluated the presence of bacteremia, abdominal compartment syndrome and icd mortality. association between inflammatory parameters and ischemia was evaluated by multivariate logistic regression. introduction: crp (c-reactive protein) has been shown to be a useful biomarker in identifying complications after major abdominal surgery. gastrectomy is a high-risk surgical procedure that requires post-operative critical care support to monitor for complications which are predominantly infective in nature. the aims of this study were to determine whether there is a relationship between post-operative crp levels and patients who developed post-operative infective complications. a retrospective analysis was performed on patients undergoing elective gastrectomy for gastric cancer at a single centre between september 2011 and july 2016. post-operative crp levels for each day following resection were analysed for all patients. roc curve analysis was used to determine which post-operative day (pod) gave the optimal cut-off. of 144 patients included, the majority were male (61.8%), mean age was 68.5 years and 53.5% had node-negative disease. a total of 84 patients (58.3%) had an infective complication, which includes those who experienced an anastomotic leak. crp levels on post-operative day 3 gave the greatest auc for the gastrectomy group (0.765). crp cut-off of 220mg/l was significantly associated with infective complications (or7.29, 95% ci 3.42-15.58, p= <0.001) and gave a sensitivity of 70% and specificity 76% (ppv 67%, npv 78%). more patients with a crp >220 on post-operative day 3 experienced an infective complication (67% vs 21%, p = <0.001) or a leak in particular (17% vs 6%, p = 0.059). a crp level of less than 220 mg/l on pod3 may be useful to predict the development or exclude the likelihood of such infective complications in this group of patients prior to clinical signs (ppv 67%, npv 78%). this may prompt and facilitate decision-making regarding early investigation and intervention or prevent inappropriate early discharge from critical care, whilst providing more assurance in identifying those who could be stepped down to ward level care. vasoplegia is commonly observed after cardiopulmonary bypass surgery (cpb) and associated with high mortality. chronic use of reninangiotensin aldosterone system inhibitors (raasi) is associated with its incidence and ensuing need for vasopressor support after cpb. renin serves as marker of tissue perfusion [1] . we examined the role of renin in the setting of raasi exposure and vasopressor needs in the peri-cpb period. prospective observational study of 31 adult patients undergoing cpb, aged 66.0±10.5 years (22 men, 9 women). blood was collected 1) post induction, pre-cpb; 2) 30 min post cardioplegia, and 3) immediately post bypass. vital signs and perioperative medications were recorded. as control, blood was collected from 5 men and 4 women aged 53.5 ±10.7, not diagnosed with lung disease and not prescribed any raasi. baseline plasma renin in cpb patients tended to be higher than in control subjects (mean=38.5pg/ml±9.2 vs. 17.1 pg/ml ± 3.5, respectively, p=0.670). 30 minutes into cpb, mean renin was increased from baseline (77.6 pg/ml±17.5, p=0.211), and remained elevated immediately post cpb (74.6 pg/ml±19.1). patients using raasi prior to cpb tended to have a larger increase in renin post cpb (delta=55.3 pg/ ml±30.9) vs. those not previously on raasi (26.9 pg/ml±9.5, p= 0.092). renin was elevated in patients requiring vasopressor support in the 24 hours post cpb vs. those not requiring pressors (41.4 pg/ ml±15.7 vs. 25.1 pg/ml±16.1 p=0.0246). in those prescribed raasi and requiring pressors post cpb, there was a tendency toward greater renin increase than those not requiring pressors postoperatively (49.9 pg/ml±49.7 vs. 8.5 pg/ml±3.9, p=0.036). this study suggests a trend toward higher renin levels, particularly during cpb, in patients prescribed raasi, and a positive association between renin and postoperative vasopressor needs. we speculate that increased renin levels may predict postoperative vasoplegia. cardiac surgery is associated with perioperative blood loss and a high risk of allogenic blood transfusion. it has been recognized that high blood product transfusion requirement is associated with adverse clinical outcomes. 1 guidelines on patient blood management therefor aim at reducing blood loss and blood transfusion requirements in cardiac surgery. 2 as there remains controversy about the advantage of minimal invasive techniques on blood loss an transfusion requirements, 2 we wanted to investigate if the average blood loss and transfusion requirement in minimal invasive endoscopic coronary artery bypass graft surgery (endo-cabg) differ from conventional technique. we assessed the influence of pre-operative anticoagulant medication for blood loss. estimated average blood loss after conventional cabg 3 is 400ml (+/-200) and transfusion requirement 3,4 units packed red blood cells 4 . we performed a retrospective cohort study of our cardiac surgical database. from 01/01/2016 to 31/12/2017, we collected data from 423 patients undergoing endo-cabg. we analyzed blood loss, transfusion as well as pre-operative use of anti-coagulants as a risk factor for blood loss. we found that mean total blood loss in endo-cabg does not differ from conventional cabg, nonetheless mean transfusion requirement was lower in our cohort. use of direct oral anticoagulant is aossciated with increased blood loss and transfusion requirements (table 1) . total blood loss is not influenced by minimal invasive technique for cabg (endo-cabg). an explanation for the lower transfusion requirements is the use of a minimal extracorporeal circulation, which is known to reduce the risk of transfusion. another important factor is the implementation of a standardized transfusion-protocol based on available evidence. reducing transfusion requirements is an important component in improving patient outcome after cardiac surgery and is related to multiple factors in perioperative care of our patients. retinal microvascular damage associated with mean arterial pressure during cardiopulmonary bypass surgery v shipulin 1 retinal perfusion corresponds to cerebral perfusion and it is very sensitive to hemodynamic disturbances [1, 2] . we investigated the association between retinal microvascular damage and hemodynamic characteristics in patients undergoing coronary artery bypass grafting surgery (cabg) with cardiopulmonary bypass (cpb). methods: 10 patients with coronary artery disease and systemic hypertension were examined. ophthalmoscopy and optical coherence tomography were performed before and 10-14 days after cabg. the hemodynamic parameters during cpb were analyzed. results: 4 (40%) patients had changes in the retinal vessels and in the ganglionic fiber structure on 10-14 day after surgery: in 30% of patients the foci of ischemic retinal oedema appeared, in 10% the decrease of the thickness of ganglionic fiber were observed. these changes may be associated with intraoperative ischemia of the central retinal artery. in 6 (60%) patients the mean arterial pressure (map) during cpb was increased up to 90 mmhg. in 4 (67%) of them the association between map and foci of ischemic retinal oedema were revealed. the ischemic retinal changes were observed significantly more often if the delta of map during cpb was over then 20 mm hg compared with the patients where the delta of map was less than 20 mm hg (p=0.035). this is probably due to an intraoperative disorders of the myogenic mechanism of blood flow autoregulation in the retinal microvasculature in patients with coronary artery disease [3] . the level of map up to 90 mm hg during cpb is associated with retinal blood flow impairment and the foci of ischemic retinal oedema. delta of map more than 20 mmhg was associated with the foci of ischemic retinal oedema and decreased ganglionic fiber thickness in 67% of cases. atrial fibrillation after cardiac surgery: implementation of a prevention care bundle on intensive care unit improves adherence to current perioperative guidelines and reduces incidence introduction: atrial fibrillation after cardiac surgery (afacs) is a very frequent complication affecting 30-50% of all patients. it is associated with an increase in morbidity, mortality and hospital and intensive care unit (icu) length of stay. we aimed to implement an afacs prevention care bundle based on a recently published practice advisory [1] , focusing on early postoperative (re)introduction of β-blockers. baseline afacs incidence and β-blocker administration practices in our centre were audited for all patients undergoing valve surgery or coronary artery bypass graft (cabg) during a 6 weeks period. the afacs prevention care bundlean easy to follow graphical toolwas subsequently introduced to the cardiac icu by a multidisciplinary team and audited following a model of improvement approach. after exclusion of patients with preoperative af, differences between pre-and post-implementation groups were compared with chisquare and fisher's exact tests for categorical, and one-way anova for continuous variables, using spss. a total of 384 patients were analysed. patient and surgery characteristics did not differ between groups. significantly more patients received postoperative β-blockers after bundle implementation (82.7% pre-vs 91.3% post-bundle, p=0.019) with a higher proportion on day 1 (36.7% pre-vs 67% post-bundle, p<0.001, figure 1 ). the incidence of afacs was significantly reduced from 35.4% to 23.3% (p=0.009), with a particularly marked reduction in the age group 65-75 years and for isolated aortic valve and cabg surgery. there was no significant reduction in hospital length of stay for this cohort. introduction of an afacs prevention care bundle using a graphical tool improved adherence to current guidelines with regards to early β-blocker administration and significantly reduced afacs incidence. future care bundles should include preoperative interventions and might reduce hospital length of stay. in neonates with univentricular physiology, there is a delicate balance between pulmonary and systemic circulations, with a tendency towards generous pulmonary blood flow, and a risk of systemic underperfusion. preoperatively, the use of hypoxic gas mixture (hm) has been advocated as a therapy to increase pvr, with the aim of improving systemic oxygen delivery. it is a therapy which has been routinely initiated in our institution in the setting of signs of pulmonary overcirculation. we performed a retrospective analysis of all patients in our institution who underwent a norwood procedure and who received hm preoperatively. we compared peripheral saturations, arterial blood gas analysis, serum lactate, regional cerebral and renal saturations and invasive blood pressure, prior to, and then 4,8 and 24 hours after hm was commenced. between 2014 and 2018 (inclusive), 49 patients underwent the norwood procedure. 18 patients received preoperative hm. average fio2 was 17% during administration of hm. average peripheral saturations were 96.1% prior to hm, and dropped to 87.4% at 4 hours, and 88% at 8 and 24 hours after initiation (p < 0.05). there was no change in any of the measured markers of systemic oxygen delivery, including regional cerebral and renal saturations, lactate, urine output or blood pressure. there was an association between an extended period of hm (> 48 hours) and the need for pulmonary vasodilator therapy post norwood procedure. hypoxic gas mixture in patients with parallel systemic and pulmonary cicrculations causes desaturation and hypoxia. it does not lead to an increase in systemic perfusion and thus an improvement in systemic oxygen delivery. its ongoing use in this fragile population should be considered. introduction: analgesia in the critical patient, and especially in the neurocritical patient, is a basic goal in all therapeutic practices. patients in the icu are frequently administered prolonged and/or high doses of opioids. multiple serious complications due to the use of infusion of opioids at large doses has been described. to reduce high doses of intravenous opioids, multimodal forms of analgesia can be used. prospective observational study of the use of tapentadol enteral and buprenorphine in transdermal patches, at low doses, for the control of pain and its effect on reducing the use of fentanyl infusion in high doses on 84 patients admitted to neuro icu of indisa clinic during 2 consecutive years (2018-19). enteral tapentadol (through ng tube) 50 mg/6 hours, was considered in patients who required intravenous fentanyl in continuous administration. buprenorphine was also added at low doses (5 ug/hr) in a weekly transdermal patch, in cases of neurosurgical spine patients, fractures and long-term neuropathic pain. pain was controlled on behavioral pain scale (bps) and visual analogical scale (vas) scores, according to the conditions of each patient. their hemodynamic, gastrointestinal complications and the appearance of delirium episodes according to cam-icu scale were recorded. results: 84 patients received tapentadol. 32 of them also received transdermal buprenorphine. all managed to maintain adequate level of analgesia, not requiring fentanyl at doses greater than 0.5 ug / kg / hr. distribution by diagnoses: neurotrauma 21 patients, guillain barre 12, spine surgery 15, hsa 18, hice 10, malignant ischemic acv 8. complications: gastric retention 12 patients (7%), hypotension 1 (1%), acute hypoactive delirium 3 (3.5%), acute hyperactive delirium 8 (9%). no drug interactions were found. the introduction of enteral tapentadol and buprenorphine patches in neurocritical patients was safe and resulted in a decrease in the use of endovenous opioids and its adverse effects. we hypothesized that changing the pain management for our post cardiac surgical patients to an assessment-driven, protocol-based approach using fast acting and easily titratable agents will significantly improve patient satisfaction by reducing pain intensity in the first 24h after surgery as suggested by society of critical care [1] guideline. we prospectively assessed 101 and 99 (05.2018 vs 06.2019) consecutive patients before and after introducing our pain management protocol. the nursing and medical team received rigorous training on the guideline as well as the correct assessment using appropriate pain scores measured at least hourly (numeric pain score, ≥ 2 is timing of beta-blocker (re)initiation versus incidence of afacs before and after prevention care bundle implementation, per post-operative day and for postoperative days 1-5 (insets) moderate to severe or critical care observation tool, > 2 is moderate to severe). we introduced a multimodal approach with a combination of fast acting iv, long acting oral opiates, regular paracetamol and rescue iv boluses for difficult to control situations and we created a prescription bundle on our electronic prescribing record. among other variables we assessed hours spent in moderate to severe pain in the first 24h after surgery and compared to the data collected before the guideline was introduced. we analysed 101 patients from 2018 and 99 from 2019. baseline characteristics were similar between the two groups. in 2018 only 41.6% of the patients spent less than 5 hours and 29.4% spend more than 10 hours in moderate to severe pain. the 2019 data showed significant improvement in that 79.5% of patients spent less than 5 hours and only 5% patients who spent more than 10 hours in moderate or severe pain. (p <0.0001, chi square) ( figure 1 ). only 9% of the patient needed rescue medications. 3% of time was the protocol inadequate necessitating other approach. introducing an assessment driven, stepwise, protocolized pain management significantly improved patient satisfaction by reducing pain intensity in the first 24h on our cardiothoracic intensive care unit. introduction: proximal femur fractures are most common fractures in the elderly and associated with significant mortality and morbidity, with high economic and social impact. perioperative pain management influence outcomes and mortality after surgery with early mobilization being possible [1, 2] . the goal of the study was to compare the efficacy and safety of the compartment psoas block for perioperative analgesia in elderly patients with proximal femur fractures. the randomized controlled study was held in medical center "into-sana" (odesa, ukraine) from january 2018 till july 2019. patients with proximal femur fractures and older than 60 years were included in the study. they were randomly allocated to 2 groupscompartment psoas block group (bupivacaine analgesia was started as soon as possible before surgery and prolonged during and after surgery with additional ischiadicus block before surgery) and general (inhalational) anesthesia with systemic analgesia perioperatively. results: 60 patients were included in this study. perioperative compartment psoas block was associated better pain control, decreased opioid consumption, better sleep quality, earlier mobilization after surgery, decreased incidence of opioid-associated vomiting/nausea and myocardial injury. there were no difference in the incidence of hospital acquired pneumonia and delirium. perioperative compartment psoas block is effective and safe for perioperative analgesia in elderly patients with proximal femur fractures, and is associated with better pain control and decreased complications incidence. parenteral olanzapine is frequently used in combination with parenteral benzodiazepines for hospitalized patients with severe agitation. the fda issued a warning for increased risk of excessive sedation and cardiorespiratory depression with this combination based on post-marketing case reports with overall limited quality of evidence [1] . the purpose of this study is to evaluate the safety and efficacy of concomitant parenteral olanzapine and benzodiazepine for agitation. this retrospective chart review evaluated agitated patients who received concomitant parenteral olanzapine and benzodiazepine within 60 minutes from 1/31/2016 to 9/1/2019 . the primary end points were rate of respiratory depression requiring mechanical ventilation and hypotension requiring vasopressors. the secondary end points were percentage of patients requiring additional sedatives for agitation during the same time frame, cumulative dose of olanzapine and benzodiazepine (midazolam equivalent) received, and rate of cardiac arrest and death. a total of 208 patients were included with notable baseline characteristics: median age of 35 years old, 59% with a history of substance abuse, and 40% with a history of psychiatric illness. for the primary outcomes, 2.9% of patients required mechanical ventilation and 0% required vasopressors. additionally, 27.9% patients received additional sedating agents to control agitation. refer to table 1 for more details. no cardiac arrests or deaths were observed. concomitant use of parenteral olanzapine and benzodiazepine within 60 minutes for the treatment of agitation appears to have a small risk of respiratory depression without significant hypotension. hip fracture is very common in the elderly,it causes moderate to severe pain often undertreated. ficb is a simple safe method, easy to learn and use. the aim of our study is to assess the efficacy and safety of preoperative ficb compared with intravenous analgesia for elderly patients with femoral fracture and hip surgery in terms of opioid consumption and perioperative morbidity methods: after informed consent obtained,54 patients 50-98 yo asa i-iii with hip fracture were randomized to receive either an us guided ficb(40 ml of ropivacaine 0,35%) or a sham injection with normal saline 30' before surgery. both groups were operated under general anesthesia. postoperative analgesia was done according to vas: vas 0-30 mm, paracetamol 1g iv at 8 h, vas 30-60 mm, ketoprofen 100 mg iv at 8 h, vas>60, morphine 0,1mg/ kgbw iv. the primary outcome was the comparison of vas score at rest over the first 30'following the procedure, at the end of the surgery and at 6h intervals for 24h. the secondary outcome were the incidence of the cardiovascular events, of the ponv and of the confusion episodes, the amount of morphine consumption for 24h results: at baseline, ficb group (a) had a lower mean pain score than the sham injection group (b). the same difference was observed over 24 h of follow-up (p<0.05). there was a significant difference between the two groups in total cumulative iv morphine consumption at 24 h and in the incidence of ponv and confusion episodes ( figure 1 ). ficb provides effective analgesia for elderly patients suffering from hip fractures, with lower morbidity and lower opioid consumption compared with intravenous analgesia. pain assessment in chronic disorders of consciousness patients with ani monitoring e kondratyeva, m aybazova, n dryagina almazov national medical reseach centre, minimally conscious research group, st petersburg, russia critical care 2020, 24(suppl 1):p166 pain and suffering controversies in doc to be debated by the scientific, legal and medical ethics communities. methods: ani (anti nociception index) monitor was used to assess pain in patients with chronic disordersof consciousness (doc) age range 22 to 56 years -9 in vegetative state/ unresponsive wakefulness syndrome (vs/uws) and 20 minimal consciousness state (mcs). average age: in mcs group 31,8±11,29 and 42,1±8,46 in vs/uws group. neurological status was assessed using crs-r scale. the average score on the crs-r scale was 5±1.4 in vs/uws and 10.45±4.5 in mcs. pressure on the nail phalanx was used as a pain impulse. ani and nociception coma scale was evaluated before the application of pain stimulus, immediately after and past 30 minutes. prolactin level was measured before the pain stimulus application and 10 minutes after. ani less than 50 indicates pain, 70-100 hypoalgesia, 30 severe pain. the mean value of the ani in mcs patients: before the pain stimulus 66.25±14.11, after the pain stimulus application 45±16.12 and 30 minutes later 66.55±18.13. prolactin level in mcs patients before pain 13.01±9.06 ng/ml; after pain 13.75±8.73 ng/ml (p>0.05). prolactin in vs/uws patients before pain 10.79±7.2 ng /ml, after pain 14.5 ±8.88 ng / ml (p> 0.05). conclusions: ani monitor revealed that vs/uws and mcs patients react equally to the pain impulse. prolactin dynamics showed poor statistical mean and can not be consider as a marker of nociception in this group of patients. it is possible that the level of pain impulse was insufficient neuroendocrine response activation or the increase of prolactin level occurs in the long term (more than 10 minutes). in all patients the total hip arthroplasty tha is one of the most common major surgical procedures associated with significant postoperative pain that can adversely affect patient recovery and could increase morbidity. effective perioperative pain management allows an accelerated rehabilitation and improve the functional status of these patients. multimodal analgesia mma combines analgesics with different mechanism of action which by synergistic and additive effects enhance postoperative pain management and reduce complications. the aim of our study is to assess if perioperative association of very low dose of ketamine, a potent nmda antagonist and dexamethasone, by antiemetic and antiinflammatory properties could decrease opioid consumption and postoperative morbidity of patients with tha. after informed consent, 58 patients scheduled for primary hip joint replacement surgery aged 55-91 yo asa i-iii were prospective randomized in two groups. both groups were operated under general anesthesia fentanyl/sevoflurane. supplementary, patients in group a received 12 mg iv dexamethasone and 8mg at 12 h and ketamine 10 mg iv bolus at induction and 10mg/h iv during surgery. postoperative analgesia was done according to vas, 0-30 mm paracetamol 1 g iv at 8 h, 30-60 mm ketoprofen 100 mg iv at 12h, vas>60 mm morhine 0,1 mg/kgbw iv. we recorded perioperative opioid consumption, the number of intraoperative cardiac events, vas score at the end of surgery and at 24 h, the incidence of ponv and persistance of chronic pain at 3 months. we obtain a significant less pain score at the end of surgery p<0.05 in group a, no significant difference at 24 h, a significant less chronic pain at 3 months, a fewer npvo and cardiovascular events in group a, p<0.05 ( figure 1 ). a multimodal approach with very low doses of ketamine and dexamethasone could be efficent in the treatment of pain for elderly patients with hip arthroplasty, decreasing postoperative side-effects and reducing chronic pain persistance. introduction: treatment in an intensive care unit (icu) often necessitates uncomfortable and painful procedures for patients. chronic pain is becoming increasingly recognized as a long term problem for patients following an icu admission [1] . throughout their admission patients are often exposed to high levels of opioids, however there is limited information available regarding analgesic prescribing in the post-icu period. this study sought to examine the analgesic usage of icu survivors pre and post icu admission. methods: 183 patients enrolled in a post-intensive care programme between september 2016 and june 2018. intensive care syndrome: promoting independence and return to employment (ins:pire), is a 5-week multicentre, multidisciplinary rehabilitation programme for icu survivors and their caregivers. patients' level of analgesia was recorded pre-admission and upon attending ins:pire, their level of prescribed analgesia was categorized using the word health organisation (who) analgesic ladder [2] . results: 33.3% of patients (n=61) were prescribed regular analgesia preadmission; this increased to 60.7% (n=111) post-admission, representing a significant absolute increase of 27.4% (95% ci: 20.2% -34.4%, p<0.001) in the proportion of patients who were prescribed regular analgesia pre and post icu. in addition, pre-admission, 22.4% (n=41) of patients were prescribed a regular opioid (step 2 and 3 of the who ladder) compared to 38.7% (n=71) post-admission, representing an absolute increase of 16.3% (95% ci: 9.8% -22.8%, p< 0.001). this study found a significant increase in analgesic usage including opioids in icu survivors. follow-up of this patient group is essential to review analgesic prescribing and to ensure a long term plan for pain management is in place. introduction: pain, agitation, and delirium (pad) are commonly encountered b patients in the intensive care unit (icu). delirium is associated with adverse outcomes, including increased mortality and morbidity. clinical guidelines suggest that routine assessment, treatment and prevention of pad is essential to improving patient outcomes. despite the well-established improvements on patient outcomes, adherence to clinical guidelines is poor in community hospitals. the aim of this quality improvement project is to evaluate the impact of a multifaceted and multidisciplinary intervention on pad management in a canadian community icu. a pad advisory committee was formed and involved in the development and implementation of the intervention. the 4-week intervention targeted nurses (educational modules, visual reminders), family members (interviews, educational pamphlet, educational video), physicians (multidisciplinary round script), and the multidisciplinary team (poster). an uncontrolled, before-and-after study methodology was used. adherence to pad guidelines in the assessment of pad by nurses was measured 6 weeks pre-intervention and 6 weeks post-intervention. data on 430 patient-days (pd) and 406 pd were available for analysis during the pre-and post-intervention, respectively. the intervention significantly improved the proportion of pd with assessment of pain and agitation at least 4 times per 12-hour shift from 68.0% to 87.5% and from 69.7% to 82.2%, respectively ( figure 1 ). proportion of pd with delirium assessment at least once per 12-hour shift did not significantly improve. a multifaceted and multidisciplinary pad intervention is feasible and can improve adherence to pad assessment guidelines in community icus. quality improvement methods that involve front-line staff can be an effective way to engage staff with pad. oversedation introduction: sedation is a significant part of medical treatment in icu patients. a too deep sedation is associated with a longer time of mechanical ventilation, lung injury, infections, neuromuscular disease and delirium, which can lead to a longer duration of icu hospitalization, as well as an increase of morbility and mortality. many patients spend a considerable amount of time in a non-optimal sedation level. a continuous monitoring system of the sedation level is therefore necessary to improve clinical evaluation. our goal was to evaluate the incidence of non-optimal sedation (under and over sedation) comparing the parameters expressed from ngsedline with clinical evaluations and to correlate oversedation and the incidence of delirium. we have studied a cohort of patients admitted to the icu of spedali civili of brescia university hospital requiring continuous sedation for more than 12 hours. in addition to standard monitoring, the patients have been studied using next generation sedline (masimo). sedation depth was evaluated through rass scale and the presence of delirium was evaluated with cam-icu scale. we collected data from 50 adult patients. our data showed high incidence of oversedation. 45 of our 50 patients had a sr>2 and 48 had a psi level<30. a logistic regression analysis was performed and it showed statistically significant association between incidence of delirium and the age of the patients (p 0.009). the association between delirium incidence and suppression rate time was at the limits of statistics significance (p 0.059) and was statistically significant for non neurocritical patients (p 0.049). our study didn't show an association between delirium and the total time of sedation. non-optimal sedation is an unsolved problem in icu, affecting lot of patients, with a major incidence of over-sedation compared to under-sedation. our study shows an association between sr levels and the incidence of delirium. predictors of delirium after myocardial infarction, insights from a retrospective registry m jäckel, v zotzmann, t wengenmayer, d dürschmied, c von zur mühlen, p stachon, c bode, dl staudacher heart center freiburg university, department of cardiology and angiology i, freiburg, germany critical care 2020, 24(suppl 1):p172 delirium is a common complication on intensive care units. data on incidence and especially on predictors of delirium in patients after acute myocardial infarction (mi) are rare. by analyzing all patients after acute mi, we aim to identify incidence and potential risk factors for delirium. in this retrospective study, all patients hospitalized for acute mi treated with coronary angiography in an university hospital in 2018 were included and analyzed. incidence of delirium within the first 5 days of care attributed to the mi and was defined by a nudesc score ≥2, which is taken as part of daily care three times a day by especially trained nurses. this research is authorized by ethics committee file number 387/19. results: 626 patients with acute mi (age 68.5±13.3 years, 260 stemi, mortality 3.4%) were analyzed. delirium occurred in 70 (11.2%) patients and was associated with a longer hospital stay (12±15.9d vs 5.5±4.3d, p< 0.001). patients with delirium were significantly older than patients without (76.3±11.14 vs. 67.5±13.10 years, p<0.001) and had more often preexisting neurological diseases (24.3% vs. 10.8%, p<0.001) and dementia (11.4% vs. 1.4%, p<0,001). multivariate logistic regression analysis suggested that odds ratio for delirium was higher in patients after resuscitation or 7.5 (95% ci 2.1-26.7), preexisting dementia or 28.9 (ci 3.1-268) and in patients with alcohol abuse or 18 (ci 2.7-120). while maximum lactate was also connected to delirium or 1.4 (ci 1.1-1.9), infarct size or type had no effect on the incidence of delirium. in patients with mi, delirium is frequent. incidence is associated with clinical instability and preexisting neurological diseases rather than infarct size. incidence and risk factors of delirium in surgical intensive care unit ma ali, b saleem aga khan university, anaesthesia, karachi, pakistan critical care 2020, 24(suppl 1):p173 introduction: delirium in the critically ill patients is common and distressing. the incidence of delirium in the icu ranges from 45% to 87%. although delirium is highly common among intensive care patients, it is mostly underreported. to date, there have been limited data available related to prevalence of delirium in surgical patients. in a study published in 2008, the risk was observed 73% in surgical and trauma patients [1] . the purpose of this study was to find out the incidence and associated risk factors of delirium in surgical icu (sicu) of a tertiary care hospital. we conducted prospective observational study in patients with age more than 18 years and who were admitted to the surgical icu for more than 24 hours in aga khan university hospital from january 2016 to december 2016. patients who had preexisting cognitive dysfunction or admitted to icu for less than 24 hours were excluded. delirium was assessed by intensive care delirium screening checklist icdsc. incidence of delirium was computed and univariate and multivariable analyses were performed to observe the relationship between outcome and associated factors. delirium was observed in 19 of 87 patients with an incidence rate of 21.8%. multivariable analysis showed that copd, pain >4 and .76] were also the strongest independent predictors of delirium while analgesics exposures was not statistically significant to predict delirium in multivariable analysis. delirium is significant risk factor of poor outcome in surgical intensive care unit. . there was an independent association between pain, sedation, copd, hypernatremia and fever in developing delirium delirium is an acute mental syndrome which may cause negative consequences if it is misdiagnosed [1, 2] . the aim of this study was to determine the incidence of delirium in different intensive care units and reveal the risk factors. the study was performed with 212 patients hospitalized in intensive care units of anesthesia, neurology and general surgery departments. written informed consent was obstained from patients or relatives. delirium screening test was performed twice daily with camicu (confusion assessment method for the icu). patients who met the study criterias, were evaluated for the possible risk factors of delirium and the data was recorded daily. patients were reevaluated after the treatment. the incidence of delirium was 32.5%. delirium was found to increase with the length of stay (p <0.001). the mean age of the patients with delirium was 67.46. this was higher than the patients without delirium (52.48) (p<0.001). visual impairment (p<0.001), hearing impairment (p=0.001), educational status (p=0.024), hypertension (p=0.002), mechanical ventilation (p = 0.001), oxygen demand (p=0.002), midazolam infusion (p=0.025), propofol infusion (p=0.042), infection (p < 0.001), sofa (p = 0.001), apache ii (p <0.001), nasogastric catheter (p=0.012), aspiration (p <0.001), number of aspirations (p<0.001), enteral nutrition (p<0.025), albumin (p=0.025), steroid (p=0.024), hypercarbia (p=0.039) hypoxia (p=0.039), sleep disturbance (p<0.001) were found risk factors for delirium. oral nutrition (p<0.001) and mobilization (p=0.003) were found to prevent delirium development. various factors are important in the development of delirium. these risk factors should be considered in reducing the incidence of delirium in intensive care units. ). an unplanned and brutal stop of alcohol consumption, as it can occur during icu admission, may lead to an alcohol withdrawal syndrome (aws). the most severe clinical manifestation of aws is described as delirium tremens (dt). there are no current guidelines available for aws treatment in icu. the study's aim was to describe the clinician's practices for dt treatment and the outcome of dt in icu patients. observational retrospective cohort study in two icus of a universityaffiliated, community hospital in france. patient diagnosed for dt during their icu stay, as defined by dsm-v classification, were enrolled in the study. results: 61 patients with dt were included between 2014 and 2017. benzodiazepines was administered to 23% of the patients in order to prevent an aws. as associated measures, vitamin therapy was administered to 83% of the patients and 59% had an increased fluid intake (mean 2.5l+/-0.7). concerning the curative approach of aws, the treatment's heterogeneity was notable. there was a high frequency of treatment's association (66% of the patients), every patient had benzodiazepines and the use of second line treatments such as neuroleptic, alpha-2 agonist, propofol was variable ( figure 1 ). complications of dt were the following: need for mechanical ventilation due to unmanageable agitation or acute respiratory distress (33% of the patients) self inflicted injuries such as pulling out of central lines, tubes, surgical drain (46%) falls (7%). seizures (33%). delirium tremens is a severe complication of an untreated aws, which can lead to serious adverse events in icu. the current lack of evidence concerning the management of aws in icu probably explains the heterogeneity of treatments. given the potential severity of aws in icu, further evidences are required to optimize care of aws in icu patients. the incidence and related risk factor of delirium in surgical stepdown unit s yoon 1 , s yang 2 , g cho 2 , h park 2 , k park 2 , j ok 2 , y jung 2 1 asan medical center, nursing department, seoul, south korea; 2 asan medical center, seoul, south korea critical care 2020, 24(suppl 1):p177 step down units (sdus) provide an intermediate level of care between the icu and the general medical-surgical wards. the critically ill patients who are in recovery after long-term intensive care or who require monitoring after acute abdominal surgery are admitted to sdus. delirium in critically ill patient is common and leads to poor clinical outcomes. it is, however, preventable if its risk factors are identified and modified accordingly. to determine risk factors associated with delirium in critically ill patients to admitted surgical sdu at asan medical center. this is retrospective study conducted on critically ill patients who were admitted to the sdu from september 2018 to april 2019 and able to express themselves verbally. delirium status was determined using the short-cam tool. data were analyzed by spss 13.0 software, using t-test, fisher's exact test and logistic regression. the incidence of delirium was 32.1%(25 of 78 patients) and hypoactive delirium(12case, 48.0%) was the most commonly assessed, followed by hyperactive delirium(9case, 36.0%), mixed type(4case, 5.1%). risk factors associated with developing delirium identified from univariate analysis were age(p=0.048), admission via icu (p= 0.041), tracheostomy (p=0.037), chronic heart failure (chf) (p=0.017), invasive hemodynamic monitoring (p=0.041), heart rate (p= 0.037). after adjusted in multivariate analysis; factors those remained statistically significant were old age (rr we identified risk factors consistently associated with incidence of delirium following admitted to surgical sdu. these factors help to focus on patients at risk of developing delirium, and to develop preventive interventions that are suitable for those patients. patients with sepsis frequently develop delirium during their intensive care unit (icu) stay, which is associated with increased morbidity and mortality. the prediction model for delirium in icu patients (pre-deliric model) was developed to facilitate the effective preventive strategy of delirium [1] . however, the pre-deliric model has not yet been validated enough outside europe and australia. the aim of this study is to examine the external validity of the pre-deliric model to predict delirium using japanese cohort. this study is a post hoc subanalysis using the dataset from previous study in nine japanese icus, which have evaluated the sedative strategy with and without dexmedetomidine in adult mechanically ventilated patients with sepsis [2] . these patients were assessed daily throughout icu stay using confusion assessment method-icu. we excluded patients who were delirious at the first day of icu, were under sustained coma throughout icu stay and stayed icu less than 24 h. we evaluated the predictive ability of the pre-deliric model to measure the area under the operating characteristic curve. calibration was assessed graphically. of the 201 patients enrolled in the original study, we analyzed 158 patients in this study. the mean age was 69.4 ± 14.0 years and 99 patients (63%) were male. delirium occurred at least once during their icu stay in 63 patients (40%). to predict delirium, the area under the receiver operating characteristics curve of the pre-deliric model was 0.60 (0.50 to 0.69). graphically, the prediction model was not well-calibrated ( figure 1 ). to predict delirium in japanese icus, we could not show the well discrimination and calibration of the pre-deliric model in mechanically ventilated patients with sepsis. introduction: delirium is a serious and common complication and in some cases it treatment is difficult. aim of the study was an evaluation of the prevalence, structure of delirium and efficacy of dexmedetomidine and haloperidol sedation in geriatric patients after femur fracture. after local ethic committee approval 207 case-records of geriatric patients with femur fracture in the period from 2017 to 2018 in the institute of traumatology and orthopedics in astana were analyzed. patients was divided for 2 groups: in dpatients with delirium treated by i/v dexmedetomidine (0.2-1.4 mkg/kg per hour), in g group patients with delirium treated by i/v galoperidol (0.10-0.15 mkg/kg). delirium was assessed by rass at day of permission and every day at 8 a.m. the prevalence, structure of delirium and efficacy of sedation were analysed. results: by anthropometric and gender characteristics of the group did not differ. the average age in the d-group with delirium was 81.8±0.9 years old, which was comparable to the g-group -79.7±0.7 years old (p = 0.06). all study participants had similar comorbidities. delirium in all patients debuted at 2.0±1.4 days, with an average duration of 2.2±1.2 days. the effect of dexmedetomidine was better and expressed in 52% decrease in the duration of delirium in compare to haloperidol (p <0.05). dexmedetomidine provided a more controlled and safe sedation compared with haloperidol. the average consumption of narcotic analgesics in the subgroup with dexmedetomidine was two times less than in the subgroup with haloperidol. thus, the average consumption of trimeperidine hydrochloride in patients of group d was 6.9 mg versus 14.1 mg in group g (p = 0.004). in gerontological patients with femur fracture treatment delirium by dexmedetomidine was more effective in compare with haloperidol. when using dexmedetomidine, the consumption of narcotic analgesics in postoperative period was 50% less than with haloperidol. live music therapy in intensive care unit mc soccorsi 1 , c tiberi 1 , g melegari 1 , j maccieri 2 , f pellegrini 2 , e guerra 1 intensive care units (icu) are not comfortable for patients, relatives or next of kin. in the last years many news approaches were described to implement the humanization of medical treatments. the positive effect of music therapy in icu is well described, especially reducing delirium risk [1] . the aim of this paper is describing the effect in patients and their family of a music live performance in icu. after ethical committee approval (procedure aou 0018356/18, italy) for three months (november 2018-january 2019) patients in icu were treated twice a week with live music therapy performed by coral vecchi-tonelli of modena, italy (fig. 1 ). data were collected all awake and conscious patients. vitals parameters, gcs, raas and cam icu were collected before, during and after the treatment, at every performance. after the treatment a feedback questionnaire were given to patients and to next of kin. results: 18 subjects were enrolled in the research with mean age of 66.66 years old, delirium rate before the treatment was 16.38% later 15.38%, raas does not show any difference. over 85% of patients were satisfied, and relatives felt less anxiety. we recorded also a satisfaction also in relatives not enrolled. the study does not demonstrate a delirium risk reduction for the small sample and the length treatment, anyway it was recorded a low delirium rate. the safety and the potential effect of music therapy are well known, surely the research underlines the feeling of patients and their next of kin: icu is the most stressful setting for admitted patients and its humanization is a current topic for medical literature. live performances could be an entertainment moment and probably create a moment of an interaction among patients, their family and medical and nurse: icu become more human. the high level of satisfaction push us to continue this experience. introduction: patients undergoing medical procedures benefit from distraction techniques to reduce the need for drugs alleviating pain and anxiety. this study investigates if medical hypnosis or virtual reality glasses (vrglasses) as adjuvant method reduces the need for additional drugs. in a prospective, randomized, interventional trial, patients undergoing procedures were stratified in four age groups, and randomly assigned into three arms by means of a closed envelope system. all patients received standard care for pain before the procedure; the control group received further drugs for pain and stress as indicated by the visual analog scale (vas; threshold 3/10) and comfortscore (threshold 14/30), two index groups received either medical hypnosis or vr glasses as a plus before and during the procedure. vas and comfort were scored continuously and analysed with the kruskal-wallis test. patients, parents and healthcare providers scored their satisfaction at the end. of 104 included patients 6 to 86 years old, 47% were female. regardless of age, pain and comfort scores were similar before and at the start of the procedure (vas 3.7-4.2; comfort 16-16.7), but as of one minute after starting the procedure, both vas and comfort reduced significantly more in both index groups compared to the control (p< 0.001), remaining far below the threshold for both pain and stress ( figure 1 ). there was no advantage of one index group over the other (p=0.43). there were no adverse effects. patients in the vr group were more satisfied than in the standard group (p=0.02) or in the hypnosis group (p=0.04). there was no significant difference in satisfaction of parents or healthcare providers. from the very start of the intervention, the application of either medical hypnosis or vr glasses significantly reduces pain and anxiety in patients undergoing medical procedures. more studies are needed but both are promising safe adjuvant tools to standard pharmacological treatment. music to reduce pain and distress due to emergency care: a randomized clinical trial ne nouira, i boussaid, d chtourou, s sfaxi, w bahria, d hamdi, m boussen, m ben cheikh mongi slim academic hospital, emergency department, tunis, tunisia critical care 2020, 24(suppl 1):p182 recent clinical studies have confirmed the benefits of music therapy in managing pain and improving quality of care in the emergency department. the aim wasto evaluate the impact of receptive music therapy on pain and anxiety induced by emergency care methods: a randomized controlled study in patients consulting the emergency department. two groups: the music therapy group; patients needed venous sampling, peripheral venous catheter or arterial catheter. will bless ten minutes music therapy by headphones and a second control group of patients with the same care without music therapy. consent was requested from all participants. the level of pain caused by the act of care was assessed by visual analogic scale. heart rate, blood pressure and the mood of the patient were assessed before and after emergency care. we assessed patient satisfaction, adverse events. patients admitted to the emergency room, patients with communication difficulties and non-consenting patients were not included results: two hundred and forty patients were included randomized in both groups, 123 with music therapy and 117 without music therapy, the results showed comparable characteristics between the two groups: demographic data, pathological history, and initial clinical presentation. after the session of music therapy a difference was noted in the evaluation of the mean vas who was in the group with music of 2.87 ± 1.82 versus 3.60 ± 2.06 in the control group p< 0.004 ci 95% [-1.23; -0.23], and the mean of diastolic blood pressure which was 71, 07 mmhg in the first group against 74.27 mmhg for the control group p = 0.048 ci 95% [-6.37; -0.25]. as for the mood, the patients were more smiling after the act of care in the group music therapy. all patients were satisfied with their experience and 93% recommend this therapy to their relatives . music therapy may reduce pain and anxiety in patients during emergency care. the music therapy is the intervention of music and/or its elements to achieve individual goals within a therapeutic.the music has proved to have positive physiological and psychological effects on patients [1] . patients admitted to the intensive care unit (icu) experience anxiety and stress even when sedated, negatively influencing recovery [2] . methods: two groups are established, a music therapy group (mg) and a control group (cg). the first one undergoes music therapy interventions, it consists of 10-minutes sessions of live music. patients of the gc will receive the usual treatment established by the service protocol for weaning management and the data are collected during the same time interval. data collection includes mean arterial pressure (map), heart rate (hr), respiratory rate (rr), oxygen saturation (sao2) and temperature (t). a total of 28 patients were recruited, of which 6 patients had to be excluded for meeting any of the exclusion criteria (n=22). of which 13 (n=13) were randomized in the gm and the rest to the gc (n=9) ic95%. regarding delirium in gm 9 (40.9%) presented a positive cam-icu, while in the cg were 13 (59.9%) (p=0.09). when analyzing the variables in the cg and gm, it was observed that there were no differences with respect to hr, rr and map variable ( figure 1 ). according to the results, we can say that music therapy as a nonpharmacological strategy for management of anxiety and delirium in patients of critical care units, might be an useful tool for the management of patients in weaning of mechanical ventilation introduction: coagulopathy and basopenia are common features of anaphylaxis, but the role of coagulopathy in anaphylaxis remains uncertain. the aim of this study is to evaluate the association between coagulopathy and clinical severity or basopenia in patients with anaphylaxis. we conducted a single-center, retrospective study of patients with anaphylaxis about their coagulopathy. levels of fibrin degradation products (fdp) and d-dimer were analyzed with the cause of anaphylaxis, clinical symptoms, medications and outcomes. we also studied the levels of intracellular histamine as a biomarker of basophil degranulation in the peripheral blood in relation to fdp and ddimer. in total, sixty-nine patients were enrolled to the study, and the levels of intracellular histamine were analyzed in 14 patients. the symptoms included respiratory failure (n=47), shock (n=56), abdominal impairment (n=20), and consciousness disturbance (n=37). thirty-two patients needed continuous intravenous vasopressors for refractory shock. the increase of fdp was significantly associated with consciousness disturbance (p=0.029) and refractory shock (p<0.0001). the increase of d-dimer was also significantly associated with refractory shock (p=0.0066). there was no correlation between the levels of intracellular histamine and either of fdp or d-dimer (p=0.13 and p=0.16, respectively). the increase of fdp and d-dimer were associated with severe symptoms of anaphylaxis, while they were not correlated with intracellular histamine. these results suggest that anaphylaxis is closely associated with coagulopathy in a mechanism which is different from basophile degranulation in anaphylaxis. cardiac manifestations of h1n1 infection in a greek icu population e nanou 1 , p vasiliou 1 , e tsigou 2 , v psallida 1 , e boutzouka 2 , v zidianakis 1 , g fildissis 1 1 agioi anargiroi hospital, attiki, greece; 2 agioi anargiroi hospital, icu, attiki, greece critical care 2020, 24(suppl 1):p185 introduction: cardiovascular involvement in influenza infection occurs through direct effects on the myocardium or through exacerbation of pre-existing cardiovascular disease [1] . the aim was to study cardiac manifestations in all pts admitted to the icu with severe influenza's attack. clinical, laboratory, electrocardiographic, echocardiographic and hemodynamic data were retrospectively recorded in all pts admitted to the icu due to influenza infection (winter 2018-spring 2019). diagnosis was established by pcr on bronchial aspirates the next 7 days after admission. myocardial injury was defined by troponin levels >116 pg/ml (10 fold uln). left ventricular systolic dysfunction was defined as ef <50% and was characterized as either global or regional. hemodynamic monitoring by fig. 1 (abstract p183) . comparison between mg and cg transpulmonary thermodilution method (picco) was recorded in pts with shock (norepinephrine >0.1 μg/kg/min). values are expressed as mean±sd or as median (ir). results: nine pts (5 males) with a mean age 49.78±17.01 years, apache ii 19±5.29 and sofa score 10.50±2.93 were assessed. icu admission was due to ards (7) and copd exacerbation (2) . icu los was 24.44± 14.19 days and mortality rate was 18%. no history of vaccination or coronary heart disease was referred. results are shown in table 1. levosimendan was administered in 2 pts with severe cardiogenic shock. in all survivors, shock and indices of myocardial dysfunction subsided till discharge. coronary angiography was performed in 1 pt showing no abnormalities. mortality was attributed to septic shock and multi-organ failure. myocardial involvement, though common in influenza pts admitted to the icu, didn't contribute to a dismal prognosis. the cardioprotective effects of levosimendan could be related to the modulation of oxidative balance. we aimed to examine the effects of levosimendan in patients with cardiogenic shock or with ejection fraction (ef) lower than 30% on cardiac systo-diastolic function and plasma oxidants/antioxidants (glutathione, gsh; thiobarbituric acid reactive substances, tbars). in 4 patients undergone coronary artery bypass grafting or angioplasty, cardiovascular parameters were measured at t0 (before the beginning of levosimendan, 0.1mcg/kg/min), t1(1 h after the achievement of the therapeutic dosage of levosimendan), t2 (at the end of levosimendan infusion), t3 (at 72 h after the end of levosimendan infusion), t4 (at the end of cardiogenic shock). the same time-course was followed for plasma gsh and tbars measurements. we found an improvement in cardiac output, cardiac index and systolic arterial blood pressure. ef increased from mean 25% to 45%. a reduction of central venous pressure and wedge pressure was also observed. moreover, indices of diastolic function were improved by levosimendan administration (e/e' from 14 to 6; e/a from >1 to <1) at early t2. it is to note that an improvement of gsh and tbars was observed early after levosimendan administration (t1), as well ( figure 1 ). the results obtained have shown that levosimendan administration can regulate oxidant/antioxidant balance as an early effect in low cardiac output patients. the modulation of oxidative condition could be speculated to play a role in exerting the cardio-protection exerted by levosimendan in those patients. table 1 . early administration of vasopressors and their use in the emergency department was associated with survival in septic shock. this seemed to be independent of median map recorded in the ed. we excluded all the traumatic or post-myocardial infarction forms. out of 83 patients, the tuberculous etiology was identified in 15 cases (18,1%), mean age was 34 years, 57,8% were men. 9 patients reported a tb contact in their environment, 5 had a medical history of pulmonary tb. after pericardiocentesis, the liquid was citrine yellow in 6 cases and hematic in 5 patients, no patient underwent surgical drainage in our serie. mycobacterium tuberculosis was found in the expectorations in 4 cases and ada was positive in 4 patients. hiv serology was negative in all our patients. a 6 months anti bacillary therapy with isoniazid, rifampin, pyrazinamide, and ethambutol was initiated in all our patients with a good evolution in 7 cases, 2 deaths, 1 chronic constrictive pericarditis, 2 small pericardial effusion and 3 lost to follow-up. althought cardiac tamponade is rarely caused by tuberculosis, this condition remains common in endemic countries such as morocco and affect younger population, hence the importance of a better knowledge of its prevalence and and multidisciplinary management and more importantly the treatment of the underlying cause using combined antibacillary medication that has shown satisfying results. 1. the main perceived limiting factor is the absence of a standardized didactic program, followed by mentor's availability in residents' perception and by mentor's experience in consultants' one. pocus teaching is present although not optimal and not homogenous in italian acc residency schools. standardisation of residents' ultrasound curriculum is suggested to improve ultrasound teaching. the study included a convenience sample of critically ill patients with supradiaphragmatic cvcs and a cxr for confirmation. us is used for direct confirmation of the guidewire in the internal jugular (ijv) or subclavian (scv) vein and visualizing the guidewire in the right atrium. to evaluate for pneumothorax, "sliding sign" of the pleura was noted on us of the anterior chest. results: 34 patients have been included, 35% of the catheters have been placed in the scv and 65% in the ijv. it was possible to confirm the position of the cvc tip for 70.6% (23 correct, 1 incorrect cxr) of 34 (figure 1 ). overall, it was not possible to identify the guide in the right atrium 11 cases (10 false negatives, 2 of them due to the presence of defibrillator leads). regarding the 1 case where an incorrect position was seen on cxr it was also detected on ultrasound: us of the inserted vein and a negative tte confirmation. in all cases it was possible to exclude a pneumothorax by us. these results show that bedside ultrasound might be a feasible technique to confirm the cvc positioning. it is important to note that the level of the operator's expertise is significant when assessing the feasibility of this method. we only had a limited sample size and the occurrence of only one misplaced catheter. these preliminary results need to be confirmed on a larger scale. central venous catheter (cvc) misplacement occurs more frequently after cannulation of the right subclavian vein compared to the other sites for central venous access. misplacement can be avoided with ultrasound guidance by using the right supraclavicular fossa view to confirm correct guidewire j-tip position in the lower part of the superior vena cava. however, retraction of the guidewire prior to the cvc insertion may dislocate the j-tip from its desired position, thereby increasing the risk of cvc misplacement. the aim of this study was to determine the minimal guidewire length needed to maintain correct guidewire j-tip position throughout an us-guided infraclavicular cvc placement in the right subclavian vein. methods: 100 adult intensive care patients with a computed tomography scan of the chest were retrospectively and consecutively included in the study. the distance from the most plausible distal puncture site of the right subclavian/axillary vein to the junction of the right and left brachiocephalic veins (= vessel length) was measured using multiplanar reconstructions. in addition, measurements of the equipment provided in commonly used 15-16 cm cvc kits were performed. the minimal guidewire length was calculated for each cvc kit. the guidewires were up to 90 mm too short to maintain correct j-tip position throughout the cvc insertion procedure in seven of nine commercial cvc kits. four of these are shown in table 1 . when us guidance is used to confirm a correct guidewire j-tip position, retraction of the guidewire prior to the cvc insertion must be avoided to ensure correct cvc-tip positioning. this study shows that most of the commonly used 15-16 cm cvc kits contain guidewires that are too short for cvc placement in the right subclavian vein. the reliability of lung b-lines to assess fluid status in patients with long period of supine introduction: ultrasound-guided cannulation is usually done using either longitudinal or transverse approach. the oblique approach utilizes advantages of both these approaches allowing visualization of the entire course of needle including tip and lateral discrimination of artery from vein [1] . the reported incidence of the complete overlap of femoral vein by the femoral artery is 8-10 percent [2, 3] . we describe the use of the oblique approach for successful cannulation of such a femoral vein which is not possible by usual approaches (figure 1 ). endothelial cells play a pivotal role in the atherogenic process. endothelial cell dysfunction (ed) is the main risk factor for cardiovascular diseases such as hypertension, coronary heart disease (chd) and peripheral occlusive disease (pod). these diseases significantly increase the risk for perioperative complications. therefore, identifying patients with ed is important and should influence our prospective perioperative strategy. however, sensitive tools to diagnose ed are still missing and do not belong to our standard of care. aim of this study was the validation of a new non-invasive method to detect ed and a correlation with a set of established an new endothelial biomarkers. the cohort includes 20 preoperative patients without anamnestic relevant cardiovascular disease and 20 patients with known peripheral occlusive disease (pod). we used non-invasive endopat® technology from itamar-medical to measure ed by changes in vascular tone before and after occlusion of the brachial artery and calculate a reactive hyperemia index (rhi). in addition, we measured established markers and alternative biomarkers potentially indicate vascular diseases such as substrates and products from the no-metabolism l-arginin, asymmetric/symmetric dimethylarginine (adma/sdma), von-willebrand factor (vwf) and sphingosine-1-phosphate (s1p). rhi was able to identify patients with pod. rhi was significant lower in patients with clinical signs and symptoms of pod (p<0.05). among other markers adma was significant higher in pod patients compared to controls and correlates with rhi. the pad technology is a helpful non-invasive functional test to measure ed and seems able in identify patients with vascular disease. in future, a combination of anamnesis, new diagnostic tools and biomarkers may further increase our sensitivity in identifying risk-patients. single-lumen 5fr and triple-lumen 6fr peripherally inserted central catheters (piccs) for cardiac output assessment by transpulmonary thermodilution s d´arrigo 1 achieving effective critical care in low-and middle-income countries is a global health goal [1] , which includes the provision of effective point of care ultrasound [2] . we sought to establish zambia's first focused critical care echocardiography training programme in a 16bedded icu at university teaching hospital, lusaka. the programme was accredited by the uk intensive care society fice programme, with teaching adapted for local disease patterns such as tuberculous pericardial effusions. parasternal, apical and subcostal windows were used to assess ventricular dysfunction, hypovolaemia, pleural effusion, alveolar interstitial syndrome and pneumothorax. zambian doctors working with critically ill patients received an intensive one-day course, followed by mentored scanning at the bedside. teaching was delivered by visiting fellows from the uk who are accredited in echocardiography and experienced ultrasound educators. patients with abnormal mean ci or hr suffer from increased hospital mortality. abnormality of mean svi was not associated with mortality. these data support accurate measurement of ci as a hemodynamic target and the normal range defined for ci. since ci also carries the hr information, ci seems to be the more important target than svi. our data cannot necessarily be interpolated to less invasive and less precise measurements of ci. an evaluative study of the novelty device with the function of auto-aspirating and pressure indicator for safety central venous catheterization ly lin, wf luo, cy tsao national taiwan university hospital, taipei, taiwan critical care 2020, 24(suppl 1):p203 previous studies have shown that 0.8% of cvc attempts resulted in arterial punctures that were not recognized by blood color. to overcome the problem, our team has developed a concept of pressure detecting syringe that can indicate the artery puncture [1] . based on previous research, different springs, the actuator of the design, have been evaluated to optimize the proposed device and reduce the risk of cvc procedure. tested devices -the inner-spring is set between the pressure indicator and plunger (fig. 1a ). three springs are tested. test condition -blood samples were simulated by glucose solution with absolute viscosities of 2 and 6 mpa-s. different blood pressures were applied to simulate the artery and vein (fig. 1b) . the response time (rt) is defined as the time required to show the indicating signal (is) which is the movement of the piston from the position in fig. 1b : a2-1 to a2-2. the rt is strongly influenced by spring (fig. 1b) but every design can show the is when pressure is higher than 50 mmhg, the assumed minimum artery pressure. the rt of s1, the strongest spring design, is about 10s in the 50mmhg-pressure and high viscosity condition. during our tests we found the user can realize the is before the position be fully changed from fig. ib : a2-1 to a2-2. thus, we believe the 10s rt, the worst case, is still acceptable. we also found the weak spring force may lead to difficulty to empty the syringe because the spring must to overcome the blood pressure and the friction between the piston and barrel. as a result, it was difficult for s3 to absolutely empty the syringe even if the blood pressure is only 30 mmhg. the spring will be compressed as fig. 1b : a2-2 and fail to push the piston when pushing the plunger forwardly, which is not acceptable in clinical use. the results indicate the feasibility of using the device to facilitate cvc and we believe the s1 or s2 are more suitable for the future application. introduction: models using standard statistical features of hemodynamic vital sign waveforms (vs) enable rapid detection of covert hemorrhage at a predetermined bleed rate [1] . by featurizing interactions between vs we can train powerful hemorrhage detectors robust to unknown bleed rates. waveforms (arterial, central venous, pulmonary arterial pressures; peripheral and mixed venous oxygen saturation; photoplethysmograph; ecg) of healthy pigs were monitored 20 min prior and during a controlled hemorrhage at 20 ml/min (n=38) and 5 ml/min (n=13). two sets of vs features were extracted: statistical features [1] and maximal pairwise cross correlations between pairs of vs within a 5s lag over various time window sizes (30s, 60s, 180s, 300s); and normalized with pre-bleed data of each given animal. for each feature set, a tree-based (ert) model [2] was trained and tested in a one-animal-out setting to mitigate overfitting on the 20 ml/min cohort, and another trained on the 5 ml/min and tested on the 20 ml/min cohort. we evaluated models with activity monitoring operating characteristics curves [3] that measure false alert rate as a function of time to detect bleeding. models using cross-correlations show no significant deterioration of performance when applied to detect bleeding at different rates than trained for, while standard models require 64s longer on average to detect hemorrhage at 1% false alert rate in the previously unknown setting ( figure 1 ). correlations between vs data encode physiologic responses to hemorrhage in a way independent of the actual bleed rates. this enables training effective hemorrhage detectors using only limited experimental data, and using them in practice to detect bleeding that occurs at rates other than used in training. we validated a dataset of 634 data lines containing hemodynamic variables and treatment options. we selected nine hemodynamic variables as inputs. furthermore, data were collected regarding underlying conditions: heart failure, septic shock, renal failure or respiratory failure or a combination. we applied datastories regression on the dataset (turnhout, belgium, www.datastories.com). six different interventions were analyzed as kpi: administration or removal of fluids, increasing or decreasing inotropes and increasing or decreasing vasopressors. finally, we elaborated and challenged 63537 predictive models to generate a decision algorithm to predict each kpi. we first looked at how each hemodynamic parameter impacts the prediction of each kpi individually and performed a standard correlation analysis as well as a more involved analysis of the mutual information content between each kpi and all other hemodynamic parameters individually. confusion matrix and variable importance was obtained for each kpi. the baseline hemodynamic parameters were: gedvi 738±218 ml/m2, evwli 11.2±5.5 ml/kg pbw, svv 14.8±8 %, mbp 77.8±16.5 mmhg, hr 94.2±23.5 bpm, ci 3.3±1.2 l/min.m2. the results of the regression analysis identified the different variables of importance for each of the different interventions ( fig 1a) . based on these results the hemodynamic variables (hr, mbp, gedvi, elwi, ci, svv) were used to develop the final hemoguide prediction model ( fig 1b) . the hemoguide app can be used to advise physicians with respect to basic therapeutic decisions at the bedside or as an educational tool for students. with the collection of new data, the accuracy of the system may grow over time. the next step of the project is to develop a more-sophisticated suite: the icu cockpit. feedback function contributes to accurate measurement of capillary refill time r kawaguchi 1 , ta nakada 2 , m shinozaki 2 , t nakaguchi 2 , h haneishi 2 , s oda 2 1 chiba university, department of emergency and critical care medicine, chiba, japan; 2 chiba university, chiba, japan critical care 2020, 24(suppl 1):p206 capillary refill time (crt) is well known as an indicator of peripheral perfusion. however, it has been reported to have an intra-observer variance, partly because of manual compression and naked-eye measurement of the nailbed color change. we hypothesized that a we developed a novel portable crt measurement device with an oled display that feedbacks weather the strength of the nailbed compression is enough and counts the time. we settled the target strength and time as 5n and 5seconds according to the study we reported before [1] . 20 examiners measured crt with and without the feedback function. the pressing strength and time during the measurement were evaluated. there was a significant difference among the pressing strength and time between the crt measurement using the device with and without the feedback function (strength: p<0.001; time: p<0.001). furthermore, intra-examiner variance was significantly reduced with the feedback function (strength: p<0.001; time: p<0.001). in all measurements without the feedback function, 41% was outside the optimal strength while the measurements with the feedback function 100% achieved the targeted range. without the feedback function, 12% could not reach the optimal time, while 100% with the feedback function did. in total, 49% of the measurements could not achieve the optimal pressing strength and time. the feedback function for crt measurements, guiding examiners to an optimal pressing strength and time, fulfilled the required measurement conditions and reduced intra-examiner variance. our novel portable device would assist an accurate crt measurement regardless of personal work experience. introduction: the aim of the study was to detect the difference of conjunctival microcirculation between septic patients and healthy subjects and evaluate the course of conjunctival microcirculatory changes in survivors and non-survivors over a 24 hours period of time. this single-centre prospective observational study was performed in mixed icu in a tertiary teaching hospital. we included patients with sepsis or septic shock within the first 24 hours after icu admission. conjunctival imaging using idf videomicroscope as well as systemic hemodynamic measurements were performed at three time points: at baseline, 6 hours and 24 hours later. baseline conjunctival microcirculatory parameters were compared with healthy control. a total of 48 patients were included in the final assessment and analysis. median apache ii and sofa scores were 16 (12-21) and 10 (7-12) respectively. 44 (92%) were in septic shock, 48 (100%) required mechanical ventilation. 19 patients were discharged alive from the intensive care unit. we found significant reductions in all microcirculatory parameters in the conjunctiva when comparing septic and healthy subjects. we found a significant lower proportion of perfused vessels and microvascular flow index (mfi) of small vessels during all three time points in non-survivors compared with survivors. in nonsurvivors we observed no significant changes in conjunctival microcirculatory parameters over time. however, survivors had significantly improved mfi of small vessels at second and third time points compared to first time point. microcirculatory perfusion in conjunctiva was altered in septic patients. over 24 hours evaluation survivors in comparison with nonsurvivors had better microcirculatory flow with incremental improvement of microvascular flow index. 16 healthy pigs were centrally cannulated for veno-arterial ecmo and precision flow probes were placed on the pulmonary artery main trunk for reference. 10ml boluses of iced 0.9% saline chloride solution were injected into the ecmo circuit and right atrium at different ecmo flow settings (4, 3, 2, 1 l/min). rapid response thermistors of standard pa-catheters in the ecmo circuit and pulmonary artery recorded the temperature change. after calibration of the catheter constants for different injection volumes in the ecmo circuit, the distribution of injection volumes passing each circuit was assessed and enabled calculation of pulmonary blood flow. analysis of the exponential decay of the signals allowed assessment of right ventricular function. calculated blood flow correlated well with true blood flow (r 2 = 0.74, p < 0.001, figure 1 panel a, individual measurements organ congestion is susceptible to be a mediator of adverse outcomes in critically ill patients. point-of-care ultrasound (pocus) is widely available and could enable clinicians to detect signs of venous congestion at the bedside. the aim of this study was to develop prototypes of congestion scores and to determine their respective ability to predict acute kidney injury (aki) after cardiac surgery. this is a post-hoc analysis of a prospective study in 145 patients for which repeated daily measurements of hepatic, portal, intra-renal vein doppler and inferior vena cava (ivc) ultrasound were performed before surgery and during the first 72 hours after cardiac surgery [1] . five prototypes of venous excess ultrasound (vexus) scores combining multiple ultrasound markers were developed (figure 1 ). the association between each score and aki was assessed using timedependant cox models as well as conventional performance measures of diagnostic testing. a total of 706 ultrasound assessments were analyzed. we found that defining severe congestion as the presence of severe flow abnormalities in multiple doppler patterns with a dilated ivc (>2cm), corresponding to grade 3 of the vexus c score, showed the strongest association with the development of subsequent aki compared with other combinations of ultrasonographic features (hr: 3. there is an increasing awareness on the consequences of fluid administration in patients leading to the development of methods that evaluate the effects of fluids loading on the cardiocirculatory system. however, most of methods used in the clinical practice investigate the effects of fluids on the cardiac function, instead of investigating those on the determinants of venous return. besides volume of fluids, the determinants of fluid loading are the blood volume distribution and the availability of vascular bed. in this study we aimed to test non-invasively the effects of fluids administration on the venular compartment in the skeletal muscle. in addition to the mean systemic filling pressure (msfp), we calculated changes in the stressed and unstressed volumes (vs, vu) and the venular bed availability. we enrolled 10 critically ill patients in our intensive care unit. we assessed volumes and pressures by the near infra-red spectroscopy on the forearm using graded venous occlusions in steps of 5mmhg from 50 to 0 mmhg. the msfp, vu and vs were measured as previously reported (microcirculation 2014; 21:606-614). the vascular bed availability was measured by changes in the volume recruited from the occlusion maneuvers. all the measures were done at baseline and after a fluid load ranging from 250 to 500 ml. values were expressed as median and interquartile range. wilcoxon test was used to compare data and a p< 0.05 was considered as significant. introduction: hypotension is a common side effect of general anesthesia (ga) and is associated with organ hypoperfusion and poor perioperative outcome [1] . post-induction hypotension (pih) is caused by the depressant cardiovascular effect of anesthetic drugs and could be amplified by hypovolemia. the aim of this study was to assess the ability of two echocardiographic fluid responsiveness markers to predict pih: the inferior vena cava collapsibility index (ivc-ci) and the velocity time integral change (δvti) after passive leg raising. sixty patients > 50 years of age and scheduled for elective surgery were included. ivc-ci and δvti were measured before ga induction. anesthesia protocol, fluid infusion and vasopressor administration were standardized in all patients. pih was defined as a mean arterial pressure (map) <65 mmhg or a relative decline from pre-induction value of at least 30% within 12 minutes of ga induction. receiver operating characteristic (roc) curve analysis was used. the optimal cutoff was selected to maximize the youden index (sensitivity + specificity − 1). the measurement of ivc-ci and/or δvti were unsuccessful in seven patients (11.6%). pih occurred in 32 patients (incidence 53 %). the areas under the roc curves ( figure 1) preload responsiveness might be detected by the changes of cardiac index (δcimini) induced by a "mini-fluid challenge" (mini-fc) of 100 ml or even by the changes (δcimicro) in response to a "micro-fluid challenge" (micro-fc) of 50 ml. however, the smaller the fluid challenge, the larger the "grey zone" of diagnostic uncertainty. we tested whether (1) micro-and mini-fc monitored by calibrated pulse contour analysis detect preload responsiveness and (2) adding 50 ml when the result of a micro-fc is within the grey zone improves diagnostic accuracy. in 30 patients with circulatory failure, we infused 50 ml saline over 30s followed by 50 ml over 60s. we measured δcimicro and δcimini by the pulse contour analysis (picco2). preload responsiveness was defined by an increase in ci (δciplr) during a passive leg raising test ≥10%. diagnostic uncertainty was described by calculating the grey zone after bootstrapping. δcimicro were larger in responders than in non-responders (5. for the micro-fc, the area under the receiver operating characteristic curve was 0.975±0.03 (threshold 1%), while it was 0.955±0.03 for the mini-fc (threshold 4%). for the micro-fc, the grey zone ranged from 0.82% to 3.47% and included 9 (30%) patients. for the mini-fc, it ranged from 2.8% to 6.8% and included 9 (33)% patients, among which 6 were already in the grey zone of the micro-fc. when evaluated by pulse contour analysis, micro-and mini-fc reliably detect preload responsiveness but with a large diagnostic uncertainty. it seems that adding 50ml more fluid to a micro-fc when its result is within the grey zone does not improve the diagnostic accuracy. the study is ongoing. the starling-sv bioreactance device (cheetah medical) reliably detects passive leg raising (plr)-induced changes in cardiac index (δci). we tested whether it can also track the small and short-time δci induced by the end-expiratory occlusion (eexpo) test, and whether shortening the time over which it averages cardiac output (24 s in the commercial version) improves the detection. in 42 mechanically ventilated patients, during a 15-sec eexpo, we measured δci (in absolute value and in percentage) through calibrated pulse contour analysis (ci pulse , picco2 device) and starling-sv. for the latter, we considered both ci starling-24 provided by the commercial version and ci starling-8 obtained by averaging the raw data over 8 s. we calculated the correlation between δci pulse and both δci starling-24 and δci starling-8 , and the area under the receiver operating characteristic curve (auroc) to detect preload responsiveness, defined by a plr test. when considering absolute values, the correlation coefficient r between δci pulse and δci starling-24 was 0.362 (p=0.02), which was lower than the one between δci pulse and δci starling-8 (rr comparison). when considering percentage changes, no correlation was observed between δci pulse and δci starling-24 . conversely, the correlation coefficient between δci pulse and δci starling-8 was 0.402 (p=0.01), but it was lower than the one obtained for absolute values (p=0.04 for r comparison). eexpo-induced δci starling-8 , both in absolute values and in percentage, detected preload responsiveness with aurocs of 0.90 (sensitivity 83%, specificity 87%) and 0.89 (sensitivity 83%, specificity 79%), respectively. shortening the averaging time of the bioreactance signal increases the reliability of the starling-sv device to detect eexpo-induced δci. moreover, the accuracy of the method is increased when absolute rather than percentage changes of ci are considered. fluids are among the most prescribed drug in intensive care, particularly among patient with circulatory failure. yet, very little is known about their pharmacodynamic properties and this topic has been left largely unexplored. there is a lack of strong scientific evidence in current guidelines for fluid administration in shock. several factors may impact the hemodynamic efficacy of fluids among which the infusion rate. the aim of this study was to study the influence of fluids administration rate on their pharmacodynamics in particular by studying mean systemic pressure (p ms ). we conducted a prospective observational study in 17 patients with circulatory failure to compare two volume expansion strategies. when a patient required a fluid bolus, 500 ml of normal saline were administered and several hemodynamic parameters were recorded continuously: cardiac output (co), arterial pressure (ap), mean systemic pressure (p ms ). infusion rate was let to the discretion of the attending physician and a "slow" and a "fast" group were determined based on the median of the infusion time. fluids effect was measured by the area under the curve (auc), maximal effect (e max ) and time to maximal effect (t max ) for each hemodynamic variable. results: p ms auc was higher in the "fast" group compared to the "slow" group (p=0.043). we observed a shorter t max and a higher e max for p ms in the "fast" group compared to the "slow" group (p=0.039 and 0.02 respectively). regarding co, t max was also shorter in the "fast" group (p=0.041). auc and e max were similar between the two groups. fluid effect dissipated within 60 minutes following the end of fluid infusion for every patient in both groups. the decreasing slope from maximal effect was comparable in the groups, for p ms and co alike. the effect of a 500 ml fluid bolus in septic shock patients vanished within one hour. a faster infusion rate increased maximal effect and shortened the delay to reach it. study is ongoing. fluid management in the control arm of sepsis trials aa anparasan, ac gordon, mk komorowski imperial college london, department of surgery and cancer, london, united kingdom critical care 2020, 24(suppl 1):p219 in the past, high-volume intravenous fluid resuscitation in severe sepsis and septic shock was common. more recently, concerns over the harmful effects of this practice have led some clinicians to adopt less liberal fluid strategies. we sought to analyse temporal trends in fluid administration in the control arms of recent adult sepsis trials and assess any correlation with patient severity and mortality. a literature search was conducted to identify relevant randomized controlled trials that reported fluid administration published post 2000. we recorded 4 outcomes: total amount of iv fluid administered in the control arms of these trials between hospital admission and hour 6 and hour 72 following trial enrolment, mortality rates at the latest reported time point and apache-ii score at admission. we computed the pearson correlation coefficient and linear regression between study dates and the 4 outcomes. we identified 9 relevant trials [1] [2] [3] [4] [5] [6] [7] [8] [9] , which recruited a total of 2,444 patients in their control arms, from 1997 to 2018. the temporal analysis revealed no obvious trend in the in the total volume of iv fluid given by hour 6 following trial enrolment (correlation p=0.94) ( figure 1 ). however, the total volume of fluid given by hour 72 decreased significantly over the period of interest (r=-0.78, p=0.02). in parallel, we observed a decrease in mortality (r=-0.6, p=0.08) but there was no evidence of decrease in illness severity over time (p=0.84). we found that in published rcts over the last two decades, the amount of intravenous fluid given to patients with sepsis in the initial 6 hours did not appear to change, however less intravenous fluid was given over the first three days. upcoming large rcts will test the safety and efficacy of restrictive fluid administration approaches in sepsis. clinical practice guidelines recommend prompt intravenous (iv) fluid resuscitation for pediatric sepsis, including an initial fluid bolus of 20 ml/kg [1] . however, recent evidence is conflicting as to the effectiveness, volume, and consequences of aggressive fluid resuscitation in septic children. therefore, we sought to determine the epidemiology of early iv fluid resuscitation in an integrated health system, specifically at community hospital emergency departments (ed). we studied a retrospective cohort of pediatric patients (ages > 1 month to < 18 years) with sepsis identified in electronic health record data at 11 community eds in southwestern pennsylvania from 2010 to 2014. sepsis was defined as 1) suspected infection (combination of fluid culture collection and administration of antibiotics and 2) organ dysfunction (pediatric sofa score ≥ 1) within 24 hours of suspected infection. fluid bolus therapy was defined as electronic documentation of administration of 0.9% normal saline iv bolus within 1 hour of the time of sepsis onset. results: among 1,247 patients with pediatric sepsis, 513 (41%) received iv fluid bolus therapy within 1 hour of time of sepsis onset. the volume of fluid administered ranged from 2 ml/kg to 67 ml/kg (figure 1 , panel a), corresponding to a median volume of 20 ml/kg (iqr 17-22 ml/kg). patients who received ≥ 20 ml/kg of fluids (n = 258, 50%) were younger (mean age 5 years, sd 5 vs. 9 years, sd 6; p<0.001), more often had blood cultures collected during evaluation (86% vs. 76%, p=0.003), and were more often transferred to another facility (48% vs. 33%, p<0.001) when compared to patients who received < 20 ml/kg of fluids (n = 255, 50%). mean fluid bolus volume within 1 hour of time of sepsis onset by hospital ranged from 12 ml/kg to 24 ml/kg (figure 1, panel b) . in a cohort of community emergency departments, 41% of septic children received intravenous fluid boluses within one hour, and of those, only one half received volumes concordant with guidelines. (figure 1 ). a wide range of fluid balance exists in septic shock patients cared for in icu. trends of serum albumin in septic and non-septic critically ill introduction: the link between hypoalbuminaemia and poor outcomes in critical care is well established [1] . limited data are available on serum albumin trends during critical illness [2] . in this study we assessed trends in serum albumin for up to 7 days in both septic and non-septic critically ill patients. we retrospectively examined the records of 1107 adult patients admitted to critical care at the royal liverpool university hospital between 2008 and 2014. we then excluded patients who did not have albumin data available for the first 7 days, leaving us with 758 patients. 506 patients (66.8%) had sepsis, and of these patients 116 had died by day 28. of the 252 non-septic patients (33.2%), 40 patients had died by day 28. albumin levels were collected for 7 days from admission to critical care, in addition to other demographic and biochemical data. statistical analysis was performed using repeated measures analysis. septic patients had lower serum albumin than non-septic patients throughout the 7 day period (p<0.001). we observed a decrease in albumin by day 2 in all groups, with levels increasing over the subsequent days. there was no difference in daily serum albumin between non-septic patients who survived or died. this is the first study, to our knowledge, to compare albumin trends in septic and non-septic critically ill patients over 7 days. further research is needed to elucidate the optimal recipients and timing of albumin therapy. introduction: burn injury is characterized by marked inflammation, capillary leakage, and profound hemodynamic alterations. early albumin resuscitation is avoided fearing a paradoxical fluid escape into the interstitium. on the other hand, administration of crystalloids in massive amounts causes tissue edema and fluid extravasation, which deteriorates tissue perfusion by increasing oxygen diffusion distance. albumin administration could reduce the amount required to maintain hemodynamic stability in this population. we investigated whether albumin improves tissue perfusion and microcirculation by reducing tissue edema. this is an observational study conducted in the burn unit of maasstad hospital, rotterdam. patients with burns higher than 15% of total body surface area (tbsa) were included in the study. sublingual microcirculation was measured at admission (t0), 4(t4), and 12(t12) hours after burn injury. total vessel density (tvd) and functional capillary density (fcd) were analyzed. fluid management was calculated according to the modified parkland formula. albumin (20%) infusion was started 12 hours after the burn insult. a total of nine patients were recruited between january and december 2019. patients were included in the study after 5.7±2.3 hours of the insult with a mean tbsa of 36±22%. the amount of crystalloid infusion was 2718±3348 ml and 8501±5230 ml at t0 and t12,respectively. within the first 12h (t12) 502±386 ml albumin was given. tvd decreased from 23.6±2.2 at t0 to 20±1.3 at t4 (p<0.05) (figure 1) introduction: spontaneous bacterial peritonitis (sbp) accounts for ≥24% of the bacterial infections that occur in patients with cirrhosis, and sbp has a high mortality rate (20% to 50%). albumin infusion has been shown to improve the outcome of sbp. the aim of this study is to examine the impact of albumin infusion on hospital length of stay (los) for cirrhotic patients with sbp. we utilized a nationwide electronic health record data set (cerner health facts®) to extract real-world data on adult patients (≥18 years old) with cirrhosis and sbp who received antibiotics and admitted between january 1, 2009, and april 30, 2018. international classification of diseases (icd-9/10) codes were used to identify cirrhosis and sbp. we used laboratory data for calculation of the model for endstage liver disease sodium (meld-na) score and vital signs data for calculation of the quick sepsis related organ failure assessment (qsofa) score at baseline for each encounter. a generalized linear model was used to assess the relationship between albumin infusion and hospital los. results: there were 2,131 encounters that identified patients with sbp and cirrhosis, of which 1,661 survived hospitalization. albumin was infused within 24 hours of admission ('early albumin') in 43% (n=718), after 24 hours in 31% ('late albumin', n=517), and not administered in 26% ('no albumin', n=426). meld-na was higher at presentation in early albumin cases versus late-or no-albumin cases (mean 24.0 and 19.5). unadjusted los was lower in patients receiving early albumin (8.7 days versus 10.4 days). risk-adjusted analysis demonstrated that early albumin led to a 17.5% reduction in los (95% ci 12.6%-22.2%, p = <0.0001). in these real-world data, albumin infusion within 24 hours of admission in patients with cirrhosis and sbp was associated with a shorter hospital stay despite more severe illness. early albumin may not only improve clinical outcomes but may also reduce the costs of hospitalization in cirrhotic patients with sbp. early albumin use in patients with septic shock is associated with a shorter hospital stay: real-world evidence in the united states introduction: septic shock is among the most common critical care illnesses and incidence is rising, with mortality in excess of 35%. septic shock predisposes patients to multiple organ failure. while albumin is effective in management of circulatory dysfunction in septic shock, its utilization in this population is understudied in the us. we evaluated the impact of albumin utilization on hospital length of stay (los) among septic shock patients. we used a nationwide electronic health record data set (cerner health facts®) to extract real-world data on adult patients (≥18 years old) with severe sepsis or septic shock, admitted between january 1, 2013, and april 30, 2018, identified by international classification of disease (icd-9/10) codes, and receipt of antibiotics and vasopressors. we calculated the charlson comorbidity index (cci) and the acute physiology score (aps) at baseline. a generalized linear model was used to examine the association between albumin and hospital los, especially accounting for the timing of albumin infusion. we identified 3,156 unique visits for septic shock patients that survived to discharge. albumin was infused within 24 hours of admission ('early albumin') in 15%, after 24 hours ('late albumin') in 20%, and not administered in 65%. both cci and aps were higher, at presentation, in early albumin cases than late-or no-albumin cases (mean: 7.49 and 7.17, and 51.50 and 43.23, respectively). unadjusted los was slightly lower in patients receiving early albumin (11.81 days versus 11.84 days). a risk-adjusted analysis demonstrated that early albumin was associated with 4.92% shorter los (95% ci 0.43%-9.22%, p = 0.0322). albumin infusion within 24 hours of admission was associated with a shorter length of hospital stay. early albumin infusion may lead to better outcomes and reduced costs in patients with septic shock. further research is being conducted to assess other potential benefits of early albumin administration in this patient population. every new septic event follows by hemodynamic instability may lead sequentially to decreased organ perfusion, multiple organ failure. acute renal failure is recognized clinical feature during sepsis (up to 40-50% in all cases). furthermore, urine output close monitoring is a cornerstone diagnostic clinical tool in each septic critically ill patient. in present study, we analyzed the dynamic minute-to-minute changes in the urine flow rate (ufr) and also the changes in its minute-to-minute variability (ufrv) during new septic event in critically ill patients. demographic and clinical data were extracted from the of 50 critically ill patients who were admitted to the icu and developed new septic event (followed by fever and leukocytosis) and analyzed. a foley catheter was inserted into the urinary bladder of each study patient. the catheter was then connected to electronic urinometer, a collecting and measurement system which employs an optical drop detector to measure urine flow. the urine flow rate variability (ufrv) is defined and calculated as the change in ufr from minute to minute. results: ufr and ufrv both decreased significantly immediate after new septic episode until beginning fluid resuscitation (ppvalues <0.001) (figure 1) . statistical analysis by the pearson method demonstrated a strong direct correlation between the decrease in ufr, ufrv and the decrease in the map (r=0.03, p=0.003; r=0.03, p=0.004) ( figure 1 ), and heart rate (r=0.12,p=<0.001) since systemic pressure starts to drop. ufrv and ufr demonstrated good clinical response to fluid administration despite the fact that systemic blood pressure did not improve (figure 1) . we consider that dynamic changes in ufrv and ufr could potentially serve as a more sensitive signals ofclinicaldeterioration during the new septic event in critically ill patients.we also suggest that those parameters mightbeable to identify the optimal end-point of fluid resuscitative measures in septic critically ill patients. diminished urinary output (uo) is largely used as marker of acute kidney injury (aki) in critically ill patients. we aimed to explore the role of urinary output on incidence and mortality of aki developed during icu admission. the study population consists of all patients admitted between 2007 and 2018 to one of the dutch icus included in the nice database with an icu length of stay of at least 48 hours, having daily measurement of creatinine and uo. only patients without renal replacement therapy that have a serum creatinine lower than 1.1 mg/dl (97.5 μmol/l) or a uo above 0.5 ml/kg/h on the day of the index icu admission were considered at risk for aki. patients were followed during their icu stay and classified according to the highest kdigo criteria reached based on creatinine alone (model 1) and creatinine plus uo (model 2) using icu admission serum creatinine as baseline. in both models, patients were classified as: no aki, renal impairment at the first day of icu admission, aki stage 1, aki stage 2, and aki stage 3. we identified 52,863 patients (60% male, mean age 63 years, median icu-los 4 days). of those, 51.2% of patients had renal impairment at the first day of icu admission. among the remaining patients, 44.4% in model 1 and 29.9% in model 2 were classified as having no aki, 2.6% and 1.4% as aki stage 1, 0.7% and 9.3% as aki stage 2, and 1.1% and 8.2% as aki stage 3, respectively. survival at 30-day markedly differed according to the aki classification model used (figure) . similarly, adjusted hrs for 30-day mortality differed among patients with and without aki compared to patients with renal impairment at the first day of icu admission ( figure 1) . among patients admitted to the icu 50% had renal impairment at the first day of icu admission. our findings suggested that uo plays an important role both on aki incidence and mortality and should be carefully interpret in the clinical setting especially in aki stage 2 classification. introduction: acute kidney injury (aki) mostly attributed to renal tubular damage, has a high morbidity and mortality outcome [1] , so a sensitive tool to assess the degree of tubular affection is needed for early detection and management of this condition. we investigated the ability of furosemide stress test (fst) (one-time bolus dose of 1mg/kg or 1.5 mg/kg if on prior furosemide-intake) to predict progression to akin stage-iii in critically ill subjects with early aki. we studied 80 subjects; 40 consecutive patients in group i receiving fst and 40 consecutive patients in group ii receiving standard medical management for aki;15 patients (37.5%) and 20 patients (50%) met the primary endpoint of progression to akin-iii in groups i and ii respectively. patients with progressive aki had significantly lower urine output following fst in the first 6 hours (p<0.033). the area under the roc curves for the total urine output over the first 2 hours following fst to predict progression to akin-iii was 0.87 (p = 0.001). the ideal-cutoff for predicting aki progression during the first 2 fig. 1 (abstract p227) . thirty-day survival according to aki classification model 1 and model 2. hazard ratios (hrs) for 30-day mortality adjusted by sex, age, type of admission, apache iv score, sofa score at day of admission (excluded renal sofa score) for patients with aki classified with model 1 and model 2 fig. 1 (abstract p226) . clinical correlation between urine flow rate variability (ufrv) and ufr and mean arterial blood pressure over new septic event (black arrows) and and after initial fluid resuscitation (red arrows). note: the ufrv and ufr decreased progressively in parallel with the falling mean arterial blood pressure and, than, rose again after the administration of fluids hours was a urine volume of less than 325 milliliters with a sensitivity of 87.1% and specificity 84.1% group receiving fst. on the other hand, statistically significant hypotension, hypo-(kalemia, phosphatemia and magnesemia) occurred in group i. the fst in patients with early aki could predict liability for progression of aki, however it should be performed under adequate monitoring. introduction: ischemia-reperfusion (ir) causes renal dysfunction and damage. ir induces renal tubular injury triggered by hypoxia and hyperoxia, mediated by oxidative stress and inflammation. furosemide inhibits na + -k + -2clcotransporter in the thick ascending limb of the renal medulla to decrease na + reabsorption, reducing oxygen consumption. we investigated if furosemide could improve renal oxygenation, function and damage by reducing o 2 consumption and oxidative stress after ir. methods: 24 wistar albino rats were divided into 4 groups, with 6 in each group; sham-operated control (c), control + furosemide (c+f), ir and ir+f. after anaesthesia (bl), 45 min supra-aortic occlusion was applied to ir and ir+f groups followed by 15 min (t1) and 2 hours of reperfusion (t2). furosemide 50μg/kg/h infusion was simultaneously administered to c+f and ir+f after ischemia. systemic hemodynamic, renal blood flow (rbf), renal vascular resistance (rvr), renal oxygen delivery (do 2ren ), renal oxygen consumption (vo 2ren ), creatinine clearance (ccr), sodium handling, urine output (uo), cortical (cμo2) and medullar (mμo2) microvascular oxygenation were measured. results: rbf was reduced in ir (2.1±1) and ir+f (2.3±1) at t1 (p<0.05) but it was further reduced in ir+f (1.9±1) (p<0.05) at t2 compared to c and c+f. rvr was increased in ir (5338±2860) and ir+f (5123±2517) at t1 compared to c. rvr was normalized in ir (2198±879) but not in ir+f (4232±2636) at t2 compared to c (p<0.05). cμo 2 and mμo 2 did not differ between groups after ir insults (figure 1 ). tissue o 2 was reduced at the medulla, but not at the cortex in ir+f group compared to ir. do 2ren and vo 2ren were reduced in ir (56±17 and 26±12 ml/ min) and ir+f (34±20 and 21±14) at t2 (p<0.05). pc was higher in ir+f (37.33±4.27) compared to ir 29.67±3.39 (p<0.05). vo 2 / tna + was increased in ir+f compared to ir. no change in ccr and uo was observed. furosemide after ir causes further impairment of renal perfusion, energy utilization and renal oxygenation resulting in renal damage. acute renal failure induced by hypoxemia: incidence and correlation study a trifi 1 , h fazzeni 2 , a mehdi 2 , c abdennebi 2 , f daly 2 , y touil 2 , s abdellatif 2 , s ben lakhal 2 1 la rabta hopital, medical intensive care unit., tunis, tunisia; 2 la rabta hopital, tunis, tunisia critical care 2020, 24(suppl 1):p230 introduction: acute renal failure (arr) is a common complication in icus and usually caused by hypoperfusion. arf induced by hypoxemia is a concept rarely reported in icu. its incidence and pathogenesis are not well understood. we aimed to study the relationship between hypoxemia and the occurrence of arf. retrospective cohort study including patients with hypoxemia whatever its etiology between january 2016 and august 2019. patients with chronic renal failure were excluded. arf was defined and ranked according to the kdigo criteria 2012. arterial blood gas, urea, creatinine and clearance were reordered on the first, third and seventh days of evolution. results: 50 patients were included and 2 groups were obtained: group of hypoxemic patients with arf (arf+, n=30): versus group of hypoxemic patients without arf (arf-, n= 20). the incidence of hypoxemie-induced arf was therefore 60%. clinical characteristics were comparable in both groups with a mean age of 47 ± 16 and a sex ratio of 1.77. the comparative study showed in arf+ group: a lower ph (7. .25], p = 0.023). the most significant correlation was showed with mdrd clearance at day 3 and p/f ratio at day 1 (rho = 0.338, p = 0.038). multivariate analysis found that septic shock and non invasive ventilation in hypoxemic patients were the factors related to arf with respectively or=11.08, 95% ci=1.56-83.84, p=0.016 and or=6.18, 95% ci=1.16-34.07, p=0.033. overall mortality was 68% (n=34) and arf was an independent factor of mortality: or=6, and 95% ci=1.35-26.64, p = 0.017. hypoxemia-induced arf is a common complication associated with excess mortality. our study suggests that renal function is correlated with the degree of hypoxemia and that this correlation is rather distinct 48 hours from hypoxemia. in preclinical models of sepsis, we have previously demonstrated that activation of amp activated protein kinase (ampk) using metformin, improves survival and organ function. thus, ampk activation is a potential therapeutic target in sepsis, and we hypothesize that exposure to metformin during sepsis is associated with decreased aki and mortality methods: retrospective analysis of a 13-hospital cohort of adult icu patients with type 2 diabetes mellitus (t2dm) who presented sepsis. we investigated if exposure to metformin during the hospitalization was associated with reduced 90-day mortality and aki. we used 1:4 propensity score matching (psm), propensity score stratification (pss) and propensity score weighting (psw) based on the probability to be exposed to metformin using 55 covariates. for psm an exact match for insulin, amputation, cardiovascular diseases, retinopathy, charlson index, egfr, hba1c, and apache iii, were used. sepsis was defined using sepsis 3 criteria, and aki as kdigo stage 2 or 3. from 164,910 patients, we found 673 diabetic adults exposed to metformin during hospitalization and 14,174 who were not. metformin exposure during hospitalization is associated with decreased 90-day mortality and aki in septic adult patients with t2dm. these findings suggest that metformin may constitute a potential therapeutic strategy in sepsis, and the potential role of ampk activation as a protective mechanism. however, studies are needed to confirm this association and the specific mechanisms of action. introduction: acute kidney injury (aki) may occur up to 50% in the intensive care unit (icu). predicting aki recovery may allow for risk stratification of patients, patient and family counseling, and early post-discharge renal care planning. however, predicting aki recovery at an early stage remains a challenge. methods: this is a retrospective study of the epanic multicenter randomized controlled trial database [1] , which was split into development (n=2194) and validation (n=2446) cohorts, and patients experiencing aki stage 3 and/or renal replacement therapy (rrt) in the icu were included [2] . aki recovery was defined as being alive, without any stage of aki, and without need of rrt at hospital discharge. a logistic regression model with backward feature elimination was developed. the model performance was assessed by discrimination, calibration, and net benefit analysis, and internally validated with ten-fold cross validation. only the results in the development cohort are reported. of the 229 patients who developed aki3, 86 patients (37.55%) recovered from aki. the multivariable model selected age, bilirubin, heart rate, mean arterial blood pressure, surgical diagnostic group on icu admission, mechanical hemodynamic support on icu admission, suspected sepsis on icu admission as aki recovery predictors. the model had a mean area under the receiver operating characteristic curve (auroc) of 0.75 (standard deviation (sd) 0.01), mean calibration slope of 1.02 (sd 0.04), and mean calibration-inthe-large of <0.01 (sd 0.01) (figure 1 ). at the classification threshold that maximized sensitivity and specificity, mean net benefit with respect to treat-none was 0.16 (sd 0.01) and mean net benefit with respect to treat-all was 0.11 (sd 0.01). by using the routinely collected clinical data, the developed prediction model can fairly identify patients with a higher chance of aki recovery at hospital discharge. introduction: acute kidney injury (aki) is a frequent complication in critically ill patients and is associated with increased morbidity and mortality. sepsis is one of the most common cause of aki. a prospective study was conducted over 6 months (january 01-june 30, 2018).we included patients with septic shock at admission or at any time during hospitalization.the aki staging was based on kdigo criteria.patients were divided into two groups, a group with aki (aki+) and a group without aki (aki-).then we compared the baseline characteristics, laboratory and physiologic data. patients with aki (aki+) were subdivided according to their prognosis. were enrolled 75 patients. the mean (sd) age was 56.43(±18) years.sex ratio was 1.91. fifty-two (70%) patients developed aki.sapsii and sofa score in admission were higher in patients with kidney injury [59 vs 44 points (p= 0.002), 6.5 vs 4 points ;(p=0.003)] respectively.the serum lactate level was significantly higher in (aki +) group patients during the first day of septic shock [6.12± 1.38 mmol/l (aki+)vs 4.11± 0.79 mmol/l(aki-);(p=0.002) ] and its clearance was lower [(32±10.99% (aki +)vs 61±13%(aki-);(p=0.001)]. a significant difference was observed in c reactive protein level [224±114 mg/l (aki +) vs 124±77 mg/l (aki-) ; (p=0.004)].among (aki+) patients, kadigo iii was observed in 59.6% of cases.nineteen (36.5%) patients received hemodialysis.a normal kidney function was recovered in 40.4% of cases.aki+ patients had a higher occurrence in disseminated intravascular coagulation (32 vs 3 patients, p=0.002),acute respiratory distress syndrome (18 vs 2 patients; p=0.023) and cardiac dysfunction (20 vs 1 patient, p= 0.001).mortality was higher in aki group (67% vs 9%; p=0.001). the development of septic aki was associated with poor outcomes and prognosis.a better understanding of sepsis induced aki pathway will enable us to develop targeted therapeutic protocols.newer tools,permitting aki early detection, may make these therapies more fruitful. this study aims to show that contrast procedures do not significantly increase the risk of renal injury and should not be deferred. traditionally ciaki is the most important cause of in-hospital renal failure after nephrotoxic drugs and shock. problem is also the non-uniform definition of ciaki proposed by three different initiatives (akin, esur and kdigo). akin, being the most rigorous, defines ciaki as an increase in serum creatinine >0.3 mg/dl or >50% of baseline within 48hours. a retrospective observational single-centre cohort study analyzed 82 patients who underwent a contrast procedure with iomeron 350. the first group underwent a ct pulmonary angiography (ctpa), and the fig. 1 (abstract p232 ). internally validated model performance: (top row) roc curve; (middle row) calibration curve; (bottom row) decision curve second a coronary angiography with pci. no patient was previously prepared (raas blockade removal, crystalloid administration etc). we studied demographics, history of ckd and comorbidities and their impact on the ciaki by the akin criteria. a total of 82 patients were divided into two groups (ctpa and pci). ctpa group (20m, 21f) all had acute pe and the pci group (28m, 13f) were treated for acs. the mean age was 69 and 65 years respectively. ckd was more prevalent in the pci group (8pt vs. 3pt) possibly explained by the more advanced atherosclerotic disease. advanced chd (nyha iii/iv) was found in 3pt (pci) vs. 2pt (ctpa) while diabetes and shock were equally distributed (11pt and 5pt) in both groups. the mean amount of contrast was significantly higher in the pci group (242.3ml vs. 60ml). the mean creatinine/egfr measured before and after contrast in the ctpa group was 87. the goal of this study was to determine whether changing the body mass (bm) with fat-free mass (ffm) in cockcroft-gault (cg) formula could provide a more accurate prediction of aki in obese patients undergoing cardiac surgery. in this retrospective study, we reviewed institutional data of patients who underwent elective cardiac surgery in a tertiary referral university hospital. baseline patient creatinine value was collected and gfr was estimated using the mdrd, ckd-epi and cg formulas. cg formula was further modified by replacing the bm with ffm derived from the bioelectrical impedance analysis. postoperative aki was defined by kdigo creatinine change definitions. accuracy of the egfr values to predict the aki was calculated with roc-auc analysis. all the calculations were performed in different categories of bmi. figure 1 ). the egfr is a poor predictor of aki in obese patients undergoing cardiac surgery. the ffm modified cauckraft-gault formula yield more accuracy in this specific group. retroaki: a ten-year retrospective study of acute kidney injury in intensive and progressive care units introduction: acute kidney injury (aki) is a frequent condition in intensive care units (icu) and progressive care units (pcu), affecting 15% to 70% of the patients, depending on the studied population and aki definition. aki has been identified as an independent risk factor of icu mortality and development of chronic kidney desease. the objective of this study was to describe the incidence of each aki stages as defined by kdigo definition (with evaluation of urine output, serum creatinine and initiation of renal replacement therapy (rrt)), in a mixed medical and surgical population of patients hospitalized in icu and pcu over a 10-year period (2008-2018). we included all patients who stayed more than 12 hours in icu or pcu of edouard herriot hospital from may 2008 to january 2019. data used to classify the patients were the urine output over a sixhour period, serum creatinine and the need for rrt, according to kdigo classification results: 18,882 hospital stays were analyzed. median icu/pcu length of stay was 3 days [iqr: 1.5-6.6]. among icu patients, 74% had at least one aki episode graded 1, 2 or 3 and 49% had at least one severe episode (stage 2 or 3). among pcu patients, 44% had at least one episode of aki and 20% a severe episode of aki. patients had an average of 1.9 episodes of aki per stay. table 1 represents the incidence of maximal aki stage during one stay. we found that urine output was the more frequent criteria to make diagnosis of aki stage 1 or 2 whereas rrt was more frequent for aki stage 3. this retrospective study reports a more important aki incidence in our icu/pcu than in previous studies. the difference could be fig. 1 (abstract p236) . when comparing auc in different categories of bmi, the mcg appeared to be the only statistically accurate formula in patients with bmi 30-34.9 explained by the difficulty to collect urine output from conventional database. serum creatinine and the use of rrt are often the only two criteria used to define and classify aki. these results confirm the high incidence of aki in icu and pcu and the importance to make an early aki screening of patients for whom preventive nephroprotective actions are needed. introduction: icu-patients with acute kidney injury (aki) requiring renal replacement therapy (rrt) are at risk for infections [1, 2] . in this study we evaluated the incidence of infection in icu patients with and without less severe aki. finally, impact on outcomes was explored. this is a retrospective study on the pdms (protection data management system) of the 4 adult icus of a university hospital. aki was assessed on kdigo criteria (creatinine (scr) and urine output), during the first 7-d of icu stay. infection was validated in the pdms by a team of icu specialists. results: during a 4-year period, a total of 7485 subjects were enrolled. aki was diagnosed in 64.7% of patients during icu stay. aki patients were older (63 vs. 59 y, p=0.001), had higher saps 2 (57 vs. 41, p< 0.001), and had more urgent icu admission (64% vs. 48%, p<0.001). more aki patients had mechanical ventilation (55% vs. 41%, p<0.001) and vasopressors on d-1 (47% vs. 23%, p<0.001). aki stage 1, 2, and 3 was present in 25.5%, 28.0% and 11.1% of patients. more aki patients had infection (57% vs. 28%, p<0.001) and increasing aki stages were associated with higher infection rates (aki-0: 28%; aki-1: 55%, aki-2: 55%, aki-3: 69%, p<0.001) (figure 1 ). we observed 2-3 times higher mortality in aki patients with infection, and a stepwise increase of mortality with increasing aki stages. after correction for infection and other confounders we found that all aki stages were associated with in-hospital mortality (ors aki-1: 1.7, aki-2: 2.0, aki-3: 3.6, all p< 0.001). over half of aki patients experienced an episode of infection and increasing aki severity was associated with higher infection rate. aki patients with infection had marked higher mortality, suggesting that infection was an important driver of outcome. however, after adjustment, aki stages had strong association with hospital mortality. several new biomarkers have been introduced to improve early diagnosis of acute kidney injury (aki). "nephrocheck" (nc; astute medical, usa) is a bedside test calculating "akirisk" (product of urinary concentration of the cell cycle arrest-markers timp-2 and igfbp7). several studies suggest the usefulness of nc in selected populations. however, the value of early routine measurement of nc is unclear. methods: therefore, we compared the prediction of a combined endpoint (cep: death <60 days and/or requirement of renal replacement therapy rrt) by nc within 12h of icu admission (nc1) and 24h later (nc2) with admission values of serum-creatinine, bun, cystatin c, urinary ngal, apache ii and sofa (roc-analysis). as a secondary endpoint we investigated the additional value of pathological measurements of nc1≥0. 3 critically ill patients showed increased relative uce in the first days of icu admission, which may be attributed to higher protein catabolism. increased relative uce was associated with arc and both had no effect on 90-day mortality. introduction: this study compared epidemiology, short-and long-term outcomes for patients with community-acquired (ca) and hospital-acquired (ha) acute kidney injury (aki). we retrospectively analyzed all episodes of aki over a period of 3.5 years (2014-2017) on the basis of routinely obtained serum creatinine measurements in 103,161 patients whose creatinine had been measured at least twice and who had been in the hospital for at least two days. we used the "kidney disease: improving global outcomes" (kdigo) criteria for aki and analyzed the first hospital admission. a total of 103161 were admitted in hospital and fulfilled the inclusion criteria. average observation period per patient was 248 days. the incidence of ca-aki among included hospital admissions was 9.7% compared with an incidence of 8.6% of ha-aki, giving an overall aki incidence of 18.3%. patients with ca-aki were younger than patients with ha-aki (64 vs 66.2y) and had significantly less comorbidities, including preexisting cardiac failure, ischemic heart disease, hypertension, diabetes. patients with ca-aki were more likely to have stage 1 aki (69,3 vs 58,4%, p<0.001) and had significantly shorter lengths of hospital stay than patients with ha-aki (14 vs 24d, p< 0.001). those with ca-aki had better survival than patients with ha-aki (figure 1; p<0 the evidence base for management of fluid removal during renal replacement therapy (rrt) is limited. a recent international survey revealed the extent of practice variation worldwide [1] . our aim was to summarise the responses from europe-based healthcare professionals who participated in the survey. the international self-administered, cross-sectional, internet-assisted, open survey was disseminated between january 2018 and january 2019 via website links and emails to members of different critical care societies. results: 485 participants from 31 european countries completed the survey of whom 365 (75%) were intensivists and 306 (63%) worked in university-based hospitals. persistent oliguria / anuria was the most common indication for fluid removal (51% responders). the parameters which guided fluid removal included hemodynamic status (47% responders), cumulative fluid balance since admission (23% responders), and 24-hour fluid balance (17% responders). 90% of participants reported using crrt with a median net ultrafiltration rate 98 ml/hr (iqr 51-108ml/hr) for hemodynamically unstable and a rate of 300 ml/hr (iqr, 201-352ml/hr) for hemodynamically stable patients. only 26% of practitioners checked net fluid balance hourly (70% nurses, 16% physicians). new hemodynamic instability, defined as new onset or worsening tachycardia, hypotension, or need to start or increase the dose of vasopressors was reported to occur in 20% fig. 1 (abstract p242 ). long-term survival patients (iqr 10.0-30.0). different strategies to re-gain hemodynamic stability were used. (figure 1 ) main barriers to fluid removal were patient intolerance (72% physicians, 85% nurses) and interruptions in fluid removal (43% physicians, 64% nurses). the majority of participants agreed that guidelines and protocols would be beneficial. the practice of fluid removal during rrt is very variable across european countries. nurses and doctors identified a need for evidencebased protocols and clear guidelines. introduction: kidney disease improving global outcomes (kdigo) guidelines suggest the use of anticoagulation in continuous renal replacement therapy (crrt) [1] . the effectiveness of the anticoagulation is important because replacing the hemofilter and tube interrupts crrt and increases total therapy time. regional citrate anticoagulation (rca) and unfractionated heparin (ufh) are most commonly using methods for crrt anticoagulation [2] . the aim of this study was to investigate the efficacy, safety and metabolic differences of the patients in icu who underwent crrt and anticoagulation method changed from ufh to rca for different reasons. after ethics committee approval (2019-14/9) 100 patients who underwent crrt between 2018-2019 at bursa uludag university hospital icu have been investigated and 11 patients who underwent crrt by both rca and ufh included in the study. we divided patients in two groups (rca, ufh), demographic data (sex, age), sofa score, creatinine, urea, mean filter life time (flt) and ultrafiltration flow (uf), platelets, electrolytes (na, k, ca, mg), lactate, nahco3 and ph of groups at beginning and ending of first rca and ufh hemodialysis collected. we used t-test and 1000 bootstraps statistic tests. in agreement with other studies [3, 4] , flt and uf was statistically significant lower in ufh group (table 1) . there was no statistically significant difference in efficiency (urea and creatinine decrease), ph, lactate, nahco3 level, platelets count and electrolytes between two groups. to our knowledge, there are no studies comparing these two anticoagulation methods in the same patients. small number of patients and retrospective evaluation are limitations of the study. our results suggest that the implementation of rca method is safe and effective as ufh method with longer flt and uf. regional citrate anticoagulation during crrt in liver failure mj jain, pk kumar g, dg govil, jk kn, sp patel, ms shafi, rh harne, dp pal, sm monanga medanta the medicity, critical care, gurugram, india critical care 2020, 24(suppl 1):p245 continuous renal replacement therapy (crrt) with regional citrate anti-coagulation (rca) is increasingly being used as a treatment modality in critically ill patients. there is limited experience of use of citrate anticoagulation patients with acute liver failure and acute on chronic liver failure who pose a tough challenge of being at a higher risk for bleeding. an institutional protocol was formulated for use of commercially available citrate solutions and the same was studied to assess filter life and safety of citrate in liver disease. the primary objective was to assess safety of citrate anticoagulation in liver disease. this study was a single centre, prospective, non-randomized, single arm, observational study. all adult patients, with acute liver failure and acute on chronic liver failure requiring crrt were included. blood ionized calcium levels of 0.9 to 1.1mmol/l was targeted throughout the therapy and total to ionized calcium ratio of less than 2.4 was maintained. rca was stopped if the ratio was more than 2.4 for 2 consecutive assessments. incidence of citrate accumulation and toxicity were assessed. average filter life was also assessed. metabolic parameters, electrolytes and strong ion gap were followed till 24 hours after completion on crrt. a total of 25 patients were included in the study. nineteen patients of acute on chronic liver failure and 6 patients of acute liver failure underwent crrt with rca. baseline average serum bilirubin, lactate and inr were 11.8 mg/dl, 6.4 mmol/l and 2.1 respectively. the average filter life was 50 hours 3 minutes. citrate accumulation took place in (n=13) patients and rca had to be stopped for ( n=6) patients due to the same. none of the patients had evidence of citrate toxicity. citrate anticoagulation was well tolerated in patients with acute liver failure in patients with or without pre-existing chronic liver disease on crrt. introduction: the intention of this study is to highlight the levels of citrate load for the general population that increases the risk of citrate complications (insufficient trisodium citrate delivery; net citrate overload and citrate accumulation) [1] . this was a prospective data collection between february and march 2019 in a fourteen bedded critical care unit. eleven consecutive episodes of crrt were collected (a new episode characterized if crrt was discontinued for 48 hours and above). one episode was excluded due to short duration (less than 4 hours). patients undergoing rca-crrt received either a fixed 25 or 35 ml/kg/h effluent dose protocol. median patient age was 59, male 100%. average time on crrt was 4.1 days (2-9). 70% of the patients had complications, although 60% were minor ( figure 1 ). all of the patients with net citrate overload had citrate loads of 13.8mmol/h or above. the main risk factors were found to be shock and liver impairment which occurred in 60% of cases of which 40% developed complications. a fixed dose effluent protocol to standardise practice can potentially lead to a higher risk of minor complications. in our experience this is likely due to a lack of appropriate monitoring for rca-crrt complications. despite this, our complication rate of citrate accumulation is in line with that reported in literature. citrate loads in our 25 ml/kg/ hr protocol were 22.6% higher than our 35 ml/kg/hr protocol and strongly related to higher complication rate that worsened in patients with risk factors for poor citrate metabolism. introduction: there is no optimal timing of continuous renal replacement therapy (crrt) in acute kidney injury (aki); however, it is based on volume overload, azotemia, hyperkalemia and severe metabolic acidosis [1] . an important reason for metabolic acidosis in aki is increased unmeasured anions (ua) [2] . delta-ph-ua (δph ua ) detects the degree of metabolic acidosis caused by ua and is calculated by using 'the partitioned ph model' [3] . in this study, we investigated whether δph ua was a predictor to start crrt in patients with aki. the study was designed as a multicentric, prospective, observational study in 2019. patients who were ≥18 years old and diagnosed with aki [1] were included. the moment aki was diagnosed, arterial blood gas, albumin, magnesium, inorganic phosphorus, urea, creatinine and δph ua values were recorded. all patients were divided into two groups as crrt(-) and crrt(+) which consists of patients performed crrt due to traditional criteria. fig. 1 (abstract p246) . incidence of complications introduction: continuous renal replacement therapy (crrt) is labor intensive and requires advanced nursing knowledge and skills. however, 40% of registered nurses (rn) are less than 2-year post-registration experiences in our unit. also there is an increasing demand of crrt from 185 crrt days in 2017 to 248 crrt days in 2018. the obstacles for crrt in our department, includes variation of regimen, complicated workflow and insufficient training of nurses. a continuous quality improvement project is carried out to standardize the regimen, enhance workflow and provide structured training to nurses in the intensive care unit, to enhance nursing competence. methods: introduction: sepsis and septic shock is a leading cause of mortality in the intensive care unit. we tried to evaluate a novel hemoperfusion cartridge through a retrospective evaluation of patient's data in our centre. we used it as an adjuvant therapy in our patients with sepsis and septic shock due to varied causes. the aim of this study was to evaluate the efficacy of therapeutic hemoperfusion cartridge (hc-foshan biosun medical ® ) in the management of patients with sepsis. we retrospectively analysed data of group 1 (n=30 sepsis) and group 2 (n=30 sepsis+hemoperfusison; sepsis treated with hemoperfusion cartridge) admitted between 2015 to 2018. group 2 had received hemoperfusion cartridge as adjuvant therapy along with standard of care. demographic data, procalcitonin [1] and leukocyte levels before and after therapeutic cytokine removal and duration of hc were recorded. while the mean duration of cvvhdf was 96.4 hours, the duration of hemoperfusion cartridge (application was 32.1±16.4 hours). among 30 patients who survived 25 patients were administered hemoperfusion cartridge within 12 hours of icu admission. there was a significant reduction in scores like apache and sofa score post hemoperfusion cartridge therapy procalcitonin and leucocyte levels after therapeutic hemoperfusion cartridge were found significantly lower than the pretreatment values (respectively p=0.001, p=0.001). retrospective analysis showed significant reduction of vasopressors, and improvement in map in group2. therapeutic hemoperfusion cartridge with cytokine removal applied with cvvhdf in septic patients have positive contributions to provide survival advantage. removal of activated leukocytes and endotoxin from the blood is a complex therapeutic effect of the device for removing endotoxin. in the main group (16 patients with abdominal septic shock) after surgery, the traditional treatment was supplemented with two sessions of endotoxin removal (2 hours each with an interval of 24 hours) using "alteco lps adsorber" (sweden). the control group consisted of 8 patients with a similar diagnosis and only traditional treatment. results: 28% of white blood cells were adsorbed in lps adsorber. among them, granulocytes (35%) were maximally extracted, then cd14 + monocytes (cd14 + mo) (33%), hla-dr + mononuclear cells (6%), monocytes (2%). il-6, il-10, procalcitonin (pct) were not adsorbed. the 28-day mortality rate in the main group was 50% and was lower compared to the control group -75%. during monitoring, in the main group 24 hours after the first removal of endotoxin, a decrease in the initially increased amount of activated cd14 + mo by 2.2 times, as well as functionally mature defensin + granulocytes (def + gran) by 1.6 times was observed. il-6, il-10, and pct decreased by 1.9; 17.8; and 1.2 times, respectively. during this period, the control group showed an increase in cd14 + mo and def + gran, while il-6, il-10 did not change, and pct increased 1.9 times. a day after the second removal of endotoxin and then 5 days later, the main group of il-6, il-10, and pct continued to decline. in the control group, only il-10 decreased after 3 days, the rest continued to grow. the cellular adsorption of endotoxin-bound cd14 + mo and mature def + gran is an important part of the mechanism of action of the endotoxin removal device. does the endotoxin adsorption of pmx column saturate in 2 hours? preliminary study c yamashita 1 in the euphrates trial, the polymyxin b-immobilized fiber column (pmx) hemoperfusion (hp) had no significant effect on 28-day mortality. endotoxin (lps) burden by endotoxin activity assay >0.90 may exceed 50 μg [1] , so the dose and duration of pmx-hp could be insufficient to lower the lps burden. to confirm this issue, we experimented in a closed-circuit with 24 h continuous lps addition, and pmx can adsorb > 50μg [2] . further, lps concentration became constant within 2 h in the single lps spike test for determining pmx-hp duration [3] . to prove our hypothesis that the single lps spike test reflects the adsorption equilibrium, and not saturation, we added lps intermittently to reaction. methods: lps (10 ng/ml) was mixed with 125 ml deactivated fetal calf serum as a reflux solution, as previously described [2] ; this concentration is much higher than that observed in septic patients. we created a closed circuit that incorporates pmx-01r at 1/14 th the amount of an adult pmx and performed pmx-hp at 10 ml/min for 5 h. lps was added in two shots (post 2 h: 1250 ng, 10 ng/ml; post 4 h: 3750 ng, 30 ng/ml). lps was measured using the limulus amebocyte lysate test at 0, 0.5, 1, 2, 3, 4 and 5 hr. after an initial decrease between 0 and 1 h, lps concentration did not decrease between 1 and 2 h after pmx-hp initiation. post lps pulse addition at 2 h, it increased and then decreased till 3 h. futher, it did not decrease between 3 and 4 h, but it increased and then decreased again after lps pulse addition post 4 h (figure 1 ). lps adsorption rates were 76.2, 43.4, and 40.7% at 2, 4, and 5 h, respectively. conclusions: lps adsorption capacity of pmx-01r was maintained even after two additional shots of lps, suggesting that the constant lps concentration in the previously reported lps spike test might be indicative of adsorption equilibrium rather than saturation. a coohort study included 65 patients admitted to three intensive care with sepsis / septic shock ( sepsis 3 criteria ) and aki ( akin score). all patients were submitted to cvvhdf with the oxiris filter (baxter, usa) . the main clinical data, il 6, procalcitonin, endotoxin ( eaa ) and sofa score were evaluated at basal time ( t0 ) and at the end of the treatment ( t1 ). all data are expressed as mean ± sd or median and iqr . anova test was used to compare the changes in the time. results: 60 patients were submitted to rrt with the oxiris filter for 46 ± 12 hours . 21 patients had aki 3 stage , 13 patients aki 2 stage and 25 patients had aki 1 stage. at t0 all groups had an high vasopressor fig. 1 (abstract 251) . lps concentration in lps pulse addition test support to maintain map ≥ 70 mmhg. il6, procalcitonin eaa and sofa total were also elevated with no difference between the groups. at t1 creatinine improved better in aki 2 ( p< 0.001 vs. t0 ) and in aki 1 ( p< 0.0001 vs t0) then in aki 3 group. map increased in aki 2 ( p< 0.01 vs t0) and aki 1 ( p < 0.01 vs t0) , but not in aki 3 group. il6, procalcitonin decreased more in aki 1 ( p < 0.0001 vs t0) then aki 3 . at t 2 sofa total was higher in aki 3 then aki 1 ( p< 0.001 ) and aki 2 ( p< 0.01 ). conclusions: aki 2 and aki 1 stage patients submitted to bp with the filter oxiris respond better then aki 3 stage patients . 2 -this transalte in a better clinical course. 3-crrt with oxiris filter is useful in septic patients with aki, but aki 3 stage septic patients represent an high risk group. a non-interventional, multicenter, non-randomized patient registry for multiple organ dialysis with the advos system multiple organ failure is a challenging problem in the icu. as an advanced dialysis system, the advos procedure can eliminate watersoluble and protein-bound substances, regulate the acid-base balance as well as fluid and temperature. in 2017, a national registry was established to collect data under "real-life" conditions of patients treated with advos without any trial-specific interventions (drks id: drks00017068). methods: data from 01/2017 to 02/2019 from 4 german hospitals (university hospitals in hamburg-eppendorf, mainz, essen, and klinikum weiden) were analyzed. clinical parameters, treatment settings and adverse events were documented. the 28-and 90-day mortality rates were compared with extrapolated rates based on the sofa score. results: 118 patients with a median age of 60 years (iqr 45-69), of whom 70 (59%) were male, were evaluated. patients had a median sofa score of 14 (iqr: 11-17) before the 1st advos treatment, which is associated with an expected mortality of 80%. the number of failing organs was 3 (iqr 2-4): cardiovascular (74%), lungs (57%), liver (47%), kidneys (74%), coagulation (69%) and cns (29%). 429 treatments with a median duration of 16 (iqr: 10-20) hours were evaluated. 87 were discontinued, of which 25 (6%) were due to a device error. 79 adverse events were documented, 13 were related to the device (all due to clotting and recovered without sequelae). significant removal of protein-bound (bilirubin: 11.2 vs 9.2 mg/dl) and water-soluble toxins (bun 32 vs 20 and creatinine 1.9 vs 1.4 mg/dl). in addition, improvement in acid-base balance was observed: ph (7.33 vs. 7.40), bicarbonate (21.3 vs. 25.5 mmol/l) and base excess (-4.5 vs. 1.0 mmol/l) ( table 1) . 28-and 90-day mortality rates were 60% and 65%, respectively. in a cohort of patients with multiple organ failure, we observed an improvement in the expected mortality rate, especially if the advos procedure was applied early. adverse events are comparable to other dialysis therapies in intensive care patients. introduction: acute kidney injury (aki) due to ischemia-reperfusion affects onethird of the patients in cardiac surgery. we investigated the potential role of cyclosporine (csa) to prevent postoperative aki and mitigate inflammatory response to extracorporeal circulation (ecc). methods: double-blind, randomized, placebo-controlled single-center study. patients (n=67) scheduled for elective cardiac surgery were randomized to 2,5 mg/kg csa or placebo before the surgery. the primary objective was to assess the role of csa to reduce the incidence of postoperative aki. the secondary objective was to study csa induced changes in the inflammatory response to ecc. results: all enrolled patients were analyzed. postoperative aki was more pronounced in the cyclosporine group compared to placebo. or=5.03 (1.76-15.74), 95% ci. the cytokine production in response to ecc was not affected by cyclosporine (figure 1) . in patients undergoing cardiac surgery, a single preoperative dose of csa does not prevent the postoperative decrease in renal function. csa does not alter cytokine release in response to extracorporeal circulation. elevated post-ecc levels of pro-inflammatory cytokine il-6 are associated with kidney dysfunction and may be predictive. new generation adsorbent such as oxiris r was introduced as novel technique in renal support for critically ill patients [1] . septic shock patients require decatecholaminization strategies emphasizing blood purification to remove catecholamine-producing mediators and evacuate overload fluid in interstitials. our 64-year-old female patient, admitted to icu after surgery with history of ovarium cancer. her septic shock was worsened with ards, hypercoagulable state and aki. vasopressors were set. patient was controlled with mode simv16,ps12,tv350 ml,peep7,fio270%. renal support was implemented by diuretic and cvvh started on the second day. at first,regular adsorbent was used, post-filter mode was set, and periodic fluid removal target was 50 ml/h. but after 24hours, no significant changes observed. oxiris r added and after 12 hours passed, requirements of vasopressors reduced, tidal volume increased, hemodynamic parameters stabilized, urine production increased. it was continued for 2 days and patient was recovered. our patient had fallen into inadequate cars stage in which not able to counter septic effects on vital organs (figure 1 ). renal would be primary target for filtration and monitoring tool. adsorbent consisted of an69 and polyethyleneimine was useful to purify blood from endotoxins conjoined with slower filtration. continuous yet cautious process in cvvh evacuate fluid and mediators while maintain steady hemodynamics. biomarkers could not be evaluated due to limited resources, but improving parameters could be signs that showed recovery process had already took place. advanced hemofiltration is a privilege. implementing and enhancing it with new generation adsorbent would increase survivors by extracting unnecessary fluids and eliminating catastrophic endotoxins and mediators. consent to publish: written informed consent for publication was obtained from the patient. analysis of retrospective cohort study data of 120 patients (pt) treated for dka at icu of kaunas clinics during 2014 -2019 has been carried out. serum kalemia, glycemia; hypokalemia, hypoglycemia episodes; rate of insulin interruption for hypo-and normoglycemia during ketoacidosis; use of nah 2 co 3 for ketoacidosis, and los in icu were analysed. spss 23.0 was used for statistic calculations. traits evaluated as significant at p < 0.05. at the beginning of dka treatment in totally hypokalemia (3.1 ± 0.3 mmol/l) was recorded in 64/120 pt (53.3 %). due to ignoring of blood ph (6.8 -7.3 (7.0 ± 0.1) kalemia was falsely misinterpreted as "normo-" or "hyper-" 3.5 -7.1 (5.1 ± 0.9 mmol/l) in 49/68 pt (72.1 %), thus disregarded so complicated by obvious hypokalemia additionally in 26/49 pt (53.1 %). in hypokalemia los in icu was 52.9 ± 29.7 vs 32.8 ± 18.6 h, p < 0.05. insulin use has caused hypoglycemia (1.2 -3.3 (2.5 ± 0.7 mmol/l)) in 22/120 pt (18.3 %), los in icu 63.2 ± 38.5 vs 38.9 ± 21.2 h, p < 0.05.insulin use was interrupted in case of normoand hypoglycemia with still persisting ketoacidosis in 39/120 pt (32.5 %), los in icu was found to be 56.5 ± 30.7 vs 37.0 ± 22.5 hr, p < 0.05. nah 2 co 3 was given for symptomatic treatment of ketoacidosis during first 10 h of dka in 33/120 pt (27.5 %) with stable hemodynamic: hco -3 buffer has increased (4.8 ± 3.3 -7.9 ± 3.1 mmol/l), p < 0.05, but it didn't control ketoacidosis, and los in icu was 55.2 ± 27.5.2 vs 39.1 ± 25.6 h, p < 0.05. hypokalemia, hypoglycemia, precocious interruption of insulin use were recorded as complications of dka treatment. all of them have prolonged los in icu. symptomatic treatment of ketoacidosis with nah 2 co 3 had no effect on it, and prolonged los in icu as well. a growing interest exists about co 2 derived parameters in shock management. central venous-arterial pco 2 difference (p cv-a co 2 ) is strictly related to cardiac output; central venous-arterial pco 2 difference to arterial-central venous o 2 content difference ratio, p cv-a co 2 / c a-cv o 2 , has been proposed as anaerobic metabolism when it's >1.4 mmhg/ml [1] . to evaluate p cv-a co 2 /c a-cv o 2 reliability in detecting anaerobic metabolism, we analyzed it in 7 consecutive patients affected by mala admitted to our icu, considering these patients as a prevalent anaerobic metabolism model. we calculated, by douglas formula, central venous-arterial co 2 content difference to arterial-central venous o 2 content difference ratio, c cv-ca co 2 /c a-ccv o 2 , as a respiratory quotient surrogate. we performed arterial and central venous blood gas analysis simultaneously at admission, we calculated p cv-a co 2 , p cv-a co 2 /c a-cv o 2 and c cv-a co 2 /c a-cv o 2 and we recorded scvo 2 . we verified relationship between p cv-a co 2 /c a-cv o 2 and scvo 2 and arterial ph, arterial lactates, sofa score at admission and c cv-a co 2 /c a-cv o 2 by linear regression analysis. pcv-aco2/ca-cvo2 greatly increases in mala (2.16 ± 0.84). pcv-aco2/ ca-cvo2 (fig.1) shows significant co-variation with ph (r2=0.618; p= 0.003) and sofa score at admission (r2=0.628; p=0.003). pcv-aco2/ ca-cvo2 has poor agreement with ccv-aco2/ca-cvo2 (r2=0.008) and disagrees with it in identifying anaerobic metabolism, in our series, in fact, ccv-aco2/ca-cvo2 is, in 3 patients, < 1 like an aerobic rq value. pcv-aco2/ca-cvo2 shows better agreement with ph, sofa score and lactate level than scvo2. in our series, p cv-a co 2 /c a-cv o 2 is good illness and acidosis severity marker, but it seems to be affected by ph value in accord with haldane effect [2] . p cv-a co 2 /c a-cv o 2 , in our study, doesn't seem to be a reliable anaerobic metabolism marker nor a rq surrogate. it is thought that early administration of basal insulin to patients with diabetic ketoacidosis (dka) may improve outcomes. small studies have shown trends towards decreases in time to closure of anion gap (tcag), rates of rebound hyperglycemia following discontinuation of intravenous (iv) insulin, rates of hypoglycemia, intensive care unit (icu) length of stay (los), and hospital los [1] [2] [3] [4] . this was a single-center, retrospective chart review of our institution's dka protocol between january 2010 and august 2019. patients that received early basal insulin within 24 hours of initiation of iv insulin and before closure of the anion gap (ag) were compared to those that did not receive early basal insulin. the primary outcome was median tcag. secondary efficacy outcomes include: time on iv insulin infusion, time to de-escalation of level of care, hospital los, and re-elevation of ag. secondary safety outcomes included incidences of hyperglycemia, hypoglycemia, and hypokalemia. a total of 334 patients were identified meeting inclusion and exclusion criteria. median tcag was longer in the experimental group (9 vs. 6 hours, p <0.01). incidence of re-elevation of ag and incidence of hyperglycemia were lower in the experimental group. other outcomes were similar (figure 1 ). early administration of basal insulin to patients with dka resulted in a longer tcag with a lower incidence of re-elevation of ag and hyperglycemia. early administration of basal insulin appears to be safe with respect to hypoglycemia and hypokalemia. glycaemic control continues to be a challenge in critically ill patients. stress induced hyperglycaemia has been associated with increased morbidity and mortality [1] . conversely, patients receiving intensive glucose control have a higher risk of death [2] . a quality improvement project was designed to develop a comprehensive insulin protocol that recognized pre-existing diabetes and reduced hypoglycaemia. data was collected prospectively in all adult patients admitted to the rah intensive care unit (icu) between october 2018 and august 2019 from the national icu audit database and electronic patient records. daily figures were collected for numbers of hypoglycaemic episodes (<4mmol/l), "in range" (4-10mmol/l) blood sugar measurements and patients with a pre-existing diagnosis of diabetes. data was collected and analysed using microsoft excel. results: 307 patients were identified; 56 patients (18.2%) had pre-existing diabetes. a total of 6908 blood sugar measurements were reviewed; 5268 (76.3%) were "in range" and 126 hypoglycaemic episodes (1.8%) occurred. there was no significant correlation between number of diabetic patients and measurements within range. of note, there was an increase in number of measurements per patient in the second half of the time period (11 vs 32). the development of this protocol has improved glycaemic control in our icu. there are considerably fewer episodes of hypoglycaemia and a large proportion of blood sugar measurements are in range. we hope to continue data collection and interrogate the prevalence of pre-existing diabetes further to reduce glycaemic variability. the optimal management of blood glucose levels for critically ill patients remains unclear. hypoglycemia, hyperglycemia and glycemic variability are associated with mortality. the time in targeted blood glucose range (tir) has been suggested to correlate with mortality depending on the status of antecedent glycemic control, but it has not been verified optimal tir and whether there is an optimal disease-specific tir. a retrospective observational study was performed at a single center. in the present study, we enrolled all critically ill patients admitted in intensive care unit from 1 january 2016 to 31 october. patients with diabetic ketoacidosis or hyperosmolar hyperglycemic syndrome and patients who had < 10 blood glucose readings were excluded. gathered information included, in part, demographics, comorbidities, severity of illness scores, diagnosis at admission, length of icu stay and hospital discharge status. the primary outcome was 28-day mortality. we analyzed to find the optimal tir for critically ill patients. several tirs were each tested for correlation with mortality. a total of 1,523 patients, 51.8% of whom had diabetes, were studied. tir 70 to 139 mg/dl (or, 0.33; 95%ci, 0.18-0.58), tir 70 to 179 mg/ dl (or, 0.33; 95%ci, 0.23-0.47) and tir 110 to 179 mg/dl (or, 0.28; 95%ci, 0.17-0.44) > 80 % was independently associated with mortality in critically ill patients respectively. the optimal tir did not differ depending on diagnosis at admission. in this retrospective evaluation, tir 110 to 179 mg/dl > 80 % was independently associated with mortality in critically ill patients, especially those with good antecedent glucose control. these findings have implications for the design of future trials of intensive insulin therapy. the prevalence of chronic dysglycemia (diabetes and prediabetes) in patients admitted to swedish intensive care units (icus) is unknown. we aimed to determine the prevalence of such chronic dysglycemia and asses its impact on blood glucose control and patient-centred outcomes in critically ill patients. in this retrospective, observational study, we obtained routine glycated hemoglobin a1c (hba1c) measured in patients admitted to four tertiary icus in sweden between march and august 2016. based on previous diabetes history and hba1c we determined the prevalence of chronic dysglycemia (prediabetes, undiagnosed diabetes and known diabetes). we compared indices of acute glycemic control in the icu and explored the association between chronic dysglycemia and icu-associated infections, mechanical ventilation, renal replacement therapy, vasopressor therapy, and mortality within 90 days. of 943 patients, 312 (33%) had chronic dysglycemia. of these 312 patients, 127 (41%) had prediabetes or undiagnosed diabetes and fig. 1 (abstract p259) . results 185 (59%) had a known diabetes diagnosis. during icu stay, patients with chronic dysglycemia had higher average blood glucose, spent less time in target glucose range, had greater glucose variability, and were more likely to develop hypoglycemia than patients without chronic dysglycemia. chronic dysglycemia was associated with greater need for renal replacement therapy (odds ratio 2.10, 95% ci 1.35-3.27) and increased 90-day mortality (hazard ratio 1.33, 95% ci 1.01-1.77) after adjustment for simplified acute physiology score 3. in contrast, chronic dysglycemia was not associated with mechanical ventilation, vasopressor therapy, or icu-associated infections. in four tertiary swedish icus, measurement of hba1c showed that 1/ 3 of patients had chronic dysglycemia (prediabetes or diabetes). chronic dysglycemia was associated with marked derangements in glycemic control during icu stay, greater need for renal replacement therapy and with increased mortality at 90 days. case report: modern antidiabetic therapie causes ketoacidosis am heiden, m emmerich krankenhaus bad oeynhausen, institut für anästhesie, bad oeynhausen, germany critical care 2020, 24(suppl 1):p263 the modern antidiabetic class of sglt2-inhibitors, that are known to reduce the risk for cardiac events [1] , are increasingly used in the last few years. a 68-year old male patient with diabetes mellitus suffered 10 days after colectomy surgery from abdominal pain and nausea. the patient had an antidiabetic therapy with empaglifozin that was paused until day 5 after surgery (nutrition start on day 5, weaning on day 6). methods: this is a case report of one male patient seen in the icu setting. daily blood values including arterial blood gases, vital parameters and clinical status of the patient were observed and evaluated. the blood gases showed this metabolic acidosis: ph 7.38; pco2 20.3 mmhg, bicarbonate 12 mmol/l, be -11.63 mmol/l, lactate 1.6 mmol/l, glucose 7 mmol/l. a ketonuria despite normal blood glucose values was noticed, so that the diagnosis of ketoacidosis was clear. after analyzing the possible causes we found out, that empaglifozin in times of catabolism and fasting can cause this severe symptomatic. we terminated the therapie with empaglifozin and under the treatment with insulin the symptoms disappeared within 3 days and the patient could be discharged from the icu on day 17 after surgery. after one episode of ketoacidosis the therapy with sglt2-inhibitors should lifelong never be started again. we recommend that intensivists should be aware of the modern sglt2-inhibitors because of the shown severe complications and the increased use of this medication. consent to publish: written informed consent for publication was obtained from the patient. while obesity confers an increased risk of death in the general population, numerous studies have reported an association between obesity and improved survival among critically ill patients. this contrary finding has been referred to as the obesity paradox. this retrospective study uses two causal inference approaches to address whether the survival of non-obese critically ill patients would have been improved if they had been obese. the study cohort comprises 6,557 adult critically ill patients hospitalized at the intensive care unit of the ghent university hospital between 2015 and 2017. obesity is defined as a body mass index of ≥30 kg/m 2 . two causal inference approaches are used to estimate the average treatment effect in the untreated (atu): a naive approach that uses traditional regression adjustment for confounding and that assumes missingness completely at random, and a robust approach that uses super learning within the targeted maximum likelihood estimation framework and that uses multivariate imputation of missing values under the assumption of missingness at random. obesity is present in 18.9% of patients. the in-hospital mortality is 14.6% in non-obese patients and 13.5% in obese patients. the marginal associational risk difference for in-hospital mortality between obese and non-obese patients is -1.06% (95% confidence interval (ci) -3.23% to 1.11%, p=0.337). the naive approach results in an atu of -2.48% (95% ci -4.80% to -0.15%, p=0.037), whereas the robust approach yields an atu of -0.59% (95% ci -2.77% to 1.60%, p=0.599). a robust causal inference approach that may handle confounding bias due to model misspecification and selection bias due to missing data mitigates the obesity paradox, whereas a naive approach results in even more paradoxical findings. the robust approach does not provide evidence that the survival of non-obese critically ill patients would have been improved if they had been obese. bowel management within an icu environment is often difficult. recent data collection from an intensive care unit at the rvi identified either loose stool or constipation on > 50% of patient days. it was postulated this could be improved with a more tightly controlled bowel management regimen. to test this hypothesis a step-wise bowel protocol was created and introduced. data was collected in the 4 month period following its implementation with the following aims: 1) assess effectiveness of the protocol 2) further observe the reasons for loose or constipated stool on an diarrhea is an important problem in each critically ill pateints [1] . we aimed to investigate the frequency and management of diarrhea in our icu. in this study 47 patient retrospectively reviewed, in our icu between 01.01.2017-03.01.2018. patients were divided into two group as diarrhea "positive" and "negative". patients with diarrhea had fluid or loose stools 3 or more times a day. each diarrhea period of the patients with diarrhea was examined separately and compared with the group without diarrhea. nutritional status, enteral product formulation, leukocyte, neutrophil, albumin values, gastric sparing, antibacterial and antimycotic use, los in hospital and in icu were compared. in diarrhea positive group, on the day of hospitalization, laxative and/or enema administration, toxin a in stool, nitrogen balance before and after diarrhea, enteral product change in diarrhea, probiotic, metronidazole or oral vancomycin use were examined. the incidence of diarrhea was 68.3%. the most common diagnosis of icu admision was respiratory failure (60-85%) in both groups. diarrhea occurred in two days after laxative and/or enema treatment. enteral nutrition was higher in both groups (≥90%). nasogastric tube feeding was significantly higher in the diarrhea group (p=0.041). there was no difference between nutritional product formulation and diarrhea development (p>0,1). antibacterial use was high in both groups (75%); however, teicoplanin use was significantly higher in the group diarrhea negative group (p=0.028). the los in icu, and hospital was higher in diarrhea group (p<0.001). no difference in mortality rates (p>0.5). many factors may cause diarrhea in icu, and diarrhea may adversely affect patient treatment and increase morbidity. we think that preventive methods are as important as the treatment of diarrhea. the use of parenteral glutamine is studied in number of rcts and systemic reviews (heyland d 2013, wischmeyer p 2014), while there is a lack of data about the use of enteral glutamine. the aim of our study was to determine the effect of enteral glutamine supplementation on the incidence of hospital infections and death. design: retrospective cohort study. inclusion criteria: males and females > 18 years of age, tbsa burned 20%-80%, nasogastric intubation.patients were divided in two groups: glutamine group (n=25) and control group (n=17). in the study group enteral glutamine was administered to the patients for 5 days after admission to the icu. baseline characteristics were well balanced between groups. no significant difference was found between groups on patients' age, sex, tbsa, need for mechanical ventilation and rate of inhalation injury. primary outcome was all-cause mortality. secondary outcome was rate of nosocomial infections (skin and skin structure infections (sssi), lower respiratory tract infections, urinary tract infections, bacteremia, sepsis). mortality rate was 6 (24%) and 7 (41%) in the glutamine group and the control group, respectively, p=0.40. rate of nosocomial infections was 14 (56%) in the glutamine group and 11 (65%) in the control group, respectively, р=0.81. rates of sssi, lower respiratory tract infections, urinary tract infections and sepsis did not differ significantly between the groups: 11 (44%) and 6 (35%), p=0.81; 6 (24%) and 7 (41%), р=0.40; 1 (4%) and 1 (6%), р=1.00; 6(24%) and 5 (29%), р= 0.97, respectively. rate of bacteremia was significantly different between the groups: 1 (4%) in the glutamine group and 5 (29%) in the control group, p=0.03. retrospective design is a significant limitation of our study. enteral glutamine supplementation may reduce the incidence of bacteremia in burn patients, but has no influence on the incidence of other nosocomial infections and mortality. further large clinical trials are needed. with outcomes were assessed with multivariable logistic regression and cox proportional hazard analyses, adjusted for baseline risk factors and randomization. in sensitivity analyses, models were further adjusted for key regulators of ketogenesis to assess whether any effect was direct or indirect. late pn increased plasma 3hb as compared with early pn, with maximal effect on day 2 (p<0.0001 for day 1 to 5 and for the "maximal effect" day in the 1142 patients). adjusted for baseline risk and randomization, plasma 3hb associated with a higher likelihood of earlier live weaning from mechanical ventilation (p=0.0002) and of earlier live picu discharge (p=0.004). as plasma 3hb replaced the effect of the randomization, the 3hb effect statistically explained these benefits of the randomization. further adjustment for key regulators of ketogenesis did not alter these findings. plasma 3hb did not independently associate with the risk of infections and mortality. withholding early pn increased ketogenesis in critically ill children, an effect that statistically mediated part of its clinical benefits. critical care patients are prone to frequent feeding interruptions for various reasons including feeding intolerance. these interruptions can lead to adverse outcomes. the aim of the study was to determine the reasons for and the duration of interruptions of enteral nutrition (en). single-center observational, cross-sectional study in a 19-bed mixed icu of a tertiary hospital. duration: 6 months. 50 patients, aged 65.4 years old (±14.6), that stayed in the icu > 48 hrs and were fed with en were included. anthropometric data, bmi, time of initiation of prescribed en, type of en formula, daily calories delivered were recorded. energy intake was calculated according to espen guidelines (25 kcal/ kg bw/day). the causes for and duration of interruption were reviewed from the patient's chart. apache ii and mnutric score was calculated for all patients. mnutric score ≤5 was used to diagnose malnutrition. all patients included in the study were endotracheally intubated. apache ii was 22.4 ±5.6. 58% of patients had increased risk of malnutrition. icu stay was 24.4 (8.0±32.0) days, and the in-hospital mortality was 24%. there were 318 episodes of en interruptions over a median icu stay of 24.4 days. median 2.5 interruptions/patient. the most common reason for en interruption was gastric residual volume monitoring followed by diagnostic and therapeutic procedures (figure 1 ). other reasons include surgery, intolerance and/or delayed feeding and extubation. the median lost feeding time was 5.4 hours/ day (3.7-7.4) for all causes, while the mean loss of total energy intake was 790 kcal/day (±321)/day. average body weight of the patients was 78 kg (±12). caloric deficit was calculated at 1950 kcal/day or 40% of the prescribed caloric goal. the results of this study showed that interruptions can lead to substantial caloric deficit, malnutrition and adverse events. an interruptionminimizing protocol could be useful in order to reduce the missing hours and to improve the clinical outcomes. relationship of goal-directed nutritional adequacy with clinical outcomes in critically ill patients pc tah there are controversies surrounding the effects of optimal nutritional intake on clinical outcomes in critically ill patients. this study aimed at investigating the relationship of goal-directed energy and protein adequacy on clinical outcomes which includes mortality, intensive care unit(icu) and hospital length of stay (los), and length of mechanical ventilation (lomv). this was a single centre prospective observational study. nutritional requirements were guided by indirect calorimetry and 24-h urinary urea.nutritional intake was recorded daily until death, discharge, or until day 14 of icu stay. clinical outcomes were collected from patient's hospital record. the relationship between the two groups (< 80% and ≥80% of overall nutritional requirement) with mortality outcomes was examined by using logistic regression with adjustment for potential confounders. terlipressin, despite being one of the main treatments for acute variceal bleeding, may lead to severe hyponatremia due to its antidiuretic activity.we aimed to identify risk factors for development of hyponatremia during terlipressin treatment. retrospective study of patients admitted to acute intermediate care unit for hypertensive upper gastrointestinal bleeding due to chronic liver disease who received terlipressin(december2011-decem-ber2018).hyponatremia was defined as a decrease in na serum levels ≥5meq and severe hyponatremia as >10meq within 3 days of treatment. we studied 191 patients, 84.3% male, mean age of 58.6 years (sd 10.8). alcohol-related liver disease was the most frequent etiology. hyponatremia occurred in 53 patients (27.7%). serum na δbetween -5 and -10meq and serum na δ>-10meq occurred in 20.4 and 7.3%, respectively (table 1) . severe hyponatremia occurred in 11 patients (5.8%) and symptoms were reported in two cases (status epilepticus and altered mental status). patients with higher baseline levels of na were more susceptible to terlipressin-induced hyponatremia and a longer length of stay was observed in patients with serum naδ>-10 meq (6.3 vs 4.4 days, p<0.022). the prevalence of hyponatremia in our study was lower than previously reported.higher serum na at admission and aih as etiology of cirrhosis were predictors of terlipressin-induced hyponatremia. neither the cumulative dose of terlipressin nor the duration of treatment appear to be related to the development of hyponatremia a δ48h-[na] >5 mmol/l was associated with larger hazards of mortality ( figure 1 ). an increase in serum sodium in the first 48 hours of icu admission is independently associated with a higher mortality in patients admitted with mild hyponatremia, normonatremia, and hypernatremia. based on our findings, it is possible that mild hyponatremia may be a protective mechanism in critical illness, which questions common practice of routinely correcting serum sodium when it is too low. introduction: acute liver failure (alf) represents a life-threatening organ dysfunction associated with increased mortality and liver transplantation represents the only definitive treatment. the aim of this study was to assess the effects of renal replacement therapy in combination with hemoadsorption in alf patients. twenty-nine patients with alf admitted to the intensive care unit (icu) of fundeni clinical institute were included in the study. after icu admission, 3 consecutive session of hemoadsorption in combination with continuous veno-venous hemodiafiltration were applied. number of organ dysfunctions and sirs criteria were recorded at icu admission. the following data were recorded before and after the 3 hemoadsorption therapies: glasgow coma scale, pao2/fio2, creatinine, 24-hours urine output, bilirubin, leucocyte and platelet count, heart rate, mean arterial pressure and vasopressor support, c-reactive protein and procalcitonine. clif-sofa score was calculated before and after the therapy. icu length of stay and 28-days outcome were noted. the mean age in the study group was 34±14 years. the median number of sirs criteria was 3 [2, 4] and the median number of organ dysfunctions was 3 [1, 6] . the use of hemoadsorption was associated with a decrease in creatinine (from 1.9±1.4 to 1.2±0.8 mg/dl, p= 0.02), bilirubin (from 14.2±12.6 to 9.2±9.1 mg/dl, p=0.05) and platelet count (96482±70913 / ul to 51275±24393 /ul, p=0.01). we also observed a decrease in clif-sofa score from 12.0±2.1 to 10.0±2.6 (p= 0.05). overall mortality was 37.9% (n=11). six patients (20.7%) underwent liver transplantation with 100% 28-days survival. the use of hemoadsorption in patients with alf is associated with improvement in liver and kidney functional tests and may represent a new therapy in bridging these patients to liver transplantation. introduction: impairment of intestinal mucosal barrier function is the initiating factor of sepsis. in order to explore the effect of lactic acid bacteria on intestinal barrier function impaired by sepsis, it is necessary to establish sepsis and lactic acid bacteria ecological models. however, how to construct these models is still unclear. co-cultures with a gradient of lactic acid bacteria and caco-2 cells were constructed. the symbiotic state was observed under an inverted microscope and lactate dehydrogenase (ldh) toxicity tests, transepithelial electrical resistance(teer) tests and western blots were used to determine effective concentrations of lactic acid bacteria in monolayer cell models. lipopolysaccharide (lps) was used to treat cells, and cell counting kit-8, quantitative reverse transcription pcr(rt-qpcr) and enzyme linked immunosorbent assays (elisa) were used to determine the appropriate concentration for sepsis models. the number of living cells decreased significantly when the moi(number of lactic acid bacteria/cell number) reached 8 ( figure 1 , panels 1a, b). the release of ldh indicated that damage to cells began to increase when the moi exceeded 10 (panels 2a, b). at an moi of 0.5, resistance values began to increase over time, whereas resistance values began to decrease when the moi reached 10 (panel 3). as the number of lactobacilli increased, the expression of tight junction protein increased and then decreased (panel 4a, b, c). in sepsis model experiments, the cell survival rate began to decrease once the concentration of lps exceeded 10^4 ng/ml (panel 5). rt-qpcr results showed that 10 2 ng/ml lps significantly increased inflammatory cytokines (panel 6), and elisa results consistently showed that tnf-α and il-6 increased significantly when lps concentrations reached 10 2 ng/ml (panel 7a, b). it is feasible to construct a cell monolayer model of lactic acid bacteria and lps. the appropriate moi of lactic acid bacteria is 0.5 and the optimal concentration of lps is 10 2 ng/ml. introduction: sepsis is associated with high mortality and morbidity. as the severity increases, physiological parameters such as ph changes are one of the most notable features in metabolic acidosis secondary to high lactate. currently there is no point of care test other than blood gas measurement that could detect these ph changes. this is challenging especially in prehospital environment. the aim of this study is to develop a novel rapid point of care testing using a sensor to detect ph change in blood. sensors were produced by screen printing graphene and silver electrodes and functionalizing the graphene working electrode with an active layer of melanin. a preclinical sensor model was produced by adding lactic acid to a citrated plasma sample thus altering its ph over a clinically relevant range. the ph sensors were exposed to modified plasma, recording any changes in the voltage. the relationship between the voltage potential and plasma ph was established using weighted least squares regression. a ph dependent change in the measured voltage, with respect to the ph of the solution, was observed with a sensitivity of -111.27 mv/ph +/-15.95 over a physiologically relevant ph range between ph7.1 and ph7.6. in this first phase proof of concept study a low cost, ph sensor was fabricated and demonstrated to be effective in measuring the ph of the plasma. this is the first time that such a sensor has been demonstrated and validated to work in this preclinical model of acidosis. the technology demonstrated here is a promising candidate for a point of care test whereby abnormal blood ph levels can be detected and monitored outside of a laboratory environment in a rapid manner. further studies are now underway to detect this change in whole blood. (figure 1) . over one year only a small proportion of patients (n=7, 4%) were classified as 'intermediate high' risk and potential candidates for reperfusion therapies. the revised national early warning score (news) with modified glasgow prognostic score (mgps) is superior to the news for predicting in-hospital mortality in elderly emergency patients t mitsunaga jikei university school of medicine, emergency medicine, tokyo, japan critical care 2020, 24(suppl 1):p286 the national early warning score (news) was developed in the ukto identify the risk of death. the previous study showed that the modified glasgow prognostic score (mgps) correlate with frailty in elderly patients [1] . the aim of this study is to evaluate the predict value of the revised news with mgps for in-hospital mortality (in 28 days) in elderly emergency patients. this study is secondary analysis and was carried out in jikei university kashiwa hospital, in japan, from 1 april 2017 to 31 march 2018. the acute medical patients aged 65 and older were included. the news was derived from seven physiological vital signs. the mgps was derived from c-reactive protein (crp) and albumin. discrimination was assessed by plotting the receiver operating characteristics (roc) curve and calculating the area under the roc curve (auc). the aucs for predicting in 28 days in-hospital mortality were 0.818 for revised news with mgps and 0.797 for the original news. the auc of the revised news with mgps was significantly higher than that of the original news for predicting in-hospital mortality (p < 0.001) (figure 1) . our single-centred study has demonstrated the utility of the revised news with mgps as a high predictor of acute phase in-hospital mortality in elderly emergency patients. the diagnostic performance of the five main emergency department (ed) triage systems has been shown to be poor in distinguishing acute coronary syndromes (acs) from mild severity diseases in chest pain patients. these ed triage systems are either clinically-based, being more sensitive or ecg-based, more specific [1] . the goal of the study was to evaluate if incorporation of cardiovascular risk factors (cvrf) into ecgbased triage could increase his diagnostic performance. cecidoc is a prospective, observational, single-center study in an academic hospital. all consecutive adult patients admitted for acute chest pain were included. we compared the ecg-based french triage system [2] to a modified system upgrading patients with a normal ecg but significant cardiovascular risk from a low acuity triage score (waiting period before medical assessment of max. 60 min.) to a high acuity triage score (waiting period before medical assessment of max. 20 min.). the final diagnosis was determined after a 30-day follow-up. we predefined as being adequate a high-acuity triage score (level 1 or 2) for acs and a low-acuity score (level 3, 4 or 5) for mild severity diseases. a total of 190 patients was enrolled over a 5-month period (age 56.8 ± 16.4; m/f ratio 1.7). triage scores of 35 patients (18.4%) with acs were compared to 103 patients (54.2%) with mild severity diseases. taking into account cvrf, the sensitivity of the triage system increased from 60 to 80% whereas the specificity decreased from 74 to 61%. area under the roc curve (auc) went from 0.69 to 0.72 (fig. 1) . for chest pain triage at ed, addition of cardiovascular risk factors into ecg-based triage increases his diagnostic performance. approximately 20% of patients presenting to hospital with an intentional overdose require admission to an intensive care unit (icu) [1] . there are currently no uk guidelines regarding the optimal use of ct head scans (cth) in this patient cohort [2, 3] . this study aims to determine whether we should be performing ct head scans in obtunded patients with suspected overdose requiring admission to intensive care. we performed a retrospective search of the icnarc database for plymouth university hospital trust, looking for patients admitted to the icu with overdose or self-poisoning as a primary diagnosis. 146 patients were identified and 86 of these patients required intubation due to obtundation(gcs<8). there were 59 males and 27 females with an average age of 38 years old. the median length of stay on the unit was 1 day. 52 of the patients has a past medical history of mental illness, and 53 overdosed on prescribed medications. the average gcs recorded on admission was 5. 52 of the 86 (64%) patients had a cth on admission, of which 5 were part of a trauma scan. 11 were known overdoses and 7 were suspected overdose as per the cth request form. the main rationale behind those requests were to exclude additional intracranial injury. none of those cth showed any signs of acute pathology (figure 1) . in this retrospective study, obtunded patients with suspected or known overdose with no history of apparent trauma or injury do not benefit from cth. in the absence of a history of trauma or focal neurological signs our conclusions are that cth provides limited value in the management of these patients. the audit was carried out to objectively investigate the problems associated with technique of folley catheterization in emergency department and 3 indoor units of internal medicine wards [1] . introduction: cellular and molecular mechanisms, epigenetic aspects of acute clozapine poisoning are studied insufficiently. the aim of this study was to identify morphological and epigenetic alteratons in brain neurons during acute exposure to clozapine combined wit ethanol. the experiments were carried out on male wistar rats weighting 200-250g (n=21). group i (control) received 0.9% nacl solution enterally; group iiclozapine 150 mg/kg in 0.9% nacl solution; group iiiclozapine 150 mg/kg in 40% ethyl alcohol. after 4 hours euthanasia was performed. autopsy included withdrawal of brain samples for histological examination (n = 21) and for determination of global dna methylation level (n = 21). the global dna methylation level (5-mc%) was determinated by fluorimetric method. inter-group comparisons were made by kruskal-wallis test. histological examination of paraffin sections of brains stained with hematoxylin and eosin was performed by light microscopy. in acute сlozapine poisoning and its combination with ethanol morphological changes in neurons of the cerebral cortex were detected. in acute сlozapine with alcohol poisoning an increase of global dna methylation level was observed. probably the identified changes have a common pathogenesis which will be clarified in our further studies. there is limited information available regarding the prevalence of adder bites and the complications of envenomation. nhs data suggests there are 100 adder bites annually in the uk with the last fatality in 1975 [1] . we performed an audit into adder bites in south west wales to identify the number attending our emergency departments, their management and clinical course as well as any environmental factors that predict increased likelihood of being bitten or the severity of the bite. a retrospective study of adder bites attending emergency departments in south west wales was undertaken (jan 2014 to aug 2019). measurements included were patient demographics, clinical presentation, type of treatment (conservative vs anti-venom) and outcome. results: 31 patients were included, age range 2-72 years ( figure 1 ). the majority of bites occurred in sand dunes (41.9%) and all bites were on extremities. anti-venom was administered to 45.2% (14/31) of patients. there was a significant positive association between the use of anti-venom and the length of hospital stay (r = 0.520; p=0.003) and a significant negative correlation between the anti-venom use and both diastolic and systolic blood pressure (p= 0.501 and 0.487 respectively p=0.01). all patients fully recovered. in this study, we demonstrated that with a full clinical assessment on presentation it is safe to decide whether anti-venom is required. the current guidelines are safe and effective in the treatment of adder bites. 98μmol/l, for pao2 < 9.9kpa and > 12.3 kpa, platelets < 165*10^9/l and > 327*10^9/l, and bilirubin > 12μmol/l. in our population of adult ed patients, the thresholds of vital values associated with increased 7-day mortality were very close to routinely used values, and most of the thresholds were included in the lowest urgency level in triage and risk-stratification scoring systems. the workload in the emergency room: direct assessment by the therapeutic intervention scoring system-76 and indirect assessment by the nasa task introduction: the number of emergency room admissions continues to increase each year, which increases the care workload of the emergency department staff, who should to use its theoretical and practical knowledge in order to provide quality care in difficult working conditions. the aim of our study was to assess the emergency room staff workload its impact on health workers and patients and to suggest an improvement strategy to decrease this workload. a prospective, monocentric cohort study with descriptive and analytic approach over one month (december 2018) conducted at the emergency department of an academic hospital. the workload endured by the emergency room staff was evaluated by the nasa task load index and on patients by the therapeutic intervention scoring system-76. there were 286 cumulative days of hospitalization in 67 consecutive patients admitted to the emergency room. the average age was 61 ±15 years. the average length of stay at the emergency room was about 103 ±48h. the average tiss-76 score was 31.7 ±14.9. factors associated with important care workload were: age ≥ 65 years, diabetes, more than 3 comorbidities, the use of intravenous antibiotics; the use of vasoactive drugs and the use of mechanical ventilation; a high tiss score was predictive of emergency room mortality. in the indirect assessment of the care workload, 41 medical and paramedical staff were interviewed, 73% of them were under 40 years old with a sex ratio of 0.58. a high level of mental and physical workload was expressed by ed staff with considerable level of frustration; the ed staff suggested mainly to improve the working conditions, communication and to redefine tasks "who does what". our study had shown a significant workload in the emergency room, a process to reduce this workload is being implemented medical simulation is a modern teaching tool increasingly used in specialties such as anesthesia, emergency medicine and obstetrics. however, it's not widely used in specialties like cardiology, althought cardiovascular emergencies are very frequent. the purpose of our study was to assess the effectiveness of simulation-based medical education in the management of cardiovascular emergencies among moroccan graduate students. we conducted a prospective, observational, multi-centrer study including the students of three moroccan universities from the 5 th to the 7 th year of medicine who underwent 6 phases: first a pre-test, then a theoretical and practical training on cardiovascular emergencies after which the students were separated in two groups, one undergoing the medical simulation training (group 1) and one who didn't (group 2), followed by a theoretical then a practical post-test on resusci anne and simman®. at last, the students were asked to answer a satisfaction survey. the reform procedure in the tunisian army consists in repairing the physical damage and deciding on the applicant's ability to continue working. terrorism increases the impact of the co-morbidity generated and the socio-economic consequences that result from it. the purpose of this work was to study the epidemiological, clinical and evolutionary profile of terrorist injuries, to specify the rates of consequent partial permanent disability (ppi) and the possibilities of returning to work. descriptive retrospective cross-sectional study of 177 reform files on military personnel injured during anti-terrorist operations from fig. 1 (abstract 296) . changes in total bcpr rate in family-and friends-witnessed ohca cases with dispatcher-assisted instruction during 20-week period after the day of disaster during three years january 2013 to september 2019. the data collection was carried out on the basis of a collection form. our 177 wounded were male, 96% of whom belonged to the army. the average age was 36 years and 3 months ± 8.869. half of our wounded were troopers. infantry and special forces were the most exposed military units. half of the accidents were recorded in the kasserine region (88 cases). chronic post-traumatic stress disorder (cptss) was found in 130 injured, followed by amputations in 18 injured. the after-effects were psychological in 32%, physical in 26% and mixed in 39% of our injured. the ppi rate ranged from 36% to 75% in 23.7% of injuries.. more than half of the injured had returned to their professional activity, 33% were put on reform for health reasons. our results showed that the esptc was the most recorded sequel, and that the ppi rate was significant in a quarter of our injuries. in our series, a third of our wounded were put on reform for health reasons. to state the importance of initial care and adequate and rigorous follow-up to recover a greater number of war wounded. introduction: the rapid response system (rrs) has been shown to decrease hospital mortality [1] . the japanese coalition for patient safety has set a major goal for hospitals to more widely implement the rrs. however, prevalence and actual circumstances of use in acute care hospitals (including small scale hospitals) in japan are as yet not well-known. web-based questionnaires were sent to acute care hospitals (of scale 75 beds-or-larger) of 17 prefectures in western japan. each participant hospital selected a certain department which answered the questionnaire. the rrs included the medical emergency team (met), the rapid response team (rrt), and the critical care outreach team (ccot). we investigated the presence and circumstances of in-hospital emergency calls, rrs and other systems, and then illuminated issues to be solved. our study suggests that delays in patient transfer to the icu after rrt activation in the wards were associated with slower physiological improvement.these findings support further and larger studies. blood and blood products use in intensive care unit m akcivan, s bozbay, o demirkiran istanbul university cerrahpasa, anesthesiology and intensive care, istanbul, turkey critical care 2020, 24(suppl 1):p304 blood and blood product (bp) transfusions are frequently used in intensive care units (icu) [1] . it is important to know transfusion epidemiology and the effect of adverse transfusion reactions and their effect on mortality and morbidity.we aimed to investigate the blood and bp transfusions in the icu. blood and bp transfusions in icu, between 2013-2017 were reviewed retrospectively. we evaluated each transfusion as a data and examined the pre-and post-transfusion laboratory values, demographic data, cause of icu admission and comorbidities. results: 284 patients who underwent transfusion in the icu, and 2188 transfusion data from these patients were included. the most frequent cause of hospitalizations were respiratory failure and sepsis. the rate of patients transfused in the five-year period decreased from 73.9% to 36.67%. the hemoglobin threshold before transfusion decreased from 8.34 g / dl to 7.91 g / dl. a total of 148 transfusion reactions were observed and the most common transfusion reaction was febrile non-hemolytic reaction. the most commonly transfused product was red blood cell suspension. transfusion reactions were found to be slightly higher in men than women in young age group(<65y) (p = 0.44 and p=0.021, respectively). transfusion reactions were found to be more frequent in emergency transfusions (p <0.01). the number of transfusions was significantly lower in patients with apache ii score <20 (p <0.01). the need for transfusion was found to be higher in patients with hematological malignancy (p <0.01). it was observed that as the mean number of transfusions increased the mortality is also increased (p <0.01). transfusion therapies are the treatments that are vital but have a serious mortality and morbidity risk. in particular, intensive care patients should be considered in detail because of their specific features. restrictive transfusion practices have positive results. association between anemia or red blood cell transfusion and outcome in oncologic surgical patients. figure 1a) . the association between rbc transfusion and adverse events also remained after adjustment (or 4.3 [2.2 -8.8] ; p < 0.001) ( figure 1b) . in oncologic surgical critically ill patients, there was an independent association between anemia (even moderate anemia) or rbc transfusion and patient outcomes. our findings highlight the need for further research to determine the optimal transfusion strategy in surgical oncologic patients. transfusion impaired skin blood flow when initially high e cavalcante dos santos, w mongkolpun, p bakos, al alves da cunha, c woitexen campos, jl vincent, j creteur, fs taccone erasme hospital, intensive care department, brussels, belgium critical care 2020, 24(suppl 1):p306 red blood cell transfusion (rbct) increases global oxygen delivery (do 2 ) and may improve microcirculation. however, the effects on blood flow have been found to be conflicting. we studied icu patients with stable hemodynamic status (mean arterial pressure (map) ≥ 65 mmhg for at least 6 hours) and without active bleeding, who received a rbct. skin blood flow (sbf) was determined (periflux system 5000, perimed, index finger; perfusion unit, pu) together with map, heart rate (hr), hemoglobin (hb), lactate levels and scvo 2 before and after rbct. sbf was measured before rbct (t0) and after (t1) for each 3 min. according to previous data indicating the lowest sbf value found in noninfected icu patients was 151 pu, all patients were analyzed according to the baseline sbf (i.e. <151 pu -low sbf vs. ≥151 puhigh sbf). the relative change of sbf (δsbf) was calculated after rbct and the responders were defined by the function of >10%. results: 63 icu patients were studied. rbct was associated with increases in map and scvo 2 but no change in sbf. at baseline, scvo 2 was lower in the responders than in the non-responders (p=0.04) and lower in patients with low sbf than in the high sbf (p=0.04). there was no difference in hb, map, and lactate, between the patients with low and high sbf. after rbct, map rose in the responders (p<0.01) and in the non-responders (p=0.03), sbf (p<0.01) rose in patients with low sbf, and sbf (p=0.02) decreased in patients with high sbf. there was a negative correlation between baseline scvo 2 (r= -0.363, p<0.01) or baseline sbf (r= -0.560, p<0.01) and the relative increase in sbf after rbct. rbct increases skin blood flow only when it is impaired at baseline. severe immune dysregulation is associated with adverse outcomes and is common in intensive care unit (icu) patients [1] . erythropoietin-stimulating agents (esas) have both anti-apoptotic and immune-modulating properties [2] . despite potential benefit, both the safety and efficacy of these agents remains unclear [3] . here we evaluate the impact of esas on morality at hospital discharge in critically unwell adult patients admitted to the icu. we conducted our search strategy in accordance with a predetermined protocol. the use of ffp is associated with an increased incidence of complications such as acute respiratory distress and infections, and the rate of complications increased with the quantities of ffp transfused [1] . pcc contain several important coagulation factors and it has been suggested that they could replace ffp. this has been shown mainly in case reports or series in which coagulation factor deficit was detected by using poc viscoelastic tests in trauma [2] or traditional hemostatic tests in obstetric patients [3] . multicenter observational study of the safety and efficacy of the prothrombin complex concentrate. a survey of anesthetists was conducted in 19 maternity hospitals at various levels of care in the russian federation. data has been collected and processed. as a result, 251 patients were analyzed. pph was determined as a volume of blood loss more than 500 ml during vaginal delivery or cs. the most significant risk factors for pph were: preeclampsia or arterial hypertension and a history of postpartum hemorrhage. 