09 39 proceedings s.z.m.c. vol: 36(1): pp. 39-43, 2022. pszmc-832-36-1-2022 1central park medical college, lahore 2department of biochemistry, azra naheed medical college, superior university, lahore. 40 comparative study of déjà-vu and associated attributes among epileptics and non-epileptics 41 comparative study of déjà-vu and associated attributes among epileptics and non-epileptics 42 comparative study of déjà-vu and associated attributes among epileptics and non-epileptics 43 comparative study of déjà-vu and associated attributes among epileptics and non-epileptics 05 19 proceedings s.z.m.c. vol: 36(1): pp. 19-25, 2022. pszmc-828-36-1-2022 1department of obstetrics & gynaecology, indus hospital, raiwind campus, lahore 2department of obstetrics & gynaecology, nishtar hospital multan. 20 controlling post-partum hemorrhage using a novel technique of multiple sponge-holding-forceps applied… 21 controlling post-partum hemorrhage using a novel technique of multiple sponge-holding-forceps applied… 22 controlling post-partum hemorrhage using a novel technique of multiple sponge-holding-forceps applied… 23 controlling post-partum hemorrhage using a novel technique of multiple sponge-holding-forceps applied… 24 controlling post-partum hemorrhage using a novel technique of multiple sponge-holding-forceps applied… 25 controlling post-partum hemorrhage using a novel technique of multiple sponge-holding-forceps applied… 02 1 proceedings s.z.m.c. vol: 36(1): pp. 1-7, 2022. pszmc-825-36-1-2022 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 2 medical students’ perception on learning anatomy online during covid-19 pandemic … p 3 medical students’ perception on learning anatomy online during covid-19 pandemic … 4 medical students’ perception on learning anatomy online during covid-19 pandemic … 5 medical students’ perception on learning anatomy online during covid-19 pandemic … cbu 1 55 6 medical students’ perception on learning anatomy online during covid-19 pandemic … n 127 7 medical students’ perception on learning anatomy online during covid-19 pandemic … untitled-2 46 proceedings s.z.m.c. vol: 35(4): pp. 46-50, 2021. pszmc-823-35-4-2021 1department of orthodontics, de’montmorency college of dentistry, lahore 2department of oral biology, post graduate medical institute, lahore 3central research laboratory, postgraduate medical institute, lahore apis mellifera 47 effect of honey on orthodontic tooth movement and osteoclastic activity in psychologically stressed animals 48 effect of honey on orthodontic tooth movement and osteoclastic activity in psychologically stressed animals p 49 effect of honey on orthodontic tooth movement and osteoclastic activity in psychologically stressed animals 50 effect of honey on orthodontic tooth movement and osteoclastic activity in psychologically stressed animals untitled-2 1 proceedings s.z.m.c. vol: 35(4): pp. 1-5, 2021. pszmc-815-35-4-2021 1department of virology, 2department of histopathology, 3department of clinical chemistry and immunology, 4department of pathology, chughtai institute of pathology, lahore 2 comparison of covid-19 igg anti-spike antibody titer after vaccination with sputnik v… 3 comparison of covid-19 igg anti-spike antibody titer after vaccination with sputnik v… 4 comparison of covid-19 igg anti-spike antibody titer after vaccination with sputnik v… 5 comparison of covid-19 igg anti-spike antibody titer after vaccination with sputnik v… file.indd 73 proceedings s.z.m.c. vol: 35(2): pp. 73-77, 2021. pszmc-797-35-2-2021 department of obstetrics and gynecology, shaikh zayed medical complex, lahore 74 postpartum intrauterine contraceptive device (ppiucd) versus interval intrauterine contraceptive device (iucd) . 75 postpartum intrauterine contraceptive device (ppiucd) versus interval intrauterine contraceptive device (iucd) 76 postpartum intrauterine contraceptive device (ppiucd) versus interval intrauterine contraceptive device (iucd) 77 postpartum intrauterine contraceptive device (ppiucd) versus interval intrauterine contraceptive device (iucd) file.indd 41 proceedings s.z.m.c. vol: 35(2): pp. 41-46, 2021. pszmc-791-35-2-2021 1department of pediatric surgery, shaikh zayed medical complex, lahore 2department of public health & community medicine, shaikh zayed medical complex, lahore 3women medical officer, bhu awan, chak-39 4department of surgery, shaikh zayed medical complex, lahore 42 knowledge, attitude and practice of breast cancer screening in medical and non-medical females students 43 knowledge, attitude and practice of breast cancer screening in medical and non-medical females students 44 knowledge, attitude and practice of breast cancer screening in medical and non-medical females students 45 knowledge, attitude and practice of breast cancer screening in medical and non-medical females students 46 knowledge, attitude and practice of breast cancer screening in medical and non-medical females students 04 14 proceedings s.z.m.c. vol: 36(1): pp. 14-18, 2022. pszmc-827-36-1-2022 1department of anesthesia, shaukat khanum memorial cancer hospital & research centre, lahore 2department of anesthesia, gulab devi hospital, lahore 15 efficacy of paediatric preinduction anxiety distraction techniques during oncologic procedures 16 efficacy of paediatric preinduction anxiety distraction techniques during oncologic procedures 17 efficacy of paediatric preinduction anxiety distraction techniques during oncologic procedures 18 efficacy of paediatric preinduction anxiety distraction techniques during oncologic procedures file.indd 83 proceedings s.z.m.c. vol: 35(2): pp. 83-85, 2021. pszmc-799-35-2-2021 department of ophthalmology, rashid latif medical college, lahore 84 acute anterior uveitis (ocular manifestation of covid-19): a case report 85 acute anterior uveitis (ocular manifestation of covid-19): a case report 86 untitled-2 13 proceedings s.z.m.c. vol: 35(4): pp. 13-17, 2021. pszmc-817-35-4-2021 1department of surgery, king edward medical university, lahore 2department of psychiatry, rashid latif medical college, lahore 3institute of molecular biology and biotechnology, university of lahore 14 initial psychological response of various population groups to covid-19 pandemic: a cross sectional study 15 initial psychological response of various population groups to covid-19 pandemic: a cross sectional study 16 initial psychological response of various population groups to covid-19 pandemic: a cross sectional study 17 initial psychological response of various population groups to covid-19 pandemic: a cross sectional study file.indd 58 proceedings s.z.m.c. vol: 35(2): pp. 58-63, 2021. pszmc-794-35-2-2021 1department of anatomy, fmh college of medicine & dentistry, lahore 2department of neurology, fmh college of medicine & dentistry, lahore 3department of anatomy, continental medical college, lahore 59 toxic effects of ribavirin on the testicular interstitium in albino rats 60 toxic effects of ribavirin on the testicular interstitium in albino rats 61 toxic effects of ribavirin on the testicular interstitium in albino rats 62 toxic effects of ribavirin on the testicular interstitium in albino rats 63 toxic effects of ribavirin on the testicular interstitium in albino rats untitled-2 25 proceedings s.z.m.c. vol: 35(4): pp. 25-29, 2021. pszmc-819-35-4-2021 1department of pulmonology, university of lahore teaching hospital, lahore 2department of histopathology, gulab devi hospital, lahore 3department of pulmonology, gulab devi hospital, lahore 4department of pulmonology, lahore general hospital, lahore 26 fnac a simple & cost effective diagnostic tool for benign & malignant pathologies … 27 fnac a simple & cost effective diagnostic tool for benign & malignant pathologies … 28 fnac a simple & cost effective diagnostic tool for benign & malignant pathologies … 29 fnac a simple & cost effective diagnostic tool for benign & malignant pathologies … full book 65 proceedings s.z.m.c. vol: 36(4): pp. 65-69, 2022. pszmc-866-36-4-2022 1department of obstetrics and gynaecology, indus hospital, raiwind campus, lahore 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 66 efficacy of oxytocin in reducing blood loss during abdominal myomectomy pvalue 67 efficacy of oxytocin in reducing blood loss during abdominal myomectomy 68 efficacy of oxytocin in reducing blood loss during abdominal myomectomy 69 efficacy of oxytocin in reducing blood loss during abdominal myomectomy file.indd 28 proceedings s.z.m.c. vol: 35(2): pp. 28-34, 2021 pszmc-789-35-2-2021 1department of pharmacology, fatima jinnah medical university, lahore. 2department of pharmacology, rehman medical college, peshawar. 3department of pharmacology, d.g. khan medical college, dera ghazi khan. 4department of pharmacology, al-aleem medical college, lahore. malus domestica 29 effect of malus domestica (apple peel) extract on peripheral blood and bone marrow cells of prednisolone treated mice ad libitum 30 effect of malus domestica (apple peel) extract on peripheral blood and bone marrow cells of prednisolone treated mice post hoc p31 effect of malus domestica (apple peel) extract on peripheral blood and bone marrow cells of prednisolone treated mice p p p p p p p 32 effect of malus domestica (apple peel) extract on peripheral blood and bone marrow cells of prednisolone treated mice p p malus domestica 33 effect of malus domestica (apple peel) extract on peripheral blood and bone marrow cells of prednisolone treated mice 34 effect of malus domestica (apple peel) extract on peripheral blood and bone marrow cells of prednisolone treated mice untitled-2 36 proceedings s.z.m.c. vol: 35(4): pp. 36-40, 2021. pszmc-821-35-4-2021 nigella sativa 1department of physiology, fatima jinnah medical university, lahore 2department of hematology, fatima jinnah medical university, lahore 3department of physiology, postgraduate medical institute, lahore nigella sativa nigella sativa nigella sativa nigella sativa nigella sativa nigella sativa nigella sativa nigella sativa nigella sativa nigella sativa 37 effect of nigella sativa on estradiol levels and menopausal symptoms in post-menopausal females nigella sativa nigella sativa nigella sativa nigella sativa 38 effect of nigella sativa on estradiol levels and menopausal symptoms in post-menopausal females nigella sativa nigella sativa nigella sativa nigella sativa nigella sativa nigella sativa 39 effect of nigella sativa on estradiol levels and menopausal symptoms in post-menopausal females nigella sativa nigella sativa nigella sativa nigella sativa nigella sativa nigella sativa nigella sativa nigella sativa nigella sativa nigella sativa nigella sativa nigella sativa nigella sativa nigella sativa nigella sativa 40 effect of nigella sativa on estradiol levels and menopausal symptoms in post-menopausal females 11 49 proceedings s.z.m.c. vol: 36(1): pp. 49-55, 2022. pszmc-834-36-1-2022 1department of pharmacology, ayub medical college, abbottabad 2department of pharmacology, post graduate medical institute, lahore 3department of pathology, punjab institute of cardiology, lahore 4department of pharmacology, sahara medical college, narowal 5department of pharmacology, shalamar medical and dental college, lahore 50 therapeutic effect of berberine versus methotrexate on joint histopathology in a rat model… berberis lycium royle (blr), berberidiaceae group group name arthritis induction treatment day 15 onwards 51 therapeutic effect of berberine versus methotrexate on joint histopathology in a rat model… p p p p p p a-e 52 therapeutic effect of berberine versus methotrexate on joint histopathology in a rat model… a-e p p p p p p p 100 100 0 12.5 75 0 0 12.5 75 25 0 0 62.5 12.5 0 0 0 25 0 0 p value ### ### *** *** ### ^ ### 100 100 0 12.5 37.5 0 0 62.5 75 62.5 0 0 37.5 12.5 0 0 0 0 0 0 p value ### ### *** *** ** # 100 100 0 25 37.5 0 0 37.5 75 50 0 0 50 0 12.5 0 0 12.5 0 0 p value ### ### *** ** ## ** # 100 100 0 12.5 12.5 0 0 12.5 62.5 37.5 0 0 62.5 25 50 0 0 25 0 0 p value ### ### *** *** # *** # p p p p p p 53 therapeutic effect of berberine versus methotrexate on joint histopathology in a rat model… 54 therapeutic effect of berberine versus methotrexate on joint histopathology in a rat model… rhizoma coptidis 55 therapeutic effect of berberine versus methotrexate on joint histopathology in a rat model… for web full book dedicated to the valiant health care warriors, medical students & researchers of pakistan bismillah hir rehman nir raheem. in the name of allah, the most beneficent and the most merciful message by the chairperson shaikh zayed medical complex & chief editor of proceedings prof. sibgha zulfiqar prof. saadia s. alam (chairperson & dean) (chief editor) we have achieved 2 years of hec y category indexation. our ojs supported proceedings website of international standards connects us to the world. the covid 19 pandemic has nearly subsided into random cases of the omicron variant while dengue has once again reared its head. in the changing scenario we steadfastly publish a diverse april to june 2022 proceedings articles on varied aspects of these and diverse medical, surgical and allied health disciplines . all articles submitted for publication have been judged on stringent international standards of plagiarism and blind peer reviews. our editions have achieved expanded viewership, with researchers in wide ranging medical fields from different institutions publishing their research in proceedings. our nexus of undergraduate and postgraduate researchers, supervisors, academicians, and reviewers remains imperative to achieving rigorous standards of research, writing and evaluation in proceedings. our desire is to build linkages with top journals and researchers both in the country and abroad in sha allah. our motto “we venture forth” pmc & hec approved proceedings shaikh zayed medical complex vol. 36 (2) pcpb/24(94) p-121/3196 apr to jun 2022 editorial board patron in chief: prof. sibgha zulfiqar chairperson & dean chief editor: prof. saadia shahzad alam phd external associate editor: dr. usman iqbal phd associate editors: prof. ayesha humayun phd dr. adnan salim fcps members-national: prof. mamoon rashid frcs (sitara-e-eisaar) prof. nadiramamoon mrcpath prof. m. ashraf phd (tamgha-e-imtiaz) prof. m.arif nadeem (fcps) prof.khalid mahmood(phd) dr. sarah ghafoorphd members-international prof.rennekoeffel phd(sui) prof.jane banaszak-holl phd(aus) prof. hisham al muhtaseb phd (ksa) dr. salma malik m.d (us) dr. sarwat shaheenm.d(us) dr. aliya ashermrcp (uk) dr. uzma nasim siddiquifcps (aus) editors: dr. shahila jalil fcps dr. samira haque fcps dr. sadiamaqsood mhpe dr. faraz bokhari m phil dr.noora hassan hezam al aqmer m phil mrs. saimamohsin msc consultant biostatician: mr. muhammad aasimm phil library & information sciences: mr. ihsan basitm phil dr.muhammad shahid soroyaphd contents asma nasir, madiha sarwar, mavra fatima, hareem noor, ayisha imran, noman aslam malik, omar rasheed chughtai, a.s. chughtai blood transfusion practice in dengue fever: a cross sectional single center study during a dengue outbreak in pakistan saadia shahzad, sarosh saleem reopening of universities for on-campus teaching in covid-19 pandemic: status of generalized anxiety in medical students alvia batool, maryam fatima, shamaila doggar, faeza rauf, samina munawar, fatima inam, samar iqbal morphometry of foramen magnum: an anatomical study saba saleem, mahrukh malik, noor ijaz, saba amjad, syedhussain raza zaidi, nabila kaukab perception of mbbs students about effectiveness of modified unobserved anatomy ospe due to covid-19 restrictions mehwish iftikhar, bushra iftikhar, mahwish shahzad, junaid iqbal, aliya aslam, mirza ameer faizan ali assessment of lipid profile in females suffering from endometriosis fahad azam, abida shaheen, hania naveed, afrose liaquat, hania ahmer, khurram irshad correlation between post exercise heart rate recovery and body composition in healthy female adults fahad ali, qazi abdul saboor, faryad ali, talha mahmud, syed ahmad ali hassan, athar iqbal association of left ventricular hypertrophy on echocardigraphy with hemorrhagic and non-hemorrhagic stroke shizra kaleemi, lubna humayun, hamna salahuddin, farah arif, asma akhtar, ayesha nasir adverse blood transfusion reaction in tertiary care hospitals an initiative towards improvement: a multi center study vishal farid raza, samiha choudhry, hina khan, sajeel ahmad, anza talal, khalid javeed khan association between depressed mood and morbid obesity: experience from a bariatric surgery program instructions for author page 1 7 14 19 23 29 35 40 46 52 the editorial board of proceedings of shaikh zayed medical complex, lahore is indebted to our honorable peer reviewers for their expert & timely reviews. advisory board basic & pre-clinical sciences prof. mateen izhar phd, mrcpath prof. sibgha zulfiqar m.phil prof. muhammad suhail m.phil prof. nasreen ehsan m.phil prof. tahira naseem m.phil dr. anwaar bashir m.phil dr. suleman dawood phd (uol) dr. nighat yasmeen phd (kemu) dr. saima chaudhary phd (uhs) medicine and allied prof. azeem taj fcps prof. abdul shakoor fcps prof. talha mahmud fcps prof. safoora aamir fcps dr. lubna riaz fcps dr. asfandyar fcps surgery and allied prof. ali rafique mirza fcps prof. muhammad ikram fcps prof. imran anwar frcs prof. shafqat mukhtar fcps dr. jamshaid rahim fcps dr. syed sajjad raza kazmi fcps diagnostic division prof. mona aziz fcps prof. saulat sarfraz fcps dr. amir khan frcr peer review board (national) prof. m. nauman ahmad ffarcs prof. naseem saud phd prof. m. ovais omer phd prof. shabbir bhatti phd prof. zahid niaz frcs prof. zamir ahmad phd prof. muhammad arif nadeem fcps prof. muhammad moin frcs prof. khalid mahmood phd prof. sadia chiragh m.phil prof. muhammad aslam fcps prof. abdul mannan m.s prof. naheed humayun fcps prof. farhat naz fcps prof. abdul hameed frcs prof. rafeea tafweez phd peer review board (national) prof. inayat thaver phd prof. ashraf chaudhry fcps prof. kashif malik fcps prof. aftabturabi phd prof. uzma hussain fcps prof. muhammad pervaiz prof. fouzia shaukat fcps prof. maryam rashid phd prof. rabeia bilal phd prof. khwaja khursheed fcps prof. tanvir akhtar butt ms prof. naila akhtar fcps prof. sajida malik m.phil dr. farooq afzal frcs dr. samanshahid phd dr. muhammad shahzad phd dr. ali hussainy zaidi m.d dr. shahzad khuram akram phd dr. uzma malik fcps dr. ayesha mallick frcp dr. tehseen haider kazmi fcps dr. khalida ajmal m.phil dr. israr ahmed fcps dr. saima batool fcps dr. saleem muhammad rana phd dr. muhammad khurram habib fcps dr. saba riaz phd dr. raaziatasadduq phd dr. nageen hussain phd dr. saadia nosheen jan fcps dr. soumble zulfiqar phd dr. yasir abbas zaidi fcps dr. muhammad zeeshan sarwar mrcs dr. muhammad imran khokar mrcs dr. nabihafarasat m.phil dr. tayyaba muzaffar m.phil dr. zaeem khalid fcps-1 peer review board (international) prof. renne koeffel phd (switzerland) prof. jane banaszak-holl phd (aus) prof. m. hisham al-muhtaseb phd (jordan) prof. mukhtiar baig phd (ksa) dr. sarwatsha heen m.d (usa) dr. zeeshan i. shaikh m.d (usa) dr. tanzeel qurat ijaz mrcp (uk) dr. hafiz sohail anjum fcps (ireland) dr. rafay azhar frcp (singapore) dr. mazhar nawaz m.d (usa) dr. ahmad azam malik phd (ksa) dr. ishtiaq ahmed fcps (ksa) dr. aamir omairphd (ksa) dr. sabina ahmed mir m.d (usa) dr. anita lamichhane m.d (nepal) dr. faiza durrani phd (uk) dr. waseem lodhi frcog (uk) dr. seerat zahra hammad m.s (ksa) dr. shafya shahid phd (u.s) dr. abdul waheed frcs (u.k) dr. humaira zareen fcps (ksa) dr. amira okud m.d (ksa) dr. asma ahmed mrcp (uk) dr. sabahat sabir mrcog (uk) dr. sarfaraz ahmad frcr (uk) untitled-2 dedicated to the researchers, clinicians & health care workers of pakistan bismillah hir rahman nir raheem. in the name of allah, the most beneficent and the most merciful message by the chairman shaikh zayed medical complex & chief editor of proceedings prof. mateen izhar prof. saadia s. alam (chairman & dean) (chief editor) we present our final edition for 2021 with encouraging news on the covid front while unfortunately dengue fever cases rise menacingly straining our health care system. in pakistan the deadly 4th wave of covid has finally ebbed with lowest infectivity rates due to increasing vaccinations alhumdolillah. we pray humanity is relieved of this deadly disease. for proceedings 2021 saw the highest number of article submissions since its inception. over a hundred research articles on wide ranging topics were submitted and vetted as per hec guidelines. the oct to dec 2021 proceedings presents articles from medical, allied and dental health professionals and wide ranging basic and applied research. all articles have been judged on stringent international criteria of plagiarism and blind peer reviews. we have achieved expanded viewership, with researchers in diverse medical fields from different institutions publishing their research in proceedings our vision, building linkages with top journals and researchers both in the country and abroad “we venture forth” quarterly proceedings shaikh zayed medical complex vol. 35 (4) pcpb/24(94) p-121/3196 oct to dec 2021 editorial board patron in chief: prof. mateen izhar phd, mrcpath chairman & dean chief editor: prof. saadia shahzad alam phd external associate editor: dr. usman iqbal phd, associate editors: prof. ayesha humayun phd dr. adnan salim fcps members-national: prof. mamoon rashid frcs (sitara-e-eisaar) prof. nadira mamoon mrcpath prof. m. ashraf phd (tamgha-e-imtiaz) prof. m. arif nadeem (fcps) dr. sarah ghafoor phd members-international prof. renne koeffel phd (sui) prof.jane banaszak-holl phd (aus) prof. hisham al muhtaseb phd (ksa) dr. salma malik m.d (us) dr. sarwat shaheen m.d (us) dr. aliya asher mrcp (uk) dr. uzma nasim siddiqui fcps (aus) editors: dr. shahila jalil fcps dr. samira haque fcps dr. sadia maqsood mhpe dr. faraz bokhari m phil dr. noora hassan hezam al-aqmer m phil mrs. saima mohsin msc consultant biostatician: mr. muhammad aasim m phil library & information sciences: mr. ihsan basit m phil dr.muhammad shahid soroya phd contents hajra farooq, omar rasheed chughtai, hijab batool, waheed uz zaman tariq, akhtar sohail chughtai comparison of covid-19 igg anti-spike antibody titer after vaccination with sputnik v in seropositive and naïve vaccinees shehrbano ali, muhammad murad murtaza, marriam nasir, muhammad ashraf chaudhry symptoms and associations of covid-19 in pakistan: a single centre, descriptive study shahzaib ahmad, muhammad abubakar shahid chishti, usman hotiana, tania a. shakoori initial psychological response of various population groups to covid19 pandemic: a cross sectional study hamna zahid, sana noreen, bahisht rizwan, shahid bashir, shahwar javaid, mujeeb ur rehman, dua waqar, ayza hashmi, mahnoor khalid, kinza kashif, osama rasheed food insecurity and its effect on mental health: a systematic review sohail anwar, amna rehman, asifa karamat, huma batool, ali afzal fnac: a simple & cost-effective diagnostic tool for benign & malignant pathologies associated with cervical lymphadenopathy sofia yasmeen abbasi, rukhsana kausar, hira naz, sadia chiragh effect of carica papaya leaf juice on blood cell count of busulfaninduced chronic bone marrow aplasia in mice shazo sana, hafiz muhammad umair, muniza saeed effect of nigella sativa on estradiol levels and menopausal symptoms in post-menopausal females arooj farooq, aatiqa abbas, arif rasheed determination of postmortem interval by estimating csf proteins concentration after death, by dye binding method at a tertiary care hospital in lahore, pakistan reem abid, muhammad sharjeel ilyas, ghazala rubi, amna ahsan, sadia shakeel, arfa ahmed effect of honey on orthodontic tooth movement and osteoclastic activity in psychologically stressed animals nabiha farasat khan, muhammad saeed, ayesha ayub, arshad kamal butt an updated insight into learning approach of government sector dental students in balochistan instructions for authors page 1 6 13 18 25 30 36 41 46 51 58 the editorial board of proceedings of shaikh zayed medical complex, lahore is indebted to our honorable peer reviewers for their expert & timely reviews. advisoryboard basic & pre-clinical sciences prof. mateen izhar phd, mrcpath prof. sibgha zulfiqar m.phil prof. muhammad suhail m.phil prof. nasreen ehsan m.phil prof. tahira naseem m.phil dr. anwaar bashir m.phil dr. suleman dawood phd (uol) dr. nighat yasmeen phd (kemu) dr. saima chaudhary phd (uhs) medicine and allied prof. azeem taj fcps prof. abdul shakoor fcps prof. talha mahmud fcps prof. safoora aamir fcps dr. lubna riaz fcps dr. asfandyar fcps surgery and allied prof. ali rafique mirza fcps prof. muhammad ikram fcps prof. imran anwar frcs prof. shafqat mukhtar fcps dr. jamshaid rahim fcps dr. syed sajjad raza kazmi fcps diagnostic division prof. mona aziz fcps prof. saulat sarfraz fcps dr. amir khan frcr peer review board (national) prof. m. nauman ahmad ffarcs prof. naseem saud phd prof. m. ovais omer phd prof. shabbir bhatti phd prof. zahid niaz frcs prof. zamir ahmad phd prof. muhammad arif nadeem fcps prof. muhammad moin frcs prof. khalid mahmood phd prof. zujaja zaheer m.phil prof. sadia chiragh m.phil prof. muhammad aslam fcps prof. abdul mannan m.s prof. naheed humayun fcps prof. farhat naz fcps prof. abdul hameed frcs prof. rafeea tafweez phd prof. inayat thaver phd prof. ashraf chaudhry fcps prof. kashif malik fcps prof. aftab turabi phd prof. uzma hussain fcps prof. muhammad pervaiz prof. fouzia shaukat fcps prof. maryam rashid phd prof. rabeia bilal phd prof. khwaja khursheed fcps prof. tanvir akhtar butt ms prof. naila akhtar fcps prof. sajida malik m.phil dr. farooq afzal frcs dr. saman shahid phd dr. muhammad shahzad phd dr. ali hussainy zaidi m.d dr. shahzad khuram akram phd dr. uzma malik fcps dr. ayesha mallick frcp dr. tehseen haider kazmi fcps dr. khalida ajmal m.phil dr. israr ahmed fcps dr. saima batool fcps dr. saleem muhammad rana phd dr. muhammad khurram habib fcps dr. saba riaz phd dr. raazia tasadduq phd dr. nageen hussain phd dr. saadia nosheen jan fcps dr. soumble zulfiqar phd dr. yasir abbas zaidi fcps dr. muhammad zeeshan sarwarmrcs dr. muhammad imran khokar mrcs dr. nabiha farasat m.phil peer review board (international) prof. renne koeffel phd (switzerland) prof. jane banaszak-holl phd (aus) prof. m. hisham al-muhtaseb phd (jordan) prof. mukhtiar baig phd (ksa) dr. sarwat shaheen m.d (usa) dr. zeeshan i. shaikh m.d (usa) dr. tanzeel qurat ijaz mrcp (uk) dr. hafiz sohail anjum fcps (ireland) dr. rafay azhar frcp (singapore) dr. mazhar nawaz m.d (usa) dr. ahmad azam malik phd (ksa) dr. ishtiaq ahmed fcps (ksa) dr. aamir omair phd (ksa) dr. sabina ahmed mir m.d (usa) dr. anita lamichhane m.d (nepal) dr. faiza durrani phd (uk) dr. waseem lodhi frcog (uk) dr. seerat zahra hammad m.s (ksa) dr. shafya shahid phd (u.s) dr. abdul waheed frcs (u.k) dr. humaira zareen fcps (ksa) dr. amira okud m.d (ksa) dr. asma ahmed mrcp (uk) dr. sabahat sabir mrcog (uk) dr. sarfaraz ahmad frcr (uk) dedicated to the researchers, clinicians & health care workers of pakistan bismillah hir rahman nir raheem. in the name of allah, the most beneficent and the most merciful message by the chairman shaikh zayed medical complex & chief editor of proceedings prof. mateen izhar prof. saadia s. alam (chairman & dean) (chief editor) we present our july to sept 2021 edition with glad tidings for all. increased covid 19 vaccinations have alhumdolillah led to a gradual reopening of academic institutions and resumption of daily activity. we pray that normalization continues and humanity is relieved of this deadly disease. for proceedings 2020 was a year of achievements alhumdolillah with development of a new website at www.proceedings-szmc.org.pk of international standards supported by the ojs/pkp platform for online submission. during the year we got recognition from higher education commission of pakistan (hec). we thank them for reposing their confidence in us awarding “y” category, an honor indeed. we are now indexed by pmc, google scholar, crossref/doi pastic and pakmedinet. efforts are underway for international indexation. the july to sept 2021 proceedings presents articles from medical and allied health professionals and wide ranging basic and applied research. all articles submitted for publication have been judged on stringent international criteria of plagiarism and blind peer reviews. we have achieved expanded viewership, with researchers in wide ranging medical fields from different institutions publishing their research in proceedings our vision, building linkages with top journals and researchers both in the country and abroad “we venture forth” http://www.proceedings-szmc.org.pk/ hec & pmc approved quarterly proceedings shaikh zayed medical complex vol. 35 (3) pcpb/24(94) p-121/3196 july to sept 2021 editorial board patron in chief: prof. mateen izhar phd, mrcpath chairman & dean chief editor: prof. saadia shahzad alam phd external associate editor: dr. usman iqbal phd, associate editors: prof. ayesha humayun phd dr. adnan salim fcps members-national: prof. mamoon rashid frcs (sitara-e-eisaar) prof. nadira mamoon mrcpath prof. m. ashraf phd (tamgha-e-imtiaz) dr. sarah ghafoor phd members-international prof. renne koeffel phd (sui) prof.jane banaszak-holl phd (aus) prof. hisham al-muhtaseb phd (jordan) dr. salman malik m.d (us) dr. sarwat shaheen m.d (us) dr. aliya asher mrcp (uk) dr. uzma nasim siddiqui fcps (aus) editors: dr. shahila jalil fcps dr. samira haque fcps dr. sadia maqsood mhpe dr. faraz bokhari m phil dr.noora hassan hezam al-aqmer m phil mrs. saima mohsin msc consultant biostatician: mr. muhammad aasim m phil library & information sciences: mr. ihsan basit m phil dr. muhammad shahid soroya phd contents fariha sahrish, abeera assad rasool, zonaira rathore, mahvish hussain, fizza waqar, samina zaman histological subtypes & staging of post-chemotherapy wilms tumour according to siop 2001 protocol: study at the children’s hospital,lahore quratulain mehdi, noaman ishaq, saba batool, kulsoom farhat, saman omer, arooj shahid efficacy of telmisartan in pristane induced arthritis rat model muhammad ali sheikh, amna idrees, jamil akhter munir ahmad, kaneez fatima, lubna riaz, amna ikram community acquired methicillin resistant staphylococcus aureus (ca-mrsa) infection in paediatric subcutaneous abscesses in pakistan abdul mudabbir rehan, fariha ahmad khan, qudsia umaira khan, rabia naseer khan, zoobia irum, sehrish zaffar, sadia chiragh malus domestica peel extract counters carboplatin induced fall in red blood cell count and bone marrow erythroid cell percentage in mice haroon javaid majid, muhammad imran anwar, muhammad zaeem khalid, shafique-ur-rehman, muhammad amir jameel predictors of mortality in necrotizing soft tissue infections in adults – experience at a tertiary care hospital in pakistan fatima khalid, sameen afzal, ghazi zafar, anila chughtai, samina zaman, akhtar sohail chughtai morphological spectrum of ovarian teratomas: a 6 years experience at a specialized diagnostic institute sumra komal, aqna malik, naheed akhtar, syed asif jahanzeb kazmi, fayyaz anjum, ayesha rida tamarix dioica (ghaz) protective potential in the carbon tetrachlorideinduced hepatotoxicity animal model ambreen anjum, hamid javed qureshi, saima tabassum, wardah anwar, rida shakil, maria anwar, muhammad hashim ghouri comparative effects of cane sugar, honey & jaggery on plasma glucose level & body weight of alloxan induced diabetic rats amna javed, muhammad ikram, shafqat mukhtar, saulat sarfraz, ayesha shahid, aman ur rehman prevalence of bacterial vaginosis during pregnancyin tertiary care hospital, lahore asifa karamat, huma batool, sohail anwar, shazia akram, atif masood, wajid ali rafai frequency and pattern of bronchiectasis in patients with chronic obstructive pulmonary disease presenting in a tertiary care hospital sadia majeed, usman aslam, muhammad ubaid ullah khan, asma inam, sadia chiragh phoenix dactylifera (ajwa date) whole fruit, flesh & powdered seed prevents anti-tuberculous drug induced hepatotoxicity in rabbits rakesh bahadur adhikari, qazi abdul saboor, mateen akram, saira fayyaz, husnain bashir, muhammad rizwan-ul-haque prevalence of right ventricular dysfunction& pulmonary hypertension and their relationship to the number of hemodialysis sessions in patients of end-stage renal disease (esrd) hira ashfaq, ahmad naseem, shabnum tariq, hina usman, tayyiba wasim, maria sharif, naheed humayun fertility-related quality of life amongst married infertile females of lahore wajida perveen, muhammad akhtar, riaz hashmi, misbah amanat ali, sahreen anwar, ayesha ghani idiopathic facial nerve paralysis & response to physiotherapy in pregnant woman; a case report syed wasif ali shah, muhammad abdul shakoor, usman amjad, haider ali, waleed imtiaz, shafqat raza hereditary multiple exostoses with ulnar hemimelia instructions for authors page 1 6 12 17 24 31 37 44 50 54 58 64 69 75 78 81 the editorial board of proceedings of shaikh zayed medical complex, lahore is indebted to our honorable peer reviewers for their expert & timely reviews. advisory board basic & pre-clinical sciences prof. mateen izhar phd, mrcpath prof. sibgha zulfiqar m.phil prof. muhammad suhail m.phil prof. nasreen ehsan m.phil prof. tahira naseem m.phil dr. anwaar bashir m.phil dr. suleman dawood phd (uol) dr. nighat yasmeen phd (kemu) dr. saima chaudhary phd (uhs) medicine and allied prof. azeem taj fcps prof. abdul shakoor fcps prof. talha mahmud fcps prof. safoora aamir fcps dr. lubna riaz fcps dr. asfandyar fcps surgery and allied prof. ali rafique mirza fcps prof. muhammad ikram fcps prof. imran anwar frcs prof. shafqat mukhtar fcps dr. jamshaid rahim fcps dr. syed sajjad raza kazmi fcps diagnostic division prof. mona aziz fcps prof. saulat sarfraz fcps dr. amir khan frcr peer review board (national) prof. m. nauman ahmad ffarcs prof. naseem saud phd prof. m. ovais omer phd prof. shabbir bhatti phd prof. zahid niaz frcs prof. zamir ahmad phd prof. muhammad arif nadeem fcps prof. muhammad moin frcs prof. khalid mahmood phd prof. zujaja zaheer m.phil prof. sadia chiragh m.phil prof. muhammad aslam fcps prof. abdul mannan m.s prof. naheed humayun fcps prof. farhat naz fcps prof. abdul hameed frcs prof. rafeea tafweez phd prof. inayat thaver phd prof. ashraf chaudhry fcps prof. kashif malik fcps prof. aftab turabi phd prof. uzma hussain fcps prof. muhammad pervaiz prof. fouzia shaukat fcps prof. maryam rashid phd prof. rabeia bilal phd prof. khwaja khursheed fcps prof. tanvir akhtar butt ms prof. naila akhtar fcps prof. sajida malik m.phil dr. farooq afzal frcs dr. saman shahid phd dr. muhammad shahzad phd dr. ali hussainy zaidi m.d dr. shahzad khuram akram phd dr. uzma malik fcps dr. ayesha mallick crcp dr. tehseen haider kazmi fcps dr. khalida ajmal m.phil dr. israr ahmed fcps dr. saimabatool fcps dr. saleem muhammad rana phd dr. muhammad khurram habib fcps dr. saba riaz phd dr. raazia tasadduq phd dr. nageen hussain phd dr. saadia nosheen jan fcps dr. soumble zulfiqar phd dr. yasir abbas zaidi fcps dr. muhammad zeeshan sarwar mrcs dr. muhammad imran khokar mrcs dr. nabiha farasat m.phil peer review board (international) prof. renne koeffel phd (switzerland) prof. jane banaszak-holl phd (aus) prof. m. hisham al-muhtaseb phd (jordan) prof. mukhtiar baig phd (ksa) dr. sarwat shaheen m.d (usa) dr. zeeshan i. shaikh m.d (usa) dr. tanzeel qurat ijaz mrcp (uk) dr. hafiz sohail anjum fcps (ireland) dr. rafay azhar frcp (singapore) dr. mazhar nawaz m.d (usa) dr. ahmad azam malik phd (ksa) dr. ishtiaq ahmed fcps (ksa) dr. aamir omair phd (ksa) dr. sabina ahmed mir m.d (usa) dr. anita lamichhane m.d (nepal) dr. faiza durrani phd (uk) dr. waseem lodhi frcog (uk) dr. seerat zahra hammad m.s (ksa) dr. shafya shahid phd (u.s) dr. abdul waheed frcs (u.k) dr. humaira zareen fcps (ksa) dr. amira okud m.d (ksa) dr. asma ahmed mrcp (uk) dr. sabahat sabir mrcog (uk) dr. sarfaraz ahmad frcr (uk) 6mavra fatima new proceedings s.z.m.c. vol: 36(1): pp. 31-34, 2022. pszmc-830-36-1-2022 pseudobasophilia: a helpful screening tool in diagnosis of dengue 1mavra fatima, 1ayesha younas, 1ayisha imran, 1asma nasir, 1nauman aslam malik, 2omar rasheed chughtai, 2akhtar sohail chughtai 1department of hematology, 2department of histopathology, chughtai institute of pathology, lahore abstract introduction: pakistan is dealing with an epidemic situation of dengue. serological testing for its diagnosis is not available everywhere across the country. so, in the current scenario, basophilia flagged by automated hematology analyzer can be a helpful screening tool in early diagnosis and prognosis of dengue in a resource limited country like ours. aims & objectives: to assess the utilization of basophilia flag as a screening tool in early diagnosis of dengue by studying the frequency of basophilia flag and it’s prognostic significance by correlating absolute basophil count with severity of thrombocytopenia. place and duration of study: it was a cross sectional study and conducted at chughtai institute of pathology from august 2021 to october 2021. material & methods: total 1007 patients who had ns1 positive confirmed dengue infection were included in the study and edta blood samples were run on mindray bc-6800 six-part hematology analyzer. basophilia flag was noted, its frequency was calculated and expressed as percentage. also, correlation of absolute basophil count with platelet count was calculated. statistical analysis was performed using spss 23.0, p value <0.05 was taken as significant. results: wbc flag showing “basophilia” was seen in 136 patients (13.6%) and a significant correlation was seen between raised absolute basophil count and thrombocytopenia using pearson test. conclusion: pseudobasophilia is an important screening tool in diagnosing dengue patients and as significant prognostic marker as increased absolute basophilic count correlates with severity of thrombocytopenia. key words: dengue, ns1 antigen, pseudobasophilia, automated hematology analyzer. introduction dengue fever is becoming more prevalent in pakistan as well as globally.1 there has been a history of outbreaks of dengue fever in country since 1994 when first confirmed outbreak of dengue fever occurred. the next one happened in 2010 and recently we suffered from this epidemic in 2021. dengue is a mosquito borne disease caused by dengue virus.2 patients commonly present with nausea, vomiting, high grade fever, joint pains (hence the name bone breaking fever) and skin rashes during febrile phase. this phase is followed by critical phase and recovery phase.3 basic testing for dengue includes complete blood count (cbc) which reveals changes in its parameters including hemoglobin (hb), hematocrit (hct), platelet count, mean platelet volume (mpv), total white cell count (tlc), differential count including neutrophils, lymphocytes, monocytes, eosinophils and basophils (percentages as well as absolute counts). differential white cell count can be helpful in diagnosis and prognosis of dengue in resource limited areas.4 certain white cell flags generated by automated hematology analyzers can be a helpful screening tool in diagnosing and assessing the severity of dengue fever.5 a high percentage or absolute count of basophils is defined as basophilia. it is commonly seen in allergic and acute inflammatory conditions and is also a characteristic feature of chronic myeloid leukemia.6 the instruments detect basophils by flowcytometry using a semi conductor laser.7 pseudobasophilia is indicated by cells other than basophils which can be seen in bone 31 pseudobasophilia: a helpful screening tool in diagnosis of dengue marrow infiltration and myeloma .8 in pakistan, the phenomenon of pseudobasophilia is appreciated owing to the presence of reactive lymphocytes.9 material and methods a cross sectional study was conducted at chughtai institute of pathology from august 2021 to october 2021.approval was obtained from the ethical and research committee of the institute. total 1000 patients, both males and females of all age groups, who had ns1 positive confirmed dengue infection were included in the study. two ml of peripheral blood sample was taken from each patient in edta tube following standard procedures. samples were run on mindray bc-6800 six part hematology analyzer. basophilia flag was noted and correlated with peripheral smear examination. suspected dengue patients who were ns1 negative were excluded from this study. statistical analysis was done using spss 23.0. frequencies were calculated and expressed as percentages. correlation of absolute basophil count and severity of thrombocytopenia was observed using pearson test. p value <0.05 was taken as significant. statistical analysis: data analysis was done using spss 23.0. frequencies were calculated and expressed as percentages. correlation of absolute basophil count and severity of thrombocytopenia was observed using pearson test. p value <0.05 was taken as significant. results 817 male, 190 female (total 1007) dengue patients were included in the study with mean age 40.77±15.58 years (table-1). cbc showed mean hb 15.75±6.5 g/dl, hct 46.9±5.3 %, tlc 5.8±3.1 x 109/l, platelet count 27.5±26.04 x 109/l, absolut basophil count (abc) 0.08±0.23 (table-2). a significant correlation (p = <0.004) was observed between increased absolute basophil count (abc) and fall in platelet count using pearson test. (table-3). our study showed 136 dengue samples with flagging “basophilia” but when peripheral smears were prepared from these samples, there was no increase in basophils (hence the term pseudobasophilia). however, reactive lymphocytes with basophilic cytoplasm were seen on the smears which were falsely counted as basophils by automated hematology analyzer. characteristics values mean age (years) 40.77±15.58 male 817 (81.1%) female 190 (18.9%) table-1: mean age and gender frequency of patients hb (g/dl) hct (%) tlc (×109/l) platelets (×109/l) abc mean 15.75 46.92 5.83 27.51 0.0889 sd 6.527 5.347 3.178 26.048 0.235 table-2: hematological parameters hb: hemoglobin, hct: hematocrit, tlc: total leukocyte count, abc: absolute basophil count platelets abc platelets pearson correlation 1 -.119** sig. (2-tailed) .000 n 1007 1007 abc pearson correlation -.119** 1 sig. (2-tailed) .000 n 1007 1007 **correlation is significant at the 0.01 level (2-tailed). table-3: correlation between abc (absolute basophil count) and thrombocytopenia discussion cbcs of 1007 dengue patients who were confirmed ns1 positive were analyzed which showed significant findings like rise in hb, hct, abc and decrease in white cell count and platelet count. along with these parameters, “basophilia” flags generated by automated hematology analyzer mindray bc6800 were analyzed. a higher hemoglobin and hematocrit level in dengue is due to increased vascular permeability; with the highest values seen on day 7.10 a study by martina et al, showed that the factors resulting in plasma leakage are cytokine storm and cross reactivity of antins1 antibodies which result in the apoptosis of the endothelial cells.11 basophilia is generally seen in conditions like infections, inflammation and also in myeloid neoplasms.12 however in dengue patients basophilia flag suggests presence of atypical or 32 pseudobasophilia: a helpful screening tool in diagnosis of dengue reactive lymphocytes owing to the infective process. hence, the instrument gives falsely raised basophil count; that’s why the term pseudobasophilia is used. to confirm this finding, peripheral smear should be prepared and examined carefully. in dengue patients, basophilia flag gives a hint of infection which can be further confirmed on peripheral smear where basophilic reactive lymphocytes can be appreciated.13 our study showed 136 dengue samples with flagging “basophilia” but when peripheral smears were prepared from these samples, there was no increase in basophils (hence the term pseudobasophilia). however, reactive lymphocytes with basophilic cytoplasm were seen on the smears which were falsely counted as basophils by automated hematology analyzer. in a study from india, 52.9% dengue cases had basophils more than 2%.14 in a research carried out in thailand, basophil count was within normal range.10 in another endemic area, 91.2% dengue cases flagged pseudobasophilia and thrombocytopenia when the samples were run on sysmex xe-2100 analyzer. however, when peripheral smears of such cases were examined; the only significant finding was reactive lymphocytes.15 the variability of the basophil count depends on the time of sample collection. researches reveal a weak synchroneity between instruments regarding the basophil count.8,16 pseudobasophilia in dengue is an underreported phenomenon which was most commonly associated with flags of “atypical lymphocytes” and “blasts” given by the hematology instruments.17 our study also shows similar results with high frequency of basophilia along with atypical lymphocytes flag on mindray bc6800. increased abc also showed correlation with severity of thrombocytopenia i.e with increasing absolute basophil count there was a marked decrease in platelet count. conclusion pseudobasophilia can be used as a helpful tool in the diagnosis of dengue in a resource restricted country like pakistan where serological confirmation of the disease is costly and not easily available across the country. also early detection of severe thrombocytopenia by using basophil flag in dengue patients can prevent them from bleeding complications. references 1. ahmed s, ali n, ashraf s, ilyas m, waheeduz-zaman tariq and chotani r a. dengue fever outbreak: a clinical management experience. journal of the college of physicians and surgeons pakistan 2008, vol. 18 (1): 8-12. 2. akram m, fatima z, purdy ma, sue a, saleem s, et al. introduction and evolution of dengue virus type 2 in pakistan: a phylogeographic analysis. virology j. 2015;12:148. 3. malavige gn, fernando s, fernando dj, seneviratne sl .dengue viral infections. postgraduate medical journal 2004;80:588601. 4. a.joshi,gayathri b.r, fazeela muneer.dynamics of differential count in dengue. international journal of advances in medicine, vol.5, no.1(2018). 5. roy, maitrayee, and akshay bali. "m2g1g2 white blood cell flag by three-part automated hematology analyzer: a hint to dengue infection in appropriate clinical context." journal of laboratory physicians. 2019;11(2):103-106 6. galli sj, metcalfe dd, arber da, dvorak am, basophils, mast cells, and related disorders. in: kaushansky k, lichtman ma, prchal jt, et al, eds. williams hematology. 9th ed. new york: mcgraw-hill education; 2016:971-972. 7. mannem c, krishanmurthy t, gayathri b. r., prabhu pd. characterization of pseudobasophilia on sysmex-xt 1800i automated hematology analyser. international journal of research in medical sciences. 2017 jul;5(7):2912-2916. 8. gibbs g, campbell g, christie i. pseudobasophilia and the advia 120. hematology 2009;14(3):159-163. 9. la russa vf, innis bl. mechanisms of dengue virus induced bone marrow suppression. baillieres clin haematol. 1995; 8(1):249-70. 10. chaloemwong j, tantiworawit a, rattanathammethee t, et al. useful clinical features and hematological parameters for the diagnosis of dengue infection in patients with acute febrile illness: a retrospective study. bmc hematol 2018; 18:20. 11. martina be, koraka p, osterhaus ad. dengue virus pathogenesis: an integrated view. clinmicrobiol rev 2009; 22(4):564-581. 12. chandrashekar v. basophil differentials as a marker for atypical lymphocyte morphologic 33 pseudobasophilia: a helpful screening tool in diagnosis of dengue characteristics. lab med. 2013; 44(02):133135. 13. kevin manuel,1 marie moses ambroise1, anita ramdas,1 and renu g'boy varghese, pseudobasophilia as a screening tool in dengue: a single center study. j lab physicians. 2021 jun; 13(2): 156–161. 14. malathesha m k, ashwini h n. hematological manifestations in dengue fever—an observation study. j evol med dent sci. 2014; 3(09):2245-50. 15. pai s. pseudobasophilia on the sysmex-xe 2100: a useful screening tool for primary dengue infection in endemic area. int j lab hematol. 2012; 34:25 16. amundsen e k, henriksson c e, holthe m r, urdal p. is the blood basophil count sufficiently precise, accurate, and specific?: three automated hematology instruments and flow cytometry compared. am j clin pathol. 2012; 137(01):86-92. 17. jácomo r h, lozano v f, da cunha neto j g, costa s s. what’s the meaning of basophilia in sysmex xe-2100? arch pathol lab med. 2011; 135(04):415. the authors: dr. mavra fatima, consultant, department of hematology, dr. ayesha younas, p.g. trainee, department of hematology, dr. ayisha imran, consultant, department of hematology, dr. asma nasir, consultant, department of hematology, dr. nauman aslam malik, head, department of hematology, dr. omar rasheed chughtai, assistant professor histopathology department of histopathology, prof. akhtar sohail chughtai, department of histolopathology, (chughtai institute of pathology, lahore) corresponding author: dr. mavra fatima consultant, department of hematology, chughtai institute of pathology, lahore. e-mail: drmavrafatima@yahoo.com 34 untitled-1 13 proceedings s.z.m.c. vol: 34(3): pp. 13-17, 2020. pszmc-755-34-3-2020 chances of pneumothorax and malpositioning of central venous catheters in internal jugular vein versus subclavian vein routes 1riffat saeed, 1muhammad naveed shahzad, 2zia qazi, 3iram qamar, 4amna javed, 5arif javed 1department of anesthesia, shaikh zayed medical complex, lahore 2department of ent, shaikh zayed medical complex, lahore 3department of anesthesia, mayo hospital, lahore 4department of gynecology & obstetrics, shaikh zayed medical complex, lahore 5department of general surgery, shaikh zayed medical complex, lahore abstract introduction: central venous catheterization (cvc) is being done all over the world. it has specific indications and should be reserved for the patient who has the potential to benefit from it. catheter related infections are an important cause of morbidity and mortality worldwide. all complications and side effects are dependent on vascular access route. international data shows malpositioning and pneumothorax related to malpositioning to be the most common complications of central venous cannulation. however there is paucity of local data regarding which of the two, ijv or scv routes are more prone to develop these complications. aims & objectives: to compare the incidence of pneumothorax and malpositioning with internal jugular vein (ijv) and subclavian vein (scv) routes of central venous catheters. place and duration of study: this randomized control trial was conducted at department of anesthesia, shaikh zayed hospital, lahore, from 812-2014 to 7-6-2015. material & methods: the non-probability purposive sampling technique was used in this study. after the approval of hospital ethical committee, 290 patients were included in this study and informed consent was obtained. demographic profile was also obtained. patients were randomly divided in two groups by using lottery method. in group a, cvc was inserted through internal jugular vein while in group b, cvc was inserted through subclavian vein. during the procedure, malpositioning and pneumothorax were monitored immediately and after 36 hours and were labeled. patients were shifted to the ward after procedure and were followed-up there. during first 36 hours, chest x-ray for placement of tip of catheter and development of pneumothorax was carried out. chi-square was used to compare complications in both groups taking p value <0.05 as significant. results: malposition was found in 18 cases, (6 from ijv group and 12 from scv group) (p-value 0.144). pneumothorax was seen in 12 cases (3 from ijv and 9 from scv group) (p-value 0.077). conclusion: our study results concluded that ijv showed fewer incidences of pneumothorax and malpositioning than scv technique. however, the difference was not statistically significant. key words: internal jugular vein: ijv, central venous catheterization: cvc, subclavian vein: scv, complications. introduction central venous catheterization is being done all over the world. it has specific indications and should be reserved for the patient who has the potential to benefit from it.1 the indications for cvc include central venous pressure monitoring, inadequate peripheral venous access, cardiopulmonary resuscitation, long term intravenous therapy (chemotherapy, hemodialysis, hyper alimentation) as an adjunct to pulmonary artery catheterization and inotropic support.2 central venous access can be achieved by different routes, utilizing femoral, axillary, internal jugular and subclavian veins, but the route favored by most centers is the internal jugular or subclavian veins. all complications and side effects depend on vascular access route.3 malpositioning and pneumothorax due to malpositioning are the most common complications of passing central lines, upto 30% of all mechanical detrimental events. varying with the number of 14 chances of pneumothorax and malpositioning of central venous catheters in internal jugular vein attempts in emergent conditions where large catheters like dialysis catheters are inserted.4 overall complication rates range upto 15%, mechanical complications reported in 19% of patients, 26% infectious complications, and 26% thrombotic complications.5 it is seen that incidence of pneumothorax varies in different studies.6 luyt et al., conducted a randomized trial and reported that the incidence of malposition was reported to be low with ijv (1.8%) as compared to scv (7.4%), however, the difference was insignificant (p=0.07) and the incidence of pneumothorax was also nil (0) with ijv as compared to scv (3.2%), (p=0.09).7 due to the presence of conflicting international data and scarcity of our own, the research was designed to determine these complications in relation to ijv & scv routes in our local setting. material and methods study design: cross sectional survey. setting: department of anesthesia, shaikh zayed medical complex, lahore. duration: six months after the approval of synopsis. sample size: sample size of 60 cases (30 in each group) is calculated in both groups with 95%confidence level, 10% margin of error, and taking expected percentage of complications in both groups i.e. 26%. sampling technique: non probability consecutive sampling. statistical analysis: all the data was entered and analyzed through spss version 20.0. the quantitative variables like age were presented as mean & standard deviation. the qualitative variable like gender, malpositioning and pneumothorax were presented as frequency and percentages. chi-square was used to compare complications in both groups taking p value <0.05 as significant. results the mean age of the patients was 45.42±15.89 years with minimum age of 18 years and maximum age of 70 years (table-1). in our study out of 290 cases 61.72% patients were males and 38.28% patients were females, (male to female ratio of 1.6:1) (fig1). the study results showed that malpositioning was found in 18(6.2%) patients and it was not found in 271(93.8%) patients (table-2). in this study the pneumothorax was observed in 12 (4.14%) patients whereas it was not observed in 278 (95.86%) patients (fig-2). malposition was found in 18 cases in which 6 patients were from ijv group and 12 were from scv group, similarly malposition was not found in 272 cases in which 139 were from ijv group and 133 were from scv group. statistically insignificant incidence of malpositioning was found in both groups (pvalue0.144) (table-3). pneumothorax was observed in 12 cases, in which 3 patients were from ijv group and 9 were from scv group, similarly pneumothorax was not observed in 278 cases in which 142 were from ijv group and 136 were from scv group. statistically the difference of incidence of pneumothorax between the two groups was nonsignificant (p0.077) (table4). data was stratified for age and it was noticed that in patients of age<40 years, malpositioning was found in 2 cases in ijv group and 6in scv group. while in patients of age ≥40 years, malpositioning was found in 4 cases in ijv group and 6 in scv group. statistically the difference was nonsignificant with a p value >0.05 (table-5). data was stratified for age and it was noticed that in patients of age<40 years, the pneumothorax was observed in 1 case in ijv group and 4 in scv group. similarly, pneumothorax was observed in 2 cases in ijv group and 5in scv group. statistically the difference was nonsignificant with p value>0.05 (table-6). data was stratified for gender and malpositioning was found in 3 cases in ijv group and 5in scv group in male participants. while in female patients, malpositioning was found in 3 cases in ijv group and 7 in scv group. statistically nonsignificant difference was found between the two groups with p>0.05 (table-7). data was stratified for gender and it was noticed that in male patients, the pneumothorax was observed 1 case in ijv group and 3 in scv group. similarly, pneumothorax was observed in 2 cases in ijv group and 6in scv group. statistically the difference was nonsignificant between the two groups with a p value>0.05 (table8). age (years) n 290 mean 45.42 sd 15.89 minimum 18.00 maximum 70.00 table-1: descriptive statistics of age (years) 15 chances of pneumothorax and malpositioning of central venous catheters in internal jugular vein fig-1: percentage. distribution of gender no. percent malposition yes 18 6.2 no 271 93.8 table-2: no. distribution of malposition study groups total ijv scv malposition yes 6 12 18 no 139 133 272 total 145 145 290 table-3: comparison of malpositioning in both groups study groups total ijv scv pneumothorax yes 3 9 12 no 142 136 278 total 145 145 290 table-4: comparison of pneumothorax in both groups fig-2: no. distribution of pneumothorax table-5: comparison of malpositioning in both groups stratified by age table-6: comparison of pneumothorax in both groups stratified by age gender malpositioning study groups p-value ijv scv male yes 3 5 0.513 no 51 52 female yes 3 7 0.175 no 88 81 table-7: comparison of mal-positioning in both groups stratified by gender table-8: comparison of pneumothorax in both groups stratified by gender discussion the choice of anatomical site of central catheter insertion should be on evidence based guidelines instead of personal preferences. the data on catheter malpositioning may have more impact on clinical decision-making. malpositioning was reported in 14% of cases even with expert practitioners.10 in our study the ijv and scv procedures were applied.overall complication of malposition was found in 18(6.2%) patients and complication of pneumothorax was observed in 12(4.14%) patients.. according to our study were did not find any significant difference between the complications (pneumothorax & malposition) among the study groups. some of the studies are discussed below showing the results in favor of our study. age mal-positioning study groups p value ijv scv <40 years yes 2 6 0.157 no 55 53 ≥40 years yes 4 6 0.491 no 84 80 age pneumothorax study groups pvalue ijv scv <40 years yes 1 4 0.183 no 56 55 ≥40 years yes 2 5 0.235 no 86 81 gender pneumothorax study groups p-value ijv scv male yes 1 3 0.335 no 53 54 female yes 2 6 0.135 no 89 82 16 chances of pneumothorax and malpositioning of central venous catheters in internal jugular vein sibylle ruesch et al concluded in their study that there was lesser incidence of catheter malposition and more incidences of arterial punctures with internal jugular vein route compared with the other access. there was no difference in incidence of hemothorax, pneumothorax or vessel occlusion.8 luyt et al., conducted a randomized trial and found that malposition was reported to be low with ijv (1.8%) as compared to scv (7.4%, however the difference was insignificant (p=0.07) as well as number of pneumothorax was also nil (0) with ijv as compared to scv (3.2%, p0.09).7 eisenhauer et al reported in their study that the rate of complications was 13.7%. there were 286 cannulations done in subclavian vein, having 12 complications out of 13 total morbidities in the study (incidence 4.2%), whereas in 248 internal jugular cannulations, there was only 1 case of morbidity (0.4%). so it is recommended on the basis of these results, that internal jugular vein route should be preferred, and the subclavian route should be reserved for cases where internal jugular approach is technically difficult or for patients requiring parenteral nutrition.16 different studies have showed results related to malposition. they have stated that jugular access was reported to have lesser catheter malposition.9,12,13,14,15 peres et al., disagreed with the results of luyt and reusch studies and found that malposition was 18.97% with ijv but 33.6% with scv which was significant (p0.01) however, pneumothorax was nil in both groups.9 mcgee, et al., investigated if the use of 16cm central venous catheters minimized intracardiac placements. 127 patients were assessed using either of the two routes. 16cm catheters were used in 102 patients and 20cm in 25 cases. in conclusion, using 16cm catheters through either of the internal jugular or subclavian routes had more incidences of safe catheterizations. it was recommended to make smaller catheter use as standard practice.17 in a case series, a technique was introduced in which the site of venous puncture was at the junction of subclavian and internal jugular veins, 2 to 3 cm above clavicle, close to the posterior border of sternocleidomastoid. it was successful in 94% cases, with complications (5%) of, 6 thoracic duct cannulations, 4 arterial punctures and 2 cases of pneumothorax in a total of 600 cases.18 non-randomized studies have reported ambiguous conclusions for complications like arterial puncture, arrhythmias, hematoma, pneumothorax or hemothorax when the internal jugular vein route is compared to the subclavian route.11,16,18 conclusion it is concluded that the incidence of malpositioning and pneumothorax is related more to the subclavian vein route as compared to the internal jugular route, however, the difference is not statistically significant. references 1. celinski sa, snafu mg. central venous catheters. in: irwin rs, rippe jm, editors. irwin and rippe's intensive care medicine. 