32.3% had no risk factors for pph. it was determined that the use of prothromplex 600 iu decreased the number of patients with transfusion ffp 12-15 ml/kg by 27.8% and increased the number of patients without transfusion by 25.9%, compared with patients without use of prothromplex 600 iu (figure 1 ). no complications were detected. the use of pcc safety and efficacy reduce use of ffp during pph. the full analysis included 45 patients on either hfc (n=22) or cryoprecipitate (n=23). the intraoperative and postoperative changes in etp and fibrinogen concentration are shown in table 1 . for fibtem a20 (intraoperatively) and fibrinogen concentration (intraoperatively and postoperatively), the mean numerical values appeared higher with hfc than cryoprecipitate. fxiii (hfc: 121.86%, 66.85%; cryoprecipitate: 115.55%, 68.68%, at baseline and 4hr after surgery start), fviii and vwf were maintained throughout surgery in both treatment groups. this was also the case for laboratory tests activated partial thromboplastin time, prothrombin time and platelet count. the forma-05 coagulation parameters analyses showed broad overlaps between hfc and cryoprecipitate, with satisfactory maintenance of the clot quality parameters, fxiii concentrations and thrombin generation parameters. the study group includes 118 men and 40 women with a mean age of 43,008 vs. 40.32 years (p=0.639) admitted with the diagnosis of multiple trauma. we found a directly proportional and highly significant statistical correlation between base excess and fibrinogen level diagnosed using the mcf/fibtem parameter(r=0.6382, p<0.0001)and an inverse proportional correlation between lactate level and fibrinogen level (r= -0.2164, p=0.0065). in the roc analysis that uses as a variable the level of base excess and as a criterion of classification the fibrinogen deficit (mcf/fibtem<12 mm) it can be observed that at a value of be<-7 mmol/l, we can diagnose a fibrinogen deficit with a sensitivity of 88.2% and a specificity of 80.6% (auc= 0.872,p< 0.0001). lactate appears to be inferior to the excess base (figure 1) , but still has a good diagnostic power, a value of 2.6 mmol/l has a sensitivity of 67.1% and a specificity of 75% (auc= 0.754,p<0.0001). the difference between the two roc curves (0.118) is statistically significant (p = 0.0028). both base excess and serum lactate can be used to diagnose fibrinogen deficiency with the mention that base excess appears to have a higher sensibility and specificity ability. based goal-directed algorithm. this approach requires further clinical validation. we conducted a retrospective study comparing transfusion strategies in patients with major trauma between 2013 and 2018. we retrieved demographic data and blood products administered from patients with at least one red-blood cell (rbc) transfusion. primary outcome was a reduction of rbc administration. secondary outcomes were mortality, icu length of stay and acute kidney injury. we included 141 patients admitted in the icu due to severe trauma (sapsii:41.5 ±21.9), and mainly after emergent surgery (68.8%). they featured a mean age of 45.3±19.3y, were predominantly male (76.6%) and 73% were in shock. in the first 24 hours of hospital admission a mean of 3.6±4.5 rbc units were administered. most patients received a fibrinogen-based protocol (fbp) (78%), with an average of 5±3g of fibrinogen and 1±3 fresh-frozen plasma (ffp) units, versus 3±4 g of fibrinogen and 6±4 ffp units in the ffp group. the fbp was associated with a decrease administration of rbcs in the first 24 hours (r = -2.6; p < 0.004), even after adjustment for severity (p=0.003) and for tranexamic acid use (p = 0.003). it was associated also with a decrease of platelet transfusion (p=0.004). fibrinogen-based protocol was not associated with a decrease in mortality, acute kidney injury or noradrenaline dose. treatment of tic in past years has progressively changed to a goaldirected fibrinogen-based approach. in our population, the use of fbp lead to a reduction of rbc administration in severe trauma patients. prospective, multicenter, randomized study comparing administration of clotting factor concentrates with a standard massive hemorrhage protocol in severely bleeding trauma patients the objective of this study was to assess the ability of the quantra® qstat® system (hemosonics) to detect coagulopathies in trauma patients. many level 1 trauma centers have adopted whole blood viscoelastic testing, such as rotational thromboelastometry (rotem®, fig. 1 (abstract 315) . study treatment plan instrumentation lab) for directing transfusion therapy in bleeding patients. the quantra qstat system is a cartridge-based point-of-care (poc) device that uses ultrasound to measure viscoelastic properties of whole blood. and provides measures of clot time, clot stiffness and a test of fibrinolytic function. methods: adult subjects were enrolled at two level 1 trauma centers which use a rotem based protocol to guide transfusion decisions. study protocols were approved by the site's ethics committee. for each subject, whole blood samples were drawn upon arrival to the emergency department and again, in some cases, after administration of blood products or antifibrinolytics. samples were analyzed on the quantra (at poc) in parallel to rotem delta (in lab). a total of 54 patients were analyzed. approximately 42% of samples had a low clot stiffness (cs) values suggestive of an hypocoagulable state. the low stiffness values could be attributed to either low platelet contribution (pcs), low fibrinogen contribution (fcs), or a combination ( figure 1) . additionally, 12% of samples showed evidence of hyperfibrinolysis based on the quantra clot stability to lysis parameter. samples analyzed on standard rotem assays showed a lower prevalence of low clot stiffness and fibrinolysis based on extem, fib-tem results. the correlation of cs and fcs vs equivalent rotem parameters was strong with r-values of 0.83 and 0.78, respectively. this first clinical experience with the quantra in trauma patients showed that the qstat cartridge detected coagulopathies associated with critical bleeding and may be useful for directing blood product transfusions in these patients. ability to perform testing at poc may provide additional clinical advantage. the objective of the study was to describe the conditions of use of fibryga® 1g, a new, highly purified, human fibrinogen (hf) recently granted a temporary import authorization for use in congenital and acquired fibrinogen deficiencies in france. observational, non-interventional, non-comparative, retrospective study conducted in 5 french hospital centres using fibryga®. data from patients with fibrinogen deficiency having received fibryga® from december 2017 to july 2019 were retrieved from their medical files. indications, modalities, efficacy and safety outcomes were recorded. indications encompassed non-surgical bleeding (nsb) either spontaneous or traumatic, including post-partum hemorrhage (pph), bleeding during surgery (sb) or administration to prevent bleeding during planned surgery. treatment success was defined as control of the bleeding or hemoglobin loss <20% for bleeding treatment and as absence of major perioperative hemorrhage for pre-surgical prevention. this analysis included 110 patients aged 56,7 ± 17.7 years and 60% were male. all presented an acquired fibrinogen deficiency requiring administration of hf. indications were nsb (n=45, 40.9%) including 15 (13.6%) pph, sb (n=31, 28.2%), and prevention of sb (n=34; 30,9%). cardiac surgeries were the main procedures associated with treatment and prevention of sb. mean total doses of fc were 2.95± 1.66g, 2.00±1.37g and 2.21±1.23g for nsb, sb and prevention of sb. success rates were 88.4% (95%ci 78.8-98.0%), 96.8% (95%ci 90.6-100%) and 91.2% (95%ci 81.6-100%) respectively. for pph, mean dose of hf was 2.53±0.74g with a success rate of 86.7% (95%ci 69.5-100%). overall, tolerance was good. fibrinogen concentrate fibryga® is mostly used for bleeding control. in one third of patients, hf was administered preventively to avoid bleeding during surgery. use of fibryga® was associated with favourable efficacy outcomes. functional testing for tranexamic acid effect duration using modified viscoelastometry t kammerer 1 , p groene 2 , s sappel 2 , p scheiermann 2 , st schaefer 2 1 ruhr-university bochum, institute of anaesthesiology, heart and diabetes center nrw, bad oeynhausen, germany; 2 ludwig-maximilans university, department of anaesthesiology, munich, germany critical care 2020, 24(suppl 1):p318 tranexamic acid (txa) is the gold standard to prevent or treat hyperfibrinolysis [1] . effective plasma concentrations are still under discussion [2] . in this prospective, observational trial using modified viscoelastometry we evaluated the time-course of the antifibrinolytic activity of txa in patients undergoing cardiac surgery. methods: 25 patients were included. modified viscoelastometry (tpa-test) was performed and txa-plasma-concentration, plasminogen-activatorinhibitor-1 (pai-1) and pai-antigen-plasma-concentrations were measured over 96h. additionally, in vitro dose-effect-curves from blood of healthy volunteers were performed. data presented as median with interquartile range (q1/q3). results: txa plasma-concentration was increased compared to baseline (t1:0 μg ml -1 ) at every time-point with a peak concentration 30min (t2) after application (p<0.0001; see fig.1a ). lysis was inhibited from 30min (lysistime tpa-test : p<0.01; lysisonsettime tpa-test :p<0.0001). maximumlysis tpa-test was decreased at t2 (t1:97% (96/97) vs. t2: 9% (6/11); p<0.0001). of note, after 24h some patients (n=17) had normalized lysis whereas others (n=8) had strong lysis inhibition (ml< 30%;p<0.05) up to 96h. high and low lysis groups differed regarding kidney function (cystatin c:1.64mg l -1 (1.42/2.02) vs. 1.28mg l -1 (1.01/ 1.52);p=0.002) and active pai-1 (93.05ng ml -1 (33.15/9100.0) vs. 16.13ng ml -1 (6.62/79.98);p=0.047). in-vitro, txa concentrations >10μg ml -1 were effective to inhibit fibrinolysis. in our trial, after 24h there was still completely blocked lysis in patients with moderate renal impairment. this could be critical with respect to postoperative thromboembolic events [3] . here modified viscoelastometry could be helpful to detect the individual fibrinolytic capacity. introduction: peri-operative coagulopathy correction based on viscoelastic hemostatic assays (vhas) and single-factor coagulation products has changed the paradigm of bleeding management in cardiac surgery [1] . in a retrospective study, we analysed patients with emergency surgery for thoracic acute aortic dissection (taad), before and after the introduction of fibrinogen concentrate in clinical practice. data were collected from paper and electronic records. the study was approved by the institutional ethical committee. 60 patients were included in the analysis, 19 operated in 2012, before fibrinogen concentrate was approved for human use, and 41 in 2018-2019. therapy was guided by a rotational thrombo-elastometry (rotem) algorithm. exclusion criteria were non-compliance with the institutional protocol and intra-operative death. we investigated allogeneic blood transfusion (abt), fibrinogen use, peri-operative bleeding (pob), surgical reexploration and post-operative complications (poc). the groups were similar in gender, age, body weight, additive euro-score and aortic cross-clamp time. fresh frozen plasma, cryoprecipitate and red blood cell transfusion were lower in the fibrinogen group, but not platelet transfusion (table). 48,7% of patients in the study group received fibrinogen concentrate and median dose was 2 g (iqr 2-3). day 1 postoperative chest tube drainage and surgical reexploration were significantly lower. there were no differences in stroke, renal replacement therapy, mechanical ventilation time and icu stay. in patients with taad surgery, rotem-guided algorithms which include fibrinogen concentrate are associated with less (pob), surgical re-exploration and abt. further research is needed to document the role of vhas and concentrated factors in reducing (poc). andexanet alfa (aa, portola pharmaceuticals, san francisco, ca) represents a modified factor xa agent which is approved antidote for apixaban and rivaroxaban. andexanet alfa may also neutralize the anti-xa effects of betrixaban and edoxaban. this study aims to compare the relative neutralization of these four anti-xa agents by andexanet alfa in different matrices. andexanet alfa was diluted at 10 mg/ml. apixaban (a), betrixaban (b), edoxaban (e) and rivaroxaban (r) were diluted in ph 8.4, 0.5 m tris buffer (tb), blood bank plasma (bbp) and in 5% albuminated buffer (ab) at 0.062 -1.0 ug/ml. anti-xa activities of all four agents were measured in three systems and the reversibility indices of aa were profiled. the reversibility index (ri 50 ) of anti-xa effects by aa was determined at 25 -100 ug/ml. each of the four agents produced varying degrees of inhibition of anti-xa at 0.062 -1.0 ug/ml, the ic 50 ranged 0.61 -1.53 ug/ml in bbp, 0.47 -1.28 ug/ml in ab and 0.49 -1.4 ug/ml in tb. andexanet alfa produced a concentration dependent reversal of all four anti-xa agents. in the bbp, the ri 50 values for a (192 ug/ml), b (32 ug/ml), e (152 ug/ml) and r (85 ug/ml). in the ab, the ri 50 values for a (140 ug/ml), b (46 ug/ml), e (176 ug/ml) and r (58 ug/ml). in the tb, the ri 50 values for a (154 ug/ml), b (79 ug/ml), e (>400 ug/ml) and r (110 ug/ml). each of the four anti-xa agents exhibit varying degrees of matrix independent anti-xa potencies in different systems, the collective order follows edoxaban > apixaban > betrixaban > rivaroxaban. andexanet alfa produced matrix dependent differential neutralization of the anti-xa effects of these agents. individualized dosing of andexanet alfa may be required to obtain desirable clinical results. the diagnostic and prognostic value of thromboelastogram (teg) in sepsis has not been determined. this study aimed to assess whether teg is an early predictor of coagulopathy [1, 2] and is associated with mortality in patients with sepsis. in total, 518 patients with sepsis on intensive care unit admission were prospectively evaluated. we measured teg and conventional coagulation tests(ccts)on preadmission and observed for development of 1, 3 days and 1, 3, 7days respectively. multivariable logistic regression was utilized to determine odds of icu/hospital mortality. the parameter of teg (maximum amplitude, reaction time; ma/r ratio) was calculated to evaluate sepsis-induced coagulopathy. the admission patients were divided into three groupsma/r0 group(ma/r= 5-14mm/min); ma/r1group(ma/r>14 mm/min)and ma/r2 group(ma/r<5mm/min). in our cohort of patients with severe sepsis, coagulopathy defined by ma/r ratio was associated with increased risk of icu/hospital mortality. introduction: blood sampling for coagulation assessment is often carried out in either arterial or venous samples in the intensive care unit (icu). there is controversy as to the accuracy of this method due to the inherent differences in physicochemical properties as well as the underlying effects of individual diseases in arterial and venous blood. clot microstructure has shown to be a new biomarker (fractal dimension-d f ) which encompasses the effects of diseases in all aspects of the coagulation system [1, 2] . in this study, we compared the effect of all these factors in venous and arterial blood to see if there is a difference in the clot microstructure and quality. 45 patients admitted to a tertiary intensive care unit and busy teaching hospital were recruited. arterial and venous blood was sampled from an arterial line and central venous catheter in situ from the same patient. standard markers of coagulation (pt, aptt, fibrinogen, full blood count), rotational thromboelastometry (rotem), whole blood impedance aggregometry and measured clot microstructure (d f ) were measured on both arterial and venous samples. no significant difference was observed in standard laboratory markers, rotem and platelet aggregation between arterial and venous blood. there were no differences in the fractal dimension (d f ) between the arterial and venous blood samples (d f 1.658 ± 0.10 vs 1.654 ± 0.08 respectively, p=0.830). samples from patients with critical illness give comparable results from either arterial or venous blood despite their underlying pathophysiological process or treatment. this confirms blood for coagulation testing can be taken from arterial or venous blood. clinicians in the emergency setting use a wide range of hemostatic markers to diagnose and monitor disease and treatment. current methods rely on the anticoagulant effect of citrate on whole blood prior to laboratory analysis. despite the well-recognized modulatory effects of citrate on hemostasis, the use of anticoagulated blood has clear analytical advantages, including repeat sampling and storage. however by altering the physiological state of the blood reproducibility and accuracy of the test is affected. recent studies have shown the potential of a novel functional biomarker of clot formation: fractal dimension (d f ), that may give an improved diagnostic accuracy. in this study we assessed the potential of this new biomarker in scientifically measuring the effects of recalcification of citrated samples. methods: 35 healthy volunteers were included. unadulterated and sodium citrate samples of blood were taken from each volunteer. citrated samples were recalcified using (1m cacl 2 ). in the study we compared unadulterated whole blood d f results to citrated d f results and repeated the citrated d f experiments 5 times for each sample over a 2 hour period to ascertain reproducibility. the d f of citrated blood was significantly lower than that of unadulterated blood (1.57±0.04 vs 1.69±0.04, p<0.001). the results of the citrate samples when tested 5 times over 2 hrs gave a coefficient of variation of 1.16%. for the first time we show that a functional biomarker of clot microstructure, d f , can precisely quantify and measure accurately the direct effect that the addition of the anticoagulant sodium citrate has on whole blood clot microstructure. the study also shows that the test is reproducible and has potential utility as a biomarker of acute disease in the emergency setting in citrated blood. this procedure now needs to be evaluated in a group of acute disease states. in this study, we analyzed the hematological abnormalities of dengue patients by thromboelastography (teg) at initial and 1-hour of fluid resuscitation. methods: this is a cross-sectional study evaluating teg readings of dengue patients with different severities presenting to the emergency department. laboratory confirmed dengue patient (positive ns1 antigen or igg/igm) was consecutively sampled. teg readings were taken at presentation and after 1-hour of fluid resuscitation. twenty dengue patients with varying severity had a median reaction time (r), α -angle, k time, maximum amplitude (ma) and lysis 30% (ly30) of 0.495 min, 68.74 ο , 3.58 min, 44.64 mm and 0.54% respectively. mean fibrinogen was normal before and after fluid infusion. there is a non-significant reduction in ma with prolongation of other teg parameters between different dengue severities. there is a statistically significant reduction of α-angle and ma between pre and post 1-hour fluid resuscitation (p=0.019 and p=0.040). normal fibrinogen with low ma, which signifies a weak clot strength, may indicate either a platelet reduction, platelet dysfunction or both. reduction in ma and α-angle post fluid resuscitation is an alarming finding. this is in contrast with previous teg studies although none of it used normal saline exclusively, studied initial fluid resuscitation in emergency department settings or studied a subject with dengue. a bigger study, especially in severe dengue is needed to validate our findings. agreement between the thromboelastography reaction time parameter using fresh and citrated whole blood during extracorporeal membrane oxygenation with teg®5000 and teg®6s m panigada, s de falco, n bottino, p properzi, g grasselli, a pesenti fondazione irccs ca´granda ospedale maggiore policlinico, intensive care unit, milano, italy critical care 2020, 24(suppl 1):p327 the r (reaction time) parameter of kaolin-activated thromboelastography (teg) may be used to assess the degree of heparinization of blood during ecmo. a teg analysis is usually performed on two types of samples: fresh (f) or citrated-recalcified (c) whole blood. teg®5000 can perform the analysis on c and f whole blood, the new teg®6s (haemonetics corp., ma, usa) only on c whole blood. aim of the study was to compare the response of r to heparin using the two types of samples and two teg devices methods: during a three months period at fondazione irccs ca' granda -policlinico of milan, teg was performed (using teg5000® and teg 6s® with and without heparinase, an enzyme that degrades heparin) on 13 consecutive ecmo patients (as part of the gatra study, nct03208270) and in 8 consecutive non-ecmo patients in whom a teg was requested for clinical purposes. bland altman analysis and lin's concordance correlation coefficient were used to assess agreement results: a total of 84 paired samples were taken (74 in-ecmo and 10 off-ecmo). ecmo patients received 19.2 (12.6-25.8) iu/kg/h of heparin. among non-ecmo patients, 5 of them did not receive any dose of heparin, two of them a very low prophylactic dose (1.6 and 2.9 iu/ kg/h, respectively), and one of them 13.1 iu/kg/h of heparin. using teg®5000, r was -21.0 (-65.5; 23.4) min shorter on c compared to f blood in patients receiving heparin (this difference disappeared using heparinase) and only -1.58 (-8.70; 5.54) min shorter in patients notreceiving heparin. r was -26.6 (-77.3; 24.2) min shorter using teg®6s (which performs the analysis only on c blood) than teg®5000 on f blood (figure 1) . when evaluating the effect of heparin using teg, clinicians should be aware that results obtained using citrated-recalcified or fresh whole blood are not interchangeable. using citrated-recalcified blood to perform teg might lead to underestimation of the effect of heparin trauma patients are at high risk for venous thromboembolism (vte). the 2002 east guidelines recommend low molecular weight heparin (lmwh) for vte prevention and antixa monitoring after initiation of the medication or after adjusting doses in certain populations [1] . studies have shown standard enoxaparin dosing of 30 mg every 12 hours may result in low antixa levels [2] . this study aims to evaluate the efficacy of a pharmacist-lead protocol for adjusting enoxaparin dosing based on antixa levels in trauma patients. this single center retrospective chart review included adult trauma patients admitted from 3/1/2018 to 9/20/2019. per protocol, patients with body mass index (bmi) ≤40 kg/m 2 were initiated on enoxaparin 30 mg twice daily, and patients with bmi > 40 kg/m 2 were initiated on enoxaparin 40 mg twice daily. peak antixa levels were drawn 4 to 6 hours after at least the third dose of enoxaparin with a goal therapeutic range of 0.2-0.4 iu/ml. the primary objective was time in days to goal peak antixa level. secondary objectives include vte occurrence, bleeding attributed to lmwh, and dosing regimens utilized. subgroups were analyzed based on body mass index (bmi). of 511 patients identified, 375 patients met inclusion criteria. median time to therapeutic antixa level was 3 days (iqr 2-6). of 337 patients fig. 1 (abstract 327) . agreement between teg®6s and r teg®5000 on citrated recalcified and fresh whole blood with bmi ≤ 40 kg/m 2 , 319 patients (94.7%) were dosed initially per protocol and 153/319 patients (48.0%) met goal antixa level at first check (table 1) . of 38 patients with bmi > 40 kg/m 2 , 24 patients (63.1%) were dosed initially per protocol and 8/24 patients (33.3%) met goal antixa level at first check. our results indicate the protocol is safe due to lack of bleeding attributed to enoxaparin, but less than 50% of patients achieved goal antixa level at first check. however, despite low rates of achieving goal antixa level, vte rates also remained low. introduction: most patients in the icu are given prophylactic anticoagulation with a fixed dose of 40 mg once daily of enoxaparin (clexane) if cct is normal and 20 mg if cct is low. studies on non icu patients have shown that afxa is below desired range for venous thromboembolism (vte) prevention. in the icu, many factors might influence afxa levels including weight, creatinine clearance (cct), shock and other medication. atxa activity was not yet reported in a big mixed icu population with variable morbidity. our study hypothesis is that enoxaparin is underdosed in most cases and routine afxa activity should be monitored in all icu patients. preventive enoxaparin (40 mg qd) was given to all patients unless therapeutic dose was needed or contraindication existed. levels of afxa activity were taken 4 hours after the 3 rd dose. therapeutic vte preventive effect was defined as afxa activity of 0.2-0.5. patient data was collected from medical files. the study is still ongoing, preliminary results were analyzed for 31 patients. 14 of 31 patients (45%) had afxa activity below normal (subtherapeutic). weight and cct were negatively correlated with afxa activity (figure 1 ). mean weight in the subtherapeutic afxa was significantly higher than the therapeutic group (83.2 vs. 72.6 respectively, p=0.031). cct in the subtherapeutic afxa was significantly higher than the therapeutic group (95.1 vs. 60.4 respectively, p= 0.003). the normal cct group (>50) had significantly more patients with subtherapeutic afxa (13 vs 8, p=0.017). in our icu, 45% of the patients receive insufficient vte prophylaxis. overweight patients and patients with normal cct should probably receive higher enoxaprin dose. afxa activity should be routinely monitored in icu patients. in this study we use a new bedside biomarker to test its ability to measure anticoagulation effects on patients who present with acute first time deep vein thrombosis (dvt). dvt requires oral anticoagulants to prevent progression to potentially fatal pulmonary embolism and recurrence. therapeutic efficacy monitoring of direct oral anticoagulants (doac) including rivaroxaban is problematic as no reliable test is currently available. advances in hemorheological techniques have created a functional coagulation biomarker at the gel point (gp) which allows quantitative assessment of: time to the gel point (t gp ), fractal dimension (d f ) and elasticity (g') [1, 2] . the prospective observational cohort study measured t gp , d f , g', standard coagulation and cellular markers in first time dvt patients at three sample points: pre-treatment and approximately 20 and 60 days following 15mg bd and 20mg od rivaroxaban respectively. strict inclusion and exclusion criteria applied. results: 40 dvt patients (mean age 64 years [sd±14.8]; 23 male, 17 female) and 177 non-dvt patients were well matched for age, gender and co-morbidities. mean t gp on admission was 267s (sd±63.3s) and 262.9s (sd±73.5s) for dvt and non-dvt respectively. doac therapy significantly increased t gp to 392.3s (sd±135.7s) after 20 days, and subsequently increased to 395.5s (sd±194.2s) at 60 days as shown in table 1 . d f , g' and standard hemostatic markers all remain within the normal range. conclusions: t gp demonstrates its utility in determining the anticoagulant effect of rivaroxaban. the significant difference in t gp between males and females needs further exploration. localized stasis as a result of transient provoking factors appears not to generate a systemic strength fig. 1 (abstract p329) . correlation of anti factor xa activity with patient cct and weight. anti fxa activity value below 0.2 (red line), was considered "non-effective prevention" introduction: trauma remains the leading cause of death all over the world. to better exploit the trauma care system, precise diagnosis of the injury site and prompt control of bleeding are essential. here, we created a nursing protocol for initial medical care for trauma. the aim of this study was to evaluate the impact of protocoled nursing care for trauma on measures of quality performance. this was a retrospective historical control study, consisted of consecutive severe trauma patients (injury severity score >16). people were divided into two groups: protocoled group (from april 2017 to march 2019) and control group (from april 2014 to march 2017). we set the primary endpoint as mortality for bleeding. the secondary endpoints included time allotted from arrival to start of ct scan and surgery, administration rate of several drugs (sedations, painkillers, preoperative antibiotics, and tranexamic acid). for the statistical analysis, continuous variables were expressed as median (interquartile range) and were compared by wilcoxon rank sum tests given a nonnormal distribution of the data. we included 152 patients in the study: 84 in the control group before the introduction of the protocol, 68 in the protocoled group. as a primary endpoint, the mortality for bleeding was similar between two groups (5% in the control group and 5% in the protocoled group). as a secondary endpoint, the time to ct initiation [group a 11 (7-16) min vs group b 7 (5-10) min; p <0.001], and emergency procedure [group a 41 (33-56) min vs group b (29-43); p <0.001] were shortened by the protocol introduction. furthermore, the administration rates of sedations, painkillers, preoperative antibiotics, and tranexamic acid were increased in the protocoled group compared with the control group. although the mortality as a patient-oriented outcome was not affected, improved quality of medical care by nursing protocol introduction may be suggested in this analysis. this single-institutional prospective study included 72 patients with uprf who were admitted to the trauma surgical intensive care unit (tsicu) and survived until discharge to home between 2012 and 2017. we evaluated the activities of daily living after the discharge using physical and mental component scores of sf-36® and defined physical dysfunction (pd) as physical function (pf-n) score of 40 or less. we divided the patients in the pd (n=34) and control (without pd, n=38) groups and compared the groups. the patients had experienced blunt injuries, including falls (38%) and pedestrian injuries (33%). the mean age was 59.9 years (men: 65.3%); the median injury severity score was 22 (interquartile range: 13-29); and the mean length of tsicu stay was 3.3 days. the average period from the injury until the survey was 34.1 months. there was no difference between the pd group and the control group in the patient characteristics, fracture type, pelvic fixation, and complications. at the time of the survey, the pd group had significantly more painful complaints than the control group (pd: 67.6%, c: 23.6%, p <0.01), and had more physical and mental problems. the sf-36®subscale score showed a significant positive correlation between physical function and body pain, mental health respectively. the percentage of those who were able to return to work was not different in both groups (pd:26.5%, c:47.4%). in the multivariate analysis of pd, only age (odds ratio: 1.039, 95% ci: 1.00-1.07, p = 0.03) was relevant. long-term pd was observed in 47% of patients with uprf. the elderly were particularly prominent, and there was an association between pain and mental health. cells (rbc) this can lead to inhibition of oxygen transport function and development of hypoxia. currently used methods for analyzing the state of rbc either do not have sufficient accuracy or require lengthy analysis and expensive equipment. the use of a simpler and more informative electrochemical approach to assessing the state of rbc is very promising. electrochemical measurements in rbc suspensions (~5 • 109 cells / l) were carried out in a special electrochemical cell [1] in the potentiodynamic mode in the potential range from -0.5 to +1.2 v using the ipc pro mf potentiostat (kronas, russia); optical measurements were performed using an eclipse ts100 inverted microscope (nikon, japan), a cfi s plan fluor elwd 60x / 0.70 lens (nikon, japan); rbc morphology was recorded in real time using a ds-fi1 digital camera (nikon, japan). when examining rbc of patients with severe multiple trauma a decrease in the ability of rbc to change their shape during electrochemical exposure was observed, indicating a decrease in deformability, which can lead to a disruption in the oxygen supply to tissues. at the same time, with the stabilization of the patient's condition a restoration of the ability of rbc to change morphology was detected which in turn could have a positive effect on the rheological characteristics of the blood (fig. 1) . the results of the analysis of red blood cells using electrochemical changes in their morphology can be used as an additional method for the diagnosis of critical conditions. severe trauma should be treated immediately. whole-body ct (wbct) is widely accepted to improve the accuracy of detecting injuries. however, it remains the problem of time-consuming. therefore, we focused on the scout image taken in advance of wbct. detecting major traumatic injuries from a single scout image would reduce the time to start treatment. a previous study suggested that even specialists could not easily find chest and pelvic injuries using wbct scout image alone. in this study, we aimed to develop and validate deep neural network (dnn) models detecting pneumo/hemothorax and pelvic fracture from wbct scouts. we retrospectively collected 2088 anonymous wbct scouts together with their clinical reports at the osaka general medical center between january 1, 2013, and december 31, 2017. we excluded incomplete, younger than 7 years old, postoperative, and poorly depicted images. the part of this dataset from january 1, 2017, until december 31, 2017, was used for validation and the rest for training dnn models. pneumo/hemothorax detection model and pelvic fracture detection model were trained respectively. accuracy, and areas under the receiver operating characteristic curves (aucs) were used to assess the models. the training dataset for pneumo/hemothorax contained 984 images (mean age 48 years; 30% female patients), and for pelvic fracture consisted of 783 images (48 years; 28%). the validation dataset for the former contained 258 images (54 years; 30%), and for the latter consisted of 186 images (55 years; 24%). the models achieved 59% accuracy and an auc of 0.57 for detecting pneumo/hemothorax, 72% and 0.62 for pelvic fracture. our results show that dnn models can potentially identify pneumo/ hemothorax and pelvic fracture from wbct scouts. increasing the number of samples, dnn model could accurately detect severe trauma injuries using wbct scout image. clinical information system (cis) is a computer system used in collecting, processing, and presenting data for patient care. it can reduce staff workload and errors; help in monitoring quality of care; track staff's compliance to care bundles; and provide data for research purpose. however, the transition from paper record format to electronic record involves changes in all kind of workflow in icu. therefore, an effective, efficient and evaluative rollout plan was required to minimize the risk that might arise from the new practice. methods: 1. small groups training were provided. a working station with different case scenarios were set up for practices. 2. individual tutorials were conducted to clarify questions. emphasis on patient care was always top priority. 3. contingency plans were available in case of server breakdown and power failure. downtime drills were conducted to prepare the staff in emergency situations. 4. step-by-step transition from paper record to electronic format was gradually carried out. a plan was discussed among cis team with clear dates and goals. 5. new items in cis were first reviewed and amended in team meeting until consensus was made; then were promulgated to all staffs during handover before implementation. fig. 1 (abstract p333) . the effect of therapy on the electrochemically induced change in the morphology of red blood cells in patients with combined trauma 6. staff compliance and outcomes were then monitored; further review and amendment would be possible if necessary. cis roll-out plan was smooth. all staffs were able to integrate cis into the daily routine. the contingency plans were well acknowledged. new items were followed as planned. ongoing enhancement in cis was put forward on nursing orders, handover summary, and integration with inpatient medication order entry (ipmoe) system. with emerging benefits cis brings along, our staff has more time to devote to direct patient care. human input in data interpretation and clinical judgment on top of cis play an irreplaceable role in patient care. the daily request for laboratory tests in intensive care units is a common practice. although common, this strategy is not supported, since more than 50% of the exams requested with this rationale may be within the normal range [1] . misconduct based on misleading results, anemia, delirium and unnecessary increase in costs may happen [2] . we have developed a strategy to reduce laboratory tests without clinical rationale. observational retrospective study, from july 2018 to june 2019. the number and type of laboratory orders requested, the epidemiological profile of hospitalized patients, the use of advanced supports, the average length of icu stay and the impact in outcomes such as mortality and hospital discharge at a private tertiary general hospital in the city of rio de janeiro / rj -brazil were analyzed. a strategy was implemented to reduce the request for exams considered unnecessary. approximately 1,300 patients underwent icu during this period. the epidemiological profile and severity of patients admitted to the unit were similar to those observed historically. there was a significant reduction (> 50%) in the request for laboratory tests and there was no negative impact on outcomes such as mortality, mean length of stay and no greater use of invasive resources. over the period evaluated, the estimated savings from reducing the need for unnecessary exams were approximately $ 150,000 per year. the rational use of resources in the icu should be increasingly prioritized and the request for routine laboratory tests reviewed. a strategy that avoids such waste, when properly implemented, enables proper care, reducing costs and ensuring quality without compromising safety. evaluating the medication reconciliation errors in 2 icus after implementing a hospital-wide integrated electronic health record system a rosillette, r shulman, y jani university college hospital, centre for medicines optimisation research and education, london, united kingdom critical care 2020, 24(suppl 1):p337 introduction: medication errors in intensive care unit (icu) are frequent [1] and can arise from a number of causes including transition of care. our aim was to investigate the impact of an integrated electronic health record system (ehrs) on medication reconciliation (mr) errors occurring at 2 critical steps: during the transition from an icu to the hospital ward and from the ward to hospital discharge. the objective was to examine the influence of icu admission on long-term medication. we performed a monocentric study in 2 icus of a university-affiliated hospital using drug chart and medical notes review to identify mr errors before, during and after icu admission. data were collected retrospectively from ehrs for 50 consecutive patients discharged from the icu between 1 june-31 july 2019, and who were newly initiated on specific drugs of interest. results: 129 drugs of interest were initiated in icu. many of these were continued after hospital discharge as shown in table 1 . there was appropriate discontinuation of all the antipsychotics newly initiated in icu. other than anticoagulants, there was no reason documented for continuation of the initiated drugs. the planned durations were documented more often after hospital discharge than icu discharge for the following drug classes (% of patients with a plan after icu discharge to the ward; % after home discharge): antibiotics (47.6%; 52.6%), and steroids (45.4%; 57.1%), but less so for analgesics (36.3%; 14.3%), insomnia (0.0%; 20.0%), and gastroprotective drugs (0.0%; 20.0%). our study has shown that medications initiated in the icu can be inadvertently continued at icu and hospital discharge due to failure in documenting indication or duration. systems are required to deprescribe icu only drugs at discharge or communicate a plan for ongoing treatment. introduction: the surviving sepsis campaign advocates the use of care bundles to guide the management of sepsis and septic shock [1] . our study aim was to assess compliance with a locally introduced sepsis pathway and to review intensive care unit admission outcomes. we carried out a prospective audit of patients admitted to the icu at royal surrey county hospital with a diagnosis of sepsis between 19/ 3/19 and 19/11/19, assessing compliance with local sepsis bundle delivery, outcome of icu admission and degree of associated organ dysfunction. results: 119 patients were identified, 71 male (59.7%), with a mean age of 65.7 (18-96). mean 1 st 24 hour sofa score on icu was 6.65 (2-15). 81% of patients required vasopressors, with 67% requiring noradrenaline >0.1mcg/kg/min, and 19% requiring an additional vasopressor/ inotrope. 36% required niv, 32% invasive ventilation and 15% rrt. icu mortality was 15%, in-hospital mortality 24%, mean icu stay 8 days (1-49), and mean length of hospital stay 28 days . in the presence of septic shock mortality was 47% with post-resuscitation lactate >4, versus 21% in patients with no vasopressor requirement or lactate <2 (p<0.05). the sepsis bundle was delivered in one hour to 61 patients (51%). where the bundle wasn't completed, antibiotics were delayed in 26% of cases and blood cultures weren't taken in 66%. where the bundle was fully delivered, unit mortality was 12% vs. 21% where it was not (p<0.05), but there was no significant difference in hospital mortality (26% vs. 30%, p>0.5) or rates of vasopressor requirement, niv, ippv or rrt. there is room for improvement in timely delivery of the sepsis bundle in our hospital and various measures are being instituted. though there was no significant difference in hospital mortality, icu mortality was significantly lower in patients when the bundle was fully delivered. surviving sepsis campaign recommends 3h and 6 h sepsis resuscitation bundle for sepsis. the study was done to assess the feasibility of the guideline and the compliance to sepsis-3 recommendations at an emergency department. prospective interventional study was conducted during one year. were involved in the study all sepsis cases with a qsofa ≥ 2. were assessed a composite of six components (measurement of serum lactate, obtaining blood culture before antibiotic administration and provision of broad-spectrum antibiotic before the end of h3 and provision of fluid bolus in hypotension, attainment of target central venous pressure assessed by cardiac ultrasonography, target lactate to normal level before the end of h6). time base line was the first medical contact at triage zone. secondary outcomes of study were the mortality rate and length of stay at intensive care unit (icu). were involved in the study, 128 patients (mean age 54±17 years, sex ration 2,3). pulmonary infections were the main cause of sepsis (37%) and urinary tracts infections (22%). at h3 components were achieved in 79% of cases [lactates (100%), blood culture (82%) and provision of antibiotics (79%)]. at h6 components were executed in 64% of cases (fluid provision achievement in 89%, ultrasonography assessment in 64% and normal lactate target achieved in 71%) (figure 1 ). the reliability-adjusted rate for completion of the 3 hours and 6 hours bundle was at 68%. patients compliant to composite bundle got the mortality benefit (odds ratios = 0.31, 95% [confidence interval, 0.11-0.72]). the study, however, did not show any benefits of mean intensive care unit (icu) length of stay. faisability of 3-6h bundle ratio was at 68%. it has shown a significant improvement in adaptation and mortality benefit without reducing mean hospital/icu length of stay. more adapted procedures are needed to improve results targeting full compliance of patients to the 3-6h bundle sepsis management. patterns and outcome of critical care admissions with sepsis in a resource limited setting m edirisooriya maddumage 1 , y gunasekara 2 , d priyankara 1 1 national hospital of sri lanka, medical intensive care unit, colombo 10, sri lanka; 2 sri jayawardenepura general hospital, department of critical care, nugegoda, sri lanka critical care 2020, 24(suppl 1):p340 introduction: paucity of epidemiological data is a major barrier in expansion of critical care services, especially in resource limited settings. we evaluated the patterns and the outcome of critically ill patients with sepsis admitted to a level 3 medical intensive care unit in sri lanka. a retrospective cohort study was performed to describe the characteristics and outcome of patients with sepsis, admitted to a medical intensive care unit. sepsis is defined according to sepsis 3 definition. we examined 360 critically ill patients admitted over a period of 6 months. sepsis was the commonest presentation, accounted for 63.6 % of all admissions. mean age was 49.2 ± 16.1 years. septic shock was present in 41.4% on admission. pneumonia (35.4%) was the commonest cause, while leptospirosis (17.9%) and meningoencephalitis (12.2%) accounted for fig. 1 (abstract p339) . sepsis 3-6h bundle components (% of goals achievment) second and third commonest causes of sepsis respectively. the sofa score on admission (8.8 ± 5.0 vs 7.5 ± 4.9, p< 0.025), occurrence of aki (62% vs 49.6%, p<0.02) and the length of icu stay (8.8 days vs 6.2 days, p <0.001), were significantly higher in sepsis than in patients without sepsis. icu mortality in sepsis (n=92) did not show a significant difference to nortality (n= 45) in those without sepsis (40% vs 35%, p= 0.5). patients with leptospirosis had a mean sofa score of 13.3, however the mortality (37.5% vs 40%, p = 0.75) was similar to others with sepsis. in contrast, mortality related to sepsis was significantly high ( 60%, p< 0.02) in the packground of immunosuppression (n= 35). respiratory failure secondary to pneumonia was the commonest cause of critical care admission with sepsis. sepsis related icu mortality was high in the background of immunosuppression. introduction: training in placement, and the subsequent safe confirmation of position, of a nasogastric (ng) tube, relies on clinicians completing an e-learning module at our trust. feeding through an incorrectly placed ng tube is a 'never event,' associated with significant morbidity and mortality [1] . analysis of these incidents reveal that the misinterpretation of chest radiographs, by medical staff, who had not received competency-based training, is the most frequent cause [2] . e-learning has revolutionized the delivery of medical education [3] , however, there are barriers to its use [4] . we hypothesized that, by taking e-learning content, and delivering it face-to-face, we would improve training rates, and thus patient safety. a questionnaire was completed by 50 critical care doctors, concerning their knowledge of the existence of the e-learning module, whether they had completed formal training in ng tube placement, and how confident they were, on confirming correct positioning, using a 5 point likert scale. all clinicians underwent training in the interpretation of ng placement, using chest radiographs. after the session they were asked to re-appraise how confident they felt. results were compared using paired t tests. confidence improved in all, rising from a pre-test average score of 3.74 (sd=0.92), to post-session 4.76 (sd=0.48), p=<0.0001. prior to the intervention, 63% of the doctors were aware of the trust guidelines, but only 17% had completed the training. after the session, 100% were aware of the guidelines, and 100% had completed the training (figure 1) . conclusions: e-learning is a useful tool, but has its limitations. by using course content, delivered with more traditional learning methods, we im-proved the number of appropriately trained clinicians, and thus the safe use of ng tubes in our unit. a systematic review of anticoagulation strategies for patients with atrial fibrillation in critical care a nelson, b johnston, a waite, i welters, g lemma university of liverpool, liverpool, united kingdom critical care 2020, 24(suppl 1):p342 there is a paucity of data assessing the impact on clinical outcomes of anticoagulation strategies for atrial fibrillation (af) in the critical care population. this review aims to assess the existing literature to evaluate the effectiveness of anticoagulation strategies used in critical care for atrial fibrillation. only 4 studies contained analysable data. anticoagulated patients had a lower mortality at 30 days and 365 days post admission to critical care, however there was an increased incidence of major bleeding events compared to the non-anticoagulated population. thromboembolic events were comparable in both cohorts. data from current literature is scarce and inferences regarding the effectiveness of anticoagulation in patients in critical care with af requires further investigation and research. every new admission to the icu prompts a handover from the referring department to the icu staff. this step in the patient pathway provides an opportunity for information to be lost and for patient care to be compromised. mortality rates in intensive care have fallen over the last twenty years, however, 20% of patients admitted to an icu will die during their admission [1] . communication errors contribute to approximately two-thirds of notable clinical incidents; over half of these are related to a handover [2] . nice have concluded that structured handovers can result in reduced mortality, reduced length of hospital stay and improvements in senior clinical staff and nurse satisfaction [3] . a checklist was created to review the information shared and to score the handover. this checklist was created with doctors and nurses and is relevant for handovers between all staff members. information was gathered prospectively by directly observing 17 handovers on the icu. there is a notable discrepancy in the quality of handovers of new patients ( figure 1 ). this is true of handovers between doctors, nurses and a combination of the two. it is also true of all staff grades. whilst a doctor may have reviewed the patient prior to their arrival, 41% (n=7) of patients weren't handed over to a doctor. the most commonly missed pieces of information were details of the patient's weight (96%, n=16), their height (100%, n=17), whether the patient has previously been admitted to an icu (78%, n=15) and whether the patient has any allergies (71%, n=12). the handover of new patients to the icu is often unstructured and important information is missed. this can be said for all staff members and grades, and for handovers from all hospital departments. post intensive care syndrome-family (pics-f) describes new or worsening psychological distress in family and caregivers after critical illness but remains poorly studied within specialist groups [1] . we aim to define the degree of pics-f within our tertiary referral cardiothoracic centre and map change over the course of 12 months. caregivers attended a 5-week multi-professional clinic alongside patients. peer support was facilitated through a café area and a caregiver group psychology session was offered with individual appointments if required. caregiver surveys were completed including: caregiver strain index; hospital anxiety and depression scale (hads); and insomnia severity index. patients also completed hads questionnaires. repeat surveys were completed at 3 and 12 months. results: over 5 cohorts, 23 caregivers attended, of which 17 were spouses (74%), 3 children (13%), and 3 others (13%), with 13 caregivers completing surveys at 12 months. patients' median apache 2 score was 17 (iqr 14-18.5) and median icu length of stay was 11 days (iqr 8-23.5). most admissions were from scheduled operations (56%). severe caregiver strain was present in 4/23 (17%) with changes to personal plans (35%) the most common sub category. hads demonstrated 13 caregivers (57%) with anxiety and 8 (35%) with depression. caregiver anxiety exceded that of patients', only reaching fig. 1 (abstract p343) . each handover was scored according to the information accurately given to icu staff similar levels at 12 months, while depression remained static ( figure 1 ). median number of nights with 'bothered' sleep was 5 (iqr 1-6.75) and 77% of caregivers expressed problems with sleep. conclusions: significant psychological morbidity in caregivers from our tertiary cardiothoracic centre is in keeping with the general icu population [2] . caregiver strain was reduced suggesting higher levels of resilience. future work should address mental wellbeing, particularly anxiety, to minimise the effects of pics-f. burnout syndrome is an illness that has increasingly affected health professionals. it is characterized by great emotional stress, physical and mental exhaustion and depersonalization of the individual. more serious cases can lead to job loss or even suicide. the described work identifies the burnout level of the multidisciplinary team through a specific questionnaireburnout syndrome is an illness that has increasingly affected health professionals. it is characterized by great emotional stress, physical and mental exhaustion and depersonalization of the individual. more serious cases can lead to job loss or even suicide. the described work identifies the burnout level of the multidisciplinary team through a specific questionnaire methods: application of a questionnaire suitable for the multidisciplinary group in november 2019. the same was answered by 85 professionals among physicians and nursing team. there was no identification of employees. after analysis of the results it is observed that 50% of the group presents initial burnout, 30% with the syndrome installed and about 5% with characteristics of greater severity. main factors found were: mental and physical exhaustion during the work day, the level of responsibility existing in the activity and the perception of disproportionate remuneration by work performed. all interviewees presented some degree of burnout or high risk to develop it. the most severe cases should be traced through occupational medicine and anti-stress measures with reorganization of work performance should be discussed in order to reduce the prevalence of this syndrome. introduction: burnout affecting the psychological and physical state of healthcare workers is recognized in the last 10 years. burnout has been shown to affect the quality of care. whilst some risk factors have been identified, there are gaps within the literature related to mental health and burnout. the aim of this study is to measure levels of burnout across 3 icu units in the metropolitan setting. to determine the level of burnout we used 2 surveys, the maslach burnout inventory human services survey (mbi-hss) and the centre for epidemiologic studies depression scale (ces-d). with the mbi-hss we analysed 3 different variables of burnout; exhaustion, cynicism and emotional exhaustion. basic demographic data and information regarding workout schedules were collected. we studied prevalence and contributing risk factors using and analysing the outcomes of the 2 self-scoring questionnaires. analysis was performed using descriptive statistical analysis. there were 90 respondents, 36% scored the threshold for depressive symptoms on the ces-d depression scale. interestingly, 40% (ci 25.4-57.7%) of those meeting the score for depressive symptoms identified as having frequent restless sleep compared with 11% (4.6-21.8 %) from those not meeting. gender did not affect depressive symptoms 35% of females and 37% of males met the threshold. with the mbi-hss for exhaustion the mean was 17.16 (sd 4.6) which is a high level of exhaustion, the second variable cynicism the mean score was 14.08 (sd 4.2), which was considered high. the final variable was emotional exhaustion the mean was 25.16 (sd 9.90), this is considered moderate levels of emotional exhaustion. fig. 1 (abstract p344) . hospital anxiety and depression scale (hads) scores for patients and caregivers at baseline, 3 months, and 12 months there was high prevalence of burnout in icu in all different categories as well as depressive symptoms. age and gender had no affect on burnout. interestingly, we identified that sleep and shift variables were linked to increased burnout. following the implementation of a fully integrated ehrs on 31 march 2019 at our university-affiliated hospital we conducted a prospective study in 2 icus by analysing pharmacists' contributions during 4 data collection periods of 5 days at 10, 15, 19 and 21 weeks post implementation. a pharmacists' contribution was defined as contacting the physician to make a recommendation in a change of therapy/ monitoring [1] . the 3 types of contribution were: a medication errorrectification of an error in the medication process; an optimizationproactive contribution that sought to enhance patient care, and a consult -reactive intervention in response to a request. a panel of experts composed of a senior pharmacist, a consultant, a nurse, and a pharmacy student assessed the impact of each contribution, scoring low impact, moderate impact or high impact. there were 160 pharmacist contributions recorded in the 4 periods. of these, 66 (41.2%) were medication errors, 93 (58.1%) were optimizations, and 1 (0.6%) was a consult ( table 1) . 37% of the contributions were assessed as having medium impact, 36% as high impact and 27% as low impact. in general, the consultant assessed fewer contributions as having high impact compared to other members of the panel, with 7 contributions assessed as high impact by the consultant versus 97 by the senior pharmacist. implementing an ehrs in combination with contributions of clinical pharmacists can prevent medication related issues. interestingly the types of incident did not change over time. introduction: most icu's are noisy and may adversely affect patients outcomes and staff performance [1] . who reports that the noise level in hospitals should not exceed 35 db at daylight and 30 db at night. the aim of this study is to evaluate the noise levels in intensive care unit, to apply awareness training to intensive care staff in terms of noise and to compare the noise levels before and after education. noise measurement areas are separated into 17 points including 12 patient bedsides, nurse desk, staff desk, wareroom, corridor and entrance of intensive care unite. measurements were performed 14 times per day. after 10 day, awareness training were given to staff in terms of harmful effects of noise. after the training, noise measurements were repeated during 10 days. after total 20 days the measurements were terminated. noise was measured with incubator analyzer (fluke model: bio-tek serial no:6050274). the mean noise values before and after the training were not statistically different from the mean average noise values (p>0.05). when the time of measurement were compared, the noise levels were higher between 10-16 hours to other measurements before and after the training statistically (p=0.001). seventeen different noise measurement areas were compared in terms of noise level, there was no statistically significant difference (p>0.05). the differences were examined at the same hours between before and after training. contrary to expectations, noise levels were found to be higher after training statistically (p<0.05). all of noise measurements were higher than the threshold values that who recommended. increased noise levels in critical care units may lead to harmful health effects for both patients and staff. our results suggest that much noise in the icu is largely attributable to environmental factors and behavior modifications due to education have not a meaningful effect. critical care medicine has focused on continuous, multidisciplinary care for patients with organ insufficiency in the face of lifethreatening illness. despite significant resource limitation low income countries carry a huge burden of critical illness. available data is insufficient to clearly show the burden and outcomes of intensive care units in these developing countries [1] . the objective of our study is to evaluate the morbidity and outcomes of patients admitted to the intensive care unit of a tertiary university hospital in hawassa, ethiopia. this was a prospective observational study. data was registered and analysed starting from patient admission to discharge during a 12 month period beginning september 2018. data regarding demographics, sources of admission, diagnosis, length-of-stay and outcomes were analysed. the total number of patients admitted to the icu was 218, with 71 patients dying over a one year period. the highest admission was from emergency medical unit, 36% and the lowest source was from pediatrics department, 8%. out of these, 69.8% were males. the mean age was 27 years (2-62). the most frequent aetiologies of morbidity in the admitted patients were traumatic brain injury (24.6%), acute respiratory distress syndrome (22.9%) and seizure disorder (8%). average median length of stay was 3.0 days (interquartile range: 1.0 -27.0). the overall mortality rate was 32.5%. the top four causes of death in the icu were respiratory illness at 24% followed by sepsis with multiorgan failure at 20%, trauma (16%) and central nervous system infection (12%). infection morbidity and mortality remains very high and needs institution of aggressive preventive strategies. the increase in frequency of trauma patients need to receive due attention. sepsis causes a high number of deaths, though overtaken by respiratory illnesses. improving the overall system of icu may achieve better outcomes in resource limited countries. introduction: icu mortality has been widely studied in the literature in relation to outcome index that primarily value organic failure [1] . however, early mortality, in the first 48 hours of admission has been little documented in the literature. the aim of this study is to analyze factors related to early mortality in icu. retrospective study at a second-level hospital. time of study was 12 months. patients who died in icu were included, patients were classified according timing of dead, including those who died within the first 48 hours of icu admission. the variables analyzed were age, sex, comorbidity, charlson index, apache ii, need for supportive treatments, more frequent admission diagnosis, origin and support treatment limitation decisions. the statistical study was carried out using the spss statistical program. 138 patients were included during the study period, 72 (52.2%) died within the first 48 hours of admission. no differences in the needs of support treatments were observed, more than 90% of patients received mechanical ventilation and vasoactive therapies. table 1 shows characteristics of patients. half of icu deaths occur within the first 48 hours of admission. severity at icu admisison was the main factor related with early mortality. severe stroke and coronary disease were the most frequent causes of early deaths in icu. in august 2018 the royal college of anaesthetists published guidelines on care of the critically ill woman in childbirth and enhanced maternal care [1] . approximately 11000 babies are born across the area covered by leicester university hospitals that includes two large maternity units and is part of the uk ecmo network. this audit sets out to assess current practice and form a basis for future planning, which will likely be representative to most major obstetric centres. a retrospective audit of all patients admitted to 'intensive care units' in leicester over a 12 month period following publication of the guidelines. the focus was on patients admitted to general adult intensive care and excludes all patients cared for in 'enhanced obstetric care' units. 9 simple standards were proposed relating to accessibility, resuscitation, follow up and multi-disciplinary learning. in total 49 women were identified with a broad range of diagnosis. the intensive care services are split across 3 hospitals and we found this led to a number of problems. the presence of trained staff to resuscitate a newborn were easily accessible, no steps to provide necessary equipment pre-emptively were present in any centre. none of our critical care units had a plan for perimortem section. on-going reviews by the obstetric and midwifery teams were very variable. contact with the infant and breastfeeding support was also poor. despite the large number of deliveries significant work needs to be done in order to come in line with the new national guidelines for critically ill woman in childbirth. clearly defined pathways around escalation of care, resuscitation of both the mother and baby, integrating care of the mother and the infant in the first few days of life, and multidisciplinary learning events are being produced de novo in response to these guidelines, some of which will be illustrated in the associated poster. interprofessional collaboration scale [3] . data were analyzed with ibm spss 25.0 results: it was found that cooperative attitudes with an average score of 49 to 75 are considered to be of average significance. interprofessional cooperation at an average score of 2,568 states that the level of cooperation is high and the quality of working life averages 125 to 150, suggesting that it is very good. as far as professional satisfaction is concerned, nurses are happy, content and satisfied with their work, despite workload and burnout conclusions: interprofessional cooperation at the icu of the general hospital of larissa is high, but satisfaction from wages, resources, working environment and conditions is low. in addition, the results showed that improvements in hospital communication between staff, has a positive impact on the quality of professional life (table 1) . contrasting with previous reports, decreased admissions per unit population in older and oldest age groups, and those with high comorbidity, suggest resource constraints may have influenced admission discussion and decision-making over the 10-year study period in wales. further investigation is warranted. icu discharge into weekends and public holidays: an observational study of mortality n mawhood, t campbell, s hollis-smith, k rooney bristol royal infirmary, general intensive care unit, bristol, united kingdom critical care 2020, 24(suppl 1):p355 introduction: up to a third of in-hospital deaths in icu patients occurs following ward stepdown [1] . discharge time seems to be associated with in-hospital prognosis, but meta-analyses have not shown a difference in weekday compared to weekend discharge [2, 3] . however, papers that examined discharge 'into' out-of-hours days, particularly on fridays, have found differences [3] . our aim was to assess whether discharge from icu 'into' out-of-hours (ooh -weekends and public holidays) is associated with in-hospital mortality or re-admission to icu, and whether these patients were seen on the wards ooh by medical staff. all adults discharged from the general icu to a ward at the bristol royal infirmary in december 2016-18 were included. in-hospital mortality rates were assessed for each day, with 'into weekdays' defined as sunday to thursday and 'into ooh' friday, saturday and the day before a public holiday. a subset of patients with data on readmission rate to icu was also examined. all available notes from patients discharged into ooh in 2018 were reviewed. the study included 1732 patients with a subset of 443 with readmission data. 117 sets of notes were reviewed from patients discharged into ooh (figure 1 ). the in-hospital mortality was significantly higher in patients discharged into ooh (5.1% vs 7.6%, p=0.012). within the subset, ooh was associated with in-hospital mortality or readmission to icu (6.8% vs 11.9%, p=0.044), though readmission rate alone was not (1.7% vs 2%, p=0.35). of patients discharged into ooh, once on a ward 64% were reviewed by a specialty doctor but 20.5% were not seen. this is the first study to examine icu discharge 'into' ooh days including public holidays. we found increased hospital mortality in ooh, similar to other studies [3] . up to a fifth of high-risk icu stepdown patients were not reviewed by a doctor on ooh days. exploring the experiences of potential donors' family members (fm) in a follow up clinic is crucial to analyze the effects of organ procurement (op) on the bereavement process, to gain insight on the reasons of family refusals (fr), and to improve family care during op. a mixed-method study involving fm at 3 and 12 months after patients' death was developed and approved by local ethics committee. fm of potential donors after brain (dbd) and cardiac death (dcd) treated in careggi teaching hospital, florence (italy) were eligible if adult and consenting. invitation letters were sent to the entitled 2 months after death and those who actively responded were involved in an encounter with a multidisciplinary group including a clinical psychologist, two nurses and two cultural anthropologists with expertise in op. organ replacement procedures such as ecmo (extracorporeal membrane oxygenation), lvad (left ventricular assist device) and dialysis are routinely used to treat multi-organ failure (mov). globally transplantation programs struggle with increasing organ shortage. patients (pts) with mov are a potential source for procurement. however, outcome data after kidney transplantation (ktx) from such donors are sparse. we retrospectively studied the cadaveric ktx at the charité berlin in 2018 and identified donors with ongoing organ replacement procedures. donor and recipient risk factors were assessed. overall patient and graft outcomes were analyzed at 12 months post-transplant. a total of 220 kidneys were transplanted. we identified 11 ktx from 7 donors with mov (6 following cardio-pulmonary resuscitation, 5 with acute renal failure -4 on dialysis) (figure 1 ). in 3 donors, a venoarterial ecmo was implanted during ecls-resuscitation. one donor needed a veno-venous ecmo due to ards, and 1 donor had a lvad implanted due to cardiac failure. the donor age was 41 ± 10.5 years (yrs). in addition, 6 donors had at least one cardiac risk factor. the kidney donor risk index averaged 0.94 (sd ± 0.14) and s-creatinine prior to ktx was 2.41 (sd ± 1.27 one way to expand the potential donor pool is donation after circulatory death (dcd), and a strategy to reduce the complications related to the ischemic time is the use of normothermic regional perfusion (nrp) with extracorporeal membranous oxygenation (ecmo) [1, 2] . we compare the use of standard nrp with an effective adsorption system inflammatory mediators (cytosorb®) in the regional normothermic reperfusion phase via regional ecmo, that involves a reduction in cellular oxidative damage, assessed as a reduction in levels of proinflammatory substances. we report a case series of 9 dcd-maastricht iiia category donors, treated in ecmo with nrp, to maintain circulation before organ retrieval, in association with cytosorb® in 5 patients. during perfusion, from starting nrp (t0), blood samples are collected 3 times, every 60 minutes (t1, t2, t3). during treatment with cytosorb®, lactate levels progressively decrease, ast and alt increase less than without cytosorb®, as sign of improvement in organs perfusion ( figure 1 ). nrp with cytosorb® might help to successfully limit irreversible organ damages and improve transplantation outcome [2] . development and implementation of uniform guidelines will be necessary to guarantee the clinical use of these donor pools. introduction: shock is a common complication of critical illness in patients in intensive care units (icus), who are undergoing major surgery. this condition is the most common cause of death in postsurgical icus. nowadays, there are different icu scoring systems for predicting the likelihood of mortality, such as apache or sofa. nevertheless, they are used rarely because they also depend on the reliability and predictions of physicians. in these sense, gene expression signatures can be used to evaluate the survival of patients with postsurgical shock. methods: mrna levels in the discovery cohort were evaluated by microarray to select the most differentially expressed genes (degs) between groups of those that survived and did not survive 30 days after their operation. selected degs were evaluated by quantitative real time polymerase chain reactions (qpcr) for the validation cohort to determine the reliability of the expression data and compare their predictive capacity to that of established risk scales. introduction: this study evaluates the prognostic ability of frailty and comorbidity scores in patients with septic shock. the 90-day mortality rate of individual medical conditions are also compared. the burden of comorbid illness and frailty is increasing in the critical care patient population [1] . outcomes from septic shock in patients with chronic ill-health is poorly understood. interstitial lung disease is a group of diseases associated with poor prognosis in the intensive care unit despite major improvement in respiratory care in the last decade. the aim of our study is to assess factors associated with hospital mortality in interstitial lung disease patients admitted in the intensive care unit and to investigate the long-term outcome of these patients. we performed a retrospective study in an intensive care unit of teaching hospital highly specialized in interstitial lung disease management between 2000 and 2014. a total of 196 interstitial lung disease patients were admitted in the intensive care unit during the study period. overall hospital mortality was 55%. two years after intensive care unit admission, 70/196 patients were still alive (36%). one hundred eight patients (55%) required invasive mechanical ventilation of whom 80% died in the hospital (figure 1 ). acute exacerbation of interstitial lung disease was associated with hospital mortality (or=5.4 [1.9-15.5] ), especially in case of acute exacerbation of idiopathic pulmonary fibrosis. multiorgan failure (invasive mechanical ventilation with vasopressor infusion and/or renal replacement therapy) was associated with very high hospital mortality (64/66; 97%). survival after intensive care unit stay of patients with interstitial lung disease is good enough for not denying them from invasive mechanical ventilation, except in case of acute exacerbation for idiopathic pulmonary fibrosis patients. if urgent lung transplantation or extracorporeal membrane oxygenation are ruled out, multiorgan failure should lead to consider withholding or withdrawal life support therapies. αgi is a malfunctioning of the gi tract in icu patients associated with prolonged mechanical ventilation, enteral feeding failure and high mortality risk. the wgap of esicm proposed a grading system for agi. four grades of severity were identified: agi grade i, a selflimiting condition; agi grade ii (gi dysfunction), interventions are required to restore gi function; agi grade iii (gi failure); agi grade iv, gi failure that is immediately life threatening. the aim was to evaluate the feasibility of using agi grades i and ii as predictors of malnutrition and 1-year mortality in critically ill patients methods: single-center retrospective cohort study in a tertiary university hospital (2015 -2017). agi grade iii and iv patients were excluded. αnthropometric data, gi symptoms (vomiting,diarrhea), feeding intolerance, gastric residual volumes and abdominal hypertension were recorded. daily prescribed caloric intake was calculated using a standard protocol and daily achievement of caloric intake was recorded. mnutric score was calculated for all patients. a score ≤5 was used to diagnose malnutrition. 200 patients (59% men, mean age 65 years) that stayed in the icu for >48 hours were included in the study. 52% were at high nutritional risk. 1-year mortality was 31%. the prevalence of agi ii was 14%. age, gender, bmi, mortality and energy intake did not differ significantly between patients with agi ii and those with agi i (table 1) . logistic the study aimed to assess the effects of icu admission on frailty and activities of daily living in the ≥80's population at 6-months. a prospective observational study with data used as a subset of the vip-2 trial [1] . research ethics committee approval from the mater misercordiae university hospital (mmuh). inclusion criteria -≥ 80 years of age and acute admission to icu from may to july 2018. data collected on 20 consecutive patients. frailty and activities of daily living (adl) were assessed using the clinical frailty score (cfs) and the katz index of independence in activities of daily living (katz). results: csf pre-admission frailty was present in 60% of patients, increasing to 93% at 6 months ( figure 1 ). 74% of survivors at 6-months had a cfs score increase by ≥ 1 point. pre-frail and frail cfs patients suffered an average 2-point deterioration in their instrumental activities of daily living (iadl). 60% of katz patients were fully functional preadmission, deteriorating to 13% at 6 months. 74% of patients declined by 1 adl at 6 months. 60% of the deceased were deemed fully functional initially. we demonstrate an association between an icu admission event and enduring functional decline at 6 months. icu admission resulted in patients acquiring on average 1.5 new iadl limitations despite their initial cfs. this is echoed in a study by iwasyna et al. who also showed similar deteriorations in iadl and cognitive impairment [2] . katz benefits may be best used in describing functional decline. 74% of patients developed at least one new limitation. however, the cfs takes into account iadl's and thus may be more sensitive in predicting the functional outcomes of an icu event at 6 months. frailty: an independent factor in predicting length of stay for critically ill t chandler, r sarkar, a bowman, p hayden medway maritime hospital, critical care, gillingham, united kingdom critical care 2020, 24(suppl 1):p366 frailty has attracted attention in the healthcare community in recent years, as it is associated with worse outcomes and increased healthcare costs [1] . our objective was to study the impact of frailty as recorded by clinical frailty scale(cfs) to prospectively evaluate the effect of frailty on hospital length of stay (los). a retrospective analysis of consecutively admitted critical care (cc) patients' data (jan'19-oct'19) was performed. electronic health records were used to collect demographics, cfs and clinical outcomes. statistical analysis was performed using stata. students t-test, simple and multiple (adjusted for age, disease severity/icnarc score) linear regression were used for comparison between groups and to see group effect. we excluded extreme outliers (los>50 days; n=13). frailty was defined as cfs>4. out of the 848 patients (male 58%), 554(66%) were emergency admissions, the rest elective (table 1) . 288(40%) were non-frail. the mean los were 15 days (d) ±12 and 10d±9 (p<0.0001) in the frail and non-frail patients respectively. for emergency patients, los were 16d(±12) and 10d(±10) for the groups, (p<0.0001). for elective patients; los were 12d(±10) and los 8d(±7), (p=0.01) for frail and nonfrail respectively. after adjusting, los was significantly higher in frail patients by 5 days (95%ci 3,7; p<0.0001), by 4 days (95%ci 1,6; p= 0.002) and by 5 days (95%ci 4,8; p<0.0001) for total cohort, elective and emergency admissions respectively. the los was 6 days higher in frail than non-frail (p<0.001) for cc survivors. frailty was associated with significantly increased los in this cohort, independent of age and illness severity. hospital capacity planning should take this into consideration when modelling bed allocation fig. 1 (abstract p365) . clinical frailty score 6-month trend robust clinical governance requires analysis of patient outcomes during an icu admission [1] . on one adult icu weekly mortality meetings are used for this purpose and aid multidisciplinary reflections on individual patient deaths. however, such reviews run the risk of being subjective and fail to acknowledge themes which may relate to preceding or subsequent deaths. this paper describes a new mortality review process in which: a) reviews are structured using the structured judgement review (sjr) framework [2] ; and b) themes are generated over an extended period of time to create longitudinal learning from death. the sjr framework has been developed by nhs improvement for the new medical examiner role, looking at inpatient deaths. we adapted this to better suit the icu creating a novel review structure. this involves explicit judgement comments being recorded, and the use of a scoring system to analyse the quality of care during the patient's stay with a focus on elements of care delivered on the icu. tabulation of this information allows analysis over time, identifying trends across all patients, and in specific subgroups. this framework has been rolled out at the st george's cardiothoracic icu weekly mortality meetings. themes that have emerged include parent team ownership, delayed palliative care referrals and inadequate documentation of mental capacity. this will continue as part of a three-month trial and following review of this trial may be extended to other critical care units in the trust. this system allows greater insight into patient deaths in a longitudinal fashion and facilitates local identification of problems at an early stage in a way that is not possible within the traditional mortality review format. the nature of the process means that key areas for change can be identified as a routine part of the clinical week. [1] . in this study, we evaluated three distinct machine-learning methods for predicting possible patient deterioration after surgery. the data was collected retrospectively from the catharina hospital in eindhoven. this dataset contained all the surgeries conducted in the hospital from 2013 up to 2017. the variables in this dataset were tested on their ability to differentiate between patients with a normal recovery versus patients with an unplanned icu admission after being admitted to the ward. the dataset contained 44 variables related to either the preoperative screening, surgery or recovery room. all variables were tested for statistical significance using a univariate logistic regression (lr), from which a subset of 34 statistically significant (p<0.05) variables was created. these variables were used to train three different types of models, namely, the lr, support vector machine (svm) and bayesian network (bn). the network structure of the bn was designed using expert knowledge and the probabilities were inferred using the data. the three models were validated using five-fold cross-validation, resulting in the following areas under the receiver operating characteristic curve: 0.82(0.79-0.86) for lr, 0.82(0.78-0.88) for svm and 0.65(0.63-0.68) for bn (fig. 1) . the results indicate that machine learning is a promising tool for early prediction of patient deterioration. the bn was included because it permits incorporating clinical domain knowledge into the learning process. however, its performance resulted inferior to the lr and svm. in future work, we will investigate alternative domainaware methods, and compare the performance with that of the clinical experts. intensive care unit (icu) admission decisions of patients with a malignancy can be difficult as clinicians have concerns about unfavourable outcomes, such as mortality [1] . a diagnosis of a malignancy is associated with an almost 6-fold increased likelihood of refusal of icu admission [1] . recent large long-term mortality studies of patients with a malignancy admitted to the icu are scarce. therefore, our aim was to compare mortality of patients with either a hematological or a solid malignancy to the general icu population, all with an unplanned icu admission. all adult patients registered in a national intensive care evaluation registry with an unplanned icu admission from 2008 to 2017 were included. subsequently, we divided these patients into 3 cohorts: cohort 1 (all patients with a hematological malignancy), cohort 2 (all patients with a solid malignancy), and cohort 3 (a general icu population without malignancy). as primary outcome, we used 1-year mortality, and as secondary outcome, icu and hospital mortality. we included 10,401 (2.2 %) patients in cohort 1, 35,920 (7.6%) patients in cohort 2 and 423,984 (90.2%) in cohort 3 ( table 1 ). the 1year mortality of patients of cohort 1, 2, and 3 was 60.1%, 46.2% and 28.3%, respectively (p<0.001). age, comorbidities, organ failure, and type of admission (i.e. surgical or medical) were positively associated with 1-year mortality in all cohorts (p <0.05). one-year mortality is higher in both patients with a hematological malignancy and patients with a solid malignancy compared to the general icu population. in addition, several factors were positively associated with 1-year mortality, i.e., age, comorbidities, medical icu admission, and organ failure. future research should focus on predictive modelling in order to identify patients with a malignancy that may benefit from icu admission. introduction: drug abuse is associated with immunosuppression in multiple mechanisms. despite that, the only study retrospectively reviewing drug abusers in the icu demonstrated less infections and better outcomes. we compared matched patient populations in order to fully understand whether drug abuse is a risk factor for infection and a predictor of poorer prognosis as is perceived by most physicians. we hypothesized that the drug abusers admitted to the icu will fare as good as or better than non-abuser icu patient populations. methods: this is a prospective study done between the years 2010-2012 on the entire patient population of the detroit medical center. after the drug abuse population was identified, controls were matched according to age and admission icu units. patients charts were reviewed and data regarding baseline demographics, infectious complication and outcome was extracted. data was retrospectively collected for 323 drug abusers and 305 matched controls. comorbidities and hospital admission diagnosis were significantly different between the two groups. disease severity scores were significantly higher in the drug abuser's patient group (dapg) on admission and during the icu stay. dapg had significantly more organ failure: more need for ventilation (30.5% vs 46.4% in the dapg (p<0.001)), more ards (1% vs 3.7%, p=0.03), more renal failure (33% vs 45.5%, p=0.002) and more need for renal replacement therapy (6.6% vs 11.2%, p<0.05) .they had longer hospital length of stay (los). there was no difference in icu or hospital mortality. multivariable modeling did not find drug abuse to be an independent risk factor for hospital mortality, icu mortality (hosp: or = 1.37, p = 0.3397; icu: or=1.43, pp = 0.07), but was a risk factor for a longer hospital los (me=1.46, p < 0.0001). drug abuse is not an independent risk factor for mortality or icu los. drug abusers should be evaluated like other patients based on baseline comorbidities and disease severity. this is a small audit which although it did not include general icu still reflects the need for encouraging clinicians and patients to speak freely regarding escalation plans. medical decsions is clinician led however this audit was carried by nursing staff as we have a duty to be advocate for our patients involvement in medical care [2] . a retrospective analysis of independent risk factors of late death in septic shock survivors c sivakorn 1 , c permpikul 2 , s tongyoo 2 (fig. 1) . the pap and katz scales seem to be adequate for predicting mortality of critically ill patients admitted to a medical icu. this finding may help in the elaboration of future icu mortality scoring systems, as well as in more rational use of resources. however, further multicenter studies are needed to better elucidate these results. adherence this last group was chosen because of its experience and specific training in the field of bioethics as a control group or reference. a total of 444 respondents participated in the study. 22.2% were emergency physicians, 14.8% intensivists, 11.2% emergency nursing, 6.2% icu nursing, 24.9% resident doctors, 13.8% medical students and 6.9% other professions. we observed variability in the responses observed not only between different groups of professionals but even within the same group reflecting the difficulty in decision making. variability was observed regarding decisions in end of life ethics conflicts. a high degree of similarity with the group of master in bioethics was observed in the responses issued by medicine students. the barriers and facilitators to framing goals of patient care (gopc) and factors motivating decision making is relatively unexplored [1, 2, 3] . a three part survey of physicians at an australian hospital in a culturally and linguistically diverse suburb ( table 1) . identification of levels of confidence and barriers and facilitators to gopc discussion and decision making was the main outcome measure. factors influencing decision-making was analysed through scenarios. results: 22 out of 96 eligible participants responded; 12 female, 10 male, clinical experience 4-31 years. level of confidence was ranked between "somewhat confident and very confident." all but one respondent had six months of icu experience. no differences in the level of confidence among physician groups. 14 barriers and 8 facilitators were identified; poor prognosis and patient or family request were most common facilitators; conflict between treating teams and the patient/surrogate and language barriers were most common barriers. factors driving gopc decision-making included clinical, value judgement, communication, prognostication, justice and avoidance. numerous barriers and facilitators were identified. factors driving decision making did not just consider clinical factors; conflict and we aimed to investigate physician-related factors contributing to individual variability in end-of-life (eol) decision-making in the intensive care unit (icu). qualitative study with semi-structured interviews with 19 specialists in critical care, (experience 2-32 years) from 5 swedish icus. data was analyzed in accordance to principles of thematic analyses. most of the respondents felt that the intensivist's personality played a major role in eol decisions (table 1) . individual variability was considered inevitable. views on acceptable outcome: respondents experienced that the possible outcome for patients was interpreted very differently and subjectively among colleagues, and what seemed an acceptable patient-outcome for one doctor, was not acceptable for another. values: most of the respondents were well aware that they might be affected by their own values and attitudes in the decision-making process. interestingly, several respondents mentioned that they thought that patients that were marginalized by society, especially drug-abusers could be at risk for receiving decisions to limit life sustaining treatments (lst) more often than others. none of the respondents thought that their own religious beliefs played any part in decision making. fear of criticism: among the less experienced respondents there was a clear sense of fear of making a questionable assessment of the patient's medical prognosis. there was a fear for criticism from colleagues that were not directly involved in the decision-making, and may have made another decision. this created a wish among younger respondents to defer or avoid participating in decision-making. physician-related, individual variability in eol decisions primarily consisted of differing views on acceptable outcome, values and fear of criticism. can (figure 1 ). within each quartile of sofa score, mortality was highest in patients with pneumonia and peritonitis and lowest in patients with cellulitis (see figure 1 ). the sepsis-3 consensus definition identified organ dysfunction as the hallmark feature of sepsis [1] . in developing sepsis-3, the sequential organ failure assessment (sofa) score was chosen for its prognostic value and relative ease of implementation clinically [2] . we propose an update based on epidemiologic data from two intensive care databases that more effectively captures organ dysfunction in the context of sepsis-3. using the mimic-iii (exploration) and e-icu (validation) databases, we extracted patients with suspicion of infection to form the study cohort. the predictive power of each sofa component was assessed using the area under the curve (auc) for in-hospital mortality. a logistic model with the lasso penalty was used to find an alternative statistically optimal score. results: by utilising alternate markers of organ dysfunction (e.g. lactate, ph, urea nitrogen) we demonstrated a significant improvement in auc for several versions of the new score, sofa2.0 ( figure 1 ). the sofa score can be updated to reflect current advances in clinical practice. using epidemiologic data, we have shown that substitution of existing components with more powerful measures of organ dysfunction may provide an improved score with greater predictive power. moreover, sofa 2.0 exhibits equivalent ease of implementation, but better reflects organ dysfunction in the context of sepsis-3. introduction: risk of acute organ failure (aof) in cancer patients(pts) on systemic cancer treatment isunknown. however, 5% of non-hematologic and 15% of hematologic cancer pts will need admission to intensive care unit (icu). ipop-sci-2017/01 is a prospective cohort study designed to ascertain the cumulative incidence of aof in adult cancer pts. single centre prospective cohort study with consecutive sampling of adult cancer pts admitted for unscheduled inpatient care while on, or up to 8 weeks after, systemic cancer treatment. primary endpoint was aof as defined by quick sofa. six months accrual expected an accrual of 400 pts to infera population risk aof with a standard error of 1%. between 08/2018 and 02/2019 10392 pts were on systemic anticancer treatment, 358 had unscheduled inpatient care and were eligible for inclusion and 285 were included. median age was 64 years, 51% were male, 52% had adjusted charlson comorbidity index (cci) &gt; 3 and hematologic cancers accounted for 22% of pts. the cumulative risk of aof on hospital admission was 35% (95%ci: 31-39); and of aof during hospital stay was 40% (95%ci: 35-44). aof was associated with older age, cci &gt; 3,hematologic malignancy, shorter median time from diagnosis and &gt; 1 prior line of therapy. on admission, 62% of pts were considered not eligible for artificial organ replacement therapy (noaort) and 34% of pts who developed aof while inhospital were judged noaort. overall, 17 (15%) of aof pts wereadmitted to icu, 31.5% for aort. median follow up 9.5 months (min 6; max 12). inpatient mortalitywas 18%, with icu mortality rate of 59%, with median cohort survival 4.5 months (95%ci: 3.5-5.4). on multivariate analysis, aof was an independent poor prognostic factor (hr 1.6; 95%ci 1.2-2.1). risk of aof in cancer pts admitted for unscheduled inpatient care while on systemictreatment is 35%, and risk of icu is 15%. aof in cancer pts was an independent poor prognostic factor. a severity-of-illness score in patients with tuberculosis requiring intensive care u lalla, e irusen, b allwood, j taljaard, c koegelenberg tygerberg academic hospital, internal medicine, division of pulmonology and icu, cape town, south africa critical care 2020, 24(suppl 1):p383 we previously retrospectively validated a 6-point severity-of-illness score aimed at identifying patients at risk of dying of tuberculosis (tb) in the intensive care unit (icu). parameters included septic shock, human immunodeficiency virus with cd 4 <200/mm 3 , renal dysfunction, ratio of partial pressure of arterial oxygen to fraction of inspired oxygen (pao 2 :fio 2 ) <200mmhg, diffuse parenchymal infiltrates and no tb treatment on admission. the aim of this study was to validate and refine the severity-of-illness score in patients with tuberculosis requiring intensive care. we performed a prospective observational study with a planned post-hoc retrospective analysis, enrolling all adult patients with confirmed tb admitted to the medical intensive care unit from 1 february 2015 to 31 july 2018. descriptive statistics and chi-square or fisher's exact tests were performed on dichotomous categorical variables, and t-tests on continuous data. patients were categorized as hospital survivors or non-survivors. the 6-point score and the refined 4-point score were calculated from data obtained on icu admission. results: forty-one of 78 patients (52.6%) died. the 6-point scores of nonsurvivors were higher (3.5+/-1.3 vs 2.7+/-1.2; p=0.01). a score ≥3 vs. <3 was associated with increased mortality (64.0% vs. 32.1%; or 3.75; 95%ci, 1.25-10.01; p=0.01)( table 1) . post-hoc, a pao 2 :fio 2 < 200mmhg and no tb treatment on admission failed to predict mortality whereas any immunosuppression did. a revised 4-point score (septic shock, any immunosuppression, acute kidney injury and lack of lobar consolidation) demonstrated higher scores in non-survivors (2.8+/-1.1 vs. 1.6+/-1.1; p<0.001). a score ≥3 vs. ≤2 was associated with a higher mortality (78.4% vs. 29.3%; or 8.76; 95%ci, 3.12-24.59; p<0.001) ( table 1) . the 6-point severity-of-illness score identified patients at higher risk of death. we were able to derive and retrospectively validate a simplified 4-point score with a superior predictive power. chronic critical illness remains a scientific challenge, from its conceptualization to its impact on patient prognosis [1] . we evaluated the long-term evolution of icu survivors by identifying the real burden of prolonged critical illness on survival, quality of life and hospital readmissions. we conducted a prospective cohort in 16 brazilian hospitals including 1616 icu survivors with an icu stay > 72h. we compared the patients diagnosed with chronic critical illness with the other patients. telephone follow-up at 3 and 6 months. quality of life was measured by the sf-12 questionnaire. it was observed that 38% of patients had some definition of chronic critical illness. chronic critically ill patients had higher mortality at 6 months (p=0.012). this difference is mainly due to higher intrahospital mortality (p=0.0001). mortality after hospital discharge was similar between groups. there was no difference in hospital readmission rate at 6 months. various scores are developed to predict pulmonary complications such as ariscat for patients at-risk of postoperative pulmonary complication [1] and lips for patients at-risk of lung injury [2] . the aim of this study was to compare these scores with ours for predicting pulmonary complications in mechanically ventilated patients in sicu. this prospective observational study was conducted in sicu at a university hospital. adult patients admitted to sicu and required mechanical ventilation >24 hours were included. primary endpoint was the composite of pulmonary complications including pneumonia, ards, atelectasis, reintubation, and tracheostomy. multivariate analysis was performed to identify risk factors of pulmonary complications and the predictive score was developed. the roc analysis was performed to compare power of ariscat, lips and our newly developed score for predicting pulmonary complications. outcomes in intensive care units have been reported to be better in higher-volume units [1, 2] . we compared outcomes for high-risk patients between low and higher volume units. audit data from irish icus is analysed and reported by the intensive care national audit & research centre (icnarc) in london. icnarc report risk-adjusted mortality rates in all patients and in low-risk patients(predicted mortality rate <20%) for each unit, using the icnarch-2015 model to predict the risk of death. we used this data to calculate the proportion of high-risk patients(predicted mortality >20%) in each unit, the mortality rate for high-risk patients, the riskadjusted mortality rate and we compared the overall risk-adjusted mortality between low and high volume units. the median number of annual new-patient admissions among 18 participating units was 390; units below this were defined as lowvolume and those above as high-volume units. the proportion of all admissions to each unit who were high-risk ranged from 8% to 54%(mean 34%). unit mortality rates for high-risk patients ranged from 33% to 69%. the ratio of observed to expected mortality(standardized mortality ratio -smr) for high risk admissions in each unit ranged from 0.87 to 1.34(mean 1.07). in fig. 1 introduction: adl weakening is often seen after intensive care and called postintensive-care syndrome (pics). this is also seen in even outside icu and proposed to be called post-acute-care syndrome (pacs), especially in elderly patients. in patients with infection, sofa score is famous for predicting in-hospital mortality, but there are no tools for predicting adl weakening during admission. to search for risk factors for adl weakening during admission other than the age, we conducted a retrospective observational study. the subjects were surviving patients with infection, aged from 16 to 89 who were admitted to our department from april 1, 2018 to may 31, 2019. information of basic characteristics, laboratory data on admission and adjunctive therapies were extracted from our database. we use barthel index (bi) as adl evaluation, and the bi at discharge were evaluated by nurses. we stratified patients by bi at discharge of over 60 or not, and investigated factors that predicted it. we compared each factor between 2 groups, and perform a logistic regression analysis with those that had a significant effect clinically or statistically. despite improved outcomes of intensive care unit (icu) patients, sleep deprivation remains a major concern after icu discharge. multifaceted causes make it difficult to treat and understand [1] . not many studies have explored sleep deprivation beyond icu. this is evidenced by findings from a recent systematic review [2] which included 8 studies with only one study [3] reporting sleep deprivation beyond icu. the aim of this paper is to present findings of sleep deprivation beyond icu from a larger study that examined the experience of critical illness in icu and beyond in the context of daily sedation interruption. hermeneutic phenomenology was used to conduct the study. 12 participants aged 18 years and above who fulfilled the enrolment criteria were enrolled into the study. the cohort comprised 7 male and 5 female participants. in-depth face to face interviews at two weeks after discharge were conducted and repeated at six to eleven months. interviews were audio taped, transcribed and thematically analysed. significant statements were highlighted and categorized for emergent themes. six participants continued to experience sleep deprivation up to eleven months after icu. two cited dreams about icu, three could not explain why they continued to fail to sleep and one stated that he continued hearing icu alarms in the silence of the night. sleep deprivation continues beyond icu due to nightmares, delusional memories and unexplained reasons. further research is needed to establish causes of sleep deprivation and explore ways to promote sleep in critical illness survivors after icu discharge. frailty is being increasingly seen as an independent syndrome. frail patients now account for an increasing proportion of hospital and critical care admissions [1] . we aimed to compare frailty and mortality in our intensive care unit. clinical frailty score (cfs) was incorporated within the electronic health record (ehr) 2019. we performed this retrospective analysis on the data collected between jan'19 and oct'19. the predictor and outcome for this study were frailty and hospital mortality respectively. all demographic data, acute physiology score, critical care and hospital outcome data were automatically collected in the ehr and recorded. we used a cut off of cfs>3 and above to define non-frail and frail respectively. chi-squared test, simple and multiple logistic regression were used. adjustment was done for icnarc score and age. total number of patients was 848, of which 140 (16.5%) died in hospital. within the patients<65 years (n=392), 79 (20%) were recorded as frail or vulnerable. the number of elective and emergency admission were 292(34%) and 556(66%) respectively. in the frail and nonfrail, mortality rates were 30% and 9.5% (p<0.001) respectively, with odds ratio of 4.14 (95% ci 2.8, 6; p<0.001) ( age is a well-known risk factor for critical care (cc) outcome and is incorporated into many prognostic tools; however, this has been criticized for assumption of normal physiology for young at baseline. in recent years, frailty in cc prognostication has been of interest, with meta-analysis correlating worsening outcomes with increasing frailty [1] . in this study, we compared the effect of frailty versus age for determining hospital survival for critically ill patients. we conducted a prospective cohort in 16 brazilian hospitals including 1616 survivors of an icu stay > 72h. we compared chronic critically ill patients (icu stay> 10 days) and the other patients. we performed psychological and functional presential assessment in patients within 48 hours of icu discharge and by telephone at 3 and 6 months. the prevalence of chronic critically ill patients was 26%. regarding outcomes, chronic critically ill patients had a higher incidence of depressive symptoms than other patients in the immediate post-icu discharge (p = 0.004), as well as a higher incidence of muscle weakness (p <0.001). however, in subsequent evaluations, we found no difference between groups regarding psychological symptoms -depression, anxiety and post-traumatic stress. higher functional dependence was observed in critically ill patients, but without difference in the quality of life score, both in the physical (p = 0.87) and mental (p = 0.84) domains. chronic critically ill patients, when compared to patients with stay> 72h, have a higher incidence of depressive symptoms at icu discharge. this difference disappears in the follow up. chronic critically ill patients present higher levels of functional dependence but without repercussions on quality of life scores. introduction: activation of the inflammatory response after cardiac arrest (ca) is a welldocumented phenomenon that may lead to multi-organ failure and death. we hypothesized that white blood cell count (wbc), one marker of inflammation, is associated with one-year mortality in icu treated ca patients. we used a nationwide registry with data from five academic icus to identify adult ca patients treated between january 1 st 2003 and december 31 st 2013. we evaluated the association between the most abnormal wbc within 24 hours of hospital admission and one-year mortality. we accounted for baseline risk of death using multivariable logistic regression (adjusted for age, gender and 24h sequential organ failure assessment [sofa] score). a total of 5,543 patients were included in the analysis. of those patients 2,387 (43%) were alive one year after ca. we plotted wbc against baseline risk of death and through graphic examination of a locally weighted scatterplot smoothing (lowess) curve found the lowest risk of death to be associated with a wbc of 12 (e 9 /l) ( figure 1 mrps were identified by a specialist icu pharmacist during this programme and classified by their significance on a scale of one to four. logistic regression was used to determine if demographic factors were associated with the occurrence of a clinically significant mrp -a significance score of two or above (figure 1) . the adjusted model included age, icu los, hospital los, apache ii, number of days of renal replacement therapy, number of days of ventilation, the number of medications prescribed at icu discharge, and the who analgesia classification at ins:pire. there were increased odds of having a clinically significant mrp for hospital los (or results: 62·8% (n=115) of patients required at least one pharmacy intervention. the median number of interventions required per patient was one (iqr 0-2); the maximum number was six. 198 mrps were recorded in this cohort. the most common intervention was clarifying duration of treatment (n=44), followed by education (n=33), and correcting drug omissions (n=27). the bnf drug class most frequently associated with mrps was neurological (n=65), which comprises analgesics (n=45) and psychiatric medications (n=20) ( figure 1 ). this was followed by cardiovascular medications (n=40), gastrointestinal medications (n=34), nutritional medications (n=25), and others (n=34). many icu survivors experience mrps. the most common class of mrp was neurological, reflecting the high incidence of chronic pain and psychiatric illness in this population following discussion with icu staff, ward staff and fy1 doctors, a formal standardized handover system was introduced. this involved a verbal handover to the appropriate fy1 by an icu doctor and the patient drug chart to be rewritten in icu at the time of handover. the next change was to display posters on the wards to alert staff that the medical team are to be contacted when a patient comes to the ward from icu and to ensure the drug chart is completed. the baseline data showed a median time delay of 4 hours, with one patient waiting 14 hours for a drug chart. following the interventions the median time delay has decreased to 0 hours within 4 months as demonstrated in figure 1 . the changes have received positive feedback from icu staff, ward staff and fy1 doctors. the aim of reducing the time delay by 50% has been achieved with the median time delay now 0 hours. this has improved patient safety by significantly reduced delays in medications and through the introduction of a standardized handover. this has also provided an opportunity for junior doctors on the wards to seek clarification regarding medications and the clinical management plan for the patient. this has established a communication channel between icu and the wards making patient care safer and more effective. telemonitoring outside the icu is scarce. but with innovative wearables measuring respiratory and heart rate wirelessly, culture on intrahospital telemonitoring should definitely change. however, culture has been known to be one of the most crucial success factors in innovation, especially in health care. human design thinking is a promising tool in health care innovation but rarely used in a multidisciplinary team to initiate an innovation culture and stimulate sustainable collaboration. the aim of this study was to initiate a pilot project with a multidisciplinary team to start using wearables for early warning score (ews) on a clinical ward. human design thinking was used to write a value proposition on wearables in clinically admitted neutropenic hematologic patients in an academic center. a multidisciplinary team was performed to cover all disciplines involved in the technical, clinical and administrative parts of the project. a vendor was chosen based on its product specifications in relation to the present hospital monitoring infrastructure. in design thinking sessions, critical appraisal of multiple telemonitoring factors was performed by sub teams and a canvas projectplan was constructed. the project team was formed of registered nurses, physicians, itspecialists, electronic health record consultants; a critical care physician was appointed as project leader. the main critical factors were: unseamlessly transmitting of both heart and respiratory rates including appropriate movements filtering to the nurse's smartphones direct uploading into electronic health record with automated ews calculation nurse driven protocol on ews follow up. philips healthcare with their intellivue guardian wearable biosensor was the chosen vendor ( figure 1 ). design thinking in a multidisciplinary health care team could positively influence the innovation culture. scientific evaluation of this wearable will focus on both nurse's acceptance and data storage and is expected in the summer of 2020. severity, readmission and lengh of stay were lower in patients receiving discharges directly to home. it seems like a safe way to discharge low-risk short stay patients. it seems to save resources and reduce costs, as well as the need for hospital beds. however, futher estudies are needed to actualy evaluate this safety. forty-four cultures were analyzed with eplex ( figure 1 ). complete agreement with conventional diagnostics was observed in 38/44 cases. no false-positive results were observed, yielding a sensitivity and specificity of 90% and 100% respectively for target pathogens. time to result was, on average, 10.4 h faster with eplex compared to conventional diagnostics. antimicrobial therapy could have been optimized in 5 patients based on the eplex result, but treatment was only changed in one case (e.coli ctx-m+) receiving meropenem 8.5 h before the antibiogram was available. the eplex blood culture panels provide high accuracy and significantly faster results. the current implementation offers substantial potential value at a minimal cost, and is a feasible approach to 24-h/ 7 days blood culture diagnostics in many hospital settings. however, efforts to increase adherence are needed. the rapid increase of extended spectrum β-lactamases (esbl)-producing pathogens worldwide makes it difficult to choose appropriate antibiotics in patients with gram-negative bacterial infection. cica-beta reagent (kanto chemical, tokyo, japan) is a chromogenic test to detect beta-lactamases such as esbl from bacterial colonies. the purpose of the study was to reveal whether cica-beta reagent could detect esbl-producing pathogens directly from urine rather than bacterial colonies to make a rapid bedside diagnosis of the antibiotic susceptibility of gramnegative pathogens. we conducted a prospective observational study from july 2019 to october 2019. patients were eligible if they were performed urinary culture tests and gram negative pathogens were detected at least 1+ from their urine samples. the urine sample was centrifugated at 200 x g for 5min. the supernatant of sample was re-centrifugated at 1500 x g for 5min and the pellet was mixed with cica-beta reagent. the test was considered positive when the enzymatic reaction turned from yellow to red or orange. (fig.1) . the bundle approach could be an effective strategy to prevent hospital-acquisition of drug-resistant pathogens in icus. fig. 1 in the aspect-np trial, c/t was noninferior to mem for the treatment of habp/vabp. we evaluated outcomes from that study in the subgroup of pts failing current antibacterial therapy for habp/vabp at enrollment. methods: aspect-np was a randomized, controlled, double-blind, phase 3 trial in which mechanically ventilated pts with habp/vabp received 3 g c/t or 1 g mem every 8 h for 8-14 days. pts with >24 h of active gram-negative antibacterial therapy within 72 h prior to first dose of study therapy were excluded, except those pts failing current treatment (i.e. signs/symptoms of the current habp/vabp were persisting/worsening despite ≥48h of antibiotic treatment). primary and key secondary endpoints, respectively, were 28-day all-cause mortality (acm) and clinical response at test of cure (toc; 7-14 days after end of therapy) in the intent to treat (itt) population. pts failing current antibacterial therapy for habp/vabp were prospectively categorized as a clinically relevant subgroup. at baseline, failing current therapy for habp/vabp was reported in 53/362 (15%) c/t and 40/364 (11%) mem itt pts, mostly piperacillin/ tazobactam (34%), 3rd/4th-generation cephalosporins (31%), fluoroquinolones (29%), and aminoglycosides (12%). baseline demographic and clinical characteristics in this subgroup, including prior therapy regimen, were generally similar between treatment arms. there were greater proportions of patients with esbl+ enterobacterales (30%) and pseudomonas aeruginosa (25%) in the c/t arm than the mem arm (20% and 13%, respectively). lower 28-day acm was seen with c/t than mem, as evidenced by 95% confidence intervals for treatment differences that excluded zero ( figure 1 ); statistical significance cannot be assumed because subgroup analyses in this study were not corrected for multiplicity. conclusions: c/t was an effective treatment for habp/vabp pts who had failed initial therapy. catheter-related blood stream infection (crbsi) is common serious infections and associated with increased mortality in intensive care units (icu). one of the most important strategy to prevent crbsi is to minimize the duration of central venous catheterization. we built a medical team consisting of doctors, nurses and pharmacists in icu to discuss whether patients needed central venous catheter (cvc) in terms of monitoring hemodynamics and administering drugs, and recommend catheter removal to attending physicians every day in april 2019. the purpose of this study is to evaluate whether our team-based approach could shorten the total duration of catheterization and reduce crbsi. this was a retrospective historical control study conducted from april 2018 to october 2019 in the icu of a tertiary care hospital in japan. every patient admitted to the icu during the study period was eligible if they were inserted cvc. patients were divided into 2 groups: conventional (from april 2018 to march 2019) or intervention (from april 2019 to october 2019). we set the primary endpoint as onset of crbsi. the secondary endpoints included the duration of central venous catheterization, the length of icu stay and hospital mortality. crbsi was defined as bloodstream infection in patients with cvc, not related to another site. we included 428 patients: 259 in the conventional group and 169 in the intervention group. the reduced, though nonsignificant, tendency of crbsi was observed in the intervention group [hazard ratio, 0.341(95% confidence interval, 0.074-1.557; p = 0.213)]. the intervention group was significantly associated with reduced duration of central venous catheterization (5 days vs 7 days; p < 0.01). no difference was observed in the length of icu stay and in-hospital mortality between groups. the team-based approach to assess cvc necessity could shorten the duration of central venous catheterization and might reduce crbsi. introduction: empiric antibiotic therapy decisions are based upon a combined prediction of infecting pathogen and local antibiotic susceptibility, adapted to patients' characteristics. the objective of this study was to describe the pathogen predominance and to evaluate the probability of covering the most common gram-negative pathogens in icu patients with respiratory infections. methods: data were collected from multiple us and european hospitals as part of the smart surveillance program (2018). mic (mg/l) testing was performed by broth microdilution, with susceptibility defined as follows for p. aeruginosa & enterobacterales: ceftolozane/tazobactam results: 78 hospitals from 24 countries provided 3384 gram-negative respiratory isolates from patients located in an icu in the us (22%), eastern europe (40%) and western europe (38%) in 2018. the 4 most common pathogens isolated were p. aeruginosa (26%), k. pneumoniae (15%), e. coli (13%), and a. baumannii (10%). among enterobacterales, 30% (588/1955) were esbl positive. figure 1 provides the probability of covering the most common respiratory gram-negative pathogens from icu patients. co-resistance between commonly prescribed first line β-lactam antibiotics is common: when nonsusceptibility (ns) of one agent was present, susceptibility to other βlactams was generally <35%. ceftolozane/tazobactam provided the most reliable in vitro activity in both empiric and adjustment prescribing scenarios compared to other β-lactam antibiotics. ceftolozane/tazobactam ensured a wide coverage of the most common gram-negative respiratory pathogens demonstrating high susceptibility levels and provided the most reliable in vitro activity in both empiric and adjustment antibiotic prescribing scenarios. further studies are needed to define the clinical benefits that may translate from these findings. evaluation of compliance of icu staff for vap prevention strategies on the outcome of patients a kaur fortis hospital, critical care, mohali, india critical care 2020, 24(suppl 1):p426 ventilator-associated pneumonia is the most common nosocomial infection diagnosed in adult critical care units. it is associated with prolonged duration of mechanical ventilation, increased icu stay and increased mortality. it continues to be a major challenge to the critical care physicians despite advances in diagnostic and treatment modalities. the primary objective of the study was to determine the compliance of icu staff towards vap prevention bundle and secondary objective was to determine the incidence, risk factors and outcome of vap patients. single center, prospective, observational study carried out from february 2017 to july 2018. patients mechanically ventilated for more than 48 hours and satisfying the inclusion and exclusion criteria were enrolled in the study. vap was diagnosed using the cdc criteria and clinical pulmonary infection score. vap preventive strategies were employed and compliance of icu staff was assessed. a total of 1617 patients were admitted to icu over the set time period and out of them 483 patients were ventilated for more than 48 hours. among them only 166 patients fulfilled the inclusion and exclusion criteria and were enrolled in the present study. excellent compliance was observed in head end elevation, sedation vacation, stress ulcer prophylaxis, and heat moist exchanger filter use, good compliance in oral care and hand hygiene and moderate to poor compliance in subglottic suctioning. the incidence of vap was 24.7% with a vap rate of 30.87/1000 ventilator days. there was a significant correlation between primary diagnosis, hemodialysis, massive blood transfusion and development of vap (p<0.05)). mean duration of ventilation (p<0.001) and mortality (p<0.05) were highly significant in vap patients. conclusions: improvement in compliance towards vap bundle and reduction of risk factors can help decrease incidence of vap and related morbidity and mortality. preventive strategies are effective in reducing ventilation-associated pneumonia (vap) in adults [1, 2] . in paediatric population there are no data about vap prevention, so we introduced a new bundle (vap-p) based on the available evidence for adults. this was designed as a before-after study. we enrolled all patients admitted to 8-bed medical-surgical paediatric icu at gemelli hospital in rome, requiring mechanical ventilation for at least 48 hours. patients with pre-existing tracheostomy were excluded. vap-p has been introduced since 2018 in order to improve quality of assistance. our bundle consisted in twice a day oral hygiene with chlorhexidine swab, daily check of oral bacterial colonization and aspiration prevention. comparison was made with an historical group including patients admitted before vap-p introduction (since 2016 to 2017). all data about demographics, antimicrobial therapy, icu stay and treatments, were collected. results: 162 patients were included (82 after and 80 before vap-p introduction). 5(6%) events of vap were recorded in vap-p group compared to 16 (20%, p=0.01) vap-p group had less vap per days of mechanical ventilation (1/100 compared to 3.3/100 p=0.01). multivariate analysis yielded an or of 0.23 (95%ci 0.07-0.81) for vap incidence after bundle introduction. mortality rate was slightly reduced in vap-p group (2.4%vs 6.2% p=ns). patients who developed vap required more days on mechanical ventilation and had higher mortality rate (12 vs 5 days p<0.001 and 14%vs 3% p=0.047, respectively). our vap-p seems effective in reducing vap incidence in critically ill paediatric population. introduction: ceftolozane/tazobactam (c/t) is a new antibiotic against mdr gramnegative bacteria infections, whose target population are the critically ill patients. even though 2/1 g dose safety administered as a 3hour-infusion has been already assessed, these patients can be under renal replacement therapy (rrt) and suffer changes in their volume of distribution (vd) that may affect antibiotic concentrations. the objective was to determine concentration reached by 3g c/t (3hour infusion) in septic patients on rrt (cvvhdf) and interdose behavior. we have used rrt machine prismaflex with oxyris filter and m100. hplc-uv method was used for simultaneous quantification of c/t. study population consisted of three obese critically ill patients with sepsis, on cvvhdf while receiving 3g c/t every 8 hours. samples were taken of prefilter, post filter blood and effluent, 30 min before infusion and 1, 2 and 4 hours after the end of it. we found great interpatient variability with the lowest cconcentration values in the patient with more hemodynamic instability using oxyris filter. even though cmax was less than reported in healthy subjects, we found similar values of auc and t ½ in comparison with healthy population studies. cmax of t was also compromised in comparison with values reported in healthy subjects, but with higher auc and t ½. cvvhdf contributes to c/t clearance. m100 filter showed the least clearance and higher values of auc and t ½. extraction rate was similar in all patients and filters (figure 1) . cmax achieved may be impaired because of the varying vd caused by obesity and rrt, but not affecting the antibiotic characteristics and behaviour. we conclude that because of the variety of clinical conditions, c-concentration is compromised particularly in hemodynamically unstable patients. however, the small sample doesn´t let us extrapolate these results. the extended infusion seems to be adequate to achieve the interdose antibiotic concentration. the use of biomarkers in sepsis is useful for early diagnosis and prognosis. the desired marker should be sensitive, specific, fast and accurate. procalcitonin (pct) measurement is approved by the fda even its efficacy is still under question. the determination of alfatorquetenovirus (ttv) could be a useful marker [1] . we analyzed 55 samples from 23 patients admitted to icu with clinical suspicion of sepsis. analytical data of c-reactive protein (crp), neutrophils and procalcitonin were collected. the sofa and apache ii scales were calculated and patients stratified according to these values in good and poor prognosis. ttv quantitative determination was carried by using a quantitative crp 2 . we calculated area under the curve (auc) of ttv plasma levels as a function of time. the statistical analysis involved u-mann-whitney and spearman test, using chi 2 for qualitative variables. results showed a not significant (ns) inverse relationship between the ttv auc and the patient proinflammatory level. a tendency (ns) was found between poor prognosis and the pct median values and crp being higher in the poor prognosis.group. a trend showed lower ttv dna count related to worse prognosis. an inverse relationship was found between pct and crp values and the ttv copies /ml plasma, ns correlation in the case of pct. there was a clear trend between the neutrophils´expansion and the regression line slope, obtained between ttv loads in the first two study steps. fig. 1 (abstract p432) . patient pk/pd measurements value>0.05), suggesting that the adsorptive mechanism wasn't primarily mediated by plasma protein. ha330 was saturated after adsorption of a total of 280.28±12.33 mg of van. the adsorptive kinetics showed an exponential reduction of van mass that reached a plateau after 30 minutes of circulation. in our study, simulating in vivo conditions of hp using ha330 during sepsis, a rapid and clinically relevant removal of van has been shown. after 2 hours of hp, we suggest to assess van plasma concentration and a loading dose of van should be considered. however, not knowing the potential interactions with other drugs, further in vivo studies are warranted to confirm these findings. assessing the volume of blood taken for blood culture and culture positivitydo we need to take less blood? it is commonly accepted that larger blood culture (bc) volumes (bcv) increase the yield of true positive cultures, and optimally 20 cc of blood should be obtained per set (2 bottles). only scarce data exists on the matter of optimal bcv. it is unknown what is the minimal volume that is acceptable for bc. the objective of this study was to determine the association between bcv and the rate of positive bc. blood taken for cultures in bd bactec plus aerobic/f negative bottles was collected from icus and acute care floors at 8 hospitals at the dmc over 6 months. blood volume was estimated automatically from blood background signal data in the bd bactec fx instrument. cultures were analyzed for each bottle. data was summarized for every month as the average volume and number of cultures taken and rate of positive bc for every unit. units were classified according to unit type (icu, medicine, surgery, mixed, emergency department (ed), organ/bmt or "other" which did not fit the previous categories) and analyzed as a group. a total of 23795 cultures were taken in 84 units. there is a positive association between bv and positive bc rate for ed and "other" units (irr=1.27, p=0.006 for the ed, irr=5.00, p<0.001 for "other" unit). all other units had no association between bv and positive bc rate (figure 1 ). secondary analysis, excluding pediatric units, gave very similar results. when comparing bv between unit types, the ed and "other" unit had significantly lower bv (2.4 ml in the ed and 3.5 ml in "other" unit compared to 4.9 ml in the icu, 4.7 ml in surgery, 4.2 ml in mixed and 7.7 ml in bmt). the correlation between bv and positive bc rate is probably limited to units taking very low bv for cultures. units taking volumes above 4 ml show no improvement in positive bc rate when higher volumes are taken. better prospective studies should be done to further establish the minimal bcv needed and spare unnecessary blood loss to hospitalized patients without compromising bc yield. de-escalating antibiotics in sepsis with the use of t2mr in a 35bed greek university icu c vrettou, e douka, i papachatzakis, k sarri, e gavrielatou, e mizi, s zakynthinos 1st icu department, university of athens, evangelismos general hospital, icu, athens, greece critical care 2020, 24(suppl 1):p440 in septic patients, the early use of appropriate empiric antibiotic therapy reduces morbidity and mortality. de-escalation refers to narrowing the broad-spectrum antibiotics once the pathogen and sensitivities are known. t2 magnetic resonance (t2mr) is a novel method of detecting eskape pathogens. we aim at investigating if using t2mr technology can expedite de-escalation of broad spectrum antibiotics. this is a prospective observational study conducted in our 35-bed university icu. inclusion criteria were critically ill patients age>18 y.o., with newly diagnosed sepsis and clinical suspicion of eskape bloodstream infection. a sample for t2mr and a blood culture (bc) sample were collected simultaneously from the patients enrolled. the t2mr bacteria panel test was run according to the manufacturer's guidelines and the bcs were processed according to the hospital standard procedures. we recorded clinical data and administered antibiotics. results: 26 patients were included in the study. mean time to culture positivity was 84 hours while mean time to t2mr result was 3.5 hours. in 20 patients the results of t2mr were in concordance with the bcs. in the remaining 6 cases, the bcs were negative while the t2 mr detected one or more eskape pathogens. there were no false negative results. de-escalation in at least one drug was applied to 8 patients (30.8%). no escalation was applied to 15 patients (57.7%) and antibiotic escalation in 3 (11.5%). conclusions: t2mr provides a quicker detection time that could shorten the time to targeted therapy. in our population this corresponded to early (within 6-12h) antibiotic de-escalation in approximately 1/3 of the included patients. antibiotic stewardship in icu. a single experience l forcelledo 1 , e garcía-prieto 1 , l lópez-amor 1 , e salgado 1 , j fernández dominguez 2 , m alaguero 3 , e garcía-carús 4 the increasing antibiotic resistance in microorganisms urged interventions such as the antibiotic stewardship programs in icu focused on reducing the inappropriate use of antibiotics by improving the antibiotic selection, the dosage, administration route and length as well as improving clinical outcomes and reducing antibiotic resistance. retrospective study where antibiotic consumption was analysed and measured in days of therapy (dots) between 2015 and 2018 in a medical-surgical icu of a university hospital where a multimodal educational program was established. specific training in infectious diseases in critically ill patients, periodic clinical and formative sessions fig. 1 (abstract p439) . correlation of blood culture positivity rate with blood culture volume by unit type were performed for icu staff and specific leaders within the icu staff designated. results: 4128 patients were admitted to icu. there was a reduction of 20,5% in dots (figure 1 ), reduction in antimicrobial resistance rates (14,36 in 2015, 7,4 in 2018 [days of resistant microorganism/1000 patientdays]) without an impact in icu global mortality (19,7% in 2015, 19,4% in 2018). the resistant bacteria registered were acinetobacter baumannii, s. aureus mr, blee and carbapenemase-producing enterobacteriaceae, pseudomonas aeruginosa mr and clostridium difficile. the safe in antimicrobial consumption was 391500€ (70% reduction). the icu stay decreased from 8,2 days (2015) to 6,9 (2018) , with no variation in mean apache ii (17,8) . the bigger decrease in antibiotic consumption was in colistin related to the reduction in resistance bacteria, in special acinetobacter baumannii, in linezolid and in piperacilin/tazobactam, even more remarkable in 2018 due to shortage of supplies which meant an increase in meropenem. the application of an antibiotic stewardship program in icu succeeded in reducing antibiotic consumption, antibiotic resistance and costs without an impact in clinical outcomes like mortality or icu stay. clinical outcomes of isavuconazole versus voriconazole for the primary treatment of invasive aspergillosis: subset analysis of indian data from secure trial p kundu, s kamat, a mane pfizer limited, medical affairs, mumbai, india critical care 2020, 24(suppl 1):p442 the secure trial was designed to compare the safety and efficacy of isavuconazole (a) versus voriconazole (v) for primary treatment of invasive mould disease caused by aspergillus and other filamentous fungi. the present analysis is aimed at comparing the indian subset of patients with that of the overall trial population and to ascertain any similarity or difference in the primary efficacy endpoint and safety/tolerability in these two groups. in secure trial, 258 patients in one group received (i) & another 258 patients received (v). the indian subset had 29 patients. we have done a qualitative analysis as the sample size of the indian subset was small. non-inferiority of (i) to (v) in terms of all cause mortality from first dose to day 42 was assessed in overall patients. the treatment difference between (i) and (v) group in the indian subset of patients was analyzed. proportion of patients who had to discontinue treatment due to teaes was analyzed. the all-cause mortality in the overall trial population met noninferiority margin (table 1 ). in the indian subset, it was higher for (i) than (v). there was a lower incidence of ocular, hepatobiliary, skin & subcutaneous tissue disorders in the (i) treated patients (see table 1 ). in indian subset, the above adverse events were less in the (i) group, but statistical inference could not be done due to small sample size. however, similar trend of less number of patients discontinuing therapy due to teaes in the (i) treated patients was seen in the overall patients & the indian subset. the all-cause mortality in the indian subset was higher in the (i) patients. a trend similar to the overall population regarding safety parameters favoring (i) was seen in the indian patients. considering the significantly higher prevalence of ia in india, suitably powered study design is necessary to draw definitive conclusions on the non-inferior efficacy & better safety & tolerability of (i) over (v) in patients of ia. introduction: ventilator-associated pneumonia (vap) is one of the most frequent healthcare-associated infections, correlated with increased mortality,extended hospital stay and prolonged mechanical ventilation. considering the latest outbreak of multiresistant a. baumannii infections in the critically ill patients with vap, there is a growing concern regarding challenges of the antibiotherapy in these patients. although ceftazidim-avibactam is considered to have limited effects on a. baumannii, it is reported to have a synergic activity in combination with other antibiotics. we performed a retrospective, observational study which included 24 icu patients diagnosed with vap(cpis > 6). oxa 23 a. baumannii was isolated from the tracheal secretions using a rapid molecular diagnostic platform(unyvero a50 system). patients were divided in two groups according to the antibiotherapy:group a meropenem + colistin and group b meropenem + colistin + ceftazidim-avibactam.statistical analysis was performed using graphpad6 applying t-test and kaplan-meier curves, having the in-hospital mortality as primary outcome and days of mechanical ventilation and hospital stay as secondary outcomes. mean age(y.o) in group a was 46 and 52 in group b and in both groups mean charlson comorbidity index was 3 points. survival percent was higher in the group treated with ceftazidim-avibactam (67 % vs 57 %, p = 0.08)(fig. 1) . length of stay was significantly decreased in group b (26.5 days vs 43 days in group a, p = 0.046). number of days under mechanical ventilation was also decreased in the ceftazidim-avibactam group (19 vs 22) but the data was not statistically significant. in light of the important thread of multiresistant a. baumannii and the lack of therapeutic measures, the synergistic activity of ceftazidim-avibactam use in combination with other antibiotics may be a promising approach to lower the mortality and hospitalization in critically ill patients diagnosed with vap. impact of patient colonization on admission to intensive care on 28 and 90 days mortality g dabar 1 , c harmouch 2 , e nasser ayoub 3 , y habli 4 , g sleilaty 5 , j infections caused by multi resistant bacteria are a major health problem, especially in icus, and it may be associated with high mortality rates. colonization precedes infection in most instances; therefore it may be a marker of a poor outcome. we tried to determine the impact of colonization on mortality at 28 and 90 days in a population of patients admitted to one medical and one surgical icu in the same institution. medical records review over three years 2016-2018 of all patients admitted to one surgical et one medical icu at hotel dieu de france hospital staying more than 24h. colonization to resistant bacteria was defined as mrsa, esbl, mdr, and vre. all patient received a nasal and rectal screen on icu admission, in intubated patients tracheal aspirate was considered as colonization in the absence of clinical respiratory tract infection. demographics, apache, sofa, immunosupression, charleston comorbidity index, length of stay, mechanical ventilation, hospitalization and antibiotic use in the previous 3 month were collected. mortality at 28 and 90 days was assessed through medical records or phone call. pearson chi-square was calculated for the association of colonization and mortality at 28 and 90 days, and subsequently odd ratio was estimated. introduction: critically unwell patients have been observed to respond unpredictably to traditional intermittent dosing (id) schedules of vancomycin, likely due to the complex physiological derangements caused by critical illness. continuous infusion (ci) of vancomycin has been suggested to overcome such problems by allowing more regular therapeutic drug monitoring and subsequent effective dose titration [1] . this study conducted at a tertiary intensive care unit, reports our experience following implementation of a continuous vancomycin infusion protocol. prospective data was collected over two consecuative periods of three months, initially capturing plasma levels for id (target level of 15-20mg/l) followed by reviewing plasma concentration levels in a ci protocol (target level of 20-25 mg/l). patients recieving renal replacement therapy were excluded. a total of 22 intermittent vancomycin prescriptions were administered and dosing levels observed. in the three month ci period, 26 patients received ci vancomycin and levels subsequently checked. the ci protocol resulted in increased blood sampling (107 samples in ci group vs. 73 samples in id cohort). two non serious incidents were reported in the ci cohort relating to preparation of vancomycin. both groups had a comparable median time to therapeutic range (48 hours). however, ci vancomycin group had a greater proportion of first samples outside the desired therapeutic range (70%vs 36%) (figure 1 ). as the therapy continued, ci vancomycin demonstrated a greater propensity towards consistent therapeutic levels than that observed with id. 83% of patients on a ci regime achieve the desired target levels compared to 77% in the id cohort (fig. 1) . it was positive for single or multiple microbes in 9(26.5%) and 22(64.7%) samples respectively. single or multiple resistance genes were detected in 5(25%) and 20(80%) samples respectively. bfpcr was positive only for bacteria in 13(38.2%), virus in 2(5.9%) and for both in 16(47.1%) cases. influenza a was found in 10(29.4%) cases. the most common organisms in community and hospital acquired pneumonia were streptococcus pneumoniae (4/12) and a. baumannii (10/22) respectively. bacterial cultures were concordant with bfpcr in 11/11 (100%) of positive cases. decisions to change antibiotics could be taken earlier based on bfpcr (p< 0.001) than if were based solely on culturesboth in culture positive (9.7± 14.3 vs 50.03±6.0 hrs) and negative cases (14.7±14.9 vs 48.0+4.3 hrs) where antibiotics would have remained unchanged. based on bfpcr antibiotics were escalated in 17(50%) patients and teicoplanin (11/ 19) was most often stopped. bal bfpcr were obtained significantly earlier, identified more organisms and bacterial resistance than culture reports and lead to more frequent and earlier antibiotic changes. severe community-acquired pneumonia (scap) is a frequent cause of hospitalization and mortality. ceftaroline is efficacious for treatment of cap (port risk class iii or iv). most severe patients were excluded from the clinical trials, so the efficacy of ceftaroline in these kind of patients is unknown methods: this is a health record-based retrospective before-after study in a tertiary care hospital. all scap patients admitted in icu between november 2017 and february 2019 receiving ceftaroline were included. control group included patients with same inclusion criteria but receiving ceftriaxone. propensity scores to adjust for potential baseline differences between groups were performed. levofloxacin or azythromicin were administered in both groups. primary outcome was the change in sofa score over the first 96h and secondary were days of mechanical ventilation, respiratory failure at 96h, need of rescue antibiotics, length of stay and mortality results: there were 28 patients in ceftaroline group and 43 in ceftriaxone group. baseline characteristics were similar except from more intubated patients in ceftaroline group (figure 1 ). there were less respiratory failure at 96h in patients with ceftaroline treatment (-73.3% vs. -51.6%; p 0,015), but no differences in other organ failures, mortality, days of mechanical ventilation or los. there were more need of rescue antibiotics in ceftriaxone group (7.1% vs .46.5%; p 0,001). we found more streptococcus pneumoniae isolation in ceftaroline group (18 (64.2%) vs 11 (25.5%); p = 0.001); more empiric use of oseltamir (16 (57.1%) vs 11 (25.5%); p = 0.012), but no more influenzae infections (11 (39.2%) vs 8 (18.6%); p = 0.101). s. aureus was detected in 1 patient in ceftaroline group and in 5 in ceftriaxone group. introduction: acute respiratory failure (arf) due to pulmonary infections is a usual cause of intensive care unit (icu) admission. immigration patterns and iatrogenic immune-suppression have made tuberculosis (tb) a common disease in western europe. severe tb requiring icu care is rare. nevertheless, mortality associated with active tb and arf is poor [1] . adult patients with tb admitted to icu from 2014-2018 were identified retrospectively. diagnosis was based on: positive cultures of sputum, bronchial aspirates or bronchioalveolar lavage fluid. demographic characteristics, reasons for admission, hiv status, anti-tb treatment and mortality were recorded. total of 25 patients with tb were admitted to icu. mean apache ii score was 20,2±6,9. sixteen were male. mean age 49,1±14,7 years. eight (32%) were hiv-positive, 3 (12%) diabetes mellitus type 2, 3 (12%) chronic liver disease. six (24%) had other causes of immunesuppression. main causes for icu admission were arf due to nonmycobacterium tuberculosis pathogens in 64%, acute liver failure in 12%, septic shock due to non-respiratory cause in 8%. overall, 52% were on anti-tb treatment at time of admission. tb involved the lung parenchyma in all patients. pleural involvement was present in 12% and lymph node in 20%. extrapulmonary sites were present in 28%: urogenital, gastrointestinal, bone marrow. pathogens identified in over-infections: 16% gram positive coccus, 20% gram negative bacilli, 16% fungal, 4% mdr-pathogen. one patient hiv-positive suffered arf due to pneumocystis jiroveci. overall, 64% died during icu stay. besides its latent evolution, mortality of tb patients admitted to icu is extremely high. arf due to over-infection seems to be the main cause for icu admission and mortality. better preventive approach of these patients may improve their outcome. introduction: human african trypanosomiasis (hat) is rarely encountered by critical care clinicians, but is an important differential for fever in the returning tropical traveler. late disease is characterized by seizures, fever and multi-organ failure [1, 2] . we present an anonymized case presenting from an endemic area in zambia referred for tertiary critical care management. the patient was too obtunded to give informed consent and his relatives could not be contacted despite extensive efforts. a middle-aged man with no past medical history from rural zambia presented to a local clinical officer post with fever and arthralgia. he was treated twice with anti-malarial medication without resolution of symptoms. two months later he was admitted febrile and obtunded to a local hospital with worsening confusion. he was transferred 8 hours by ambulance to our facility in lusaka, which is the only public tertiary critical care unit in zambia results: gcs on arrival was e3m4v2 without localizing neurology. microbiology investigations were negative, including for toxoplasma, cryptococcus, hiv or malaria. the patient suffered a generalized seizure followed by a sustained gcs of 3 and was admitted to the icu for invasive ventilation and seizure control. peripheral blood smears demonstrated trypanosomes consistent with hat secondary to trypanosoma brucei rhodesiense. he was commenced on melarsoprol but rapidly deteriorated, with signs of melarsoprol-induced arsenic encephalopathy and subsequent tonsillar herniation. his death was confirmed by neurological criteria. conclusions: icu management of fulminant hat involves supportive neurocritical care plus melarsoprol, a toxic arsenic compound with common side effects of hepatotoxicity and dysrhythmia. arsenic encephalopathy occurs in 10% of late hat, with a fatality rate of 70% [1] . early diagnosis is associated with a 95% survival rate in developed world travelers repatriated from endemic areas [2] . lithium chloride to prevent endothelial damage by serum from septic shock patients (in vitro study) a kuzovlev 1 the aim of the study was to investigate into effectiveness of lithium chloride (licl) as agent that prevents damage to the monolayer of endothelial cells under the action of serum from multiple trauma patients with septic shock. methods: serum from 5 pts with septic shock (sepsis-3) and 5 healthy donors was withdrawn. monolayer of ea.hy926 endothelial cells were incubated for 3 hrs at 37°c with healthy person's serum and with septic patient's serum without licl and with it at concentrations of 0.01 mmol, 0.1 mmol, 1 mmol, 10 mmol. licl was added 1 hour before the change of serum. after incubation cells were washed and fixed with 2% paraform solution and permeabilized with 1% triton x-100 solution. fixed cells were stained with primary antibodies to vecadherin and then incubated with secondary antibodies conjugated with oregon green 488 fluorescent dye as well as with phalloid red and hoechst dye 33342. images were processed by fluorescence microscope and imagej 1.44p and metavue 4.6 programs. western blotting was used to detect antibodies to ve-cadherin, claudin and gsk-3beta. statistics included mann-whitney test and chi-square test. incubation of a monolayer of endothelial cells with 5% serum of septic shock patients led to loss of ve-cadherin contacts and decrease of claudine. preincubation with licl 0.01 mmol did not prevent dismantling of claudine, actin, ve-cadherins; 0.1 mmol licl prevented it (p>0.05), but at higher concentrations (1 mmol, 10 mmol) almost completely protected endothelial monolayer from destruction of intercellular contacts (p<0.05). serum had almost no effect on the phospho-gsk-3β level after 5 min, 15 min, 30 min and 1 hr, but caused a significant (60%) decrease in its level after 2 and 4 hrs. licl (1 mmol) caused a significant increase in phospho-gsk-3β already 15 mins and up to 4 hrs after exposure. licl prevents septic damage to the monolayer of endothelial cells in vitro in a gsk-3beta mediated way. introduction: the autonomic nervous system (ans) controls both heart rate and vascular tone, which are known to be impaired during septic shock (ss) . acute inflammation is presumed to increase arterial stiffness of large arteries in experimental studies [1] . the objectives of this work are to verify if standard ss resuscitation modulate mechanical vascular properties and to verify if alterations in these vascular properties and ans activity are correlated. a protocol of fecal peritonitis septic shock and standard resuscitation (fluids and noradrenaline) was applied on 6 pigs. the arterial blood pressure waveform was recorded in the central aorta and in the femoral and radial arteries. the characteristic arterial time constant tau was computed at the three arterial sites, based on the twoelement windkessel model [2] . the total arterial compliance (ac) and the total peripheral resistance (tpr) were also estimated. baroreflex sensitivity (brs), low frequency (lf, 0.04-0.15 hz) spectral power of diastolic blood pressure, and indices of heart rate variability (hrv) were computed to assess ans functionality. results: septic shock induced a severe vascular disarray, decoupling the usual pressure wave propagation from central to peripheral sites, as shown by the inversion of pulse pressure (pp) amplification, with a higher pp in the central aorta than in the peripheral arteries during shock. the time constant tau together with ac and tpr were independently decreased. a decrease in brs, lf power, and hrv describe an ans dysfunction. after the administration of fluids and noradrenaline, both vascular and autonomic dysfunction persisted and these were found to be significantly correlated. measures of mechanical vascular function and ans activity could represent an useful end-point to guide further clinical investigations and refine our understanding of ss mechanisms, especially under medical treatment. introduction: lipopolysaccharide (lps), is a component of gram-negative bacteria known for its activation of the host immune system. the phospholipid transfer protein (pltp) has previously been shown to promote the binding of lps to lipoproteins, to limit inflammation and to lower mortality following injections of lps or bacterial infection. the aim of the present study was to investigate the role of pltp and lipoproteins in the detoxification of lps from the peritoneal cavity. injection of lps intra-peritoneally (ip) (1mg/kg) to wild type (wt) and pltp knocked-out mice (pltp-ko) (n = 9 per group). mass concentration and activity of lps were quantitated by lcmsms analysis of 3-hydroxymyristate and lal bioassay, respectively. lipoprotein fractions in plasma were separated by ultracentrifugation (n=10 vs n =12). following intra-peritoneal injection, clearance of intra-abdominal lps was faster and plasma neutralization was more efficient in wt than in pltp-ko mice ( figure 1) . indeed, lps found in plasma of wt mice was proportionally less active, sustaining a higher capacity for wt mice to neutralize lps (figure 1b) . quantitative dosage of lps in portal blood, 15 minutes after ip injection, revealed that plasma lps associates rapidly with the lipoprotein fraction (hdl plus ldl), and in higher proportions as compared to pltp-ko mice (66 [62-72] % vs 50 [41-54] %, respectively; p < 0.01). in line with previous studies, these observations now indicate that, lps readily associates with lipoproteins in a neutralizing process pltp mediated. finally, even with a heavy lps load (25 mg/kg), the bulk of lps was still found in the lipoprotein fraction (80 [80-90] %), suggesting that lipoproteins plus pltp in wt mice have a high capacity to detoxify intraperitoneal lps. in a model of peritonitis, lipoproteins and pltp were found to constitute key playors for peritoneal clearance and neutralization of lps. it emerges as a key pathway for the resolution of the inflammatory response in peritonitis. introduction: autotaxin (atx, enpp2) is a secreted enzyme present in biological fluids that catalyses the production of lysophosphatidic acid (lpa). lpa is a bioactive phospholipid evoking various cellular responses in most cell types. upregulated atx levels have been reported in various chronic inflammatory diseases. given the established role of lpa in the inflammatory response, we investigated a possible role for the atx/lpa axis in lps-induced endotoxemia. methods: lps was injected intraperitoneally (20 mg/kg) in mice producing 50% atx levels (atx df/+ , heterozygous null mutant mice), in mice producing 20-30% reduced atx levels upon inducible inactivation (r26creer t2 /enpp2 n/n mice) and in mice expressing 150-200% increased atx levels (enpp2-tg mice). kaplan-meier survival analysis was performed. atx activity was measured using the toos activity assay. results: atx df/+ mice that produce almost 50% reduced serum atx levels show increased survival compared to their littermate controls. for the inducible inactivation of atx, enpp2 n/n targeted mice were crossed with the r26cre-er t2 mice and tamoxifen induction enabled temporal control of floxed gene expression. r26creer t2 /enpp2 n/n mice were more protected against lps-induced endotoxemia compared to control mice. enpp2-tg mice overexpressing autotaxin and showing a 2-fold increase in plasma levels do not display improved survival rates compared to control group. conclusions: atx participates in systemic inflammation, as reduced atx levels in circulation decrease lethality of mice from caused by lps. the excess amount of circulating atx does not exacerbate the systemic inflammatory response to lps. introduction: pneumonia (pn) is a prevalent and severe infectious lung disease. host genetics plays an essential role in the pathogenesis of infectious diseases including pn [1] . the aim of the study was to analyze the variability of genes associated with neutrophil activation in pneumonia. to identify differential expressed genes (degs) in communityacquired (cap) and hospital-acquired pneumonia (hap) dataset «genome-wide blood transcriptional profiling in critically ill patients -mars consortium» (gse65682) from gene expression omnibus was analyzed (logfc≥2.0, fdr-corrected p-value<0.05). degs associated with neutrophil activation were selected according to gene ontology go:0042119 («neutrophil activation»). with the use of gtex portal and blood eqtl browser, we searched for esnps (expression single nucleotide polymorphisms) in whole blood for neutrophil activation genes differentially expressed in cap/hap. these esnps were further analyzed for their association with pn via the global biobank engine (gbe). a total of 46 degs from gse65682 correspond to go:0042119 genes (43 up-and 3 down-regulated) of which 39 genes were common to cap and hap. functional enrichment of 46 degs based on disgenet detected top-5 diseases associated with these genes (fdr-corrected p-value<0.05): myeloid leukemia, chronic; sepsis; asthma; lung diseases; allergic asthma. for these 46 genes 1366 esnps common to gtex portal and blood eqtl browser were identified. more than half of all variants were located on the second chromosome and influenced the expression of tnfaip6 and il18rap genes. among all esnps we identified variants associated with pn in the gbe (table 1) . we identified genes related to neutrophil activation, genetic variability of which was associated with pneumonia. sepsis was induced in wild-type c57bl6 mice (n=41) and cse knockout mice (n=41) by i.p. injection of 10 8 cfu/mice mdr p. aeruginosa. similar experiments were repeated after cyclophosphamide induced neutropenia. survival was recorded for 7 days. mice were sacrificed for determination of bacterial load and myeloperoxidase (mpo) activity as a surrogate marker of myeloid cell recruitment. cytokines were measured in serum by legendplex inflammatory panel. total leukocytes from mice spleens, with or without pretreatment with the h 2 s donor gyy3147, were incubated with 1 x 10 4 cfu/ml mdr p. aeruginosa. bacterial clearance was recorded. we observed a significant decrease in survival of cse -/mice as compared to cse +/+ mice (12% vs. 47%; p: 0.025). this survival advantage was eliminated in neutropenic mice (17% for both groups, p: 0.873). cse -/mice had increased pathogen load in the liver (6.57 ± 0.13 vs 5.26 ± 0.50, p: 0.029) and lung (6.70 ± 0.17 vs 5.29 ± 0.55, p: 0.035). mpo activity was lower in cse -/mice in the liver (634 ± 71 vs 1029± 179, p: 0.048) and lung (7627 ± 585 vs 11121 ± 1468, p: 0.34). cse +/+ mice had increased serum levels of il-23 (121.13 ± 33.68 vs 31.41 ± 7.02 of cse -/-, p: 0.001); mcp-1 (4769.91 ± 908.83 vs 1940.37 ± 1062.65, p: 0.026) and gm-csf (22.91 ± 4.66 vs 8.11 ± 1.92, p: 0.004). phagocytic activity of leukocytes from cse -/mice was reduced compared to cse +/+ mice. this deficit was eliminated after gyy4137 pretreatment (fig. 1) . deficiency of host-derived h 2 s leads to increased susceptibility to mdr p. aeruginosa infection due to an inefficient neutrophil chemotaxis and neutrophil mediated phagocytosis. acknowledgement funded by the itn horizon 2020 marie-curie european sepsis academy introduction: neuroinflammation often develops in sepsis along with increasing permeability of the blood-brain barrier (bbb), which leads to septic encephalopathy [1] . the barrier is formed by tight junction structures between the cerebral endothelial cells [2] . we investigated the expression of tight junction proteins related to endothelial permeability in brain autopsy specimens in critically ill patients deceased with sepsis, and analyzed the relationship of bbb damage and measures systemic inflammation and systemic organ dysfunction. case series included all adult patients deceased with sepsis in the years 2007-2015 with brain specimens taken at autopsy available. specimens were categorized according to anatomical location (cerebrum, hippocampus, cerebellum). the immunohistochemical stainings were performed for occludin, zo-1 and claudin. patients were categorized as having bbb damage if there was no expression of occludin in the endothelium of cerebral microvessels. results: 38% (18/47) developed multiple organ failure before death. 74.5% (35/47) had septic shock. the deceased with bbb damage had higher sofa maximum scores (16 vs.14, p=0.04), and had more often procalcitonin levels above 10 (56 % vs.28 %, p= 0.045). bbb damage in cerebellum was more common in cases with c reactive protein above 100 mg/l as compared with crp less than 100 (69% vs. 31 %, p=0.025). absence of zo-1 expression in cerebral meningeal samples associated with bbb damage (17 % vs. 0 %, p=0.046). positive blood cultures (n = 22) were associated to absence of zo-1 expression in cerebellar glial cells (92 % vs. 44 %, p=0.018). in fatal sepsis, damaged bbb defined as loss of cerebral endothelial expression of occludin ( figure 1 ) is related with severe organ dysfunction and systemic inflammation. loss of zo-1 in endothelial cells associates with bbb damage, and sepsis contributes to zo-1 loss in cerebellar glial cells. oxylipins are oxidative breakdown products of cell membrane fatty acids. animal models have demonstrated that various vasoactive oxylipin pathways may be implicated in septic shock pathophysiology but these have been poorly studied in humans. oxylipin profiling was performed on serum samples collected on enrolment to the vanish (vasopressin vs. norepinephrine as initial therapy in septic shock) trial. samples were analysed with liquid chromatography-mass spectrometry. patients were followed up until 28 days. results: samples were collected from 154 of 409 (37.7%) patients on inclusion to the trial and 39 (25.3%) had died by 28 days. non-survivors were found to have higher levels of a number of oxylipins including: 14,15-dihydroxyeicosatrienoic acid (dhet) (p<0.01), 11,12-dhet (p=0.03), 15(s)-hydroxyeicosatetraenoic acid (p=0.02), 14-hydroxyoctadeca-pentaenoic acid (p=0.04) but lower levels of the precursor eicosapentaenoic acid (p=0.012). when corrected for multiple comparisons with the benjamini-hochberg test, only 14,15-dhet remained significant (p=0.025). although there was a difference in median 14,15-dhet levels between survivors and non-survivors, many values were below the level of detection (n=84/154 (54.5%)). as such, we also analysed 14-15-dhet as a binary variable (figure 1 ). patients with detectable 14,15-dhet were more likely to die (hr 2.4 [95% ci 1.2-4.6], p< 0.01) and have a higher median lactate (p =0.01) and total sofa score (p< 0.01) than those patients where baseline 14,15-dhet was undetectable. our study suggests the oxylipin 14,15-dhet may be associated with septic shock severity and 28-day mortality. these results are consistent with the known vasodilatory actions of this class of oxylipin. more work is needed to confirm its exact role in septic shock and whether this pathway is amenable to therapeutic intervention. introduction: activation of neutrophils is a mandatory stage and a sensitive marker of systemic inflammatory conditions that can lead to the development of multiorgan failure. the aim of the study was to investigate into the antiinflammatory effects of lithium chloride on human neutrophils in vitro. study was carried out on neutrophils isolated from the blood of 5 healthy donors. 50% of neutrophils were activated by 100 mkm fmlp, 50% -by 100 ng/ml lipopolysaccharide (lps); then their activity was evaluated by fluorescent antibodies to cd11b and cd66b degranulation markers. intact and activated neutrophils were treated with a solution of lithium chloride (9 mmol). immunoblotting was used to assess gsk3b activity in neutrophils. mann-whitney criterion and p<0.05 were used for statistics. results: lithium chloride 9 mmol decreased the level of expression of cd11b on intact neutrophils by 16% (p=0.07), cd66b by 15% (p=0.07). fmlp increased cd11b expression on neutrophils by 2.6 times (p=0.0007), cd66b by 2.5 times (p=0, 0022). addition of lithium chloride solution to fmlp activated neutrophils reduced the expression of cd11b (p= 0.0317) and cd66b (p=0.0079). lps increased cd11b and cd 66b expression by 2.1 times (p=0.0007, p=0.0022, respectively); addition of lithium chloride reduced the expression of cd11b (p=0,0317) and cd66b (p=0.0079) on neutrophils. fmlp led to a dephosphorylation of gsk-3b by 47% (p<0.05), lithium chloride increased its phosphorylation by 387% (p <0.05). adding lithium chloride to activated fmlp neutrophils restored the level of gsk-3b phosphorylation by 277% compared to controls (p<0.05). lithium chloride modulates the inflammatory activation of neutrophils by bacterial components through the phosphorylation of gsk3b in neutrophils. human host immune responses to lipopolysaccharide: a comparison study between in vivo endotoxemia model and ex vivo lipopolysaccharide stimulations using an immune profiling panel dm tawfik introduction: sepsis, a leading cause of mortality among critically-ill patients in the icu, recently recognized by the who as a global health burden. patients that suffer from sepsis exhibit an early hyper-inflammatory immune response which can lead to organ failure and death. in our study, we assessed the immune modulations in the human in vivo endotoxemia model and compared it to ex vivo lipopolysaccharides (lps) stimulation using 38 transcriptomic markers. methods: eight healthy volunteers were challenged with intravenous lps in vivo. in parallel, blood from another 8 volunteers was challenged with lps ex vivo. blood was collected before and after 4 hours of lps challenge and tested with the immune profiling panel (ipp) prototype using the filmarray® system. the use of ipp showed that markers from the innate immunity dominated the response to lps in vivo, mainly markers related to monocytes and neutrophils. comparing the two models, in vivo and ex vivo, revealed that most of the markers were modulated in a similar pattern (68%). some cytokine markers such as tnf, ifn-γ and il-1β were under-expressed ex vivo compared to in vivo. t-cell markers were either unchanged or up-modulated ex vivo, compared to a down-modulation in vivo. interestingly, markers related to neutrophils were expressed in opposite directions, which might be due to the presence of cell recruitment and feedback loops in vivo. the majority of ipp markers showed similar patterns of expression post-lps challenge in both models, except for several markers related to neutrophils and t-cells. the ipp tool was able to capture the early immune response in the human in vivo endotoxemia model, which is a translational model mimicking immune host response in septic patients. introduction: serum levels of tyrosine kinase receptor mer and its ligand gas6 predict mortality in septic patients in the intensive care unit. however, whether their early measurement at emergency department (ed) presentation also predicts mortality and organ failure still needs to be clarified. in this multicentre observational study, septic patients admitted to 5 italian eds were included [1] . at ed presentation blood samples were taken for routine biochemical analyses and serum mer and gas6 measurement. urinalyses, blood gas analyses and chest x-ray were routinely performed. mortality at 7 and 30 days, as well as the presence of organ damage such as acute kidney injury (aki), thrombocytopenia, pt-inr derangement and sepsis-induced coagulopathy (sic) were evaluated according to baseline levels of mer and gas6. in conclusion, neither mer nor gas6 are early predictors of mortality in septic patients at ed presentation. however, mer independently predicted the development of sic, thrombocytopenia and pt-inr derangement in this population. glycocalyx shedding correlates with positive fluid balance and respiratory failure in patients with septic shock n takeyama, y kajita, t terajima, h mori, t irahara, m tsuda, h kano aichi medical university, department of emergency and critical care medicine, aichi, japan critical care 2020, 24(suppl 1):p463 endothelial hyperpermeability would play a major role in septic shock related organ failure. the aim of this study is to clarify the relationship between glycocalyx shedding and respiratory failure, sofa score, plasma angiopoietin (ang)-2 level and patient survival. methods: plasma samples were collected from 30 septic shock patients from admission to icu discharge and 10 healthy volunteers. plasma syndecan (syn)-1 and ang-2 were measured and clinical data was also collected. septic shock patients were classified into 3 groups according to the time-course change of syn-1 levels. excess syn-1 (>400 ng/ml) during 0 to 3 days and remaining high following 4 to 7 days were assigned to group i. excess ang-2 during 0 to 3 days and decreased following 4 to 7 days were assigned to group ii. moderate increase (<400 ng/ml) during 0 to 7 days were assigned to group iii. results: plasma syn-1 levels are positively associated with increased ang-2 levels (r2=0.41, p= 0.005), suggesting that ang-2 is involved in endothelial hyperpermeability. fluid balance and ventilator-free days (vfd) are significantly increased in group i as compared with group iii. sofa score, apache ii and patient outcome does not show any differences between groups i, ii, and iii. the positive correlation between glycocalyx shedding and fluid balance indicates plasma syn-1 may be a valuable marker for endothelial hyperpermeability. the negative correlation between glycocalyx shedding and vfd indicates plasma syn-1 may be a valuable marker for respiratory failure. the plasma level of syn-1 for prognosis and organ failure excluding ards in patients with septic shock requires further investigation. serial procalcitonin measurements in the intensive care unit at hiroshima university hospital k hosokawa, s yamaga, m fujino, k ota, n shime hiroshima university hospital, department of emergency and critical care medicine, hiroshima, japan critical care 2020, 24(suppl 1):p464 introduction: serum procalcitonin (pct) is a promising biomarker for differentiating bacterial infections from other inflammatory states. moreover, including serial pct measurements in the management of acute respiratory infection reduces the duration of antibiotic therapy without increasing the mortality. however, limited real-world information is available regarding the use of pct in intensive care units (icus). we extracted and analysed data from january 1 to december 31, 2018 from all the orders and results of pct measurements in the icu (26 beds) at hiroshima university hospital. a total of 1,252 pct measurements from 409 icu patients were included. in 170 patients, pct was tested ≥3 times during a single icu stay. serial pct measurements showed a fade-out pattern (76 [45%] patients), a second day-peaked decrease pattern (35 [21%] patients), and a series of negative patterns (30 [18%] patients). compared to patients who demonstrated the fade-out pattern, those who demonstrated the second day-peaked decrease pattern had higher mortality rates (3% vs. 20%, p < 0.01). approximately one-third patients in the icu who had decreasing serial pct values demonstrated the second day-peaked decrease pattern. since this group of patients had poorer survival, further studies are needed to clarify the association between a late rise in pct levels and delayed therapeutic intervention. the research was performed on 200 full-term newborns; no clinical signs of bacterial infection were diagnosed. on the 1, 5, 20 days the plasmà concentration of il-1ß, il-6, il-8, tnf-α, g-csf, sfas, fgf, no was determined by capture elisa; cd3cd19, cd3cd4, cd3cd8, cd69, cd71, cd95, hla-dr, cd34, cd14, cd3cd56, lymphocytes in apoptosis -immunophenotype analysis. by applying the statistical cluster population analysis of the immunological criteria under study we have evaluated the feasibility of sepsis diagnostics at the admission to the intensive therapy unit. the diagnostic rule for sepsis has been formulated by applying the "decision tree" approach to the "r" statistic medium. the cluster analysis confirms the presence of two clusters (presence of absence of sepsis: these two components explain the 60.81% of the point variability). the diagnostic rule for the early diagnostics of sepsis is as follows: disease develops providing during the first 48 hours cd95≥16.8%, no≤9.6 mkmol/l or cd95≤16.8%, cd34≤0.2%, cd69≥4.12% or cd95≤16.8%, cd34≤0.2%, cd69≤4.12% and lymphocytes annexinv-fitc+pi-≥12.3%. 45 newborns featured the confirmed sepsis development. the accuracy of this diagnostics amounts to 95.41%; sensitivity to 97.06%; specificity to 94.67%; diagnostic false positive share to 5.33%; diagnostic false positive share to 2.94%; positive result accuracy to 89.19%; negative result accuracy to 98.61%. the aggregate determination of cd95, cd69, annexinv-fitc+ pi-, cd34 and the plasma concentration of no enables the pre-clinical diagnostics of sepsis development. efficacy of pancreatic stone protein in diagnosis of infection in adults: a systemic review and metaanalysis of raw patient data j prazak 1 , p egimann 2 , i irincheva 3 , mj llewelyn 4 , d stolz 5 , lg de guadiana-romualdo 6 , r graf 7 , t reding 7 , hj klein 8 , ya que 1 fig. 1 (abstract p469) . impact of 24h lactate and bio-adm values in patients with elevated lactate level at admission. the green curve in the left km-plot illustrates data from 75 patients with 5 events; the red curve 70 patients with 18 events. the green curve in the right km-plot illustrates data from 28 patients with 4 events; the red curve 96 patients with 48 events. of note, differences in numbers between admission (n=328) and 24h (n=269) is related to initial mortality introduction: adrenomedullin (am) is a peptide synthesized in vascular endothelial cells and cleared by the lungs. the use of am as an inflammatory biomarker and his predictive value has been studied in critically ill patients, but not yet in veno-venous extracorporeal membrane oxygenation (ecmo). the purpose of this study was to describe the plasmatic levels of am in patients supported with ecmo for acute respiratory failure methods: am (normal values <0.55 nmol/l) was measured at 5 time points: immediately before (t0), 24-h (t1) and 72-h after (t3) ecmo initiation and immediately before (t4) and 72-h (t5) after ecmo removal, in consecutive patients with severe respiratory failure supported with ecmo enrolled in the gatra study (nct03208270) at fondazione irccs ca' granda -policlinico of milan. data are reported as median (25 th -75 th percentile). statistical analysis was performed using logistic and random effects regression models (to account for repeated measurements within individuals) results: a total of 131 measurements were taken in 32 consecutive patients. am (nmol/l) decreased along the course of ecmo: t0=2.0 (1.5-6.4), t2=2.0 (1.5-6.4), t3=1.6 (1.1-3.1), t4=1.3 (0.8-2.0), t5=0.9 (0.6-2.1) (mean diff.= -0.65, 95%: ci -0.96, -0.35). am was lower in patients with viral compared to bacterial ards (mean diff.= -2.7, 95%ci -5.2, -0.2) (figure 1 ). am was higher in more severe patients (sofa>= 10, n=14) compared to less severe patients (sofa< 10, n=18): 5.7±4.8 vs 1.4±0.8 nmol/l, respectively p<0.001. basal values of am could not predict mortality at 28 days (or=0.8, 95%ci: 0.5-1.2) after conditioning for sofa score and respiratory failure etiology conclusions: am plasmatic values seem to be higher in more severe patients and in patients with bacterial ards. am decreased along the ecmo course but could not predict mortality in our group of patients fig. 1 (abstract p471) . plasmatic adrenomedullin during ecmo heparin binding protein (hbp) is released from activated neutrophils upon stimulation of b2 integrins. this pro-inflammatory effect generates the hypothesis that it can be a sepsis biomarker for patients admitted at the emergency department (ed) methods: the prompt study (clinicaltrials.gov nct03295825) took place at the ed of six greek hospitals. participants were admitted with suspected acute infection and at least one vital sign change. hbp was measured by an enzyme immunosorbent assay in plasma. sepsis was diagnosed by the sepsis-3 criteria. the primary study endpoint was the sensitivity for the diagnosis of sepsis. outcome prediction was the secondary endpoint. a total of 371 patients were enrolled; 166 had sepsis. the most common infections among patients without and with sepsis were upper respiratory tract infections in 30.2% and 1.2%; community-acquired pneumonia in 6.8% and 28.3%; and acute pyelonephritis in 9.3% and 28.3%. median hbp was 24.0 and 32.7 ng/ml respectively (p: 0.027). following analysis of the area under the curve (auc) it was found that the best discriminatory cut-off for sepsis was 19.8ng/ml. the comparative diagnostic performance of hbp versus qsofa score is shown in figure 1 . the odds ratio for sepsis with hbp above 19.80 ng/ml was 2.07 (p: 0.001). at the same cut-off point the sensitivity, specificity, positive predictive value (ppv) and negative predictive value (npv) for the prediction of early death after 72 hours was 100%, 35.7%, 4.1% and 100% respectively. hbp is more sensitive but less specific than qsofa for the diagnosis of sepsis in the ed. the rule-out prediction of early death seems the great merit. chronobiological and recurrence quantification analysis of temperature rhythmicity in critically ill patients introduction: rhythmicity and complexity of several circadian biomarkers, such as melatonin, cortisol and temperature have been found to be modified by critical illness. we examined the potential alterations of core body temperature (cbt) fluctuations and complexity in three groups (n=21): patients with septic shock upon icu admission (group a, n=10), patients who developed septic shock at icu hospitalization (group b, n= 6) and controls (group c, n=5). the hourly, average cbt was computed for 24 h upon icu admission and discharge in groups a and c, as well as during septic shock onset in group b. cosinor analysis of cbt curves was performed leading to the estimation of mesor (mean value), amplitude (the difference between peak and mean values) and acrophase (phase shift of maximum values in hours). complexity of cbt signals was evaluated with recurrence quantification analysis (rqa). no significant alterations in any circadian feature within groups were found, except for amplitude. controls exhibited increased entry cbt amplitude (0.45 ± 0.19) compared to groups a (0.28 ± 0.18, p < 0.05) and b (0.32 ± 0.13, p < 0.05). higher entry cbt amplitude in groups b and c was related with lower saps ii (r = -0.72 and -0.84, p < 0.003) and apache ii scores (r = -0.70 and -0.6, p < 0.003) respectively, reduced icu and hospital stay in group b (r = -0.62 and -0.64, p < 0.003) and entry sofa score in group c (r = -0.82, p < 0.003). recovery cbt time series appeared more periodic in relation with icu entry, for all groups. a more random cbt signals pattern upon results: among 23.011.601 individuals, 159.691 received inpatient treatment for sepsis. 41% had severe sepsis. 