7th ed. philadelphia: lippincott williams & wilkins; 2011: p 19-37. 2. ray br, mohan vk, kashyap l, shende d, darlong vm, pandey rk. internal jugular vein cannulation: a comparison of three techniques. journal of anaesthesiology, clinical pharmacology. 2013; 29(3):367. 3. kumar a, gupta k, bhandari s, singh r. folding back of central venous catheter in the internal jugular vein: methods to diagnose it at the time of insertion? indian journal of anaesthesia. 2013; 57(1):104. 4. marik pe, flemmer m, harrison w. the risk of catheter-related bloodstream infection with femoral venous catheters as compared to subclavian and internal jugular venous catheters: a systematic review of the literature and meta-analysis. critical care medicine. 2012; 40(8):2479-85. 5. sayed aa, johar a, nazir a, qasim m. central venous pressure line. professional med j. 2009; 16:44-7. 6. rezende-neto j, hoffmann j, al mahroos m, tien h, hsee l, spencer netto f, et al. occult pneumomediastinum in blunt chest trauma: clinical significance. injury. 2010; 41(1):40-3. 7. luyt d, mathivha l, litmanovitch m, dance m, brown j. confirmation of the safety of central venous catheterisation in critically ill infants and children-the baragwanath experience. south african med j 1996; 86(5suppl):603-6. 8. ruesch s, walder b and tramèr m. complications of central venous catheters: internal jugular versus subclavian access—a systematic review. critical care medicine, 2002; 30(2), pp.454-460. 9. peres p. positioning central venous catheters – a prospective survey. anaesthesia and intensive care, 1990; 18(4), pp.536-539. 10. gladwin mt, slonim a, landucci dl, gutierrez dc, cunnion re. cannulation of the 17 chances of pneumothorax and malpositioning of central venous catheters in internal jugular vein internal jugular vein: is postprocedural chest radiography always necessary? critical care medicine. 1999; 27(9):1819-23. 11. sznajder j, zveibil f, bitterman h, weiner p and bursztein s. central vein catheterization. failure and complication rates by three percutaneous approaches. survey of anesthesiology, 1986; 30(4), p.196. 12. barrera r, mina b, huang y, groeger js. acute complications of central line placement in profoundly thrombocytopenic cancer patients. cancer. 1996; 78(9): 2025-30. 13. kaiser cw, koornick ar, smith n, soroff hs. choice of route for central venous cannulation: subclavian or internal jugular vein? a prospective randomized study. journal of surgical oncology. 1981; 17(4):345-54. 14. litmanovitch m, hon h, luyt d, dance m, mathivha l. comparison of central venous pressure measurements in the intrathoracic and the intra-abdominal vena cava in critically ill children. anaesthesia. 1995; 50(5):407-10. 15. reed c, sessler c, glauser f, phelan b. central venous catheter infections: concepts and controversies. intensive care medicine. 1995; 21(2):177-83. 16. eisenhauer ed, derveloy rj, hastings pr. prospective evaluation of central venous pressure (cvp) catheters in a large city-county hospital. annals of surgery. 1982; 196(5):560. 17. mcgee wt, moriarty kp. accurate placement of central venous catheters using a 16-cm catheter. journal of intensive care medicine. 1996; 11(1):19-22. 18. haapaniemi l, slätis p. supraclavicular catheterization of the superior vena cava. acta anaesthesiologica scandinavica. 1974; 18(1):12-22. 19. molgaard o, nielsen m, handberg b, jensen j, kjaergaard j, juul n. routine x-ray control of upper central venous lines: is it necessary? acta anaesthesiologica scandinavica. 2004; 48(6): 685-9. the authors: dr. riffat saeed assistant professor, department of anesthesia, shaikh zayed medical complex, lahore. dr. muhammad naveed shahzad anesthetist, department of anesthesia shaikh zayed medical complex, lahore. dr. zia qazi assistant professor, department of ent, shaikh zayed medical complex, lahore. dr. iram qamar assistant professor, department of anesthesia, mayo hospital, lahore. dr. amna javed assistant professor, department of gynecology & obstetrics, shaikh zayed medical complex, lahore. dr. arif javed assistant professor, department of general surgery, shaikh zayed medical complex, lahore. corresponding author: dr. riffat saeed assistant professor, department of anesthesia, shaikh zayed medical complex, lahore. e-mail: zahidhnch@hotmail.com 50 proceedings s.z.m.c. vol: 35(3): pp. 50-53, 2021. pszmc-808-35-3-2021 prevalence of bacterial vaginosis during pregnancy in tertiary care hospital, lahore 1amna javed, 1muhammad ikram, 1shafqat mukhtar, 3saulat sarfraz, 1ayesha shahid, 2aman ur rehman 1department of obstetrics and gynecology, shaikh zayed medical complex, lahore 2department of histopathology, shaikh zayed medical complex, lahore 3department of radiology, shaikh zayed medical complex, lahore abstract introduction: pregnancy is a condition usually associated with increased vaginal discharge including vaginal infections like bacterial vaginosis, leading to adverse perinatal outcomes due to disturbance in vaginal ecosystem that warrants further evaluation and timely management. aims & objectives: the study was conducted to determine the prevalence of bacterial vaginosis in pregnancy and its contributing factors. place and duration of study: out patient department of obstetrics & gynecology, shaikh zayed hospital, lahore. duration of the study was six months. material & methods: 120 pregnant females with age 20-40 years with singleton pregnancy of more than 12 weeks’ gestation were included. women who were immunosuppressed and history of diabetes and hypertension and recent history of use of antibiotics were excluded. vaginal swab specimen was taken & bacterial vaginosis was labelled after fulfilling amsel criteria. data was analyzed using spss version 21. results: in this study, 120 cases were taken and mean age of participants was 27.55±3.97 years. the mean duration of gestation was 18.76±6.14 weeks. bacterial vaginosis (bv) was seen in 24 (20%) of the cases. bv was seen in 19 (20.65%) cases with age 20-29 years’ vs 5 (17.86%) cases in age group 20-40 years with p= 1.0. bacterial vaginosis was nearly significantly high in multiparous women where this was seen in 13 (27.65%) cases as compared to 11 (15.06%) cases in primiparous women with p= 0.11. out of 20 cases of bacterial vaginosis, low education was seen in 12 (60%) of the cases. conclusion: the prevalence of bacterial vaginosis in pregnancy at local setup is high and is seen nearly in 1 out of 5 cases. this demands health educational programs to enlighten women regarding screening and early detection & treatment to prevent complications during pregnancy. key words: amsel criteria, bacterial vaginosis, female genital health, pregnancy, vaginal flora. introduction normally, vaginal ecosystem is characterized by the presence of gram-positive lactobacillus which plays an important role in maintaining female genital health. disturbance in vaginal ecosystem and simultaneous replacement of normal flora with anaerobic bacteria leads to bacterial vaginosis causing malodorous vaginal discharge.1 pregnancy is a condition usually associated with increased vaginal discharge including vaginal infections like bacterial vaginosis, leading to adverse perinatal outcomes including risk of late miscarriages, preterm labor, risk of chorioamnionitis and postpartum wound infections or endometritis.2,3 the prevalence of bacterial vaginosis ranges from 5% to 58.5% in different population and in pregnancy, its prevalence is 11-16% in developed countries.2,3 recent studies showed significant reduction in adverse outcomes with the help of appropriate screening and treatment.4 due to adverse perinatal outcomes associated with bacterial vaginosis especially in pregnancy, pathological vaginal discharge warrants further evaluation and timely management. in spite of 20% prevalence of bacterial vaginosis in pregnancy, evidence of epidemiology of bv in pregnancy in developing countries is still scarce.5 this study aims to provide data on prevalence of bacterial vaginosis among pregnant females and its contributing factors for early screening and detection which will be beneficial to prevent adverse perinatal outcome and improve quality of health. material and methods a cross sectional study was done in out patient department of obstetrics & gynecology, shaikh zayed hospital, lahore from 01-04-2019 to 01-10 51 prevalence of bacterial vaginosis during pregnancy in tertiary care hospital, lahore 2019 enrolling 120 pregnant females with age 20-40 years with singleton pregnancy of more than 12 weeks gestation via non probability consecutive sampling technique. women who were immunosuppressed and history of diabetes and hypertension and recent history of use of antibiotics (within 4 weeks) were excluded. after the approval of ethical committee of shaikh zayed hospital (szh), lahore an informed consent was taken from each patient fulfilling the inclusion criteria. socio demographics and clinical data like age, parity, monthly household income, level of education and duration of gestation at presentation was taken and recorded on a specially designed proforma. all cases underwent vaginal examination by cusco’s speculum and a sterile swab specimen was taken from vagina and was examined by using amsel criteria: (three of the following four criteria must be present)6: presence of homogenous whitegrey vaginal discharge, presence of clue cells under microscope & a fishy amine odor of the vaginal discharge before and after addition of 10% koh (positive whiff test); a vaginal ph of > 4.5. statistical analysis: data was analyzed with the help of spss version 21. quantitative variables like age and duration of gestation at presentation were documented in terms of mean ± sd (standard deviation). frequency & percentages were calculated for parity, monthly household income and bacterial vaginosis. effect modifiers were controlled through stratification of age, parity, monthly household income, educational status, trimester of pregnancy to see the effect on bacterial vaginosis. p-value ≤ 0.05 was taken as significant. results in this study there were total 120 cases and the mean age of the participants was 27±3.97 years and mean duration of gestation was 18 ±6.14 weeks as in table-1. frequency of parity and monthly household income is depicted in table-2. bacterial vaginosis (bv) was seen in 24 (20%) of the cases (table-2). bv was seen in 19 (20.65%) cases with age 20-29 years vs 5 (17.86%) cases in age group 30-40 years with p=1.0. bacterial vaginosis frequency was increased in multiparous women i.e. 13 (27.65%) cases as compared to 11 (15.06%) cases in primiparous women with p= 0.11. also it was high in women with low education 19 (22.35%) as compared to 5 (14.28%) with higher education (p=0.34) there was no significant association of bv with monthly household income and trimester of pregnancy (table-3). age (years) duration of gestation (weeks) mean 27 18 std. deviation 3.97 6.14 table-1: age & duration of gestation at presentation in study subjects (n= 120) factors frequency n=120 percentage parity primiparous multiparous 73 47 60.83% 39.17% monthly household income <rs. 20,000/ >rs. 20,000 72 48 60.00% 40.00% bacterial vaginosis present absent 24 96 20.00% 80.00% table-2: frequency and percentage of factors in study subjects (n=120) sociodemographic bacterial vaginosis total p value yes no age 20-29 30-40 total 19 (20.65%) 5 (17.86%) 24 (20%) 73 (79.35%) 23 (82.14%) 96 (80%) 92 (100%) 28 (100%) 120(100%) 1.0 parity primiparous multiparous total 11 (15.06%) 13 (27.65%) 24 (20%) 62 (84.94%) 34 (72.35%) 96 (80%) 73 (100%) 47 (100%) 120(100% 0.11 monthly household income <rs 20,000/ >rs. 20,000/ total 14 (19.44%) 10 (20.83%) 24 (20%) 58 (80.56%) 38 (79.17%) 96 (80%) 72 (100%) 48 (100%) 120(100% 1.0 educational status under-graduate graduate&above total 19 (22.35%) 5 (14.28%) 24 (20%) 66 (77.65%) 30 (85.72%) 96 (80%) 85 (100%) 35 (100%) 120(100% 0.34 gestational age (trimester wise) 2nd 3rd total 14 (17.5%) 10 (25%) 24 (20%) 66 (82.5%) 30 (75%) 96 (80%) 80 (100%) 40 (100%) 120(100% 0.61 table-3: association of bacterial vaginosis with sociodemographic factors. (n= 120) discussion globally bacterial vaginosis is an emerging gynaecologic concern especially in women of childbearing age because this is considered as one of 52 prevalence of bacterial vaginosis during pregnancy in tertiary care hospital, lahore the risk factors for adverse perinatal outcomes including risk of miscarriages, preterm labour, preterm premature rupture of membranes, and postpartum wound infections.7 the methods for diagnosis of bacterial vaginosis also have a great impact on the variation of bacterial vaginosis prevalence. although the gold standard for diagnosis of bv is proposed by nugent. but the clinical criteria by amsel6 is suitable for daily routine as it does not require laboratory facilities hence, no delay in reporting. it is arduous to evaluate all of these criteria for diagnosis in busy practice; and requires the ability of the gynaecologist to analyze wet mount microscopy.8,9 in the present study bacterial vaginosis (bv) was seen in 24 (20%) out of the 120 cases. these results were comparable to the findings of the previous studies where almost similar burden of the disease was found. according to a study done by ibrahim sm et al, the bacterial vaginosis was seen in 17.3% of pregnant cases.10 slightly higher percentages were noted by the results of other studies, where they revealed this percentage in more than 30% of the cases. romoren m et al. revealed bv in 38% of the pregnant ladies.11 while in another study by habib a & siddiqui i revealed its prevalence as 35.3% of cases.12 this variability of the results can be due to difference in criteria for diagnosis. in another study regarding the prevalence and it contributing factors this was noted in 19.4%, in their study by mangistie z et al.13 the incidence of bv is 30-35% in a study conducted by ries aj.14 however, study by gul f, et al. showed frequency of bv of about 2.5% in women presenting to antenatal clinic of a tertiary care hospital in peshawar.15 furthermore, regarding various contributing factors leading to bacterial vaginosis, mengistie z et al. found higher number of cases having bacterial vaginosis in younger age (21-29 year) where this was seen in 21.2%, low educational status (less than 12 grade) in 23.1% and 3rd trimester pregnancy in 14% of the cases.13 these results were also comparable to the present study findings, where bv was seen in 19 (20.65%) cases with age 20-29 years’ vs 5 (17.86%) cases in age group 20-40 years with p= 1.0; though this difference was statistically insignificant. and among various contributing factors leading to bacterial vaginosis, low education was seen in 19 (22.35%) of the cases while 3rd trimester of pregnancy had 10 (25 %) cases and younger age group was seen in 19 (20.65%) cases each. in another study conducted by chaudhury and colleagues, the peak age of bv was 25-34 years.16 in index study the peak age of infection was 20-29 years. there were few limitations of this study as this study did not look for concomitant contributing factors like urinary tract infection which was an important confounder and also did not look for the feto -maternal outcomes. however, there were many strengthening points as well as this study highlighted a very underrated entity. conclusion the prevalence of bacterial vaginosis in pregnancy at local setup is high and is seen nearly in 1 out of 5 cases. among various contributing factors low education was the commonest. this demands for health educational programs to enlighten women regarding screening and early detection & treatment to prevent complications during pregnancy. references 1. machado d, castro j, palmeira-de-oliveira a, martinez-de-oliveira j, cerca n. bacterial vaginosis biofilms: challenges to current therapies and emerging solutions. frontiers in microbiology. 2016 jan 20; 6:1528. 2. afolabi bb, moses oe, oduyebo oo. bacterial vaginosis and pregnancy outcome in lagos, nigeria. in open forum infectious diseases 2016 jan 1 (vol. 3, no. 1, p. ofw030). oxford university press. 3. aduloju op, akintayo aa, aduloju t. prevalence of bacterial vaginosis in pregnancy in a tertiary health institution, south western nigeria. the pan african medical journal. 2019; 33. 4. mcgregor ja. bacterial vaginosis in pregnancy. obstetrical & gynecological survey. 2000 may 1; 55(5):1-9. 5. ibrahim sm, bukar m, galadima gb, audu bm, ibrahim ha. prevalence of bacterial vaginosis in pregnant women in maiduguri, north-eastern nigeria. nigerian journal of clinical practice. 2014 mar 26; 17(2):154-8. 6. amsel r, totten pa, spiegel ca, chen kc, eschenbach d, holmes kk. nonspecific vaginitis: diagnostic criteria and microbial and epidemiologic associations. the american journal of medicine. 1983 jan 1; 74(1):14-22. 7. muzny ca, schwebke jr. pathogenesis of bacterial vaginosis: discussion of current hypotheses. j infect dis. 2016; 214(suppl 1):s1. 8. nelson tm, borgogna jl, brotman rm. vaginal biogenic amines: biomarkers of bacterial vaginosis or precursors to vaginal dysbiosis? front physiol. 2015; 6:253. 9. shayo pa, kihurwa a, massinde an, mirambo m, rumanykia r, ngwaalida n, et al. prevalence of bacterial vaginosis and associated factors among 53 prevalence of bacterial vaginosis during pregnancy in tertiary care hospital, lahore pregnant women attending at bugando medical centre, mwanza, tanzania. tanz j health sci. 2012; 14(3):1-7. 10. ibrahim sm, bukar m, galadima gb, audu bm, ibrahim ha. prevalence of bacterial vaginosis in pregnant women in maiduguri, north-eastern nigeria. niger j clin pract. 2014; 17(2):154-58. 11. romoren m, velauthapillai m, rahman m, sundby j, klouman e, hjortdahl p. trichomoniasis and bacterial vaginosis in pregnancy: inadequately managed with the syndromic approach. bull world health organ. 2007; 85:297–304. 12. habib a, siddiqui i. frequency of bacterial vaginosis in patients with vaginal discharge. khyb med uni j. 2016; 8(1):32-34. 13. mengistie z, woldeamanuel y, asrat d, adera a. prevalence of bacterial vaginosis among pregnant women attending antenatal care in tikuranbessa university hospital, addis ababa, ethiopia. bmc research notes. 2014 dec; 7(1):1-5. 14. ries aj. treatment of vaginal infections: candidiasis, bacterial vaginosis, and trichomoniasis. j am pharm assoc (wash) 1997; 37(5):563-9. 15. gul f, faiz nr, raziq f, malik l, sherin a, kazi bm, et al. frequency of vaginal discharge and its association with various sexually transmitted diseases in women attending antenatal clinic. j postgrad med inst 2005; 19(1):86-91. 16. chowdhury mn, jabbar fa, kambal am. isolation of gardnerella vaginalis from high vaginal swabs. trop geogr med 1985; 37(4):328-33. the authors: dr. amna javed assistant professor, department of obstetrics and gynecology, shaikh zayed medical complex, lahore. prof. muhammad ikram head, department of obstetrics and gynecology, shaikh zayed medical complex, lahore. prof. shafqat mukhtar head, department of obstetrics and gynecology, shaikh zayed medical complex, lahore. prof. aman ur rehman head, department of histopathology, shaikh zayed medical complex, lahore. dr. saulat sarfraz head, department of radiology, shaikh zayed medical complex, lahore. dr. ayesha shahid pg trainee, department of obstetrics and gynecology, shaikh zayed medical complex, lahore. corresponding author: dr. amna javed assistant professor, department of obstetrics and gynecology, shaikh zayed medical complex, lahore. e-mail: amnamuddassar@gmail.com numbering.indd 58 proceedings s.z.m.c. vol: 34(4): pp. 58-63, 2020. pszmc-772-34-4-2020 radiological analysis of hind paw joint in murine rheumatoid arthritis model treated prophylactically or therapeutically with cassia fistula versus naproxen 1hassan farooq, 2mariyam iftikhar piracha, 3muhammad usman, 1rabia tariq, 4saadia shahzad alam 1department of pharmacology & therapeutics, khawaja muhammad safdar medical college, sialkot, 2department of pharmacology & therapeutics, akhtar saeed medical college, lahore 3department of pharmacology & therapeutics, shalamar medical & dental college, lahore 4department of pharmacology & therapeutics, shaikh zayed medical complex, lahore abstract introduction: cassia fistula has anti-inflammatory activity. rheumatoid arthritis (ra) is an autoimmune disease, causing inflammation and disability of the affected joints. it is mostly treated symptomatically with nsaids, which themselves have a lot of adverse effects. in this study the anti-arthritic effect of cassia fistula versus naproxen has been observed radiologically. aims & objectives: the present study was radiological analysis of anti-arthritic effect of cassia fistula compared to naproxen in complete freund’s adjuvant (cfa) induced rat model of rheumatoid arthritis. place and duration of study: this study was conducted at the animal house of university of veterinary and animal sciences, lahore for a period of three months. material & methods: the study was carried out on 96 male rats divided into 12 groups of 8 rats each. single 0.2ml dose of complete freund’s adjuvant (cfa) was injected in the right hind paw of each rat in all groups except group 1 (negative control). group 2 was positive control. the prophylactic (3-7) and therapeutic (812) groups were given naproxen (25mg/kg), anthraquinone extract (250 &500mg/kg) or methanolic extract (250 & 500mg/kg) of cassia fistula orally bd on days 1,2 &3 (first dose preceding cfa injection by 30 minutes) and on days 9,10 &11 for therapeutic groups respectively. on day 15, x-rays of right ankle joints of all groups were taken as a confirmatory investigation on the final status of the arthritis. results: our results showed both anthraquinone and methanolic extracts of cassia fistula have dose dependent (500˃250mg/dl) prophylactic and therapeutic anti-arthritic potential albeit lesser than naproxen 25mg/kg, to reduce the swelling of hind paw and deformity of interphalangeal joints on radiological examination. conclusion: the efficacy of cassia fistula makes it a potential weaker candidate to naproxen in prevention and treatment of rheumatoid arthritis key words: cassia fistula, anthraquinone, ra, cfa, nsaid, anti-arthritic effect. introduction cassia fistula known as golden shower and amaltas, has great variety of uses ranging from constipation to glandular tumors in traditional medicine.1,2,3 each part of this plant has therapeutic potential but specifically its fruit pulp has antiinflammatory property and has demonstrated useful activity in various joint diseases.4 the common active principle found in all parts of the plant is a phenolic antioxidant: anthraquinone.5 rheumatoid arthritis is an autoimmune long lasting disease characterized by pain, swelling and stiffness of small joints which ultimately results in their deformity and disability.6,7 it has a better prognosis if treated early and militantly. the aim of treatment is to reduce pain and swelling, prevent bone deformity and improve quality of life. nsaids and disease modifying anti-rheumatic drugs (dmards) are usually used. nsaids are the most commonly prescribed drugs to subside pain and swelling throughout the world.1 naproxen (nsaid) inhibits tissue cyclo-oxygenase and decreases pro-inflammatory prostaglandins, the important mediators of pain and inflammation.8,9 thus, alleviates symptoms of rheumatoid arthritis but has some adverse effects.10,11 fruit pulp of cassia fistula has been used traditionally in many herbal medicines and shown to possess antiinflammatory activity.12 our previous experiments showed reduction in ra factor levels and ankle caliper measurement with cassia fistula given prophylactically and 59 radiological analysis of hind paw joint in murine rheumatoid arthritis model treated prophylactically... therapeutically.13 the present study was carried out to further verify those effects radiologically. this research was unique, as comparison of antiarthritic activity of the standard drug naproxen with cassia fistula methanolic fruit pulp extract and anthraquinone extract has not been done radiologically before to the best of our knowledge. material and methods after approval from the ethical review board committees of shaikh zayed federal postgraduate medical institute, lahore, this comparative study was completed over a period of six months at uvas, lahore. collection of cassia fistula bark and fruit pulp was done from botany department of university of punjab, lahore. cassia fistula extract preparation: the extraction process of cassia fistula bark and fruit pulp was performed in the labs of pcsir, lahore by following method.14 the extract was utilized after confirmatory anthraquinone test. anthraquinone extraction: methanolic extract preparation: confirmatory test for anthraquinones: cassia fistula bark and fruit pulp extract was boiled for 5 minutes after addition of 10ml of 1% hcl. sample was filtered and allow`ed to cool at room temperature. then using equal volumes of 10% ammonia and chloroform partition of the cooled filtrate was done twice and the layer was allowed to separate. the presence of combined anthraquinones was indicated by rose pink colour.13, 14 experimental setup: ninety six male wistar albino rats weighing 170200gm, acclimatized for a week, fed with standard laboratory diet, maintained at 25±2oc with relative humidity of 45-55% under 12 hours light and dark cycles were kept at uvas animal house, lahore. preparation of rat model of rheumatoid arthritis: to induce arthritis a single 0.2ml dose of cfa was injected in the right hind paw of each rat on day 1.15 a gradual increase in the swelling around injection site was noticed within few hours. the clinical evidence of arthritis was observed with gradual increase in the swelling around injection site and decreasing mobility of rats during 9th to 15th days of post cfa injection. 1% carboxymethyl cellulose was used as suspending agent for making water suspension of test extract and standard drug. extracts were given orally bd on days 1, 2 and 3 to the prophylactic groups (3-7) preceding cfa injection, and on days 9, 10 and 11 to the therapeutic groups (8-12).16 at the end of study on day 15 x-rays of right ankle joint were taken to evaluate bone deformity and disease progression. after careful numbering of rats, division into twelve groups was made with eight rats each. group 1: healthy male rats, not treated. given normal saline in equal amount. group 2: diseased control rats left for natural recovery after induction of rheumatoid arthritis with single 0.2ml cfa injection in right hind paw. prophylactic groups: group 3: tab. naproxen 25mg/kg group 4: anthraquinone extract at250mg/kg group 5: anthraquinone extract at 500mg/kg group 6: methanolic extract at 250mg/kg group 7: methanolic extract at 500mg/kg therapeutic groups: group 8: tab. naproxen 25mg/ kg group 9: anthraquinone extract at 250mg/kg group 10: anthraquinone extract at 500mg/kg group 11: methanolic extract at 250mg/kg group 12: methanolic extract at 500mg/kg radiological analysis: x-ray apparatus (siemens-60ma, germany) and industrial x-ray film (fuji photo film, japan) were used to take radioghraphs with operating conditions of 220v with peak of 40v, exposure time of 0.2 seconds and a 60cm tube to film distance for anterior-posterior projection. grading of radiographic images was done as follows: mild: diminished swelling of hind paw, no deformity of interphalangeal joints. moderate: minimal swelling of hind paw, low deformity of interphalangeal joints. intense: increased swelling of hind paw, highly deformed interphalangeal joints.17, 18 powdered 30gm fruit pulp of cassia fistula + ethanol 150ml (1:5) in soxhlet apparatus heated for 24hours at boiling point of solvent obtained 9% anthraquinone extract stored with desiccant after being concentrated and dried cassia fistula powdered bark extraction in soxhlet apparatus with double distilled water & methanol 9% concentrated extract was produced in rotary evaporator and refrigerated till further use 60 radiological analysis of hind paw joint in murine rheumatoid arthritis model treated prophylactically... results x-rays showed no swelling (ns) of hind paw and no deformity (nd) of interphalangeal joints of group 1, 3 and 8 (fig-1,3&8). swelling (s) of hind paw and deformity (d) of interphalangeal joints of group 2, 6 and 11 (fig-2,6&11). low swelling (ls) of hind paw and low deformity (ld) of interphalangeal joints of group 4, 7, 9 and 12 (fig-4,7,9&12). low swelling of hind paw and no deformity of interphalangeal joints of group 5 and 10 (fig-5&10). fig-1: group 1 healthy control shows no swelling (ns) of hind paw and no deformity (nd) of interphalangeal joints. fig-2: group 2 disease control shows swelling(s) of hind paw and deformity(d) of interphalangeal joints. fig-3: prophylactic group 3 naproxen treated rats shows no swelling (ns) of hind paw and no deformity (nd) of interphalangeal joints. fig-4: prophylactic group 4 anthraquinone 250mg/kg treated rats shows low swelling (ls) of hind paw and low deformity (ld) of interphalamgeal joints. fig-5: prophylactic group 5 anthraquinone 500mg/kg treated rats shows low swelling (ls) of hind paw and no deformity (nd) of interphalamgeal joints. fig-6: prophylactic group 6 methalonic extract of cassia fistula 250mg/kg treated rats showing mild swelling with less deformity of interphalangeal joints. fig-7: prophylactic group 7 methalonic extract of cassia fistula 500mg/kg showing low swelling of hind paw and low deformity of interphalangeal joints. improvement in joint space and cartilage deformity. ld l s n d n s s d 61 radiological analysis of hind paw joint in murine rheumatoid arthritis model treated prophylactically... fig-8: therapeutic group 8 naproxen 25mg/kg treated rats showing no bony destruction and swelling of joint with reduced edema and swelling of soft tissues. fig-9: therapeutic group 9 anthraquinone 250mg/kg treated rats showing reduced progression of ra with low joint swelling and less narrowing of joint space. fig-10: therapeutic group 10 anthraquinone 500mg/kg treated rats showing significant improvement of joint swelling by less soft tissue swelling. fig-11: therapeutic group 11 methanolic extract of cassia fistula 250mg/kg treated rats shows swelling (s) of hind paw and deformity (d) of interphalangeal joints. fig-12: therapeutic group 12 methanolic extract of cassia fistula 500mg/kg treated rats shows low swelling (ls) of hind paw and low deformity (ld) of interphalangeal joints. discussion rheumatoid arthritis, which is an autoimmune disease of progressive bones and cartilage damage, is associated with severe disability and has systemic effects as well.6 naproxen, a nonsteroidal antiinflammatory drug (nsaid) is used to treat rheumatoid arthritis but itself has many side effects.11, 12 plants have been used to treat different ailments including arthritis since ages.2 remarkable effects were produced by cassia fistula in earlier researches in murine model of arthritis.13,14 the current research was conducted to further verify this effect radiologically. radiological studies of hind paw joints: the radiological studies further verified the dominant anti-arthritic effect of methanolic and anthraquinone extracts of cassia fistula proven in our earlier experiments. clinically the diagnosis of rheumatoid arthritis, which requires therapeutic monitoring, is undeniable and it remains the standard method in assessing the disease progression. the reduced joint space, apparent in xrays, is a hallmark of arthritis. radiological studies of hind paw joints of disease control groups showed swelling of hind paw and deformity of interphalangeal joints as shown in fig 2. when treated by naproxen (both prophylactic and therapeutic groups; 3 and 8) there was almost complete reversal of radiological changes as compared to disease control group 2, with no deformity of joints and no swelling of hind paw as seen in fig-3 & 8. anthraquinone at the dose of 500mg/kg in group 5 and 10 had similar but lesser effects to that of naproxen group 3 & 8 with no joint deformity and low swelling of hind paw as shown in fig 5 & 10. anthraquinone has anti-inflammatory activity, inhibits production of superoxide anions from neutrophils and has antioxidant effect as well l s l d nd l s 62 radiological analysis of hind paw joint in murine rheumatoid arthritis model treated prophylactically... due to inhibition of lipid peroxidation.19,20,21 lowest or negligible improvement on rat hind paw joints radiological features was with that of 250mg/kg dose of methanolic extract of cassia fistula in group 6 & 11 as seen in fig-6 & 11. these radiological findings were novel and provided insight into the development of rheumatoid arthritis in the murine ankle joints and its resolution with cassia fistula extracts. conclusion in this innovative study, joint x-rays of both the prophylactic and therapeutic models were taken as a confirmatory investigation on the final status of arthritis which verified our findings regarding cassia fistula anthraquinone and methanolic extracts anti-arthritic potential in comparison to naproxen. therefore, the efficacy of cassia fistula make it a potential weaker candidate in prevention and treatment of rheumatoid arthritis. furthermore, anthraquinone extract in the dose of 500mg/kg was found to have more potent anti-arthritic effect. references 1. kala cp. ethnobotany and ethnoconservation of aegle marmelos (l.) correa. indian j traditional knowledge. 2006; 5(4): 537-540. 2. gupta ak, tondon n, sharma m. quality standards of indian medicinal plants, medicinal plants unit. indian council med res.2008;2: 47-53. 3. ayurvedic pharmacopoeia of india, part 1, vol.5, new delhi, government of india publication, 2001. page no. 8, 9. 4. indian herbal pharmacopoeia revised new edition, 2002, indian drug manufacturers association mumbai, page no.106-113. 5. nadkarni km. indian materia medica-2, 3rd ed. popular prakashan, bombay, gupta. pharm biol. 2009; 47(3): 195-202. 6. wollenhaupt j, zeidler h. undifferentiated arthritis and reactive arthritis. curr opin rheumatol 1998; 10(4): 306-13. 7. swash m, glynn m. hutchison's clinical methods. edinburgh. saunders elsevier, 2007. 8. zumora ra, billar tr. inducible nitric oxide synthase and inflammatory disease. mol med 2000; 6: 347-56. 9. lichtenberger lm, romero jj, dial ej, moore je. naproxen-pc: a gi safe and highly effective anti-inflammatory. inflammopharmacology. 2009; 17(1):1-5. 10. hoyle mg. naproxen and elevated liver enzyme [2013] available at traditional knowledge. 2006; 5(4): 537-540. 11. cooper k, bennett wm. nephrotoxicity of common drugs used in clinical practice. arch intern med. 1987; 147:1213-1218. 12. j.anitha and s.miruthula. anti inflammatory and phytochemical analysis of cassia fistula fruit pulp extracts. ijp. 2014; 1(3): 207-215. 13. farooq h, piracha mi, alam ss, sultan f. comparative anti-arthritic study of cassia fistula with naprexen in rheumatoid arthritis murine model. proceedings, szmc. 2018; 32(3): 35-39. 14. gupta v, agarwal a, tiwari hp (1990). isolation and characterization of two flavonol and a xanthone glycosides from the stem bark of cassia fistula linn. ind. j. chem. b. 28: 282-4. 15. a.m bendele. animal models of rheumatoid arthritis. j.musculoskeletal neuron interact. 2001; 1(4):377-385. 16. peter c. taylor, edward c. keystone et al. baricitinib versus placebo or adalimumab in rheumatoid arthritis. n engl j med 2017; 376: 652-62. 17. gupta a, singh s. evaluation of antiinflammatory effect of withania somnifera root on collagen-induced arthritis in rats. pharmaceutical biology. 2014;52(3):308-20. 18. fausto salaffi, m. c. m. d. c., 2016. conventional radiography in rheumatoid arthritis. int j clin exp med, 9(9), pp.17012-27. 19. kelly, s. et al., eur. j. neurosci of pharmacology. 2007. 20. siddhuraju p, mohan ps, becker k (2002). studies on the antioxidant activity of indian laburnum (cassia fistula l.): a preliminary assessment of crude extracts from stem bark, leaves, flowers and fruit pulp. j. agric. food. chem. 79: 61-7. 21. raju ilavarasan, moni mallika and subramanian venkataraman. antiinflammatory/ anti-oxidants activities c.fistula bark extracts. afr. j. trad cam. 2005; 2(1):70-85. the authors: dr. hassan farooq assistant professor, department of pharmacology & therapeutics, khawaja muhammad safdar medical college, sialkot. 63 radiological analysis of hind paw joint in murine rheumatoid arthritis model treated prophylactically... dr. mariyam iftikhar piracha assistant professor, department of pharmacology& therapeutics, akhtar saeed medical college, lahore. dr. muhammad usman senior demonstrator, department of pharmacology & therapeutics, shalamar medical & dental college, lahore. ms. rabia tariq pharmaceutical chemist, department of pharmacology & therapeutics, khawaja muhammad safdar medical college, sialkot. prof. saadia shahzad alam hod, pharmacology & therapeutics, shaikh zayed medical complex, lahore. corresponding author: dr. hassan farooq assistant professor, department of pharmacology, khawaja muhammad safdar medical college, sialkot. e-mail: hassan_oldravian@gmail.com 24 proceedings s.z.m.c. vol: 34(3): pp. 24-28, 2020. pszmc-757-34-3-2020 a detailed study of antimicrobial sensitivity pattern of penton valentine leucocidin gene positive and negative staphylococcus aureus from pus samples asma akram, 2mateen izhar, 2anwaar basheer, 1mariya ali, 3asma yaqoob, 4asim saifullah 1department of pathology, continental medical college, lahore 2department of microbiology, shaikh zayed medical complex, lahore 3department of pathology, fatima memorial medical and dental college, lahore 4department of pediatric orthopedic surgery, children hospital, lahore abstract introduction: staphylococcus aureus harboring panton valentine leucocidin gene are emerging and spreading worldwide. pvl gene was first identified by noel panton and francis valentine in 1932 who ic pakistan only limited data is available regarding the effect of pvl gene on sensitivity pattern of staphylococcus aureus. therefore, this study was conducted to understand the antimicrobial sensitivity pattern of both pvl positive and negative staphylococcus aureus isolates. aims & objectives: this study was conducted to understand the antimicrobial sensitivity pattern of both pvl positive and pvl negative staphylococcus aureus isolated from pus samples received from various indoor and outdoor departments of a tertiary care hospital of lahore. place and duration of study: microbiology and molecular biology laboratory shaikh zayed hospital lahore. duration of study is one year after the approval of research topic. material & methods: a total of 384 staphylococcus aureus isolates from skin and soft tissue infections were identified and selected. their antimicrobial sensitivity testing was done by kirby disc diffusion method using muller hinton agar. results: frequencies of pvl gene in mrsa and mssa were 51% and 44% respectively. frequency of pvl gene was also found to be high in ciprofloxacin sensitive, gentamicin sensitive, erythromycin resistant and fusidic acid resistant isolates. conclusion: almost half of staphylococcus aureus isolates were found pvl positive. they were mostly multidrug resistant. the pvl positive staphylococcus aureus isolates showed high resistance against antibiotics than pvl negative isolates. key words: pvl (panton valentine leucocidin), mrsa (methicillin resistant staphylococcus aureus) introduction in genus staphylococci there are more than 26 species included till date. staphylococcus aureus is one of the most critical and harmful pathogen among these staphylococci.1 it is the most widely recognized microbe that causes skin and soft tissue diseases in both children and adults.3 staphylococcus aureus also cause life-threatening infections, for example, hemorrhagic pneumonia and endocarditis.4 staphylococcus aureus colonize different body sites however nasal pit is the main site of colonization. pathogenic factors produced by staphylococcus aureus render this organism exceedingly pathogenic.5 they include surface proteins, toxins and enzymes.6 in the past, staphylococcus aureus caused high mortality due to its pathogenicity. this mortality was reduced to sufficiently low levels due to discovery of penicillin in 1940s. but relieve from penicillin was brief as penicillin resistant staphylococcus aureus (prsa) emerged rapidly. methicillin, a beta lactamaseinsensitive beta-lactam prepared in late 1950s was a good alternative to prsa but methicillin resistant staphylococcus aureus that were resistant to commonly available beta lactamase from hospital settings. it caused an increased use of antibiotic of last resort vancomycin as a treatment option for mrsa. initially, the rates of mrsa increased slowly and they were mostly isolated from hospital settings but later dramatic increase in frequency of mrsa occurred2 and it was followed by isolation of mrsa from community settings from the people that were previously healthy and had no or very little chance of carrying mrsa.7 community acquired staphylococcus aureus infection was previously caused by methicillin sensitive 25 a detailed study of antimicrobial sensitivity pattern of penton valentine leucocidin gene positive staphylococcus aureus.8 it was also observed that in many areas of world ca-mrsa had become more common than ca-mssa.9 moreover, ca-mrsa is more infectious than ha-mrsa. the clinical picture of ha-mrsa infection is less severe than ca-mrsa but it is difficult to treat as compared to ca-mrsa.2 methicillin resistance in staphylococcus aureus is mediated by acquiring meca gene that encodes altered penicillin binding protein that has low affinity for beta lactams. both mrsa and mssa can carry pvl gene (panton valentine leucocidin gene) which makes them more aggressive and pathogenic. it was discovered by van develde in 1894 who found its capability of lysing leucocytes and is named after sir philip noel panton and francis valentine who related it with skin and soft tissue diseases in 1932.5 pvl gene is acquired by staphylococcus aureus by viruses called prophages which carry various genes between bacteria.10 these toxins destroy the outer membranes of white blood cells by the combine action of 2 secretory proteins named s and f by making pores in the membranes of cell and causing leakage of cellular contents through that pores resulting in cell death.11 purified pvl is only toxic to white blood cells and macrophages in humans and rabbits, they are not toxic for erythrocytes.12 in nepal a study was carried out in 2014 with sample size of 73 including mrsa and mssa and they were isolated from various samples; among them pvl gene was 26.1% in mrsa and 51.9% in mssa.13 while in india a study was conducted in which overall prevalence of pvl was 62.85% and among mrsa and mssa it was 85.1% and 48.8% respectively.14 limited work has been done on pvl gene and effect of harboring gene on organism’s sensitivity pattern in pakistan. so, this study was planned to observe the drug resistance pattern of pvl positive and pvl negative staphylococcus aureus collected both from indoor and outdoor. abbreviations: ca-mrsa: community acquired methicillin resistant staphylococcus aureus; clsi: clinical and laboratory standards institute; ha-mrsa: hospital acquired methicillin resistant staphylococcus aureus; mdr: multidrug resistant; pcr: polymerase chain reaction; pvl: panton valentine leukocidin; material and methods approval and consent to participate ethical approval to conduct the study was obtained from the institutional review board (irb), federal postgraduate medical institute, shaikh zayed hospital, and lahore. collection and sample processing pus samples from various indoor and outdoor departments of shaikh zayed hospital lahore were collected. data related to samples was noted on standardized proforma. samples were processed and staphylococcus aureus was identified according to laboratory sops. antibiotic susceptibility testing was performed according to csli guidelines edition 2016. following drugs were used; cefoxitin, erythromycin, ciprofloxacin, gentamycin, fusidic acid, penicillin, vancomycin and linezolid. staphylococcus aureus showed resistance to at least three antimicrobial drugs were categorized as multidrug resistant. results table-1 shows 72% resistant in staphylococcus aureus while pvl gene frequency was high in ciprofloxacin sensitive cases but relationship between drug sensitivity and pvl gene is not significant (p=0.246). while table-2. shows 63% resistant in staphylococcus aureus against erythromycin while pvl gene frequency was high in resistant cases. table-3 shows high sensitivity against gentamycin while pvl gene frequency was high in sensitive cases. table-4 shows high sensitivity against fusidic acid and pvl gene frequency is also high in sensitive cases. table-5 and fig-1 describe that mostly isolates were multidrug resistant and pvl gene frequency was also high in mdr cases while table-6 and fig-2 explain that most of staphylococcus aureus were mrsa and half of those mrsa were pvl positive. ciprofloxacin total n=384 pvl+ve pvl-ve p value sensitive 92 (24%) 50 (54.3%) 42 (45.65%) 0.246 intermediate 15 (3.9%) 5 (33.3%) 10 (66.6%) resistant 277(72.1%) 131(47.2%) 146(52.7%) table-1: penton valentine leukocidin gene and ciprofloxacin erythromycin total staph aureus n=384 pvl+ve pvl-ve p value sensitive 130 (33.9%) 61 (47%) 69 (53%) 0.625 intermediate 11 (2.9%) 4 (36.36%) 7 (63.6%) resistant 243 (63%) 121 (49%) 122 (51%) table-2: erythromycin and pvl gene relation 26 a detailed study of antimicrobial sensitivity pattern of penton valentine leucocidin gene positive gentamycin total n=384 pvl+ve pvl-ve p value sensitive 235 (61.2%) 120 (51%) 115 (49%) 0.428 intermediate 2 (0.5%) 1 (50%) 1 (50%) resistant 147 (38.3%) 65 (44.3%) 82 (56%) table-3: penton valentine leukocidin gene and gentamycin fusidic acid totalstaph aureus n=384 pvl+ve pvl-ve p value sensitive 240 (62.5%) 120 (50%) 120 (50%) 0.429 resistant 144 (37.5%) 66 (46%) 78 (54%) table-4: panton valentine leucocidin gene and fusidic acid drug resistance total staph aureus n=384 pvl+ve pvl-ve p value mdr 290 (75.5%) 143 (49.3%) 147 (50.7%) 0.555 non-mdr 94 (24.5%) 43 (46%) 51 (54%) table-5: relationship of mdr and pvl gene fig-1: relation between mdr and pvl gene methicillin total pvl+ve pvl-ve p value sensitive mssa 121 (31.5%) 53 (43.8%) 68 (56.2%) 0.228 resistant mrsa 263 (68.5%) 133 (50.6%) 130 (49.4%) table-6: relationship between methicillin sensitivity and pvl gene fig-2: relation between methicillin sensitivity and pvl gene discussion antimicrobial sensitivity of staphylococcus aureus isolates was tested for following purposes: (i) to know mrsa and mssa (ii) to find out the correlation of ciprofloxacin sensitivity with pvl gene, as it has been considered that pvl gene ratio is high among ciprofloxacin sensitive staphylococcus aureus16 (iii) to know frequency of mdr isolates (iv) any unusual pattern in ast. all isolates were sensitive to vancomycin and linezolid while 97.4% of samples were resistant to penicillin. in current research, among 384 samples, 68.5% were mrsa and 31.5% were mssa. among mrsa 50.6% have pvl gene while in mssa 43.8% have pvl gene. suberna roy et al. from india have reported 85.1% pvl gene in mrsa and 48.8% in mssa which indicated a higher prevalence of pvl gene than our findings.14 another study conducted in pakistan in 2016 reported 44% mrsa and 55% mssa while pvl gene frequency in mrsa was 31% and in mssa it was 18%15 indicated a lower prevalence of pvl gene than our findings. so, current research work indicates that the frequency of both mrsa and pvl is increasing in our setup with the passage of time. this may be due to unnecessary and excessive use of antibiotics in our setup that is making isolates more antibiotic resistant and causing poor infection control and selection of resistant isolates in the community. it has been assessed that pvl gene frequency is usually high in ciprofloxacin sensitive staphylococcus aureus.16 in 2008, guidelines were published in the uk regarding the treatment of pvl positive staphylococcus aureus infections that emphasized that only ciprofloxacin sensitive cases should be sent to the laboratory for evaluation of pvl gene.17 we found high percentage of pvl gene 0.00% 20.00% 40.00% 60.00% 80.00% sensitive mrsa resistantmssafr eq ue nc y in p er ec en ta ge multidrug resistance and pvl gene relation total pvl+ve 0.00% 20.00% 40.00% 60.00% 80.00% total pvl+ve pvl-vef re qu en cy in p er ce nt ag e relationship between methicillin sensitivity and pvl gene sensitive mrsa resistantmssa 27 a detailed study of antimicrobial sensitivity pattern of penton valentine leucocidin gene positive in ciprofloxacin sensitive staphylococcus aureus but the difference of percentage was not so high and it was also not statistically proved. in current investigation, ciprofloxacin resistance in staphylococcus aureus is 72% while it had been reported as low as less than 3% from england and wales.6 these results reflect that prevalence of ciprofloxacin resistance and pvl gene varies greatly between geographical locations and populations. our investigation shows that more than half of staphylococcus aureus were resistant to erythromycin and pvl gene frequency was high in erythromycin resistant staphylococcus aureus. these results are in accordance with the results in nepal.5 current results show that 38% of staphylococcus aureus were resistant to gentamycin and frequency of pvl gene was high in gentamycin sensitive cases. gentamycin resistance in staphylococcus aureus was reported 20% in research from england. moreover, 37% of staphylococcus aureus were resistant to fusidic acid while pvl gene frequency was high in fusidic acid sensitive. fusidic acid resistance in staphylococcus aureus was reported 33% in research from england and wales in 2005.6 due to sensitivity testing of isolates, it was easy to find out mdr and non-mdr frequency in these isolates.pvl gene frequency was high in mdr. in current study, 75% staphylococcus aureus were mdr and 25% were non-mdr while a study from south india reports 91% mdr isolates.18 conclusion almost half of staphylococcus aureus isolates were found pvl positive. they were mostly multidrug resistant. the pvl positive staphylococcus aureus isolates showed high resistance against antibiotics than pvl negative isolates. acknowledgement: i am grateful to all the staff of microbiology department, shaikh zayed hospital lahore specially our supervisor prof. mateen izhar for his guidance from topic selection to thesis compilation and for funding in this research. all samples were collected and processed in microbiology laboratory shaikh zayed hospital lahore. references 1. aboud sa, el-yamani a, hussain ma, ahmed a. prevalence and antibiotics sensitivity of staphylococcus aureus skin infection in children in khartoum, sudan. curr res micro. biotechnol. 2015; 3(4):690-3. 2. boyle-vavra s, daum rs. community-acquired methicillin-resistant staphylococcus aureus: the role of panton–valentine leukocidin. laboratory investigation. 2007 jan; 87(1):3. 3. mcneil jc, hulten kg, kaplan sl, mason eo. decreased susceptibilities to retapamulin, mupirocin, and chlorhexidine among staphylococcus aureus isolates causing skin and soft tissue infections in otherwise healthy children. antimicrobial agents and chemotherapy. 2014 may 1; 58(5):2878-83. 4. sina h, ahoyo ta, moussaoui w, keller d, bankolé hs, barogui y, stienstra y, kotchoni so, prévost g, baba-moussa l. variability of antibiotic susceptibility and toxin production of staphylococcus aureus strains isolated from skin, soft tissue, and bone related infections. bmc microbiology. 2013 aug 8; 13(1):188. 5. bhatta dr, cavaco lm, nath g, kumar k, gaur a, gokhale s, bhatta dr. association of panton valentine leukocidin (pvl) genes with methicillin resistant staphylococcus aureus (mrsa) in western nepal: a matter of concern for community infections (a hospital based prospective study). bmc infectious diseases. 2016 may 15; 16(1):199. 6. holmes a, ganner m, mcguane s, pitt tl, cookson bd, kearns am. staphylococcus aureus isolates carrying panton-valentine leucocidin genes in england and wales: frequency, characterization, and association with clinical disease. journal of clinical microbiology. 2005 may 1; 43(5):2384-90. 7. herold bc, immergluck lc, maranan mc, lauderdale ds, gaskin re, boyle-vavra s, leitch cd, daum rs. community-acquired methicillin-resistant staphylococcus aureus in children with no identified predisposing risk. jama. 1998 feb 25; 279(8):593-8. 8. vandenesch f, naimi t, enright mc, lina g, nimmo gr, heffernan h, liassine n, bes m, greenland t, reverdy me, etienne j. community-acquired methicillin-resistant staphylococcus aureus carrying pantonvalentine leukocidin genes: worldwide emergence. emerging infectious diseases. 2003 aug;9(8):978. 9. moran gj, krishnadasan a, gorwitz rj, fosheim ge, mcdougal lk, carey rb, talan da. methicillin-resistant s. aureus infections among patients in the emergency department. new england journal of medicine. 2006 aug 17; 355(7):666-74. 10. supersac g, prévost g, piémont y sequencing of leucocidin r from staphylococcus aureus 28 a detailed study of antimicrobial sensitivity pattern of penton valentine leucocidin gene positive p83 suggests that staphylococcal leucocidins and gamma-hemolysin are members of a single, two-component family of toxin, infect immun, 1993, vol. 61 (pg. 580-7) 11. prévost g, supersac g, colin d, et al. freerj, aitken r, alouf je, et al. the new family of leucotoxins from staphylococcus aureus: structural and biological properties, zentralbl bakteriol suppl 24, 1994 stuttgart gustav fischer verlag (pg. 284-93). 12. finck-barbançon v, duportail g, meunier o, colin da. pore formation by two-component leukocidin from staphylococcus aureus within the membrane of human polymorphonuclear leukocytes, biochem biophys acta, 1993, vol. 1182 (pg. 275-82). 13. shrestha b, singh w, raj vs, pokhrel bm, mohapatra tm. high prevalence of pantonvalentine leukocidin (pvl) genes in nosocomial-acquired staphylococcus aureus isolated from tertiary care hospitals in nepal. biomed research international. 2014. 14. kaur h, purwar s, saini a, kaur h, karadesai sg, kholkute sd, roy s. status of methicillinresistant staphylococcus aureus infections and evaluation of pvl producing strains in belgaum, south india. jkimsu. 2012 jul 1; 1(2):43-51. 15. madzgalla s, syed ma, khan ma, rehman ss, müller e, reissig a, ehricht r, monecke s. molecular characterization of staphylococcus aureus isolates causing skin and soft tissue infections in patients from malakand, pakistan. european journal of clinical microbiology & infectious diseases. 2016 sep 1; 35(9):1541-7. 16. ellington mj, perry c, ganner m, warner m, smith im, hill rl, shallcross l, sabersheikh s, holmes a, cookson bd, kearns am. clinical and molecular epidemiology of ciprofloxacinsusceptible mrsa encoding pvl in england and wales. european journal of clinical microbiology & infectious diseases. 2009 sep 1; 28(9):1113. 17. health protection agency. guidance on the diagnosis and management of pvl-associated staphylococcus aureus infections (pvl-sa) in england. 18. rajan v, schoenfelder sm, ziebuhr w, gopal s. genotyping of community-associated methicillin resistant staphylococcus aureus (ca-mrsa) in a tertiary care centre in mysore, south india: st2371-sccmec iv emerges as the major clone. infection, genetics and evolution. 2015 aug 1; 34:230-5. the authors: dr. asma akram senior demonstrator, department of pathology, continental medical college, lahore. prof. mateen izhar head, department of microbiology and virology, shaikh zayed medical complex, lahore. dr. anwaar basheer associate professor, department of microbiology and virology, shaikh zayed medical complex, lahore. dr. mariya ali senior demonstrator, department of pathology, continental medical college, lahore. dr. asma yaqoob senior demonstrator, department of pathology, fatima memorial medical & dental college, lahore. dr. asim saifullah senior registrar, department of pediatric orthopedic surgery, children hospital, lahore. corresponding author dr. asma akram senior demonstrator, department of pathology, continental medical college, lahore. e-mail: asmaasim26@gmail.com untitled-1 35 proceedings s.z.m.c. vol: 34(3): pp. 35-40, 2020. pszmc-759-34-3-2020 therapeutic effects of canagliflozin and zinc sulphate alone and in combination on pancreatic histology in type-2 diabetic rat model 1ayela eman zia, 1bushra suhail, 3zujaja zaheer 1department of pharmacology, university college of medicine and dentistry, university of lahore, lahore 2department of, pharmacology, king edward medical university, lahore abstract introduction: diabetes mellitus is a common metabolic syndrome characterized by persistently elevated blood glucose levels. canagliflozin is an sglt-2 inhibitor that controls hyperglycemia by reducing the reabsorption of filtered glucose and excreting it in the urine. zinc sulphate exhibits some beneficial role in diabetes mellitus but has not been compared to canagliflozin individually and in combination. aims & objectives: to observe the effects of treatment with canagliflozin and zinc sulphate on pancreatic histology in streptozotocin induced type-2 diabetic rat model. place and duration of study: the study was conducted in the department of pharmacology, kemu and pgmi, lahore for the period of two months. material & methods: it was an animal experimental study of eight weeks duration in which 48 adult healthy albino rats of male gender were divided into six groups and were provided the high fat diet throughout the study period. groups a and b were maintained as healthy and diseased controls respectively. groups b, c, d, e and f were administered single i/p dose streptozotocin (35mg/kg) at day 22 for inducing diabetes. upon confirmation of diabetes after a week the rats were further treated as per group designation orally for 4 weeks , individually or in combination with full or half doses of canagliflozin (10 mg /kg/day, 5mg/kg/day ) and zinc sulphate (30mg/kg/day, 15mg/kg/day) . all animals were euthanized at the completion of study duration. the pancreatic tissue was taken out and examined for the histopathological changes (size and number of pancreatic islets, karyolysis and ballooning degeneration). results: there was a marked improvement in the size and number of islets as well as the inflammatory changes in the combined treatment group (with canagliflozin in full as well as half dose of zinc sulphate) as compared to the groups given zinc sulphate and canagliflozin separately. conclusion: combined treatment with canagliflozin and zinc sulphate has a better protective effect on the pancreatic tissue in diabetes than either of them used alone. key words: diabetes mellitus, canagliflozin, zinc sulphate, histopathological, streptozotocin introduction there has been an alarming increase in the number of individuals having diabetes during the last two decades, which poses a threat to increase the health expenditure for diabetes worldwide and specifically in the developing countries, where the rate of rise is more.1,2 the current treatment options provide control of hyperglycemia but the associated side effects worsen the patient’s compliance, so there is a dire need of a therapy which can provide a good glycemic control with minimal side effects.3 in this regard, canagliflozin, which belongs to the class of sglt-2 inhibitors seems to be a good option since it is well-tolerated in the body and has additional benefits of lowering blood pressure and body weight. it promotes urinary excretion of glucose via decreasing its reabsorption at the proximal tubule and thus causing a reduction in the renal glucose threshold.4 zinc regulates the functions of many tissues and has the potential to normalize the hyperglycemia.5 limited studies have shown the efficacy of zinc combined with oral hypoglycemic agents in combating diabetes mellitus.6 the high-fat diet/streptozotocin (hfd/stz) rat model is an example of the experimentally induced type 2 diabetes model. it is prepared by combining a fatrich diet to produce insulin resistance and then treating with stz (a toxin of the beta cells), that leads to greater decrease in the functional mass of the pancreatic beta cells.7 36 therapeutic effects of canagliflozin and zinc sulphate alone and in combination on pancreatic histology the present study is designed to observe the role of zinc and canagliflozin alone and in combination on pancreatic histology in type-2 diabetes mellitus and to hypothesize that the combination therapy has a better effect than each given separately. material and methods study design: animal experimental study. setting: the study was conducted in the department of pharmacology, kemu and pgmi, lahore. sampling technique: simple random sampling. sample size: forty eight rats were divided into six groups by lottery method. inclusion criteria: male sprague-dawley rats, weighing 120g to 180g. exclusion criteria: rats showing signs of any disease. 