21% of sepsis and 28% of severe sepsis patients had an explicitly coded hai. the proportion of hai was higher in patients that received icu-treatment than in patients without icu-treatment (35% in icu/14% in non-icu sepsis, 37% in icu/17% in non-icu severe sepsis patients). tab. 1 shows the foci of explicitly coded hai. nosocomial pneumonia was the most common hai in all patient groups. clabsi occurred more frequently in icutreated patients; 21% were affected. cauti and c. diff infections were more common among non-icu-treated sepsis patients. more than one quarter of non-icu-treated sepsis patients had a c. diff infection. hai are common causes of sepsis and pose a significant healthcare burden. the proportion of patients affected and the distribution of foci differ between non-icu-and icu-treated sepsis patients with important implications for sepsis management within hospitals. impact of sepsis protocol triggered by ramathibodi early warning score (rews) in ipd sepsis on clinical outcomes s matupumanon 1 , y sutherasan 2 , d junhasawasdikul 2 , p theerawit 3 sepsis is now early identified and managed during triage in the emergency department. however, there is less focus on the effect of patients' management at the ward level. we aim to evaluate the impact of the implementation of the sepsis protocol on clinical outcomes in in-patients with new-onset sepsis. we conducted a prospective observational cohort study among adult medical patients admitted to the general wards in a university hospital. a 25-month pre-protocol period (august 2016 to august 2018) was assigned to a control group, and a 14-month protocol period (september 2018 to october 2019) was allocated to a protocol group. an in-patient sepsis protocol comprised nurse-initiated sepsis protocol by ramathibodi early warning score (rews)≥ 2 plus suspected infection, prompt antibiotic, lactate measurement, and fluid resuscitation was implemented. (table 1) . the implementation of in-hospital sepsis protocol was associated with significant improvement in patients' outcomes, namely lactate measurement, starting antibiotic within 1 hr, fluid management, and the shorter length of icu stay. icu routine nursing procedures interfere with cerebral hemodynamics in a prolonged porcine fecal peritonitis model sl liu 1 , dc casoni 2 , w z'graggen 3 , d bervini 3 , d berger 1 , sj jakob 1 routine nursing procedures (np) can interfere with blood pressure and cardiac output and may therefore alter cerebral hemodynamics in critical illness. this may be risk factor of sepsis-associated encephalopathy. methods: 20 sedated and mechanically ventilated pigs were randomized to fecal peritonitis or controls (n=10, each). after 8 hours of untreated peritonitis, the animals were resuscitated for 76 hours (resuscitation period). np [assessment of sedation (as), tracheal suctioning (ts), change in body position (cp), lung recruitment maneuver (rm)] were performed at baseline and 8h, 32h, 56h and 72h after start of rp. systemic and cerebral hemodynamics and o 2 saturations were recorded continuously. shock is the most common cause of death in the postsurgical icu, including septic shock and hypovolemic shock, reaching the 50-60% mortality in septic shock. the inadequate response of the immune system to the infection triggers a potent inflammatory cascade, where the c-reactive protein (crp) is an essential key in the amplification and maintenance of this cascade. the gene encoding to crp is located on the proximal long arm of human chromosome 1 (1q32). the gt polymorphism in the promoter sequence of crp gene (rs2794521) has been associated with invasive pneumococcal disease. thus, we analyze the relationship between rs2794521 polymorphism and the risk of developing septic shock in postsurgical patients. an observational, retrospective and single-center study was conducted on a sample of caucasian patients undergoing major abdominal surgery, of which one part developed septic shock and another part developed systemic inflammatory response syndrome, who were used as control. the rs2794521 polymorphism was analyzed by vasoactive medications are commonly used in sepsis treatment but may correlate with peripheral ischemia and the well-publicized complication of limb and digit loss. yet, the association between limb and digit threat and the intensity, duration, and pattern of vasopressor exposure are unknown. we studied adults (2010-2014) at 12 hospitals in an integrated health system who met criteria for sepsis-3. we identified the time to clinically apparent limb or digit threat using clinical adjudication among those with vasopressor-dependent sepsis (i.e. >1 hour of vasopressors at sepsis onset) who had a surgical evaluation within 28-days of sepsis onset. we defined daily vasopressor intensity as 0 to 4 vasopressors administered. then, we created a time-dependent model for threat with mortality as a competing risk with a weight function to estimates the varying contribution of vasopressors over time. we determined the subdistribution hazard (sh) ratio of threat for various patterns of vasopressor exposure and intensity, adjusted for age, baseline risk factors, and sequential organ failure assessment (sofa) score at sepsis onset. of 110,621 adults with sepsis, 13,147 (12%) were vasopressordependent (age, 66 [iqr, 56-77]; 7,040 [54%] males; max sofa score, 8 [sd 5] ). of these, 3,664 (28%) died and 117 (0.9%) had evaluations for limb or digit threat 4 [iqr, 1-8] days after sepsis onset. the model-based weight function showed the contribution of vasopressors to threat was stable over time ( fig 1a) . overall, a 1 unit increase in cumulative vasopressor exposure was associated with risk of threat (sh ratio, 2.60 [95%ci, 1.60-4.23], p<.001). for various patterns of vasopressor exposure, greater intensity associated with increased risk of threat ( fig 1b) . compared to constant exposure, an increasing and peak pattern associated with the greatest sh (fig 1c) . cumulative vasopressor exposure was associated with an increased risk-adjusted hazard of limb or digit threat following sepsis. fig. 1 (abstract p509) . relationship between vasopressor exposure and limb or digit threat following vasopressor-dependent sepsis. panel a demonstrates the estimated contribution of daily vasopressor intensity prior to surgical evaluation for limb or digit threat, with mortality as a competing risk. panel b and c explore the relationship between threat and both cumulative vasopressor exposure and the pattern of exposure following sepsis onset. (b) the maximum cumulative vasopressor exposure was associated with the highest risk of limb or digit threat (shr 17.5) when compared to reference exposure pattern (shr 1.0, reference). (c) increasing (shr 1.2) and peak (shr 1.2) patterns of cumulative exposure were associate with an increased sh of limb threat, while a decreasing pattern was associated with a lower risk (shr 0.8) when compared to constant intensity (shr 1.0, reference). abbreviations: shr: subdistribution hazard ratio proportion of encounters transitioning from phenotype at presentation within 24hrs, by arrival phenotype assignment and probability of membership. (c) tsne plots for α-type, ß-type, y-type, and ∂-type, with core (dark), marginal (light), and non-members (grey) in plots on the left and core, marginal, non members, and transitioning members (black) on the right fig. 1 (abstract p005). isolated microorganisms critical care references: 1. wertz et al. critical care explorations 1: e0058 the process investigators choosing wisely guidelines for the provision of intensive care services, version 2. ics structured patient handovers references: 1. care of the critically ill woman in childbirth the proqol manual: the professional quality of life scale:compassion satisfaction, burnout & compassion fatigue/secondary trauma scales references: 1. shimabukuro-vornhagen a et al. ca the code: professional standards of practice and behaviour for nurses, midwives and nursing associates p415 introduction: the aim of this study was to compare factors associated with the icu mortality for vap due to multidrug-resistant (mdr) klebsiella spp. in case of monobacterial (mo) vs polibacterial (po) origin. methods: retrospective data analysis of patients treated in icu with mdr klebsiella spp. strains as pathogens of vap during three year period was carried out. results: data of 71 patients were evaluated. mo vs po of mdr klebsiella spp. vap cases was found to be 35 (49.3 %) vs 36 (50.7%), p = 0.906. the icu mortality was 9/35 (25.7%) in mo, and 17/36 (47.2%) in po one, p = 0.060. statistical significant differences of survivors vs non-survivors in mo and po vap due to mdr klebsiella spp. were found in medians of neutrophilosis 78 p430 introduction: we study the population structure and resistome of mdr enterobacterales and pseudomonas aeruginosa isolates, c/t-susceptible or -resistant, recovered from low respiratory, intraabdominal and urinary tract infections of icu patients of 11 portuguese hospitals (step study results: in e. coli, two vim-2 producers were found (st131-b2-h30-o25:h4-ctx-m-15 and st88-c-h39-o22:h4) (c/t-mic=0.5/4-1/4 mg/l). a kpc-3-st5463-cladev-h160-o164:h56 (16/4 mg/l) was also detected. the most frequent esbl-e. coli clone was st131 cpr klebsiella pneumoniae (32 patients), candida spp. (21 patients). the comparison subgroup consisted of 217 patients with bacteremia caused by non-escape pathogens. we evaluated the days of mechanical ventilation, duration of antibiotic therapy (amt), icu length of stay (los), hospital los and mortality (table 1). results: mortality in patients with bacteremia caused by non-eskape pathogens was 23.5%, candida spp vancomycin mass removal over 120 minutes of hemoperfusion using ha330. bars refer to vancomycin mass (mg): blue (experiment 1) and red (experiment2) bars using blood while green (experiment 3) bar using balanced solution. yellow dashes are mean mass values of the three experiments (with standard deviations) and yellow line represents the reduction curve over time table 1 (abstract p438). results. * p-value versus non-eskape subgroup mechanical ventilation p453 translational value of the microbial profile in experimental sepsis studies sp tallósy 1 , a rutai 1 , l juhász 1 , mz poles 1 , k burián 2 , d érces 1 , a szabó 1 , m boros 1 invasive hemodynamic monitoring and blood gas analyses were performed on anesthetized animals between 18-24h of sepsis. the respiratory, cardiovascular, renal, hepatic and metabolic dysfunctions were evaluated with the species-specific sequential organ failure assessment (sssofa) score, the microbial profile was determined with selective media and maldi-tof ms in the initial inoculum and in the abdominal fluid taken 20h after sepsis induction. results: strong correlation was found between the initial dose of the inoculum (cfu) and the sssofa scores for organ dysfunction (rats: r = 0.656, p=0.0186; pigs: r=0.570, p = 0.0391) p466 introduction: pancreatic stone protein (psp) has shown promise as a biomarker of infection however, its diagnostic potential has not been systematically evaluated. we performed a systematic review and meta-analysis of available data on psp to evaluate its value for detecting infection in adults and determining a plasma or serum threshold value. methods: the pubmed and cochrane library database were searched for studies on psp in adult patients and their raw data were analyzed to estimate the best psp cut-off value that could detect infected patients using the youden's index. the cut-off sensitivity, specificity, positive predictive value (ppv) and negative predictive value (npv) were computed and compared to those for procalcitonin (pct) and c-reactive protein (crp). finally, we explored the potential value of a model combining all three biomarkers to detect infection. results: from a total of 44 potentially eligible published studies, 5 containing 631 patients were included in quantitative analysis. among them, 370 patients suffered from a clinically confirmed infection. the median appropriate statistical tests were used using spss 23. cd 64 was expressed as % age of neutrophils expressing positivity. results: sixty patients were analyzed. all parameters were compared between survivors and non survivors. demographics were comparable. most common source of sepsis was lungs and majority were admitted due to medical reason. non-survivors had significantly increased number of days with septic shock. at day 8 median values of all the biomarkers and the sofa score were significantly higher in the nonsurvivor group (p<0.05). there was a decreasing trend of all 3 biomarkers and sofa score amongst survivors. on multivariate logistic regression analysis, increased cd64 and crp levels between baseline and day 8, increased days with septic shock and increased sofa references: 1 introduction: we characterized the association of c-reactive protein (crp) with extracellular vesicles (evs) in plasma from sepsis patients and assessed a commercial crp adsorbent (pentrasorb, pentracor, hennigsdorf, germany) to deplete free and ev-associated crp. in addition, we characterized the potential pro-inflammatory effects of ev-bound crp on monocytes and endothelial cells monocytes and human umbilical vein endothelial cells (huvecs) were stimulated with isolated evs (20,000 g, 30 min) monocyte il-8 secretion was quantified by elisa; the activation of huvecs was assessed by their expression of icam-1 and e-selectin using confocal microscopy. results: septic plasma (n=30) contained 227.0±88.6 mg/l crp vs. 0.7±0.4 mg/ l for healthy controls (n=5). both, total evs and crp + evs were significantly elevated in septic plasma as incubation of septic plasma with pentrasorb resulted in depletion of free crp (247.2±72.6 mg/l before vs. 1.8±0.7 mg/l after adsorption) as well as in a significant reduction in crp evs from crp-depleted septic plasma induced significantly lower il-8 levels. huvec icam-1 or e-selectin expression, however, did not increase upon stimulation with septic evs. conclusions: treatment of septic plasma with pentrasorb efficiently removes free crp and detaches crp from the ev surface, resulting in reduced proinflammatory effects flow cytometry confirmed the association of monocytes with platelets and platelet-derived evs as well as the uptake of evs by monocytes. conclusions: storage of isolated monocytes induces a shift towards cd16 expressing proinflammatory monocytes, which seems to be mediated by residual platelets and platelet-derived evs. it remains to be clarified whether evs released from activated platelets can also trigger a shift towards proinflammatory, intermediate monocytes in vivo ethical approval was provided by ucl research ethics committee (5060/001). paired parametric analyses were performed and data displayed as mean +/-95% ci. results: plasma calprotectin concentration began to increase 1.5hours after endotoxin administration, was significantly higher than baseline by 2 hours (356.7ng/ml vs. 737ng/ml, p <0.01), peaked at 4 hours (mean 1373ng/ml, figure 1) and normalized by 24 hrs. calprotectin peaked earlier than comparator soluble mediators (procalcitonin 8hrs, crp, 24hrs) and exhibited 100% sensitivity; all participants demonstrating a minimum 2-fold increase from baseline (mean 3.84x). calprotectin displayed greater baseline variability (sd 147.9ng/ml) than either crp or procalcitonin. conclusions: our results indicate the potential of plasma calprotectin as a biomarker for bacterial infection. it increases earlier and peaks more rapidly than standard biomarkers. whilst higher baseline variability was observed p501 a multicenter randomized controlled study on landiolol for the treatment of sepsis-related tachyarrhythmia: subanalysis of the j-land 3s study o nishida kagoshima university graduate school of medical and dental sciences, department of emergency and intensive care medicine methods: we analyzed a retrospective cohort of electronic health records from adult sepsis patients at 12 upmc hospitals from 2010 to 2014. we defined sepsis-3 by i.) suspected infection (e.g., administration of antibiotics or body fluid culture) & ii.) organ dysfunction (e.g., 2 or more sofa points) in the first 6 hours of care. data were organized by hour and included vital signs, lab values, and treatments (e.g., total hourly iv fluids (ml) and norepinephrine equivalent dose). for each hour we describe, i.) available data elements, ii.) presence of sepsis-3, and iii by hour 6, most patients had vital signs (99%; n=70,559), basic labs (88%; n=62,719), fluid cultures (94%, n= 66,995), while serum lactate was completed in 24% (n=17,818) conclusions: early sepsis care patterns are variable. iv fluids were given during early hours, when uncertainty about sepsis was greatest, while vasopressors were administered after sepsis-3 elements were present. p504 effects of abdominal negative pressure treatment on splanchnic hemodynamics and liver and kidney function in a porcine fecal peritonitis model sl liu 1 department of intensive care medicine splanchnic hemodynamics and laboratory parameters were measured at baseline (bl, start of rp), and 24h, 48h and 72h after start of rp. two/three-way rm-anova or mixed-effects analysis, and student t tests were performed. results: npt in controls had no effect. after sepsis induction, mean arterial pressure (map) decreased by 15 (7-22) mmhg, cardiac output (co) by 1.3 (0.7-2.2) l/min, and arterial lactate increased by 0.2 (0.1-0.5) mmol/l. sepsis and resuscitation was associated with increasing hepatic and renal arterial flows (p≤0.002, both), and increasing prothrombin time npt in sepsis resulted in numerically less noradrenaline administration (0.3±0.3 ug/ min/kg in sepsis with npt vs. 0.8±0.7 ug/min/kg without npt, p= 0.310) and positive fluid balance (2.8±0.4 ml/h/kg with npt vs. 3.1± 0.4 ml/h/kg without, p=0.241). conclusions: in our experimental fecal peritonitis model, npt did neither impair splanchnic hemodynamics nor abdominal organ function. whether npt helps to reduce noradrenaline and volume administration in abdominal sepsis should be evaluated in further studies. p505 association between a c-reactive protein gene polymorphism (rs2794521) with the risk of develop septic shock in postsurgical patients of major abdominal surgery p martínez-paz 1 valladolid, spain; 2 hospital of medina del campo notably, the three groups received a comparable pro kg dose of acetaminophen. no difference was found between groups in term of toxic effects. patients carrying the cyp3a5p showed a more pronounced effect on body temperature in respect of wt and ugt1a1p °c respectively, but it does not reach statistical significance (fig.1b). only 50% of the patients reach a temperature <38°c at t2 and only 20% <37.5°c. conclusions: polymorphisms in enzymes involved in the metabolism of acetaminophen are relatively common. cyp3a5p seems to lead to higher peak plasmatic concentration and a slightly increased efficacy in fever control panel a: variations of acetaminophen plasmatic levels after 30 minutes (t1) and 2 hours (t2) after administration of an iv dose of 1g of paracetamol in wt patients and patients carrying mutation; panel b: body temperature variations in wt patients and patients carrying mutations clinical research, investigation, and systems modeling of acute illness (crisma) center, department of biostatistics we determined phenotype cohesiveness using probability of assignment at presentation, defining core members as ≥90% and marginal as <90% probability. we determined how members transitioned to other phenotypes over 24hrs using t-distributed stochastic neighbor embedding (tsne) plots and determined the odds (95%ci) of transition. results: we studied 37,198 adult sepsis encounters (median age 69 1c) the odds of ever transitioning from presenting phenotype increased significantly for marginal members vs publisher's note springer nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations we thank the department of education of the basque government (piba 2019-57) and the university of the basque country upv/ehu (ppg17/65, giu 17/32) for their financial support. a great disaster affects the family-and friend-performance of bcpr by diminishing the willingness of family and friend bystanders to follow the instruction provided by dispatchers. the experimental method ifitem could be an alternative of fibtem in cases when internal coagulation pathways assessment is prioritized (i.e. heparinized patients on extracorporeal supports). patients undergoing limitation of life-sustaining therapy had lower karnofsky scale scores. therefore, this scale may be useful to guide end-of-life decisions in the future, but further studies with larger number of patients are needed. readmission after discharge home from critical care: a qualitative study c robinson 1 , f nicolson 1 , p mactavish 1 , t quasim 2 , jm mcpeake 1 1 nhs greater glasgow and clyde, nhs greater glasgow and clyde, glasgow, united kingdom; 2 university of glasgow, nhs greater glasgow and clyde, glasgow, united kingdom critical care 2020, 24(suppl 1):p392 readmissions to acute care occur in a high number of critically ill patients within 90 days of hospital discharge [1] . biomedical drivers such as frailty and pre-existing co-morbidities have been identified as drivers for readmission. however at present there is limited data on the influence of social problems on readmission. this study, using a grounded theory approach, sought to understand from a patient/caregiver perspective what the drivers for readmission to acute care were. ethical approval was granted from the west of scotland research ethics service (19/ws/0091). a grounded theory approach was used to explore from a patient and caregiver perspective what the drivers for readmission are [2] . using a clinical database, we identified those patients who had an icu admission ≥ 3 days who were readmitted to acute care within 90 days of hospital discharge. the researcher attended the ward and after discussion with the direct care team conducted a semi-structured interview with patient and/or caregiver. the interview was recorded and transcribed verbatim. the transcripts were analysed to generate initial codes, followed by the development categories and sub-categories. theoretical sampling was undertaken. results: 15 participants were interviewed.10(66.6%) were patients and 5 (33.3%) were caregivers. the themes that have emerged from the data were: pain and polypharmacy; lack of social support and/or isolation; strained relationships with primary care providers and information provision across the patient journey. subsequent theory development is underway to understand how this learning could help reduce readmissions in future. in conclusion, both social and biomedical drivers are likely to contribute to acute care readmission in this group. future interventional work is required in order to identify modifiable factors to reduce this burden for patients and the healthcare service. frailty has shown to have prognostic relevance for patients with critical illness. since a wide range of tools has been described to screen for frailty, we aimed to describe the association of two frailty screening tools, the clinical frailty scale (cfs) score and the modified frailty index (mfi) in critically ill patients. we performed a post-hoc analysis of a multicenter cohort of patients admitted to six canadian intensive care units (icu) between february 2010 and july 2011. frailty was identified using the clinical frailty scale (cfs) and the modified frailty index (mfi). concordance of the frailty screening tools was evaluated with partial spearman rank correlation and intraclass correlation (icc). discrimination and predictive ability of the tools for hospital mortality, 1-year mortality, hospital readmission and adverse events were compared using concordance statistic (c-statistic) and calibration plot adjusting for age, sex, sequential organ failure assessment (sofa) score and icu admission source, respectively. the cohort included 421 patients. prevalence of frailty was 32.8% (95% confidence interval [ci] 28.3%-37.5%) with the cfs and 39.2% (95% ci 34.5%-44.0%) with the mfi. concordance between the two tools was low [(icc of 0.37; 95% ci 0.29-0.45) and partial correlation coefficient of 0.38 (95% ci 0.29-0.47)], even after adjustment. hospital and 1-year mortality were greater for frail compared to non-frail patients using of both tools. similarly, both tools found frail patients were less likely to be living independently after hospital discharge, and more likely to be rehospitalized when compared to non-frail patients. while the cfs and mfi show low concordance, both showed good discrimination and predictive validity for hospital mortality. both tools identify a subgroup of patients more likely to have worse clinical outcomes. the post-intensive care syndrome (pics) is a myriad of physical, psychiatric and cognitive disorders secondary to critical illness, leading to a decreased quality of life and an important socioeconomic burden. this study aimed to identify if the conformity to a pics prevention bundle was able to reduce the incidence of the syndrome at icu discharge. all patients admitted to the icu from january 1 st to december 31 st 2018 were included. the conformity to each of the ten components of the pics prevention bundle was assessed daily, and the patients were evaluated for anxiety, depression, cognitive dysfunction, muscular weakness, mobility impairment and nutritional risk at icu discharge and at a 3-to-6-months follow-up consultation. the patient cohort was divided in terciles according to bundle conformity for the analysis. results: from the 1145 enrolled patients, 352 (31%) were evaluated at icu discharge, and 103 (29%) attended to the follow-up consultation. there was no difference in baseline characteristics between the cohorts. there was no correlation between the prevalence of pics at discharge and bundle conformity during icu stay (90% vs. 87% vs 87%, p 0.834), though there was a decrease in nutritional risk and days in mechanical ventilation (table 1) . after 3 to 6 months there was a reduction on the prevalence of any kind of pics, mobility impairment, muscular weakness and nutritional risk. the patients that developed pics were older and had a higher simplified acute physiology score iii at icu admission. a higher adhesion to a pics prevention bundle was not able to prevent the occurrence of the syndrome. post intensive care syndrome (pics) is well recognized following general icu care [1] . intensive care syndrome:promoting independence and return to employment (ins:pire) is a multidisciplinary complex intervention designed to address pics [2] . with a paucity of evidence on pics after cardiothoracic intensive care, we aim to evaluate pics and the feasibility of the ins:pire intervention in this population. those attending the clinic received 5 weeks of intervention including individual appointments with icm nurse, physician, pharmacist, and physiotherapist. a café area facilitated peer support alongside psychology group sessions. primary outcome was quality of life measured by eq-5d-5l. further surveys included: pain, mental health, and selfefficacy. questionnaires were taken at baseline, 3 and 12 months. results: over 5 cohorts, 27 patients attended, 67% male, median age 66 years (iqr 61-75), median apache 2 score of 17 (iqr 14-18.5), and median icu length of stay was 13 days (iqr 9-21). a total of 14 (53%) patients completed surveys at one year. scheduled admissions represented 56% of those attending. mean euroqol eq-vas score was 70/ 100 (sd +/-18) at baseline increasing to 78/100 (sd +/-16) by 1 year (table 1) . those with problems in at least one domain of eq-5d-5l fell from 92% at baseline to 73% at 1-year with the breakdown shown in table 1. severe problems were seen in 22% falling to 18% at 1 year. hads demonstrated an anxiety or depression rate of 44%. brief pain inventory identified 14 patients (52%) with ongoing chronic pain. mean self-efficacy was 32/40 (sd +/-6) at baseline and 34/40 (sd +/-5) at 1 year. cardiothoracic intensive care patients have ongoing and persistent features of pics with significant effects on health-related quality of life. further, the ins:pire multi-professional complex intervention is feasible within this specialist group. screening approach might be implemented whenever screening of the total icu population is not deemed feasible. influenza is an acute viral illness with a significant financial burden. point of care testing for influenza is available and has demonstrated accuracy [1, 2] , the current gap in knowledge is the question around the opportunity cost of influenza testing. if poct is financially a less costly test this could free up scarce resource. the study adopts a cost minimisation approach. the point of care test is the roche cobas® liat® machine which can detect flu a/b and is compared with the west of scotland specialist virology centre's established in house multiplex real time pcr assay.the model was developed using microsoft excel and has 2 arms comparing analysis of the above mentioned tests. the model estimates that the total cost of poct per patient tested is £3926.33 compared with £4053.92 for lab testing ( figure 1 ). this is a saving of £127.60 per patient when poct is used. the result swings in favour of the lab test when poct specificity falls to 95.72%. if the lab could provide the result of influenza testing within 12 hours the result would swing in favour of lab testing. zanamivir which will potentially be used increasingly in the intensive care setting can more than double the difference between the 2 tests in favour of poct. this research suggests that poct offers potential cost savings in the icu setting. this is the case as long as poct specificity is higher than a threshold of 97.52% and the lab take longer that 12 hours to return the result. the sensitivity analysis should allow for external validity given the usual variations in icu practice. the aim of the present study is to describe the demographic, clinical, microbiological aspects and the outcome of patients with intensive care unit-related (icu-related) bacteremia. moreover, we aimed to study the patient outcome in association with colistin susceptibility. retrospective, single-center study in a 16-bed icu for 12 months, from 1/10/2018 to 30/9/2019. icu-related bacteremia was defined as bacteremia in patients with icu stay >48 hours or icu readmission (first admission ≥ 1 month before). only the first episode of bacteremia was considered. the primary outcome was 28-day mortality. data regarding clinical, demographic and outcome characteristics were retrieved from the patient files. the hospital's ethics committee approved the present protocol. moreover, the patients with bacteremia due to colistin-resistant pathogens were compared with the patients affected by colistin sensitive microbes. forty episodes of gram-negative icu bacteremia were collected during the aforementioned period in 40 patients (61.5% male) with a mean age and apache ii of 60.6±16.8 years and 18±8.1, respectively. the event had taken place at an average of 13.7 days. the responsible isolates were resistant to carbapenems in 82.5% of the episodes. the majority of the events were due to a single isolate (85%). acinetobacter baumannii and klebsiella pneumoniae presented the majority of the implicated microbes (35% and 37.5%, respectively). the crude 28-day mortality was 30%. finally, we could not detect any difference in mortality between the colistin sensitive and the colistin-resistant pathogens ( figure 1 ). the present study denotes that, in a setting of extremely drugresistant pathogens with limited treatment options, gram-negative bacteremia in the icu is associated with increased mortality. image 1 : characterization of resistance mechanisms affecting ceftolozane/ tazobactam in enterobacterales and pseudomonas aeruginosa icu isolates using whole genome sequencing (step study) m hernández-garcia 1 , cc chaves 2 , jm melo-cristino 3 , ds silva 4 , ar vieira 5 , mp f. pinto 6 , jd diogo 7 , eg gonçalves 8 , jr romano 9 , rc cantón 1 1 hospital ramón y cajal-irycis, microbiology department, madrid, spain; 2 introduction: clostridium difficile infection (cdi) is the main cause of hospital acquired diarrhoea [1] . the aim of this study was to compare characteristics of cdi during yr 2011 and 2015. a retrospective observational study was carried out in lithuanian university of health sciences hospital -the largest teaching facility of tertiary care in country. according to department of infection control records, patients (pt) with (w.) diarrhoea and the first positive stool test for c.difficile toxin a/b were included. age, charlson comorbidity index (cci) score, profile of hospital department (medical (md), surgical or icu) where cdi was diagnosed, type of cdi (healthcare-associated (ha), hospital or community-acquired) and rate of risk factors (rf) have been estimated in both 2011 and 2015. ibm spss 23.0; pearson's chi-square, fisher's exact tests were used for statistics. p < 0.05 was statistically significant. results: in total 7 pt from 2011, 72 from 2015 were enrolled. in 2011 n=4 (57%) pt were ≥65 yr old, in 2015 -n=45 (63%), (p=0.045). in 2011 cci>5 was estimated in n=6 (86%) pt in comparison of n=46 (64%) in 2015, (p=0.025). in 2011 n=0 (0%) of cdi cases were ha, in 2015 -n=12 (17%), (p=0.01). in 2011 n=5 (71%) of cdi were diagnosed in md in comparison of n=60 (83%) in 2015, (p=0.01). in 2011 12 weeks prior to cdi n=5 (71%) pt have been admitted to hospitals, n=7 (100%) have been treated w. antibiotics, n=4 (50%) -w. ppis, n=5 (36%) -w. h2 antagonists, n=3 (43%) -w. immunosupressants in comparison of n=51 (71%), n=69 (96%), n=36 (57%), n=26 (71%) and n=21 (29%) in 2015, respectively, (p>0.05). overall rate of cdi cases among in-hospital patients increased tenfold by yr 2011 and 2015. in 2015, more elderly patients had cdi and severe comorbidities were less frequent in comparison with 2011. in 2015, more cases of cdi were hospital-acquired and have occured in medical departments. rate of risk factors of cdi remained unchanged.these results indicate a possible relationship between ttv dna count and immunological alteration. the ttv quantitative determination could be useful as a proinflammatory marker in sepsis, with some benefits: low cost, easy determination and good correlation with immune system functionalit. it will be necessary to perform a larger study to check our hypothesis and to establish a ttv level threshold that may allow to anticípate the disease prognosis. introduction: acute kidney injury (aki) is a serious complication in sepsis and associated with high morbidity and mortality. the combination antimicrobial regimens with vancomycin (vcm) and broad-spectrum betalactams (bsbl), such as piperacillin tazobactam and cefepime, have been identified as potentially nephrotoxic combinations, but existing studies have not provided sufficient evidence. the aim of this study was to evaluate detailed association between the combination antimicrobial therapy and the risk of aki in septic patients. this investigation was a post hoc analysis of 2 prospective nationwide cohorts enrolling consecutive adult patients with sepsis in intensive care units in japan. in this study, progression of aki was defined as one or more elevation of renal sub-score in sequential organ failure assessment score from day 1 to day 4. we regarded anti-pseudomonal penicillins, fourth generation cephalosporines, and carbapenems as bsbl. multivariable logistic regression analysis including a two-way interaction term (vcm x bsbl) was performed to assess the add-on effects of each antimicrobial agent on the progression of aki. the final study cohort comprised 1837 patients with sepsis. among them, 45 received vcm without bsbl, 1055 received bsbl without vcm, 249 received both vcm and bsbl, and 488 received other type of antimicrobials. the administration of vcm was associated with an increased risk of aki in patients with bsbl [odds ratio (or), 1.57 (0.96-2.57); p=0.072]. however, the tendency was not evident in patients without bsbl [or, 0.23 (0.03-1.56); p=0.133]. the interaction effect on the progression of aki between vcm and bsbl were statistically significant (p for interaction=0.038). the regression model including two-way interaction term suggested that the combination of vcm and bsbl might synergistically increase the risk of aki in patients with sepsis. increasing resistance to carbapenems due to carbapenemase productionone of main actual problems of antibacterial resistance in burn icu. production of several types of carbapenemases (kpc, ndm and oxa-48) is common in k. pneumoniae strains. carbapemenase production is a marker of extreme antibacterial resistance. the aim of our study was to investigate the epidemiology of nosocomial infections caused by producing kpc, ndm and oxa-48 k. pneumonia strains in burn icu. total of 26 patients with nosocomial infections caused by 26 carbapenem resistance strains of k. pneumoniae were included in the study, from whom 3 had lower respiratory tract infection, 23 had skin and skin structure infection. initial identification of isolates was performed in laboratory by automatic microbiological analyzer. for all of k. pneumoniae isolates presence of bla ndm , bla oxa-48 and bla kpcgenes were examined by pcr method. baseline characteristics of patients: me (iqr) of age -50 (39; 64) years, me (iqr) of tbsa -40 (29; 52) percent, me (iqr) of icu los -30 (21; 35) days. inhalation injury was diagnosed in 10 (38.4%) patients. total of 11 patients died, mortality rate was 42.3%. all patients were diagnosed with nosocomial infection caused by k. pneumoniae. from 26 k. pneumonia strains 1 (3.8%) were found to be producing kpc, 3 (11.5%)producing ndm and 20 (76.9%) -producing oxa48. only 5 (19.2%) carbapenem resistance k. pneumoniae isolates were not producing carbapenemases. from 20 patients infected by oxa48 producing k. pneumoniae 20 patients died, mortality rate was 40%. from 23 patients infected by oxa48 or ndm producing k. pneumoniae 10 patients died, mortality rate was 43.5%. from 5 patients infected by non-carbapenemase producing k. pneumonia no one died. carbapenemase producing strains are widely spread among carbapenem resistance strains of k. pneumoniae in burn icu. mortality of patients infected by producing oxa48 or ndm k. pneumoniae strains reaches 43.5%. the rationale for blood purification as adjunctive therapy during sepsis involved the capacity in removing endogenous and exogenous toxins, but currently no recommendations exists [1] . a critical point may be the potential interaction with antimicrobial therapy, which remains the mainstay of sepsis treatment. the aim of our study was to investigate the vancomycin (van) removal during blood purification using an in vitro model of hemoperfusion (hp) with ha330 cartridge (jafron, zhuhai city, china), most widely used in china and actually available in europe. this is an experimental study. three independent experiments were performed: we injected 250 mg of van in 500ml of whole blood from healthy donors (experiment 1 and 2) or in 500ml of balanced solution (experiment 3) in order to assess membrane saturation. a closed-circuit (blood flow of 250ml/min) simulating hp ran using ha330. samples were collected from arterial line at 0, 5, 10, 15, 30, 45, 60, 90, 120 minutes; van plasma concentrations were measured and removal was evaluated using mass balance analysis. differences in mass removal was assessed using kruskal-wallis test. results: figure 1 shows van mass at each timepoints. we observed no difference between in blood and in balanced solution experiments (p the aim of this study is to determine if routine bbv testing in the icu contributes to the discovery of undiagnosed bbv infections. icu patients may require renal replacement therapy (rrt). sharing rrt equipment carries a risk of bbv transmission, which mainly relates to hepatitis b (hbv), hepatitis c (hcv) and hiv. since 2012, all glasgow royal infirmary icu patients undergo routine bbv screening, with rrt machines allocated for patients with specific bbv statuses. routine bbv testing is beneficial to both the individual and society. hcv is a pertinent health issue in scotland. the scottish government aims to eliminate hcv by 2030 and is researching innovative and costeffective methods to identify undiagnosed infections. this single-centre retrospective observational study examined prospectively collected clinical data from 1069 icu admissions. proportions were compared using a two-proportion z-test and a logistic regression model was carried out to determine if deprivation quintile was independently associated with the seroprevalence of bbvs. the bbv seroprevalence in the cohort studied: 0.45% (hbv), 11.7% (hcv), 0.91% (hiv). the seroprevalence of hbv in the cohort studied was similar to that of scotland (p=0.11), but the seroprevalence of hcv (p<0.001) and hiv (p=0.01) were statistically significantly higher than that of scotland. due to the small number of reactive test results for hbv and hiv, the relationship between deprivation and bbv seroprevalence was explored for hcv only. the only independent variable associated with a reactive anti-hcv test result was "current or previous illicit drug use" (adjusted odds ratio of 40.2; 95% confidence interval of 21.1-76.4; p<0.001). this study shows that routine bbv testing in the icu is useful in discovering new bbv infections. this is the first observational study focusing on the value of routine bbv testing in an icu setting to our knowledge. continuous infusion vancomycin protocol is a safe, acceptable and effective alternative to intermittent dosing of vancomycin in critical care. ceftaroline is an efficacious treatment in patients with severe cap, admitted in icu. it relates to earlier resolution of respiratory failure and less rescue antibiotics. we need an adequately pragmatic trial to confirm our findings organ dysfunction in scrub typhus, incidence and risk factor a sarkar 1 , a guha 2 , r dey 3 [1, 2, 3, 4, 5] . its preads by bite of larval stageof thromboculid mites or chigger [1] . clinical features may include fever, headache, myalgia, lymphadenopathy, eschar, skinrash. it may also cause pneumonia, renal failure, shock, meningoencephalitis, multiple organ failure [1, 2] . our study aims to discuss the incidence of organ dysfunction in a comprehensive way taking the overall population of patients with identified scrub typhus infection. there is lack of data in eastern india regarding the incidence and risk factors of developing multiorgan dysfunction syndrome (mods) in scrub typhus. in this retrospective study we studied the incidence of various organ involvement and the risk factors associated with the development of mods in scrub typhus. we collected data from december 2016 to november 2019 in tertiary care hospital at kolkata. we have included all patients who are having fever, scrub typhus igm antibody positive, age more than 14 years. sofa score was used in evaluating patients with mods. exclusion criteria involves patient who are having coinfectional ong with scrub typhus. in a cohort (n=114), patients with multiorgan dysfunction syndrome was seen in 27 patients (23.68%), the mean age in group of patients with mods was 50.0+/-14.96 years (mean+/-sd). in group of patients with mods, fever duration in days was of 11+/-3.58 days (mean+/-sd), interval from treatment to defervescenc in days was 5.11+/-2.39 days (mean +/-sd). among patients with mods, hematologic involvement was seen in 7 patients (25.92%), hepatic involvement was seen in 19 patients (70.37%), renal involvement was seen in 17 patients (62.96%), neurologic involvement was seen in 24 patients (88%), respiratory involvement was seen in 14 patients (51.85%), cardiovascular was seen in 8 patients (29.63%), icu shifting was necessary in 20 patients (74.07%), mechanical intubation was needed in 14 patients (51.85%) in multiorgan dysfunction syndrome patients. hospital mortality in patients with mods was 3 patients (11.11%). no mortality was seen in patients without mods. other parameters were evaluated among patients with mods. they include eschar in 1 patient (3.7%), seizure in 7 patients (25.93%), hepatoslenomegaly in 26 patients (96.3%), leucopenia in 3 patients (11.11%), leucocytosis in 13 patients (48.14%), thromnbocytopenia in 7 patients (25.92%),decreased hemoglobin in 22 patients (81.48%), transaminitis in 19 patients (70.37%). the risk factors associated with the development of mods are platelet counts, bilirubin, transaminitis, glasgow coma scale, time interval from treatment to defervescence, hemoglobin, total leucocyte count and fever duration. scrub typhus is an important cause of acute febrile illness in this part of the country and is frequently associated with organ dysfunction. however, the overall mortality is low which is similar to other studies done before [2] . score at baseline were significant (p<0.05) predictors of mortality.highest area under the roc curve was obtained for number of days with septic shock (0.857) followed by increased cd64 between baseline and day 8 (0.798). though serial pct levels significantly increased amongst non-survivors, it did not predict mortality. serial level of biomarkers in icu patients may predict mortality. larger trials are needed to confirm the results. plasma strem-1 levels were retrospectively measured at day 1-2, 3-4 and 6-8 in 116 septic shock patients from the immunosepsis cohort (nct02803346), included between 01/2016 and 12/2018, using a validated elisa method. the associations between strem-1, mhla-dr, 28-day survival status, and occurrence of icu-acquired nosocomial infection (ni) were assessed. neither strem-1 nor mhla-dr levels at d1/2 were associated with the occurrence of icu-acquired ni. however, 28-day mortality was significantly higher in patients with d1-2 strem-1 value superior to the median (39.6% vs 11.3%, p=0.0103; median=539 pg/ml). a significant inverse correlation was found between mhla-dr at d6-8 and strem-1 at d1-2 (sp -0.378, p<0.0001) and at d6-8 (sp -0.382, p<0.0001). at d6-8, when stratifying patients based on strem-1 (400pg/ml) and mhla-dr (5000 ab/c), patients combining elevated strem-1 and low mhla-dr presented with significantly higher 28day mortality (47.6% vs 8.7%, p = 0.0003, chi-squared test) and ni incidence (31.8 vs 12%, p=0.044) compared with patients with low strem-1 / high mhla-dr. this study shows for the first time that trem-1 pathway activation is associated with septic shock-induced immunosuppression, as shown by an inverse correlation between strem-1 at baseline and mhla-dr expression at d6-8. persisting high strem-1 values and low mhla-dr expression in septic shock patients are significantly associated with higher rate of icu-acquired infection and mortality. introduction: sepsis mortality remains high [1] . the surviving sepsis campaign (ssc) recommends to guide resuscitation on normalization of lactate levels [2] , however this is debated [3] . we have shown that plasma levels of bio-adrenomedullin (bio-adm) were associated with patient outcome during sepsis [4] . we therefore aimed to evaluate the added value of bio-adm to lactate measurement in the adrenoss cohort. this is a post-hoc analysis of the adrenomedullin and outcome in severe sepsis and septic shock (adrenoss) cohort study. the adre-noss study is a prospective observational study conducted in twenty-four centers and included 583 septic patients [4] . we studied the relationship between the association of initial evolution of lactate plasma levels and bio-adm level at 24h and outcome in patients for whom both markers were available at admission and one day later ("24h"). bio-adm levels below 70 pg/ml were considered as low, and high if greater than 70 pg/ml [4] . in patients with high lactate levels (>2 mmol/l) at admission (n=328), lactate normalization (<2 mmol/l) at 24h was associated with better outcome than in patients with persistently high lactate at 24h (28day mortality 15.9% vs 41.9% respectively, hr 3.3 [2.0-5.3], p<0.001) ( figure 1 ). among patients with decreasing lactate, high and low bio-adm levels at 24h identified patients with different outcomes (28day mortality 7% vs 26% for low vs high bio-adm respectively, hr 4.4 [1.6-11.7], p<0.005). high and low bio-adm levels at 24h also differentiated outcome of patients with persistently elevated lactate (hr 4.5 [1.6-12.3], p<0.005). in patients with low initial lactate, neither lactate or bio-adm had no added prognostic. our data suggest that measurement of bio-adm in addition to lactate may help physicians to refine risk stratification and therefore to guide resuscitation during sepsis. the effect of fluid replacement in sepsis, severe sepsis and septic shock in first 24 hrs in clot quality and microstructure s pillai 1 , g davies 2 the inflammatory response in sepsis can lead to a spectrum of coagulation system defects [1] . sepsis and severe sepsis is associated with a hypercoagulable state where the clot microstructure is known to be a tight and highly elastic clot, which is potentially resistant to fibrinolysis ( figure 1 ). conversely, septic shock is associated with a hypocoagulable state where the clot microstructure is loose and structurally weak. the study aim to investigate the effect of fluid resuscitation and replacement in clot microstructure over 24 hours. methods: 100 patients (50 sepsis, 20 severe sepsis and 30 septic shock) were included in the study. all these patients received standard fluid replacement therapy with crystalloids. blood samples were collected at 0 hours, 4 hours and 24 hours. clot microstructure, standard markers of coagulation and inflammatory markers were measured. in sepsis group following fluid administration, the d f reduced initially and then remained stable (1.78-0 hours, 1.74-4 hours, 1.73-24 hours, normal d f range 1.73 ± 0.04). in severe sepsis group, the d f reduced initially, then increased (1.80-0 hours, 1.71-4 hours, 1.76-24 hours) and in septic shock, the df was very low to start with and there were only slight increase with fluid administration (1.66-0 hours, 1.68-4 hours, 1.67-24 hours). the hypercoagulable state and clot quality in both sepsis and severe sepsis group improved with fluid resuscitation, however despite an early improvement in clot quality, ongoing fluid resuscitation resulted in markedly reduced functional clot with very low clot strength and functionality. this study demonstrates that d f as a marker of clot quality and function may have potential in fluid and component replacement in critical illness and injury. this study analyses the prognostic ability of white blood cell count (wbc), neutrophil:lymphocyte ratio (nlr) and c-reactive protein (crp). hypo-and hyperimmune responses have been associated with increased mortality from septic shock [1] . patients with septic shock (sepsis 3.0) admitted to queen elizabeth hospital birmingham, between december 2017 and july 2019 were included. the primary outcome was 90-day mortality. data was tested for normality and presented as median (iqr) and analysed using a mann whitney u test. categorical data was presented as % and analysed using a chi-squared test. a p value of < 0.05 was used to determine significance. a multivariate binary logistic regression analysis was conducted using age, apache ii, charlson comorbidity index, performance status, and initial lactate as covariates. a hosmer lemeshow test of >0.05 indicated good fit. results: 474 patients were admitted with septic shock. the majority (61%) were male, with a median age of 64 (55-75) and a 90-day mortality of 37%. on day 1, wbc was lower in patients who died compared to patients who survived (9 [7] [8] [9] [10] [11] [12] [13] [14] [15] patients who died of septic shock had a lower wbc, nlr and crp response early on compared to survivors. this may represent early immunoparesis that allows infection to propagate unchecked. however, this was not independently associated with mortality when confounding factors were accounted for. a specific metabolite of mitochondriaitaconic acid is formed upon proinflammatory activation. the attempts of various researches to find the itaconic acid in peripherical blood of patients with sepsis were unsuccessful [1] . some phenylcarboxylic acids (phcas) are known to be microbial metabolites and sepsis biomarkers; they also affect the mitochondrial functions [2] . concentrations of phcas (phenyllactic, p-hydroxyphenylacetic, phydroxyphenyllactic acids) and mitochondrial metabolites (succinic, itaconic acids) in 48 serum samples from 8 patients on the 1 st day of diagnosis of sepsis and 35 serum samples from 22 patients with late stages of sepsis (sepsis-3) were measured by gas chromatographymass spectrometry; control group -20 donors. results: itaconic acid was found in low concentrations (0.5-2.3 μm) only at early stage of sepsis. the multiple increase in levels of phcas and mitochondrial metabolites were detected in patients with late stage of sepsis in comparison with early stage and donors, p<0.001. increased succinic acid (up to 100-1000 μm) concentration is the result of succinate dehydrogenase inhibition by microbial metabolism intermediates (phcas), which was confirmed by in vitro experiments in isolated mitochondria (fig.1) . itaconic acid may be a promising marker in early stage of sepsis, which needs to be proved. prediction of severe events in clinical sepsis is challenging. for such prediction we aimed to compare the novel biomarker calprotectin in plasma, with routine biomarkers. in a prospective study, blood samples were collected from consecutive patients who triggered the sepsis alert in the emergency department in our hospital. c-reactive protein (crp), procalcitonin, neutrophils, and lymphocytes were analysed according to routine practice. p-calprotectin was analysed using a specific particle enhanced turbidimetric assay (gentian diagnostics as). the composite endpoint, which was termed severe event, was defined as death or admission to the intensive care unit (icu)/high dependency unit (hdu) within 48 hours from arrival. the study included 367 patients with written informed consent, of whom 335 were considered to have infection (defined as obtained blood culture and subsequent antibiotic therapy for at least 4 days or until discharge or death), and 32 had no infection. seventy-four patients (22%) with infection developed a severe event. mean pcalprotectin was 2.99 mg/l (standard deviation (sd) 2.10) among patients with infection and 2.35 mg/l (sd 2.64) among patients without infection (p=0.02). in patients with infection mean p-calprotectin was 3.81 mg/l (sd 3.18) among those with and 2.75 mg/l (sd 2.50) among those without a severe event (p=0.006). analysis of area under the receiver-operating characteristic (roc) curve for prediction of severe events showed superiority for p-calprotectin compared with procalcitonin and neutrophil-lymphocyte-ratio, both regarding all sepsis alert cases and regarding the patients with infection (p< 0.05 for all comparisons), fig 1. in addition, there was a trend toward superior performance compared to crp (p=0.10 and 0.15). in sepsis alert patients, p-calprotectin was elevated in those who subsequently developed severe events. p-calprotectin was superior to traditional biomarkers for prediction of severe events. introduction: rapid diagnosis of acute infections and sepsis is critical in emergency departments (eds). current tests have slow turnaround times, low sensitivities, and/or signals from contaminant or commensal organisms. empirical antimicrobial treatment may result in severe adverse events and contributes to antimicrobial resistance. diagnostics to distinguish bacterial from viral infections and noninfectious etiologies support clinicians in efforts toward antimicrobial stewardship. in a prospective, non-interventional study in the eds of 6 sites in greece (prompt study nct03295825), we evaluated hostdx sepsis, a host response test for suspected acute infections and suspected sepsis. hostdx sepsis measures 29 human mrna targets and employs advanced machine learning to differentiate patients with bacterial and viral infections, and noninfectious etiologies. adult patients presenting with suspected acute infection and at least one vital sign change were enrolled. whole blood rna was quantified using nano-string ncounter. predicted probabilities of bacterial and viral infection were calculated (bvn-1 algorithm). patients were adjudicated in a retrospective chart review by 3 independent infectious disease specialists blinded to hostdx sepsis results. among 396 patients adjudicated as bacterial (56), viral (45), noninfected (1), or indeterminate (294) the area under the receiver operating characteristics (auroc) of hostdx sepsis for predicting bacterial vs. viral/non-infected patients was 0.92, and auroc for viral vs. bacterial/non-infected patients was 0.87 (fig.1) . our results indicate that hostdx sepsis distinguishes bacterial from viral infections and other etiologies with high accuracy. hostdx sepsis is currently developed as a rapid point-of-care device with a turnaround-time of less than 30 minutes. hostdx sepsis may therefore assist ed doctors in making appropriate treatment decisions earlier, towards the ultimate goal of antimicrobial stewardship. we studied the diagnostic value of a leukocyte deformability assay that rapidly quantifies the immune activation signatures of sepsis in an undifferentiated population of adults presenting to the ed. ed clinicians must balance the benefits of early intervention against the risks of indiscriminate use of resource-intensive interventions. there are no currently available rapid diagnostics with acceptable performance to achieve this balance. we prospectively enrolled adult patients within 5 hours of presentation with signs of suspicion of infection in two eds in the usa. edta-anticoagulated blood was drawn and analyzed using deformability cytometry [1] . procalcitonin (pct) levels were also measured. patients were retrospectively adjudicated for sepsis-3 by physician committee using the entire medical record. diagnostic performance characteristics and receiver operating curves were used to examine the diagnostic performance of the assay as well as pct. of the 190 patients enrolled, 17.4% were adjudicated as septic. the leukocyte deformability assay demonstrated 91% sensitivity, 68% specificity, and 97% negative predictive value for a single cutoff. the auc was 0.86 ( figure 1 ). pct with a cutoff of 0.5 ng/ml had 55% sensitivity, 87% specificity, and 90% negative predictive value. the auc for pct (as continuous variable) was 0.8. the leukocyte deformability assay of immune activation signatures demonstrated superior diagnostic performance for sepsis when compared to pct. the assay's diagnostic performance and rapid turnaround time of 5 minutes may positively impact patient outcomes while minimizing indiscriminate use of valuable resources in the ed. it is already known in literature that high levels of midregional proadrenomedullin (mrproadm) are related with organ disfunction in infections despite of source and pathogens [1] . similarly, microcirculatory impairment has been reported in sepsis. we examine the correlation between microcirculatory disfunction and mrproadm as a sign of early organ failure. we included 20 consecutive adult patients with suspected infection, sepsis or septic shock admitted to our intensive care unit (icu) as first hospital admission with an expected icu stay of > 24hours. mrproadm was measured daily during the first five consecutive days and sublingual microcirculation was assessed with incident dark field (idf) technology at t1, t2, and t5. we collected information on saps ii, apache scores, and sofa score for each timepoint. results: ten patients had septic shock, 5 sepsis and 5 infection. three patients died during icu stay. a mrproadm clearance of 20% or more between t1 and t2 was found associated with the improvement of mfi (mann-whitney u test, median increase 12.35% versus 2.23%, p= 0.005) (figure 1) . a mrproadm >1.42 nmol/l at the icu admission was associated with a worse sofa score at all the timepoint. moreover, mrproadm levels at admission was found significantly related with icu mortality (auc 0.941 [0.819-1]; p=0.017). mrproadm shown no relation with absolute value of mfi. the study shows a good correlation between the clearance of the biomarker and the improvement in mfi. moreover, our results support previous findings on the prognostic value of mrproadm in terms of sofa and icu-mortality. clinical performance of a rapid sepsis test on a near-patient molecular testing platform r brandon 1 , j kirk 2 , t yager 2 , s cermelli 2 , r davis 2 , d sampson 2 , p sillekens 3 , i keuleers 3 , t vanhoey 3 1 immunexpress, seattle, united states; 2 immunexpress, immunexpress, seattle, united states; 3 biocartis nv, biocartis, mechelen, belgium critical care 2020, 24(suppl 1):p481 the purpose of this study was to clinically validate a new, rapid version of the septicyte™ assay on a near-patient testing platform (biocartis idylla™). septicyte™ lab is the first-in-class sepsis diagnostic to gain fda-clearance but has a complex workflow and a turnaround time (tat) of~6 hours. the assay in idylla™ cartridge format is called septicyte™ rapid. septicyte™ lab was translated to the biocartis idylla™ near-patient testing platform and analytically validated. for this study, 0.9ml of peripheral blood paxgene tm solution from previously collected patient samples was pipetted directly into the cartridge and inserted into the idylla™ reader. patients were part of an independent cohort (n=200) from intensive care units located in the usa and europe. septicyte™ rapid results were reported as a septiscore™ between 0 and 10 with higher scores representing higher probability of sepsis. assay performance determined included technician hands-on-time (hot), assay tat, failure rates, and area under roc curve based on comparison to retrospective physician diagnosis. average hot was 2 minutes, and average tat was 65 minutes. clinical samples could be processed immediately with septicyte™ rapid and did not require 2 hour pre-incubation of paxgene blood, greatly improving tat. correlation of septiscore™ values between lab and rapid, based upon a subset of samples run on both platforms, was very high (r 2 >0.97). estimated roc auc performance for discriminating sepsis from non-infectious systemic inflammation (nisi/sirs) was similar to that previously reported for septicyte™ lab. this is the first demonstration of a validated, fully-integrated, rapid, reproducible, near-patient, immune-response sepsis diagnostic, providing actionable results~1 hr, to differentiate sepsis from non-infectious systemic inflammation / sirs. accuracy of septicyte™ for diagnosis of sepsis across a broad range of patients r brandon 1 , k navalkar 2 , d sampson 2 , r davis 2 , t yager 2 1 immunexpress, seattle, united states; 2 immunexpress, immunexpress, seattle, united states critical care 2020, 24(suppl 1):p482 the purpose of the study was to demonstrate sepsis diagnostic performance of the biomarkers of septicyte™ in subjects other than critically ill adults, and in hospital locations other than icu. septicyte™ lab was the first immune-response sepsis diagnostic assay to gain fda-clearance (k163260) and, as part of gaining this clearance, clinical validation was performed on adult patients admitted to intensive care (icu) only [1] . we therefore performed an in silico analysis across a broad range of patients using the septicyte™ host immune response biomarkers and algorithm. peripheral blood gene expression data, including public and private datasets, were chosen based on quality, annotation, and clinical context for the intended use of septicyte™. multiple comparisons were performed within datasets to better understand the diagnostic performance in certain cohorts including healthy subjects. diagnostic performance was determined using area under curve (auc). results: table 1 shows some characteristics of the selected datasets and patients, including number of datasets (n=22) and comparisons (n=55), number of cases (n=2234) and controls (n=2089) used in comparisons, patient category and hospital location. septicyte™ aucs for the three groups of adults, adult / pediatric and pediatric / neonates were 0.88, 0.85, and 0.87 respectively, which is similar to that previously reported (0.82 -0.89) [1] . these results suggest that the septicyte™ signature has diagnostic utility beyond adults suspected of sepsis and admitted to icu. this signature has now been translated to the near-patient testing platform biocartis idylla™ (as septicyte™ rapid) which promises rapid (~1 hour) diagnosis of sepsis in a broad patient population following further validation. introduction: especially extracorporeal cardio pulmonary bypass (cpb) is known to induce severe inflammation. postoperative inflammation is associated with a sepsis like syndrome including endothelial barrier disruption, volume depletion and hypotension. sphingosine-1-phosphate (s1p) is a signaling lipid regulating permeability and vascular tone. in septic humans decreased serum-s1p levels could be identified as marker for sepsis severity. we addressed three main issues: (1) are serum-s1p levels affected by cardiac surgery? (2) are potential alterations of serum-s1p levels related to changes of acute-phase proteins, s1p sources or carrier? (3) is the invasiveness of the surgery a factor that may influence serum-s1p levels? methods: 46 elective major cardiac surgery patients were prospectively enrolled in this study. serum samples were drawn pre-, post-procedure and on day 1 and day 4 after surgery. we analyzed s1pand its potential sources: red blood cells (rbc) and platelets. we further quantified levels of other inflammatory markers and documented other clinical parameters. median serum-s1p levels in all patients before the procedure were 0.77 (iqr 0.61-0.99) nmol/ml. serum-s1p levels decrease after surgery, whereas all other inflammatory markers increase. serum-s1p levels dropped by 58% in the on-pump and 31% in the off-pump group. changes of serum-s1p levels are associated with s1p sources and carriers: albumin, hdl and vwf:ag activity. patients with a full recovery of their serum-s1p levels after surgery compared to their individual baseline presented with a lower sofa score (p>0.05) and shorter icu stay (p<0.05). serum-s1p levels are disrupted by open heart surgery and levels might be negatively affected by endothelial injury or loss of s1p sources. low serum-s1p levels may contribute to prolonged icu stay and worse clinical status. future studies may investigate the beneficial effects of s1p administration during cardiac surgery. the aim of study is to measure and correlate the expression of ncd64, mhla-dr, pct (procalcitonin) and qcrp (quantitative creactive protein) to predict development of sepsis and its outcome. in this tertiary centre based longitudinal cohort study, a total 110 patients were enrolled in whom sepsis was suspected on the basis of clinical diagnosis and supported by lab investigations. they were divided into two groups sepsis/case and non-sepsis/control. disease severity in icu was assessed by sequential organ failure score (sofa). blood samples for routine lab investigations and biomarkers were taken at the time of admission in icu before administration of first dose of antibiotics at time d 0 /d 1. assessment of biomarkers was done simultaneously with tlc at d 0 /d 1 , d 3 and during follow up of patients till their final outcome. there was no significant (p>0.05) mean change in pct, qcrp, sofa, ncd64, mhla-dr from day 1 to day 3, however, mean change was higher among cases than controls.on comparison of mhla-dr between the groups across time periods, mhla-dr was significantly (p=0.0001) lower among septic patients than controls at both day 1 and day 3. all biomarker correctly predicted cases among different percentage of patients with different sensitivity and specificity. there was no significant (p>0.05) association of mortality with the study biomarkers except for pct. in our study, diagnostic value of pct in differentiating sepsis from non-sepsis was similar to ncd64 among all biomarkers studied. no advantage of ncd64 or mhla-dr was found over pct in diagnosis and correlation with disease progression and mortality. introduction: aqp4 is a water channel protein contributing to astrocyte and immune cells migration, blood-brain barrier maintenance and cell survival [1] [2] . aqp4 genetic variants represent biomarkers associating with outcome after traumatic brain injury and intracerebral hemorrhage [3] [4] . linking aqp4 genetic polymorphism to the course of sepsis has not been studied. methods: study cohort included 124 icu patients diagnosed according to sepsis-3 consensus. aqp4 rs11661256 polymorphism was studied by analyzing pcr products in a 2% agarose gel using an aqp4 specific polynucleotide tetraprimer set. data were analyzed by log rank test (medcalc 18.11.3), and odds ratios/hazard ratios were computed. statistical significance was determined by fisher test (ft) or mann-whitney test. results: 23 of 124 sepsis patients had the minor mutation a for snp rs11661256 located within the regulatory 3' region of the aqp4 gene. septic shock occurred more frequently in homozygotic carriers of aqp4 c allele vs. patients with aa or ca genotype: or=3.75 (95%ci: 1.47-9.56), p=0.006 (ft). lethality in septic shock patients, n= 85, significantly increased compared to sepsis patients with no shock, n=39 (82% vs. 20%, p=0.001, ft). maximum sofa values were significantly lower in patients with minor allele a compared to cc carriers of (9.6 vs. 12.0, respectively, p=0.008). in post-surgery group of patients, carriers of ac or aa genotypes had significantly increased survival compared to patients with cc genotypes: chi-square=5.804; hr=0.455 (95%ci: 0.24 -0.863) for lethality; p=0.016 (figure 1) . association of minor allele a of aqp4 snp rs11661256 with survival in sepsis patients seems secondary to linking the snp to decreased development of multiorgan failure and septic shock that contribute to mortality. validation of presepsin as a biomarker of sepsis in comparison to procalcitonin, il-6 and il-8 v chantziara 1 , f kaminari 1 , c sklavou 1 , s fortis 2 , p kogionou 2 , s perez 2 , a efthymiou 1 1 saint savvas hospital, icu, athens, greece; 2 saint savvas hospital, cancer immunology and immunotherapy center, athens, greece critical care 2020, 24(suppl 1):p486 sepsis is an everyday challenge for the intensivist and biomarkers are useful tools for identification and treatment of this syndrome. we sought to validate presepsin as a biomarker of sepsis in comparison to pct(procalcitonin) and interleukins (il-6,il-8). we enrolled 25 patients,18 men and 7 women average age 58 (39.5-74) years old, apache ii 16 (13.5-20.5), saps ii 48(40.5-58.5), sofa 8 (6.5-9). 11 patients were septic on admission (according to surviving sepsis campaign: international guidelines for management of sepsis and septic shock: 2016), 9 had a septic episode during their hospitalization in the icu while 5 patients never endured sepsis. we measured presepsin, procalcitonin, il-6, il-8 during sepsis and on remission. results: all septic patients had increased values of presepsin, pct, il-6 and il-8 during sepsis with a cutoff value for presepsin 800pg/ml, while the values of these biomarkers were significantly decreased during remission or in comparison to non-septic patients(presepsin p = 0.002, pct p≤ 0.001, il-6 p≤ 0.001, il-8 p= 0.004. all patients who were not septic survived while among septic patients 8 died (40% mortality). presepsin correlated significantly with pct, il-6 and il-8 (p<0.05). presepsin is a valid biomarker of sepsis and correlates significantly with all the other values of pct, il-6 and il-8. clinical sepsis phenotypes are proposed at hospital presentation. these phenotypes, biomarker profiles, and outcomes are not yet reproduced in prospective data. even less is known about the biologic mechanism the drives these distinct groups. thus, we sought to validate clinical phenotypes and to determine markers of innate immunity, coagulation, tolerance and tissue damage in a prospective cohort. we prospectively studied patients with sepsis-3 criteria within 6 hours of presentation at 12 hospitals in pennsylvania (2018-2019) using automated electronic alerts. using 29 clinical variables, we predicted phenotypes (α, β, γ, δ) for each patient using euclidean distance anchored to published seneca phenotype centroids. discarded blood was analyzed in a subset (n=160) for markers of innate immunity (e.g. il-6, il-10), coagulation (e.g antithrombin iii, eselectin), tolerance (e.g. ho-1, igfbp7), and tissue damage (e.g. serum lactate, bicarbonate) results: among 549 patients, α-type was present in 146 (27%), β-type in 140 (25%), γ-type in 168 (31%) and δ-type in 95 (17%, figure 1a ). on average, β-type was older and more comorbid (mean 73, sd 9 yrs; mean elixhauser 4.6, sd 2.2) with renal dysfunction (median creatinine 1.8 [iqr 1.1 -2.9] mg/dl, p<0.01 all). the δ-type had more acidosis (mean hco3 -20.1, sd 4.7 meq/l), higher serum lactate (median 1.8 [iqr 1.0-3.5] mmol/l, p <0.01 both) and inpatient mortality (13%, figure 1b) . the γand δ-type had greater markers of innate immunity and abnormal coagulation (e.g il-6, icam p<0.01 both), while markers of increased tissue damage (lactate) and poor tolerance (ho-1) were present in δ-type, compared to α-type (figure 1c) . the distribution and characteristics of clinical sepsis phenotypes were reproduced in a prospective validation cohort. similar to the seneca study, distinct biomarker profiles of tissue damage, innate immunity and poor tolerance were present for the δ-type. the effect that neoadjuvant chemotherapy and hyperthermic intraperitoneal chemotherapy (hipec) may have in the postoperative kinetics of biomarkers remains unknow. some studies demonstrate that neoadjuvant chemotherapy and hipec do not invalidate the use of inflammatory markers in postoperative patient monitoring, but none have compared biomarkers kinetics between patients who underwent hipec or only cytoreduction surgery. our main purpose was to identify a difference pattern in c-reactive protein (crp). we conducted a single-center observational study from january 2015 to november 2019, including all patients who underwent cytoreductive surgery with or without hipec. crp was measured daily until seven post-operative day. we compared patients with and without hipec. a total of 19 patients were included, 15 were female. mean age was 63 yrs (44-76). no clinical and demographical differences were observed between groups. no documented infection was found. after surgery crp increased markedly in both groups. crp time-course from the day of surgery onwards was significantly different in hipec patients (9.78 ± 3.95 mg/dl vs 14.80 ± 5.63 mg/dl; p=0.035). multiple comparisons between hipec and non hipec patients were performed and crp concentration was significantly different on the 5th and 7th pod (figure 1 ). no differences were found in other biomarkers (leucocytes and platelets) neither in body temperature. after a major elective surgical insult crp levels markedly increase independently of hipec. serum crp time-course showed a higher pattern in hipec patients despite no infection detected. decreased thrombin generation potential is associated with increased thrombin generation markers in sepsis associated coagulopathy d hoppensteadt 1 , f siddiqui 1 , e bontekoe 1 , r laddu 1 , r matthew 2 , e brailovsky 3 , j fareed. introduction: sepsis associated coagulopathy (sac) is commonly seen in patients which leads to dysfunctional hemostasis in which uncontrolled protease generation results in the consumption of clotting factors. the purpose of this study is to determine the thrombin generation potential of baseline blood samples obtained from sac patients and demonstrate their relevance to thrombin generation markers. baseline citrated blood samples were prospectively collected from 49 patients with sac at the university of utah clinic. citrated normal controls (n=50) were obtained from george king biomedical (overland park, ks). thrombin generation studies were carried out using a flourogenic substrate method. tat and f1.2 were measured using elisa methods (seimens, indianapolis, in) . functional antithrombin levels were measured using a chromogenic substrate method. the peak thrombin levels and auc levels were lower in the sac patients in comparison to higher levels observed in the normal plasma ( table 1 ). the sac group showed much longer lag time in comparison to the normal group. wide variations in the results were observed in these parameters in the sac group. the f1.2 and tat levels in the sac group were much higher in comparison to the normal. the functional antithrombin levels were decreased in the sac group. these results validate that thrombin generation markers such as f1.2 and tat are elevated in patients with sac. however, thrombin generation parameters are significantly decreased in this group in comparison to normal. this may be due to the consumption of prothrombin due to the activation of the coagulation system. thus, persistent thrombin generation with simultaneous consumption of clotting factors such as prothrombin contributes to the consumption coagulopathy observed in sepsis patients. introduction: procalcitonin (pct) is used in the icu as an inflammatory marker to monitor bacterial infections and guide antibiotic therapy. whether pct can predict bacteremia and therefore could prevent expenses attached to bloodcultures is unknown . we investigated whether pct can predict the outcome of blood cultures in the icu and reduce expences. a single centre observational cohort study was performed in a dutch community teaching hospital . adult patients who were staying in the icu and were suspected of bacteremia were included. simultaneously with drawing of blood cultures, samples for pct measurement were obtained. expenses for pct measurement and bloodcultures were calculated. in the study period of one year, a total of 120 patients were included. three patients were excluded because of incomplete data. out of the 117 included patients, ten patients had positive blood cultures. there was a significant difference in pct levels between patients who had positive bloodcultures versus patients with negative bloodcultures (8.01 ng/ml vs 0.71 ng/ml) ( figure 1 ). the negative predictive value for negative blood cultures is 97% when pct is below 2ng/ml, there was no difference in crp levels between the two groups (148 mg/l vs 179 mg/l, p= 0.83).a set of negative blood cultures in our centre costs 35 euros. positive blood cultures however costs significantly more depending on the micro-organisms found. pct only costs 8.50 euros per measurement. so when blood cultures are omitted when the pct level is below 2ng/ml, a cost reduction of 38% can be achieved. a pct value below 2 ng/ml is a good predictor of a negative blood cultures in icu patients suspected of bacteremia. pct guided bloodculture management in these patients could lead to a significant cost reduction introduction: level of cfdna in plasma is a promising prognostic candidate biomarker in critical illness [1] . oxidized cfdna (ocfdna) have not been studied as a biomarker although its functional role in cellular stress have attracted attention of researches [2] . the goal of our study was to assess the early prognostic value of plasma cfdna/ocfdna for sepsis in a nicu setting. the cohort included 115 nicu patients diagnosed with stroke, intracerebral hemorrhage (ich), anoxia, encephalopathy. cfdna was isolated from day 1 plasma and stained with picogreen. oxidized dna was determined using dna immunoblotting with anti-8-oxo-desoxiguanosine antibodies. genotyping of allelic variants of the tlr9 rs352162 gene was performed using a pcr and designed allele-specific tetraprimers followed by electrophoretic separation of the products statistics was performed by the fisher test and mann-whitney test. results: sepsis was diagnosed by sepsis-3 criteria in 35 patients (30.4%). average nisu staying was 8,8±11,1 days. circulating dna plasma levels on day 1 predicted the future sepsis development (figure 1 ): or for cfdna was 7.54 (95%ci: 3.03-18.76), p<0.001; or for ocfdna was 5.57 (95%ci: 1.64-18.97), p=0.008. power of both performed tests with alpha=0.05: 1.0. log rank test demonstrated better predictive value of cfdna vs. ocfdna (figure) . concentrations of cfdna, but not ocfdna, on day 1 significantly positively correlated with maximum sofa values during hospitalization, day 3 and pre-outcome leukocyte count and neutrophil-to-lymphocyte ratios in a limited cohort of nisu patients with tlr9 rs352162 cc genotype and not in other patients with genotype tlr9 ct+tt. increased level of plasma cfdna better then ocfdna predicts sepsis development in nisu. further studies are warranted to clarify the fig. 1 (abstract p490) . pct values in patients with positive blood cultures and patients with negative blood cultures possible utility of tlr9 rs352162 polymorphism determining for sepsis risk stratification early on nisu admittance. admission was related with higher severity of illness and extension of icu stay for all groups. reduced cbt fluctuations upon icu admission was found to more severely ill patients with worse clinical outcomes, while the more periodic cbt patterns were correlated with high cbt rhythmicity and better outcome. the impact of sex on sepsis incidence and mortality have been elucidated in previous studies, and sex is increasingly recognized as one key factor in sepsis [1] . some studies indicate that women have better immunologic responses to infections [2] . later investigations assume this advantage is linked to immune modulating genes located on the x-chromosome [3] . the purpose of this study is to reveal sex differences in incidence of and mortality of sepsis in a large population-based cohort. methods: 64049 adult participants in the hunt2 study (1995-97) were followed from inclusion through end of 2011. incident bloodstream infections (bsi) from all local and regional hospitals in nord-trøndelag county were identified through linkage with the mid-norway sepsis register, which includes prospectively registered information on bsi used as a specific indicator of sepsis. we estimated age-adjusted cumulative incidence of first-time bsi and compared the risk of a first-time bsi and bsi mortality in men and women using age-adjusted cox proportional hazard regression. during a median follow-up of 14.8 years 1840 individuals experienced at least one episode of bsi, and 396 died within 30 days after a bsi. cumulative incidence and cumulative mortality curves are shown in fig. 1a introduction:the proportion of hospital-acquired infections (hai) among sepsis patients is unknown in germany. systematic differences in hai foci between sepsis patients with and without icu treatment are insufficiently described. retrospective cohort study based on nationwide health claims data of the german statutory health insurance aok. incident inpatient sepsis cases were identified in 2013/2014 among insured persons >15y without preceding sepsis in 24 months prior to index hospitalization. sepsis was defined according to explicit sepsis icd-10-codes (incl. severe sepsis/septic shock). hai were defined based on specific icd-10-codes for surgical site infection, catheterintroduction: elevated renin is associated with an increased risk of death in patients with vasodilatory shock (vs). recent data show that patients with vs and elevated renin levels have improved survival when treated with angiotensin ii (ang ii) + standard care (sc) vs placebo + sc. patients with acute respiratory distress syndrome (ards) can develop angiotensin-converting enzyme (ace) defects that can lead to elevated renin levels and insufficient endogenous ang ii production. we hypothesized that patients with severe ards and elevated renin shock would have improved survival when treated with ang ii + sc vs placebo + sc. in the randomized, placebo-controlled, double-blind athos-3 study, 321 patients with severe vs receiving >0.2 μg/kg/min of norepinephrine or the equivalent were randomized to intravenous ang ii (n= 163) or placebo (n=158). in a post hoc analysis, we assessed the subset of patients with elevated renin (defined as a renin level greater than the median value of the overall athos-3 population) and ards (defined by a pao2/fio2 ratio <300) at the time of randomization. survival to 28 days was compared between the ang ii group (n=41) and the placebo group (n=61). in patients with elevated renin and ards, baseline age, acute physiology and chronic health evaluation ii score, and blood pressure were similar in the ang ii and placebo groups. the median serum renin level was 459.5 pg/ml (iqr: 285.8-1036.0) compared to the normal range for serum renin: 5-58 pg/ml. a significantly higher proportion of patients receiving ang ii survived to day 28 compared to those in the placebo group (51% vs 31%; p=0.01). elevated renin identified patients with vs and ards who were most likely to gain a survival benefit from ang ii. elevated renin is likely caused by an ace defect and may describe an important subset of patients with a biotype that responds well to ang ii therapy. introduction: elevated renin levels have been shown to be associated with an increased risk of death and more severe acute kidney injury (aki) in patients with vasodilatory shock (vs). recent data show that patients with vs and elevated renin levels have improved survival when treated with angiotensin ii (ang ii) + standard care (sc) vs placebo (pbo) + sc. we hypothesized that vs patients with severe aki and elevated renin levels would have improved survival and enhanced renal recovery with ang ii treatment. in the randomized, pbo-controlled, double-blind athos-3 study, 321 patients with severe vs received >0.2 μg/kg/min of norepinephrine or the equivalent and were randomized to intravenous ang ii + sc (n=163) or pbo + sc (n=158). in a post hoc analysis, we assessed the subset of patients with elevated renin (defined as a renin level greater than the median value of the overall athos-3 population) and severe aki (defined as those with aki requiring renal replacement therapy [rrt] at baseline). survival and renal recovery were assessed in patients treated with ang ii + sc (n=45) and pbo + sc (n=60). in patients with elevated renin and severe aki, baseline age, acute physiology and chronic health evaluation ii score, and blood pressure were similar between ang ii + sc vs pbo + sc. the median baseline serum renin level in the whole group was 352.5 pg/ml (iqr: 115.9-785.4; normal range for serum renin: 5-58 pg/ml). a significantly higher proportion of patients receiving ang ii + sc vs pbo + sc survived to day 28 (43% vs 22%, respectively; p=0.03). ang ii recipients also had a higher rate of discontinuation from rrt by day 7 (43% vs 12%; p=0.04). in this study, elevated-renin shock patients with aki treated with ang ii + sc gained a survival benefit and earlier discontinuation from rrt compared to those receiving pbo + sc. elevated renin is likely caused by an angiotensin-converting enzyme defect and may identify those patients with a biotype that responds well to ang ii therapy. most clinical trials conclude the ineffective use of anticoagulation for sepsis-induced coagulopathy [1] . however, post hoc analyses of randomized control trials report positive results [2] , suggesting anticoagulation is effective in specific populations exhibiting coagulopathy. further, anticoagulants should be administered in the early phase [3] ; however, methods for precisely predicting the progression of sepsis-induced coagulopathy are not established. this study aimed to create and evaluate a prediction model of coagulopathy progression using machine-learning techniques. we performed a subgroup analysis of data from a retrospective cohort study involving adult septic patients in 40 japanese institutions from january 2011 to december 2013 and used the japanese association for acute medicine disseminated intravascular coagulation (dic) score as a dic severity index test. the predictive ability of δdic ([dic score on day 3] -[dic score on day 1]) was evaluated using various statistical methods. using variables available at the outset, we compared the predictive ability of random forest (rf) and support vector machine (svm) with that of multiple linear regression analysis. a total of 1110 adults with sepsis were included in the analysis. the root mean square error in δdic score for the multiple linear regression analysis model was 2.1168 compared with values of 1.6508 and 1.9394 for rf and svm, respectively. thus, the rf method predicted the progression of sepsis-induced coagulopathy more accurately than multiple linear regression analysis. conclusions: rf, a machine-learning technique, was superior to multiple linear regression analysis in predicting the progression of sepsis-induced coagulopathy. this prediction model might enable us to use anticoagulation in an early phase. this study examined the efficacy and safety of landiolol, an ultrashort-acting β1-blocker, for treating sepsis-related tachyarrhythmia, according to patient background characteristics. the j-land 3s study (japiccti-173767) was conducted in patients with sepsis, diagnosed according to the sepsis-3 criteria, and tachyarrhythmia (atrial fibrillation, atrial flutter, or sinus tachyarrhythmia). the patients had a mean heart rate of ≥100 beats/min and required catecholamine administration to maintain a mean blood pressure of ≥65 mmhg. the efficacy endpoint was the percentage of patients whose heart rate could be controlled within 60-94 beats/min at 24 h of registration. the safety endpoint was the incidence of adverse events within 168 h of registration. subgroup analyses of efficacy and safety were performed after stratifying the patients according to various patient background characteristics. a total of 151 patients were randomized, 76 to landiolol and 75 to the control group. the efficacy endpoint, percentage of patients with a heart rate of 60-94 beats/min at 24 h of registration, was significantly higher in the landiolol group (54.7% vs 33.3%; mantel-haenszel test: p = 0.0031). the incidence of adverse events was 63.6% and 59.5% in the landiolol and control groups, respectively, and there was no difference between the two groups. most adverse events were related to sepsis or septic shock. the subgroup analyses showed that no patient background characteristic clearly affected the efficacy and safety of landiolol. landiolol is a well tolerated and effective therapeutic agent for controlling heart rate in patients with sepsis-related tachyarrhythmias; its safety and efficacy were not affected by the patient background characteristics investigated. tissue oxygenation monitoring in sepsis r marinova, at temelkov umhat alexandrovska, anesthesiology and intensive care, sofia, bulgaria critical care 2020, 24(suppl 1):p502 near-infrared spectroscopy (nirs) was proposed as a concept in the end of 20th century. this method offers noninvasive monitoring of oxy-and deoxyhemoglobin in tissues.nirs could be measured on the thenar or forehead within few santimeters of the skin. it was first applied as a monitoring in cardiovascular surgery. patients with sepsis have changes in the microcirculation which are important target for therapy. invasive monitoring of oxygen delivery and consumption has been used in patients with sepsis but as every invasive technique such a monitoring hides risks. nirs offers a noninvasive method for tissue oxygenation monitoring (sto2) and could be useful in patients with sepsis and septic shock. the aim of the study is to compare noninvasive tissue oxygenation monitoring with hemodinamic monitoring and lactate values in patients with sepsis methods:the study includes 19 critically ill patients in icu of umhat alexandrovska, sofia. 10 of the patients fullfil the criteria for septic state. the other 9 patients do not have sepsis. in both group of patients are measured tissue oxygenation with invios monitor, mean arterial pressure, oxygen saturation in mixed venous blood and lactate values during 72 h after icu admission. patients with sepsis are reported with significantly lower values of tissue oxygenation, compared to patients without sepsis. the values of tissue oxygenation correlate well with the mixed venous blood oxygenation, mean arterial pressure and lactate values but not significantly with apache scores. conclusions: nirs when used for tissue oxygenation monitoring correlates well with the hemodinamic monitoring and lacate values in patients with sepsis and could be used as an noninvasive monitoring for guiding teurapeutic strategies. tissue oxygenation monitoring has no linear correlation with the severity of illness in patients with sepsis and could not be reccomended as a guidance in the early ressuscitating stage of sepsis. further investiganions in these field are needed.the sequenom´s massarray platform and a recessive inheritance model was selected (cc vs tt/ct). the possible association between the cc recessive form of the rs279451 polymorphism and the septic shock risk was analyzed, demonstrating a statistically significant relationship (p=0.02) between both conditions. among patients who developed septic shock, 79.2% presented a recessive inheritance pattern while 54.5% showed the ct/tt genotype. on the other hand, those patients with the recessive form of the rs279451 polymorphism were selected and a statistical analysis was performed comparing those patients who developed septic shock from those who did not develop it, obtaining a statistically significant relationship (p=0.036) between the presence of the recessive form of polymorphism and the likelihood of developing septic shock. the recessive form of rs279451 polymorphism is a risk factor for septic shock in post-operative patients of major abdominal surgery. introduction: sepsis remains one of the major causes of morbidity with mortality rates as high as 50 % worldwide, representing significant clinical challenge to confront highly intangible therapeutic needs. rnabased structures are emerging as versatile tools encompassing a variety of functions capable to bypass the current protein-and cellbased therapies. rna aptamers act as disease-associated protein antagonists. here, the effects of an aptamer, apta-1, were evaluated in animal models that mimic systemic inflammation in humans. high dose of lps endotoxin was used to induce systemic inflammation in mice and in non-human primate animal models. apta-1 was administered intravenously in two doses post lps infection. animals were monitored and blood samples collected up to 72 hours after apta-1 administration. healthy-and lps-only treated animals served as control groups. complex analyses of clinical parameters, hematology, serum biochemistry, inflammation and tissue damage markers were performed. results: apta-1 increased survival of endotoxin challenged animals up to 80% in a dose-dependent manner and exerted profound effects on wellbeing and recovery of healthy eating habits. administration of apta-1 led to delayed coagulation and enhanced fibrinolysis; maintained the complement cascade activated while preventing it from further amplification. expression of pro-inflammatory cytokines was reduced while anti-inflammatory increased. endogenous pro-inflammatory molecules (damps), secreted from injured cells, were preserved at healthy level in animals treated with apta-1. systemic inflammation and sepsis lead to severe dysregulation of several arms/axis of innate immune response. our studies showed that apta-1 affects various components of this system and restores the organism's control over its dysregulated immune response. thus, apta-1 might be a promising potential therapeutic candidate to treat life-threatening conditions such sepsis. several preclinical studies demonstrated beneficial effects for methane (ch 4 ) administration in various inflammatory conditions. our aim was to investigate the consequences of post-treatment with inhaled ch 4 in a clinically relevant intra-abdominal sepsis model. anesthetized minipigs were subjected to fecal peritonitis (0.6 g/kg, 5-9x10 6 cfu i.p.; n=22) or sham-operation (sterile saline i.p; n=5). invasive hemodynamic monitoring with blood gas analyses was started between 16-24 hours, organ dysfunction parameters (pao 2 /fio 2 ratio; mean arterial pressure; lactate, bilirubin, creatinine; urine output and platelet counts) were determined according to a modified porcinespecific sequential organ failure assessment (ps-sofa) score system, the perfusion rate (pr) of sublingual microcirculation was measured by incident dark field illumination imaging. the animals were divided into non-treated septic or septic shock groups (n=6-6) and ch 4treated septic or septic shock (n=5-5) subgroups, ch 4 inhalation started from the 18th hr (2.2% ch 4 in normoxic air; 500 ml/min). despite the standardized induction, heterogeneous severity of organ damage was evolved. in septic and septic shock groups the median values of ps-sofa score reached 5 (4.75-5.65) and 13 (11.75-14), respectively. septic shock was characterized by significant elevations of creatinine and bilirubin levels, while the platelet count decreased (from 332 to 76 *10 9 /l). inhalation of ch 4 increased the sublingual pr by 22% in the septic group, the creatinine and bilirubin levels were decreased by 28% and 80%, respectively. ch 4 post-treatment significantly decreased the ps-sofa score (to 1; 0.5-2.75) and resulted in lower values in septic shock group (to 10; 9.5-12.4). methane post-treatment effectively influences sepsis-related end organ dysfunction. up to a severity threshold it may be a promising additional organ protective tool. evaluation of sepsis awareness among various groups in turkey: a survey study s erel, o ermis, ö nadastepe, l karabıyık gazi university school of medicine, anesthesiology and intensive care, ankara, turkey critical care 2020, 24(suppl 1):p510 introduction: sepsis is a common life-threatening condition in critically ill patients [1] . public awareness is important for early recognition of sepsis and improvement of outcomes [2] . we aimed to evaluate sepsis awareness among different groups of people. methods: prospective paper-based surveys were issued between 1st july and 1st august 2019 to patients, the relatives of the patiens, hospital staff and general public who gave consent to participate in the study. the questionnaire included ten questions about demographic informations, occupational informations of hospital stuff and sepsis awareness. a total of 588 participated in the survey. of these participants, 87 (14.3%) were patients, 50 (8.5%) were relatives of patients, 134 (22.8%) were physicians, 125 (21.3%) were medical students, 49 (8.3%) were nurses, 51 (8.7%) were other hospital stuff and 92 (%15.6) were other people. of these participants, 425 (72.3%) had heard of the word "sepsis". 206 (35.0%) responded correctly regarding the definition of sepsis. 325 (55.3%) of the participants heard the word "sepsis" during their education, but only 53 (9%) heard it through the media. in the groups of high school graduates, university graduates and postgraduates, the rate of hearing the word sepsis and correctly identifying sepsis is significantly higher than the primary school graduates or illiterate groups. (p<0.05). physicians, nurses and medical students were heard of the word "sepsis" significantly more than other groups (p<0.005). physicians and medical students responded more accurately to the definition of sepsis than other groups (p<0.05). public awareness of sepsis is limited compared to healthcare workers. increasing public knowledge of sepsis through education and through media may contribute to raising public awareness and improving outcomes. the association between clinical phenotype cohesiveness and sepsis transitions after presentation jn kennedy 1 , eb brant 1 , km demerle 2 , ch chang 3 , s wang 4 , dc angus 1 , cw seymour 5 1 key: cord-015021-pol2qm74 authors: nan title: third international congress on the immune consequences of trauma, shock and sepsis —mechanisms and therapeutic approaches date: 1994 journal: intensive care med doi: 10.1007/bf02258437 sha: doc_id: 15021 cord_uid: pol2qm74 nan this issue of the journal contains the abstracts for the third international congress on the immune consequences of trauma, shock and sepsis -mechanisms and therapeutic approaches. we hope that the information contained in this special issue will stimulate you to participate in the congress, to contribute to the knowledge being developed in this field and to use this information to help you in providing better care for your patients. we thank the editors and the editorial board and publishers of the journal for their interest and support in preparation of this special issue. we also, on behalf of the scientific committee, welcome you to the third international congress in munich on 2-5 march 1994. when, in the mid-1980s, we thought of having a worldwide congress, we hoped to bring together investigators to discuss this theme. the explosion of knowledge occurring around that time provided an excellent background against which the first conference in 1988 provided stateof-the-art information and consensus on factors involved in injury and sepsis. in 1991, the second congress was held at the time of another resurgence of research, study and information on injured and operated patients. it seemed then that there would be a lull in the development of new information and therapy, and that another state-of-the art conference might not be necessary until 1995 or 1996. however, the explosion in molecular biology has continued. the wonderful world of cytokines has gone from ill to il-6 to il-11, il-12 and il-13 and beyond. the vast amount of information about mediators and their importance in disease is impressive. this has all suggested a magic bullet that might be used to alter or block inflammatory responses. this has not happened, however, and the question is "why not"? our science is powerful, but our therapy is still weak. what are the issues, then, in 1994, to be dealt with at this symposium and congress? (1) proposals for new terminology. there have been a number of proposals for new terminology and new classifications of injury, sepsis, inflammation and various other problems related to human illness. the question is whether this is the way to go. will this contribute to better clinical trials, information basis and better research? the pros and cons of this development will be reviewed by those making the proposals and those questioning the need for and wisdom of this effort. (2) magic bullets: the prospect of a magic bullet to deal with inflammation in injury and infection seemed highly promising earlier. many preclinical trials and a lot of animal research suggested the possibility of a great breakthrough in clinical care. what has become, then, of all the expensive and extensive multi-institution randomized, placebo-controlled, double-blind clinical trials of agents that block mediators and endotoxin. many such studies have yielded equivocal, marginal or negative results. the reasons for this and the future of clinical research will be the subject of presentations and discussions to set the stage for further work. (3) should future clinical trials be based on new classifications of illness such as mods, sirs, apache iii, sap ii, mrm, etc., or should trials be dedicated to specific diseases -urinary tract infections, pneumonia, trauma patients, cardiac surgery and other specific problems, rather than generalized problems of sepsis, the sepsis syndrome and other classifications? in other words, should we now begin to have clinical trials on specific diseases with causes that are known and can be attacked? the causality of disease becomes an important consideration in this regard. (4) a multitude of potential therapeutic agents has been proposed on the basis of animal studies. how should we decide which of them should be brought to clinical trial? the possibilities are endless as we develop new clinical information about the mechanisms and pathogenesis of human disease. (5) information on the pathogenic mechanism of disease states and of injury continued to emerge in an explosive fashion, and in light of our gathering knowledge we can look forward to working out a cohesive system of response to injury. (6) additional information will be provided in plenary sections, many symposia and free communication sessions and posters, which will update the participants on a variety of relevant topics presented by many of the leading in-iv vestigators in these fields. topics will range from molecular mechanisms, such as signal transduction, through the explosive growth of information on the role of cytokines and pathophysiology, to practical considerations in the design of immunomodulatory therapeutic regimens. these merely touch on a few areas, from the basic to the clinical, which will be the subjects of those symposia. all this information will fit into the jigsaw of this exciting area and its stimulus to further research study. this promises to provide an exciting, educational programme with experts and participants from all over the world. we hope it will set the stage for many years to come and will increase our understanding of trauma, shock and sepsis and help us to provide better therapy for those of our patients who are affected by such problems. a. the clinical syndrome of mods versus mof will be reviewed in detail by those who have made these proposals. b. an extensive review of the design and interpretation of clinical trials in patients with shock and injury will be provided. the reasons why so many clinical studies in the recent past have been negative will be reviewed. the therapeutic strategies that are being developed for the treatment or prevention of mods or mof will be the subject of another panel discussion by experts who have been involved in and contributed to this area. a consensus conference or controversy conference will be presented about various aspects of mods or mof, including the benefits of supernormal oxygen delivery, bacterial translocation, parenteral nutrition, the immune response and other aspects. the successes and failures of completed clinical trials will be presented by those who are involved in these clinical trials, with a refreshing review of the problems related to that injury. there will be late news about studies just being completed at present or after the beginning of 1994 and where they stand. c. the mechanisms and biochemical profiles of specific organ dysfunction or failure will be reviewed. what are the definitions? what are the mechanisms? how can organ dysfunction and/or failure be defined? an extensive review of the biological mechanisms involved in production of injury by mediators will be presented. a session will be devoted to how future ongoing trials might be better designed and what can be done about the studies recently completed, many of which are negative. d. the immunological or inflammatory pathways resulting in organ injury will be reviewed in detail in presentations and a panel discussion. we look forward to welcoming you to an exciting and rewarding conference, which undoubtedly possesses the potential to become a landmark event and major reference point for any scientific discussion about the complex of host defense dysfunctions following trauma, shock and sepsis. studies over the past 25 years have established that the contact system, which forms bradykin/~, is gax important mediator in hypotensive septicemia. in addition to hradyk{nln, another product of the contact system, kailikrein, can mediate inflammation by virtue of its chemotaetic mad neutrophj/activating properties. using functional and immunochemical tech~2ques, we have demonstrated activation of the contact system in the adult respiratory distress syndrome in typhoid fever and clin/cal sepsis. we have also been able to inhibit the hypotension but not the disseminated intravaseular coagulation in a model of primate sepsis by the use of a monoclonal antibody directed agsi~st factor xii, the initiating protein of the contact system, in volunteers given e. coil endotoxin, who did not develop hypotension, we were also able to demonstrate activation of the contact system with a rise of alpha-2macrogiebulin-kalllkrein complex. we have also examined, j~ an i~tensive care situation, patients with sirs. we found that serial measuremezzts of the contact system were useful in eva~u~ting prognosis+ these studies suggest that inhibition of kalllkrein a~d l e r bradykinin actions might be useful i~ obviating many .of the features seen in sepsis and septic shock. dextran sulfate (dxs) activates the contact system and, in vivo, produces transient hypotension. in order to better define the mechanisms underlying the dxs-induced hypotension, we investigated the effects of either the plasma kallikrein inhibitor, des-pro2-iarg]5]aprotinin (bay 4620) or the b2 kinin antagonist, hoe 140 on the hypotensive response to dxs. in the first study, anesthetized miniature pigs (5 pigs/group, randomly assigned) were given one of the following treatment protocols: 1) dxs (5 mg/kg), 2-5) dxs plus bay 4620 (45, 90, 180, or 360 rag), or 6) saline. dxs alone produced a profound but transient systemic arterial hypotension with a corresponding reduction in plasma kinin-containing kininogen. circulating kinin levels, complement fragment c3adesarg and fibrin mom)mer were all increased. bay 4620 produced a dose-dependent delay or attenuation in these effects with the highest dose completely blocking dxs-induced hypotension and elevations of kinin, c3adesarg and fibrin monomer levels. thus, the effects of dxs are solely dependent on contact system activation and this activation is sensitive to bay 4620. llowev~:r, contact system activation is known to produce changes in a variety of vasoactive mediators, all of which can affect blood pressure. in a second study, two groups of pigs (3/group) were given either dxs alone (2 mg/kg) or dxs 10 minutes after a bolus injection of hoe 140 (30 #g/kg). dxs alone produced transient hypotenmon. this response was completely blocked by hoe 140 pretreatment. both groups had identical reductions in kinin-containing kininogen. we conclude that dxs-induced hypotension is produced by activation of the contact system which results in the production of bradykinin. liberation of bradykinin is both necessary and sufficient to produce all of the hemodynamic changes observed. dr. matthias siebeck, department of surgery, university of munich, klinikum lnnenstadt, nussbaumstrasse 20, d-80336 munich, germany in experimental animals exposed to i.v. injection of endotoxin accumulation of leukocytes in various organs as lungs and the liver is a prominent feature. as a part of these morphological changes damages of endothelial ceils are regularly seen. this process, which is a part of endothelial-cellular interaction, leeds to exposure of the sub-endothelial basement membran. the basement membran is known f6r its capacity to activate the contact system of plasma. during this cascade activation, coagulation factor xii is converted to the active factor xii. this activation might produce increased plasma kallikrein activities and thereby give release of the vasoactive substance bradykinin. using a porcine model we have noticed that endotoxin infusion (0,01 mg/kg) induces elevated plasma kailikrein activities within two hours after the start of the infusion. this enzyme activity remained increased during the next hours and reached value of up to 50 u/1. in patients with sepsis we also have observed elevated plasma kallikrein activities with enzyme activities up to 200 u/1. in order to further elucidate the significance of these elevated enzyme activities, we prepared human plasma kallikrein and injected it intravenously in anaesthetized pigs (1). when very small plasma kailikrein activities (0,3 u/kg bodyweight) were given intravenously a 50% decrease in arterial blood pressure was seen in the animals. in the patients with sepsis also decreases in prekallikrein values and functional plasma kallikrein inhibition are frequently seen. furthermore, degradation of high molecular weight kioinogen is found in these patients indicating formation of bradykinin. these experimental and clinical studies underline that contact activation in sepsis might results in the release of very powerful mediator substances which can be of pathophysiological importance in this disease. a number of pathological disorders as reperfusion injury, bone marrow transplantation, polytrauma and septic shock are associated with capillary leakage. as the activation of the complement system and the contact phase play a major role in these diseases we investigated whether cl-lnhibitor (c1-inh), which inactivates cl-esterase, kallikrein and clotting factors xii and xl, could abolish vascular leakage. a capillary leakage was induced in rats by the administration of interleukin-2 (5 x 106 iu/kg). the increased vascular permeability was monitored for one hour as the extravasation of fitc marked rat serum albumin from a mesenterial vessel by a video-image processing system. ci-inh (berinert®, behringwerke) given as a single i.v. bolus in concentrations of 100, 250 or 500 u/kg dose-dependently prevented the capillary leakage. carrageenaninduced inflammation in the rat leads to vascutar leakage and to edematous swelling of the paw. ci-inh in this model leads to a dose-dependent decrease in paw edema formation. finally, we investigated the effect of ci-inh (infusion (100-125 u/kg x h) on a lps-induced shock in the rat by combination therapy with the antithrombotlc agents antithrombin ill (kybernin®) or rec. hirudin (both substances from behringwerke). in this animal model mortality was 90 % in the untreated control. both antithrombotic agents decreased mortality rates by inhibiting formation of dic; a further significant improvement of survival was achieved by the treatment with ci-inh. thus+ it could be concluded that c1-inh has a beneficial effect in diseases associated with a vascular leakage. iclb and laboratory for experimental and clinical immunology, university of amsterdam, the netherlands; 2thrombosis research center, temple university, penn., usa; 3oklahoma medical research foundation,. ok. city, usa. to evaluate the contribution of the contact system to activation of other mediator systems in an experimental model of sepsis, we investigated the effect of mab c6b7 which inhibits activation of factor xli, on activation of complement and fibrinolytic cascades and activation of neutrophils in baboons suffering from a lethal sepsis. activation of the complement system was assessed by measuring circulating levels of c3b/c and c4b/c, and a significant reduction was observed in 5 animals that had received a lethal dose of e. coli together with mab c687 (treatment group), compared to 4 animals that had received a lethal dose of e. coil only (control group). activation of the fibrinolytic system as reflected by circulating plasmin-=2antiplasmin complexes and tissue plasminogen activator, and activation of neutrophils, assessed by measuring circulating elastase-=l-antitrypsin complexes, was also significantly less in the treatment group. we conclude that activation of the contact system protein factor xll during the inflammatory response to a lethal dose of e. coil in this baboon model, modulates directly or indirectly activation of the complement and fibrinolytic systems and that of neutrophils. in a prospective study, plasma levels of c3a, c3, and c5a were measured in 30 patients from an internal intensive care unit. 20 patients were clinically septic defined by the criteria of bone et al.(l) . the remaining 10 patients were critically ill but didn't fulfill the clinical criteria of sepsis. from both groups of patients blood samples were taken over a l0 days period. during the first 3 days blood samples were drawn every 8 h, on day 4-6 every 12 h and the last 4 days once daily. mean plasma concentrations of c3a within the first 32 h after clinical onset of sepsis were 1019 + 618 pg/ml, whereas non-septic-patients exhibited mean values of only 595 +_ 312 p_g/m/. c3 levels were lower for septic-patients (840 + 480 lag/ml) than for non-septic-patients (1340 _+ 770 lag/ml). the most profound difference between both groups was found, when the c3a/c3 ratio was compared (1.84 + 2.28 for septic-patients and 0.4 _+_ 0.18 for the control group). no significant differences between both patient groups were observed in c5a plasma levels (5.9 + 2.1 ng/ml in septic-patients vs. 5.6 _+ 1.5 ng/ml in control patients). in 13 of 20 cases of clinically defined sepsis causative organisms like bacteria, protozoa or fungi could be cultured from blood, bronchoalveolar lavages and/or section materials. application of the complement parameters to survivors (n=9) and non-survivors (n=l 1) within the septic-group revealed, that the c3a/c3 ratio could also be used as a prognostic parameter for clinical outcome. the possibility of rapid and easy measurement of c3a and c3 in only 20-25 minutes (2) and the significant difference of the c3ajc3 ratio between the septic and non-septic group renders this parameter a good candidate for early diagnosis of sepsis in the intensive care unit. hirudin, a single polypeptide chain composed of 65 amino acids with 6 cysteine residues (mr 7000 daitons), is the most potent and specific thrombin inhibitor, which is now available as a genetically engineered product (rec. hirudin -hbw 023, behringwerke; marburg). the aim of our study was to establish a rabbit model of tissue factor (tf) induced activation of the extrinsic pathway of coagulation and to evaluate the therapeutic efficacy of rec. hirudin. coagulation was induced in female nzw rabbits by infusion of 0.5 p.g/kgxh thromboplastin for 7 hours. development of disseminated clotting was manifested by a decrease of fibrinogen and platelets to 27.0 % and 33,0 % respectively, and by an increase of fibrin monomers from 13.2 to > 85.0 ~tg/ml. we administered rec. hirudin to rabbits in 3 different concentrations (0.5, 1.0 and 2.0 mg/kg); treatment started simultaneously with the infusion as an i.v. bolus. rec. hirudin significantly prevented the decrease of fibrinogen, platelets and the increase of fibrin monomers. this effect was dose dependent and long lasting, even 7 hours after the administration of rec. hirudin, clotting was still significantly reduced. as could be drawn from the plasma levels, rec. hirudin had been cleared from plasma at this time. in a post-treatment study we administered rec. hirudin (0.5, 1.0 and 2.0 mg/kg i.v. bolus) as late as 2 hours after the start of tf infusion. at this time there was already a prominent activation of coagulation. even in this post-treatment regimen rec. hirudin significantly prevented disseminated clotting. hence, it was concluded, that rec. hirudin by inkihiting thrombin could be effective in the prevention of coagulation disorders including disseminated intravascular clotting (dic) induced by a septic disease. research laboratories of behringwerke ag, 35001 marburg, germany $3 novel protease inhibitory activities of the second domain of urinary trypsin inhibitor (r-020) and its effect on sepns-lnduced organ injury in rat atsuo murata 1, hitoshi toda 1, ken'ichi uda 1, hidewaki nakagawa 1 , takesada mori 1, hideaki morishita 2, tom yamakawa 2, jiro hirese 2, atsushi ni~ 2, nariaki matsuura 3 1osaka university medical school, osaka, 2mochida pharmaceutical co. ltd. tokyo, 3wakayama medical schoof, wakayama, japan inhibitory-activities of the second kuntz-type inhibitor domain of human urinary trypsin inhibitor (uti) and its effect on sepsis-induced organ injury in rat were investigated by using the recombinant protein. uti is a glycoprotein with a structure in which 2 kunitz-type inhibitor domains are linked in a row. we isolated the gene encoding the second kunitz-type inhibitor domain of uti, and then constructed expression plasmids by ligating it to the e. coli phoa signal peptide gene. these plasmids expressed the second domain in e. coil strain je5505 which lacks the membrane lipoprotein. the recombinant second domain (r-020) innb[ted trypsin, plasmin, neutrophil elastase and chymotrypsin. in addition it inhibited blood coagulation factor xa and plasma kallikrein in a concentration dependent and competitive manner. the in vivo effect of the recombinant r-020 was investigated in a rat model of septic shock induced by cecal ligation and puncture. the administration of r-020 significantly improved the survival rate of the rats and attenuated the pathological changes of lung and iiver. we found out the novel protease inhibitory activities of the second domain of uti and its protective effects on sepsis-induced organ injury. macrophages are known to secrete lysosomal proteinases,mainly cathepsin b and cathepsin l, and also ~-proteinase inhibitor (pi),related to acute phase proteins.disturbances of proteinases/ proteinase inhibitors correlates with inflammatory process,leading sometimes to noncontrol "pathglogical" proteolysis (jochum et ai.,1990) . the cathepsin l-like and cathepsin b-like activity were measured in serum of 42 patients with chronic bronchitis (14 -with obstructive, 28 -with nonobstructive bronchitis),acute bronchitis (15) and 35 healthy persons.simultaneously the level of~pi was determined in the same groups.cysteine proteinases were measured with help of fluorogenic substrates,as was presented earlier (korolenko et ai.,1991) , ~pi with help of immune enzyme method. it was shown increase of cathepsin l-like and cathepsin b-like activities during aggravation of chronic bronchitis comparatively to the controls (2-4 fold) .after treatment there was a tendency to normalization of indices,but the increase was about 30-40% more than the control values.~pi level in this group was also increased (two-fold),in patients with acute bronchitis -2-4-times more comparatively to the control.it is possible to conclude that chronic bronchitis induced increased secretion both cysteine proteinases and d{pi into blood. some peculiarities of ratio were noted in patients with emphysema. endotoxins are microbial products derived from the outer cell membrane of gram negative bacteria. the active component of endotoxin is lipopolysaccharide (lps), a complex macromolecule consisting of polysaccharide covalently bound to a unique lipid, termed lipid a. now recognized to embody the endotoxic principle of lps, lipid a consists of a/31-6 diglucosamine backbone, both ester and amide linked fatty acids, some of which are acyloxyacylated, and charged constituents such as phosphate, phosphorylethanolamine and 4 amino arbinose lps, exerts its biological effects in vivo by noncytotoxic interactions with a variety of host inflammatory mediator cells, primarily the mononuclear phagocyte and the endothelial cell, although other host cells also participate. these interactions are modulated by lps-specific binding proteins found in plasma, including lps-binding protein (lbp) scd14 and perhaps other proteins as well. specific receptors for lps have been identified on mammalian cells which mediate signal transduction via multiple pathways. lps-activated host cells are stimulated to secrete or express multiple proinflammatory mediators, including tnf-a, illa, il-1/3, ifn-a, il-6, il-8, il-10, paf, pge, ltb 4 and procoagulant activity. the overproduction of these proinfiammatory mediators results in the manifestations of endotoxemia, observed experimentally as fever, hypotension, disseminated intravascular coagulation and death. modulation of activity of these mediators protects animals against lethality. similar pathways are thought to be operative in gram negative sepsis, and control studies with human volunteers support such conclusions. immunotherapeutic approaches in clinical gram negative sepsis have, to date, been less successful. in vitro experiments and studies in animal models have recently shown that several proteinaceous bacterial exotoxins can evoke cytotoxic effects that ultimately lead to cardiovascular collapse and shock. since the possible relevance of bacterial exotoxins in the pathogenesis of septic shock has received very little attention in the past, an attempt will be made here to provide a brief overview of this generaily neglected topic. protein toxins act intracellularly or they dz~nage the integrity and function of the plasma membrane. major representatives of the former group are the adenosine diphosphate (adp)-ribosylating toxins, e.g. cholera and cholera-like toxins, diphtheria toxin), and the neurotoxins. most medically relevant toxins of this category have been studied in great detail. although often responsible for severe and sometimes fatal disease, their association with septic shock is rare. in contrast, experimental evidence is accumulating for a role of membrane<lamaging toxins in the pathogenesis of inflammatory lesions. formation of transmembrane pores is probably the most widespread mechanism via which such physical membrane perturbation can occur (1,2), the present discussion will be restricted to this category of toxins. the author shall first discuss basic properties of pore-forming proteins, and consider the factors that direct their attack to specific targets. thereafter, attention will be drawn to diverse functional consequences that may follow membrane damage so that a general concept of how pore-forming toxins may contribute towards the development of shock will evolve. i. bhakdi, s. and tranum-jensen. j. 1987 gram positive bacteria produce exotoxins that, at least in part, exhibit the unusual properties of superantigens. superantigens (sag) activate v/3 selectively t cells to produce lymphokines including i1-2 and tnf/lymphotoxin. systemic application of the sag staphylococcal enterotoxin b (seb) to d-galactosamine sensitized mice causes lethal shock within hours. seb reactive v/38 + t cells accumulate within 90-120 min mrna for the ii-2, i1-4, tnfai~ and ifn-3,. parallel in time high concentrations of bloodborne i1-2 and tnf are noted. anti tnf t~//3 neutralizing mab protect mice against seb induced t cell dependent lethal shock thus indicating a key role of tnf in effecting lethal toxicity. within 12-18 h distinct levels of tolerance to seb become discernable on ligand reactive v/38 t ceils, dependent of the dose of seb used. these levels include anergy, t cell receptor downregulation, loss of cd2, cd4, cd8 accessory molecules and programmed cell death. m.freudenberg, y.yaegashi, p.nielsen, c.galanos. the sensitivity towards endotoxin (lps) is genetically determined, however it may be increased under different experimental conditions. these include treatment with hepatotoxic agents such as d-galactosamine, and with live (infection) or killed bacteria. it is well established now that d-galactosamine sensitizes to lps by increasing the susceptibility of the host to toxic activity of lps-induced tnf~. sensitization by bacteria, especially by gram-negative once is of especial interest because it implies that infecting microorganisms not only produce lps, but als0 sensitize the host to its lethal activity. sensitization to lps by bacteria proceeds in all lps-sensitive (lps n) mouse strains that have been investigated so far. it also proceeds in lps-resistant (lps d) c3h/hej mice. the absence of sensitization to lps in lps d c57bl/10sccr mice was identified as being due to their inability to produce interferon-7 (ifn?). administration of exogenous ifn? to these mice conferred sensitivity to lps. conversely, administration of anti-ifn? to lpsn; mice inhibited the development of sensitization'by bacteria. it may be concluded therefore that ifn7 is the mediator of sensitization to lps by bacterial infection. the ifn? defect of c57bl/10sccr mice concerns only the ifn7 response to bacteria, since the response to the t-cell mitogen con a and to cd3 monoclonal antibodies was not impaired. the ifn?producing cells themselves were shown to be intact. on closer investigation the impaired ifn? response was identified as the result of a defective accessory function of macrophages. macrophages of c57bl/10sccr mice are incapable of producing interferon-~ (ifn~) which we could show to be obligatory for the production of ifn? in response to bacteria. although antibiotics are required to clear bacteremia, they do not reverse evolving shock, because endotoxin free in the bloodstream or exposed on the surface of circulating bacteria continues to activate the production of inflammatory mediators, furthermore, antibiotics have been shown to induce release of endotoxin from gram-negative bacteria during the treatment of severe infection. in an in-vitro study, using live escherichia coil, we have observed the release of endotoxin upon treatment with several antibiotics like cefuroxim, imipenem, ceftazidime and aztreonam. incubation with imipenem or ceftazidime induced an early lyeis and a mild rise in endotoxin levels, whereas incubation with cefuroxime or aztreonam resulted in the formation of filaments with a late but dramatic rise in endotoxin levels. in a second study we studied the influence of different antibiotics on the tnf-= production of e.coli stimulated human monocytes. we found again great differences in the production of this important cytokine among the different antibiotics according to their capacity to induce liberation of endotoxin. these differences in the amount of endotoxin release are probably caused by penicillin-binding-proteins (pbp) specificities of the antibiotics with resultant differential morphological changes, in a rat sepsis model treatment with imipenem or ceftazidime resulted in a transient increase in il-6 levels, whereas treatment with aztreonam resulted in progressively increasing levels of il-6 and a significant increase in mortality. the increased mortality in pbp-3 specific aztreonam treated rats might have been the result of filament formation in the abdominal cavity of the septic rats. the endotoxin sequestered at local sites may account for the precipitation of gram-negative shock. finally, we've also demonstrated that in patients under treatment for septic shock endotoxin release can be related to the administration of antibiotics. carbapenem-and cephalosporin-lnduced release of lps from smooth and rough pseudomonas aeruginosa: in-vivo relevance j. jackson and h. kropp, merck research laboratories, rahway, nj b-lactam (cell-wall active) antibiotics are generally deemed the class of antibiotics most responsible for the release of free endotoxin (lps) from gram-negative bacteria due to their cell lytic actions although all antibiotic classes studied thus far induce release of endotoxin. we have recently shown in-vitro that antibiotics within the b-lactam class (i.e. imipenem , meropenem and ceftazidime , with equivalent mics) differ in their propensities to release excessive amounts of lps from both smooth-(s-) and rough-(r-) lps laboratory and clinical and antibiotic-sensitive and -resistant p. aeruginosa isolates and that lps release is independent of cell lysis. additionally, variations in the induction of endotoxin release is antibiotic concentration dependant and correlate with antibiotic penicillin-binding protein (pbp) affinities which result in morphological changes. these studies also show that lps differences measured via chromogenic lps (c-lal) assay correlate with lps lethality in lps-sensitive mice. release of lps was compared to changes in cfus, cell mass, morphology and antibiotic pbp-specificity. corresponding in-vivo studies in cd1 mice against s-lps p. aeruginosa isolates show that, despite equivalent in-vitro protection, antibiotic efficacy in mice differ as much as 100-to lo00-fold. to establish a possible in-vivo role for antibiotic-induced release of free lps we compared antibiotic efficacy results in mice challenged with virulent parent s-lps and its relatively avirulent r-lps mutant (lacking only its 0 side chain) p. aeruginosa isolates. results show the relevance of quantity and physiochemical composition (bioreactivity) of free lps to virulence and in-vivo antibiotic efficacy. in other in-vivo studies chemical agents that destroy cells (m~) which mediate lps lethality in-vivo were used to pretreat mice prior to antibiotic therapy and bacterial challenge with wild type pseudomonas. we conclude that circumstances in which antibiotic-induced filamentation occurs are also conditions that yield excessive lps release, the in-vivo effects of which are shown through the requirement of larger amounts of antibiotic to afford equivalent protection. bacterial endotoxins have been reported to play a significant role in the pathogenesis of gram negative sepsis by their interaction with, and stimulation of, host inflammatory mediator cells to produce cytokines, biologically active lipid intermediates, and procoagulant activity (tfa). although both microbe-associated and free endotoxin are capable of stimulating mediator production, studies from our laboratory suggest that free endotoxin may be the more potent stimulus for tnf and tfa. further, our studies suggest that the majority of the proinflammatory activity released from antibiotictreated e. coli coisolates with lps by two different fractionation techniques. to assess whether actions of endotoxin directly contribute to lethality in gram negative sepsis, an experimental model was developed in which mice were made hypersusceptihle to the lethal effects of endotoxin by treatment with d-galactosamine (dgaln). challenge of cf-1 dgaln-treated mice with e. coli olll:b4 resulted in an lds0 of approximately 2.5 x 104 cfu. resolution of the peritonitis and bacteremia by treatment with cell wall-active antibiotics resulted in an increase in the ldso. ceftazidime and imipenem, which differ in their mechanism of action and in their capacity to effect endotoxin release in vitro, also differed in their in vivo capacity to provide chemotherapeutic protection (3 fold vs 8 fold respectively, p< .01). parallel studies with endotoxin hyporesponsive c3h/hej mice indicate significantly greater levels of protection with imipenem (> 80 fold vs saline controls). collectively these data suggest that endotoxin may contribute directly to the pathogenesis of experimental gram negative sepsis. bacterial lipopolysaccharides (lps) are the endotoxins of gram-negative bacteria and represent their major surface antigens. lps is made up of three chemically, biologically and genetically disctinct regions, i.e, the o-chain, the core region and the lipid a moiety whereby the latter represents the endotoxic center. it is our current understanding that lps is responsible for many of the pathophysiological events observed during gramnegative infections and that one of the major mechanisms leading to shock and death is the lps-induced activation of macrophages resulting in the production and release of lipid and peptide mediators, among which tumor necrosis factor seems to be the most important. therefore, in the fight against the lethal outcome of gram-negative infections, modern strategies, in addition to antibiotic treatment, aim at i) the neutralization of tumor necrosis factor, ii) the inhibition of the production of tumor necrosis factor or iii) the neutralization of the activation potential of lps for macrophages by monoclonal, preferably human antibodies. the latter approach, to be effective against a broad spectrum of gram-negatives, must be directed against common structures of lps (lipid a and core region). the molecular basis of this approach and the controversy in this field will be discussed. passive immunotherapy has been used since 1893, when von behring described the administration of immune horse serum to treat a patient with diphteria infection. even if this therapy was sometimes successful in bacterial infections, it has been largely replaced by antibiotics. however, antibiotics have their limitations, especially in critically-ill patients. to improve outcome, adjunctive therapies such as immunotherapy with polyclonal and monoclonal antibodies particularly against endotoxin are again considered. the role of humoral immunity in host defenses against bacterial infections is weu known. for instance, tile importance of antibodies in the defense against gramnegative infections has been established clinically by studies relating the outcome of patients with gram-negative bacteremia to tilers of antibodies directed at the offending pathogens at the onset ofbacteremia (mccabe 1972; pollack 1979) . ever since we know the role of endotoxins in the pathophysiology of sepsis, antibodies against the s-and r-lps have also been detected in sepsis patients. the aim of the administration of iv/g to the sepsis patient is as follows: 1 ) enhancing of opsonization and phagocytosis(antibactericidai activity) 2) synergistic effects with [3-1actam antibiotics 3) neutralization of endotoxin, the main pathogenic mediator of gram-negative sepsis 4) modulation and/or inhibition of cytokine release the enhancement of opsonic-and phagocytic-activity especially with igg via fc and c3 receptors has been well documented. monoclonal antiendotoxin antibodies, proven in clinical studies, do not appear to neutralize endotoxin in vitro and are not reproducibly protective in animal models of sepsis. also they can not suppress endotoxin-induced tnf-~, il-6 release in mice (baumgartner 1990, corriveau and danner 1993) . in conlrast, recent studies of a polyclonal immunoglobulin preparation, containing high levels of antibodies against gram-negative bacteria and their o-antigen of lps in igg, igm and iga classes (pentaglobin®) provide evidence to neutralize endotoxin. this effect is demonstrated in vitro (berger 1990 (berger ,1993 , in animal models (stephan 1985 , berger 1993 and also in prospective, randomized, controlled clinical trials (schedel 1991 , poynton 1992 , behre 1992 . furthermore mortali b' was reduced statistically in patients with septic shock and endotoxemia by using this preparation, as has been demonstrated by sehedel. anti-core lps monoolonal antibodies: binding specificity and biological properties f.e. di padova, r. barclay, e.th. rietschel. bacterial lps and cytokines are responsible for the pathological processes of gram-sepsis and are suitable targets for therapeutic interventions. chemical characterization and structural analysis of different lps have revealed common features. the inner core region of lps shows a high degree of similarity among e. coli, salmonella and shigella. among a large number of broadly cross-reactive murine anti-core lps mab one of these igg2ak) has been selected and chimerized into a human igglk (sdz 219-800). in elisa and in immunoblots on purified lps both sdz 219-800 and wni 222-5 show a strong reactivity with all smooth lps from e. coli and salmonella. reactivity with all the known complete core structures from e. coli and salmonella (ra) is evident. reactivity with re structures or free lipid a is not observed. this mab cressreacts with all clinical e. coli isolates from blood, urine and feces and with other enterobacteriaceae. sdz 219-800 and wni 222-5 have biological activity as they inhibit the lal assay and the secretion of monokines (il-6 and tnf) by mouse and human macrophages. moreover, sdz 219-800 and wni 222-5 inhibit the release of il-6 and tnf in vivo. in vivo sdz 219-800 as well as wni 222-5 neutralize the pyrogenic activity of e. coli lps and protect mice from lethality in d-gain-sensitized mice. the possibility to use wni 222-5 as a capture antibodies in the immunolimulus assay opens the possibility to differentiate the origin of the lps in patients with endotoxemia. franco di padova, sandoz pharma ag, ch4002 basel, $chweiz $7 presentation of lps to cd14 by lps binding protein peter s. tobias, julie gegner, katrin soldau, lois kline, loren hatlen, douglas mintz, and richard j. ulevitch. the activation of myeloid cells by lipopolysaccharides (lps) has been shown to require the serum glycoprotein lps binding protein (lbp) and binding of lps to membrane bound cd14 (mcd14). other cells such as human umbilical vein endothelial cells (huvec), smooth muscle cells, and some epithelial cells, which do not express mcd14 but nevertheless respond to lps in the presence of serum, have receptors for complexes of lps with the soluble form of cd14 (scd14). these complexes of lps with scd14 are only formed efficiently in the presence of lbp. we have begun to characterise the mechanisms by which lbp enables lps to bind to cd14, either soluble or membrane bound. with the use of fluorophore and radiolabelled reagents we have developed procedures for quantitative measurement of the association of lps with lbp and of lps-lbp complexes with cd14. these results show that the delivery of lps to scd14 is catalysed by lbp, i.e., lbp is not included with the lps-scd14 complex. in contrast, on the surface of cells, lbp does not dissociate from the cells after lps binds to mcd14. the kinetics, equilibria and stoichiometry of these reactions will be discussed in the context of models for cellular activation by lps and cellular uptake of lps. supported by nltt grants gm37696, ai32021, ai15136, gm08172, and assistance from the pharmaceutical research institute of johnson and johnson. the scripps research institute, imm-12, 10666 n. torrey pines rd. la jolla, ca usa 92037. modulation of endotoxin-induced cytokine production by lps partial structures h.-d. flad, h. loppnow, t. mattern, and a.j. ulmer department of immunology and cell biology, forschungsinstitut borstel, d-23845 borstel lipid a constitutes the active moiety of endotoxin (lps) of gramnegative bacteria. it activates mononuclear phagocytes to produce cytokines, such as tnf, i1_-1, and il-6, which are the major mediators of the endotoxic effect of lps in vivo. lipid a precursor la (synthetic compound 406) does not induce cytokines, but is able to specifically antagonize lps-or lipid a-induced mediator production in human mononuclear cells, vascular endothelial cells, and smooth muscle cells. furthermore, we present evidence for the first time that t-lymphocytes proliferate in response to lps and express mrna for interleukin-2 and interferon-~ and that these responses are also antagonized by synthetic lipid a precursor la. when comparing the agonistic and antagonistic activity of lipid a and different partial structures at the functional and binding level, the number and length of the fatty acids and the number of phosphoryl groups were pound to be of crucial importance. unexpectedly, lipid a precursor la, although biologically inactive, turned out to be both the most potent antagonist and competitor in inhibiting the binding of lps. taken together, our results provide evidence for a model in which lipid a partial structures compete with lps for specific cell surface receptor(s). in this sense, biologically inactive lipid a analogues may be good candidates as therapeutic agents for the prevention of gram-negative septic shock. two mammalian lipid a-binding proteins have been identified that are believed to have important roles in mediating the host response to endotoxin: lipopolysaccharide-binding protein (lbp) and bactericidal/ permeability-increasing protein (bpi). human lbp shares a 45% amino acid sequence identity with human bpi. despite the sequence homology, the two lipid a-binding proteins have very different functional activities. lbp is an acute phase serum protein that markedly potentiates the proinfiammatory host response to gram-negative infection by a mechanism which involves binding of the lbp-lps complex to cd14 receptors on monocytes, neutrophils and endothelial cells. in contrast, bpi is a neutrophil granule protein with potent bactericidal and lps-neutralizing activities. the divergent functional properties of these two lps-bindlng proteins can be explained by the inability of bpi-lps complexes to bind to cell-surface cd14 receptors. a recombinant protein (rbpi23), corresponding to the amino terminal 23kd fragment of human bpi, has been shown to retain the potent biological activities of the hdlo protein and may represent a novel therapeutic agent for the treatment of gram-negative infections, sepsis and endotoxemia. for therapeutic effectiveness in many clinical situations, rbpi23 will have to successfully compete with relatively high serum levels of lbp (5-60 ~g/mi) for binding to endotoxin and gram-negative bacteria. to evaluate this issue, experiments were conducted to compare the relative binding affinities of rbpi23 and human recombinant lbp (rlbp) for lipid a. the binding of both proteins to iipid a was specific and saturable with apparent kd's of 2.6 nm for rbpi23 and 58 nm for rlbp. in a competition assay format rbpi23 was approximately 75-fold more potent than rlbp in inhibiting the binding of nsi-rlbp to lipid a. these results demonstrate that rbpi23 has a significantly higher affinity for endotoxin than does rlbp and may explain the potent inhibitory activity of low concentrations of rbpi23 in a variety of in vitro functional assays for lps activation of cells despite the presence of high lbp levels. for example, rbpi23 at 0.2 ~tg/mi was able to totally inhibit lps-induced tnf release from monocytes despite a 100-fold weight excess of rlbp over rbpi23. and for heparin binding. three separate domains which inhibit the lal reaction to lps and bind to heparin were identified in amino acid regions 17-45, 65-99 and 142-169 . a single synthetic peptide (85-99) was bactericidal. these results suggest that rbpi23 contains three separate functional domains which may contribute to its high affmity interaction with gram-negative bacteria and heparin. the individual activity of each domain and the cooperative interaction among domains provide the basis for developing rbpi23 analogues with increased biologic efficacy. a considerable body of experimental data has accumulated implicating tumour necrosis factor (tnf) as a principal mediator of the pathophysiological features of septic shock. these data prompted the development of clinical strategies designed to limit excess (inappropriate) tnf production. monoclonoal antibodies (mobs) were developed and a phase ii dose escalation trial in 80 patients confirmed that the mab was safe, and suggested that it was having a beneficial effect on certain parameters. preliminary results of a large phase iii study indicated that (a) the mob was safe; (b) that it was of no discernible benefit in non-shocked patients; (c) that it reduced mortality in shocked patients, especially during the first 3 days. an alternative strategy was to take advantage of the high binding affinity of soluble receptors for tnf (stnfr). stnfr-iggfc constructs were made for both the p55 and p75 receptors. both were effective in animal models of lps challenge, but when a clinical trial was done with the p75 stnfr-fc there was unexpected mortality in the treated arm. using an animal model of live e.coli sepsis, we have shown that this may have been due to the release of bound tnf from the construct. plasma enhances while bpi inhibits lps-induced cytokine production from peripheral blood mononuclear cells (pbmc). pseudomonas species produce cytokine-inducing substances which are different from lps as indicated by the fact that polymyxin b blocks only 40% of the cytokine-inducing activity of these pyrogens. we now tested the effect of plasma and bpi on the il-1[3-inducing activity of pseudomonas maltophilia -derived pyrogens (pmp). bacteria were cultured to the log phase and filtered (300kd) to obtain prop. dilutions of pmp or lps were added to pbmc alone or to pbmc in 5% plasma +/-bpi (500 ng/ml). pbmc were incubated for 24 hours at 37°c and total il-i~ was measured by ria. results: il-i[~ in ng/ml (n=7, mear~+sem, *p<0.05 vs control). 0 control 0.1_+0 + bpi 0.1+0 5% plas. 0.2_+0 + bpi 0.2_+0 pmp (ng/ml) lps (ng/ml) 60 600 3000 1 10 3.2_+1 8.0_+1 16.3_+3 1.2_+. 2 5.2+.6 0.2_+.1 3.3_+.9" 13_+3" 0.1_+0" 0.1_+0" 2.2_+. 5 5.7+1 14_+2 5_+.6* 14_+2" 0.6+. 4" 5.3+1 15-+2 0.1-+0" 0.5_+.3" cba, c57bl/6, balb/c, akr, dba, swiss mice, guinea pigs, rabbits have been used in research work. the toxicity, immunogenicity, mitogenic and immunomodulating activity of lps have been studied. the possibility of reduction of the toxic activity of lps on macroorganism by bioglycansimmunomodulators obtained from sea invertebrates anymals (crenomytilus grayanus, stromhus gigas) have been investigated too. lps has been shown to induce specific antibody response of laboratory animals. cba mice are high responsive to lps. lps stimulates humoral immune response of mice to tdependent and t-independent antigens and suppresses intensity of the delayed hypersensitivity. the small doses of lps stimulate functional activity of macrophages, the large doses of lps -decrease one and show the cytotoxic effect. the bioglycans enhance the resistance of mice to the lethal effect of lads and provide protection 59-75% of mice. one opens possibility to use of bioglicans for reduction of toxinemia in generalizated forms of pseudotuberculosis. thus, lps from y.pseudotuberculosis is immunogen and immunomodulator wich has influence on humoral and cellular factors of immunity and plays the important role in immunopathogenesis of infection. endotoxaemia is implicated in the pathophysiology of obstructive jaundice. the lirnulus lysate (lal) assay is the gold standard method for measuring endotoxin concentrations, but inherent biochemical and technical problems limit the usefulness of this assay. the endocab elisa is a novel assay which measures endogenous antibody (igg) to the inner core region of circulating endotoxins (acga). objectives we evaluated the significance of endotoxaemia in biliary obstruction using the endocab assay and subsequently the specificity of the humoral response to endotoxin compared with an exogenous antigenic challenge [tetamls toxoid (tt) ]. materials and methods in experiment i three groups of male wistar rats (250-300g) were studied [no operation (n=39) , sham operation (n=40), and bile duct ligation for 21 days (bdl)(n=50)]. plasma was collected and assayed for bilirubin, endntoxin(lal) and acga(endocab). in experiment ii 30 rats were actively immunised with tetanus toxoid ('it) and then randomised to have no op(n=8), sham op(n=8) or bdl(n=i4). blood was taken at this time (to) and 21 days later(t20 at sacrifice for acga concentrationslendocab] and igg produced to tt(ttab) [elisa] . antibody concentrations are expressed as % increase from control values.results in bdl rats, acga concentrations were significantly increased compared with controlslp<0.001, mann-whitney]. endotoxin concentrations were sporadically elevated in the jaundiced rats but the rise was not significant. in experiment [i there was no difference between the acga or ttab concentrations in the fllree groups at to, bdl rats had a significant rise in acga concentrations by t21 [p<0,001,paired t-test] and humoral response to tt was significantly impaired in bdl rats compared with control groupslp<0.05, paired ttest data plasma endotoxin was measured by means of an endotoxinspecific endospecy test after pretreatment of the plasma with a new perchloric acid method that we developed. the normal value of plasma endotoxin is less than 9.8 pglml. polymyxin b was administered at a dose of 12,500 u every 6 hours. plasma endotoxin rapidly decreased to the normal range in 40 of the 42 patients. body temperature fall significantly. apache ii scores were also significantly improved. tumor necrosis factor-o~ and interleukin 6 decreased in survivors, while in high values tended to persist in patients died. no side effects were observed in any of the patients. in conclusion, intramuscular injection of minute of polymyxin b was useful in the treatment of endotoxemia. 19-1 uchimaru, morioka 020, japan. l e v a n t g r a m n e g a t i v organisms. m e t h o d s : u n d e r general anesthesia, 12 n o r w e g i a n b r e d landrace pigs (17-30 kg) of either sex, 6 pr group, u n d e r w e n t t r a c h e o s t o m y a n d w e r e v e n t i l a t e d on a 50/50 air a n d o x y g e n m i x t u r e a i m e d at m a i n t a i n i n g a n o r m a l p h a n d a isocapnic level. ventilation w a s not readjusted d u r i n g the observation period. the anesthesia w a s k e t a m i n e 0.6 m g / k g h a n d d i a z e p a m 2.4 m g / k g h i n t r a v e n o u s l y . h e m o d y n a m i c m o n i t o r i n g of m e a n aorta, p u l m o n a r y artery, central v e n o u s a n d p u l m o n a r y capillary w e d g e pressures w a s p e r f o r m e d w i t h a 5f s w a n -g a n z catheter a n d an aorta catheter. a continous infusion of r i n g e r ' s acetate ( 1 0 m l / k g h ) w a s g i v e n intravenously. w h e n stabilised, the a n i m a l s w e r e g i v e n 2.5 x l09 cfu of e colt intraperitoneally as a bolus in 100ml saline, the a n t i b o d y g r o u p received in a d d i t i o n 5 m g / k g e5 a n t i e n d o t o x i n i n t r a v e n o u s l y over 1 h o u r via a n infusion p u m p at the start of the observation period. the a n i m a l s w e r e observed for 6 hours. results : a t 5 a n d 6 hours, the o x y g e n c o n s u m p t i o n increased by 30 % in the a n t i b o d y treated g r o u p w h e r e a s there w a s a significant fall of 30 % in the sepsis group. in the a n t i b o d y group, the arterial p h a n d the cardiac index were also significantly h i g h e r at the s a m e p o i n t s in time. there w a s no significant difference in arterial po2. in severe bacterial infections it would be beneficial to neutralize the plasma endotoxin content with complex forming compounds. the phenothiazines are able to form complexes with endoto×in and the existence of these complexes were already shown in differential speetrophotometry and animal experiments, however, the mechanism of partial neutralization was not clarified. therefore some representative phenothiazines and structurally related compounds were tested for anti-endotoxin activity. the endotoxin neutralizinb effects of several benzophenothiazines were investigated in differential speotrophotemetry, tnf induction and in the conventional limulus test. in animal experiments some beneficial effect of complex forming compounds was found. the benzophenothiazines were not able to inactivate the biological effect of endotoxin in the limulus test. the recent findings indicates that a multifocal effect can be responsible for "anti-endotoxin action in vivo". effects of tnf inducing effect of endotoxin in leukocytes and bypotensiv action in experimental animals were reduced by some phenothiazine derivatives. monophosphoril lipid a was without effect. of microbiology, albert szemt-gydrbyi medical university, odm t~r lo, h-6720 szeged~ hunbary 46 involvement of streptococcus pyogenes erythrogenic toxins in the induction oflstreptococcal toxic shock syndrome 3 heide mgller-alou~* , joseph e. alouf , die [er gerlach , ~atherine fitting., and jean-marc ca~aillon . unit6 des toxines microbiennes and "unit6 d'immuno-allergie, institut pasteur, 28, rue du docteur roux -75015 paris (france) ; institut f~r experimentelle mikrobiologie, jena (germany). superantigen erythrogenic toxin a (eta) is thought to be involved in toxic shock syndrome in humans by inducing massive release of cytokines by patient immune cells. the cytokineinducing capacity of eta w~:s £:ompa~ed to that of lps, a gram-negative bacterial cell wall component. eta elicited weak production of il-1 d and ~, tnf ~ and il-6 in purified human monocytes whereas lps stimulated the production of high amounts of these cytokines. in the presence of t cells, eta elicited the production of significant amounts of il-i~, il-i~, il-6 and il-8. however, the most preponderant cytokine was tnf~, which peaked at i0 ng/ml after stimulation with i0 ~g eta. comparable amounts of tnfd (ca 4 ng) were induced by 0.i ~g eta and 0.i ~g lp$. in contrast to lps, eta was a strong inducer of tnf~ which was produced only in marginal amounts by lps. these results suggest that the septic shock induced by gramnegative bacteria (lps) and by gram-positive bacteria {extracellular superantigens) follows different pathogenic pathways. lps-induced shock is mainly mediated by monocytes and monocyte-produced cytokines (il-i and tnf). the eta-induced shock is mediated by t-cells or depends on t cell help for the production of monocyte-liberated cytokines. production of t cell cytokines such as tnf~ and interferon 7 in addition to the other cytokines contribute very likely to the severity of the toxic-shock resulting from s. auzeus and s. pyogenes infections in humans. the present study was utidertakc~l to cvalu~tlc the effect of soluble chemically modified giucan during septic shock. carboxylnethyl-b-i,3-glucan (ram 120000) was injected twice 48 and 24 h before the shock i.v. in a dose of 50 ing/kg. shock was induced in u~?esthetizcd (sodikm~. l)mntobarbital) rats by i.v. injection of endotoxin of escherichia colli 0127 bs, 5 mg/kg. aiiofcmg pretreated ruts survived during first 24haher ¢ndotoxine, while in controi shock group the lethality was 90%. the concentration of ~col)terin in serum was significantly elevated 24 hafterthc second cmginjection (appare~tly 80 % if compare with the control rats), but didu't chartged 60 rain and 1 s0 rain after endotoxin injectjom cardiac output in cmogroup was higher a* the i20 and 180 min after endotoxine onset ( 8i % trod 72~, respectively of initial level) than in the control shock group (64 % and 52% at the same time). pretreatment of rals with soh~ble giucan w~ts associated with beneficial effects o~ the hepatic c~ergy $ia[tls after 3 h after challenge of endotoxiae: the tissue level of lactale was ahnost twice lower than in the control ruts, me~mthne the tissue atf in cmg pretreated group was higher at 40 %. twice injected macrophage stimuhttor soluble glucan can prevent the endotoxic shock, and extremely ir~creased survival rate after endotoxine injection. the national committee of surgical infections of the spanish association of surgeons have produced a computer program for the collection and analysis of information on surgical infections. the program is suitable for ibm compatible hard disk personal computers and works through the ms-dos system. the main menu is called up on the screen when the operating disk has been installed; it reads as follows: i. new record; 2. modify records; 3. erase records; 4. searches; 5. reports; 6. configure; o. ouit. if you ask fdr a new record the screen will prompt you to enter the number of case, record number, hospital, age and sex. the next screen will come up and the words "topographic diagnosis" will flash. a menu of 34 areas or organs will be displayed. then, the words "type of pathology" (inflammatory, neoplastic, traumatic and other). days of postoperative period. type of surgery (programmed and emergency). type of operation (clean, clean contaminated, contaminated and dirty). duration of surgery. this is followed by "order of operation" and the "type of anaesthesia (general, regional or local). you are then required to supply the "diagnostic code of who" (icd9) and the "procedure code of who. analytic and concurrent illnesses (total proteins, albumin, haemoglobin, haematocrit, leucocytes, red corpuscles, glucose and bilirubin). the next screen asks for "risk factors" (obesity, uraemia, neoplasia, malnutrition, urinary catheter, distant infection, artificial valve, immunosuppressive drugs, over 65 years and anergy. this is followed by a screen headed "postoperative complications". "evolution" (the questions asked are drainage, systemic antibiotics, and on each ocasion a choice of 31 antibiotics is displayed), local antiseptics, reoperation, etc. under "microhiology" is a choice of 37 organisms and the chance of identifyin 3 organisms. finally, "sepsis score". our recent work had shown that renshen-fuzi-chaihu mixture could increase the survival rate in experimented study. the purpose of this study was to determine the effect of combined administration of renshen-fuzi-chaihu mixtuer and antibitics (sa) in patients with septic shock. the result showed that, in sa group (40 cases), the total effective rate was 87,5%, in the contral group (combined administration of gentamycin and dexamethasone, 25 cases) the total effective rate was 84%. however the obviously effective rate in sa group 70% was significantly higher than in contral group 44% (p<o.os). sa group appeared to be more obvious than the contral group in raising arterial pressure, recovering consciousness and pulse, improving cold lilmbs and reducing fever (p<o.o5). it was found that sa could inhibit the pathogeneses changes of septic shock such as the decreases of superotide dismutase (sod) and glutathione peroxidase (gsh-px) in erythrocytes and the increases of plasma free hemoglobin (phb), plasma malondialdehyde (mda) and 13-glucuronidase (13-gc) occured in septic shock. sa had stronger effect than gd in inhibiting the changes mentioned above. hence, these results suggest that sa has beneficial effect in the treatment of septic shock. sa could diminish the decrease of the antioxdase activities and inhibit lipid peroxidization and stabilize cellmembrance. this may be one of the principal mechanisms of the beneficial effect of sa in the treatment of septic shock. donna l. lehman, heather a. childers, irshad h. chaudry and alfred ayala. the thymus is thought to be a privileged sight for tcell generation. here the t-lymphocytes are either selected for their potential to recognize and react competently to foreign antigens or de-selected, due to their potential to react to auto-antigens, de-selected. tcells with this latter capacity are thought to be deselected through a process referred to as programmed cell death or apoptosis. while it is known that thymic apoptosis is elevated by a variety of stressers associated with infection and inflammation, little or no information is available concerning its presence in polymicrobial sepsis. it was, therefore, the aim of this study to determine whether thymocytes exhibit increased apoptosis during sepsis. to assess this, thymocytes were harvested from c3h/hen mice at i and 24 h following cecal ligation and puncture (clp; to induce sepsis) or sham-clp (sham). thymic yields were assessed and the extent of apetosis determined by staining the cells with propidium iodide (a dna intercalating dye) for facs analysis or by examining the extracted dna electrophoretically for the endogenous endonuclease activity the results (n=6/grp) indicate that at i h post-clp there was no marked changes in any of these parameters. however, at 24 h the thymic cell yield from clp mice was significantly decreased (sham:3.6!0.2 x l06 vs clp:0.3i0.1 x 106* cells; *,p<0.05), the % of cells apoptotic by facs analysis increased from 1.7i0.1% in sham to 8.7±3.2%* in clp, and the septic mouse genomic dna exhibited dna degradation these results indicate that clp, but not simple laparotomy, induces marked thymic apetosis, which may contribute to the lymphocytic immune deficiency seen in these mice the purpose of this retrospective study was to evaluate effectiveness of irrigation to treat septic knee joints from longterm results. from 1982 to 1992 31 patients with septic knee joints have been treated. treatment in acute cases started with asp#at[on of the effusion to evaluate for cultures. without awaiting the result treatment was continued with immediate joint irrigation under arthroscopic control. three redon tubes ( ch 16 ) were introduced for continuous irrigation and for intermittent distention of the joint cavity. in case of chronic joint infection additionally all foreign and synthetic material was removed. systemic antibiotics were given. patients could be re -examined with a mean follow-up of 6 years. re-examination showed that immediate arthroscopic lavage started at the onset of septic sings had best results (less signs of osteoarthr[tis, less synovitis, less pain while loading and less range of motion loss ). 21 patients (80%) out of 26 patients with s tre_=.tsd acute se.~tic knee joint had excellent functional recovery, whereas all 5 patients with chronic septic knee joints showed poor results. additionally, in 3 of these 5 patients with chronic septic knee joint after irrigation a further procedure was necessary due to recurrent septic knee joint. the concept that bacteria can translocate across the intestinal mucosa under a variety of circumstances is well established. however, due to the inherent limitations of in vivo studies, the cellular mechanisms underlying the process of bacterial translocation (bt) across the epithelial barrier are poorly understood. this is especially true in those circumstances in which there is no merphologic evidence of mucosal injury. consequently, we have utilized an in vitro cell culture model system to investigate the cellular events involved in the translocation process. in this model system, a polarized monolayer of intestinal cells (caco-2 cell line) is grown on a 3 micron filter in a two compartment system. after tight junction integrity is established, bacteria are placed in the apical surface chamber and bt across the caco-2 monolayer is measured by culturing the basal chamber. the results of our studies utilizing the caco-2 system indicates that; l) several strains of non-pathogenic bacteria will translocate across the caco-2 monolayer in a time-dependent and dose-dependent fashion. however, the incidence and magnitude of bt are highest for those strains of bacteria (gram-negative enterics) that are most commonly cultured in vivo. 2) caco-2 cells are actively involved in the transcellular passage of bacteria across the caco-2 monolayer, since inhibition of caco-2 microtubule or microfilament function decreases the magnitude of st. 3) lectin but not integrin receptors are involved in bacterial translocation of e. col~ across the caco-2 cell monolayer. 4) caco-2 cells have the ability to ingest and kill certain strains of bacteria. the mechanisms involved in caco-2 bactericidal activity appear similar to that of pmns to the extent that both are oxidant but not nitric oxide mediated. these in vitro studies suggest that caco-2 cells can function as "nonprofessional" phagocytes and that both bacteria and enterocytes are involved in the translecation phenomenon. bacterial translocation due to gut barrier dysfunction is considered to be a major cause of septic complications and multiple organ failure following trauma, burn injury, hypoxia, shock and shock-like states. the invasion of bacteria into the circulation from the gastrointestinal tract or even from other sources, however, should not necessarily result in systemic infection or organ cohinisation, as long as the humoral and cellular defense mechanisms are not impaired. thus, a reduced res clearance capacity and hepatic clearance function of bacteria and toxins can be observed under the same conditions which enable bacterial translocation from the gut. in order to evaluate the influence of circulatory, metabolic and humorul disorders on the elimination of bacteria out of the blood circulation, a series of experiments were performed in anesthetized and ventilated rabbits which received defined numbers (1.3 x 108 colony-furming units) of escherichia coli as a bolus injection. in unaffected control animals the intravenously injected bacteria are eliminated about 99.9% within the first 15 minutes and are totaiy cleared within 90 minutes. 57% offue total cfu counted in the cultures of the organ homogenates were found in liver and spleen, whereas only very low numbers could be detected in the lungs and kidneys. systemic injury of the animals by hemorrhagic shock, endotoxinemia, hypoxia, coagulation disorder, systemic vasoconstriction by norepinephrine and other types of injury reduced the elimination rate of bacteria and changed the pattern and degree of their dissemination and tissue distribution. the bacterial clearance was delayed mostly in rabbits subjected to hypoxemic hypoxia in which about 100 cfu of viable e.eoli per ml blood could still be counted 3 hours following their injection. the number of viable bacteria was some ten powers higher in organs of hypoxic rabbits in comparison to control ardmais, and an excessive colonisation of the lungs and kidneys with 25% respectively 11% of the total cfu of the cultared organ homoganates was observed. unilateral iscbemia of the kidney prior to the intravenous injection of e.coli only moderately delayed the bacterial clearance, bowever, caused an intense eohinisation of the affected reperfused organ. in contrast to this, inhalation injury did not promote the accumulation of bacteria in the lungs. conclusion: shock, systemic and local affections reduce the clearance of bacteria from the blood circulation and enable their dissemination and accumulation in vital organs. the degree of this reduction and the organ pattern of the bacterial distribution varies with the type of injury. thus, these factors seem to be essentially involved whether or not bacterial translocation and multiple organ failure will follow the invasion of bacteria into the circulation as in the ease of gut barrier dysfunction. movement of enteric baaterla from gut to extralumeaal sites (bacterial tra~sloeation) m~y play a role ~. nraltipl~ orga~ failure. traumatic stress (shock, hffectiort, and im~lammation) and severe illness are common alateeedents. tnt~st~aal lleus is e common proximme causal factor. we have exanlined the hypothesis that bacterial ttanslecation from the gut in expe~me~tal paaerestitis is due to bacterial overgrowth as a cuas~ucnee of a deere~ae in intestinal transit. ne~rotit_.~g pancreatitls fbliowlng bile duct iigation in the opossum is associated with colonization of the pa,se ~re, aa by gut derived organisms or salmonella of biljary origin in infected animals (previously reported). to ti~rther define the mechanism of transloemdon in panereati* inflammation, pancteatitis was induced hy iigatlon of t~e lower bile duet distal to the pancreatic duets m the rat. intestinal transit, enteric bacteriology, and tromsloeatien of bacteria to mesanterie lymph nodes, blood stream and splanelmie organs wen deletmhaed at 24 (n~ 7), 4g (n ~ 10), and 96 (n~ 10) hours with the fallowing results: ly~testinai,..transfi". geometric mean of fluorescent marker -25 % at 24 hour~ with return to 51% of gut leugth (similar to ~m) at 96 hour~. bacterial overgrowt_hff -increase in total bsctetia ~ad gram negative rod.~ at 2,; hours with return to sham control at 96 hours (9 log cfu/g veraus 6 log cfu/g ia ileum). translocation to mesenteric nodes -71, 90, and 30 percent at 24, 48, and 9fi hours compared to 49b in sham controls (n=l of 26). conclusion -bacterial translocation following panetoatieobiliary duct ljgation induced panereatitls ha the rat is tighlly linked to intestinal trausit and bacterial overgrowth within the gut lumen. speclrjlation -the ilens in in/~arm~ation may be related to activation of eytokiaes and mediated by nitric oxide. preliminary evidence for this notion will be discussed. in a series of studies we investigated: a) the time course of bacterial translocation (bt), b) the kinetics of lps and cytokine appearance (tnf, il-6, soluble tnf receptor [stnf-r] ) in the circulation, and c) the role of lps and/or tnf with regard to haemorrhagic shock (hs) related pathophysiologie alterations and mortality in baboons and/or rats. method: baboons were subjected to femur fracture, soft tissue trauma (acute experiment), and hs (40 mmhg mean arterial pressure: map for 2-4 h terminated at an accumulated oxygen debt of 180-200 ml/kg followed by resuscitation). a chronic model of hs was set up by administration of zymosan activated plasma (zap) before and after hs but without trauma. hs was induced in rats by bleeding the animals to map of 30-35 mmhg 90-180 rain followed by resuscitation. results and conclusion: in rats, lps increased in the systemic circulation, plasma tnf levels were found to be significantly elevated at 90 min and were still markedly increased at 150 rain postshock. plasma tnf, il-6, and stnf-r levels increased already during the shock period in baboons, while tnf was not detectable. our data suggest that haemorrhagic shock may lead to early bt in the intestinal wall (at 30 min following resuscitation in rats subjected to hs for 90 min, similarly at 3 h after the end of oxygen debt period in baboons ) and transient access of gut-derived lps into the circulation (levels became significant at 120 min, while in baboons higher levels up to 300 pg/ml were found at the end of shock and 1 h after reinfusion, partially accompanied by bacteremia). in addition, since the treatment of rats with anti lps measures or anti tnf therapy significantly improved the 48-hour survival rate it can be assumed that endogenous lps and lps-induced mediator(s), in particular tnf, may lead to organ injury and mortality following haemorrhagic shock. alteration of the mesenteric blood flow and the consequent ischemia / reperfusion injury to the intestine appear to be one of the earliest events in the systemic inflammatory response following severe thermal injuries. the causative relationship between the subsequent disruption of the intestinal integrity and the potentiation of the translocation of indigenous bacteria and/or endotoxins to remote body organs and the systemic circulation has been deeumented in several studies. among other effects, endotoxemia solely or acting synergistically with thermal injuries has been shown to promote bacterial translocation. as these sequences continue without adequate intervention, multiple organ failure and eventually death may become inevitable. hence, the crucial need of treatment modalities with the capacity of modulating the intestinal blood flow and preventing the manifestations of these pathological incidents. although the underlying mechanisms of this process have not yet been fully elucidated, there is accumulating evidence that various interacting vasomediators are involved. the actions of these mediators can probably be modulated by either nonspecific or selective agents. among the nonspecific agents, the composition, volume and timing of the resuscitation fluids appear to play an important role in this process. nitropmsside, a nonspecific vasodilator, has been shown to prevent the decrease of the postbum mesentefic blood flow when selectively infused in the mesenteric artery. the nonspecificity of various vasodialators with their possible undesirable systemic effects emphasizes the importance of the identification and modulation of selective vasomniiators. thromboxane a2 (txa2) and angiotensin ii (a-ii) appear to be the primary mediators of the postburn vasoconstriction. both mediators were found to be elevated following thermal injuries. in a previous study, pretreatment with oky-046, a thromboxane synthetese inhibitor was found to attenuate the postbarn mesenteric vasoconstriction. in a bum/sepsis porcine model the efficacy of dup753, an a-11 receptor antagonist as a posthum treatment modality was evaluated. treatment with dup753 prevented the early posthum and the late posteodotoxin elevation in the mesenteric vascular resistance and subsequently abrogated the fall in the mesenteric blood flow. the incidence of burn & endotoxin induced bacterial translocation was significantly reduced by dup753 from 86% to 29% (p<0.05, fisher's exact test). the indigenous flora of the gastrointestinal tract exerts a potent influence on the developmental maturation of normal systemic immunologic responsiveness. moreover, both oral and intraportal administration of experimental antigen is associated with a systemic state of immunologic hyporesponsiveness, known as oral tolerance. since trauma and critical illness are associated with changes in the flora of the gut and with altered systemic immune reactivity, we have hypothesized that the interactions of an altered gut flora with immune tissues in the gut and liver play a role in the pathogenesis of the immunologic derangements of critical illness. prolonged feeding of two clinically relevant killed organisms-pseudomonas and candida-to a rat results in significant impairment of cutaneous delayed hypersensitivity (dh) reactivity. conversely, measures directed against gram negative bacterial overgrowth in experimental peritonitis result in partial restoration of impaired dh reactivity. to ascertain the role of the liver in the pathogenesis of these alterations, we studied the differential immunologic sequelae of portal versus systemic (ivc) infusion of killed bacteria. experimental infusion of live or killed gram negative organisms into the portal vein produces dh suppression in rive, and profound suppression of mitogen-driven lymphocyte proliferation in vitro; systemic administration of the same organism has no effect on these phenomena. suppression is tgf3-dependent, and mediated by a soluble factor released by fixed tissue macrophages of the lung and spleen. the suppressive influence appears to be mediated at the level of interactions between il-2 and its high affinity receptor. release of this factor can be induced by a second circulating factor present in the plasma of portally-, but not systemically-infused animals two hours after bacterial administration. these studies suggest that the release of inducible immunoregulatory factors from the liver, and possibly from the gut, may play a role in the pathogenesis of the profound derangements of systemic immune homeostasis that accompany trauma and critical illness. acad.hospital st. radboud,postbus 9101,6500 lib nijmegen introduction. the central question being tested in this study was whether liposome encapsulated dichloromethylene-diphosphonate (ci2mdp) mediated elimination of liver and splenic maorophages would influence zymosan induced systemic toxicity (st) and bacterial lranslocation (bt). materia|s and methods. 90 male swiss (icr) mice were used weighing 21-25 g elimination of liver and spleen macrophages was accomplished by intravenous injection of 200/11 suspension of ci2mdp-liposome suspension. control mice received an i.v. injection with 200 pt phosphate-buffered saline (pbs) . two days later all mice were challenged with an i.p. injection of zymosan suspended in paraffin (z) at a dosage of either 01 mg/g, 0.5 mg/g or 1.0 mg/g body weight (n=8 in each group), signs of st and bacterial translocation bt were measured 24 hours later the st was assessed by blindly grading the degree of lethargy, conjunctivitis, diarrhea, and ruffled fur of each mouse on a four point scale. bt to the mesenteric lymph nodes (mln) and systemic organs (liver, spleen, lung and blood) was tested by culturing on blood and macconkey's agar at the time of sacrifice sections of the distal ileum were taken for histology. an additional two control groups (n=6 in each group) were tested to verify that neither paraffin nor pbs or ci2mdpqiposome suspension alone induced bt or st. furthermore survival over a 12-day period was assessed in a control and macrophage-depleted group with a dose of 1d mg/g z. results. neither the paraffin nor ci2mdp-tiposome suspension induced bt or st the incidence of bacterial translocation to the mln was similar between macrophage depleted and control groups. however, macrophage depletion was associated with a significantly higher incidence of systemic spread of bacteria. data expressed as positive tissues/total tissues tested: 11/32 vs. 1/32 at 0.1 mg/g zymosan (p~o.01); 19132 vs. 10132 at 05 mg/g zymosan (p~q05); 23/32 vs. 11/32 st 1.0 mg/g zymosan (p~o.o1).the magnitude of bt however did not show any differences. although systemic bt was greater in the macrophage depleted groups, the systemic toxic response was significantly less at doses of 0.5 mg/g zymosan (3.14 -+ 0.69 vs. 7.13 -+ 3, p~o.01) and 1.0 mg/g zymosan (4.38 -+2.50 vs. 11.25 -+ 2.5, p<_.o.01) . the 12 day mortality after 1.0 mg/g zymosan was 27% in the control group and 0% in the macrophage depleted group (p=o.05). histology did not show any differences between the control and macrophage depleted groups. conclusion. the lethal and toxic effects of zymosan in this model appear to be more related to the excessive activation of macrophages than to the systemic spread of bacteria. bacterial translocation (bt) in hemorrhagic shock was studied using a porcine model. in this study three groups were randomized: one control (n=3) and two experimental (n=6 each). a portal vein catheter was placed and remained in situ 48 hrs. a femoral artery and vein catheter were in place 12 hrs. the arterial eaanula was used for monitoring mean arterial pressure (map) in all animals and to bleed the experimental animals and the venous cannula to repelfuse with ringer's lactate (rl) or autologous blood (ab). baseline samples and values were taken in this period. control animals were observed for a sham shock period of 6.5 hrs., given 2 more hours of anesthesia, and studied for further 48 hrs. 40% of the estimated blood volume was withdrawn from each experimental animal over 30 min. these animals were kept in shock for 6.5 hrs. at the end of the shock one group was perfused with rl and the other with ab. two hours later they were extubated and further studied for 48 hrs. samples from portal blood for cultures and measurement of leukotriene b4 (ltb4), prostaglandin e2 (pge2), and nitric oxide (no) metabolites were taken from all animals at intervals beyond baseline up to 48 hrs. after 48 hrs. under general anesthesia a second laparotomy was performed and samples for culture from mesenteric lymph nodes (min), liver and spleen were taken; as well as samples for histologic study with light and scanning electron microscopy were taken from jejunum, ileum, and colon. animals were then euthanized. results: significant shock was induced as indicated by map and arterial blood gases. significant bt to min was observed in all groups. no significant bt was observed in cultures from liver and spleen in any of the groups. increased values for ltb4, pge2 and no metabolites were found during the period of shock. these results suggest that the isehemic phenomenon triggers a significant inflammatory response, and that bt to min may be an epiphenomenon without apparent significant influence on the inflammatory response. objective: to study the potential effects and mechanisms of action of systemic administration of monoclonal antibodies anti-endotoxin (ha-1a) in a animal model of gut-origin sepsis. materials and methods: exoeriment a. balb/c mice were transfused with allogeneic blood (from c3hb-iej mice). five days post-transfusion the animals were gavaged with lxl09 viable escherichia coil and randomized into 3 groups (n=22/group) to receive a sham burn (sb group) or a 20% thermal injury immediately followed by the systemic administration, via a tail vein, of monoclonal antibodies (6 mg/kg) (ha-1a group) or a 20% burn injury and administration of equal volume of sterile saline (c group). all 3 groups were resuscitated by intraperitoneal injection of 0.3 ml of saline. the animal survival rate was observed for 10 days post-burn. experiment b, transfusion and burn procedures were identical to experiment a but the mice (n=8/group) were gavaged with 109 viable e.coli labeled with mmndium oxine. four hours after burn the mesenteric lymph nodes (mln), liver, lungs and blood were harvested to determine plasma endotoxin levels and the magnitude of transtocation of labeled bacteria as measured by the residual radioactivity (dpm) in the organs. circulating endotoxin levels were determined by limulus colorimetric assay. diamine 0xidae(dao) is an enzyme existing in the mucosa of small intestines, but its activity is low in the plasma. it is evoked in the blood with the intravenous injection of much heparin. it is well known that da0 declines under the chronic mueosal injury when the small intestines are cut off or the mucosae become atrophied. aim of this paper is to confirm that da0 goes up to the measureable level by the intravenous injection of a small amounts of low molecular heparin(lmh objectives: the aims of this study is to investigate the inhibitor" effect of the prolease inhibitor, urinastatin, on endotoxin induced bacterial transleeation in mice. materials and methods:lcr mice were divided in six groups as the fotlows, group i: control mice receiving intraperitoneal injections (ip) of saline 24.5 and 24 hr prior to sacrifice, group ih treated mice receiving ip of utinastatin 24.5 hr and endotoxin (5mg/kg) 24hr prior to sacrifice,group ilk sham4reated mice receiving ip of saline 24.5 hr and endotoxin (5mg/kg) 24hr prior to sacrifice,group iv: saline control mice receiving ip twice during 30 rain intervals,group v: mice received ip of utinasmtin, and 30 min later they were received ip of endotoxin (5mg/kg), group vi: mice received ip of saline,and 30 min later they received ip of endotoxin (5mg/kg). in group l,ii and ill ,the mice were killed by cervical dislocation.using stetile techniques,a middle-line incision was made and the mesenteric lymph nodes were harvested in order to culture on selective agars for quantitation of bacterial trauslocation. in group iv, v and vi, survival was recorded for 3 days. unnastatin pretreatment could rednce mortality significantly. first surgey, shiga university of medical science, seta ,ohtsu 520-21,japan increased gut permeability correlates with inflammatory response in multiple trauma aptients. pape, h.-c. dwenger, a. grotz, m. regel, g. purpose: disturbances of intestinal permeability have been advocated as a pathogenetic factor of posttranmatic multisystem organ failure (mof). a close relationship with impairment of the stationary reticulocndothelial system (res) and a systemic inflammatory response (sir) was discussed. these tactors were investigated prospectively in patients with multiple trauma showing posttraumatie complications (multiple organ failure, mof). methods: 31 polytranmatized patients were studied prospectively. according to a mof-score (goris) those with posttrattmatic complications were selected (>7 points at 2 days), others were excluded. every second day gut permeability according to the ratio of urine concentrations of lactulose and mannitol (l/m) was evaluated (enteral application). at parallel time points res clearance capacity (k-value, invasion constant, normal range 0.6 -0.8 mind) was studied after i.v. injection of 100 mbq 99rotehuman albumin. liver perfusion was calculated from these data, total serum bilirubin (/zmol/l) was documented. serum elastase (#g/l) levels were determined enzymatieally. results 86.0 148+ 239+ liver perfusion did not ehangu, bilirubin showed progressive worsening indicating mof. a positive correlation was present between l/m and k (r=0.88) and between l/m and ela (r=0.71). conclusions: there is a positive correlation between the time pattern of intestinal permeability dysfunction and res hyperactivity as well as between intestinal permeability and the systemic intlammatory response (elastase levels). the results speak in favor of an interaction between intestinal and extraintestinal inflammatory systems, which in eombiuation are likely to be responsible for post~anmafic complications. endotoxemia, il-6 release and consecutive acute phase reaction are observed as a host response to surgical trauma. as well vasodilative prostaglandins (pg) and thromboxane (tx) are released after abdominal meaenteric traction (mt). the following hypotension and acute hypoxeraja are duo to prostacyelin (pgiz) arm can be avoided by perioperative cyclooxygenase inhibition. we therefore focused on the effect of pg and tx liberated following mt on the induction of endotoxemia. methods: in a prospective, randomized double-blinded protocol 50 patients, who were scheduled for major abdominal surgery (pancreatic or infrarenal abdominal surgery), were studied. ibuprofen (400 mg i.v.) or a placebo equivalent was administered 15 minutes before skin incision. mt was applied in a uniform fashion. baseline values were obtained before induction of anesthesia. further measurements followed before the incision of the peri[onenm (tl) and 5, 30, 90, 180 min, . the plasma concentrations (,pc) of 6-keto-pgft,, txb: and-ki-12-pgf ~ (stable metabolites of pgi2, txa2 and pge~) were determined by ria. we measured endotoxin pc by limulus-amoebocyte-lysate test and il-6 levels by elisa. data are given as mean+sem (* p< 0.05 placebo vs. [ibuprofen] ). results: endotoxin plasma levels increased before incision of the peritoneum tl both in the ibuprofen pretreated and in the placebo group. peak pc were observed 180 minutes after mt. endotoxin pc were significantly higher in the ibuprufen treated group (t5 0.16+0.02e[0.32+0.06] eu/ml). il-6 pc demonstrated an increase continuously from t4 to t7 (t7 115+12 [91 + 15] ng/l) in both groups. after intentional abdominal mesenteric traction we observed a marked increase of 6-keto-pgf~,, pc up to 6h after mt in untreated patients with a peak of 2450*[60] ng/1 at tl. also txb: and kh2pge 2 pc showed a considerabe increase up to 6h after mt in the placebo group. in ibuprofen pretreated patients the pg and tx pc remained within the normal range. discussion: our data clearly indicate a significant endotoxemia and il-6 release following major surgical trauma which is not initiated either by prostaglandin or thromboxane release. moreover endotoxemia is accentuated by ibuprofen pretreatment. therefore we hypothesize that in major abdominal surgery prostacyclin release-after mt may play a crucial physiological role in maintaining splanclmic microcirculation and thus preserving gut mucosal barrier function. objectives of the study it has been shown recently that parenteral and certain euteral diets promote the translocation of gut flora to the mesenteric lymph nodes (mln) and systemic organs, a process termed bacterial translocation (bt). in chow fed rats bt usually does not occur without further promoting factors. the goals of the present study were to determine whether the provision of defined amounts of standard lab chow during iv-tpn administration wotfld redane the incidence of bt, materials und methods male spf spragnle-dawley rats were divided into 6 groups. group 1 received standard laboratory chow feeding ad lib. in group 2 a central venous catheter was placed, ligated and secured by a spring coil tether attached to a swivel allowing free movement in the housing cage and chow was fed ad lib. in group 3 50% of the calculated daily required calory intake (drci) (307/kcal/kg) was given by iv-tpn (28% glucose, 4,25% amino acids) and 50% by limited chow administration. groups 4 and 5 received 70% and 90% of the drci by i.v. tpn and 30% and 10% respectively by chow feeding. group 6 received iv-tpn only. after 7 days the rats were sacrificed and the mln, liver, spleen and cecum removed aseptically, homogenized and cultured for bt samples of distal ileum were taken for light microscopy. the group with the least amount of chow shown to be protective against bt received the amount of non-fermentable fiber of that chow regimen during iv-tpn feeding and bt was studied. 