48 adult healthy albino rats of male gender were purchased from uvas (university of veterinary and animal sciences) and kept in the animal house of pgmi (post-graduate medical institute), lahore. animals were divided randomly into 6 equal groups having 8 rats in each group. the rats were exposed to natural day and night cycles at room temperature of 22 ± 2 °c with 50 ± 5 % humidity throughout the experiment. they had a free access to rat chow and water ad libitum. an interval of seven days was given to them to get acclimatized before the start of the experiment. preparation of doses & sampling: the calculated dose for individual rat, i.e. 10mg/kg/day of canagliflozin8 and 30mg/kg/day of zinc sulphate9 were weighed and dissolved in 1ml of distilled water. high fat diet was prepared by combining 1.5g of cholesterol, 1g of sodium deoxycholate and 8ml of coconut oil in every 100g of normal rat chow.10,11 hyperlipidemic rats were then injected intraperitoneally with streptozotocin (35mg/kg) at day 22, dissolved in 0.1m sodium citrate buffer at ph 4.5 (prepared by adding 46.5ml of citric acid to 3.5ml of sodium citrate solution and making it up to 100ml with distilled water).12 after a week from injection of stz, blood samples obtained from the lateral tail vein were used in determining the blood glucose value using glucometer and those having blood glucose > 180 mg/dl were labeled as diabetic. grouping of animals: forty eight rats were divided into six groups randomly by lottery method, with 8 rats in each group. these groups were labelled as a,b,c,d,e and f. each of the group was kept in a separate iron cage. rats in group a (normal control group) were fed with standard rat diet (normal rat chow), throughout the study period of 8 weeks. rats in all the other groups (b, c, d, e and f) were given high fat diet throughout 8 weeks and streptozotocin after 3 weeks for the induction of type-2 diabetes. rats in group b (disease control group) were only given streptozotocin to induce diabetes. rats in group c were administered zinc sulphate orally from 4th to 8th week after the induction of diabetes. rats in group d were administered canagliflozin orally from 4th – 8th week after the induction of diabetes. rats in group e were administered full doses orally of canagliflozin (i.e. 10mg/kg/day) and zinc sulphate (30mg/kg/day) from 4th – 8th week after the induction of diabetes. rats in group f were administered half doses orally canagliflozin (i.e. 5mg/kg/day) and zinc sulphate (15mg/kg/day) from 4th – 8th week after the induction of diabetes. euthanization: twenty four hours after last dose administered the rats were sacrificed at the end of week 8. pancreas of each rat was identified and dissected out en-bloc and preserved in formalin separately. histological examination: after removing the pancreas, it was stretched on filter paper and fixed in 10% buffered formalin (ph 7.4) at room temperature. the fixed specimens were sliced, processed and embedded into paraffin blocks. the blocks were then cut into 4μm paraffin sections by a rotator microtome. clean glass slides were put in hot air oven for 24 hours and were properly numbered and 4μm thick representative sections of tissues were mounted. de-waxing of tissues was done by dipping them in xylene. slides were hydrated by passing through decreasing concentrations of alcohol and were dipped in hematoxylin. afterwards, these were washed in the running water. then, these slides were placed in 1% acid alcohol, again washed under tap water and put in 1% ammonia solution followed by water wash. then, the slides were put in eosin for 5 minutes and washed under tap water. the stained sections were mounted with dpx and examined under microscope to evaluate for the presence or absence of the structural changes like signs of inflammation (karyolysis and ballooning) and atrophy (changes in the size and number of islet of langerhans).13 the size and number of pancreatic islets were measured with the help of a deca-head microscope (nikon imaging software package-d) and mean was taken for further calculations. statistical analysis: all the data was entered in spss version 23 for qualitative data and graph pad prism version 8 for quantitative data. quantitative data (size and number of islets) was expressed as mean ± s. d. 37 therapeutic effects of canagliflozin and zinc sulphate alone and in combination on pancreatic histology mean plots were used for graphical presentation to see the changes in the parameters. the data was evaluated by one way analysis of variance followed by tukey’s multiple comparison tests. qualitative data (karyolysis and ballooning degeneration) were scored as numbers and expressed as percentage of changes in groups. the significance of differences was measured through kruskal-wallis anova and mann-whitney u test. p-value of less than 0.05 was considered significant. results 1) size of islets of langerhans (μm): gro up a gro up b gro up c gro up d gro up e gro up f 0 50 100 150 200 250 size of islets of langerhans siz eo fis let s(( μm ) ** *** ### ### ### ### *** ### fig-1: comparison of mean size of islets of langerhans of groups a, b, c, d, e and f ### indicates p-value is <0.001 as compared to group a. *** shows p-value is <0.001 ** means p value <0.01, when compared to group b. group a=normal control, group b=disease control, group c=zn treated, group d =cana treated, group e = zn+cana (full dose), group f = zn+cana (half dose). 2) number of islets of langerhans: gro up a gro up b gro up c gro up d gro up e gro up f 0 10 20 30 number of islets of langerhans no .o fis let s ### *** *** ### ### fig-2: comparison of mean number of islets of langerhans among groups a,b,c,d,e and f ### shows p-value is <0.001 when compared to group a. *** means p-value is <0.001 as compared to group b. group a=normal control, group b=disease control, group c=zn treated, group d =cana treated, group e = zn+cana (full dose), group f=zn+cana (half dose). 3) karyolysis: fig-3: comparison of karyolysis among groups a,b,c,d,e and f. group a=normal control, group b=disease control, group c=zn treated, group d =cana treated, group e=zn+cana (full dose), group f=zn+cana (half dose). 4) ballooning degeneration: fig-4: comparison of ballooning degeneration among groups a, b, c, d, e and f. group a=normal control, group b=disease control, group c=zn treated, group d =cana treated, group e = zn+cana (full dose), group f=zn+cana (half dose). 38 therapeutic effects of canagliflozin and zinc sulphate alone and in combination on pancreatic histology fig-5: group a (normal control) histology of pancreas showing normal islets of langerhans (40 x; h&e) fig-6: group b (positive control) severe reduction in the size and number of islets in stz-induced diabetic rat (40 x; h&e) fig-7: group c (hfd+stz+zinc) some restoration in size and number of islets with moderate karyolysis and ballooning (40 x; h&e) after treatment with zinc alone fig-8: group d (hfd+stz+canagliflozin) some restoration in size and number of islets, ballooning and karyolysis still present after treatment with cana alone (40x; h&e) fig-9: group e (hfd+stz+zinc+full cana) restoration of size and number of islets with mild karyolysis and ballooning (40x; h&e) fig-10: group f (hfd+stz+zinc+halfcana) restoration of the size and number of islets with mild karyolysis and ballooning (40x; h&e) discussion histopathological study with the light and deca-head microscopy revealed some gross changes in the size, number and structure of the islets of langerhans. stz administration led to the shrinkage and 39 therapeutic effects of canagliflozin and zinc sulphate alone and in combination on pancreatic histology disruption of the structure of islets and the degenerative changes were pronounced, like karyolysis of the nuclei and ballooning degeneration, causing the cells to have a highly vacuolated cytoplasm. upon injection with stz, atrophy of islets was prominent causing a visible reduction in the size and number of the islets of langerhans. similar findings were reported in the recent works using the rat model of stz induced diabetes.13,14 after the administration of zinc for four weeks, there was some reversal of these histopathological changes in the pancreatic sections of the rats of group c. a significant restoration of the size of islets as well as the reversal of karyolysis and ballooning degeneration was observed on histopathological examination, whereas no significant difference was observed in the number of islets with zinc treatment. these results resemble those of another study in which zinc supplementation given to stz-induced diabetic rats was associated with significant rejuvenation of the islets histology.5 zinc preserves the pancreatic architecture probably through its anti-oxidant property.15 canagliflozin was also found to exert a beneficial effect in restoring the histological morphology of pancreas (karyolysis and ballooning degeneration) but to a lesser extent than zinc. this observation is in accordance with a study done earlier in which the treatment of zdf (zucker diabetic fatty) rats with canagliflozin minimized the degenerative changes in diabetic induced islets.16 another study indicated that dapagliflozin (closely related to canagliflozin and both belong to sglt-2 inhibitors) has a role in the preservation of pancreatic islets morphology of hfd fed diabetic rats17. in diabetes, increase in demand of insulin secretion occurs which can cause stress of the endoplasmic reticulum, leading to increased apoptosis of the beta-cell. by decreasing hyperglycemia through a non–insulin-dependent procedure, sglt-2 inhibitors reduces the demand on the beta cells to produce insulin and thus prevents excessive apoptosis of these cells as seen in diabetes.8,18 even better results were seen in the combination groups e and f, where the addition of zinc to canagliflozin further improved the morphological changes in the islets. the degenerative signs (karyolysis and ballooning) almost disappeared and the number and size of the pancreatic islets were comparable with the negative control group. no study to date has suggested this potentiating effect of zinc with canagliflozin. conclusion this study has demonstrated that the combined administration of zinc and canagliflozin has exerted a stronger effect on restoring pancreatic histology back to normal in type-2 diabetic rat model as compared to both of these drugs given alone. references 1. zimmet pz, magliano dj, herman wh, shaw je. diabetes: a 21st century challenge. lancet diabetes endocrinol 2014; 2:56-64. 2. nanditha a, ramachandran a, snehalatha c, chan jc, chia ks, shaw ej, et al. diabetes in asia and the pacific: implications for the global epidemic. diabetes care. 2016; 39(3):472-85. 3. chaudhury a, duvoor c, reddy dendi vs, kraleti s, chada a, ravilla r, et al. clinical review of antidiabetic drugs: implications for type 2 diabetes mellitus management. front endocrinal. 2017; 8(6):1-12. 4. brocavich ss, hilas o. canagliflozin (invokana), a novel oral agent for type-2 diabetes. p&t. 2013; 38(11):656-60, 65-66. 5. barman s, srinivasan k. zinc supplementation alleviates hyperglycemia and associated metabolic abnormalities in streptozotocininduced diabetic rats. can j physiol pharmacol. 2016; 94(12):1356-65. 6. burki z, hussain m, burki s, farooqi w, zeb a, ahmad s. effect of zinc supplementation on serum fasting blood sugar and hba1c in adult diabetics on oral hypoglycemic agents. gomal j med sci. 2017; 15(1):8-11. 7. skovso s. modeling type 2 diabetes in rats using high fat diet and streptozotocin. j diabetes investing. 2014; 5(4):349-58. 8. liang y, arakawa k, ueta k, matsushita y, kuriyama c, martin t, et al. effect of canagliflozin on renal threshold for glucose, glycemia, and body weight in normal and diabetic animal models. plos one. 2012; 7(2):e30555. 9. nazarizadeh a, asri-rezaie s. comparative study of antidiabetic activity and oxidative stress induced by zinc oxide nanoparticles and zinc sulfate in diabetic rats. aaps pharm sci tech. 2016; 17(4):834-43. 10. balasubramanian mn, muralidharan p, balamurugan g. anti-hyperlipidemic activity of pedalium murex (linn.) fruits on high fat diet fed rats. int j pharmacol. 2008; 4(4):310-13. 40 therapeutic effects of canagliflozin and zinc sulphate alone and in combination on pancreatic histology 11. desu bsr, saileela ch. anti-hyperlipidemic activity of methanolic extract of rhinacanthus nasutus. int j res pharm chem.2013;3(3):708-11 12. marie m, arafa n, sara abdullah mubarak a. effect of canagliflozin or metformin on metabolic disorders in obese diabetic rats. afr j pharm pharmacol. 2015; 9(46):1071-9. 13. abunasef sk, amin ha, abdel-hamid ga. a histological and immunohistochemical study of betacells in streptozotocin diabetic rats treated with caffeine. folia histochem cytobiol. 2014; 52(1):42-50. 14. omar nm, atia gm. effect of nigella sativa on pancreatic β-cell damage in streptozotocin induced diabetic rats. the egyptian journal of histology. 2012; 35(1):106-16. 15. zhu k, nie s, li c, huang j, hu x, li w, et al. antidiabetic and pancreas-protective effects of zinc threoninate chelate in diabetic rats may be associated with its antioxidative stress ability. biol trace elem res. 2013; 153(1-3):291-8. 16. kuriyama c, xu jz, lee sp, qi j, kimata h, kakimoto t, et al. analysis of the effect of canagliflozin on renal glucose reabsorption and progression of hyperglycemia in zucker diabetic fatty rats. j pharmacol exp ther. 2014; 351(2):423-31. 17. macdonald fr, peel je, jones hb, mayers rm, westgate l, whaley jm, et al. the novel sodium glucose transporter 2 inhibitor dapagliflozin sustains pancreatic function and preserves islet morphology in obese, diabetic rats. diabetes obes metab.2010;12(11):1004-12 18. eizirik dl, cardozo ak, cnop m. the role for endoplasmic reticulum stress in diabetes mellitus. endocr rev. 2008; 29(1):42-61. the authors: dr. ayela eman zia senior demonstrator, department of pharmacology, ucmd, university of lahore, lahore. dr. bushra suhail assistant professor, department of pharmacology, ucmd, university of lahore, lahore. prof. zujaja zaheer head, department of pharmacology, king edward medical university, lahore. corresponding author: dr. ayela eman zia, senior demonstrator, department of pharmacology, ucmd, university of lahore, lahore. e-mail: ayelaemanz@gmail.com 31 proceedings s.z.m.c. vol: 35(3): pp. 31-36, 2021. pszmc-805-35-3-2021 morphological spectrum of ovarian teratomas: a 6 years experience at a specialized diagnostic institute fatima khalid, sameen afzal, ghazi zafar, anila chughtai, samina zaman, akhtar sohail chughtai department of histopathology, chughtai institute of pathology, lahore abstract introduction: ovarian teratomas include mature cystic teratoma (mct), immature teratoma (it) and monodermal teratoma (mot). malignant transformation (mt) can occur in mct and mot, which remains a diagnostic challenge. aims & objectives: to discuss the morphological spectrum of ovarian teratomas reported during the last six years at our specialized diagnostic institute and determine the frequency of mt in mct and mot. place and duration of study: the study was carried out at chughtai institute of pathology from january 2015 to december 2020. material & methods: a retrospective study was performed by retrieving 1018 cases of ovarian teratomas reported in last 6 years. spss version 20 was used for data analysis. results: mct was most frequent ovarian teratoma with frequency of 95.6% (n=973), 2.26% (n=23) being it and 2.16% (n=22) being mot. it were mostly seen in pediatric age group. mt in benign mct was seen in 0.72% of mct (n=7). squamous cell carcinoma was most frequent malignancy 71.4% (n=5). one case of mot also showed mt into papillary thyroid carcinoma, classic type, with frequency of mt in mot being 4.76% (n=1). all mt in mct had age >30 years which was a significant finding (p <0.001). conclusion: ovarian masses in postmenopausal patients with suspicious radiological features, irrespective of lesional size warrant a thorough gross examination, extensive sampling and a careful microscopic evaluation. key words: ovary, teratoma, malignant introduction ovarian teratomas are germ cell tumors (gct) that arise from ectopic pluripotent stem cells that fail to migrate from yolk sac endoderm to the urogenital ridge during embryogenesis. they comprise elements from all three embryological germ layers i.e. endoderm, mesoderm and ectoderm.1 teratomas include mature cystic teratomas (mct), immature teratomas (it) and monodermal teratomas (mot). mct are well differentiated relative to the germ cell layers while immature teratomas are incompletely differentiated and contain embryonic tissue along with neuroepithelium.2,3,4,5 the it are graded from i to iii on the basis of quantity of neuroepithelium.6,7 mot are composed of a single tissue type and usually contain thyroid tissue, in which case they are referred to as struma ovarii.8 malignancy can also arise in struma ovarii. the age of presentation of malignancy in struma ovarii is usually in the 5th decade of life, with thyroid type malignancies being the most common.9,10,11 presence of a concomitant malignancy arising in a teratoma is a diagnostic challenge not only for clinicians but also for pathologists. the preoperative risk assessment of teratomas with malignant transformations (mt) are challenging and it is of great importance that optimal management options be selected carefully.12 we hereby discuss the morphological spectrum of ovarian teratomas reported in the last six years (january 2015-december 2020) at the chughtai institute of pathology. material and methods after approval from institutional review board, a retrospective study was performed in all histologically diagnosed cases of ovarian teratomas in females of all ages, from january 2015 to december 2020 at the department of histopathology, chughtai institute of pathology. all diagnosed cases of mct, it of all grades, mot and malignancies arising in mct were included in the study. specimens received unfixed or received from an outside laboratory were excluded from the study. non-probability consecutive sampling was used. 32 morphological spectrum of ovarian teratomas: a 6 years experience at a specialized diagnostic institute the data was retrieved using institutional software nexus pro. a total of 1018 cases of teratomas were retrieved. clinico-pathological parameters including age, tumor size, histological type and laterality were obtained from pathology reports. statistical analysis: data was analyzed using spss version 20. the mean and standard deviation were calculated for quantitative variables including age and tumor size. mt was noted in both mct and mot. qualitative variables including histologic type and laterality has been presented in the form of frequencies and percentages. effect modifiers like age and tumor size have been controlled through stratification (for mt). post stratification chi-square test was applied by taking p value of 0.05 as significant. results a total of 1018 cases were included in the study. mean age was 30.7+/10.2 years. 46.6% (n=474) of the cases were right sided, 37.2% (n=379) were left sided while 7.0% (n=71) were bilateral (table-1). 39.7% (n=404) of the cases were between age group of 20-29 years (table-3). mean size of the tumor was 8.1 +/-3.6 cm, with 60.5% (n=616) cases being between 5 to 10cm. a total of 95.6% (n=973) were mct with 94.9% (n=966) diagnosed as benign mct (table-2). 0.72% (n=7) of the cases showed a malignancy arising in mct. 5 cases were of squamous cell carcinoma (71.4%), 1 was mucinous adenocarcinoma, 1 was high grade b-cell nonhodgkin lymphoma (nhl). mean age of cases with malignancy arising in mct was 45.1 years and mean size was 9.73cm (table-4). all malignancies arising in mct had age >30 years with 3.05% chance of malignancy over the age of 40 years compared to overall chance of malignancy of 0.72% for all age groups. this finding was significant with a p value of <0.001 (table-5). no correlation with size and malignancy was noted (table-6). 2.10% (n=21) cases were diagnosed as mot. 1 case of mot showed papillary thyroid carcinoma arising in it, while one case was of strumal carcinoid. 2.26% (n=23) were it (n=2 for grade i, n=9 for grade ii and n=12 for grade iii). 73.9% (n=17) of it were diagnosed in patients under 30 years of age. age age (years) 30.7 +/-10.2 years size size (cm) 8.1 +/-3.6cm frequency and percentage of laterality right 474 (46.6%) left 379 (37.2%) bilateral 71 (7.0%) not specified 94 (9.2%) different histologic subtypes mct 973 (95.6%) g1, it 2 (0.20%) g2, it 9 (0.88%) g3, it 12 (1.18%) mot 22 (2.16%) table-1: break down of results (n=1018) tumor size histologic type mct malignancy in mct g1, it g2, it g3, it mot malignancy in mot <5.0cm 166 0 0 0 1 5 0 5-10.0cm 585 5 2 6 7 10 1 >10.0cm 215 2 0 3 4 6 0 table-2: frequency of different histologic subtypes and tumor size stratification age group histologic type mct malignancy in mct g1, it g2, it g3, it mot malignancy in mot <10 years 4 0 1 2 4 0 0 10-19 years 83 0 1 1 2 1 0 20-29 years 391 0 0 4 2 7 0 30-39 years 309 1 0 2 2 4 0 40-49 years 133 3 0 0 2 4 0 >50 years 46 3 0 0 0 5 1 table-3: frequency of different histologic subtypes and age group stratification sr. no. diagnosis laterality age size 1 moderately differentiated squamous cell carcinoma right 42 5.0cm 2 high grade b-cell nhl right 40 8.2cm 3 moderately differentiated squamous cell carcinoma left 53 6.9cm 4 poorly differentiated squamous cell carcinoma right 55 18.0cm 5 moderately differentiated squamous cell carcinoma left 40 8.0cm 6 well differentiated squamous cell carcinoma right 51 12.0cm 7 mucinous adenocarcinoma left 35 10.0 8 papillary thyroid carcinoma in struma ovarii left 55 6.5cm table-4: malignancies arising in teratomas 33 morphological spectrum of ovarian teratomas: a 6 years experience at a specialized diagnostic institute age group benign mct malignancies in mct < 40 years 787 1 788 ≥ 40 years 179 6 185 966 7 973 (grand total) p=<0.00001 (significant=<0.05) table-5: age group and malignancies size benign mct malignancies in mct total <5.0cm 166 0 166 5-10.0cm 585 5 590 >10.0cm 215 2 217 total 966 7 973 (grand total) p=0.16 (significant=<0.05) table-6: size correlation and malignancies in mct discussion teratomas are one of the most common germ cell tumors with mature or immature histological features. mct are composed of well differentiated derivations from at least two of the three germ layers (ectoderm, endoderm and mesoderm) and are benign in nature whereas it, are rare tumors comprising incompletely differentiated tissue found either in pure form or as a component of a mixed germ cell tumor and occurs essentially during the first two decades of life.1,2,3,4,5 they clinically have a malignant behavior and are characterized by the presence of immature (neuroectodermal) tissue. immature teratomas are typically larger than mature cystic teratomas and are graded as i to iii according to the percentage of neuro epitelium. the higher the grade of the mass, the more aggressive behavior is expected.5,6 monodermal teratomas, as the name indicates, are composed predominantly or solely of one tissue type. there are three main types of ovarian monodermal tumors: struma ovarii, ovarian carcinoid tumors and neural tumors. struma ovarii is the commonest monodermal teratoma8 and has an excellent prognosis until malignant transformation occurs. malignant transformation most commonly occurs in the form of papillary thyroid carcinoma (ptc), however rarely follicular carcinoma, poorly differentiated carcinoma and anaplastic carcinoma can also arise in this tumor type.9,10,11 in this study we determined the morphological spectrum of more than a thousand ovarian teratomas (n=1018) diagnosed in a span of six years at chughtais institute of pathology, lahore. benign mct (fig-1a) were most frequent, comprising 94.9% (n=966) of total cases. mean age of the patients was 30.7 years with most of the cases (37.4%) presenting within the age group of 20-29 years. these findings are consistent with a study conducted by trabzonlu et al. in which mean age of the patients 34.5 years.13 average size of the teratomas was 8.01cm with 59.8% of the cases being in between 5-10cm. these findings are partially consistent with a study performed by rathore et al14 and trabzonlu et al. with an average size of 7.60cm and 8.37cm respectively. in our study majority of the teratomas were right sided (46.6%) with 7% being bilateral. right side predominance has also been reported by ismail et al. and chun et al.15,16 however, a study conducted by khan et al from peshawar showed left sided predominance.17 a. benign mature cystic teratoma b. well differentiated squamous cell carcinoma arising in mature cystic teratoma c. moderately differentiated squamous cell carcinoma 34 morphological spectrum of ovarian teratomas: a 6 years experience at a specialized diagnostic institute d. papillary thyroid carcinoma arising in monodermal teratoma fig-1: different morphological spectrum in teratomas. malignant transformation (mt) in a mct is a known phenomenon but is a rare occurrence. numerous studies have demonstrated a frequency of mt in mct to be not more than 2% and usually effecting patients in the post-menopausal age group.8,9,12a variety of malignant transformations are documented till date, the commonest being squamous cell carcinomas (88.3%), followed by adenocarcinoma, fibrosarcoma, rhabdomyosarcoma, malignant melanoma, urothelial carcinoma, adenosquamous carcinoma, endometrioid carcinoma, carcinosarcoma and papillary thyroid carcinoma.18-25 in our study, malignancy arising in mct, was seen in 7 cases, which comprises 0.72% of all included cases. a study conducted by ud din n et al. showed frequency of epithelial malignancies arising in mct as 1.9%.12 another study conducted by ulker et al. also showed malignant transformation in 1.9% of the total cases.20 mt was mostly seen in patients of peri-menopausal and postmenopausal age group with 6 cases out of a total of 7 being ≥ 40 years of age (85.8%). the mean age in our study for these mt cases was 45.1 years compared with 30.7 years for all cases of mct. this difference in age between benign mct and mt in mct was statistically significant with a p value of <0.001. our findings are consistent with these national and international studies. the average size of the teratomas with mt in mct was 9.73cm. there was no significant correlation between sizes of mt in mct and benign mct. previous studies have identified some risk factors for malignant transformations in mcts; these factors broadly include older age at presentation, post-menopausal status, and raised serum tumor markers; especially serum ca-125 levels. other findings consistently seen in patients with mt in mct include larger tumor masses with grossly visible areas of hemorrhage and necrosis.12 our findings are consistent with the general notion of a larger lesional size in case of mt in mct as compared to benign mct.12,13,21 most common malignancy reported to arise within the mct is squamous cell carcinoma and it usually comprises more than 70% of the malignancies.12,13,20-23,26 in our study, squamous cell carcinoma was the most frequent mt in mct comprising 71.4% (n=5) of all cases showing malignant transformation (fig-1b and 1c). one case was of mucinous adenocarcinoma while another case was of a high grade b-cell non-hodgkin lymphoma (nhl). our case with a mucinous adenocarcinoma arising in mct of the ovary was confirmed to have a germ cell origin confirmed by its reactivity for ck-20 immunohistochemical (ihc) stain and negativity for ck-7 ihc stain.27,28 ovarian b cell lymphomas are very rare and usually are of primary ovarian origin, without any associated teratoma. the high grade b-cell nhl included in our study was associated with an ovarian mct.29 overall frequency of immature teratomas was 2.26% (n=23) with mean age of 20.8 years, mean size of 8.62cm and were right sided predominantly (n=12, 52.2%). most of immature teratomas were of grade 3 (n=12, 52.2%). a study conducted by gallion et al. also showed majority of cases of immature teratomas being diagnosed in patients under 25 years of age.30 a study conducted in india showed a median age of 19 years. in this study, grade ii (it) were most frequent, comprising 46% of the cases while grade iii (it) comprised 25% of the cases.5 these results were in contrast to our findings in which grade iii (its) were the most frequent. 22 mot were included in the study with an average age of 36.3 years and a mean size of 7.29cm. 1 case (4.55%) showed papillary thyroid carcinoma (ptc) arising in the background of mot (fig-1d). the age of that patient was 55 years with tumor size being 6.50cm. one case was of strumal carcinoid (4.55%). the frequency of ptc and strumal carcinoid arising in mot is reported to be in between 5% to 10%.31,32,33 conclusion ovarian teratomas have a diverse morphological spectrum including the mature cystic, immature and monodermal subtypes. malignancies arising in each one of these subtypes is a rare phenomenon. however, ovarian masses in postmenopausal patients with suspicious radiological features, 35 morphological spectrum of ovarian teratomas: a 6 years experience at a specialized diagnostic institute irrespective of lesional size warrant a thorough gross examination with extensive sampling of the specimen and a careful microscopic evaluation. correlation with serum tumor markers and intraoperative consultation in such cases should be sought to determine the best modality of treatment for these patients. references 1. outwater ek, siegelman es, hunt jl. ovarian teratomas: tumor types and imaging characteristics. radiographics. 2001 mar; 21(2):475-90. 2. peterson cm, buckley c, holley s, menias co. teratomas: a multimodality review. current problems in diagnostic radiology. 2012 nov 1; 41(6):210-9. 3. wetherell d, weerakoon m, williams d, beharry bk, sliwinski a, ow d, manya k, bolton dm, lawrentschuk n. mature and immature teratoma: a review of pathological characteristics and treatment options. med surg urol. 2014; 3(124):2. 4. abiko k, mandai m, hamanishi j, matsumura n, baba t, horiuchi a, mikami y, yoshioka s, wakasa t, shiozawa t, konishi i. oct4 expression in immature teratoma of the ovary: relevance to histologic grade and degree of differentiation. the american journal of surgical pathology. 2010 dec 1; 34(12):1842-8. 5. deodhar kk, suryawanshi p, shah m, rekhi b, chinoy rf. immature teratoma of the ovary: a clinicopathological study of 28 cases. indian journal of pathology and microbiology.2011oct 1;54(4):730. 6. cecchetto g. gonadal germ cell tumors in children and adolescents. journal of indian association of pediatric surgeons. 2014 oct; 19(4):189. 7. schneider dt, terenziani m, cecchetto g, olson ta. gonadal and extragonadal germ cell tumors, sex cord stromal and rare gonadal tumors. in rare tumors in children and adolescents 2012 (pp. 327402). springer berlin heidelberg. 8. kondi-pafiti a, mavrigiannaki p, grigoriadis ch, kontogianni-katsarou k, mellou a, kleanthis ck, liapis a. monodermal teratomas (strumaovarii). clinicopathological characteristics of 11 cases and literature review. european journal of 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in mature cystic teratoma of the ovary: a retrospective study of eight cases and review of literature. przegladmenopauzalny= menopause review. 