4 5,5+0,9 4 12 23,9 -2,3 16 ,7 2/12 63+11 125~1 "7,7-+0,7 6,7-+1 5 12 24~6 -3,5 67 8/12 241+~243 170+163 8_+0,7 7+1,1 6 18 25, 5 -4 88,8 16118 2061-+3224 2211+4015 8,4~0,7 8,1-+1~1 conclusions: the administration of 30% of drci by chow feeding during iv-tpn significantly reduced the incidence of bt and maintained gut barrier function. the addition of the respective amount of dietary fiber of this group did not prevent iv-tpn-indueed bt. dr. med. m naruhn., dep. of general surgery, eberhard-karls-university, hoppe-seyler-str. 3 previous experimental studies have suggested that a disturbed ecology of the enteric bacterial population might contribute to the development of bacterial translocation from the gut in acute liver failure (alf). in the present study, the effect of oral administration of lactobacillus reuteri r2lc and oat fiber on bacterial overgrowth and translocation was investigated in rats with acute liver failure induced by subtotal (90 %) liver resection. the oatmeal soup base was anaerobically inoculated with lactobacillae and fermented for 15 hours, after which the animals were fed with either fermented or unfermented oatmeal or saline daily for 6 days prior to the operation. bacterial translocation to mesenteric lymph nodes (mln) and the systemic circulation was determined, as well as the intestinal bacterial flora and enterocyte protein content. the incidence of bacterial translocstion to the systemic circulation was nit in rats subjected to sham operation and saline treatment and 17 % in animals subjected to 90 % bepatectomy and lreatment with fermented oatmeal, while 80-90 % and 34-50 %, respectively, in rats subjected to hepatectomy and treatment with either saline or unfermented oatmeal. only one rat with fermented oatmeal demonstrated bacterial growth in mln (p < 0.05 vs hepatectomy and treatment with saline or unfermented oatmeal). the enterocyte protein content significantly decreased (p < 0.01) in salinetreated animals following 90 % hepatectomy, while there was no significant difference between bepatectomized animals with oral administration of fermented or unfermented oatmeal. the number of anaerobic bacteria, gram-negative anaerobes and lactobacillus significantly decreased and the number of e.cnli increased in the distal small intestine and colon in hepatectomized animals with enteral saline or unfermented oatmeal as compared with animals subjected to sham operation or bepatectomy with fermented oatmeal. our results thus show that the occurrence of bacterial translocatiou from the gut in 90 % hepatectomy-induced alf could be prevented by enteral administration of fermented oatmeal, maybe partly due to a positive effect on the enteric bacterial ecology. 190_+20" 16+_3" 11.7" data=mean_+sd, * stats anova p<0.05 vs control. l+air and lap groups, both exposed to exogenous i.ps shnwm:t m significant increase (p<.05) in lps gut translocation compared to control and l+co2. this correlated with a significant increase in peritoneal inflammatory responses (o2-,tnf) above that of the control and l+co2 groups, while mac-1 and cr3 opsonized phagocytosis were significantly impaired. the absence of significant differences between l+air and lap groups indicates that lps rather than wound factors is the principle mediator. thus, lps plays a significant role in regulating peritoneal responses in the early post-operative period dept of surgery, rcsi, beaumont hospital, dublin 9, ireland brlke e, berger d, staneseu a, buttenschsn k, vasilescu c, seidelmann m, beger hg in 32 patients undergoing a colonoscopy, endotoxin, endotoxin neutralizing capacity (enc), thromboxane b o (stabile metabolite of tbmomboxane ~), 6-keto-prostaglansin, leueotriene c4, interleukin 6 and the incidence of bacteremia were determined before and then every five minutes during the procedure. twenty-one of 32 patients showed a significant increase of endotoxin plasma levels during colonoscopy (p=0.003), whereas only one patient had a positive blood culture with bacteria obviously derived from the gastro-intestinal tract. the enc decreased significantly five minutes after the beginning of eolonoscopy and was diminished further thereafter. the baseline values were reached after 24 hours. ~hromboxane b o levels also increased after five min. from 89 to 376 pgyml peaking at 30 min. with 1332 pg/ml. 6-keto-prostaglandin,leucotriene c 4, ii-6 and crp remained unchanged. a control group of i0 volunteers who were not subjected to endoscopy, were prepared for eolonoscopy by orthograde lavage. the blood sampling procedure remained identical. no differences were seen in all described parameters for the controls. these data show that the gut barrier can be compromised by mininml invasive procedures, at least, concerning bacterial products. living bacteria, on the contrary, do not pass the gastro-intestinal wall. endotoxin, when determined by enc, is more sensitive than the conventional limulus-amebocyte-lysate test. no acute-phase reaction was induceri by the observed endotoxin translocation. it can be speculated from the dramatically enhanced thromboxane b 2 levels, together with its hemodynamie effects, that the thromboxane release may support translocation of bacterial products. sepsis is common after hemorrhagic shock. this study aims to demonstrate that hemorrhagic shock alone can promote translocation of gut bacteria from intestinal tract to its regional nodes and subsequently to blood. one hundred twenty mice, divided into 4 groups were subjected to 30, 60 and 120 minutes of 20%, 30% and 40% of hemorrhagic shock. on the specified time, blood cultures were taken and mice were sacrificed. the intestinal tract were histologically examined for any changes which allows translocation and its regional nodes were quantitatively cultured for translocated bacteria. there was a direct relationship between duration and degree of hemorrhagic shock and incidence of translocation (p 0.05). there was a high incidence of gut bacterial translocation to the mesenteric and mesocolic nodes in all degrees of shock (p 0.05). bacterial growth in the regional intestinal nodes increased and blood cultures were positive in direct proportion to degree and duration of shock. histologic evaluation of segments of git showed submucosal congestion to allow bacteria normally contained within the gut to cause systemic infections. translocation of gut bacteria in untreated hemorrhagic shock is clearly shown in this study on animal models. in this study, guotobiotic rats with 5 known species of bacteria were subjected to total parenteral nutrition(tpn) and subsequent hemorrhagic shock. the purpose of the study was to observe the impairment of gut barrier function following tpn and hemorrhagic shock and to study the mechanism of enterogenic infection induced by tpn and shock.the results were as follows: 1.long term(8-12 days) tpn induced impairment of gut barrier function, evidenced by atrophy of intestinal mucosa, significant decrease in diamine oxidase activity of intestinal mucosa and blood, and marked microecologic imbalance of the intestinal mucosa flora with dorminant growth of aerobes and relative decrease in anaerobes. the degree of mucosal damage were proportional to the duration of tpn. 2.in tpn+shock groups, failure of gut barrier function was found. ri,~ere were further damage in the mucosa, with a large number of gramnegative organisms invading mucosa and submucosa and a significant decrease in dao activity as compared with each relative tpn groups. these changes were significantly correlated with enhanced bacterial translocation, elevation of lps and mda levels in the plasma. these findings suggested that long term standard tpn impaired the gut barrier function, precipitating posttraumatic gut barrier failure. thus infec. fion following shock might be oi'iginated from the gut and it was obviously related to the impaired gut defence resulted from antecedent tpn. the determination of plasma dao activity might provide a valuable tool for the ear. ly diagnosis of gut injut;y during tpn and after trauma. in our earlier studies we have investigated the dynamics of granuloayte infiltration of the ischemic/reperfused s~all intestine (g. illy~s, j. hamar int. j. exp. 9athol. 73. 161. 1992.) . there was a increasing infiltration of the mucosa c111m~nating at the 3d to 4th hours of reperfusion. in the present series we have studied sc~e of the conseqn/ences and the possible role of this cellular reaction. 45 ~in isehemia was followed by a 4 hour reperfusion in the anesthetized rat. arterial ~/ad mesenteric venous blood samples were collected at 5 m_in, i, 2~ 3, and 4 hours of reperfusion. elastase and lactate concentrations were determined and hamoculture was carried out from the blood samples, and tissue pieces from the heart, lung, liver and kidney were collected for histological analyses at the above mentioned times of reperfusion. all blood samples were free of cell bacteria. staphylococci appeared only occasionally at the 4th hour in the arterial blood .and at the 3d and 4th hours in the venous blood, respectively. arterial and venous elastase activities were high throughout the reperfusion, venous concentrations being higher at all times. lactate concentrations of the arterial and mesenteric venous blood samples increased during shock. ~ranuloeyte infiltration of all organs studied appeared during the 2d hour and it increased at later times of reperfusion. it is concluded that heavy infiltration of the intestinal mucosa can block bacterial translocation in most of the cases during reperfusion. granulocytes activated either by the reperfused area or by the released cytokines infiltrate other organs contributing by this way to the mesenteric shock s!rndrc~e. intestinal motility plays an important role for maintaining nutrient transport and absorption and for balancing the enteric bacterial population. disturbances of intestinal motility may be one of the earliest notable changes in intestinal function. in the present study, we aimed at determining early alterations in intestinal transit time following ischemia-reperfusion injury induced by occlusion of the superior mesenteric artery in the rat. intestinal ischemia was induced for 20 and 30 minutes by applying a microvascular clip on the superior mesenteric artery followed by reperfusion 2, 4 and 6 hours after clip removal. intestinal transit time was measured by the propulsion of a radiolabelled solution (cr51). light microscopy was performed on intestinal samples. macroscopical pathological changes were not observed. however, microscopically, mucosal epithelial oedema, degeneration or slight ulceration occurred in rats 6 hours after reperfusion in ischemia-20 rain group and 4 and 6 hours after reperfusion in the ischemia-40 rain group. delayed small intestinal transit time was seen from 2 hours and on after intestinal ischemia for both 20 and 40 rain ischemia followed by reperfusion. the distribution of radioactivity demonstrated that most radioactivity was accumulated in the first two segments following intestinal ischemia and reperfusion, significantly differing from what was seen in animals subjected to sham operation (p < 0.01). the distribution of radioactivity in segments 4 and 5 in the group with repeffusion 6 hours after intestinal iscbemia for 20 rain was significantly higher than that noted in the group with repeffusion 6 hours after intestinal ischemia for 40 min (p < 0.01). q'he results indicate that a delayed intestinal transit time may be one of the earliest pathophysiological alterations noted, associated with duration of gut ischemia, and a potential factor for the development of bacterial overgrowth, gut barrier failure and bacterial translocation, in hypovolemic conditions. bacterial infections still constitute a major cause of morbidity and mortality in patients with acute liver failure. the present study aimed at evaluating the effect of ethylhydroxyethyl cellulose (ehec) on bacterial translocation following surgically induced acute liver failure. acute liver failure was induced by subtotal hepatectomy (90 %) in the rat. water-soluble ehec was administered orally 1 and 12 hours prior to hepatectomy. the incidence of bacterial translocation from the gut to mesenteric lymph nodes (mlns) and systemic and portal circulation was evaluated and the number of isolated bacteria from these samples and from intestinal content were determined. intestinal transit time, bacterial adherence onto the intestinal surface, intestinal mucosal mass, bacterial growth and dna synthesis, bacterial surface characteristics (hydrobiology: hydrophobicity, hydrophilicity and neutrality; surface charges: positive, negative and neutral) were also determined. hepatectomized animals showed a 80-100 % translocation rate to mlns or blood 2 and 4 hours after operation, while only 0-1% of rats subjected to sham operation or animals with 90 % hepatectomy and pre-treatment with ehec (p < 0.01). bacterial overgrowth, increased bacterial adherence onto the intestinal surface as well as decreased intestinal mucosal masses were observed in animals with subtotal liver resection alone, alterations that were prevented by enteral ehec treatment. a delay in intestinal 2-hour transit time occurred in both groups with subtotal liver resection, with or without enteral ehec. ehec inhibited bacterial growth and dna synthesis, and altered bacterial surface properties following 1hour incubation with bacteria. in conclusion, the findings in the present study imply that ehec alters enterobacterial capacities for metabolism, proliferation and invasion by effects on e.g. bacterial surface characteristics. furthermore, ehec seems to possess a trophic action on the intestine, rather than exerting its effect by enhancing intestinal motility. department of surgery, lund university hospital, s-221 85 lund, sweden disturbances in intracellular calcium signalling can potentially result in impairments of cellular responses vital to the functional integrity of both immune and non-immune cells, and thus contribute to a decrease in host resistance against infection and to multiple organ system failure during sepsis. studies in our laboratory have focused on assessments of intracellular ca 2÷ regulation and ca~+-depended cellular responses in the liver, skeletal muscle and splenic tlymphocytes harvested from rats subjected to gram-negative intraabdominal sepsis. cytosolic ca 2+ concentration, [ca2*]i, and ca 2+ fluxes were measured by the use of fluorescent ca 2+ chelating dyes (fura-2 or indo-1) and 45 ca respectively. to assess sepsis-related changes in ca 2+ dependent cellular responses, we measured the acute phase protein response in the liver, the regulation of protein and sugar metabolism in the skeletal muscle, and the proliferation response in the splenic tlymphocytes. altered ca2+ i signalling with sepsis was correlated with an exaggerated inappropriate acute phase protein response (100% ¢) in the liver, and a blunted insulin mediated sugar utilization (60% 4) and increased proteolysis (50% ~) in the skeletal muscle. in t-lymphocytes, a decrease in mitogen induced elevation of [ca2+]i by 35-40% was correlated with a significant depression in their proliferative capacity. these studies clearly suggest that altered calcium signalling is correlated with disturbances in cellular responses in both immune and non-immune cells during sepsis. the altered cellular responses adversely effect the outcome of the septic injury. (supported by nih grant gm 32288). alfred ayala, ping wang and irshad h. chaudry. changes in macrophage capacity to respond to foreign pathogens are thought to be central to the developing immunosuppression associated with traumatic injury. in this respect, the suppression seen in m~ functions following hen (a common component of traumatic injury) may be mediated by the direct or indirect inhibition of their capacity to perceive external stimuli (e.g., opsonized & non-opsonized bacteria, and their cellular components, etc.} due to the breakdown of the receptormediated signal transduction system. results of a number of studies by our laboratory and others indicate that this inability to respond to external stimuli is in part due to the loss of cell surface receptors. decreases have been documented for not only la antigen, but also c3b, fc, and tnf receptors following hem in mice. furthermore, studies which have examined second messenger generation in these cells indicate that m~ derived from the peritoneum and spleen exhibit a decreased capacity to mobilize ca +2 from intracellular stores. this protein kinase dependent process of [ca+2] i mobilization appears to be linked to the inability to synthesize inositol triphosphate. of interest, the depression in ca +2 signal generation appears to be inversely related to presence of elevated levels of camp in m~ from hen mice. we have reported that m~ priming agents, such as ifn-7 (which exhibits salutary effects on m~ function following hem), appear to restore cell signal transductive capacity while reducing the levels of camp. nonetheless, the extent to which depressed receptor signal transduction in hem, is due to receptor loss~dysfunction or elevated antagonistic second messenger levels remains to be determined. conclusions: significant impairment of calcium signaling occurs at all time-points prior to and following pha stimulation in trauma patients. tcell activation failure can, in part, be explained by the inadequacy of this essential intracellular second messenger system. restoration of immunocompetence following trauma will have to address strategies to better assess and restore this vital step in the activation sequence leading to proliferation during the antigen recognition process. patrick a. bseuerle institute 01 biochemistry, albert-ludwigs-university, hermann-herder-str. 7, d-79104 freiburg, germany the active form of the transcriptional activator nf-~b is a heteredimer composed of a 50 and 65 kda polypeptide. in this form, nf-'<b can bind with high affinity to decameric sequence motifs (consensus: 5"-gggpunnf'ypycc-3") found in enhancers and promoters of many genes activated upon a wide variety of pathogenic conditions, including viral and bactedai infections, acute phase response, oxidative stress and uv and gamma irradation. clinically important target genes for nf-k:b encode inflammatory cytokines (il-1b, tnf, [l-6, il-8, gm-csf etc), cell adhesion molecules (icam-1, elam-1, vcam-1), acute phase proteins and immunoregulatory ceil surface receptors. in most cell types, nf-~:b is sequestered in the cytoplasm and, therefore, unable to initiate transcription of these genes. the cytoplasmic form of nf-~:b contains one additionai subunit, referred to as h,;:b. i~b can reversibly suppress dna binding and nuclear uptake of nf-~b by virtue of its association with p65. upon stimulation of cells, ikb is proteolytically destroyed, allowing nf-kb to enter the nucleus and activate genes upon binding to transcriptional enhancers. we have demonstrated that various antioxidative compounds inhibit the activation of nf-~b in response to many inducers. moreover, nf-~:b was shown to be activated upon treatment of cell cultures with p.m-amounts of hydrogen peroxide. these data suggested that nf-~b prinicipaliy is an oxidative stress-responsive transcnption factor. the notion is supported by numerous reports on the induction of oxidative stress by many nf-nb-inducing agents. nf-~b activition can likewise be suppressed by protease inhibitors. these drugs apparently inhibit a yet unidentified protease responsible for the inducible decay of inb. we propose to use nf-nb inhibifors as novel drugs with a very broad application under acute phases el inflammation, injury and infection. northern blot experiments revealed that expression of the recently discovered lipopolysaccharide binding protein (lbp), a 58/60 kd glycoprotein, in the liver rises substantially during the acute phase response. experiments with an in-vivo acute phase model using new-zealand-white rabbits and also in-vitro experiments employing stimulated hep-g2 cells showed that lbp transcripts could be induced 10 to 100 ford within 24 hours after induction with cytokines. by using a newly established nonradioactive nuclear run-on technique we show that induction of lbp during the acute phase is transcriptionally regulated. furthermore we screened a human genomic library and were able to clone the lbp gone, containing the promoter 1.5 kb upstream. several liver-specific and "acute-phase-typical" potential binding sites were found fn the promoter region and reporter gone assays were set up. several caat-boxes, the il-6 responsive elements hstf-hsp 70.5 and h-apf-1 rs as well as the transcription factor hnf-5 and the glucocrticoid-responsive elements oct-2-d and gcre were found, which correlates with the induction pattern of lbp mrna in hep-g2 cells by il-6, il-1 and dexamethasone. pcr-based analysis of the exon-intron pattern of the lbp gene revealed similarities with the genes of two other lps binding proteins, namely bpi and cetp. further analysis of this novel acute phase protein, that also has an important role in endotoxin recognition and immunomodulation will help in understanding the complex acute phase mechanism and potentially lead the way to new therapeutic strategies. lps activation of nucle?/r fa-ctor:~b " (nf:~bj"in cd14.transfected fi'broblasts does not appear to require tyrosine kinase activity d. t. golenbock, r. delude, r. savedra, s. vogel and m. j. fenton lipopolysaccharide (lps) is the major constituent of the outer leaflet of gram-negative bacteria. during the course of serious infection, shock may occur as a result of the interaction of lps with macrophage lps receptors. cd14, a 55 kda glycosyl phosphatidylinositol (gpi)-linked protein, functions as an lps receptor. a critical issue in lps activation concerns the mechanism by which cd14, which has no transmernbrane domain, transduces a sig,nal after lps binding. evidence exists which suggests that cd14 may signal cells after lps binding by interacting with an lps signal transducer with tyrosine kinase (tk) activity. wild-type chinese hamster ovary fibroblasts (cho) normally do not respond to lps, but can be activated following transfection with cd14 (cho/cd14). stimulation of cho/cdi4 with as little as 1 ng of lps/ml resulted in the release of 3h-arachidonic acid (3h-20:4) into the culture supernatant. in addition to 3h-20:4 release, we examined lps-imiuced activation (transl0cation) of the transcription factor nf-~b. similar to lps-induced 3h-20:4 release, these responses were observed only in cho cells lransfected with cd14 closely resembling the same response in the marine macrophage cell line raw 264.7. maximum nf-~cb expression occurred at 30 minutes. in contrast to lps-induced 3h-20:4 release, dexamethasone, cycloheximide, and actinomycin d had no effect on activation of nf-r,.b in cho/cd14, suggesting that nf-r,b activation begins early after lps binding to cd14 and does not require transcription or translation. we then examined cells for lps-induced tk activity by western blotting cellular ly~ates with anti-phcsphot-'rosin:~ anl',bodie~" although tk activity was seen in lps-stimulated raw cells and phorbol ester stimulated cho/cd14, lps-induced tk activity in cho/cd14 was not observed. although the tk inhibitor herbimycin inhibited the release of 3h-20:4 in cho/cd14 and raw cells, neither herbimycin nor genestein effectively blocked nf-r,b activation in either cell type. these results suggest that tk activity is involved in a portion of the signaling pathway that is distal to initial signal transduction. the absence of observable lps-induced phosphotyrosine residues in cho/cd14 cells as well as the failure of tk antagonists to inhibit lps-induced nf-~cb activation suggest that the proposed cd14-related lps signal transducer in cho/cd14 is not a tymsine ldnase. dimished cardiac inotropic function and response to 13-adrenergic receptor (13-ar) stimulation are frequently seen in septic patients, these cardiac defects are also seen in animal models of entoxemia. to determine the characteristics of the decreased transmembrane signal associated with the decreased beta adrenergic response we measured the force and rate of contraction of atria harvested from sprague-dawley rats exposed to endotoxin (0:2 gm/kg). to stimulate various components of the transmembrane signal cascade of 15-ar was isoproterenol (i3-ar), acetylcholine (m2ach receptor), naf(gs and gi proteins), forskolin (adenylate cyclase), and calcium (intracellular contracttile signal) were used. to eliminate the transmembrane control of calcium and test the intrinsic contractile response, hypothermia (22°c) was used the results showed alterations in the inotropic function with no significant difference in chronotropic response. the inotropic reponse for the endotoxin exposed atria and controls is shown below these results show that there is a transmembrane signal defect both for the i~-ar signal and for ca +. these data indicate that the level of the defect appears to be at the g proteins. ischemia-reperfusion of the rat intestine produces an injmry both to the intestine and to distal organs, notably the lungs and liver. prior studies have shown that 1 h of intestinal ischemia leads to a nemtrophil independent reperfusion injury of the gut. thus, rats rendered neutropenic by pl~l antiserum have been shown to express a severe local gut injury when subjected to ischemia and reperfusion. this gut injury is postulated to he mediated at least in part by the complement system. the soluble (s) cri receptor which binds to c3b and cdb was given to rats at a dose of 12 mg/kg by intravenous bolus, 15 min before reperfusion of the ischemic gut. a control group was given pbs buffer. at 4 h of rmperfmsion the animals were sacrificed. pbs treatment led to a significant activation of both classical and alternate complement pathways while scri inhibited both pathways (fall to zero of chbo ). intestinal mucosal injury score, assessed by histological grading was reduced by scri (2.49±0.22 to 1.73_+0.17,p<0.05) . intestinal neutrophil sequestration estimated by assay of myeleperoxidase was also reduced (0.05 ± 0.01 to 0.02 ± 0.01 u/g). c3 was detected in the gut hy i~unohistochemistry. remote organ injury was modified that is lung permeability (ration of concentration of radiolabelled albumin in broncho-alveolar lavage fluid to that in blood) was reduced (11.3±1.4 x i0 3 to 3.9±0.8 x 103, p<o.05). however, neutrophil sequestration by the lungs was unaffected. this remote protection is thougth to be related to prevention of ic3b deposition on imng andotheli~ which activates adherent neutrophils. these findings support the hypothesis of complement mediated local and remote injury. we postulate that complement fragments may be deposited either as a result of the ischemia induced loss of membrane bound complement regulatory proteins, such as decay accelerating factor or membrane cofactor protein, which allow complement fragments to accumulate on the endothelial cell surface. alternatively, increased expression of p-selectin on the endothelial cell surface may, by means of its complement binding domains, act as a lattice for complement deposition. indeed, p-selectin has been detected on the surface of endothelial cells ans platelet-neutrophil aggregates recovered from the intestine of these animals. ischemia (4hr) followed by reperfusion (dhr) of rat hind limbs resuits in local (muscle) injury as well as in remote (lung) injury, which is characterized by increased vascular permeability (leakage of j2sl labeled albumin) and neutrophil accumulation (tissue content of myeloperoxidase, mpo). within 60 minutes of reperfusion following ischemia, tumor necrosis factor-o~ (tnfc 0, interleukin-i (1l-i) and il-6 plasma levels increased significantly, reaching maximum levels after 2 hours ofreperfusion. polyclonal antibodies to tnfet and 1l-i provided significant protection against muscle and lung injury. muscle and lung vascular permeability decreased in anti-tnfct treated rats by 41% and 74% respectively and mpo was reduced by 72% and 83%. antml-1 yielded a protection in muscle and lung vascular permeability of 24.7°,4 and 52.4% respectively whereas muscle mpo was reduced by 46.4% and lung mpo by 25.9°,4. these results were confirmed by the use of soluble tnf~ receptor and il-i receptor antagonist (il-ira). when icam-i expression in vivo was examined using 125mabeled antmcam-i, constititive expression of icam-1 was evident only in lung but not in muscle. during reperfusion icam-1 expression increased by 98.2%. treatment with anti-tnfcx or il-ira reduced icam-i upregulation by 73.9°,4 and 74.8% respectively. frozen sections of normal rat lung revealed no evidence of e-selectin expression, whereas lung sections obtained after ischemia and reperfusion immunhistochemically demonstrated expression of e-sdectin. lungs from rats pretreated with anti-tnfct were negative for e-selectin staining. our data indicate a role for cytokines in the upregulation of adhesion molecules in ischemia/reperfusion injury. adherence of neutrophils to the coronary endothelium is associated with significant myocardial injury following 90 min of ischemia (mi) and 4.5 h of reperfusion (r). p-selectin is expressed by activated endothelial ceils and platelets within 10 min and tethers polymorphonuclear leukocytes (pmns) to the endothelium. we examined the cardioprotective effects of a monoclonal antibody (mab) designated pb1.3 to p-selectin in a feline model of mi+r. pb 1.3 (1 mg/kg) administered 80 rain post-occlusion (10 min prior to reperfusion) significantly attenuated myocardial necrosis compared to a non-blocking mab (nbp1.6) for p-selectin (15 + 3 vs 33 + 5% of area-at-risk, p<0.01). endothelial dependent relaxation to acetylcholine was also significantly preserved in ischemic-reperfused coronary arteries isolated from cats treated with pb 1.3 compared to nbp1.6 (67 + 6 vs 11 + 3, p<0.01). furthermore, pmn adherence to ischemic-reperfused coronary endothelium was significantly attenuated in pb1.3 treated cats (64 + 7 vs 136 + 12 pmns/mm z, p<0.01). also, normal coronary endothelium activated with thrombin (2 u/mt) had increased pmn adherence under conditions of shear stress, an effect which was significantly (p<0.01) blocked by pb1.3, but not by nbp1.6. furthermore, p-selectin is markedly upregulated 10-20 min post-reperfusion in coronary arterial and venous endothelinm. similar results were obtained in a rat model of mesenteric ischemia/reperfusion, including upregulation of p-selectin on mesenteric vascular endothelium. these results demonstrate that tethering of neutrophils to endethelium by p-selectin is an important early consequence of reperfusien injury, and a specific monoclonal antibody to p-selectin exerts significant endothelial preservation and cardioprotection in ischemia and reperfusion states. as recent interest has increased in the role of extracellular matrix proteins in inflammation, we examined the role and relationship of laminin and fibronectin to the processes of cell infiltration in a rat model of heart isografts. changes occurring in this system are on a nonimmunological basis and may be an effect of the initial surgical manipulation and the warm and cold ischemic times of transplantation. lewis (lew) donor hearts were transplanted into lew recipients, naive lew acted as naive controls (nc). grafts were harvested 2, 4, 8, 16, 24, 48, 72, 120 and 168h after transplantation (n=6/time point) . snap frozen sections of organs were stained for cd-4 (w3/25), cd-8 (ox8), t-lymphecytes (tl) (ox-19), macrophages (ed1, ed2), neutrophils (pmn) (mom), icam-i (iap29), laminin and fibrinogen. results were evaluated visually by counting cells/field of view (cf) for w3/25, ox8, ox19, ed1, ed2 and mom, staining for icam1, lain and fib was assessed visually using a scale (ve=0-10). as early as 2h after transplantation, fibrinogen and laminin expression increased in i, peaking 8h after transplantation (ve= 4.5; nc: ve=i.5) . at 16 and 24h in i, on both vessels and interstitial cells laminin and fibrinogen expression had declined to baseline (ve=i.5) but rose again at 48h, peaking at 72h (ve=4) and remaining elevated thereafter (ve=3). the high expression at 8h correlated well with the maximal pmn infiltration at that time (cf=6) in i returning to baseline at 24h (cf=0.5) and thereafter. the second peak at 72h correlated with the onset of macrophage and t-lymphocyte infiltration. thus, laminin and fibrinogen seem to guide leucocyte infiltration following graft ischemia during the time of reperfusion. background. reperfusion of ischemic kidneys with xenogeneic blood leads to activation of endothelial cells and circulating leucocytes with the final consequence of microvascular thrombosis. in concordant systems, the mechanisms of rejection can be studied due to cross-reactivity of mediators with anti-human monoclona~ antibodies. aim of the study. to obtain information about the kinetics of proinflammatory cytokines and production of soluble adhesion molecules in the acute phase of reperfusion, eight kidneys from rhesus monkeys were perfused ex-vivo with human blood (group b/o) for one hour in a closed system. blood levels of il-lb, il-6, tnf-a, soluble icam and e-selectin were measured in elisa-technique under steady-state conditions. results. cytokine levels rose significantly within the 60minute interval (il-lb: 6,1__+2,6 to 161,1__+98,5 pg/ml; il-6:30.24-7,7 to 274,2__+75,8 pg/ml; tnf-a: 544,2__+363,6 to 1651,0+25,7 pg/ml; p<0.05). immediately after the beginning of reperfusion, soluble icam-1 and selectin levels were abnormally high and constantly rising throughout the observation period, reaching significance at 60 minutes. discussion. high levels of proinflammatory cytokines may lead to an induction of adhesion molecules, thus upregulating the leukocyte-endothelial interaction in an complement-independent mechanism. specific pretreatment with monoclonal antibodies against icam-1, lfa-1 or other soluble mediators may be useful in downregulating hyperacute rejection in transspecies transplantation. prolonged ischemia has been associated with later dysfunction of solid organs as it may cause upregulation of adhesion molecules, production of cytokines on vascular endothelium and migration of host cells into the graft. these early events may lead to irreversible organ injury. this study evaluates the effects of global ischemia on early immunohistological changes in cardiac grafts transplanted in rats. isografted hearts (lewis->lewis) were were divided into ischemic and non-ischemic groups (n=30/group). all donor hearts were flushed immediately with cold saline. non-ischemic hearts were then transplanted within 30 rain, ischemic hearts were stored in cold ringer's solution for 3 hours before revascularization. representative grafts were removed after 12. 24, 72hrs, 10 and 100 days, and evaluated immunohistologically (cells/field of view=c/f). restitution of ventricular activity was significantly delayed in ischemic grafts (5 vs 1 rain). after 12 hrs, all ischemic grafts exhibited an extensive interstitial edema, declining slowly thereafter. at the same time, numbers of pmn peaked (3 vs 1 c/f in non-ischemic grafts), whereas edl+macrophages (9 vs 2 c/f) and tnfe expression peaked by 24hrs. by 72 hrs t-lymphocytes began to enter ischemic myocardium and icam-1 was moderately increased. after 10 days cellular infiltration had returned to baseline, and no differences were seen among both groups after 100 days. global myocardial ischemia inhibits initial graft function, and engenders a brisk inflammatory reponse, primarily pmn and macrophages, with increased mhc class ii and cytokine expression. leukocyte -endothelial interactions are the result of endothelial activation, leukocyte activation or combination of both, which are accompanied by nee-expression, upregulation or shedding of adhesion molecules (selectins, inlegrins). such interactions differ with regard to the stimulus (e.g. thrombin or histamine for p-selectin, endotoxin or tnf/il-1 for e-selectin), the time course of response (minutes versus hours) and the localisation in different organs. recently assays are available for circulating soluble fragments of the cell bound adhesion molecules e.g. se-seleetin was found to be increased in plasma concurrent with high circulating endoloxin and cytokine levels. the importance of adhesion molecules for the sepsis event is evident, while effectiveness of anti-adhesion inolecu]e therapy is controversial e.g. beneficial anti-e-selectin therapy in baboon bacleremia but deleterious effects of amti-cd18 treatment in the same model. in other species similar controversial results with anti-cd18 therapy in sepsis were reported. steven l. kunkel,theodore standiford* and robert m. stricter. the migration of leukocytes to the lung during endotoxemia is dependent upon the coordinated expression of lung vascular adhesion molecules and the subsequent production of appropriate leukocyte chemotactic proteins. in experimental animals, neutrophils accumulate within the lung soon after the administration of endotoxin, while mononuclear cell infiltration occurs in a more distal manner. a kinetic analysis of lung leukocyte levels revealed a 3-fold increase in neutrophil numbers associated with dispersed lullg tissues 2 hours after lps treatment, while macrophage levels increased by 4-fold at the 24 hour time point. thus, the recruitment of different leukocyte populations to the lungs during endotoxemia is likely directed by different mechanisms. recent studies have identified a supergene family of small inducible chemotactic cytokines (chemukines) which possesses chemotactic and activating properties for neutrophils. the prototype of this family is interleukin-8 (il-8). interestingly, a related supergene family has been identified which possesses activity for recruiting mononuclear cells. examples of this group of inflammatory chemukines are monocyte chemotactic protein-i (mcp-i) and macrophage inflammatory protein-i alpha (mip-i). in initial in viva studies we examined whether mip-i was expressed systemically or in a compartmentalized fashion post lps challenge. assessment of plasma cytokine levels revealed maximal tnf levels occurred i hour post lps administration, returning to baseline by 4 hours, while mip-i levels were maximal at 2 hours (2,5 ng/ml), with a second peak at 24 hours after lps challenge. interestingly, aqueous extracts of liver homogenates from lps treated animals demonstrated no mip-i levels, while aqueous extracts of lung revealed a 4-fold increase in mip-i levels over control lungs. immunohistochemical analysis of the lungs from 24 hour lps treated animals demonstrated the alveolar macrophage was a rich source of mip-i protein. cell-associated mip-i was also expressed by blood monocytes adherent to the pulmonary vascular endotheliun, however the expression of monocyte-mip-i was observed by 2 hours post lps administration. immunohistochemical analysis also demonstrated that mip-i antigen is associated with the extracellular matrix on the interstitial side of the endothelium. this suggests that the extracellular matrix, which is produced during inflammation, can bind mip-i and this may serve as a depot for the prolonged presence of nip-1. in additional studies we have demonstrated that the intratracheal instillation of rmui [ip-l(loong) activation of polymorphonuclear leukocytes by inflammatory stimuli may contribute to the development of multiple organ failure in septic patients. thereby pmnl are proposed to avidly adhere to vascular endothelium causing damage by the subsequent release of toxic agents. as cellular adhesion is primarily mediated by 62-integrins and lselectins, the present study compares the expression of these adhesionmolecules on pmnl in septic patients and healthy volunteers. methods: expression of 62-integrins and l-selectins on pmnl was measured in whole blood by flow cytometry using the monoclonal antibodies ib4 and dreg 200, baseline values were determined immediatley after drawing blood. in addition cells were incubated 45 min at 37°c to allow for spontaneous regulation of adhesion molecules. 55 blood specimens from 8 septic patients were obtained during the course of their illness. control values were determined in 12 healthy volunteers. results: baseline expression of 62-integrins and l-selectins was not signifcantly different in septic and in healthy subjects. in contrast, there was a significant upregulation of g2-integrins and shedding of l-selectins of pmnl in septic patients (sp) compared to healthy volunteers (hv). the local or systemic production of inflammatory cytokines, such as tumor necrosis factor alpha (tnfc~), can serve to modulate multiple aspects of neutrophil function. the ability of neutrophils to leave the circulation and migrate to areas of infection is one essential component of host defense. l-selectin, a leucocyte-associated adhesion molecule, is responsible for the initial reversible contact between neutrophils and endothelium and the subsequent roiling action of neutrophils along the vessel wall. in contrast to other adhesion molecules, l-selectin expression is rapidly down-regulated after neutrophil activation. the loss of l-seleclin may thus be a critical determinant of how neutrophils become unbound from their endothelial attachments and enabled to proceed towards an underlying extravascular area of infection. we hypothesize that the shedding of l-selectin is a strictly controlled process, occurring primarily at localized sites of inflammation, which may be modulated by tnf~, a flow cytometric method of staining neutrophhs by monoclonal antibodies in whole blood is described whereby the kinetics of l-selectin shedding may be followed in real time. the dose response and time course of in-vitro l-selectin shedding by neutrophils from normal human subjects was assayed after exposure to n-formyl-methionylleucyl-phenylalanine (fmlp) and tnfc~. either singly or in combination, our results show that l-selectin shedding can be reliably followed over time. a significant percentage of cells shed l-selectin after exposure to 2000 pg/ml tnfc~ or 1000nm fmlp (but not at 400pg/ml tnfc~ or 200nm fmlp). greater numbers of cells were able to shed their l-selectin when fmlp and tnf~x were presented in combination rather than alone. high levels of tnfc~ did not appear to alter the threshold concentration of fmlp required to induce shedding, we conclude that the extent and rapidity of l-selectin shedding may be modified by different combinations of ligands and that shedding, by vidue of the high concentrations of cytokines or chemotactic factors required, is a process localized to sites of infection or inflammation. we prospectively studied 23 patients with severe sepsis syndrome; group a : septic shock with or without adult respiratory distress syndrome lards) (n =7, bacteremia = 6); group b : sepsis syndrome without septic shock (n = 16, bacteremia = 9). serial plasma samples obtained on day 0, 1, 3, 6, 10 and 14, were assayed using elisas method (british biotechnology), normal control levels of soluble icam-1 and e-selectin, obtained from 11 healthy volunteers, were respectively 205 ± 19.4 ng/ml and 45 ± 5.7 ng/ml (mean _+ se), acute lung injury was quantified dally on a tour-point score system (murray, am rev respir dis,1989) . compared to control mean values, initial levels of groups a and b were significantly higher for icam-1 (p < 10 -4) and e-selectin (p < 10-3). comparisons of group a and [] (* = p<0.05; ** = p<0.00t) soluble icam-1 levels of group a enhanced significantly (p<0.05) during the first 24 hours, and a sustained high levels was of bad prognosis (25% of survivors at day 6). the evolution of soluble icam-1 and e-selectin levels were significantly correlated with murray's score (spearman test : p <0.001). conclusion: these results suggest that endothelial adhesion molecules are released into the plasma of patients with severe sepsis syndrome. soluble icam-1 and e-selectin are correlated with endothelial lung damage, and loam-1 seems to be a better indicator of the severity of endothelial injury. introductory remarks to anti-adhesion molecule strategies as a therapeutic modality ch wortel, repligen corporation, one kendall square, building 700, cambridge, ma 02139, usa. the development of antimicrobial therapy represented a major breakthrough in the struggle against disease. it strengthened the notion that disease could be overcome by eliminating foreign invaders threatening the host. this paradigm has proven to be very successful, the threat of many infectious diseases has significantly changed, some have even been eradicated. nevertheless, sepsis has remained a severe condition, increasing in incidence while mortality remained very high. more recently, it has become increasingly clear that besides the nature and treatment of an exogenous agent, the reaction of the host defense itself plays a pivotal role in the outcome of the event. endogenous mediators, such as tnf, il-i, il-6 and il-8, govem many of the actions of the host defense system. while the expression of these cytokines more often than not benefit the host, (over)-expression can cause severe damage. based on this hypothesis,anticytokine strategies, such as those targeted against tnf or il-1, have been evaluated for the treatment of sepsis. results of these early studies have not yet indicated success in improving the outcome of the disease. it has been difficult to define a patient population where a benefit could be reproducibly shown. furthermore, it has been documented that synergy between cytokines occurs, but detailed knowledge of the cytokine network is not yet available. it is conceivable, that neutralization of one cytokine prompts the induction of another which will evoke the intended response in the host. recent data obtained in human endotoxemic volunteer models seem to confirm this. if this turns out to be the case, neutralizing a single cytokine may not be a successful approach. cytokines in tum, induce various adhesion molecules, such as icam-1. such molecules regulate for instance the neutrophil-endothelial cell interactions, which are thought to play an important role in the pathogenesis of systemic organ injury. the potential for monoclonal antibodies to adhesion proteins to reduce vascular and tissue damage has been studied in a large number of experimental models. protective effects have been observed in a wide variety of inflammatory, immune, and ischemia-reperfusion injuries. thus, altering the host response by modulating the function of adhesion molecules may attenuate the inadvertent injury caused by inappropriate behavior of host defense cells. targeting cellular surface interactions has been added to the efforts to change the outcome of disease. modulation oftheseprocesses seems very promising, but may temporarily leave the host without effective defense mechanism. great care therefore, must be exerted when studying this powerful two-edged sword in a clinical setting. our knowledge of the role of adhesion molecules in the intlammatory response has increased rapidly due to the availability of new reagents and mice geneticly deficient in adhesion molecules. these molecules are important in interactions of leukocytes with endothelial cells, other leukocytes, platelets, and epithelial cells. when these molecules are engaged, they can also play a role in activating leukocytes and their effector functions. in the venules of the systemic circulation, adhesion often occurs through a series of sequential interactions. initial interactions are mediated by members of the selectin family to loosely associate the leukocytes with the endothelium and are followed by firm adhesion requiring members of the integrin and immunoglobulin family. later interactions with endothelium may require pecam. adhesion molecules are usually required for leukocyte emigration in response to extravascular stimuli and for neutrophil-mediated endothelial cell injury. they are critical for host response in many diseases including infections. however, when the inflammatory response results in damage to host tissues, patients may benefit from blocking the leukocyte response. anti-adhesion molecule agents are an important potential antiinflammatory therapy. the focus of anti-adhesion therapy may be at any step of the sequence. diseases where anti-adhesion molecule therapy may benefit patients include ischemia/reperfusion injury in many organs, ards and mof, and transplantation, both to protect the donor organ from ischemia/reperfusion injury and to inhibit graft vs host disease. many strategies have been considered and include: 1) blocking the ability of adhesion molecules to recognize their ligand using antibodies that have been humanized or soluble receptors linked to igg to prolong their circulating halflife, 2) blocking the ligands for adhesion molecules using soluble adhesion molecules, peptide analogues, or oligosaccharides, and 3) blocking the production of the adhesion molecule using anti-sense oligonucleotides. because the synthesis of adhesion molecules is usually regulated by cytokines, inhibiting the action of cytokines is another potential site for interrupting the adhesion process. although important issues of safety must be evaluated, the potential for modulating the inflammatory response make this an exciting area of improvement in health care delivery. claire m. doerschuk, m.d.; riley hospital for children, room 2600; 702 barnhill drive; indianapolis, in 46202 usa. modulation of neutrophil-endothelial cell adhesion with anti-cdl i/cd18 monoclonal antibodies as a therapeutic modality. ch wortel, repligen corporation, one kendall square, building 700, cambridge, ma 02139, usa. the central role of inflammatory cells in the pathogenesis of lung and systemic organ injury is well recognized. binding of neutrophils to endothelial cells and migration into the parenchyma are largely regulated by complementary adhesion molecules. the leukocyte integrins are glycoproteins expressed on the neutrophil surface and in the cytoplasmic granules. integrins consist of a common beta2 or cluster differentiation (cd)18 chain covalently linked to one of three different alpha chains (cdlla, cdllb, cdilc) and exist on the cell surface as three distinct heterodimers. cdlla/cd18 is expressed on all leukocytes, whereas cd 11 b/cd 18 and cd 11 c/cd 1.8 are restricted to cells of myeloid origin. cd i 1/ cd 18 interacts with intracellular adhesion molecule-1 (icam-i), its ligand on endothelial cells. the potential for monoclonal antibodies to adhesion proteins to reduce vascular and tissue damage has been studied in a large number of experimental models. protective effects with anti-cd18 antibodies have been observed in a wide variety of inflammatory, immune, and isehemia-reperfusion injuries, such as arthritis, burns, endotoxic shock, bacterial meningitis, autoimmune diabetes, nerve degenemrion, allograft rejection, allergic asthma, acute lung inflammation, skin lesions, and ischemia-reperfusion models of the intestine, myocardium, lung, skeletal muscle, and central nervous system. protective effects have also been observed in animals resuscitated following hemorrhagic shock. blockage of cd18, however, would affect all leukocytes, as would antibodies to cdlla/cdi8. targeting cdllb/cd18 would affect cells of the myeloid lineage only, which could prove to be beneficial. cd11b/cd18 is not only involved in transendothelial migration, but is also implicated in adherencedependent formation of reactive oxygen species. blocking cd1 lb/cd18 may therefore not only reduce the numbe r of leukocytes accumulating in the tissue, but also attenuate the oxidant stress of infiltrated neutrophils. anti-cd 11 b treatment has been used effectively to reduce tissue injury initiated by ischemia-reperfusion, complement activation and endotoxemia. altering the host response by modulating the function of adhesion molecules may attenuate the inadvertent injury caused by leukocytes, but may also temporarily leave the host without effective defense machinery. overall, animal studies suggest that it may be safe to inhibit neutrophil adhesion for a limited period of rime. these observations will have to be confirmed in carefully designed clinical trials. c, arbobydrams are ubiquizom constir~uts of cell sv.rfaees, and possess many c~xssfies ttm~ m~,e ~em ide~. canaidates for r~ognifioa mole~ule& in m~y systems whe,~ cer udhesioa ~lays a critical ro~ car~hydram l:~dtag ~otegas have been shown to b~ad tocell surfa~ earbohydzaxes ~nd pzrl~pate in cell-ceil lumtaefion& such sys,.ems include ~rti~za~io=, deveaopmeat, l~thoge~-hcet reeog--ition ~d i~zmmadon_ in particular, tb.z recent di%~ve~ of lhe selec~ and th~ impo.~a~c~ in teukccy~udo~lelium adh~ion has -~f~m av.c~on ok l~in m~ted cell adhe~on. s~vere/poten~s/cs.rbohydr~ l~ga~s hrve ~e~l ~u~ilied for ~he s~lcc~ins. the,~ c~u be broadly di,,sded la~o ~wo m'oups -sibyl l~wis x m~ mh~.~l oligo~chadd~s, ~d sf/~ ca~ohydmma, all ~:~ ~l~dns bind m siflyl l~wis x (sie$ o!igos~ccb.e.rkms, zlthou~ w~ differing avi~re~. 'we have i~¢n~ed the functional g~oups 0a s~ex ~n~ med/a~ ~he b~u ~di~g of ~h~ c~b0hydmm = e-se/sedm we have used ~hat iv.formation to sya~esize sle ~ '~mt0gs r.he, t focus on replacing slslic ~sd ~nd fuc0s¢ wi~ simpler, more stable strunt~es. a[~ou~a ~ proeer~ is ongoing, we hve been ~ucee,.~ful a~ rep~aein8 t.ke si~ic a~id. residue wi~ std.fzte. ~ce~ or la~c amd groupa we t'we ex2aninad &e ten, bunion of ezed~ hydroxyl group of the fizeose residue ~ billding of e-, l-~nd p-selees..u. we have also found m~fi~fio~ of the reducing end ~¢.cha'i~ ~z increase mtagovsst activity. the, m¢ond. group of figs,rids a.r eontzin su~a~ 0u a ea.rbohydr~t¢ support,, und seem to bi~.d to t~e sele~ti~s wi~ dlf:ferem characteristics c0.an does sle:, s=h compounds are m2ogniz~d by l-selects. md p-selectia, bur., in genera/, not e, selecti~ these dam may mdicam r.hat l-and p-s~ ¢at~ h~d via o, second ~te thaz operates lu~.ead of, or in conjunction with ~tc sle" b~ding ~iite. dam rela~&~g to ±e, se two types of ,ml~ liga~ds have beam t~ed to desig~ potential the2~peutics for i~fi~anmat0ry disease. lr:rng maimai models of acute lung lu3ury we can demo~trate that eompmmds that inhibit seleetiu birding ~ ~i~o hzve ber~ficial effects when uc~d in rive. progressive microvascular damage in the tissue adjacent to a cutaneous burn injury results in extension of burn size. the role of leukocytes in the pathogenesis of microvascular injury was investigated by inhibition of their adherence to the microvascular endothelium using monoclonal antibodies directed to leukocyte cdi 8 or its endothelial ligaud, intercellular adhesion molecule-1 (icam-1, cd54). a model of thermal injury was developed using new zealand white rabbits. two sets of three full-thickness burns separated by two 5 x 30-mm zones were produced by applying brass probes heated to 100°c to the animals' backs for 30 sec. cutaneous blood flow determinations carried out with a laser doppler blood flowmeter were obtained for 72 hours. there were five experimental groups: controls given saline alone; animals given monoelonal antibody to the cd 18 r 15.7 prior to burn injury (pre-r 15.7); animals given r 15.7 30 min after burn injury (post-r 15.7); animals given a monoclonal antibody to icam-i, r 6.5 prior to burn (pre-r 6.5); and animals given the r 6.5 30 min postburn injury (post-r 6.5). blood flow in the marginal "zone of stasis" between burn contact sites was significantly higher in the antibody-treated animals. administration of the antibodies 30 min after injury was as effective as preburn administration in preserving blood flow. at 72 hr post-burn all antibody -treated animals had blood flow in the areas at risk for progression (i.e., the zone of stasis) at or above baseline levels while the control animals had levels equal to 34.7 _+ 12% of baseline (p < 0.05 by analysis of variance and mann-whitney u test). these results indicate that leukocytes play an important role in the pathogenesis of burn wound progression, and that this progression can be attenuated by moduiating adherence to endothelial cells. a wealth of information now supports the hypothesis that inhibition of cell adhesive mechanisms will nter the course of immunologicand inflammatory processes. what remains unclear is whether inhibition of specific mechanisms wfl[ be of therapeutic benefit in any specific human disease. current data derived from animal models are not inconsistent with the hope of therapeutic benefit, but techniques for inhibition (e.g., antibodies, antisense oligonucleotides, inhibitory peptides, inhibitory carbohydrates, smaii synthetic inhibitors, etc), tissue and species differences in the relative contributions of adhesion molecules to the inflammatory process, and the cascade model of adhesive interactions are all confounding issues, making predictions of therapeutic benefit in any specific human disease process very difficult. additional concerns involve the potential roles of adhesive mechanisms in host resistance to infection. as human therapeutictdals are initiated, more exact information on the roles qf specific adhesion molecules in human disease should emerge. inhibition of leukocyte adherence to endothelial cells can represent a novel therapeutic approach to septic shock. we performed a pilot study to evaluate the safety and tolerability to cy-1787, a monoclonal antibody against human e-selectin, in patients with septic shock. septic shock was defined by clinical signs of sepsis, a documented source of infection, and fluid-resistant hypotension requiring the use of vasopressors. eleven patients entered the study, but 2 patients who died during the first 24 hours were excluded, as this was part of the protocol. cy-1787 was administered as a single intravenous bolus of 0.1 mg/kg (n=3), 0.33 mg/kg (n=3) or i mg/kg (n=3) mg/kg. the antibody was well tolerated. none of the 9 patients died during the 28 day follow-up period. organ failure was assessed for 5 organs (cns, lungs, liver, kidneys and coagulation). the mean number of organs failing, which was initially 2.7 ± 1.2, decreased to 0.2 ± 0.4 at the end of the study (p<o.o01). these results are therefore encouraging. administration of an antibody to e-selectin in patients with septic shock requires further study. the importance of cytotoxic t lymphocytes (ctl) in the host defense mechanism against viral as well as parasitic and bacterial diseases is becoming increasingly appreciated. the design of vaccines to generate cytotoxic t ceils from nonviable or subunit constructs, however, poses challenges due to the unique characteristics of antigen processing and presentation by class i major histocompatibility complex (mhc) molecules. since the final antigen recognized by ctl are short peptides capable of high affinity binding to class i mhc molecules, we have designed strategies: a) to define the structural characteristics of peptides that are required for their high affinity binding to mhc; and b) to formulate these peptides into immunogenic vaccines that are capable of generating cytotoxic t lymphocytes. the strategy and results to accomplish these two goals will be discussed. amnon altman and erich gulbins ras proteins represent essential intermediates in receptor-mediated growth and differentiation pathways. they couple signals initiated by receptor or nonreceptor protein tyrosine kinases (ptks) at the cell surface to cytoplasmic targets which coordinate signal transduciion to the nucleus. ras also participates in t lymphocyte activation initiated by the antigen-specific t cell receptor (tcr)/cd3 complex, which leads to lymphokine production and proliferation. receptor ligation causes activation of associated ptks of the src and syk families as the earliest identifiable event in this pathway. the importance of ras in t cell activation is indicated by the findings that an oncogenic ras protein activates the interleukin-2 (il-2) gene promoter; conversely, a transdominant inhibitory ras mutant inhibits this event and the activation of map kinases. the rate-limiting step in ras activation is the exchange of bound gdp for gtp, which is catalysed by guanine nucleotide exchange factors (gefs). we identified the sh2/sh3 domain-containing vav protein, which is selectively expressed in hematopoietic cells, as a ras-specific gef coupled to several hematopoietic receptors. vav can be activated by two pathways, i.e., by ptk-mediated phosphorylation or by binding of diglyceride second messengers to a cysteine-rich, protein kinase c (pkc)homologous vav domain. these pathways are independent as demonstrated by structure/function analysis and the use of pharmacological inhibitors. the two activation pathways may enable vav to integrate signals from distinct hematopoietic cell receptors, and couple them to ras. t cells express another, ubiquitous ras gef, i.e., sos, which is known to be coupled to activated tpk receptors. t cell activation leads to membrane association of a signaling complex consisting of sos and two other signaling proteins, grb-2 and shc, via direct binding of the shc sh2 domain to the tyrosine-phosphorylated chain of the tcr/cd3 complex. thus, multiple receptors, ptks and ras activators participate in signaling pathways that lead to hematopoieitc cell growth and differentiation. the interactions and function of these various signal transducers will be reviewed. phagocytes. immunity is mediated by specific t lymphocytes, cytokines produced by these specific t cells activate antimicrobial capacities in mononuclear phagocytes, thus contributing to protection. other immune mechanisms also participate in the acquisition of resistance. on the other hand, t cells are also crucial for many pathologic sequelae of intracellular bacterial infections. although ifn-y is central to macrophage activation, synergistic and antagonistic effects with other cytokines occur. the cd/3 t cells, representing the major t cell population in experimental mice, are the main mediators of antibacterial immunity; an auxiliary role for y/~ t cells appears likely. the c~//3 t cell population further segregates into cd4 t cells which recognize microbial peptides in the context of gene products of the major histocompatibility complex (mhc) and cd8 t cells which see their peptide in the context of mhc class i. the -y/~ t cells are generally double-negative. knock-out mice in which the genes for various t cell receptor chains, for mhc class i or mhc class ii molecules, for cytokines or cytokine receptors had been deleted, have provided extremely helpful tools for analyzing the role of t cell subsets and cytokines in antimicrobial immunity. using these mutant mice, the role of distinct t cell subsets and cytokines in bacterial elimination and granuloma formation was analyzed in detail. these studies underline the central role of a//3 t cells and ifn-y in antibacterial immunity and, furthermore, show a distinct function of 7/$ t cells. moreover, these studies show that the relative contribution of cd8 or cd4 t cells to antimicrobial immunity varies in response to different pathogens. apoptosis is an active form of cell death in which the cell participates in its own demise, providing the biochemical pathways that trigger and mediate the complex events of the process. although this phenomenon has been studied for many years, it is only recently that significant progress has been made towards the elucidation of the molecular mechanisms that control apoptosis. once such mechanism came as something of a surprise: in a number of eases, apoptosis can be promoted by the action of the c-myc protein, a protooncogene product that had previously been associated only with cell proliferation (and presumably, survival) . expression of c.myc in cells can cause them to enter an apoptotic pathway or proliferate, mad this "decision" depends upon additional signals, such as growth factors that inhibit apoptosis and allow the ceils to proliferate. thus, c-myc directs cells out of rest and the choice of proliferation or death depends upon the presence or absence of survival signals. other survival signals are generated by oncogene products that can cooperate with my¢, including bcl-2 and abl there are implications of this interplay between apoptotie and anti-apoptorlc signals in cell iransformation and the resistance of some tumor ceils to therapeutic agents capable of inducing apoptosis. oncogene interactions therefore control the life or death of transformed ceils but also have important roles in normal, development processes. in the immune system, developing t ceils are "screened" for potential self-reactivity by a process of negative selection, in which activation via the t cell receptor (tcr) induces apoptosis. this process can be mimicked in t cell hybridomas, and appears to depend upon the expression of the c-mye protein. evidence that this represents a requirement for the myc/max heterodimer will be presented. as above, these cells are presented with the "choice" to proliferate or die and this depends not only upon myc but also other signals. thus, expression of functional v-an kinase in these ceils can render them resistant to the activation of apoptosis via tcr ligation, surprisingly, bcl-2 does not appear to be capable of blocking this form of apoptosis, and the reasons for this will provide new insights into the development of the immune system and the process of apoptosis. although the oncogene products discussed here probably do not directly participate in the biochemical process of apoptosis, they represent molecular controls on this complex mechanism. as such, they gave us new ways to look at the life and death of a cell recent attention has focused on macrophage release of cytokines, such as il-1, il-6 and tnf as important mediators of septic shock. however, serum cytokine levels have correlated poorly with outcome of septic shock. the purpose of this study was to compare the functional status of macrophages to serum cytokine levels in early sepsis by an analysis of splenic macrophage protein synthesis during abdominal sepsis. macrophage total protein distribution and biosynthetic activity was assessed by large format 2-d page, autoradiography of 35s-labeled proteins and computer assisted densitometric analysis. sepsis was induced in sprague dawley mts by cecal ligation and puncture (clp). untreated and sham operated rats served as controls. splenic macrophages were harvested at 3 and 24 hrs. following clp. serum il-6 levels were determined at 3,12, and 24 hours by the 7 tdi bioassay. by 12 hrs. 100% of the clp mls exhibited multi-microbiaj bacteremia. control or sham operated rats did not show bacteremia. clp resulted in a significant 286% elevation (p < 0.05) in serum il-6 levels at 12 hrs. there was no difference in il-6 at 3 and 24 hrs. when compared to the control groups. 2d page studies examined splenic macrophage total protein distribution and biosynthetic activity at 3 and 24 hrs. post-clp. at 3 hrs. macrophages from clp rats showed a substantial decrease in total protein pattern (353 protein spots vs 503 protein spots) when compared to non-septic control macrophages. a decrease in the number of lower molecular weight proteins (< 40 kd) was observed. at 24 hrs. post-clp, splenic macrophages from clp rats showed a substantial increase in the synthesis of low molecular weight (< 40 kd) proteins, when compared to non-septic controls. we conclude that: l) fulminate abdominal sepsis induces an early (3 hrs.) staining with cd14 and cd16 mabs will identify two monocyte subpopulations in human blood: a major population of regular monocytes, which strongly expresses the cd14 antigen (cd14++) and a minor population with weak expression of cd14 and expression of the cd16 antigen lcd14+/ cd16+ cells). in healthy control donors(n=35) the latter cells account for 45+22 cells/mm 3 and 9%+5% of the monocytes. in sepsis patients, the cd]4+/cd16+ cells can become a major population with more than 50% of all monocytes in 3 of 18 patients and with more than 500 cells/mm 3 in 4 of 18 cases. there was no correlation of cd14+/ cd16+ cells to any clinical parameter except for cd14+/cd16+ percentage and body temperature (p= 0.013). the cd14++ monocytes showed a substanttial decrease in cd14 antigen density in 9 of 11 patients. three-color-if shows that the cd14+/ cd16+ cells in sepsis patients when compared with the cd14++ monocytes exhibit a higher level of class ii antigen and a lower level of cdiib and cd33 antigens, consistent with a more mature nature of the cd14+/cd16+ cells. levels of il-6 were increased in sepsis patients: 3 of 5 patients with high numbers of cdi4+/cd16+ cells ( 200/mm 3) had high levels of il-6 (250u/ml). these data suggest that septicemia may lead to substantial changes in blood monocyte composition and this may be related to elevated levels of cytokines such as il-6. human peritoneal macrophages were exposed to increasing doses of lipopolysaccharide (lps) or a synthetic lipid a analogue (mrl 953) and production of the cytokines il-lb, il-6, tnf-a and g-csf was assessed at the protein-and mrnalevel. cells were also stimulated with low doses of lps and mrl 953 to study their "adaptation" to a secondary challenge with high doses of lps. tnf-a production after stimulation with lps could be almost completely relieved by preincubation of cells with low doses of lps and similarly, il-6 production was suppressed. surprisingly, however, "adapted" cells were able to synthesize even larger quantities of of g-csf and il-lb when exposed to a secondary lps challenge. the changes in tnf-a, il-6 and g-csf protein synthesis could a/so be detected on the mrna level, whereas transcript levels for il-lb remained unaltered as assessed by northern blot analysis. high doses of the synthetic lipid a analogue mrl 953 were also able to "adapt" macrophages for a secondary lps challenge by downregulating tnfa-and il-6-production, while priming secretion of g-csfand il-lb as well. taken together, here we describe for the first time the discordant adaptation of human peritoneal macrophages to a secondary lps stimulus in vitro. these findings appear to bear ramifications for the in-vivo endotoxin response during inflammation and gramnegative septicemia. shock states with inadequate cellular oxygen delivery may contribute to macrophage (mo) activation or dysregulation. in this study we compared the effect of transient hypoxia and endotoxin pretreatment (lps 1) on mo mediator release with a second endotoxin stimulus (lps2). methods: in vitro cultures of marine peritoneal exudate mo were exposed to 2 hr. of hypoxic or normoxic conditions, then incubated 22 hr under identical normoxic conditions _+ 10 ng/ml of lps 1 pretreatment. during the final 24 hours all m~l were exposed to a range of lps 2 concentrations. m~i supernatants were assayed for tnf, il-1, il-6, pge2, nitric oxide (no), and superoxide release. results: lps 1 markedly inhibited mo tnf release by lps 2 but hypoxia had no effect on lps 2-triggered tnf release. hypoxia increased mo il-6 production in the absence of lps 1, but inhibited lps 1 augmentation seen under normoxic conditions. pretreatment with lps 1 increased no production from lps 2 under normoxic conditions, but bypoxiainhibited this effect. pretreatment with lps1 signifcantly augmented mo release of il-i and pge 2, but hypoxia had no significant effect (data not shown) superoxide production was increased by lps 1 under normoxic conditions, but hypoxia significantly inhibited superoxide release (not shown a. lepape, c. docile, f. arpin, m. c. gutowski, v. banssillon and j. bienvenu. i aim of the study. increased levels of tnf are inconsistently correlated with the severity of sepsis. one possible explanation is a decrease ability of ceils to produce eytokines. the objectives of this study were m compare at different levels of of clinical severity the responsiveness of cytokine producing cells. this was evaluated by the response to a stimulation by lps as well as to the inhibitory effect of ptx. the technic used is a whole blood ex-vivo model, as described by desch et al. (1) . h materials and methods. different groups of icu patients, postoperative (n=12), sepsis (n=12), septick shock (n=13), and healthy control (n=10), were studied. blood was drawn in endotoxio free heparinized robes. stimulation was achieved by addition nf75 pl oflps (10ng/ml) to 675 id of whole blood in presence of various concentrations of ptx (0, 10 "5, 10 -4, 10 "3, i0"2m). after 5 hrs incubation at 37°c, the tubes were centrifuged and supematants frozen at -80°c. tnfa and/l6 were measured by elisa (medgenix r and immunotech r respectively). monocytes were quantified on a technicon h6000. the results are expressed as median values. non parametric tests are used for testing differences between groups. hi results, tnftx and 11,6, corrected for monocytes count, are given as % of their initial value, the baseline before stimulation or inhibition being 100%, 1) stimulation by lps. the control group is much more stimulated (>21000% for il6, >8000% for tnfa). blood samples taken postoperatively and in patients with simple sepsis are significantly less stimulated (>1500% for il6, >600% for tnfa ). the lowest stimulation was observed in patients with septic shock (median = 168%), some patients being not stimulated at all. 2)effects of ptx.the inhibitory effect of ptx on tnftx production is effective in all groups at 10 -3 m (reduction to less than '¼ of the median values), and is almost complete at 10" 2 m. the septic shock group has a decreased sensitivity to ptx. il6 production exhibits a lesser reduction at 10 -3 m (~ 'a to ½ of the median values), further increased at 10 -2 m. the septic shock group is again less sensitive to ptx. iv conclusion: the reduced ability of circulating monocytes to produce cytokines during severe infections is confirmed here. ptx is able to reduce significantly tnfc~ at 10 -3 m and the inhibition is nearly complete at 10 -2 m. surprisingly, there is a lesser, but significant suppressive action of ptx on il6, not found in experiments using purified monocytes. one possible explination could be the interplay between cytokines production. (1) lymphokine research (1989) cdna sequencing constitutes a powerful method of measuring steady-state mrna levels for all genes transcribed in a given cell or tissue at a particular stage of differentiation. by comparing transcript abundance both prior to and following differentiation, individual genes can be identified whose transcription is regulated both positively and negatively. in order to examine monocyte activation, the human monocyte line thp-1 was induced with phorbol ester (48 h) and activated for 4 h with lipopolysaccharide (lps) after which polya + rna was purified. the rna from control and lps-treated cells were each used to construct a cdna library under identical conditions, and all resulting clones were selected for cdna sequence analysis. each clone sequence was evaluated by matching with both genbank and our own gene databases. very different patterns of gene expression were seen in the two libraries, the latter reflecting very high levels of known inflammatory mediators such as il-1 and tnf. a second set of libraries were made from umbilical vein endothelial cells (huvec), both with and without lps stimulation, and were analyzed in a similar fashion. the effects of lps induction on specific gene transcription in both cell types will be discussed. t. tadros, md, th wobbes, me) phd, rja goris, md phd to investigate whether the preactivation of regional macrophuges by liposomes containing muramyl tripeptide (mtp-pe) can counteract the detrimental effect of blood transfusions on both anastomotic repair and host susceptibility to infections. methods eighty lewis rats received lmg/kg of either empty or mtp-pe encapsulated liposomes, intraperitoneally (ip). twenty-four hours thereafter, the animals underwent resection and anastomosis of both ileum and colon, and received 3 ml of either saline or blood from brown norway donors,iv. the animals were killed 3 or 7 days after surgery and examined for septic complications and anastomotic repair. the average anastomotic strength, as assessed by bursting pressure (+sd), was significantly diminished in the transfused animals, as compared to the non-transfused animals (ileum;day 3; 44-+8 vs 99+7, p<0.001). transfused animals pretreated with mtp-pe encapsulated liposomes showed a significant improvement of their anastomotic bursting pressure (93+9, p<0.001 vs transfusion). pretreatment with mtp-pe encapsulated liposomes decreased significantly the incidence of anastomotic abscesses in transfused animals ( from 90% in ileum on day 3 to 20%, p<0.001). conclusions preactivation of regional macrophges by intraperitoneal administration of mtp-pe encapsulated liposomes prevents the detrimental effects of transfusions on anastomotic repair and reduces the incidence of intraabdominal sepsis. academic hospital nijmegen, dept of general surgery, pb 910 i, 6500 hb nijmegen, the netherlands. leukemia cell line, teip-1. robin s. wa, gner*, perry v. halushka "~, and james a. cook*, departments of physiology , pharmacology "l" and medicine "t, medical university of south carolina, charleston, s.c. 29425. adherence of monoeytes to endothelium and extracella/ar matrix proteins is essential for accumulation at sites of inflammation. txa2, an arachidonic acid metabolite, inhibits human monocyte chemotactic responses suggesting that txa 2 may alter monocyte adhesiveness. we selected the thp-1 cell line, a human monocytic leukemia cell line to further investigate the effect of txa 2 on adhesion. we tested the hypothesis that txa 2 alters lpsinduced adhesion of thp-1 cells and that txa 2 exerts its effect on adhesion via a camp dependent mechanism. thp-i cells were exposed to s. enteritidis endotoxin (lp.g/ml) _+ the cyelooxygenase inhibitor lndomethacin (in), the txa 2 mimetic i-bop (0.11.tm,) or txa 2 receptor antagonists bms180291 and l657925 (10~m). cells were allowed to adhere for 24 hours and adherent protein/well was determined. lps-induced a significant (p<0.05;n=7) increase in adherence of thp-1 cells (basal, 2.4+0.85gg protein/well; lps, 20.4+_6.0p.g protein/well). the amino acid glutamine is an essential compound for synthesis of purine and pyrimidine basis and therefore necessary for rna-and dna synthesis. in human plasma the concentration of glutamine is between 0.5 -0.6 mm, and is reduced in septic patients up to 80 % (0.1 -0.2 mm). monocytes play a central part in the inunune system and it was of interest, whether glutamine is involved in the modulation of cell surface markers and phagocytosis of these cells. human peripheral blood mononuclear ceils were obtained from 500 ml heparinized blood of apparently healthy donors by ficoll-paque density gradient and isolated by counterflow elutriation. the puritiy was more than 90 %. subsequently cells were cultured in phenolred-free rpmi 1640 medium with various concentrations of glutamine (0.05, 0.1, 0.2, 0.3, 0.6, 1, 2 mm) in teflon-fluorinated ethylene propylene bottles to exclude cell adhesion and possible cell activation. aider seven days culture, cell viabilty was determined by trepan blue exclusion and varied between 70 and 90 %, independent of glutamine concentrations. cell surface markers were detected by flow cytometry, noaspecifie phagoeytosis was measured with latex beads and specific phagocytosis with opsonizied e.eoli using a facscan. lower concentrations of glutamine decreased the expression of hla-dr and icam-1/cd54 on monocytes in a dose-dependent manner. the receptor for fc'/rucd64 as well as the receptors for complement cr3/cdllb and cr4/cdllc were down-regulated. cr1/cd35 which is only slightly expressed on monocytes was not influenced. furthermore, no effects on the expression of cdi4, the receptor for transferrin cd71 and fc'friii/cd16 were seen. our data indicate, that lower concentrations of glutamme influence the phenotype of monocytes. we are now interested to study whether glutmnine influences non-specific phagocytosis, or whether specific phagocytosis correlates with the decreased expression of fc'/r and complement receptors. we investigated immunologically more than 300 patients who were admitted to icu because septic syndrom during the last four years. patients were immunologically followed up 2-3 times per week until release from icu. the expression of hla-dr antigen on monocytes turned out to be the best prognostic parameter. the persistence (> 5 days) of low hla-dr expression (< 30%) predicts fatal outcome (mortality > 85%). the altered phenotype was associated with a functional deactivation of monocytes (diminished apc, ros formation, cytokine secretion). we called this phenomenon "immunoparalysis". ifn-gamma and gm-csf were able to restore the altered phenotype and function in vitro. however, addition of autologous plasma from septic patients with "immunoparalysis" to these cultures prevented the cytokine-induced restitution. the inhibitory activity could not be removed by dialysis. therefore, we started a study to prove the therapeutic efficacy of plasmapheresis. indeed, [7 of 33 patients recovered from "immunoparalysis" following repeated plasmapheres; 16 of them survived (46 %). 7 patients recovered temporarely and 9 patients did not respond (all 16 died). the survival rate in the control group of septic patients with persistent "immunoparalysis" was 4 of 38 (11%; p<0,01). in summary, plasmapheresis in association with immune monitoring may be an alternative strategy to improve survival rate in severe sepsis. taurolidine, a synthetic taurine-formaldehyde derivative has antiadherent, bactericidal and anti-lps properties functioning primarily through binding of the lipid a region of the lps molecule. the active derivative of taurolidine, taurine, modulates calcium channel activity, critical to the initiation of a number of immunostimulatory pathways. we hypothesised that taurolidine may have direct immunostimulatory activity. the aim of this study was to investigate the immune effects of taurolidine on peritoneal macrophage (pmo) function and then determine the role of taurine in this response. study 1: in vivo stimulation:cd-1 mice (n=35) were randomized to receive taurolidine (200mg/kg bw/i.p.) or saline cor~trol. peritoneai cells were harvested after 24 hours and were assessed for pm0 function [superoxide anion generation (o2-), nitric oxide (no), tumor necrosis factor (tnf), fc/cr3-mediated phagocytic function (phago) study 2: control pm0 were harvested and cultured in vitro with taurine (1.0mg/ml for 10 hrs), after which time they were assayed for 0 2-and tnf release. in vivo stimulation with taurolidine taurolidine has specific immunological effects on m0. release of the inflammatory mediators 0 2-and tnf, and fc/cr3-mediated phagocytosis were significantly increased, while release of the endothelial relaxing factor no was significantly reduced. in addition, the amino acid taurine, which is released as a byproduct of taurolidines breakdown has an immunostimulatory effect on pmo and may be the active moeity of the compound tanrolidine. in sepsis, a number of mediators which affect vasomotor tone and cardiovascular function are produced. inasmuch as sepsis causes decrease in systemic vascular resistance (svr), attention is usually focussed on vasodilators such as lactate, tumor necrosis factor, interleukin-i & 2, and nitric oxide. but injury and inflammation als0 cause production of several vasoconstrictors whose effect may not be evident in changed svr, but may significantly affect organ blood flow or function in the paracrine environment. endothelin (et) is a 21 amino acid peptide vasoconstrictor produced by ischemic or injured endothelial cells (ec's). et is also a potent constrictor for renal mesangial and coronary vessels, an endocrine regulator, and a negative cardiac inotrope. systemic et levels increase significantly in hypoperfusion and ischemia. while et is principally produced by ec's, we asked if human monocytes might also produce et and thereby regulate vasomotor tone in areas of inflammation. monocytes from healthy donors were separated on ficoll, resuspended in rpmi 1640 +5% fetal calf serum and stimulated with i0 ug/ml endotoxin (lps). et was measured by radioimmunoassay. lps-stimulated monocytes produced 87 ! 6 fm of et/106 cells (vs. unstimulated controls of <25). this calculates to 15-30% of the amount of et observed in patients with low cardiac output, sepsis or ischemia. we conclude that et is a cytokine produced by both ec's and monocytes with potent effects on numerous cells and organs in the critically ill. wuppertal 5, germany we and other authors showed that fatal outcome in septic disease is associated with a decreased capacity of peripheral blood monocytes for the in vitro production of proinflammatory cytokines, especially tnf-alpha. we found that this monocytic deactivation is completed by a persistent and marked decrease of hla-dr expression on monocytes (< 30 % hla-dr+ monocytes) and a diminished antigen presenting activity whereas the capacity to form the antiinflammatory il-1 receptor antagonist remains high. in order to evaluate the in vivo situation and to determine at which level tnfproduction/secretion is altered we assessed the tnf-alpha mrna expression in freshly isolated peripheral blood mononuclear cells (pbmnc) from septic patients. tnf-mrna was onty rarely detected by semiqaantitative polymerase chain reaction in pbmnc's from septic patients with monocyte deactivation. meanwhile, it was found in almost all pbmncs from septic patients without monocytic deactivation. we wondered, whether il-i0, which ,is known to depress monocytic proinflammatoly response and mhc class ii expression, could be one of the mediators in fatal sepsis. in fact, we found that il-10 message in pbmncs of septic patients peaked in the beginning phase of monocytic deactivation. in further investigations we found that tnf-administration can induce monocytic deactivation in a murine model/n vivo and provoke il-10 message in human pbmncs in vitro. these results support our hypothesis that an excessive delivery of proinflammatory cytokines in a first phase can induce an overwheiming inhibitory feedback, mediated by immuninhibitory mediators like il-l0, which leads to often fatal monocytic deactivation in a second phase. interferon-gamma which is known to counteract il-10 production and the effects of il-10 on monocytes restores the function and phenotype of monocytes from septic patients with monoq, te deactivation in vitro and could be a possible therapeutic agent in otherwise fatal sepsis. our laboratory previously reported that lps dependent macrophagederived tnf-a production can be enhanced by pretreatment with lps at substimulatory lps priming doses coincident with a suppression of lps dependent nitric oxide (no) production (zhang and morrison, j. exp. med 17:511, 1993) . in order to extend the characterization of these lps priming effects in mouse macrophages, we examined the capacity of substimulatory lps to modify lps dependent il-6 production. macrophages were obtained from peritoneal exudate of thioglycollate treated c3heb/fej mice and cultured in rpmi 1640 medium containing 10% fetal bovine serum. macrophages were pretreated with various subthreshold stimulatory concentrations of lps (olll:b4) for 6 hours, washed three times, and then stimulated with the effective stimulatory concentration of lps for 18 hours. the amount of il-6 in the supernatant was measured by il-6 dependent cell line (b9 and 7td1) proliferation assay. il-6 was produced by macrophages at lower threshold doses of lps than those required for tnf-o~ or no production. subthreshold doses of lps modulated il-6 production in a biphasic manner characterized by an initial suppression and then potentiation. higher doses resulted in secretion of il-6 during the initial incubation with lps and subsequent desensitization. il-6, like tnf-~ and no, is, therefore, also affected by lps pretreatment. moreover, tnf-a and il-6 shared the similar potentiational pathway, but differed by the fact that only il-6 was inhibited. (supported by r37 ai23447 and po1 a54474.) department of microbiology, molecular genetics and immunology and the cancer center, 1000 wahl east, university of kansas medical center, kansas city, ks 66160-7832. korolenko t.,urazgaliev k.,and arkhipov s. the role of macrophage (mph) stimulation in mechanism of protective effect of new immunomodulators yeast polysaccharides -heteropolysaccharide cryelan and homopolysaccharide mannan rhodexman (both produced by petersburg chem.-pharm. inst.) was studied. in vitro according to nst test incubation of murine peritoneal mphs with cryelan or rhodexman, 150 ~g/ml,30 min was followed by increase of potencial microbicidic activity of mphs. in vivo mph stimulation by immunomodulators studied included increase rate of carbon particles phagocytosis during single i.v. or i.p. mode of administration to mice 1-14 days after (peak at 2nd day for i.v. and 5th day for i.p. mode of administration of the same dose of 5 mg/100 g b.w.).the preliminary injection of cryelan (5 mg/100 g, 24 or 48 h before) to mice with acute cold stress (-10 ° c, 1 h) revealed protective effect restorating the value of depressed phagocytosis up to the normal level;the positive effect on ultrastructure of hepatocytes was noted also.there was no changes of plasma corticosterone level between group with acute cold stress and mice with cryelan + acute cold stress (several fold increase comparatively to the control mice).as was suggested, the mechanism of protection can include mph stimulation and secretion of some acute phase proteins responsible for positive effect of immunomodulators. new yeast polysaccharides cryelan and rhodexman can be used for macrophage stimulation,especially in pathological states. immunomodulators were shown to increase production and secretion of lysosomal enzymes (like zymosan). secreted enzymes,especially cysteine proteinasescathepsins b and l -involve in the process of inflammation;however, excessive release of these enzymes may lead to noncontrolled proteolysis followed by tissue degradation (assfalg-machleidt et al.,1990) .the effect of zymosan,bcg and new immunomodulator carboxymethylglucan (cmg), second fraction on secretion of lysosomal enzymes by murine peritoneal macrophages was studied. zymosan increased the secretion of n-acetyl-~-d-glucosaminidase and ~-galactosidase into the culture medium (3-4 fold); bcg possessed similar effect.cmg in the same concentrations (50 /~g/ml) increased release of these enzymes only saightly (1.6 times).it's known that zymosan-induced secretion reflects the enzyme release from formed lysosomes (warren,1989) .it was suggested that cmg activated macrophages via interaction with scavenger-receptors,followed by weak secretion of lysosomal enzymes and as a result decrease of tissue damage. in vivo zymosan induced stimulation of mononuclear system of phagocytes followed by increase of cysteine proteinases activity in liver at the 5th day. in the same time in blood n-acetyl-~-d-glucosaminidase and n-acetyl-~-d-galactosidase activity increased 2-5 fold. it was concluded that in drug design it's possible to select such immunomodulators,e.g. cmg,which can activate mononuclear system of phagocytes and do not damage tissue. endothelin-i (et-i) is produced by injured/ ischemic endothelium, mobilizes intracellular ca ++ and is a potent vasoconstrictor. it is also a ca ++ agonist for anterior pituitary or renal mesangial cells and monocytes. et-i causes monocytes to produce interleukin-l, 6, 8, prostaglandin e2, and substances which trigger neutrophil superoxide production. et-i levels increase in shock and et may play a role in activating leukocytes post shock causing reperfusion injury. but blood flow experiments suggest splanchnic circulation changes more profoundly in shock than peripheral circulation. we therefore asked if et-4 (or vic), the et which predominates in splanchnic vessels, had any effect on monocyte cytokine production. human monocytes from health~ blood donors were separated on ficoll. 0.5 x 10ucells/ ml in rpmi 1640 + 10% fcs were incubated i0 min., 6 & 24 hrs. with 10 -8 m et-i, 10 -8 m vic or i0 ug/ml of lps. supernatants were assayed by elisa. we have shown that low dose endotoxin pretreatment (lps 1 ) for 24 hrs markedly inhibits the macrophage (mo) release of tumor necrosis factor (tnf) and increases interleukin-1 (il-i) in response to a subsequent endotoxin stimulus (lps2). in this study we examined the kinetics of lps 1 inhibition of tnf and augmentation ofil-1. methods: murine peritoneal exudate mo from balbc mice were exposed in vitro to medium or 10 ng/ml of lps1 for intervals of 0 to 24 hours. culture medium was then replaced with 0, 10 or 1000 ng/ml of lps2 for 24 hrs. tnf and il-1 in mo supernatants were measured by specific bioassays. during sepsis endotoxin (lps) activates macrophages (mo) to release mediators such as tumor necrosis factor (tnf), interleukin-6 (il-6), interleukin-i (il-i) and prostaglandin e2 (pge2). we showed that preexposure to lps (lps 1) alters the response of murine m~i to subsequent lps stimulation (lps2). we hypothesized that in vitro cytokine release by lps2 in human monocytes (mo) is also be altered by preexposure to lpsi. methods: human peripheral blood mo were obtained from healthy volunteers (n=8), cultured in vitro 24 hrs, then pretreated 24 hr _+ lps 1-cultures were then stimulated with lps 2 and mediators in mo supernatant measured: tnf, il-i, and il-6 by specific bioassays, pge2 by immunoassay kit. serum cytokine levels (specific elisa kits) were compared to in vitro supernatant levels. data is expressed as % control_+sem, lps2= 1000 ng/mh the table shows that all mediators were increased, in the absence of lps 1. pretreatment with lps 1 resulted in complete inhibition of lps2-triggered tnf release. in contrast, lps 1 significantly increased mo secretion of il-1, il-6 and pge 2 (data not shown). serum cytokine levels were as follows: tnf 407_+21, il-i 241+208, and il-6 8.0-+2.5 ng/ml. these serum levels were low, showed an extremely wide variation, and did not correlate with in vitro lps2-triggered mediator production. conclusion: human monoeyte mediator production is differentially regulated by preexposure to lps 1. provocative in vitro testing of monocytes may ultimately be clinically useful to identify prior in vivo lps exposure or mo macrophages release numerous secretory products involved in host defense and inflammation. activated macrophages with cytokines produced have been implicated in tissue damage in sepsis and multiple organ dysfunction. aimed to elucidate the organ-association phenomena,this study is to compare peritoneal macrophage(pm),alveolar macrophage(am), and kupffer cells(kc) during sepsis in terms of cellular protein contents as symbol of activation by flow cytometry analysis. sepsis were produced by cecal ligatien and perforation (clp) in wistar rats weighing 210-290g.pm were obtained by peritoneal lavage,am by bronchial lavage and kc by incubating the collegenase digested liver with pronase-e. leukocytes have been implicated as a mediator of the microvascular dysfunction associated with reperfasion of ischemic tissues. a role for ieukocytes is largely based on observations that rendering animals anutropenic with anti-neutrophil serum or preventing leukocyte adhesion with monoclonal antibodies attenuates the increased fluid and protein leakage from the vaseulature that is normally observed in postischemic tissues. we have recently undertaken studies designed to determine the relationship between leukocyte-endothelial cell adhesion and albumin leakage ia rat mesenterlc venules exposed ~o ischemia-reperfusion (i/r). leukocyte adherence and emigration as well as albumin extravasafion were monitored in single postcapillary venules using iatravital fluorescence microscopy, lschemia was induced by complete occ!usion of the superior mesenteric artery and ~dl parameters were monitored at various intervals following reperfusion. the magnitude of the leukocyte adherence and emigration, and albumin leakage elicited by i/r was positively con-elated with the duration of ischemia. the albumin leakage response was also highly correlated with the number of adherent and emigrated leukocytes. monoclonal antibodies against the adhesion glycoproteins cd18, cdllb, icam-1 and l-selectin, but not p-or e-selecdn, reduced i/r-induced leukocyte adherence and emigration as well as albumin leakage. phauoidln, an f-aetin stabilizer, largely prevented the emigration (but not adherence) of leukocytes and greatly reduced, the raicrovascular protein leakage. plateletleukocyte aggregates were formed in postischemic vemdes; the number of aggregates was reduced by antibodies against p-selecdh, cdilb, cd18, and icam-1, but not e-selectin or lselectin. a significant fraction of the mast ceils surrounding the posteapillary venules degranulated in response to ischemia/repeffusion, but mast cell stabilizers did not afford protection against the albumin leakage elicited by i/r. these results indicate that reperfusloninduced albumin leakage is tightly coupled to the adherence and emigration of leukocytes in posteapillary venules. this adhesiomdependent injury response is primarily mediated by cdllb/cdi8 on activated neutrophils and icam-1 on venular endothellum, and appears to require l-selecda dependent leukocyte rolling. mast cell degranulation does not appear to conwibate to the vascular pathology associated with i/r. m.d. rod=iek, boston, ma, usa the polymorphonuclear neutrophil (pmn) has long been known to pa~tlcipats in the inflammatory rebpons~ as a phagocyte and killer of invading organisms, but little attention has been given to its potential as a participant in the in~une interaction of lymphocytes and macrophages. we and others have shown that the pmn may have i~m~/nomcdulatory effects both in vitro and in vlvo. more recently it has been proven that the pmn can make mrna for and secrete the proinflammatory oytokines illa, il-ib, tnfs, il-6 and il-8 as does the other major circulating phagocyte, the monocyte/macrophags. furthermore it has been shown to make the potentially autoregulatory oytokines gcsf and gmcsf. these functional capabilities suggest that the pmn is not an wend cell ~, but one which has a potential role in regulation cf ~he immune response and that this potential ~cle should no longer be ignored when considering the immune abnormalities existing in patients following majo~ injury or surgery. we have investigated the proinflaznmatory oytokine secretion patter~ by pmn in patients following major ~hermal or tra~matic injury and in volunteers fellowinq endotoxemia. ?ollowing major injury there is variable pmn secretion of these cytokines when stimulated in vlero. following endotoxemia in a group of human volunteers pmn showed a hypo=esponsivenesa to lps 4 hrs following endotoxin infusion followed at 24 hre by an overshoot. pretreatment with steroids modulated this overshoot phenomenon, suggesting that receptors for steroids are involved in the regulation of cytokin® secretlon by fmn. these results sugges~ that the pmn, the most numerous cell in the circulation and the first to respond to an ins~l~ may be a so~rce of the prolnflammatory cytokine cascade following injury that has been recognized as significant in the process which often leads to multiple o;gan failure, the immunosuppresslon which occurs following major thermal injury may predispose these individuals to infection and sepsis, which remain a significant cause of morbidity and mortality. included among the many immune aheratlons are the p2 integrln (cdlla, b,c/cd18) dependent activities of adhesion, chemotaxls, diapodesls, and phagocytosls. our investigations indicate that, following major thermal injuries, the expression of the [~2 integrlns, but not cd14, is significantly decreased on neutrophlls (pmns). it remains unclear if pmns from thermally injured patients respond normally to lps, the effects of treatment in vitro with lps and f-met-leu-phe (fmlp) on the expression of cdtlb was examlned on pmns from the peripheral blood of healthy volunteers and non-septic burn patients (>30~; total body surface area, >ls~ full thickness), the pmns were incubated with lps (]ng-10p.g/ml) or f'mlp (10 "1 1 to 10"6m) et 37oc for 30 mln, in 1~; human ab serum, the expression of the 132 ]ntegrins was detected using monoclonat antibodies and flow cytometry. lps and f'mlp resulted in a slight increase (3 fold) in the expression of cd11b on pmns from burned patients compared to an 8 and 14 fold increase, respectively, on pmns from healthy individuals. this inability of lps or fmlp to increase cd11 b expression was not due to the amount of lps bound to the two cell populations. because the same defect is seen after either lps or fmlp stimulation, it is speculated that the defect must be in the amount of preformed cd1 ] b or its transport to the plasma membrane. platelet-activating factor (paf) and neutrophils have been implicated in the patbophysiology of ischemia-repeffusion injury, in addition, paf stimulates neutrophi[ (pmn) oxidative metabolism in vitro. the present study examined the potential role of paf in repeffusion injury in an in viva rabbit model. eight anesthetized rabbi~s underwent retroperitoneal exposure of the infrarenal abdominal aorta after percutaneous insertion of a catheter through the jugular vein into the infrahepatic inferior vena cava. doppler flow probes were placed around the abdominal aorta and the right common femoral artery to assess flow through these vessels. an occlusive ligature was placed around the abdominal aorta (superior to the flow probe) at t = 0 and total occlusion of blood flow to the lower extremities was maintained for g0 mins., after which the ligature was released allowing for reperfusion of the ischemic lower limbs. effluent blood from the ischemic hind-limbs was collected through the ivc catheter at the times indicated below and assayed for paf by a direct radioimmunoassay. in addition, neutrophil h202 production was determined by a previously described 2'7'-dichlorofluorescein flowcytametric assay. 185 _+ 53 amean _+ s.e.m, pg/ml blood; brelative fluoresenee (% of baseline); caortic and femoral artery flow (% of baseline); *p < 0.01 vs. baseline; 1"p < 0.05 vs. baseline. a significant elevation of paf was observed in ischemic hind-limb effluent blood at 15 min. after release of the aortic ligature during the repeffusion phase, as compared to baseline levels. in addition, pmn h20 2 production was increased by 2.3-fold above baseline values by 1 hour after ligature release during the reperfusion phase. both of these elevations were transient and returned toward baseline by 2 hours post-isehemia. tatar occlusion of hind-limb flow was achieved as evidenced by the absence of aortic or femorat flow at 90 rain. post-ischemia, however after release 01 the ligature a significant reactive hyperemia was observed by 15 mln. into the rapeffusion phase. histolog[c examination of reper[used gastrocnemius muscle revealed moderate pmn infiltration into the interstitium. in conclusion, these data indicate that paf is released into the circulation during repeffusion, and is likely involved as a mediator in the observed pmn oxidative burst activity, thereby contributing to reperfusien injury. following thermal injury and infection granulocyte function ts abnormal. to elucidate the mechanism by which thermal injury and infection affect the granulocyte's ability to polymerize and depolymedze actin, we serially measured f-actin levels in granulocytes from 14 burned patients (mean age 42,1 +_ 17.7 years, mean burn size 39.8% _+ 22.2%) during the first s weeks post injury. six of the patients had infections during the course of the study, (septicemia, wound invasion and pneumonia). actin levels in granulocytes from eleven healthy volunteers (mean age 34 years) were measured repeatedly and served as controls. lysecl white blood cell preparations were brought to 370c and incubated with n-formyl-met-leu-phe (stim) or with dulbecco's phosphate unbuffered sellne (unstim). the cells were concomitantly stained and fixed with formaldehyde, lysoleclthln and fiuoresceln phafioidin. actin depolymedzation (depol) was measured by incubating stimulated cells at 0°c before the stain-fixative was added. baseline (base) f-actln levels were assessed by adding stsln-fixatlve to icecold unstimulated cells. fluorescence was estimated in a facscan and expressed as ilnesr mean channel fluorescence_+ sem (mcf). figure 1 displays granulecyle fectln levels in infected and uninfected patients as compared to controls. f-actln levels were consistently lower in control cells than in those from burned or burn-infected patients under all measured conditions. granulocytes from infected burned patients demonstrated a significant decrement in their ability to depofymerlze f.actin compared to both uninfected burned patients and controls, while there were no significant differences between infected and 1,~ uninfected patients in the baseline, unstlmuleted and stimulated conditions. those results indicate la5 that grsnulocytas from burned and bum-infected patients contain higher levels of polymerized actln than ~ ,2s control cells. in order to study tumor necrosis factor (tnf) receptor sensitivity in septic critically ill patients we investigated 20 blood samples of such people in reaction of leucocyte migration inhibition. migration of their polymorphonuclear leucocytes (pmns) was studied with stimulation with human recombinant tnf in concentration of 166.7 u/ml (recommended by manufacturer is the range of 10-200 o/ml) and without such. ten healthy blood donors formed control group. the results obtained showed diminished pmn reactivity to tnf in patients (migration inhibition was absent) oscaring with significantly increased migration ability of their pmns (107.2% of that in control group). at the same time normal pmns in control group did show migration changes upon tnf stimulation. considering all the above we come to a conclusion that externally added tnf fails to activate pmns in critically ill patients more than they are by their endogenous tnf. moreover, this tnf no longer serves a positive chemotactic factor for such pmns. these findings may suggest that in critically ill septic patients reactivity of pmns to tnf is deeply altered. tnf receptors of pmns are either exhausted as such by excessive stimulation with endogenous tnf or further transmission of their message is impossible due to "fatigue" of the cell's activation mechanisms. we express our gratitude to reanal factory of laboratory chemicals for generously providing us with a tnf com~rcial sample. ~-sanguis medical, ekaterineburg russia; s-urals med.lnst. activated neutrophils infiltrating the local site of inflammation following trauma release high amounts of destructive lysosomal enzymes into the extracellular space. cytokines were discussed to be involved in regulation of this early process. the task of this investigation was to evaluate the possible regulatory role of interleukin-6 (il-6) and its potential immunosupressive opponent, the transforming growth factor-&, in regulation of neutrophil degranulation. we analysed the concentration of the al-proteinase-inhibitor complex of the lysosomal elastase as marker for the degranulation of neutrophils as well as the levels of il-6 and tgf-61 in the plasma probes of patients undergoing multiple trauma and severe surgeries. the time courses of il-6 and elastase were found to be highly correlated, wheras the concentrations of the cytokine tgf-e~ were found to be not significantly altered in comparison to the control group. this close temporal correlationship was confirmed by investigation of fluids derived from sites of inflammation. interstingly, the inhibitory potential (~zcproteinase inhibitor, antithrombin iii) was dramatically reduced in the early inflammatory phase. to prove this in vivo findings, the effects of il-6 and tgf-i~ 1 on the degranulation of isolated human neutrophils of healthy donors was investigated in vitro. pathological high concentrations of rhll-6 up to 104u/ml (as detected in fluids derived from local inflammatory site) were found to be capable to induce a significant release of lysosomal elastase in a concentration-dependent manner, whereas the degranulation of neutrophils was uneffected by tgf-61. in conclusion, these data suggest a contribution of il-6 in regulation of neutrophil activation at sides of inflammation. the immunosuppressive cytokine tgf-i&~ seems to have no direct regulatory effect beside its described chemotactic function on neutrephils. postirradiation chan~es of adhesive properties arid supercoiled nucleoid dna structure of blood leukocytes were studied in macaca nemestrina andrats. the dynamics of membrane chan~es after nonlethal irradiation of rats demonstrated the temporary increase of the leukocyte adherence at 24 h followed by return of this parameter to normal levels at 48 h. after lethal irradiation of both animal species the increase in adhesive leukooytes fraction was detected as early as at 3 h. this hi~her index persisted until the end of experiments (5 days). the early (3-5h) temporary loosin~ of supercoiled dna structure was demonstrated in the leukocytes of nonlethally irradiated animals. this phenomenon seems to be connected with the lymphocyte fraction chan~es. this process was not dependent on altered adhesive properties of leukocyte membranes. the membrane chan~es of leukocytes preceded decondensation of supercoiled dna after lethal irradiation of animals, in this case loosin~ of supercoiled dna pro-~ressively increased at 24 h and at the later terms of postirradiation period. the systemic inflammatory response syndrome (sirs) involves many inanunological reactions of the host including acfivatinn of inflammatory mediator cascades and depression of cellular reactivity in t-lymphecytes (1). there are reports of nentrophil dysfunction in inflammatory disorders of the skin (2), are there dysfunctions concerning the unspecific host defense in sirs, as well? in this study, we examined the reactivity of neutrophil granolocytes from patients suffering from sirs. twenty-one patients (apache ii-score 21±5) with diagnosis of sirs entered the study. granulocytes were prepared as reported previously (2) . in parallel, granulocytes from 20 healthy individuals were tested. two granulocyte functians were studied in vitro: 1. migration of the ceils in a boyden chamber through a filter matrix following stimulation with 5 different receptor dependent stimuli (c5a, intefleukin-8, platelet-activating-factor, leukotrien b4, fmlp). 2. release of 13glucuronidase following stimulation with the aforementioned activators. the results demonstrate, that the release of 13-glucuronidase in patients suffering from sirs was comparable to the enzyme release of granulocytes prepared from healthy individuals. each stimulant induced release of p-glucuronidase in a characteristic dose dependent fashion. all granulocyte preparations from the healthy donors showed a positive chemotaxis response in the migration-assay. in contrast, only ten out of twenty-one patients had granulocytes migrating after stimulation. the two groups of patients displaying reactive or non-reactive granulocytes differed clinically: the nonreactive group consisted of patients with multiple organ failure (10/11) and nonsurvivors (8/11), whereas 8/10 patients in the reactive group survived. thus, the in vitro chemotaxis of granulocytes is impaired in a subgroup of patients with sirs. this defect of the non-specific host defense may contribute to poor prognosis and outcome of these patients. dermatol. 85: 194-198, 1985 klinik ffir an~isthesiologie und operative intensivmedizin der cau kiel, schwanenweg 21, 24105 kiel, germany. objectives of the study: major emphasis has been given to the analysis of interactions of antibiotics with microorganisms. effects of antibiotics on cells of primary host defense mechanisms, such as the neutrophils, are less well known. therefore, attention has been focused on clindamycin, a member of the lincoseamide family. materials and methods: the effect of clindamycin (i -i00 ~g/ml) on 8 granulocyte functions (healthy volunteers) such as random migration, chemotaxis (agarose method), ingestion (radiometric assay), superoxide (cytochrom c reduction) and hydrogen peroxide production (phenol red oxidation), lucigenin-and luminol-amplified chemiluminescence (luminometry) and degranulation (turbidometry with micrococcus lysodeicticus) were investigated in vitro. results: motility and degranulation were inhibited, ingestion of saccharomyces cerevisiae, zymosan-induced lucigenin-and luminol-amplified chemiluminescence, superoxide and hydrogen peroxide production were stimulated in a dose dependent fashion. conclusion: clindamycin has granulocyte function modulating properties. recognition of immunomodulating effects of antibiotics may have therapeutic significance, especially in patients with long-term antibiotic therapy or immune deficiencies. the intense muscle activity (ea) of rats resulted in increase of neutrophil influx in muscles during the recovery. we investigated neutrophil proteinases involvement in neutral proteinases balance of skeletal muscles by na. the rats were submited to swim with the load (8% of body mass) till exhaustion. immediately after na the neutrophil antiserum was injected i.p. to rats of experimental group. saline was injected to control animals° injections were repeated in 12 h of the recovery and cytosol proteolytic activity (ph 7.4; fitc-casein) was determined. isolated soleus muscles were incubated also in vitro and proteolytic activity of incubation media was measured. it was found that there was 2-3 -fold proteinases activity increase in cytosols of all investigated muscles (soleus, white and red portions of quadriceps) of control animals by 6 h of the recovery (the comparison was done with the sedentary rats). in 12 h cytosol proteolytic activity decreased and then increased again by 24 h of the fast. antiserum injections resulted in relible decrease of the proteolytic activities at every investigated time. when incubating m. soleus in vitro the activities of proteinases in incubation media turned out reliably less if soleus muscles were isolated from the animals to which antiserum was injected. the conclusion is that neutrophil proteinases can be involved in the balance of rat skeletal muscle neutral proteinases after ~a. a lot and new clinical problems complicating the outcome of polytrauma, burn and septic patients in surgical intensive care units, have arisen as the care improvement prolonged the patient's survival: a progressive degradation of organ and system functions often develops, usually making its first clinical appearance by ards, followed by the other organ failure (mof) and sepsis symptoms. the clinical picture is polymorphic, the end result of a complex systemic pathophysiological reaction trigg~ed off by trauma consequences (tissues disruption, hypo~xygenatiun and necrosis). nowadays there is not a preventi~ or specific therapy to lower the mortality rate (70-80%) and-'mdy-a~ early, aggressive surgical approach .-evacuating haematomas, stopping bleeding, toileting all septic, necrotic foci and restoring anatomic continuity-, seems to be of some help this complex clinical entity has not an univocal denomination yet. the proper labelling of an illness should come from the full understanding of its pathopysiology and suggest the proper treatment choice. clinical and experimental studies demonstrated that pathophysiologic mechanisms involved in the past-traumatic illness, share the same anatomo-pathological elemem: the interstitial edema, due to a generalised endothelial micro circulatory injury. this alteration, as constantly seen in polytrauma patients, develops in a few hours after trauma as a consequence of the deregulation of the homoeostatic and immune mechanisms. in fact the overproduced oxygen free radicals and r~ombinam cytokines (il1,tnf), together with the complement degradation fragments, the proteolytic enzymes and many other mediators are all strongly h~l ~ ,_he e,,j,yheha! ceils. our~osect, atim~,-bnsed on examination of autopsical specimens from 85 polytraanm patients, showed that such endothelial damage, supporting the interstitial edema, is widely and simultaneensly distributed, ensues shortly arer trauma and shows its effects in different organs at different times, only because each apparatus has different fimctienal reserves: the lung is the first organ to fail just because its ah, celocapillary membrane is one of the most delicate bodily structure, and its function is irroplace~le. we think it will be of a great help, in planning a preventive therapy, to chose a denomination focusing the physician's attention on the earl)" generalized endothelial injury and its effects, as in trauma patients it is present -even if latenflysince the first few hours. we would like to see the generalised endothelial microcircolatory injury properly highlighted when considering the best definition and the optimal nomenclature for the post-traumatic s3mdrome. the presence of interleukin (il)-8 in bronchoalveolar lavage fluid of critically ill patients correlates clinically with the development of the adult respiratory distress syndrome lards), and inhibition of il-8 in animal models can attenuate lung injury. collectively, evidence to date suggests that il-8 attracts and activates neutrophiis (pmn), which are then responsible for the capillary leak of ards. however, an alternative explanation is that il-8 is directly toxic to the endothelial cell (ec). in this study, we have hypothesized that il-8 can disrupt endothelial integrity independent of pmn. meth ods: human umbilical vein (huv) ec monolayers were cultured to confluency on collagen-coated micropore filters. to assess ec integrity, 1251.albumi n leak was quantitated by measuring the 1251 counts which crossed the monolayer, using a gamma counter. il-8 (lpg/ml) was incubated in the culture medium with 1251.albumi n for 21 hrs. the il-8 dose was not cytotoxic. to determine the involvement of protein synthesis in this process, selected monolayers were pretreated with cycloheximide (ch) prior to 11.-8 addition. statistical analysis was performed using anovmfisher plsd. we have previously shown that platelet activating factor (paf) enhances cdt8 expression and primes pmn's for subsequent 02generation. both are important steps in pmn mediated injury and are assumed to occur in concert. following major trauma non-specific pmn inflammation is activated, however, unbridled systemic pmn activity needs to be minimized. since circulating catecholamines are high early post-injury, we hypothesised that they downregu/ate cd 18 expression and pmn priming via the [3 adrenergic signal transduction pathway. methods: normal human pmns were primed with paf (10ng/ml for 5 min) or pre-treated with 10-5m of isoproterenol (i) or forskoklin (f) for 5 rain and then primed with paf. cd18 expression was measured by flow cytometry (fig.l) and 02-generation in response to 10-rm fmlp was determined as sod inhibitable reduction of cytochrome c ( fig.2 holler** and georg w. bornkamm* lymphocyte-endothelial interactions are crucial for various immune responses, including cytokine driven inflammatory processes. protein kinase c (pkc)-inhibitors on the other hand are discussed as potential cytokine antagonists. in the present study we investigated the influence of the pkc-inhibitor gf109203x on cytokine-and endotoxin induced expression of intercellular adhesion molecule 1 (icam-1) and on adhesion of lymphocytes to cytokine activated endothelial cells. we found that tumor necrosis factor alpha (tnfo0-and lipopolysaccharide (lps)-induced icam-1 expression on human endothelioma celts (eahy926) were unaffected by the pkc-inhibitor and thus appeared to be independent of pkc activation. in contrast, gf109203x significantly reduced icam-1 expression induced by interferon-y (ifn-?) and interleukin-1 (il-1). the functional relevance of these findings was evaluated in an adhesion assay using human umbilical vene endothelial cells (huvec) and peripheral blood mononuclear cells (pbmc). in fact, the ifn-? and il-1 induced adhesion of pbmc to cytokine treated huvec could be downregulated by the pkc-inhibitor, whereas tnfc~-and lps-mediated adhesion was not influenced. additionally, the il-1 driven icam-1 expression on eahy926 cells as well as the il-1 induced adhesion of pbmc to huvec was found to be tnf-dependent, since both effects could be inhibited by an anti-tnf monoclonal antibody (195f) . these in vitro data further support the idea of examining pkc-inhibitors, such as gf109203x, for their biological relevance in cytokine related dysregulations. seiffge, d., bissinger, t., laux, v., during inflammation there are some key processes, which occur in the microcirculation: the release of mediators from various cell types, the migration of inflammatory cells towards a chemotactic stimulus in the tissue, the expression of adhesion molecules on different cells, and the extravasation of plasma proteins. the aim of the present study was to elucidate the mediator induced interaction of leukocyte adhesion and plasma leakage in postcapillary venules. using an analogous video-image analysing system we have studied the effect of different mediators on leukocyte adhesion and macromolecular permeability in the mesentery of the rat. the increase in permeability was measured as changes in optical density. we found that topical administration of leneotriene b 4 (ltb4, 5x10 "6 tool/l) or intravenous injection of interleuldn-2 (il-2, 5-106 iu/kg b.w.) and lipopolysaccharide (lps, 15 mg/kg b.w.) resulted in a significant extravasation of fitc-labelled rat serum albumin (fitc-rsa) in venules but not in arterioles. we could correlate the changes in vascular permeability with a locally increased number of rolling and sticking leukocytes in venules. both effects were dose dependently inhibited by different drugs. pentoxlfylline inhibits lps-indueed fitc-rsa extravasation and leukocyte adhesion at a dose of 3 mg/kg b.w., superoxid-dismutase (sod, 10.000 iu/kg b.w.) was able to decrease the ltb 4 effect, and the immuumodulating drug leflunomide (hwa 486) exerted inhibitory effects on il-2-induced permeability at a dose of 3 mg/kg b.w.i.v. the obtained results demonstrate that lps, ltb 4 or il-2 induced extravasation of fitc-rsa is mediated by activated leukocytes and can be deminished following administration of different drugs. platelet-endothelial cell adhesion molecule-i (pecam-i), a member of the immunoglobulin superfamily, is constitutively expressed at high levels on the endothelial cell surface. in vitro data have suggested that pecam-i functions as a vascular adhesion molecule, specifically in neutrophil transmigration across the endothelium. this current work is the first demonstrating the in vivo role of pecam-1 in neutrophil migration. blocking antibodies to human pecam-1, in which the antibodies are crossreactive with rat pecam-1, were able to block the movement of neutrophils into the rat lungs after igg immune complex deposition. furthermore, when human foreskin was transplanted into mice with severe combined immunodeficiency and the site injected with tnf-alpha, anti-pecam-i blocked neutrophil emigration into the dermal interstitium. it has already been established that neutrophil recruitment is dependent upon selectin mediated rolling, followed by firm adherence that is icam-1/ integrin mediated. these data suggest, for the first time, that a third endothelial adhesion molecule (pecam-i) is involved in the coordinated recruitment of neutrophils in vivo. to test whether trauma causes generalized activation or priming of pmns, cdi8 adherence receptors were measured with iinmunomonitoring in whole blood after lps stimulation ex vivo. anesthetized (fentanyl) mongrel pigs (15-25 kg) were subjected to 40% arterial hemorrhage + soft tissue injury and after liar, resuscitated with all the shed blood + supplemental fluid. blood was collected at 24hr intervals from unanesthetized animals with indwelling catheters, pmns were counted, and lps was added (0,1,5,10 i.tg/ml) ex vivo. after 3hr incubation at 22-24°c, %cd18 (+) pmns were determined with fitc-ib4 and flow cytometry from mean channel fluorescence histograms. 49±8 # p<0.05 vs baseline * p<0.05 vs sham $p<0.05 vs no anesthesia these observations provide direct evidence for time-dependent changes in pmn priming following major injury because cd18 expression was depressed for at ]east 24hr after trauma relative to sham but by 72hr, was enhanced, relative to sham, and because fentanyl anesthesia at 72hr had a greater effect on cd18 expression in trauma vs sham. neutrophil (pmn) adhesion to vascular endothelial cells (•c) is a key element in the inflammatory response and tissue injury. inflammatory mediators such as lps (exogenous) and tnf (endogenous) can promote pmn-ec interaction which is believed to be responsible for capillary leakage and subsequent organ injury. however, the mechanism of this injury remains unclear.we hypothesised that the mechanism of tissue injury is due to ec necrosis with release of toxic products and that activated pmn are responsible. human pmn were obtained from healthy donors, separated by density gradient, and activated with lps (50ng/ml), tnf(10ng/ml), and lps/tnf(25ng/10ng/ml). cultures of the human ec tine(ecv-304) were used as surrogates of the microvasculature, were exposed to either lps, tnf, lps/tnf and pmn activated with lps, tnf, lps/tnf and incubated for 6, 12, 18, and 24hrs. ec necrosis was assessed by a 51cr release cytotoxicity assay. pmn activation was assessed by cd1 lb receptor expression and respiratory burst activity 6hr 0_+0 1.7-+ 1 0-+0 3.2_+ 1 0_+0 4.4_+ 1 0_+0 2.3_+0.9 12hr 0+0 3.24_3 0_+0 9.3_+3 0_+0 11_+3" 0+_0 12+--2.1" lghr 04-0 0.9_+2 0+_0 284-12" o:fo 25,12" 0~0 30+-9.5* 24hr 0_+0 4.14-4 0-+0 33+_3* 0_+0 30_+9* 0_+0 32_+10" data = ec % necrosis mean_+sd stats: student's t-test with significance (*) set at p<0.05 vs control. ( our previous studies have indicated that despite the increased cardiac output and maintenance of tissue perfusion, hepatoceliular dysfunction occurs during early sepsis. nonetheless, it remains unknown whether vascular endothelial cell function (i.e., the release of endothelium-derived relaxing factor/nitric oxide) is depressed under such conditions and, if so, whether endothelial cell dysfunction also occurs at the microcirculatory level. to determine this, rats were subjected to sepsis by cecal ligation and puncture (clp), following which these and corresponding shams received 3 ml/100g bw normal saline. at 5 hr after clp (hyperdynamic sepsis) or sham operation, the thoracic aorta was isolated, cut into rings, and placed in organ chambers. norepinephrine (ne, 2xi0 -7 m) was used to achieve near-maximal contraction. responses for an endothelium-dependeut vasodilator, acetylcholine (ach, via nitric oxide), were determined. in additional studies, the small gut was isolated at 5 hr post-clp. after pre-contraction of blood vessels in the isolated gut with 5xl04 m ne, vascular responses to ach (5x10 6 m) and an endotheliumindependent vasodiiator, nitroglycerine (ntg, 5xl0 -7 m), were determined. total vascular resistance (tvr, mmhg/mi/min/100g) was then calculated as pressure/ perfusinn rate. ach-induced relaxation (%, n=6/group) in the aortic rings were: ach lxl0 i~s, st-in ~ ~ significantly at 5 hr post-clp (i.e., increased *p(o 05 vs. sham; n-4 per group. tvr) in the absence of any changes in ntginduced relaxation (fig. a) . thus, the vascular endothelial cell dysfunction observed in the aorta in early sepsis also occurs at the microcirculatory level. introduction: the cytokine-mediated adherence of leulcooytes to vascular endothelium is considered as an early step in the cascade of pathologic reactions culminating in the "systemic inflammatory response syndrome" (sirs); the purpose of this study was to evaluate the influence of interleakin-1 on leukooyteendothelial cell-interactions and microoirculation in the liver after hemorrhagic shock by means of intravital microscopy. methods: in anesthetized female sprdrats co.w. 200-230g) shook was induced by fractionated withdrawl of arterial blood within 5 rain and maintained for 1 h (map at 40 mm hg, cardiac output 50 % of baseline). rats were adequately resuscitated with 60% of shed blood and twice the volume in ringer's solution additionally. following 5 h of reperfusinn (map >100 mm hg, co >120% of baseline) the microcirculation in liver lobules was examined by intravital fluorescence microscopy after labelling of leukocytes. continuous administration of il-lra (synergen, boulder, colorado, 5mg/kg/h) was started at different time points in a randomized and blinded manner. the animals in group p (n=6) received the il-lra as pretreatment beginning 5 min prior to shock induction. in the group t (n=6) the application of il-lm started at the beginning of the reperfusion period with a bolus injection of 5 mg/kg and was followed by continuons administration of 5 mg/kg/h. the control group c (n=4) received equal volumes in nac10,9%, the sham-operated group s (n=4) was not exposed to shock. results: macrohemodynamics were comparable in all shook groups. the increased percentage of permanendy adherent leukocytes after hemorrhagic shook (s: 9,1% +1,1%; c: 42,1% _+5,4%) was significantly reduced by pretreatment or treatment with il-lra (p: 16,9% -+1,9%; p<0.001, t: 28,8% -+3,6%, p<0.01, anova). temporary adhesion of leukocytes was unaffected by application of il-lra. liver microcirculation measured by volumetric blood flow in liver sinusoids and sinusoidal diameters was impaired after hemorrhagic shock in all groups and was not affected (c: 31993iam3/s +40971um3/s, p: 32768llm3/s +4445}am3/s, t: 326211ams/s -+2998 lam3/s, s: 390201am3/s -+39041am3/s). di.seu~sinn: the results demonstrate that permanent adherence of leukocytes to endothelium is in part regulated by il-1. pathological adherence could be reduced by application of illra, even given at die time of resuscitation. the effect of ll-lm on permanent adhesion is a specific event and might be caused by reduced expression of specific receptors on sinusoidal endothelial cens and leukocytes. objectives of the study. the adhesion of activated neutrophils (pmn) to endothelial ceils (ec) and the concomitant production of reactive oxygen metabolites (rom) initiates organ damage after trauma, sepsis, shock and organ reperfusion. aien of this study was to investigate the effect on adhesion and rom production of the highly water-soluble, membrane-permeable and physiological ascorbic acid (asc). materials and methods. adhesion of pmn to nylon fiber (cell count) and simultaneous rom production (chemiluminescence-cl-response) were measured up to 6 retool/1 asc as well as adhesion, rom production and ec damage (lllln-release from labeled ec) of endotoxin-activated pmn to cultered ec moanlayers. in an in vivo animal model (sheep with lung lymph fistulas) the effect of asc (1 g/kg bw bolus, followed by 0.2 g/ kg-h infusion) on the endotoxin-induced (0.5 ixg/kg bw) neutropenia (cell count), lung capillary permeability damage (lung lymph protein clearance) and rom production of neutrophils (zymosan-induced cl response) was measured. results. asc scavenged rom dose-dependently during adhesion of pmn to nylon fiber (p<0.0005 at 6 mmol/l asc), adhesion itself was unchanged. during the activated pmn/ec interaction asc scavenged rom (p<0.025 at 6 mmol/l asc) and reduced the adhesion dose-dependently (p<0.0005 at 6 mmol/l asc); ec damage was also reduced (p<0.0005 at 6 retool/1 asc). in the in rive model asc increased the endotoxin-induced blood pmn decrease (p<0.05), decreased the protein clearance (p<0.05) as well as the zymosan-induced rom production (p<0.03), indicating the asc-mediated reduction of adhesion, rom production and lung tissue damage processes. conclusions. by in vitro and in rive experiments ascorbic acid reduced the adhesion-and rom production-initiated tissue damage. therefore, i.v. administration of ascorbic acid is recommended for oxidative stress-associated states after trauma, sepsis, shock and organ reperfusion. for neut rophi l-accumulat ion and activation. we investigated the influence of or to the activation and the expression of lecam-i and cdiib,cdi8 on neutrophils and lymphocytes. methods: from blood samples (n=5) all white blood cells (wbc) and neutrophils (nc) were isolated and cultured. or were produced via the xanthine oxidase/hypoxanthine system. after 0,5, 15,30,60 and 120 minutes a giemsa-staining to determine the granulation of neutrophils (n: normal, r : reduced ) and a facs-analysis with monoclonal antibodies detecting cdiib,cdi8 and lecam-i was performed. results: under the influence of or a degranulation of neutrophils starting at 15min was observed in wbc-cultures (n/r: 0min 91/9, 15min 76/24, 30min 68/32, 60min 52/48, 120min 49/51). these data were confirmed in the dot-plots of facs-analysis. only in wbc-cultures or induced a significant increase of lecam-i expression on neutrophils up to 30min followed by a decrease to normal values at 60min. lecam-i on lymphocytes disappeared totally during the observed period. cdllb,cdl8-expression was not altered. conclusion:increased lecam-i expression on neutrophils due to or could enhance the 'rolling' of neutrophils along the endothelium which is a prerequisite for neutrophil sticking and migration. further or are able to activate neutrophils without endothelium. these changes seem to be mediated by other wbc. introduction. multiple organ failure (mof) has been hypothesized to be the result of an excessive uncontrolled autedestructive inflammatory response. since the complement system is an important mediator and initiator of the inflammatory response, interruption of this cascade could theoretically lead to an attenuation of mof. in order to test this hypothesis we evaluated the response of c5-delicient mice in a model of zymesan indt~ed mof. materials and methods. 25 c5-deficient b2d10/oid and 25 c5-sufficient b2d10/new mice were used in this study. on day 0 all mice received an intraperitoneal injection with zymosan suspended in paraffin in a dose of 1 mg/g body weight. between day 0 and 12, biological parameters (temperature, body weight and clinical condition) were measured daily and mortality was monitored. clinical condition was assessed by blindly grading the degree of lethargy, conjunctivitis, diarrhea, and ruffled fur of each mouse on a two point scale (maximum score=4). on day 12 all surviving mice were sacrificed and relative organ weights of lungs, liver, spleen and kidneys (relative organ weight= (organ weight/body weight)x100) wore calculated. earlier experiments with our model have shown a good correlation between histological organ damage and relative organ weights. statistical analysis of biological parameter was performed using the koziol curve analysis. analysis was divided in an acute phase (day 0-4) and a late phase (day 8-12). relative organ weights were analyzed using wilcoxon's test and mortality rate using fischor's exact test. results. all zymosan injected mice showed a typical triphesic illness. deterioration of the clinical condition as indicated by the symptom score and the decrease in temperature and body weight in the acute phase were all significantly lass severe in c5 deficient mice (all p<0.005). in the late phase no differences could be noticed in the courses of biological parameters. overall mortality was 2/25 (8%) in c5 deficient mice and 8/25 (32%) jn c5 sufficient mice (p=0.049), a difference mainly due to a difference in the acute phase. organ damage assessed as the relative organ weights did not show any statistical differences for any organ between both strains. conclusion. complement factor c5 appears to play an important role in the acute hyperdynamic septic response in this model but deficiency of c5 could not prevent organ damage in the late mof phase. this suggests that other factors could be more important in the development of the inflammatory response leading to mof. proinflammatory cytokines are thought to play a critical role in the pathophysiology of multiple organ failure (mof). in mice, zymosan-lnduced generalized inflammation (ztgi) leads to mof. therefore we performed a sequential study into plasma levels of, and macrophage production capacity for, four cytokines during the development of mof in the zigi model. male young-adult c57bl/6 mice received zymosan (1 mg/g body weight) intraperitoneally. groups of 14 animals were killed after 3, 6, 8, 18 and 24h and subsequently at each day until day 12. plasma was collected and peritoneal macrophages were isolated and cultured overnight with or without lipopolysaccharide (lps). interleukin 1-ct, and -6 (il-lc~,~,), and tumour necrosis factor-o~ (tnf-c 0 were measured in plasma and culture fluid by means of a ria (detection limit 0.1 ng/ml). interleukin-6 (il~) levels were assayed using the b9 hybddoma cell proliferation assay. zymosan induces a three-phase disease in mice. after an acute phase the animals recover. around day 7, they start to develop clinical signs which resemble mof. plasma tnf-~ peaked within 3h after zymosan injection and disappeared within 8h. from day 8 onwards, tnf levels started to rise again. plasma il-6 behaved almost similarly in the acute phase, but in the mof phase plasma il-6 remained low. no circulating il-16 could be detected at any time point. macrophage lps-stimulated production of il-lcq il-11~ and tnf--c~ was suppressed immediately after zymosan injection. production of il-16 and tnf-~ was normalized within 6h, while production of il-lc~ remained lower than that in macrophages from untreated control mice. only at day 6 did production of il-i~ reach control values. il-16 production was higher than control values from day 3 onwards. il6 production was similar to that of ili-il the production of tnf-ct was strongly elevated between days 1 and 6 and again during days 10 to 12. the development of mof-like symptoms during zlgi in mice is accompanied by increased plasma levels of tnf-ct without enhanced il-16 or il-6. also, the ability of macrophages to produce excessive amounts of il-16 and tnf--~, as well as the suppressed capacity to produce il-lcq could be important mechanisms in the pathophysiology of mof. when conjugated to an asialoglycoprotein, dna and oligonucleotides are specifically taken up by the hepatocytes via the asialoglyccprotein receptor which is unique to the liver. human asialoglycoprotein (~1-acid, asgp) was derivatized with low molecular weight poly(l)lysine(pll) and complexed with 5 antisense dna's (as) complementary to the 5' region of the il-6 gpl30 receptor. the antisense were 5'-agtttagggatgagg-3' (asl), 5'-atcttcatcttctgaat-3' (as2), 5'-aagtgaatgattaaaacact-3' (as3), 5'-aaacctttataggcg-3' (as4), and 5'-cgttctacaactgcaacgt-3' (as5). using hepg2, the biological effects of these antisense complexes on the high affinity il-6 receptor were evaluated by scatchard analysis, cellular proliferation, and acute phase protein expression by radioimmunoprecipitation and two dimensional gel electrophoresis. scatchard analysis demonstrated that high affinity receptor expression was inhibited by incubation of cells with asgp-pll-asi for 24 h. underivatized asl was less effective and the complex, asgp-pll-as2, had minimal effects on high affinity binding. when the cells were treated with the conjugates and stimulated with il-6 (i00 units) asgp-pll-asi alone showed a dose dependent (0.i-0.4 ~m) inhibition of 3ss fibrinogen synthesis. two dimensional gel electrophoresis showed that expression of other acute phase proteins was also blocked. these results indicate that the targeted delivery of antisense molecules via conjugates recognized by the asialoglycoprotein receptor can block the cytokine stimulated acute phase protein response in hepatocytes, this approach may be relevant to the therapeutic management of patients with severe injury and sepsis. it has been established that immune cells are able to express neuropeptide genes and to release products that were considered to be of neuroendocrine origin. we have shown that proenkephalin (penk), a neuropeptide encoding gene, is expressed in lymphoid cells in culture. to study the physiological significance of these observations we have used the model of experimental endotoxemia. in this model, a disease state is induced by bacterial lipopolysaccharide (lps), that activates the immune system, the adrenocortical axis and the nervous system. we found that the expression of penkmrna is markedly enhanced in vivo immediately after lps injection both in the adrenal glands and in the lymph nodes. in situ hybridization analysis combined with immunohisto-chemistry indicated that the induced penk expression is confined to macrephages within the lymph nodes and chromaffin cells in the adrenal medulla. furthermore, this expression in lymph nodes is modulated by ligands of the adrenergic system. our results strongly support the notion that immune derived opioids participate in the bidirectional communication between the nervous and immune systems. of neurology hadassah university hospital, jerusalem 91120, israel. objectives of the study: multiple-organ-failure is recognized as the most severe, and often lethal, complication after multiple trauma. however there is no adeqate animal model available. our goal was to develop an animal model, in which reproducable irreversible failure of parenchymal organs is achieved in the late phase after insults in the early phase (trauma). materials and methods: l0 female merino-sheep were included (mean weight: 30kg). day 0: hemorrhagic shock (mean arterial pressure (map) 50mmhg for 2 hrs.), closed femoral nailing (ao-technique), day 1-5: bolusinjection of endotoxin (et) (0,75~tg/kgbw) und zymosan-activated plasma (zap) (20ml) every 12hrs., day 6-10: observation. bronchoalveolar lavage (bal): day 1, 6, 10. the course of representative parameters of organ function was documented: cocardiac output (i/min), svr -systemic vascular resistence (dyn ~ s cm-5), pap -putm.art.pressure (mmhg), pap2 -arterial oxygen pressure (mmhg), bill -bilirubin (;xmov1), crci -creatinin clearence (ml/min) statistics: data as means+sem, *significant from baseline (wileoxon test; p<0005) results: baseline day 4 day 6 day 8 day 10 heart: co 4,96_+0,32 5,33_+0,36 5,4_+0,2 7,30_+0,77* 7,98_+0,66* svr1520_+134 1395+141 1403_+89 1119+_122" 1089+-91" lung: pap 17,0_-20,7 19,6_+1,6" 22,0+-1,2"25,8+2,0" 28,8+-1,7' pap2 103,1+1,5101,5+-5,7 97,3_+4,7 91,9+-5,6 89,8+_3,9* liver: bill 2, 94_+0, 344, 82_+1, 34' 5, 13_+0, 68'6, 13_+0, 7" 7, 19_+0, 91" kidney:crcl 86, 5+_30, 9 109, [8] [9] [10] [11] [12] [13] [14] [15] [16] [17] [18] [19] 4 74, 7_+10, 868, 8+14, 1 53, 1+17, 6 histologic specimens showed all signs of fulminant mof. combination of hemorrhagic shock, femoral nailing, et und zap (insults in the early phase) lead to an irreversible organ failure in the late phase. prostaglandin e (pge) levels are elevated by trauma, shock or sepsis and can profoundly affect the immune response. pge2 is produced by many cell types including fibroblasts, macrophages, monoeytes, follicular dendritic cells, and epithelial cells and is induced by il-i, bacterial lps, components of the complement cascade, tnf, il-6 and crosslinking of surface fc receptors for igg, iga and ige. our research has shown that pge inhibits b cell activation (specifically enlargement, class ii ~c and fc~ rii expression), proliferation, igm and igg3 responses, t cell proliferation, and il-2 synthesis in the mouse model. in contrast, pge greatly promotes class switching to ige,the isotype responsible for type i allergic hypersensitivity. thus, our model mirrors th~ general immunosuppression and elevated ige titers of the trauma or sepsis patient. pge increases the number of cells secreting ige and iggl, acts on surface igm positive b cells, synergizes with il-4 and lp$ to induce preswitch germline transcripts, and induces more rapid expression of mature vdj~ mp~a than in eontro~ pge intracellular signalling occurs through cyclic adenosine monophosphate (camp) levels and can be mimicked by camp-inducing agents and blocl~ed by an inhibitor of campdependent protein kinase a. pge action requires de novo protein synthesis and candidate pge-inducible regulatory proteins have been identified by 2d gel eleetrophoresis. thus, pge inhibits a number of immune mechanisms while promoting ige production. a deeper understanding of pge immune regulation may lead to more effective treatment of immune perturbations as sequelae of trauma, shock or sepsis. during infrarenai aortic surgery mesetueric traction (re.t.) results in prostacyclin (pgi:) release and consecutively in hemodynamic disturbances (decreased systemic vascular resisteace, mean arterial pressure; increased cardiac output, heartrate). these symptomes are bypassed by cyclooxygenase inhibition. hemodynamic symptoms vanish after 30-45 rain even without cyclooxygenase inhibition although pgi2 levels remain elevated. to study the endocrine vasopressor system in a prospective double blinded protocol, we investigated 26 patients undergoing major abdominal surgery as compared to 26 ibuprofen (400 rag, i.v.) pretreated (ibu) patients. the surgeon applied m.t. in a uniform fashion. we chose a general anesthesia combined with a supplemental thoracic epidural anesthesia. at the points in time 5, 15, 30, 45, 90, 180, 360 , 1440 rain after and before (to) mesentzrie traction we determined the plasma concentrations (pc) of 6-keto-pgf~o~pr~, epinephrine, norepinephrine, dopamine, renin, aldosterone, adh and cortisol. pc of 6-k-pgf~,tp~, peaked 5 minutes after m.t. (2274_+284, ibu: 102_+12, to: 60+i ng/l) and declined monotonously over 6 h (203 +_42, ibu: 84_+ 12 ng/1). catecholamine pc "s did not exceed the reference range during the observation period. reninpc peaked after 45 rain (66_+23, ibu: 18+3, to: 16-+2/~u/ml); aldosteronc also presented a maximum after 45 rain (110 + 12, ibu: 66-+ 15, to: 56 +-7 pg/ml), whereas cortisol demonstrated irrespectively of circadian rhythms a maximum 6 h after m.t. (26+_1, ibu: 24-+3, to: 10+_1 ~g/1). adh pc peaked 5 min after m.t. (71+7, ibu: 20-+6, to: 5+_1 pg/ml) and showed analogously to 6-k-pgft~j~ pc a monotone decline over the observation period. our data demonstrate a counteractive reaction to pgiz mediated vasodilation via adh secretion. the second regulative is the renin-angiotensin-aldosterone system (raas), which is activated 45 min after m.t., the aldosterone pc does not paratlel the cortisol pc, which peaked post operafionem in both groups, probably due to the end of anaesthesia. a regulative release of catecholamines could not be documented. the activation of adh and raas after mt is not a hormonal response primaryly related to surgical trauma and/or stress but a counterregulation to systemic vasoditafion induced by prostacyclin. although adh and raas support systemic circulation, angiotensin and vasopressin may compromise local organ blood flow (e.g. splancimic vascular bed). insfitut f. klin. chemic, anaesthesiologie ~, chirurgie l*, univ. ulm, 89070 elm, expression of c-fos protein in rat brain following occlusion of superior mesenterie artery. takanobu there is general agreement that neurologic abnormalities are seen in sepsis. the aim of this study is to examine what effect does the brain receive in case of sma occlusion by immunohistochemistry using antibody to c-fos, an immediate early gene, which is recently recognized as a genetic marker of activated neurons. moreover, we investigated the correlation between c-fos induction in the brain and plasma endotoxiu level. 20 rats of them received sma clipping and others wee used as control. control and treated rats at 2, 4, 6, 8 hours were perfused and fixed. the brain were sectioned at 20 pm and stained by abc method using c-fos antibody. plasma endotoxin level of 5 rats were measured at 0, 2, 4, 6, 8 hours after the treatment by chromogenic limulus method. immunohistochemical study showed scarcely no immunoreactivity in control rat brain. in treated rat brain, the significant expression of c-los was detected in specific nuclei including the habenula, some hypothalamie nuclei, amygdala, locus ceruleus and nucleus tractus solitarii. such immunoreactivities were increased in time curse, which well corresponded plasma endotoxin levels. the mean plasma endotoxin level of 0, 2,4, 6, 8 hours after the treatment were 6.48 ± 5.57, 15.0 _-4-4.37, 10.0 _+ 4.18, 14.4 ± 3.52 and 58.4 ± 28 .6 pg/ml, respectively. the results indicate that limbic and hypothalamic-brainstem systems are involved in sma occlusion, and suggest that such neuronal actival.jon may precede the elevation of plasma endotoxin icy.el. systemic vascular resistance and increased cardiac output accompanied presumingly by a increased pulmonary shunt (qs/qt). this response is induced by prostacyclin (pgi2). we examined oxygen transport after traction on the mesentery root and the transpulmonary prostacyclin levels in a prospective placebo controlled study with intravenous ibuprofen. methods: with approval of the human [nvestigadon review board we studied 52 patients in a prospective, randomized double-blinded protocol who were scheduled for major abdominal surgery. ibuprofen (400 mg i.v.) or a placebo equivalent was administered 15 minutes before skin incision. pulmonary artery thermodilution and radial artery catheters were placed after induction of anesthesia. mt was applied in a uniform fashion. baseline values preceded the incision of the peritoneum (to). fulther assessments followed 5, 15, 30, 45, . tile plasma concentrations (pc) of 6-keto-pgft, (stable metabolite of pgi2) were determined in arterial and mixed venous blood by radioimmunoassay. at all points in time we measured arterial and mixed venous blood gases. qs/qt was calculated by standard formula. data are given as median (p < 0.05 placebo vs. [ibuprofen] [117] mmhg (*p< 0.0i). these changes were accompanied by a marked increase of 6-keto-pgf~ pc up to 180 rain after mt in arterial and mixed venous blood of untreated patients with a peak of 2450*[60] ng/l tl (*p< 0.00ol). there was no difference between arterial and mixed venous pc. ibuprofen pretreated patients (n=zr) demonstrated stabile qs/qt and pao2 while 6-keto-pgf~ pc remained within the normal range. discussion: our data clearly indicate that mesenteric traction response includes a critical rise in qs/qt followed by significant decrease of paov stable oxygen transport determinants following cyclooxygenase inhibition signify an action mediated by prostacyclin. an indicative transpulmonary gradient for 6-keto-pgft~ was not detectable. a splanchnic vascular source for pgi2 release seems to be likely, but could not be proved by our current data. department of anesthesiology, cliu. chemistry * and surgery*; university clinics uim, prittwitzstral]e 43, 89075 ulm, germany it is unclear whether injuries like bums, in general, directly result in alterations of cell-mediated immunity that, in turn, promote endotoxic and bacterial translocation or, alternatively, whether these conditions allow increased bacterial invasion that, in turn, inhibits cmi. aim: to determine whether infectious challenge, as clp alone or combined with ti causes further immune abnormalities in the days following clp. study plan: on day 1, two groups of n=36 8 week old aj mice were subjected to either a 20% scold burn (ti), or were untreated (c) n=18. on day 10, 18 mice (ti+clp) and 24 mice (clp) were subjected to clp. the two other groups (ti and c) were untreated. at days 11, 12 and 13 after thermal injury splenocytes (sp) were harvested and cultured with cona for an assay of il-2 and adherent splenocytes (as) were cultured with lps for il-1, tnf, il-6 and pge2. results: either ti + clp or clp alone result in significantly decreased secretion of all cytokines tested. in the ti group almost every cytokine production determined was elevated in comparison to ti + clp and prosmcyclin (pgi2) has been implicated in the pathophysiology of septic shock. however, pgi~'s role in the inflammatory response to sepsis is not well-defined. the purpose of this study was to identify which acute septic events are mediated by pgi 2 during graded bacteremia. methods: eleven ~nesrhetized, hemodynamically monitored adult swine were infused iv with aeromonas h. (109/ml) at rates increased incrementally from 0.2 to 4.0 mi/kg/hr over 4 hours. animals were studied in two groups: septic control (sc), graded bacteremia only (n=6); pga (n=5), graded bacteremta plus anti-pgiz antibody, 50 ml/hr iv, beginning at 2 hours. mean systemic (map) and pulmonary arterial (pap) pressures and arterial po2, mmhg, cardiac index (ci), l/min/m2, oxygen delivery index (do2i) and consumption index (vozi), ml/min/m2, and oxygen extraction (er), %, )latelet aggregometry (plt), %max., plasma pg6-keto f1 alpha ; in the first instance~ peak values of lt ~4 after i~ hrs post infarction were 6 times higher than in the controls and excess leucocyte infiltration was noted at the infarction zone. in second instance two levels of lt b4 led to weak infiltration of the infarction zone by leucocytes. a. mo~e~o, in~.~p~siolo~,d~t.e~.cardiolo~,bogotsolets4, ~ev252024, ukrmne systemic lesion$of erythron in traumatic disease and possibilities of their regulation by opioid peptides. redkin y. v., fominih s. g. using clinical (121 patients) and experimental material(128 rats and 74 dogs) we revealed general regularities of erythron lesions after hard mechanical trauma of various genesis as well as some mechanisms of development of posttraumatic anemia and possibilities of its correction with preparations of opioid peptides. the condition of central and peripheral compartments of erythron was studied with unified morphologic, immunogematological, biochemical and radiological methods. it was revealed that irrespective of the experimental animal species (dogs, rats) or in clinical experiments (patients) and irrespective of the injuring factor type (skeletal trauma, craniocerebral trauma, loss of blood) in erythron can be observed one-directed unspecific reaction realized by the considerable lowering of hemoglobin concentration, erythrocytes number and hematocrit. in the initial period (1-7 days) in the system of erythron prevail processes of distraction and elimination of er~zthrocytes relatively to the general production of stimulated erythropoiesis. the primary alterating factor is the prolonged intensification of peroxydation of membrane iipids of erythrocytes with simultaneous lowering of reserves of reduced glutathione. the distraction of erythrocytes is supported by the developing phenomena of autoallergization of organism that becomes apparent by the appearance of sensitized t cells and antierythrocyte antibodies. the intensified production of erythropoietin rules to the realization of he program of fetal and terminal (reserved) erythropoiesis. failure of erythropoiesis function is supported by disturbances of the processes of the injuring of cell metabolic apparatus. using of dalargin (15 microgram per kilogram of body mass intrap'eritoneally within 5 days after the trauma) showed the precise pharmacotherapeutic effect revealed by the diminishing of anemia of experimental rats, more . fiberbronohoscopic procedures are known to produce "peep-like" effects and to increase pulmonary artery (pa) resistance [2] . peep can affect rv function by reducing preload and ejection fraction (ef) [3] . since changes of rv function during bronchoscopy in septic patients are not reported, we measured rv parameters before, during and after fiberoptic bronchoalveolar lavage (bal). method: this 34-year-old patient (apache-ii:24) developed a hyperdynanlic septic state due to staphylococcus aureus (blood culture). we inserted a "fast response" thermistor pa-catheter (baxter-edwards) to evaluate rv performance [4] . the therapeutic procedure included volume replacement, vasopressors (dopamine 5, dobutamine 3 gg/kg/min. iv) and analgosedatior/. before bronchoscopy (olympus bf-30, od=6 mm) the patient received pancmonium for muscle relaxation. ventilation was not changed during the procedure (endotracheal tube: id=8 ram, bennett 7200a, pressure controlled mode, pm~x=25 mbar, peep=8 mbar, i:e=i:i, fio2=1.0). we measured rv enddiastolic volume (edv), stroke volume (sv), ef, heart rate (hr), cardiac index (ci) and mean pa pressure (mpap gerlach h, gerlach m, clauss m, falke kj renal hypoxia and/or ischemia initiates the development of a deteriorated medullary perfusion based on fibrin deposition in the peritubular capillaries, vasoconstriction, and perivascular edema, which is followed by a swelling of the tubular epithelial ceils, intraluminal tubular obstruction, and a backleak of fluid through the injured tubules into the renal interstitium, finally leading to an acute tubular necrosis (atn) [1 ], clinically diagnosed as acute renal failure (arf). one important pathway for induction of enhanced vascular procoagulant activity and permeability is based on the synthesis and expression of macrophage-derived cytokines, which bind to specific endothelial cell surface receptors. we recently described the identification and purification .of a new 55,000 dalton polypeptide, which is synthesized and expressed by murine macrophages after stimulation with lipopolysaccharide, and exerts procoagulant activity on cultured endothelial cells [2] . in the presented study, we demonstrate that the new polypeptid is also synthesized by macrophages under hypoxic conditions. the protein binds to specific receptors, which are expressed by endothelial cells dependent on the environmental oxygen tension. animal studies were performed after approval by the local committee for animal safety; the animals were anesthetized, treated and supervised in accordance with the guidelines of this committee. in contrast to other authors, who performed long-term hypoxia experiments in awake animals, we preferred to implement the studies under anesthesia for ethical reasons, although regulatory functions for ventilation might be influenced. animal studies demonstrated that the intravenous injection of the polypeptide initiates fibrin formation in the peritubular vessels. keeping the animals under hypoxic conditions induces similar effects, which are reduced by a rabbit-antiserum against the new protein. in conclusion, the new polypeptide obviously contributes to the pathogenesis of acute renal failure by tubular necrosis during and after hypoxic events. the use of verapamil as cardioprotective agents for management of patients with acute ischemic/reperfused heart is based on the assumption that the increased intracellular ca+ level is a key factor in causing cell death. our in vitro study was designed to focus on effects of verapamil on the metabolic potential of cardiac slices after reversible ischemia in rats. the material consisted of two main groups : group a (non ischemia/reperfusion group) and group b (ischemia/reperfusion group), each is subdivided into two subgroups (a and b). each subgroup included 10 rat hearts. group aa is the control group, group ab is verapami] added group. group ba is ischemia group without verapamil. group bb is verapamil added group. ischemic cardiac slices were obtained from rats subjected to 15 min. haemorrhage to induce reversible global ischemia. both nonischemic and ischemic cardiac slices were placed in well oxygenated krebs ringer phosphate buffer containing 150 mg% glucose & 5 gm% bovine albumin and incubated in dubnoff shaking water bath for 60min at 37°c the results revealed that there was an enhancement in release of free fatty acids (ffa) (240%) and lactate (163%) and in glucose uptake (160%) in group ba as compared with group aa. these metabolic alternations produced by ischemic cardiac slices were reversed by verapamil addition (200 ml%) but in group ab verpamil did not alter the release of ffa & lactate from non-ischemic cardiac slices, whereas it inhibited glucose uptake from these slices by 67%. the improvement of the metabolic intervention of ischemic myocardium indicates that verapamil may be of importance in reducing the extent and severity of acute myocardial ischemic injury in acute haemorrhage. severe endothelial dysfunction occurs following injury to carotid arteries which is characterized by a decreased ability of these arteries to dilate when challenged with ach or a23187, but not with a direct vasodilator (nano2). this failure to relax to ach and a23187 reflects an inability of endothelium to generate edrf, but relaxation recovers gradually to control values by 2 weeks. exogenous no donors (e.g., c87-3754 or spm-5185), accelerate the recovery of the injured endothelium in rat carotid arteries. intravenous infusion of an no donor (30 p.g/day) with an implanted osmotic pump significantly accelerated the recovery of regenerated endothelium to produce edrf at 7 days. rat carotid artery rings relaxed only 17 + 5% and 15 + 5% to 10 gm ach in vehicle treated rats and in inactive no donor treated rats respectively 7 days following injury compared with 67 + 6% in no donor rats (p<0.001). relaxation to 100 gm nan02 was normal in all groups indicating that the differences in relaxation were not the result of damage to vascular smooth muscle. contraction to l-name (1 mm) was markedly reduced by injury, but was protected by no donors (p<0.01). thus, exogenous no donors enhance the ability of the endothelium to regenerate and to release edrf in response to endothelium-dependent vasodilators. this may be due to an anti-proliferative and anti-mitogenic effect of no on vascular smooth muscle cells, allowing the endothelium to regenerate without intimal thickening. no also has been shown to inhibit platelet aggregation, and to attenuate neutrophil adherence and activation. the superoxide scavenging effect of no is not the basis for these effects since hsod is inactive in preserving endothelial function in injured arteries. thus, no exerts a variety of cytoprotective effects which may be of importance in protecting against vascular injury. much evidence has now accumulated to show that the excess production of the vasodilator nitric oxide (no) in sepsis is an important contributor to the hypotension and multiorgan failure characteristic of this condition. various cytokines play an important role in this process through their ability to induce the production of one of the enzymes responsible for no synthesis, the inducible no synthase (inos). we have studied the effects of cytokines on the induction of this enzyme both in vitro using vascular smooth muscle cells, and in a murine model of gram-negative sepsis. tn smooth muscle ceils, the cytokines il-1, ifnq', and tnf-oc show strong synergy with one another in the production of inos. in order to define the molecular basis for this synergic effect, we have linked the promoter of the inos gene to a "reporter" gene, chloramphenicol acetyl transferase (cat), and transfected these constructs into vascular smooth muscle cells. assays of cat activity reflect the activity of the promoter in this system, and by generating sets of deletion mutants of the promoter sequence we have been able to define the area within the promoter which mediates the synergic effect of these cytokines. in addition to stimufatory effects on inos production, certain cytokines are able to down-regulate the production of inos in vascular smooth muscle cells, and the effects of these counterregulatory cytokines will be discussed. the interaction of these cytokine effects in the whole organism has been studied in a murine model of gramnegative sepsis. widespread induction of inos occurs in this model as assayed by enzyme activity and through use of specific antisera to inos. neutralizing antibodies to tnf-~ and tfn-y are both able to prevent death in this model, but it is only the anti-ifn-y which attenuates the induction of inos assayed in the liver. clearly there is some redundancy in the effects of cytokines on the production of inos in sepsis, and greater understanding of the most important factors in inos production is required in order to target anti-cytokine therapy most appropriately. effects of nitric oxide on hepatocyte metabolism in inflammation. j. stadler, department of surgery, tu mqnchen, frg hepatocellular nitric oxide (no) synthesis is induced by proinflammatory mediators such as tumor necrosis factor, interleukin-1 and interferon gamma or by bacterial toxins such as lipopolysaccharide. stimulation of the hepatocytes (hc) with a combination of these agents leads to an output of no in quantities which are not seen in any other celltype. it has been demonstrated by various investigators that important effects of these cytokines and bacterial toxins on hc metabolism can be attributed to the action of no. in contrast to other celltypes hc seem to be relatively resistant to suppression of basic metabolic functions such as energy metabolism by no. therefore, cell damage has not been described to a significant extent following exposure to no. however, no does inhibit total protein synthesis. the exact biochemical mechanism of this phenomenon has not been uncovered yet, but it has been demonstrated for some specific proteins that their production is inhibited at a posttransscriptional level. as in many other celltypes cgmp generation is elevated in hc by no through activation of the soluble guanylate cyclase. cyclic gmp may possibly exert a plethora of metabolic functions, but it is interesting to note that most of the cgmp seems to be transported out of the cell. some very specific effects of no on hc metabolism include the inhibition of the glyceraldehyde-3-phosphate dehydrogenase (gapdh) and the cytochrome p450 (cyp) enzymes. inhibition of gapdh activity is mediated through nitrosylation of critical domains of the enzymes by no which enhances auto-adpribosylation. this effect on gapdh activity might be responsible for the inhibition of gluconeogenesis by no, which has been described recently. finally, no-mediated inhibition of cyps may help to explain the suppression of hiotransformation processes which is a characteristic featur,'~ r ~ "~flamed liver. nitric oxide (no) is an endogenous inhibitor of polymorphonuclear leukocyte (pmn) adhesion which limits pmn-endothelial cell interactions under normal conditions. we have previously demonstrated that following ischemia, no production by the vascular endothelinm is dramatically reduced. accordingly, we investigated the effects of no-donors on pmn accumulation and tissue injury following hemorrhagic shock and ischemia. hemorrhagic shock was induced in anesthetized rats by bleeding to 35 mmhg for 3 hours followed by reperfusion. segments of superior mesenteric artery (sma) were isolated and suspended in organ baths. in rats receiving saline sma relaxation to acetylcholine (ach, 100 nm) was reduced by 80% compared to control sma segments (p<0.01) while relaxation to sodium nitrite (100 gm) was unaffected. in addition, mesenteric tissue pmn accumulation as determined by myeloperoxidase (mpo) activity was significantly elevated compared to controls (p<0.0l). interestingly, treatment with the no-donating agent, s-nitroso-n-acetylpenicillamine (snap) significantly preserved sma relaxation (p<0.01), attenuated mesenteric mpo (p<0.05) activity, and significantly improved survival compared to saline vehicle. in anesthetized, open-chest dogs we investigated the cardioprotective actions of a novel no-donor, spm-5185 (schwarz pharma), following regional myocardial ischemia (1 hour) and reperfusion (4.5 hours) . treatment with spm-5185 (500 rim) significantly reduced myocardial necrosis by 70% (p<0.01) compared to an no-deficient analog of spm-5185, spm-5267. furthermore, mpo activity within the ischemic-reperfused zone was also significantly (p<0.01) reduced following treatment with spm-5185 compared to spm-5267 (1.6 + 0.5 vs. 3.8 + 0.3 u/100 mg tissue). these data strongly suggest that no is a potent inhibitor of pmn-mediated tissue injury following hemorrhagic shock as well as in acute myocardial ischemia-reperfusion injury. overproduction of nitdc oxide (no') may contribute to sepsis-induced hypotension. during septic shock, excess no" is produced by an isoform of nitric oxide synthase (nos) which is induced by inflammatory mediators. nonselective nos inhibitors have been proposed as a new therapeutic approach to treating hypotension in septic shock. we studied the differential hemodynamic effects of n~-methyi-l-arginine (l-nma), a nos inhibitor, in normal canines versus those challenged with endotexin (lps) and compared the activity of this drug across the venous, pulmonary and systemic vascular beds. awake canines were challenged with lps (0 mg/kg, n=15:8 mg/kg, n=21 ; or 16 mg/kg, n=30) and treated with l-nma (0, 1,2, 4,10 mg/kg/hr) for 22 hours following a 0, 10, or40 mg/kg loading dose. animals were resuscitated with iv ringers solution (10 ml/kg/hr). hemodynamic data were collected at 0, 2, 6, 10, 14, and 22 hours using intravascular catheters and radionuclide heart scans and analyzed by anova. in both normal and endotoxemic animals, l-nma at all doses studied similarly increased mean arterial pressure (p=0.07), and systemic vascular resistance index (p=0.ol) and decreased cardiac index (p=0.05) and oxygen delivery index (p=0.01). in contrast, the effect of l-nma on mean pulmonary artery pressure, central venous pressure, pulmonary capillary wedge pressure, and pulmonary vascular resistance index was greater in lps-challenged canines compared to normal animals (p<0.05), but this differential effect on the venous and pulmonary circulation occurred, >6 hours after lps challenge. l-nma did not significantly increase survival rates or times at any of the doses studied (1, 2, 4, or 10 mg/kg/h) in either the low (8 mg/kg) or high dose (16 mg/kg) lps-challenge groups. a nonsignificant (p>0.2) trend toward a beneficial effect on survival ol low dose l-nma (1 mg/kg/h) in animals given the 8 mg/kg lps-cha[lenge was not enhanced by increasing the lethality of the model or by administering higher l-nma doses. at the highest l-nma dose used in this study (10 mg/kg/h), survival time decreased significantly for both the low and high dose lps-challenge animals (p<0.05). this increased mortality was not explained by changes in plasma concentrations of either lps or tnfc~. thus, l-nma did not have a greater effect on the systemic arterial circulation in endotoxemic compared to normal canines. however, in the venous and pulmonary vascular beds, the effect of l-nma increased with time after endotoxin-challenge these data suggest the induction of nos activity by endotoxin in canines may be relatively greater in venous and pulmonary vessels compared to systernic arteries. l-nma, a nonselective nos inhibitor, did not decrease mortality in endoloxemic canines and the highest dose studied was harmful. pulmonary hypertension (ph) and arterial hypoxemia are characteristic features of the adult respiratory distress syndrome (ards). reducing pulmonary vascular pressures may promote the resolution of pulmonary edema. intravenously infused vasodilators lower ph in ards, but, as a result of their general vasodilatatory effects, systemic mean arterial pressure may also decrease. furthermore, blood flow may be increased to non-ventilated or poorly ventilated lung areas resulting in a rise of intrapulmonary shunt, thus causing a further fall in pad 2. recently, short term inhalation of low concentrations of the gas nitric oxide (no), an endogenous endothelium derived relaxing factor, which is rapidly inactivated in blood by hemoglobin, was reported to decrease ph without causing systemic vasodilation in sheep [1]. similar changes have been observed in patients with severe ards during repeated short term inhalation of no (18 and 36 ppm), which rapidly and selectively decreased the mean pulmonary artery pressure (pap) and, in contrast to intravenously infused prostacyclin, induced a remarkable increase of pad2 [2] . this improvement in oxygenation was caused by a redistribution in blood flow away from intrapulmonary shunt areas to normal ventilated lung regions. continuous no inhalation (5-20 ppm) consistently lowered the pap and augmented the pao2/f.o 2 for up to 53 days. no negative side effects were observed during the whole time span examined. in particular methemoglobin levels always remained below 1.5%. following these investigations, it could be shown that these effects may also occur using concentrations in the parts per billion range [3] , which may reduce possible toxic side effects. however, in the same study it was demonstrated that the dose-response curves for pa02 and pap have different patterns. whereas pap presented a continuous dose-dependent downward tendency with an eds o of approximately 2-3 ppm, the improvement of oxygenation had a maximum at 10 ppm and, at higher doses, drifted back towards the baseline data. the ed~o was estimated at approximately 100 ppb, i.e. more than ten times lower than for the reduction of pap. in conclusion, inhalation of no by patients with severe ards may result in persistent and reproducible decreases in pap associated with an evident improvement in pad2, thus allowing reduction of the f.o 2. no inhalation should be performed using low concentrations which are less toxic, although any possible risks still have to be considered carefully. dose-response studies for the individual patients are recommended urgently. finally, controlled randomized studies are required to demonstrate that additional no inhalation is able to reduce mortality of ards. inhibition of the activity of glyceraldehyd-3-phosphate dehydrogenase (gapdh), an enzyme of the glycolysis/gluconeogenetic pathway, through adp-ribosylation is promoted by nitric oxide (no). since no is produced in the septic liver and hypoglycemia is a major problem of late sepsis, it was investigated whether no interferes with gluconeogenesis of hepatocytes. hepatocytes (hc) were isolated from sprague-dawley rats using a collagenase perfusion technique and differential centrifugation. exogenous no was applied by incubation with the no-donors s-nitrosyl-acetylpenicillamine and sodium-nitroprusside. endogenous no synthesis was induced by incubation with cytokines (tnfcq il-1, ifnj and lipopolysacchafide (lps). 24 hrs later the incubation medium was changed to a solution containing lactate, ornithine, lysine, ammoniumchloride and glucagon for optimal conditions of gluconeogenesis. after 3 more hrs glucose and nitrite levels were determined spectrophotometrically. gapdh activity was measured by the nadh-dependent conversion of 1,3-diphosphoglycerate to glyceraldehyde-3-phosphate. incubation of hc with no-donors led to a concentrationdependent inhibition of gluconeogenesis and gapdh activity. however, gapdh activity was about 5 times more sensitive to the inhibitory effect of exogenous no. incubation of hc with cytokines and lps induced nq synthesis as measured by an increase in nitrite concentrations. endogenously produced no suppressed gluconeogenesis by 49_+3%. in contrast to exogenously applied no, the effect of endogenous no synthesis was less on gapdh activity resulting in an inhibition of only 32_+12%. in conclusion, exogenous and endogenous no inhibited gluconeogenesis as well as gapdh activity. however, there was no correlation between the extent of inhibition of these two parameters of hepatocellular glucose metabolism. we have shown that inhibition of hepatocyte (hep) synthesis of nitric oxide (no) potentiates cell injury in a model of acetaminopheninduced oxidative stress and the extent of damage was paralleled by depletion of reduced glutathione (gsh) stores. to clarify the role of no in modulating the redox state of hep, we studied the effect of inhibition of cytokine-mediated no production on hep gsh stores, in a system of isolated rat hep in primary culture, no synthesis was induced (stim) by exposure to il-1, tnf, ifn, and lps for 16 hours. 20, 60, 120 and 200 ~m of n-monomethyi-l-arginine (nmma), a specific inhibitor of no synthesis, was added. cells incubated in media alone served as controls (cont). the no metabolite (no2); aspartate aminotransferase (ast), an indicator of cell injury; and gsh were assayed. (data presented as mean + sem; n=4.) gsh (nmovma orotein) ..~ (nmol/ma orotein) cont 14.1 + 0.3 32 + 4.0# stim 13.4 + 0.6 139 + 12 stim+2o tzm nmma 12.7 + 1.1 88 + 5.0# stim+60 ~m nmma 9.9_..+ 0.4* 68 + 4.5# stim+120 pm nmma 7.6 + 0.6* 57 + 3.0# stim+200 )lm nmma 5.0 + 0.5* 37 + 2.5# anova 0,0002 0.0001 (* p < 0.05 versus stim, # p < 0.01 versus stim; anova with neuman-keuls) gsh in cont+120 i~m l-nmma was equivalent to that of cont (13.3 vs.14.1). ast release was equivalent in all treatment groups. these data show that inhibition of hep synthesis of no depletes intracellular stores of reduced gsh. we conclude that hepatocyte no production modulates cellular gsh homeostasis and as a result, may be hepatoprotective in oxidative injury. nitric oxide (no) is a modulator of immune response and may be involved in the changes in immune reactivity after major trauma and operations. we investigated no-generation in rat and mice spleen cells (sc) after partial hepatectomy (ph). c57bl/6 mice and lew rats underwent a 50% and 70% ph, respectively. sc were prepared 1-6 days after ph and plated at 1 to 2 x 10ecells per well. after 20 h incubation at 37°c, no-production was measured as nitrite levels (griess reagent). normal mouse sc did not produce no, neither basal nor in response to lps or con a starting at the second day after ph, we found a substantial production of no. in rats, also sc from control animals were able to generate no; both basal and stimulated no-generation were further enhanced after ph (table, values expressed as mean --se). after shame operation, there was only a modest elevation of noproduction in rat and mouse sc. in first experiments we could demonstrate no-production also in phagocytes from a patient 3 days aider liver partial resection (1.4 nmol nitrite/106 cells) enhanced no-production in macrophages may contribute to the changes of immune reactivity after partial hepatectomy. nitric oxide (no) is recognized as an important mediator in endotoxemia and sepsis. increased synthesis of no has been demonstrated in septic humans and animals, and no inhibitors have been used in the treatment of septic shock. recent reports have, however, suggested that this form of therapy may cause serious organ damage. in the present investigation circulatory and metabolic changes in the liver were studied during treatment with the no-synthase inhibitor n-nitro-l-arginine-methyl ester (l-name) in endotoxemia. methods: juvenile pigs were randomized to one of the following treatment groups: 1) encletoxin and l-name, 2) endotoxin, 3) naci and l-name, 4) nach preliminary results from groups 1 (n=7) and 2 (n=8) are presented. catheters for pressure measurement were introduced into the aorta, hepatic and portal veins and ultrasonic transit time flow probes were placed on the hepatic artery and portal vein. a catheter was introduced into the pulmonary artery. endotoxin (1.7 gg/kg/h) was given as a continous portal infusion over the entire observation period of 8 hrs. l-name (25 mg/kg) was given as a bolus after 3 hrs. of endotoxemia. results: endotoxin transiently reduced portal vein flow (pvf) by 25%* and hepatic artery flow (hal e) by 45%*, while l-name caused a further and lasting reduction in flow (pvf 78%, haf 90%)*. transhepatic (portal-hepatic vein) vascular resistance increased to 3 times baseline value during endotoxemia while l-name caused a further marked increase in resistance to 12 times initial value. portal oxygen saturation (so2) decreased by 60%* during endotoxemia. l-name caused a reduction in portal so2 by 87%*. arterial so2 was unchanged in both groups. hepatic oxygen uptake was not changed by endotoxin, but was markedly reduced after addition of l-name. endotoxin caused a 27% reduction in cardiac output (co). the addition of l-name reduced co by a total of 71%*. *: p < 0.05. conclusion: is the present model of endotoxemia treatment with the nitric oxide synthase inhibitor l-name markedly reduced liver perfusion and portal oxygen supply. this might explain the increased liver damage reported in previous studies using no-inhibitors. the increase in transhepatic resistance found after l-name treatment will tend to cause pooling of blood in the splanchnic veins, resulting in reduced filling of the heart and thus contribute to the observed reduction in cardiac output. institute for surgical research, rikshospitalet, the national hospital, university of oslo, 0027 oslo, norway. we have investigated the role of tumour necrosis factor (tnf) and interleukin-i (il-i) in the induction of nitric oxide synthase (nos) by bacterial endotoxin (lipopolysaccharide; lps; 2 mg kg -i i.v.) in vivo. in anaesthetized rats, pretreatment with a monoclonal antibody for tnf (tnfab; 20 mg kg -i s.c., at 16 h prior to lps) or with an il-i receptor antagonist (il-ira; 16 mg/kg bolus and 2.4 mg/kg/h infusion) ameliorated the fall in mean arterial blood pressure (map) at 90-180 min after lps. for instance, endotoxaemia for 180 min resulted in a fall in map from 114-+6 (control) to 79-+4 mmhg (p<0.01; n=8). in contrast, animals pretreated with tnfab or il-ira prior to lps injection maintained significantly higher map at 180 min when compared to lps-control: 118-+3 mmeg (n=5) and 103-+7 mmhg (n=5), respectively (p<0.01). three hours of endotoxaemia significantly reduced the contractile effects of noradrenaline (na) in the thoracic aorta ex vivo. the hyporeactivity to na was partially restored by in vitro treatment of the vessels with ng-nitro-l-arginine methyl ester (l-name, 20 min, 3x10 -4 m). pretreatment of rats with tnfab or il-ira significantly (p<0.05) prevented the lps-induced hyporeactivity of rat aortic rings ex vivo. l-name did not alter or only slightly enhanced the contractions of aortic rings obtained from tnfab or il-ira treated lps-rats, respectively. at 180 min after lps there was an induction of calcium-independent nos activity in the lung (5.14-+0.57 pmol citrulline/mg/min, n=8), which was attenuated by tnfab and !l-ira by 37-+6% and 46-+6%, respectively (n=5; p<0.05). thus, the production of both tnf and il-i contributes to the induction of nos by lps in vivo. the protective effect of agents which inhibit the release or action of tnf or il-i in shock may be, in part, due to inhibition of nos induction. neal garrison, md objective: sepsis is often accompanied by organ dysfunction, in part due to impaired microvascular perfusion. recently, nitric oxide (no) has been described as an important mediator of the hemodynamic changes of sepsis, and no synthase (no-s) inhibitors have been advocated for treatment of septic shock, but their visceral microcirculatory effects are inadequately characterized. we postulated that no-s inhibition would exacerbate the impaired organ perfusion of sepsis. methods: six groups ofdecerebrate rats were studied. bacteremia was induced with 109 live e. coli, which consistently increased cardiac output 15-20% above baseline (bl). the no-s inhibitor nm-nitro-larginine methyl ester (l-name,1 mg/kg iv), prevented this increase and elevated map by 25-30%. in the first 3 groups, total hepatic blood flow (thbf, ml/min by time transit flowmetry) and microvascular perfusion (mi-ibf, ¼ bl by laser doppler flux) were measured. in the other 3 groups, in vivo videomicroscopy was used to observe renal microvascular responses (ila=interlobular artery, aff=afferent arteriole, eff=efferent arteriole; % bl for all). results: data are 60 rains after e. cob. n=5-7/group. * p<0.05 vs bl by remanova and § p<0.05 vs e. coli alone by anova. ec+l-name 11-+2 -57_+10" § -45_+4* § -89_+3* § -33+5* -20+5* § conclusions: l-name administration in controls decreased renal blood flow, indicating no contributes to basal renal tone. bacteremia decreased mtlbf but not thbf, and mi-ibf was further impaired by no-s inhibition. e. coli caused renal preglomemlar, but not postglomerular constriction and reduced flow. l-name exacerbated these e. coli-induced alterations and caused eff constriction. these data indicate that no-s inhibition exacerbates bacteremia-induced impairment of renal and hepatic blood flow, suggesting that no is an importam compensatory dilator mechanism in these organs during sepsis. irf (iron responsive factor) is the central regulatory protein of intracellular iron metabolism able to bind to responsive rna elements (ires) present atthe 5'untranslated region (utr) of ferritin mrna and 3'utr of transferrin receptor mrna. binding of irf to ires results in repression of ferritin mrna translation and increased stability of transferrin receptor mrna leading to enhancement of transferrin receptor translation. we describe here that either tetrahydrobiopterin dependent stimulation as well as cytokine (ifn-~)/lipopolysaccharidemediated induction of nitric oxide synthase activates irf, which is due to direct interaction of nitric oxide with the iron-sulphur-cluster of irf. this was shown by gene expression studies using a plasmid containing a ferritin ire and a cat indicator box which was transfected into k562 myelomonocytic cells, which were shown to have a constitutive form of nitric oxide synthase (nos). furthermore, the increased binding of 1re to irf due to irf activation of irf by nitric oxide was demonstrated by gel shift assays. irf activity was much more increased in cellular extracts from murine macrophages (j774) where a cytokine inducible form of nos has been characterized earlier as compared with irf activity in k562 cells, where nos was stimulated by increasing the availability of the essential nos cofactor 5,6,7,8-tetrahydrobiopterin. we then demonstrated that activation of irf by nitric oxide is accompanied by alterations in ferritin translation as checked by metabolic labeling and immunoprecipitation. these results suggest a reasonable mechanism for the regulation of iron disturbances under chronic inflammatory disorders, characterized by increased concentration of immune activation parameters like ifn-5or neopterin and low serum iron and hemoglobin concentrations. taken nitric oxide, no, the putative endothelial derived relaxant factor, edrf, has been shown to be a potent inhibitor ofplatelet aggregation in vitro. in vivo evidence however, is scarce. accumulation of platelets in the lungs has been shown to occur during extracorporeal circulation. the aim of the present study was to investigate the effect of inhaled no on this reaction. materials and methods: the animals were divided into two groups, each consisting of 7 pigs. platelets were selectively labelled with luln-oxine. dialysis was instituted via catheters in the femoral vessels. in group 1, no, 50 ppm, was added to the inhaled gas from the start of dialysis. in group 2 no was not given. the activity over the lungs was followed dynamically with a gamma camera. central hemodynamics was monitored via a swan -ganz catheter. results: the activity was significantly lower in group 1, from 2 minutes after start of dialysis and onwards, indicating diminished accumulation of platelets in the lungs. parallel to this the hemodynamic response in terms of increased pulmonary artery pressure and pulmonary vascular resistance was blunted in this group conclusion: inhaled no in this model seems to affect pulmonary platelet sequestration. an associated attenuation of the changes in central hemodynamics was also seen. previous studies from our laboratory have demonstrated that vascular contractility decreased in endothelium-intact blood vessel rings in early and late stages of sepsis. although endothelium removal in early sepsis restored vascular contraction, the depressed smooth muscle contractility observed in late sepsis was not restored by endothelium removal. this indicates that impairment of smooth muscleper se may be responsible for such dysfunction in late sepsis. the aim of this study, therefore, was to determine whether or not smooth muscle-derived nitric oxide (no) plays a role in producing vascular smooth muscle dysfunction during late stages of sepsis. to study this, rats (250-300g, n=4-5/group) were subjected to sepsis by cecal ligation and puncture (clp). septic and shamoperated rats then received 3 rrd/100g bw normal saline. the animals were killed at 10, 20, or 35 h post-clp (10 h post-clp=early sepsis; 20-35 h post-clp=late sepsis), and thoracic aortic rings were prepared for contraction studies using organ chambers. the complete removal of endothelial cells was tested by the absence of any significant acetylcholine-induced vascular relaxation. contractile responses to norepinephrine (ne, 10 9 to 10 -5 m) were determined in the aortic rings without intact endothelium. ng-monomethyl-l-arginine (l-nmma, 300/~m, an inhibitor of no synthase) was then added to the organ chamber and ne-induced peak contraction was determined before and after the addition of l-nmma. the peak contraction (rag/rag tissue, mean_+sem) is shown below: the results indicate that the addition of l-nmma did not significantly affect ne-lnduced peak contraction in endothelium-denuded vessel rings at 10 and 20 h after clp. in contrast, l-nmma administration produces an 18% increase (p<0.05) in peak contraction during late sepsis. therefore, the vascular smooth muscle contractile dysfunction observed at 35 h post-clp is partially due to smooth muscle-derived no over-production. thus, unlike macrophages in which inducible nitric oxide synthase (inos) is observed in early sepsis, the inos in vascular smooth muscle appears prominent only in the late stages of sepsis. in three cases of human septic shock in which ng-monomethyi-l-arginine, (l-nmma) a nitric-oxide-synthase-inhibitor was applied, we isolated three completely different types of pathogens: candida, pseudomonas aeruginose and multiresistant coagulase-negative staphylococci. this observation suggests that endotoxin alone is not the main factor triggering hypotension in septic shock by the nitric oxide pathway. in a 68-years-old woman in severe septic shock due to a candida and pseudomonas aeruginosa infection complicated by adult-respiratorydistress-syndrome conditions deteriorated despite adequate conventional therapy. in this trial, effects of l-nmma on cytokin-levels were investigated. the study-protocol was approved by the ethical committee of the department of surgery. after two boll of 200 mg of l-nmma, a continuous infusion was installed (0.05 mg/minute and kg body weight l-nmma). as expected mean arterial blood pressure rose (62 to 134 mmhg}, heart rate stayed stable (124 + 4 b/rain), systemic vascular resistance increased (225 to 354 dyne.sec/cm5), cardiac output decreased (17 to 15.2 l/rain), and cardiac index declined (9.94 to 8.63 l/min/m2}. before and after 90 minutes while the infusion of l-nmma, blood samples for immunological measurements were taken and processed together. pulmonary-shunt-volume was observed before the application of l-nmma, after one hour and after 140 matutes. neopterine increased from 16.51 to 32.55 ng/ml, tumour-necrosis-factor-a increased from 24.16 to 36.61 pg/ml and intedeukin-6 increased from 61.9 to 98.2 pg/ml. immunoglobulines a, g, and m (3.2 to 2.9, 8.1 to 7.6, 1.1 to 0.9 g/i), complement factor c-3c and c-4 (0.54 to 0.50, 0.22 to 0.23 g/i), alpha-l-antitrypsine (3.86 to 3.83 g/i), c-reactive-protein (111.1 to 105.6 rag/i), interleukin-1 (0 pg/ml) and soluble interleukin-2 (2128 to 1983 units/ml) did not change significantly. pulmonary-shuntvolume decreased from 54.1% to 35.4% within one hour and to 36.6% after 140 minutes. in septic shock blocking nitric oxide as an intervention at the end of a not ~,et ful!y understood cascade might have important influences on pulmonary-shunt-volume and inter-cell-communication. department of surgery, pharmacy* and immunology**, university hospital of zurich, r~imistrasse 100, 8091 zurich, switzerland we previously reported that hypoferremic cba mice had an increased resistance to salmonella infection, and that injection of ammonium ferric citrate (afc) to these mice led to enhanced infection (ganthier et at. 1986. microbiol.immuno130:425) . because nitric oxide (no) is involved in the antimicrobial activity of routine macmphages towards various inttacellular pathogens, we investigated the influence of iron on the bactericidal activity of cba mouse macrophages towards s.typhimurium and on the production and activity of reactive nitrogen intermediates (rni). peritoneal macrophages hum cba mice were cultured in the presence (or not) of afc 50 ,um, ifn-,/250 u/ml, lps 1 fig/m/, ngmonomethyl-l--arginine (mmla) 2ram. nitrite (no2-) content of the supematants was determined by a standard griess reaction, and h202 release was measured by the peroxidese dependant oxidation of phenol red. for intracellular killing, macrophages monolayers were infected, and, at various intervals, lysed by triton x-100, and surviving bacteria enumerated by colony counting on agar. for in vivo experiments, mice were infected ip with 0.5 ml of a suspension of 5.10 ~" s.typhimurium, strain c5, and injected with aminoguanidine (ag) 1 mg/ml in saline. our results show that the rn[ inhibitor ag strongly accelerates the mortality of infected mice, the survival rate decreasing from 60% in the control group to 20% in the treated group, 10 days after challenge. correlatively the rni inhibitor mmla induces in vitro a decrease in the rate of bacterial killing, fxom 78% to 57%, in macrophages triggered with ifn-? + lps. the cultivation of macrophages in the presence of afc leads to a decreased no 2-accumulation, 4.7 nmole/well v.s. 21 nmole/well. conversely h202 production is enhanced from 09 nmole/well up to 2,33 nmole/well. nevertheless, macrophages cultivated in the presence of afc exhibit an increased tale of intracellular killing, 77% in iron exposed macrophages v.s, 68% in control macrophages. when triggered with ifn-~, alone, macrophages have a reduced antibacterial activity (57% v.s. 68%) whereas the addition of afc to these macrophagas restores an elevated (72%) rate of killing. in conclusion, the results show that bactericidal activity of cba macrophages towards s.typhimurium depends on the production of no by these macrophages ; but they also demonstrate that no is not the only reactive species involved in the intracellular kil/ing of s.thyphimurium ; indeed afc which strongly inhibits rni production, stimulates h20 2 release by these macrophages and increase their bactericidal activity in vitro. nevertheless afc may promote bacterial growth in vivo. crssa. unit4 de microbiologie. bp 87. 38702 la tronche cedex france. henning jahr, ulrike noack, karin braun the large amounts of no produced by the inducible no synthase in rat macrophages have direct antimicrobial effects, but inhibit the activation of the lymphocyte-dependent host defense system. the aim of this study was to investigate if complement activation influences no-generation. spleen cells from lew rats were incubated at 37°in tcm-199/5% fcs, with or without additional rat serum. after 20 h, nitrite (end product from no metabolism) was measured by oriess reagent. in rat spleen cell preparations, most of the no is produced by macrophages. complement activation in vivo was carried out by i.v. injections of 240 u cobra venom factor/kg b.w. at days 0 and 2. significantly higher (p<o.o1) lps-stimulated no-generation was found in spleen cells prepared at day 4 (13.0 4-1.3 nmol nitrite/106 cells, n=6) compared to spleen cells from non-treated animals (3.6 ± 1.3 nmol, n=6) complement activation was effective also in vitro. when incubated in 25% zymosan-activated rat serum, both basal and lps-stimulated noproduction were enhanced in spleen cells (table, values increased production of nitric oxide (no) is associated with sepsis, however, the effect of no inhibition on survival during sepsis is unclear. we examined the effect of no inhibition on host's ability to eliminate bacteria and survival in mice sepsis model. sixty female balb/c mice were injected with e.coli (10qbody) into peritoneal cavity. the treated group received n-ultro-l-arginine-methyl-ester (l-name), an inhibitor of nos, one hour before bacterial challenge. thirty mice were observed for survival after e.coli challenge and the other mice were sacrificed at 4 hours. blood was obtained by cardiac puncture and peritoneal lavage fluid (plf), liver and lung were harvested. the number of peritoneal exudative cell (pec) was counted and colony forming units (cfld of bacteria in the tissues, blood, and plf were also determined. in addition, pec was cultured in vitro with or without lipopolysaecharide (lps). tnf levels in the blood, plf, and supematants of cultured pec were measured by l929 bioassay. survival rate (%) qrql/,p n 8 hr 24 hr 4 dav control (normal saline 0.1ml/body i.p.) 10 100 50 40 n10 (l-name 10mg/kg i. administration of l-name before e.coli injection increased the number of bacteria in plf and tnf level in plasma, the result suggests that nos inhibitor may depress the host's ability to kill the injected bacteria and that it may induce greater systemic tnf production. we conclude that nos inhibitor is detrimental to animals in sepsis. the hypothesis that sod prevents reperfusion injury to the liver by protecting the endogenous endothelium derived vascular relaxing factor (no) from being inactivated by oxygen free radicals should be investigated. male wistar rats were subjected to 60 min of partial liver ischemia (70%) and 30 min of consecutive reperfusion in situ. some rats were pretreated with the no synthase inhibitor nitro-l-arginin (nla: i0 mg/kg). sod, when used, was given as mn-containing preparation at a dose of 6000 u. i.v. i0 min prior to ischemia. results (sod/ nla+sod/ ni~,, respectively): bile flow (~i/g/min) : 0.93±0.11"#/ 0.52±0.08/ 0.48±0.05. alt (u/2) • 171±40~#/ 477±173/ 506±182. gldh (u/l): 5.2±1.2e#/ 10.7±4.2#/ 24.3±9.1. in absence of nla, sod enhanced postischemic bile flow and reduced leakage of alt and gldh into the plasma, while the effects of sod disappeared, when endogenous 140~synthesis was inhibited by nla. nla had no ~ffect on untreated animals submitted to the same protocol. these results may suggest, that oxygen free radicals interfe~:e with the endogenous vasodilator no and that the benefit of sod can be attribuated in large part to a protection of no during reperfusion. nitric oxide (no) reduces pulmonary vascular resistance and increases oxygenation by inhalation in ards patients [1] . animal studies on the endogeneous no production proved that no is exhaled after synthesis in the respiratory tract [2] . with approval by the hospital ethics committee, we studied healthy volunteers and ventilated patients by measurement of inand exhaled no by no/no2-chemiluminescence in segments of the upper respiratory tract. we found that no is synthesized predominantly in the nose and in the upper nasopharynx, leading to inspiratory tracheal no concentrations of 0.07-0.13 parts per million in non-smoking volunteers; data during mask ventilation per nose were as follows: non 0.089 0.0422 0.0721 0.0311,2 parts per million (ppm) no, mean, n = 10; ~ p < 0.05 (t-test) between smokers and non-smokers; 2 p < 0.05 (t-test) between in-and expiration. about 50% of no is resorbe.d by the lung, and the exhaled no -in contrast to former presumptions -is the remaining part of the autoinhaled no. intubated and ventilated patients are deprived of no autoinhalation, and the exhaled no in the trachea decreases more than 95% {o.043 ppm before intubation vs. 0.003 ppm after intubation, ~, < 0.01), whereas the nasal no concentration increases by cumulation. hence, low-dose no inhalation in ards patients might be considered as a no replacement, especially since the tracheal no concentrations we measured in volunteers are similar to those which could be demonstrated to be effective for ards in former studies [3] . the data demonstrate that no is synthesized in the nose, and inhaled and resorbed by the lung. this phenomenon of no autoinhalation, which is reduced in smokers and in ventilated patients, does possibly contribute to the regulation of the ventilation-perfusion ratio in the lung. kikuchi 1, yoshihiro inoue 1, masao yoshida 2 1 critical care and emergency center, and 2department of bacteriology, iwate medical university. nitric oxide (no) is produced by macrophages and vascular endothelial cells. it is thought to be the true form of endotheliumderived relaxing factor, and to be involved in the fall of blood pressure during septic shock. we have reported previously that endotoxin (lps), tnf-c~ and il-2 contribute to the occurrence of septic shock (circ shock 38:264; . the present study investigated the in vitro effects of lps, tnf-o~, il-2 and ifn-'i on the production of no by macrophages. peritoneal macrophages were cultured with lps, il-2, tnf-~ and ifn-¥. in culture with lps, no was produced in a dose-dependent manner, and the production was suppressed by a no production inhibitor, l-nmma. ifn-y, il-2 and tnf-c~ used alone did not promote the production of no, but helped lps to facilitate no production. a combination of il-2 and tnf-c( enhanced lps-facilitated no production to a greater extent than il-2 or tnf-c~ alone. the above results suggest that these cytokines are involved in endotoxin shock through the mechanisms that facilitate no production. 19-1 uchimaru, morioka 020. japan. t. yolk 1,2, i. ioannidis 1, w.j. kox 2 and h. de groot 1 objectives: nitric oxide (no.) is known to play an important role in neurotransmission, vasoregulation, and bactericidal as well as tumoricidal cellular defense systems. by means of no-donating agents we studied the mechanism of no-mediated cytotoxicity against rat liver microvascular endothelial cells. materials and methods: 3-morphoiinosydnonimine-n-ethylcarbamide (sin-l) and sodium nitroprusside (snp) were used as no. donators, ldh release and trypan blue exclusion were applied to indicate cell viability. results: sin-1 (5mm), which releases both no. and o2-., caused a time-dependent cytoreduction of 60% and 80% after 3 and 6 hrs, respectively. this effect was not inhibited by superoxide dismutase (sod), which removes 02--to form h202 and 02. in contrast, the addition of catalase (cat), which removes h202 to form h20 and 02, diminished the cytotoxic effect ot sin-1 to 30%, equivalent to high concentrations of snp (20mm), which solely releases no.. in addition, the amount of h202 formed by 5mm sin-1 equaled the amount ot enzymaticany produced h202 by 5mu/l glucose oxidase (god). whereas god in this concentration and snp in low concentration (5mm) alone were not toxic to endothelial cells, the combination caused an 80% reduction of viability, comparable to the effect observed with sin-1 (5mm) alone. moreover, toxicity mediated by high concentrations of snp (20mm and 50ram) was reduced by 20% under hypoxic conditions indicating synergism with no-and 02. conclusions: we conclude, that no.-induced toxicity against endothelial cells is enhanced slightly in the presence of oxygen and is not due to an interaction with 02-. however, toxicity increases dramatically in the presence of h202. this may possibly occur either by a chemical formation of more potent reactive hydroxyl radicals or by independent actions on different cellular targets. although it is becoming increasingly clear that nitric oxide (no) is associated with otxan dysfunctions in septic patients, little is known as to die kinetics of no production in these patients to clarify these kinetics, we have investegated serum arid monocyte associated no in septic patients. five patients who had undergone gastrointestinal surgeries mid were complicated postoperatively with severe sepsis served as the subjects in this study (sgroup), using twelve healthy volunteers as a control group (c group). serum no2 and no3 levels were measul*d sliectrophotometricallymonocyte cultures were done for 24 itrs with or without lps+ifn-t stimulation mid no2 and no3 levels in the supernat,'utts were also measured spectrophotometrically. furthermore,using an no selective electrode,time course of the lps+ifn-t induced no production by monocytes was studied. l) serum no2 levels in s group were slightly lfigher than dmse in c group,bnt there were no significoatt differences between the two. 2)both no2 levels in the supematmlts of monocytes cultured with lps+ifn-t mid no2, no3 levels ill lhe supernatants of monocytes cultured without lps+ifn-y were significantly higher in s group. further,the time required for the no production by enltnred monoeytes was siglfificantly shorter also in s groap. conclusions l)in severe septic patients, monocyte no productions wei~ not only primed, but also activated concomitantly, suggesting that these monocyte activations m'e strongly associated with organ dysfunctions in sepsis. 2)based on the restdts of serum no nteasuremeuts, it can be deduced that no p~'oduced by monocytes in septic patients affects tissues and org~s ioeoregiomflly. objectives of the study: nitric oxide (no) is an important messenger which accounts for a wide spectrum of physiological and pathophysiological processes, e.g. mediating vasodilation in endoloxin shock investigating the signal lransducfion pathways involved in the regulation of the inducible nitric oxide synthase (inos), we report the involvement of phosphatidylcholinespecific phospholipase c (pc-plc) and proteinkinase c (pkc). materials and methods: no-production was induced in the murine macrophage cell line j774 with lipopolysaccharide (lps) [lljg/ml] and interferon ;f (ifny) [200u/ml]. after 18 hours of incubation no-release was determined as nitrite (no2) by using a colorimetric assay based on the griess-reaetion. results: preincubation of cells with the pkc-inhibitors staurosporine (stp), chelerythrine, bisindolylmaleimide (bim) and d609, that recently has been identified as a selective inhibitor of pc-plc, diminished significantly lpsand ]fnt-induced no2-production (p<o,001). although we observed no toxic effect on cell viability, no2-production was related to protein concentrations to exclude any inhibitory effect on celt growth. celts preincubated with stp [lo0nm] released 20% less no 2 in response to lps and ifn,[ compared to control cells cultured without inhibitor. chelerythrine [30}jm] and bim [ioijm] reduced the no2-formation by 50% and 85%, respectively. interestingly, the most potent inhibitor of no-production turned out to be the xanthate d609 cells pretreated with d609 [301~g/mt] released 96% less no 2 than stimulated controls the inhibitory effect on no 2production decreased the later the inhibitore were added after stimulation; e.g. bim added 6 and 12 hours after stimulation reduced no2-production only by 50% and 12%, respectively. accordingly, inqs activity present in stimulated cell lysates supplemented with l-arginine and co-factors was not suppressed by the inhibitors tested. conclusions: our findings suggest, that the induction of inos but not its activity is a pkc and particularly pc-plc dependent process. dr. k tschaikowsky, institut for anaesthesiologie, universital erlangen--nqrnberg, krankenhausstr 12, 91054 erlangen interleukin-1 (il-1) affects nearly every cell type by increasing the expression of a variety of genes associated with the promotion of inflammatory processes. these include upregulation of cyclooxygenase and nitric acid synthases. the il-1ri transmits a signal(s) through the cytoplasmic domain which is structurally related to the fruit fly toll protein. we have recently cloned the promotor most proximal to the 5' utr of the human il-1ri gene. the genomic sequences reveal a lack of tata and caat boxes but rather a considerable (75%) gc rich region. the translation of the type i il-1r appears under a significant suppression and may limit the biological activities of il-1 by low level of expression. on the other hand, the type ii il-1r, lacking a significant cytoplasmic domain, does not transmit a signal and acts as a decoy receptor preventing the binding to the type i receptor. there are several approaches to reducing il-1 activity; inhibition of il-1 converting enzyme which cleaves pro-il-l[3, anti-il-1ri, the il-i receptor antagonist (il-1ra) and soluble (extracellular) il-1ri and il-1rii. il-1ra is structurally related to il-1 and binds at 4°c to the il-1ri with near equal affinity as that of bona fide il-i; however, il-1ra does not transduce a signal. il-1ra has been given to humans in pharmacologic levels (mean plasma levels of 30gg/ml) without evidence of agonist activity. in addition, there has been no affect on normal homeostatic parameters, suggesting that il-i does not play a role in health. in healthy humans given intravenous endotoxin, il-ira reduced endotoxin-associated neutrophilia by 50% and completely reversed endoloxin-iuduced immunosuppression, il-tra is produced naturally and is elevated in the circulation in several diseases. in a variety of diseases, plasma levels of il-1ra are in 100-fold molar excess to those of il-i [3. it is unclear whether these endogenous levels of il-1ra are sufficient to block il-1 activity in disease. a single injection of ]l-6 or ifna in humans does not induce il-1 but rather high levels of il-1ra (10ng/ml). prof.dr.l.a. aarden interleukin 6 (il6) is a multifunctionai cytokine with a central role in host defense mechanisms and consequently in inflammation. il6 is thought to be involved in the pathogenesis of various diseases. therefore, specific inhibitors could have therapeutic value. a human-mouse chimeric monoclonal antibody to il6 has been applied in a phasei clinical trial in patients with multiple myeloma and in primate models of sepsis. no toxicity was observed and the most remarkable outcome of this study was the build up in the circulation of il6-anti-il6 complexes, suggesting that plasma il6 measurements represent a gross underestimation of il6 production in the patient. until now no natural antagonists of il6 have been described. the biological activities of il6 results from binding to a lowaffinity il6-receptor (il6r). the il6/il6r complex induces disulfide-linked homodimerization of the signal transducing receptor component, gp130. studies on the structure-function relation of il6 revealed that disruption of the gpl30-interaction site on il6 gives rise to a mutant il6 that, by virtue of its intact binding to the il6r, acts as a potent antagonist on human hepatocytes and on human b cells. in vivo studies using this molecule will be initiated in the near future. the evolution of infla~ation involves a dynamic series of cellular events controlled by specific signals which are important to the initiation, maintenance, and final resolution of the inflammatory response. the various phases of inflammation are influenced via the expression and subsequent regulation of a number of key mediators which play a predominant role during each particular stage of the developing lesion. for example, the initial elicitation of blood born leukocytes to on inflamed area is a complex system that is dependent upon the expression of early response cytokineso these proximal mediators, including both interleukin-i (il-i) and tumor necroisis factor (tnf), are important in the initiation phase by activating both immune and non-immune cells and establishing a series of cytokine networks, thus, the above proximal mediators can stimulate endothelial cells, epithelial cells, smooth muscle cells, and fibroblasts to generate more distal cytokines. these latter cytokines include interleukin-8 (il-8), macrophage inflammatory protein-i (mip-i), and monocyte cbemoattractant protein-i (mcp-i). the importance of the above chemukines can be found in the role they play in leukocyte elicitation, as well as wound repair. numerous investigations have shown the diverse cellular sources of il-8 and the ability of il-8 to elicit neutrophils in vitro at pm to nm concentrations, however, recent studies have sho%~ an additional role for il-8 during the resolution phase of the inflammatory process. using corneal pocket animal models of angiogenesis/neovascularization, il-8 was found to be a potent angiogenic factor at concentrations ranging from 2-40ng/ml. sequential fluorescein angiograms demonstrated that il-8 induced neovascularization by 14 days, which regressed by 6 weeks. in further analyses, both conditioned media recovered from either inflamed human heumatoid synovial tissue macrophages or lps-stimulated peripheral blood monocytes were found to possess potent angiogenic activity, which was effectively blocked by neutralizing antibodies directed against human il-8. in addition, an il-8 antisense nligomunclentide blocked the expression of a functionally active angiogenic factor from lps treated monocytes. the above data demonstrate that il-8 has multifumctional activities during the initiation, maintenance, and resolution of a local inflammatory response. inhibits specific t-cell responses to soluble protein antigens and alloantigens. the proliferative responses of th0, thl and th2 cd4 ÷ t-cell clones and cytokine production by these t-cell subsets are equally well inhibited. the inhibitory effects of il-10 are indirect and related to inhibition of antigen-presenting capacity and accessory function of the antigen-presenting cells (apc). however, il-10 also directly inhibits t-cell proli/eration induced by cross]inked anti-cd3 or anti-cd2 mabs (e.g. in the absence of professional apc), by specifically inhibiting il-2 gene transcription and il-2 protein production. il-10 also inhibits the production of the pro-inflammatory cytokines il-l-a, ~, il-6, and tnf-a and the chemokines il-8 and mip-i-u, which are strong chemo attractants for neutrophils and macrophage, respectively. in contrast, il-10 enhances the production of the il-1 receptor antagonist, a cytokine with anti-inflammatory activity. il-l0 produced by monocytes downregulates class ii mhc expression and il-10 production by these cells in an autoregulatory fashion. collectively, these in vitro data indicate that il-10 is a powerful suppressor factor for immune-and inflammatory responses and therefore may have potential clinical utility as an anti-inflammatory agent. this notion is supported by recent studies which indicated that il-10 also prevents lethal lps-induced toxic shock in mice. five of the chamekines (hip-i~,era~tes. ip-io and il 8) also chemoattraet t lymphocytes, while others act on mast cells. we have studied the effaces of these chemokines on t cell subsets. il 8 preferentially chemoattracts uns~imulated t cells. raises ~ttracte resting as well as activated cd4 and cd8 memory t cells. ip-10 chemoattracts only preactivated (not resting) cd4 or cd8 t cells. hip-l~ preferentially chemoattracts gd8 t calla, while hip-18 more readily attracts the cd4 t call subset. the 8 chemoklnes and ip-io were also sho~rn to promote the adherence of ~he same subsets of anti-cd3 activated t cells to ili activated human umbilical vein endothelial cells (hitvec). this was not associated with a~y i~crease in the number of lymphocyte adhesion molecules, bu~ could be inhibited by neutralizing monoclonal antibodies re lfa-i and vla-4. these chemokines also rapidly increased ~ha adhesion of resting t l~phoeytes to plates coated wi~h 8 integrlns (i-cam or v-cam) or matrix proteins such as f~bronectln. this induction of adhesion began wiehln 30', peaked by 1 hr and was completed in 3 hr. this suggests that chemoklnee rapldly increase the binding affinity of adhesion prote~ns on t cells. we also recently observed ~hat mcaf/mcpi and rantes chemoattract unatimulated mast calls. furthermore, igs actlvared mast calls were chemoattraeted by mcaf, ra~tes, pf4 and mip.i~. however. ~he chemoklnee did not induce mast cells ~o release histamines. presumably, chemokines as regulators of the adhesion, migration and homing of all kinds of leukocytes participate in many types of inflammatory diseases. natural killer cell stimulatory factor or interleukin-12 (il-12) is an heterodimerie cytokine formed by two covalently linked glycosylated chains of 35kd and 40 kd. il-12 was originally purified from the supermatant fluids of ebv-transformed human b ly~phobiastoid cell lines. however, in addition to b cells, phagocytic cells, i.e. monocytes, macrophages, and neutrophils, have been identified as the major physiological producers of il-12. phagocytic cells produce il-12 in response to bacteria, bacterial products such as lps, and intracellular parasites. the cell types on which il-12 exert biological activity are t and nk cells. the major direct biological function of il-12 on these cell types are the induction of cytokine production, primarily ifn-y, the enhancement of a generation of cytotoxic cells, and a mitogenic effect on antlgen-activated lymphocytes. in part thromgh its ability to induce ifn-y production, il-12, produced in response to infections, represents an important functional link between effector cells of innate resistance, phagocytic cells and nk cells, and effector cells of adaptiye resistance, t and b lymphocytes. the ability of il-12 to induce ifn-y production from circulating. nk cells and at least a proportion of t cells determines a rapid, antigen-independet production of ifn-y during infections. ifn-y acts as a potent enhancer of phagocytic and bacteriocidal activity of phagocytic cells, participating early in infection to activate some of the inflammatory mechanisms that represent the first innate resistance against infection. this antigem-independet activation of phagocytic cells that involves, in addition to il-12, other monocyte derived cytokines such as tnf and il-l, has been shown to be important in experimental animals for the resistance against infection by microorganisms such as limteria monocytogens and toxoplasma gondii. in addition to this role, early in infection in the antigen-independet phagocytic cell activation, il-12 has a major effect in the ensuing antigenspecific admptive immune response by facilitating the generation of t helper type 1 (th-i) response and suppressing a th-2 response. the presence of il-12 during antigen stimulation in vitro using human peripheral blood lymphocytes, or both in vivo and in vitro in experimental animals, induces generation of th-i cells producing ifn-y and il-2, and inhibits the generation of th-2 cells producing il-4. il-12 induces a direct differentiative effect on th-cells, priming them for high ifn-y production. ~is priming effect is stable and maintained even when t cell clones are cultured for extended periods in the absence of il-12, however, il-12 needs to be present during the first few days after stimulation with antigen or mitogen and established th clones are resistant to the priming effect of ifn-y. unlike the generation of high ifn-y producing clones, the il-12-mediated inhibition of il-4 producing cells in probably due to selective mechanisms, which may include a selective mitegenic effect of il-12 for thl cells and an inhibitory effect of ifn-y on th2 cell proliferation. in several experimental systems, it has been shown that the enhancing effect of il-12 on th-i responses is dependent on production of if~-y and on the presence of nk cells, possibly needed for the early production of ifn-y. the importance of phagocytic cells and nk cells in determining the characteristics of the th response during antigenic stimulation indicated that the mechanisms of innate resistance have a determining influence on the subsequent antigen-specific immune response: il-12 appears to have a key role in mediating the interaction between phagocytic cells. nk cells, and t lympocytes in these proccessem. trinchieri, g. 1993, interleukin-12 and its role in the generation of t e 1 cells, imm~ol. on peripheral blood monocyte/macrophages, il-13 behaves like il-4 in a variety of assays, and shows both similar and contrasting, activties with either il-10 or ifn-y. il-13 reduces inflammatory monokine and il-10 production. it reduces the pyrogen-induced expression of tissue factor (herbert et el, 1993, febs lett. 328, 268) . it mpregulates manmose receptor and ~c class ii expression, while reducing cdi4 expression. il-13 produces morphological changes (extensive cell processes, aggregation, giant cell formation) which have previously been observed with il-4. it inhibits niv-i production (montaner et el, 1993, i. exp. mad. 178, 743) , and interferes with hiv-induced cell fusion. the activities of il-13 and il-4 differ on t-ly•ocyte and large granular lymphocyte population. in particular il-13 does not exhibit the suppressive effects of il-4 on thl-type-helper functions (ifny production) and cytotoxic functions (perforin production). data will be presented showing that il-4 and il-l3 share a common receptor on a nu~er of cell types, but not on t-lympocytes (see also zmrawski st el. 1993, embo j.7. 2663-2670), explaining both thai common and divergent activities. the levels of il-13~a in activated peripheral blood lymphocytes are greater than or equivalent to those of il-4 mp~ja and the two ml{nas are expressed by different t lympocyte population: in particular, t8 cytotoxic lymphocytes express markedly higher levels of il-13~rna. in vivo, il-13 may thus be contributing, significantly to some of the activites previously ascribed to il-4 il-10. recently, we cloned the human cdna encoding interleukin-13 (il-13). human il-13 is a non-glycosylated protein of 132 aa with a molecular mass (mr) of 10 kd and has biological activities on monocytes and b cells, il-13, like il-4, strongly enhanced the expression of class ii mhc antigens on monocytes and induced expression of cd23 (fc~rii). furthermore, il-13, like il-4 and il~10, inhibited the production of pro-inflammatory cytokines il-1, ]l-6, tnfo~ and chemokines miplc~ and tl-8 by lps activated monocytes. both il-13 and il-4 enhanced the production of il-lra by monocytes. in contrast, il-13 lacked the t cell stimulating activities of il-4. the responses of monocytes to il-13 and il-4 were not additive or synergistic, supporting the hypothesis that il-13 and il-4 receptors are complex and share a common component which is important for signal transduction, taken together, these data indicate that il-13, il~4 and il-10 have important anti-inflammatory activities on human monocytes. in addition, these results indicate that il-13 is unique in that it can modify inflammatory reactions without stimulating (as does il-4) or inhibiting (as does il-10) t cell responses. transforming growth factor beta i (tgf-fll) belongs to a family of proteins that have potent immunoregulatory properties ranging ftom effects on the growth and dfflerentiafion of primitive stem cells to the differentiated functions of immune system effector cells. tgf-i~i is synthesized as a large secretory precursor polypeptide of 390 amino acids that is inactive until processed to a disulfide linked homodimeric molecule of 1t2 amino acids each. recently it has been found that tgf-[~i can have autocrine as well as paracrine effects on the immune system indicating that immune cells can activate the inactive secreted form of tgi~-~i. in addition, tgf-i~i has differential intraceuular effects on cell surface receptor modulation, tyrosine phosphorylation and cytokine gene transcription as well as cell mediated cytotoxicity. the systemic administration of tgf-131 has proven beneficial in a variety of animal models such as septic shock, allograft rejection and experimental forms of autoimmunity including collagen induced arthritis and experimental autoimmune encephalomyelitis. moreover the increased levels of tgf-ill and other tgf-13 isoforms found in several disease states associated with immunosuppression such as different forms of malignancy, chronic degenerative diseases and aids implicate the involvement of tgt~-i~ in the pathogenesis of some diseases. hopefully, well designed clinical trials will define the potential roles of the tgf-[3 family of proteins in the treatment of diseases of man. patient development of systemic inflammatory response syndrome (sis) after severe trauma is accompanied by a wide range of immune aberrations and altered cytokine production. in particular, t lymphocyte proliferation and ifn, t production is suppressed while monocyte (mo) tnfc~, il-6, pge2 and tgf~ production is massively increased concomitant to decreased antigen presenting capacity. recently, two new cytokines, il-10 and il-12, have been characterized as critical in regulation of mo tnfa and il-6, as well as in induction of t lymphocyte proliferation and ifn 7 production. in this study, peripheral blood leukocytes (pbl) from severe trauma patients (injury severity score >35) were analyzed for their il-10 levels, their in vitro proliferation to mitogen (pha) and their response after il-12 addition. since il-10 produced either by mo or by t lymphocytes can depress m~ antigen presenting capacity, inhibit t cell ifn,/production and directly diminish t cell proliferation, it might be suggested that immunosuppressed patients' mo and/or t lymphocytes would have increased il-10 levels. increased patient il-10 production might also be resulting from the high levels of tnfa a known stimulator of il-10. conversely, since il-12 augments mo antigenpresenting capacity, thl induction and proliferation, post-trauma leukocytes might be il-12 deficient. pbl of trauma patients were compared to normals' pbl, either unstimulated or ptta induced, and their levels of il-10 found to be dramatically and significantly reduced. patients' isolated m~, either stimulated with the bacterial cell wall analogue, mdp, or unstimulated, also had depressed il-10 production concomitant to elevated tnfa production when compared to normals' mo. mechanisms for the depressed patients' mo il-10 were explored. increases in tgf[3 may have partially contributed to the patients' depressed il-10 level, but elevated pge2 had no effect. addition of il-12 to patients' pbl significantly increased their mitogen responses. these data imply that sis is characterized by disruption in the interactions between mci and t lymphocytes so that patients' m~i produce excesses of some mediators (tnfa, il-6, pge2) and a dearth of other monokines (il-12, il-io). t lymphocytes are not activated and, therefore, unable to function in both immune defense and monocyte regulation. it is known that lge receptor-mediated or ca-ionophore-induced activation of mouse bone marrow-derived mast cells (13mmc) may result in the production of different cytokines including the interleukins (il) 3, 4, 5, and 6 as well as gm-csf and tnf-a. in the present study we analyzed the effects of exogeneously applied pro-inflammatory cytokines (il-1, 1l-6, tnf-c 0 as well as various mast cell growth factors (il-3, il-4, il-9, il-10, ngf, kl (kit ligand)) on cytokine production in primary mouse bmmc using a standard activation protocol (lxl06 bmmc/ml; ll.um ionomycin; 24-48h). the actixdties of bmmc supernatants were assessed in specific biological (il-3, il-4 il-6, 1l-9) and/or elisa assays (il-5, il-9). here we show that homogeneous populations of bmmc (>97%alcian blue+/safranln-; in vitro age: 4 weeks) generated in the presence of recombinant (r) rail-3 from normal balb/c mice produced modest amounts of 1l-6 and low or undetectable levels of il-3, -5, and -9 after induction with lp.m ionomycin only. however, a dramatic increase (5-to 10-fold) of these cytokine activities was noted, when in addition to ionomycin also human (11) rll-la was provided during the induction period. this il-1 effect was dose dependent with a maximgm at 2-10 u/ml hrll-la and specific, as pre-incubation (lh) of bmmc with 20 ng/ml hrll-1 receptor antagonist abolished the action of 2 u/ml hrll-lcc similar effects were noted with hrll-lg or rurll-lb (lng/ml, respectively), but not with rhll-6 or rmtnf-~. both mrll-10 and hrll-10 substantially enhanced ionomycin-induced 1l-6 production of bmmc in the absence or presence of il-1. il-10 significantly enhanced il-6 and il-9 production while decreasing il-3 activities to abont 50-90% of control levels, when il-i0 was provided in the presence of il-l/ionomycin. a monoclonal anti-nfil-t0 antibody (ascites 1:200) abrogated the effects of mrll-10. other mast cell-active cy~okines (]1,-3, il-4, 1l-9, ngf, or kl) added to ionomycia-or 1l-1/ionomycin-treated bmmc had no major effects on cytokine production. il-1 and il-i0 did not induce significant cytokine release in the absence of ionomycin suggesting tlmt cadependent signalling was required. at doses of 10"6m, dexamethasone, corticosterone, or hydrocortisone almost completely abolished ionomycin/il-1/ll-10induced cytokine production. the inducer cocktails per se did not interfere with the cytokine bio-assays. in case of il-9 inducibility of this cytokine in bmmc was confirmed at the mrna level by northern blot analysis. hence our data show that activated mast cells are a source of il-9 previously recognized as a product of th2type lymphocytes only. moreover, our study reveals novel functional roles for i-l-i, il-10, and ghicecorticoids in the regulation of cytoldne production in mast ceils. accumulating data suggests that cytokines, peptides involved in regulation of both physiological and pathological immunological responses, predominantly are produced at the local site of antigen stimulation. a new method was used to detect cytokine-producing cells in haman tissue at the protein level. single-cell production of 17 different httman cytokines, ilia, ill [3, illra, il2, il3, il4, il5, il6, ils, ill0, gm-csf, tnfa, ifn 7 and tgf[31.3, was identified by indirect immunohistochemical staining procedures and use of carefully selected cytokine-specific mab's. frozen sections were fixed with 4 % paraformaldehyde and permeabilized by 0.1% saponin treatment, eluting cholesterol from the membranes. the intracellular presence of all cytokines except ill, illra (late) and tfg[31_3, could be demonstrated by a characteristic perinuclear configuration in producer cells. in addition, the immunoreactivity extended over a large extracellular area encompassing the producer cell. a localization of the cytokine to the golgi-organelle was established by use of two culour staining including a haman golgi complex specific mab. this staining pattern was only evident in producer cells because injection of recombinant human cytgkines into the tissue caused a membraneous and extracellular staining pattern. both the extra-and the intracellular types of staining reaction could, however, be blocked by preincubating the cytokine specific mab with pure human interleukins. oxygen radicals (or) directly induce lipid peroxidation, indirectly they trigger adhesion and activation of pmn leukocytes. we investigated whether or also lead to a release of acute-phase response cytokins such as tnf-alpha, il-i beta or il-6 in whole blood cultures to maintain the induced inflammatory reaction. methods: blood samples from healthy volunteers (n=10) were incubated at 37°c. or were produced by the xanthine oxidase (xo)/ hypoxanthine (hx) system. after 0,30,60,120,240 and 360 minutes plasma levels of tnf-alpha, il-i beta and il-6 were determined with elisa kits. results: under the influence of or tnf-alpha plasma levels increased from 0,5 pg/ml at 0min to 14pg/ml, 236pg/ml, 343pg/ml after 60, 120 and 240 min. il-ibeta (0,3pg/ml, 0,6pg/ml, 1,5pg/ml, 81pg/ml and 169pg/ml after 0,60,120,240 and 360min) and il-6 (0,2pg/ml, l,lpg/ml, 4,1pg/ml, 58pg/ml and 72,9pg/ml after 0,60,120,240 and 360min) plasma levels were increased 60min later than tnf-alpha. summary: these data suggest that or do not only play an important role in initial accumulation and activation of pmn leukocytes but also lead to a stimulation of monocytes to produce the acute phase reaction cytokins tnf-alpha, il-i beta and il-6 to maintain and strengthen the inflammatory reaction. department of general surgery, steinhsvelstr.9, 89070 ulm, germany jan k. horn md, greg a. hamon md, robert h. mulloy md, greg chen bs, rebecca chow bs, and christof birkenmaier md. transforming growth factor-i~l (tgf-131) is released from inflammatory ceils following injury and in sepsis. in vitro experiments have confirmed that low concentrations of tgf-131 (0.1-1.0 ng/ml) are chemoattractive for monocytes, whereas higher levels of tgf-131 (>5.0 ng/ml) potentiate production of the immunedepressive prostaglandin e 2. other investigators have shown that tgf-]31 can cause the appearance of cd16 (fc immunoglobulin receptor) on monocytes exposed to 10 ng/ml of tgf-[~i for 24 hours. monocytes also express on their surface a glycoprotein that binds complexes of lipopolysaceharide (lps) and lpsbinding protein (lbp). such binding is associated with generation of proinflammatory cytokines such as tumor necrosis factor alpha. we have shown that cd14 is depressed in septic patients and therefore we hypothesized that tgf-131 could account for the down-regulation of cd14 observed in these individuals. we incubated normal human monocytes with platelet-derived tgf-[31 for 2 and 24 hours at 37°c and examined ceils for cd14 and cd16 expression using flow cytometry after immunnfluoreseent staining with appropriate monoclonal antibodies. monocytes were selected on the by usual criteria for size and granularity. non-viable ceils were excluded with the use of propidium iodide. two populations of monocytes could be found afcer incubation at 37°c alone. one displaying high density of cd14 had increased fluorescence over the homogeneous expression of cd14 in cells maintained at 4°c (baseline). the other population displayed decreased cd14 expression relative to the baseline cells. tgf-i~i (10-50 ng/ml) caused a shift of ceils from the high density into the low density cd14 population. this trend was observed within 2 hours of incubation and was complete by 24 hours. we observed a net decrease in cd14 expression 0f32% for all subjects studied (p<0.05 vs controls). phorbol myristate acetate (100 ng/ml) also caused down-regulation of cd14 to a similar degree as tfg-i~i. we also confirmed that monocytes could be induced to express cd16 after incubation with tgf-131 (10 ng/ml) for 24 hours. these studies demonstrate that monocytes incubated with immunodepressive levels of regulation of cd14 by tgf-131 deplete their surface expression of cd14 while generating cd16. this down-regulation of cd14 by tgf-131 correlates with our clinical observations of lower cd14 expression on monocytes obtained from septic patients. for over 25 years, activated t lymphocytes have been considered to be the cellular source of mif. we recently isolated and cloned the murine homolog of mif after identifying the specific secretion of this protein by lpsstimulated pituitary cells in vitro and in vivo. however, further experiments showed that mif protein is detectable both in t-cell deficient (nude) and hypophyseetomized mice, suggesting that yet additional cell types may produce mif in vivo. since monocytes/macrophages are a major source of the cytokines that appear in response to lps administration, we examined the possibility that mif also is expressed in cells of the monocyte/macrophage lineage. we found that mif is expressed constitutively in the murine macrophage-line raw264.7 and in thioglycollate-elicited peritoneal macrophages. significant amounts of mif mrna (rt-pcr) and protein (western blotting) were observed in cell lysates. in raw 264.7 cells, mif secretion was induced by as little as 10 pg/ml of lps (e.coli 011 l:b4), peaked at 1 ng/ml, but was not detectable at lps concentrations >1 txg/ml. similar data were obtained with elicited macrophages, but higher lps concentrations were required, unless the cells had been preincubated with ifn 7. production of mif by lps-stimulated (l ng/ml) macrophages peaked at 12 hr. expression ofmif mrna and tnf mrna by lps-stimulated raw 264.7 macrophages was investigated by rt-pcr. as expected tnf mrna expression increased over the range of lps concentrations (1 pg/ml to 1 p_g/ml). in contrast, levels of mif mrna correlated inversely with lps concentration. by competitive pcr, mif mrna was observed to increase approximately 2-fold after lps induction (100 pg/ml). mif secretion also was induced by tnfoc (1 ng/ml) and ifn? (10 iu/ml), but not by il-113 and il-6 (up to 10 ng/ml). lps and ifn 7 had additive effects in inducing mif secretion. in separate experiments, macrophages stimulated with recombinant mouse mif (1 gg/ml) were found to secrete bioactive tnf~ (>750 pg/ml by l929 cytotoxicity). we conclude that the macrophage is an important albeit overlooked cellular source of mif in vivo. mif secretion is induced by lps, tnfc~ and ifn?. mif also stimulates macrophages to secrete tnf. taken together with previous observations that anti-mif antibody protects against lethal endotoxemia, these data implicate mif as a critical mediator of inflammation and septic shock. inflammation is characterized by an exacerbation of proinflammatory cytokine production. cytokines such as il-4, il-10, and tgf8, have been identified as anti-inflammatory mediators thanks to their ability to down regulate the production of il-1, il-6, il-8, tnfc~ by activated monocytes / macrophages. however, other cells, including polymorphonuclear cells (pmn) do contribute to the release of pro-inflammatory cytokines. we investigated the capacity of the so-called anti-inflammatory cytokines to control the release of il-8 by activated neutrophils. human pmn were purified following glucose-dextran sedimentation and ficoli-hypaque centrifugation. the cells were cultured at 37°c for 24h in the absence or presence of lipopolysaccharide (lps) or tnfa. il-8 release was measured in the supernatants using a specific elisa. among tested cytokines, il-10 was the most efficient inhibitor of il-8 production by lps-activated pmn. il-4 was also active, whereas no down regulation was noticed with tgfp~i. when tnfa was used as a triggering agent, none of the cytokine could prevent il-8 production. northern analysis are under investigation to precise the level of the il-4-and il-10-induced inhibition of il-8 production by pmn. our data illustrate that il-10 and il-4 possess the capacity to down regulate the production of il-8 by both monocytes and pmn, whereas tgfb has a more limited inhibitory activity. ciliary neurotrophic factor (cntf), a member of the il-6 superfamily, has recently been shown to promote axonal growth and neuronal healing. cntf production is also increased during neuronal and muscle damage, associated with soft tissue injury or trauma. we postulated that production of cntf may explain the loss of skeletal muscm protein that occurs in inflammation. 40 female, wistar (175-200 gm) rats received either 250 or 25 pg/kg bw s.c. injections of recombinant rat cntf for seven days, or received sham injections and were freely-fed. additional animals were pretreated with 10 mg/kg ibuprofen lp prior to 250 pg/kg bw cntf. rats treated with 250 ,ug/kg bw cntf lost 6.8_+0.4 gms bw as compared to freely-fed controls which gained 17.0_+3.1 gms (p<o.001 by anova and lsd), and ibuprofen treatment had no effect on cntf-induced weight loss (-5.8_+0.7). animals pair fed to the high dose cntf gained body weight (10.2-+3.5), although weight gain was less than seen in freely-fed controls. rats treated with 25 pg/kg bw had reduced body weight gain (+7.8+_4.4 gin). food intake was also reduced by 29% in 250 pg/kg cntf (from 23.7+_0.7 gm/day to 16.8_+0.6 gm/day; p<o.05) and was also unaffected by prior ibuprofen treatment (18.4+-0.6 gm/day). despite the loss in body weight noted in the high dose cntf animals, there was no appreciable hepatic acute phase response, since liver weight (8.6+0,1 gms vs. 9.4_+0.3 gins), total protein (54.5+-2.9 gms/l vs 44.8+-3.6 gins/l) and albumin (29.0+1.5 gms/l vs. 26.0+_1.6 gms/l) were not different between cntf and freely fed animals. we conclude that cntf causes anorexia, and increases body weight loss in excess of the associated anorexia. unlike the cachexia associated with il-1 and tnf infusions, which is associated with an acute phase response, cntf acts predominantly on food intake and carcass weight through a prostaglandin-independent mechanism and has only minimal acute phase inducing properties, n. joseph espat, md. university of florida, department of surgery, j-box 100286, gainesville, fl. 32610. of cytokines by neurotrophic factors, the neuroendocrine system and phenotiazines. cytokines, including tnf and il-1, have a pathogenetic role in various diseases related to infection and inflammation. the action and/or production of cytokines can be regulated by drugs or endogenous mechanisms that involve the cetral nervous system (cns) we found that the antipsychotic chlorpromazine (cpz) potently inhibits tnf production and protects mice against endotoxic shock. cpz also inhibits brain tnf production in mice intracerebrally injected with endotoxin, whereas cpz analogs that do not cross the blood-brain barrier inhibit only perypheral, not brain, tnf. tnf production and susceptibility to the toxicity of endotoxin, tnf and il-1 are increased in adrenalectomized mice, indicating that endogenous corticosterone (cs) controls cytokine production and toxicity, in a feedback fashion since endotoxin and cytokines increase serum cs. increased tnf production was also observed in mice where the hypothalamus-pituitary-adrenal axis was blocked by chronic administration of dexamethasone, following a two-day washout. administration of cytokines acting through the gp130 signal transd,uction protein, including il-6 and ciliary neurotrophic factor, cntf potentiated il-l-induced elevation of cs. il-6 and cntf also induced hepatic acute-phase proteins. thus il-6 and cntf might have antiinflammatory activity, by potentiating the feedback responses of the neuroendocrine axis. these data indicate how complex can be the interregulatory relationships between the brain and the immune system, how they can be used to pharmacomodulate cytokine action and how their disregulation might exacerbate il-1-and tnf-mediated pathologies. lipopolysaccharide (lps) constitutes a well established activator of monocytes/ macrophages (mizi) and murine b lymphocytes. the hydrophobic part of lps, termed lipid a, represents the endotoxic principle of lps. we now demonstrate that lps is also capable of inducing proliferation of human t cells at a dose of 10 to 1000 pg/ml. the specificity of this stimulation was analysed by the following experiments: i) the synthetic hexaacyl escherichia coiltype lipid a (compound 506) also stimulated human t cells; ii) the synthetic tetraacyl lipid a precursor (compound 406) or the 1phosphono-oxyethyl analogue pe-4, structures known to antagonize lps-induced monokine production in human mz, also inhibited lpsinduced t-cell proliferation. the t-cell proliferation expressed the following features: i) cd4 ÷-as welt as cd8 ÷ t cells proliferate in response to lps, ii) limiting dilution analyses reveal that the frequency of responding cells was between 1/500 to 1/1000 cells; iii) purified t cells alone cannot be stimulated by lps, but need the help of monocytes. this help cannot be replaced by b cells or fixed monocytes. furthermore, for activation a direct cell-to-cell contact between t cells and monocyte is neccessary; iv) t cells stimulated with lps express mrna for il-2 and ifnf, but not for il-4 or il-5 (determined by pcr). in supernatants, ifnf was detectable (elisa); v) lps-activated cd4 ÷ as well as cd8 + t cells express cd25, the high affinity il-2 receptor. our results indicate that in contrast to previous reports human t cells respond to lps with il-2 receptor expression, lymphokine production, and proliferation. the reactive cells appear to belong to the t,1 cell type, the finding that human t cells can be activated by lps is of important relevance for the understanding of the pathomechanisms of infections by gramnegative bacteria. this may lead to new strategies for the therapy of sepsis. we have recently shown that human eosinophils produce mrna for interleukin (il)-8 when stimulated with calcium ionophore (eur. j. immunol. 23 (1993), 956). we now present evidence that human eosinophils contain preformed il-8 protein although they do not express mrna for il-8 as measured by rt-pcr. il-8 protein expression was determined using specific anti-human il-8 monoclonal antibodies by western blotting, facs analysis and immunohistochemistry. moreover, preformed il-8 was re/eased into supernatant by 99% pure eosinophils after priming with gm-csf or il-5 and subsequent 25-min stimulation with paf, rantes, ionomycin or pma, however not with il-1 or il-2, as measured by elisa. this observation implies that activation of protein kinase c and/or intracellular calcium mobilization are necessary events for il-8 release in human eosinophils. the determined amounts of preformed il-8 protein was higher in patients with asthma compared to normal individuals suggesting a role for eosinonhi! derived il-8 in asthma. since the eosinopnit is the ,,r~:=z~mmant cell in the asthmatic airways, we determined il-8 concentrations in bronchoaiveolar lavage (bal. ~ids from normal !i~.~ijvtduals and asthmatic patients. indeed, il-8 concentrations in bals from both groups were similar to those seen in the supernatants released by activated eosinophils. the role for il-8 in asthma remains to be determined. based on a well established rat model named activity-induced ulceration (asu) or activity based anorexia (aba), we have developed a rat model for chronic stress. male rats were exposed to a feeding schedule in order to reduce body weight by 20-30% within 14 days. while one group of rats was kept sedentary, the other had access to a running wheel. food restricted rats allowed to exercise developed highly increased activity as compared to ad libitum fed rats in wheels (ca. 15 versus 2 kin/day). they developed elevated plasma corticosterone levels (30_+5gg/dl) at their circadian peak, increased adrenal weight and decreased thymus and spleen weight as compared to control rats and weight matched sedentary rats, establishing this paradigm as a model of chronic stress. no differences in plasma acth were seen among the various groups, suggesting increased sensitivity of the adrenal glands to acth. the effect of chronic stress accompanied by hypercorticism on baseline and endotoxin-induced cytokine levels was studied in this model. under baseline conditions,splenic il-1 a was suppressed in both food-restricted groups. this was more pronounced in rats with access to a running wheel. il-6 and tnf activity in plasma was not affected. after administration of 1001.tg/kg lipopolysaccharide (lps), tnf activity in plasma was suppressed by food restriction but there were no differences between sedentary or exercising rats. in addition, no differences were found among groups for corticosterone, spleen il-lc~ and plasma il-6 activity. we conclude, that the food restriction-induced hyperactive rat model is a useful model for studies on the effects of chronic stress. in this model, under basal conditions il-le~ in tissues may be subject to downregulation by glucocorticoids, whereas after lps-stimulation of cytokine production only tnf activity is regulated by fast acting feedback loop between cytokines and the hypothalamo-pituitary-adrenal axis (hpa-axis). this is supported by strong correlations between plasma tnf activity and corticosterone in stressed rats. cytokine antagonists modulate cytokine actions and it appears that a balance between production of cytokines and cytokine antagonists is important to control of host responses. recent observations show that during myocardial infarction there is an increase in circulating cytokines such as tumor necrosis factor (tnf) and interleukin 6 (il-6). we tested the idea that compensatory increases in cytokine antagonists likewise occur and investigated changes in plasma concentrations of tnf and interleukin i (il-1) and those of their specific antagonists interleukin-1 receptor antagonist (il-1 ra) and soluble tumor necrosis factor receptor type i (s tnf r-i). we also measured plasma concentrations of ~-melanocyte-stimulating hormone (~-msh), an endogenous neuropeptide that occurs in pituitary, brain, skin, gut and other tissues. when administered to laboratory animals, this peptide has potent antiinflammatory and antipyretic activity, perhaps by mimicking an endogenous modulatory influence on cytokine actions. samples were obtained from 20 patients with acute myocardial infarction at presentation in the coronary unit, every three hours for 24 hours, and daily thereafter until discharge. we found elevations in the plasma cytokines il-1 and tnf only in a proportion of subjects whereas cytokine antagonists c~-msh, il-lra, and stnfr were elevated in all patients, o~-msh was elevated at presentation but it decreased progressively in successive tests, reaching normal values within 24 hr whereas stnfr and 1l-lra peaked at 3-6 hr or later. there were significant correlations between plasma concentrations of cytokine antagonists and enzymes released from injured myocardium, including creatine kinase (ck), aspartate serum transferase (ast), and lactate dehydrogenase (ldh). the data show that cytokine antagonists increase in the circulation of patients with acute myocardial infarction likely to modulate prninflammatory effects of cytokines. objects of the study: interleukin-6 (il-6) is a multifunctionai cytokine known to be involved in important homeostatic processes. il-6 levels are increased in many pathological situations, and correlates with disease activity and patient outcome. to exert its effect il-6 binds to its receptor on the membrane of its target cell. this receptor is also present in a soluble form in plasma and in urine. in this study we quantitatively measured the availability of soluble il-6 receptor (sll-6r) in biological fluids in healthy volunteers. materials and methods: blood, urine, cerebrospinal fluid (csf), and synovial fluid (sf) are obtained from healthy volunteers, sil-6r and il-6 are measured using elisa's. both elisa's are tested for interference of sil-6r mid il-6. the effect of sll-6r in the il-6 bio-assay (b9) is tested. results: baseline levels are (mean + sd): 76.6 -+ 19.3 ng/ml in serum, 3.7 -+ 1.3 ng/ml in urine, 1.64 _+ 0.4 ng/ml in csf and 11.6 + 3.3 ng/ml in sf. there is no interference of sll-6r in the il-6 elisa or bio-assay and no interference of 1l-6 in the sil-6r elisa. conclusions: in all investigated biological fluids sll-6r can be found. these data provide baseline values for investigations concerning pathological situations. both elisa's described are useful tools to do these investigations. trauma-associated immune aberrations coincide with augmented release of immunoregulatory and proinflammatory cytokines. the present study examines the effect of thermal trauma on the capacity for specific (receptormediated) response of lymphoid cells to interleukin 1 (il 1) and to turnout necrosis factor ~x (tnfc0. methods. bum patients (n=24, 25->75 % total body surface area) were studied weekly up to 42 days post-injury. the limulus amoebocyte lysate (lal) test was used to measure plasma endotoxin levels. the percentage of il 1~-and tnfcz-binding t(cd 5) lymphocytes was assessed by flow cytometry analysis. levels of il 1 receptor antagonist (il lra) in patients' plasma and cultures of peripheral blood ceils (pbc) were determined by immunoassay. results. plasma endotoxin concentrations were significantly (p<0.05) increased up to 3 weeks post-bum (means 3.5+1 in non-surviving and 2.6+ 0.6 u/ml in surviving patients vs < 1u/ml in the control). within 2 weeks of bum, the percentage oft ceils expressing receptors for tnfa and il 1[~ constitutively was elevated (by 10-15 fold). in contrast, the capacity for de novo receptor expression by activated pbc was reduced. serum levels of il ira were significantly increased (range 2.5-40x10 j pg/ml vs <0.2x10 j pg/ml in the control). in all patients, high concentrations of il lm were released spontaneously in unstimulated cultures of adherent ceils (range 20-30x10 -3 pg/ml vs 4-8x10j pg/ml in the control). however, its secretion was decreased in lps-stimulated parallel preparations. conclusions. in the bum patient, susceptibility to the immunoregulatory effect of tnfcz and tl 1~ may be modulated by infection-related products. alterations in the capacity for receptor expression and secretion of 1l lra may affect il 1-regulated biological responses including specific immune reactions. while studies suggest that il-10 is an important lymphokine involved in cell-mediated immunity, little is known about this mediator's role in hem-induced immunesuppression. our aims, therefore, were to determine: i) if il-10 contributes to depressed t-cell responses seen following hem; and 2) how other agents, known to play a role in hem, effect il-10 release. to study this, c3h/hen mice were bled to and maintained at a map of 35 mmhg for 1 h and then adequately resuscitated. mice were killed 2 h post-hem to obtain splenic t-cells (nylon-wool purified). il-10's immunosuppressant role was demonstrated by the ability of monoclenal antibody (mab) to il-10 to markedly improve the t-cell proliferative response [2.5 #g the marked increase in capacity of t-cells from hem mice to produce il-10 was significantly reduced by treatment with either ibu or mabs. since ibu, tgf-~, as well as il-6 are all reported to directly/indirectly influence prostanoid synthesis, this implies that eicosanoids play a major role in inducing il-10 release by t-cells following hem which depresses t-cell function. the mechanisms underlying immunosuppression induced by thermal injury and alcohol ingestion are in part due to cytokine dysregulatinn. il-10 down-regulates production of eytokines by maerophages and may be an important regulator of the initiation of the immune response. il-10 has also been demonstrated to inhibit the production of no by macrophages. this study examined the alterations in eytokine production and effect of inhibition of no production on immunologic function in a routine thermal injury model. methods: balb/c mice (n=40) were randomized to 4 groups: saline-sham(ns-sham), alcohol-sham(etoh-sham), ns-bum, etoh-bum. animals received 20% etoh or ns daily for 14 days by gavage. a 30% full thickness bum was induced 4 hrs after the last dose of etoh or ns. animals were resuscitated, then sacrificed 4 days post bum. splenic lymphocytes were cultured for 5 days with lps, and lps with two concentrations of n-monomethyl-l-arginine, a nitric oxide inhibitor (l-nmma 2.5ug/ml, 10ug/ml). splenocyte production of il-10, interferon-gamma, il-2, pge2 were measured, and lymphocyte proliferative response examined. results: il-10 production was significantly suppressed in thermal injury. exogenous l-nmma normalized the suppression of 11.-10 in a dose-dependent manner, indicating nitric oxide may modulate il-10 and interferon-gamma production in thermal injury. il-10 production is normal in etoh-burn animals. conclusion: il-10 and interferon-gamma production is altered in this murine thermal injury model, and may contribute to this injury-induced immunosuppression. inhibition of no synthesis normalizes il-10 production and should be investigated further as an immanomodalator in thermal injury. surgery, infection and inflammation results in the production of pro-inflammatory cytokines which mediate metabolic and immunologic host responses. the aim of this study was to characterise the elaboration of cytokine release following a variety of surgical procedures. twenty one patients undergoing elective intermediate, hip, knee and major gastrointestinal surgery were studied. levels of interleukin-1 (i1-1), interleukin-6 (i1-6), the interleukin-1 receptor antagonist (i1-1ra) and the acute phase c-reactive protein (crp) were measured in bloods drawn 0, 1, 2, 4, 6, 12, 24 and 48 hours following operation. a portion of the results are shown (mean -+ sem). 135+37 1738-+843 145_+24 one and two factor anova; *p<0.0002, #p<0.0001, §p<0.0009, ¶p<0.0014, for differences between groups i1-1 was not detected at any time point. both ii-ira and i1-6 increased after surgery. maximum responses occurred following major git and hip surgery, minimal responses were seen after intermediate and knee surgery. ii-ira levels increased within two hours and remained elevated for 24 hours; the b-ira increase was a thousand fold greater than the rise in i1-6 levels. i1-6 levels increased up to 24 hours after surgery. crp levels reflected maximum ii-ira and i1-6 levels (r2=.676, p<0.0001 and r2=.282, p<0.04 respectively). high ii-1ra and i1-6 levels reflect major surgery, however the ii-ira response is more rapid and of greater magnitude. the strong i1-1ra correlation with crp may indicate that this regulatory cytokine is itself a mediator of host responses to surgery. dept. of surgery, meath/adelaide hospitals, heytesbury st., dublin 8, ireland. change of il-6 and soluble il-6 receptor levels after surgery s. hisano, k. sakamoto, s. mita, t. ishiko, m. ogawa [objectives] under surgical stress, il-6 plays a main role in producing acute phase proteins and contributes to host defense mechanism. soluble il-6 receptor (sll-6r) is considered to be agonistic to il-6, unlike other soluble type receptors of cytokines. here we measured il-6 and sll-6r levels in the serum and drain fluid from surgical field in order to investigate the changes of il-6 and sll-6r after surgery and their origins. [materials and methods] serum and drain fluid samples from 21 cases (6 of esophagectomy and 15 of gastrectomy ) were serially collected before and after surgery. il-6 and sll-6r levels were measured by elisa. [results] (1) serum il-6 : all cases reached the maximum level on pod-l, more precisely 3-6 hours after operation. (2) il-6 in the drain : maximal il-6 levels in the drain were recognized 3-6 hours after operation, at almost the same time as serum il-6. furthermore the il-6 values in the drain were much higher, about 100 times, than those in serum. (3) sll-6r in the serum : all cases reached minimum levels 3-12 hours after operation and recovered to the preoperative levels a few days later (decrease ratio : 43.9+5.6~,, range : 9-84~'). (4) sll-6r in the drain : sll-6r levels in the drain showed almost the same value and change as serum sll-6r. [conclusions] (1) il-6 is produced from the cells gathering around operative fields whereas sll-6r is considered to be produced in the cells which do not gather around the operative fields. (2) there may be a mechanism that down-regulates sll-6r in the early stage of surgery. [objectives] il-6 plays an important role in host defense in the early stage after surgery. in the present study, we examined changes in il-6 concentration after major thoracoabdominal surgery and elucidated the effect of surgical trauma and factors influencing postoperative elevation of serum il-6. [materials and methods] thirty-eight patients undergoing elective surgery of the thoracoabdomen were classified into 6 groups according to the location of the operation. bloods and drain fluids were serially obtained and samples were frozen until measured, keukocytes were simultaneously collected for northern blot analysis. concentration of il-6 was measured by elisa and il-6 mrna was detected by northern blotting after total rna was extracted by the acid guanidium phenol chloroform method. [results] (1) serum il-6 levels reached the maximum concentration on the 1st postoperative day in all patients. (2) the il-6 peak was significantly correlated with surgical trauma as defined by the operation length and the volume of blood loss during operation (r=0.554, p<0.01, r=0.427, p<0.01, respectively). (3) the peak concentration of serum il-6 in patients undergoing esophagectomy was significantly higher than in those undergoing pancreaticoduodenectomy (p<0.05), despite a similar degree of surgical trauma. (4) peak 1l-6 concentration observed in a patient who underwent esophagectomy was about 100 fold greater in the drain fluid of thorax than in the peripheral blood. (5) il-6 mrna was demonstrated in leukocytes from thoracic and abdominal exudate at 6, 24 and 48 hours after surgery. in contrast, il-6 mrna could not be detected in leukocytes from the peripheral blood. [conclusion] il-6 is mainly produced in the operative field and subsequently enter the peripheral blood to induce cytokinemia. the operation length, volume of blood loss and thoracotomy are factors influencing the concentration of cytokine in the blood. zaragoza spain age may be an important factor influencing the function of immunocompeteut cells releasing cytokines after both accidental and surgical trauma the aim of the present paper is to ascertain if patients (pts) over 60 years old show a different serum level cytokine pattern than pts under 60 after a standard surgical procedure considered as a "medium strength trauma". patients and methods: 33 pts(22 females 11 males)with gallstone disease were perspectively studied, pts were allotted in two groups: gr.a:17 pts under 60 years(mean age:49.5+-7)gr.b:16 pts over 60 years(mean age:65.5_+5). all pts underwent cholecystectomy and cholangiography. 6 pts in gr.a and 7 pts in gr. b underwent common duct exploration. 1 spbintercctomy was performed in each group. on the day of surgery (pre) and on the 1st and 7th postoperative day(leo, 7po) : percentages of cd19, cd5, cd4, cd8 and cd16 cells we measured by means of flow cytometry using moab. and levels of il-1, il-4, il-6 and tnf "in vivo" by elisa using moab. results: ere: cd16% was 5.3_+3 in gr.a and 7. 5 objectives of the study. after surgery for esophageal cancer multiple organ damage has been reported to be caused by polymorphonuclear leukocyte (pmn)-mediated injury. we measured serum granulocyte colony-stimulating factor (g-csf) and interleukin 8 (il-8) levels to determine a role of g-csf and il-8 in pmn function after surgery for esophageal cancer. materials and methods. peripheral pmn counts, peripheral pmn chemiluminescence, serum g-csf levels, and serum il-8 levels were measured before and after surgery in 12 patients with esophageal cancer (ec), and 12 patients of gastric cancer (gc). esophagectomy with thoracotomy and laparotomy were performed for patients with ec, while subtotal gastrectomy with laparotomy were performed for patients with gc. results. peripheral pmn counts (p<0.01) and peripheral pmn chemiluminescence (p<0.05) of patients with ec were significantly decreased compared to those of patients with gc at 4 and 8 hours after surgery. serum g-csf levels of patients with ec were significantly (p<0.01) increased compared to those of patients with gc at 4 and 8 hours after surgery. serum il-8 levels of patients with ec were significantly (p<0.01) increased compared to those of patients with gc at 4, 8 and 24 hours after surgery. significant inverse correlations (p<0.0l) between peripheral pmn count and serum g-csf and il-8 levels were seen at 4 hours after surgery. conclusion. these results suggest that many circulating pmns, which are excessively activated by g-csf and il-8, may adhere to the endotherial cells and then migrate into the tissues, and cause multiple organ damage after surgery for esophageal cancer. immunnogical changes in patients with severe brain trauma receive increasing attention since morbidity and mortality ere still high. interleukin-6 (il-6) was previously detected in the cerebrospinal fluid (csf) during different pathologies of the nervous system (1, 2, 3). in our study we monitored il-6 and nerve growth factor (ngf) production in the csf after human brain trauma. since astrocytes within the brain constitute one of the major cell type contributing to the inflammatory response through the release of cytokines and other factors after injury, we investigated the functional relationship of il-6 and ngf on a single cell niveau using cultured astrocytes. methods csf was obtained from patients with severe brain injury (glasgow coma score (gcs) <8 and ct abnormatities or gcs <8 over 6 hours) after implantation of intraventricular icp monitoring device for therapeutic purpose and collected over 24 hours csf and serum. il-6 and ngf were assayed by elisa. astrocytes were isolated from neonatal mouse brain as described (4) . ngf production by cultured astrocytes was measured by elisa in the presence of csf, il-6 and il-6 antibody. astrocyte migration was tested in a chemstaxis chamber. results 17 head trauma patients were included in this study (approved by the university hospital medical ethics board) and the csf was obtained through intraventricular catheters. high levels of il-6 were detected in the csf of these patients when compared to serum during the first days after brain trauma. furthermore ngf could be found inside the intracerebral compartment. csf containing high levels of il-6 could stimulate ngf production in cultured astrocytes. this effect could be [nhibited partially by il-6 antibodies, purified il-6 exposed to cultured astrocytes in vitro, stimulated the migratory activity of these cells in a dose response fashion. il-6 was found in the csf of brain injured patients, suggesting a role for this cytokine in the pathophysiology of brain injury. since astrocytes are involved in maintaining the homeostasis of the brain, we further investigated the possible role o1 il-6 on astrocyte functions, il-6 promoted ngf production in vivo and in vitro, thus contributing to neuronal cell survival and regeneration. furthermore il-6 stimulated astrocyte migration in a dose response fashion, potentially contributing to astrocytosis following brain injury and inflammation, these results show that il-6 represents a key cytokine in traumatic human brain injury with possible systemic effects, which are at preserlt under investigation. we studied a) the role of tnf and b) the therapeutic effect of a mab to tnf with regard to haemorrhagic shock (hs) related ,pathophysiologic alterations and mortality in rats. method: a prolonged hs was induced by bleeding to a blood pressure of 30-35 mmhg for 180 pin followed by reinfusion of shed blood (sb) and resuscitation with two times of sb volume of ringer's lactate over 50 rain. 15 animals received a bolus dose (20 mg/kg) of tnf mab (celltech, berkshire, uk) at 15 min after resuscitation (tn3). the control group (n = 15) was treated similar to the tn3 group but received ringer's lactate (con). results: at 180 min the prolonged hs resulted in a metabolic acidosis indicated by a significant decrease of blood ph (7.16 + 0.04), hco3-(16.3 ___ 2.0 mm), and base excess (-12.5 + 1.9 ram) values with pco2 (47.9 + 7.7 mmhg) and po2 (136.2 + 20.1 mmhg) in the tn3 with no difference to the con group. immediately after resuscitation (230 min) plasma endotoxin levels were found to be increased in both groups (48.8 + 73.3 in tn3 vs 30.1 _ 25.7 pg/ml in con group) . prior to the treatment with tnf mab (230 min) there was also no difference between plasma tnf levels of the two groups (42.1 + 60.7 in tn3 vs 60 + 141.8 pg/ml in con group). treatment with the tnf mab at 65 rain post-hs improved the 48hour survival rate to 73.3 % as compared to 26.7 % in the control group. macropathologic evaluations revealed frequency of intestinal bleeding in oniy 2 animals in the tn3 vs 11 in the con group. no bleeding in the kidneys was found in the tn3 but in 4 rats in the con group. the significant increase in lung wet weight observed in non-survivors in the con (n = 11) was prevented in animals which died in the tn3 (n = 4) group ((9.3 +_ 2.4 vs 6.3 +_ 1.0 g/kg). conclusion: our data suggest that tnf formation induced by hs in rats is an important mediator for pathophysiologic alterations leading to multi organ failure and lethality. antibodies to tnf might be a useful agent in the treatment of haemorrhagic shock related disorders. 55-+8 n=ll*$ 15-+5 n=7 78_+22 n=5* * p<0.05 vs baseline :~p<0.05 no anesthesia vs anesthesia thus 1) tnf production increased 2-3 fold by 48-72 hrs following trauma in unstimulated blood, but was reduced or not changed after lps stimulation, so circulating leukocytes are probably not an important source of tnf post trauma; 2) anticd18 had no obvious effect on tnf production in unstimulated or lps stimulated blood, relative to vehicle, which suggests that the protective mechanism of anticd18 does not involve tnf suppression; 3) fentanyl anesthesia at 72 hrs following trauma unexpectedly decreased lps-evoked tnf production, which suggests that anesthesia alone can influence an inflammatory response. proinflamrnato~ cytokines have been shown to play a signific~t role in the pathogenesis of sepsis, which is a very common occurrence in born injury. tnfa is infrequently detected in the blood of burned patients, the ability to detect the shed receptors of stnfg has not been determined. serial serum mmples from burn patients were collected from the time of admission until death from septic shock. these samples were analyzed using an enzyme-linked immunosorbent assay (elisa) for stnfr, l-ira, tnf-a, and il-ib. the patients ranged in age from 26 to 72 yeas of age. the percentages of bum ranged from 40% -90%. cytokine concenlrntions vmled from patient to padent irrespective of bum size. tnfa levels were consistentiy in the range of30pgjml -150pg/ml. peaks in the tnfa values were above 90pg/ml and were also associated with a peak in the stnfr levels. these levels began at <10,000pghnl within the in,st 6 ins of injury and gradually increased with time. clinically. ti~ appearance of eytoklnes was independent of positive wound, blood, or respiratory cultures however peak values in tnfa and stnfr were ~ialed with a fluid requirnmenl levels of il-i ra were also elevated independent of clinical findings as well as extent of injury. in pl5 there is a significant corresponding peak in il-trn (>40~8/ml) at the same time as t/~:a and stnfr levels. we aimed to characterise the pattern of secretion of interleukin-1 beta 0l-ii3), intefleukin-6 (il-6) and tumour necrosis factor alpha (tnfa) in multiply injured patients and to relate these results to their clinical condition and outcome. two hourly blood samples were taken from ten patients from the time of injury until 48 hours. cytokine levels were measured using sandwich enzyme-linked immunosorbent assays (elisas). injury severity scores (iss) were calculated and haemorrhage was assessed from the blood transfusion requirement over the 48 hours. patients' ages ranged from 17 to 86 years. iss varied from 9 to 50 and transfusion requirement from 0 to 14 units. five patients died after the study period. ]1,-6 was raised in 9/10 patients (max level 12,500 pg/ml) but was unrelated to condition or outcome. 5/8 showed a rise in il-1b (max level 90 pg/ml) which was negatively correlated to iss (i=-0.789, p<0.02). tnfa was raised in 2/10 (max level 95 pg/ml). peak tnfc~ was positively correlated with iss (1=0.676, p<0.05) and haemorrhage (i=0.602 but p<0.5). il-ib and tnfa production was mutually exclusive. there was no common cytokine profile for these patients. unlike elective surgery there was no correlation between peak 11,-6 and severity of injury: tissue damage may not be the stimulus for the cytokine response to multiple injury. periods of ischemia or hypoxia produce endothelial damage in peripheral organs. tumor necrosis factor-alpha (tnf) plays a central role for regulation of endothelial physiology during septic events, taking influence on vascular permeability and coagulant activity [1] . animal experiments demonstrated a synergism between hypoxia and septic shock on letality, leading to the hypothesis that low oxygen tension leads to enhanced sensitivity of target cells for tnf [2] . radioligand binding studies with ~261odid-tnf on cultured human endothelial cells were performed after incubation in several environmental oxygen tensions (pc2) for 12 hours. data were achieved by nonlinear regression of an idealized saturation curve according to the equation: b = n " k./(1 + k,); b = totally bound tnf; k,: association constant (concentration for half-maximal binding); n: number of binding sites per cell. p_o0o (mm h¢i): _k, (nm}: n (molecules/cell): 130 -150 1.83 ± 0.15 2600 _+ 500 55 -70 1.27 ± 0.15 3600 + 500 25 -35 0,41 ± 0.13 4800 -+ 500 10-15 0.18 + 0.07 5900 -+ 300 presented are calculated values on the idealized curve + 95 % percentiles. hypoxia induces enhanced binding of tnf to specific receptors on the endothelial cell surface in a time-and dose-dependent manner by a mechanism, which is not dependent on oxygen radicals, as shown by additional protocols with radical-scavenging drugs. with respect to former findings about a correlation between growth and tnf receptor affinity [3] , these data lead to the hypothesis that enhanced tnf binding during hypoxia is due to a biochemical conversion of the receptor protein from the low affinity to the high affinity state, possibly by posttranslational phosphorylation of the binding protein by intracel)ular kinases. the proposed involvement of tnf-dependent pathways in pathogenesis of organ dysfunction and multiple organ failure after hypoxia/ischemia may provide a basis for understanding the initiation of hypoxic vascular injury, as manifested by increased permeability and prothrombotic tendency, and, thus, merits further attention. the levels of activity of circulating cytokines (ill, il-6 and tnf-alpha) which are believed to play important regulatory role in response to trauma are determined (by hioassays and respective anti-cytokine antibodies) in mice and rats subjected to scald injury ion c,10 see, °0v bsa, ld 50) and (80 c, 30 see, 20 ~ b ~^)~ , respectively. biphasic increase of cytokine activity was noted in mice: initial increase of il-i and il-6, 1-3 hr following injury and of try activity 6hr after scald, followed by elevated levels of il-i and il-6 at 12hr, with tendency of decrease of activity at later time points. increased activity of tnf was noted 24 hr following injury, in rats, initial, short-lived increase of il-i and tnf activity was detected lhr following injury, folowed by increase on days i and 3 postburn. il-6 increase peaked 3-12 hr after scalding and levels remained elevated 3-5 days following injury. similar kinetics of appearance of proinflammatory cytokines (il-i and tnf-alpha) both in lethal and ncnlethal injury concomitant with differential profile of circulating il-6 activity (early,short-lived increase and later slow decrease of activity in lethal burn injury) with late persistent high levels of activity in nonlethai injury demonstrated in the present study highlight the need for investigation the relationship of these cytokines in burn-injury induced inflammation. zikica jovicic,lnstitute for medical research, mma,crnotravska 17,11002 belgrade~yu. asadullah k (1), woiciechowsky c (2), liebenthai c (1), doecke wd (1), volk hd (1), vogel s (2), v. baehr r (1); depts. of med. immunology (1) and neurosurgery (2) , medical school (char#d), humboldt university berlin, frg in patients after polytrauma or major abdominal surgery a hyperinflammatory phase seems to be followed by the development of a phase of monocyte inactivation. the latter is charaeterised by a decrease of monocytic hla-dr expression and a shift to anti-inflammatory cytokine production. as shown, by us and others, this phenomenon indicates severe immunodepression with a high risk of infection. however, the mechanisms leading to monocyte inactivation in the above mentioned syndromes may be multiple. to elucidate the influence of a selective, sterile trauma to the central nervous system (cns) on immune reactivity the neurosurgieal patient is an interesting model. initially, 30 patients who developed a systemic inflammatory response syndrome following neurosurgery were analysed. in all of them a marked decrease of monocytic hla-dr expression was observed soon after the operation. these results suggest that neurosurgery alone can induce immunodepression and lead us to conduct a prospective study, in which we closely monitored l0 patients undergoing neurosurgery from the first preoperative day until at least day 6 after the operation. hla-dr expression was decreased hi all patients to various extent only hours after surgery. in one patient only we found a persistently reduced hla-dr expression and this was the only patient to develop sepsis syndrome. this suggests that a prolonged, postoperatively decreased hla-dr expression is predictive of infection following cns trauma. in order to assess, whether a decrease of hla-dr expression was associated with a preceding inflammatory response, local cytokine release in the cns was compared with systemic cytokine release. for this purpose, paired samples of earebrospinal fluid (csf) from a vantricle drainage and peripheral blood plasma were obtained. in the csf extremely elevated futerleakin (il)-6 levels, peaking already a few hours after the operation were found. in plasma, by eontrast, il-6( and tnf-alpha) was detectable not until days later and only if infection was present. the antiinflammatory ili-ra, on the other hand, was also present in csf but peaked after il-6 and was detectable in peripheral plasma too. we believe there is an association between the inflammatory response in the cns and the following depression of hla-dr expression on peripheral blood monocytes. our results suggest that even a sterile cns-trauma by itself may contribute to general immunodepressinn leading to septic complications. the aim of this study was to evaluate the effect of haemorrhagic shock (hs) a) on total capacity of the host, and b) the circulating blood cells to produce tnf immediately after bleeding. in vivo studies: baboons were subjected to a limited oxygen deficit (180-200 ml/kg) hypotension phase (mean arterial pressure = map of 35-40 mmhg for 2-4 hours followed by adequate resuscitation). rats subjected to hs (map of 30-35 mmhg for 90 rain followed by reinfusion of shed blood and fluid resuscitation) were challenged with endotoxin (1 ~g/kg i.v.) at the end of shock (rhs group). the control group (rco) received the same dose of endotoxin as rhs group but without prior bleeding. in vitro studies: whole blood (wb) obtained from both baboons and rats before and at the end of hs were incubated with endotoxin (100 ng/ml) for 2 hrs at 37 °c. results: at 90 min post-lps challenge we found significantly higher plasma tnf levels in rats that were subjected to hs prior to the endotoxin challenge as compared to the control group (956 _+ 585 vs 276 + 199 pg/ml) . after hs the tpc was significantly decreased in in vitro stimulated cbc of both rats (12 + 4 post-hs vs 164 + 89 ng tnf/ml pre-hs) and baboons (8 ± 16 post-hs vs 233 ± 188 pg tnf/ml pre-hs). in contrast, the il-6 productive capacity was increased in baboons cbc (not yet analysed in rats) stimulated at the end of hs (224 ± 106 pre-vs 1545 ±_ 1254 pg il-6/ml post-hs). conclusion: from our data we suggest that despite of down regulation of the cbc to produce tnf the overall tpc is enhanced at the early stage of i-is. with regard to the related literature (chaudry's group) it can be assumed that among the macrophage/monocyte populations, as the main source only the kupffer cells (kc) exhibit enhanced tnf production capacity following haemorrhage. the mechanisms of down/up regulation of cytokine response of cbc and/or kc following hs remain to be examined. d. eg~er, s. geuenich °, c. dertzlin~er °, e. schmitt*, r. mailhammer, h ehrenreich #, p. drrmer, and l. h01mer gsf-instimt fox experimentelle h~znatologie, °medizinische kliulk iii, klinikum groghadern, munich, *institut for immunologic, johannes gutenberg universit/it, malnz, and #psychiatrische k/in& der georg-aagust-universi~t, grttingen, germany. it has been shown previously (ehranreich et al., 1992, new biol. 4: 147 ) that mouse bone marrow-derived mast cells (bmmc) synthesize and secrete endothelin-1 (et-i) and express eta-type endothelin receptors (eta). so far, however, no functions of et-1/et a in bmmc have been described. in the present study we investigated the effect of exogeneously administered et-1 on the release of histamine, serotonin, and leukotriene c 4 (ltc4) by primary mouse bmmc (in vitro age: 4 weeks) caltured with different recombinant mttrine cytokines (interleukin 3 (il-3) and/or kit ligand (kl) in the presence or absence of il4) for two weeks prior to activation. et-1 (5x10 -8 to lxl0 -6 m) induced an extremely rapid (_<lmin) and dose-dependent release of histamine and serotonin (up to 20% and 16% of total mediator content, respectively, comparable to ige/ag-indueed mediator release) from bmmc cultured with il-3 and il-4. only when cultured in the additional presence of il-4 rather than in medium containing kl or kl plus il-3 alone, bmmc released histamine and serotoaln upon challenge with et-1.furthermore, et-1 stimulated ltc 4 biosynthesis up to 4-fold in bmmc cultured in the presence of il-4. in contrast, the peptide was without major effect on ltc4 biosynthesis in bmmc culturd without il-4. the release of histamine and sereotonin was not altered if bmmc were preincubated for i h with il-4 prior to activation with et-i, suggesting that a differentiation process rather than a functional priming effect had been initiated by ]l-4. et-1 (10"6m) -induced histamine and serotouln release from bmmc was inhibited by an eta-specific antagonist (cyclic id-asp-pro-d-val-leu-d-tel ) in a dose-dependent manner, with a complete inhibition observed at an antagonist concentration of 10 -8 m. crude peritoneal cells (containing 2-3% serosal mast cells) obtained from normal balb/c-mice released 87% histamine upon challenge with et-1 (10 -6 m; 20 rain). taken together, these resu/ts demonstrate that et-1 can directly function as a histamine and serotohin secretagogue and as a stimulator of ltc 4 production in mast cells. il-4 appears to be involved in the differentiation of immature mast cell prec~trsors towards an et-l-reactive functional phenotype. when inflammatory reactions are advanced, type ii phospholipase a2 (type ii pla2) is produced in high levels by various cells at the site of inflammation. cytokines such as tnf-a and il-i activate type ii pla2 and promote the production of eicosanoid, which is one of the mechanisms by which they enhance the inflammatory reaction. in the present study, the blood levels of type ii pea2, endotoxin, tnf-c~, il-6 and il-8 were determined in patients with sepsis to examine the relationship between these levels and the patients' pathological conditions. the levels of type ii pla2 and tnf-a in these patients were 218.3 ± 179.9 ng/ml and 98.5 ± 92.1 pg/ml, respectively, the two parameters being significantly correlated (r = 0.6460, p = 0.0001 ). there were also a significant correlation between the level of type ii pla2 an il-6 (r = 0.4262, p = 0.0188). there was no significant correlation between the level of type ii pla2 and il-8. these in vivo findings suggest that tnf-a may be involved in the production of type ii pla2. a. filzmaver, g. reisbach, e. schmitt °, r. mailhammer, and l. hiittner. gsf-iustitut ff~r experimentelle h~imatologie, munich, and °iustitut fttr immunelone , johannes gutenberg universit~t, mainz, germany the product of c-kit, a transmembrane tyrosin ldnase receptor, and a corresponding kit ligand (kl) are critically involved in the control of different hemopoietic cell lineages including the proliferation, maturation, migration, and function of mast cells. it has been reported previously that interleukin (il)-4 down-regulates kit receptors in human primary myeloid leukemic cells and in a human mast celt line (sinaber et al. 1991, j. immunol. 147: 4224) . transforming growth factor (tgf) gl, was also found to down-modulate c-kit mrna and kit receptor proteins in human aml blasts, a mechanism probabely contributing to the anti-proliferative effect of tgfih on kl-stimulated aml ceils in vitro (de vos et at. 1993, cancer res. 53: 3638) . in porcine endothelial cells transfected with a retroviral construct carrying the human (hu) e-kit gena, exogenous hu kl transiently down-regulated kit receptors (blume-jeusen et al. 1991, embo j 10: 4121) . in the light of these previous findings we analyzed the effects of these exogenously applied cytokines (il-4, tgfgl, kl) on kit receptor expression and kl-mediated proliferation and chemotactic migration of primary mouse bone marrow-derived mast cells (bmmc) (in vitro age: 4 weeks). our results show that in bmmc cultures initiated with recombinant (r) marine (mu) il-3 the additional presence of rmull-4 for 3 or 14 days did not change the expression of kit protein (assessed by facs analysis with the anti-mouse c-kit antibody ack-2) nor kl-mediated proliferation or chemotaxis. in contrast, il-4 syeergistically increased kl-stimulated growth of bmmc in a short-term proliferation assay (mtt-tes0. pre-incubation of bmmc with rhutgfl~ 1 (5.0, 1.0, or 0.1 ng/ml) for 24h had no effect on kit receptor expression, although tgffil at concentrations of 0.5 -5.0 ng/ml completely suppressed the proliferative action of kl on these ceils. similar anti-proliferative effects of tgfg 1 were observed in various factor-dependent mast cell lines stimulated with different mast cell growth factors (il-3, 1i,-4, il-9, and/or il-10). moreover, pre-incuhation (24h) of bmmc with tgf-gl (>500 pg/ml) significantly enhanced spontaneous undirected cell movement (chemokinesis) and synergistically increased il-3-or kl-induced chemetaxis. when bmmc were preancuhated with rmukl (200 ng/ml) for 1, 3. or 5 days, a transient down-modulation of kit receptors with a maximum effect on day 1 was demonstrated by facs analysis and correlated well with a decreased chemotactic response of these cells. in conclusion our results show that neither il-4 nor tgfi31 affect expression of kit receptors in primary murine bmmc. it is reasonable to suggest that c-kit expression is controlled in a cell type-specific manner.interestingly, tgfgl is obviously able to dissect the proliferative from the migrational signal transducted by kl in these cells. objectives of the study: antisense strategies using dna-otigonucleofides (odn) to modulate the cytokine response are presently under investigation. odn are thought to act very specifically with little or no relevant negative side effects. we now report that odn unspeeifically protect wehi 164 cells from tnf-mediated cytolysis. material and methods: wehi 164 subclone 13 ceils (5 x 105), that are highly sensitive to the cytolytic activity of tnf, were grown on 96-well culture plates in rpm11640 medium. after 24 hours, phosphorothioate(ps)and partially ps-modified-odn as well as phesphodiester-odn (20-29bp) were added (0.1, 1 and 10 pm). four hours after incubation with odn, ce(i lysis was induced by recombinant murina tnf. after 18 hours the plates were washed and stained with crystal violet cell lysis was determined by reading the absorbance (abs) at 590 nm. results: wehi 164 ceils incubated with tnf (1-8ng/ml) were completely lysed after 18 hours (0% abs). interestingly, wehi cells incubated with tnf and odn resisted complete lysis, eg cells incubated with 2.5ng/ml tnf and 11jm odn showed still 30% of the absorbance observed in control ceils without tnf (100% abs). the protective effect of odn started at 0.1pm, reached a maximum at 1,um, and diminished at 101jm. with increasing amounts of tnf the protective effect of qdn decreased and no protection was detectable at 8ng tnf per ml conclusions: dna-oligonucleotides were found to unspecifically inhibit tnf-induced cytolysis. we hypothesize, that this protective effect of qdn results from an inhibition of the binding of tnf to its receptor, or from interference of odn with the subsequent signal transduction mechanisms. as a consequence, to discriminate the specific effect of odn in biologic systems, several control odn should be used. secondly, whether dna released by degradation of tumor cells or leukocytes can significantly impair tumor-and immune-defense mechanisms merits further investigation dr. med. michael meisner, institut for anaesthesiologie der universitat erlangen-nqmberg, krankenhausstral~e 12, d-91054 erlangen. in this study we investigated the involvement of serine protease and free radical generation in the systemic release of tumor necrosis factor-alpha (tnf) and interieukin i(il-1), in the sepsis model of lipopolysaccharide (lps, 5mg/kg i.p.) induced hepatitis in galactosamine (gain, 18rag/mouse, i.p.) sensitized mice. treatment of gain-sensitized mice with lps (gain/lps) led to dramatic increase in serum cytokine (tnf and il-i) ievels and transaminase activity at 3 hr and 8 hr respectively. pretreatment of serine protease inhibitor, c~jantitrypsin (a j-at, 50mg/kg i.p.), 30 rains prior to gain/lps treatment, fully protected the animals against the hepatotoxic challenge with significantly reduced serum tnf and il-1 levels. in order to block and scavenge superoxide generation, the mice were pretreated with xanthine oxidase inhibitor, allopurinol (al, 2 x 100mg/kg i.p.) and pyran polymer-conjugated superoxide dismutase (sod, 2 x 200 unit/mouse i.v) r6spectively. pretreatment with al and sod (24 and 1 hr prior to gain/lps) prevented gain/lps hepatitis and blocked lps induced released of tnf and il-1 into serum of the mice. the protective agents like cq-at or al/sod did not protect the mice against th~ hpp~totoxi£ ch~llpn-e indllee4 b'~ th~ recombinant mmlse tnf-o' (0.3 ~/rno~e j.p.) ~d oi~lps 1~ caln-.~dlfa%aed mlce. it-l cett~aged la tnf (x/gain treated mjde was not detectable in animals pretreated with oq-at or al/sod. our study suggests that a serine protease sensitive to cq-antitrypsin is responsible in regulating tnf release, possibly by proteolytic cleavage of a tnf-precursor or membrane bound tnf. in addition our evidence suggest that the balance of extracellular protease/antiprotease activity may be regulated by free radical generation, possible superoxide anion, resulting in inactivation of the antiprotease. il-1 release may be subsequent to tnf release. objective: during sepsis one can observe a dramatically impaired production of proinflammatory cytokines like the tumor necrosis factor alpha (tnf-a), interleukin i-alpha (il-la), intedeukin i-beta (il-i&) and interferon gamma (if~) upon in vitro stimulation of circulating cells. however there is also evidence of a decreased ability to produce cytokines in other immuno-deficient states. in this study we compared the capacity to secrete proinflammatory cytokines upon in vitro stimulation of patients in severe sepsis and patients with malignant tumors. methods: heparinized blood samples of ten patients (62+ 14 years) in severe sepsis (sepsis score > 15 according to e}ebute and stoner) were drawn at onset of disease, from fifteen patients with solid growing carcinoma (59+19 years) blood was drawn at diagnosis prior to any therapy. controls were obtained from fifteen healthy volunteers. 50 pl of whole blood were incubated either with 450/4 of a standard medium or with 400 pl of a standard medium and 50 pl of phytohemagglutinin (pha) a potent mitogen. after an incubation period of 24 hours plasma concentrations of tnf-a, il-la, il-16 and if-~ were determined by elisa. comments: our results suggest that down-regulation of cytokine secretion or of cell responsiveness to non-specific mitogens during sepsis has occurred. we observe a similar phenomenon for the group of carcinoma patients vs control significant for stimulated tnf-a and stimulated if-t. sustained immunological interactions between tumorcells and cytokine producing cells could effect responsiveness of the latter, a general increased immuno-tolerant state in patients with carcinoma has to be discussed. however we found significant differences between sepsis and cancer concerning the in vitro capacity of responsable cells to produce il-la and il-i#. the dramatically decrease of the ability to produce il-i upon in vitro stimulation could be more sensitive for a septic state than stimulated tnf-a or if-3,. objective: tumor necrosis factor alpha (tnf-a) has been implicated as a central mediator of sepsis and its sequelae. increased systemic levels of this cytoklne seem to be correlated with severity of sepsis and outcome. however mechanism of action and metabolism of tnf-g are not fully understood. in most studies blood samples for tnf-a determinations are obtained either by peripheral venipuncture, a central venous catheter or by an indwelling arterial catheter. very often blood samples are taken in different manners within the same study. in this study we measured circulating tnf-a and the amount of tnf-a released upon in vitro stimulation in arterial and central venous blood. methods: heparlnized arterial and central venous blood samples of ten patients (6 males, 4 females, mean age 62+_14) with severe sepsis (sepsis score > 15, elebute and stoner} were drawn on day 1,3,6,8, and 14 of disease. blood was immediately placed on ice and processed within 1 hour. 50 pl of whole blood were incubated with 450 pl rpmi-medium supplemented with antibiotics and l-glutamlne or with 400 pl of rpmi-medium and 50 pl phytohemagglutinin (pha) a potent mitogen. after an incubation period of 24 hours samples were centrifuged and plasma was harvested and stored at -30 ° celsius before assessment of tnf-a concentration by elisa. statistical analysis was performed with the paired student-t-test. results: we found a significant difference (p < 0,05) for circulating mean arterial tnf-a concentration (237 pg/ml _+ 34 sem} and central venous tnf-a (203 pg/ml +_ 29 sem). upon in vitro stimulation there was also a significant difference (p < 0,002) between released arterial tnf-~' {746 pg/ml _+ 88 sem) and venous tnf-a (637 pg/ml +_ 76 semi. conclusions: these results are difficult to interprete but could reflect the influence of pao 2 and sao 2 on tnf a release. it could also be the result of different concentrations of tnf-o release influencing factors like for example endotoxin, interferon-f or prostaglandin. a possible pulmonary and/or a hepatic metabolism of tnf-n and tnf-a producing cells cannot be ruled out. however for better interpretations of tnf-a release in septic states it is necessary to use either arterial or venous blood samples. early inflammatory processes following trauma and/or infections were found to be associated with the secretion of high amounts of proinflammatory cytokines. besides intedeukin-t (il-1), tumor necrosis factor-a (tnf-c 0 and interleukin-8 (il-8) the multifunctional cytokine intedeukin-6 (il-6) was described to be a central regulatory element of the primary cellular and humeral defence reaction. the previously described close temporal correlation of pathologically elevated il-6-concentrations and the extracellulary release of lysosomal enzymes from activated pelymorphnuclear neutrophils suggests, that il-6 may be a potential substrate of these preteases. the serine preteases elastase (ec 3.4.21.37 ) and cathepsin g (ec 3.4.21.20) derived from the azurophilic granules were assumed to be mainly involved in unspecific proteolysis at sites of inflammation by cleavage of structural as well as soluble proteins at random sites, if the inhibitory potential is decreased. the possible proteolytic activity of elastase and cathepsin g toward the proinflammatory cytokine interleukin-6 (il-6) was investigated. the addition of purified neutrephil elastase and cathepsin g to recombinant human il-6 leads to a rapid sequential degradation in vitro. at least two intermediate products could be detected by silver staining and western blotting following protein separation under reducing conditions. the serine protease inhibitor g-anitrypsin was shown to prevent the proteolytical degradation of intedeukin-6. furthermore the loss of the biological activity of both, recombinant and natural human il-6, was demonstrated by determination of the capacity of protease-treated il-6 to stimulate hybddoma growth (7td1 bioassay). these data suggest a possible downregulation of pathologically elevated il-6 levels by proteolytic activity of extracellulary released enzymes at sites of inflammation. the aim of the study was to compare circulating levels of three cytokines -il-1, il-6, 11_-8 -between critically ill subjects who developed gram-negative sepsis and who did not. materials and methods: the patient population consisted of 39 patients admitted to an intensive cars unit, with different underlying diseases. sepsis diagnosis was given according to pre-estabilished cdteda. nineteen cases were enrolled in sepsis group, twenty in control group. serum sampling was collected in sterile tubes at study entry and every three days until study dismissal. serum concentrations of il-1, 11_-6 and il-8 were measured using commercially available test kits, based on the dual immunometric sandwich principle. results: the causative patogens of sepsis were: pseudomonas aeruginosa, acinetobacter, eseherichia co~i, serratia marceseens, proteus mirobilis and citrobacter freundl the time of observation was equal to 12 days, for a total of four tests performed (to, tl, t2, t3). i1.-1 was not detected in any samples. the serological profiles of the two cytokines 11.-6 and 11_-8 were similar; augmented levels were found at study entry and throughout the observation period, peaking at t3 and decreasing at t4. however, in patients with sepsis, il-6 and 11_-8 concentrations were significantly higher in respect to control group. conclusion: our observations shown that in icu patients increased il-6 and il-8 release may be induced by cdtical illness; however, in subjects in which sepsis occurred, il-6 and il-8 production appears more significantly elevated, suggesting a role of il-6 and 11_-8 in the pathophysiology of sepsis. the fact that ii. objective: to check whether continuous veno-venous haemofiltration (cvvh) could remove the cytokines, namely tumour necrosis factor alpha (tnfc 0 and interleukin 6 (il-6) from the circulation of critically ill patients with sepsis ad multiple organ failure (mof). setting: the intensive therapy unit of the medical school teaching hospital. patients: nine critically ill patients with sepsis and mof treated with cvvh. methods: blood samples were collected before the cvvh had been started. then, blood and ultrafiltrate samples were collected simultaneously after 4 hours and every 24 hour. tnfct and il-6 levels were measured using the bioassays with cell lines wehi-164 ci13 and 7td1, respectively. other data were recorded from the patient notes and intensive therapy unit charts. results: no measurable concentrations of tnfct were detected in either blood or ultrafiltrate samples. il-6 was found in all the patients' plasma samples and five patients' (55.5%) ultrafiltrate samples. the il-6 blood level ranged from 17.4 to 3061.7 u/ml (mean 518.2, sd 686.8). the il-6 level in positive ultrafiltrate samples ranged from 21.0 to 1150.2 u/ml (mean 252.2, sd 401.8). conclusions: our preliminary results suggest that il-6 is present in bloodstream of septic patients. we assume we could not detect tnfa in any sample because we usually started observations when septic state had developed. cvvh could extract cytokines from the circulating blood. it remains under discussion, whether that extraction may be beneficial to patients with mof. the pattern of some significant cytokines tnf, il-1 and il-6 and their pharmacomodulation were evaluated in an experimental model of polimicrobial sepsis induced in cd-1 mice by cecal ligation and puncture (clp) in order to understand their roles. this model of sepsis, which resembles the clinical situation of bowel perforation, was also compared with that induced by administration of pure endotoxin (lps). tnf was detectable in serum and tissues during the first 4h with a peak 2h after clp at a significantly lower level than after lps. il-1 was measurable in serum only after 24h, significantly increased in spleen and liver after 4 and 8h and in mesenteric lymphonodes from 8 to 24h after clp compared with shammice. il-6 was significantly increased in serum throughout the first 16h after clp. pretreatment with dexamethasone (dex), ibuprofen (ibu) and nitro-l-arginine (n-arg) significantly reduced the survival time while chlorpromazine (cpz) and tnf did not affect it. only the antibiotics and pentoxifylline (ptx) significantly increased the survival in clp. however cpz and dex protected from lps-mor~ality. in conclusion, by inhibiting tnf with dex, cpz, ptx a reduced, unchanged and increased survival time was observed and by increasing tnf with ibu and tnf administration the survival was decreased or unchanged respectively suggesting that the modulation of this cytokine does not seem to play a significant role in clp unlike lps_ moreover the negative effects of ibu and n-arg suggest an important and protective role by prostaglandins and no in clp. to gain more insigths on the contribution of tnf~, il-i~ and if 7 to lps toxicity, we explored the time-course of the cytokine production in ealb/c mice given different doses, from the lethal (= 3 ld50) to the sublethal (= 1/3 ld50) of three different lps (e.coli oiii:b4 and 026:b6; p.aeruginosa r261) endowed with different degree of toxicity cytokines were measured in serum and organs with specific elisas up to i0 h after lps administration. results demonstrate that i) circulating and organ levels of tnf~ do not reflect lps toxicity. in fact, the lethal dose of lps 026:b6 induced as much tnf~ as the sublethal dose of lps 0111:b4; furthermore, lps r261, whose cytokine inducing capability is far lower than that of lps from e.coli, induced higher tnf~ levels at the sublethal than at the lethal dose. in addition, policlonal anti tnf ab, that were able to protect mice from e.coli lps induced mortality, failed in mice treated with lps r261 2) circulating il-i~ levels are generally low and increase significantly only in muribond animals. on the contrary, in spleen and lung very high levels of il-i~ are persistent from i to 8 h post lps administration moreover, the treatment with 6 mgr of neutralizing policlonal anti il-i~ ab, did not modify survival in lps challenged mice. 3) circulating and organ levels of if 7 are proportional to the dose and degree of toxicity of all the administered lps even if lps r261 was again a less efficient cytokine inducer than lps from e.coli. csa is an immunos~ppressive drug, able to inhibit gene expression for many cytokines, including if 7. to study the effect of cytokines modulation on lps toxicity, csa was administered to mice twice at the oral dose of i00 mg/kg before the challenge with lps. mice were monitored in terms of mortality and tnf~, il-i~ and if 7 production. together with the total ablation of if7, the strong reduction of tnfu and unmodified il-i~ levels, a significant increase of lps toxicity was also observed. these results suggest the hypothesis that the numerous factors that jointly mediate lps toxic effects, can also be protective, the final outcome depending on their relative ratio rather than on the absolute amount interleukin-1 (il-1) mediates the septic shock syndrome and affects intestinal secretion in vitro. we studied the intestinal production of il-t and its effects on diarrhea during endotoxic shock. cd-1 mice were randomized to 15 mg/kg e.coli 0111:b4 lps or saline infusion (i.p. or i.v.). diarrhea invariably occurred following lps infusion. mice were sacrificed at 0, 30', lh, 2.5h, 4h, 6h, 12h, and 24h (3 mice/group/time-point). the small bowel was compressed and the intestinal contents were weighed and expressed per g sb weight. the small (sb) and large bowels (lb) were eventually frozen, weighed, and homogenized for either cytosolic protein or total rna. il-i~ (cell-associated agonist) was measured with a radioimmunoassay specific for mouse il-l~ (detection limit 100 pg/ml) and expressed as ng/g weight + sem (lowest detectable amount 1 ng/gwt). northern analysis of total rna and in sfu hybridization of paraformaldehyde-fixed frozen tissue were done with [0~-32p]-iabeled mouse il-lc~ cdna probes. only sb had il-i~ constitutively present (6.2 + 1.6 ng/gwt). lps i.p. or i.v. induced elevation of il-lc¢ in both organs in a biphasic pattern; lps i.v. induced 3-fold more il-i~ than lps i.p. following lps i.p., il-i~ in sb was 19.2 + 0.6 ng/gwt at lh, reached maximal levels at 2.5h (31.8 -+ 0.5 ng/gw-i) and returned to baseline at 24h. saline controls maintained their constitutive il-i~ levels. sb had 2fold more il-1 ¢ than lb and identical kinetics, but lb showed a clearer doseresponse. northern analysis of sb-total rna showed induction of il-i~ mrna by lps in correlation with il-lc¢ kinetics. il-i~ mrna producing cells were mononuclear cells in the lamina propda and epithelial cells at the bottom of the crypts of ueberkuhn. mucus and fluid were increased in the small bowel post-lps in correlation with intestinal il-lc~ kinetics (r2=0.9). separate mice were pretreated with saline i.p. orthe il-1 receptor antagonist (irap, 30 mg/kg bolus i.p.) and were challenged 20 rain later with 1.5 mg/kg lps i.p. or saline i.p. specific blockade of il-1 by irap decreased intestinal secretion at 4h and 6h post-lps challenge (p<_. 0.05, student's-t-test). these data indicate that local (intrinsic) intestinal il-i~ mediates sepsis-induced intestinal changes. inflammatory cytokines initiate the host response to endotoxemia, causing severe physiological and hemodynamic changes which may lead to septic shock. among the regulatory systems that play an important rote in controlling host inflammatory responses is the pituitary. it has been known for many years for example, that hypophysectomized animals are extremely sensitive to lps lethality. while investigating the possibility that protective, pituitary mediators might explain this phenomenon, we identified the cytoldne mif to be a specific secretory product produced by pituitary cells in vitro and in vivo after lps challenge. analysis of serum mif levels in control, t-cell deficient (nude), and hypophysectomized mice revealed that pituitary-derived mif contributes significantly to the rise in serum mif that occurs after lps administration. of note, pituitary mif content (0.05% of total pituitary protein) and peak serum mif levels (80-340 ng/ml) were determined to be within the range observed for other pituitary hormones that are released after pituitary stimulation. to investigate a possible beneficial role for mif in septic shock, we co-injected mice with purified, recombinant murine mif (rmif) together with lps (15 mg/kg). surprisingly, rmif markedly potentiated lps lethality compared to control mice that were injected with lps alone (85% vs. 35%, p = 0.003). to confirm these results, mice were treated with anti-rmif antibody prior to injection of a high dose of lps (17.5 mg/kg). anti-rmif antibody fully protected mice against lps lethality, increasing survival from 50% to 100% (p = 0.0004). serum levels of tnf,~, the first cytokinc that appears in the circulation after lps challenge, were reduced by 38.0 _+ 9.5% in anti-rmif-treated mice. we conclude that pituitary derived mif contributes significantly to circulating mif in the post-acute response in endotoxemia and may act in concert with other pituitary mediators to regulate both pro-and antiinflammatory effects. moreover, mif may play a critical regulatory role in the systemic host response in septic shock. our results suggest that anti-rmif antibody might be of potential therapeutic use in the treatment of septic shock. although anti-interleukin-1 (il-1) antibodies and il-1 receptor antagonist have been shown to improve survival in animal models of endotoxemia and abrogate the lethal effects of tnf, the presence of il-1 in the serum does not correlate well with outcome. we hypothesized that this may be because il-1 acts mainly in a paracrine fashion and is metabolized before it diffuses into the circulation. methods: we measured the il-i~ mrna expression with the differential reverse transcription polymerase chain reaction (rt-pcr) using g-actin as internal standard in the peritoneal macrophages and lung tissue in normal controls and mice after cecal ligation and puncture (clp). clp resembles human intra-abdominal sepsis in that it is characterized by very slight elevations of serum il-1 levels. results: il-lg mrna levels after clp are expressed as % of normal (mean+sem, n=5 in several experimental models of infection exacerbation of disease was observed, when infected animals were depleted of tuajor necrosis factor (tnf). after sublethal cecal ligation and puncture (clp) leading to peritonitis and sepsis the survival of mice also critically depends on tnf as demonstrated in earlier studies, when clp-treated mice injected with anti-tnf antibody died, whereas mice injected with a control antibody survived after clp (echtenacher et al. 1990, j. inununol. 145: 3762) . from a panel of different cell types (macrophages, neutrophils, t lymphocytes, natural killer cells, mast cells) able to produce tnf upon activation~ the mast cell is apparantly the only one capable of storing in cytoplasmic granules preformed tnf-ct which is rapidly released following challenge. in the present study-we analyzed serum tnf after lps injections as well as the outcome of clp in severely mast cell deficient mutant mice (wav v) as compared to syngeaeic wild-type littermates (+/+). we proposed that concentrations and/or kinetics of serum tnf should be different between wavv mutants and wild-type mice, if mast cell-derived tnf significantly contributes to the rise in serum tnf levels following systemic stimulation with endotoxin. although similar levels of increased tnf were detected in the sera of both genotypes after 1 and 2 hours of lps injection (100 btg/ 0.5 ml / mouse i. p.), mast ceil-deficient mice indeed showed decreased serum tnf levels 30 iron after injection amounting to only 12 to 25% of the concentrations observed in the corresponding sera of normal wildtype mice. in the clp model of septic peritonitis we found that mast celldeficient mutant mice were dramatically more sensitive to clp than syngeneic normal mice resulting in 92% mortality in w/w v versus 8% mortality in +/+ mice 2.5 days after initiation of clp. further experiments with w/w v mutants selectively reconstituted with cultured bone marrow-derived mast cells from normal syngeneic wild-type mice and the use of an antibody specifically blocking the action of tnf tn vivo should clarify a potential protective function of mast cells in this model of septic peritonitis. interleukin-10 (il-10) inhibits cytokine production, including tumor necrosis factor (tnf), by lipopolysaccharide (lps)-aetivated maerophages. we recently observed that lps injection (e.coli 055:b5, 100 gg ip) into balb/c mice induces the rapid release of circulating il-10 (17±9 u/ml at 90 min). blocking endogenous il-10 using monocional antibody (jes5-2a5, 2 mg, 2h before lps) resulted in a massive increase in tnf production (107±47 in lps+anti-il-10 treated mice vs 6±2 ng/ml in lps alone, p<0.05, n=4 to 5 mice per group) and an enhanced lps-induccd lethality (53% vs 7% in anti-il-10+lps or lps alone respectively, p=0.01, n=15 mice per group). irrelevant igg1 rat monoclonal antibody (lo-dnp) did not influence neither tnf production nor lethality associated with endotoxin shock. this led us to study the production of il-10 during human septicemia. plasma samples were obtained from 65 patients with gramnegative (gns, n=22) or gram-positive septicemia (gps, n=43) and from 20 healthy volunteers. among these patients, 17 suffered from septic shock at the time of sampling. il-10 levels were measured by elisa (detection limit: i 1 pghrd). we found that 35 patients (54%) had increased il-10 plasma levels (range 12 to 2740 pg/nd). patients with gps had il-10 levels similar to the ones observed in gns (median: 12 vs 17.5 pg/m, respectively). patients with septic shock had higher il-10 values (median: 58 pg/ml) than septicemic patients without shock (11 pg/ml, p=0.002). no il-10 was detected in plasma from healthy volunteers. we conclude that il-10 is produced daring human septicemia. our experimental data suggest that il-10 might be involved in the control of the inflammatory response induced by bacterial products. dr arnand marchant, immunology department, hopital erasme, 808 route de lennik, 1070 brussels, belgium. to provide information about the role of tnf in sepsis and mods we measured tnf and stnfr-i levels in septic patients and investigated if there is a relation between plasma concentration of these molecules and the severity of sepsis evaluated by two scores (apache 1i and sss). patients and melhods: 20 septic patients fullfilling sepsis criteria of american college of chest physician and society of critical care medicine were studied. tnf-cc and stnfr-i (60kda) were measured by enzyme immuneassays (norms1 values = 13+8 pg/ml and 1.06_+0a ng/ml respectively). results: the mean tnf and stnfr-i values for each patient (mean+sd) were 93+64 pg/ml and 11.1+5.5 ng/ml respectively. these values are approximately seven and ten times greater than those observed in normal healthy volunteers (p<0.001). mean tnf concentrations for each patient were significantly greater in non survivors (112+52 vs 64_+74 pg/ml p<0.05); stnfr-i levels also were greater in this group, but the difference was not statistically significant (12.6+5.2 vs 8.8_+5.5 ng/ml). plasma tnf and stnfr-i concentrations were significantly correlated (r = 0.65 p<0.005). mean tnf levels were significantly correlated with apache ii (r = 0.49 p<0.05) and sss (r = 0.54 p<o.01); stnfr-i levels were likewise correlated with both scores (r = 0.80 p<0.001 and r = 0.76 p<0.001 respectively). tnf and stnfqgi levels increase with the number of dysfuctional organs; in particular tnf and stnfr-i increase when hemodynamics deteriorate and kidney failure ensues. conclusions: the correlations between tnf and stnfr-i levels and sepsis severity scores suggests that tnf is involved in sepsis and may influence the occurrence of mods and finally clinical outcome. tnf and stnfr-i are in fact especially increased when mods occurs. in particular their concentrations are elevated when cardiovascular system and kidney failure ensue, suggesting that tnf can have a direct role in hemodynamic derangements caused by sepsis and that kidneys are involved in tnf and stnfr-i excretion. lif is a pleiotropic cytokine that has been implicated in the pathogenesis of sepsis in animal models. we conducted a prospective study in 33 septic patients to correlate lif plasma levels with patient's clinical and biological data (among them il-6). plasma was drawn daily from day one to .ten and lif presence was determined with a specific elisa and with the dai,a bioassay (sensitivity of 100 pg/ml and 1 ngml respectively). risk factor associated wftt~ with elevated lif plasma level was determined by an univariate analysis. a multivariate stepwise logistic regression analysis was subsequently performed with a bmdp statistical software. lif was detected with the elisa and with the bioassay in 11 and 2 out of the 33 patients, respectively. lif peak plasma levels were statistically associated with high creatmin levels, temperature and il-6. multivariate regression analysis showed that high serum creatinin level was the major independent risk factor associated with elevated lifplasma levels. this correlation was also found for il-6. peak plasma levels of both lif and il-6 correlated inversely with patients survival duration. cox's analysis for plasma levels of lif >480 pg/ml yelded a hazard ratio of [exp (1.86)=6.44]. our study indicates that lif levels were associated with clinical and biological parameters of illness severity and significantly increased (cut-off value 480 pg/mi) in patients with fatal outcome. current consensus exists about the central role of tumor necrosis factor (tnf) alpha in initiating the systemic inflammatory response syndrome (sirs). a correlation with sirs has inconsistently been found. tnf effects its pleiotropic reactions upon two distinct cellular receptors. soluble extracel]ular fragments of the human 55 kda tnf receptor (stnfri) and the 75 kda receptor (stnfrii) are detectable in the circulation. the kinetics of these endogenously produced tnf-inhibitors were measured to evaluate their role in patients with sirs. fourteen patients of an operative icu were included with the diagnossis of sirs (mean apache ii score: 20 points). serial blood samples were obtained within 12 h after diagnosis of sirs, every 6 hrs for the first 48 hrs and every 12 hrs thereafter until patients died or recovered. soluble tnfri and stnfrii were assayed by an enzymed-linked immunological binding assay. soluble tnfri and ii could be detected in all samples with a significantly higher level (p<o,o01) compared to healthy controls (normal value: tnfrh 1,5 ng/ml; tnfrii: 2,3 ng/ml). the serum concentration of tnfri (14,1 ng/ml) as well as tnfrii (18,2 ng/ml) were significantly higher in nonsurvivors (n=6) compared to survivors (8,6 and 9,5 ng/ml; n=8) at the onset of sirs and during the course of the inflammation response (p<0,05). a positive correlation exists between both receptors (r-0,57; p < 0,01 ). increasing levels and maximal values of stnfri (59 ng/ml) and i1 (82 ng/mi) were detected only in patients who died. the serum concentrations of patients who recovered did not change and are remaining at the initial level. tnf alpha bioactivity was detected in 10 out of 14 patients. no significant correlation exists between the concentration of both receptors and tnf alpha as well as the apache ii score at the onset of sirs. elevated serum concentration of stnfri and 11 are found in patients with sirs. these naturally occuring antagonists reflect the inflammatory process. the measurement of soluble tnf receptor levels may be useful in monitorina sirs and in the prediction of outcome. 11-2 is given to patients with advanced melanoma or renal carcinoma. however, this therapy is accompanied by severe side effects, including the vascular leak syndrome (vls) that closely resembles septic shock. to investigate the involvement of cytokines and phospholipase a z in the pathogenesis of the vls we collected serial plasma samples from patients during the first 24 hours after administration of il-2. in these patients we monitored temperature, blood pressure and heart rate and assessed levels of the cytokines tnf, il-6 and il-8 using immuno assays. in these plasma samples we also measured phospholipase a 2 activity using an enzyme activity assay, since this enzyme is beleived to play a major role in the generation of lipid mediators. we demonstrate that during il-2 therapy the cytokines tnf, il-6, and il-8 are produced and that the release of these cytokines is followed by an induction of phospholipase az activity in the plasma of these patients, we further discuss the role of these mediators in the development of the vascular leak syndrome observed in patients receiving il-2. objectives of the study: sepsis caused by candida albicans (ca) is no longer a clinical rarity but a common consequence of immunosuppression and surgery, and is associated with high mortality. tumor necrosis faetor-c¢ (tnf-<~), interlcukin-lg (il-lg) and interleukin-6 (il-6) are thought to play an important role in the pathogenesis of the sepsis syndrome. we therefore studied the in vitro production of tnf-<x, il-ib and il-6 by human peripheral blood mononuclear cells (pbmc) stimulated by ca compared to stimulation by bacterial lipopolysaccharide (lps). materials and methods: pbmc were isolated from venous blood of 5 healthy volunteers and cultured at a concentration of 2.5 • 10 ~ eells/ml in culture medium with a dose-range of either heat-killed ca or lps. supernatants were harvested after 24 hours. using elisa, tnf-c~, il-ib and il-6 concentrations were measured. ca was isolated from a patient with ca sepsis and cultured under sterile conditions. results: ca and lps stimulate the production of tnf-~t, il-11] and il-6 by human pbmc. we observed a dose-dependent synthesis of these three cytokines in human pbmc. tnf-c¢, il-ib and il-6 in ng/ml as mean _+ standard error. ca/ml 0.0 1. (2) we constructed a single-chain fv-fragment (sc-fv) from the variable domains of a neutralizing antibody to human tumornecrosisfactor-alpha (tnf) because sc-fv should have some advantages in vivo. the sc-fv has a similar affinity (3,8x10-9m) as the fab-fragment (2,7x10-9m) but bind the antigen with a eightfold lower avidity compared to the parental mab (4,8x10-10). the parental mab as well as its fragments can compete the binding of rhutnf to both tnf-receptors. this was shown by competitive binding to the cell line hl-60 and to immobilized rtnf-receptors. in the nutralization assay on the cell line, l 929, the sc-fv can neutralize tnf but in comparison to the mab or its fabfragment, the capacity was three-fold lower as expected from the molar ratio of the dissociation constants. the in rive-neutralizing capacity was tested in a mice receiving toxic dose of rhutnf. the sc-fv showed a 15-fold lower neutralizing capacity in comparison with the fab-fragment. this could be explained in different manners: i) partial instability of scfv at 370c, ii) shorter half-life because sc-fv but not fab-fragments are eliminated via kidney, iii) elimination of tnf-immunocomplexes may be improved by opsonization mechanisms which requires some structures of the constant region (as in fab or mab). in summary, despite the encouraging in vitro experiments, the in vivo data suggest that sc-fv are not the right strategy for neutralizing tnf in vivo. tnf production in endotoxin non-responder mice, effect of a glucocorticoid antagonist g. iazgtr jr, e. duda, j. ol~th, e. husztik, g. iazar glucocorticoids inhibit lipopolysaccharide (lps)-induced tnf production and tnf can stimulate pituitary adrenocorticotropin secretion, resulting in the release of corticosterone. earlier studies (1) reveales that the glucocorticoid antagonist ru 38486, nufepristone, sensitizes both endotoxin responder and nonresponder, c3h/hej, mice to endotoxin lethality. we have also shown that the glucocorticoid antagonist ru 38486 sensitizes endotoxin-tolerant (endotoxin-pretreated) and normal mice to the lethal effect of septic and endotoxin shock. on this basis, we set out to study the influence of ru 38486 on tnf production induced by endotoxin in endotoxin responder and non-responder mice. one and 2 hrs following lps injection, ru 38486 significantly increased the tnf concentration in the serum, liver and spleen of treated animals as compared with the control and methylprednisolonetreated groups. in tissue cultures, ru 38486 induced the tnf synthesis of myeloid cells and increased the tnf production of genetically modified hela cells, which synthesize tnf constitutively. normal and tumor cell cultures exhibited an increased sensitivity toward tnf in the presence of ru 38486. however, the same dose of lps did not induce tnf production in the serum, liver and spleen of endotoxin non-responder mice and this was not influenced by concomitant ru 38486 treatment. these studies suggest that ru 38486 aggravates lps lethality via factors other than tnf in endotoxin non-responder, c3h/hej mice. the cytokine response to a novel exnacellular mitogenic factor, mf, produced by the group a streptococci (gas), and to the streptococcal pyrogenic exotoxins (spe) a and b, was determined by in vitro stimulation of peripheral blood mononuclear cells (pbmc) obtained from healthy blood donors. the induction and kinetics of 14 different cytokines was studied at single-cell level using cytokine specific monoclonal antibodies and intracellular immunofluorescent staining. all three mitogens induced a massive ifny and tnfi3 response in 10-16% of the pbmc after 48-96 hours of cell stimulation. in contrast, il2 and tnfc~ containing cells was only detected in 1-3% of the pbmc. the induction ofil3, il4, il5, and ill0, was weak or completely absent. thus, the response indicates activation of th1 cells. however, illc~, illl3, illra and il8, were consistently found and gradually produced, peaking at 24 hems in approximately 5-8% of the pbmc. mf induced an equally extensive cytokine as well as proliferative inducing capacity, as did spea and speb, which suggests that also mf is a superantigen. a marked inter-individual variation could be noted between lymphocytes isolated from different individuals, both in the toxin induced proliferation and cytokine induction. this may be one explanation for the varying clinical severity noted in patients after infection with clonal gas strains. introduction -tumour necrosis factor (tnf) is released by mononuclear cells in response to inflammation and sepsis. it has been implicated as a possible mediator in inflammatory bowel disease (ibd) but its pathogenic role remains uncertain. this study assessed the relationship between tnfct and disease activity by measuring plasma and faecal tnf concelnmtions in an experimcntai model of ibd. methodologo' -colitis was induced in male wistar rats by intmcolonic instillation of 30rag 2,4,6-trinitrobenzenesulphunic acid in 0.5ml 50% ethanol (n=12). control animals received an isovohlmetric instillation of normal saline (n=12). faecal pellets were collected before induction of colitis (day 0) and throughout the experiment. after 8 days the colon was excised, ueighed and the colon macroscopic score (cms) assessed plasma tnf (ptnf) samples were assayed by bioassay and elisa. faecal samples were vortcxed in water and the supernatant removed for estimation of protein and faecal tnf (itnf) content (elisa). there is a significant increase in itnf on day 4 -#p=0 02 alld the total ftnf measured during the study correlatcs with the cms oll day 8 -p=0.04 there is no correlation between ftnf and ptnf. conclusions -therc is evidence of coloific tnf excretion by macroscopically normal animals and following induction of colitis there is increased faecal tnf which correlates with disease actmty. there is no significant elevation in bioactive or imnmnoreactive plasma tnf in colitie animals this data would suggest that faecal tnf is a marker of colonic inflamrnatiort and serial tnf assessment inay be useful as an indicator &severity in ibd. to study the effect of in-vivo hypoxia/reoxygenation on cytokine elaboration by routine peritoneal macrophages. materials and methods: adult male cba mice (20-25g) were primed intraperitoneauy with either lipopolysaccharide (lps) 10ug]anlmal or saline. five days later, the animals were subjected to hypoxia (8%02) for 160, followed by 20 of normoxia. harvested peritoneal macrophages were plated in-vitro, culture supernatants were collected at 20, 40, and 60 and assayed for tumor necrosis factor (tnf), prostaglandin e2 (pge2) and nitric oxide (no). control animals underwent the same procedures, but were maintained in normoxia (21%o2 disease 5, berlin, germany the precise mechanisms of reactivation of latent human cytomegalovirus (cmv) infection are still unknown. apart from the permissive effect of diminished cellular immunity there is evidence of a cytokine mediated cmv reactivation as it is known for other systemic virus infections (e.g. hiv). we found that tumor necrosis factor-alpha (tnf) -in contrast to all other cytokines tested -can stimulate the activity of the cmv-ie-enhancer/promoter region in the human monocytic cell line, hl 60. we wondered whether our in vitro results were actually reflected by the incidence of cmv reactivation in diseases which go together with high tnfalpha levels. cellular immunodeficiency as well as at least temporarily increased tnf levels are known to occur in septic disease. we tested for the presence of cmv infection in peripheral blood mononuclear cells (pbmnc) by means of 24h centrifugation culture, immunocytological detection of different cmv antigens (apaap-technique), and pcr technique (cmv-ie dna). in striking contrast to healthy controls almost all examined septic patients were positive, irrespective of the method for cmv-ddtection applied, including the less sensitive immunocytology. follow-up studies showed that cmv-antigen positive pbmnc ceased to be detectable in patients with good outcome once septicaemia had been overcome. to further back up our hypothesis, we looked for a coincidence of enhanced tnf-alpha serum levels and cmv antigenaemia (immunocytology) in some other diseases which are known to have either enhanced tnf serum levels or increased incidence of active cmv infection and found the majority of patients with b-cell chronic lymphocytic leukemia, liver cirrhosis, and common variable immunodeficiency to be positive for these two parameters. we now wondered whether, apart from cmv reactivation, tnf-alpha could also cause cellular immunodeficiency this way promoting cmv replication and spreading and eventually causing cmv disease. in summary, we were able to show that a diminished cellular immune response results from/n vitro or in vivo methylprednisolone (mps) is used to suppress both inflammatory and immune responses. il-1 receptor antagonist (il-lra) and tnf binding protein (tnfbp) are naturally occuring anti-cytokine proteins, possibly modulating inflammatory and immunological responses. to define mps modulation of cytokine and anticytokine responses, we examined effects of mps on il-lra production and tnfbp generation as well as il-i~ and tnfa production by human peripheral blood mononuclear cells (pbmc) stimulated with lipopolysaccharide (lps). pbmc were purified from the blood of healthy volunteers, then incubated with lps (10ng/ml) supplemented with different concentrations of mps (0, 500pg/ml, 50mg/ml). after 24hrs incubation, the supernatants were collected for rias of secreted cytokines. the cell lysates obtained by 3 cycles of freeze-thaw, were also collected for r ias of cell-associated cytokines. both the supernatant for secreted cytokines and the cell lysates for the cell-associated cytokines were subjected to rias for il-i~, tnfa, il-1 ra, and tnfbp-i (tnfsrp55). il-la, il-1 ra, and tnfbp were produced mainly as cell-associated forms in response to lps, whereas most tnfa was secreted into the supernatant of lps-stimulated pbmc. mps supplement resulted in reduction of il-la, il-lra, and tnfa production. il-la production was reduced up to 5.4+1.6%(50mg/ml) by mps in a dose dependent fashion, whereas about 50% decrease in tnfc, and il-lra production was observed at both high and low dose mps supplement. however, total tnfbp generation was not reduced significantly by mps supplement. furthermore, tnfbp secretion was increased 12 times more in the supernatant of lps-stimulated pbmc with 500gg/ml of mps. these data suggest that mps could effectively block tnf activity by both reducing its production and up-regulating tnfbp generation and that mps works as an anti-inflammatory drug as well as an immunosuppressant by modulating cytokine and anticytokine responses. in severe gram-negative sepsis, excess of proinflammatory cytokines is associated with increased morbidity and the mortality. we have found that immunocytes possess functional 13-adrenergic receptors (~-ar) which, when activated, down regulate the lipopolysaccharide (lps) induced gene expression and subsequent secretion of tnf-~ in vitro. in the present study, we used a lethal endotoxic model in mice to test the hypothesis that 13-ar stimulation will down regulate the gene expression thus decreasing the circulatory levels of proinftammatory cytokines and improve the survivals in severe endotoxicosis. the animals (iso group) were injected 3 times with !3-ar agohist isoproterenol (0.5 mg/kg i.m. and 0.5 mg/kg s.c.) 4, 2 and 0 hours prior to a lethal dose of lps injection (25 mg/kg i.p.). control group (ctl) received same volume of saline at the same times before the lps insult. at the appropriate time points after lps injection animals were sacrificed and venous blood was collected from the inferior vena cava. the plasma levels of tnf-cc and il-loc were determined by elisa. in the mortality study, each grou the data showed that isoproterenol significantly decreased the mortality and the circulatory levels of tnf-c~ and il-lcc (fig. 1) . these data suggest that 13-ar activation of immunocytes down regulates cytokine gene expression, decreasing circulatory eytokine levels thus reducing endotoxicosis mortality. these results provide a new pharmacological approach to reduce the excess of proinflammatory cytokines and mortality in sepsis. introduction: cytokines released from monocytic cells are thought to play an important role in the pathophysiology of sepsis. during the last few years several investigations have been performed aimed at modulating this mediator response. in the present investigation using a full blood model we have studied the effect of a standard immune globulin and an antitipopolysaccharide (lps) immune globulin preparation and a monoclonal anti-lps antibody in modulating the endotoxin induced cytokine response. methods: citrated blood from healthy human donors was incubated at 37 °c in rotating polystyrene tubes. samples were taken before addition of 1 x 103 ng/l e. col± endotoxin and after 2 hours. plasma was assayed for the cytokines minor necrosis factor alpha (tnf-~) and interleukin-6 (il-6) using elisa methods. the eodotoxin concentration was assayed by lal and end-point chromogenic substrate technique. the blood was preincubated with standard immune globuline (human igg from pooled plasma, gammagardr, baxter) or with an anti-lps immune glubuline (human anti-lps igg obtained by screening of blood donors, novo nordisk) in the concentrations of 4mg/ml or 8 mg/ml or with the monoclonal lipid a igm antibody ha-1a for ten minutes before the endotoxin was added. results: two hours after endotoxin addition markedly elevated tnf-~ and il-6 values were found.in the plasma. cytokine values at 2 hours: tnf-ec 1689 pg/ml (mean) and 1l-6 1123 pg/ml (mean). preincubalion with 8 mg/ml standard immune globulins reduced il-6 values by 49 % (mean) while there were no effects on tnf-c~ values at 2 hours. a slight reduction in il-6 values was also found with the lower concentration, 4mg/ml, but this was not statistically significant. preincubation with anti-lps immune globulins had no effect on il-6 nor on tnf-cc values. preincubation with the monoclottal lipid a igm antibody ha-1a in variable concentrations had no effect on cytokine release in this model. in this full blood in vitro model standard immune globuline reduced endotoxin induced 1l-6 release while tnf-cc values were unchanged.the anti-lps immune glubuline preparation and the monoclonal lipid a igm antibody ha-1a had no effect on the il-6 or tnf-ct release. the effect of anti-tnf treatment in severe haemorrhagic/traumatic shock was investigated in a conscious rabbit model. the jugular vein (for administration of substances and blood withdrawal) and right femoral artery (for measurement of bp) were cannulated and an aortic thermistor probe inserted into the left; femoral artery for the measurement of cardiac output (co). the following day, upto 40% of the estimated blood volume was withdrawn over 80-40 rain, to reduce bp to 40-50mmttg and co by 2/8 mad withheld for 60 rain. shed blood (60%) was then reinfused followed by lactate solution to give a total potential dreulat%ag fluid volume of 120%. zymoeen activated plasma (0,36gg/ml) was given 10 rain prior to haemorrhage and during blood r~n%eion, all ~,~ir, ak were killed at 72h and o~gane removed for histology. rabbita received either monoclonal antirabbit tnf (r6, rat igg1, 8mgkg i.e. n=7) or emine (nffi6), 60 rain prior to haemorrhage. the cardiovascular and biochemical changes during the haemorrhage and hypotensive phase (is. haemorrhagic stimulus) were similar in both groups. upon reinfusion, haematccrit and white cell count remained sigutfia~utly (p<0.05) higher in saline compared to r6-treated animals, indicating a relative haemoconcentration (i,e. reduced circulating volume) in saline animals. the lettkocytosis persisted in the saline animals upto 72h. plasma glucose, lactate and asparteto .m~-o transferase all rose similarly in both gorups but plasma ¢rea~!~+-e levels were markedly (p<0.05) higher in ealine ve r6 animals at 48 and 72h indicative of kidney damage, using a macro and micro. pathological scoring system, major organ damage was seen in the lung, liver and kidney which was significantly (p<0.05) attenuated in r6 treated animals. in the hmg and liver, this was primarily due to a reduced bleeding and pmn infiltrate and to an additional maintenance of tubular integri~:y in the kidney. hence, in tlais model anti-tnf treatment markedly reduced the biochemical and histological indices of severe ,, ~gan damage during liaemorrhage and ranerfn~ion. thermal injuries and systemic bacterial sepsis produce a wasting diathesis with anorexia and marked loss of lean tissue and body fat. endogenously produced cytokines, including interleukin-1 (il-1 ), are thought to mediate many beneficial as well as pathologic host changes that occur during burn injury and infection. to evaluate whether bluckade of il-1 in vivo would affect survival and attenuate the anorexia and cachex[a associated with a thermal injury and chronic fulminant gram negative infection, 24 sprague-dawley rats were studied. anesthetized rats were divided in 3 groups, implanted with alzet r minipumps and randomized to receive the following treatment for 14 days:i: 25 gg/kgbw/min of il-lcc; ii: 25 ~tg/kgbw/min of il-1 receptor antagonist (il-ira); iii: buffered vehicle. the animals were subjected to a 30% bsa, full thickness scald burn and the wounds then infected with 10 + cfu/ml pseudomonas aeruginosa. eight additional rats were anesthetized, but were not burned, and served as healthy controls. . il-hx reduced food intake transiently but was otherwise without effect however, il-lra significantly attenuated weight loss over the first 8 days and improved food intake between days 5 and 8. we conclude that blockade of il-1 after a thermal injury with superimposed infection does not adversly affect the progression of the disease, but does attenuate the anorexia and weight loss associated with this model. these data indicate that il-i receptor blockade may offer a means to minimize the morbidity associated with catabolic illness. tumor necrosis factor (tnf) and its downstream induced mediators have pivotal roles in the pathogenesis of sepsis and septic shock. the suppressive effect of pentoxyfillin (ptx) on tnf production may be of clinical importance. when peripheral white blood ceils were stimulated with either lps or staphylococcus aureus, pentoxyfillin inhibited tnf production. in contrast, no inhibitory effect was observed on the induction of il-6. ptx inhibited not only the tnf production of monocytes but also the tnf secretion of both granulocytes, and unseparated whole blood. the facs analysis revealed that the expression of cd14 antigen on monocytes was not influenced by pentoxyfillin. the in vitro tnf and il-6 producing capacity in septic patients were higher than in the control group, but decreased on subsequent days especially in fatal cases. administration of ptx (200 mg/day) in septic patients resulted in tnf and il-6 productions similar to those found in control donors. it also subsequently led to an improvement of the clinical status. a further improvement in apache score could be achieved by administration of pentaglobin ° (biotest)( 5ml/kg/day ). the prevention of lpsinduced in vitro tnf and il-6 production by pentaglobin ° could be demonstrated in in vitro experiments involving the use of whole blood rather than purified lymphocytes, very probably because of the presence of lps binding protein in the plasma. the level of soluble icam-1 in sera of septic patients was significantly higher (800-1200 ng/ml) than in normal individuals (50-150 ng/ml), but it decreased following the ptx and pentaglobin ° therapy. it is presumed that ptx and pentaglobin ° can antagonize eytokine production at different levels, resulting synergistic action being beneficial in the treatment of sepsis. albert szent-gy6rgyi medical university, institute of microbiology, szeged, hungary h-6720 ddm t. 10. e~i~im~/tii, septic sh0~ '. tnf alfa/pge, and somatostatln lands ji., alvarez j., gram m., llanos k., alcalde j., sanchez ja., balibr~d jl. taurine, a semi-essential sulphur-containing t:~-amian acid, promotes neutrophil phagocytic activit3' against yeasts and enhances intracellular killing of e.coli. paradoxically it protects against hypochlorous acidmediated biomembmne oxidatio~ and abrogates the pyrogenic effects of intracerebral endotoxin. it is therefore possible that taurine mediates some of these effects by modulating the cytokine cascade. aim to assess the cytekine responses in a rat model of intraperitoneal sepsis, pretreated with supplenmntary taurine, by measuring tnf and il-6 concentrations. methodology female wistar rats (wt 160-200g) (n-6 per group) were prefed with 2% tanrine, 3.33% glyciuc ( in tap water) or tap water (tw) for 15 days prior to caecal ligation aud puncture (clp). normothermia was maintained intraoperatively and the animals received 0.9% saline subcutaneously (3 mls/100g) post-procedure. animals were venosected by direct cardiac puncture at 6 or 18 hours post-operation and the blood placed in endotoxin-free tubes. centrifilgation and storage of plasma at -70°c was completed within 2 hours. tnf and il-6 were measured using wehi and b9 bioassays respectively. results (means ± sem) bioactivc tnf concentrations were not significantly different between clp or sham groups. however plasma il-6 estimation revealed a statistically significant reduction at 6 hours in tanrine-treated animals compared to glycino and tw controls ( objective: to evaluate the effects of allogeneic blood transfusion, thermal injury and bacterial garage on interteukin 4 (il-4), tumor necrosis factor alpha (tnf) production and host mortality and to study if the administration of thymopentth (thy) could affect these events. materials and methods: balb/c mice were transfused with allogeneic blood (from c3h/hej mice) and treated daily with thy (i mg/kg)(t+thy) . control animals were transfused but not treated (t). after 5 days systemic blood was harvested to determine the circulating levels of il4 and tnf. a second group was treated with thy for 5 days and then received a gavage wide lxl09 escherichia coli and a 20% burn injury (bg+thy). one hour post-bum, blood was collected to measure cytokins. control animals received the same treatment but thy (bg). a third group was transfused, treated with thy for 5 days and then received gavage and burn (tbg+thy). one hour post-burn, blood was collected to measure cytokins. control animals received the same treatment but thy (tbg). all treated and control groups had 8 mice/group. in separate groups (n=20/group), receiving the same treatments, mortality was observed for 10 days post-burn or transfusion. the production and release of oxygen radicals by phagocytic leukomytee is one of the most relevant and important functions of these substances in biology. when neutrophilic granulocytes or certain macrophages phagocytize they produce super oxide and hydrogen peroxide and form secondary products of these activated species. all of these substances are released in to the phagosome. it has been appreciated since 1977 that the amount of oxygen consumed by phagocytes in producing free radicals is prodigious. what has not been totally appreciated is the significant contribution this 02 consumption contributes to total body oxygen consumption. our understanding of increased oxygen consumption following sepsis is compounded by a paradox of increased cardiac index and a narrow av02 difference. it was mclean in a study reported in 1967 that showed an increase in cardiac index and a narrowing of the avo z difference in humans following sepsis. other studies documented similar changes in humans and it has recently been speculated that increasing o~ delivery might have a favorable impact on outcome. at least three different explanations have been put forward as to why there are possible alterations in energy metabolism during sepsis. these include decreases in oxygen supply, peripheral shunts and direct action o~ cellular mediators on o~ delivery, a provocative review by hodgkiss and nark in 1992 shewed that in experimental animals sepsis did not cause any evidence of bioenergatic failure in muscle i liver, heart or brain tissue. they further showed that the increase in serum lactate is mot necessarily due to cellular hypoxia. they conclude that cellular hypoxia and defects in energy producing metabolites are not the cause of metabolic abnormalities in sepsis. we set out on a series of experiments to try to explain the paradox in cellular metabolism and whether or not white cells might contribute to total body oxygen consumption during sepsis. the first set of experiments involved growing rat cardiac myocytes on tissue culture. pyruvate was added to the medium as substrata and physiologic levels of peroxide were also added simultaneously, carbon 14 labeled co~ production was measured as was nucleotides and degradation products from the cell. the peroxide induced a significant inhibition of pyruvate dehydrogenase. in addition, there was a loss of cellular atp and a corresponding rise in the release of inosine and adenine from the cell. we concluded from this first sst of experiments that physiologic levels of peroxide are a potent inhibitor of pyruvate dehydrogenase in isolated cardiac mitochendria as well as the cultured myocyte. we further showed that pyruvate dehydrogenase inhibition blocks the aerobic oxidation of glucose and leads te adenine nucleotide metabolism with adenosine and inosine release from the cell. other enzymes within the tca cycle did not appear to be specifically blocked by peroxide. this would explain the increase in lactate and the ability of the cell to continue to make high energy phosphate by entry of metabolites in to other entry points of the tca cycle, the second set of experimbnts was designed to estimate the magnitude of whole body reactive oxygen generation via nadph oxidase following granuloeyte activation invivo. using a guinea pig model baseline oxygen consumption was determined. ~iaphenyl iodinium (dpi) was used as a specific inhibitor of granulocyte nadph oxidase. animals were divided into two groups; those that received dpi and those that served as control, all animals were then injected with phorbcl myristate acetate (pma) intravenously. pma is a potent stimulus for granulocyte activation and subsequent reactive oxygen generation. whole body oxygen consumption measurements were then repeated after the p~la injection. in addition, arterial blood gas amalysis was performed, circulating neutrophils were collected and assayed for their ability to generate hydrogen peroxide and the r~ght lung was removed for wet and dry weight measurement. total body oxygen consumption increased by 25 percent after pma injection. arterial oxygen tension fell significantly in the control animal. neutrophil hydrogen peroxide generation rate doubled and lung water in the untreated animals increased by 25 percent. in this animal model we could show that granulocyte activation substantially increases whole body oxygen consumption to approximately 120 percent of control values. the nadph oxidase inhibitor, dpi, decreases granulocyte hydrogen peroxide generation invivo and prevents the pulmonary injury induced by pma. using this animal modal it is possible to transpose this to the human septic state. if we assume there are 5,000 neutrophils per cubic millimeter of blood, a 70kg man would have ~1.75xi0 u circulating neutrophils. when activated ixl0 ~ neutrophils generate one nanemole of hydrogen peroxide per minute. t~erefore, the circulating neutrophil pool could generate -1.75xi0 nanomoles of hydrogen peroxide per minute. this corresponds to an oxygen consumption of -4.2ml/ojmin by the circulating neutrophil pool alone. this does not take into account the production of oxygen by macrophages or the amount of oxygen consumed to produce nitric oxide. manipulation of oxygen delivery in human sepsis may or may not be beneficial. irshad h. chaudry, ping wang, and alfred ayala life threatening blood loss, due to soft tissue and bony injuries, is a frequent complication in trauma victims. although numerous organs and cells are adversely affected by hemorrhage, the focus here will be to discuss whether or not there are any differential alterations in splenic and hepatic immune functions. in particular, m~ antigen presentation (ap) and associated processes will be described since one of the important functions of the m6 is to activate resting t-cell lymphocytes by processing and presenting foreign antigens in a mhc class ii-restricted fashion. studies have shown that splenic m6 and kupffer cell (kc) ap was significantly depressed following hemorrhagic shock and remained so for at least 96 h alter resuscitation. the presentation of antigenic fragments associated with mhc class ii antigen itself was not decreased following hemorrhage, but the catabolism of antigens in antigen-presenting cells was decreased. this is likely due to a significant loss of cellular atp. although m6 ap was depressed in splenic, as well as kc, only kc showed an enhanced capacity to produce inflammatory cytokines, il-1, il-6, and tnf. this difference in response between kc and splenic md may be due to differences in the microenvironment in which these cell populations are found, as well as potential differences in the effects that hemorrhage has on the environment from which these cells are obtained. splenic m6 from hemorrhage-resuscitated mice exhibited a significantly reduced cytotoxic response, whereas kc showed an enhanced cytotuxic capacity. the increased kc cytotoxicity is probably mediated through the increased expression of cell-associated tnf and the release of reactive nitrogen. the enhanced production of reactive nitrogen may play an important role in the clearance of microbes translocated from the gut following hemorrhage. however, excessive release of reactive nitrogen by kc may have deleterious effects on the adjacent hepatncytes. such an effect could, in part, explain the reduction in active hepatocellular function, which is seen following hemorrhage-resuscitation. thus, hemorrhage induces differential effects on splenic and kc m6 populations; it decreases splenic m6 but increases kc capacity to mount a cytotoxic response. the depressed splenic md and kc ap was, however, significantly improved by posttreatment of animals with atp-mgcia, chloroquine, diltiazem or interferon-'/. these agents also decreased the susceptibility to sepsis following hemorrhage. thus, the use of atp-mgci~, dfltiazem, chloroquine or interferon-7 offers new therapeutic modalities in the treatment of immunosuppression and for decreasing the susceptibility to sepsis, which is observed following severe blood loss. the production of macrophages from bone marrow precursor cells is a dynamic and highly regulated process. the differentiation of myeloid lineage-restricted progenitor cells is initially controlled by interleukin-3, which drives the progenitors along a maturation pathway that leads to their response to specific lineagerestricted colony-stimulating factors(csfs) we have previously hypothesized that thermal injury can initiate a self perpetuating cycle of production of defective immune cells: thermal injury can initially affect circulating immune cells. then these cells, by unbalanced production of colony stimulating factors and other mediators, can cause a derangement of hematopoiesis resulting in an abnormal production profile of tnf, il-i, and il-6 and cytotoxic activity of bone marrow derived cells. the results of our studies so far have supported parts of this hypothesis. il-3 and il-3+m-csf treatments increased the production of tnf in burn 4 day macrophages and progenitor cells compared to normal cells. il-3+m-csf+il-10 increased the production of il-6 by burn progenitor cells compared to normal macrophagee. preliminary results for the ratios of tnf/~ actin mrna indicate that il-3+m-csf increased the mrna for tnf in the 4-day macrophages derived from burned animals compared to macrophages derived from normal animals. ratios of il-6/~ actin mrna indicated that il-3, il-3+m-csf, and m-csf increased the il-6 ~rna in progenitor cells from burn animals compared to progenitor cells from normal animals. these preliminary results support our hypothesis that bone marrow derived macrophages and progenitor cells from burned animals act differently compared to cells from unburned animals regarding the production of inflanunatory cytokines. more specifically, the results suggest that the different cell types are regulated in different ways by cytokines, and cells from burned animals are regulated differently than cells from unburned animals. it i,~ to ~ssume that the translneatinn is due to a ixx~h,v functioning ~astroiutestlnal surface pro|ection system, which leads to an unacceptably high load ¢ln the imrsunc dcfcncc system, partlcul~ly the local system in the gastrointestinal wall, leading tu exh~ustiou of ihi~ system. tlae g&~lrointesliual i]'ac( can be regal'deal a.s a t~2servojf of appr 200 m ~ separating 10 i~ eucayotic ceils of the hosl ttom l0 ~4 bacterial culls. in the human body the the myriad of cndngan¢nts micrix)rganistns, exogenous itl,~ttdittg micro~ganisms altd taxies m-'e sepia"areal li'otll lhe rest of the body by a simple epithclhd layer. the barrier function is heavily tlcpcnding tm =~ proleclive layer cmtsisting ie. surl~:emts, nluclts, probiotic bacteria, and ,~ub~tances with strong antioxid~lt, antipfotease al~d other i~rotective eft'ecru, the surfaet~ml htycr consists at tile besl if1 i(.) bimolecular layers, fl is thus very iltil~, the epilhelial layer is renewed every 24 hours , and tile tanuwer of the mucus is ~so fa,~t, all attdphy of die epithelial layei induced by g&sttointestinal st~'vation does also include the mu~usprndocing cells, gnhlet cells, which leaduhg lu slmnage aad luwer quality of gi mucus, altered viscoelas,jc properties and reduced protection, ]~'obioiic bactcrla keeps the ppm's low, produce nutrients for the intestinal mllcosa, eliminates tuxia~ and stimulates the local immune system, the gl surface is conslanlly under hc wv wearing by ingested bacteria and their enzymes, ingested substaucas, chefftie,'ds at~d toxins. 'll~e n~tective flora is reduced in disease, by stress and by cm'clcss antibiotic therapy. wc have made attempts tn recondition the i,,.astroiniesli!l~d. tnucosa by rcstlpply of surfactants or sttrfactanfliko glyc~lipids, gels with pseudomuein [until.oil, and probiotl¢ laetobaeilli. by supply of glycolipids we have bt:cn able m prevent and heal intlammatory and tlleerallve injuries (and io p)'cveut microbe transhteatkln) in uppe~' and lower (.31 u'acl, this c~al be fu~'ther augmentented by shnultaueos supply el' a p.~uedomucin,likc gel like pectin. ()at contain~ one hundred times more ot membrane lipid$ th~a =nay other food, much of fiber, ~spceially watcrsnlnhlc betaglucaal~, at~d has an close to ideal amino acid cornpo~ition, <')at, termented by species-specific lactobacilli, has a strol~ ability to prevent local inflammatory and ulcerative chunges,transh)cadnn and sepsis in experimeut~d attimals mid to pl~ven~ revert mof in ba,nan.~.lt seems us, that it the mucos~lnucus/probiotic bacterial filnclinns cat'= 1%' restated by mucosa reconditioning, even in severely sick iudividuals) the hx:~d iuuuuue ~y,~teme will be capable tu pr~)tec! the ix~y ag~lls~ sepsis ~nd mof. the impaired immune response associated with multiple system organ failure (msof) has been most widely studied following surgery for inflammatory disease and trauma. the majority of late deaths following trauma are due to infection and msof. not all patients with multiple organ failure have concomitant infection present, yet most have been septic at some time in their hospital course. their generalized inflammatory condition often persists whether or not organisms have been recovered. immune failure probably precedes msof with or without overt infection and indeed may perpetuate such widespread sepsis. macrophage tumor necrosis factor-alpha (tnf) production is thought to represent an important pathogenic mechanism by which this is mediated. cecal ligation and puncture (clp) is a clinically relevant murine model of intra-abdominal infection and sepsis. serum tnf and endotoxin levels, and peritoneal macrophage tnf production are only slightly elevated in this model even in endotoxin sensitive animals. mortality in this model, therefore, does not appear to be attributed to overwhelming endotoxemia and/or tnf production. it has therefore been postulated that tnf and other cytokines may act in a paracrine fashion to cause local injury. tnf messenger rna (mrna) expression was therefore measured and found to be increased in the lung and liver following clp. a different pattern of expression was seen after endotoxin administration, characterized by a more rapid rise and fall, and enhanced tnf mrna expression in the spleen as well. route of spread of infection as well as the type of stimulus may determine the organ specific cytokine response. these data support the concept of locally produced tnf as a contributing factor in tissue damage and multiple organ failure during sepsis. the estimation of histamine's role in sirs changed over the years based on increased knowledge in shock pathogenesis, evidence later found to be faulty, and the discovery of other, more fashionable mediators. over the decades, the prevailing view has been that histamine is a noxious mediator contributing to the fatal outcome of shock. the analysis of experimental data on exogenously administered, endogenously released and/or newly formed histamine in septic/endotoxic shock suggests that histamine contributes to the early cardiovascular changes via hi-receptor vasoconstriction thus excerbating (directly and indirectly) the effects of catecholamines. in the later phase of septic shock (low out-put, high resistance states), however, it can be assumed that histamine exerts strong vasodilator and positive inotropic effects via h2-receptors. these effects are considered beneficial in counteracting or attenuating the effects of sympathetic responses of catecholamines and other mediators. thus, a blockade of the early reaction with hi-antagonists and a stimulation of the h2-receptors by h2-agonists are worthwhile therapeutic approaches. shock produc~s ~ij alterations, an encrgy crisis and eventual c~ll damage, however, shock in itse|f do~ not lead freqoenfly to r~mote organ damage. in animal ~xpcrhnents and clinical expariez:ces in korea and vietnam, shock i~r s¢ was not associated with lung damage, renal failure or ards. the same is true with g.l bleeding. however, when shock is due to trauma or associated with tlssus injmy, then organ malfunction m~d damage is frequent. shock is bclleved to increase the frequency of infection, but clinical evidcnc~ for this is scant. hemonhsgio shock akme is an unusual o~orrenee dinica[ly. immune dysfolletion dons occur witl~ experhnental hypovolemic shock and in pailcnts with hypotcnsion or after receiving blood ~ansfosions. tbc most significant factor affecting reoorrcnce of resented ca,lorectal liver mclastasis was hypotensivc episodes during operation. blood transfosions depress immune function, improve transplant ~raft survival and increase ttll'rmr rec~r'tcnos ill co[eli and head and nc~k cancers. in experimental hemorrhagic shock, we found decreased response of [ymphocytes to mitogens, el:clad mixed lymphocyte reactilln and i[,-2 decreased. othe~.x hsvc found decreased macrophage and t and b ¢¢lt function, deci~ased re,ease of/l~i, increased poe v depressed macrophagc antigen presanlatiota, increased ien y, decreased il-2, 3 and 5, and shared mscrophags funclion with loss nr f and c 3b r~ccpto~s. this is inhibited by chlornqoine, ibuprofen, dihiazem, and atp. shock may activate ncutrophils contributing to ilappitlg in cspillarics, no refluw, increased adhesion, cytotoxicity and organ damage. this may be related to decreased clearance of bacteria after shock. t~lls shock, hypotcnsion and decreased microcirculatory blued flow initiate or set the stage for immune dysfmtction. they, along with tissue injury trigger inflammatory cascades. this also contributes to immunologic dysfonctitm, which is associated with increased lufoction. thus altered bh,od flow and mediator cascades are bolh involved, the question now is can wc rapidly improve microcirculatory blood flow, and dg'masa inflsrmnatory fosponss, which is also necessary for survival the effe~ of ~nterferon ~amma @n wour~ healing was ewaluated ~n a routine mo~el. ~mma ~n~erferon we8 g~ven in doses of ~37.5 to ~2,500 u~%$ synchronous with ereatio~ of a pa~splnal woumd then daily for is ~lays. ~mteet~on ~amma impaired tensile strength a% all ~oses relative to control. m0zphology suggested ~reas~ ¢oll~gen deposition and r~du~-~ neovag~ulaeit¥ ~n t~eat~4 animals, interferon ~amma was a2so ~valuated in a murine mo~el of cecal llgatlon and punneure. one hundred to 22,s00 uni~ vere ~iv.n following ~nju~ an~ than daily. interferon qamma was associated with inco:ea~ed mortalit~ and earlier death a~/a~n ~n a dose dependant manner (p< 0,05). afmlnistra~ion the proinfiammatory cytokines tumor necrosis factor-c~ (tnf-c0, interleukin-16 (il-ii3), interleukin-6 (il-6), and interleukin-8 (il-8) have been implicated in the pathogenesis of the multiple organ dysfunction syndrome (mods) following shock and trauma. these mediators which are released in high concentrations by activated macrophages, endothelial cells, and connective tissue cells cause a systemic inflammatory response syndrome (sirs), thus leading to a shock-like state and tissue destruction. recently, a number of proteins have been characterized in in vitro and in vivo studies demonstrating potent anti-inflammatory properties. these latter cytokines include interleukin-4 (il-4), interleukin-10 (il=10), transforming growth factor-6 (tgf-j3), and the newly sequenced interleukin-13 (il-i3). they are considered antiinflammatory, since in vitro studies using purified macrophage cultures or whole blood assays revealed an inhibitory effect of these antmnfiammatory cytokines on ~he synthesis and release of tnf-cl and il-u3. in addition, they reduced the severity of disease and reduce plasma levels of tnf-c~ and il-ii3 when administered to animals with infection or inflammation. furthermore, naturally occurring substances have been described that specifically neutralize the bioactivity of il-1b and tnf-a. the il-1 receptor antagonist (il-i ra) is related structurally to il-i and binds to the same cell-surface receptors, but does not stimulate the cell. in animal models ore. cob-induced shock, inflammatory bowel disease, and peritonitis, injection of il-lra significantly reduced the severity of disease. the cytotoxicity of tnf-c~ can be inhibited through two types of soluble tnf receptors (stnfr) type i (p55) and type ii (p75). these stnfrs are proteolytic cleavage products of the cell-bound tnfreceptors. when given to baboons to combat e. coil-induced shock, soluble receptm's to the tnf type [ receptor decreased the severi~ of the shock-like state. it has been well accepted that shock and severe injury result in an increased release of proinflammatory cytokines with a high incidence of sirs and mods. our studies investigating plasma levels of anti-inflammatory mediators (il-4, il-lra, stnfrs) in patients after severe injury further suggest that shock and injury lead to an activation of anti-inflammatory processes with altered plasma levels of these mediators. however, while plasma levels of il-4 and 1l-lra were significantly increased in trauma patients compared to healthy volunteers, stnfrs did not differ from control samples. thus, shock and severe injury result in different activation patterns of anti-inflammatory processes. m,l. rodrick, boston, b~usa following severe thermal injury significant suppression of the immune response has been demonstrsfed. these changes are associated with increased susceptibility not only to co,on human pathogens, but to organisms not normally pathogenic in the uncompromlsed host. early observations included prolonged graft rejection and skin test anergy in patients following burn injury. work from our laboratory and that of numerous others has shown that the~e is a profound £mmunoeuppresslon following severe thermal injury. i~mune deficiency has been identified in these patients hyz ii lack cf t nell responses to mitogens, 2) early decreases in total t and helper t cells in the peripheral bloo~ 3} lack of response to tetanus toxoid i~uniza~io~ in spite of increased non-speoifi~ i~k~unoglobulin praduction and 4) severe and prolongei deficiency of interleukin-2 (il-2) prcductlon by peripheral blood mononuclear cells (pbmo). this il-2 deficiency could be an underlyin~ cause of all the afo~ementloned immune defects by failing to activate both helper and suppressor t cell functions. mo~a recent work has focused on the molecular mechanisms of this il-2 deficiency end shown that the defect lies post-transcrlptionally since mrna for il-2 is also depressed following thermal injury. we have also investigated the potential of a defect in ii-2 receptor status on peripheral blood mononuclear uolls from patients fqllowing burn injury and in a murine model. in both oases no decrease in il-2r was found either by phenotypic markers or by funational assays. another hallmark of thermal injury is hyperao~ivatlon of proinflammatory ~ytok~ne secretion, which may play a significant role in multiple organ failure following burn injury. therapy o~ major burn injury may reasonably be aimed, therefore, at up-regulating t cell ~unction and down-regulating hyperactive secretion of proieflaam~ntory cytokines. the majority of patients dying of bums succumb to sepsis which coincides with dysfunction in the specific and non-specific host defences. generally, concepts relating to the mechanism(s) of specific immune failure in thermal trauma imply that inhibition of t (cd4+) cell responses is imposed by the injury or infection-related factors. however, the same factors elicit strong biological reactions within the lymphoid compartment. recent evidence indicates that systemic activation is inherent in the burn patient. to establish the relation of molecular and cellular events to the development of post-bum anergy we examined the state of and the capacity for t cell activation with regard to: 1) occurrence of activation-induced cell death (aicd), 2) activation-related phenotypic and functional_ diversity in the pool of peripheral cd4+ t cells. initially, (up to 7 days post-bum), peripheral blood lympliocytes from severely injured (> 35% total body surface area) patients exhibited high levels of constitutive expression of surface receptor for ]l 2 (cd 25) and spontaneous blastogenesis. the presence of activation-related t cellproducts in bum plasma was also apparent. subsequent impairment of the t cell receptor (tcr)-regulated t cell responses in vitro was accompanied by significantly increased dna fragmentation that is associated with cell death by the mode of apoptosis. using molecular markers we established that flesh peripheral blood ceils from immunosuppressed patients also contain large numbers of apoptotic cells. fluctuations in the number of viable (pi-) peripheral blood lymphocytes involved primarily cd4+/cd45ro+ (memory) subset of t ceils. the above observations suggest that thermal trauma-associated t cell anergy develops through aicd, a phenomenon commonly associated with the tolerogenic activity of bacterial superantigens. persistence of staphylococcal infections in the burn patient may support this assumption. response following trauma jane shelby, ph.d. the immune system is integrated with other physiologic systems, and is exquisitely sensitive to changes in nervous and endocrine systems changes following traumatic stress challenge. the immune, nervous and endocrine systems interact via both direct and indirect pathways which utilize neuro and endocrine hormones, neurotransmitters, neurepeptides and immune cell products. it is now known that the immune system may be affected by all of the neuroendocrine products produced during a stress response, with evidence for innervation of iymphoid organs, lymphoid cell receptors for neuroendocdne products, and leukocyte production of chemicals which are virtually identical to certain neuroendocdne peptides (acth, endorphins). trauma induced alterations in the equilibrium of various neuropeptides and neuroendocdne hormones have a significant impact on immune response potential, affecting control of proliferation, differentiation and function of immune cells. for example, the neurohormone melatonin is thought to be a natural antagonist to counteract glucocorticeid associated immunosuppression resulting from stressful challenges, such as surgery and trauma, plasma melatonin levels are known to be significantly reduced in burn patients. the administration of exogenous me[atonin improved cellular immune response following burn injury in an animal model. melatonin was also shown to have in vivo cytokine regulatory activity, increasing the potential for il-2 secretion and downregulating excessive il-6 and ifn~ in burn injured, stress susceptible mice. the regulatory interactions between the immune, nervous and endocrine systems provide mechanistic pathways for trauma associated immune dysfunction. increased knowledge of these interactions will enhance the potential for the design of novei clinical interventions to improve immune response and decrease the risk for infection in trauma and surgical patients. . animals receiving e were given a single dose daily of either 2.4 g/kg of e in a 20% solution by garage (ge), or 1.25 g/kg of sterile ive in saline. four hours following the last dose, bum animals were subjected to a 30% body surface area bum injury to their dorsum. twentyfour hours following injury, the animals were sacrificed and spleen cells were harvested for assessment of lymphocyte function. splenocytes were prepared by mincing the spleen, followed by incubation on glass petri dishes to remove adherent macrophages. non-adherent cells were then tested for proliferative response to t-cell mitogen concanavalin a (con a) and b-cell mitogen lipopolysaccharide (lps). data were analyzed by anova. results: chronic alcohol exposure and burn injury independently inhibit lymphocyte response to con a but not to lps. the combination of e plus bum injury, however, pmfouedly decreases this response to both con a and lps as outlined in the this data clearly identifies the synergistic impairment of immune function produced by ethanol and bum injury. it is furthermore apparent that ibis effect is gut mediated and that gastrointestinal exposure to alcohol is necessary to produce this effect. further studies will work to identify cellular and subcellular mechanisms to explain this effect. in experimental animal studies and investigations on human volunteers endotoxin infusion is mgulary accompanied by the release of the cytokine tumor necrosis factor a (tnf-~) determined by elisa technique. in patients with menigococcal sepsis also elevated tnf-a values have been found using a functional assay. we have studied the role of tnf-et in surgical icu patients with sepsis. using functional technique, we were not able to detect tnf-~ activities in the patient plasmas. when this cytokine, however, was determined by immunochemicai technique (el1sa) elevated tnf-e~ values where frequently oberserved. in order to further elucidate these observations, we studied shedding of tnf receptors in the patients. in these studies, we noticed that shedding of tnf receptors oecured regulary in the patients. at the time of diagnosis, soluble tnf receptor p55 and p75 were both 2-3 fold higher than values found in plasma samples obtained prior to die diagnosis of sepsis. we also observed that the sepsis patients revealed higher maximum values of p55 and p75 during the icu stay compared to values found in surgical icu patients without sepsis. these observations indicate that soluble tnf receptors are available in sufficient amounts to bind tnf-ot which is released in surgical patients developing sepsis. this mechanism may explain why functional tnf-c~ was not detected in the patients. institute for surgical research, rikshospitalet, the national hospital, university of oslo, 0027 oslo, norway. decker, d., sch6ndorf, m., bidlingrnaier, f., hirner, a., yon rfcker, a. the advantage oflaparoscopic cholecystectomy over conventional open surgical approaches in the treatment of symptomatic cholelithiasis has been shown convincingly by clinical studies. in order to facilitate comparisons of different surgical approaches, we evaluated the cell biological characteristics of tissue trauma by measuring changes in various cell surface markers on leukocytes and eytokines in plasma as a possible means to assess tissue trauma in choleeystectomy. patients recruited into our study had experienced at least one typical bifiary colic, had ultrasound-proven cholelithiasis (stages 0-ii according to me sherry), were 35-55 years old, and presented for elective choleeysteetomy. patients could choose between laparoscopic and conventional eholeeystectomy after being informed about the advantages and disadvantages of each procedure. cell surface markers on leukoeytes were determined using whole blood techniques with the help of commercially available fluorescent monocloml antibodies and flow cytometry. shed cell surface markers in plasma and cytoldnes were measured with the help of sandwich-elisa kits. blood samples were drawn 24 h before surgery, immediately before incision (after anaesthesia), 2 h and 24 h after incision. seventeen cell surface markers were examined on different cell populations and cellular subsets in 18 laparoscopic and 15 open-surgery patients. three soluble cell surface markers and six cytokines were monitored. by statistical analyses (multivariate regression analysis, student's t test, wilcoxommann-whituey's rank sum test) the six markers/cytekines that best distinguished open surgical from laparoscopic procedurea were determined. these were 1. the interleuldn-2 receptor and im soluble form (cd25/scd25); 2. the activation antigen fd-1 and its soluble form (cd30/scd30), a member of the nerve-growth-factor receptor family; 3. the cd45ro epitope which characterizes t memory ceils; 4. the trausferrin receptor cd71; 5. the soluble adhesion molecule icam-1; and 6. the cytokines interieukin-6 and interleuldn-8. on the basis of these results, a tissue trauma activation (tta) index was calculated by combining the marker/cytoldne measurements by simple multiplication. anaesthesia and pre-ineision maneuvers did not significantly change cell marker or cytokine levels in either surgical approach as compared to 24 h before surgery. 2 h after incision the tra index in open cholecystectomy showed a distinct 22-37 fold increase, whereas in laparoseopic surgery a mere 2-3 fold increase was noted. 