2018 jun; 17(2):63. 15. ismail sr. an evaluation of the incidence of rightsided ovarian cystic teratoma visualized on sonograms. jdiagnostmedsono.2005jul;21(4):336-42 16. chun s, jeon gh, cho hj, ji yi. comparison between incidence of right-and left-sided ovarian cystic teratomas. journal of reproductive endocrinology. 2012 apr 1; 4(1):43-8. 17. khan mm, sharif n, ahmad s. morphological spectrum of mature ovarian teratoma. gomal j med sci 2014; 12:76-80. 18. bal a, mohan h, singh sb, sehgal a. malignant transformation in mature cystic teratoma of the ovary: report of five cases and review of the literature. archives of gynecology and obstetrics. 2007 mar; 275(3):179-82. 19. ulker v, numanoglu c, akbayir o, akyol a, tuncel a, akca a, aydin o. malignant transformation arising from mature cystic teratoma of the ovary: a report of six cases. j obs gynaecol res. 2012 may; 38(5):849-53. 20. hackethal a, brueggmann d, bohlmann mk, franke fe, tinneberg hr, münstedt k. squamouscell carcinoma in mature cystic teratoma of the ovary: systematic review and analysis of published data. the lancet oncology.2008 dec 1;9(12):117380 21. m.m. desouki, o. fadare, b.k. chamberlain, n. shakir, a. kanbour-shakir, malignancy associated with ovarian teratomas: frequency, histotypes, and diagnostic accuracy of intraoperative consultation, ann. diagn. pathol. 19(june (3)) (2015) 103-106. 22. l. dos santos, e. mok, a. iasonos, k. park, r.a. soslow, c. aghajanian, et al., squamous cell carcinoma arising in mature cystic teratoma of the ovary: acase series and review of the literature, gynecol. oncol. 105 (may (2)) (2007)321–324. 23. m. sakuma, t. otsuki, k. yoshinaga, h. utsunomiya, s. nagase, t. takano, et al.,malignant transformation arising from mature cystic teratoma of the ovary: aretrospective study of 20 cases, int. j. gynecol. cancer 20 (july (5)) (2010)766–771) malignancy in post-menopausal patient. 24. kudva r, ayachit gs, ayachit a. malignant melanoma arising in an ovarian mature cystic teratoma-a rare entity. journal of clinical and diagnostic research: jcdr. 2015 apr; 9(4):ed14. 36 morphological spectrum of ovarian teratomas: a 6 years experience at a specialized diagnostic institute 25. sanz-baro r, alvarez g, fernández-aceñero mj, di fiore ha. ovarian rhabdomyosarcoma arising from mature cystic teratoma. 26. sharma ivy, saikia cj. squamous cell carcinoma arising in mature cystic teratoma – a rare case report. paripex-indian j res. 2015; 4:168-9. 27. kerr se, flotte ab, mcfalls mj, vrana ja, halling kc, bell da. matching maternal isodisomy in mucinous carcinomas and associated ovarian teratomas provides evidence of germ cell derivation for some mucinous ovarian tumors. the american journal of surgical pathology. 2013 aug 1; 37(8):1229-35. 28. snir ol, buza n, hui p. mucinous epithelial tumours arising from ovarian mature teratomas: a tissue genotyping study. histopathology. 2016 sep; 69(3):383-92. 29. afzal s, zaman s. high grade b-cell nonhodgkin’s lymphoma arising in a mature cystic teratoma of the ovary: a case report. international journal of molecular and cellular medicine. 2017; 6(4):239. 30. gallion h, van nagell jr, donaldson es, hanson mb, powell df. immature teratoma of the ovary. american journal of obstetrics and gynecology. 1983 jun 15; 146(4):361-5. 31. szczepanek-parulska e, pioch a, cyranska-chyrek e, wolinski k, jarmołowska-jurczyszyn d, janickajedynska m, majewski p, zabel m, ruchala m. the role of immunohistochemical examination in diagnosis of papillary thyroid cancer in strumaovarii. folia histochemicaetcytobiologica. 2019;57(1):35-42 32. wei s, baloch zw, livolsi va. pathology of strumaovarii: a report of 96 cases. endocrine pathology. 2015 dec; 26(4):342-8. 33. nishida m, kawano y, yuge a, nasu k, matsumoto h, narahara h. three cases of immature teratoma diagnosed after laparoscopic operation. clinical medicine insights. case reports. 2014; 7:91-4. the authors: dr. fatima khalid consultant histopathologist, department of histopathology, chughtai institute of pathology, lahore. dr. sameen afzal consultant histopathologist, department of histopathology, chughtai institute of pathology, lahore. dr. ghazi zafar consultant histopathologist, department of histopathology, chughtai institute of pathology, lahore. dr. anila chughtai consultant histopathologist, department of histopathology, chughtai institute of pathology, lahore. dr. samina zaman consultant histopathologist, department of histopathology, chughtai institute of pathology, lahore. dr. akhtar sohail chughtai consultant histopathologist, department of histopathology, chughtai institute of pathology, lahore. corresponding author: name: dr. fatima khalid consultant histopathologist, department of histopathology, chughtai institute of pathology, lahore. e-mail: faatimaa1988@gmail.com 6 proceedings s.z.m.c. vol: 35(3): pp. 6-11, 2021. pszmc-801-35-3-2021 efficacy of telmisartan in pristane induced arthritis rat model 1quratulain mehdi, 2noaman ishaq, 3saba batool, 4kulsoom farhat, 5saman omer, 4arooj shahid 1department of pharmacology, quetta institute of medical sciences, quetta 2department of pharmacology, bakhtawar amin medical and dental college, multan 3department of pharmacology, cmh multan institute of medical sciences, multan 4department of pharmacology, army medical college, rawalpindi 5department of pharmacology, mohtarma benazir bhutto shaheed medical college, mirpur ajk abstract introduction: rheumatoid arthritis is one of the most common systemic inflammatory diseases characterized by progressive damage to the joints. there is rising evidence that renin angiotensin aldosterone system signaling is also involved in the inflammatory response in rheumatoid arthritis and its blockers possess anti-arthritic properties. telmisartan is an angiotensin receptor blocker and ppar-γ agonist and its anti-arthritic effects were evaluated. aims & objectives: this experimental study was designed to evaluate the anti-arthritic efficacy of telmisartan in pristane induced rat model of arthritis in adult female rats. place and duration of study: the study was conducted in the department of pharmacology, army medical college, rawalpindi, in collaboration with national institute of health and armed forces institute of pathology from july 2020 to august 2020. material & methods: twenty four (24) adult non-pregnant female sprague dawley rats were divided in three groups (n=8) designated as group a (normal control), group b (arthritic control) and group c (telmisartan group) & maintained on standard diet and water adlibitum. rheumatoid arthritis was induced in each rat of groups b &c by a single intradermal injection of 0.5ml pristane at the base of its tail on day 0 and the disease developed in two weeks. all 3 groups were given distilled water 2.5 ml/kg from 2-4 weeks and group c was additionally given dissolved telmisartan orally at 2 mg/kg/day. anti-arthritic efficacy was determined by assessing arthrogram score and total leukocyte count on day 0, 14 and 28 along with histological examination done at the end of the study. data analysis was done using spss version 25. results: healthy rats in group a maintained a unremarkable arthogram & histogram score & tlc count of 6675±350/μl during the entire study period. telmisartan administration in group c for two weeks after pristane induction resulted in significant reduction in arthrogram score (as) 9.5±3.66, total leukocyte count (tlc) 7350±550/μl and histological score (hs) to 6.88±1.24 as compared to (as) 14.50±2.07, wbc 10150±350/μl & (hs) 10.75±2.05 in group b, left untreated with pristane alone. the intergroup comparison showed significant p value < 0.05 respectively. conclusion: anti-arthritic effect was shown by telmisartan as it was able to ameliorate the changes induced by pristane. key words: anti-arthritic efficacy, telmisartan, pristane, rheumatoid arthritis introduction rheumatoid arthritis (ra) is one of the most common inflammatory disorders occurring due to multiple factors including environmental and genetic risk factors.1 clinical presentation of symmetrical joint involvement comprises of arthralgia, swelling, redness and limited mobility. the risk of development of extra articular manifestations such as keratitis, rheumatoid nodules, pericarditis and pleuritis is increased with poor control or severe disease. it is responsible for continuing disability, premature death and increased socioeconomic burden.2 several strategies are employed in managing the disease. surgical approaches help in improvement of functional activity and structural misalignment of joints but considered as a last resort. physical and occupational therapy helps in relieving the muscular spasm and related joint stiffness.3 current therapies include non steroidal anti-inflammatory drugs (nsaids), glucocorticoids (gcs) and biological agents which improve pain, fatigue and disability with more focus on controlling synovitis.4 treatment modalities with fewer side effects are required for improvement of therapy. 7 efficacy of telmisartan in pristane induced arthritis rat model the renin angiotensin aldosterone system (raas) is well known for its function as regulator of blood pressure but there is accumulating validation that its signaling is involved in reaction to inflammation identified in various disease states. angiotensin ii type 1 receptor (at1r) can be presumed as a potential therapeutic target as rodent models of arthritis have massively expressed its presence in cultured ra-fibroblast like synoviocytes (fls) as well as in hyperplastic synovium. the role of angiotensin converting enzyme inhibitors (aceis) and angiotensin receptor blockers (arbs) in ameliorating the laboratory and clinical parameters has also been documented.5 telmisartan showed betterment in the parameters of arthritis in a rat model of ra. it exerts anti-inflammatory actions by restraining tumor necrosis factor α (tnf-α) production and inhibiting pro-inflammatory mediators such as interleukin-6 (il-6) and nuclear factor kappa beta (nf-κβ).6 various experiments and studies are being carried out to explore the anti-inflammatory properties of telmisartan with regards to various conditions. the current study is intended to demonstrate the antiarthritic effects of telmisartan in rat model of pristane induced arthritis (pia). material and methods study design: laboratory based experimental study. ethical approval: the ethics review committee of “center for research in experimental and applied medicine (cream)”, army medical college gave the approval for study. setting: it was carried out in department of pharmacology and therapeutics, army medical college (amc), rawalpindi in collaboration with national institute of health (nih), islamabad and armed forces institute of pathology (afip). study was conducted for duration of one month from july to august 2020 and the animals were kept in animal house of nih islamabad. sampling technique and size: twenty four (24) adult non-pregnant female sprague dawley rats weighing 250-300 grams were randomly divided into three groups of eight animals each. experimental setup: the rats were acclimatized for a week before the start of experiment. they were retained in wire topped cages and standard environmental conditions were maintained with temperature ranging between 25±5˚c and 50±10 % humidity. free excess to clean drinking water and standard rodent diet ad libitum was provided during the study duration. twenty four (24) adult nonpregnant female sprague dawley rats were divided in three groups (n=8) and were assigned as group a (normal control): comprised of healthy rats. intradermal injection of 0.1 ml normal saline was given at the base of tail at day 0 and rats were given distilled water 2.5 ml/kg/day orally from 14 to 28 days. group b (arthritic control): rheumatoid arthritis was induced with pristane on day 0 and rats were given distilled water 2.5 ml/kg/day orally from 14 to 28 days after induction. group c (telmisartan group): rheumatoid arthritis was induced with pristane on day 0 and rats were treated with telmisartan manufactured by pharm evo (pvt) ltd in a dose of 2 mg/kg/day orally for two weeks after induction.7 induction of rheumatoid arthritis: induction of ra was done by a single intradermal injection of 0.5 ml of pristane purchased from martin dow marker ltd at the base of tail of each rat in group b and group c.8 arthritis developed gradually over a period of two weeks. assessment of arthritic progression by arthrogram score: the assessment of arthritic progression was done by clinical scoring of joint inflammation on all four limbs on alternate days from week 0 to 4 weeks after pristane injection. it ranged from 0-4, where 0=no swelling and redness; 1=single joint involvement; 2=involvement of two joints; 3= involvement of more than two joints; 4=severe arthritis involving whole paw and digits. the collective minimum score for all four limbs would be 0 and maximum would be 16.9 the arthrogram score at day 0, 14 and 28 were statistically analyzed. hematological analysis: blood was collected from the lateral tail vein on day 0, 14 and 28 and was stored in ethylene diamine tetra acetic acid (edta) tubes for the analysis of total leukocyte count (tlc).10 it was done by using sysmex kx-21 automated analyzer. joint histopathology: all the rats were euthanized by chloroform at the end of the study on day 28. the proximal part of tibia was sliced by using 5.5" angled bone cutter and right ankle joints were removed. slides were prepared with eosin and hematoxylin stains. histological scoring was done according to a set criteria as described by alabarse and his colleagues. for synovial inflammation, five high‐power magnification fields were scored for the percentage of infiltrating mononuclear cells as follows: 0, absent; 1, mild (1-10%); 2, moderate (11-50%); 3, severe (51-100%); for synovial hyperplasia: 0, absent; 1, mild (5-10 layers); 2, moderate (11-50 layers); 3, severe (>20 layers); for 8 efficacy of telmisartan in pristane induced arthritis rat model extension of pannus formation: 0, absent; 1, mild; 2, moderate; 3, severe; for synovial fibrosis: 0, absent; 1, mild (1-10%); 2, moderate (11-50%); 3, severe (51-100%); for cartilage erosion, that is, the percentage of the cartilage surface that was eroded: 0, absent; 1, mild (1-10%); 2, moderate (11-50%); 3, severe (51-100%); and for bone erosion: 0, none; 1, minor erosion(s) observed only at high‐power magnification fields; 2, moderate erosion(s) observed at low magnification; 3, severe transcortical erosion(s).11 statistical analysis: the statistical package for the social sciences (spss) version 25 was used for data analysis with quantitative data expressed as mean + standard deviation (sd). one way of analysis of variance (anova) was applied to observe the mean difference among control and experimental group and compared by post hoc tukey’s test. the difference between two observations was considered significant if the p value was equal to or less than 0.05 (p ≤0.05). results the arthrogram score was zero in all the groups at day 0. it remained zero in group a throughout the study duration whereas in group b it reached a mean of 13.5±2.97 by day 14 and reached a mean of 14.5±2.07 by day 28. fig-1 shows a rat of group b with arthrogram score 12. arthrogram score of group c reached a mean of 12.5±3.96 by day 14 and was declined after treatment with telmisartan by day 28 with a mean of 9.5±3.66. fig-2 shows a rat of group c with arthrogram score 04. fig-1: rat of group b with hind paws showing edema and redness fig-2: rat of group c with improved arthrogram score. the mean tlc of group a, b and c was 6600±300/μl, 6562±450/μl and 6488±360/μl respectively on day 0. it remained almost constant in group a on day 14 and day 28 with a mean of 6600±310/μl and 6675±380/μl. tlc was increased on day 14 in the other two groups with mean of 9988±510/μl and 9987±710/μl. group b was left untreated and its mean tlc on day 28 was 10150±350/μl whereas a dosing schedule of 2 mg/kg/day of telmisartan led to a decrease in levels of tlc to 7350±550/μl. the histological examination of group a revealed minimum score as it was disease free and did not show any microscopic changes. the synovium lining was smooth with no penetration of infiltrating cells. maximum score was seen in the rats of group b which ranged from 8-13. marked percentage of infiltrating mononuclear cells was seen in four out of six slides while rest showed moderate infiltration. two slides showed severe synovial hyperplasia but was moderate in other slides. the extension of pannus formation was maximum in two slides while the others showed mild to moderate pannus formation. six out of eight slides showed moderate synovial hyperplasia. all the slides showed cartilage erosion. mild to moderate presence of bone erosion was seen in all slides except one. mean score of this group was 10.75±2.05. fig-3 shows a slide of group b with a score of 10 showing bone erosion and inflammation with infiltrating cells. slides of group c showed improvement. the score ranged from 5-9. two slides showed moderate infiltrating mononuclear cells, two showed no infiltration while four showed mild infiltration of mononuclear cells. mild to moderate synovial hyperplasia was present in all the slides. mild pannus formation and synovial fibrosis was a feature of all the groups. five slides showed mild cartilage erosion, whereas the remaining slides showed moderate erosion. mild bone erosion was seen in five slides and moderate in two slides. the mean score of this group was 6.88±1.24. fig-4 shows a score of 07 with surface corrugation. 9 efficacy of telmisartan in pristane induced arthritis rat model fig-3: photomicrograph of ankle joint of group b fig-4: photomicrograph of ankle joint of a rat of group c treatment with telmisartan for two weeks decreased the arthrogram score, tlc and also showed improvement on histological examination. the intergroup comparison of the three groups of all the parameters depicted a statistically significant p value ˂ 0.001 as shown in table-1. parameter group a (n=8) group b (n=8) group c (n=8) p value anova mean±sd mean±sd mean±sd arthrogram score 0 14.50±2.07 9.5±3.66 ˂ 0.001* total leukocyte count (per μl) 6675±380 10150±350 7350±550 ˂ 0.001* histopathology score 0 10.75±2.05 6.88±1.24 ˂ 0.001* * significant p value ≤0.05 table-1: intergroup comparison of arthrogram score, total leukocyte count and histopathology score on day 28. discussion ra is one of the common types of arthritis in the elderly population affecting the quality of life. the main pathological characters include persistent chronic synovitis, dysplasia of synovial membrane and pannus formation eventually causing cartilage and joint obliteration.12 pia is one of the extensively used models for research purpose due to appreciative resemblance to the chemical characteristics of ra.13 it is similar to the condition seen in humans due to the presence of symmetrical joint involvement, presence of rheumatoid factor, bone and cartilage destruction as well as dependence of specific immunity seen in such models.14 in the present study the results indicated that the development of arthritis correlated with the increase in arthrogram score, tlc and histological score in group b as compared to the group a (normal control) till day 14. these findings are backed up by a number of studies conducted by researchers. chen and his fellows in 2019 also observed the signs of severe arthritis spreading to ankle joints two weeks after injecting pristane.15 regarding tlc, tissue homeostasis is maintained by the monitored movement of leukocytes between lymphoid and peripheral tissue through various stromal compartments. this trafficking is disturbed in ra.16 feng and qiu in 2018 successfully created a model of ra by subcutaneous injection of emulsion. the results indicated higher leukocyte count as well as damaged cartilage and considerable infiltration of inflammatory cells.17 we followed a therapeutic model by commencing the treatment with telmisartan on day 14. it significantly lowered the aforementioned parameters when compared with the group b (arthritic control). group c (telmisartan group) displayed the absence of swelling, redness and an overall decrease in the mean arthritic score. these findings were in accordance with the work of hasanin and mohamed who studied the anti-inflammatory effects of telmisartan in a rat model and saw a significant decrease in arthritic parameters.6 two weeks treatment with telmisartan also displayed a decrease in tlc and histological score. angiotensin ii (ang ii) has a central role in leukocyte migration and infiltration at molecular and cellular level at the sites of inflammation.18 it is accountable for regulating proliferation of cells and apoptosis by binding to at1r. ang ii exhibited from synovial tissue regulates synovial perfusion and growth by its actions on synovial at1r.19 thus the decrease in tlc can be justified by its at1r blocking potential. other than this, olmesartan, an arb like telmisartan was used orally in a model of zymosan induced ra. it decreased the tlc with specific reduction in neutrophil count as well as showed improvement in histological score.20 this signifies that our experimental drug telmisartan may also have anti-arthritic effect against ra. limitations of study: this study has also some limitations. although the study showed that telmisartan has anti-arthritic properties, the preventive role of telmisartan should 10 efficacy of telmisartan in pristane induced arthritis rat model also be evaluated in such a model. more over future studies should also be conducted using this drug focusing on adverse effects. conclusion oral administration of 2 mg/kg/day of telmisartan for two weeks significantly lowered the mean arthrogram score, total leukocyte count and histological score in a rat model of rheumatoid arthritis. therefore, it can be concluded that telmisartan has anti-arthritic effects. acknowledgement: authors are thankful to dr. hussain ali, head of research animal facility, national institute of health, islamabad for his supervision regarding induction of rheumatoid arthritis. a special thanks to army medical college, rawalpindi for facilitating us to conduct this project. references 1. abhishek a, doherty m, kuo cf, mallen cd, zhang w, grainge mj. rheumatoid arthritis is getting less frequent-results of a nationwide population-based cohort study. rheumatology. 2017 may 1; 56(5):736-44. 2. guo q, wang y, xu d, nossent j, pavlos nj, xu j. rheumatoid arthritis: pathological mechanisms and modern pharmacologic therapies. bone res. 2018 apr 27; 6(1):1-4. 3. bullock j, rizvi sa, saleh am, ahmed ss, do dp, ansari ra, ahmed j. rheumatoid arthritis: a brief overview of the treatment. medical princpract. 2018; 27(6):501-7. 4. xu q, zhou y, zhang r, sun z, cheng lf. antiarthritic activity of qi-wu rheumatism granule (a chinese herbal compound) on complete freund’s adjuvant-induced arthritis in rats. evid based complement alternat med. 2017 jan 1; 2017:1960517 5. terenzi r, manetti m, rosa i, romano e, galluccio f, guiducci s, ibba-manneschi l, matucci-cerinic m. angiotensin ii type 2 receptor (at2r) as a novel modulator of inflammation in rheumatoid arthritis synovium. sci rep. 2017 oct 16; 7(1):1-1. 6. hasanin ah, mohamed rh. telmisartan alone or in combination with etanercept improves anemia associated with rheumatoid arthritis in rats: a possible role of anti-inflammatory and renoprotective effects. pharmacol rep. 2020 apr; 72(2):379-88. 7. yanagihara h, ushijima k, arakawa y, aizawa ki, fujimura a. effects of telmisartan and olmesartan on insulin sensitivity and renal function in spontaneously hypertensive rats fed a high fat diet. j pharmacol sci. 2016 jul 1; 131(3):190-7. 8. faisal r, gul m, anwar a. low doses of pristane in comparison with high doses in induction of arthritis in female-spraguedawley rats. khyber med univ j. 2016 dec 24; 8(4):185-8. 9. gul a, kunwar b, mazhar m, faizi s, ahmed d, shah mr, simjee su. rutin and rutin-conjugated gold nanoparticles ameliorate collagen-induced arthritis in rats through inhibition of nf-κb and inos activation. intimmunopharmacol. 2018 jun; 1(59):310-7. 10. faisal r, chiragh s, popalzai aj, rehman ku. anti inflammatory effect of thymoquinone in comparison with methotrexate on pristane induced arthritis in rats. j pak med assoc. 2015 may 1; 65(5):519-25. 11. alabarse pv, lora ps, silva jm, santo rc, freitas ec, de oliveira ms, almeida as, immig m, teixeira vo, filippin li, xavier rm. collagen‐induced arthritis as an animal model of rheumatoid cachexia. journal of cachexia, sarcopenia and muscle. 2018 jun; 9(3):603-12. 12. cai p, jiang t, li b, qin x, lu z, le y, shen c, yang y, zheng l, zhao j. comparison of rheumatoid arthritis (ra) and osteoarthritis (oa) based on microarray profiles of human joint fibroblast‐like synoviocytes. cell biochemistry and function. 2019 jan; 37(1):31-41. 13. wang b, zhao p, zhou y, meng l, zhu w, jiang c, wang l, cai y, lu s, hou w. increased expression of th17 cytokines and interleukin-22 correlates with disease activity in pristane-induced arthritis in rats. plos one. 2017 nov 28; 12(11):e0188199. 14. ruiz jt, luján l, blank m, shoenfeld y. adjuvantsand vaccines-induced autoimmunity: animal models. immunologic research. 2017 feb 1; 65(1):55-65. 15. chen q, zhang x, xiong y, chen c, lv s. the cd25+/cd4+ t cell ratio and levels of cii, cix and cxi antibodies in serum may serve as biomarkers of pristane-induced arthritis in rats and rheumatoid arthritis in humans. comparative biochemistry and physiology part c: toxicology & pharmacology. 2019 mar 1; 217:25-31. 16. buckley cd, mcgettrick hm. leukocyte trafficking between stromal compartments: lessons from rheumatoid arthritis. nature reviews rheumatology. 2018 aug; 14(8):476-87. 17. feng fb, qiu hy. effects of artesunate on chondrocyte proliferation, apoptosis and autophagy through the pi3k/akt/mtor signaling pathway in rat models with rheumatoid arthritis. biomedicine & pharmacotherapy. 2018 jun 1; 102:1209-20. 18. piqueras l, sanz mj. angiotensin ii and leukocyte trafficking: new insights for an old vascular mediator. role of redox-signaling pathways. free radical biology & medicine. 2020 feb11:157:38-54 19. wu y, lu x, li m, zeng j, zeng j, shen b, zeng y. renin-angiotensin system in osteoarthritis: a new potential therapy. international immunopharmacology. 2019 oct 1:75:105796. 20. guerra gc, de menezes ms, de araújo aa, de araújojúnior rf, de medeiros ca. olmesartan 11 efficacy of telmisartan in pristane induced arthritis rat model prevented intra-articular inflammation induced by zymosan in rats. biological and pharmaceutical bulletin. 