24 h after incision, the tra-index returned to near pre-surgery levels. in conclusion, our results demonstrate that changes in cell surface markers and cytokines can help evaluate the magnitude of tissue trauma in diffei'ent surgical approaches. the relationship between lymphocyte subpopulation changes after thermal injury and the increased susceptibility of burned patients to infection is unclear. in this study, we have attempted to correlate such subpopulation changes with the presence of infection in burned patients. peripberal blood from 19 patients was monitored for lymphocyte subpopulation changes three times weekly for three weeks postburn and weekly thereafter for three additional weeks. mean bum size was 44.7% (range 24%-84%) of total body surface and mean age was 38 years. infection was diagnosed by carefully defined clinical and laboratory criteria and its presence or absence noted each time blood was drawn. samples taken when patients had wound infection, bacteremia, or pneumonia were compared with samples taken in the absence of systemic infection. whole blood samples were stained with four monoclonal antibodies, the red blood cells lysed and the leukocytes fixed and analyzed by flow cytometry. for each patient sample, the proportion of lymphocytes falling within the light scatter gates was determined as the percentage of cells negative for cd14 and most strongly positive for cd45. this percentage was used to correct each sample for the presence of debris or nonlymphocytic cells. the proportion of cd4 and cd8 positive cells was slightly greatc~ in the samples from infected patients, while the proportion of b cells (cd20+) was unchanged and nk (cd16+) cells were decreased by ahnos[ 50% compared to sampie~ li'om uuiuleclcd patients. the percentage of cells positive for cdilb (c~ integrin) decreased sharply and cd4ro (memory cells) decreased slightly in samples from infected patients while the expression of the lymphocyte homing receptor and cd29 were unchanged. cd25 (il2 receptor) and cd69 (early activation marker) were significantly increased in the samples from the infected patients while hladr was unchanged. these changes in lymphocyte phenotype correlate with the presence of infection. if they closely precede or occur during the early development of infection they may be valuable clues to the mechanism of susceptibility following thermal injury. trauma patients are subjected to an immediate massive impact on their host defense integrity due to the combined effect of tissue trauma, shock and endotoxemia. cytoldnes are playing a crucial role within the course of an impaired cell mediated immune response (cmi) resulting from a disruption of intact m%/tcell interaction. the current study was undertaken to further elucidate the mechanisms of dysfimctional cmi following major burn and mechanical trauma -via comparative analysis of mrna expression and protein release. the major regulatory levels for different cytokines were determined in mitogen, respectively lps stimulated peripheral blood mononuclear cell (pbmc) cultures of trauma patients on consecutive days (13) t, 3, 5, 7 and 10 post injury. we analyzed the cumulative data for interleukin-1 beta (il-i[3), il-2, il-8 as well as tumor necrosis factor alpha (tnf-~) and saw a considerable impairment of the protein release in the stimulated pbmc cultures until d5 post-trauma and recovery thereafter. *p < 0.05, ** p < 0.01 vs control comparing the autoradiographies of the specific cytokine mrna expression with the protein release in the supernatants, we saw a good correlation between mrna signal intensity and protein synthesis for il-8 and 11,-2, suggesting that for these cytokines the main regulatory mechanisms are located at the pre-/transcriptional level. for the other cytokines investigated one has to suppose posttranseriptional mechanisms. the analysis of our data clearly indicates a severe impairment of forward regulatory immune mechanisms following trauma. most likely the regulatory mechanisms, that are involved are greatly different among the cytokines investigated. it may be concluded, that depressed cmi responses post-trauma are partly due to an impaired pro-inflammatory cytokine production. the severity of the injury (iss) correlated with the development at multiple organ failure (mof-score; r=0.612). the levels of mediators and markers of the inflammatory response were generally higher in the more severely injured group (iss>30, n=16). i1-6, 11-8, g-csf, fpa, and c3a -levels differed significantly (p<0.005) between the iss-groups (>-< iss 30) at the time of admission, whereas on day 3 tnfa, c3a, 11-6, and ealpi showed significant differences. beyond the first week, major differences were restricted to pge 2 and c3a. the formation of two groups with respect to later multiple organ failure (mof < 3; mof > 2 n= 14) yielded similar results. leukocyte-facs analysis revealed significant differences mainly in the cd14 (monocytes), cd3/cd25 (i1-2r + t-cells), and cd29/cd4 (th 2calls) populations. summarizing our findings we were able to detect some alterations in the surface antigens of immunocompetent cells. the inflammato d response, however, seemed to be more pronounced and correlates wi~ the further clinical course. using an experimental bum model in rodents, we have demonstrated that administration of a full thickness, scald burn involving 20% or more of the total body surface area (tbsa) elicits systemic responses which are characterized by numerous alterations in t-ceu function (i.e., lymphokine production and contact hypersensitivity (ch) responses) plus an enhanced susceptibility to bacterial infection. in the present study we questioned whether the apparent systemic effects mediated by large burns would be elicited as site-specific alterations in immune function following administration of small area burn trauma (5% tbsa). following a 20% tbsa burn, ch responses to contact sensitizing antigens were found to be altered. the depression in ch responses could be induced independent of the site used for topical skin sensitization. following a 5% tbsa thermal injury, development of ch responses were affected in a site-specific manner. immunization of 5% tbsa thermally injured mice in a site near the position of the burn resulted in depressed responsiveness, whereas immunization through a contralateral site resulted in responses that displayed both the intensity and kinetics of a ch response equivalent to sham-bumed mice. similar systemic and site-limited changes in lymphokine production were observed with 20% and 5% tbsa thermal injuries, respectively. a 20% tbsa injury affected the lymphokine producing potential of all cells regardless of which lymphoid tissue the cells were isolated from. the effect of a 5% tbsa burn was significant but site-specific. thus, ceils from lymph nodes receiving drainage from thermally injured tissue were specifically affected, whereas lymphokine production by cells from lymphoid organs receiving drainage from unaffected skin was normal. it was concluded that modulation of lymphokine production and cellular immune responses may be a normal consequence of burntrauma regardless of the size of the burn. changes in immune competence can be mediated either regionally or systemically in direct proportion to the area of skin exposed to the burn injury. this work is supported by phs grant gm46899 and the office of navy research n00014-92-j-1612. division of cell biology and immunology, department of pathology, university of utah school of medicine, salt lake city, ut 84132. post spleneetomy septic sequelae may be fatal, but the mechanisms remain unclear. the objectives ef this study were to assess the mortality from concomitant splen-'etomy and ]~eritoneal bacterial challenge and to elucidate the local cetkdar responses. 50 cd-1 mice were randomised to receive laparotomy and sham splenectomy (l) or splenectomy (s) with simultaneous ca'-cal ligation and "):mcture and the survival patterns assessed. subsequently, 150 cd-1 mice were randomised into control (c), l or s groups and peritoneal cells studied at 24 hours for bacterial phagocytosis and killi:~g, superoxide (02-) and tumour necrosis factor (tnf) production and macrophage activation vsing mac-i(cd-11b) receptor in~.ensity expressed es mean channel of fluorescence (mcf). these resides indicate that sf!enectomy predisposes to nrortal~ty from bacterial sepsis ia the early pos~ operative period compared to sham operated animals. failure ~f p'.acrophages to kill bacteria in the splenectomv group '~:cured in t?~e absence of impairment of oxygen freeradical or tnf pred:~ctien. the macrovh~ge ac!ivotion marker mac-1 was significantly reduced in both l and s groups and impaired phagocytosis of bacteria oceured in both operative groups compared to controls. laparotomy a!one reduces macrophage activity in terms of surface re:eptor mac-1 expression and !ingestive capacity. splenectomy however s~gnificantiy ~mpairs r-acrophage-wediated l~,acterial killing and this qefect rttav co~tribut~ sig~ifjcav'ly to th-~ dissemination of local infection and to n':ortalit). depts of haem~ tology & surgery, beaumont hosoital, dub!in 9,eire. introduction: loss of cell membrane integrity appears to be a common pathway of injury to tissues subjected to high-voltage electrical shock. the cell membrane is the most heat labile structure in the cell, and is also the most vulnerable to externally-imposed electrical forces. skeletal muscle and nerve cells are particularly susceptible to electroporation by clinically relevant electric fields. restoration of membrane integrity is essential for cell survival in victims of electrical shock. we have studied the effect of non-ionic triblock copolymers ( poloxamer class) on the transport properties of isolated rat skeletal muscle cells following electroporation-induced membrane disruption. 1-3 mm long adult skeletal muscle fibers were isolated by enzymatic digestion from the rat flexor digitorium brevus and maintained under standard culture conditions. they were loaded with the calcein-am dye and placed in a 37,c chamber for recording by real-time video confocal microscopy. the cells were subjected to 4 msec, 150 v/era, 1 a field pulses with a low duty cycle to allow thermal relaxation. peak temperature rise was 0,2.c. the uye content of the cell was monitored in real time. experiments were carried out in calcium-free phosphate buffered saline, with 1 mm mg%. experiments were repeated with 4 mm neutral dextran ( the aim of the present paper is to ascertain if thuracotomy induces a different pattern of variations of cytokines, immunocompetent cells and antibodies from laparotomy in the early postoperative period. 60 patients (34 males 26 females,mean age: 59.5_+5) 30 with gallstone disease and 30 with non neoplastic pulmonary disease were studied. none of these patients received blood transfusion, biological response modifiers, radiotherapy or surgery for at least 6 months before being included in our study. anaesthetic procedures were similar in all patients and none were matnourished. on the day of surgery and on the 1st and 7th postoperative days (pre, lpo, 7po) percentages of cd19, cd5, cd4, cds, cdi6 were measured by means of flow cytometry using moab., and levels of ig a, lgg, igm, ige. by nephelometry cytokine levels in peripheral blood(il-1, il-2, il-4, il-6, tnf) were measured in 10 pts. of each group by means of elisa using moab. _r. esults:variations of il-2 and il-4 were not s.s.. il-6 increased but differences between groups were not statistically significant (s.s). il-i decreased on 1po and increased on 7po in both groups but were only s.s. in the th.g., and therefore, the differences between groups were s.s (p<0.05).tnf decreased in the l.g. and increased in the th.g. on the 1po, the difference was s.s(p<0.05); on 7po, tnf decreased in the l.g. and decreased in the th.g. but these variations were not s.s. cell percentages decreased an lpo and increased on 7po, except for %cd19 cell that increased on lpo and decreased on 7po ,in both groups of pts. differences were not s.s. ig a, igm decreased and ige increased in both groups (p< 0.0i), but differences between them were not s.s. in contrast, igg decreased on 1po (p<0.01) and increased on 7po in both groups, but the decrease iu the th.g. was greater than in the l.g. twenty male children,aged from six months to 3 years,admitted for elective inguinal operation were studied. the operations were performed under balanced combined anaesthesia (fentanyl,thiopemtone,vecuronium, 70% nitrous oxide in oxygen) and blood samples were collected before flunitrazepam premedication,after anaesthesia,4 and 24 hours after anaesthesia. cells from the wound were collected with cellstick sponge which was removed from the wound 4 or 24 hours after anaesthesia. the study was approved by the local ethical committee. the percentage of neutrophils was increased and that of lymphocytes was decreased in perpheral blood after the operation.the values in the wound were close to the values found in peripheral blood. the percentage of t-lymphocytes (cd3) and helper-t-cells (cd4) decreased in peripheral blood being lower in the wound than in peripheral blood after the operation. the percentage of t-eytotoxic cells (cd8) also decreased in peripheral blood and was similar to that in the wound. b-lymphocyte (cd19) percentage was increased in pe~pheral blood after the operation and was higher than in the wound. the percentage of activated t-cells (cd3+hla-dr-positive cells) in peripheral blood increased while that of natural killer cells (cd3+cd16+leu19-pos) was increased just after anaesthesia being decreased at g and 24 hours after the operation. spontaneous lymphocyte proliferative responses didn't change while phytohemagglutinin a and concavalin a induced responses were decreased in peripheral blood samples 4 hours after the operation with recovery at 24 hours.pokeweed mitogen induced lymphocyte proliferative responses were decreased at 4 hours (p<o.01) and 24 hours after the operation we previously reported that plasma ige increased after traumatic injury. in this study, we measured plasma ige ievels in 40 trauma patients on hospital admission and 3x weekly in the intensive care unit to evaluate ige kinetics. patient characteristics were: mean age 36-+ 16 years (33 m, 9 f), mean injury severity score (iss) 26+ 10, sepsis (19 patients), sepsis syndrome (9 patients), ards (4 patients), renal failure (5 patients). ige increased in 28 patients (70 %). the mean increase was 527 ng/ml (range 0 to 7362 ng/ml; 95% ci was 160 to 895 ng/ml). increase in plasma ige was significantly greater in patients with sepsis syndrome (p=0.021) and renal failure (p<0.001). although mean plasma ige was higher with ards, pneumonia, hepatic dysfunction, and shock, these differences were not significant (p>0.05). plasma ige increase was not related to severity of injury by iss score (p =0.42). the mean day to highest ige was 9.1-+6.5. the day sepsis was first observed preceded the day of highest ige by 4.2+6.6 days. there was a significant association between the day of sepsis onset and the day of highest ige (p=0.018). eight of nine patients with sepsis syndrome had > 100% increase in plasma ige from admission. one patient's ige levels were normal (20-100 ng/ml) for 24 days and then increased to 7300 ng/ml over the next 12 days, after onset of sepsis syndrome. changes in ige plasma levels may reflect the action of cytokines, such as il-4, which concurrently regulate production of ige and il-1 receptor antagonist in a response to sepsis. sepsis remains a leading cause of late mortality in trauma and hs. although hs-induced bacterial translocation is supposed to be the major cause of sepsis and mof, depression of the res increases susceptibility to infection after injury. the purposes of this study were: a) to evaluate the res in the lung, spleen and liver after hs and subsequent hypertonic saline (hsl) treatment, and b) to document the patterns of phagocytic activity in these organs during 24 hrs. adult male wistar rats (250+_23 gin) were submitted to hs (sbp 40 tort) and after t hr (shock i hr) and 24 hrs (shock 24 hrs) hsl (nac17.5%, 3.5 ml/kg) treatment, 1131 e. coli (i 131) was injected into the portal vein 10 ~tci (n_>12). twenty minutes later, the lungs, spleen and liver were harvested and scintilographic counts obtained. data is depicted as mean_%+sem * p<0.05, ~" p<0.001 and statistical analysis was performed by analysis of variance and wilcoxon tests. one hr after treatment, lung 1131 uptake was increased and liver and spleen 1131 uptake were reduced compared to sham. twenty four hrs after treatment, all organs, except lung uptake, returned to normal values. radioautographic histological analysis revealed radiolabeled particles inside phagocytic cells of all organs. we conclude that pulmonary phagocytic activity increases after 1 hr of hs hsl reatment, diminishing by 24 hrs although still above normal values. in contrast, res suppression occurs in liver and spleen after 1 hr hs hsl treatment, returning to normal values by 24 hrs. these results may explain lung complications and immunosuppression after trauma. infusion of endotoxin as well as major surgery is followed by lymphopenia in peripheral blood. the purpose of this study was to investigate to which tissues the lymphocytes are redistributed in response to endotoxaemia and major surgery. in addition changes in lymphocyte subpopulations and expression of mecii was measured. lymphocytes were isolated from peripheral blood of 30 rabbits, labelled with 111indium-tropolene and reinjected intravenously into the rabbits, i0 rabbits received an infusion of escherichia coli endotoxin 2 ~g/kg, while i0 rabbits were subjected to a major sham operation and i0 rabbits served as a control group. the redistribution of lymphocytes were imaged with af gamma camera, and calculated with an interfaces computer before, and 2, 4 and 6 hours after major surgery or infusion of endotoxin or saline. interleukin-l~ and serum cortisol were measured. in addition we followed cd2, cd3, cdlla/b, cdis, cd20, cd44, mhcii and cd4/cd8 ratio. following endotoxaemia interleukin-lf~ increased significantly, following endotoxaemia as well as major surgery serum cortisol increased significantly. following major surgery as well as endotoxaemia there was significant lomphocytepenia in peripheral blood with a decreased cd4/cd8 ratio while the cd3 positive subpopulation increased. in addition there was a decrease in the expression of mhcii on the lymphocytes peripheral blood. the radioactivity of the lymphatic tissue in and around the intestine increased to 129% of initial values following endotoxaemia and to 125% following major surgery. the results indicate that endotoxaemia as well as major surgery induces redistribution of lymphocytes from peripheral blood to lymphatic tissue. among the lymphocytes staying in peripheral blood there was a decreased expression of mhcii and a relative decrease in cd4 cells compared to cd8 positive lymphocytes. in order to analyze the effects of immune suppressive substances on expression of mrna of interleukin-2(il-2) and interleukin-2 reeeptor(il-2r), this study was carried out. twenty male rabbits with comminuted fracture were used in the study. ten ml blood were taken at 0, i, 2, 4, 6 days after injury. the sera were tested for the effects on lymphocyte blastogenesis and induction of il-2 stimulated by concanavalin a(con a): the sera from the rabbits 2 days after injury were analyzed with sds-page gel eleetrophoresis, and divided into three groups by ultrafiltration (ufpi ttk, 30kd,milipore; centricon-10,10kd,amicon), that are less than 10kd, between i0 and 30kd, and more than 30kd. each group of the substances also was tested for the expression of il-2 and il-2r by the dot blot hybridization. the results showed that: i) all sera from the rabbits after injury had significant suppression on lymphocyte proliferation and secretion of il-2 by the con a-stimulated splenocyte in mice; 2) the sera from the rabbits 2 days after injury had more profound suppression than other injured sera; 3) there was a marked band at about 10kd in sera from the rabbits 2 days after injury, but nothing at the same position in normal sera analyzed with electrophoresis; 4) the substance with molecular weight of about iokd had more obvious suppressive action on expression of mrna of il-2 and il-2r than other groups substances, of which molecular weights are more than 10kd. it is concluded that: i) the sera from the injured rabbits can reduce immune response; 2) there is kind of substance, of which molecular weight is about 10kd, it is probable the main factor involved in the pathogenesie of postinjury suppression immune; 3} the substance can depress the expression of mrna of both il-2 and il-2r. research institute of surgery daping, chongqing, 630042 p. r. china acute ethanol uptake prior to injury modulates monocyte tnfo~, production and mononuclear cell apoptosis. g. szabo, b. verma, p. mandrekar, d. catalano monocytes (mo) have been shown to contribute to immunosuppression after both major injury and alcohol consumption. we reported that acute ethanol exposure of m(3 results in decreased antigen presentation, induces tgf-13 and pge2 while inhibiting inflammatory monokine production. we also showed that post-trauma immunosuppression is mediated by hyper-elevated mo tnfc~ and il-6. consequently, here we investigated rnonokine production in trauma patients (n=10) who had elevated (>o.lmg/dl) or had no blood alcohol level (n=t0) at the time of emergency room admission. none of the patients had chronic alcohol use history. met tnfc~ production from trauma patients with prior alcohol uptake was undetectable during days 0-3 post-injury in contrast to patients without alcohol exposure. furthermore, decreased tnf~x levels were found in alcoholic patients' mci after mdp or ifny + mdp induction. however, mcl tnfc~ levels during the 4-8 days post injury period became higher in alcoholic trauma patients. furthermore, over 9 days post-injury, alcoholic trauma patients showed significantly elevated mci tnfo~ production after adherence isolation, mdp, or ifn+mdp stimulation compared to patients without alcohol. these results suggest that acute ethanol uptake prior to injury decreases tnf(x inducibility in the early post-trauma period, but these patients' mo produce hyper-elevated tnfa levels later post-injury, thereby prolonging their cytokine shock risk. tnf ng/ml 0-3 days post-injury 9 days post injury stimulus ale. pt. pt 9.3 11.9 5.6 4.0 immunosuppression might also be increased by the elevated apoptotic activity found in trauma patients' mononuclear ceils, which was even greater in alcoholic trauma patients' cells. in non-alcoholic trauma patients' preactivated mo, in vitro acute ethanol (25-150mm) exposure resulted in a significant down-regulation of tnfc~ (p<0.001) and il-6 (p<0.01) production. in contrast, in vitro ethanol exposure increased the production of inhibitory monokine, tgfi]. these results provide both in vivo and in vitro evidence for the effect of acute ethanol exposure increasing immunosuppression and cytokine shock. the 'systemic inflammatory response syndrome' (sirs) with consecutive septic multi-organ dysfunction represents the major cause of late death following major mechanical and burn trauma. systemic hyperinflammation and concurrent depression of cell mediated immune response (cmi) render the traumatized host anergic, resulting in profound susceptibility to opportunistic infection. monooytes/macrophages (mo) play a central role within the host defense system in developing and manifesting states of injury, shock and sepsis. the mechanistic scrutiny of the synthesis patterns of crucial cccytokines appears to be a helpful tool to further analyse mo behaviour in the compromised individual. the objective of this study was to further dissect the characteristics of cytokine regulation in pbmc under stressful conditions, via analysis of the expression of cd14+ receptor, the proinflammatory mediator il-6, the macrophage activating factor ifn-5,, and neopterin (npt) a metabolite of activated mo. we investigated pbmc's on consecutive days 1, 3, 5, 7 and 10 after mechanical trauma of 9 and after bum trauma of 5 patients (mean age 38~17 years; mean iss 34±2 pts). in trauma patients we saw a massive increase of pha induced neopterin synthesis compared to controls. however, when discriminating the npt levels in the supernatants for the amount of mo stimulated, the npt output of the individual cell was lower compared to mo of nontraumatized individuals. interestingly there was a contrary coarse in the cumulative protein release patterns of il-6 and ifn-7 in mechanical versus burn trauma patients. wheras in burn patients ifn-y was decreased significantly (16+5 u/ml) compared to controls (58+6 u/ml) as well as mechanical trauma (59+12 u/ml). il-6 showed a significant suppression following mechanical trauma (662+58 u/ml) vs control (1266+91 u/ml) and bum patients. the rt~,na signal intensity for beth eytokines was in concurrence with the protein release in more than 85% of the individual patients investigated. from these data we can conclude that the inadequate low npt synthesis predominantly in bum patients appears to be a sign of cellular immaturity and is probably partly due to low t-cell ifno t signals. in addition we could state that the quality of trauma is apparently responsible for the different synthesis patterns of ]l-6 and ifn-q,. it has been postulated that bacterial invasion or endotoxemia are necessary for cytokine production following burn injury. we studied the organ distribution and kinetics pattern of il-fc~ (cell-associated il-1 agonist) in eutrophic rats subjected to either 20% tbsa cutaneous scald injury (bi), muscle scald injury of equivalent 20% tbsa (mbi), sham muscle bum (resection of skin only, up to 20% tbsa) (smbi), and sham cutaneous burn (sbi), followed by saline resuscitation (15 mukg i.p.). separate rats were infused with 15 mg/kg e.coli 0111:b4 lps or saline lv. unmanipulated rats were baseline normal controls. liver, lung, spleen, ileum, thymus, kidney, skin, and plasma were harvested at various time-points within the first 24h. tissues were frozen, weighed, homogenized, the homogenates centrifuged and the supernates assayed with a radioimmunoassay specific for rat il-l(z (detection limit 150 pg/rnl). il-lc~ was expressed as ng/g weight + sem (lowest detectable amount 1.5 ng/gwt). il-lo~ was constitutively present only in the skin (102 + 16.4 ng/gwt). cutaneous burn and sham cutaneous bum induced biphasic elevations of il-lcc in the liver and lung only, with maximal levels at 2.5h (in the liver, bi = 16.5 _+ 6.2 ng/gwt, sbi = 1.7 + 0.1 ng/gwt, p _< 0.05; in the lung, bi = 10.3 + 1.3 ng/gwt, sbi = 1.9 + 0.8 ng/gwt, p -< 0.002). of note, both bi and sbi rats had detectable il-i~ in the liver at timepoint 0 already (2 min real-time). these levels increased in parallel until 30 min and became eventually different by 1 log at 1 -2.5h. all other organs as well as plasma were below detection limits. muscle burn injury and sham muscle burn (skin resection) induced similar elevations of il-10~ in the liver at lh, indistinguishable from each other and from cutaneous burn. in contrast, lps challenge induced dramatic elevation of il-t~ in all organs tested except for the kidney; the spleen was the most responsive organ to lps-induced il-lo~ production. these data indicate that thermal or mechanical injuries induce very early and organ specific production of il-1 c~ in vivo by mechanisms other than endotoxemia. injury-induced complement and platelet activation may be involved as well as the neuro-endocrine axis, which may explain the low levels of il-lo~ induction observed in all rats at the very early time-points. trauma services, massachusetts general hospital, and department of surgery, harvard medical school. fruit, st, boston, ma 02114. j. f. schmand *#, a. ayala* and i. h. chaudry* studies indicate that i.v. infusion of the colloid hes in normal animals does not adversely affect non-specific immunity. it remains unknown, however, if lies affects cell mediated, specific immune functions after trauma and hemorrhage (hem). to study this, non-heparinized c3h/hen mice underwent midline laparotomy to induce trauma and were then bled to and maintained at a bp of 40 mmi-ig for 60 rain. the animals were then resuscitated with either 4 times (x) the shed blood vohune as lactated ringer's solution (lrs) or 2x lrs + lx 6% lies. sham mice were neither hemorrhaged nor resuscitated. at 2 or 24 hours post hem serum, peritoneal (pm~) and splenic macrophages (sm~) were obtained. bioassayes were employed to assess the levels of ii-l, il-6 ( alternatively pmqb showed no differences in il-1 release between all groups at 2 and 24h, while sm~ from the lrs + hen group showed a depression at 24h. tnf production by pm~ was depressed in all groups at 2h and remained so in the lrs + hes group at 24h. sm~b showed decreased tnf release values in both hem groups at 2 and 24h. in summary, the levels of inflammatory cytokines (particularly the values of circulating il-6) after trauma/hem are positively influenced by the administration of hes. this might be due to a protective effect on pmqb and sm~, but also on other cytokine producing cells, e.g. kupffer ceils. we conclude that hes is not only a safe, but also beneficial agent in the resuscitation of patients atler trauma/bemorrhagic shock. this study investigated endotoxemia and consecutlve immune response in 39 patients with multiple trauma (median injury severity score = 20,5). blood samples.were collected shortly after injury and after 0,5, 1, 3, s and l0 days. endotoxin was measured with limulus-amebocyte lysate test and the specific antibody content (sac) against endotoxins of the classes igg, igm and lga by elisa-technique. five antigens were used: lipopolysaccaride (lps) of e.coli (ec), lipid a of e.coli (la), lps of pseudomonas aerog. (pa), lps of vibrin cholerae (vc) and cx-hemolysin of staphylococcus anreus (oth). a nephelometer indicated the total concentrations of igg, igm and iga. differences were checked with wilcoxon-test and p<0,0s was considered significant. cross-reactivity was calculated with rank correlation coefficients. results: endotoxemia peaked shortly after injury (0-3h) at 0,425 eki/ml (median), decreased thereafter to 0,04 eh/ml at day s and remained on this level. sac oflgmclass increased to all endotoxins and peaked at day 3 revealing the lfighest level to la followed by pa (= 80% of la-sac), ec (= 65% of la-sac) and vc (= 40% of la-sac). lga antibodies increased as well but only slightly and not significant (exception: sac to la was elevated significantly at day 5). igg antibodies increased similar to iga class only slightly and again only sac to la was significantly higher at day 5 and 10. however sac to (xh of all ig-classes remained continuously on the same level troughout the observation time. correlation analysis revealed strong cross-reactivity (r>0,5; p<0,01) most often between antibodies of igm-elass (73%) followed by igaclass (40%) and lgg class (2%]. conclusions: multiple trauma is associated with temporary endotoxemia. endotoxins probably translocated from the gut cause specific increase of anti endotoxin antibodies in blood of the igm-class. endotoxins cause no increase of antibodies to gramposilave bacteria. igm antibodies are most unspecific. during cardio-pulmonary bypass, as well as postoperatively, high levels of endotoxin, interleukin-6 (ii-6) and c-reactive protein (crp) were measured in 30 patients. i0 female and 20 male, ageing from 30 to 73 with a median age of 59. blood sampling was done preoperatively, immediately after induction of anaesthesia, after thoracotomy, after cannulation of the aorta and right atrium after the first half of the reperfusion phase, after closure of the thorax, 2 and 4 hours after the operation and then every morning until the 5th postoperative day. blood was drawn into heparinized tubes (i0 iu/ml) which were free of endotoxin. crp levels were determined through the use of the behring nephelometer. 11-6 levels were measured by using commercially-available elisa test. the endotoxin level was determined by a chromogenic modification of the limulus amebocyte test. the statistical analysis was done using the wilcoxon ranks test and correlation analysis. a significant increase {p 0.01) in endotoxin plasma occurred during surgery, culminating in a peak (median value of 0.795 eu/m!) during reperfusicn. plasma levels of endotoxin continued to be slightly raised till the 5th day after surgery, whereas those of interleukin-6 rose at the end of the operation and were at their highest 4 hours later (median value of 217.5 pg/ml). crp levels were also high postoperatively with a median value of 114 mg/l, and were markedly raised on day 2 (191 mg/l). a definite, statistically significant correlation between the plasma levels of endotoxin and 11-6 during the operation was establisthed (p 0.05), leading us to conclude that the endotoxin liberated during cardiac surgery acts as the main trigger in the releasing of 11-6, and thus induces the postoperative acute phase reaction. there was no evidence of a correlation between crp and endotoxin or 11-6 plasma levels. impaired immune function is well described following trauma and hemorrhagic shock (hs). prior studies have utilized peripheral blood or spleen cells to index immune function following hs. however, changes in mucosal immunity are not weii characterized in this setting. gut origin sepsis is thought to be an important cause of organ failure and death following trauma. a rodent model was utilized to allow comparison of mucosal-associated immune function vs, systemic compartments after hs. fischer rates underwent hs (map 35±5mm hg) for 60 minutes followed by resuscitation with shed blood and lr. sham animals were instrumented only. rat tears were collected at 24 and 72 hours following hs for quantitation of slga by ria. animals were sacrificed at 24 hours and spleen (spl), peripheral lymph nodes (pln), and mesenteric lymph nodes (mln) harvested for cell population analysis using flow cytometry and mitogen stimulation analysis. cell marker expression analysis revealed no changes in t or b ceil populations following hs. mitogen mucosal immune function appears relatively spared following hs. the mechanism(s) for this variability in immune function requires further investigation. we have found that transplantation of bone marrow in a hind-limb graft to syngeneic lethally irradiated recipient is followed not only by rapid repopulafion but also overpopulation of bone marrow cavities. the question arises whether this unexpected phenomenon could be the result of stimulation of stem cells by factors (cytokines) released from surgical wound at the site of anastomosis of graft with recipient. aim of the study was to investigate which tissues damaged during the procedure of limb transplantation may be a potential source of humoral factors accelerating in vivo bone marrow proliferation. methods. experiments were carried out on lew rats in 5 groups. in group i, the hind limb was transplanted orthotopically to a syngeneic recipient; in group ii, sham operation was performed; in group iii, a four-cm long cutaneous wound was made on the dorsum; in group iv, limb skin was harvested, fragmented and implanted into peritoneal cavity; in group v, bm from femur and tibia was implanted intraperitoneally. bm, lymphoid tissues and blood were sampled 10 and 30 days later for cell concentration and phenotype evaluation. results. the yield of nucleated cells from tibia was on day 10 in the control 29.3 + 3.2, in group 1 58.1 +2.0, in group ii 47.1 + 1.6, in group iii 50.8+2.0, in group iv 48.5_+2.0, in group v 37.4_+4.9x10(6). the evident increase in bmc yield in all groups continued until day 30. increase in weight and total cell count of spleen and mesenteric lymph nodes in all but group iii was also found. no differences in percentage of maturing erythroid cells, but higher of mature myeloid cells and lower of lymphocytes were observed. conclusions. trauma of skin, muscles, and bone brought about an increase in bone marrow cellularity and acceleration of maturation of myeloid lineage. transplantation of bm ceils alone did not produce this effect. transplantation of bm in limb graft is a good model for studies of natural factors reaulatin~ bm hemormesis. this study sought to determine a relationship, if any, between the degree of hypochclesterolemia upon trauma patients' admission and their subsequent outcome. all blunt and penetrating trauma patients admitted to a level i facility from 1987 through 1991, and who had serum cholesterol assayed during the first 24 hrs were retrospectively studied for development of death or significant organ dysfunction. the mantel-kaenzel chisquared test was used to determine significance of data at the p< 0.05 level. results: 2909 trauma patients were admitted during the four-year period who had serum cholesterol assays performed in the first 24 hrs. 24 patients had cholesterol levels less than 50 mg/dl; 8 of these (33.3%) died, 3 (12.5%) developed ards, 5 (20.8%) developed acute renal failure, and 8 (33.3%) developed multisystem organ dysfunction; hypocholesterolemia in these patients was not due to liver injury or massive fluid administration. the risk of death was 5 times greater and risk of multi-organ failure 3 times greater in this group than in those with a normal serum cholesterol (>if0 mg/dl; 1820 patients; p<0.05). conclusions: admission serum cholesterol level in the trauma patient serves as a powerful marker for those at risk of subsequent organ failure or death. hypocholesterolemia in this setting may result from organ hypoperfusion and humeral mediator release. lung tissue contains many immunocompetent cells. resection, therefore, is expected to activate extensively inflammatory mediators such as pmn-elastase, pmstanoids and pteridines. in a prospective clinical study we compared 35 patients (pts) undergoing either thomcotomy with or without lung tissue msectioh and tboracoscopic lung resection concerning activation of inflammatory response. material & methods: group a pts (n=8) had thoraantomy but no lung tissue injury; group b pts (n=ls) had thoracotomy and lung tissue resection due to benign diseases; group c (n=9) represents group b tissue resection but using a thomcoscopic procedure. the following parameters were determined pre-, peri-, and postoperatively: elastase and crp as indicators of activation of pmn-leukocytes and injury severity; prostacyclin (pgi 2) and thromboxane (txa~) as parameters of lung endothelial response; prostaglandin f2~ (pgf~) and pgm representing pulmonaly metabolic activity; pge a and neopterin as proof of macmphage activation. statistics were performed using analysis of variance for repeated measures. results: group b pts revealed postoperatively an increase in crp (p<0.001) indicating a higher injury severity in comparison to the thoracoscopic procedure (c). both, controls (a) and group c pts did not show pmn-activation, whereas group b demonstrated a reversible increase in elastase. surgical trauma caused in all groups a release of pgi z and txa 2 which was more pronounced in c (p<0.05) and most in b (p<0.01). similar results were found for pge~ and pgf2=. there was no activation of maerophages since neopterin did not increase. apparently, metabolic lung function was not impaired because there was no marked rise in pgm except in b (p<0.05 vs. c). discussion: our results demonstrate that lung tissue injury aggravates the mediator release induced by thoracic traum. these mediators among others are able to increase capillary pressure and hence lung edema formation. impairment of lung function, however, seems dependent on the extent of the liberation. therefore, the maximal release reactions occured in group b and c after lung tissue resection, whereas the controls showed the highest levels immediately after the incision. we conclude that thoracoscopic procedures are superior in reducing the resection trauma per se and hence might prevent severe mediamr-induced (pulmonary/systemic) sequelae. in a prospective study we investigated 45 patients using radiochemical method according to sch~dlich (s) and photometric method according to hoffmann (h). serum of severly traumatized patients was withdrawn directly after admission at our emergency room and in narrow time intervals during first 24 hours after trauma. follow up control samples were taken daily until day ten. whereas no elevated pla-ca was found during first 24 hours, a peak was regularly observed around day four. there was high correlation between pla-ca and iss (r=0.71, p<o.o001) except patients suffering from thoracic trauma, thoracic trauma {tt) provoked similar gons-score-and pla-ca-values compared to multiple trauma (mt) despite significantly lower iss: pla-ca (h) mt 32.9 + 6.4 tt 28,7 +_ 9,5 n.s. pla-ca(s) mt 11.7 +_ 2.6 tt8.3 _+ 2.6 n.s. goris mt 3.5 4-0.7 tt3.6 _+ 1.3 n.s. iss mt 35 4-3 tt21 4-3 p<0.01 we found delayed elevation of pla-ca after severe trauma, however detection of delayed pla-ca in the serum of traumatized patients cannot give reliable information about the exact role of pla in the inflammatory reaction after trauma as latest results show that pla is secreted early but bound at lipophilic sites of vascular membranes. hern~mdez m j, amiguet ja, monreal ji, vid~n jr, jim6nez f j, l6pez-vivanco g acute phase reactant proteins increment in serum of patients with traumatisms and inflammation has been previously reported. alpha-1 antitrypsin (aat), alpha-2 macroglobulin (amg) and ceruloplasmin (cp) are evaluated in the following of 33 patients, 27 males and 6 females (mean age: 43 years), with compound fractures. 57 % of them were located in tibia and fibula. intercurrent infection developed in 4 patients and internal fixation rejection in one. measurements were made by radial immunodiffusion on days 0, 3, 10, 17, 24, 31 and 38. a control group of 30 healthy volunteers was also evaluated. aat and cp showed increased values from day 3 which kept elevated untill the end of the study. amg elevated from day 24. when infection developed, aat and cp values were even higher whereas amg values decreased. fixation rejection caused aat, amg and cp increment. aat and cp increment in non complicated fractures might be due to accompanying inflamation, by means of cytokines release and aprps hepatic synthesis induction. therefore, increment is higher when infection or rejection develop. amg behaviour at early stages of evolution is secondary to hiperconsumption, considering mean life shortening after proteases ligation. amg modifications in fixation rejection remain obscure. aprps increment modulates inflammation and bony callus formation by means their antyprotease and antioxidant properties. clinically, aprps might be useful parameters in determining complications onset in fractures evolution. hemorrhagic shock (hs) is a common complication of trauma, as well as some major surgical procedures. the alteration of the immune system by hs is thought to contribute to the increased septic morbidity and mortality seen in these patients. multiple mediators are released following hs. il-1, il-6, tnf, pge 2 and tgf-b have all been implicated in these immune system changes. in vitro, pge 2 causes many of the immune system changes following hs, yet the time course of pge 2 release has not been well deliniated. the aim of this study was to determine the time course of pge 2 release by peritoneal and splenic macrophages following hs. c3h/hen mice were bled to a mean bp of 35+5 mm hg for 60 minutes. the animals were resuscitated with the shed blood and normal saline. control animals were cannalated, but blood was not withdrawn. the animals were sacrificed at 2, 12, 24, 48, and 72 hours following hs. peritoneal and splenic macrophages were harvested from each animal and cultured with lps for 24 hours. the supernatants were collected and frozen until assayed. pge 2 was assayed using an elisa (cayman chemical). results are shown below for the peritoneal macrophages: pge 2 release by peritoneal macrophages was significantly elevated at 24, 48 and 72 hours over control. it was maximal at 48 hours, but by 72 hours it had started to decline. splenic macrophage pge2 release was much more variable. (data not shown.) there was less consistency between time points and no clear trend was seen. in conclusion, pge 2 is significantly elevated at 24 hours following hs, and reaches a peak at 48 hours. pge 2 release may be responsible for the immune system changes seen in the first few days following hs. splenic and peritoneal macrophages respond differently in their release of pge 2 following hs; which may be due to differences in their milieu. there is an obvious association between the altered immunocompetence of patients following traumatic injury of various types and the increased riskto develop complications. the present investigation was undertaken in order to analyze the value of both neopterin (n) and c-reactive protein (crp) in monitoring disease course in patients with multiple trauma. we were interested to compare nconcentrations as marker of t-cell/macrophage activation with crp levels as indicator of the inflammatory acute-phase response daily serum concentrations(n and crp) were measured in 12 patients (9 male, 3 female, mean age 31.42), admitted to our icu following serious trauma (mean iss=34) and in 10 control subjects. the clinical course of each patient was evaluated with apacheii score according to knaus. mean stay in icu was 9.25 (range 5-13). 5 patients didn't develop organ failure(nof-group), 4 patients survived, developing mof (s,mof-group), 3 patients died with mof (ns, mofgroup). three patients(one s and two ns) showed signs of septicemia and septic shock. the serum samples were measured for neopterin (immu-test-ria, henning-berlin) and for crp (immunoturbidimetric method) -at the university hospital for internal medicine, innsbruck, austria. the highest crp levels were measured 48h after trauma, but they were not accompanied by significant increase in n-values. we found significantly elevated n-concentrations from the days 5-6. n and crp levels showed a parallel release peaks on the days 6-7 and the values discriminated significantly among the three outcome groups(the second crp peak is lower in comparison to the first posttrauma peak). by the ns-group we found constant and parallel increasing crp and n levels, reaching extremly high values 24-48h and preceding clinical signs of mof and sepsis. the values increased dramatically before death. the serum levels by all 10 control subjects were upper the normal limit for n and crp.. the parallel occured peak changes in n and crp concentrations showed a significant correlation with clinical status, using a disease activity score (apac h eli). our findings suggest that both-cellular and humoral mediated immune mechanisms are activated after multiple trauma and that simultaneous determination of n-crp may be of advantage as diagnostic and prognostic parameter in polytrauma-patient monitoring panel. it is apparent, that measurement of very high levels may be of value as a srong indicator in developing of mof or septic complications and thus offer the possibility for earlier therapeutic intervention. the modification of t-cell/macrophage and acute-phase responses may be potentially useful in the treatment of icu patients, resuscitated after severe trauma, and suggest that n-crp may be relevant indicator for testing experimental immunomodulating therapies. although several in vitro studies suggested the catalytic action of iron in oxygen free radical generation, few data are available concerning iron-metabolism following ischemia-reperfusion injury and hemorrhage in vivo. aim of the present studies was to evaluate iron metabolism following mild and severe hemorrhage in the presence and absence of intraperituneal hematoma. methods. male lewis rats (300 g) divided in 4 groups (observation time 120 hrs): a: mild hemorrhage by sequential blood sampling (0.9 ml each)(0, 2, 4, 6, 24, 48, 72, 96, 120 hrs). b: a with hemoperitoneum (hp)(2 ml/100g), c: hemorrhagic shock: blood withdrawal of 3 ml/100g over 30 min and resuscitation with ringer's lactate (2 x shed blood) and isogenic donor blood; d: c with hp. parameters: serum iron ([g/dl], ferrozin-method); 59fe labeling of erythrocytas (application of 1 ci 59fe i.a. into a donor rat, 5 days later bleeding of the rat and injection intraperitoneally in experimental rat; plasma transferrin ([g/ml], ria). t-tests and mann whitney. data are expressed as mean + sem. results. compared to baseline, mild and severe hemorrhage caused a significant increase in serum iron at 24 and 48 hrs, respectively (212+30 vs. 248+35; 185+_28 vs. 255+_22). hp reduced this increase (209+_17 vs. 177+21; 194+25 vs. 197+-48, p<.05). i-ip significantly increased hemoglobin between 24 to 96 hrs in mild (12.1+_.8 vs. 14.2+.7) and severe hemorrhage (10.3+.6 vs. 13.1+.6). in mild and severe hemorrhage comparable maximal resorption (59fe labeled erythrocytes in circulating blood) of intraperitoneal hematoma occured at 24 hrs (mild: 318+_71 vs. severe: 364+-114 counts/rain). hp increased survival-rate following hemorrhagic shock (no hp: 10/17; hp: 10/13). in mild hemorrhage no alterations of plasma transferrin concentrations. in shock group without hp, there was at 96 hrs a significant higher transferrin level than with hp alone (3.2+_.5 vs. 2.23 + .2). in conclusion, hemoperitoneum with hemorrhage prevents increase in plasma iron levels by increasing hemoglobin through resorption of erythrocytes and thereby suppressing iron mobilization from endogenous iron resources, e.g. liver. this suggests that intraperitoneal hematoma not necessarily promotes iron-mediated oxygen free radical production, especially since survival in these groups was improved. increased transfarrin levels in group c at the end of the experiment suggest increased iron turnover which was not observed in shock groups associated with hemoperitoneum. primary immune response to thymus-dependent (srbc) and thymus-independent (vi-polysaccharide) antigens was determined during 30 days after cct. antigenic challenge was made on day i, 5, 15 or 30. number of spleen plaque forming cells (pfc) and serum haemagglutinin (ha) titre were studied on day 5 after immunization. cct stimulated immune response to t-dependent, but not to t-independent antigen. enhanced response was noted when srbs were given in posttraumatic day i, it was higher than preceding one after injection on day 5 and reached a maximum when antigen was presented on day 15 after trauma (pfc number 2.4 times exceeded the level of immunized, but nontraumatized rats). the rise of pfc number in spleen correlated with increased ha level. thymectomy had been performed 30 days before cct completely abolished posttraumatio stimulation of immune response to t-dependent antigen. standard thoracotomy also enhanced humoral response when srbc were administered on day 15 after the operation. thymus trauma modeled by pincett squeezing of its right lobe and performed at the same time with thoracotomy fully abrogated as well as thymectomy stimulation of immune reaction. thus, different experimental chest traumas are able to increase antibody production via thymus-dependent mechanisms. endothelin is a 21 amino acid peptide produced by ischemic or injured endothelial cells. in vitro and in animal studies, et is also a potent constrictor for renal mesangial and coronary vessels, a negative cardiac inotrope and an endocrine regulator. our lab has shown that et is an agonist for monocyte production of interleukins i, 6 & 8, prostaglandin e 2 (pge2) , and substances which trigger superoxide production. systemic et levels increase in hypoperfused and septic patients, and et may be yet another important cytokine in critical illness. but while et has been shown to be a potent vasoconstrictor in healthy dogs, systemic vascular resistance does not increase in critically ill patients with high et levels. (we hypothesize that this might be due to pgez-mediated vasodilation predominating over et-mediated vasoconstriction.) we next asked what happened to systemic et levels in patients with major burns (>20%.) ten hemodynamically stable patients resuscitated by a modified parkland formula to a urine output >30 cc's per hour had et levels drawn on admission, at i, 6 12, 24, and 48 hrs. et levels were measured by radioimmunoassay. mean levels were elevated at 205±28 pg/ml at all time points versus levels in healthy controls of 39±9. in summary, systemic et levels increase significantly in patients with major burns. et may be yet another cytokine playing a significant role in the immune, inflammatory and multiorgan dysfunction observed with major burns. restoration processes in an organism after ischemic damage are realized through ~n~lammatory mechanisms~ the intensity of which is significantly defined by blood levels of neuropeptides. myocardial infarction (mi) was chosen for studyin 9 these processes since it eradicates the influence of infectious factc~rs. 32 dogs~ in whom mi underwent different forms o¢ healer, g; bhn~ed ~h~t during the acute phase of the disease there was a characteristic rise of ne!~ropeptides in the blood. these neuropeptides had nociceptive and antinociceptive effects. particularly substance p and -endorphins triggered off the development of compensatory and adaptive mechanisms and defined the intensity of inflammatory reaction at the zone of ischem~t: damage-notable fall in substance p levels after an ~nitial increase, while the ~-endorphins stayed high was an important condition for non complicated healing of mi. on the other hand high levels of substance p with low ~-endorphin concentrations lead to increased infiltration o~ neutrophils into the infarction zone and weakened the activity of synthetic processes~ thereby leading to left ventricular aneurysm. at the same time low intitial levels of substance p slowed down the development of necrotic processes which lead to delay in refunctioning of the heart and complicated the healing process. thus, regulation of the levels of neuropeptides in the blood in trauma forms a perspective method of its treatment. of laparascopic versus open choleocystectomy c. schinkel, s. zimmer, v. lange, d. fuchs, e. faist the impairment of immune function due to surgical trauma may be followed by deleterious septic sequelae. compared to open abdominal surgical procedures (lap), laparaseopic surgery (lsc) is associated with a decrease in hospital stay and in accelerated patient recover. the aim of the study was to evaluate the sensitivity of the immune sermn parameters of il-6, saa and neopterin, the percentage of cd14+ cells, the in-vitro il-2 synthesis after mitogen stimulation and lymphocyte proliferation, in order to purposefully discriminate differences in the severity of trauma. we investigated the blood of 27 patients with cholecystolithiasis undergoing either laparascopic (16) or open (i 1) cholecystectomy on consecutive perioperative days -1, 1, 2 and 3. there was no significant difference between the two groups concerning age and sex. patients with clinical signs of acute cholecystitis were excluded from the study. operation time and hospital stay were obviously longer in lap patients (67 versus 145 minutes, 5 versus 20 days) compared to the lsc group. concerning the unspecific acute phase reaction we could show no difference in the increment of senun amyoid a (saa) synthesis in the lsc group (d-i 40 + 1 lng/ml, d1 362 + 43ng/ml) versus lap group (d-1 61 + 19ng/ml, d1 457 + 45ng/ml), while in serum il-6 levels we saw a less steep increment in the lsc group (7-fold from d-1 to d 1) compared to the lap group (10-fold from d-1 to d 1). the analysis of cd14+ receptor expression and serum neopterin did not reveal any difference between the groups. lymphocyte function showed an impairment of proliferation to antigen stimulation in lap (d -1:24.000 + 3.330 cpm, d 1:17.300 + 3.000 cpm) compared to the lsc group (d -h 22.500 + 4.200 cpm, d h 26.000 + 2.900 cpm). in both groups il-2 synthesis was decreased post-operatively. our data indicate that laparascopic cholecystectomy reusults in a less distinct unspecific acute phase reaction post-trauma compared to that following lap. neopterin serum levels and cd14 receptor expression show that these parameters apparently are less useful markers to detect differences of surgical trauma severity while it appears that the impact of lap is reflected most impressively on the lymphocyte compartment. trauma alters the host resistance of organism and is accompained by appearence of excgenic and endogenic proteins in the body. to understand the molecular mechanisms of host resistans disorders in trauma, as a first step, the genetic regulatory mechanisms of immune response after antigen injection has been studed. the appearence of specific protein factors (13-19 and 25 kda), in the nucleus of rat splenic and brain cells, accordingly, was shown after immunization with sheep erythrocytes. the stimulatory effect of these factors on the il-2 mrna and il-2 production was detected. the nucleotide sequences of the human il-2 gene regulatory region bounding by the splenic nuclear proteins were determined between +39-141 b.p. the il-2 trans-factors shows the affinity to splenic and thymic lymphocytes in vitro. thus, the antigen causes the appearence of specific protein factors in the cells,which act on the gene level,stimulate il-2 production and the host resistance. these results cause the next step of experiments using the same model, but after trauma. these investigations will let us verify the hypothesis that the protein il-2 gene trans-factors may play a definite role in the decrease of the cell immune responce after trauma. confronted with the routine procedure of prophylactic treatment of candidates for surgery in a rural african hospital, we initiated studies on the fre'quency of post-surgical malaria. in tanzania 195 non-pregnant patients from rural areas were followed. of preoperative patients 10% had a parasitaemia and those maintaining it showed no increase or complaints. nine percent of patients without detectable parasitaemia before surgery came down afterwards and one-third had malaria-like complaints. spinal and general anaesthesia were equally applied in these last patients. in burkina faso we studied 120 patients of which 16% had a parasitaemia on admission and 6% had postoperative malaria. half of the surgical patients came from rural areas, whilst only 14% of those with malaria lived in the city (with much less exposure and immunity). 57% underwent major surgery and 43% minor. bloodtransfusions (4% with parasites) never evoked a parasitaemia in recipients. post-surgical malaria is thus a reality in about 10% of the adult cases, both in east and west africa. surgery evokes a cascade of factors, varying from cortison to interleukines and acute phase proteins; immune responses may temporarily be suppressed. clinical attacks of malaria in otherwise immunes could be evoked by one of these factors. though malaria can easily be cured, the differential diagnosis is difficult because of post-surgery fevers; we found that 16% was treated without justified indication. the involvement of "student-doctors" a. this study examines glucose uptake and hexose monophosphate (i~ip) shunt activity in normal human peripheral lymphocytes and polymorphonuclear leukocytes (pmn). glucose uptake was determined by measurir,g the uptake of tritiated deoxyglucose, a non-metabolized glucose analogue. adsorption of co2 derived from [i-14c] glucose was used to determine knp shunt activity. in vitro assays were carried out in hormone concentrations approximating normal and elevated trauma blood levels. (normal -cortisol 0.12 ~g/ml, glucagon 80 #g/m1, epinephrine 20 ~g/ml, insulin 18 t~u/ml; traumaeortisol 0.40 ~g/ml, glucagon 500 /*g/ml, epinephrine 420~g/ml, insulin 36 ~ij/ml. analysis of twenty subjects showed a reduction of 207° ~mp shunt activity by lymphoeytes and a ]3% reduction in glucose uptake by p~n in normal vs. trauma hontc,nes p < 0.05. lymphocyte glucose uptake was also reduced by trauma hormones p~0.05. it ha~ be.ea~ suggested thgt idiopatno pulmonary fibrous (y.pf) [s a consequence of severe alveolar epithelial injury and is associated with an nveolar irnammamry reactio~ and the presence 0f.neutr0phils. there~bre, neutr0pk~ chemoattra~ant~ are probably important in the genegs oft.he infial lesions of ipf. the obse,"wson that stimulated macrophages are or~n histologically promin~t in fibmfio [-~gs ~.nd am capable of p~oducmg a v~dery 0f flbrogenic pep'ides also a~gues for their role ~n the pathogenic prc~e~ oflpf. the observation that stimume~ maerophages ere often histologica[iy prominent in fibrotio lungs and ~re ~pable of producing a varie~, offibroge.~e peptide~ also argues for tkek role in the pathogenic process, therefore, we ha-~e tested the potentn for iater!eukln-8 (i1..-8) and mo~tocyte chemotacde pop, de 1 (x¢cp-1) to induce neutro~hil ~d mononuclear phagocyte accumuhdon in lungs of pafient~ with pulmonary .~r~idosis and i~f. brenet~o.alveolar lavabo (bal) fluids from 12 ipf and 15 sar~qidosis patient were conexntratea by reversed-phase chromatography, ~d ii.4 arid mcp-i asso.~ed by ells& ehemotaxis mad enzyme-reieasing ~ssas's on msnocyte~ and neatrophiis. elisa revealed significenfly elevated b al-eoneentrations o£mcp-i (20.1 ng]mg aibumm) in purisms with p~monary sarcoidodis artd in ipf (41.8 ng!mg) in comparises to 1.1 normal individuals (4.24 ng/mg) and to 15 patients w~th obreic bronentis (cb) (~, 16 rig/rag). similarly, chemota*dc ac~a~' for monocles (mcp-1 e.qu/va]ent) was strongly increased in sareoidosis (86.03 ngjmg) as well as ~n 12f pag,nts (54.47 ng/mg). norra.al indlvidu~s and cb patiants hzd a 7. or 3-fold lower ~cn%i~y, re~peefively. patients with ipf and sarcoidosi~ also h~l eievated il-8 ievei~ (15.5 and 26.0 rig/rag, respe~veiy; nomzls: 2.14 rig/rag; cb: 4.23 ng/mg) mad nvatropmi ohemotax~ (60.25 ~'~d 49.68 nnmg, res!z~ztiveiy; aormals: 0.25 ng,'mg; cb: 1l06 ngmg). these data suggest that increased ievels of born mcp.1 ~d il-8 may be oharacted~tie for ~arcoidosis or ipf_ it appears iikely that both ehernoattraetants ~ontribute to the influx ofmonocytes and neutrophils into the pulmonary alveoius and interstit~um in these dlsea~es. we have recently shown that the combined administration of noninjurious doses of lps and paf in the rat produce ards-like lung injury characterized by neutrophil adhesion to lung capillary venules, neutrophil accumulation in lung parenchyma, pulmonary edema, and increased protein and neutrophil count in bal fluid. this new paradigm of lung injury was associated with elevated serum tnfc~ and pretreatment with anti tnfa mab dose-dependently prevented these responses. also, the combined administration of lps and paf induced lung mrna levels of tnfe~ (5 fold vs. lps or paf alone), ll-lg (80 fold), kc (60 fold) and il-6. taken together, these data suggest that this new paradigm of lung injury is cytokinemediated and that lps/paf in vivo can functionally couple to the activation of gone expression of a multi-cytokine network system, all of which may be involved in the pathogenesis of ards. materials and methods. the sheep model included hemorrhagic shock and closed femoral nailing at day 0, 12hourly injections of e. coli endotoxin and zymosan-activated autologous plasma at clays 1-5 and further observation and measurements at days 6-10. from venous blood and bronchoalveolar lavage(bal)fluid of ten merino sheep (mean weight 30 kg) neutrophil counts (10e6 pmn/ml blood or epithelial lining fluid-elf-), the elf/ plasma ratio of albumin (r), and the zymosan-induced (stim) and non-induced (spont) chemiluminescence response (cl) of blood (10e6 cpm/250,000 pmn), and of blood-and bal-isolated pmn (10e6 cpm/25,000 pmn) were measured. for statistical calculations the wilcoxon test was used. data of the changes in polymorphonucleur leukocyte (pivinl) metabolism have been suggested to play a pivotal part in the post-traumatic systemic inflammatory response syndrome. the underlying cellular mechanisms which control this response are not yet completely understood. since the 'ca 2+ second messenger'-system has been shown to be involved in regulation of pmnl-'respiratory burst', we investigated changes in pmnl-ca z÷ regulation in relation to oxygen free radical mediated injury. methods. in 12 polytranmatized patients (mean injury severity score = 32) arterial and venous blood samples during 15 days. daily evaluation of horowitz-quotiant (po2/fio2), plasma lactate (mg/dl) and body temperature ( results. body temperature peaked at day 6 and 7 (day 0: 36+.4; day 7: 39.8+.4). plasma lactate was significantly increased at day l (36+2) and day 7 (16.2+2). hurowitz-quotient (day 0: 344+43) was low at day 4 (199+19) and day 7 to 10 (178+25)(p<.05). at day 7 a substantial rise in venous pmnl-superoxide production (day 5: 1.9+_.45, day 7: 3.3+.7, day 9: 1.72+_.8), oecured with significant increase in plasma lipid peroxidation (day 1:0.6+0.2 ; day 7: 1.3+0.2). pivin~-myeloperoxidase activity was high at day 2 (2.3+--.5) and then continuously declined (day 7: 1.3+.3). plasma antiexidant activity (glutathione pemxidase) was reduced by 34% at day 7 (day 1: 5.3+.3; day 4: 4.4+_.2; day 7: 3.5+.2). whereas basal ca 2+ concentration remained unchanged (day 4: 68+_17, day 7: 68+_15), fmlp-stimulated cytosolic ca 2+ mobilization increased at day 7 (day 4: 457+89, day 7: 11084410, day 9: 670+262). conclusion. the present study in polytraumatized patients shows, that seven days after injury the agonist-induced pmnl ca 2+ mobilization is significantly enhanced. at the same time, pmnl-oxygen free radical release and phagocytotic activity, systemic fever response and lactate concentrations were maximal. these observations were accompanied by post-tranmatic respiratory failure and in some patients by clinical signs of multiple organ failure. preliminary data from an ongoing study using hes-and dextran-infusions in these patients show attenuation of this inflammatory response. stefan rose, m.d., trauma surgery, univ. of saarland, 66421 homburg/saar 1donnelly sc, 1haslett c, 1dransfield i, 2robertson ce, 3grant is, 4carter c, 4ross ja, 5tedder tf. 1dept's of respiratory medicine, 2accident & emergency, 3intensive care, 4surgery, university of edinburgh, scotland and 5dept. tumor immunology, dana farber cancer institute, boston. the selectins are a family of adhesion molecules (l-selectin, e-selectin, pselectin), all of whom are implicated in inflammatory cell transendothelial migration. they, as a family can be proteolytieally cleaved from their parent cell and exist in a soluble form within the circulation. ards is a disease state in whic neutrophils and neutrophil transendotheliat migration have been implicated. in this study we wished to investigate whether the levels of these circulating soluble receptors from patients at-risk of ards at initial hospital presentation, correlated with subsequent ards progression. eighty-two patients were enrolled (pancreatitis (n=19), perforated bowel (n=12), and multiple trauma (n=51)), of whom 14 progressed to ards. assays for soluble l,p & e-selectin were performed on collected plasma samples via a sandwich elisa. (ns = not significant, **** = p<o.0001) we have found a highly significant inverse correlation between a low circulating soluble l-selectin (and not soluble e & p-selectin) and subsequent ards. these results further our understanding of neutrophilendothelial interactions in the early stages of ards pathogenesis and offer the promise of sl-selectin in combination with other markers of inflammatory events being used to identify individuals at particularly high risk of ards progression on initial hospital presentation. it has been suggested that polymorphonuclear leukocytes aggregate in the lung capillaries of patients developing the adult respiratory distress syndrome (ards) and account for the endothelial damage while releasing toxic metabo-iites such as o.a. free oxygen radicals. among many antioxidants which have been investigated in in vitro systems and in animal models, n-acetylcysteine (nac) has been established to be useful as a free radical scavenger in vivo. to assess the influence of n-acetylcysteine (nac) on the activation of neutrophils in patients undergoing cardiopulmonary bypass (cpb) surgery with extracorporeal circulation (ecc), we evaluated the plasma levels of elastase and myeloperoxidase (mpo) throughout the operation and up to 24 hours after surgery. four hours after surgery also a bronchoalveolar lavage was performed. a spectrophotometric method was used for the detection of plasma elastase activity and mpo levels were measured using a radioimmunoassay. we found that pretreatment with intravenous nac prevented the increase in elastase activity (632 _+ 231 pmol/i vs 1666 _+ 187; p -0.027), but not the release of neutrophil related mediators (2.20 _+ 0.56 ng/i vs 1.80 _+ 0.17; p=0.63). besides, nac had the capability to prevent the enhancement of neutrophil numbers in lavage fluid as is normally seen during cpb (1.4 + 0.8 % vs 25.0 _+ 14.3; p =0.04). although not significant, nac prevented also the increase in elastase levels in lavage fluid (19.0 + 18.5 pmol/i vs 172.7 + 101.4; p = 0.12). it is concluded that nac is able to protect against the inactivation of a 1-proteinase inhibitor, the main inhibitor of elastase activity, and that nac interferes with adhesion and migration of neutrophils. therefore nac may be useful in the prevention of tissue damage. this study is supported by inpharzam belgium. sodium nitroprnsside (snp) has previously been shown to attenuate the neutrophil induced lung injury associated with limb ischaemia and repeffusion. glyceryl trinitrate (gtn), already widely used in aortic surgery, also increases nitric oxide but has a less marked splanchnic "steal" effect. we examined the effect of gtn on lung injury, neutrophil infiltration and activation in a model of aortic occlusion (30 rain) and repeffusion (120 min). sprague dawley rats were randomized into: control (n=i 1); ischaemia repeffusion (ir) (n=12); and ir treated with gtn (2ug/kg/min) during repeffusion (n=10). myeloperoxidase activity (mpo) measured pulmonary neutrophil influx. pulmonary endothelial permeability was measured by wet to dry weight ratio (w/d), broncboalveolar lavage protein (bal) and neutrophil counts (bal pmn). neutrophil superoxide release (mean channel fluorescence mcf) was measured by flow cytometry in a further ir v gtn experiment (n=6 per group). control 573_+t02"* 992_+125 579_+54** bal pmn/mm 3 36l+-126 3"219+1087# 820_~221 *p<0.05, #p<0.02 v control/gtn;**p<0.01 v ir. students t test the significant increase in mpo activity produced by ir was attenuated by gtn. the increase in pulmonary microvascular leakage after reperfusion was also prevented by gtn. neutrophil superoxide release increased significantly from 9.4+0.76 to 13.9_+1.92 mcf after 40 minutes repeffusion (p<.01). this increase in superoxide was prevented in the gtn treated group. these results indicate that gtn inhibits neutropbil superoxide release during limb reperfusion, thus preventing pulmonary vascular leakage and neutrophil infiltration. these data suggest that the beneficial effects of gtn in aortic surgery may be due in part to a protective effect onpulmonary microvasculature. department of surgery, beaumont hospital, dublin 9, ireland. lipton adult respiratory distress syndrome (ards) is marked by increased vascular permeability of the lung to white blood ceils (wbc). indeed, influx of wbc into the lung is believed to be the central mechanism in acute lung injury of ards. evidence is developing that the neuropeptide c~-melanocyte stimulating hormone (c~-msh), a proopiomelanocortin derivative, inhibits inflammatory reactions in animai models of inflammatory responses in humans. to test the influence of a-msh on experimental ards, the peptide was administered to rats after intratraeheal infusion of endotoxin (s. typhosa, 500 #g in 0.25 ml saline). three groups (n=10 each) received: intratracheai infusion of endotoxin plus ip injections of o~-msh (100 ~tg in 0.2 ml saline) at 0, 2, and 4 hr post-endotoxin treatment; intratracheal endotoxin infusion plus saline injections; control intratracheal saline infusion plus saline injections. the bal data showed overall differences among groups (f2,29=6.2, p<.003), o~-msh treatment inhibited endotoxin-induced wbc migration into the pulmonary tree (p<.05). circulating wbc counts were markedly lower in both groups given endoroxin than in the control group (p<.01), but there was no difference in wbc count between these two endotoxin-treated groups. the results indicate that c~-msh can reduce wbc migration in experimental lung injury. in further experiments we tested the hypothesis that c~-msh, administered alone or in combination with an inhibitor of gram-negative bacterial growth, increases survival in a model of peritonitis/endotoxemia/septie shock. cecal ligation and puncture were performed under sodium pemobarbital anesthesia (75 mg/kg, i.p.) in female balb/c mice (16-20g) that were then randomly assigned (15-17 mice per group) to receive at time 0 and 3 hr later: control saline thjectioas (i.p.), injections of ~-msh (100 ~tg), gentamicin sulfate (i0 mg/kg), or both ~-msh and gentamicin. one ml of normal saline was given s.c. to each animal for fluid replacement. both ix-msh and gentamicin treatments administered singly improved survival; when given in combination survival was even greater these findings suggest that the anticytokine ot -msh molecule might be useful for treatment of systemic inflammation, perhaps particularly when used in combination with agents that inhibit bacterial growth. previous studies showed a close relationship between xanthine oxidase activation, membrane damage and postischemic cardio-respiratory failure following aortic clamping and reperfusion. aim of the present study was to evaluate alterations of polymorphonuclear leukocyte (pmnl)-metabolism and signal lransduction systems during ischemia/reperfusion induced by aortic clamping in man. methods, in 14 patients (3 female, 59+4 years) with elective reconstruction of infrarenal aortic anearysms, arterial (a) and venous (v) blood samples were obtained before clamping and declamping, and 60 min after declamping. arterial pla 2 concentrations were higher than venous (113+32 vs. 71+30). while cytosolic basal pmnl ca 2+ concenlxation was not altered at the end of ischemia (v: 52.3+8; a: 42_+6) and during reperfusion (v: 60+8; a: 69+12), fmlp-stimulated cytosolic ca 2+ mobilization showed a significant arterial increase as compared to venous (1652_+601 vs. 638+85, p < .05). these presented data indicate that the reperfusion syndrome after infrarenal aortic clamping is characterized by 1) pulmonary membrane damage possibly due to pmnl-degranulation, 2) activation of pmnl-superoxide radical production with release of activated pmnl in arterial circulation, and 3) a significant arterial increase of pmnl cytosolic ca 2+ mobilization after fmlp-stimulation parallel to pmnl-activation. in conclusion, ischemia/reperfusion in man induces pulmonary damage and activation of pmnl superoxide production possibly due to an altered pmnl-ca 2+ messenger system by inflammatory mediators (e,g. ii-8, tnf, paf). the septic lung diseases (i.e. ards) generally show distinct alveolar damage and surfactant disturbances. it is thought that alveolar macrophages are involved in specific release products like h202 and cytokines like tnf. in this pathway the type ii alveolar epithelial cell (p2 cell) is not only an important target, but as recently shown it is also capable of producing h202. the aim of this study was to investigate the influence of endotoxin on h202 release of p2 cells. we obtained p2 cells from lungs of female wistar rats and investigated the effect of lipopolysaccharid (lps) on h202 release of fresh isolated cells and on cells cultured for 2 days in a fibronectin matrix. furthermore we conditioned isolated alveolar macrophages for 16 hours with 1 p.g/ml lps and tested the conditioned medium (mcm) on cultured p2 cells. p2 cells 5 h ex vivo were _> 70 % pure, _> 95 % vital and had an basal h202 release of 64.5 _+ 1.8 nmol h202 per hour and million cells. adding 1 p.g/ml lps to the incubation medium the h202 release decreases slightly but significantly to 54.2 _+ 2.3 nmol. adding 0.3 p.g/ml phorbol myristate acetate (pma) to the basal incubation medium the h202 release increased 3-fold to 148.3_+ 26 nmol. pma induced h202 release decreased to 128.3 + 41.5 nmol after addition of 1 p.g/ml lps. after 2 culture days the p2 cells were _> 98 % pure and showed a pma inducible h202 release of 174.8 _+ 9.9 nmol addition of 1 p.g/ml lps had the inverse effect as on freshly isolated cells as it increased the h202 release up to 216.2 _+ 2.4 nmol. addition of mcm to cultured p2 cells increases pma-stimulated h202 release to 223.3 +_ 10.7 nmol. the release decreased to 191.9 _+ 2.4 nmol when an murine anti-tnf-alpha antibody was added. vitality of cultured cells was > 96 % in all experiments. the results show that lps has an direct effect on p2 cells cultured on fibronectin. we conclude that the observed additional stimulatory effects of mcm seems to depend on tnf-alpha. the induction of h202 release of p2 cells could be important for generating internal oxidative stress in p2 cells before external oxygen radicals exceed. the produced h202 did not necessarily damage p2 ceils, but it can effect surfactant metabolism, especially when extracellular h202 release of alveolar macrophages following an immune response is increasing. introduction: primary stabilization of femoral shaft fractures in patients with multiple trauma is beneficial. however, in patients with associated lung contusion we have found an increased incidence of ards, apparently associated with primary reamed femnral nailing (rfn). previous animal studies revealed, that perioperative disturbances of lung ftmetion appear to be related to the reaming procedure, ix~ssibly due to pulmonary embolizafion of bone marrow fat. in a prospective clinical analysis we compared effects of intrameduuary nailing with and withont reaming on parameters known to be related to ards-pathoganesis. in order to gain further insight into the role of endotoxin and cytokines in the pathogenesis of the adult respiratory distress syndrome (ards), we enrolled 23 patients with severe lung injury after sepsis (17) or polytrauma (6) and obtained multiple blood samples (14 days) for endotoxin, tumor necrosis factor e (tnfa), interleukin 18 (il-18) and interleukin 6 (il-6) determination. to evaluate the cytokine releasing capacity of the blood, plasma concentrations of tnfe, il-l8 and il-6 were also determined after the "in vitro" stimulation of the whole blood samples with lipopolysaccharide (lps, 0.1 ng/ml) for 2 hours at 370 c (stimulated values). the difference among stimulated cytokines levels and the basal plasma concentrations were defined as "delta values", an expression of the cytokine releasing capacity of the blood. the pao2/fiao2 quotient was used as an index of the severity of lung injury (sli). the endotoxin plasma level was significantly higher in patients with sli < 200 (0.288 ± 0.038 eu/ml, mean values ± sem) versus the patients with a sli > 200 (0.175 ± 0.023 eu/ml, p<o.01). the basal plasma concentration of ll-6 also correlates with the sli index (p<o.o01). the delta tnfe and delta il-6 values were s~gnificantly decreased in patients with a severe lung injury. compared with the survivors (n:11), the deceased patients (n=12) showed higher endotoxin level, a reduced tnfa and il-6 releasing capacity of the blood and higher basal il-6 levels. we can conclude that endotoxemia, high il-6 plasma level and a decreased capacity of the blood to release tnfa an il-6 under endotoxin stimulation associates with the severity of lung injury. [objectives] experimental and clinical findings implicate the involvement of inflammatory cytokines in the pathogenesis of the decreased oxygenation in the lung after major surgery, though the mechanism remains unclear. in the present study, we elucidated the involvement of il-8 in the pathogenesis of lung injury defined by deterioration of pulmonary oxygenation after esophagectomy. [materials and methods] seventeen patients underwent subtotal esophagectomy through a right thoracotomy. in all patients, tbe alveolar-arterial oxygen tension difference (aado2) was measured everyday and bronchoalveolar lavage fluid (balf) samples were obtained from a single segment ($4 or $5) of the right and left lung through a bronchoscope on days l, 3, and 5 days after surgery. they were then examined for cell analysis, measurement of cytokines by elisa, and measurement of neutrnphil elastase (ne) by elisa. [results] aado2 unexceptionally deteriorated postoperatively and the mean aado2 in 17 patients reached the maximum (mean ± sem; 372 _+ 23 mmhg) on the 3rd postoperative day. the mean concentrations of il-8, ne and neutrophil count in balf also increased postoperatively and reached their maximum level (2137 + 618 pg/ml, 1603 -+ 493 gg/l, and 1561 ± 477 x 103 cells/ml, respectively) on the 3rd postoperative day. these increases in balf were recognized in both the right and left lung. a highly significant correlation was observed between il-8 and ne levels in balf (r=0.82, p=0.0001). also, significant correlations between aado2 and the neutropbil count, and the concentration of ne or 1l-8 were observed. in contrast to il-8, neither il-6, il-113 nor tnfa was detected in balf. [conclusion] major surgical trauma such as esophagectomy may induce recruitment of neutrophils to the lung and cause lung injury. il-8 increased in the lung is an important mediator of neutrophil recruitment and activation in the lung. phorbol myristate acetate (pma) is commonly used to cause edema and other tissue damages in the lung. lung injuries induced by the administration of pma has been shown to be mediated mainly by neutrophils. neutrophils recruited to the lower respiratory tract may damage lung tissues by releasing reactive oxygen species, neutral proteases and lysosomal enzymes. the present study was conducted to investigate whether c~tocopherol, entrapped in dipalmitoylphosphatidylcholine liposomes and delivered directly to the lung, could counteract some of the pma-induced lung injuries. plain liposomes or c~tocopherol-containing liposomes (8 mg c~-tocopherol/kgbody wt.) were instilled into the lungs of rats 24 h prior to pma exposure (25 pg/kg, intratracheally) and treated rats were killed 3 h later. lungs of control animals exposed to pma developed increases in lung weight and lipid peroxidation as well as decreases in lung angiotensin converting enzyme (ace) and alkaline phosphatase (akp) activities. pma treatment also caused an increase in myeloperoxidase (mpo) activity in the lung, suggestive of neutrophi] infiltration. pretreatment of pma-treated rats with plain liposomes had no effect on pma-induced injuries. in contrast, pretreatment of rats with liposomal c~-tocopherol, 24 h prior to pma administration, resulted in a significant elevation of pulmonary a-tocopherol concentration, accompanied by a concomitant reduction in mpo activity and reversal of pmainduced changes in lung edema, lipid peroxidation, ace and akp activities. these results appear to demonstrate that the intratracheal administration of a liposome-associated lipophilic antioxidant, such as a-tocopherol, can significantly ameliorate the toxic effects of reactive oxygen species, released from pmastimulated pulmonary target cells and infiltrating neutrophils. jk wojcik, g palasiewicz, w roszkowski immunology department,institute of tuberculosis and lung diseases~ warszawa, poland, m~larhospital, eskilstuna, sweden. introduction:the critical point in neutrophil migration to the alveolar space in the course of ards is the attachment between giyeoproteins of neotrophil membranes(sialyl lewis x carbohydrate epitope containing u-l-fucose) and lectin domains of endothelial p and e selectins (gmp-140,elam-i). a potential beneficial therapeutic strategy may be designep to suppress neutrophil recruitment to the lungs by preventing their rolling and attachment to the pulmonary endothelial cells. objeclive:to investigate if tz-l-fueose prevent experiments pulmonary hypertension in ards as effective as metylpredniselone. methods:only healthy rabbits (n-24) weighing 5.d-5.5 kg oaselinepao>11 kpa and mean pulmonary arterial pressure (mpap)<i.5 kpa were included in this study. anaesthesia was induced with thiopental 20-25 mg/kg bw and "blind" jntubation was performed. a 5f swan ganz catheter was introduced via the left jugular vein into the pulmonary artery. pma, as was used in our experiments activates neutrophils and produces acute respiratory failure with pulmonary hypertension and pulmonary edema. eight rabbits serving as control animals received pma 25 ug/kg bw iv only. sixteen other animals were divided into two groups receiving either ~-l-fucose 100 mg iv or metylprednisolone 150 mg iv 15 min before pma admininstration. results:after pma administration notable physiological changes including marked increase in mpap(2.0-+o.~6 kpe3 ..lere found. the increase of npap was observed to be completely blocked in both groups of animals which received ~-l-fucose or metylprednisolone. conclusion:in aur rabbit model ards cz-l-fucose pretreatment prevents pulmonary hypertension after pma administration. the effect is comparable with high dose metylprednisolone. the possible explanation is that g-l-fueose molecules block iectin domains of gnp-140 or elam-i preventing the adhesion of the neutrophils to the pulmonary endotheliom. bartens c, elmadfa i, steltzer h, fischer m, druml w introduction: various micronutrients and vitamins are particulary effective in modulating immune functions and host defense. the nutritive ant±oxidants vitamin c, e and b-carotene, as well as selenium lower the burden of reactive free radicals and protect the structural integrity of cells and tissues of the immune system, as well as other systems in the body. certain cells of the immune system like polymorphonuclear leucocytes (pmn's) use free radicals as a weapon to destroy invading pathogens. these reactive molecules, when released into the surrounding area, mediate lipid peroxidation and tissue injury. there is growing evidence that pmn's are activated in vivo by complement or immune complexes in disorders such as ards. the respiratory host defense mechanisms of patients with adult respiratory distress syndrome (ards) may be defective. the aim of this study was ta evaluate the status of the free radical scavengers and their activity in ards and to investigate the respiratory burst of ards patients. methods: seven ards patients (3 sepsis, 4 trauma) with an fie 2 of 0.59+0.(36, a peep of 8.50±0.99, and a murray score of 2.83 ±0.18 were included in this study. 15 healthy adults served as controls. superoxide anion production in granulocytes was measured photometrically, and the hydrogen peroxides by fluorimetric assay. vitamin a, e, and g-carotene concentrations were assessed by hplc, and plasma vitamin c photometrically. plasma selenium was determined by dectrothermal aas. plasma lipid peroxidation pruducts, expressed as malondialdehyde (mda), were determined by thiobarbituric acid assay and hplc. results: mda-plasma (/xmol/l) 0.74~: 0.46* 0.37 ± 0.16 *= <0.01, **=p<0.001 condusion: ards patients showed significantly decreased plasma concentrations of vitamin c, e and g-carotene, as well as selenium. these nutritive ant±oxidants are consumed during increased oxidative stress. mda, a final product of lipid peroxidation, is increased. however, a difference in rates of release of hydrogen peroxides and superoxide-anion of pmn could not be detected. therefore, oxygen radical accumulation is more likely a result of excessive inspiratory oxygen concentrations (f.io2), pro-oxidant drugs, and a high settlement of pmn in the lungs, and not an increased release of free radicals of the single pmn. kd glycoprotein secreted by the alveolar type ii cells. recent study showed that sp-a exists in the sera from normal healthy adults and that the significantly elevated serum sp-a levels were observed in the patients with interstitial pneumonia and pulmonary alveolar proteinosis. these findings means there may be a mechanism that the sp-a produced in the alveoli escapes into the bloodstream. here we examined the serum sp-a levels in the patients with critical illness including sepsis and ards. ]clinical subjects & methods] a total of 99 serum samples were collected from 39 patients hospitalized in the division of critical care medicine(icu), sappom medical college hospital. serum sp-a levels were measured by an enzyme-linked immunosorbent assay using monoclonal antibodies to human sp-a. ]results] there was no significant difference between the septic (n=12) and non-septic (n=27) patients. patients with ards (n=10, 52.0+4.85ng/ml) and with respiratory disease other than ards (n=12, 49.6+-5.35ng/ml) showed significantly higher levels of serum sp-a than those with non-respiratory disease (n=17, 27.4_+4.86ng/ml). as to the ards patients, it was likely that the serum sp-a levels were getting higher in their clinical courses. however, there was no significant correlation between the serum sp-a levels and the pa%/fio 2 ratio. ]discussion] at the present time, the exact mechanism of the escape of sp-a into the bloodstream remain to be unclear. our results may suggested that there was some relation between this leakage of sp-a and the alveolar-capillary permeability because of the higher sp-a levels observed in the ards patients. however it is also likely that higher serum sp-a level represents the proliferation of alveolar type ii cells as the regeneration of damaged lung. further studies are now being done. neutrophils contain a number of enzymes within intracellular granules,potentially damagingto lung tissue.release of these enzymes into the lung tissue from the sequestred activated neutrophils is felt to play significant role in lung injury in the course of aros. using 6-hours acute animal model of aros the activity of cathepsins,beta-glucuronidase and acid phosphatase was determined in the lung tissue. results of this research were compared with neutrophil proteases activity in serum and broncho-alveolar lavage.also hemodynamic,respiratory and biochemical investigations were performed before beginning of experiment,and in a two-hours time intervals of its duration.after 6 hours was determined total and free activity of cathepsins,beta-glucuronidase and acid phosphatase in full homogenate of the lung tissue,mitochondriallysosomal fraction and the final supernatant. in the course of our experiment we observed an increase (30.i-49.6%) both total and free activity of neutrophil proteases in all fractions of the lung tissue.generally accepted current pharmacological treatment of aros only attenuated this increase but never caused reversion to the level before beginning of our investigations. the presence of aros was histologically proved.an activity of acid phosphatase in serum and the lung tissue fractions was compared with histochemical pictures of the lung. results of our research indicate that neutrophil proteases are a very important mediators in aros and they are a cause of the lung injury. in addition neutrophil-derived proteases can amplify the inflamatory process by enzymatitally activating of many other mediators and they also may to increase an oxidant induced injury by imparing of antioxidant defenses. oepartment of general surgery, sk~odowskiej 24a 15-276 ~ia~ystok, poland, tel/fax +48 85 619742. in vitro studies demonstrated that paf activates polymorplionuclear leukocyte (pmn) 5-1ipoxygenase, but the generation of leukotrienes (lts) is critically dependent on exogenous arachidunic acid (aa) disposal (f. grimminger et at., mol. pharmacol. 41:757, 1992) . we investigated the features of paf-evoked lt synthesis in a model of pulmonary microvascular leukostasis, in the presence and absence of intravascular n-3-and n-6 -prescursor fatty acid supply. human neutrophils were infused into isolated, ventilated and perfused rabbit lungs to achieve microvascular sequestration of ligand-responsive leukecytes. leukotrienes (lt) and hydroxyeicosatetra(penta)enoic acids (het(p)e) were quantified by itplc techniques. intravascular application of paf (5 um) or arachidonic acid (aa;ioum) provoked the generation of limited quantities of 4-series lts and 5-hete (total sum of 5-1ipoxygenade products rd. 7 and rd. 27 pmol/ml.; both in pmn-preloaded and non-preloaded lungs). combined administration of pat r and aa caused amplification of 5-1ipoxygenase product formation with predominance of cysteinyl-lt synthesis both in lungs without (total sum rd. 67 pmol/ml) and, much more strikingly, with pro-application of neutrophils (total sum rd. 308 pmofml). eicosapentaeooic acid (10 um) elicited exclusive generation of 5-series lts and 5-hepe (total sum rd. 82 pmol/mi). dual stimulation with paf and epa provoked amplification of epa-derived 5-1ipoxygenase product formation, again with predominance of cysteinyl-lts, in lungs without (total sum rd. 224 pmul/ml) and in particular in those with preceding microvascular pmn entrapment (total sum rd. 545 pmol/ml). we conclude that the paf-evoked 5-1ipoxygenase product formation in the neutrophil-harbouring lung capillary bed is critically dependent on intravascular precursor fatty acid supply, with epa representing the preferred substrate as compared to aa. pmn-related transcellular eicosanoid synthesis is suggested to underly the predominant generation of cysteinyl-lts. these findings may be relevant for lipid mediator profiles provoked by infusion of n-3-versus n-6enriched lipid emulsions in diseases with pmn-related inflammators events. adult respiratory distress syndrome (ards) involves damage to the alveolar epithelium and microvascular endothelium. products released from neutrophils (pmn) are thought to be among the chief causes of this damage, while the role of mononuclear cells (mnc) is uncertain. we studied patients at risk of or with acute ards. we measured the concentration of myeloperoxidase (mpo) and elastase (el) within circulating pmn as an index of pmn activation, elastin degradation products (edp) in the plasma as an index of tissue damage, as well as the cytotoxici~y of pmn and mnc to endothelial cells (ec) in vitro. cells and plasma were prepared from venous blood of healthy controls; or systemic arterial blood of patients on ventilators in intensive care but not at risk of ards; at risk of ards because of sepsis; at risk because of multiple transfusion/major trauma; or with acute ards. lung injury, multi-organ failure and apache h scores were recorded. to measure cytotoxicity, human umbilical vein ec were labelled with cr-51, incubated with either pmn or mnc for 18 hours, and the supernatant counted. the adherence of the pmn and the mnc to the ec was also measured. standard assays were used to measure mpo and el in the pmn, mad edp in the plasma. the mnc cytotoxicity did not differ between the groups. the pmn cytotoxicity was higher in the transfusion/trauma patients compared to each other group, which did not differ from each other. there was no correlation between cytotoxicity and adherence for any group. the mpo and the el concentrations were significantly decreased in the pmn from each of the patient groups, including the patient controls, compared to healthy controls. the edp were only increased in patients at risk of or with ards. thus pmn degrannlation occurred in patients not at risk of ards, but tissue damage as measured by increased edp only occurred in patients with or at risk of ards, suggesting that factors additional to pmn activation and degranulation are required for the progression to ards. there was no correlation between the parameters measured and the indices of disease activity, or outcome. these measures of pmn and mnc activation and function are not of value as prognostic indicators in patients with or at risk of ards. the liver is made up of several different cell types which subserve distinct functions in vivo. in addition to the different functions of the distinct cell types, it is now evident that even within the population of hepatocytes substantial functional heterogeneity exists. this heterogeneity occurs in a very organized fashion in the normal liver based upon the position of the hepatocyte in the acinus. although it has been proposed that the acinar heterogeneity of hepatocytes arises from migration of immature hepatocytes from the periportal region to the perivenous region where they become senescent and die, many of the heterogeneous responses can be accutely reversed by reversal of the oxygen gradient via retrograde perfusion. under normal conditions in which the acinus is exposed to unidirectional flow, the periportal hepatocytes are exposed to relatively high levels of oxygen, substrates and hormones. as these substances are extracted along the aeinus their concentration decreases leaving relatively low concentrations for the perivenous hepatocytes. in the normal liver, the heterogeneity of response is attenuated by extensive communication via gap junctions. the major liver gap junctions are made up of hexamers of connexin 32 (cx32) which forms channels between cells capable of pas.~ing any molecule smaller than 1000 d. coupling is sufficient to allow diffusion of molecules such as atp or camp from one hepatocyte into > 20 adjacent hepatocytes within 60 seconds. during stress conditions such as endotoxemia or zymozan inflammation, expression of cx32 is markedly decreased while the secondary gap junction protein cx26 is either unchanged or even increased. while cx 26 readily effects electrical coupling, molecules > 350 d pass only very slowly. this would result in restriciton of transmission of moecules the size of atp or camp. since inhibition of gap junctions also attentuates metabolic response to hormone or nerve stimulation, it is evident that modulation of hepatocyte hetereogeneity by gap junctions must be considered in determining the mechanisms of metabolic alterations during stress. already minor haemorrhage decreases portal venous blood supply to the fiver and the reduction in portal blood flow becomes more pronounced with more profound btood loss. severe hacmorrhagic hypovolemia also reduces hepatic arterial blood supply which, however, is maintained over a vide range of haemorthage. the net effect of blood loss is a reduction in liver oxygee supply and this reduction is in proportion to the vulume iossed. however, oxygen supply to the liver exceeds the demands of the normal liver and this is the ca~ stilt following reduction of 30 % of blood volume. the situation in sepsis is more complicated. po~l venous supply to the liver is redur.~i fairly early following normovolemic sepsis while hepatic arterial blood supply is maintained at le,~t initialiy, oxygen saturation might be maintained in arterial blood but may also be slightly reduced during sepsis, oxygen saturation of portal venous blood is significantly reduced during sepsis due to increased extraction of the intestines. therefore oxygea delivery to the liver during sepsis becomes sigalfkzntly reduced. at the s,~ne time and for mai.v.ly unknown reasons the need for oxygen becomes significantly increased in the ~-~ptic liver. as a consequence liver oxygen consumption becomes flow dependent and the liver is likely to suffer from ischemia during septic conditions. $94 although liver failure is well recognized in sepsis, it is generally thought to be a late complication following pulmonary and renal failure. jaundice, hypoglycemia, encephalopathy and bleeding secondary to low levels of liver-synthesizing clotting factors are, however, signs of rather severe end-stage hepatic failure. furthermore, elevated liver enzymes (sgot and sgpt) represent hepatucyte damage and not hepatocellular dysfunction. in view of this, a more sensitive indicator of hepatic function is desirable in order to detect early hepatic abnormality. in this respect, indocyanine green (icg) is a tricarbocyanine dye that possesses several properties which makes it particularly valuable inthe assessment ofhepatic function. this dye is bound m albumin and is cleared exclusively by the liver through an energydependent membrane transport process and is nontoxic at lower doses. we propose that maximal velocity (vm~,) of icg clearance is a valuable measure of active hepatocellular function, since the total concentration of functioning receptors is directly proportional to vm~. we have utilized a fiber optic catheter and an in vivo hemoreflectometar to continuously measure the administered icg in vivo and consequently determine its clearance without the need of blood sampling. using this technique, we have found that in the early stages of sepsis (i.e., 2 and 5 h following cecal ligation and puncture), the vm~ and kinetic constant (k=) of icg clearance was significantly depressed. it should be noted that at this stage of sepsis, there was no elevation in serum enzyme levels. furthermore, hepatic blood flow and cardiac output increased at the above mentioned time points. thus, the extremely early depression in active hepatocellular function in sepsis, despite the increased hepatic blood flow and cardiac output, may form the basis for cellular dysfunctions leading to multiple organ failure during sepsis. additional studies indicated that following hemorrhage, active hepatocellular function was markedly depressed. this returned to prehemorrhage levels after ringers lactate resuscitation, however, this function was not maintained and decreased significantly after fluid resuscitation. nevertheless, the depressed active hepatocelinlar function following hemorrhage was markedly improved by post-treatment of animals with either atp-mgci2, peutoxifylline or diltiazem. thus, the use of icg clearance provides an early sensitive indicator of hepatic abnormality during sepsis and following hemorrhage and this method should be used, not only experimentally, but also in the clinical arena for the early detection of hepatocellular abnormality. although multiple organ dysfunction syndrome (mods) remains a major cause of mortality and morbidity in intensive care units, very little is known about the mechanisms that precipitate its development. since an episode of inadequate tissue oxygenation is considered to be the trigger for mods, we have proposed that a primary localized injury such as ischemia/reperfusion may be sufficient to cause a change of gene expression of remote and apparently unaffected organs. such modulation of remote organ gene expression may decrease the organ's tolerance to a subsequent stress contributing to the development of mofs. to test this hypothesis, rats were subjected to hepatic regional ischemia by clamping the blood flow (hepatic artery and portal venous inflow) of the left and median liver lobes. intestinal congestion was prevented by allowing flow through the smaller right and caudate lobes. after 60 minutes of ischemia, the clamp was removed and the blood flow restored. the animals were allowed to recover for 1, 4 and 24 hours. kidneys were removed, total rna was isolated and poly(a) ÷ selected by affinity chromatography on oligo(dt) columns. message was in vitro translated using rabbit reticulocyte iysates in the presence of radioactive amino acids. the gene products (radiolabeled polypeptides) were fractionated by two dimensional gel electrophoresis, and visualized by fluorography. analyses of the two dimensional fluorograms indicate that there is a dramatic change in the electrophoretic pattern of in vitro translated products in samples corresponding to kidneys obtained after 60 minutes of hepatic ischemia and 24 hours of reperfusion with respect to kidney samples obtained after sham operation or from control rats. the latter were not subjected to any surgical manipulation. these studies suggest that the gene expression of the kidneys is specifically modified after a remote organ injury (hepatic ischemia/reperfusion). we speculate that this change of gene expression in kidneys after an indirect injury may be part of the early events leading to the development of mods. a priming event, e.g. local ischemia, in combination with a second insult, e.g. sepsis, may amplify a host's response and lead to multiple organ failure. to better understand the mechanisms involved in the pathophysiology, male fischer rats were subjected to 20 min of hepatic ischemia followed by reperfusion (rp) and injection of 0.5 mg/kg salmonella enteritidis endotoxin (et) at 30 min of rp. et injection potentiated the postischemic liver injury as indicated by histopathology and an increase of plasma alt activities from 870+122 u/l (i/rp only) to 3900+470 u/l at 4h rp. inhibition of kupffer cells (kc) with gadolinium chloride (7 mg/kg) attenuated liver injury in this model by 70%, however, monoclonal antibodies (cl26, wt3) directed against adhesion molecules (132 integrins, cd18) on neutrophils had no effect on the injury despite the substantial accumulation of neutrophils in the liver at that time (467+52 pmns/50 hpf; baseline: 13+3). isolation of kc and neutrophils from the postischemic liver indicated a 10-fold increase of the spontaneous superoxide formation only in the kc fractions [3.85+0.68 nmol o2-/h/10%elts (kc2); 6.82_+0.92 (kca) ] at 4h rp compared to control cells. in addition, stimulation with phorbol ester or opsonized zymosan revealed a substantial priming of kc for reactive oxygen formation. in contrast to the short-term experiments (4h), the antibody wt3 (4 mg/kg) attenuated liver injury by 90% at 24 h of rp and improved survival. conclusion: liver injury during the early rp phase is mediated mainly by kc generating excessive amounts of reactive oxygen while neutrophils are primarily responsible for organ damage during the later rp period. (es-06091 and gm-42957) tumor necrosis factors (tnf) are cytokines which are cytotoxic towards some tumors in vivo and certain tumor lines in vitro. moreover, these polypeptides are powerful immunomodulators and have been found to be distal mediators in several models of septic shock and septic organ failure. one of the best-characterized experimental systems is the hepatitis caused by lps or tnf in galactosamine (galn)-sensitized mice. here we describe a cell culture system, in which the direct toxicity of tnf towards mouse hepatocytes was examined. the toxicity of tnf, as determined by ldh-release or formazan-formation, was dose-and time-dependent. the threshold of toxicity was 10 ng/ml, which corresponds to serum concentrations found in mice after lpsinjection. toxicity was only observed in hepatocytes sensitized with transcriptional inhibiters such as galn, actinomycin d (actd) or cxamanitin. sensitization was neither observed with different translational inhibitors nor with various other metabolic inlaibitors or toxins. inhibitors of protein synthesis or protein processing such as cycloheximide, puromycin, tunicamycin and ricin protected actdsensitized hepatocytes from tnf-induced cytotoxicity. tnf induced apoptotic changes and dna-fragmentation in sensitized hepatocytes which is in line with the above findings that cell death is dependent on protein synthesis. thus tnf may be a trigger of programmed cell death during inflammatory organ damage. with the purpose of studying the role of complement activation in tissue injury after ischaemia and reperfusion we blocked the complement cascade in a model of rat liver isehaemia and reperfusion, either by administration of soluble human complement receptor type 1 (scri), 25 mg/kg iv after vascular occlusion (n=8) or by depleting the complement system using cobra venom factor (cvf), 0.5 mg im, 24 and 6 hours before ischaemia (n=10). non-ischaemic rats (n=8) and ischaemic non-treated rats (n=15) were used as controls. the experimental procedure consists of the temporary interruption of arterial and portal blood flow to the left lateral and medial lobes of the liver during 45 minutes, followed by reperfusion, recording the liver blood flow and haemoglobin saturation with a laser doppler flowmeter and photometer during one hour after declamping; alt levels were assayed and immunoperoxidase stainings for c3 and c9 were performed. there were statistically significant differences between the experimental ~roups and the untreated ischaemic control group in terms of post-isehaemic blood flow (p<0.001) and haemoglobin saturation (p<0.001). c3 and c9 were present in the endothelium of the ischaemic control group. no deposits of c3 or c9 were found in the cvf group. few c3 and no c9 were found in scri treated rats. these results show that the effect of reperfusion injury in the rat liver is ameliorated either by depleting complement with cvf or by regulating complement activation with scri. hepatic dysfunction, a major cause of mortality following hemorrhagic shock, has not yet been well characterized. the present study was designed to assess the effects of liver blood flow and cytokine levels on hepatic function following resuscitation from severe hemorrhagic shock in normal and cin-hotic rats. methods: aftor pentobarbltal anesthesia, 23 control and 38 cirrhotic sprague-dawley rats were subjected to severe hemorrhage to reduce their systolic blood pressure to 50 + 5 mm hg. this level of hypotension was maintained until the skeletal muscle transmembrane potential (era) depolarized by 25%.; the animals were then resuscitated with ringer's lactate solution in three times the volume of the shed blood. serial blood samples for tumor necrosis factor (tnf) determination (a modified flow-cytomeuic wehi cell bioassay) were obtained at baseline, during hemorrhage and following resuscitation. liver blood flow measurements by low dose galactose clearance (glc) and functional bepatocyte mass (fhm; defared as galactose elimination capacity [gec] from the zero order portion of the plasma disappearance curve following an intravenous galactose bolus [500 mg/kg], divided by liver weight) were measured before shock and after resuscitation. results: higher survival rates (p < .05) were observed in control as compared with cirrhotic rats. shock produced a significant reduction in gec (to < 0.01); fhm (19 < .01); and liver blood flow (p < 0.01) in normal and cirrhotic rats. decreases in gec and fi-im were greater (p < 0.05) in cirrhotic rots. tnf levels were higher (p < 0.05) in cirrhotic rats at baseline and during induction of shock. pre gap junctions provide pathways for metabolic signals between cells. in the liver, the majority of gap junctions are composed of connexin 32 (cx32) polypeptide subunits, and are regulated by gluconeogenic hormones. since sepsis and other inflammatory states alter hepatic glucoregulatory control, we have evaluated the contribution of gap junctional conductance to the metabolic dysregulation in the liver. an acute inflammation was induced in rats by injection with e. coli endotoxin (lps lmg/kg). northern blot/hybridization analysis of total rna isolated from livers after endotoxin injection show a decrease in the steady state transcript levels of cx32 to 18% of sham controls. immunostaining of liver sections using anti-cx32 revealed punctate fluorescent staining on the plasma membrane at regions of call-cell contact in saline injected animals, whereas, staining was only observed in cytoplasmic vesicles 6 hrs after animals were treated with lps, suggesting the internalization of cx32 without replacement on the cell surface. the staining was quantitated and expressed as % of pixels above threshold. at 6hr post injection 3.6% ofpixels exceeded threshold, compared to 16.2% in sham controls. functional gap junctional communication was assessed by dye coupling using lucifer yellow in an isolated perfused liver under intravital fluorescence microscopy. dye diffusion was markedly decreased 6hr after endotoxin injection. this suggests that decreased metabolic coupling after lps injection results from decreased gap junction abundance. the present data suggest that metabolic dysregulation during sepsis may arise in part from changes in intercellular communication caused by a decrease in gap junctional expression and communication. given the marked metabolic heterogeneity of hepatocytes with respect to acinar location, metabolic signaling via gap junctions most likely serves to moderate this heterogeneity, contributing to a coordinated metabolic response. altered cellular ca 2÷ regulation might be a critical step in organ dysfunction during sepsis and ischemia/reperfusion events. the aim of the present study was to evaluate hepato-ceuular ca 2÷ regulation in isehemiah'eperfusion after hemorrhage and to assess effectiveness of tnfc~-monoclonal antibody (tnfo~-moab). methods. male sprague-dawley rats (250g, n>_6/group; pentobarbital 50 mg/kg) with hemorrhage for 60 rain at 40 mm hg. reperfusion by ringer's lactate (2 x maximal bleed out/60 min) and 60% of citrated shed blood. tnfcz-moab (tn3, ceutech, 20 mg/kg in 0.9% nac1) infused during flrst 5 min of reperfusion. at baseline, end of ischemia and 60 min of reperfusion, hepatecyte isolation by liver collagenase perfusion. " 45 hepatocyte incubation (30 mg w.w./ml) with caci 2 (0.05 2+ 2+ 2+ mbq/ml) for 60 rain (ca influx [slope, 1/mini; ca uptake [nmol ca /mg protein]) w/ and w/o epinephrine (epi, 100 nm). hepatecyte resuspension in radioisotope-free medium and farther incubation (exchangeable ca 2+ (ca2+ex) [nmol ca2+/mg protein]; ca 2+ membrane flux [nmol ca2+/mg protein'min]). during incubation, aliquots (100 ~tl) were centrifuged through oil/lanthanum gradient and acivity measured by scintillation counting. statistics: anova. mean + sem. results. hepatocyte ca2+ex and membrane ca 2+ flux were significantly increased at both, the end of ischemia (14.3+.9; 0.41+.05) and reperfusion (16.4+.9; 0.50+.2), as compared to sham-operated animals (7.6+_.5; 0.09+.02)(19<.05 ). tnfc~-moab treatment significantly prevented reperfusion-induced increase of ca2+ex (10+.8) and membrane ca 2+ flux (0.17+.03)(p<.01). fast ca 2+ influx was significantly increased by epinephrine in hepatecytes from sham-operated rats (0.23+.03 vs. epi: 0.52+.1, p< .05). this hormone effect was not observed in isehemia (0.47+.05, epi: 0.6!-_.2) or reperfusion (untreated: 0.13+.06, epi: 0.07+.01; tnft~-moab: 0.26_+.04, epi: 0.33+.07). conclusion. the present study clearly demonstrated hepato-cellular ca 2+ overload in ischemia and reperfusion as a result of hemorrhagic shock. analysis of membrane ca 2+ fluxes and hormone ca 2+ mobilization suggests disturbances of membrane ca 2+ transport mechanisms, e.g. through ca2+-atpases. reperfusion-induced oxygen free radical generation which affect exchange kinetics of cellular ca 2+ buffering compartments might also be operative. prevention by tnfct-moab indicates the pivotal role of tnf as an early inflammatory mediator of hepatocellular alterations in signal transduetion mechanisms and cellular homeostasis. although the precise mechanism has not yet been elucidated, bacterial translocation and endotoxin absorption have been frequently shown after burn, and have been postulated to be one of the underlying processes of sepsis. the purpose of the current study is to define the hemodynamic response of the liver to endotoxin release in burns, in correlation to bacterial translocation. twelve female minipigs, weighing 20-28 kg, underwent a laparotomy & transition time ultrasonic flow probes were positioned on the portal vein, the common hepatic artery, and the superior mesenteric artery. 6.5 fr catheters were inserted in the superior mesenteric vein and the left hepatic vein. a jejunostomy was also performed. after five days all animals were anaesthetized and randomized to receive 40% of tbs a third degree burn. eighteen hours after burn. 100 gg/kg e. coli lps was intravenously administered over 30 rain. ali animals were studied for additional 24 hours and then sacrificed. several recent data suggest that in severe injuries, such as shock state, the gradual activation of kupffer cells and the excessive release of destructive and immunosuppresive products from macrophages may contribute to the development of "multiple organ failure". in in vivo experiments in mice, the effect of kupffer cell phagocytosis blockade on the correlation between the tissue distribution of lps, endotoxin sensitivity and lps-induced tnf production was investigated. to depress the activity of the kupffer cells, gadolinium chloride (gdc13) or carrageenan was used. th~e studies indicate the dissociation of tissue localisation of cr jllabelled endotoxin and endotoxin lethalithy. both gdc13 and carrageenan depressed kupffer cell activity, but endotoxin sensitivity was enhanced only by carragenan treatment. however, there was a close correlation between the sensitivity to lps and lps-induced tnf production as measured in the serum, since lpsinduced tnf production was enhanced only by carrageenan treatment. on the other hand, gdc13 pretreatment significantly increased tnf production in the spleen. these results support our earlier findings that gdc13-indueed kupffer cell phagocytosis blockade leads to activation of the spleen, and may explain some of the immunological effects of gdc13. inositol(l, 4, 5) triphosphate (ip3) has been proposed as a second messenger for calcium mobilization. the addition of ip3 at low concentration has been shown to cause calcium release from intracellular microsomal store in rat hepatocytes. the effects of sepsis on the ip3 binding from microsomal fraction of rat hepatocytes during sepsis were investigated. sepsis was induced by cecal ligation & puncture (clp). control rats were sham-operated. three microsomal fractions (rough, intermediate and smooth) were isolated from rat liver. study of ip3 receptor binding was performed with tridium label ip3. the results shewed that the ip3 binding was significantly depressed by 40-47% (p<0.05) during late sepsis (18 hrs after clp), but not in early sepsis (9 hrs after clp). the ip3 binding depression during late sepsis was most significant on rough and intermediate endoplasmic reticulum (p<0.05), but not on smooth subfraction. since ip3 binding plays an important role in the regulation of intracellular calcium homeostasis in hepatocytes, an impairment in the calcium release due to depressed ip3 binding on smooth and intermediate endoplasmic reticulum during late sepsis may have a pathophysiological significance in contributing to the development of altered hepatic metabolism during septic shock. septic organ failure is currently recognized as an overactivation of the nonspecific immune system by bacterial stimuli giving rise to proinflammatory mediators. little is known about the mechanisms of the resulting cellular injury. here, a synergism is described between tnf as a major mediator of septic organ injury released by macrophages and hydrogen peroxide (h202) as a representative of reactive oxygen species as formed by e.g. neutrophils. rat hepatocytes are only slightly sensitive to either agent alone. when treated with a conbination of tnf and h#09 a stronq synergistic toxicity was found, especially w~e~ tnf-treatment preceeded challenge with h~o~. we have recently described a coculture model bfzrat liver macrophaqes and hepatocytes where lps induces hepatocyte cell death partially mediated by macrophage tnf release. when h202 was also employed in fhis more complex cellular system a similar synergism was found: the ecc~ of lps was 62 consecutive patients with liver cirrhosis admitted to the department of surgery over a 1 year period from january to december 1992 were studied for their complement profiles in relation to other parameters of liver function, the aim of the study was to determine if a direct correlation existed between low complement levels and end stage liver cirrhosis. cirrhotic patients were divided into child's a, b and c categories using child's classification. complement levels (c3, c4) were measured and functional assay for complement (ch50) were performed in each of these groupings in addition to normal blood donor controls. these results show that the qualitative c3, c4 and the functional chs0 complement assays have good predictive values in assessing deteriorating liver function• in particular, the functional assay for complement (ch50) showed marked impairment in child's c patients (p<0.000001) confirming the impaired immunological status of these patients. sera from this group of patients (child's c) were titrated with pig red blood cells (rbcs) in a haemolytic assay. the results showed that there were significantly less haemolysis of pig rbcs in these patients (p=0.0177) as compared to the controls. this findings strongly support an impaired immunological status in child's c liver cirrhosis and may explain the high incidence of sepsis as a terminal event in these patients. aim:kupffer cells(kc) have an importamt play to cause hepatocellular injury in sepsis, because these cells release many kinds of substances. we reported that oxygem radicals released by kcs stimulated by lipopolysaccharide (lps) caused hepatocellular injury. aim of this study is to investigate the relationship between imtracellmlar calcium(ca) concentration of cultured rat kcs stimulated by lps and release of oxygen radicals, and effect of prostaglandin e~ (pge~) on imtracellular ca concentration. production of acute phase proteins (c-reactive protein, crp, transferrin, tf) and £erritin (f) in rat hepatocytes (hps) and its dependence on extracellular matrix components were studied. hps isolated from the liver by collagenase perfusion were cultured at ~o5 per 0.5 ml medium fi2+dmem (1:1) with 10% fetal calf serum for 2 days on uncoated or type i collagen coated plastic surface or in the presence of dextrane sulphate in the medium. hps were stimulated by conditioned medium (gm) from i~ia-p or e. coli lps preineubated human blood mononuclear cells. production of crp, tf and f by hps was detected by elisa. it was found that both cms decreased tf synthesis in hps by 15-50% (p<o.os). the level of f synthesis didn't change or only slightly decreased. the addition of cms led to the enhancement of crp synthesis more than 2.5 times. the most reproducible results were obtained when plastics had been coated with collagen. dextran sulphate markedly increased crp synthesis. thus using this model of an acute phase reaction in vitro we found the enhancement of crp synthesis and the decrease of tf synthesis. our data correspond with nublished materials on the acute phase in vivo and point to relevance of the proposed model. metabolism of hepatocytes under traumatic illness has not been sufficiently investigated. therefore we have applied absorption and fluorescent cytophotometry techniques to study the content of rna, glycogen and its fractions in hepatocytes of patients with severe mechanical trauma, both with and without endointoxication at various stages. for these purposes slides were prepared from the repeated aspiration biopsy material according to special techniques (kudryavtseva et al., 1983) . slides were stained by gallocyanin-chromalum to measure rna or underwent fluorescent pas-reaction to measure glycogen and its fractions (kudryavtseva et al., 1970 (kudryavtseva et al., ,1974 the ability of 02 metabolites derived from the xanthine-xanthine oxidase system to inhibit mitochondrial function was examined using freshly isolated rat liver mitochondria. under uncoupled conditions, mitochondria exposed to free radicals exhibited a significant decrease in 02 consumption supported by nad+-iinked substrates, but showed almost no change in 02 consumption in the presence of succinate and ascorbate. the activity of electron transfer complex i (nadh oxidase and nadh-cytochrome c oxidoreductase) and the energy-dependent reduction of nad+ by succinate were unaltered by oxidative stress. exposure to free radicals also had an uncoupling effect at all three coupling sites. the degree of mitochondrial swelling was closely correlated with the inhibition of state 3 oxidation of site i substrates and with the increase in state 4 oxidation of succinate. the immunosuppressive agent cyclosporin a (cya) completely prevented the mitochondrial damage induced by oxygen free radicals (swelling, ca2+ release, sucrose trapping, uncoupling, and selective inhibition of the mitochondrial respiration of site i substrates). the same protective effect was found when ca2+ cycling was prevented, either by chelating ca 2+ with egta or by inhibiting ca2+ re-uptake with ruthenium red. these findings suggest that the deleterious effect of free radicals on mitochondria in the present experimental system was triggered by the cya-sensitive and ca2÷-dependent membrane transition, and not by direct impairment of the mitochondrial inner membrane enzymes. correspondence should be addressed to: naoshi takeyama the aim of this study was to clarify the critical role of reduced glutathione (gsh) on the progression of lipopolysaccharide (lps)induced liver damage of mouse. the conditions of gsh-depletion and -richness were produced by intraperitoneal administration of buthionine sulfoximine (bso), a specific inhibitor of gsh synthetase, and of gsh-monoethyl ester (gsh-me), respectively. gsh-me, which was esterified the caboxyl group of the glycine residue, is readily transported into cells and is hydrolyzed intracellularly; this leads to greatly increased the cytosolic and mitochondrial levels of gsh. bso and gsh-me were administered intraperitoneally 3 mmol/kg and 10 mmollkg, respectively, and lps (2 mg/kg) given 1 h later. hepatocytes were isolated by in situ perfusion of the liver with collagenase 6 h after lps injection. the degree of hydrogen peroxide (h202) synthesis and mitochondrial membrane potential were measured by flow cytometry using fluorescence probe dichrolofluorescein diacetate and tetrametyl rhodamine ethyl ester, respectively. the degree of mitochondria membrane permeability transition (mmp) was assessed by [014]sucrose entrapment. we found that lps treatment results in a decrease in hepatic gsh level and mitochondrial membrane potential, and an increase in h202 synthesis and mmp. lps administration to bsq-pretreated mouse worsened at[ these parameters. in contrast, gsh-me pretreatment ameliorates. all together, gsh may acts an important role in cellular defence against lps-mediated liver damage, and mmp may result in the decrease in mitochondrial membrane potential. it has been shown, up to now, that disturbances of enzymatic processes in the cell organella are the basic factor of the changes taking place during endotoxin shock it has been observed that lysosomes are biochemically changed as result of the effect of lipopotysacharides of gram negative endotoxic bacteria. the purpose of the experiment was: 1.evaluation nfthe course of the ees due to the biochemic changes 2.determination of the influence of the ees on the activity of lysosomal enzymes in liver cell 3.deterrmnatian of the influence of h and d on the activity of some lysosomal enzymes in liver cell during ees material and method: examinations ware carried out on 30 dogs. the shock was evoked by i.v. administration of endotoxin e.coli. the dogs were divided into 5 groups: icontrols, i[ -dogs with untreated shock, iii-dogs in shock treated with h, ivdogs in shock treated with d, v -dogs in shock treated with h and d. the following parameters were determined: 1.activity of acid phosphatase in the venous blood serum. 2-6.total and free activity of acid phosphatase and catepsins in the full liver cell homogenate, in mitochondrial-and-lysosomal fraction, and in the superuatunt (all after 4 hours of the experiment). conclusions 1. essential increase of activity of lysosomal enzymes was observed in the ees. 2. increased activity of lysosomal hydrolases in the liver cell homogenate corresponds, as a result of the endotoxin effect, with increased enzymatic activity in the peripherial blood. extensive investigations from our laboratory of the pathophysiology of hepatic no-flow ischemia (45 min) -reperfusion injury demonstrated substantial kupffer cell activation with reactive oxygen formation during the first few hours of reperfusion as indicated by increases in plasma glutathione disulfide (gssg) levels in vivo (am. j. physiol. 260: g355, 1991) and enhanced superoxide formation by kupffer cells (kc) isolated from the postischemic livers (free radic. res. commun. 15:277, 1991) . the early kc-induced injury initiated the later, primarily neutrophilmediated reperfusion injury (faseb j. 4: 3355, 1990 ). to test whether or not similar mechanisms are operative during hepatic ischemia induced by hemorrhagic shock, male fischer rats (230-250 g) were subjected to 60 rain of hemorrhage (mean arterial pressure 40 mm hg) followed by resuscitation (rs) (reinfusion of shed blood). hepatic atp levels declined from 2.3+0.2 txmol/g to 0.24_+0.08 (60 min shock) indicating severe ischemia, and recovered to 1.04--0.2 at 90 min rs. to test for potential oxidant stress during rs, plasma gssg conc. were measured. although gssg levels increased by 120% compared to baseline levels (0.69!-_0.31 i.tm), there was no significant difference to shamoperated controls. spontaneous superoxide formation of isolated kc was 0.32+0.05 nmol o2-/min/10ecells (kc2) and 0.51+0.11 (kc3) in controls and did not change significantly after shock and 90 rain rs. addition of phorbol ester or opsonized zymosan to isolated kc did not indicate a priming of these cells. conclusion: in striking contrast to our previous findings with hepatic no-flow ischemia, there is no evidence for a significant kc activation after 60 min of hemorrhagic shock and up to 90 rain of resuscitation. objectives-this study investigates morphometric changes in kc nltrastmcture which might account for this diminution in phagacytosis. materials and methods -three groups of wistar rats were studied: control, sham-operated [sham] and bile duct ligated [edl] . after 3 weeks animals were sacrificed and livers were "perfusion fixed" with 3% glntaraldehyde and processed for transmission electron microscopy and image analysis. results -in the the bdl group kupffer cell numbers were greatly increased. biliary ductular proliferation was evident and sinusoidal spaces were compromised. eleven nuclear and cytoplasmic parameters were measured and statistically significant results are summarised in the results: ascorbic acid levels were slightly elevated after the hs and returned to basal values after 120 rain. glutathione concentrations were increased significantly after the hs and the gsh levels kept rising continuously to as much as 4-fold of basal levels at the end of 120 min. mda showed no significant variation before and after the hemorrhagic shock. plasma concentrations of ratine, teucine, isoleucine, phenylalanine, proline, threonine and serine showed significant increase over basal levels during the whole ischemic period. glutamic acid was slightly elevated at the onset of hs and remained the same for the rest of ischemic period. hydroxyproline was significantly below the basal value at the mid-point of ischemic period. alanine, glycine, histidine and aspartate did not change significantly throughout experimental time course. conclusions: (1) oxidative stress might not be severe in the systemic hemorrhagic shock for pigs since no significant variations were observed for ascorbic acid and mda (2) the increase of gsh might be that it was released from hepatocytes when the animal was under systemic ischemia. (3) celzain amino acids might be acting as transmitters in the event of ischemia. however, fm"lher investigations are needed to clarify the detailed mechanism in regard with antioxidants and amino acids. dr. fang-ku p'eng taichung veterans general hospital talchung, taiwan 407, r.o.c. jia shi yong, huang zhi oiang and men xian jun. in patients underi~ent major hepatic surgery,obstructive jaundice has been implicated with fnoreased susceptibility to sepsis and liver failure. jaundice was said to he associated with derangement of iamune function and result in expression of pr,:.inflar...aatory cytoki~es that cause hepatooellular damage. this study is ~.o investigate the effects ,.,t lip,~polysaccharide(lps)rats. jaundice were produced in healthy ~istar rats of either sex weighing 180-220 m~ by common hi l~ duet l igation(bl)l).seven days post l igation, rats were given lps(236.gg.10)0.7rag/kg intraperitoneally. at the end ,:,f l,gand 5 hours after lp$ ehanllenge, liver were removed and prepared for forzen sections immediately. i)emonstration of inf in liver parenchyna were performed by method of abu i~munohistochemistry using r~onoolonal antibody against tnf, leve of mrna for tnf ~ere measured by in-situ hybridization using biotin-labeled edna probe. results sho'~ed that tnf stained granules appeared in kup~'fer eell~ and polymorphonuclear leukocytes(pnnl,but not in hepatoeytes nor endothelial eel is. they ~'ere located within cytoplasms and peaked at 3-5 hours after lps ohal lenge. there was vocomitant tnfmrna expression but peaked earlier 1 hour post lps challenge. tnfmrna vcere detected notably in necrotic area and barely in bdl rats ~ithout lps challenge. it is therefore postulated that production of tnf by inflammatory effeetor cells-kupffer cell and phn in site, in the obstructive jaundice rats during sepsis may atribute to the hepatocellular damage. it also provide evidence for the beneficial effects of early release of biliary obstruction. in the present study, an animal model of a0c in wlstar rats was developed by tigatlng the common bite duct and injecting d.3mt solution of e. goli % b, (g×10' efm/mt) into the bite duct. the mortality rate was 40~ at 48h after aoc. at 6, 12, 24, 48h after aoc, kcs were isotated and cultured atone or cocuttured with hepatocytes to evaluate the modulation effect of kgs on hepatoeyte protein synthesis. the results showed that tactodehydrogenase leakage was increased progressively as the injured k~ function and structure developed. tnf and il-i tn the supernatant were detected with the peak values at 12h after aoc. at 48h after aoc, ~-teueine incorporation was obviousty decreased in the normal hepatocytes eocuttured with stimulated kcs compared to the uormai hepatocyte cultured group (p<8.01), but it was slgnificantiy increased in the group cocultured with normal kcs. in the a0c group, 'h-teuclne ineorporatien was decreased progressively in the injured hepatoeytes cocuttured with stimulatedkgs and it was markedly elevated when cocuttured wlthnormat kcs at 48h after a0c (p<0. og). it is concluded that the impaired hepatocyte proterin synthesis was directly mediated hy the stimulated kc function during aoc. such a mechanism possibly may be involved in the patho0ensis of hepatic dysfunction and m0f following h06. " the project supported by nsfc. in the past few years it has become evident that cytokines are implicated in various pathophysiological mechanisms. therefore measurement of cytokines and their potential inhibitors in biological fluids may be helpful in diagnosing and monitoring of diseases. however, dependent on the type of assay used investigations performed in different laboratories have often yielded conflicting results. in order to assess reliability and reproducibility of cytokine measurements two comparative studies for the determination of cytokines in biological fluids were launched by several investigators interested in this field: one national austrian ~tudy and one european study in the context of the european workshop for rheumatnlogy research. serum and synovial fluid samples were distributed by the organisers, and participants had to measure one or several of the following cytokines in all samples: il-1, il-2, 1l-6, i58, tnf-alpha, ifn-gamma, gm-csf, and the soluble receptors for il-2 and tnf; both commercially available immunoassays and home-made assays were allowed. so far, the main conclusions emerging from these preliminary studies are: (1) the same immurzoassay (elisa) yielded comparable results in different laboratories; (2) immunoassays from different manufactureres usually measured the highest concentrations in the same samples, yielded similar relative values but disparate absolute values; (3) assays for soluble receptors yielded less discrepant results; (4) some assays seem to be more suitable for measuring cytokines in biological fluids than others. the mean retrieval of exogenous recombinant human cytokines was approximately 50% for most cytokines tested. however, it must be borne in mind that natural and recombinant cytokines may behave differently in immunoassays. this raises the question as to the necessity of setting up international standards for all cytokines. possible interferences by serum components such as (lipo)proteins, complement, rheumatoid factors, etc., which may either decrease assay sensitivity or yield false positive results, must also be considered. in addition, natural cytokine binding proteins (e.g. soluble receptors) might interfere with cytokine determination in immunoassays. the problem of immunoassays versus bioassays has not yet been approached, but is without any doubt worth indepth investigation. thus, these studies are a promising first approach to deal with the difficulties of cytokine analysis in biological samples, and it is hoped that they will help to overcome some of the major methodological problems in the near future. tumor necrosis factor (tnf) is a pleiotropic cytokine which plays a key role in a number of immunologically mediated disorders like septicemia or cachexia. tnf receptors are found on the surface of a variety of cells, where they provide molecular signals for the cytokine effect. there exist two types of soluble tnf receptors (stnf-rs), tnf-r p55 and tnf-r p75. these stnf-rs can compete with the cell surface receptors and block their activity. the expression on and shedding from the cell surface of stnf-rs is enhanced by interferon gamma. increased concentrations of stnf-rs can be found in patients with infectious as well as malignant disorders. in patients undergoing immune stimulatory therapy with interleukin-2 and interferon alpha a simultaneous increase of tnf and its soluble receptors can be seen. in patients with hiv infection a strong correlation exists between the levels of stnf-rs and markers of immune activation like neopterin or beta-2-microglobulin. likewise patients with hematological disorders show elevated stnf-r levels. patients with weight loss have higher concentrations than patients with stable weight. the final value of stnf-rs within the complex network of cytokines is not yet clear. however, there exist striking parallels between infectious and malignant disorders pointing to a key role of endogenous immune activation. biochemicatly, d-erythro-neopterin derives from guanosine triphosphate. in vitro studies show that large amounts of neopterin are produced by human monocytes/macrophages stimulated with interferon-j (ifn-j). neopterin is biologically stable, and its halflife in the blood seems to solely depend on renal excretion. specimen for neopterin determination can be stored without special precautions. increased concentrations of neopterin have been described in body fluids of patients suffering from diseases which are apparently associated with an activated cellular immune response and with enhanced endogenous formation of ifn-~', e.g,, 1 ) increasing neopterin levels predict rejection episodes in allograft recipients 2) virus infections induce increased neopterin concentrations, and the degree of neopterin elevation predicts prognosis in these patients which is particularly true in hiv-1 infection 3) in autoimmune disorders such as systemic lupus erythematosus or rheumatoid arthritis neopterin levels reflect activity of the disease 4) increased neopterin concentrations in patients suffering from certain types of cancer indicate poor prognosis. significant associations between changes of neopterin and decrease of hemoglobin as well as the development of weight loss and cachexia, e.g., in cancer patients and in patients with hiv-1 infection, support the concept that endogenously formed cytokines such as ifn-~" and tumor necrosis factor-~( are involved in the pathogenesis of such symptoms. in conclusion, neopterin monitoring is a sensitive tool to detect the activation status of the cell-mediated immune system in vivo. repair and healing or perpetuation of acute inflammation is characterized by a complex network of interacting cellular and humoral defence mechanisms. of the numerous inflammatory mediators/effectors investigated hitherto, the proteolydc lysosomal enzyme pmn elastase has turned out to be highly destructive when released extracellularly thus contributing considm:ably to organ dysfunctions in severe posttraumatic and postoperative courses. besides various cytokines, elastase in complex with its main antagonist c¢ 1-proteinase inhibitor (a 1pi) is also supposed to induce the shedding of intercellular adhesion molecules from inflammatory cells (pmns, monocytes/macrophages, endothelial cells). these soluble adhesion molecules may modulate the inflammatory process in many different ways, e.g. via acting as chemotaxins, blocking neutrophil activation or competing with the membrane-bound forms in cell-cell adhesion. thus, measuring circulating or local levels of elastase-a 1pi and soluble adhesion molecules (icam, e-selectin,-pselectin, l-selectin) may be a helpful tool in judging the severity of an acute inflammatory process. in several clinical studies on surgical patients suffering from multiple trauma and/or sepsis we could demonstrate that the measurement of these parameters in consecutive plasma samples and local body fluids was highly valuable for early diagnosis and prognosis of multiple organ failure. prof. dr. m./ochum, institut fdr klinischechemie und biochemie, lmu miinchen, nufibaumstrage 20, 80336 m0nchen, germany procalaitonin (proct) a 116 aa peptide is dramatically increased in the blood of patients with various sepsis and infections. the increase begins as soon as 3 hours after the andoto:edn release and readied maximal level with a 200 to 80b-fold increase 24 h after li~ edrmrdstration. prcc, alcitorfin response was temporally different from these of tnf~ and ii-6, that reached peak levels at 2 h and 3 h respectively. at the opposite of eytokine (tnfcz, ild, j, is), proct is a very stable molecule. the half iife of proct in ~he blood is surprising if we consider the half llfe (about nine mirtute) of mature ¢alcitonin (co. we have found that it is due to the binding of proct with a protein on the n-termktal part. thin complex of about 80 000 daltons is unable to cross the kidney and the c_n9 barriers and the half life is at least elghteen hours. this long half llfe is very useful in the evaluation of a sepsis. a very sensitive and specific sandwich in'ununoassay has been developped. the results cart be delivered in two hours. at time, we know that in the healthy adults, the values are less thau 0.1ng/ml. it is also clear that proct is not increased ~n patients wlth inflammatory dleeasea without lnfectlon. for instance, proct is not signlffcat3vely increased in purpura rhumatom, arthritis, crohn's disease, rectocolitis, also in various cancer patients with inflammatory components. am other interesting finding, is the absence of increase or a very low increase in the viral infections. at the opp0site, in all the patients with bacterial sepsis or in the malaria pat/ants, progt was dramatically increased, from i00 to 10 000 fold. p~oct can be useful to the follow up of patients with sepsis. actually, we don't know the origin of this proct production, in the united states, will also be discussed. the septest portion of this study will eventually enroll over 500 patients tentatively diagnosed as septic. the results of septest will be compared with commonly accepted symptoms and criteria associated with sepsis in order to determine the clinical utility of this endotoxin assay. preclinical results of patients and normals be presented as well as data on the time course of endotoxemia and clinical outcome of selected septic patients. preparation of dna libraries clifford w. schweinfest, ph.d. a fundamental tool of the molecular biologist today is the ability to work with purified dnas representing genes of interest with respect to the investigator's field of study. originally applied almost 20 years ago to the genetic dissection of simple organisms such as viruses and bacteria, molecfllar cloning was rapidly applied to study the more complex genetics of the mammalian cell. today, virtually any study of cell physiology which depends upon specific gene expression is approachable using the tools of the molecular biologist. therefore, stress induced gene expression (such as the heat shock gene, hsp70) or the regulation of the nitric oxide synthetase gene or the induction of cytokines and growth factors as a result of trauma or injury lend themselves to investigation at the level of the gene. during these workshops, we will discuss the means by which cdnas and genes are isolated, amplified and identified. we will discuss the strategies and methods for cloning cdnas and genomic dnas, for organizing such cloned libraries, for finding specific genes and for characterizing those genes. this speaker will focus on the techniques and considerations involved in the synthesis of cdna libraries. topics include rna isolation, mrna quality, reverse transcription, choice of cloning vectors and library quality. genomic dna library synthesis will also be discussed with respect to the vector/host systems available and applications which are specific to genomic dna. finally, polymerase chain reaction (pcr) will be discussed briefly with regard to its impact on dna cloning. the identification of the genes or gene products that have a role in cellular processes is fundamental to our understanding of genetic control and methodologies to achieve this aim are currently available. all levels of the various signal transduction pathways can b~ molecularly analyzed to define the mechanism by which critical steps in each pathway are achieved. the questions that are unresolved in each area of investigation serve to direct the scientist towards analyses of gene products or gene regulation of the gene itself. thus, selection of the type of library (genomic or cdna) to be used is dependent upon the specific goals of each investigator. to facilitate resolution of this question, an overview of the uses for the different types of libraries will be presented. at least four methodologies are currently available for screening a library, which are dependent upon specific interaction between nucleic acids, nucleic acids and proteins, antibodies and antigens or between different proteins. the use and advantages of each of these methodologies will be discussed along with a description of probe preparation. sequence methodologies required to begin characterization of gene clones will be presented. hopefully, the participants in the workshop will help define areas of emphasis and allow time for directed interaction to discuss specific applications based upon their own experimental systems. alterations of physiological conditions, such as those occurring after shock and sepsis, induce changes in gene expression of cells composing the major organ systems. the challenge is to detect and characterize these changes in geae expression and relate them to the injury. the development of cloning techniques has emerged as the methodology of choice. changes in gene expression are usually characterized by variations in the concentration of cellular messages because synthesis of the final product "the polypoptide" is usually proportional to the concentration of mrna. due to the rapid regulation of gene expression the cellular concentration of messages changes very quickly. this is commonly referred to as steady-state levels, a balance between transcription and degradation. steady-state levels of mrna can be detected in a single cell (in situ hybridization) or in total rna isolated from cells or organs and separated in agarose gels (northern blots). although analysis of mrna steady-state levels are very useful, they do not provide information about the mechanisms that regulate gene expression. changes in gene expression primarily occur at the level of transcription; characterization of the rna species being synthesized at a given time is performed by the nuclear run-off technique. when there is no a priori information about the gene that may be regulated after a particular situation such as sepsis, the total pool of messages present in cells at given time can be characterized by a combination of in vitro translation of the cellular messages and fractionation of the in vitro translated products by two dimensional gel electrophoresis. such approach permits the visualization of the general pattern of gene expression, a "finger print", of the physiologic state. in summary, researchers now have access to a great variety of techniques to identify and characterize genes expressed in response to a physiological condition that may be utilized as "molecular tools" to study the organism's responses to shock and sepsis. the acute phase proteins are a set of plasma proteins with greatly increased plasma concentrations during acute inflammations and after tissue trauma, e.g. after major surgery. the proteins counteract the source of injury, facilitate clearance of infectious particles by phagocytes, assist immune responses and initiate wound healing. the corresponding genes are expressed in liver hepatocytes and regulated by the inflammatory mediators interleukin 1 and interleukin 6 (ill, il6). class1 acute phase genes are mainly controlled by ill and by combinations of ill plus il6; class2 genes mainly by ii,6. the human c-reactive protein(crp), serum amyloid a(saa) and complement c3 genes will be discussed as examples of class1 genes, and rat c~2 macroglobulin as a prototypical class2 gene. both classes of genes use different types of ¢ytokine response elements in their promoter-proximal transcription control regions; class1 genes use the transcription factor nf-il6 or c/ebp as the key factor to mediate cytokine responsiveness; class2 genes use a different, so far unidentified factor to achieve responsiveness to il6. ongoing efforts to purify this factor, called the il6-response factor (il6-rf), from rat liver nuclear protein extracts will be described. the il6-rf apparently mediates the effects of il6 in a broad range of cell types, including b lymphocytes and other hematopoietic cells. it is therefore likely to be of equal general importance for gene regulation by il6 as nf-il6. the importance of transcription factors as potential targets for novel bioactive substances and possibly therapeutic agents will be discussed with the help of several recent examples. new methods for altering the germ lines of mice provide powerful tools for understanding the roles of individual genes in normal and pathological processes, and for reproducing specific genetic mutations that result in congenital disease. three approaches will be discussed. conventional transgeulc mice are frequently constructed using artificial transgenes that express genes from a promoter other than the normal gene promoter. this paradigm is used to produce very high levels of a gene product, for example, with a viral promoter, or to attempt to regulate gene expression using an inducible promoter. alternatively, normal promoter sequences can be hooked to a gene whose expression is lethal to the cell (e.g., diptheria toxin a chain) to ablate all cells expressing that gene. another approach uses an intact gene with its normal regulatory sequences. here, an elevated level of the gene product is delivered from the correct cells in the correct tissues at the correct developmental stages or in response to natural signals. this approach has been refered to as "genetic pharmacology," and provides a precise delivery of the gene product to the target. however, to be most effective, the transgene should contain arl regulatory sequences as welt as the entire genomic segment containing gene coding regions. the introduction of conventional transgenes is limited to roughly 50 kilobases, while genes containing all regulatory sequences frequently stretch over dozens or even hundreds of kilobases. to overcome this problem, procedures have been developed recently to introduce yeast artificial chromosomes (yacs) into mouse embryonic stem (es) cells. yacs can include over 2 mb of human dna, large enough to encompass the largest known human genes. es cells are also useful for targeted gene deletions and mutations. homologous recombination can be targeted to any known gene in es cells. when these modified cells are injected into a mouse blastocyst, they can be incorporated into all tissues of the resulting mouse, including germ cells, so subsequent generations will pass the genetic modification as an inherited trait. assessment of the effects of a null allele on development can provide valuable insights into its normal role. ultimately, these technologies may be adapted to human cells -not for embryonic intervention, but to provide modified stem cells for various tissues (e.g., gut, eye, hematopoietic system) which could then be applied to somatic therapies. with major illness/injury that respirswy faihne fi'equently followed sepsis, tilney el al nse, d the term "soqueutial syste~ failure" for patients with renal failure after solmir of ruptured abdominal aortic ~aeurysms, i then reviewed our experiences after inju~ and oporstion and suggested that multiple, progsesslve or sequential systems or organ failure was a syndrome to be reckoned with in the, 70's. eiseman et al then wrote about "multiple organ failure", especially with liver failure. polk el m found that renmte orgau failure often signalled occult anita-abdominal infection. fry et al ampbasized the role of uncontrolled infection in organ failure, border et ai described metabolic alterations with mof and used the term multiplesystems organ feilmo (msof). other early conltibutots to our knowledge of mof include faist, trunkey and miller, marshall and dimic&, cassone, catlleo, meakins and marshall, (}otis et at., mater and many others. mof or msof were used eommoifly. cerra coined the terms "post-trsumatic septic sfndromc" and "hyper metabolism otgau failure complex", and border et al, used lhe ex3~ression "gut origin seplic states" to illdioate important eurrem aspects of abe gt.'nerlc problem of organ failure and death, other terms include acute organ-system failure, multiple organ injury syndrome, post-traumatic multi-system organ failure nod post-~aumatic organ system infection s~dmmc (fi~sis), i bdieve the latin "maltiple organ failure" is clean, neat and says it all, now there is a proposal for a new set of torminolugios -mods and sirs. will they be helpful in the study and esro of patients? our speakers in this session will review thcsa proposals and the evidence as we strlvo for eoasensns. it has been well demonstrated that the administration of pro-inflammatory cytokines, and especially tumor necrosis factor (tnf) can result in a picture similar to septic shock with secondary multiple organ failure. it is also clear that tissue hypoxia can lead to organ damage. we believe that it is the interaction between the two phenomenons that can lead to mof. on the one hand, the inflammatory response is amplified in the presence of hypoxia. on the other hand, the release of the mediators of sepsis can increase tbe oxygen demand ef the tissues, alter their oxygen extraction and decrease myocardial contractility, and the combination of these elements accounts for the development of septic shock. this hypothesis is supported by two lines of evidence. first, mof is usually preceded by an episode of acute circulatory failure (even though frank circulatory shock may not be evident). second, recent experimental and clinical studies indicated that immunotherapy (especially antibodies to tnf) is particularly effective in the most severe forms of sepsis, associated with circulatory shock. hence, an effective way to prevent mof may be a combination of aggressive cardiovascular management and immunotherapy. development of the multiple organ dysfunction syndrome (mods) in the critically ill follows an heterogeneous group of stimuli including infection, sterile inflammmion, extensive tissue injury, ischemia, intoxication with a variety of substances, and immune system activation. whether the resulting physiologic abnormalities reflect a common pathologic process, or are simply a non-specific manifestation of tissue injury is unknown. moreover, the lack of clearly-defined functional criteria for the characterization of the syndrome on the one hand, and of reliable biochemical markers of its evolution on the other, has made attempts to define its epidemiology and natural history difficult. using a numeric scoring system to quanfimte the clinical severity of mods, we demonstrated that, although mods is more common in patients who have significant infection at the time of icu admission, a much stronger correlation is evident between the severity of mods and the subsequent development of nosocomial icu-acquired infection. furthermore the severity of mods correlates strongly with the severity of the clinical septic response, and with the degree of immunologic compromise evident on delayed type hypersensitivity skin testing. to determine the relative importance of the initial stimulus, and the host response to that stimulus, to the subsequent emergence of mods, we undertook a matched cohort study of 109 critically ill patients admitted to the icu with infection, matched by age, sex, and apache ii score to 109 uninfected controls. organ dysfunction scores were higher in patients admitted with infection, but were also significantly associated with the expression of a septic response at the time of icu admission, independent of the presence of infection. by stepwise regression analysis, the magnitude of the septic response was the most powerful predictor of the severity of mods in both infected and uninfected patients. these data suggest a model of mods in which a stimulus (eg infection) and the host response to that stimulus (eg the septic response) result in a state of altered systemic homeostasis (manifested in this example by immunologic dysfunction). moreover they suggest that the principle determinant of the emergence of mods is the response of the host, rather than the nature of the stimulus, and are consistent with the hypothesis that a stereotypie systemic response to an heterogeneous group of injurious stimuli underlies the pathogenesis of mods. arthur e. baue, m.d, ever since the attorapt 10 build the tower ef babel, as desclibcd in the old testament of the biblc, it has been difficult to got agreement o;x common definitions or language, although mof has served well, there will always he now classifications proposed with good reasons for them, thus mods and sirs are not the final oxpressinns in the lexico~aphic sweepstakes. attcmpts to develop standard animal shock and scpsis models (hemorrhage, endotoxin, foes[ pellets, cecal ligation and puncture, e. colt hlf0sioi0 to evaluate therapy have net been suoca.ssf~. clinical syndromes of mof, sepsis, sepsis syndrome and athers have not been aeacpte.d by all. our problem is that we arc tryittg tu d¢ffino a non-emily. mof or mods or sirs is net a disease or oven a syndrome. it is a series of complications of injury, disease and intensiva care which loads to death for many patients in intensive care units, it is not a fixed entity, but an ewilving picture. we have lumped (ugethet for therapy and definition a number of diseases and problems according to clinical manifestatieos rather than the eause of ihe problem, the search for unified mechanisms has not bean successful. will we benefit oleo from a more precise classification and definition of a non-outjty, a hodgu-podge of human disorders which have charsmeristic.s and manifestations of tissue injury, inflammation and infactian? re.hi clinical trials of potential "magle ballets" suggest that we should go in a different direction--instead of lumpors, we should be splitters, to seek effective therapy, we must fucus on specific disorders such as trauma, specific infections, inflammatory disease and chronic or acute organ damage (copd -renal failure, etc.), i will say more aboet this later ha the me.ethag. thcre is cue potentially important pert of the mods proposal, and that would be do~mcntation of organ dysfunction before fsiiure occurs and before tile need for external supporl dovelope. this could lead to hatter methods of preventing organ failure. preveution is ultimately the only answer to organ failure, m mof, mods and sirs. have wo reached a consensus?--only that we arc dealing wilh a diffl~x:lt probtolr looking for solutlons. acute lung injury (ali) and its most severe form, acute respiratory distress syndrome (ards) characterized by severe hypoxemia, diffuse infiltration of both lungs, a reduction of compliance and a wedgepresure lower than 18 mm hare related to direct or indirect injury. when aliresults from direct injury (toxic agents, lung contusion, pneumonia), the lesions of epithelial barrier and the activation of lung macrophages are the first events leading to the release of inflammatory mediators wh:ch are responsible for alteration of the membranar permeability and for invasion of the alveoli by fluids, proteins and cells. an inflammatory reaction develops, with injury to endothelial cells, adhesion of polymorpbonuclear leucocytes (pmn) and the release in blood stream of intlammatory mediators dispersing the inflammatory reaction to other organs and tissues. the failure of lung function also results into hypoxia in other organs, leading to tissular ischemia, triggering an inflammatory reaction leading to organ dysfunction. frequent complications of direct lung injury are septic pneumonia with endotoxin release, phagocytes activation and cytokine production disseminating inflammation. by this way, direct lung injury can be at the origin of the multiple organ dysfunction syndrome (mods), the frequence of which having been estimated to 40% and is increased when all has occured in patients alreadypresenting an organ dysfunction (immunodepresslon, cancer...). when ali results from redirect or extrathoraclc injury (trauma, infections, hypovolemic shock, acute pancreatitis...), inflammatory mediators and micro-aggregates released in particular organs or tissues reach the lung via the blood and lymph streams, are filtered in the organ or trapped in the lung capillaries, and are responsible for a local inflammatory reactaon (endothelial cell activation, pmn trapping and adhesion, platelet aggregation, release of mediators). sepsis acts by the way of endotoxiu and sequential eytokines release (tnf, ill,...) while trauma with important blood loss acts especially by the way of hypoperfusionreperfusion phenomena leading to a disseminatedinflammatory reaction and to a possible bacterial transloeation when intestinal hypopeffnsion is present. in these conditions of indirect injury, all (or ards) is a iocal early (often the first) manifestation of a general phenomenon of altered membrane permeability and inflammatory reaction, easily and rapidly recognized by its clin~cai signs. the iung is one of the numerous organs participating to the mods, in these conditions, the mortality rate is high, reaching more than 80% when sepsis is present or when at least 3 organs are implicated in mods. by direct or redirect injury, lung is so a target organ for mods and injured lung can be the cause of mods or simply a participant to this syndrome. jorge cervts-navarro main determinants for brain dysfunction are breakdown of the blood-brainbarrier and raise of intracranial pressure (icp), in second place decrease of systemic blood pressure, disorders of blood coagulation and respiratory function. the blood-brain-barrier of cerebral blood vessels can be opened by stroke, cerebral trauma, inflammation, convulsions, acute hypertension and changes in the blood osmolarity. the consequence is brain oedema, increase of lop and decrease of the perfusion pressure. a close correlation between intraventricular pressure and the fate of patients with severe brain injuries has been established. regional cbf values of 16 to 18 ml/100g/min are reflected in isoelectric eeg, and values about t5ml/100g/min, result in loss of somatosensory evoked potentials. for ionic pump failure the threshold has been determined by values of about 10 ml/100g/min and is reflected in massive release of intracellular potassium. in stroke and in brain trauma when the oedematogenous condition is initially localized the tissue pressure variance within and between the hemispheres lead to tentorial and falciform herniations are the common cause of death even before the critical threshold of intracranial pressure is reached. the increase of the icp over the level of the perfusion pressure leads to the arrest of the cbf. if this persists {or periods exceeding 6 seconds of global brain ischemia loss of conciousness and developing seizures appear. if it exceeds 10 min irreversible injuries of the brain appear. following cerebral ischemia and after severe head injury a tendency to increased coagulation can be detected. vascular occlusion may develop either as a result of local thrombus formation or from emboli caused by circulating platelet aggregates. the formation of microthrombi is triggered by the plateletactivating factor a mediator in central nervous injury also responsible for aggravation of posttraumatic brain edema. acute hemorrhagic leucoencephalitis and brain purpura are known to arise as complications of acute viral infections and following gram-negative septicaemia. institute of neuropathology, free university of berlin, hindenburgdamm 30, 12200 berlin, germany undoubtedly, attempts to quantify as objectively as possible the degree of dysfunction of the various organs is very valuable. however, cardiovascular failure has a peculiar place in the context of sepsis. the development of acute circulatory failure (shock) is of paramount importance as a cause rather than as a consequence of organ failure. it is therefore essential to clearly separate patients with and without circulatory failure. once this has been done, attempts to list the cardiovascular system among the other systems have been unconvincing. in a number of studies, cardiovascular complications are assimilated to a low systemic vascular resistance (svr), but since svr is basically the ratio of blood pressure over cardiac output, and since hypotension is already included in the definition of shock, then a low svr is basically equivalent to a high cardiac output, which is a characteristic rather than an ominous complication of severe sepsis. development of a low cardiac output unresponsive to fluids is usually a terminal event. it is not advisable to list other problems such as severe arrhythmias, myocardial infarction or tamponade as complications of severe sepsis. host defense dysfunction following trauma, shock and sepsis e. faist, g. f. babcock * major accidental, burn and operative injury often precipitates a profound multicentric immunologic depression that is believed to predispose patients to become anergic and thus to develop towards a higb probability of infection. anergy or a lack of immunologic reactivity stands for a total elimination of skin test reactivity and recall antigen responses in the inununocompromised patient. anergy following overwhelming trauma or major septic conditions results from a massive impairment of cell mediated immunity (cmi) together with a whole body malignant inflammationiike state. we and others have demonstrated clearly that the skeration of cmi following trauma is mainly due to the disruption of intact monocyte (moo) / t-cell interaction. within this phenomenon we see a shift of the cell ratio in the compartment of peripheral blood mononuclear cells (pbmc) with a considerable increase of prostaglandin e 2 synthesizing m~ and a simultaneous decrease of functionally competent cd3+ and cd4+ lymphocytes. t-cell dysfunction in states of profound stress is characterized by impaired synthesis of two crucial lymphokines -interleukin-2 (il-2) and gamma-interferon (y-ifn). the inability to produce adequate amounts of il-2, most likely attributable to alterations in the transmission of signals from the cell membrane to the nucleus, results in incomplete proliferative t-cell responses to antigenic stimuli, while a lack of ?-ifn results in inadequate m~ antigen processing and presentation. the role of the two functionally distinct t-helper subsets, t-helper-1 (thl) secreting principally il-2 and ?-ifn and t-helper-2 (th2) acting principally in inducing antibody formation with a secretion of il-4, il-5, il-10 and tnf-[3 has still to be established within the context of major trauma. antibody formation to common protein antigens is impaired, the predominant finding is a shift from igm to igg synthesis. although excessive secretion of prninflammatory cytokines (il-u3, tnf-c~, il-6, il-8) has been considered to be deleterious for the host in states of major inflammation and infection, in-vitro and whole blood investigation of me function has clearly revealed that there is initially a marked suppression of moo from septic patients to synthesize and secrete proinflammatory cytokines to endotoxin. this probably will result in immunodeficiency of traumatized as well as septic patients, since these cytokines in low concentrations are involved in the upregulation of the essential humoral and cellular immune functions. abnormalities of pmn function noted after serious injury have been interpreted by some investigators as signs of unresponsiveness of this cell population, others have demonstrated signs of pmn byperactivation. latest findings revealed that there is a clear pmn dysfunction in circulating blood: 70% of all pmn's in this compartment show downregulated or non existent ~-integrins within 72 hours posttrauma -these cells do not phagocytose, have a reduced respiratory burst and appear to be completely degranulated. restoration of these cell functions within 7 days post-trauma is significantly correlated to survival of traumatized patients. other reports however stress increased expression of cr3+ receptors (cdll/cd18 complex) on circulating pmn's, increasing the ability of these cells to adhere to intercellular adhesion molecules, thus contributing to play a major role in inducing the pulmonary capillary leakage. our knowledge about the perturbation of host defenses associated with serious injury and sepsis is continuously increasing and represents the precondition for the design of effective therapeutic measures to prevent and counteract the resistance to invading microorganisms. dept. of surgery, klinikum grosshadern, ludwig-maximilians-university, munich, germany. * dept. of surgery, university of cincinnati, cincinnati, ohio, usa lg thijs~ ce n~ck sepsis is the most common cause of acute disseminated intravascular coagulation (dic) and coagulation disorders as well as abnormalities of fibrinolysis are common in septic patients. some clinical and experimental studies indicate that dic is a pathogenetic factor for the development of multiple organ failure. endotoxin and various mediators (tnf, il-i) enhance tissue factor and decrease expression of thrombomodulin on endothelial cells in vitro. also, tpa activity is suppressed and release of pai-i is stimulated. in such a way the endothelial surface becomes procoagulant whereas fibrinolysis is inhibited. following administration of endotoxin to normal volunteers levels of prothrombin fragments (fi+2) and thrombin-antithrombin (tat) complexes increase, indicating thrombin generation. also the fibrinolytic system is activated as evidenced by a rise of levels of tpa and plasmin-antiplasmin (pap) complexes but this is counteracted by a subsequent release of pai-i. in severely ill septic patients levels of tat complexes are increased and concentrations of the natural inhibitors of coagulation (at iii, protein c) are usually decreased. also, tpa levels snd pap complexes as well as concentrations of pai-i are elevated. the severity of many of these alterations is related to mortality. thus, both coagulation and fibrinolysis are activated in sepsis, but not in a balanced way, thereby promoting coagulation. one of the hallmsxk pathological alterations associated with sepsis is the development of microvascular thrombosis, an entity ~lmre~erizod by microvas~tlar plugbing with leukoeyies (predominately nentrophils) and fibrin deposits. preranably, ischemia remlt~g fibm ve~-'ttlac ow,.luwion contributes sisni.ficantly to end orgen darnaje and consequent dysfatlctlon. p, er~t srldies haw focused pmdominately on the role of adhesion molecules in the pathogenesis of neu~ophil sequestration during infection and s~sis. this preparation will review the mechanisms underlying intravak-'v.ler i~brin deposition and its contribution to organ injury. the normal endothelial cell sure is generally conddered 1o be anticos~am. this state is maintained by two major anticoag~lam molecules on the cell so,ace: hepax'an su,lfale and thmmbomodulin. studies performed over the past decade have do~mentod a general shii~ towards a procoa~qtlant staw during gram negative hffeodon, aft e~| mediated mainly by changes in the endothalial cell surface. antt-tf antibodies have been shown to be pro~ective during l~'ml endotexemia. in summary, ~erova~'~ler fibrin deposition, in ~rt mediated by ~rcgulation of cellular tf, is a consistemt patholo~cal flndin~ during sepsis and contributes to organ injury. strategies designed to abrogate this event may n~nimjze the magnitude of erg~ dysftm~on. n, v, christou md phd, department of surgery, megill university, montreal, quebec, canada interactions between immune calls and the endothelium vie adhesion molecules serve to modulate immune cell traffic between the blood strum and the extrmvascular space, these families of molecules acting in a cascade and closely integrated with the oytoklna network, blood coagulation and the complement system, are responsible for the homing of leukocytes to areas of inflammation and the realrculatlon of tymphocytes. there are three types of immune ceils that must exit the blood stream into the extravascular space: lymphocytes ; polymorphonuclear leukocytes; and non-lymphocyta mononuclaar cells (monooytes, basophlls, aoslncphils). b lymphocytes exit the blood stream and recircu}ate mostly via the high endothelial venule (hev) in lymphatic tissue, rarely, in chronic }nfectlon=, they may recirqulate vie hev in non-lymphoi:t tissue. t-lymphocyte recircutation on the other hand can occur via hev in lymphoid tissue, as well as, the endothalium of the skin, returning to peripheral lymph nodes via the lymphatic channels. lymphocyte recirculation is the mechanism which allows ¢ontaot between the immune repertoire and pathogens, and homing delivers these cells to the appropriate environment for activation and for effeotor function, pmns exit the blood stream through endotbelium in close proximity to the site of inflammation after appropriate endothelial membrane receptor expression. they must reach the site of pathogen tnveslon quickly in order to be effective. their exudation to the inflammatory site can be modulated by their intravascular pdmlng, which results from appropriate receptor expression. because endotoxin leak from the gut has been postulated as an important trigger mechanism for the systemic inflammatory response syndrome seen after serious injury, the immunologic and metabolic effects of bacterial endotoxin infusion into normal volunteers can be expected to shed considerable light on the validity of this hypothesis. we and ethers have used tbis model to show that endotoxin causes a temporary paralysis in the ability of circulating monocytes to prudce the proinflammatory cytokines interleukin-i (il-i) and tnf-a. endotoxin, however, causes increased production of pge2 by the same cell population. endotoxin infusion causes significant suppression of circulating t-cell numbers and a profound suppression in the ability of the circulating t-cell population to proliferate and to produce interle~in-2 (il-2) upon in vitro stimulation. after endotoxin circulating t-cells are also more sensitive to the inhibitory action of exogenous pge2. administration of cyclooxygenase inhibitors at the time of endotoxin infusion largely abrogates the effect of endetexin on t-cell proliferation and il-2 production. endotoxin infusion also causes the release of increased levels of circulating catabolic hormones including cortisol. administration of cortisol alone or cortisol along with endetowin to volunteers has allowed dissection of cortisol effects from those of endotoxin itself. cortisol infusion alone causes a diminution in the circulating t-cell population but the remaining ceils continue to produce il-2 after appropriate stimulation. cortisol infused along with endotoxin blocks the overshoot in monocyte proinflammatory cytokine production seen 24 -48 hours after endetoxin infusion. at present, one may conclude that administration of sublethal doses of bacterial endetoxin to human volusteers produces alterations in cytokime production and tlymphocyte activation which are both similar and dissimilar to those seen following serious injury. hemodynamic support is based first on intravenous fluids and second on vasoactive agents. colloids are usually preferred to crystalloids, for their more rapid hemodynamic effects and their lesser passage in the interstitial space. in the presence of hypotension, the pharmacological profile of dopamine makes it the first agent of choice. it is only when high doses of dopamine are insufficient to restore a sufficient tissue perfusion pressure that norepinephrine should be added. even when cardiac output is not decreased, a dobutamine infusion is often indicated to counteract the myocardial depression, prevent vasoconstriction, and increase oxygen delivery to the tissues. the benefit derived from the administration of "renal" doses of dopamine is doubtful, but stimulation of dopaminergic receptors may increase blood flow also to the gut. in any case, none of these catecholamines has been shown to significantly improve the oxygen extraction capabilities of the tissues. agents with antiinflammatory properties but also with some vasodilating properties, such as n-acetylcysteine, prostaglandin el or pentoxifylline represent more attractive options to increase oxygen extraction. cardiovascular support should aim not only at the restoration of a sufficient arterial pressure but also at the maintenance of an optimal oxygen delivery to the cells. it should thus be guided also by measurements of cardiac output, mixed venous oxygen saturation and oxygen extraction, and indexes of cellular hypoxia, such as blood lactate levels, gastric intramucosal ph or vanearterial pc02 gradients. supranorma[ 02 delivery contributes to the prevention of tissue hypoxia tissue hypoxia is considered to be the common final pathway in the pathogenesis of multipte systems organ failure. it may directly contribute to cellular injury and organ dysfunction as well as enhance further mediator activation and release by a positive feed back mechanism. prevention of tissue hypoxia, is therefore, one of the most important aims in the supportive therapy of critically ill patients, especially in those with sepsis and septic shock. mismatch of cellular o= supply and demand in these patients may resuit from the impairment of the cardiocircu[atory system on all levels. 1. myocardial depression with a drop in 02 delivery (do~) 2. redistribution of blood flow between the different organ systems 3. inadequate nutritive blood flow due to tissue edema and endothelial damage (gic, leucocyte swelling etc.) these pathological changes may go along with increased metabolic needs. cellular and organ 02 supply may therefore be inadequately matched with cellular 02 needs in patients with normal and even supral normal doz. hyperdynamic circulation is often present in adequately volume resusucitated patients. it is most likely that one of the bodys main compensatory mechanisms would maintain cellular o= supply in the face of increased metabolic needs and impaired oz extraction capabillities. this pathophysiology may explain the findings in several clinical investigations that patients with normal or even supranormal doz can have signs of inadequate regional tissue oxygenation. it is also consistent with the results of different studies which demonstrated that the achievement of supranormal doz levels, either spontanuously or due to adequate supportive therapy, resuits in better patient outcome. any attempt to prevent or treat tissue hypoxia in these patients, however, has to take into account the effects of therapy not only on global dgz but especially on regional and microcirculatory blood flow. r. rossaint, d. pappert, k. lewandowski, h. gedach, k. slama, k.j. falke klinik for anaesthesiologie und operative intensivmedizin, ukrv, freie universit §t berlin, germany important contributory factors to the high mortality of severe acute respiratory distress syndrome lards) are the aggressive therapies required, such as mechanical ventilation with high inspiratory oxygen concentrations and airway pressures. as specific therapies for reducing or preventing the general inflammatory reaction of the lung resulting in severe hypoxemia is unknown, today's therapy is limited to procedures which predominantly support or maintain pulmonary function, i.e. pressure limited ventilation with peep and permissive hypercapnea, avoiding or treatment of fluid overloading, and positional maneuvers. extracorporeal lung assist combined with low frequency positive pressure ventilation has been recommended, when conventional therapy fails. today, extracorporeal gas exchange may be carried out using a system internally coated with covalently bound heparin. this allows for a lower level of systemic heparinzation reducing the risk of severe hemorrhagic complications of extracorporeal respiratory support. from april 1989 to august 1993 89 patients, aged from 15 months to 60 years, were transferred to our intensive care unit (icu) for treatment of severe ards. all fulfilled at least the slow entry criteria of the u.s. national ecmo study. due to hypoxemia 13 patients required veno-venous extracorporeal membrane oxygenation (ecmo) immediately after transfer to our icu. the other 76 patients were first treated with pressure controlled mechanical ventilation, permissive hypercapnea, prone position, and dehydration, if necessary. in 23 out of these 76 patients, in which the gas exchange did not improve in the following days, veno-venous ecmo with heparin-coated systems was additionally performed. heparin was given to achieve a partial thrombin time between 40 -55 s and an activated clotting time between 120 -150 s. if surgery was performed during ecmo, heparin infusion was interrupted. no severe bleeding complications related to anticoagulation for the extracorporeal bypass could be observed, however, in some patients, after three weeks of ecmo thrombi in the drainage cannula were observed. a problem of of membrane leakage shortened the life span of the membrane lungs to a mean of about four 4 days. in the conventionally treated group 87% survived and in the additionally with ecmo treated group 58% survived, representing a 75% survival rate in patients whose risk of death is considered more than 50%. on the basis of these experiences, we conclude that the described strategy, including veno-venous ecmo with heparin-coeted systems, may improve the survival in patients suffering from severe ards. abnormally high skeletal muscle po 2 in septic patients and its normalization during recovery: evidence against tissue hypoxia but for impaired oxygen metabolism of skeletal muscle? p. boekstegers, k. werdan department of medicine i, gro6hadern university hospital, munich, germany a pathological oxygen uptake supply dependence was suggested to indicate tissue hypoxia in sepsis-a hypothesis which is discussed controversially. because the detection of reduced oxygen transport to tissue and &tissue hypoxia would have important implications for therapy, skeletal muscle oxygenation was studied in patients with sepsis. intermittent and continuous determination of skeletal muscle po 2 by polarographic needle or catheter probes provided no evidence for skeletal muscle hypoxia even in severe stage of sepsis (boekstegers et al., crit care med 1994) . in contrast, mean skeletal muscle po 2 was found to be abnormally high in patients with sepsis (48 _+ 10,4 mmhg, n=39) compared to patients with limited infection (28, 4 + 4, 7 mmhg, p<0.001, n=16) , to patients with cardiogenic shock (22,3 + 6,2 mmhg, p<0.001, n=ls) and to healthy subjects (33,3 _+ 8,1 mmhg, p<0.001, n=10). furthermore, serial measurements in patients with sepsis showed that skeletal muscle po 2 was higher (44,1 _+ 10,3 mmhg) on days with severe sepsis than on days with intermediate state (30,1 _+ 8 mmhg) and than on days without sepsis (26, 8 _+ 5, i mmhg) . thus, a decrease of abnormally high skeletal muscle po 2 was related to improvement of sepsis. this was also demonstrated by comparing the decrease of abnormally high skeletal muscle po 2 with the decrease of apache ii score or elebute score as well as interleukin-6 serum levels in a separate study (boekstegers et el., shock, in press). in summary, our data provide no evidence for skeletal muscle hypoxia in patients with sepsis but support the assumption &impaired oxygen metabolism in severe sepsis. neutropnlic emigration from the vascular space into the extracellular matrix is important for the response to tissue injury or contamination by providing a large recruitable pool of tissue-based phagocytes. the consequences of neutrophil exposure to intravaseular chemoattractants, likely relevant for il-8 and c5a in sepsis, trauma, and in burn injury, are suppression of neutrophil attachment to endothelial cells and matrix proteins. it is probable that proteoglycan-bound attractants are of considerable biologic relevance, and may result in enhancement of responses to subsequently encountered cytokines. in the presence of connective tissue proteins expressing integrin-specific binding ligands, neutrophils respond to tnf-cq gm-csf and other cytokines by producing large amounts of hydrogen peroxide. this response is likely important in the digestion of contaminated or injured tissue by facilitating activity of neutrnphil granular proteases and inactivating plasma anti-proteases. this matrix-dependent oxidative response is important in defining potential novel targets for therapeutic intervention. the response appears to involve signalling by integrin-linked tyrosine kinases. the net effect of matrix protein injury through this mechanism is enhancement of the fibrotic response which underlies much of the prolonged ventilator dependence of patients with the adult respiratory distress syndrome. this adherence dependent response has also been implicated in direct hepatocyte injury, and may represent an early component of the syndrome of multisystem organ failure. study purpose: in this patient (pat.) population with a considerable sepsis morbidity and mortality, parameters proposed for sepsis assessment (elebute sepsis score [ele], sirs) were investigated with regard to their use in the early classification of pop. pat. at risk for developing sepsis. patients and methods: in a previous observational study on 110 consecutive pat. after elective open-heart surgery, we had determined ele daily for at least 3 pop. days (in pat. with a longer stay: until icu discharge). after publication of the newly proposed sirs definition, these criteria were retrospectively calculated and compared to ele. results: on the total of n =515 patient-days, both ele and sirs showed significant (p<0.0001) correlations with parameters reflecting the general and sepsis-related severity of the pop. clinical course. compared to sirs, the ele correlations were better (icu mortality: 0.59 vs. 0.32; icu treatment days: 0,51 vs. 0.32; days on mechanical ventilation: 0,54 vs. 0.32; days with vasopressor requirement: 0.59 vs. 0.33; number of microbiological findings: 0.44 vs. 0.29), the optimal classification criteria for the mainly sepsis-related outcome gave maximal youden indices of 0.87 for ele (ele >_12 on >_2 days, accuracy: 94%) compared to 0.42 for sirs (sirs present on >3 days, accuracy: 67%). accordingly, ele roc curve areas for both overall (0.92) as well as sepsis-related prognostic evaluation (0.99) were significantly (p<0,05) larger compared to sirs (0.79 and 0.86, resp.), this higher overall accuracy of the ele criterion was primary due to a more valid assessment already on the first and second pop. day, where sirs still had a high false positive classification rate (46% and 53%, compared to 7% and 1%, resp.). conclusion: in the early postoperative course after cardiac surgery, the sirs definition displayed a high false-positive classification rate (low specificity) for subsequent sepsis-related mortality compared to better classification results obtained by the elebute sepsis score. from the departments of medicine i and of "cardiac surgery, grosshadern university hospital, marchioninistr. 15, d-81377 munich, frg. correlation between physiological and immunological parameters in critically ill septic patients. ma rogy, h oldenburg, r trousdale, s coyle, l moldawer, sf lowry a relationship between physiological parameters of severe sepsis and immunological function has not been established. in an effort to assess such a relationship we prospectively evaluated nine severely ill septic patients. physiological risk was assessed by the apache iii score 1, while one component of immunologic function was evaluated by peripheral blood mononuclear cells (pbmc) eytokine production after in vitro lps stimulation 2. four of the nine patients died. apache iii scores at 72 h were lower in survivors (s) than in non-survivors (ns), (51-+13 vs 111-+1 5 p<0.05), while apache iii scores at admission were not significant different between s and ns (82-+13 vs 95-+13). down regulation of cytokine production by pbmc upon lps stimulation was a transient event in s. while s demonstrated an 3 fold increase of tnf-a bioactivity with[r~ 72 hours, ns did not demonstrate any increase at all. a similar pattern was demonstrated for il-1[3 and il-6 immunoactivity. tnf was measured by wehi bioactivity, il-1[~ and il-6 immunoactivity were determined by elisa. the sensitivity was 35 pg/ml for tnf, 30 pg/ml for il-ll3 and 35 pg/ml for il-6, respectively. in conclusion, both physiological as well as immunological functions of severe critically ill septic patients demonstrate predictive value for ultimate survival. while patients biological status seems to be more predictable by apache iii at day 3, p<0.05, the pattern of cytokine production by pbmc upon lps stimulation over the first 72h might be a reliable predictor as well. introduction: therapy of sepsis and its sequelae depends largely on its early recognition. many studies have investigated the change of certain mediators during sepsis and their potential to predict multiple organ failure and outcome. it was the objective of this study to investigate whether the onset of sepsis can be predicted by alterations of levels of interleukin-6 (il-6), tumour-necrosis-factor (tnf), pmn-elastase and c-reactive protein (crp). materials and methods: over a one year period, 40 polytraumatized patients were prospectively studied (mean age 42y, 71% male, iss 28). serum and edta-plasma samples were taken in 12h intervalls until the patient left the icu. il-6, tnf, elastase, and crp were determined immunologically. sepsis was defined according to the criteria of 'systemic sepsis' (veterans" administration study, 1987) with at least 4 of 7 clinical signs: (1) tachycar-dia> 100/min, (2) temperature >38,9°c, (3) blood pressure < 100mmhg, (4) mechanical ventilation, (5) leukocytosis > 15.000 /ml, (6) thrombocytopenia < 100.000/ml and (7) presence of an obvious septic focus. clinical parameters, sepsis severity and serum levels were documented on a daily basis, beginning on day 2 after trauma. results: 21 of 40 patients developed a systemic sepsis (52.5%), and 5 died. all mediator levels were elevated under septic conditions. the clinical severity of sepsis correlated well with the respective levels of mediators. in patients, who developed a sepsis the following day, il-6 (510 vs. 195 ng/l; p=0.029), crp (160 vs. 121 mg/l; p=0.035) and tnf (32 vs. 15 ng/l; p=0.045) were significantly increased as compared to those patients who remained non-septic. elastase levels were considerably elevated but did not reach the level of significance. we conclude that il-6, tnf and crp appear to be sensitive markers for prediction of septic complications in polytraumatized patients. objectives of the study: the assessment of liver function in polytraumatized patients who are at risk of developing mof is too inaccurate and late by using conventional biochemical parameters. methats: the injury severity of the patients (n=26) was determined by the injury severity score (iss). lidocaine is given at a dose of 1 mg/kgbw over 2 rain. i.v. and is metabolized in the liver by a cytochrome p-450 mechanism to monoethylglycinexylidide (megx). the metabolite is measured by a fluorescence polarization immunoassay. serial determinations of the test were performed between the 1 ~t and the 11 ~ day after trauma and were compared with other liver function tests (bilimbin, gldh, alt, ast). the systemic inflammatory response syndrome (sirs) is still a challenge concerning early diagnosis, therapy and prognosis. therefore, evaluation of inflammatory and disease activity becomes more important. c-reactive protein (crp) is a well established acute phase protein in chronic inflammatory diseases. recent reports suggest an induction of crp by interteukin-6 (il-6), a cytokine involved in the mediator cascade of sirs. on the other hand, tumornecmsisfactor alpha (tnfcx) is a very early released mediator in sirs removed very rapidly from circulation. in addition, soluble tnf receptors (stnfr~5, stnfr75) are released into circulation in the acute phase response. this study examines the kinetics of five acute phase proteins (crp, il-6, tnfot, stnfr55 , stnfr75 ) in patients suffering from sirs. eighteen patients entered the study after diagnosis of sirs. blood samples were drawn every six hours during the first two days and every twelve hours thereafter. crp was measured in an routine turbimetric assay. il-6 was detected in an biological assay using the/l-6 dependent 13-cell line 1313/29. detection of tnfc~ was performed in an elisa system using a monoclonal antibody" for tnfo~. soluble tnf receptors were also measured by elisa. crp levels were elevated (>10 mg/l) in all patients and at all time points. crp values did neither differ significantly in patients with (143±33 mg/l) or without (129a:39) multiple organ failure (mof) nor in survivors (133±41) or non-survivors (147:t:44). in contrast, 1l-6 was elevated in patients wilh mof (mean 740 pg/ml, range 0-8900 pg/ml). il-6 levels correlated especially with lung dysfunction. tnf(x levels were consistently elevated in patients with mof. crp, il-6 and tnfoc did not correlate with each other. in contrast, levels for both stnfr showed a positive correlation (r=0.7). patients could be divided into two groups by values for stnfr~5 and stnfr75: the group with higher soluble tnf receptor levels showed increasing values combined with a poor prognosis. the group with lower levels of soluble tnf receptor consisted of patients surviving mof or without mof. in conclusion, crp does not monitor the course of sirs adequately. in contrast, il-6 correlates with mof and episodes of high disease activity. high stnfr levels may indicate poor prognosis. klinik f0r an/isthesiologie and operative intensivmedizin der cau kiel, schwanenweg 21, 24105 kiei, germany. ch. waydhas, md; d. nast-kolb, ivid; m. jochum, phi); l. schweiberer, mi) objective: to evaluate the irfflarranatory response after different types of orthopedic operations and compare them with the systemic effects of accidental trauma of varying severity. patients: in 135 consecutive patients with multiple injuries (iss 40.6) the inflammatory response to trauma was prospectively studied. the patients were divided into 4 groups according to their iss points. additionally, the alterations after 79 secondary operations (>24 hr) were determined (msteosynthesis of the femur (n=28), pelvic girdle (n=ll) and spine (n=8), facial reconstruction (n=13), smaller osteosynthesis (n=13) and others (n=6)). methods: specific and unspecific parameters of the inflammatory response were determined in the trauma patients every 6 h, beginning on admission of the patient to the emergency room for a period of 48 hr, and in the operative patients on the morning of the operation, at the end of the procedure and every 6 hr during the first two days. results: lactate, neutrophil elastase, heart rate, po2/fio2-ratio, and other parameters discriminated significantly between the 4 injury severity groups during the first 48 hr (kruskal-wallis-test, p<.05). the degree of postoperative changes differed significantly (kmskal-wallis-test, p<.05) between the 5 types of operations for lactate, heart rate, po2/fio2-ratio, nitrogen excretion and showed a strong discriminating tendency for neutrophil elastase and c-reactive protein. the extent of changes were highest after operations of the pelvic girdle, followed by procedures on the femur, spine, smaller bones, and the facial region. the postoperative changes after osteosynthesis of the femur or pelvis were comparable to the alterations noticed after smaller (iss 1 to 24) or moderate (iss 25 to 40) accidental trauma for neutrophil elastuse, heart rate, po2/fio2-ratio and parameters of the coagulation system. conclusions: there is a considerable inflammatory response to operative procedures that varies with the type of surgery. large operations cause changes in the body homeostasis that resemble those after multiple injuries. it remains to be established whether the inflammatory sequelae of surgical trauma are additive to the changes caused by accidental trauma. objective of the study: we retrospectively compared characteristics of elderly patients (~65 years) and yeunger patients admitted to a surgical {sicu) and a medical intensive care unit (micu). we further studied the relations between advancing age, chronic disease, sepsis, organ system failure (osf) and mortality in the elderly group. material and methods: during a 1-year period, 538 patients were consecutively admitted into the 8icu; and during a 2-year period, 487 patients were consecutively admitted to t~mich. criteria for chronic disease, sepsis, osfsi.e. cardiovascular (cf), pulmonary (pf), renal (rf), neurological (nf), haematological (hf), hepatic (lf), and gastrointestinal failure (gf)-were derived from the literature. results: patients from the sicu and~cu were similar in age, number of osf, and length of stay. however, when compared to sicu patients, micu patients had more cf (p<o.o1), sepsis (p<o.o01), and mortality rate (27% vs. ii~, p<o.ool). in contrast, sicu patients had more hp and nf, and prior chronic disease (all, p<o.o01). of the total group of 1,025 patients, there were 406 (40%) patients 65 years of age or older. elderly and younger patients had comparable length of stay in the icj, but elderly patients had significantly more chronic disease, sepsis, and number of osf (all, p<o.o02). all osfs, except hf and nf, were more ~o~mon i~ mdeljl, eompard to you-ger patients. ~he mortality is also higher in elderly patients (29% vs. 11% in younger patients). after multiple logistic regression, only number of osf increased mortality by a factor of 2.48 (95% confidence limits, 1.97-3.12), age did not increased [odds ratio 1.05 (i.01i.i0)], while admission to the sicu reduced risk of death by a factor of 0.15 (0.08-0.30). conclusions: based on these results, we conclude that age does not have any impact on icu outcome in the elderly. the only prognostic factor is the extent of acute disease, as measured by the number of osf. hence, prevention of oaf may influence outcome in elderly patients with critical illness. the lower risk of death in sicupatients may be explained by the large proportion of postoperative elective patients with relative mild chronic disease, and the lower incidence of sepsis. department of surgery, free university hospital, de boelelaan 1117, 1081 ~v amsterdam, the netherlands. septic shock and low output syndrome h.miyakawa, t.noguchi, s.hattori, a. mizutani, m. mori and n.honda objectives: cytckines are thought to cause the acute phase reactions under several critical conditions. we had investigated the relationships between cytokines network which included il-6,1l-8,acth and cortisol,and the outcome of the patients with septic shock or low output syndrome (los). materials and methods: in the septic group and los group,nine and four patients were enrolled respectively. furthermore, the patients in the septic group were divided into two groups,survival (n=5) and non-survival group (n=4).tn these patients,ll-6,1l-8,acth and cortisol had been measured every day after diagnosis of septic shock or los.the total number of measurements were 12 points in survival septic group, l l points in non-survival septic group and 10 points in los group.statistical analysis was used student's t-test to compare the mean of each group,and the changes were reported as clinicat significant if p<0.05. results: il-8 levels in non-survival septic group were higher than in both survival septic and los group.ll-6 levels in non-survival septic group were more significant than in both survival and los group.otherwise,there were no differences with respect to acth and cortisol among three groups. conclusions: we assumed that los would make several organs dysfunction as well as septic shock. but our results showed septic shock, especially non-survival,had different cytokins network mechanism for organs failure from los. lasson ,~, g/3ransson j, jijnsson k. eady diagnosis of complications after trauma is imperative to decrease morbidity and mortality. for this purpose an easy, cheap and fast diagnostic method is needed. the aim of this study was to analyse if complications after surgical trauma of various extent could be diagnosed earlier using preoperative or daily postoperative measurements of various plasma proteins than by using climcal signs of complications. material and methods: two acute phase proteins (c-reactive protein and antichymotrypsin), lbur main plasma protease inhibitors (alpha-2-maeroglobulin, c 1estemse inhibitor, antithrombin ill and alpha-2-antiplasmin) and one collagen precursor (procollagen iii peptide) were measured preoperatively and then once dally for 7 days postoperatively. for the plasma protease inhibitors immunorcactive as well as functional levels were measured. haemoglbin, albumin and the white blood cell count were also measured. measurements were done preoperatively in 256 patients and continued postoperatively in 78 patients, 70 % of the patients were operated on for malignancy. resulls: preoperative plasma c-reactive protein levels were higher in patients developing postoperative complications. this discrimination increased when plasma was sequentially analysed postoperatively, when a remaining high level ora second increase in plasma levels depicted a complication 1-3 days earlier than clinical signs. high levels were seen both in patients developing local as well as in patients developing general complications. there was no clear correlation between plasma protease inhibitory levels and complications, neither pre-nor postoperatively. procollagen iii paptide levels were slightly (but not significantly) higher postoperatively in patients developing complications. contusions: the acute phase response, especially concerning c-reactive protein, predicts postoperative complications better than both plasma protease inhibitors or collagen precursors. preoperative plasma c-reactive protein levels indicate the risk of postoperative complications, while daily postoperative measurements more readily depicts a complication, usually preceeding the clinical signs of complication by 1-3 days. dept. of surgery m'alm6 general hospital, s-2 i4 01 malmr, sweden. mof is a major threat to the extensive burned patients and associated with a high mortality rate. the role of endotoxemia in the pathogenesis of mof in burned patients remains controversial. in this study, we examined the relationship of plasma endotoxin levels to development of mof, and outcome in patients with thermal injury. methods: a prospective cohort study of 17 patients admitted with burns covering more than 70 per cent of body surface area was undertaken. circulating endotoxin concentrations were measured by modified limulus amebocyte lysate assay in serial samples of plasma. results: 7 out of 17 burned patients developed mof according to multiple criteria. the plasma endotoxin concentrations of patients with mof were 0.512--1.127 eu/ml, significantly higher than that of 10 patients without mof (0.216-0.553 eu/ml) on 3, 14, 21 and 28 days postburn (p< 0.05--0.01). significantly more incidence of positive endotoxin test (>_ 0.120 eu/ml) was found in patients who developed mof as compared to that of non-mof during the observation period (p<0.05). as the mean endotoxin levels increased, the prevalence of mof and death also increased (see table below), persistent endotoxemia carried a poor prognosis. conclusions: the present investigation provide further evidence that endotoxemia in severely burned patients commonly occur. cimulating endotoxin has also been found to be strongly associated with development of mof and mortality following major burn injury. multiple hemostatic changes occur in sepsis mad multiple organ failure (mof). to evaluate the role of platelcts in patients with sepsis and mof, we examined changes in surface glyeoproteins on circulating platelets of t4 patients with suspected sepsis and mof. the severity of sepsis and mof was assessed by eiebute and apache 1i scoring system, respectively.using flow cytometric techniques and platelets specific monoclonal antibodies, platelet surface expression of fibrinogen receptor on gpiib-iiia, ofvon willebrand receptor gpib, and of granula glycoproteins (thrombospondin, gmp-140, and gp53) was measured. receptor density of gpiib-illa mad gpib on circulating platelets was not affected by sepsis or mof. in septic patients surface expression of activated fibrinogen receptor (libs1 expression) was significantly elevated (p<0.05) and correlated well with severity of disease (f0.597). no significant change in surface expression ofthrombospondin, gmp-140 or gp53 was noted in septic patients. in contrast, degranulation ofgraanle glycoproteins was significantly elevated in mof (!0<0.05) that correlated well with severity of mof (gmp-140, r=0.611; thrombospondin, r=0.643).we speculate, that platelets in sepsis circulate in a hyperaggregable (fibrinogen receptor activation ) but still reversible state that results in increased risk of microthrombotic events. in the course of the disease, irreversible platelet degranulation might occur and may play an important role in development of mof. abdominal sepsis is still associated with high morbidity and mortality. the present study aimed at evaluating patients with abdominal sepsis treated at our surgical intensive care unit during a 10-year period with the aim of identifying potential prognostic factors, bacteriological cultures, diagnostic procedures, treatment and outcome. during the period 1983-1992 i66 patients with abdominal sepsis were treated at the icu at our university hospital. 60 patients were women and 106 men with a mean age of 64 (17-101) years. in 74 cases, the abdominal sepsis occurred as a postoperative complication. the patients were scored according to apache ii and bacteriological cultures and the occurrence of organ failure were noted. the patients were hospitalized in median for 40 (-273) days out of which 7 (-178) in the intensive care unit. 46 out of 166 patients (28 %) died in median after 18 (1-194) days. the primary cause of mortality was multiple organ failure (38/46; 83 %). apache ii scoring could not predict a fatal outcome. abdominal bacterial cultures were dominated by bacteria of enteric origin (83 %) and in 60 % cultures grew multiple bacteria. 134 patients bad organ failure and 64 multiple organ failure. 29/166 patients (17 %) had abdominal sepsis due to diffuse peritonitis despite a morphologically intact gastrointestinal tract and the absence of localized abscess formation. mortality in this group was significantly higher as was the percentage of positive blood cultures and the occurrence of multiple organ failure. abdominal sepsis is still associated with a high mortality, predominantly caused by multiple organ failure. abdominal culture findings are dominated by bacteria of enteric origin. in about 1/5 of patients with severe abdominal sepsis a diffuse peritonitis with intact gastrointestinal tract without localized abscess formation was found. in this group the mortality was increased as well as the risk of developing multiple organ failure. during the period from january 1985 to september 1993 56 patients, mean age 24+4 years were referred to our department of resuscitologywith the diagnosis of eclampsia. all the patients were delivered by cesarian section and were mechanically ventilated for 7.3_+5.7 days. diagnosis of sepsis was confirmed in 6 cases by clinical and microbiological methods. patients were divided in two groups: lnon septic patients, 2-patients with sepsis, the control group consisted of 20 patients after cesarian section without symptoms of eclampsia or infection. we determined plasma concentrations of immunoglobulins a,g,m(a,g,m), complement factors (c3,c4), alphal-antitrypsin (aat), trausferrin (trf) and albumin (alb) using beckman (usa) analyzer,protein concentration, using kone (finland) analyzer. a(mg/dl) g(mg/dl) m(mg/dl) c3(mg/dl) c4(mg/dl) k 203+-31 971+47 205_+21 134+12 30+-3 1 154-+29" 568-+48* 142_+24" 81-+14' 17_+4" 2 102+_21'* 420-+30** 96-+21"* 67-+10"* 11_+3" in a prospective study we investigated serum of 12 severly traumatized patients withdrawn directly after admission at our hospital (tr i). follow up controls were taken daily until day ten after trauma (tr ii). two control groups were performed: serum of healthy volunteers (co, n =24) was investigated as. well as serum of patients undergoing elective herniotomy (n=12) 24 hours before (op i) and 24 hours after operation (op ii). serum bactericidal index (sbi) was determined using a hemolytic e.coli strain 08:k87:h19. 200/11 suspension with a final concentration of 250 -400 cfu were incubated with l oopl serum. after overnight incubation sbi was calculated according a special formula. results: co 74.8 _+ 6.3 opi 77.8 _+ 6.4 opii 68.2 _+ 7.7* tri 38.4 _+ 9.1"* trii 57.6 + 8.9** (*:p< 0.05; **:p<o.01) sbi represents a simple and reproducible method to detect immunobiological alterations after trauma and elective surgery. patients undergoing elective surgery showed slight but significant diminuation of sbi 24h after operation. deterioration of sbi was observed in serum of traumatized patients directly withdrawn after admission at our hospital. ten days after trauma significant improvement of sbi was found. further investigations have to be done to proof sensitivity and prospectivity of this simple but reliable test. a sepsis like syndrome (sls) occurs in i0 % of our patients following cardiac surgery. sls is characterized by a severe decrease in systemic vascular resistance requiring the use of vasopressors to maintain adequate hemodynamics in the presence of negative blood cultures. in order to determine whether preoperative factors predict the occurrence sls we retrospectively studied 62 patients with sls (group i: age 61.1 ~ 10.8 years); they were compared to 52 control patients (group 2: age 58 ± 14.3 years). treatment of sls consisted of aggressive fluid replacement and norepinephrine infusion. rydrocortisone (200 mg/24 hrs) was given on the first day. antibiotics were switched prophylactic to cover gramnegative bacteria in 48 pts. preoperatively pts in group 1 revealed a significantly lower cardiac index (2.5 ± 1 i/min/m 2) compared to group 2 (3.1 ± 1.2 i/min/m2; p < .02) higher left atrial pressure (group i: 17.8 ~ 8.1 mmhg; group 2: 13.1 ~ 6.1 mmhg; p < .005) and lower ejection fraction (group i: 57.9 z 16.3 %; group 2 65.2 ± 10.9 %; p < .03). bypass time, minimum and mean blood pressure on bypass and aortic clamp time were not different between the groups. immediately postoperatively (pop), the white blood count was elevated (group i: 15,700 ± 8,200/ui; group 2: 10,200 ~ 4,400/ui; p < .0005) and core temperature 12 hours pop was higher (group i: 38.3 7.6 °c; group 2: 37.6 ~ 8.4 °c; p < .004). serum lactate was already elevated on arrival on the icu pop (group i: 5,0 z 1.9 mmol/l, group 2: 2.2 ~ 0.9 mmol/l) but dropped within 24 hours (p < .0005). icu stay (group i: 3.7 ! 4.3; group 2: 1.6 ± 1.9 days; p < 0.001) and mortality over 3 months (group i: 19.1 %; group 2: 3.1 %; p < .005) were significant higher in group i. sls is observed more frequently in patients with preoperative cardiac congestion and results in longer icu stay and higher mortality. although intraoperative hemodynamics are similar in both groups, the white blood count and serum lactate levels suggest an intraoperative cause for sls. further investigations will focus on possible intraoperative causes of sls. the objectives of the study were to estimate the problem of stress ulcers in critically ill patients and to compare ranitidine and omeprazole to determine their efficacy in the prophylaxis of stress ulcers. 33 patients who were admitted to the intensive care units between january and july '92 with polytrauma, sepsis or those requiring ventilatory support for more than 48 hours were selected for the study. the patients were randomized into 3 groups to receive ranitidine, omeprazole or a placebo. the changes in gastric ph were monitored 4 hourly and the patients underwent an upper gi endoscopy 48 hours after the start of the study to look for stress ulcers and hemorrhage. the drugs were withdrawn one week after the start of the study. of the 33 patients selected for the study, 14 had undergone cardiac surgery, 6 had polytrauma and 13 had sepsis. the change in average pre and post drug gastric ph was +1.1 in the omeprazole group, +0.5 in the ranitidine group and -).9 in the placebo group (p < 0.05). endoscopic evidence of stress ulcers was seen in 62% of patients not on prophylaxis, 18.1% of those on omeprazole and 54.5% of those on ranitidine. while all the patients in the placebo group who had stress ulcers showed endoscopic evidence of hemorrhage, 82% of the patients with ulcers in the ranitidine group and none of the patients with ulcers in the omeprazole group showed hemorrhage (p < 0.01). in critically ill patients the incidence of stress ulceration is high. an alkaline gastric ph is protective against stress ulcers. omeprazole when used for prophylaxis against stress ulcers in critically ill patients or those on short term ventilation will reduce the incidence of formation and hemorrhage from stress ulcers. the early and accurate diagnosis of sepsis is of key importance for critically ill patients survival. many studies have shown that tumor necrosis factor ~x (tnf ~) and interleukin-6 (il-6) are mediators of the inflammatory response in sepsis. bacterial antigens interact also with antigen specific t cell receptors and the activation of t ceils takes place. activated t cells release soluble interteuldn-2 receptors (sil-2r). the aim of this study was to evaluate: the significance oftnf c~, il-6 and sll-2r in the diagnosis of bacterial sepsis and to evaluate the correlation of clinical and laboratory parameters with serum levels of tnf ~x, il-6 and sll-2r. we examined 19 patients with clinical signs of sepsis treated in the intensive care units of university medical centre ljubljana. venous blood was drawn from each patient within 24 hours after the first clinical signs of sepsis. clinical data (body temperature, blood pressure), laboratory dam (peripheral white blood cell count with differential, platelet count, c-reactive protein), and quantitative blood cultures were recorded. serum levels of tnf c~, 11,-6 and sil-2r were measured in 19 septic patients and in 20 adult healthy controls. tnf ct, il-6 and sil-2r serum levels were measured by commercially available ria and elisa (amersharn and eurogenetics) kits. the differences in serttm levels of tnf ~, il-6 and sil-2r between healthy control and patients and correlation between clinical and laboratory parameters were calculated. we found that only sll-2r serum levels were significantly (p<0.005, student t test) increased in patients in comparison with healthy controls. of all indicators of sepsis that we compared, only hypotension was significantly correlated with tnf ~x levels and leukocytosis with ]1,-6 levels. we conclude that only the increased serum sil-2r levels were predictive of bacterial sepsis within 24 hours after the first septic signs. tnf c~ and il-6 levels were not significantly increased at that time in lifts small group of patients. in order to create high precision prognostic system for patients with sepsis and septic shock we have made prospective and retrospective analis of 200 cases of sepsis. patients were included in this analis according bone's criterions of sepsis. comparing with apache-h, we have excluded such phisiologic variables as:temperature,na+ ,k+ ,}it, ph,which had a little influence on prognos of illness. we have added the other, more important variables: biilirubin, plateles,pti. we took into account: the seat arrangement, methods of respiratory support, comorbid conditions as: caneer, diabets,obersity. in general new score has 5 headings: -physiologic variables, -age, -seat arrangement, -chronic comorbid conditions, -methods of respiratory support. roc -analis have showed a greater precision of our score, compared with apache-ii score. there is significant difference between the roc-curve of apache-ii scare. the k.p.tit~v.,i.e.gurmanchuk. objectives of the study. sepsis is a severe complication of the local infection, th e fever and the systemic inflammatory response syndrome may take place in such a case as manifistations of the inflammation induced by bacteria. the systemic fever observed in infection is known to develop due to the action of certain cytoldnes. other systemic manffistation of inflammation are the production of acute phase reactants, acute phase proteins, the activation of the complement system. ivratertals and methods. we observed 35 children with light and 45 children with severe course(l-3 class of serionuess) of the sepsis. the ftmetional activity of the complement system (ch50, level of c 1-c5, factors b told d, c3a), level of c-reactive protein (crp} and tumor necrosis factor (tnf) were investigated, results. the level of crp was increased in the children with more severe class of the septic process in both group (60%-1 st and ioo%-2 nd}( with light and hard eours of disese) of patients. parameters of the complement system -the activity of cl, c4, c3 components, activity of b and d factors-were increased in the seram of children with light course of sepsis. in children with severe course of sepsis these parameters were signffieantty decreased, especially in the group of chiidrellwith higer level of crp. we found the negative correlation of the tnp-alfa concentration with the functional activity of classical pathway components (cl~3) and the positive onewith the funfional activity of the alternative pathway factors b-and d of the complement system. conclusions. thus, in children with different course of sepsis certain changes in levels and functional activity of an acute phase o[ inflammation proteins -crp and the complement system ones -&ire characteristic. the relationship between the tnf-alfa level and the activity of classical and alternative pathways proteins indicates on its role in the complement proteins genes expression. schr6der j, braun j, rennekampff ha, schr6der dw various alterations of the immune response are known in trauma, but the course will not be complicated by multiple organ dysfunction syndrome (mods) in all patients. phenotyping of cells offers a rapid system of evaluation certain aspects of the immune response. different subsets of lymphocytes and neutrophils were determined to evaluate their prognostic role in developement of mods. in 16 trauma patients with an injury severity score (iss) > 20 (mean iss = 32; mean age 41 years) lymphocyte and neutrophil phenotypes cd 3 (t-cells), cd 4 (t-helper cells), cd 8 (t-suppressor cells), ratio cd 4/cd 8, cd 11b (receptor for cr 3) and cd 16 (fcriii) were measured on day 1,2,3,5,7 and 10 post trauma. the expression of class ii histocompatibility antigen (hladr) on monocytes (hladr+ cd 14) and il 2-receptors on t-helper cells (cd 25/cd 41 were determined as well. the percentage of cells was monitored by immunofluorescence using monoclonal antibodies and three color cytometry. the percentage of hladr+ cd 14 were significantly lower an day 2,5,7 and 10 in patients who developed mods (p<0,05) compared to patients without mods and a healthy control (p<o,o01). the expression of il 2receptors on cd 4 was significantly different only on day 2. no differences could be measured in lymphocyte phenotypes cd 3, 4, 8 and cd 4/cd 8-ratio as well as in expression of cd 11b and cd 16 on neutrophils. class ii histocompatibility antigen (hladr) on monocytes offers the best ability to reflect cellular immunosuppression in trauma. this parameter allows the best differentiation of patients with and without mods. we retrospectively studied the relative importance of age, prior chronic disease, sepsis and organ system failure (osf) to determine outcome in critically ill patients with acute renal failure (arf). arf was defined as serum creatinine > 280/zmol/i, a twofold creatinine rise in prior renal insufficiency or the need of acute renal replacement therapy. definitions for prior chronic disease and other osfs -i.e. cardiovascular (cf), pulmonary (pf), neurological (nf), haematological (hf), hepatic (lf), and gastrointestinal failure (gf)-were derived from the literature and described previously. of the 1025 consecutively admitted patients to a surgical and a medical intensive care unit during 1 -ye0r period, 136 (13%) had arf. arf mortality was 52%. ninety-eight percent had other osf. overall, cf, pf, gf, and nf was significantly more common in nonsurvivors than in survivors (all, p<o,02). although both the number of osf before and after arf was higher in nonsurvivors than in survivors (p<0.02), the number of osf before was higher compared to after arf in both groups of patients (p<0.001). furthermore, age, cf and pf before the ontset of arf, nf thereafter, and renal replacement therapy were related to mortality (all, p<0.03). however, prior chronic disease was not related to mortality and chronic renal insufficiency was inversely related to outcome (p<0.002). after multiple logistic regression, advancing age, cardiopulrnonary failure and sepsis before arf, and nf after arf remained significant. these results show the high prevalence of arf in critically ill patients and that arf rarely occurs alone. the prognosis of arf in critically ill patients is poor, especially if associated with advancing age, additional osf, particularly cardiopulmonary failure and sepsis before arf, and nf after arf. hence, prevention of cardiopulmonary and sepsis before the outset of arf may influence outcome in arf in patients with critical illness. the search cominnes for a magic bullet ta help critically ill petienta, bat recent elini ',.ml 1rials of agents that showed ixomise in animal studies raise questions about such trials. expcrlmemai uvidan~ :in anlmats and human volanteet~ for concepts, mechanisms and treatment of inju~ ur illness can be substentlal, l~rauasive and exciting but the positive effects of potential therapy suggested by such animal and ctinieal research may be difficult to prove in sick patients. efficacy of a new txeatment can be difficult to prove became elinie.al situations, hi spi~e of important biologic phenomena, are v~iable and conrplex. there is redundancy and overlap of mediators ~d problems of timing of therapy. a majt~r cause of this dilemtns is not with the trials or hypotheses, but rather the promem of the cause or the causability of the human dise~.se that is trebled. this is/hu "causa nets" or '*specific cause" as described by stehbens, when prevention or tre.atmcnt is based em the sauna nero, extinetiem of the disease is ix~ssible as with saul/ pox. only elimination of the cause wl12 cure the disease. careful animal studit~s evaluate single pcrturbmiuns for survival or other changes. an ldso preparation may be infiuancad aignificantly by a ~mall clangs which may be insignific~lt ht pstiants, most human experiments ate carried out in normal volunleers with a single porlurbation, such as a low-level, non-lethal dose of a substance. such studies are huportant in defining mechanisms attd mediators of disease, in our world of injm'ed, operated, infected and inflslned patients, there are many diseases, mof is not one of lhem. it isn't even a syndrume, it is the final pathway for a number of diseaaes~ each of which has a cause. mecormock believes thal a "multi-causal" etiology is a euphemism for ignoranc# or a synonym fo~ unknown etiology. admission iv sn icu, a high apache seer% the sepsis syndrome, mof, muds, or sirs and having predictors indicating potential mortality, are not diseases. the inmpors are getting negative results in trials of agents on eiek or septic icu patients and aru now becoming splitters, dafining subgaoups at higher risk for death. such an approach may produce positive results if the diseases are identified and the treatmeul is specific for them, a major challenge in deterraintng the efficacy of new therapies for sepsis nnd shock is identifying patients whose e#temtc inflammatory response is appropriate from those in whom the mediator| are contributing to end organ system damage end death, traditional and recently revised categorical patient descriptions such as sepsis syndrome, sepsis syndrome with shock, multiple organ system failure, or the recently proposed systemic inflammatory response syndrome (sirs) may not be sufilcient since they identify individuals at varying levels of risk for short term mortality, the efficacy of new lmmunotherapy may be directly related to the patient's severity end risk of mortality at entry to the study. we recently reported (jama 1993;270:1z33-1~41) a comprehensive risk assessment approach for patients with sepsis syndrome, a common eategarlcal description used as an entry eriterlon for many trials. by screening s database of 58,737 recent admissions to 107 hospitals in the united states and western europe, we identified 1,195 patients who met sepsis syndrome criteria but were not enrolled tn clinical trinh, we then developed a survival model end severlty assessment method to estimate their 2g.day mortality risk. k..ente physiologic abnormalities were the most important prognostic factors (82% of total chl square) influencing outcome. the specific disease resulting in ice admission and the days the patient spent in the hospital and icu before qualification were independent risks, as were age and a clinical history of cirrhosis. the predictive model was cross-validated within the original 1,195 patients with a receiver d_perator characteristics (roc) area of 0.76 and in two independent databases, the first independent database contained 295 hospitalized patients with gram-negative infection meeting criteria for sepsis syndrome (roc=0.80); in the second, the 302 placebo patients withtn the recently completed phase ill e~aluatlan of il-1ra (rocffi0.76). in all databases the risk model was able to accurately risk stratify patients throughout the range of risks that varled from a very low 1% to a very hlglt 99%. by drawing on the ready avatlabillty of large clinically aeuurate, current databases and using the power of modern statistical techniques to model the bodfs response to sepsis, we are able to produce very aeettrate pre-treatment risk estimates, these prospectively defined and continuous risk estimates reduce reliance on multiple subgroups and data dredging that can compromise a stady's integrity. they cart provide a nnlform method for patient description across clinical trials of different attempts at immmlotherapy. risk calculations can also be useful in developing entry criteria for phase ii evaluations in order to initially test efficacy on a limited number of high risk patients, fonowing sueeessinl completion of a definitive trial, the risk estimates can be used to develop specific |ndlcattons for s drug's use and can monitor the impact of both new and multiple compounds on patient outrome, when dealing with therapeutic trials for the treatment of sepsis, there are 2 ways of checking the comparability of the groups (placebo and treated groups) -to use several description items such as age, sex, preexisting disease, number and nature of organ failures, physiology score, such as saps il (1) -to use a model for risk of death either from a score (apache ii, lii, (2) saps ii) or directly (mpm ii system (3)) before using the model for risk of death in septic patients, is it mandatory to validate it (by calibration, i.e. goodness of fit, and discrin'dnation, i.e., roc curves) first on a data base using the same definition for sepsis as in the trial and secondly on the placebo group of the trial ? or is it better to use a specific model for each trial ? the general models usually do uot fit for septic patients. there are 2 methods to make them fit : either to add variables to the original score (model for sepsis using the apache iii system (4)) or to customize the model. for saps ii the customization is applied to the equation regression, using the same saps ii score. for mpm 1i each component of the model can be customized. several remarks : only one model is published to be applied at the icu entry ( mpm ii 0) before any therapy. 2 -most of the published scores are calculated from the lsl icu day. applying these scores to the let day of sepsis could have a very different significance. 3 -is the accp classification of sepsis useful at the bedside to select oatients ? ( the pathophysiology of sepsis involves a dynamic interaction between the host and the infecting microorganism(s). a multitude of biochemical and hemodynamic interactions occur which function in concert to determine the ultimate outcome of the septic episode. the complexities of the septic process preclude outcome analysis by in vitro systems or computer models, animal studies have become indispensable in evaluating new therapeutic agents in the treatment of sepsis. these studies provide valuable information on safety, pharmacokinetics, relative efficacy and dosing strategies for potential therapeutic agents in the treatment of sepsis. there are three basic types of animal models: (1) toxicity models with injection of bacterial endotoxin or exotoxins; (2) live or killed bacterial challenge models; (3) actual infection models. all models employ some form of external manipulation under controlled conditions to induce sepsis and to analyze potential therapeutic efficacy. these experimental requirements may limit the ability of animal models to accurately predict the clinical value of new agents in human sepsis. four major end points are analyzed in the various animal models: (1) hemodynamic parameters; (2) modulation of host-derived inflammatory mediators; (3) resolution of end organ injury; or (4) lethality. each animal model has certain advantages and limitations. species-specific differences in physiologic and immunologic features make it difficult to directly extrapolate the results of animal experiments into clinical medicine. while no ideal animal model exists, consistent benefit of a new immunotherapeutic agent in multiple animal systems provides the most compelling preclinical evidence of its therapeutic potential in septic patients. consensus-assisted development of a study protocol on sepsis: an important difference from previous randomized trials there have been several, and perhaps too many metaanalyses of cliuical trials on surgical infections, but their results have only rarely been incorporated into the design of new randomized trials in sepsis. metaanalysis is retrospective. it seems more important to analyse and to criticize the design of tria/s before they are ongoing. incorporation of that into development of a study protocol is the aim of the following procedure: the time-schedule of about two years should include further cell culture and animal experiments after the first successful studies showing effectiveness. exhaustive evidence for a dose-dependent cost-banefit and near complete explanations of the pathomechanism should not be awaited for development of the protocol of the clinical study. however, a discussion forum of experts should be performed in a recently published, almost perfect trial in order to see the frontiers of the present research in cell and molecular biology, clinical pharmacology, statistics and decision making. this should be followed by a metaanalysis of at least 10 study designs on sepsis with methods of formalized medical decision making (decision tree). clinical algorithms -developed by objective methods including nominal group processes -should be developed to standardize any concomitant treatment in the centers considered for a multinentre trial. the latter is usually necessary in the field of sepsis. finally, a pilot study should be performed to demonstrate not only feasability, but to learn about all possible types of interventions which modify endpoints as response variables. especially mortality is not free from such effects. the time for the individual phases of this consensus-assisted process of protocol development have to be shortened by their temporal hiterloeldug. due to the fast discovery of new chemical factors in sirs it wilt otherwise be impossible to come up with results of randomized trials which are fascinating enough to be incorporated in daily routine treatment. severity scoring systems are intended to provide a population-based estimate for the risk of an unwanted outcome resulting from some disease process. in the area of infections and the "septic" response, this has routinely been defined as death. for anti-infective trials, there has been a reasonable correlation of severity (apache ll) with outcome measured as persistent or recurrent infection, but this is not as robust as the association with mortality. it is expected that non-lethal failure would not immediately correlate with disturbed physiology: the basis for antiinfective failure depends on a variety of factors not measured in severity scoring, such as type of infecting organisms, density and susceptibility to administered antimicrobia[s, and anatomic features of infection (e.g., pancreatic). severity scoring is important in differentiating these "categorical" variables from continuous (physiologic) variables. there are, additionally, a series of problems related to the cause of death in relation to severity scoring. in those tdals in which cause of death is analyzed, high severity scores did not routinely predict death as an immediate sequelae of sepsis/septic shock. many of the predicted deaths occurred remotely of progressive background diseases. other important problems with severity scoring have to do with lead-time bias: hew long patients were ill before entry into the study. statistical techniques for condensing various categorical and continuous variables, as well as factoring in information requiring the pharmacodynamics of agents which affect outcome, are being developed to further refine deviation from predictive equations as an outcome measure. stepwise logistic regression analysis has identified the presence of shock, intm-abdominal or unknown source of infection, hospital acquired infection, age greater than 70 years, neutropenia and inappropriate antimicmbial therapy as independent variables associated with increased mortality in sepsis. maldistribution or" these variables is a concern in targe randomized clinical studies. it should be noted that only the selection of antimicrobiai therapy could be addre.~'sed and altered at the time of enrollment in a sepsis clinics] trial. inappropriate antimicrobial therapy has been noted in 41%-66% of septic patients. well designed, randomized sepsis trials do not assure the absence of imbalance between treatment and control groups with regard to inappropriate antibiotic therapy, especially with subgroup analysis. statistical manipulation to correct for any imbalance is appropriate but avoiding imbalance is ideal. selection of antibiotics in septic patients requires such considerations as clinical setting (history, physical examination and underlying disease), appropriate laboratory tests (such as gram stain), identification of source and associated bacterial flora, antibiotic susceptibility patterns for that hospital and likelihood of resistant organisms. determination of inappropriate antibiotic therapy is made retrospectively after culture results are available. in some circunmtances the organism and antibiotic sensitivities cannot be predicted, while in others it can. leibovici et al identified hospital acquired infection, antibiotic treatment before a bacteremic episode, endotracheal intubetion and thermal trauma as independent variables which predicted isolation of a multiresistant organism. the same authors developed a predictive index for resistant organisms, which when compared to prescriptions of attending physicians, improved antibiotic selection in critically ill patients. including a standard antibiotic regimen for all septic patients in a clinical trial is impractical. there is too much interhopsital variability in flora and antibiotic susceptibility. in addition, the aggressive broad spectrum coverage necessary for some patients is inappropriate for others. a protocol which guides a prescribing physician through a logical chain of reasoning might improve antibiotic selection in critically ill patients and could be included in the design of a clinical trial in sepsis. although this approach might minimize potential for maldistribution of inappropriate antimierobial therapy, it would likely create multiple problem areas. will the patient's physicians agree to the protocol choice? what is the availability of speciftc antibiotics at a participating hospital? what is the aplpiieability of study results in typical clinical situations? if there is agreement that thin is an appropriate antibiotic protocol, should it not be used in all seriously ill septic patients in the hospital? in other conditions such as ards, the success of protocol therapy is dependent upon measurable goals such as pao 2 and highest plateau pressure, while measurable goals are not as clear in choosing antibiotics. based on the above confounding issues, a complex and extensive algorithm covering many potential possibilities of error and specific antibiotics seems impractical; however, an algorithm focusing on three to four major common errors in antibiotic selection and dealing with classes of antibiotics is plausible for inclusion in sepsis clinical trials. in this session we will attempt to outline the methodology of such a future study, emphasizing the immense difficulties involved in its proper conduction including: design, selection of patients, surgical considerations, supportive treatment, the "human factor" and ~nd points. only a close cooperation between a few dedicated surgeons, obsessively studying a large number of well selected and stratified patients over years, could make such a study possible. algorithmis have frequently been mistaken for the graphic format in which they are presented. however, an algorithm is neither a flow chart nor a list of instructions; it is a step-by-step approach to solving a problem. likewise, a clinical algorithm is a step-by-step approach to solving a clinical problem. cas are deterministic, which does not mean that they are rigid. they are practical rather than mathematical algorithms, that tan be used only with clinical judgment, and must be tailored to each patient. there are a number of types of, or uses for, cas that can improve sepsis management, including: diagnostic or severity scorm cas, a management ca for managing sepsis constructed according to recently published standards for use in teaching research protocol algorithms that must be used to collect data or guide implementation of a clinical trial aimed at validating one or more dicisions in the management ca; finally, protocol-style cas drawn from the management ca and should be validated using matching outcome studies. organisational problems peculiar to multicentre trials the organisation of any randomised clinical trial is fraught with difficulties, and that of a multicentre trial particularly so. there are problems associated not only with the principles on which the investigation is based but also with the detailed organisation and analysis. are all the participants truly unbiased about the relative merits of the regimens being studied? is like being compared with like -are all the end points and the standards for measuring them clearly defined so that they cannot be misunderstood? are all eligible patients being studied, and their results reported? is truly informed consent being sought that will satisfy not only the patient's right to choose what happens to him, but also the law of the land? and, finally, how can the participants be sure that the trial will be stopped at the right time, to ensure that no treatment is given to any patient that might be harmful? these questions may seem insoluble, but until we tackle them we shall never be certain. dr.m. ev~s, scar~mu~ hospital, scarborough, nonhyo~s~ y0126ql, u.k. increasing knowledge of the pathogenesis of sepsis and septic shock has led to the development of many new therapies and technologies capable of preventing, blocking or even reversing the deleterious cycle of events. recent results of subsequent multicenter trials testing some novel therapeutic drugs, however, did not confirm the promising sometimes overwhelming effects obtained experimentally. it is postulated that this is largely due to inappropriate design and conduct of multicenter trials as currently performed. one of the shortcomings emphasized in this paper is the "treatment mix" problem. multicenter trials in sepsis are performed because no single center is able to recruit the necessary number of eligible subjects within a reasonable time. each single center (icu) has its own individual policy which cannot be standardized for the trial (treatment mix). even if we ascribe that each icu did the "best" for their patients, it is highly probable that the outcome differs between the icus. this translates to the effect of the new therapy under investigation. ]t is therefore worthwhile to consider that icus must first demonstrate a certain standard based on the patients' outcome (audit) before becoming accepted as a participating center. this prerequisite ensures comparable quality between participating centers, led to the reduction of "noise level" and increases the probability of positive study results. the riyadh-icu program based on standard mortality ratios is recommended as an audit instrument. during the last decade most clinical trials of potential therapeutic agents in systemic sepsis have failed to demonstrate benefit from the drug under study. although in many instances it is entirely possible that the agent in question does not have clinical efficacy, an equally plausible explanation for the multitude of negative results is that the studies were improperly designed and thus could not show therapeutic benefit, if it existed. problems which have been identified in these various trials include inappropriate or unrealistic definitions of response to the drug; inadequate sample size, with attendant insufficient statistical power to demonstrate a difference, if it existed; insufficient standardization of customary therapeutic maneuvers in septic patients; imprecise criteria for patient entry into the study, which creates unacceptable inhomogeneity between control and treatment groups and invites unjustified post-study subgroup analysis; the lack of a formal sequential monitoring procedure for interim data analysis, which could potentially affect patient safety; and a primary analysis of study results that excludes protocol deviants and is not according to intent-to-treat. these and other problems have impacted upon the validity of the various trials conducted to date and may well have prevented the resolution of controversies surrounding the use of certain agents in the treatment of septic patients. the complexity of bacterially-induced septic shock involves a cascade of events that evolve over time, beginning with the proliferation of offending organisms and followed by the release of toxic mediators from the bacteria. endotoxins [e.g. lipopolysaccharides) from gram-negative bacteria initiate septic shock by precipitating a systemic inflammatory response syndrome (sirs) through release of a broad spectrum of hostderived mediators. over the last century, hundreds of these endogenous mediators have been proposed to serve as pathophysiological substances in this "alphabet soup" of cytokines, eieosanoids, biogenic amines, kinins, peptides, ions and hormones. an evaluation of the literature on septic shock leaves the investigator with a sense that, although many pieces of the septic shock puzzle have been presented, no clue was provided to indicate how these pieces fit together, in an attempt to assess objectively the causal role of individual mediators, we recently completed a collective meta-analysis of 24 mediators that were individually reviewed and subjected to koch-dale analysis of causality by distinguished authorities in the field (1). in summary, our attempt analyze available evidence to support a cause-and-effect relationship for putative mediators of septic shock revealed that only eight of the 24 mediators analyzed could be strongly inferred as playing a causal role in septic shock. evidence supported the involvement of endotoxln, endorphins, complement, interleukins, tumor necrosis factor, eieosanoids, platelet activating factor and calcium. other mediators either lacked adequate supportive evidence or were not conclusively involved. posttraumatic outcome may be reduced by an amplified stress response mediated by neural and humoral stimuli. the classical hormonal catabolic response is predominantly mediated directly or indirectly by neural stimuli, while most changes in inflammatory responses such as leukocytosis, acute phase proteins and changes immunofunction are mediated by humoral mediators. clinical experience from controlled studies suggest afferent neural blockade and modification of stress response to improve organ function and postoperative outcome, and limited experimental studies also suggest neural blockade to be of beneficial value in shock states. of special value may be the improved gastro-intestinal motility after neural (visceral) blockade. no outcome studies are available from patients with major truma, multiple organ failure or sepsis, conditions where humorai mediators may be more important to modify than neurally mediated responses. future studies should investigate the clinical outcome effect of combined neural and humoral mediated blockade, as well as the potential role of an initial neural blockade in elective trauma (first hit) to improve the metabolic scene and outcome if a postoperative complication (second hit) should occur. neutrophil elastase is the predominant protease of the human neutrophil. this enzyme, and a variety of other enzymes, including neutrophil collagenase, are released by activated neutrophils in the setting of high concentrations of reactive oxygen metabolites. in addition, in this extracellular environment, the normal antiprotease defense mechanisms of the organism may have been severely inactivated oxidatively especially along capillary endothelial cells. accordingly, release of neutrophil elastase which mediates inflammation and degrades most extracellular matrix proteins, including elastin, fibronectin and many forms of collagen, may contribute to tissue injury and organ failure in a variety of diseases ranging in severity and acuity from pulmonary emphysema to the adult respiratory distress syndrome (ards). as a result, significant efforts by the pharmaceutical industry have been directed not only toward the development of human neutrophil elastase inhibitors, but also toward their characterization in a variety of animal models of neutrophil mediated diseases, and their evaluation as potential therapeutics for the treatment of a variety of human clinical conditions. data from animal models of organ injury and failure along with data collected from a series of patients at risk for ards and with ards will be presented and discussed as a rationale for inhibition of neutrophil elastase. parameters that need to be monitored to obtain success in future clinical studies will also be presented in this context. have turned out to initiate and maintain organ dysfunctions in severe posttraumatic and postoperative courses. such proteolysis-induced disorders are especially rendered possible by the concurrently arising consumption of inhibitory regulators (e.g. a vproteinase inhibitor=a 1pi, antithrombin iii=at iii) via cofnplex formation and/of proteolytic/oxidative inactivation. besides the well-known destructive potency against protein substrates, proteinases such as elastase or thrombin (active or in complex with the serpin inhibitors ~ 1pi or at iii) are also supposed to increase the inflammatory reaction by inducing cytokine release or shedding of soiuble adhesion molecules from various inflammatory cells (pmns, monocytes/macrophages, endothelial cells). those cell-derived mediators may provoke further cell activation through an autocrine/paracrine loop thus increasing significantly the proteinase burden at the inflammatory focus. therefore, this noxious circle might be interrupted by a convenient proteinase inhibitor therapy to ameliorate the inflammatory process. to verify this hypothesis, high dosis of at iii (mean plasma levels up to 140 %) were applied in first controlled randomized studies on surgical patients suffering from multiple trauma or severe sepsis. in control patients we could clearly demonstrate the sequential appearance of proteinase/serpin-complexes (fa 1pi, tat), cytokines (il-8, il-6), and soluble intercellular adh6sion molecules (icam, elan, p-selectin) in the circulation. these factors turned out to be a helpful tool for early diagnosis and prognosis of forthcoming organ dysfunctions. interestingly, m at iii-treated patients a significantly reduced release/production of inflammatory mediators became obvious concomitant with the improved clinical course in comparision to an increasing deterioration in control patients. soluble mediators of inflammation are cytokines (e.g. il-1, il-6 and tnfe) as well as breakdown products of complement proteins (e.g. c5a and terminal complement complex). therefore, attenuation of both, release and generation of these synergistically functioning mediators together with stimulation of release of antagonists including soluble forms of receptors are potentially beneficial in all kind of inflammatory diseases. intravenous immunoglobulins (ivigs) are known to have an anti-inflammatory effect in humans (nih consensus conference on wig, 1990) . the mechanism of action becomes more and more clear. in single cells stimulated by anti-cd3 mab wig was able to down-regulate production of il-2, il-10, ifn% and tnfj~ significantly. igg coated solid-phase stimulates release from monocytes of interleukin-1 receptor antagonist (il-lra) but not of il-1; this observation is very much in contrast to the effect of endotoxin in the same in vitro system. furthermore, naturally occurring anti-cytokine antibodies can be demonstrated in apparently healthy individuals and in wig prepared from plasma pools. some of these antibodies can be detected by antigen/antibody reactions in fluid phase (anti-ll-le, anti-il-6), some only by solid-phase detection systems (anti-ll-8, anti-ifn% anti-tnfc 0. infusion of wig to patients suffering from inflammatory disorders have resulted in a decrease in il-6 in circulation. during infusion wig might induce an acute but transient rise of tnfo~, il-6, and il-8. self limitiation of this process might be due to demonstrable instant release of il-1 ra and soluble tnfo~ receptors. in vitro ivig has also been shown to attenuate complement activation via the classical pathway. furthermore, acid haemolysis of erythrocytes in paroxysmal nocturnal haemoglobinuria could partially be prevented by ivig. ivig was able to attenuate forssman shock in guinea pigs. we have seen a patient with congenitally elevated turnover of alternative pathway of complement who repeatedly showed an increase of haemolytic titres after administration of ivig. thus, wigs apparently have multiple potencies including attenuation of soluble mediators of inflammation and might therefore be of benefit in trauma, shock, and sepsis. significance of sinusoidal liuing cells and of humoral factors on hepatic function following sepsis and major surgery hirata k, katsuramaki t, yamaguchi h, mukaiya m, yamashiro k. regeneration of the liver after hepatic necrosis or partial removal has been extensively studied, but uncontrolled mechanisms to irreversible hepatic failure following sepsis or major surgery of liver or with hepatic dysfunction have so far not been well documented. we suggested the importance of the intrasinusoidal aggregation between sinusoidal lining cells and intrasinusoidal running ceils in the mechanisms of hepatocyte dysfunction. thus, the purposes of this study was to investigate the changes in each sinusoidal lining cell during this process and the effect of cytokine on hepatocyte under various oxygenation. [ materials and methods ] ( 1 ) clinical study : thirty two patients with histologically proved malignant tumor underwent major hepatectomy were devided in two groups, i_e. the cirrhotic liver group and the non-cirrhotic [aver group. most of the former were the patients with hepatocellular carcinoma and most of the latter were the patients with metastatic carcinoma of colorectal cancer. five patients were complicated septic conditions with hepatic failure. following operation in each patient, a verous blood sample was taken for measurement of serum il-1, il-6, tnf and c-reactive protein concentration. and hepatic biopsies were sometimes undergone in patients with anomalous hepatic function. (2) experimental study : kupffer ceils, sinsoidal endothelial cells and hepatocytes were separated from mouse ( c3hhen ) liver. and also polymorphonuclear cells and platelets were purified from blood and peritoneal macrophages were from peritoneal cavity of mouse. co-cultures between or among these cells with iipopolysaccharide were performed. cytokine concentrations in media and hepatocytic function were compared. [ results and conclusion ] our findings in above experiments demonstrated the cleanly different concept for the mechanism of hepatocyte dysfunction following sepsis or major surgery. namely, the effect of hepatoeytie function by cytokines or superioxide were significantly affected under low oxygenation, but not significant under good oxygenation. hirata m.d.,nishi 17,hokkaido,japan $120 470 immune complexes as .mediators of sepsis and immune dyafumcffon laa k horn, chxistof birkenmaier, and greg h2mon departmen~ of surgery, san frandsco general hospital, urdversr 3, of calif(~rrda, san frandsco. immune complexes (ic) a;:e commonly found circulating in parians during infection and sepsis; and following traumm, bums, and massive l~ensfusion. clearance of 1c occurs by phago~'tmsis of esll-bou_nd or opsonized p~tlcles. monocyte activation du~jng phmgocytos/s could lead ~o release of cytokilies ieletexiot~ to the hosh to examine tb/s phenomenon, we formed insoluble ic by precipitating iow-endotoxin bovine serun~ albumin (bsa) with rabbit ant/-bsa igg. we have ~town that these ic are ingested by monocytes and concomitantly stimv/ate re~pizatory bu~t activity measuxed by assay of in%racellul~ peroxldaffon. we have shown that ic ingesffon produces signlqcant el,era,lore in the monoey~e phenotype. spedacally, cdi4 is down-regu/ated, along wl~ cd 32 and cd64. thus responsiveness to lipopolysacchmdde (128) a~xd i=m~unoglobulin fe st~mu/mtlon is reduced. in cert,+rest, the complement rote.p tot cdc42 is u~-regulamd, indicaling mcremsed re~ponalx=~ness to opsonized pat,rotes, we examined monocyte superna~n~s for production of hzmor necrosis factor alpha (tnfg) following ic stimulation and showed that ic are capable of provoking ~elaase of hrmaunl)reac~ve tni~. j[c also increase mono=yte produchon of prnstaglandin e2. in summary, ~/~ese results demonstrate lhmt ic are capable of indudng production of ¢ytokines and the imrau-nosuppressive prostaglandin pge2. ic also allez ~he surface expression of important receptors involved in actlvaffon by lp~ and phagocyiosis. thus, ic aze competent for indud/on of the mepl/e s!0otlse. by examining m0n0cyte phenotypms, it may be possible to d~mrmkne extent ohn'u~u~e complex activation and predict immune depression iu criffcally ill pa~ents. a great deal of experimental work has focused on preventing and/or treating sepsis and organ failure following injury. many of these strategies have tried to attack mediator substances released during the systemic inflammatory response following injury. sugermann has demonstrated the improvement of pulmonary function, but not eappillary leak using [buprofen in the porcine model. in a clinical trial, faist demonstrated the effectiveness of indomethacine in amelierating the post trauma of cellular amenity using in vitro parameters. morbidity and mortality, however, were not different between control and treatment groups. baue und chaudry have suggested that atp magnesium chloride may have protective effect in renal failure um kupfer cell dysfunction seen after shock. pentoxiphyllin has been found to be effective in improving tissue oxygenation and oxygen consumption following hemorrhage, and there may be promise for this drug in the ischemia reperfusion injury. monoclonal antibodies against endothxin and inflammatory mediators seemed promising at first glance; however, preliminiary clinical data of the treatment of sepsis with either ha-ia or e5 monoelonoal antibodies against bacterial cell wall have been disappointing. initial studies with il-i receptor antagonist also appeared encouraging, but no difference was seen in the most recent trial. currently studies are ongoing with monoclonal antibodies to tumor necrosis factor which may have more efficacy given the fact that it is a macrophage mediator released by endotoxin. although we seem closer to understanding and preventing the problems of sepsis and organg failure after trauma, we must continue to appreciate the complex interrelationships and delicate balances inherent in the host defense system. overzealous attempts at restoring immunity in the absence of understanding pathophysiology may be just as dangerous at post-traumatic immlmospremsion. severe acute pancreatitis was caused by different etiologic factors, but once pancreatic tissues were infected, patients show a similar pattern of aggravation and develop organ failure. remarkable elevation of serum il-6 was unexceptionally observed in patients with severe acute pancreatitis. the serum levels were more than 500 pg/ml (normal: less than 4 pg/ml) during the first one or two days after onset. there was a significant correlation between the peak serum il-6 level and the number of organs showing failure (r=0.883). tnf-a production by isolated peritoneal macrophages following lipopolysaccharide (lps) stimulation was significantly increased in cerulein-induced pancreatitis rats. serum tnf-a activity was elevated following intraperitoneal administration of lps as the septic challenge both in pancreatitis rats and in control rats, being significantly higher in the former (p<0.05). histological findings and liver function tests revealed that lps induced more severe liver damage in pancreatitis rats than in control rats. these results indicate that increased tnf-a production by peritoneal macrophages in acute pancreatitis augmented lps-induced liver injury. organ failure in severe acute pancreatitis could be caused, at least in part, by increased cytokine production. it is clear now, that in mods, organ injury is not directly due to exogenous factors, such as bacteria or toxins, but instead is largely a consequence of the host's own endogenously produced mediators. there is an extensive body of literature on the inhibition of mediator production, release and action in different cascade systems by glucocorticoids. they can serve as a host protection against overactive defense reactions if used adequately. a new understanding of the regulation of cytokine synthesis, especially tnf, may lead to an insight of the conflicting findings between the benefits of glucocorticoid therapy in animals and its current failure in recent clinical trials. glucocorticoid shares its role with other novel therapeutic approaches also shown to be successful only in experimental settings. a mete-analysis of steroids in sepsis, however, revealed a beneficial effect in the subgroup of patients with gram-negative infections. there is now a series of arguments to reevaluate the indication for steroid thera-py and/or prophylaxis in mods and sepsis. honjo i-1-1, kumamoto 860, japan in septic shock j. briegel, m. halter, h. forst, w. kellermaun, m. bittl, k. peter imrodnetlea and methods: hydrocortisone (hc) at an infusion rate similar to the maximal secretory rate of cortisol in man improves hemodynamics in human septic shock (1). we hypothesized that the hc-induced hypercortisolemia prevents a harmful overshoot of immune reactions. to test this hypothesis we prospectively investigated 12 patients admitted to the icu in refractory septic shock. after resuscitation (mechanical ventilation, fluid resuscitation, titration of vasopressors, and initiation of antimicrobial therapy) hc was started with a loading dose of 100 mg/ 30 rain, followed by a continuous infusion (10 mggh). with hemodynamic improvement (dopamine < 2 gg/kg/min) the infusion rate nfhc was tapered over a period of 3 days. phospbolipase a 2 (pla2), c-reactive protein (crp), and elastase-proteinase inhibitor complex (e-pi) were deierminated daily. results: according to our previous results hemodynamics improved during hc infusien. after 4 days dopamine requirements decreased to less than 2 gg&g/min in all patients. this corresponded to an attenuation of the systemic inflammatory response syndrome (sirs) indicated by the abrogation of fever and the decrease in heart rate and inflammatory mediators. pla 2 activity and e-pi concentrations decreased to near normal values and crp concentrations decreased as well. after cessation of hc infusion (between day 5 and 7) a reinforcement nfthe sirs was observed despite cortisol levels within the normal range. this was first indicated by e-pi, followed by fever, increases in heart rate, crp, pla2, and finally by the relapse of septic shock in four patients on days 7,8 9, and 1o. a second infusion of hc again resulted in shock reversal and in a decrease in pla2, crp, and e-pi in the patients under study. discussion: there is growing evidence that the endogenous steroid hc and proinflammatory cytokines integrate a complex immunoregulatory feec~ack circuit. the elaboration of tnf, il-16, il-6, and il-8 is attenuated by-100 rag hc prior to endotoxin challenge in humans (2, 3) . cytokines like tnf, il-16, il-6, and il-8 are potent proinflammatory signals for acute reactants (--, clip), neutrophil degranulation (--, e-pi), and pla 2 synthesis and secretion. our data provide evidence that hc at an infusion rate similar to the maximal secretory rate of cortisol in man attenuates the systemic inflammatory response in human septic shock. to evaluate the feasibility of a nigh-dose regimen of antithrombin iii (at iii) treatment in patients with multiple injuries regarding blood levels of antithrombin iii and undesired side effcts and to analyse effects of the initibitortherapy on the systemic inflammatory response. patients and methods: 20 patients with an iss nf more than 28 points who could be treated with at iii administration within 180 rain after trauma were randomized to receive either at iii or placebo. at iii was given to reach an antithrombin iii inhibitory capacity in plasma of 140% by using the following calculation: at iii to administer = (140% -at iii measured) x body weight. at iii was given every six hours for a period nf48 hours and on the 4th day. patients were followed up throughout their stay in the icu but at least 14 days. in this abstract the data of the first 10 patients are included. results: the patients' median injury severity score was 41 points with a median age of 42 years. the patients were equally distributed between verum and placebo groups with respact to age , iss, prehospital treatment, blood pressure, hemoglobin and at nmevel on admission, and time from the accident until the test solution was given. the patients of the at iii group received 18,100 units of the inhibitor on average during the first 4 days. with our regimen, the at iii levels could be kept between 125% and 140% of normal, whereas control patients an at hi concentration of 40% to 60% was observed. patients with at ni treatment required less red blood cells bur received the same amount of ffp and fluids. there were no differences with respect to platelet count, partial thromboplastin time, l~rothrombin time and prothrombim. we did not observe bleeding disorders or any other side effect with peak concetrations of up to 206% of normal. white blood count, pmn--elastase, interleukin 6 and 8 and c-reactive protein tended to be lower in the at iii group. there were no differences with respect to renal and hepatic function parameters but the duration od mechanical ventilation was shorter in the at ii1 group (8.5 vs. 15.8 days) conclusions: we conclude that our treatment protocol (a) was able to restore at iil levels to the desired range of 140%, (b) did not lead to coagulation disorders, (c) did not increase transfusion requirements, (d) did not reveal other undesired side effects, and (e) showed a tendency towards reduced posttranmatic inflammation and mechanical ventilation requirements department of trauma surgery, 45122 essen, germany antithrombin-lll (at-ill) acts as an inhibitor of thrombin, further proteases and the alternative pathway of the complement system. inhibiting thrombin, at-ill works as a functional antagonist of paf~ therefore, in state of hypevolemic-traumatic shock, continuous supranormal serum -and tissue -concentrations of at-ill may be efficient to reduce the local posttraumatic reactions resulting from complement and pmn-activation. 20 patients with severe multiple trauma (7 treatment [at-ill] --13 controls [c]) were investigated. study cdteda were age > 16 and < 65 years, injury severity (iss) > 30 points and glasgow-coma-scale > 8 points; randomization and treatment has to be started within 6 hours after trauma. permission for the clinical study was given by the local ethic committee. bradykinin (bk) and related kinins are potent inflammatory peptides which possess the ability to induce, vasodilation, increased vascular permeability and hyperalgesia. cp-0127, a novel homodimer bk antagonist has previously been shown to increase survival in rat and rabbit models of lethal endotoxin shock and is now in clinical trials for sepsis. we have now evaluated the effect of cp-0127 in other models of inflammation. male rats were precannulated with a catheter in the carotid artery. 24h later bk was injected ia and the pain score ranked from 0 (no responses) to 5 (vocalization). cp-0127 at 3.6 umoles/kg completely inhibited the pain responses for a period of 2.5 -3h. cp-0127 at 3.6 umoles/kg s.c. was also found to inhibit the increase in paw volume and hyperalgesia induced in rats over a 3-4h period by an intraplantar injection of 0.1% carrageenan. the abdominal constriction response 1o an intraperitoneal injection of kaolin was inhibited in a dose-dependent manner by cp-0127. when 50 ul of 0.5% formalin was injected into the paw of a mouse a characteristic licking response was observed which was biphasic in nature. cp-0127 significantly inhibited both the first (0-5min) and second (15-30 min) phase responses. ]n a rat burn model, where the hind paw is immersed in water at 55°c for 15 sec the increase in paw volume was significantly reduced by pretreatment with cp-0127, 3.6 umoles/kg s.c. finally cerebrai edema was induced in rats by applying cold (-78°c for 10 sec) to the dural surface following a craniectomy. cp-0127 at 3.6 umoles/kg s.c. produced a significant reduction in the amount of edema compared with sham controls 24 h later. these data suggest that bk is an important mediator of inflammation and hyperalgesia and that the bradykinin antagonist, cp-0127, may be useful in the treatment of such inflammatory, hyperalgesic disorders. partial hepatectomy in humans is associated with a considerable morbidity due to hemodynamic and metabolic derangements, which increase the risk for organ failure and mortality. we hypothesized that endotoxemia may play a pivotal role in these complications. we therefore, investigated whether peri-operative infusion of rbpi23, a recombinant protein of the human neutrophil bpi with bactericidal and endotoxin-binding capacity, could prevent postoperative derangements following partial hepatectomy. male wistar rats (230-250 g.) received a 70% liver resection (phx) or a sham operation (sh), and a continuous intravenous infusion of either 0.2 mg/kg/hr rbpi23 (phx-bpi, n=8; sh-bpi, n=7) or the (iso-electric, iso-kd) control protein thaumatin (phx-con, n=8; sh-con, n-8). various parameters were measured 4 h after the resection or sham operation. mean arterial pressure, cardiac output and heart rate were significantly decreased in phx-con rats compared with sh rats, which effects were not observed in phx rats treated with rbpi23. blood ph was significantly decreased in the phx-eon group, whereas the leucocyte count, hematocrite and il-6 levels were significantly increased compared to sham levels. in the phx-bpi group, these parameters were restored to near sham levels. in vitro experiments with rat plasma and human mononuclear cells (mncs) revealed that plasma of phx-con rats is highly capable of activating mncs, accompanied by the release of cytokines. this activation is attenuated with phx-bpi plasma. in vitro added acd14 or polymyxin b was able to reduce the activation by phx-con rat plasma to the levels of phx-bpi rats thus, these data suggest that systemic endctoxemia, possibly of gut origin, is a major cause of postoperative hemodynamic and metabolic derangements following phx and that rbpizz can prevent these changes. more recently we reported a transient appearance of both endotoxin and tnf in the circulation of rats subjected to the haemorrhagic shock (hs) already at 90 -120 rain. similar to bpi, recombinant bpi was found to bind lps and inhibit tnf formation in vitro. the aim of this study was to investigate the effects of rbpi21 (kindly provided by xoma corporation, berkeley, ca) against haemorrhage related endotoxemia and mortality in rats. method: a prolonged hs was induced by blood withdrawal to a mean arterial pressure of 30-35 mmhg for 180 rain followed by reinfusion of shed blood (sb) and resuscitation with two times of sb volume of ringer's lactate over 50 rain. rbplg. 1 was administered at a total dose of 5 mg/kg i.v. (2.0 mg/kg at the16-eginning followed by two doses of 1.5 mg/kg each at end of shock and the end of resuscitation). the control group was treated similar to the bpi group but received thaumatin as a protein control preparation at the same dose as rbpi21. results: imrffe?diately after resuscitation (230 min) the detected plasma endotoxin levels in the control group (mean = 61, range = 0-120 pg/ml) were almost neutralized by rbpi21 treatment (mean = 13, range = 0-53 pg/ml) . plasma tnf levyis were not significantly influenced by rbpi treatment at the two time points 230 and 320 min of experiment (means: 65 and 51 in bpi vs 49, 65 pg/ml in the control group). the 48-hour survival rate was improved from 6/16 (37.5 %) in the control to 11/16 (69 %). conclusion: these data suggest that haemorrhagic shock may lead to bacterial translocation and/or transient endotoxemia with concomitant cytokine formation that may play an important role in the pathogenesis after shock and trauma, rbpi21 might be a useful therapeutic agent against endogenous bacterfal/endotoxin related disorders in hemorrhagic shock. morbidity and mortality after hypoxia of the vital organs had been correlated to the production of oxygen radicle which is mediated by xanthine oxidase activity, in this study we have evaluated the survival rate after allopurinol. 42 rabbits weighed 900 + 200 grams divided into two groups. group i included 22 tabbits were treated with allopurinol 80 mg/kg for seven days before induction of haemorrhage. group ii as a control included 20 rabbits. all rabbits were subjected to 25% arterial blood loss through the central ear artery for one hour then resusciatation was done by the heparinized withdrawn blood through a marginal ear vein. during the experiment blood pressure and heart rate were monitored through the central ear artery. also uric acid, lactic acid, glutathione activity were estimated. animal survival was followed for 10 days. postmortem vital organ histochemistry and histopathology examinations were done. in group i the survival after three days was 10 out of 22 while in group ii it was two out of 20. our conc|usion, allopurinol had increased the survival in aiiopurinol pretreated rabbits which may indicate the value of allopurinol premedication for patient prepared for elective bloody surgical intervention . h2 receptor antagonists are commonly used for stress ulcer prophylaxis, but their actions on the septic response are largely unknown, in an experimental model, 26 pigs were first anesthetized, then injured with 100 joules of energy to the posterior thigh, then hemorrhaged 30-40% of their blood volume. after i hr of shock, all the shed blood plus 2x the hemorrhage volume as lactated ringers was infused. following resuscitation, ranitidine (1.5 mg/kg iv twice daily) or saline placebo was begun. the treatment group was randomly assigned in a blinded fashion. after 72 hrs, a septic challenge was administered (15 bg/kg of e. coil endotoxin (lps)). serial gastroscopy, gastric ph, hemodynamics, abg's, physiologic dead space ventilation, leukocyte counts, and tumor necrosis factor (tnf) levels were recorded for 180 min. baseline values and units were cardiac index 102 _+ 9 ml/min/kg (ci), arterial po2 228 + 11 mmhg(pao2), base excess 6.5 -+ 1 meq (be), physiologic dead space fraction 23 +_ 4% (pds), and tnf 0.4 + 0.1 units/ml. baseline gastric ph was 3.9 -+ 0.4 and 4.8 _+ 0.5 in the placebo and ranitidine groups, respectively. the gastritis following hemorrhage was marginally attenuated in the ranitidine group. following lps infusion the following were obtained: ci pao2* be* gastric* pds* peak* 120 rain 60 rain 120 rain ph 120 min tnf ranitidine 167_+17 118_+15 -4.4±1. bum injury results in hypermetabolism, fever and nitrogen wasting. endotoxin (lps) has been proposed to mediate these effects, either directly or via activation of macrophages to produce cytokines such as interleukin-6 (ii-6). this study was designed to clarify the role of lps and 11-6 in the metabolic response to bum injury. twenty-five burn patients (49 -+ 17%; 16 + 15% ft bsa burn; 32 _+ 16 years old) were studied serially for three weeks post bum. patients underwent partitional calorimetry to assess metabolic rate and compartmented heat loss. nitrogen was assayed using chemiluminescence. lps and i1-6 were measured with limulus amebocyte lysate assay and elisa. patients were excluded if they suffered smoke inhalation, showed any sign of sepsis or failed to rapidly meet their nutritional needs via the enteral route. ten patients received intravenous polymixin b (5,000 u/kg/day to bind lps). these patients did not differ for the remainder. all patients were hypermetabolic and febrile in proportion to the size of their bum wound but were not endotoxemic (3.1 +_ 5.6 pg/ml; normal <40 pg/ml). i1-6 did demonstrate a significant correlation with cole temperature (tr~ = 37.6 + 0.21ogi1-6, p=0.04) and with nitrogen excretion (nou t = -5.2 4-0.091ogi1-6 + 0.14tr, p=0.01). administration of polymixin b had no effect on metabolic rate, temperature or i1-6 levels but did reduce nitrogen excretion resulting in more positive nitrogen balance (0.t grn/day vs. -0.1 gm/day, p=0.04). although bum injury does not produce an obligatory endotoxemia, i1-6 does appear to play a role in the fever and nitrogen wasting seen with such injuries. the effect ofpolymixin b on nitrogen excretion suggests that lps may play a role either locally or in the portal system. introduction: there is substantial evidence that release of inflammatory mediators by activated kupffer cells contribute to the course of a systemic inflammatory process, e.g. after shock or lrauma. besides the systemic effects of mediators such as tnf, paf or interleukines, local actions on hepatic microvasculature and hepatic inflammatory response have to be considered. our aim was to assess the role of tnf and paf by blocking their effects using anti-tnf monoclonal antibody, pentoxifylline and a paf antagonist. methnds: in anesthetized sprd-rats, hemorrhagic shock was induced by withdrawl of arterial blood within 5 rain and shock state was hold for 1 h at a map of 40 mm hg (cardiac output of 50 %). following adequate resuscitation with 60% of shed blood and twice of this volume as ringer's solntion, animals recovered to map >1130 mm hg and co >120%. hepatic microcirculation and sinusoidal leukocyte-endothelium interactions were examined by intravital epi-fluorescence microscopy at 1, 3, or 5 hours after resuscitation. in a blinded fashion, a rat-specific monoclonal anti-tnf antibody [2 mg/kg, celltech, uk) , pentoxffylline (ptx, 50 mg/kg, hoechst, d), and a paf antagonist (web 2086, boehringer, ingh., d) were given either as pretreatment or at the time of resuscitation (n=6-8 group bolla. k*., duchateau, j., hajos, gy., mbzes, t., hern~di, f. prevention of temporary/secondary immune deficiencies or reduction of their severity and/or duration as well as the reduction of the perifocal inflammatory processes belong to the rational targets of posttraumatic/pedsurgical medication. such a targeted medication can result in less frequently occurring nosocomial infections, and in reducing the duration of the intensive care and convalescence period. the results of in vitro studies performed with the amino acid sequence 32-35 of thymopoietin, i.e., with thymocartin in whole blood and peripheral mono-nuclear celi(pbnc) cultures clearly show some characteristic effects of this immunomodulator. preincubation with the tetrapeptide significantly (p<o.ol) and concentration-dependent reduced the endotoxin(lps)-induced tnfalfa production from 95,7 to about 20 pglml, but did not abolish it. the inhibition of the lps(0.1 ug/ml)-induced tnf production occured already in presence of 0,1 pg peptide/ml and peaked at the concentration of 0,01 ng/ml. higher concentrations did not increase this effect. in contrast to the other two small thymic peptides (tp-3 and tp-5), thymocartin did not enhanced the pwm-induced pge 2 production in pbmc cultures up to a concentration of 1 ng/ml, inclusive. the selection of thymocartin (tp-4) for perisurgical medication and/or for treatment of trauma-induced temporary immune depression has been strongly supported also by targeted animal experiments. thus, mortality of about 90% registered in mice infected with pseudomonas aeruginosa after bum injury could be reduced with the tetrapeptide treatment to a level which did not differ significantly from that of the infected non-burned controls. moreover, observations gathered first of all with the inflammatory model of air poach/croton oil grenuloma (selye) clearly showed, that a very definite, antiexudative effect can be achieved with this thymic peptide. as to this effect, thymocartin has been confirmed, e.g., to be equipotent with locally administered fluocinolone acetonide and/or s.c. injected prednisolone. the interim results of two explorative double-blind clinical studies (involving more than 260 patients after polytrauma and hip-fracture untill now, respectively) correspond to the expectations. the treatment significantly reduced the occurence of nosocomial infections, the days spend in intensive care and on antibiotics as well as it shortened the bed-out time and reduced the additional medication. objective of this prospective randomized study was to further delineate the mechanisms leading to these alterations and to investigate the effects of immunomodulatory intervention. patients and methods: 20 patients (pts) formed control group a. 20 group b pts received 3x50 mg indomethacin (indo) from the first (dl) to the fourth postoperative day (d5) intravenously to block prostaglandin e2induced suppression of cmi response as well as 50 mg thymopentin (tp-5) subcutaneously preop., on d2 and d4 to augment cmi reactivity. in vitro investigations preoperatively, on dl and d7 included measurements of the percentage of cd4+ t-helper (th) cells, pbytohemagglutinin (pha)induced synthesis of interleukin (il)-2 as one cytokine primarily produced by thl-cells, and pha-induced synthesis of il-6 as one cytokine primarily produced by th2-cells. delayed type hypersensitivity (dth) skin response to an antigen skin test battery and specific antibody (ab) production following tetanus vaccination preoperatively and on d7 were used as in vivo tests for thl-and th2-induced immune response respectively. results: postoperatively, group a pts showed a persistent significant reduction of cd4+ th cells, il-2/il-6 ratio, and dth skin response as compared to baseline values (bl), while ab production increased (p<0.05 vs.bl). in group b pts no change in cd4+ th cells, il-2/il-6 ratio, or dth skin response was observed (p<0.05 vs.group a), and postoperative ab production increased significantly (n.s. vs. group a). conclusions: 1.these results clearly indicate a significant suppression of th1 induced cmi response, while th2 induced response remains normal following ohs. 2.these alterations may gain clinical significance whenever a postoperative infection requires a th1 response and may explain the susceptibility to opportunistic microorganisms observed in pts after ohs. 3 the aim of this study was to find out whether the establishment of administration of thymostimulin (tp-i) in jaundiced and anergic rats would return the rats to the state of immunocompetence. material and methods. male wistar rats weighing 250-300 g were injected with haemocyanin (klh). all rats with a positive response were included in the study and we evalu ated the t cell subpopulations and il-2. they underwent laparotomy and the common duct was ligated and divided. those rats whic become jaundiced and anergic one week after the operation, were divided in two groups: control group ( objective of study. the goal of this study was to prove the necessity of thymus derived immunomodulators treatment in acute period of infantine sepsis, materials and methods. immune status was investigated in 11o infants. clinical conditions of children were defined by syndrome of multiple polyorgan]c insufficiency. the heaviness nf condition was estimated as the sum of points according to efforts needed to restore of basic living functions -respiration, hemodynamjcs, hemocoagulatlon, metabolism, functions of liver and kidneys. it was expressed in form of clinic condition classes from 1 to 4. results.it was found that in 100% of infants with 3-4 classes of heaviness immunode[iciency took place simultaneously with ii and iii stages of hemocoagulattve syndrome. amounts of t cells and t helpers were decreased more then 2 times, ratio th/ts was reduced to 1.5 or lower. concentration of lgg and igawas reduced almost a hull in comparison to control. phagocytic and metabolic activities nf neutrophils were depressed. complement system state was changed: the functional activity of the alternative pathway factors b, d and the level or c3a subcomponent were increased but the level of c2, c3,, c4 and c5 components of the classic pathway were decreased. during the development of coagnlopathy the direct correlation between chso, levels of plasminogen and antithrombine iii was found. in greater extent the function of immune system was disodered in accordance with 2 point metabolic violations and hemocoagulative disturbances considered as disseminated inmtravaseular clotting syndrome on ii or ili stages. conclusions. the rehabilitation of the immune system functions was promoted by taetivine and thymaline in doses of i-3 mcg/kg per day and 2 mg/kg per day accordingly during a week in children with 3 -4 classes el heaviness. aiter 11-14 days tactivine had advantage. when eoagulopathy took place thynaaljne promoted restoration of (1-3)-~-d-glucan (bgc) is a major cell wall compornent of pathogenic fungi and shows many immunopharmacological activities on experimental animals. lps is also derived from gram-negative bacteria and have in~ttnomodulating activities on immune systems positively or negatively. because of difficulty to cure contaminating lps from bgc preparations, it is very hard to evaluate the exact activities of bgc. in this study, we compared the immunological activities of lps ans highly purified bgc on murine system. materials and methods: c3h/hen or c3h/hej mice, 6 to 12 wkolds of both sexes were used through experiments. highly purified (1-3 )-{5-d-glucan (gurdran; bgc)was courteous gift from seikagaku kogyo co. tokyo, japan and resolved in endotoxin-free ultra-pure water. escherichia cell lps was purchased from sigma chemicals, usa. mitogenic activities of bgc or lps on splenic cells were measured by mit dye assay. in vitro activation of peritoneal exudate macrophage (pen) have moniotored by phagocytosis and intraphagocytic killing of pseudomonas aeruginosa (z~b-139. induction of cytokine productions from pem or spleen cells induced by bgc or lps were detected by cytotoxic assay. results and (bnclnsions: i ) in vitro cultures of pem with bgc have induced enhanced phagocytic and bactecidal activities in c3h/hen and c3h/hej mice, whereas lps only induced such activities in c3h/hen mice. most effective dose of bgc was 10 ug/ml. 2) bgc-dependent proliferative response of c3h/hen splenic cells was not detected under the culture condition used. 3) induction of tnf~ prodction was apparently observed in bgc-stimulated c3h/hej pd4 cultures, but not in lps stimulated one. these findings indicate that activation mechanisms of pr94 and splenic cells by bgc are different from that by lps. to investigate the bsc-induced p~4 activation, intracellular signaling pathways of p~4 by bgc-stimulation are under consideration. yamagami k, uedono y, kawakami k, kitazawa y and tanaka t according to the latest reports of use of il-2 in a burned-sepsis model, the dose was too high to adjust for human therapy. adoptive transfer of l~ymphokine-activated killer (lak) cells has been demonstrated as a means of enhancing therapy in malignant tumors. we therefore attempt to use lak therapy on burned-infected mice instead of il-2. under anesthesia, 8-weekold male c3h-hej mice were subjected to a 20% scald or sham burn and then resuscitated. sham burned mice served as controls. scald burn mice were subjected to peritonitis by intraperitoneal innoculation of 105 organisms of p. aeruginosa 24 hr after burn. burned-infected-mice were divided into two groups. one group had no further treatment, and the other group received lak cells (2x105) intraperitonelly after septic challenge. the mice were scored for survival daily. on day 7 after burn, using moabs dual-color flow cytometry, we studied lymphocyte expression in mice with use of blood and spleen. simultaneousely we studied the alteration of il-1 and il-6 in their serum using immunoassy kits. sham burned animals had no mortality. the mortality of the lak-treated mice was significantly reduced when compared with that of the burned-infected mice. the most consistent changes observed were depressions in percentages of thyl, 2 positive cells and a remarkable depression of nk cells in spleen. after lak cell treatment, those two percentages of cells significantly increased. all the data of assay of ill and [l 6 were not detected on sham burn mice serum. due to burn and septic challenge the levels of illand il 6 (n=6) were raised to 45.9_+17.4 and 3866_+1045 pg/ml, while the levels in lak treated mice were reduced to 4.23_+2.37 and 437_+110 pg/ml, respectively. the results reported here demonstrate that lak therapy is able to improve cell-mediated immunity in burned-infected mice. this is associated with a significantly improved survival rate, since ill has been demonstrated to mediate immune and inflammatory responses. also a cause effect relationship between elevated endetoxin levels and increases of [l6 has been recently established. the aim of our present study was to investigate the effect of parenteral indomethacin on the immune system of patients under major operations. our study included 20 operated patients, 10 of which received 100mg indomethacin before and 1,2 and 3 days after surgery (group a) and 10 patients received no antiinflammatory therapy (group b). we measured total t-cells helper (cd4), suppressor (cd8), nk-cells and interleukin-1 and tnf production by pha or endodoxin (lps) stimulated peripheral blood mononuclear cells at day 0 and 1, 3 and 7 days after operation. in group a we detected a significant (p<0.05) increase in cd4/cd8 ratio on day 3 while no differences were ~oted in nk-cells or cytokine production in both groups (table 1) . it seems, therefore, that parenterai indomethacin may have a benefitial effect on the immune system of patients under major operations by increasing the t-helper /t-suppressor cell ratio while having no effect on the production of cytokines by peripheral blood mononuclear cells. this study was undertaken td dete~nnine the role of prostaglandin synthesis inhibitor on survival post traulna sepsis. analysis of the effects of prostaglandin synthesis inhibitor on survival of four groups of 15 mice each were made after laparatomy and intravenous administration of live e.coli. post trauma sepsis, group a received no treatment; group b received antibiotics im; group c received prostaglandin synthesis inhibitor im and group d received both antibiotics and prostaglandin synthesis inhibitor im. survival time was longest in group treated with prostaglandin synthesis inhibitor and antibiotics post trauma sepsis. the singular use of either antibiotic or prostaglandin synthesis inhibitor did not alter the outcome on survival. mortality rate was lowest in the group treated with combined prostaglandin synthesis inhibitor and antibiotics. use of this combination showed a reduction in bacterial growth in peritoneal and blood samples, mild morphologic changes secondary to sepsis in the heart, lungs, liver, kidney and stomach and marked enhancement of mean level of activity. the study indicates that addition of prostaglandin synthesis inhibitor in treatment of trauma-sepsis has clear beneficial effects. interferon-? (ifn-y) is a potent immunostimulant which has been shown to be detrimental when administered during septic shock. blockade of this cytokine however is beneficial during the acute septic response. the aim of this study was to evaluate the cellular effects of this cytokine and its blocking monoclonal antibody (moab) in lethal sepsis following injury. 100 female cd-1 mice were randomized into two groups: septic=lps vs injury=hind limb amputation (hla vs igg. nstats= anova=p<0.05 vs lgg ifn-y was detrimental when given to control septic animals. however its blocking moab was protective (p<0.05). conversely ifn-y was proteetive in injured septic animals compared with anti-ifn-? (p<0.05). these findings demonstrate that the immune stimulant ifn-? has therapeutic potential in the immunosuppressed injured host predisposed to sepsis, while blockade of this cytokine is required for protection in the septic host with a normal immune response. dept of surgery, rcsi, beaumont hospital, dublin 9, ireland. oxidants play a role in physiology. the potential noxious oxidant biochemistry is restrained by chemically distinct antioxidants. these antioxidants, which may reside in different cellular compartments, can act synergistically. in normal physiology an oxidant/antioxidant bai&nce exists. unbalance in favour of the oxidants is called oxidative stress. oxidative stress has been implicated in many pathologies including lung diseases(e.g, doxorubicin induced cardiotoxicity), ischemia/reperfnsion damage and central nervous system trauma. transition metals such as iron are involved in the early events leading to oxidative damage in these pathologies. this has been underlined by the finding that postischemic cns pathology closely resembles that caused by a chemical oxidative insult (e.g. iron microinjection). moreover, a protective effect of oxygen-radical scavening agents or compounds that inhibit lipid peroxidation (antioxidants) in brain ischemia/trauma is apparent. some known drugs (e.g. anti-arrhythmics or histamine h2-antagonists may act as non-specific antioxidants. howe~er, recently, interesting new membrancespecific antioxidants (e.g. 21-aminosterdids) have been designed. subt&e effects on iron redox chemistry contributes to the potent antioxidant activity of these 21aminosteroids. since a few years, one area that greeted enthusimastlcally in clinical medicine is that of reoxygenatlon injury or ischemia-repeffusion, a phenomenon accepted to make a majo:" contribution to organ dysfunction in trauma, shock and sepsis through the formation o( oxygenated free radical species. however, thei detection in vivo always remains a difficult challenge. efforts have been made recently to directly establish the formation of free radicals in biological samples as complex as blood, plasma or tissue with the use of electron spin resonance (esr) spectroscopy. using spin trapping agents, the presence of reactive carbon-and oxygen-centered radicals has been demonstrated in animals blood submitted to heart, renal and intestinal ischemia-repeffusion or to liver transplantation. recently, the in vivo formation of free radicals in the cerebra; extracellnlar fluid during cerebral isehemia-repeffusion has been assessed by the spin trapping technique coupled to brain microdialysis. esr investigations have also been conducted to detect endotoxin-induced free radical generation in rat or baboon liver and oxygen free radicals (ofr), produced both at intra-and extra-cellular levels, seem to play a major role in the pathophysiology of tissue injury and organ dysfunction in sepsis, through induction of lipid pemxidation in cell membranes. oxidative damage can result both to an ofr-overpmduction or to a depletion of endogenous antioxidant defenses, which are represented by scavenger enzymes (e.g. sod), hydrosoluble and liposoluble antioxidants (e.g. ascorbate, vitamin e), and other mechanisms such as metal-ion sequestration. in animal models, oxidative damage has been proved by detection of increased levels of lipoperoxidative products, and a depletion of antioxidant defenses was found both in plasma and tissues. in the few clinical studies a similar results have been reported, and oxidative damage appears to correlate with dic, with organ dysfunction and with red blood cell deformability. in a group of critically ill septic patients we found increased levels of conjugated dienes (cd) (bioproducts of lipoperoxidation) and decreased plasma levels of alpha-tocopherol and coenzyme qlo (coqi0)(endogenous liposoluble antioxidants). a relationship between cd and uric acid (p<0.004) was found, may be related to xantine-dehydmgenase to xantine-oxidase conversion. antioxidant depletion is due both to overconsumption and to a nutritional deficiency, even if a reduced synthesis can also be present. in fact one more relationship between coqi0 and plasma cholesterol (p<0.0005) demonstrates the commun hepatic biosynthetic defect. is there any role for antioxidative therapy in sepsis? can it achives a significant improvement in the septic course, organ dysfunction, and final outcome? several antioxidant drugs, have been evaluated in experimental and clinical sepsis: scavenger enzymes, natural antioxidants, alloporinol, 21-aminosteroids or lazaroids, have been proved to reduce lipopemxidative damage, to protect organ dysfunction, and in some cases to improve survival. several questions must be addressed in evaluating safety and utility of antioxidative therapy. more specific and standardized methods must be applied to detect and quantify the lipoperoxidative damage. antioxidant drags must act selectively on ofr-production, without interference with other than that are beneficial for the organism. antioxidants must be able to achive in adequate amount the tissue or cellular compartment where their action is required. many antioxidants have also a pro-oxidative effect which can he detrimental in some conditions. the timing of administration, the dosage used and the association wih others antioxidant drugs must be defined. nutrition plays an important role as antioxidative therapy, since it supplies all compounds needed to maintain adequately levels of endogenous antioxidant or to support their synthesis. thiobarbituric acid reactive substances (tbars) concentration in serum has been determined in a large population of healthy subjects and in patients suffering acute hepatitis and chronic cases of hepatitis c, as well as several other processes. the correlation with different physiological and clinical parameters in these populations is also presented. treatment with interferon of the chronic active hepatitis c patients, 5x10 u three times a week during two months, led in those patients whose sgpt activity normalized in serum, to a concomitant decrease in serum tbars content. the possible theoretical involvement of peroxidation and antioxidants in this beneficial effect of i~terferon in hepatitis c patients is discussed. the results presented confirm the value of tbars as laboratory test in the management of disease and as a useful tool for the study of pathogenic and/or therapeutic mechanisms. 4-hydroxyalkenals are among the different substances measured by the tbars assay. these compounds show several biological effects that have been demonstrated mainly in different experimental models. we have studied the capacity of different tissues to conjugate these compounds patients surviving the initial subarachnoid hemorrhage (sah) from a ruptured in~raeranial aneurysm are prone to develop cerebra] ischemia from vasospasm which is associated with significant morbidity and mortality. among the many treatments that have been prol~osed to treat this condition, none has been shown to be satisfactorily effective. recently, a new drug, namely the 21-aminosteroid tirilazadmesylats, has been studied in a large, prospective, multicentor trial for its effectiveness to improve the outcome of patients with aneurysma! said. the study was conducted in europe and australia in 41 neurosurgical centers and included 1023 patients. all patients received presently accepted standard treatment, including nimodipine. patients were randomized to four different groups: placebo and 8 groups of tirilazad in escalating dosage (0.6 -6 mg/kg/day). the clinical outcome was compared for these 4 groups and cerebral a~giography was performed in 75 % of patients on day 7 -11 follovang sah, to study the effect 0f'the dflf~ off cerebral vasospasm. the study reached its planned endpoint 6 months ago. preliminary results ~how a significant improvement with regardto mortality in those patients receiving the highest tirilazad dose as compared to placebo and the two lower dose groups. additional beneficial effects were seen in the 6 rag/day groups with regard to the occurence of symptomatic vasospasm and good clinical outcome. although the final analysis of all data is still pending, these results suggest that tirilazad-mesylate could represent a major improvement for the treatment of patients with sail. experimentally lipidperoxidation products (lpo) are increased in acute pancreatitis (ap) due to oxygen radicals (or). the purpose of this clinical study was to determine the antioxidant status and lpo in patients suffering from ap and to evaluate the effect of antioxidant treatment with sodium selenite (se) thus improving the function of the se dependant glutathione peroxidase which is the major detoxifying system for or. materials and methods: in z0 patients sufferin s from severe ap (c-reactlve protein >120 me/l) we determined on day 1 and day 8 after admission the lpo ma!ondialdehyd (mda), conjugated dishes (cd), reduced (gsr) and oxidized (gssg) glutathione, the vitamins a,c,e and se. moreover the patients were evaluated clinically using the ranson and the apache ii score. i0 patients were randomly treated with 600 ug/day of se for 8 days. results: all patients suffered from a severe depletion of antioxidants,especially a low concentration of se (only 1/3 of normal). thereby the increase in lpo correlated with the clinical course. during se treatment lpo decreased and the levels of antioxidant vitamins improved. se had no influence on leth-slity the lenl or the chan in rs or ap ii. background: since reperfusion injury occurs when oxygen is reintroduced into ischemic tissue, the ideal timing for administration of therapeutic compounds aimed at ameliorating oxygen radical mediated injury is at the time of initial fluid resuscitation. currently used colloid or crystalloid preparations do not provide optimal, or even significant, anti-oxidant protection. systemic iron chelation affords protection against the iron catalyzed components of oxygen and lipid radical mediated tissue injury. the conjugate resulting from chemical attachment of the clinically approved iron chelator, deferoxamine (dfo, desferal ®, ciba), to hydroxyethyl starch (hes) represents a novel approach to colloid based fluid resuscitation. hes-dfo contains 85% hes and 15% chemically bound dfo. the polymer-drug conjugate has a lower molecular weight than that of hes in order to allow more rapid excretion. results: preclinical and initial clinical trials indicate that hes-dfo is well tolerated, even at high doses. in animal studies, fluid resuscitation with hes-dfo does not significantly improve central hemodynamic recovery beyond that observed with hes, but hes-dfo seems to afford better protection of microcirculation in organs at risk (lung, liver and gut), possibly by decreasing neutrophil sequestration. in a burn model, total fluid requirements are lower and oxygen utilization higher in hes-dfo treated animals compared to hes controls, suggesting decreased vascular leak and improved tissue perfusion. conclusion: hes-dfo represents a means by which potent antioxidant protection can be administered at resuscitation. iron has been suggested to play a pivotal role in oxygen flee radical mediated tissue injury. in vitro experiments indicated its critical role as a katalyst in hydroxyl free radical generation 0fenton-reaction). since iron chelator deferoxamine administered in shock alone demonstrated severe side effects, a hydroxyethylstarch (hes)daferoxamine (dfo)-conjugute was used to modulate oxygen free radical injury during the ischemia/reperfi~ion syndrome induced by hemorrhagic shock. methods. female lewis rats (190-215 g, n>6; pentobarbital anesthesia 50 mgjkg), in hemorrhagic shock ( the aim of the study was to elucidate (1) whether the generation of or would affect lung and kidneys as primary shock organs in the very early phase of sepsis and (2) whether dfo-hes could prevent this tissue damage. methods: in 67 rats sepsis was induced by cecal ligation puncture (clp) peritonitis. the animals were randomly assessed to 2 groups: one group was treated with 3 ml dfo-hes (100 mg/kg iv), the other rats received solely 3 ml of the carrier starch solution. 30, 60, 120, and 240 min after induction of sepsis respectively, the animals were sacrificed, the organs collected, and tissue contents of glutathione (gsh), malondialdehyde (mda), myeloperoxidase (mpo) and conjugated dienes (cd) determined. plasma samples were obtained for analyses of endotoxin (chromogenic lal test). blood pressure (map) was measured via a carotid artery catheter. results: clp caused sepsis with high (> 1.84 eu/ml) endotoxin levels. map in both groups decreased slightly but significantly during sepsis regardless any treatment. in the lungs mpo concentration was increased (p<0.05) in the lies group already 30 min after sepsis induction. concomitantly, tissue gsh level decreased and lipid peroxidation was pronounced as shown by elevated mda and cd levels. dfo-hes diminished tissue pmn accumulation and mpo concentration. moreover, at each time point lung mda and cd levels were lower (p<0.02). histomorphological examination showed marked micro-atelectases, destruction of the alveolar septa, and splicing of the basal membranes in the lies group. in contrast, in dfo-hes treated rats the alveoli remained well-ventiiated and only some enlarged reticular fibers without splicing were observed. almost similar results were found for the kidneys. mpo levels differed neither within nor between both groups. the slight decrease in gsh levels seen after 60 min in the dfo-hes group seems to demonstrate an oxidative stress to a lesser degree. the most impressive effect of iron chelation, however, was revealed by the lipid peroxidation products. at each time point, mda and cd levels were lower (p<0.02) compared to the hes group. light and electron microscopic examination disclosed tubulotoxic and mitochondriat damages while dfo-hes lxeatment prevented that alterations. conclusion: both the biochemical and histological results of this study reveal an early and remarkable generation of or in peritonitis-induced sepsis. thereby, these or obviously cause pulmonary and renal tissue damages, intravenous application of dfo-hes may, however, benefit by preventing early lipid peroxidation of the tissue. the proteolytic irreversible conversion of xanthine dehydrogenase (xd) to xanthine oxidase (xo) is triggered by calcium flux. the aim of our study is to clarify ~he link between intracellular ca2+ levels and xo activity determined by uric acid release, and to evaluate the efficacy of verapamil, on the generation of hydrogen peroxide associated with reperfusion by assaying lactate & pyruva~e release and the levels of cytosolic free nad /nadh ratio. experimental protocol consisted of :(a) non ischemic/reperfused experiment in which normal cardiac slices of rats were perfusated with oxygenated kreb's ringer phosphate buffer containing glucose (150mg%) and bovine albumine (5gm%) for 60min at 37°c.it composed of 3 groups, group aa (control group), and groups ab & ac (perfusate supplemented with verapamil in the dose of loo&200 mi% respectively). (b) ischemic reperfused experiment in which ischemic cardiac slices were obtained from rats subjected to 15 min ~aemorrhage.lt was also divided into two groups; group ba and bb (verapam~/ 200mi% added to perfusate}. verapamil stimulated uric acid release from normal rat cardiac slices were 267% in group ab and 767% in group ac(dose related). rates of uric acid release is enhanced by verapamil in group bb. moreover, rates of uric acid release in groups ac & bb are insignificant. in verapmil added groups (group ab, ac & bb), increase uric acid release is associated with an enhancement in pyrurate release and with increase levels of cytosolic free nad+/nadh ratio, although it is not evident ~ ischemic group (group ba).it is concluded that the conversion of xd to xo is calcium independent. eicosanoids like thromboxane a2, leukotriene b4 and leukotriene c4 are known as promoters of initial inflammatory reactions. we investigated whether oxygen radicals (or) are able to induce a release of these eicosanoids in whole blood. blood from healthy volunteers was incubated with xanthine oxidase/hypoxanthine to generate oxygen radicals. after 0,5,15,30 and 60 minutes plasma levels of thromboxane b2 (txb2), leukotriene b4 (ltb4) and leukotriene c4 (ltc4) were determined via elisa technique. another volunteer had taken 1000mg aspirin one day before taking the blood sample (no7). results: txb2 plasma levels increased from 14pg/ml at 0min to 144pg/ml, 350pg/ml, 370pg/ml and 660pg/ml at 5,15,30 and 60min (p<0,01) . ltb4 and ltc4 plasma levels showed an increase during the first few minutes (ltb4: 0min: llpg/ml,5min: 87pg/ml; ltc4: 0min: 23pg/ml, 5min: 97pg/ml (p<0,05)) followed by a decrease to normal values at 15min. in the sample no7 the cyclooxigenase-pathway was completely inhibited, the txb2 plasma-levels did not alter at all, whereas ltb4 and ltc4-plasma levels weren't affected. opallogeneic blood transfusion jane shelby, ph.d., and edward w, nelson, m.d, there have been numerous investigations dudng the last two decades examining the effect of surgery, anesthesia, blood loss and transfusion on vadous immune parameters in humans and animal models. there appears to be concurrence among several well controlled studies that transfusion of whole blood (containing leukocytes), has regulatory effects on immune ceil function which include decreased cell mediated immune response, and inhibition of il-2 secretion. these effects occur following transfusion alone and in con.cart with the distinct immune effects of surgery, trauma and anesthesla, the clinical consequences of this immune modulation by transfusion include decreased allogeneic response to transplanted organs, which has been exploited clinicelly in renal transplant patients. additionally, there is evidence for a strong association with increased risk for infection in transfused patients following surgical procedures. aiiogeneio blood transfusions have been shown to inhibit cellular anti.bacterial mechanisms, causing increased susceptibility to bacterial pathogens, in humans and in animal models. there is also concern that allog~neic transfusion may adversely affect cancer patients, resulting in decreased disease-free survival. several stategies have been proposed to minimize the adverse effects of blood transfusion. there is evidence that the risk of immune mediated infectious complications associated with transfusion may be greatly minimized wlth the use of autologous blood and leukocyte free allogeneic blood.products in surgical and trauma patients, it also appears that the inhibition of cellular immune response and il-2 productiorl following atlogeneic blood transfusion may be mediated by increased prostaglandin e2 secretion, and that immune response may be preserved in allogeneio whole blood transfused subjects receiving c3lc~oxygenase inhibitors such as ibuprofen. among these are various alterations in immune function. efforts have therefore been made to utilize alternatives to homologous transfusions. these include the use of autologous predonation, supplemental iron therapy, and recombinant human erythropoietin. although initially considered innocuous, these therapies are now recognized to have potential deliterious immune sequelae. erythropoietin, by its ability to lower serum iron levels, can impair both lymphocyte and nk cell activity. autologous donation impairs nk cell function. finally, supplemental iron therapy can stimulate bacterial growth and increase the rate of infectious complications. this talk will present a discussion of these factors as well as a weighting of their importance. r.l rutan, rn;bsn, shriners burns institute and the university of texas medical branch, galveston tx, usa the serious sequelae of homologous blood transfusions have resulted in vigorous efforts at identifying alternate therapies for correcting red blood cell (rbc) deficits. erythropoietin (epo) was hypothesized to exist in the early 20th century, however the protein was not isolaled until 1977. the human gene was identified and cloned in 1983, which permitted the production of epo through recombinant techniques. the earliest clinical trials were performed in anemic end-stage renal failure palients on hemodialysis. treated patients experienced increases in erythropoiesis with normalization of hematocrit and hemoglobin levels, cessation of lrans-fusion requirements and improvement in general wellbeing. these studies, however, identified side effects of epo treatment such as hypertension, seizures and ee deficiency. volunteer trials have established that the hypertension is not a direct pressor effect but rather the result of abnormally rapid increases in red cell mass in the face of the incompetent volume-controlling mechanisms of the end stage renal failure patient. lower doses of epo and the subsequent gradual increases in red cell mass are associated with significantly lower incidences of hypertensive complications of epo therapy. likewise, seizure activity is not the result of a direct epileptogenie effect but parallels the incidence of hyper-tensive-related sequelae during high.dose epo treatment. in cross-over designed studies, pre-existing iron deficiency has been demonstrated to decrease or negate stimulated erythropoiesis but effective-hess can be restored with appropriate fe supplementation. exogenous epo is effective whether given by iv or sq routes and dose response curves do not vary with route of administration. increases in rbc mass are directly related to the dose of epo, both in amount and frequency of administration although there is a 3-5 day time lag between the first epo dose and laboratory indications of its action (i.e. increase in the number of reticulceytes in peripheral wood). epo is currently labelled for use in the treatment of anemias associated with end-stage renal disease and aids. however, its use in the surgical population has been explored because of its unique direct dose-response, epo has been used to effectively increase the blood harvest amounls in autologous pre-donation, significantly increase hematocrils in children following thermal trauma and successfully increase red blood cell mass following essential surgical procedures in patients with religious aversion to transfusion. by blood transfusion in colorectal cancer surgery mm heiss md, ch delanoff md, r stets md, j hofinann, e faist md, kw jauch md, fw schildberg md allogeneic blood transfusions are associated with an increased risk for postoperative infections in colorectal surgery when compared with autologous blood transfusions. attribution of this effect to immunomodulation was suspected in our previous study (lancet 1993; 342:1328-33) . task of the recent investigations was to analyze which specific effector systems were affected in-vivo by this transfusion-associated modulation. for global in-viva assessment of cell-mediated immunity (cmi) multiple recall skin-reactions were applied prior and post-operative. the specific humoral immune mechanisms were investigated by applying tetanus-toxoid one day preoperatively and deterimnating the quantitative igg-response. for indication of macrophage stimulation in-vivo tnf-levels were determinated by bioassay. dth-responses were significantly suppressed (p<0.05) in patients receiving allogeneic blood (n=36) or operated without blood transfusions (n=17). dthresponses were not suppressed and tendentiously increased in patients with autologous blood transfusions (n=24). in contrast, specific igg-levels increased sigmficantly (p<0.05) in patients receiving allogeneie blood (from 1.6 + 4.4 to 9.7 _+ 18.4 ie/ml) whereas in patients receiving autologous blood a smaller increase (from 1.9 + 4.1 to 4.0 + 6.7; p=0.068) was observed. tnflevels demonstrated a similar pattern with a higher increase in patients receiving allogeneic transfusions (l 1.4 + 12.0 to 28.5 + 11.4 u/ml) compared to those patients with autologous blood (8.7 + 12.5 to 17.1 + 23.0). in conclusion these data indicate that allogeneic blood transfusions lead to a remarkable macrophage/rhs stimulation. this is corroborated by the boostered humoral igg-response which was initiated before onset of surgical trauma and blood transfusion. concerning cmi this caused a substancial suppression probably due to a stimulated secretion of immunosuppressive monokines. objective: firstly, to analyse the concentrations of the cytokines tumor necrosis factor (tnc), interleukin-1 (il-i), interleukin-6 (il-6) and coagulatioo/fibrinolysis parameters in postoperatively retrieved blood from a surgical area, secondly to characterize the correspanding cytokine patters in the patients and thirdly to study cytokine concentrations in the initial portion of drainage blood from a surgical area. materials and methods: blood retrieval was performed in a closed-loop system without anticoagulant during 4-6 hours after surgery in 10 patients undergoing arthroplasty (9 hips and 1 knee). 7kf, il-1, it-6, thrembin-antithrombin complexes (tac) and antithrombin (at) ~ere determined in shed blood. patient plasma tn v, il-i and il-6 concentrations ~ere analysed at the beginnlqg and end of the 4-6 hour blood retrieval period. in a separate study (5 hip arthroplasties) f~f, il-i and il-6 ~ere determined in the initial portion of drainage blood. cytekine analyses ~re performed usiog ipmuooassays. an omidolytic method was used for at determinaf.ion and tac was analysed by elisa. n~n-poram~tric tests was used for the statistical comparison. results: the patient plasma il-6 coocemtratiems rose from a median value of 0 to 116 pg/ml, p<o.01, during the blood ret.rieval period. in c was detectable in 4 patients, range:15-50 pg/ml. il-i was not detectable in the patients. in retrieved blood tag was >200 mg/ml in all samples (ref:<4.1 mg/ml) and at was 0. 16-0.51 units/ml (ref:o.80-1.20) . the il-6 concentrations in retrieved blood was >1000 pg/ml in all samples. tn v or il-i was not detectable. in the separate study, (n=5), characterlzing eytokine content in the initial portiere of drainage blood, in= (range: 11-277 pg/ml) and il-i (range:5-125 pg/ml) ~re present in all samples but ii-6 (range:o-25 pg/ml) was detectable in o.qly one semple. conclusion: theses findings indicate that hypereoagulability and hic~ ccrcentratioos are present in retrieved blood. the cytokine pattern in the initial portion of blood from a surgical area differed from these observed in retrieved blood and in the systemic circulation. to identify the role of both autologous and homologous blood on postoperative infections in elective cancer surgery. materials and methods: 159 patients with colo-rectal cancer submitted to curative elective surgery were prospectively studied. on hospital admission the following nutritional measurements were assessed: serum level of albumin, cholinesterase, delayed hypersensivity response , total lymphocyte count and weight loss, as were age and sex, duration of operation , operative blood loss, amount and type of blood given, pathological dukes' stage of the disease and the attending surgeon were also recorded. results : eighty-four patients (52.8%) were perioperatively transfused. thirty-six (42.8%) patients were given autologous blood , while 48 (57.2%) received homologous blood. no patients received both autologous and homologous blood. twenty eight (17.9%) patients developed postoperative infections. non transfused patients had a 9.3 % infection rate , those receiving autologous blood had a 13.9 % infection rate, whi]e in the homologous blood group the infection rate was 33.3% (p < 0.001). univariate analysis showed that infections were significantly related to operative blood loss (p<0.01), length of operation (p<0.05) blood transfusion (p<0.05) and attending surgeon (p<0.05) . multivariate analysis identified homologous blood transfusion as the only variable related to the occurrence of postoperative infections , while the other variables failed to reach statistical significance. blood transfusion (bt) remains an essential life-saving treatment for surgical patients. however, besides the beneficial short-term impacts, negative longer-term effects are observed, which include various alterations in the immune responsiveness. in surgical patients these alterations may contribute to the increased risk for infections and cancer recurrence. since relatively few data demonstrate immunologic changes occurring in other lymphoid compartments than blood after bt, we studied the effect of et on the frequency and responsiveness of immune cells in bone marrow (bm), spleen (spl) and blood (b) in a rat model. normovalemic, 2 month old rats were transfused intravenously with syngeneic heparinized venous blood (3x2ml, every other day), and 3, 7 and 14 days after the last transfusion bm cells ( leh is an experimental oxygen-carrying resuscitation fluid. since leh is cleared from the circulation primarily by the mps, its effect on the development of sepsis and the nature of its relationship with the mps remain a major concern. preliminary in vivo data from our laboratory failed to show any leh effect on the hemodynamic and hematologic responses to endotoxin lipopolysaccharide (lps) in the rat. in contrast, leh exacerbated the lps-induced tnfa production and early mortality. the exacerbation of early mortality by leh was attenuated by pretreatment with the tnfu synthesis inhibitor rolipram. ex vivo, peritoneal macrophages from rats treated with leh and lps have shown increased il-lg mrna signal as compared to lps alone. also, leh increased tnftx production by peritoneal macrophages in response to lps stimulation in vitro. additionally, recent pilot studies indicate that leh attenuates pma-induced superoxide production from rat peritoneal macrophages and that leh augments fmlp-induced migration of human monocytes. taken together, these data strongly support possible interactions of leh with the mps and therefore the nature of such interactions should be further explored. over the last decade, we have developed liposome encapsulated hemoglobin (leh) as an artificial oxygen carrying fluid, or blood substitute. our efforts have focused on studies to define the safety and efficacy of this resuscitative solutions. leh consists of distearoyl phosphatidylcholine, cholesterol, dimyristoyl phosphatidylglyeerol, and alpha tocopherol in a 10:9:0.9:0.1 mole ratio and can encapsulate hemoglobins of different origin (bovine, human, recombinant human). leh is fabricated using hydrodynamic shear to create an average particle size of 0.3 microns. leh can be lyophilized using disaccharides and stabilized in the dry state and easily reconstituted before administration. histopathology and clinical chemistries indicate that leh rapidly accumulates in tissue resident macrophages in small animals injected in the tail vein, principai1y in the liver and spleen. the consequences of accumulation in the reticuloendothelial system are manifest by transient increases in liver transaminases (ast, alt), bilirubin, and bun over 24-48 hours with no change in biliary function (ggt, ap) . clearance through the liver and spleen is observed over the course of 1-2-weeks. more recent attention has been focused on secondary consequences of leh administration especially with regard to inflammatory eytokines. leh does not elicit expression of tumor necrosis factor in vivo and in isolated macrophage cultures, but does result in a transient increase in serum il-6. we have also examined the interaction of leh with lps in vitro macrophage culture to further understand how this blood substitute may effect the immune system. we have labeled leh with technetium-99m (99mtc) to study the biodistribution of leh non-invasively in anesthetized rabbits. rabbits were infused with a 25% topload of leh (200 mg of phospholipid, 2.5 g of hemoglobin per kg of body weight) and imaged continuously with a gamma camera. at 20 hours, images were again acquired. animals were then sacrificed and tissue counts obtained, images revealed an initial rapid uptake bythe liver, 8% at 30 minutes and 15% by 2 hours. the spleen accumulated activity at a slower rate, 3% at 30 minutes and 7% at 2 hours. at 20 hours, autopsy biodistribution studies revealed that approximately 42.6% of the dose is in the blood pool, 15.4% in liver, 18.1% in spleen, 3.2% in lungs, 2.4% in muscle and 1.6% in urine, with trace levels in kidney, brain and heart (<1 °/o). in a hypovolemic model, rats were 10% or 50% exchange transfused with 99mtc-leh. in the 10% exchange model, 99mtc-leh was rapidly taken up by the liver and spleen with minimal activity in the circulation at 20 hours. with the 50% exchange, 50% of the leh was in circulation at 20 hours. the interaction of leh with platelets labeled with indium-111 was also studied. after infusion of leh, the labeled platelets rapidly moved from the circulation to the lungs and liver. over the next 30 minutes, the platelets gradually returned to circulation. this effect was not seen with iiposomes of the same lipid composition but containing no hemoglobin. non-invasive imaging is proving to be a very useful tool for the investigation of leh. the need for a safe, efficacious and commercially viable blood substitute is unequivocal. of the several strategies pursued to invent an adequate blood substitute, liposome entrapped hemoglobin (leh) has been already established as a leading possibility. major advances in liposome technology have already resulted in liposome preparations compatible with clinical use for drug delivery. recent technological advances made by the u.s. naval research laboratories resulted in the capacity to entrap hemoglobin into liposomes in a way which secludes hemoglobin from interacting freely with biological systems. the leh produced has already been tested in in vivo systems and was foun.d to be well tolerated. moreover, the leh originally produced as a solution can be transformed into a lyophilized form which can be reconstituted and delivered as a fresh solution. while important milestones in leh development for a practical blood substitute have been achieved, several issues remain to be explored. most notably, the long term consequences of leh on host defense mechanisms and, in particular, immune cell function. in addition, it is important to understand more fully the metabolic fate and repercussions of leh delivered at clinically relevant dose/schedule regimens. finally, while leh is a highly promising strategy for a blood substitute, the present formulations consist of human hemoglobin derived from human blood, to improve the safety profile, a recombinant preparation for liposome entrapment will be much desired, aa-ginine, a semi-essendai dietary amino acid, possesses several unique and potentially pharmacologic properties. argirdun is a potent secretagogue for pituitary growth hormone and prolacfin and for pancreatic insulin and glueagon; it modulates host protein metabolism by increasing nkmgen retention and enhancing wound collagen synthesis. it also is a potent t call function regulator. ait of these effects coupled with its relative lack of toxicity and safety make it an a~antive nulritionai pharmacologic agem (t). rodents fed supplemeutal arginine exhibit increased thymsc weight which is due to increased numbers of thymic lymphocytes present in the gland. thymic lymphocytes from animals fed supplemental ar~e demonstrate increased blastogenesis in response to coma. and pha (2) . peripheral blood lymphocytes from humans given supplemental arginine also have heightened mitogunic responses to mitogen or antigens (3) . in postsurgery padents supplemental arginine abrogates or diminishes the deleterious effects of trauma on lymphocyte responsiveness and restores peripheral blood lymphocyte responses much faster than observed in controls. overall host immunity is also enhanced by arginine. allograft rejection is enhanced and septic animals survive longer when given supplemental arginine (4) . tumor bearing urginine-supplemented animals have decreased tumor growth and enhanced survival (i). lastly, asgmine can induce t cell maturation and t cell mediated responses in athyrnic nude mice. arginine also has remarkable effects on host nitrogen metabolism post-injury. in increases nitrogen retention in healthy human volunteers and in surgical patients. this beneficial effect on overall nitrogen metabolism is accompanied by a unique effect on the healing wound. supp]emental arginine increases wound collagen synthesis which also translates into increased wound breaking strength (5) . arginine has no effect ou epithelialization. douglas w. wilmom, m.d. boston, ma gintamine is the most abundant amino acid in the body, but it has long been considered a nonessential amino aeid because it is synthesized in many tissues. fohov~g st,~'vation~ injury or infection, skeletal muscle pmteln inoresses its net tale of degradation and releases amino acids into the blunds~mm at an aocelerared rate. app~o)~mately one-third of the amino nitmgea is ghitamine, which is metabolized by the kidney where it parth:~pates in acid-base homeostasis, is the primly ~ for lymphocytes, mac~optmgcs and untexocyms, and contm'butcs to the synthesis of giumth~une. olmamine degrades slowly while in ~olu~ou, especially at usual room teml~mtums. because giulamine was considered nonessential, it has beer absent r'om nil intravenous and most gluts.mine should be considered a cendittona]ly essential nutrient for individuals with serious ilinesses, uspccially those confoanded by infcctinn and inflammation. over the uc~:t 5-7 years, glutamine will be incorgorated into most feeding formulas designed for patients with critical illness. 526 o]~ga-3 pufa there continues to much interest in the application of the 0mega-3 pufa in clinical nutrition. the basic principle has been that the 0mega-3 pufa will displace arachidunic acid and result in a decrease in eic0san0id production. in addition these changes in pufa will after the physical characteristics of the membrane including flujdity, receptor function and transmembrane signals. animal studies have shown that there is omega-3 incorporation with continuou~ enteral feeding both in control and endotoxic animals within 3 days. this includes the liver, spleen, circulating and alveolar marc0phages and the lung. this incorporation resuls in significant changes in the eicosan0id production including pgf 2 and ket0-pgflalpha. there is improvement in the cardio-vascular reep0nse of these animals with ~ecreamed lactic acidosis and improved cardiac contractility. as well there is improved immune function with improved t cell response to mit0gens. the ~ of a mumber of pharmacological agents blocking cicosanoid production can enhance the cell effects of 0mega-3 pufa. clinical studies using short term entsral nutrition with 0mega-3 either alone or with other enteral supplements in a number of clinical settings have shown significant 0mesa-3 incorporation and decreased eicosan0id production. these positive results must be discussed with the additional evidence that long term omega-3 supplementation decrease eic0san0id production but als0 induce a state of immune suppression that is capable of increasing transplant sunvival. these 10ng te~ inune effects may benefit clinical conditions including rheumatoid arthritis and cr0hn' disease early enteral nutrition instituted i~mediately afte~ injury will decrease the entry of bacteria into the intestinal wall and decrease the number of bacteria that translocate into the portal blood. these reductions are associated with & decreased catabolic response, decreased plasma cortisnl levels, end decreased vma excretion in the urine and prevention of mueosal atrophy. sdecific nutrients also affect the transloeation process. addition of arginlne to the diet significantly improves the ability to kill translocated organisms. however. translooetion across the gastrointestinal barrier is not affected. in contrast, glutamine diminishes the rate of translooation across the imtestinal barrier and also improves killing of the beetarla that do translooate. the omega 3 fatty acids in the form of fish oil slightly decrease the rate of translocation but more significantly increase the ability of the animal to kill translo~ated organisms, all three dietary additives, i.e. argini~e, glu=amine and fish nil. significantly improve survival, hut adding glyoine or medium chain triglyeeridem do not, combinations of srginine and glutamlns, glutamine and fish oil, and fish ell end arginine each improve survival, and to a greater degree than a combination of all three. these studies add further evidence that translocation is an important determinant of survival after injury, early feeding with immunonutrlent enriched dices will improve survival and dsarease transloeation to varying degrees, depending upon the nutrients provided. objectives: we studied effects of supplementing a commercial enteral diet, impact r (imp, sander nutr lnc), with fiber (imp/fib) or alanyl-glutamine (imp/ag, exogenous glutamine (gln) 14 gms/l) on influencing the incidence of bt to mesenteric lymph nodes (mln) in burned mice. fiber has been shown to improve gi integrity under certain stress/treatment conditions. the dipeptide ag is a water-stable source of gln, which is a specific fuel for many cells including enterocytes. traumacal (trcal), a high-protein, high-fat enteral diet (mead johnson iuc), was also studied, as well as rodent chow (harlan teklad inc), which contains very high protein & fiber. methods: anesthetized cf-1 mice aged 8-12 wks received 32% tbsa fullthickness dorsal burns & were resuscitated with 2 cc ip saline. diets were allowed ad lib; caloric intakes were comparable in all gps except fasted gp (fast 24 hrs, chow 24 hrs). at 48 hrs postburn mln were sterily removed, homogenized and plated on heart brain infusion agar; cfu/g mln tissue were determined. bt was analyzed by fishers exact test, cfu/g by anova-bonferroni. * p<0.01, ** p<0.05 compared to imp and burn-fast gps. background. infectious complications following trauma, major operation, or critical illness adversely affect hospital cost and length of stay (los). some key nutrients have been shown to possess immune enhancing properties. this multicenter trial was conducted to determine if early administration of an enteral formula supplemented with arginine, dietary nucleotides and fish oil can decrease los and infectious complications in icu patients. methods. this was a prospective, randomized, double-blind study of 326 adult icu patients who required enteral feeding for > 7 days. patients entered the study within 48 hr of the event, were stratified by age and disease, and were randomized to receive either the supplemented formula (impact®) or the conventional formula (osmolite ® hn). feedings were initiated at full strength and advanced to at least 60 ml/hr by 96 hr after event. results. both groups tolerated administration of formula well. for patients fed > 7 days, the median los was 25% shorter (p=o.ol) for the--supplemented group (21 days) compared to the conventional group (28 days). the incidence of most infectious complications was lower in the supplemented group, but this difference reached significance only for urinary tract infections (p=o.o05). the supplemented group had a significantly shorter los from onset of infectious complication until discharge for patients with pneumonia (19 vs. 27 days) and skin/soft tissue infection (19 vs. 37 days). conclusions. administration of the supplemented formula was safe and well tolerated. when fed > 7 days, it reduced the incidence of most infectious complications, and significantly reduced los. materials and methods: twenty-seven patients were randomised into 3 groups ( n= 9 each) to receive either a standard enteral formula, the same formula enriched with arginine, rna and omega 3 fatty acids (enriched group) or isonitrogen, isocaloric parenteral nutrition. early enteral nutrition was started within 12 hours following surgery (10 ml/hour). it was progressively increased reaching a full regimen on day 4. on hospital admission and on post-operative day 1 and 8, the following parameters were assessed: serum level of transferrin , albumin , prealbumin, retiool binding protein (rbp), cholinesterase. delayed hypersensitivity response, igg, igm, iga, lymphocyte subsets and monocyte phagocytosis ability were evaluated on admission and on post-operative day 1, 4, 8. the three groups were comparable for sex, age, cancer stage, type and duration of surgery, intra-operative blood loss and amount of blood transfused . in all groups a significant drop in all the nutritional and immunological parameters was observed on postoperative day 1. comparing post-operative day 1 versus day 8 a significant increase of prealbumin (p<0.02) and rbp (p<0.005) was found only in the enriched group. with respect to immunological variables an increased phagocytosis ability (p<0.001) and a significant recovery in delayed hypersensitivity response (p< 0.005) was observed only in the enriched group. conclusions : these data are suggestive for a more effective post-operative recovery of both. nutritional and immunological status in cancer patients fed with enriched enteral formula. gastrointestinal intolerance was equivalent (18% in each group) and laboratory screening confirmed that both diets were safe. when analyzing clinical outcome for all patients, there were no significant differences in septic complications (immun-aid = 41% vs vivonex ten = 35%), mean mof score (immun-aid = l.b vs vivonex ten = 3.6), or mortality (immun-aid 0% vs vivonex ten = 6%) . kowever, when analyzing the subgroup of patients with severe injury (iss or ati _> 25), patients receiving immun-aid appeared to have fewer septic complications (33% vs 50%) and their mean mof was significantly lower (1.8 _+ 0.7 vs 5.8 + 1.7, p = 0.05, student's t-test) . these preliminary data indicate that immun-aid is tolerated well when aggressively delivered immediately postinjury. the ultimate affect on clinical outcome appears ~avorable for immun-aid, but needs to be confirmed in larger patient groups. kemp?n, m., neumann, h.a., he i[michh b: as both increased, normal and reduced phagocytic capabilities of polymorphonuclear leukocytes (pmn) and monocytes in acute batterial infections have been reported, the role of phagocytes in patients with severe sepsis is less clear.we examined pmn and monocytes from 10 patients in septic shock and 17 heailhy votunteers for phagocytic function. phagocytosis was determined by flow cytometry (facscan) and was measured by the ability of pmn and monocytes to phagocytose e.coli marked with fluorescent antibodies. a septic shock was defined by the presence of a ~ource of i, nfoctiqn with a known bacteriology, distinct signs of a systemic response and defined minimum scores in 3 icu scoring systems indicating the presence of a multiple organ failure. additionally we examined how phagocytosis is influenced when a new enteral diet formulation containing substrates suggested to improve immune function or arginine, one of its major compononts, is added in vitro in defined concentrations and incubated for 10 minutes. pmn (p{o,05) and monocytes (p<o,01) of the septic patients showed a significantly enhanced phagocytosis compared tolhe phagocytes of the healthy group. the incubation with the enteral diet in vitro was followed by a higher dose-related rate of phagocytosis, especially in the healthy group, that was not statistically significant. after addition of l-argintne we atso observed an iqcrease in phagocytosis, that was lower than in the group with the complete diet. using a modern immunefluorescence-cytometric essay we founfl an improved phagocytic function in septic shosk. there are signs for an influence of nutrient substrates on phagocytosis which seems to be complex and should be further investigated. arg and gln were lower in the 49%-bcaa vs 16%-bcaa group; arg was related directly to arg dose and inversely to bcaa dose (p<o.o01 for all). clearances of bcaa increased with increasing bcaa dose (p<o.o01); arg and gln clearances were directly related to the bcaa clearances (rz=o.65, p<o.o0i). relationships between arg, gln and other aa seem to be ruled by patterns involving specific intracellular aa transport systems. regulation of arg and gln clearances through bcaa administration may be related to an increased flux of aa into synthetic processes. intestinal segments from rats immunized by infection with the nematode trichinella spiruus undergo intestinal anaphylaxis or type i hypersensitivity when challenged in vitro with parasite antigen. immunized rats exposed to t-radiation (7gy) and sustained on rat chow,fail to fully express type i hypersensitivity up to 14 days post irradiation (dpi). we hypothesized that enteral nutritional support immediately following irradiation may be effective in preserving mucosal immune responsiveness or in reducing the recovery time after radiation-induced injury. rats were infected with 3x103 t.spiralis larvae. thirty to fifty days later rats received 7gy t-radiation from a co source as total abdominal irradiation (tai). immediately following tai, rats were fed an elemental amino acid diet (eaad) and at various dpi sacrificed and tested in ussing chambers for local anaphylaxis, expressed as antigen-induced ci secretion. the normal ci secretory response to antigenic challenge which is suppressed after irradiation,was restored by 3 dpi when rats were placed on the eaad. antigen-induced c[ secretion is dependent on the interaction o~ antigen with specific ige antibodies on the surface o5 mucosal mast cells and their subsequent degranulation. mast cell-derived 5-ht,histamine and pg, together effect ci secretion.in irradiated rats on rat chow,the failure to respond to antigenic challenge appears to be associated with the destruction of mast cells. they are undetectable with standard staining procedures and mucosal histamine levels are drastically reduced. also,el secretion can be induced by direct stimulation of epithelium with cr secrctagogues and circulating ige levels are normal. despite a more rapid recovery of immune responsiveness in irradiated rats receiving eaad,mast cells were not restored. we conclude that the eaad contributes to an adaptation that supports the expression of local anaphylaxis in the absence of mast cells. total parenteral nutrition (pn) and bowel rest may influence the host response to infection. this study examined the different host response to infection in parenterally and enterally fed animals. forty-three rats were divided into pn group and total enteral nutrition (en) group. they received identically constituted isocaloric isonitxogenous diets for seven days. animals were then challenged intraperitoneauy with 3xl(y escherichia coli and survival were observed. in other experiments, they were sacrificed before and two hours after bacterial challenge. peritoneal cavity was lavaged with 10ml saline and peritoneal exudative cells (pec) were harvested and cultured in vitro for 24 hours with and without lipopolysaccharide (lps). colony f(m, ning units of bacteria (cfu-b) in the peritoncal lavaged fluid (plf) were determined and tnf in the plf, serum and pec culture supernal,ants were measured by l929 bioassay. twenty-fonr hour survival rate in en group was significantly greater than that in pn group (60% vs 0%). the data suggest that local immune responses of pn-treated animals may be depressed with consequent disturbance in the clearance of bacteria. this may lead to a greater systemic cytokine response and resulting in a poor survival after bacterial challenge in pn group. the effects of intragastfic tube feeding of diets enriched with m-6 polyunsaturated fatty acids (pufa) from safflower oil (so) or (o-3 pufa from menhaden oil (mo) on eicosanoid production, and onthe membrane phospholipid fatty acid composition were investigated in a severe acute septic shock model. the percent calories from fat maintained at 30% in the diets was adjusted to provide 10% each of saturated, monounsaturated, and pufa using olive oil and palm oil as filler fats. after feeding for 5 days, lipopolysaccharide (lps, 9mg/100g) was injected through the tail vein. the percent of animals surviving in the mo group was 67% (14/21), and did not differ significantly compared to 57% (12/21) in the so group. the plasma concentrations of tnf ct for the groups of animals fed so (1.2+3.0 ng/ml) and those fed mo (1.8+1.1) did not differ significantly. the levels of prostaglandin(pg)e2, 6-keto-pgflc~ and tumor necrosis factor-ct (tnf-c 0 were determined 90 min after lps injection. the fatty composition (relative mole%) of the liver was determined before (lps-) and 3h after the lps administration (lps+). the data (mean_+sd; n=6) are as follows. group these data indicate that a marked reduction in the plasma levels of poe2 and 6-keto-pgflct for rats fed mo diet was associated with a reduction in aa which was displaced by epa in the membrane phospholipids. further, in these rats, we observed that there was a 2.8% reduction in the percent of epa following lps challenge compared to the basal levels. however, for the group of rats maintained on so diet, the liver membrane phospholipid fatty acid composition before and after lps administration was unaltered. these finding suggest that although a reduction in aa was associated with a decrease in the production of pro inflammatory eicosanoids after 5 days of continuous mo feeding, there was no marked effect on the survival following lps challenge. animal models have been used to examine the effects of various nutrients and nutrient combinations upon the immune system. we studied the effects of standard and specialized enteral formulations on the response of mice to infectious challenge with listeria monocytogenes, influenza a or candida albicans in order to test the efficacy of specialized ingredients. cf-1 outhred female mice (12-15 g) were fed purina #5002 chow, commercially available osmolite hn ®, perative ® or impact*. mortality and tissue burden of pathogen were examined during the 21 d period following induced infection. the results indicate that there were no differences between the groups fed the liquid formulas with regards to mean survival time or % survivors in these models of infection. examination of spleens in the groups challenged with l. monocytogenes and kidneys in the groups challenged with c. albicans revealed no differences in tissue burden of pathogenic organisms. alterations in the length of pre-feeding from 0 to 8 days prior to infection did not affect these results. these data demonstrate that, contrary to previous reports, the use of specialized nutrients did not improve resistance to disease in mice challenged with l. monocytogenese, influenza a or c. albicans as compared with mice fed osmolite hn. animals fed standard chow tended to be more resistant to infectious disease than those fed the liquid formulas perhaps due to the form of diet or complexity of ingredients. the results indicate that these animal models of infectious challenge may be of limited use to distinguish effects of modified nutrient composition of enteral formulas. laboratories the beneficial effects of sesame oil (ses) have been attributed to the presence ofsesamin, a non-fat constituent which is claimed to iultibit a-5 desaturase activity. we investigated the effects of ad libidum feeding of ses enriched diet on fatty acid composition of phospholipids from plasma, liver, on eicosanoid production, and on the protective effects in mice after cecal ligation and puncture (clp) and compared with safflower oil (so) diets. olive and palm otis were added as filler fats to the so diet to maintain the ratios of saturated, and unsaturated fatty acids similar to ses diet. thus, while the total fat remained at 15wt%, the only difference between the two diets is the presence ofsesamin in ses diet. the animals were fed for 3 weeks. the incorporation of dihomo-y-linolealc acid (dgla: 20:3 o)-6) in the phospholipids from plasma and from the liver for the group of mice fed so diet ranged between 1-3% compared to the undetectable levels for those fed so diet. there were no significant differences in the incorporation of other fatty acids either in plasma or in the cell membranes between the two groups. following an intraperitoneal administration oflps (10p.g/kg body wt), the plasma levels of both pge2, and txb2, were significantly decreased (p<0.02) by nearly 50% for the group of animals maintained on ses diet compared to those maintained on so diet. these findings suggest that sesamin inhibited the release of arachidonic acid (aa) which upon accumulation is retroconverted to dgla which could reflect a modest increase in the content of dgla. failure to decrease the formation of aa could mean that the effects of sesamin on chain elongation process may be after aa is formed, and that sesamin may not inhibit a-5 desaturase activity. on the other hand, our data suggest that sesamin present in sesame oil inhibited the activity of cyclooxygenase which converts aa to its metabolites), or alternatively could block the activity of phosphalipase a 2 (pla2) (which releases aa from membrane phospholipids) as is evident from a marked decrease in eicosanoid production. the percentage of animals surviving after cecal ligation and puncture in rite group of mice fed ses diet was 65% (13/20) which was markedly higher (p<0.01) compared to only 20% (4/20) for the so group. increase in survival in the group of mice fed ses diet was associated with nearly 50% reduction in the production of tnf in response to lps i.p. challenge, for ses group compared to so fed animals. our data suggest that sesamin, present in sesame oil, decreased the production tnf~, inhibited the activity ofpla 2 and consequently offered a significant protection against clp. thus sesamin or sesame oil appear to be novel antiinflammatory agents which are present naturally in many edible products. [dept. surgery, ~e. deasoness hosp.,harvard medical school, boston, usa] stress causes alterations in the homeostasis of an organism with major changes in various organs, lifespans of cells, protein turnovers, metabolic pathways of activation or stimulation, energy mobilization, etc. punctual changes have been described in several organs and various cell types of the nervous, endocrine and immune systems. many stored or newly synthesized proteins are released to the bloodstream to be transported to target organs or to be disposed of. as stress alters protein synthesis and requirements in the targets, the bloodstream is a useful system for monitoring the relevant changes. we present here the pattern of newly synthesized or released serum proteins in c57bl/6 mice that have been stressed by immobilization. the proteins have been labelled with 3ssmethionine in vivo and the study was performed by two dimensional gel electrophoresis based on the method originally described by o'farrel. the electrophoretic run was carried out in an isodalt apparatus. a molecular dynamics laser scanner and the kepler image analysis system (lsb, rockville, usa) were used for modelling the radiofluorographic images. the radiofluorographic images of the gels from serum samples obtained from mice injected with i mci of 3ss-methionine reveal about 440 protein spots, from which a small fraction -about 20are altered in both qualitative and quantitative manner. the major changes are in the range of 25 to 60.000 d. the analysis displays a highly reproducible pattern, where clear differences between stressed and non-stressed conditions are shown. differences between patterns attributed to chronic vs. acute stress will be presented. patients with severe trauma or major surgery are known to acquire alterations in neutrophil functions which contribute to their enhanced susceptibility towards microbial infections. we analyzed neutrophil functions from fourty patients admitted for major surgery 6 days and 1 days preoperatively and on the ist and 6th postoperative days (study-days i, 2, 3, and 4) in a randomized placebo-controlled double-blind study. patients were divided into two groups, one ~eceived 6 days preoperatively an enteral nutrition enriched with omega-3 fatty acids, arginiee, and rna (impact), the other an isocaloric, not enriched control nutrition (placebo). meutrophils were isolated from the peripheral blood and stimulated with the ca-ionophor a23187 (7.3 pm). the capacity of the respective neutrophils to generate leukotrienes was analyzed by rp-hplc. neutrophils from impact group generated significantly higher amounts of ltb5 (mean values 6.6 ng/l x i~ cells) compared to the placebo group (2.6 ng) at study-day 2 [p, 0.01). in contrast, the capacity of neutrophils to produce ltb4 was not significantly different in both patients populations at study-days 2 and 3. the omega-oxidation of ltb4 to its biologically less active metabolites oh-ltb4 and cooh-ltb4 was not significantly different in both groups. however, neutrophils from group f patients generate more ltc4 at study-day 2 (p(0/5). the capacity of the patients'neutrophils to generate reactive oxygen radicals upon stimulation with fmlp, pma or the caionopbore a23187 exhibited patient dependent differences but no correlations to the repective nutritional supplementation as was measured by luminol dependent ohemiluminescense= these data indicate that an enteral nutrition enriched in omega-3 fatty acids lead to alterations in distinct parameters of neutrophil functions. clinical patient characteristics and mean caloric intake were similar between the two groups. both formula were tolerated well and the incidence of diarrhea was low. the number of infectious and wound complications as well as the apache ii score was evaluated for the two study groups. although the number of complications in the group supplemented with arginine, rna and omega-3 fatty acids was lower no significant difference in the occurance of infectious and wound complications has been observed between the two groups. however, the apache ii score indicated a significantly improved clinical outcome in patients with supplemented diet. in conclusion, early enteral nutrition with an arginine, omega-3 fatty acids and rna supplemented diet helps to recover more rapidly after surgical total parenteral nutrition (tpn} is associated with intestinal atrophy and dysfunction attributed to the absence of the non-essential amino acid glutamine in current parenteral nutrition solutions. in this animal study with 18 male wistar-rats we tested the hypothesis that glutamine-dipeptide supplementation during tpn for 10 days would restore small bowel atrophy and tpn induced dysfunction. a conventional tpn solution (250 kcai/kg bw) was compared to an isocaloric and isonitrogenous tpn (0,5 g n/animal/d) supplemented with alanin-glutamin (ala-gin) dipeptide (0,15 g n derived from ala-gin =30% gin-n). an enteral fed cqntrol group was included. mucosal thickness, villous height and architecture, absorption of water and glucose as well as dissacharidase activity of maltase and alkaline phosphatase were evaluated. total parenteral nutrition in rats causes villous atrophy, a significantly reduced absorption rate and a decreased activity of villous enzymes compared to an enteral fed control (p<o,01). addition of ala-gin to parenteral nutrition solutions prevents intestinal atrophy and restores functional alterations with a significantly increased rate of water and glucose absorption as well as a higher dissacharidase activity compared to the standard tpn group (p<0,01). there was no significant difference found between the enteral fed control and the dipeptide supplemented animals concerning intestinal structure and absorption, the enzyme activity was significantly decreased in the ala-gin supplemented group compared to the enteral fed control (p<0,01). in this rat model supplementation of parentera] nutrition solutions with ala-gin dipeptide restores tpn induced intestinal atrophy and dysfunction. boston. ~ usa injury, infection and major o1~'ations s~nulate a variety of factors whi~ initiate the body's response to th~ st~..sses. 'i"hese reactions are m~liated by a cbmage ia the neta'al hormonal, eac.ranmemt, by the elaboration of eytokines and tl~ougb the stimu/ation of other inflammatory mediators. this eatabolio setting zesulis in body protein loss, which g-taduai1y erodes both fimetianal and stm~ tissue. wiu3_b the normally nom'ished individual can withslzod "d~ response for a brief period of time, ircolonged eatabollsm l~ associated with ~,nmunosuppmssion, poor wound healing, and proioagcd convalescence. surgeons have rej2ed on intraveaous or eater~ feedings to ~everse this process. a va~ety of data now indicates that it is extremely di~edt to replete protein-containing tissue d~g the eataholie state of illness; the umutt response to hypercaloise feedings in this setting is the incxeased accretion of body fat md water. administration of gxow~ ho~moae ~cesults i.a n shift of the body's oxidative machinery to fat rather than eazbohydmte utiq~afic~; ¢oncomitaatiy, protein synthesis is stimulated. this metabolic state may have clinical utility in p~e.n.ts who need to heal large wotmds, in those who need to gain strength in order to be w~ed 5ore veatilatory support, or those patients who ~ve mulfiorgan failure and nee~ to ealaaace protein syatsesis to in.suze recovery. 235als are now in progress to dcterm~e the benefit of growth hormone in these and other disease state.s, rn the fature, the increased use of growth hormone, will occur;, this and other growth factors will serve to su~2po~ the host and shorten convalesceace following catabolic diseases. our objective was to study effects of pre-trauma growth hormone (gh) treatment on liver and skeletal muscle metabolism in a trauma model in piglets. twenty-four piglets were randomized to three treatment groups; one group was pretreated with gh 24 iu daily three days before the experiment (gh-3), another group treated with gh 24 ie at the start of the surgical procedure (gh-1) and one group served as untreated controls (con). they underwent a standardized surgical procedure to place sampling cathethers in the portal, hepatic and femoral veins and doppler flow probes around the portal vein, the hepatic and the femoral arteries. all pigs recieved a primed constant infusion of u-14c-glutamine. substrate fluxes were measured one (lh) and five (5b) hours after termination of the surgery. nitrogen excretion was significantly decreased in the gh treated animals (gh-3:30.8+4.2 mg/kg, gh-i: 37.6+_.5.8 mg/kg, con: 55.4+-3.1 mg/kg, p=0.004). in the hindieg, there was reduced net glutamine efflux due to decreased absolute glutamine release (gh-3:0.9 + 0.8gmol/min at lh and -2.4+1.4 in.tool/rain at 5h, gh-i: -3.4+1.5 i.tmol/min at lh and -1.4+1.2 pmol/min at 5h, con: -8.7+_2.8g mol/min at lh and -11.6+3.4/amol/min at 5h, p=0.005, p=0.006), whereas the absolute uptake of glutamine in hindleg was unchanged (p=0.84). glucose uptake in the hindleg was decreased (p=0.0006). net glutamine uptake in liver was attenuated (p=0.03 at 5h), whereas net glutamate release from liver was increased (p=0.005). thus, pre-trauma treatment with gh improved nitrogen economy, attenuated net glutamine loss from hindieg due to decreased absolute release, and attenuated hindleg glucose uptake. liver net glutanaine uptake was decerased and net glutamate release increased. patients with major abdominal surgery exhibit a considerable catabolic response with negative nitrogen balance and protein breakdown, which may have detrimental effects on outcome. we investigated the effects recombinant human growth hormone on substrate metabolism in parenterally fed patients. 48 metabolic healthy patients were randomized to receive either placebo or hgh in a dosage of 0,075; 0,15 or 0,3 i.u. per kg bw. substrate oxidation rate, energy expenditure as well as short life proteins were measured daily. six patients were excluded from analysis as drop outs, due to surgical complications. we could find a dose-dependend effect of growth hormone on resting energy expenditure, carbohydrate oxidation, fat and proteine oxidation. with increasing growth hormone resting energy expenditure increased from 20.9 calories per kg body weight to 26.6 calories. this was mainly due to an increased carbohydrate and fat oxidation, while proteine oxidation decreased. this was in accordance with a decreased urea.production rate and an improvement in cumulative nitrogene balance, which increased from minus 12.7 to plus 8.5 g n / 7 days. in consequence short life proteins were also increased. the only negative side effect was an increased blood glucose concentration in some of the patients, making insuline administration mandatory in 3 patients. our results are in accordance with other studies showing improvement of nitrogen balance, maintainance of lean body mass and muscular strength. if growth hormone administration may have any impact on morbidity, mortality and length of hospital stay in these patients it should be evaluated in a larger number of patients. in 30 patient after liver transplantation (oltx) the effect of recombinant human growth hormone (rhgh) on protein metabolism and hormonal changes was studied. patients and methods: the underlying disease in all patients was end stage liver cirrhosis, non of the patients included in the study was suffering from malignancies. the patients were randomly allocated to receive either 81u of rhgh bid (sc) or no alternative medication. the study period lasted 14 days. from daily 24 hrs udne collection urinary nitrogen loss and daily loss of urea were determined. body compostion analysis (bca, bioimpedance, akern-ltaly) was performed preoperativly and at the postoperative days 7 and 14 to evaluate changes in body water and body cell mass (bcm). the effect on resting energy expenditure (ree) was analysed using indirekt calorimetry wrhin the same intervalls (metabolic monitor, datex). blood levels of gh, igf1 and igf2, igfbp3 were measured daily during the study period, on day 3 a 24 hour profile of gh was.performed. samples were collected each 20 minutes. results: cumulative udnary nitrogen and urea loss were not signifcantly different for the whole study period but for the first 4 study days. bia indicated for the rhgh-group a loss of bcm to a lesser extent than for the controls (n.s.), changes in bodywater where similar for both groups. ree changes (kcal/kg bcm) were not significantly different for the two groups. blood levels of gh and igf1 differed significantly between the groups for the whole study period, but not igf2 and igfbp-3. the circadian rhythm of endogenous gh-excretion had not been altered by gh, but absolute levels of gh were increased. conclusion: during the first 4 days after oltx we could proof the protein sparing effect of gh treatment. although the differences between the two groups were not significantly different for the whole study period the results showed on all days a less catabolic situation for the rhgh treated patients. to possibly improve protein balance after major abdominal surgery we studied the effects of gh on protein metabolism after ltx. excluding malignant diseases, 29 candidates for ltx were randomized to either postoperatively receive the usual treatment (co=control group, n=14, age range 25-59 yr, bmi 23.9+5.7 kg/m 2) or for 14 days additional gh (2x8 i.u.s.c.) (gh group, n=14, 21-59 yr, 21.8+2.2 kg/m2). metabolic studies (calorimetry and lsc-leucine infusion) were performed 5 (test a) and 11 days (test b) after surgery. tracer analysis was done by gas chromatography-mass spectrometry. during the 2-week study clinical parameters did not differ significantly between groups; however, there was a tendency for increased glucose levels, insulin need, and energy expenditure at the time of test b in the gh group. leucine oxidation (x+se) during test a;b was 117+10; 111+15 (co) and 110+7; 100±12 (gh) pmol/kg ffm/h (n.s.). protein breakdown was 134±16; 131+30 (co) and 156+14; 178+30 (gh) pmol/kg ffm/h (n.s.). nonoxidative-leuine disposal was 119±9; 127+16 (co) and 139±11; 177+20 (gh) pmol/kg ffm/h (p<0.05 for the rise in the gh group). tracer data were supported by cumulative urea nitrogen excretion (days 1-4: gh 52+3 vs co 66±7 g; p<0.05). we conclude that after major abdominal surgery gh lowers nitrogen excretion by increasing total body protein synthesis. it is not known which organs contribute to this effect. in several investigations it has been shown that the protein saving effect of recombinant human growth hormone (rhgh) in the postoperative state is accompanied by raised igf-1 levels in plasma. it was concluded that the anabolic effects of rhgh were probably, at least in part, mediated by igf-1. this study was aimed at detecting the efficacy of igf-i on modulation of the protein metabolism in the catabolic state. patients and methodes: 38 patients (both sexes, age: 40-75 years, bmi 17-30 kg/m 2) with major upper gastrointestinal surgery (gastrectomy or partial gastrectomy) received 80 ug rhlgf-1/kg body weight/twice daily or placebo during 5 treatment days beginning on the first postoperative day in a double-blind, randomized study. all patients received hypocaloric and hyponitrogenous total parenteral nutrition (3g chng, o.6g aa/kg) troughout the whole study period. cumulated nitrogen balance, 3-methyl-histidin exretion in urine as well as serum-igf-1 levels have been determined. the two tailed wilcoxon test was used for statistical analysis results: first results will be presented previous studies have suggested that growth hormone (gh) potentiates various activities of leukecytes. however, it is not clear whether gh can improve survival of animals with gram-negative sepsis. we investigated the effects of gh on host response to infection in septic mice model. methods balb]c mice were randomized to 3 groups (n=10/gronp): gh-high (4+8mg/kg/day), gh-iow (0a8mg/kg/day) or control (saline). following subcutaneous treatment (every 8 hours for 6 days) with gh or saline, mice were challenged i.p. with e.coli (lxl08/body). survival rate was recorded every 2 hours. in the next experiment, gh-high and control animals were sacrificed 4 hours after bacterial challenge. peritoneal cavity was lavaged with 3ml saline and the peritoneal exudative cells (pec) were harvested and counted. colony forming units (cfu) of bacteria in the peritoneal lavaged fluid (plf), liver, lung and blood were determined. pec were cultured in vitro for 24 hours with lipopolymccharide (10p.g/ml in normotensive adults growth hormone (gh) rises when clonidine challenge is performed. recently evidence was shown for gh to accelerate wound healing. while gh secretion patterns are inconstant the gh-dependant insulin like growth factor (igf-1) and the insulin like growth factor binding protein (igfbp-3) reflect the integrated gh secretion over days. since we administer clonidine to patients with acute alcohol abuse we determined plasma levels of igf-1 and igfbp-3. materials and methods: 21 patients sheduled for esophagus resection were investigated once they had given written informed consent. 8 patients showed acute alcohol abuse and were treated with clonidine postoperatively• 13 patients with no history of acute alcohol abuse served as controls. besides liver enzymes igf-1 and igfbp-3 were determined. results: both groups had a comparable hepatic capacity of synthesis with decreased cholinesterase levels as to be expected after a major operation. regression analysis of igf-1 and igfbp-3 levels showed no differences between the groups. discussion: igf-1 and igfbp-3 plasma levels are not increased in normotensive patients who are treated with clonidine for prevention of postoperative alcohol withdrawl syndrom. further studies are required to discriminate whether our results are due to impaired postoperative liver function or if clonidine has not any impact on the gh-igf-axis in patients with alcohol abuse. recent studies have revealed that igf-i has several immanoregulatory roles. however, little is known about its effects on septic animals. we tested whether igf-i could increase the survival o f mice challenged intraperitoneally (i.p.) with e. coli. m~thqd$ balb/c mice received either high dose igf-i (n=10, 24mg/kg/day), low dose igf-i (n=10, 2.4mg/kg/day) or saline (11=10) every eight hours subcutaneously for six days. animals were then challenged i.p. with lx108 e. coli. survival was observed every two hours. in the other experiment, mice that received high dose igf-i or saline were sacrificed four hours after bacterial challenge. peritoneal cavity was lavaged with 3ml saline and the peritoneal exudative ceils (pec) were harvested and counted. colony forming units (cfu) of bacteria in the peritoneal lavaged fluid (plf), liver, lung and blood were determined. in addition, pec were cultured/n vitro for 24 hours with lipopolysaccharide (10).tg/ml). tumor necrosis factor (tnf) in the supernatants of pec culture, plf and serum were measured by l929 bioassay. results igf-i improved the survival rate at a dose of 24mg/kg/day. severe chronic neutropenia (scn) is a disorder characterized by severe neutropenia secondary to either a maturational arrest of myelopoiesis at the level of promyelocytes (kostmann-syndrome; scn) or regular cyclic fluctuations in the number of blood neutrophils with a median absolute neutrophil count (anc) of less than 500/~tl (cyclic neutropenia). patients suffering from scn have an impaired acute inflammatory response to injury and an increased susceptibility to infections, i.e. bacterial or fungal infections, resulting in chronic oral inflammation, severe pneumonia, abscess formation and bacteraemia. we have treated 36 patients (32 scn, 4 cyclic neutropenia) with r-methug-csf (filgrastim). thirty of 32 patients with scn and all 4 cyclic neutropenia patients responded to this treatment with an increase of the median anc to greater than 1000/~tl. the dose of r-methug-csf needed to achieve and maintain the response varied between 0.8 and 120 ~tg/kg/d. long-term treatment did not exhaust myelopoiesis: the anc remained stable after up to 6 years of treatment and antibodies to r-methug-csf were not detected. the increase of anc was associated with dramatic clinical responses: significant reduction of severe bacterial infections, reduction of intravenous antibiotic treatment, and reduction of hospitalizations. no severe bacterial infections occurred in any of the r-rnethug-csf responders during longterm treatment. severe adverse events, most likely due to the underlying disease, included the development of mds/leukemia in two patients, and osteopenia/osteoporosis in 12 patients. these results demonstrate the beneficial effects of r-methug-csf treatment in severe chronic neutropenia patients. the newborn is predisposed to mortality during infections due in large part to developmentally immature host defenses. recently cytokines have been identified that regulate the development of these defenses. one such cytokine is specific for neutrophils and is termed granulocyte colony stimulating factor (g-csf). in the studies to be discussed, we have attempted to impact hematopoiesis in the fetus using exogenous rhg-csf delivered through the pregnant dam. our rationale was that an enhanced myeloid system may result from such in utero treatment leading to, perhaps, enhanced protection to microbial insult. rhg-csf crossed the placenta and reached peak fetal serum concentrations 4 hours after administration. these levels were 1000-fold lower than the levels detected in the adult dams serum. white blood cell counts were elevated approximately 2-4-fold over control newborn blood. this increase was due to circulating numbers of neutrophils. the marrows from these pups were hyperplastic consisting of predominately immature and mature neutrophilic cells. no changes were seen in the thymus or livers. pups born to mothers treated with rhg-csf since day 15 of gestation (parturition in rodents occurs on approximately day 21) survived a lethal challenge of group b-13 hemolytic streptococcus. in contrast their untreated yet infected counterparts did not survive. recent clinical developments have led to trials of rhg-csf in certain neutropenic states including cyclic and chronic neutropenia. we have identified several of these patients who were pregnant during treatment with rhg-csf. biologically and anfigenically identifiable rhg-csf was detected at increased levels in the cord serum of these neonates. we will discuss our findings with these patients in light of our animal model of neonatal myeloid development. we have previously demonstrated decreased g-csf production and g-csf mrna expression from activated mononuclear cells from newboms compared to adults (cairo, pediatr res 31:574, 1992) . we have also recently observed that administration of a single dose of rhg-cse to one-day-old newborn rats induced significant neatrophilia, while administration for 7 days induced neulrophilia and increased the bone marrow (bm) neutrophil storage and progenitor pools (cairo, blood 76:1788 , 1990 cairo, pediatr res 27:612, 1990) . we embarked on a phase i trial exploring the safety, pharmacokinetics, and biological efficacy of g-csf in newborns with presumed sepsis. 42 infants <72 hrs old with a diagnosis of presumed sepsis were stratified into three groups: very preterm (26-30 wk), preterm (31-35 wk), and term (>36 wk) and randomized to receive either a placebo or 1, 5, and 10 gg/kg/qd or 5 and 10 p.g/kg/bid of rhg-csf infused by pump over 1 hour for 3 consecutive days. cbcs were obtained at 2, 6, 24, 48, 72 and 96 hrs. tibial bone marrow aspirations were performed 72 hrs after study entry and differential counts and cfu-gm pools were determined. c3bi expression was determined at 0 and 24 hrs after rhg-csf, and g-csf pharmacokinetics were performed after the first dose of rhg-csf utilizing a sandwich elisa. a significant increase in the anc was observed at 48, 72 and 96 hrs following administration of both 5 and 10 ~tg/kg/d of rhg-csf. the maximum increase in the anc occurred 72 hrs after 5 and 10 ~tg/kg/d (397 -116%) (p<0.01) and (621% -+ 200%) (p<0.001), respectively. there was a significant dose-dapendeat increase in the bm neutrophil storage pool (18 _+ 4% vs. 62 + 6%) (p<0.01) (placebo vs. 10 ~tg/kg/d). there was no significant difference in the nantrophil proliferative pool. an increase in cfu-gm and cfu-gemm was seen at all doses tested, compared to placebo (53.5 _+ 8.6 vs. 87 -+ 15) (colonies/l(p cells/plate). c3bi expression was significantly increased 24 hrs after 10 bg/kg/d of rhg-csf (233 + 81% vs. 83 +-26%) (p<0.012). peak serum g-csf levels occurred at 2 hrs and were dosedependent. the half-life of rhg-cse was 4.44 + 0.4 hrs. most importantly, there was no observed toxicity from g-csf in all patients studied. 