2016 nov 1; 39(11):1793-801. the authors: dr. quratulain mehdi demonstrator, department of pharmacology, quetta institute of medical sciences, quetta. dr. noaman ishaq assistant professor, department of pharmacology, bakhtawar amin medical and dental college, multan. dr. saba batool assistant professor, department of pharmacology, cmh multan institute of medical sciences, multan. dr. kulsoom farhat associate professor, head, department & pharmacology, army medical college, rawalpindi. dr. saman omer demonstrator, department of pharmacology, mohtarma benazir bhutto shaheed medical college, mirpur ajk. dr. arooj shahid demonstrator, department of pharmacology, army medical college, rawalpindi. corresponding author: dr. quratulain mehdi demonstrator, department of pharmacology, quetta institute of medical sciences, quetta. e-mail: qurat.mehdi84@gmail.com 58 proceedings s.z.m.c. vol: 35(3): pp. 58-63, 2021. pszmc-810-35-3-2021 phoenix dactylifera (ajwa date) whole fruit, flesh and powdered seed prevents anti-tuberculous drug induced hepatotoxicity in rabbits 1sadia majeed, 2usman aslam, 3muhammad ubaid ullah khan, 4asma inam, 2sadia chiragh 1department of pharmacology, continental medical college, lahore 2department of pharmacology, al-aleem medical college, lahore 3allama iqbal medical college, lahore 4department of pharmacology, azra naheed medical college, lahore abstract introduction: hepatotoxicity induced by anti-tuberculous medicine is known due to their oxidative stress. ajwa dates may have a role to protect liver from oxidative stress aims & objectives: to assess the preventive effect of ajwa date on hepatotoxicity induced by anti-tuberculous drugs in rabbits. place and duration of study: post graduate medical institute, lahore for three months, from may 2014 to july 2014. material & methods: thirty rabbits were distributed into five groups. rabbits of group a and of b were fed on normal diet in form of pellets. group c, d and e were provided diet containing one whole ajwa date, flesh of one ajwa date and powdered seed of one ajwa date respectively in each 100 grams of diet throughout study. group b, c, d and e were administered 50mg/kg isoniazid and 100mg/kg rifampicin orally for 14 days. serum levels of liver enzymes alanine transaminase (alt), aspartate transaminase (ast) and alkaline phosphatase (alp) and bilirubin were estimated on day 0 and 14. one way anova followed by post hoc tukey’s test and t-test were applied for statistical analysis using spss 20. results: baseline lfts were normal in all groups. significant hepatotoxicity was observed after 2weeks of inh and rifampicin administration in disease control group b (alt 200.2±19.3 & alp 231.0±21.3 iu/l, ast 139.0±22 & bilirubin 0.48±0.046mg/dl, (p value < 0.001) as compared to healthy control group a (alt47.2 ± 6.7 & alp78.2 ±5.0 iu/l, ast 43.0 ± 9.7, bilirubin 0.10± 0.00mg/dl). (p value < 0.001). concomitant ajwa intake during the same period resulted in an equipotent significantly similar improvement in lfts in groups c (whole date) alt55.7 ± 4.7&alp 91.5 ±5.0iu/l, ast, 59.0 ± 15.3 &bilirubin 0.09 ±0.02 mg/dl): d (flesh) alt89.8 ± 6.3 & alp111.3 ±9.4 iu/l, ast73.7 ± 8.3 & bilirubin0.12± 0.04 mg/dl & e (seed powder) alt85.8 ± 8.6 iu/l &alp 92.8 ±11.4 iu/l, ast57.5 ± 5.3 & bilirubin 0.12 ±0.04 mg/dl) versus group b (p value < 0.001). and near normalization of liver function close to that of healthy control group conclusion: co-administration of ajwa date whole fruit, flesh and seed powder are equipotent and effective in preventing isoniazid and rifampicin induced hepatotoxicity key words: ajwa date, hepatoprotective, rifampicin, isoniazid. introduction tuberculosis has remained the prime culprit behind mortality among people owing to a single infectious agent,1 with over 10 million people getting infected along with 1.5 million deaths occurring in 2018.2 in 2019, pakistan alone contributed 5.7% of cases to the global burden of tb.3 for the treatment of drug susceptible pulmonary tuberculosis, isoniazid (inh), rifampicin (rif), pyrazinamide (pza) and ethambutol (emb) are prescribed during the intensive phase while isoniazid and rifampicin are continued into the continuation phase of anti-tubercular treatment (att) for 8 weeks and 18 weeks, respectively.4 liver is the prime organ involved in metabolism, activation and detoxification of almost all compounds reaching it through circulation. many natural and synthetic compounds metabolised by liver may produce acute and chronic heterogeneous responses, the responses being outlined as druginduced hepatotoxicity or drug-induced liver injury (dili),5 where dili being strictly defined by ast levels greater than 5 times the upper normal limit.6 the significance of dili in respect of treatment is that of it being the sole adverse indication for halting an ongoing treatment or withdrawal of a drug from consumer market. for this reason, 59 phoenix dactylifera (ajwa date) whole fruit, flesh & powdered seed prevents anti-tuberculous drug… prevention of some diseases, including tuberculosis, has been problematic.7 anti-tubercular drugs i.e., isoniazid and rifampicin, along with pyrazinamide, are known to have adverse side effects including drug-induced hepatotoxicity.8 it has been demonstrated that a prolonged isoniazid and rifampicin combination therapy can induce hepatic stellate cells and fibrotic changes through oxidative stress.9 to ensure the progress of treatment avoiding hepatotoxicity, reduction in drug-induced oxidative stress can prove beneficial. a study illustrated how administration of antioxidants like vit e and vit c can reduce the levels of att induced oxidative stress by elevating levels of anti-oxidant enzymes.10 phoenix dactylifera is a flowering plant belonging to palm family, arecaceae. the fruit of which is an edible date, ajwa being the best variety. ajwa date has been used for its several benefits which include hepatoprotective,11 nephroprotective,12 anti-cancer13 and antioxidant and anti-inflammatory effects14 among many others. ajwa date has shown to possess bioactive compounds that halt lipid peroxidation and inhibit cox1 and cox2.14. our literature review has given us insight into hepatoprotective effects of ade. we aim to explore this effect against anti-tuberculous drugs in a rabbit model using natural product instead of extract. material and methods this randomized control study was conducted at post graduate medical institute (pgmi), lahore after approval of the institutional ethical committee for basic sciences from may 2014 to july 2014. sample size of 6 rabbits in each group was calculated using 90% power of study and 5% level of significance.15 adult healthy male rabbits, 30 in number were selected according to inclusion criteria of 1.2 to 1.5 kg. they were bought from the local market kept in cages for 15 days for acclimatization and randomly divided into five groups (a, b, c, d and e). one group (a) was administered distilled water in a volume equivalent to drug volume administered to the rest of the four groups. the other four groups (b, c, d and e) were administered isoniazid 50 mg/kg (schazoozaka pharmaceuticals, pakistan ltd) and rifampicin 100 mg/kg (schazoozaka pharmaceuticals, pakistan ltd).16 both drugs were dissolved in distilled water and kept in dark bottles at 4°c. they were administered as a single daily dose in the morning for a period of fourteen days. group a and b rabbits were given a preparation of diet pellets constituted of split chickpeas 500g, dry fodder 500g and jawar 500g. plain flour 300g was used with water to bind all these ingredients in the form of pellets, which were then air dried and refrigerated. diets for group c, d and e were prepared by mixing flesh and seed powder (group c), flesh only (group d) and seed powder only (group e) of one ajwa date in 100gm of abovementioned diet. diet pellets and tap water was provided to all groups ad libitum. blood samples of about 2 ml were collected from the marginal ear vein while the animal was kept in a restraint cage, by using a 3 ml disposable (bd, pakistan) syringe with a 23-gauge needle. samples were collected on day 0 and 14. blood was collected and put in a clot activator vacutainer (biovac, pakistan). after half an hour it was centrifuged at 2500 revolutions per minute for 10 minutes and serum was separated and stored at -20°c. serum samples were analyzed for estimation of alanine aminotransferase (ast), aspartate aminotransferase (alt), alkaline phosphatase (alp) and bilirubin. on day of estimation, stored samples were thawed to room temperature and kept in incubator (din-12880-k1, germany) for 5 to 10 minutes at 37°c. serum alanine aminotransferase (ast), aspartate aminotransferase (alt) and alkaline phosphatase were estimated using diagnostic kits (diasys, germany) and serum bilirubin was estimated using diagnostic kit (human, germany) using spectrophotometer (slim by seac, italy). significant rise above baseline and normal control (p value <0.05) was considered induction of hepatotoxicity. statistical analysis: data was analyzed by spss 20. the variables were expressed as mean ± sd after confirmation of normal distribution at baseline. one way anova followed by post hoc tukey’s test was applied for comparison among groups of rabbits. paired sample t test was used for comparison between day 0 and 14 in each group. p value<0.05 being considered significant. results thirty adult healthy male rabbits weighing 1200 1500 grams were used in this study. they were divided into five groups (a, b, c, d and e). each group contained 6 animals. hepatotoxicity was induced by isoniazid and rifampicin. they were followed for 14 days with no mortality in any group. body weight was measured weekly to adjust the dose of the drugs. effect of ajwa date on serum alt, ast, alp and bilirubin levels is presented as figure 1,2 3, and 4 respectively. table 1 shows mean 60 phoenix dactylifera (ajwa date) whole fruit, flesh & powdered seed prevents anti-tuberculous drug… difference in values between day 0 and 14 with pvalue of all the groups. at baseline values were remarkably similar in all groups. after 14 days values were very significantly higher in att group (group b) as compared to all other groups. difference among the treatment groups themselves as well as with the normal control group was nonsignificant. when we compared the rise in alt level at the end of the study from baseline it was significant in groups b, d and e. the rise was four times higher in group b and less than twice to the baseline in groups d and e. when we compared the rise in ast and alp levels at the end of the study from baseline, they were significant in groups b, c, d and e. the rise was three times higher in group b and less than one and half times in group c, d and e and numerically lowest in group c receiving whole ajwa date. when we compared the rise in serum bilirubin levels at the end of the study from baseline it was significant in group b only. in group a (normal control) the percentage increase in serum alt was 0.4%, serum ast was 1.2%, serum alp was 0.4% and serum bilirubin was 0%. in group b (anti-tuberculous drugs) the percentage increase in serum alt was 295%, serum ast was 217%, serum alp was 184.4% and serum bilirubin was 380%. in group c (att + whole ajwa date) the percentage increase in serum alt was 10.3%, serum ast was 23.7%, serum alp was 14% and serum bilirubin was 25%. in group d (att + ajwa date flesh) the percentage increase in serum alt was 87%, serum ast was 47.4%, serum alp was 35% and serum bilirubin was 0%. in group e (att + ajwa date seed powder) the percentage increase in serum alt was 75%, serum ast was 39.6%, serum alp was 22.1% and serum bilirubin was 7.6%. fig-1: effect of ajwa date on alt (mean ± sd) of att induced hepatotoxic rabbits (n=6) group a: normal control, group b: anti-tuberculous drugs (att), group c: att + whole ajwa date, group d: att + ajwa date flesh, group e: att + ajwa date seed powder ***p value < 0.001 vs all other groups fig-2: effect of ajwa date on ast (mean ± sd) of att induced hepatotoxic rabbits (n=6) group a: normal control, group b: anti-tuberculous drugs (att), group c: att + whole ajwa date, group d: att + ajwa date flesh, group e: att + ajwa date seed powder ***p value < 0.001 vs all other groups fig-3: effect of ajwa date on alp (mean ± sd) of att induced hepatotoxic rabbits (n=6) group a: normal control, group b: anti-tuberculous drugs (att), group c: att + whole ajwa date, group d: att + ajwa date flesh, group e: att + ajwa date seed powder ***p value < 0.001 vs all other groups fig-4: effect of ajwa date on serum bilirubin level (mean ± sd) of att induced hepatotoxic rabbits (n=6) group a: normal control, group b: anti-tuberculous drugs (att), group c: att + whole ajwa date, group d: att + ajwa date flesh, group e: att + ajwa date seed powder ***p value < 0.001 vs all other groups 61 phoenix dactylifera (ajwa date) whole fruit, flesh & powdered seed prevents anti-tuberculous drug… groups serum alt serum ast serum alp serum bilirubin difference (mean ± sd) p value (ttest) difference (mean ± sd) p value (ttest) difference (mean ± sd) p value (ttest) difference (mean±sd) p value (ttest) group a 0.2 ± 0.7 0.611 0.5 ± 0.8 <0.203 0.3±0.2 0.363 0.00 1.000 group b 149.5 ± 20.7 0.001 95.2 ± 18.7 < 0.001 149.8±25.5 < 0.001 0.38±0.02 < 0.001 group c 5.2 ± 5.8 0.083 11.3 ± 3.4 < 0.001 11.3±3.5 0.003 0.03±0.00 0.203 group d 41.8 ± 4.9 0.002 23.7 ± 6.1 <0.003 28.8±4.8 0.002 0.00 1.000 group e 36.8 ± 6.3 0.002 16.3 ± 4.4 <0.001 16.8±3.8 0.010 0.01 0.363 table-1: mean ± sd of difference in serum alt, ast, alp (iu/l) and bilirubin (mg/dl) level between day 0 and day 14 (n=6) group a: normal control, group b: anti-tuberculous drugs (att), group c: att + whole ajwa date, group d: att + ajwa date flesh, group e: att + ajwa date seed powder discussion anti-tubercular therapy is notorious for its hepatotoxic side effects. hepatotoxic effects remain the main reason for cessation of att and many complications that arise in result. it is known that anti-tubercular drugs like rifampicin and isoniazid damage the functional and structural integrity of liver by inducing oxidative stress, as indicated by increased levels of ast, alt, alp and bilirubin. a study suggested that rifampicin causes liver injury by inducing oxidative stress, elevated toxic metabolites caused by cyp induction and upregulation of ppar gamma pathway.17 studies have also shown that metabolism of rifampicin and isoniazid dangerously deplete the glutathione reservoirs in liver and thus pave the path for lipid peroxidation, which ultimately leads to dili.18 taking this finding into consideration, it was hypothesized that ajwa date can be put to use as a hepatoprotective agent on part of its antioxidant properties based on high phenolic and flavonoid content and dpph scavenging activity, as compared to other varieties of dates19. to observe the hepatoprotective effect of ajwa date fruit, an experimental study was designed based on rabbit model. thirty rabbits were divided into 5 groups (a, b, c, d and e) of 6 rabbits randomly assorted into each group. administration of isoniazid and rifampicin raised level of liver enzymes and bilirubin three to four times above baseline (p value < 0.001). similar results were seen in an identical study on rabbits using same dosage of inh and rifampicin.16 the results of this experiment manifested that ajwa date (as whole, flesh and seed powder) was able to significantly bring down the elevated levels of ast, alt and alp induced by anti-tubercular therapy in groups c, d and e receiving ajwa date. whole date, flesh and seed have shown statistically similar benefit. these results are in line with our hypothesis that ajwa date fruit is able to counter the att induced hepatotoxicity. the hepatoprotective effect of ajwa date against various hepatotoxic compounds in different animal models has been demonstrated in several studies. shamim et al. found that ajwa date fruit extract (adfe) is protective against monosodium glutamate induced oxidative stress.20 other studies also found hepatoprotective effects of adfe against liver toxicity induced by ccl4,21 ochratoxin a22 and diclofenac.23 one of these studies estimated the levels of liver enzymes as well as found higher hepatic reduced glutathione content and raised serum levels of superoxide dismutase, catalase and glutathione-s-transferase, after ajwa extract treatment of hepatotoxic rats, indicating antioxidant potential of ajwa date extracts22. the extract of ajwa date fruit was used in previous studies, our results build on existing evidence that the whole date as well as the flesh and seed powder are also able to exhibit the same nature of results as adfe. the hepatotoxicity that emerges as a result of antitubercular drug intake severely hampers the goals of treatment as treatment needs to be stopped in order to avoid further complication. our findings signify that this adverse effect can be countered by adding naturally occurring ajwa date to the diets of patients receiving the treatment without adding any other drug to the regimen, which may overwhelm the liver already striving drug induced oxidative stress. as ajwa seed powder has also shown promising results, it may be safely used in diabetics, being low in sugar content as compared to flesh.24 findings of this study were limited by estimation of liver enzymes only. estimation of markers of oxidative stress would have put light on targets of action, as antioxidant property of ajwa date was basis of hypothesis of this study. also, histopathological studies alongside biochemical tests, would have given a deeper insight and meaning to the results. the practical implication of this study faces a difficulty in adoption as ajwa date being exotic to pakistan, is an expensive fruit which is affordable to a few in the community. however, alternative date fruits of the same family of palms 62 phoenix dactylifera (ajwa date) whole fruit, flesh & powdered seed prevents anti-tuberculous drug… more affordable to the general population can be studied for the same hepatoprotective effects. conclusion ajwa date flesh and seed powder are effective in preventing the rise in liver enzymes induced by isoniazid and rifampicin administration to rabbits. whole fruit being only numerically superior to flesh or seed powder. acknowledgement: gratitude to pgmi, lahore for providing financial and technical support. references 1. global tuberculosis report 2019. who/cds/tb/ 2019.15. [internet]. who. 2019. p. 2020–2. available from: www.who.int/tb/data 2. macneil a, glaziou p, sismanidis c, date a, maloney s, floyd k. global epidemiology of tuberculosis and progress toward meeting global targets — worldwide, 2018. mmwr morb mortal wkly rep. 2020; 69(11):281-285. 3. chakaya j, khan m, ntoumi f, aklillu e, fatima r, mwaba p et al. global tuberculosis report 2020 – reflections on the global tb burden, treatment and prevention efforts. int j of infect dis. 2021; doi: 10.1016/j.ijid.2021.02.107. 4. nahid p, dorman s, alipanah n, barry p, brozek j, cattamanchi a et al. executive summary: official american thoracic society/centers for disease control and prevention/infectious diseases society of america clinical practice guidelines: treatment of drug-susceptible tuberculosis. clininfect dis. 2016; 63(7):853-867. 5. fisher k, vuppalanchi r, saxena r. drug-induced liver injury. arch pathol& lab med. 2015; 139(7): 876-887. 6. wang n, huang y, lin m, huang b, perng c, lin h. chronic hepatitis b infection and risk of antituberculosis drug-induced liver injury: systematic review and meta-analysis. j chin med associat. 2016; 79(7):368-374. 7. holt m, ju c. drug-induced liver injury. handb exp pharmacol. 2010; (196):3-27. 8. el hamdouni m, ahid s, bourkadi j, benamor j, hassar m, cherrah y. incidence of adverse reactions caused by first-line anti-tuberculosis drugs and treatment outcome of pulmonary tuberculosis patients in morocco. infection. 2019; 48(1):43-50. 9. biswas a, santra s, bishnu d, dhali g, chowdhury a, santra a. isoniazid and rifampicin produce hepatic fibrosis through an oxidative stressdependent mechanism. intj hepatol.2020;2020:1-12 10. wardhan h, singh s, singh v. a study of the oxidative stress and the role of antioxidants in att induced hepatotoxicity in tuberculosis patients. indian j public heal res dev. 2016; 7(2):243–249. 11. sheikh by, elsaed wm, samman ah, sheikh by, bin ladin a-mma. ajwa dates as a protective agent against liver toxicity in rat. 2014; 3(2): 358-368. 12. ali a, abdu s. antioxidant protection against pathological mycotoxins alterations on proximal tubules in rat kidney. func food health dis. 2011; 1(4):118. 13. khan f, ahmed f, pushparaj p, abuzenadah a, kumosani t, barbour e, et al. ajwa date (phoenix dactylifera l.) extract inhibits human breast adenocarcinoma (mcf7) cells in vitro by inducing apoptosis and cell cycle arrest. plos one. 2016; 11(7): e0158963. 14. zhang c, aldosari s, vidyasagar p, nair k, nair m. antioxidant and anti-inflammatory assays confirm bioactive compounds in ajwa date fruit. j agric food chem. 2013; 61(24):5834-5840. 15. abdu s. b. the protective role of ajwa date against the hepatotoxicity induced by ochratoxin a. egyp j natural toxins, 2011; 8(1,2): 1-15. 16. kalra bs, aggarwal s, khurana n, gupta u. effect of cimetidine on hepatotoxicity induced by isoniazidrifampicin combination in rabbits. indian j gastroenterol. 2007 jan-feb; 26(1):18-21. 17. kim j, nam w, kim s, kwon o, seung e, jo j, et al. mechanism investigation of rifampicin-induced liver injury using comparative toxicoproteomics in mice. int j mol sci. 2017; 18(7):1417. 18. pan y, tang p, cao j, song q, zhu l, ma s, et al. lipid peroxidation aggravates anti-tuberculosis druginduced liver injury: evidence of ferroptosis induction. biochem biophys res commun. 2020; 533(4):1512-1518. 19. khalid s, ahmad a, kaleem m. antioxidant activity and phenolic contents of ajwa date and their effect on lipo-protein profile. funct foods heal dis. 2017; 7(6):396-410. 20. shamim r, iqbal z, tariq a, zulfiqar s, ara n, khan f. protective effects of phoenix dactylifera on monosodium glutamate induced histomorphological changes in hepatocytes of adult male albino rats: a light microscopic study. med forum mon. 2018; 31(1):87-90. 21. elsadek b, el-sayed es, mansour a, elazab a. abrogation of carbon tetrachloride-induced hepatotoxicity in sprague-dawley rats by ajwa date fruit extract through ameliorating oxidative stress and apoptosis. pak j pharm sci. 2017;30(6):2183-91. 22. al-khafaf a, jwad s. study for the preventive effects of ajwadates alcoholic extract on thehepatic tissue functions and lipid profileregression induced by ochratoxin a in male albino rats. int j pharm res. 2018; 10(3):391-398. 23. aljuhani n, elkablawy m, elbadawy h, alahmadi a, aloufi a, farsi s, et al. protective effects of ajwa date extract against tissue damage induced by acute diclofenac toxicity. j taibah univ med sci. 2019; 14(6):553-559. 63 phoenix dactylifera (ajwa date) whole fruit, flesh & powdered seed prevents anti-tuberculous drug… 24. khalid s, ahmad a, masud t, asad mj, sandhu m. nutritional assessment of ajwa date flesh and pits in comparison to local varieties. j anim plant sci. 2016; 26(4):1072-1080. the authors: dr. sadia majeed assistant professor, department of pharmacology, continental medical college, lahore. dr. usman aslam senior demonstrator, department of pharmacology, al-aleem medical college, lahore. muhammad ubaid ullah khan final year student, allama iqbal medical college, lahore. dr. asma inam associate professor, department of pharmacology, azra naheed medical college, lahore. prof. sadia chiragh head, department of pharmacology, al-aleem medical college, lahore. corresponding author: dr. usman aslam senior demonstrator, department of pharmacology, al-aleem medical college, lahore. e-mail: dr.usmanaslam@yahoo.com 44 proceedings s.z.m.c. vol: 35(3): pp. 44-49, 2021. pszmc-807-35-3-2021 comparative effects of cane sugar, honey & jaggery on plasma glucose level & body weight of alloxan induced diabetic rats 1ambreen anjum, 2hamid javed qureshi, 3saima tabassum, 1wardah anwar, 4rida shakil, 5maria anwar, 6muhammad hashim ghouri 1department of physiology, al-aleem medical college, lahore 2department of physiology, akhtar saeed medical college, lahore 3department of physiology, niazi medical & dental college, sargodha 4pakistan institute of medical sciences, islamabad 5department of physiology, independent medical college, faisalabad 6institute of physiotherapy, gulab devi hospital, lahore abstract introduction: diabetes mellitus is characterized by hyperglycemia with impaired metabolism of carbohydrates, fats, and proteins resulting from insufficiency of secretion or action of insulin. the interplay of genes and environmental factors is believed to stress glucose homeostasis. simple dietary and lifestyle modifications have a role in prevention and slow progression of the disease. the choice and role of sweeteners (sugars) such as ordinary cane sugar, honey, and jaggery have always been a difficult decision for patients with type 2 diabetes. aims & objectives: evaluation and comparison of the effects of cane sugar, honey, and jaggery on plasma glucose in the sera and body weights of alloxan-induced male wistar rats with diabetes. place and duration of study: the study was conducted at the animal house of services institute of medical sciences lahore for a duration of 20 months approximately. material & methods: in this experimental study 120 male wistar (albino) rats were randomly allocated into 4 groups of 30 each. after acclimatization, freshly prepared 2% alloxan monohydrate was administered to all the rats via intraperitoneal injection in a single dose of 120mg/kg body weight. blood sample via tail tipping method was analyzed for fasting serum glucose at 4th post-diabetes induction day. rodents having ≥ 200mg/dl serum glucose were regarded as type 2 diabetes models for further study. all animals had access to usual pelleted diet ad libitum. diabetic controls in group i were fed with equivalent amount of distilled water. group ii, iii and iv rats were fed freshly prepared sweetener containing cane sugar, honey and jaggery solutions respectively in a dose of 1g/kg body weight dissolved in 2ml distilled water on pre-scheduled time of the day for 28 days via oral syringe-dosing. final serum glucose was estimated at 29th day of study by terminal blood sampling and final body weights were recorded. data analysis was performed using spss version 20. results: group iii (honey fed) rats had least hyperglycemia of 34.53±77.31mg/dl, followed by group ii (cane sugar fed) rats (79.31±73.03 mg/dl) then group i diabetic control rats (88.11±92.56mg/dl) and maximal rise in jaggery fed rats with value of 134.50 mg/dl. significant reduction (p=0.049) in body weight (-40.50g) was noted in group ii rats whereas maximum weight gain occurred in group i (127.57 ±72.475), than those of group iv rats (69.13 ± 92.88) followed by ii rats (38.42±89.43). conclusion: the result demonstrates that honey in type 2 diabetes mellitus possess antidiabetic properties via minimum hyperglycemic and weight gaining effects than processed cane sugar. as it was presumed, minimally processed jaggery has not shown significant beneficial effects on serum glucose and body weight. owing to the advantageous effects on metabolic profile, honey may be regarded as a relatively safer sweetener in diabetics. key words: alloxan induced diabetes, hyperglycemia, type 2 diabetes mellitus, diabetes mellitus,cane sugar,honey,jaggery introduction diabetes mellitus (dm) describes a group of chronic progressive disorders of the metabolic system characterized by high glucose levels, with impaired metabolism of carbohydrates (cho), fats, and proteins resulting from insufficiency of secretion or action of endogenous insulin.1 in this heterogeneous syndrome genes and environment interact, eventually stressing glucose homeostasis. dietary choices have a decisive role in the expression of genetic susceptibility to diabetes.2 worldwide diabetes epidemic has been associated 45 comparative effects of cane sugar, honey & jaggery on plasma glucose level & body weight… with the way of life-related risk factors such as obesity, sedentary lifestyle, and low-quality dietary choices.3 neuroscientists scientifically established the notion of innate preference of sweet taste for humans since antiquity. sugars, utilized as sweeteners are an important ingredient of human food but there has been ample debate regarding their contribution to metabolic syndrome and diabetes.4 evidence shows that extensive utilization of sweeteners is robustly related to a variety of interrelated negative effects and development of type 2 dm. reduced insulin sensitivity has been linked with excessive use of common table sugar (sucrose) and different artificial sweeteners. considerable research has displayed that majority of type 2 dm cases can be prevented through simple dietary and lifestyle modifications.5 nutritional recommendations by diabetes associations have emphasized to attain individualized glycemic, blood pressure, and lipimeic control along with achievement and maintenance of body weight together with delay or prevention of diabetic complications. it also persuades people with type 2 dm to abate their energy intake while maintaining a healthy eating pattern to encourage weight loss and improved glycemic control. it has been acknowledged that judicious usage of sweeteners in modest amounts is acceptable especially those which minimally derange metabolic control and body weight.6 one of the most exploited sweetening agent by mankind; white odorless crystalline processed table sugar has been linked closely to the mounting epidemic of obesity, dm, metabolic syndrome, dyslipidemia, fatty liver, and various other related negative health effects.7 clinical researchers are continuously struggling to explore other sugar alternatives possessing less harmful impact on overall human health. so, currently, there is a resurgence of scientific interest in natural medicinal sweeteners such as honey, by researchers, the medical community, and even the general public.8 a relatively safer traditional sweetening agent, honey offers incredible antiseptic, antioxidant, antimicrobial, antiplatelet, antinociceptive potentials, cariogenic protection, and immune-boosting properties for the goodness of human health.9 besides the dominant sugar content, honey contains the modest amounts of more than 180 organic and inorganic constituents, vitamins, minerals, proteins and amino acids. it also contains trivial amounts of antioxidants, enzymes, organic acids which impart beneficial health effects to this sweetener.10 before the popularity of refined sugar and other sweeteners, jaggery (derived from sugar cane) was considered one of the dominant forms of cane sugar utilization but today it is just a marginal food on the world market, although still significant in some of the producing countries including pakistan and india (south asia).11 it is a natural mixture of sugar and molasses. if pure clarified sugar cane juice is boiled, what is left is solid jaggery possessing sucrose (6585%), whereas white sugar contains purely sucrose (99.7%).12 this traditional low priced minimally processed sweetener has been claimed to have health and environmental advantages in comparison with refined cane sugar. although scarcity of high quality scientific research exists, yet few studies point out significant positive health effects of jaggery. the impact of jaggery and honey on metabolic variables has shown conflicting results.13,14 presumed anti-diabetic effects of these two sweeteners are much complicated and have not been convincingly compared with the ordinary table sugar. on these grounds, the objective of this investigation was to determine and associate the effects of honey, cane sugar plus jaggery on plasma glucose levels and body weights in alloxan-induced male albino diabetic rats with the purpose to identify the safest sweetening agent for patients with diabetes. material and methods study design and setting: this experimental study was conducted at department of physiology, services institute of medical sciences (sims), lahore pakistan from march 2013 – november 2015. ethical considerations: the study protocol and procedures had the approval from the research advisory / evaluation committee of college of physicians and surgeons (cpsp), for basic medical sciences islamabad. animal research: reporting of in vitro experiments (arrive) guidelines were followed for the conduct of research on lab animals. animals: 12–14 weeks old 120 male albino rats of wistar strain obtained from the national institute of health, islamabad were housed in polypropylene cages at the animal house of services institute of medical sciences, lahore pakistan. housing conditions were thermostatically maintained at controlled ambient temperature 26±2oc and an aerated animal room with a 12 hours’ dark and light cycle. commercially available standard pelleted food ad libitum and tap water in clean bottles were fed to the animals. rats were selected as per following criteria: 46 comparative effects of cane sugar, honey & jaggery on plasma glucose level & body weight… inclusion criteria: healthy, adult male albino rats, weighing 200-300grams. exclusion criteria: rats who were inactive or with some disease (symptomatic illness) or anomaly. rats that didn’t develop diabetes after receiving alloxan injection. randomization: rats were selected through initial purposive sampling and randomly grouped into 4 groups of 30 rats each using a random number table. animals were placed in their respective cages with mentioned group name and identification tags. procedures and interventions: for one week these rats had access to a normal diet before the commencement of the experiment allowing them to be acclimatized. after seven days, rats fed on a normal diet were deprived of food overnight (for about 12 hours) and were weighed, manually restrained, and then anesthetized with chloroform in a glass dome. freshly prepared 2% alloxan monohydrate, as a sole dose of 120mg/kg body weight of individual rat was administered through intraperitoneal injection to all the rats of group i, ii, iii, and iv.15 rats with serum glucose ≥200mg/dl were regarded as alloxan-induced rats with diabetes and used as type 2 diabetes models for further study. confirmation of dm was established by estimating fasting plasma glucose at 4th postdiabetes induction (about after 96hours) employing a portable glucometer via tail tipping method.16 these blood hyperglycemic values by which diabetes in rats was confirmed were recorded as “initial serum glucose” for all diabetic animals in each group respectively. according to the type of feeding/supplementation of sweetener solutions, groups were named as group i diabetic control (only distilled water fed), group ii (cane sugar), group iii (honey) and group iv (jaggery) respectively. three different sweetener solutions were prepared daily from