31 of 42 patients were on ventilators prior to administration of rhg-csf and there was no increase in pulmonary toxicity. these preliminary data suggest that rhg-csf is well tolerated at all gestational ages in newborns with presumed sepsis. a multi-center phase ii/iii randomized double-blindad placebo controlled trial is required to determine the efficacy of rhg-csf in this clinical setting. we investigated the effects of recombinant canine granulocyte-colony stimulating factor (g-csf) on survival, cardiopulmonary function, serum endotoxin levels and tumor necrosis factor (tnf) levels in a canine model of lethal bacterial septic shock (clinical research.41:240, 1993) . methods: awake 2 ylo beagles had serial cardiopulmonary and laboratory studies before and for up to 10 days after intraperitoneal placement of an e. celi infected clot. nine days before and daily until 3 days after clot placement, animals received high (n=17) or low dose (n=17) g-csf or protein control (n=20) subcutaneously. results: survival in high dose g-csf animals (14/17) was significantly improved compared to low dose (10117) and controls (12120) (p<0.04 wilcoxon). high dose g-csf also improved cardiovascular function evidenced by a higher mean left ventricular ejection fraction (day 1 after clot, p<0.001) and mean arterial pressure (day 2, p<0,02) compared to low dose and controls. high dose rcg-csf increased (p<0.001) peripheral neutrophil numbers both before and after clot implantation (2 hours to 4 days) compared to low dose and controls. in addition, high dose rcg-csf produced a more rapid (p<0.0001) rise (day 2) and fall (day 4) in alveolar neutrophils determined by bronchoalveolar lavage compared to low dose and controls. lastly, high dose rcg-csf decreased serum endotoxin (2 to 8h, p<0.002) and tumor necrosis factor (tnf, 2h, p<0.02) levels compared to low dose and controls. discussion: these data suggest that therapy with g-csf sufficient to increase peripheral neutrophil numbers during peritonitis and septic shock may augment host defense and endotoxin clearance, reduce cytokine levels (tnf) and improve cardiovascular function and survival. the use of g-csf in sepsis prophylaxis in neutropenic patients is well established and has been ascribed to accelerated recovery in granulccyte counts. here, an additional sepsis-prophylactic property could be demonstrated in healthy volunteers: eleven volunteers were employed in a sinqle-btind, controlled study and were given 480 uq g-csf or saline placebo via subcutaneous injection. blood was withdrawn immediately before and 20 or 44 hours later. lps-inducible tnf, il-1, stnf-r p75 and il-lra were assessed in the supernatant of whole blood incubations stimulated with 10 ug/ml lps from salmonella abortus equi. similarly to previous animal studies, lps-inducible tnf was attenuated by about 50% 20 hrs. after treatment. the same was true of il-lb. in contrast, lps-inducible stnf-r p75 which was indetectable in blood incubations from untreated donors increased dramatically 44 hrs. after g-csf treatment. il-lra found after lps challenge was increased tenfold by g-csf treatment. it is concluded that g-csf treatment switches peripheral leukocytes to an antiinflammatery state characterized by an attenuation of il-i and tnf releasing capacity and an augmentation of the release of cytokine antagonists. this findinq minht offer a novel concept in septic shock prophylaxis. objective.the aim of the study was to investigate the effect of recombinant human g-csf (rhg-csf) on survival, bone marrow neutrophil myelopoiesis, neutrophil counts, levels of bacteria and some important sepsis mediators in a model of rat abdominal sepsis. lethal peritonitis was induced with a 4 mm coecal perforation (cp) in male wistar rats. rhg-csf was administered as 10 /.tg/kg iv every 12 h, first dose at sepsis induction. bone marrow neutrophi] progenitors were determined as blast colonies, cfu-gm and cfu-g. neutrophils and bacteria were determined in peripheral blood and peritoneal fluid. lps, tnf, endothelin 21 and lactate were measured in blood from femoral vein. mortality rates were registered with g-csf treatment starting either 1 or 7 days before or 4 hours after cp. results. mortality was reduced from 90% to about 50% with rhg-csf intervention and there was no difference between the pretreatment and treatment groups. bone marrow blast colonies were not influenced while neutrophil myelopoiesis was augmented at the stages of cfu-gm and cfu-g. neutrophils in blood and peritoneal cavity were enhanced and numbers of bacteria in the same compartments were substantially reduced. circulating lps, tnf, endothelin 21 and lactate were attenuated the first 24 hours after cp. neutrophil myelopoiesis is augmented with increased number of neutrophils in blood and peritoneal cavity, resulting in enhanced clearance of pathogens. lps, tnf, endothelin 21 and lactate are suppressed the first 24 hours during sepsis course. a. wendel, j. barsig, g. tiegs gm-csf stimulates the proliferation and differentiation of granulocytic and monocytic progenitor cells. in addition the hemopoietic cytokine activates the inflammatory response in mature leukocytes. the priming effect of gm-csf towards lipopolysaccharide (lps)-induced cytokine production in vitro has been described, but little is known about proinflammatory gm-csf effects in vivo. we detected gm-csf in plasma of lps-challenged mice with kinetics similar to tnf, reaching peak levels 2h after lps administration. gm-csf pretreatment (50 ~tg/kg i.v.) enhanced mortality in mice challenged by a sublethal dose of lps. plasma levels of tumor necrosis factor (tnf) and interleukin-6 (il-6) were significantly enhanced. a monoclonal antibody, which neutralizes gm-csf bioactivity, rendered mice less sensitive towards lethal lps-challenge. tnf-and il-6-tevels were reduced in these mice compared to control animals without antibody treatment. in addition, severalfold potentiation of lps-induced cytokine release by gm-csf was observed in vitro in murine bone marrow cell cultures. these data demonstrate the proinflammatory capacity of gm-csf and suggest that the hemopoietic cytokine plays also a role as an endogenous modulator of lps toxicity. immune dysfunction, developing in the wake of multiple trauma, overwhelming infection and other forms of critical surgical illnes% is associated with increased infections, morbidity and mortality. the mechanisms responsible for alterations in immune regulation are incompletely understood but monocyte appear to play a central role. polymorphonuclear leukocytes (pmn) are known to play a central role in the inflammatory response of the host toward invading microrganisms. reports of defects in all the aspeots of pmn function have been accumulated in recent years. the possible role of gm-csf in modifing the state of immuno suppression detected in severe intraabdominal infected pt~. inspite of surgical appropriate procedures and in reducing the expected mortality is investigated. the safety of rh-gm-csf administration in sepsis is also evaluated. a double blind randomized study is proposed. this study include 60 icu patients who do not exhibit signs of shock and/or ards, with clinical signs and symptoms of abdominal infection. immunodepressed patients-aids, chronic chemotherapy or chronic steroid administration do not partecipate to the study. patients will receive rgm-csf (l~g/kg/day) or placebo in 24 hs. continuous infusion for 15 days. safetyandefyieacy will be assessed till to day 30. the apache ii score is adopted for risk stratification of patients because it is reliable and validated, objective and composed of information that is indipendent of diagnostic criteria. patient's entry criteria is apache ii > 16 (score 16 corresponds to expected mortality rate of 38%).in this protocol the surgeons report the judgement of the efficacy of surgical procedure to remove or not the focus of infection. objectives: infections and subsequent septic responses remain the leading cause of death among surgical intensive care (sicu) patients despite tmprovetaunts in supportive care and brond-epectrum antibiotics. usually invading bacteria are efficiently cleared by neutrophil granulocytes. however, during sepsis various neatrophil dysfunctions have been demonstrated, leading to impaired host defense. granulocyte colony-stimulating factor (g-csf) induces a sustained increase in circulating neutrophils and enhances various noutrophil functions. it was the purpose of the present study, to evaluate the safety and efficacy of g-csf (filgrastim) in sicu patients at risk of sepsis. materiel a.d methods: the study was designed as an open-label phase-ll study of filgrastim. ten consecutive slcu patients, with a therapeutic interveotion score greater than 30, were included in the study. filgrastim was given by daily continuous intravenous infusion for 7 days or discharge from the sicu. apache ll-score, multiple-organ-failure (mof) score, definitions of infections, sepsis, systemic inflammatory response syndrome (sirs), and acute respiratory failure were applied daily. a response to filgrastinl th_erapy was defined as an improvement in disease severity quantified by a decrease of > 4 apache 1i score points on day 4 after onset of treatment. results: none of the 10 patients developed a sepsis or mof later on and no patient died during hospitalization. specific postoperative complications occured in one patient ~jth a leekage of the oesophagou-gastric anastomosis after oesophageus resection. at study entry the leucocytes amounted to 10.110 + 2.000/~tl (mean + sem) and reached a level of 29.280 +_ 5.810/tal at day 6 after onset offilgrastim therapy. the apache ii score initally was 14 + 1.57 (mean + sem) and as an indicator of filgrastim response a decrease of 5 points ~dthin 4 days oceured in 7 out ot 10 patients. filgrastim was well tolerated, side effects were not noted. growth of solid tumors might be modulated by the activity of inflammatory and/or immune effector cells of undefined specificity. in this study patients undergoing surgical treatment for gastric (n= 41) or colorectal (n= 47) cancers were evaluated for endogenous serum levels of granulocyte colony-stimulatingfactor (g-csf) during a pre-and postoperative time period. from the same blood specimens mononuelcar cells (mnc) were prepared. the release of ifn-%, and il-2, which are secreted by thl cells, were stimulated in vitro by pha during a cell culture period up to 48 hours. the patients were further classified for their immunreactivity by responses in dth skin testing to seven different antigens (e.g. tetanus toxoid, ppd, diphtheria toxin, trichophyton, streptococcus, candida and proteus antigens). dth testing has been repeated in each patient two remarkable results were obtained. the serum levels of endogenous g-cse showed a biphasic increase with maximum values of 900 pg/ml (preoperative < 70 pg/ml) on day 1 and day 3 to 5 after surgical treatment. similar patterns of g-csf production were found in both groups of patients with gastric or colorectal cancers. high serum levels of g-csf were significantly (p < 0,01) correlated with infectious complications in patients whh gastric cancer (n= 17/22). secondly patients could be arranged into two groups according to an anergic (n= 25) or normergi¢ (n = 63) responsiveness in dth testing. the frequency of anergi¢ responsiveness was similar in both patients with gastric (n= 11/41) or colorectal (n= 14/47) cancers. interestingly we found a significant correlation (p < 0,01) between low serum levels of g-csf and anergy during the postoperative period in both groups. stimulation of mncs from anergic patients (n= 30) within the pre-and postoperative period resulted in reduced mean values (about 50%) for ifn-ff release (preoperative means llo0 pg/nfl), if compared to patients with normergic dth (n= 63, preoperative means 1960 pg/ml). similar, but less significant results were obtained for il-2 secretion. our results confirm a correlation between infectious complications and g-csf in the postoperative period, however elevated levels were also found in some patients without any signs of infections. more interestingly there might be an association between cytokine (c~csf, ifn-% and il-2) release and dth, which is known to be mediated by activated thl calls. to recognize anergic dth as a possible higher risk in the postoperative outcome of cancer patients extended periods of observation are needed. objectives of the study effects of recombinant huraan granulocyte colony-stimulating factor(rhc-csf)a galnst severe septic infections were investigated by its single use or by its corn b{nation with cephera antibiotlcs.we examined its effects on the mortality,and circulating blood neutrophyis counts and functlons,such as phagocytic activity and h 2 02 production using the rat severe septic model. rats were subcutaneously administsrd rhc~csf(s0orl0o ~ g/k~ body wt)after on set of peritonitis brought about by cecal ]igation and one puncture withe2-gaug e needle once a day for three days.in addjtlon,cefmetazol na(cmz)(50m$/k9 bo dy wt)was injected intrarnustularly to the rats tv~ce a day for three days. cirehlatlng blood neutrophyls counts were determoned electronically with a hem ocytometer,and blood smears stained with may~runwaldm.qlemsa~taln. neutrophyls functions in vltro,such as phagocytic activity and h 20 2 producti on using the rat severe septic model was analyzvd by automated flow cytometri c single cell-analysis methods. the reortallty rate after 2 weeks was significantly decreased by administratlon of rh~-csf(p<0,01).ln addjtion,a combination therapy of rhg-csf wlte cephern ant~biotics(cmz)showed a significantly survive] advantage and the rate had b een reached 57.1%. nextly,treatn%ent wlth rhg-csf(s0 ~ $/k9 body wt)increased the nuzaber of the peripheral blood neutrophjls slgn[fieantly(p<0.01). iv~oreover,functions of neutrophlis which were phagocytic activity and h2 02 p roduction were remarkably enhanced by admlnlstratlon of rhg-cs~(50 ~ 9/ks b ody wt) (p<0.(15). these findings suggest that combination therapy of rhcrcsf with cephern antib iotlcs(cmz)is an efficient regime against severe infectlons.and the increased ne utrophils counts and enhanced neutrophiis functions were played a important ro le about the survival advantage. granulocyte macrophage colony-stimulating factor (gm-csf) is a haematopoietic growth factor active on neutrophils and macrophages. leukopenia often occurs following renal transplantation and can be associated with infection and/or the myelosuppressive effect of azathioprine. aim: we report the use of gm-csf in 17 renal allograft recipients with leukopenia. nonglycosylated recombinant gm-csf was obtained from e. coli transvected by human gm-csf gene. m~terial ~,nd methods : written informed consent was obtained from all patients. patients were suffering from toxic neutropenia (neutrophils < 500/mm 3) with medullar hypocellularity on bone marrow aspiration, or leukopenia (neutrophils < 1000/ram 3) with cytomegalovirus infection requiring ganciclovir administtation. gm-csf was given subcutaneously at a dally dose of 2 to 5 mcg/kg/day, according to renal function. results : in all cases, neutrophil counts returned to normal levels within 1 to 4 days. in most of them, spectacular correction was observed within 24 hours, with a single injection. adverse events due to gm-csf at this dose were mild and easily managed (2 cases of bone pain treated with paracetamol). one acute rejection episode was observed after correction of leukopenia. conclusion : on the basis of this study, it appears that gm-csf at a dose below 5 mcg/kg/day is an effective treatment for renal transplant recipients with leukopenia associated with cmv infection or toxic neutropenia. department of nephrology, 161, rue de s~vres, hopital necker, 75743 paris, france. changes in serum g-csf and il-6 after surgical intervention hitoshi toda 1, atsuo murata 1, hidewaki nakagawa 1 , takesada mori 1, nariaki matsuura 2 1osaka university medical school, osaka, 2wakayama medical school, wakayama, japan we measured serum immunoreactive interleukin 6 (il-6) and granulocyte colony-stimulating factor (g-csf) levels of the patients undergoing major thoraco-abdominal surgery for esophageal cancer. serum samples were collected from eight patients on the day before surgery, at the time of operation, and thereafter at suitable intervals for one week. il-6 and g-csf were measured by means of enzyme linked immunoassay. the normal range of serum ]l-6 was less than 2 pg/ml and g-csf less than 4 pg/ml. values between groups were compared with linear regression analysis. both serum g-csf and il-6 levels reached their maximal levels at the first postoperative day and decreased thereafter. the correlation between g-csf (y) and il-6 (x) was y=3.28x+6.16 (r=0.786, n=86, p<0.01 ), showing a significant correlation. in the case who suffered from aspiration pneumonia and ards at the second postoperative day, the peak level of il-6 was 50 pg/ml and g-csf 69 pg/ml respectively. the estimated value of g-csf was 171 pg/mi by the regression equation. this means the real g-cse level was less than half of the estimated value. it suggests that low responsiveness of g-csf is one of the reason of immunodeficient state after the major surgery, neutrophils from injured patients ingest and kill bacteria less efficiently as compared to those of healthy individuals, probably reflecting the suppression in respiratoly burst which occurs after severe trauma. one of the main mechanisms of killing bacteria by neutrophil granulocytes is production of oxygen radicals (respiratory burst). granulocyte colony-stimulating factor (g-csf), a 20kilodalton cytokine, leads to a sustained, dose-dependent increase in circulating neutrophils. thus, it was investigated whether filgrastim (recombinant human granulocyte colony-stimulating factor, rhg-csf) therapy fits for prophylaxis of sepsis in postoperative/posttraumatic patients, and whether, besides an expected increase in neutrophil count, filgrastim would also augment neutrophil function. material and methods: this study was designed as an open label, prospective phase ii study of filgrastim and performed in a surgical intensive care unit (sicu) (university hospital). 10 postoperative/post-traumatic patients with a therapeutic intervention scoring system (tiss) score greater than 30 were treated with filgrastim (0.5 -1 l.tg/kg/day) for prophylaxis of sepsis on 7 days or until discharge from the sicu. production of oxygen radicals can be quantified by analysis of fmlp-and zymosan-induced chemiluminescence. neutrophil oxygen radical production was tested by fmlp-and zymosan-induced chemiluminescence by the polymorphonuclear cells (pmn) of these patients in multiple blood samples over a period of up to 14 days. results: none of the patients treated with filgrastim for prophylaxis of sepsis developed sepsis. in vitro fmlp-induced (10 -7 reel/l) neutrophil oxygen radical production was significantly increased under therapy with filgrastim by a maximum of 797% +-570% (217% -1826%) compared to pretreatment values of 100%. tapering of filgrastim resulted in a reduction of fmlp-induced neutrophil oxygen radical production within 48 hours. in contrast, zymosan-induced neutrophil oxygen radical production was not affected by filgrastim treatment. conclusions: besides its quantitative effect on neutrophil counts enhanced neutrophil function, documented here as increased fmlp-induced oxygen radical production, may account for the beneficial effect of filgrastim for prophylaxis of sepsis in posttraumatic/post-operative patients. granulocyte colony stimulating factor (g-csf) and granulocytemacrophage colony stimulating factor (gm-csf) have been recently introduced in the treatment of chemotherapy-induced neutropenia. effects of these csfs on cellular immune system were evaluated in 38 neutropenic gynecological cancer patients during chemotherapy. g-csf and gm-csf were equally able to induce a rapid recovery of white cell count within one or two days. g-csf treatment resulted in a significantly higher concentration of leukocytes measured in the peripheral blood although by gm-csf a sufficient effect was achieved (p<0.05). before initiation of csf treatment urinary neopterin was similar in both groups of patients (283+/-38 and 267+/-40 lamol/mol creatinine for gm-csf and g-csf respectively expressed as mean +/-one sd). in g-csf treated patient only a marginal induction of neopterin was observed. on day 4 the mean value was about 40% above the basal level (p<0.05). on the other hand gm-csf treated patients were characterized by a pronounced increase in urinary neopterin levels. in comparison with the basal level a more than 2 fold induction was noted and the difference between g-csf and gm-csf was highly significant (p<0.01). this effect was confirmed in vitro by investigating the effects of these csfs on interferon-gamma mediated pathways in thp-1 human myelomonocytic cells. results suggest activation of immune effector cells by gm-csf which may help the organism to overcome infections. however, activated macrophages produce several growth factors which may increase malignant proliferation, and augmented neopterin production as sign of macrophage activation has also been associated with poor prognosis m several malignancies. more data are therefore necessary to clarify whether csf mediated immune activation is beneficial or deleterious for cancer patients but considering our results caution in applying csfs in oncology seems advised. from a historical perspective, the development of humoral immunity to bacterial endotoxin has assumed a prominent position in the spectrum of therapeutic approaches which have been explored for the treatment of gram negative septic shock. predicated upon the fact that rough strains of bacteria manifest lps containing exclusively conserved structural features common to lps from all gram negatives, specific antibodies were elicited which conveyed cross protective immunity in experimental models of bacteremia and endotoxemia. such studies culminated in a well-conducted, randomized, double-blind placebo-controlled clinical trial using passively administered human polyclonal antiserum to treat patients with suspected gram negative sepsis. the efficacy of treatment established in that trial spurred efforts to develop monoclonai reagents which, to date, have not been uniformly successful in reproducing those earlier studies with polyclonai antibodies. nevertheless, the numerous successes which have been documented in experimental models of endotoxemia continue to foster promise for this immunotherapeutie approach. several recent studies with human polyclonalimrnunoglobulin preparations containing antibodies reactive with lps and lipid a have yielded promising results in treatment of patients with sepsis. in addition, the recent development of an antiidiotypic monoclonal antibody which reflects an internal image of a kdo specific monoclonal antibody has provided an alternative experimental approach to generate anti-lps antibody. immunization of mice with the antiidiotype provides significant protection against subsequent lps lethality consistent with the development of circulating immunoglobulin specific for lps. thus, the use of polyclonal immunoglobulins contrives to provide an alternative and potentially cost effective method for the treatment of endotoxin shock. supported by r37 a123447 and pot ca54474. john holaday, anne fortier, shawn green, glenn swartz, john madsen, carol naey, and jan dijkstra entremed, inc.. rockville, md, 20850. at the time of diagnosis, the signs and symptoms of septic shock are an indication that the systemic inflammatory response is well underway; thus, it has been argued that the endotoxin "cat is out of the bag", and that subsequent passive immunization may be too late to achieve therapeutic benefit. our approach has been to evaluate active immunization as a prophylax~s against sepsis. mice were inoculated twice (two weeks apart) with liposomes containing dmpc[i.8], dmpg[0.2], cholesterol [1.5] , and monophosphoryl lipid a [20-200 gg/txmole phospholipid] by several routes (i.p., i.m.), and serum was collected 10-11 days after each inoculation. after a single injection, highest tilers of ab were produced in mice inoculated i.p., but mice inoculated by all routes produced anti-lipid a ab. following the second injection. ab levels were roughly equivalent in mice inoculated by all routes, regardless of lipid a concentration. mice vaccinated i.p. with liposomes containing 0, 20 or 200 gg lipid a were treated with cyclophosphamide to produce neutroperda and then challenged with e. cole in an infection model of gram negative sepsis. the lds0 for control (liposomes with no lipid a) mice was 3x10 bacteria; ld50 for mice vaccinated with 20p.g was 24x103 (8-fold increase in resistance) and with 200~tg was 48x103 bacteria (16-laid increase in resistance). mice vaccinated as before were also treated with actinomyein d to increase sensitivity to lps (salmonella minnesota) challenge in an endotoxemia model of grain negative sepsis. the ld50 for control (liposomes with no lipid a) mice was 30 ng lps; the ld50 for 20gg lipid a was 70rig lps (2-fold increase in resistance) and for 2001xg was 570ng lps (19-fold increase in resistance). mice were similarly vaccinated and challenged with an aggressive gram negative pathogen, francfsella tularensis. the ld50 of franciseua in normal mice or mice inoculated with liposomes without lipid a was 20-40 bacteria. in contrast, mice vaccinated with liposomal lipid a (200ggl survived challenges as high as 30,000 bacteria, (3 logs of protection). the impressive protective capacity of this vaccine did not correlate with ab liter in any of the sepsis models, nor did it correlate with classic nonspeeific events, such as macrophage activation. maerophages harvested from the peritoneum of mice vaccinated and protected against sequelae of gram negative infections did not spontaneously kill the bacteria in vitro, but could be activated by ifn-y for antimicrobial activity equivalent to that of macrophages from unt#eated mice. research is underway to defme the protective mechanism(s) activated by this liposomal-lipid a vaccine. intervention by monophosphoryl lipid a in septic shock jon a. rudbach, ribi immunochem research, inc., hamilton, montana, usa monophosphoryl lipid a (mla), the clinical form of which is called mpl®-immunostimulant, has been tested extensively as an intervenient material in septic shock. mla is protective when given to experimental animals prior to a live microbial challenge or challenge with lethal doses of microbial products or certain cytokines. this is shown with gram negative and gram positive bacteria, gram negative bacterial endotoxins, and gram positive bacterial exotoxins. furthermore, animals treated with a regimen of mla which results in a refractory state to a lethal dose of gram negative bacterial endotoxin concomitantly display increased resistance to a live bacterial challenge. thus, both endotoxin tolerance and nonspeciflc resistance to infection can be manifested simultaneously. also, prophylactic doses of mla do not interfere with other therapies given subsequently; an additive or a synergistic protective effect can be demonstrated with certain combinatorial treatment regimens, such as mla followed by antiendotoxin monoclonal antibodies. the preclinical studies were extended to human trials wherein the safety of agonistic doses of mla was verified. furthermore, when mla was administered to human volunteers 24 hr before challenge with a pharmacologically active dose of reference endotoxin, febrile, cardiac, tnf, il-6, and il-8 responses were all decreased significantly as compared with the responses of subjects pretreated with a control solution and challenged with endotoxin. human trials with mla are being extended into patient cohorts which have high probabilities of developing septic shock; this will expand the safety base and establish clinical efficacy for mpl®-immunostimulant. a considerable body of in vitro evidence supports the concept that the effects of lps on cells of the immune/inflammatory systems are controlled by interactions of lps with cd14. to evaluate if blocking lps-cd14 interactions has potential as a therapeutic in septic shock we have evaluated the effect of anti-cdi4 monoclonal antibody (mab) on lps-induced cytokine production and physiologic changes in an experimental model of endotoxin shock performed in cynomolgus monkeys. a novel model has been established where animals were treated with interferongamma for three days prior to infusion of highly purified lps over an eight hour period. in this model lps challenge resulted in marked release of eytokines in the blood, substantial hemodynamic changes, release of liver enzymes and alteration in lung permeability observed over a 24 hour period. to evaluate the effect of treatment with anti-cd14 mab, animals were given either nothing, an isotype control or anti-cd14 mab (5mg/kg) 30 rains, prior to the beginning of the lps infusion. evaluation of physiologic changes including mean arterial blood pressure and cardiac output, quantitative analysis of eytoldne levels including tnfct, il-113,1i,-6, il-8 and il-10, and liver enzymes during a 24 hour period revealed that treatment with anti-cd14 mab markedly attenuated all parameters of injury including decreased mean arterial blood pressure, increased cytnkine levels and the release of liver enzymes observed in animals given the isotype control mab or those not treated. administration of anti-cd14 mab to interferon-gamma treated animals not challenged with lps did not induce any detectable physiologic changes or increases in cytoldnes. these studies suggest that strategies to block lps-cd14 interactions will have utility in diseases such as septic shock or ards where lps plays a central role in initiating injury. preclinical studies with recombinant bactericidal/permeability increasing proteins (rbpi and rbpi23). p.w. "frown, dept. of preclinical science, xoma corporation, berkeley, california, usa. bactericidal/permeability increasing protein (bpi), from neutrophils, binds to and neutralizes lipopolysaccharide (lps); it also specifically kills gram-negative bacteria (gnb). these properties, which reside in the n-terminal half of the molecule, indicate potential therapeutic application in the treatment of gram-negative sepsis. the gene for human bpi has been cloned and recombinant holoprotein (rbpi) and a 23 kd n-terminal fragment (rbpi;3) have been produced in sufficient quantities for preclinical studies. both rbpi and rbpi23 bind to lipid a and neutralize the biological activities of lps derived from a variety of organisms, rbpi23 has equivalent antibacterial activity to bpi against rough gnb but is up to 30x more potent than bpi vs. serum-resistant and smooth gnb. rbpi and rbpi23 compete with lps-binding protein (lbp) for binding to lps under physiological conditions. consequently, both rbpi and rbpi23 block the cd14-dependent lpsinduced synthesis of the cytokines tnf, il-1, el-6 and il-8 in vitro. rbpi23 has also been shown to inhibit the lps-induced synthesis of reactive 02 metabolites, endothelial adhesion molecules and the procoagulant molecule tissue factor. in animals, rbpi has been reported to increase survival of endotoxin-challenged rats and mice, to inhibit the dermal schwartzman reaction in rabbits and to increase survival of neutropenic rats with pseudomonas bacteremia, rbpi23 increases survival and decreases cytokine production in endotoxin challenged mice and rats. it normalizes lps-induced changes in hemodynamic, pulmonary and/or metabolic parameters in lps-induced rats, rabbits and pigs. treatment with rbpi23 also increases survival and decreases cytokine production in bacterial challenge models in rats and mice. rbpi23 was not toxic to rats after 10 daily consecutive i.v. doses of 10 mg/kg. this combination of properties indicate that recombinant bpi may be useful in the treatment of sepsis. phase i/ii clinical trials of rbpi23 have begun. the discovery of lps binding protein (lbp) and subsequent identification of cd14 as a receptor for lps or lps-lbp complexes has resulted in a new understanding o£ how lps responsive ceils are stimulated. cd14 is found either as a glycosylphosphatidyl-inositol (gpi)-anehored membrane glycoprotein (mcd14) of myeloid cells or as a soluble serum protein (scd14) lacking the gpi-anchor. binding of lps to mcd 14 triggers cell activation while binding of lps-scd14 complexes to cells such as endothelial or epithelial cells that normally do not express mcd14 activates these cells. these pathways are shown in schematic form below. 4 ~di mcd14 plays a crucial role in presentation of lps to additional membrane components that make up a functional lps receptor. an immediate consequence of engagement of this functional receptor is protein tyrosine phosphorylation. the molecular mechanisms leading to these events will be discussed. understanding of these pathways will lead to the development of new therapeutic approaches to controlling host responses to lps. pretreatmen t posttreatment (before or after tnf peak) d) with different antibody dosages: 15 mg/kg ---0.1 mg/kg pretreatment with anti-tnfab prevented death in most model situations (except peritonitis), but also posttreatment up to 4h after sepsis induction was successful in the few studies performed. there is additional evidence that low-dose tnfab is partially effective. especially baboon anti-tnfab studies provided many insights into the pathophysiological sequences of sepsis induction, due to crossreactivity with human reagents. those events include the cytokine sequence with tnf-dependent il-i, il-6, or il-8, but also il-lra or stnf receptor release. granulocyte as well as endothelial cell activation were shown to be partly tnf related, and the procoagulatory response was influenced by anti-tnf treatment. from many animal studies the concept that tnf plays a pivotal role in sepsis is clearly evident and therefore anti-tnf therapy is a major candidate tbr clinical studies. the beneficial or harmful effects of tnf-mediated inflammatory responses depend on the clinical context. decreasing exaggerated tnf-mediated inflammatory responses may be useful in some patients with organ failure. tnfr:fc (immunex, seattle, wa) is a recombinant human protein composed of two identical extracellular p80 tnf receptors linked by the fc region of iggl. it neutralizes tnf with an affinity for tnf<z of 10-10 m and has a t1/2 of • 63h in normal humans. methods: 18 normal volunteers were randomized to receive either tnfr:fc vehicle (n=6), or low dose (ld) tnfr:fc (10 mg/m 2 iv, n=6), or high dose (hd) tnfr:fc (60 mg/m 2 iv, n=6) infused over 30 rain prior to endotoxin (e) (escherichia coil 0:113; 4 ng/kg iv) administration. plasma cytokines were measured using elisa and tnf bioactiv'dy. systemic hemodynamics were evaluated with indwelling arterial and pulmonary artery catheters and radionuclide heart scans and compared before and after fluid loading in subjects given vehicle with hd tnfr:fc. results: endotoxin-related symptoms were unchanged after ld or hd. peak temperature occurred at a later time point (4-5h) in subjects given tnfr:fc compared to vehicle (3h) (p=.004). at 3h, 5h (post fluid load ), and 8h, subjects given vehicle developed increased cardiac index and heart rate and decreased systemic vascular resistance index (all p=.0001 ). hd tnfr:fc did not alter this hyperdynamic cardiovascular response. ld and hd inhibited the e-induced teukopenta at i h post endotoxin (p<.03) and later leukocytosis (p<0.05). peak tnf bioactivity in subjects given vehicle was 154 + 44 pg/ml and <10 pg/ml in the ld or hd tnfr:fc groups (p<.001). two immunoassays for tnf were performed because the detection of receptor bound tnf varies with elisa. tnf (biosource) was decreased in ld vs vehicle or hd (p=.0001) whereas tnf (r&d systems) was increased after hd or ld vs vehicle (p<.001). decreased levels of il-8 (p=.00t), granulocyte colony stimulating factor (p=.03), and il-1 receptor antagonist (p=.001) were found after ld or hd vs vehicle. il-6 levels were lower after ld vs vehicle or hd (p=.006). il-10 levels were similar among subjects given vehicle, ld, and hd tnfr:fc. conclusions: ld and hd tnfr:fc delayed the febrile response to e but did not alter the early hyperdynamic cardiac response. this suggests that mediators other than tnf play an important role in the initial cardiovascular response to e in humans. tnfr:fc attenuated the release of some e and tnf-induced cytokines. ti~e antiinflammatory responses of tnfr:fc may be useful in some patients with disease processes resulting from excessive tnf-mediated inflammatory responses. the biosynthesis of tnf within macrophages is regulated both at the transcriptional and the translational levels. the 5' flanking region of the tnf gene contains several transcriptional control elements, including two kb enhancers similar to those of immunoglobulin genes and a "y box" similar to that of the mhc class ii promoter. these elements are critical for the enhancement of transcription noted after stimulation of macrophages with lps. despite these and other elements, transcriptional regulation alone accounts neither for the absence of tnf protein during normal homeostasis nor the profound increasein tnf production following stimulation with lps or other secretagogues. the translation of tnf mrna into protein is also tightly regulated via mechanisms involving the octameric "ttatttat" motif present in the 3'untranslated region of tnf, human inducible no synthase, and several other cytokine genes. this region not only destabilizes mrna, but also confers a translational blockade so that tnf protein is not normally synthesized despite leaky basal transcription. lps stimulation releases translational blockade and, together with enhanced trancriptional rate, accounts for significantly increased tnf secretion. opportunities for inhibition of tnf production are available at each of these two levels. agents which increase intracellular camp (pentoxif¥1ine, amrinone) inhibit accumulation of tnf mrna; in contrast, corticosteroids act primarily at the level of translation, inhibiting lps induced translational activation. understanding the regulation of tnf biosynthesis may assist in clinical strategies designed to regulate tnf secretion. the serine protease thrombin is formed by enzymatic conversion from its proenzyme form after coagulation activation and may enhance the inflammatory response in various ways. in the last step of the coagulation cascade, thrombin cleaves its substrate fibrinogen, thus forming fibrin. the fibrin clot may prevent phagocytosis of bacteria and promote local bacterial growth. thrombin can also stimulate a specific th.rombin receptor that exists on a variety of cells. for example, minute amounts of thrombin ma), stimulate platelets to develop tissue factor and boost the activation of more thrombin, thus initiating a positive feedback loop. other cellular effects of thrombin receptor stimulation include increase in intracellular calcium of platelets and monocytes, contraction of smooth muscle cells, increase in endothelial permeability, thromboxane generation by neutrophils and lymphocytes, pulmonary vasoconstriction, and enhanced cytokine production. animal experiments have been conducted with a variety of thrombin inhibitors including heparin, antithrombin ili, hiradin, and hirudin-like peptides. an alternative approach is the inhibition of the coagulation cascade at an earlier point in the cascades, or administration of the anticoagulant enzyme protein c work from our own group in a porcine model of lipopolysaccharide-(lps)-induced shock with disseminated intravascular coagulation has shown that prelreatment with recombinant desulfatohirudin (r-h) or with a preformed complex of human donor antithrombin iii with heparin or post treatment with r-h are able to block the degradation of fibrinogen and the formation of fibrin monomer. in addition, after pretreatment with r-h it was found that lps-induced lung injury was reduced. since both natural hirudin as well as r-h were well tolerated by healthy volunteers, adminislration of r-h may be a promising therapeutic approach in patients with sepsis and sirs. corticosteroids have been used in the management of various shock syndroms including sepsis. the data concerning the effects of this therapy is, however, conflicting and in large prospective studies, the use of early administration of high doses of corticosteroids have been found to give no beneficial effects in patients with sepsis. we have particularly been engaged in studies on possible effects of corticosteroids on the plasma cascade systems and on the mediator release from leukocytes. in in vitro studies, high doses of methylprednisolone were found to facilitate endotoxin induced generation of plasma kallikrein. furthermore, plasma prekallikrein and kallikrein inhibitor values decreased together with formation of kallikrein-c1 inhibitor complexes (1). in this in vitro plasma model we also found that methylpredoisolone alone in doses of 1 mg/ml induced contact activation with the formation of kallikrein-c1 inhibitor complexes. methylprednisolone was also found to have an additive effect on endotoxine induced decreases in kallikrein inhibitor functions. in the same experimental model on endotoxemia methylprednisoione was found to give an additive effect on activation of the initial part of the complement cascade (2) . activation of the terminal part of complement, however, was inhibited by the same dose of methylprednisolone. addition of methylprednisolone alone to plasma was also found to activate the initial part of complement. using a full blood model, we studied the effect of methylpredoisolone in modulating the endotoxin induced cytokine response. when 103 ng/l e. coli endotoxin was added to citrated blood from healthy human donors preinkubation with 20 gg/ml methylprednisolone significantly reduced the release of tumor necrosis factor et (79 %) and interleukin-6 (63 % ) at two hours after exposure to endotoxin. in conclusion, these studies show that corticosteroids might facilitate contact activation of plasma and reduce the function of major protease inhibitors. furthermore, we also observed that this drag in in vitro conditions facilitated activation of complement. on the other hand, corticosteroids were found to reduce endotoxin induced cytokine release from leukocytes in whole blood. these studies also showed a dose dependence of the effects of corticosteroids on endotoxin induced cellular and cascade system activation. different predisposing conditions like extensive trauma, infection or pancreatitis result in a remarkably similar inflammatory response. although thls inflammatory response primarily aims at the removal of devitalized tissues, destruction of bacteria and reestablishment of the integrity of host tissues, it may potentially become unregulated and generalized and thereby producing deleterious effects to vital organs or the body" as a whole. cytokines play an important role in the development of the systemic inflammatory response. most of their biological effects, however, are not directly mediated but exerted through other mediator systems. when the inflammatory response results in the formation of capillary leak syndrome and refractory hypotension, the unregulated activation of complement and contact systems appears to be the cause. both systems are almost exclusively regulated by c1-1nhibitor. am unbalanced degradation of the inhibitor protein was found to be associated with the onset of capillary leakage in various conditions such as bone marrow transplantation, severe burns, and septic shock. these observations suggest a relative deficiency of biologically active c3-1nhibitor in these septic shock like syndromes. therapeutic intervention studies were initiated using high doses of c1-1nhibitor concentrate. first results are promising, as in most patients the administration of the drug was followed by a quick and marked improvement of the patient's hemodynamics. whether the administration of c1-1nhibitor alone or in combination with other drugs is also sufficient to improve long-time survival has to be investigated in double-blind, placebocontrolled studies. salutary effect of human soluble complement receptor type-1 in models of adult respiratory distress syndrome g. feuerstein, m. dimartino, and *r. rabinovici, smithkline beecham pharmaceuticals, king of prussia, pa, usa, and *jefferson medical college, philadelphia, pa, usa adult respiratory distress syndrome (ards) is a severe pulmonary insufficiency syndrome associated with diverse diseases and injuries. ards is believed to be the consequence of a massive acute inflammatory reaction consisting of activated neutrophils infiltration into the lung. neutrophil activation is likely the result of multiple factors of which complement is believed to play a major role. a potent complement regulatory system in the form of the complement receptor type-1 (cr-1) exists on many cells where protection against complement injury is provided by inhibition of both the alternative and classical pathways for c3/c5 convertase formation. we have recently purified the human recombinant cr-1 in a truncated form (brl55730, tp-io, scr-1) and tested this soluble protein for protective effects in several models of ards. specifically, we have used the following rat models: 1) endotoxin-induced ards in paf primed animals; 2) 11.-2induced ards; 3) thermal injury (30% body surface)-induced ards; 4) acid aspiration-induced ards. ards-like features included edema (increase in lung wet weight and water content), neutrophil accumulation (histological inspection and myeloperoxidase assay) and increase in cells and protein in the bronchoalveolar lavage (bad. scr-1, 1-25 mg/kg, i,v., given as prophylactic (and in some cases, therapeutic) treatment significantly inhibited edema formation and leukocyte infiltration and, in some cases (e.g., endotoxin/paf or il-2), virtually completely. these comprehensive studies strongly support the therapeutic potential of scr-1 in ards due to diverse injuries. the xanthine derivative pentoxifylline (ptx) and several analogs of ptx have been evaluated for their protective effect in various animal models of septic shock. data from these in vivo studies demonstrate that (1) ptx suppresses lps-induced tnf release from activated macrophages; (2) ptx prevents tnf-induced pmn activation; (3) ptx attenuates lps-or tnfinduced acute lung injury as evidenced by prevention of increased pulmonary vascular permeability and detoriation of gas exchange; (4) ptx exerts its .protective effects even when administered following initiation of sepsis. the sum of these actions suggest that administration of ptx in sepsis may have therapeutic potential. hans hoffmann md, dept. of surgery, klinikum grosshadern, ludwig-maximilians-universit~.t, marchioninjstrasse 15, d-81377 m0nchen, germany. pentoxifylline [pof] and related xanthins are drugs of known haemorrheologieal activity and widely used clinically. recently, new pharmacological aspects of these established drugs could be demonstrated. it was found that pof selectively inhibits the formation of tnf but not il-6 most likely by causing accumulation of intraceliular camp via inhibiting phosphodiestersse activity, and, secondly, by inducing prostacyclin in different cell types. furthermore, pof is able to counteract tnf-mediated adherence of neutrophils to vascular endothelium and to inhibit fmlp-indueed respiratory burst in neutrophils. we and others have, therefore, studied its effect in different animal models. it was found that pof protected from increased pulmonary vascular permeability and sequestration of neutrophils into the lung in different animal models of acute lung injury. moreover, in pigs with induced faecal peritonitis the drug improved haemodyrmmle variables as well as pulmonary compliance and reduced the number of pulmonary neutrophila and lymphocytes. consequently, as a general outcome, pof could be found to improve survival in different models of haemorrhagie and endotoxie shock. the protective effect of pof was dose-dependent and observed even when pof was given up to 4 hours after endotoxin. in order to evaluate these pharmacological effects of pof in humans, we studied pof under controlled conditions of endotoxlnemia in volunteers. we found that pof selectively inhibits the lps-indueed endogenous formation of tnf without affecting il-6 and il-8. moreover, pof counteracts the lps-indueed initial leukocytopenia by interfering with the adherence of neutrophils in the microvasculature. meanwhile the inhibition of endogenous formation of tnf by pof could be demonstrated under different clinical conditions of acute and chronic cytokine release syndromes, {okt3 first-dose reaction, cancer-or tuberculosis-induced cachexla}. it appears worthwile and promising to investigate wether pof exhibits beneficial effects also in septic patients. objectives: to determine the specific role of paf in both lethal primate bacteremia and nonlethal human endotoxemia. materials and methods: two double-blinded placebo controlled studies were performed. first, ten baboons (14.5 +.7 kg), were randomized to a control group receiving 101° cfu/kg of intravenously administered e. co/i (n=5) and a treatment group (n =5), pretreated with .1 mg/kg of paf receptor antagonist ro24-4736 two hours prior to e. co/i infusion. in a second study, ten healthy male volunteers were randomized to a control group receiving 4 ng/kg of intravenously administered endotoxin (lps) (n=5) and a treatment group (n=5) pretreated with 10 mg of ro24-4736 eighteen hours prior to lps administration. physiologic parameters and cytokine levels were measured continuously in both studies for 24 hours. results: baboons pretreated with the paf antagonist had improved oxygenation (96 + 11 mm hg v 59 -+ 6 in controls, p < 0.05) and creatinine clearance 12 hours post e.coli (39.8 + 23.4 ml/min v 0.1 _+ 0.1 in controls, p < 0.05). similarly, volunteers pretreated with the paf antagonist experienced fewer symptoms, including rigors and myalgias at 1 hour post lps. this was in concert with a diminished peak cortisol (668 +_ 107 mmot/l v 959 +-159 mmol/l in controls, p < 0.05), epinephrine secretion (1057 + 165 nmol/l v 2029 + 431 nmol/l in controls, p < 0.05). hemodynamics and circulating tnfa levels (data not shown) were unaffected in either study by prior paf antagonism. concluslons: paf influences some components of the multi-organ failure syndrome in lethal gram negative sepsis and the counter-regulatory hormonal system in human endotoxemia through tnf-independent mechanisms. paf antagonists may serve as adjuncts to cytokine blockade in the treatment of gram negative sepsis. the mediator role of platelet-activating factor in gramnegative septic shock pierre g. braquet, david hosford and matyas koltai institut henri beaufour, le plessis robinson, france considerable evidence has been accumulated to support the concept that a paf/cytokine autogenerated feedback network is responsible for priming and amplification of inflammatory mediator release in septic shock. cytokines, such as tnf~x, il-1 and other interleukins interact with paf which then amplifies cytokine production, therefore, paf may be the principle mediator in endotoxin shock. a single factor identified as tnf is suggested to play a pivotal role in the fatal phase of septic shock. presumably excess tnf release is the result of amplification process primarily triggered by paf. one may assume that the high tnf concentration in the blood of 'non survivors' is the consequence rather than the cause of cell death. the dubious results of clinical trials with anti-tnf antibodies and il-1ra, a naturally occurring il-1 antagonist protein, have risen debate around the exceptional role of cytokines in the pathomechanism of septic shock or systemic inflammatory response syndrome (sirs) induced by gramnegative microorganisms. there are similar problems with the interpretation of the results obtained in clinical trials of monoclonal igm antibodies against e. coli endotoxin, ha-1a and e5. future studies will answer the question as to the effectivity of inefficiency of this treatment. perhaps when the plasma concentrations of lps and cytokines exceed a critical level, treatment fails to save the patients life. with regard the causal role of lps in septic shock, the putative role of bacterial porins may have to be taken into consideration. the marked release of paf induced by endotoxin leading to excess txa2 production may be responsible for the microvascular failure and death in septic shock. antagonists of paf markedly protect against the release of txa2, and may interrupt the vicious circle of amplification process. no produced by the inducible no synthase plays a pivotal role in causing vascular paralysis in lps-induced sirs. to explore the relationship of paf to no synthesis will provide better understanding of the pathomechanism of septic shock. several paf antagonists, including bn 52021, have undergone phase ii and phase iii clinical trials. the strategy in the treatment of sirs, however, remains multifactorial, since little is known about the sirs caused by microorganisms other than gram-negative bacteria. evidence has begun to accumulate which suggests that in sepsis excess production of lps and cytokines can lead to uncontrolled production of inos and that this might in part explain the severe unresponsive hypotension seen in some septic patients. a number of strategies have been explored in an attempt to limit excess no production. a favourite target has been competitive inhibition of nos by arginine analogues such as l-nmma. in animals, some investigators have shown that the haemodynamic effects of lps challenge can be attenuated by l-nmma, but there is a narrow toxic-therapeutic ratio. in a model of live e. cell sepsis, we have been unable to reproduce these findings. localisation of no production by immunohistochemistry suggests that no production is compartmentalised, and more subtle methods of regulation may be required if this approach is to have clinical valuc. anti-adhesion molecule strategies ch wortel, repligen corporation, one kendall square, building 700, cambridge, ma 02139, usa. the neutrophil has been implicated as a pivotal player in the pathogenesis of multiple organ dysfunction. an important first step in the process of neutrophilmediated organ injury involves the binding of neutrophils to endothelial ceils. this process is largely regulated by complementary adhesion molecules, some of which are present constitutively on the cell surface and others that can be upregulated in response to chemotactic and pro-inflammatory stimuli. several different adhesion molecules have been described. l-selec tin is expressed on the plasma membrane of neutrophils and is shed from the surface early in the inflammatory process. it is therefore thought to mediate the initial interactions with endothelial cells. e-selectin and p-selectin are endothelial adhesion molecules. e-selectin is not constitutively expressed but is upregulated by inflammatory mediators, particularly il-1 and tnf. p-selectin is present in the weibel-palade bodies within the endothelial cytoplasm and can be upregulated rapidly in response to thrombin, histamine, and oxidants. all selectins recognize oligosaccharides, particularly sialylated lewis x antigen. this carbohydrate moiety is expressed on many proteins, including the selectins themselves. the leukocyte integrins are glycoproteins expressed on the neutrophil surface and in the cytoplasmic granules. integrins consist of a common beta2 or cluster differentiation (cd)i 8 chain covalently linked to one of three different alpha chains (cd11 a, cd1 lb, cd1 lc) and exist on the cell surface as three distinct heterodimers. cd1 l a/cdi 8 is expressed on all leukocytes, whereas cd1 l b/ cd18 and cd1 lc/cd18 are restricted to cells of myeloid origin. cd1 i/cd18 interacts with intracellular adhesion molecule-1 (icam-1), its ligand on endothelial cells. icam-1 is a member of the immunoglobulin family of adhesion molecules. it is constitutively expressed on endothelial cells and can be upregulated by cytokines. the potential for monoclonal antibodies to adhesion proteins to reduce vascular and tissue damage has been studied in alarge number of experimental models. protective effects have been observed in a wide variety of inflammatory, immune, and ischemia-reperfusion injuries such as arthritis, bums, endotoxic shock, bacterial meningitis, autoimmune diabetes, nerve degeneration, allograft rejection, allergic asthma, acute lung inflammation, skin lesions, and ischemia-reperfusion models of the intestine, myocardium, lung, skeletal muscle, and central nervous system. protective effects have also been observed in animals resuscitated following hemorrhagic shock. since modulating the function of adhesion molecules may temporarily leave the host without effective defense machinery, safety evaluations are of utmost importance in evaluating this therapeutic approach. treatment of gram-negative septic shock with an immunoglobulin preparation: a prospective, randomized clinical trial ingolf schedel, ursula dreikhausen; birgit nentwig; marion hockenschnteder, dirk rauthmann, salim balikcioglu, rolf coldewey, helmuth deicher, md endotoxin may play a major role in the pathogenesls of muitiorgan failure in the context of the toxic process in septicemia induced by gram-negative pathogens. the hypothesis which has led to a prospective clinical study consisted in the idea of inhibiting the biological effects ofendotoxin during the early phase of septic process, and thereby attampting to attain a positive effect on the clinical course of the disease. -objecave: to evaluate the effectiveness ofa polyclonal immunoglobulln (ig) preparation containing igg, lgm, and iga as an adjunctive therapy for septic shock. -desigru prospective, randomized clinical trial. patients: fifty-five patients w~th septic shock were randomly allocated to two groups according to criteria of septic shock. -intervention: one group of patients (n = 27) received a commercially available immunoglobulin preparation (containing high titers of antibodies specific for determinants to bacteria1 endotoxin) during the first 3 days after inciosion in the study. the other randomized group (n = 28) did not receive any immunogiobulin preparation. -measuremems and main resmts: during the period of <6 wks after the beginning of clinically apparent septic shock, death related to the septic process occurred in one (4%) of 27 patients who received immunoglobulln. by comparison, nine (32%) of 28 control group patients died during this period (p <. 01 ). within the first 48 hrs after onset of the clinically apparent septie process, significaptly increased activity of circulating endotoxin and simultaneously decreased specifie igg serum titers to lipid a were detected in the group of nonsurvivors. the rationale for the use of polyvalent intravenous immunogiobulins (ivig) to treat severe infections stems from in vitro and animal studies documenting that high-dose igg enhance opsonization and phagocytosis of pathogenic bacteria. moreover, recent clinical studies have shown that in septic patients the phagocytic function of circulating polymorphonuclear neutrophils (pmn) is defective; the degree of such impairment correlates to the severity of the septic state, the beneficial effect of administering high dose igo in septic patients can be two-fold: i) ivig provides large quantities of antibodies against invading pathogens; the igg bound to the pathogens can opsonize their phagocytosis; 2) the opsonization of phagocytosis by exogenously administred polyvalent igo would be of special importance in those patients who present a significant defect of their phagocytic function, when they are challanged by a large bacterial invasion. a prospective, randomized, double-blind study was carried out comparing the administration of ivig (sandoglobulin@) vs, paeebo (human albumin) in severely septic surgical patients, only patients with gepis score >_20 (meaning a mortality risk >70%) were accepted into this protocol. patients were randomized to receive 0.4 g/kg of ivig or placebo on day 0 (when they reached sepsis score>20), repeated on day +1 and +5. at the beginning of icu treatment, the two groups of patients were similar for severity of sepsis, age, concomitant disease, type of surgical procedures, antra and perioperative procedures, antibiotic administration. the results of the study indicated a significantly reduced mortality in patients with severe surgical sepsis treated with ivig as compared to placebo control patients (mortality: 38% vs, 67% respectively; p< 0,05). in conclusion, the results of our study in patients with severe surgical sepsis were the following: 1) ivig plus multimodal treatment of sepsis, including antibiotics, reduce mortality significantly', 2) the reduction of mortality seems to be due to a decreased incidence of lethal septic shock. despite substantial clinical research, the avallable data regarding the effectiveness of supplemental immunoglobulin (ig) treatment in sepsis in adult patients do not yet allow definitive conclusions. in view of the persistently high sepsis mortality there is a need to continue clinical investxqations regarding supplemental sepsis treatmen~ in general, as well as concerning ig administration in particular. we present and discuss the protocol of the ongoing ,,score-based-immuneglobulin therapy of sepsis (sbits)" study. the protocol (theoret surg 8(1993)61-83) of this multicenter, randomized, prospective and double-blind trfal relies on the results of an observational trial on i.v. igg treatment in 163 patients with sepsis and septic shock (infection 19~1991)216-227), carried out as a prerequisite for the present trial. using microcomputer-based bedside routine score monitoring, we regard quantitative measures of severity of disease and sepsis: only patients with a certain degree of both severity of disease (apache ii score 20-35) and severity of sepsis (elebute sepsis score 12-27) will be included. by observing these previously validated inclusion criteria, this trial snould iqentify a priori and include patients with potentially optimal response to therapy, consisting o~ either placebo (0.i % albumin) or polyglobin n" -12 ml (0.6g)/kg on day 0 and 6ml (0.3 g)/kg on day i. with an anticipatedpopulation size of 800 patients the study should comply with the statlstical requirements (estimated mortality: 30%, with a 33 % reduction in 28-day mortality in the treatment groupl to prove or disprove the question of igg effectiveness in sepsis in terms of improved prognosis. up to november 1993, more than 460 patients had been included; patient enrollment will be finished in 1994. previous studies have demonstrated rhll-i ra, a naturally occurring antagonist of il-1, increases survival in animal models of andotoxemia and eschehchia coli bacteremia and attenuates the decrease in mean arterial pressure resulting from challenge with both gram-negative and gram-positive bacteria. previously, in 99 patients, rhll-lra was demonstrated to increase survival in patients with sepsis syndrome and septic shock in a dose-dependent manner. methods: a randomized, double-blind, placebo-controlled, malticenter, clinical trial enrolled 893 patients at 63 academic medical centers in europe aad north america. eligible patients received either placebo (vehicle) or rhil-lra (anakinra) 1.0 or 2.0 mg/kg/hr by continuous intravenous infusion for 72 hours. the presence of organ dysfunction (i.e., ards, dic, renal, and hepatic) at study entry was determined prospectively by a clinical evaluation committee using definitions which were developed a-priori. survival time was evaluated over 28 days utilizing a linear dose-response model, assuming a log-normal distribution. results: 563 patients had one or more sepsis-induced organ dysfunction(s) at study entry. a dose-related increase in survival time was observed with rhll-lra compared to placebo in patients with ards, dic, and renal dysfunction (p --< 0.04 endotoxin infusion releases platelet-activating factor (paf), a potent phospholipid mediator which leads to an autocatalytic amplification of cytokine release. bn 52021 (ginkgolide b), a natural paf receptor antagonist, has provided significant protection against sepsis in different animal models• a randomized, placebo-controlled, double blind, multicenter trial on efficacy (mortality at d28) and tolerance of bn 52021 (2 iv infusion of 120 mg x 2/day over 4 days) in severe sepsis has enrolled 262 pts. the 28day mortality rate was 51% for the placebo group and 42% for the bn 52021 group (p = .17). the efficacy of bn 52021 was greater in pts with gram-negative sepsis: the 28-day mortality rate was 57% for the placebo group and 33% for the bn 52021 group (p = .01). bn 52021 also reduced mortality among pts with gram-negative septic shock (mortality was 65% for placebo vs 37% for bn 52021; p = .01). using statistical adjusments for pronostic factors, the relative risk of death of the bn 52021 group was 0.61 (0.34 -1.08, 95% confidence interval; p = .09). this risk corresponds to an adjusted reduction in mortality of 39% for pts receiving bn 52021. no differences in mortality rates were found between the placebo and the bn 52021 groups in the absence of gram-negative sepsis• there were no differences in adverse events between the placebo and the bn 52021 groups. bn 52021 is a safe and promising treatment for patients with severe gram-negative sepsis. a confirming study, focused on gram negative sepsis, is in progress. v~3lliam a. kanus m.d. and the rhll-lra it has been traditional within the field of infection and sepsis to think in terms of specific indications for drugs based on the type of infecting organisms, advances in antibiotic therapy now control or ltnflt the growth of bacteria. the majority of deaths are now caused by either an initial overwhelming response to infection or subsequent multiple organ system failure attributed, in part, to the effects of intrinsic biologic responses of the host. type of organism, therefore, may not be as critical as determining the exact severity of the host's severity or risk of death from infection. we also know that both the relative benefit of a new treatment across groups and its absolute benefit for an individual patient will vary with their risk in a predictable fashion. we recently iuve~iguted the relationship between one measure of host response, the acute risk of death as prospectively estimated by u comprehensive risk mode[ for 28-day mortality (jamb. 1993; 270:12,33-1241) , by its retrospective application to the results from the phase in evaluation of recombinant human intcrlenkin-1 receptor antagonist (rhll. ira). we found that there was a significant interaction between the patient's predicted risk of mortality at the time of entry to the study and the ability of rhil-lra to prolong survival time (x2 = 7.6, p [] 0.02, log.normal) for all 893 patients in the trial• survival benefit began st approximately 24% baseline risk of 28-day mortality. for the $80 patients with a predicted risk > 24%, there was a 22% reduction (p=0,00$ log normal). when we examined the variation in patients above and below the 24% risk level with hazard functions, i.e., their daily risk of death during the study period, we found that placebo patients with < 24% risk had lltile acute daffy risk during the hlltial two days follawh~g study entry and this risk was little affected by rhil-lra, in contrast, patients with > 24% risk had high daily mortality risks during the tuttlal two days that high dose rhtl-lro substantially reduced. these results are compatible with our current understanding of outcome from sepsis and the proposed mechanism of action o£ immunotherapy, the earliest deaths from sop sis are secondary to an immediate inflammatory response followed closely by deaths secondary to multiple organ system failure, later deaths (after 14 days) are not as closely related to the acute effeete of the inflammatory cascade. because of the timing and action of most proposed tmmunotherapy, they may be capable of preventing mortality primarily in these initial two phases. in this study, an independent predicted risk of mortality reflected this mortality pattern ned illustrated the potential benefit of immtmotherapy. use of a predicted risk of mortality in the design and analysis of clinical trials could improve our understanding of the clinical benefit of these new therapeutic approaches. the systemic inflammatory response syndrome (sirs) is a term recently proposed to describe patients with systemic inflammatory responses to insults such as infections (sepsis), trauma, burns, pancreatitis, and other initiating events. patients with sirs may have similar activation of inflammatory mediators and similar outcomes independent of the initiating event. these outcomes include organ dysfunction and failure, shock, and death. challenges to the successful conduct of clinical trials in sirs include the complexity of illness in these patients and the important--but limited--clinical benefits of novel compounds that may be limited to selected patient subsets. addressing these challenges will require new tools and approaches. these will include more sensitive and appropriate endpoints, and the use of methods such as baseline risk adjustment, to allow detection of drug risk interactions not captured adequately by categorical definitions, such as sepsis syndrome. on the basis of supportive preclinical and phase i safety studies, we have initiated phase ii clinical trials of a novel bradykinin antagonist, cp-0127, in four sirs subcategofies: sepsis, multiple trauma, burns, and pancreatitis. each of these studies is designed to measure the effect of cp-0127 on mortality, organ dysfunction and failure, and activation of mediators. in addition to investigating rates of organ failure using standard definitions--a new endpoint--a continuous summary measure of organ dysfunction (the acute physiology score of apache tm iii) is being used to quantify the degree of organ dysfunction and the speed and pattern of recovery of physiologic stability. in the sepsis study, another new approach--a study specific risk model based on the apache ill database--has been developed which will be used to assign a pre-treatment baseline risk to each patient enrolled. the primary outcome variable will be risk adjusted survival time to 28 days. this type of risk-adjusted analysis may allow for more efficient and powerful trials and more accurate and useful indications for use. study purpose: in post-cardiac surgical patients (pat.) at risk for sepsis, the efficacy of early i.v. immunoglobulin (ig) treatment was compared to a matching historical control (con.) population. postoperative risk assessment: using apache ii scores lap) (first postoperative [pop.] day) in a pilot study phase, we were able to differentiate between the large population (95.2%) of pop. low-risk pat. (ap< 19; mortality: 1%) and the small groups of pop. pat. at risk lap= 19-23) and high risk lap_24) with a significantly higher mortality (14% and 76%, mainly due to sepsis). subsequently, among 1341 consecutive pop. pat. we prospectively identified and treated these pat. iq treatment reqimens: first study period (n =41): (gg (psomaglobin n a, tropon biologische pr~parate, cologne, frg, day 1:8 ml/kg, day 2:4 ml/kg). second study period (n=25): iggma (pentaglobin r, biotest, dreieich, frg, 5 ml/kg on days 1 to 3). results: ig pat. and con. were comparable in demographic data, operation characteristics and baseline disease severity lap and elebute sepsis scores). in contrast to con. (risk: n=21, high-risk: n-21), the ig pat. showed a marked improvement in disease severity (fall in ap), especially in the high-risk group (igg, n=26: p<o.05; iggma, n=13: p: 0.08). in this group, ig led to higher (p<0.05) response rates, defined as an ap score decrease ->7 within four days (igg: 54%, iggma: 62%; con.: 19%), and reduction in mortality (igg: 46%, iggma: 46%; con.: 76%), statistically significant (p<0.05) for ig treatment as a whole (igg and iggma). conclusion: given the good comparability of the study groups, our results indicate, despite the non-randomized design, that early supplemental ig treatment can improve disease severity and may improve prognosis in prospectively apache ii score-identified high-risk patients after cardiac surgery. objective. elevated plasma levels of endothelin (et) have been demonstrated in both experimental and human sepsis. et has been proposed as a sepsis mediator leading to vasoconstriction with tissue hypoperfusion and organ failure. the aim of the study was to determine the effects of sepsis treatment with volume resuscitation, antibiotics and the anti-lps monoclonal antibody es® on big et and active, 21 aminoacids et (et 21) in rat abdominal sepsis. methods. lethal peritonitis was induced with a 4 mm coecal perforation (cp) in male wistar rats. plasma levels of big et and et 21 were determined with amersham tm endothelin rias 4, 8 and 12 h after sepsis induction. experimental groups: 1. cp control, 2. volume replacement (vr); 0,9% saline 10 ml/kg/h continous iv infusion started after 2 h, 3. antibiotic; imipenem 40 mg/kg iv after 2 h, 4. e5®; 10 mg/kg iv after 2 h, 5. vr + imipenem + es® after 2 h. results. high concentrations of both big et and et 21 could be demonstrated after 2 h and lasting for 12 h after cp. neither volume replacement nor imipenem did influence the elevated plasma et. e5® significantly reduced et 21 both 4, 8 and 12 h after sepsis induction, but did not reduce big et. when es® was combined with vr and imipenem, reduction of et 21 was the same as for e5® alone. these results strongly suggest that bacteria and hypovolemia per se are not decisive stimuli for et production during sepsis. e5® reduces circulating lps and tnf which is the probable mechanism of the suppressed et 21 synthesis. the unaltered big et fraction after e5® treatment indicates conversion of big et to et 21 as the site of action responsible for reduced et 21. conclusion. lethal peritonitis in the rat is followed by elevated plasma levels of big et and et 21. e5® anti-lps antibody significantly reduces plasma et 21 while volume resuscitation and antibiotics failed to do the same. es® did not reduce plasma big et. pmx treatment on severe endotoxemia with multiple organ failure was safety and effect in prognosis, and sepsis related parameters. it was certified that reduction of plasma endotoxin was effective in severe endotoxemia. a. lechleuthner,s. aymaz, g. grass, c. stosch, s. dimmeler, m. nagelschmidt, e. neugebauer. ii. dept. surgery, university of cologne, germany. introduction: the cardiovascular therapy of hypodynarnic shock states is a challenging problem. in clinical as well as experimental studies beneficial functions of a new hg-agonist bu-e-75 in congestive heart failure has been demonstrated 03aumann, 1989). therefore, we investigated the effect of bu-e-75 in hypodynamic shock in pigs. materials and methods: pigs (deutsches hausschwein, pitrain, [20] [21] [22] [23] [24] [25] were anesthesized with fentanyl/dormicum, ventilated (n20:o 2 = 2:1) and cardiovascular parameters were monitored with a complete icu-eqnipment. the hypodynamic model was established in a pilot study (4 animals) to evaluate the effective concentration of bue-75 in healthy and endotoxin (lps)-treated animals. endotoxic shock was induced by continous infusion of 10 ~g lps/kgkg/h (055:b5, fa. difco). the hypodynamic state was defined as a decrease of cardiac output by 30 % of steady state levels. a wedge pressure of 10-12 mmhg was kept constant by volume resucitation during the experiment. in a subsequent randomized controlled trial (rtc) 3 groups with 6 animals per group were studied. the groups were treated as follows: group i, lps and 0,9 % nac1; group ii, lps and bu-e-75 (100 #g/kgkg/h); group iii, famotidine (h2-blocker) pretreatment (1 mg/kgkg), lps and bu-e-75. results: the pilot study in healthy pigs revealed, that bu-e-75 had positive inotropic effects. these effects were inhibited by the h 2antagonist famotidin. bu-e-75 however had no beneficial effects in the hypodynamic phase of endotoxic shock in the rct. cardiac index (ci) and the oxygen delivery (do2) were not significantly influenced by bu-e-75 application (group i versus group ii). bu-e-75 did not ameliorate the negative inotropic effect measuring left ventricular stroke work (lvsw) in hypodynamic shock phases. on the contrary, bu-e-75 led to a further significant decrease of lvsw (p < 0,05). famotidin pretreatment did not affect the response (group iii versus group ii). conclusion: in hypodynamic shock states the h2-agonism seemed to have no beneficial effect under these experimental conditions. receptor down regulation or changes of signal transduction under septic conditions may be responsible. cellular studies may help to identify these mechanisms. objectives. antithrombin iii inactivation of proccagulant proteases is so far the only inhibitory therapeutic approach to disseminated intravascutar coagulation (dic). we therefore set out to investigate whether cll substitution reduces coagulation activation in an endotoxin induced rabbit dic model. materials and methods. 29 male rabbits chbb:hm(spf) were randomty assigned to one of the following groups. group k : naci 0.9% (control without endotoxin, n=10). group e : endotoxin 120 tjg kg "1 bolus i.v. + naci 0.9% (control with endotoxin, n=10). group c : endotoxin 120 pg kg -1 bolus i.v. + cll 400 u kg -1 bolus + 400 u kg "1 4h "~ i,v. (treatment group, n=9). all animals were anesthetized and mechanically ventilated. blood samples were drawn prior to endotoxin administration (m1) and after 120 (m2) and 240 rain. (m3). thereafter, lung and liver tissue samples were taken intravitatly in a standardized fashion for h&e microscopic fibrin quantification using a triple score (fibs). from all blood samples the prothrombin time (pt), activated partial thromboplastin time (aptt), fibrin monomers (fm), and d-dimers (dd) were measured. for statistical significance of differences between the groups anovas and the wilcoxon test (fibs) were performed. results. fibs for lung/liver were significantly different (p<0.05) between group e (lung 180, liver 159) and c (lung 89, liver 0) (group k : lung 11, liver 0). , a synthetic serine proteinase inhibitor, has an anticoagulant activity in the absence of" antithrobim iii. gabexate has been reported to be useful in the treatment of disseminated intravascular coaguiation due to neoplastic diseases. in this study, we investigated gabexate therapy for the treatment of dic due to sepsis in the postoperative critical patients. materials and methods: from july 1992 to june 1993, 15 patients in the surgical intensive care unit met the criteria of dic or pre-dic. eleven were male and four were female with the mean age of 66.7 years. all these patients suffered from some complication of operations which led to the development of sepsis. foy was administered at the rate of 2mg/kg/hr untii the coagulation profile retumed to normal or the patient died. the coagulation parameters were monitored before and on the 1st, 3rd, 5th and 7th day. results: fourteen of these fifteen patients died despite transient improvement of the coagulation parameters in five patients. these patients suffered from sepsis resulting from surgical complications which could not be well controlled. the only survival was a case of recurrent intrahepatic duct stone with biliary tract infection complicated with sepsis and dic. after choledocholithotomy and the use of foy, the patient recovered gradually. conclusion: dic is a late manifestation of sepsis in the critical surgical patients. the most important thing is to eradicate the cause of sepsis. if the underlying septic focus cannot be controlled, dic will persist despite the use of gabexate mesilate. emergency surgery, taipei veterans general hospital, taipei, taiwan. there are 2 main types of bradykinin (bk) receptor, namely bk~ and bk z. the bk 2 receptor is constitutive. the bk 1 receptor is also constitutive but in the majority of cases is inducible and involved in chronic inflammatory syndromes such as sepsis, hyperalgesia and airways hyperreactivty in animals. the mechanism(s) involved in the upregulation of the bk 1 receptor is unclear, however a variety of agents including lps, e coil and ill are particularly efficacious in vitro and in vivo. ill and bradykinin acting at their respective receptors are believed to be involved in sirs/sepsis. we have investigated the effect of antagonists at ill (antril), bk 2 (bradycor [cp-0127]),bk~ (cp-0298) and bkz/bk 1 (cp-0364) receptors on the de novo generation of bk~ receptors (reflected by hypotensive responses to a bk 1 agonist) in the lps-treated (10ug iv) rabbit. in lps treated rabbits hypotensive responses to bk~ but not bk 2 agonists increased with time and at time 210min appeared maximally induced. constant iv infusions of cp-0127 blocked bk 2 but not bk~ and cp-0298 bk~ but not bk 2 responses. cp-0364,cp-0127+cp-0298 and antril+cp-0364 blocked both bk 2 and bk~ responses. antril alone had no effect on bk 2 or bk~ responses. within 30-60 min after stopping the infusions of antagonists the responses to bk~ and bk z agonists were the same as those in nonantagonist infused rabbits. these results indicate, at least in the lps-treated rabbit, that neither bk2,bk ~ or ill receptors alone or in combination, are involved in the de novo generation of bk 1 receptors. in vitro studies demonstrated that beth bradycor and cp-0364 (but not antril) were antagonists at both bk z and bk~ receptors. if both bk z and bk 1 receptors are significantly involved in chronic inflammatory situations in man such as sirs/sepsis then the rationale for the use of compounds such as bradycor or cp-0364 is clear. infection is a major cause of or contributor for morbidity and mortality in liver transplant recipients. effectiveness of prophylactic and therapeutic protocols is important for the success of liver transplantation ( olt ). sdd is used as prophylaxis for reduction of infection caused by gram negative or fungal microorganisms. between september 1988 and july 1993 400 olt's in 368 patients were performed at our department. the actuarial 1-year patient survival is 90 %. infection prophylaxis is started with sdd and ciprofloxacin once the patient is accepted as an olt candidate. perioperatively metronidazol, tobramycin and cefotaxim, postoperatively cotrimoxazol are prescribed additionally. the table shows pneumonia, peritonitis, major wound and urinary tract infection are common nosocomial infections following severe injury. in a series of severely injured patients from the university of louisville hospital, pneumonia was the most common infection followed by peritonitis, intra-abdominal abscess formation and burn wound infection. pneumonia is actually the leading cause of death from nosocomial infection. these are defined as occurring from 48 to 72 hours after hospital admission. this definition has important implications for antibiotic therapy because the likely pathogens and their respective sensitivities are different for community acquired pneumonia. the diagnosis of nosocomial pneumonia is difficult following major injury as many patients will have pre-existing fever, leukocytosis, tachypnea, and chest x-ray changes. reliance on sputum gram stain and culture is important and best obtained by a bronchoalveolar lavage or protected specimen brush during bronchoscopy. predisposing risk factors include severe head injury, emergent intubation and shock, and such patients have been shown to benefit by early tracheostomy. staph aureus has been the most common pathogen isolated from the sputum and the remainder gram-negative organisms with pseudomonas aeruginosa, and klebsiella pneumonia predominating. bacteria recovered by site as well as by intensive care unit is published in the six month antibiogram which also includes recent antibiotic sensitivities. this aids in empiric antibiotic selection against such nosocomial organisms. in a series of severely injured patients (iss -30), mean temp. was 101.4f, leukocytosis was 16k, pan2 was 87, fin2 was .47, and peep was 5.1 at the time of diagnosis (ards excluded). there was marked reduction in class ii histocompatibility antigen (hla-dr) density on peripheral and bal monocyte/macrophages which recovered over time with resolution of pneumonia. immune suppression occurred prior to development of pneumonia, was especially localized to the infected tissue, but recovered with clinical improvement. specific immune modulation targeted to pulmonary white cells may hasten clinical recovery and minimize pulmonary dysfunction. -clinical experience j. tnllemar amphntericin b remains the drug of choice for many systemic fungal infections. its advantages include a broad spectrum of activity and intravenous administration. the major disadvantages of amphoterlcin b is its severe side-effects, especially the nephrotoxicity. to decrease the toxic side..cffccts various liposomal amphoteficin b formulations have been produced. it was found that these liposemal formulations were as effective as amphotericin b but in contrast had a low incidence of toxicity. at present there are three ~different variations of lipid formulations under assessment: amphotericin b lipid complex (ablc), amphotericin b coloidal dispersion (abcd) or true liposomes. the ablc has a ribbon like structure. it has been shown to have a reduced toxicity and an efficacy ranging from being as effective to four times less effective that conventional amphotericin b. regarding abcd the particles have a disk-like structure with a diameter of around t00 am and a thickness of 4nm. the ami-fungal efficacy is 2-4 times less than that of conventional amphotedcin b. both ablc and abcd are presently investigated in phase ii/iii studies in the us. ambiseme is currently the only commefieally available true lipesome. ambiseme is a spherical small unilamellar lipesome with a diameter less than 100 nm with a mutina ld50 of > 175 mg/kg. it has been used in dosages up to 6 mg/kg/day in compassionate based studies with good tolerability. the mycological efficacy range from a 83% response rate for invasive candida infections to 41% response rate for aspergillosis. ambisomc have been evaluated as anti-fungal prophylaxis in randomized trials in bone marrow (bmt) and liver transplant (ltx) recipients. it was well tolerated. in bmt recipients the incidence of proven fungal infections was 8% among placebo treated patients compared to 3% for the ambisome treated patients (ns). in ltx recipients ambisome prophylaxis was effective, significantly reducing the incidence of deep fungal infections from 16% to 0% ill placebo and ambisome treated patients respectively (p<0.01). prospective randomized trials comparing these various amphotericin b preparations with conventional amphotericin b is needed to determine their future place in the therapeutical arsenal. two patlentgroups ere particularly at risk to develop serious cmv disease: cmv seronegative transplant recipients of seroposltlva donors and those patlants treated for rejection with anti t-ceil preparations, we have evaluated the value of prophylactic anti-cmv immunoglobulin (cytotect", biotest pbarma gmbh, dreieich, frg) administration in high risk heart and kidney transplant recipients, in a double blind placebo controlled study 40 kidney transplant recipients, treated for biopsy proved re)action with rabbit atg, received globullntplacebo infusions. the preparatlons were given i,v, in a dose of 100 mg/kg at day 0,7,14,35,56 and 77 after the initiation of anti = rejection therapy, passive immunization completely prevented cmv related death, although it did not reduce th~ incidence of cmv isolation, viraemia or disease, this effect was mainly observed in cmv saronegativa recipients of a serop0sitive donorktdney. seroposltive recipients did not benefit from treatment and seronegatlve recipients of a seronegetlye donor were not et risk for cmv infection at e!l. in a open study the incidence of cmv infection and disease was evaluated in 143 consecutive i~eart sllograft recipients. sixty-five patients were cmv seronagatlve and they all received passive immunlzation according to the dosage schedule used in the kidney patients, but starting on the day of transplantation, this scheme resulted in median snti-cmv igg titers of 1200 elisa units during 3 months. cmv infection occurred in 21/65 ~eronegetlve and in 40/81 seropositive recipients (n,s,), in ssronegetive donor-recipients pairs the incidence was significantly lower (3/34] , the passively immunized seronegstive recipients of e seroposltlve donorheart showed comparable incidence of cmv infection f18t29) vs the seropositive recipients. primary infection more often resulted in disease than secondary infection (11121 v8 10/40), but no difference in incidence of disease (11165 vs 10/81 ) or severity in symptoms was noted between the immunoglobulln treated serone(]ative patients and the seropositiva recipients. apparently passive immunization induces anti-cmv immunity which crossly resembles naturally acquired resistance. abdulkadirov k.,chebotkevich v., moiseev s. the incidence of infection is still high in patients underwent bmt. this complication is the major cause of mortality if it is not recognized and treated promptly and properly. our data showed that from 21 patients with different types of leucemia after autologous and allogenzc bmt 19 had the episodes of fever. in the ma i ority of these episodes the bacterial etiolog$ gram negative bacflli and gram positive cocci) can be proved. on the other hand, in 62% of the fever cases we detected also viral respiratory (corona-, adeno-, rs-and other) infection. our previous investigations showed that even in healthy persons the viral infection has influence on antibacterial immunity, in the cases of model experimental reaction in volunteers we found the decrease of delayed hypersensitivity 10-14 days after intranasal inoculation of influenza virus a (h3n2-3) to bacterial (staphylococcal, streptococcal and pneumococcal) and ~iycoplasma pneumoniae antigens in the leucocyte migration inhibition test. these results showed that respiratory viruses may be the important pathogenic factor in the development of bacterial infection in posttransplanted period. we consider the constant control of latent and visual respiratory viral infection in bmt patients to be very important. ficcb the ~ter£~li of the nation~l institute of trad/~atoloqy in budapest 1.00 consecutive cases of revision hip grafting were carried out arthroplasties wlth hemoloquous bone between the years 1990 and 1993. in the same period of time 732 pri~ total hlp replacen~nts were performed under i4entieal technical conditions. the average septic rate for the 'total hip althroplasties was less than 1%. in the selected i00 cases the septic rate was 4% indicating the role of bone grafting° homografts were prepared by deep freezing~ it .is recognized that the cells of the hl~grafts become destroyed by the ium~unological, response of the host~ and the patients develop ~ti-hl~, ar~tib'o~ies. the dead ~trix, however, has a bone-inducing capacity that stimulates host osteoblasts to recolonize the *i~/trix which serves as scaffolding. the sequence of events favours the infections. for this reason, beside preventive perioperative systemic ant/biotic treatment, local ~ntibioties were also applied in the form of antibiotic-//npregnated cement. the role of age and the .immune status of the patients .is discussed.. the purpose of this study is to evaluate the rate of toxemia in patients with acute panereatitis and to find this coudition to the activation of cascade systems that are encountered in the subsequent complications of the disease. we studied a series of 45 patients with acute pancreatitis, the severeness of which was evaluated by the ranson's criteria and the apach-ii scoring system. all of them were considered to have severe acute puncreatitis. the determination of toxemia was made using the limulus test (lal test). we also determined the levels of the third (c3) and fourth (c4) complement components as weu as the coagulation factors, iibrinolysis faeters and kimns by serial measurements. the severity of the disease was serially determined by the apach-ii scoring system. it was found that complement activation ( which was also assessed using a graphically illustrated method by a aggregometer ) was followed by an increase of morbitity and mortality .we also detected that toxemia (positive lal-test) was closely correlated with complement activation and more of the ranson's criteria. a clear relation existed between the number of ranson's signs and the enmplieations' rate (1"= -0.766, p < 0.001). the documentation of toxemia and the complement activation cannot predict the kind and the severity of complications. the study of coagulation, fibrinolysis and kinms systems didn't reveal any results with statistical significance. necrotizing pancreatitis still represents a life-threatenthg disease. infectious complications dominate among the causes of death. differences in the individual immune response could possibly explain different clinical courses even in patients with comparable pancreatic morphology. to explore the inflammatory response in acute pancreatitis, the following investigation was performed. methods: peripheral-venous blood was withdrawn on admission and furthermore twice weekly in as yet 36 patients with acute pancreatitis and tested for the parameters mentioned below. in parallel, polymorphounciear granaiocytes were isolated using density gradient centrifugation and assessed for superoxide anion and hydroxyl radical producing capacity using electron spin resonance techniques. results: total leukocyte cotmt and total lymphocyte count did neither reflect the clinical course nor predict complications. this comes tree also for serum igg, igm, iga, c3, c4, crp, alpha-l-antitrypsin and neopterth as well as for plasma il-la, il-ib, il-1ra, il-2, il-2r, il-6r, tnf-ct, tnf-~r (p75) and icam-1. in contrast, pmn-elastase, il-6 and il-8 closely correlated to the clinical course. isolated pmn's in vitro capacity to produce oxygen radicals depended on the respective radical species and was slightly elevated (superoxide anions) or decreased (hydroxyl radicals), respectively. patients with a cd4+/cd8+ ratio below i were seen at risk of developing septic complications. in contrast, a percentage of monocytes of 30 % or more among total mononuclear cells indicated an uncomplicated course, in general. conclusions: the immune status of the individual patient may significantly influence the course of acute pancreatitis. the cytokine pattern in peripheral blood is very complex and most parameters are of little use for the clinician. the pmn-elastase, il-6 and il-8, however, closely correlate to the clinical course and may prove valuable for follow-up. the cd4+/cd8+ ratio was found the best predictor of septic complications, but it failed in non-septic patients. a percentage of 30 % or more of monocytes among total mononuclear ceils indicated a rather mild course. the reduced ability of the pmns to produce hydroxyl radicals may help to explain the frequent development of septic complications in severe necmtizing pancreatitis. peroxidation of membrane lipids contributes to ceil injury in pancreatitis. overwhelming release of toxic metabolites by infiltrating neutrophils is regarded a major pathogenetic factor, too. as yet little is known about the mechanisms by which oxidative stress and leukocytes damage pancreatic cells. the present study examines (i) the susceptibility of pancreatic acinar cells to attacks by oxidants and leukocytes and (2) the potential of antioxidants to prevent such damage in order to better understand the cellular mechanisms of pancreatic injury in inflammatory states. methods: freshly isolated rat pancreatic acinar ceils were exposed to a model system of oxidative stress consisting of 20 mu/ml xanthine oxidase (xod), 500 mm hypoxanthine (hx), 50 mm fec13 and 75 mm edta. in a second set of experiments, acinar cells were exposed to excess autologous neutrophils or neutrophils obtained from patients with acute pancreatitis. neutrophils were stimulated by zymosan a, pma, and il-8. cell viability was assessed by both cellular uptake of trypan blue (tb) and by release of ldh. results: the xod/hx system caused a time-dependent acinar cell injury. this injury was effectively prevented by catalase (cat) and gfutathione peroxidase (gpx). in comrast, superoxide dismutase (sod) enhanced cell injury. addition of both sod and cat abolished the damage seen with sod alone. the non-enzymatic scavengers mannitol, dmso, dmtu and the iron chelator deferoxamine were not protective and at a higher concentration even accelerated cell decline. the newly developed antioxidants of the lazaroid type effectively prevented oxidative acinar cell damage. stimulated neutrophils, both autologous and heterologous, did not damage healthy acinar cells but had even protective effects. conclusion: pancreatic acinar ceils are very susceptible to oxidative injury. a combination of catalase and sod prevented cell damage effectively. sod when given alone may rather damage than protect aelnar cells when h202 is generated in concentrations overwhelming the capacity of endogenous catalase. therapeutic approaches to pancreatic disease using antioxidants should, therefore, include combinations of protective substances. the lazaroids seem to be candidates for clinical use as antioxidants in pancreatitis. the results argue against direct toxic effects of stimulated neutrophils to pancreatic acinar cells. are ch~act~z~ by the presence of a polymicrobial flora, the pmtotyi~ cffthese inf~ons is secend~,y bacterial pedtonitlw, whereby a pathololoeal process in the ~trointesfimd tract r~ful~ in tim disrup~on ofi~ inteffrlty and ¢ollseqtlent sptl]nge of inte~.i,o~.l gontents into the peritoneal c~iry. the ensuing infection invariably contains a mixtm~ of gt~m negative enteric bacilli, gram positive b~eria and anaerobe& experimental and clinical =t~ies have de~ed the eantrlbution of each of th¢~ components to ti~ ovemu virulence of these in~ons, gram negative enteri~ such as f.veher~chla coil ere endowed with a virulent l~l~x~lyse~haride ptill~ly t~sponsible for lethality, by contrast, bacteroldes sl~cles, which rarely c~se death, prornot~ abscess fonllation, a uniqm~ capsul~ polyseccluu'ide, particularly on b.j~ogiljs slrai~, oontributes to tjtis erect, several mecltanims have bccn pml~ed whereby or~ microorganism mi~t interact with its microbial ~net to augment the overall virulence of a r~xed im~edan. these include: l) provision of nutrients by one apexes which stimulates the growth of its ~opathoge& 2) inhibition of host deletes by one of the migroorganisms so that the other microbes might persist and exert their virulence, 3) the trant~ of vim.©n~e traits between ~renr~a.,dsms and 4) the ~.mizatian d the mi~oe~vironmental con~tion$ by one d the baetez'isl pa#, so that the other might persist. exampl~ for each of these m~banisms imv~ been provided by experimental ttudies i~stigating e.co!l-b.p~flls synergistic in~ra~ons. byproducts ofg.coli metabolim l~¢ovide essential short ebath fatty acids £~ optimal b,frosili~ ga'owth. fm-ther, oxygen ¢ons~tmption by kcelt lowers oxygen tension end redox potantial to levels eomlucive to b#a#lts gro~h. coawr~ely, b,~agtlis rolea~s proteases and fatty acids wl~¢h impair pl'tsgocy~¢ ~lt rmctlon tnd permit f-..¢oli proliferation and expression of its intrinsic virulent. in summaxy, interactions among the separate microbial cemponents of mixed infections heighten the overall virttienee of these lafectiot~, this knowledge provides ~r rationale for targetting of antibiotic therapy against the knowa eantributors of these synergistic pro~¢sses, intraabdominal abscess formation and the macrophage william g. cheadle, m.d., department of surgery, university of louisville school of medicine, louisville, ky 40292 inflammation of the peritoneal cavity following bacterial contamination has been classified into primary, secondary and tertiary, the last two relating to bacteria originating from the gastrointestinal lumen. the natural history of such infection is either resolution without clinical sequelae, which is uncommon, abscess formation, or generalized peritonitis, which occurs as a result of failure of peritoneal host defenses. early clearance of microorganisms by peritoneal fluid circulation and filtration througti subdiaphragmatic lymphatics into the thoracic duct and systemic circulation occurs as well. simultaneously peritoneal macrophages and the omentum approach the area of inflammation and lead to neutrophil influx and abscess formation adjacent to the affected viscus. we have found a shift in peritoneal macrophage function from antigen presentation to proinflarnmatory cytokine production that occurs early after experimental peritonitis produced by cecal ligation and puncture. this is also reflected by reduced class ii histocompatibility antigen expression on peripheral blood mononuclear cells and peritoneal macrophages. this is accempauied by an influx of both neutrophils and macrophages into the peritoneum and subsequent abscess formation. interestingly, there is little serum endotoxin or tnf seen in this model despite tnf mrna expression in peritoneal macrophages. we believe this model is more clinically relevant than other models of endotoxemia or bacteremia in which different patterns of cytokine expression are seen. newer agents aimed at reduction of systemic manifestations of sepsis originating from intra-abdominal infection such as monoclonal antibodies against cytokines or il-1 receptor antagonists may need to be directed against remote organ macrophage populations while preserving peritoneal macrophage function. inflammation is a complex process involving microcirculatory changes, extravasation of fluid and a cellular influx in the affected body area. in our communication, we will only consider the regulation of the cellular infiltrate which plays a major role in the defense of the peritoneum against microbial invasion. until recently, it was thought that the influx of leukocytes in the abdomen was induced by bacterial products, local humeral factors and secretions of resident macrophages. there is now increasing evidence that this view is too simplistic. many other cell types present in the abdominal cavity or composing the peritoneal membrane (mast-cells, mesothelial cells, fibroblasts) are able to release or secrete vasoactive or chemotactic substances such as histamine, prostagtandines, or cytokines. they are most likely to play a role in the regulation of intraperitoneal inflammatory reactions. the emigration of leukocytes towards the abdominal cavity is also modulated by a previous contact with gram negative bacteria. in the rat, this intriguing phenomenon is long lasting, cannot be transferred by serum and seems independent from t lymphocytes. the clinical relevance of these various regulating mechanisms has still to be determined. kinnaert paul, h6pital erasme, route de lennik 808, 1070 bruxelles belgium generalized response in secondary peritonitis the clinical course of an intraabdominal infection may depend on a variety of variables including the capacity of host defense mechanisms and the degree of the inflammatory response. if local defense mechanisms fail to restrict the inflammation to the abdominal cavity a generalized inflammatory reponse will result. in a first stage generalized signs of a local inflammation become detectable whereas the second stage comprises the overwhelming systemic inflammatory response. the extent of this systemic response determines the outcome. sometimes it may appear to be unrelated to the severity of the intraperitoneal findings. the activation of plasma systems and cellular elements leads to a fast release of cytokines, inflammatory mediators and other substances. these parameters precisely reflect the degree of the generalized response. inflammation of the peritoneum causes significant morbidity. objektives: to test the hypothesis that peritoneal mesothelial cells play a role in regulating inflammatory responses within the peritoneal cavity, we examined neutrophil-chemotactic activity (interleukin 8) and monocyte-chemotactic cytokine (mcp) release by sytokine-etimulated mesothelial cells. confluent human peritoneal mesothelial cells were exposed to varying concentrations of phorbolmyristate-acetate (pma) and the cytokines tumorneerosis factor a (tnf a) and interleukin i~ (il-i~). the supernatant was examined for il-8 by elisa and for mcp by investigating the ehemotactic activity for isolated human monocytes. mesothelial cells express low levels of il 8 and monocyte chemotactic activity when cultured. these activies were significantly increased (2-fold) after stimulation with either tnf a or il-i~. additionally macrophage inflammatory protein was detected. these observations provide a probably important mechanism whereby peritoneal mesothelial cells respond to imflammatory stimuli released during peritonitis and how leucocyte recruitment by liberation of chemotactic cytokines is regulated. the perioperative course of lps, tnfa and il-6 in 19 patients with bacteriologic proven abdominal infection (intraabdominal abscess 10, diffuse peritonitis 4, pancreatic necrosis 2, pancreatic abscess 3) was followed prospectively and evaluated for possible correlation with septic state and organ function. methods: patients were studied in a 6 to 10 hours period during their first surgical intervention because of intraabdominal infection. all were monitored for their cardiovascular, respiratory, hepatic and renal function. plasma samples for lps. tnfa and il-6 determination were drawn preoperatively, intraoperatively, and until 2h postoperatively in regular intervals (min 7/pat), results: preoperative apache ii was 12 in median (rain 4, max 25). 10 patients fulfilled the criteria of sirs. 4 of them were in septic shock.there was a significant correlation between preoperative tnfa and apache ii (p=0,000 i, spearman coefficient). preoperative cardiovascular (systol. rr<90 mmhg) and respiratory (pao2<75mm hg) dysfunction were associated with significantly elevated tnfa (cardial: p=0,000 i, wilcoxon; pulmonal: p=0,0003) and il-6 (cardial: p=0,2; pulmonal: p=0.0001) overall, lps, tnfa and il-6 values varied considerably during the observation period. however, tnfa was markedly higher in patients with sirs and septic shock (group a: n= i 0, mean 182 pg/ml) than in those who did not fulfill these criteria (group b; n=9, mean 24 pg/ml; p=0,00 i, wilcoxon). il-6 was significantly higher in group a (mean 2169 pg/ml) than in group b (mean 191 pg/ml; p=0,0o i wilcoxon). conclusion: perioperative tnfa and il-6 were shown to correlate significantly with preoperative organ function, apache ii and the severity of sepsis. these results could help to define patients that might benefit from further therapeutic strategies, e.g. antibody administration. department of surgery, university vienna, akh wien, wahringer gurtel 18-20, 1090 wien. aim of the study: the purpose of this pilot study was to establish and to prove a standardized reproducible animal model of intraperitoneal sepsis induced by e.coli-endotoxinaemia in lew.lw-rats in order to investigate early immunoserological responses to find a mediator based evaluating system of peritonitis sepsis. materials and methods: in 30 lew. lw-rats, diffuse peritonitis was induced by intraperitoneal injection of a mixture of e.coli (khu +) and autogenous haemoglobin solution. in the control animal group (n= 12) an intraperitoneally injection of physiological saline solution was done. blood samples were obtained by heart puncture after 6 hours. stastistieal calculations were performed on a personal computer with the spss programm vers. 4.01 (correlation with pearson's r, mann-whitney-u-test, descriptives statistics, discriminant analysis). results: in contrast to the sham treated rats, the peritonitis animals showed significant differences in the concentrations of endotoxin, interferon-gamma (wn-y), the pteridin derivate biopterin and serum pla2-activities [endotoxin range from 0.064 eu/i, sd=0.19 to 102.38 eu/1, sd-145.4 (p < 0 0001), ifn-¥ levels, range from 147.9 pg/ml, sd-141.6, to 4591 pg/ml, sd=6541 (p < 0.0001), circulating pla2-activities range from 116.2, sd=45.4 to 185.4 u/1, sd=105.4 (p < 0.01) and biopterin range from 54.4 nmol/l sd=17.2 to 127.8 nmol/l, sd=76.8 (p < 0.001)]. for the peritonitis group we found strong correlations between the degree of endotoxinaemia to elevated levels of ifn-'~ (rp = 0.72, p < 0.0001) and bioptefin synthesis (rv= 0.82, p < 0.0001). the increase of ifn-t levels was correlated to the regulatory synthesis of biopterin (r = 0 73 p < . .. p • , . 0.0001) and to the pla2-actwtues (rp = 0.50, p < 0.005). the biopterin synthes~s correlates slightly with the pla2-actn,ities (rp= :0.38; p < 0.05). using the para, meters of endotoxin, ifn-y levels, biopterin and the pla~ -activities only, the statistical procedure of the linear discriminant analysis makes it possible, to distinguish between non-septic animals and septic animals correctly at a rate of 87 %. anaerobes were found in 53.4%, anaerobes were isolated in 22.8%. there were aerobic and anaerobic associations in 12.1% and microflora was not found in 11.6% of the cases. express method of anaerobes discovering let to receive information on 1 -3 days early than in generally accepted nethods. intraaotal transfusion of oxygenate blood and laser irradiation of blood reduces the duration of anaerobic sow, disminishes intoxication and accelerate the patients recovery. patients with abdominal sepsis are subject to long periods of hospitalization and high associated morbidity and mortality rates. this category of patients is thus consuming extensive facilities and costs. as the age-related outcome of abdominal sepsis is not fully known, the aim of the present study was to investigate abdominal sepsis in the elderly. out of 166 patients with abdominal sepsis treated at the surgical intensive care unit during a 10-year period, 72 (43 %) had an age of 70 years or more. 28 were women and 44 were men, a sex distribution not differing with patients younger than 70 years. the patients were scored according to apache ii and septic severity score (sss) upon arrival to the intensive care unit. bacterial cultures, the occurrence of organ failure, hospitalization and outcome was noted. in median two operations were performed for both "younger and elderly" patients. the median time of hospitalization in the elderly was 29 (-183) days including in median 10 days in the icu. figures in patients less than 70 years of age were comparable (34 (-293) days out of which in median 10 days in the icu). apache ii and sss-scores did not significantly differ (14.2 vs 13 and 23.3 vs 26.5, respectively), between the groups. neither did the incidence of organ failure differ (57/73 vs 77/94). however, the incidence of multiple organ failure was significantly lower in elderly patients (21/72 vs 42/94 (p < 0.001)). the mortality rate, however, did not differ between the groups (21/72 vs 26/94). in conclusion, severe abdominal sepsis in the elderly was not associated with an increase in mortality, incidence of organ failure or hospital stay. with the help of light transmissional scanning electron microscopy morphology of erythrosytes of peripheric blood was studied in patients with different stages of diffuse peritonitis before and after intravascu!ar irradiation of blood with heliun-neon laser. peritoneal morphology was investigated in patients who died from peritonitis, it was established that in all phases of peritonitis occured stomatocytoric and echinocytoric transformation of erythrocytes which progressed simultaneously with increase of intoxication. it combined with strongly pronounced vessels variability of microcirculatory peritoneal bed which displaied by erythrocytes aggregation, stasis and microtrombogenesis. in intravascular laser irradiation of blood number of erythrocytes which underwent to stomatocytoric and echinooytorie transformation was lower than in patients without laser irradiation. it indicated that the intravascular irradiation of blood with helium-neon laser can prevent development of severe alterations of rheological property of blood and consequently variability of microcirlatory peritoneal bed in patients with diffuse peritonitis. abdominal sepsis is still associated with high morbidity and mortality rates, frequenfly caused by multiple organ failure. it has been reported that changes in capillary permeability play a role in the pathogenesis of multiple organ failure. the present study aimed at evaluating the influence of intraabdominal sepsis induced by cekal ligation and puncture on capillary permeability in multiple organs and tissues. 96 adult male sprague-dawley rats were subjected to laparotomy with separation of the cekum (sham operation) or induction of intraabdominal sepsis by cekal ligation and puneatre (n--48 in each group). at 3, 6, 12, 24, 48 and 72 hours (n=6/timepoint), the animals were evaluated concerning mortality and capillary permeability as determined by the passage of :25i-labelled albumin from capillaries to the peritoneum, the proximal and distal small intestine, cekum, colon, spleen, kidneys, lungs. the mortality rate in rats with intraabdominal sepsis was 33 % both at 48 and 72 hours. capillary permeability in the peritoneum, cekum, colon and kidneys significantly increased from 6 hours and on in rats with intraabdominal sepsis. in septic animals, capillary permeability in the lungs and spleen increased from 12 hours and on and in the proximal and distal small intestine from 24 hours and on. different types of alterations in capillary permeability seem to appear: 1) a temporary short increase e.g. in the proximal small intestine and spleen; 2) a temporary longer increase e.g. in the colon and kidneys; 3) a persisting increase e.g. in the peritoneum, cekum, distal small intestine and lungs. we conclude that experimentally induced intraabdominal sepsis induces early alterations in capillary permeability in multiple organs and tissues. such changes may contribute to explain the development of sepsis-induced multiple organ failure. despite a number of significant advances in the care of burn and non-burn traumatic injury, infection and sepsis remain major causes of morbidity and mortality. the severe immunosuppresslon often seen in patients with severe trauma or large burns may predispose these patients to life threatening infections. included among the many immune alterations are changes in the functional capabilities of neutrophlls (pmns). we have examined the expression of the p 2 integrins (cd1 l a, b,c/cd18), and the fc'?r (cd16, cd32, and cd64), as well as several functional parameters, on pmns from thermal and non-thermal traumatic injury, pmns were obtained from patients sustaining severe trauma (initial apache ii score >10) or thermal injury (>30~ total body surface area, 15% full thickness), and healthy controls. the expression of cd11 b and c and to a lesser degree cdi 1 a was significantly reduced on pmns. the expression of cd16 and cd32 but not cd64 was also significantly reduced. pmns displaying this reduction in receptor expression have a significantly reduced ability to phagocytose bacteria and undergo the oxidative metabolic burst response. thermal and traumatic injury result in global reduction in the expression of 132 integrins and for which may lead to decreased functional capabilities, these abnormalities may in turn account at least in part for the increased rate of infection in these patlems, institute, dept. of surgery, 2~1 8ethesda ave, cincinnalt, oh, usa, 45267-0s5b, antibiotic-phagocytic cell interactions: their effect on endotoxin release. c g c-emmet1, dep[baeteriolog.z, univer_sitv of glasgow, scotlan~_d increasingly it is recognised that pathogenic bacteria are capable of surviving intracellularly within phagocytic cells in addition to their capacity to produce disease whilst in the extracellular milieu. as well as providing protection from certain antibiotics which fail to penetrate the phagocyte, such intraceltular bacteria may be transported from the initial site of infection to a distant more vulnerable body site wherein they may proliferate. it is also known that some antibiotics are capable of becoming concentrated within phagocytic cells mid displaying bioactivity therein. such bioactivity might be responsible for the release of endotoxia #orn gram-negative bacteria which when liberated from the celt could ~gger the cytokine cascade. anfib,.'otic-induced damage to the ultrastructure of bacteria can also occur when the target bacteria are exposed to low (sub-mic) concentrations of certain drugs. such bacteria may present quite altered surface components m host-defense cells as well as releasing biologically active ceil wall components such as endotoxin. the nature of these interactions at the cellular level as well as the consequences for the host will be discussed. new jersey medical school: umd, newark, nj 07107 a technique of physiologic state classification has been developed based on the m~itlvariable analysis of patient derived data sets of seventeen physiologic variables. these multivariable data sets obtained from critically ill patients requiring intensive care, were aormallsed by the mean and the standard deviation of recoverin~ trauma patients who were not critically ill, the resulting normalized seventeen variable sets were then clustered. seven independent data groupings were developed. the normal stress response hyperdynamic state seen post-trauma and in compensated sepsis (a stets)/ metabolic insufficiency seen in septic decompsnsation (b stste}; early (c,) and late (e2) respiratory insufficiency associated with ards; cardlogenlc dscompensation (n state); post-trauma hyvolemla without shock (r stats). the stats closest to a new patient's values allows patient classifi0atlon with regard to his previous physiologic state. classifying observations f~om patients who lived or died who fell into these physiologic states enables a probability of death (p death) to be obtalned. utilizing this criteria for the staging of severity in 44 recent trauma patients the physiologic states accurately and significantly predicted the likelihood that the patient had an increased circulating level of the eytoklnes tnf and il-1. the probability of death (p death) as well as the cytoklne levels appear to be a function of the physiologic b state with the highest levels being seen in the b state of metabolic insufficiency and the c~ state of oombined respiratory and metabolic insqffioienoy characteristic of septlc ards. the increase in the magnltude of metabolic abnormalities associated with the transition from non-sepsls to septic a, septic b, or septic c z states was associated with an increasing probability of death (p denth)(mean a state =.28, mean b state = .57, mean ~ state = .61). the accuraay of this estimate was prospectively analyzed in this group of 44 m~itlple patients of whom 72% had sepsis and 36% had ssptlo ards. the 22 survivors had a mean p death of 0.39 and the 22 deaths had a mean p death of 0.63. the severity of post-trauma sepsis can be quantified by probability analysis and stra~ifie~ by physiologic state. serologic tests have not been extensively tes'~ed in surgical patients but seem to be of limited value. we use nystatin as the main form of chemoprophyhxis. patients "~'ith signs of infection who do not rapidly improve with antibacterial therapy are candidates for anti-funsal therapy, amphoteradn b remains the first llne of therapy although combination therapy '~'ith flueonazole is use;l with increasing freque~;c)', the recovery of c~dida from an antra-abdominal site represents a challenging problem, anti~ngal therapy in such patients depends on the underlying disease, the nature of the infected material and overall patient risk. role of neural stimuli and pain principles and practice of anesthesiology effect of combined prednisolone, epidural analgesia and indomethacin on the systemic response after colonic surgery arginine: biochemistry, physiology and therapeutic irnplications immunosfimulatory effects of arginine in normal and injured rats arginine stimulates lymphocyte immune response in heahhy humans rote of arginine in trauma, sepsis and immunity arginine enhances wound healing in humans if labrecque t, gv campion t, and the rhll-lra phase i//sepsis syndrome study group the cleveland clinic foundation a murine-anti-human tnf-monoclonal antibody known as cb6 was the first anti-tnf mab which was studied in a phase ii multinational trial in the treatment of patients with severe sepsis.this was an open-label, dose-escalation trial consisting of 80 patients who were enrolled into one of four treatment groups: (1) 0.1 mg/kg of anti-tnf mab, (2) 1.0 mg/kg, (3) 10 mg/kg or (4) 1.0 mg/kg at study entry and the second dose 24 hours later. the small sample size in each group (n=20) precludes detailed statistical inference in this study. nonetheless, a considerable amount of useful information was obtained from this investigation. irst, this study demonstrated the clinical feasibility of specific anticytoldne therapy in septic patients. second, the measurement systemic levels of tnf proved to be an elusive target; interleukin-6 may prove to be a more useful indicator of cytokine activation. third, immunologic reactions including tnf: anti-tnf mab immune complexes and human anti-routine antibodies were frequently found in these patients. despite their apparent lack of overt toxicity in this study, these immunologic reactions may complicate this form of anticytokine therapy. additionally, the potential benefits of anti-tnf mab therapy occur within the first 48 hours of therapeutic administration in these septic patients. infecting organisms differ in their potential to induce tnf in vitro and these differences correlate with circulating tnf levels observed in septic patients. rapid methods to define those patients most likely to respond to anticytokine therapy are needed to determine the ultimate therapeutic potential of these agents in clinical medicine. wherry, j., abraham e., wunderink r., silverman h., perl t., nasraway s., levy h., bone r., wenzel r., balk r., allred r., pennington j. and the tnfa mab sepsis study group.tnfa mab (bay x1351) is a murine monoclonal antibody raised against human tumor necrosis factor. tnf~ mab has been shown to reduce morbidity and mortality in animal models of septic shock and has been safely administered to septic and non septic patients.to evaluate the efficacy and safety of tnf~ mab in patients with sepsis syndrome, a prospective, multicentered, double-blind, placebo-controlled trial was conducted in 31 hospitals in north america. patients were prospectively stratified into shock or nonshock groups and then randomized to receive a single intravenous infusion either of 15 mg/kg tnf~ mab, 7.5 mg/kg tnf~ mab or placebo (0.25% human albumin).patients received standard aggressive medical/surgical care during the 28 day post dosing period.the three treatment arms were well balanced with respect to demographics, apache ii score and other parameters. for all infused sepsis syndrome patients, those who received tnf~ mab had slightly reduced 28 day all cause mortality compared to placebo. among shock patients there was a more pronounced trend towards efficacy at day 28 post dosing with lower mortality rates in both active treatment arms. among nonshock patients tn~ mab did not appear beneficial. the initial clinical experience with a chimeric anti-tnf monoclonal antibody, ca2, was undertaken in septic patients. the objectives of the study were to determine the safety, pharmacokinetics and effects on cytokine levels of ca2. as a single infusion or in combination with ha-1a in septic patients. the study was conducted with the intent to progress to an efficacy trial based on the information collected.the trial was conducted in three stages. stage 1 was an open label trial in which 4 groups of 5 patients each with the clinical diagnosis of sepsis received ascending doses of ca2 (0.1, 1, 5, 10 mg/kg). stage 2 was a randomized, double blind study in which patients received a single dose of ha-1a (100 mg) and placebo or one of 3 doses of ca2 (1,5,10 mg/kg). stage 3 was a randomized, double blind study in which patients received a single dose of placebo or one of 3 doses of ca2 (0.1, 1, 10 mg/kg). in addition to usual laboratory tests, the following assays were performed: chimeric anti-tnf concentration, anti-chimeric antibody, endotoxin, tnf, il-1, and il-6 levels.a total of 141 patients were enrolled from 13 clinical sites (20 in stage 1, 60 in stage 2 and 61 in stage 3). primary analyses were performed on patients in stage 1 and 3. there were 65 patients who received ca2 exclusively and 16 patients received placebo. administration of ca2 was well tolerated at doses up to 10 mg/kg. no patient discontinued treatment due to adverse events. human anti-chimeric antibody responses were positive in 61% (30/49) of evaluated patients. mean cma × and auc increased proportionally with increasing doses of ca2. the mean half-life was -70 hrs (58-96 hrs). a dose related decrease in tnf concentration was observed 1 hr post infusion of ca2. tnf is considered to be one of the central endogenous mediators for the inili'ation of the pathophysiological changes in patients with sepsis and septic shock. high tnf levels were demonstrated to correlate with patient outcome. blocking or neutralising tnf with specific antibodies was effective in preventing death in some animal modets of sepsis. in a placebo controlled prospective randomized study we tested the mur~ne derived antibody mak 195f. it is a f(ab')2 fragment. the fragment rather the complete antibody was selected in order to reduce the potential immunogenicity and to facilitate tissue penetration. 122 patients with severe sepsis or septic shdck were enrolied in the study, three different doses of mak 195f or placebo were administered (0,1; 0,3 and i mg/kg) over a perid of 72 hours in random order. the patients were evaluated for side effects, hemodynamics, organ dysfunction, cytokines (il 6, il 8 and tnf), and outcome. at this time only an interim analysis of 60 patients is available i indicating that mak 195f in all 3 dosage groups resulted in a decrease in il 6. this contrasted to a further in crease of il 6 in the placebo patients. no serious side effects have been reported so far. a more detailed analysis on all 122 patients in the study will be presented and discussed.$152 s154 staubach,k.h., otto, v., kooistra,a,, rosenfeid,j.a., bruch, h.p., univ. lfibeek, germany once endotoxinemia occurs in sepsis a vieieus cycle with translocation of et can be established. increasing the clearance capacity for et would therapeutically be the ulimate aim. we developed a new et on-line adsorption (ad) system in whole blood by means of polymyxin b (pb) coupled eovalently to a matrix (acrylic particles) via a 13 atom-chain spacer. the detoxification capacity was 150 ug[et/ml column material. the biocompatbility resulted in 90~ platelet recovery. the column contained 7 ml of admaterial and was sterilized by high steam autoclave, anticoagulation was achieved by heparine 1.000 iu/h in the inflowline after bolus injection of 5.000 iu. hp was performed on 8 pigs at a rate of 50 ml/min by means of a roller-pump until the animals succumbed (h). 8 animals served as controls (c). serum et levels rose from 3.40 pg/ml to 74,9 pg/ml after 5 hours in the c and from 2.77 pg/ml only to 28 pg/ml in the h group after 7 hours whieh was highly significant. survival time could be extrended from 216 to 313 min. results are listed in the following l. blinzler, p. zaar, m. leier, r. b(2rger, d. heuser clinic of anaesthesiology , city hospital nuremberg, germany sepsis and multiple organ failure (mof) are still related with poor prognosis inspire of pharmacological and technical progress. impressed by revealing reports about blood purification the continuous veno-venous hemofiltration (cvvh) was used as supporting treatment beside the critical cam basic therapy of mof. from 1989 to 1991 78 consecutive patients were treated by cwh. mof was caused by hemolrhagic-traumatic noxa in 21°,4 and by septic-toxic event in 79%. all patients required mechanical ventilation (fio2>0, 5) . 86°4 showed hyperdynamic shock. 85% had renal and 53% hepatic failure. medium appache ii score amounted to 29,8 points. cvvh was performed in postdilution mode with a polyamide membrane (fh 66) and high volume exchange (50 l/die). anticoagulation was done with heparin. hemofiltration in mof was installed, when critical cam basic therapy including adequate respiratory and hemodynamic management, pamnteral nutrition, antibiotic treatment, etc., failed to stabilize organ functions. during consequent application of cvvh most of these patients showed improvement of their clinical course. pulmonary stabilization was seen in 71%, hemodynamic in 68% and renal in 66% of the cases. 42% of the patients survived and were discharged from hospital. 10 of 45 non-survivors (58%) died because of fatal mof within 24h after admission to icu. patients with early application of cvvh in mof showed a better survival rate.mediators of mof, i.e. products of the complement cascade measured in blood and nitrafiltrate by elisa, were partially removed by cvvh. the testing ultrafiltrate by hplc demonstrated decreasing spikes ofpolypeptides during hemofiltration. mof seems to be generated by cascade-activation of immune competent cells and plasmatic mediators (e.g. bmdykinin, eicosanoides, cytokines, anaphylatoxins, etc.). therapeutic approaches aim to inactivate or eliminate single substances. cwh with high-flux membranes in combination with high-volume exchange allows elimination of many mediators with different molecular weight and therefore may contribute to improve the prognosis of mof. other significant advantages of this teqalnique like adequate nutrition, optimized fluid balance and control of body temperature should not be negicctod. introductioni pseudomonas (p) aeruginosa has to be considered an important pathogen of nosocomial pneumonia and septic organ failure. the lung seems to be the predominant target organ for the pore-forming p. aeruginosa cytotoxin, thus inducing microvascular injury. with respect to therapeutical consequences, the potential protective effects of paf-antagonist (web 2086), cyelooxygenase inhibitor (diclofenac) and specific and unspecific antibodies on cytotoxin-induced pulmonary vascular reaction and mediator release were studied in the isolated perfused rabbit lung. methods: cytotoxin (6p_g/ml) was administered into the perfusion fluid in all groups, either in the absence of inhibitors (n=6), or after pretreatment with web 2086 (5xl0-gm, n=6), or diclofenac (10#g/ml, n-6). furthermore, the application of specific antitoxin (mg/ml, n=6) was tested in comparison with the unspecific immunoglobulins (venimmun®, behring, 7.5 mg/ml) (n=6) and the combination of immunogiobulins, web 2086 and diclofenac (n=6). six experiments without toxin served as controls. the arterial pressure mad the weight gain as an indicator of edema formation were continuously monitored during the three hour peffusion phase. arachidonic-ucid metabolites, as well as lactate dehydrogenase (ldh) and k + concentrations were determined at 30 rain intervals. results: cytotoxin caused a gradual increase in pulmonary arterial pressure, reaching a maximum value of 2.5 times higher than the control, starting after 1 min and a delayed onset of edema formation resulting in a mean weight gain of 20 g after 120 min. this was paralleled by a significant increase in prostacyclin generation and a continuous release of k + and ldh. thromboxane synthesis exceeded about 10 times that of controls in the toxin treated lungs. pretreatment with web 2086 or diclofenac significantly attenuated the pressure response and edema formation evoked by cytotoxin. the addition of the unspecific immunognbulin preparation alone induced a transient pressure increase within the first minutes, but mean values remained below those of the cytotoxin group in the continuing observation period. mmost complete inhibition of the pressure reaction, the edema formation and the metabolic alterations was achieved mainly by the combination of immunoglobulin, web 2086 and diclofenac and to lesser extend by the specific toxin antibody. conclusion: the current results point towards the crucial role of paf and aa-metabolites as mediators of cytotoxin induced microvascular injury. the systemic or local application of cytotoxin antibodies or even unspecific immunoglobolins in combination with paf-antagonist and diclofenac appears to be a promising therapeutic approach in the case of infection with cytotoxin-preducing strains. cytokines have long been shown to be of particular importance in the metabolic derangements occurring in lps-induced shock. recent studies strongly imply the involvement of platelet aggregating factor (paf) in the pathogenesis of gram-negative bacterial sepsis. an autocatalytic feedback network has been postulated to exist between paf and tumor necrosis factor (tnf), a key cytokine involved in septic metabolic cascade, leading to an uncontrolled amplification of inflammatory mediator release. we have previously shown that st 899 (4-n,n,n trimethylammonium-(r)-3-isovaleroyloxy-butanoic acid z-3-(5-chlorphtalidiliden) ethyl ester bromide) was quite effective in inhibiting the "in vitro" binding of 3h-paf (ki=7.5x10 -8 m) to rabbit platelets. the present study shows that pretreatment of c57bl/6 mice with st 899, administered by different routes, dose-dependently and significantly reduces the lethality induced by endotoxin (e.coli 055:b5 injected at 30 mg/kg intraperitoneally). very interestingly, st 899 administered at the same doses as above (i.e. 1.25, 2.5, and 5 mg/kg body weight) results to be significantly effective in reducing the endotoxin-induced release of serum tnf. the reported dual activity of st 899 (i.e. paf antagonism and decreased circulating tnf levels) may turn out to be greatly beneficial, in combination with current therapies, in the treatment of diseases that involve overproduction of tnf and paf such as septic shock. introduction: recently, we reported that prophylactic whole body hyperthermia (42.5°c) induces heat shock protein ('asp) 72 and increases smvival 2-4 fold in a mouse endotoxin model (am. j. physiol. in press). other investigators reported that prophylactic pharmacologic induction of hsp-72 by sodium arsenite improves survival in a rat sepsis model (abstract a95 am. rev. resp. dis. vol. 147, 1993) . the effects of heat are complex and in addition to formation of lisp-72 include release of cytokines, changes in cellular ph etc. thus, the protective mechanisms of heat may differ from those due to pharmacologically induced . the purpose of this study was to compare the protection of heat vs the protection of pharmacologically induced hsp-72 in a mouse endotoxin model to determine if different protective mechanisms were likely to be involved.. i%'lethods: both sodium arsenite (10 mg/kg) and ethanol (400 ~1 of 39% ethanol) caused marked induction of hsp-72 in lung, gut, kidney, and liver, which was comparable to heat-induced hsp-72. female nd4 mice weighing 22-25 gms were pretreated with arsenite or alcohol 8 hours prior to challenge with escherichia coli endotoxin (-ld 80) and survival was compared to control mice. results: survival at 24 hrs. for arsenite treated and alcohol treated mice was 96% and 88% respectively and was statistically different from the 40% survival for control mice. (p<0.01) (n=25 mice per group). however, at 7 days post endotoxin, there were no differences in survival in the 3 groups, i.e., ~ 4% survival for all 3 groups. in contrast, the protective effect of hyperthermia remains present at 7 days, i.e., ~ 70% survival vs 20% survival control. conclusion: the protective effect of heat is probably due to other factors such as the effect of hyperthermia to release il-lc~ and is not due solely to hsp-72 formation. it was the aim of the study to examine whether bacteria play a causative role in the pathogenesis of anastomotic insufficiency following gastrectomy in man.the study was carried out in form of a prospective, randemised, double-blind, multicenter trial. primary endpoints were the rate of anastomotic insufficiencies, infectious-and uncomplicated postoperative courses. all pat. received a periop, i.v. prophylaxis with cefotaxim. identical numbered vial either contained placebo or polymyxin b, tobramycin, vancomycin and amphotericin b . the vials were administered 4x per day from the day be ~ fore the operation until the 7th postop, day. insufficiencies were detected by gastrographin swallow and recorded by x-ray on day 7 postop.. evaluation was carried out on an "intention to treat'basis. statistical analysis was done with the pearson's chi square and fisher's exact tests~ results: interim analysis was carried out in 3/93 after 137 pat. had been recruited. along with a significant reduction of s.aureus and enterobacteria there was a reduction in the rate of anastomotic insufficiency of the esophago-jejunostomy from 10.6 % in the placebo-group to 3.5 % in the treatment group. the difference was not yet significant. the rate of nosocomial infections (e.g. respiratory tract infection and uti) were significantly reduced from 33.3 % in the placebo-group to 16.4 % in the treatment-group (p ~ 0.0281;fisher's exact test). in march 1994 final results with more than 200 patients will be presented for the first time. (= po2 <80 mm hg, b s-creatinin >2 mg%). respiratory insufficiency was the most frequent systemic complication followed by sepsis and respiratory insufficiency. etiology of pancreatitis and initial serum increase of pancreatic enzymes predicted neither complications nor outcome. only 2 of 14 deaths occurred during the 1st week, all other deaths occurred late (after 4-12 weeks), generally as the consequence of septic complications and multi-organ failure. high levels of crp were correlated with a compliacted course and a fatal outcome. although same cytokines (e.g. 11--6) were found increased in severe disease, the predictive value of these markers was not better than the combination of ctinical scores (ranson, imrie, apache ii) with gt or crp. conclusions: intensive care medicine can often control the inital shock situation in severe pancreatitis. thus. only 15% of deaths today occur eady in the course of the disease, whereas this percentage varied between 40-70% just 10 years ago. nowadays, most deaths are caused by late septic complications and multi-organ failure. ranson-and ct-scores as well as serum crp predict a course with systemic complications; they are less helpful for prediction of sepsis and late mortality. it is doubtful whether measurements of cytokines will help to better predict the late outcome. as yet, only careful and continuous monitoring of patients (e.g. by apache scores) may help to early identify those who develop septic complications and multi-organ failure. the classic description of severe acute pancreatitis has hinged upon the release of large volumes of activated enzymes into the peritoneal cavity and thertce the lymphatics and blood stream. these activated enzymes escape from the pancreas due to disruption of cells with associated ischaemia and occasional infarction of tissue. for 20 to 25 years it has been postulated that the bocly's defence system to activated pancreatic enzymes required supplementation iu the form of anti-protease support either in the vascular space or in the peritoneal cavity. all controlled studies have shown that this is either impracftcal or unnecessary.hore recently release of a large number of cytokines from monocytes, macrophages and neutrophils have been considered to be harmful to the body and various agent~ which oppose the action of tnf alpha, paf and similar cytokines are being examined in experimental anim~is and certain clinical trials, it has clearly been shown that higher levels of cytokines are released in the patients with objectively graded severe acute pancreatitis than in those with milder disease. we now seem to be moving into an exciting phase of potentially beneficial therapy in acute pancreatitis which has had no specific effective therapy through studies utilising aprotinin, gabexate mesilate and fresh frozen plasma. inflammation cascades may play a role in the pathogenesis of acute pancreatitis. to evaluate the status of the cellular immune system we examined serum concentrations of immune activation markers in 24 patients with acute pancreatitis (14 males, 10 females; median age: 56 years, range: 32-81 years). concentrations of neopterin, serum soluble tumor necrosis factor receptor (stnf-r) and serum soluble intercellular adhesion molecule type 1 (slcam-1) were determined using immunoassays (henning, bender, t cell sciences). 13/14 had increased concentrations of stnf-r compared to the 75th percentile obtained in healthy controls (>4.2 ng/ml), and 9/24 patients had increased neopterin (> 8.7 nmol/i), 9/24 presented with elevated slcam-1 (>440 u/i). all patients with increased neopterin also had increased stnf-r, 4 patients had concentrations of all three markers outside the normal range. there existed a significant correlation between neopterin and stnf-r (rs = 0.728, p < 0.001 ). weak associations between age and stnf-r (rs=0.435, p=o.05) or neopterin (rs=0.456, p = 0.044) were also found. our results demonstrate activation of the cell-mediated immune system taking place in a sub-group of patients with acute pancreatitis. the finding of increased neopterin and stnf-r levels implies that activated monocytes/macrophages are involved in the pathogenesis of the disease. further data are necessary to evaluate potential associations between changes of marker concent-rations and the course of the disease. pancreatic injury after heart surgery was reported as soon as 1970 ( 1,2) and characterized by increased serum or urine amylase levels (in about 33% of patients) in the 5 fi~t postoperafi.'ve days. this pancreatic injury, which sometimes led to acute pancreatitis, was atreaay at~buted to inappropriate perfusion of this organ. in the 198ffs, 3 studies were published dealing with pancreatic suffering alter heart surgery, in large series of patients, concluding ~n~at panc~a~c injury (with a low incidence of pancreatifis) is more common than previously recognized and is a potential source of complication after camliac surgery (3, 4, 5) . in a recent study (6), evidence of pancreatic cellular injury was found in 80 out of 300 patients undergoing cardiac surgery, with 22 out of these 80 patients presenting abdominal signs or symptoms and 3 developing severe pancreafitis. this injury was associated w~th preoperative renal insufficiency, valve surgery, ~..stoperalive hytxxension, calcium administered periopuratively and length of bypass. we studied 300 patients submitted to cardiopulmunary bypass (cpb) for heart surgery and used the measurement of un:~sin, pancreatic iso-amylase and lipase in plasma for biochemical characterization of pancreatic cellular injury. blood samples were obtained before surgery, directly aller surgery (return to inte~ve care unit), 8 hours alter surgery and in the folfowing days alter surgery (days 1, 2, 3, 4 and 8). computed tomography scan of pancreas was performed in patients presenting hi~ levels of amylase on day 1. we measured abnormal levels of trypsin and pancteatic iso-amylase in 30% of patients and observed 2 simultaneous releases of these 2 enzymes, the fi,'st one in the 24 hours after surgery and the second more intense from day 4 and pa~icularly on day 8 after smgery. this second release was concomitant with abnormal levels of llpase. these biochemical observations were accompanied by radiological and clinical signs of pancreatic injury in about 2% of our patients : pancrealic abnormalities were revealed by scan in 7 patients and acute pancreatitis in i patient. more pronounced pancreatic suffering was observed in patients undergoing valve replacement than in patients undergoing coronam-anrtic bypass grafm~g. analysis of trypsin and pare're, tic 1so-amylase are sw.cific of pancreatic cellular injury and their simultaneous ir~rease in plasma alter cpb in our padents confirms the presence of an exocrine pancreatic injury. the presence of a simultaneous peak of lipase mcaezse~ the specificity of overt pancreatic injtu 7 diagnosis. the precise cause of th/s injury could he related to hypoperfnsion leading to ischemic injury of foe splancbnic area, pancreas being largely sensible to hypoperfnsion (7). this hypoperfosion could he responsible for the ftmt release of pancrealac enzymes observed in our patients and would contribute to the deterioration of other organs leading to an inflammatory reaction developing in the following days and responsible for the second release of pancreatic enzymes observed in our patients. patients with necrotizing pancreatitis show a heigh rate of pulmonary, renal and septic complications, whereas the course in acute interstitial pancreatitis is generally very mild. we have prospectively analysed the value of endotoxin, interleukin-6(il-6) and transferrin in compare with c-reactive protein(crp) for the early assessment of the severity of acute pancreatitis. patients aud methods: the values of endotoxin(measured by limulus-lysate-test), ii-6(elisa), transferrin and crp (nephelometry) were analysed daily along the first i0 days of hospitalisation by 28 patients with acute pancreatitis admitted to our hospital from 4/1991 to 4/1993 . it was judged whether the patients have either interstitial (aip) (n=9) or necrotizing (anp) (n=lg) pancreatitis. 5 patients with anp have died during the course of pancreatitis (mortality=17.5%). results: 1-severity o~ pancreatitis: signifcant differences (p<o.05, mann-whitney test) could be calculated between aip and anp for endotoxin, il-6, transferrin and crp during the i0 days of monitoring. 2-mortality: except of il-6 on days 2, 3, 4 and 5 after ad-mission there were no significant differences between the values of analysed parameters in patients who survived and those values in patients who died. 3-organ failure: the patients with pulmonary, renal or multiorgan failure have significantly heigher values of endotoxin, il-6 and crp along the i0 days of monitoring in compare with those patients without an organ failure. conclusions: endotoxin, il-6 and transferrin, an important endotoxin carrier, are promising parameters to differentiate between the severe and mild form of pancreatitis comparable with crp. this parameters may be helpfull in the early clinical assessment of patients with acute panereatitis. the determination of cytokines may elucidate the pathophysiologic mechanisms that produce early systemic complications in acute interstitial (i) or necrotizing (n) pancreatitis (ap). the appearence of respiratory insufficiency (ri) is used to appraise the early systemic complications and is compared to the results of cytokine blood levels. in a prospective clinical trial from 10/92 -8/93, 23 patients with ap were recruited and blood samples taken for cytokine determination with commercial elisa-kits. the differentiation between i (n=12) and n (n=l 1) ap was made through ct-scan in all and laparotomy for drainage and necrosectomy in 13 patients. the etiology of ap was gallstones in 9, alcohol abuse in 7, hyperlipidemia in 2 and other or unknown causes in 5 patients.five of 11 patients with nap died early (n=l) or of late septic complications. none died of iap.the peak of cytnkine level in the first three days of hospitalisation was used for calculation. ri was diagnosed in the first week of hospitalisation, if oxygenmask or iutubation for at least 3 days was nescessary to treat arterial hypoxemia. for statistical analysis u-test of wilcoxon, mann and whitney was applicated.the il-6 concentration in blood reached up to 2600pg/ml in the first days depending on the severity of ap and dropped to almost zero in the next days, independently of the clinical course. the differentiation of i-versus nap, using a cutoff-line of 600 pg/ml was correct in 20 patients (87%, sensitivity (se): 82%= specificity (sp): 91%, (z:0,001). high levels of il-6 over 500 pg/ml correlated well with the prognosis ifri in the first week (accuracy: 87%, se: 80%, sp: 100%, c~: 0,001). the blood levels of il-8 reached a maximum of 1381 pg/ml in the first days, depending on the severity of ap, and showed a correlation to the clinical course in the following days. the peak of 1l,-8 blood levels indicated correctly the severity of ap in 18 out of 23 patients using a cut offtevel of 200 pglml (accuracy: 78%, se: 82%, sp: 75%, ~: 0,01). there was no correlation between cytokine blood levels and mortality, also there seemed to be no correlation between the levels of il-6 and il-8. in the bloodsamples of five patients with i-or nap no c~-tnf was detectable.the blood levels of il-6, and to a lesser extent of [l-8, can predict the severity and early systemic complications of ap in men. the excessive rise of cytokines can be explained by the stimulation of immunological cells ( macrophages, lymphocytes and endothelial cells) in the course of ap. objectives: in an opossum model, ligation of the sphincter of oddi or separate ligation of the bile and the pancreatic duct together leads to severe haemorrhagic pancreatffis while single ligation of the pancreatic duct causes only an exocdne atrophy. since bo has been discussed to be a contdbutor t¢, res dysfunction, this may depdve the organism of a potent defensive mechanism thus promoting the course of pancreatitis. the present study wa,,; carded out tc develop a new method for measuring dynamic liver res func.. tion and to test the hypothesis that bo causes a res dysfunction. material & methods: 18 opossums were randomly placed in three groups. all received a central venous line. after midline laparotomy, a specially designe0 tourniquet was placed around the common hepatic duct above its junction with the pancreatic duct (group a, n=6), around the pancreatic duct (group b, n=6) or around both ducts (group c, n=6). after one week, the tourniquets were closed. using a scintillation camera, the liver uptake of nanocoll, a 99mtcalbumin colloid with a diameter of 25 nm, was measured before, 2,5 and 6 days after the obstruction. the curves were analysed by a computer and two parameters (r, s) were calculated using natural logarithmic regression. as a common measure for res function, the corrected granulopexic index (a) wa,,; determined, after day 6, the pancreas of each opossum was searched for necrosis by morphometdc methods. results: all animals survived till day 6. the opossums in groups a & 8 had a macro-and microscopic normal pancreas, while those in group c showed a severe hemorrhagic pancreatitis. the granuinpexic index showed a significan~ change to the worse starting at day 5 in group a & b (p<o,05) and at day 2 it~ group c (p<0,ol). at day 6, res function in group c was significantly worse than that in group a & b (p<0,01). the regression parameter r showed no significant change in groups a & b, but a highly significant decrease in group c starting at day 2 (p<0.005), thus showing a dysfunction of the hepatic res. conclusions: obstruction of the hepatic or pancreatic duct alone does no{ result in immediate dysfunction of the hepatic res, while obstruction of beth ducts does. because only ligation of both ducts is followed by a severe hereof.. rhagic pancreatitis, res dysfunction could be an important factor in the pathogenesis of pancreatitis and resulting organ failure.logarithmic regressinrl has proven to be a valuable tool to analyse dynamic hepatic res function. (b6) in lewis rats weighing 220-250g. there were five groups: group a (n=14) were untreated controls; group b (n=7) were given isotonic saline intraperitoneally and observed for 48 hours; group c (n=7) 2 ml x 108 e. coli intraperitoneally and observed for 48 hours; group d (n=7) were given 4 ml x 108 e. coli and observed for 48 hours; group e (n=7) 4 ml x i08 e. coli and observed for 24 hours, the rats were than killed, the spleens were removed and heparinised blood taken for immunological tests. total t-ceils, helper t-ceils, suppressor t-cells, monocytes, and the interleukin 2 receptor were determined by monoclonal antibodies, indirect immunfluorescence, and flow cytometry (facs analyser 11). statistical analysis was by the t test on rank transformed data.in group b (isotonic saline) the total t-ceils increased in the blood (p=0,003) and spleen (p=0,0001 )com pared with the untreated rats (group a), in the groups with peritonitis (c, d, and e) we found highly significant rises in suppressor t-cells in the blood and spleen (both p<0,0001 ) compared with the non-infected control groups a and b. this increase was significantly higher in group d than in groups c and e. as well as other alterations, we found a uniform increase in the number of t suppressor ceils in our model of acute peritonitis that was both time and dose dependent, department of surgery, university vienna, akh wien, w~hringer gurtel 18-20, 1090 wien. d~'na~'aics of som~ l~mnunologic indices in children with abdo;~tinal shock v.z.i~ioskalenko, s.f.vesiol$,t.v.p~bina serum (iga,:~i,g, circulating imaune co!~plexes, co:apleaentary serum activity, antimesothelialvisceral and parietal antibodies) and cellular (i-l~q~hocytes, ~s/th,b-lyaphoeytes,i nlaunoleukosis to aesothelial allergens; spontaneous blast cell transformation to phytohemaaglutinin and .nesoth<-~lial ~,togen; phagooftic neutrophil activity) im:~une indices v.'ere studied in 60 children operated v:ith symi~to'as of abdo~ainal shock. ,'~!-~ of them had appendicnlar(26-1ocalized jo-diffuse, 9-~eneralized) and 5-hemoperitoni-~is (dull abdominal trau;na with injayy of 'the spleen-4, the spleen and liver-q), flo patients tjere operated on for co~iplete iieus (~-intussnsc.sption, 5-com:lissural ileus). 2he follo~'~ing serum factors: cireula:;ing im~aune complexes and ~).esothelial antibodies are the aost i~portant ~ar~ers of the abdominal shock course. :yith the progress of the process the indices increase irresponsive of t.l~e cha±.aeter of pathology. in case of ileus regress and hemoperitonotis an abrupt drop in ~he titer of anti~aesothelial antibodies is noted. in bacterial peritonitis a high ~iter of anti:aesothelial parietal antibodies in great dilutions ("+++","++++"-q ; 720) persisted even in the period of clinical recovery. cellular factors cham'acterize dfnaaics of the abdominal shock coulees less specifically llast cell transfor~aation and phagocyue neut~ophil activit 2 can inderectly characterize transition of shock f~orn reversible into irreversible phases. the past so years have brought a number of major advances in burn care. the initial advance was the discovery that burn victims required large volumes of resuscitation fluid in the initial 24 hours to maintain hsmodynamic stability, this was followed hy the dsvelopmsnt of topical antimicrohlal agents to prevent end treat infections. next, was the reporting that early excision of burn wounds, with autografting of the excised wounds was possible.finally, was the identification of the importance of aggrsssive nutritional support for the hypermstabolic burn patient.these advances have markedly increaesd survival rates for given burn mimes, s~ch that burns which would previously have been considered to be aniformly fetal, are now readily survivable.thsre remain two unsolved problems in burn care, which are limiting survivability cf burn patients.the first ie ths treatment of inhalation injury. ths sscond is how to obtain coverage of ths maaaivsly burned patient, who has limited skin graft donor sites available.this ssssion will focus on the new treatments being developed to obtain permanent coverage of burn wounds.these include the use of various tcplcal growth £aotore which can speed the rate of reepithelialization of wounds, both the beneficial and dstrimental effects of using cultured karatinooyte preparations, to obtain in excess of one squats meter of a pseudoskin from a single fifteen square centimeter biopsy of unburned skin, will be discussed, finally, the used for artificlal dermal preparations will be discussed. integumentary wound healing consists of simultaneous epithelializalion and contraction. not long ago, it was thought that healing proceeds at an optimum fixed rate and could only be delayed. the recent explosion in molecular biology and recombinant gene techniques have produced information on the biological activity of compounds previously believed to be biologically inert. many current attempts to modulate the rate of wound healing center on the topical application of various growth factors produced by integumentary cells or agents designed to modulate growth factor expression or response. epidermal growth factor (egf), plateiet derived growth factor (pdgf), transforming growth factor (tgf) and fibroblast growth factor (fgf) have been demonstrated, both in vitro and in vivo, to stimulate the proliferation and diferentiation of their designated cells types. egf and pdgf have been clearly demonstraled to increase the rate of wound closure in partial and full thickness wounds. pdgf also has demonstrated efficacy in the closure of recalcitrant pressure sores, whereas tgf increases the tensile strength of incisiona[ wounds and fgf stimulates angiogenesis. it appears that epidermal keratinocyte migration during wound healing depends on integrin-mediated interactions with compounds and ceils extracellular matrix. additionally, the activities of extracellular glycoproteins such as fibronectin, fibrinogen, laminin and thrombospondin are coordinated by multiple integrins with specific distributions and binding affinities. currently, agents which incorporate specific protein sequences essential for the interaction of the glycoproteins with the cells of the extracellular matrix are being investigated. it is has been clearly established that the rate and nature of wound healing can be influenced by the application of various agents and that sufficient quantities of these agents can be manufactured. the future challenge is to develop techniques to now the objective evaluation of the clinical efficacy of these various agents and to employ the appropriate agents in a cost-effective manner. transplantation; and 4) the immune status/manipulation of the host.multiple interactive variables will determine the effect of employing allogeneic cells and tissue in wound coverage design and a batter understanding of the dynamics of the wound-graft microenvironment will facilitate the dove opment of clinicauy beneficial graft composites. cadaver allograft skin (cas) is the "gold standard" for wound coverage, but has limitations including varied availability & quality, immunogenicity leading to rejection, & potential for disease transmission. our development of a temporary living skin replacement (tlsr) addresses these issues; the tlsr consists of highly screened human neonatal fibroblasts (hf) cultured in biobrane a, a bilayer dressing consisting of nylon mesh laminated to a thin silicone membrane which controls water-vapor transmission. studies in our lab indicated that "fresw ~ tlsr grafts reproducibly close full-thickness wounds in mice & perform better than bb controls. we studied wound adherence & structural/molecular properties of fresh & cryopreserved (cryo) tlsr. methods: tlsrs were prepared by seeding 4000/co 2 hf in bb nylon mesh in dmem. hf quickly attached to the nylon mesh & were allowed to proliferate 3-6 wks. fibroblast mitochondrial activity was assayed by mtt assay. matrix proteins were identified by immunostaining, mrnas for specific growth factors were identified by reverse-transcription polymerase chain reaction amplification, & quantitated by gel scans. bb structure was studied by scanning electron microscopy & stretching measurements pro/post cryo. 80 grafts were cryo'd in 10% dmso, quick-thawed & sutured onto 2x2 cm excised full-thickness dorsal wounds on athymic mice. 40 "fresh" grafts were similarly placed. controls grafts were cas and aeellular bb. adherence was quantitated by a device which peeled grafts from wounds at intervals following placement, using a serve motor and a strain gauge. results: postcryo, storage and subsequent thawing tile tlsrs maintained >60% cell viability by the mtt assay, indicating continued mitochondrial activity, and bb structure & stretchability were maintained. multiple matrix proteins secreted and deposited in the bb nylon mesh (types l/iii collagen, decorin, fibroneetin) were identified by specific immunostaining. growth factor mrnas in the tlsrs (afgf, bfgf, kgf, tgf~,p~,) were present in 10-10,000x higher levels in fresh/cryo tlsrs than in adult hcs. grafts adhered to wounds on mice through 40 days of followup. histologic exams on days 6-40 showed excellent vascular ingrowth and minimal inflammation. adherence of tlsrs to wounds was >cas adherence. burn wound coverage in the massively burned patient remains a difficult problem. although cultured keratinocytes have been utilized for burn wound coverage, their impact on the patient with burns greater than 80% total body surface area has not been spectacular, with poor graft take and unstable epithelium.current investigations have been directed toward dermal replacement beneath either very thin split-thickness autografts (stag) or utilizing cultured keratinocytes. current products include: collagen dermal replacement with thin stag (burke, et al). collagen dermal replacement with cultured keratinocytes and fibroblasts (boyce, et ai). allograft dermis with cultured keratinocytes (cnno, et al). allograft dermis with thin stag (life cell). polyglactin acid mesh and neonatal human fibroblasts with thin stag (hansbrnngh, et al).investigations regarding culture media, use of growth factors, topical nutrients and antibiotics, and melanocytes for pigmentation as well as safety and efficacy are needed before any of the current products become viable options for coverage of the massively burned patient. the~ is a growing world-wide problem with the ujc of cadaver tissues and ocgans bae, au~ of the tren~m~s~km of dilemma such a; cmutzfeldt.jukob disease and iiiv as we1] as ready availability of urdform lis~ue~. on dec~mt~r 14, 1993, the fda assumed control of as1 tissue bar~s in the uldtod st=tea in an attempt to bflng ~s difficult problem of dise~s~ transmission under ¢onlrol. in europe, ~om¢ of the governments are consldofll~ a c~mplcte bat) on the use of cadaverlc fissu~s such as ddn, 'this |ncroam in regulation of cadavefle ~s,quct will incmar¢ the difficulty of obtain~g and dlslflbulmg them. however, thc nc~ for these tissues contlnue~ m incrcaso, we will discuss ~'l¢ solulion to this important pmbl~n: tissue engineering. tlssu~ engineering is an in~rdisdpllnary field that applies pdnclplc~ of angin~edng and die life sclcnce~ reward the development of ~olok~¢al sub~dtute,~ ih= mslom, maintain, or improve tissue function, "13ssuc ongln~cdng can provide ~ho nccassary tlssuoa for wound repair ~d ibe assuranoe fl'~t the lissuos are d.ls¢~¢ free. in addition, a ds~uo-cng~ne~n~l wound covering will bo u~lvemally acceptable and evntlublc as "off g~o shell", consis~t products, them are several approaches to restating thls function in a large wound, 'l'nosc i~elud~ tmmcdiete long term coverage, short t=nn coverage, uandtl~el coverage and compost= dssu¢ coverage, "flssuo onglncrcd wound coverings that meet those vaflous ne,.cds will he r~vlowod.cllni~:sl and experimental d~la in venous ulcer, dlabctl¢ ulcers, prossur~ ulcers and bum wounds wgj be mvlcw~, a~ welt as new approacl~s u~ csrtilag¢, bone, liver and bone marrow it~suos. c oomplon, k nadirs, w press, g wetland, j fallen iv, shrtners burns institute and massachusetts general hospital, boston, ma~schusetts, usa the clinical "take" rate o? cultured epithelial autografts (cea) has been observed to increase with transplantation to allodermls, but the reasons for the improved clinical performance have not yet been defined. the aim of this study was to determine the biological impact of normal human dermis on cea differentiation and maturation, biopsies of cea transplanted to engrafted and de-opldermlzed human homograft dermis have been compared to nopsles of cea transplanted to granulation tissue in tullthickness burn wound beds on the same patient, each patient serving as hls or her own control. paired test and control biopstes from six patients have acquired from as early as one week postgrafting to as late as 3 years postgrafting (one patient) and analyzed histopathologlcally, ultrastructurally and immunoh[stochemloally, results demonstrate more rapid normalization of differentiation markers (e,g., involucfln, fllaggrln, cytokeratln profiles) in the cea transplanted to allodermls compared to their corresponding controls by in all patients, the proliferation rate within the basal layer ot the epidermis as determined by ki-67 (proliferation-associated antigen) is seen to norh~altze more quickly in the cea transplanted to allodermls in every case, persistence of allodermal matrix can be dooumented in all patients by elastic tlssue-trichrome stain, allowing visualization of the dermal elastin network. the popu;atlon densities ot intraepldarmal langerhans cells are conslstently and signlflcantly higher in cea transplanted to ,allodermls, possibly reflectlng an immunologlcal reaction to the underlying allogenlc tissue. overall, these preliminary results indicate that transplantation to a normal human dermal matrix accelerates the maturation of cea-deflved epidermis, wound closure continues to be a major problem in patients who have sustained a major thermal injury, cultured epidermal autografts (cea) have been utilized extensively since 1984 when galllco et el reported theh'use in two brothers with greater than 90% total body surface area burn. unfortunately, cea take rate varies widely and the resultant skin coverage is often fragile and the cosmetic results are less than optimal however the overall take rate and durability of the coverase can be markedly improved by using nn allodermls base as the recipient bed. a review of cea applications performed by physicians using cultured outologens epithelium obtained from blusurfaoe teclmology, inc. shows a marked discrepancy in the results obtained utilizing different methods of wound bed preparation. tgf-b is an important modulator coordinating complex physiological events associated with growth and development. it is assumed that tgf-b is also involved in the well-coordinated process of cutaneous wound healing by regulating proliferation, differentiation, chemotaxis and matrix deposition. the purpose of our study was to analyze the spatial and temporal pattern of tgf-b expression during granulation tissue formation in patients with accidanutl surgical trauma (monotraumata mid polytraumata) and bum wounds. after debridement (day 0), the full thickness wounds were covered with epigard, a synthetic dressing until day 10. after this time the granulated wounds were closed by transplantation of mesh graft. biopsies of the wound center were taken from 30 patients at the beginning of surgical treatment (day 0) and after 3, 6, and 10 days. cryosections were stained with antibodies against tgf-fi s using the apaap technique and -for standard histology -with hematoxylin-eosin. for identification of the cell type expressing tgf-6, double staining immunofluorescence experiments were conducted using antibodies specific for monocytes/macrophages, polymorphoanclear neutropkils and fibroblasts. the results showed a characteristic pattern of tgf-t~ distribution during wound development. tgf-fi appearence was mainly cell-associated znd the absolute and relative number of cells that were positive increased with lime. infiltrating cells and developing blood vessels were most prominently stained; epithelial and t-cells showed no immuno-reactivity. a delay of emergence for tgf-b during the time course could be seen in one patient group. this might reflect various regulation patterns depending on the type and severity of injury.(1) pharmatec gmbh, frankfurt (2) institut fiir immonologie and serologic, heidelberg ( immune cells extravasating specifically in skin recognize and eliminate the invading antigens (bacteria, viruses, etc.) either in situ or transport them to regional lymph nodes. they also participate in the process of skin wound healing. cells which traffic through the skin can be harvested from efferent lymph drained from a given area of skin. the type of migrating cells changes after trauma, heating and infection. we have developed a method for collection of human afferent lymph in lower limbs. the method allows obtaining immune cells from normal and injured skin and their characterization. aim of the study was to characterize skin immune cells in situ and in skin lymph with use of immunohistological methods (staining, facs). results. group 1, cells migrating through skin: 75+4% t lymphocytes (cd3), 6+3% langerhans and dendritic cells (cdla, hla dr, s100), 41+9% cd4, 18+6% cd8, no b cells (cd22, 23), 74% cd45r0 (memory cells), 6+3% il2r. approximately 90% cells possessed cdlla and 18 antigens. cd1 lc was expressed only on large cells. the frequency of all phenotypes was different from the blood populations. group 2, cells in skin: langerhans cells were found only in epidermis, cd3,4 and 8, cd45r0, rb, ila/18 cells around venules, cd68 (macrophages) uniformly dispersed, no il2r and b cells. hla dr positive were endothelial and some dispersed mononuclear cells. group 3, one, three and thirty days after surgical wound (simple varicous vein extirpation): high density of epidermal langerhans cells, hla dr positive keratinocytes and all endothelial ceils, few il2r cells, perivenular infiltrates of cd68, 45r0 but less cd3 cells, high density of cdlla/18 cells. classic staining of isolated and in situ located ccl!s with mgg or he did not allow to follow kinetics of changes. conclusions. this study presents the first in the literature quantitative data of immune cell traffic through normal and injured human skin. in the controlled release of biological response modifiers for soft tissue regeneration. alan s. rudolph, helmut speilberg, mariam monshipouri, and florence rollwagen, and barry j. spargo. we have employed lipid microstructures as controlled release vehicles for the delivery of growth factors in wound repair. traditional liposomes as well as novel lipid based microcylinders have been examined for their in vitro kinetics of the release of transforming growth factor beta (tgf-b). in vitro reiease has been examined by setting up models with examine the physical release of iodinated tgf-b as well as a cell based bioassay (based on the ht3 bioassay). the hollow lipid microcylinders (50 microns in length and i micron in diameter) show an initial burst (5-10 ng) followed be zero order kinetics which result in the release of approximately i ng tgf/day. this release behavior can be modified by temperature based on the phase behavior of the lipid bilayer which comprises the microcylinder.we have also examined the cellular response to lipid microcylinders applied in vivo. the lipid microcylinders are mixed in agarose and implanted as a composite hydrogel block under the flank of a mouse. the blocks are removed 1, 3, and 5 days following implant and the cells analyzed by facs sorter analysis. the observed pattern of ceil recruitment to the blocks mimics that seen in a local inflammatory response. cell surface phenotype studies included the determination of cd3 and cd4, mac-l, and ig bearing cells. we have also begun to examine the change in cell surface phenotype and kinetics of recruitment following the inclusion of tgf-beta in the lipid microcylinders.center for biomolecular science and engineering, code 6900, naval research laboratory, washington, dc. 20375-5348. expression pattern of heat shock proteins in acute, good healing and chronic human wound tissue. abstract: wound healing is a complex biologic process that is well characterized at the histological level, but its molecular regulation is poorly understood. after clot formation, inflammatory cells are rapidly drawn into the wound, followed by migration of fibroblasts and epithelial cells that divide and repopulate the wound area. during the last decade peptide growth factors and cytokine are thought to play a key role in initiating and sustaining the phase of tissue repair. these factors which are released from different cells appear to initiate the cascade of events that lead to healing. different studys described the rapid activation of a family of proteins,named heat shock proteins (hsp) in differnt tissue that were exposed to various forms of stress (heat, toxic agents, mechanical). in this context hsp's have the ability to regulate protein folding and assembly, to transport proteins across cytoplasm and membranes, to disrupt protein complexes, to stabilize, degrade and regulate the synthesis of proteins and to take part in dna replication and repair. we now attempted to find out if hsp-gene activation is also involved in injury and wound healing, which likewise resemble a stress situation for cells. therefore we collected tissue samples during operation and single biopsies from chronic wounds (decubitus for example) and granulation tissue. after rna preparation from these samples we used rna-pcr and nothern analysis to study the expression of objectives of the study chronic, non-healing cutaneous tflcers are a challenging clinical and socioeconomic problem. several animal studies have shown that cytukines (e.g. egf, pdgf, fgf, tgfb) accelerate the healing process and tissue repair in general. results from first clinical trials indicate a promising value of cytokines in the treatment of chronic non-healing diabetic and venous ulcers. recent reports in the literature indicate that the biological activity of the solution of platlet derived wound healing formula (pdwt~) released from c~-granules (mainly pdgf & tgfi~) is greater than the activity of the recombiant single factors like e.g. pdgf-bb (robson, lancet 1992) . the aim of our study was to determine whether a correlation exits between the concentration of tgfi~ & pdgf and the time course of wound healing. materials and methods pdwhf was prepared from 200ml of auto]ogous patient blood and diluted with a special buffer to a final concentration of 30 ng/ml g-thromboglobulin. the concentrations of pdgf and tgfg were determined by elisa-tests developed in our laboratory. 22 patients with chronic non-healing ulcers have been evaluated alter treatment by topical application of pdwhf. pdfg and tgff~ concentrations of the topical solution were measured and two patient groups formed for analysis the time course of wound healing was regularly and meticulously documented and evaluated by photography and casting. the time from initiation of treatment instil o wound volume reduction to 50 go of the origional size (t50%) was noted• results: healing of extensive burn wounds can be accelerated by grafting cultured autologous or allogeneic keratinocytes. the stimulation of granulation tissue formation and reepithelialization is presumably based on growth factors and cytokines released by keratinocytes. we wanted to prove this hypothesis by investigating the bfgf expression during wound development, bfgf is mainly described as an angiogenic protein with mitogenic activity on various mesodermal and ectodermal cell types pointing to its stimulating potential in wound heating. in the present study we compared the pattern of human bfgf m-rna expression and the localization of bfgf protein during the first days of wound healing. biopsies were taken from 5 juvenile human bum patients, immediately after wound debridemerit mad on day 8 after transplantation of cultured allografts. biopsies were snap frozen and cryosected. the pattern of bfgf expression was assessed by in situ hybridization of the bfgf m-rna with a digoxigenin-labelled antisense-rna and the parallel detection of the mature protein with an anfi-bfgf monoclonal antibody. our study revealed typical patterns of bfgf-m-rna-expression and intense bfgfprotein deposition during granulation tissue formation and reepithelialjzation of healing bum wounds. 'it, is known that major thermal injuries cause early impairment of wound healing followed by decreased influx of granuiocytes st. the site of injury. the role of granuiocytes in the process of wound healing is not ~"~ "" elucidated, it is now assumed that they are not merely phagocytic cells but active participants in ~n~*'1~.,.,a+~o~: processes secreting_ a number of various cvt-;kines, in order to investigate the effect of there is accumulating evidence that neuropeptides could be involved in the pathogenesis of several inflammatory reactions. vasocactive intestinal polypeptide (vip) and substance p (sp) have been detected by immunohistochemistry in normal as well as inflammed skin mostly in perivascular and periglandular location. both vip and sp are involved in vasodilatation, mast cell degranulation and irnmunomodulation.we determined the influence of sp and vip on the proliferation of lymphocytes in patients with psoriasis and healthy individuals. peripheral blood t-lymphocytes of 6 psoriatics and 6 healthy controls were isolated by density gradient centrifugation and passage over nylon wool. cell enrichment was controlled by facs analysis, lx105 t-lymphocytes were then incubated alone or in coculture with 2x104 irradiated autologous lymphocytes in culture medium containing 10 -7 mol/i sp or vip. cell proliferation was measured semiquanfitatively by tdr uptake in a betacounter. significance was tested by the wilcoxon signed-rank test.our results show that sp and vip exert only an effect on unstirnulated t-cells. in healthy individuals but not in patients with psoriasis sp increases significantly proliferation of t-cells. vip, however stimulates significantly the blastogenesis of t-lymphocytes only in psoriatics.our results confirm the psychoneuroimmunologic component in inflammatory reactions and vip and sp could be partially implicated in their pathogenetic mechanisms. moreover psoriatic lymphocytes show an altered reaction to sp and vip. this might be due to a preexisting (genetic?) or more likely to an epiphenomenal receptor defect. the adhesive interactions between endothelial cells and circulating ~enkocytes in shock and innammatory vondltions is mediated by several distinct families of ce11-surface determinants. of particular importance are the leukocyte integrins cdib / cdlla-c. in this study monoclonal antibodies to two of the u chains (cdlla & cdiib) and the common [~ chain (cdib) have been used to investigate leukocyte-dependent and leukocyte-independent plasma leakage in tee skin of rabbite. plasma leakage was measured as the local accumulation of t2si-hsa over a 30 rain period, the chemotac~c peptide imlp (0.1. 30ng) and bradykinin were used to induce cell.dependent and cell-5ndependent leakage respectively, the antibodies used were 6.5e (cdis), nri85 (cdlla) and antibody 198 (cdllb). ]ntradermal in~ections of bradyklnin and ~dlp both caused a dose dependent increase in plasma extravasatien (15.~. ffi 2.1p.l to 54.4 z b.bttl and 16.4 ,-1.8~ to 59.8 z 6.71d respectively. 6.5e (0.0072-2.4mf,/k~ iv) caused a dose dependent inhibition of imlp-induced but not bradyldnin.inducecl plasma exudation. at 0.24mk/kg, the plasma leakage was completely inhibited, antibody nr185 produced similar results, treatment with antibody 198 did not cause inhibition o£ plasma leakage due to either tnedi~tor. in vitro, the irmnune system ex~nination in persons with bone, chest and abdominal traumatic injury (i group . 22 patients without infectious coz~lications and 2 group -53 patients with wound infections development) was carried out. to restore found immunity disorders and host defense to infection 23 patients of the 2 group were treated with thymalin-the biologically active peptides prepared from bovine thymus. the examination on t~e i-3 days after injury revealed a considerable decrease of lymphocytes, ed3",$d4 ~ and cd8 cells amo~it in the blood, cd4 /cd8 ratio and indexes of let~ocyte migration inhibition test in both groups of patients. the imm~lity disorders recovered to norm on the 7-10 days in pateents of+the i group. but stable ~eple$ion of cd3 and cd4 cells amount, lower cd4 /cd8 ratio and indexes of leukocyte migration inhibition test in patients of the 2 group were observed~ besides that, these persons showed higher cd19cells amount and ig level in the blood. after thymalin therapy valid ii~rovement of inun~e status was discovered. also good clinical effect of immunotherapy and best wo~id healing observed in 80% of cases. these results allow us to propose that the thymus involution and the reduction of cell-mediated immunity responsiveness with disturbances of immu_uoregulatio~ on the level of restriction of activated cd4 tho cells play the most important role in the pathogenesis of wound infections development in persons with traumatic injury.dept. of immunology, military-nedical academy, lebedeva str. 6, 194175, st.petersburg, russia a severe impairment of neutrophil (pmn) function often occurs following severe thermal or non-thermal traumatic injury. our laboratory has previously reported that following severe burn or non-burn traumatic injury the expression of the p2 integrlns (cd11a,b,c/cd18) and the fw receptors (cd16, and cd32) were significantly decreased on pmns, in this study, the effects of gm and g-csf on the expression of the f~ r and the ~2 integrln family on pmns were examined, pmns were obtained from severe trauma (initial apache ii score ;z 10) or thermal injury (>30~; total body surface area, >15~ full thickness) and incubated /n v/tro with gm or g-csf. the j32 integrins or fcyr were detected with monoclonal antibodies and flow cytometry. gm end g-csf induced a sllght increase in the percentage of pmns expressing cd1 lb, cd16, and cd18 while gm bur not c-csf induced an increase in the percentage expressing cdi 1 a, cd1 lc, and cd32, gm-csf and to a lesser extent g-csf induced an increase in the density (1,5 fold) of the ~2 integrlns on pmns from normal, burn, and trauma patients, these data suggest that cytoklne modulation with csfs could have a role clinically in certain situations. institute, dept. of surgery, 231 bethesda ave, cincinnati, oh, usa, 45267-0558. funl~al infections after solid organ transplantatlon(sot) lewis flint, md and ed,~-afd e. etheredge, me) dept. of surgery tullrte univ. school of medicine new orleans. louisiana infections contribute to increased gra5 loss and mortaliw following sot. pr~isposing facton include diabetes, hepatitis, leukopenia, cc.¢xistem infection, and intense, especially triple drug, immunosuppression. funga] infections occur ~s isolated conditions in 6% and in association with bacterial infection(l 1%), viral infection(3*/.), and combined infections(it%), candida sp. is the most common fungus recovered but aspecgillus, coccidiodies, cryptococcus, histoplasma, mueor~ ghizopus, tinea, and toruiop~is s?. also are pathogens. clinical syndromes vary among orga.aizms or may be variable with a single p~tthogen, for ~ample, with aggressive immunosuppression, candlda my be localized esophagitis or cystitis or systemically iavaslve with an associated high mortality. aspergilius presents ~ a diffuse pneumonia while cryptococcus causes pulmonary and centrad nervons sy'stem infection, clinical examination, ct scanning and aggressive sampling for c'ultures a.s wall as serologic tests contribute to diagnosis. empiric the~py is ind',cated where there is a high level of suspicion. preventlon of ca.adlda izfection is ~ci~itated by early remov-a.1 of central }ants, ca~hetess and stents as well as by the use of oral nystatin. amphotericin ]~ remains the drug of choice for treatment of in.save fungd infection, surgical resection of infectious loci in the lung and brain is indicated in selected patients. the main problems of diagnosis in lower respirator-), tract infection are the differentation of infection from colonization or contamination, and the isolation of a reliable and true pathogen. expectorated sputum may be unreliable in pneumonia, because of contamination by oropharyngeal flora. although blood cultures may be negative, they provide a precise diagnosis and should be obtained in all pneumonias. other more invasive procedures are transtracheal needle aspiration, fibrobronchoscopic techniques including protected specimen brush and bronchoalveolar lavage with quantitative culturing and cytological analysis, transthoracic needle aspiration, thoracoscopy -guided biopsy and open lung biopsy. recently m. e1-ebiary, a. torres et al, 1993 reported quantitative cultures of endotracheal aspirates for the diagnosis of ventilator-associated pneumonia offering reliable results in these patients and should be further investigated. any invasive procedure in a severely ill patient should be carefully directed weighing the risks as well as the benefits, whilst taking the underlying diseases and expected survival into consideration. -current therapeutic approach is based mainly on monotherapy with broad spectrum antibiotics. combination therapy is apparently indicated only in p. aeruginosa infections and severe s. aureus pneumonia. graft infection can lead to fulminant graft failure or rapid progressive cirrhosis. for prevention of graft infection immunoprophylaxis, i,e. administration of human polyclonal anti hbs hypedmmunoglobutin (hig), starting in the anhepatic phase during operation, has proved to be at least partially succesful when performed on a long term basis.from a total of 415 olt in adult patients 100 olt were performed for hbsag positive liver disease (cirrhosis n=86, fulminant liver failure n=8, retransplantation n=6) in 94 pat. all pat. received 10.000 u hig in the anhepatic phase and 2.000 u/per day for the first week. a small group of 9 pat. received hig only for i week (short term immunoprophylaxis), in all other pat. hig is administered on a long term basis to keep anti hbs serum levels above 100 uii or until graft infection occurs (long term immunoprophylaxis);one-year survival rates are 100% in pat. who were transplanted for fulminant hepatitis, 96% in pat. with cirrhosis and long term prophylaxis, and 78% ir~ pat. with short term prophylaxis. all fatalities were related to hbv graft infection. the total rate of graft infection was 100% under short term prophylaxis and was independent from preoperative hbv dna status, under long term prophylaxis graft infection occurad in 28% in pat, negative for hbv dna. in hbv dna positive pat. infection rate was 56%, the total rate of reinfection for all pat. with long term prophylaxis was 36%the results of liver transplantation in hbsag positive pat. are comparable to other indications, graft infection with hepatitis b virus ist the major risk factor for these patients. under long term therapy with hig the rate of graft infection can be significantly reduced. the crucial cellular element for mods-mof: monocyi'f_./m acrophaoe ronald v. meier, m,d., f.a,c,s. the severely :injured or crldcally ill surgical patient is at high risk for immune dysfunction. a major consequence of this immune dysfunction is multiple organ dysfunction and failure leading to death, the underlying etiology is now recognized to be an uncontrolled, unfocused, disseminated activation of the host normally protective inflammatory. ,, cascades.. the resultant "mahgnant' systemic" inflan'a'natlon produces d~ffuso multiple organ bystander injury !eading to progressive organ dysfunction and failure. systemic malignant inflammation involves diffuse actlvatton of all components of the humoral and cellular inflammatory host response. of these various components, the macropha~e is the crucial central cellular element. the tissue fixed macrophage is ideally located diffusely throughout the various organs injured to orchestrate the inflammatory process. the macrophage is long-lived and highly metabolic, the macrophage regulates both the extent and the dissemination of the inflammatory processes. the macrophage is an exu'emely active c¢11 capable of producing and releasing not only directly eytotoxlc agents, s irnil~, to the neutrophil, including oxidants and numerous proteases out also the multitude of other cytokines and initiators of the interacting inflammatory cascades. the macrophage is the central source for ehemotactic agents (il-8, ltb4, c5a) for neutrophils and other inflammatory cells, production of vasoaetive arachidonie acid metabolites (tx, pgi2, poe, lt's), complement components (c3a, csa), thrombotic agents (pca, tx), metabolic and physiologic modulators (il,1, il-6 or tnf), and immunosuppressivc agents (poe2, il-10). these products of the macrophage are highly effective in enhancing and augmenting the inflammatory response. disseminated activation otthe macrophage is critical to the induction of the long-term diffuse activation of inflammation necessary to induce multiple organ injury and failure. our ability to elucidate the molecular mechanisms that control the macrophage will lead to our ability to conu'ol the maerophage response and prevent mods-mof.flarborview medical center, 325-9th ave za-16, seattle, wa 98104 usa