day 1 of experimental study to 28th day, freshly by gentle and thorough mixing of the respective sweetener in 2ml distilled water. each experimental animal was fed with the respective sweetener solution in a dose 1g/kg body weight on pre-scheduled time of the day for total 28 days of the study period via oral syringe dosing method. whereas rats in group i, which served as diabetic controls were given equivalent amount of distilled water. fig-1: rat readily drinking sweetener solution (1 gm/kg) by the oral syringe-dosing statistical analysis: the data were evaluated using ibm (international business machines) statistical package for social sciences (spss), version 20. the normality of data was assessed using the shapiro-wilk test. the level of significance was set at p < 0.05. for arithmatical calculations of quantitative variables, mean ± sd (standard deviation) was presented for variables that were distributed normally, and median with iqr (interquartile ratio) were calculated for variables non-normally distributed. for normally distributed variables, parametric tests were used for inferential statistics. one-way anova was applied to observe group mean differences of quantitative variables and post hoc tukey’s test was applied to see which group mean differs. for non-normally-distributed variables, non-parametric tests were applied. kruskal wallis test was applied to observe the group mean differences of quantitative variables and the mann whitney-u test was applied post hoc. repeated measure analysis of body weight and serum glucose was carried out by using paired samples t-test for normally distributed data and wilcoxon matched-pairs signed-ranks test for non-normally distributed data. results effects of sweeteners i.e. cane sugar, honey, and jaggery were evaluated through changing serum glucose levels and body weights in alloxan-induced rats with diabetes. the difference in change of serum glucose after 28 days was highly significant (p=0.001) between the groups. rise in serum glucose at the termination of study was maximally seen in rats of group iv (jaggery) as 134.50 mg/dl (84.25-186.75), followed by group i (diabetic 47 comparative effects of cane sugar, honey & jaggery on plasma glucose level & body weight… control) rats (88.11±92.56mg/dl) and then in those on cane sugar supplementation (79.31±73.03 mg/dl) while least rise in levels of serum glucose was observed in the rats belonging to group iii (honey) (34.53±77.31mg/dl) (fig-2). fig-2: comparison of change in serum glucose after 28 days of supplementation in the groups, i (diabetic control), ii (cane sugar), iii (honey), and iv (jaggery). avalues are given as mean ± sd for normally distributed variables and median (iqr) for non-normally distributed variables. b p-value is generated by paired samples t-test. cp-value is generated by wilcoxon matched-pairs signed-ranks test. (p=0.001 is considered highly significant and p˂0.05 is considered significant). fig-3: comparison of change in body weight after 28 days of supplementation in the groups, i (diabetic control), ii (cane sugar), iii (honey), and iv (jaggery). avalues are given as mean ± sd for normally distributed variables and median (iqr) for non-normally distributed variables. b p-value is generated by paired samples t-test. c p-value is generated by wilcoxon matched-pairs signed-ranks test. (p=0.001 is considered highly significant and p˂0.05 is considered significant). the initial and final weights of rats significantly (p<0.001) differed in four groups at the end. among the total 120 type 2 dm rats, in 28 surviving rats of group i, at the 29th experimental day, there was highly significant (p=0.001) augmented body weight. {final body weight– initial body weight (grams)}. similarly, this highly significant increased body weight trend was observed in the 30 rats of group iv with a p-value of 0.001, but only a significant increase of weight was observed in 26 surviving rats of group ii with a p-value of 0.040. on the contrary, bodyweight of rats in group iii was significantly (p=0.049) reduced with a difference in their final body weight – initial body weight as -40.50g (-94.50-27.75) (fig-3). the control group exhibited a maximum bodyweight gain, followed by the rats on jaggery supplementation and lastly by those given cane sugar. contrary to this, honey fed rats had shown significantly reduced the body weight. discussion our study evaluated the effects of various categories of sweeteners in optimal dose on serum glucose and body weight levels in alloxan induced male diabetic rats. among these; artificial cane sugar (sucrose), minimally processed jaggery and naturally occurring honey were fed to rats in their respective groups and effects on mentioned parameters were studied compared with the control group. minimum rise in levels of serum glucose were observed in the rats belonging to group iii (honey) in our study which are well supported by the demonstration of results by al-waili in patients with diabetes when compared with the effects of dextrose and sucrose along with greater elevation of insulin.17 our study has not explored the mechanisms by which honey consumption caused mentioned hypoglycemic effects in type 2 dm rats. even the current scientific literature could not fully explain the mechanisms by which honey reduces hyperglycemia, weight gain, and other anti-diabetic effects. however, some of the recent studies suggest that the blend of various carbohydrates particularly fructose and oligosaccharides might be the contributory factors. honey rich in fructose, a naturally occurring sweetener enhances glycemic control much better than sucrose. it has a glycemic index of 19, compared to glucose which has 100 and sucrose (refined sugar) with 60. it suggests that glucose-lowering or antidiabetic properties of honey might be attributed to diminished rate of intestinal absorption, extended gastric emptying duration, and less food intake.18, 19 48 comparative effects of cane sugar, honey & jaggery on plasma glucose level & body weight… significantly differed weight changes among the four groups were observed after 28 days of experimentation. highest weight gain in diabetic control rats could be due to the polyphagia and increased consumption of the rat chow due to diabetes. this assumption is supported by the evidence of altered balance of hypothalamic neuropeptides, intraneuronal signaling mechanisms in insulin deficiency states.20 results of the current study illustrating positive impact of honey on lowering body weight is consistent with the findings by chepulis and starkey in a long term feeding trial who compared the effect of honey against commonly employed sucrose on diabetic rats.21 although it was hypothesized by us that consumption of minimally processed sugar of jaggery would cause relatively less elevation of plasma glucose and minimal weight changes in comparison with the artificial cane sugar but on the contrary significant hyperglycemia and weight gain were observed by the animals in this group. this could be attributed to high glycemic index of jaggery compared with other sweeteners.22 in the literature, diverse and contradictory reports are available regarding the metabolic effects of jaggery (and other sugarcane-based minimally processed sugars) in individuals with diabetes. in a closely related remote study, brekhman and his coresearchers explored the influence of varying amounts of refined white sugar and (organic) brown sugar from common precursor of sugar on certain indicators of cho and fat metabolism lasting for about 90 days. the results of this study about the comparative effects of dietary sucrose and muscovado unrefined sugar (very much similar to jaggery) showed minimal derangement in glucose and body weight in the high dose brown sugar fed group compared with sucrose fed rats, suggestive of a potential beneficial nutritive effect of brown sugar (jaggery) on human metabolic health.23 both these contrary studies are quite old and this motivated us to explore and validate the results of experimentation taking honey in account as well. one probable justification for our failure to corroborate the nutritional work of brekhman might be adoption of unconventional ways of sugar administration as well as different approach for diet formulation. limitations and recommendations: the results of present animal based study could be extrapolated to type 2 diabetic human individuals in a meaningful way, if the number of subjects would have been more. additionally, the promising effects of honey seen in the present experimental study need to be further investigated in well designed, controlled clinical studies to determine whether these can be duplicated in actual clinical situations. thus, clinical trials with larger patient populations are needed to replicate the results of this study. moreover, the chemical characterization (both quantitative and qualitative) of sweeteners used in this research work was not sought out due to technical and financial restrains. this possibly might have given the logical inkling of active constituents of a particular sweetener and its role in type 2 dm in the study subjects. this study underscores the need for additional studies to elucidate the mechanisms by which honey improves the metabolic control in type 2 dm. the promising effects of honey seen in the present experimental study need to be further investigated in well designed, controlled clinical studies on human populations to determine whether these can be duplicated in actual clinical situations. furthermore, studies that unravel the true long term effect of minimally processed jaggery in context of dm are recommended. conclusion the findings of this study validate that natural sweetener i.e. (group iii) honey in type 2 dm has the least weight gaining and hyperglycemic effects than chemically processed cane sugar and jaggery. whereas the minimally processed jaggery exhibited considerable hyperglycemia and weight gain as compared to the highly processed table sugar. it suggests the beneficial effects of honey on the overall metabolic profile of subjects with diabetes. references 1. american diabetes association. classification and diagnosis of diabetes mellitus. diabetes care 2017; 40 (suppl 1):s11–s24. 2. ericson u, hindy g, drake i, schulz ca, brunkwall l, hellstrand s, almgren p, orho-melander m. dietary and genetic risk scores and incidence of type 2 diabetes. genes & nutrition. 2018; 13:13. 3. romao i, roth j. genetic and environmental interactions in obesity and type 2 diabetes. j am diet assoc. 2008; 108(4):s24–s28. 4. toop cr, gentili s. fructose beverage consumption induces a metabolic syndrome phenotype in the rat: a systematic review and meta-analysis. nutrients 2016; 8, 577. 5. chong s, ding d, byun r, comino e, bauman a, jalaludin b. lifestyle changes after a diagnosis of type 2 diabetes. diabetes spectr. 2017 feb; 30(1): 43-50. 6. evert ab, boucher jl, cypress m, dunbar sa, 49 comparative effects of cane sugar, honey & jaggery on plasma glucose level & body weight… franz mj, mayer-davis ej, neumiller jj, nwankwo, verdi cl, urbanski p, yancy ws. position statement: nutrition therapy recommendations for the management of adults with diabetes. diabetes care 2014; 36: 3821-42. 7. laverty aa, magee l, monteiro ca, saxena s, millett c. sugar and artificially sweetened beverage consumption and adiposity changes: national longitudinal study. int j behav nutr phys act. 2015 oct 26; 12:137. 8. bobiş o, dezmirean ds, moise ar. honey and diabetes: the importance of natural simple sugars in diet for preventing and treating different type of diabetes. oxid med cell longev. 2018; 4757893. 9. meoa sa, al-asiri sa, mahesa al, ansari mj. role of honey in modern medicine. saudi journal of biological sciences. 2017 july; volume 24:5,975-8. 10. cortés me, vigil p, montenegro g. the medicinal value of honey: a review on its benefits to human health, with a special focus on its effects on glycemic regulation. cien inv agr 2011, 38: 303-17. 11. walter r. jaffe. health effects of non-centrifugal sugar (ncs): a review. sugar tech 2012; 14:87-94. 12. waheed s, ahmad s. instrumental neutron activation analysis of different products from the sugarcane industry in pakistan – part 1: essential elements for nutritional adequacy. aoac 2008; 2: 392-9. 13. galvez l, kwon yi, genovese mi, lajolo fm, shetty k. antidiabetes and antihypertension potential of commonly consumed carbohydrate sweeteners using in vitro models. journal of medicinal food 2008; 11: 337-48. 14. okabe t, toda t, inafuku m, wada k, iwasaki h, oku h. antiatherosclerotic functions of kokuto, okinawan noncentrifuged cane sugar. journal of agriculture and food chemistry 2009; 57: 69–75. 15. jain s, bhatia g, barik r, kumar p, jain s, dixit vk. antidiabetic activity of paspalum scrobiculatum linn in alloxan induced diabetic rats. j ethnopharmacol. 2010; 127: 325-8. 16. guidelines for rodent blood withdrawal and tail biopsy. the univeristy of north carolina at chapel hill. 2002. 17. al-waili ns. natural honey lowers plasma glucose, c-reactive protein, homocysteine, and blood lipids in healthy, diabetic, and hyperlipidemic subjects: comparison with dextrose and sucrose. journal of medicinal food. 2004 apr 1; 7(1):100-7. 18. erejuwa oo, sulaiman sa, wahab ms. oligosaccharides might contribute to the antidiabetic effect of honey: a review of the literature. molecules. 2012 jan; 17(1):248-66. 19. bobiş o, dezmirean ds, moise ar. honey and diabetes: the importance of natural simple sugars in diet for preventing and treating different type of diabetes. oxid med cell longev. 2018 feb 4; 4757893. 20. namkoong c, kim ms, jang pg, han sm, park hs, koh eh, lee wj, kim jy, park is, park jy, lee ku. enhanced hypothalamic amp-activated protein kinase activity contributes to hyperphagia in diabetic rats. diabetes. 2005; 54:63-8. 21. chepulis l m, starkey n. the long-term effects of feeding honey compared with sucrose and a sugarfree diet on weight gain, lipid profiles, and dexa measurements in rats. journal of food science 2008; 73: 1-7. 22. uma p, hariharan rs, ramani v, seshiah v. glycaemic indices of different sugars. int. j. diab. dev. ctries. 1987; 7: 78-82. 23. brekhman i i, nesterenko i f. brown sugar and health. oxford: pergarnon press, oxford, new york, toronto, sidney, paris, frankfurt. 1983. the authors: dr. ambreen anjum assistant professor, department of physiology, al aleem medical college, lahore. prof. hamid javed qureshi department of physiology, akhtar saeed medical college, lahore. dr. saima tabassum assistant professor, department of physiology, niazi medical & dental college, (sargodha) dr. wardah anwar assistant professor department of physiology, al aleem medical college, lahore. dr. rida shakil medical officer, pakistan institute of medical sciences, islamabad. dr. maria anwar assistant professor, department of physiology, independent medical college, faisalabad. muhammad hashim ghouri iastm institute of physiotherapy, gulab devi hospital, lahore. president, pakistan research and welfare society corresponding author: dr. ambreen anjum assistant professor, department of physiology, al aleem medical college, lahore. e-mail: draanjum28@yahoo.com 24 proceedings s.z.m.c. vol: 35(3): pp. 24-30, 2021. pszmc-804-35-3-2021 predictors of mortality in necrotizing soft tissue infections in adults – experience at a tertiary care hospital in pakistan 1haroon javaid majid, 2muhammad imran anwar, 3muhammad zaeem khalid, 3shafique-urrehman, 2muhammad amir jameel 1department of general surgery & surgical oncology, shaikh zayed medical complex, lahore 2department of surgery (unit-ii), shaikh zayed medical complex, lahore 3department of surgery (unit-i), shaikh zayed medical complex, lahore abstract introduction: necrotizing soft tissue infection is a potentially fatal ‘flesh-eating’ disease that requires prompt intervention to save a patient’s life. identification of parameters that indicate worsening of the disease (predictors of mortality) is an important part in management that guides a surgeon towards rapid surgical treatment. aims & objectives: this retrospective study aims to identify factors that are associated with mortality in adults with nstis. this study also aims to propose solutions for the better management of these infection to improve surgical outcome and survival. place and duration of study: opd & emergency of surgical unit i and ii of shaikh zayed hospital, lahore over a period of 10 years between the years 2010 to 2021. material & methods: this is a cross-sectional epidemiological, retrospective study conducted on patients presenting in the outpatients department, as well as the emergency of surgical unit i and ii. a total of 72 patients meeting the inclusion criteria were enrolled in this study. spss version 23 was used for data analysis. results: out of 72 patients, 31.94% of patients were receiving some form of immunosuppressive treatment (n=23), out of which 7 patients lost their life (30.43%, p=0.000). there was a preexisting immunosuppressive disease in 48.61% patients (n=35), and in this subset, a total of 10 patients didn’t recover (28.57%, p=0.000). 13 out of 17 patients who died received first debridement after 12 hours from presentation (p=0.021). high mortality was seen in patients who had the presence of shock at hospital admission. conclusion: delay in diagnosis and surgical treatment, the presence of shock at admission accompanied by evidence of organ dysfunction, increasing comorbidity index are reasons for the higher mortality seen in these patients of nstis. the timing of the first surgery is of the essence thus highlighting the importance of the “golden hour” for the management of nstis. key words: necrotizing soft tissue infections (nstis), early diagnosis, mortality, debridement introduction necrotizing soft tissue infections (nsti) are rapidly progressive skin and soft tissue infections that cause widespread tissue necrosis and are associated with systemic illness and a high mortality rate and remain a challenge not only for the most experienced surgical team but also require full time involvement at the multidisciplinary level if lives are to be saved.1,2 a potentially massively destructive infection, nsti has popularly been named as a 'flesh-eating disease', which if diagnosed or treated late is associated with a grave outcome.3,13 early diagnosis, adequate resuscitation with simultaneous administration of broad spectrum antibiotics followed by prompt surgical intervention forms the basis of improving the outcome in these critically ill patients.1,2,3,4,5,6 nstis are variously classified on the basis of the pathogen, the site of the infection and the extent of tissue involvement.4 the classification system according to microbiology categorizes, type 1 which is the commonest and is defined as the presence of multiple microbes, usually involving anaerobes, while on the other hand, type 2 nsti have only one documented pathogen, most commonly group a streptococcus or less commonly s. aureus. clostridia can also be isolated in type 2 infections.20 type 3 is very rare and caused by vibrio vulnificans (seawater organism), and type 4 i.e., fungal.7,15,16,17 certain monomicrobial infections include communityassociated mrsa. different terms can be used to 25 predictors of mortality in necrotizing soft tissue infections in adults – experience at a tertiary care hospital describe nstis of various anatomic sites, for example necrotizing infection of the perineum is called fournier’s gangrene. diabetic foot infections are usually polymicrobial and can progress to a necrotizing pattern due to the presence of concomitant microvasculature injury along with the infection. the classically described meleney's synergistic gangrene is a rare form of necrotizing bacterial infection of the abdominal wall which can develop following abdominal surgery.18 if not identified early and treated promptly, it can lead to extensive fatal gangrene. last but not the least, nsti are also classified according to the depth of necrosis. ‘necrotizing cellulitis’ is a term used when the infection of the subcutaneous tissue extends into the dermis. ‘necrotizing fasciitis’, as the name suggests, involves the fascia and the term ‘myonecrosis’ or ‘necrotizing myositis’ is used when the infection involves the muscles with or without skin manifestations.4 despite better understanding and advances in management, the fatalities from nstis have remained constant at around 30% for the past thirty years or more.4,5,6 thus, we conducted this retrospective study at our institute to identify factors that were significantly associated with mortality in adults with nstis. it is postulated that mortality can be predicted in patients with nstis due to the presence of certain indicators on hospital admission and during the course of treatment in one individual setup. material and methods this is a cross-sectional, retrospective study done on patients presenting in the outpatients department, as well as the emergency of surgical unit i and ii of shaikh zayed hospital, lahore between the years 2010 to 2021. inclusion & exclusion criteria: adult patients (age >18 years) admitted with necrotizing soft tissue infection (nsti) were included in this study. all patients were ensured for complete records. both males and females were taken into account. four patients who died before surgical intervention could be carried out were excluded from this study because of incomplete records. all patients who had any previous intervention from any setup before presenting at shaikh zayed hospital were also excluded from the study. type i diabetics were not taken into account as there is a lack of evidence of it causing immunosuppression. patients with two or more concomitantly affecting variables (for example cancer patients with diabetes) were also not included in this study. method patients were identified with and diagnosis was confirmed by a detailed review of medical records. standard treatment was given to all patients and included early emergency and later scheduled repeated debridement as required, broad-spectrum antibiotic cover, and physiologic and nutritional support followed by early soft tissue reconstruction by the plastic surgery teams. a total of 72 patients who met the inclusion criteria were taken into account and a detailed review of medical records and outcome was analyzed retrospectively for the indicators of fatalities in patients with necrotizing soft tissue infections. diabetes and immunosuppression: only type-ii diabetics who had diabetes for greater than 10 years were included. recent studies suggest that long term insulin resistance causes microvascular and macrovascular complications and an abnormal t cell response causing immunosuppression23. cancer induced immunosuppression: studies suggest that tumor cells neutralize our host immune response, specifically il18, hence reducing the ability of our immune system to fight off infections24. chronic liver disease: chronic liver disease is an immunocompromised state that predisposes the patient to infections by gram negative bacteria and is associated with 30% mortality. this state is characteristically defined as ‘immune paralysis’ that leads to an increase in anti-inflammatory cytokines and a decrease in pro-inflammatory ones.25 statistical analysis: the authors used statistical package of social sciences version 23 for data analysis and the data compilation and input was carried out by the authors themselves. results the total number of patients 72 (males:41, females:31). mean age: 47 years (range 22 – 71 years). there was evidence of immunosuppressive treatment in 31.94% (n=23) of patients, out of which 7 patients lost their life (30.43%, p=0.000). there was a preexisting immunosuppressive disease in 48.61% patients (n=35), and in this subset, a total of 10 patients didn’t recover (28.57%, p=0.000) while 15.2% patients had no comorbidities. table-1 shows details of the predisposing conditions. although im/iv injection sites were the commonest infectious entry pathways, 8% of the necrotising infections were found to be in post-operative surgical wounds (table-2). 26 predictors of mortality in necrotizing soft tissue infections in adults – experience at a tertiary care hospital patient's condition at presentation total number of patients mortality p values immunosuppressive treatment 23 0.00 corticosteroids 10 4 nsaids 11 2 chemotherapy 2 1 immunosuppressive disease 35 0.00 diabetes mellitus 31 7 cancer 1 1 chronic liver disease 3 2 pvd 3 0 no comorbids 11 1 0.00 table-1: pre-existing conditions infectious entry pathway n= number (% percentage). total patients 72 abscess or wound n= 27 (38%) trauma n = 6 (8%) postoperative infection n = 6 (8%) im or iv injection site n = 33 (46%) table-2: possible source of infection lower limbs were the most affected anatomical site, the trunk/torso was involved in 30.5% of the patients (fig-1). fig-1: (percentages of anatomical sites affected with necrotizing soft tissue infections) high mortality was seen in patients with primary or secondary involvement of the trunk (11 out of 38 patients, mortality 29%), nsti involving postoperative surgical wounds (4 out of 6 patients, mortality 67 %) and those who had clostridial nsti (3 out of 9 patients, mortality 33%). any combinations of these criteria were associated with more than 90% mortality. the 17 patients who died in this group fulfilled at least two or more of these criteria. the mean time from initial presentation to first surgical intervention was calculated as fourteen hours, ranging from 4 42 hours. on average, patients who recovered had a mean of five debridements, ranging from 2 10 debridements. additionally, 23.5% of the total mortalities were operated before 12 hours from the time of admission while 76.5% of the total mortalities were operated after 12 hours or beyond. late diagnosis, delayed presentation and the time from admission to surgical management of more than 12 hours was associated with a higher rate of mortality (p=0.021 – table-3). time of surgical intervention after presentation total mortalities in the subset p value >12 hours 13 0.021 <12 hours 4 table 3: time of first surgical intervention patients who were received in shock and had the following number of patients received total mortalities high white cell count (> 25,000 × 103/μl) 5 1 raised serum creatinine (> 2 mg/dl) 6 1 heart disease 4 2 high white cell count and raised serum creatinine 6 4 high white cell count, raised serum creatinine and heart disease 2 1 total 23 9 table-4: patients received in shock four patients had to undergo amputations (6%) to control the extent of infection. most patients spent a total of 35 days on the surgical floor on average, ranging from 7 60 days, notably spending a mean of 4 days in the intensive care unit (range 2-17days). the mortality rate was calculated to be 24% (17 out of 72 patients included in the study). most of these patients were found to be in shock on initial evaluation and a high white cell count (> 25,000 × 103/μl), raised serum creatinine (> 2 mg/dl) and/or heart disease. nine out of 23 patients meeting these criteria died (39%). mortality was 16.32% in patients who were not received in shock (8/49 patients). details are shown in table-4. 27 predictors of mortality in necrotizing soft tissue infections in adults – experience at a tertiary care hospital culture swabs were taken from each patient included in the study, 61 out of 72 patients had at least one pathogen positive in the reports (85%). multiple microbes were documented in 28 patients (39%) and 11 patients received negative results. (15%). s pyogenes was the most common pathogen isolated from the cultures (n=23; 32%), either alone (n=16), or along with s. aureus (n=7). 39 patients (54%) had type i nsti and 22 (31%) had type ii nsti. bacteraemia was found in 38% of patients. clostridial, mrsa, acinobacter and fungal infections were associated with the highest mortality. discussion diabetes mellitus, immunocompromising conditions, immune-suppressive therapies, vascular disease, and obesity are common comorbidities in patients with nstis which have been pointed out by many authors and our experience showed similar results as there was evidence of immunosuppressive treatment in 31.94% and immunosuppressive disease in 48.61% of our patients.1-7 such patients usually present with signs and symptoms of sepsis and reduced survival is noted specially in patients who do not receive prompt surgical treatment under broad spectrum antibiotic cover and accelerated resuscitation.8,13 however, it is interesting to note that 33% of our patients had absolutely no comorbidities. although increasing comorbidity index is associated with increased mortality by most authors1-7, even in these patients with no comorbidities, delayed diagnosis and surgical treatment leads to significant morbidity and mortality and thus adverse outcomes9,13. thus, a high index of suspicion even in these previously fit patients cannot be over emphasized for a favourable outcome.9,10 undoubtedly, nstis are best treated in large tertiary care centers where all intensive care/ multidisciplinary facilities and experience is available. the effect of transfer to such centers and the resulting delay in treatment has been studied by other authors who have found that transfer status is associated with a higher mortality rate in nstis.11,13 we share a similar observation although our data on this aspect is compromised. we feel that a prospective study is warranted to illuminate further what factors contribute to the higher mortality rate in these very ill patients. although injection sites (im/sc/iv) were the commonest infectious entry pathways (46%) in our series, 8% of the necrotizing infections were found to be in post-operative surgical wounds and were associated with a very high mortality (4 out of 6 patients, mortality 67 %). nstis complicating surgical wounds have been reported by various authors, are difficult to recognize and manage and associated with a significant morbidity and mortality.12,14 the 4 patients with necrotizing postoperative infections in our series who died suffered from delay in diagnosis and/or transfer delays leading to a delay in surgical intervention and thus a poor outcome. three of these patients had been referred from other hospitals and were too unstable for prompt surgical debridement and had to be resuscitated for more than 12 hours before they could be shifted to the operation theatre. two of these had clostridial infections (fig-2 & 3). one of the four patients with post-operative nstis who died included one of our own patients who had undergone an open necrosectomy for infected pancreatic necrosis and in whom the fungal necrotizing surgical site infection was recognized too late (fig-4). this was the case in three other patients among those with compromised immunity and with fungal nstis proven at some stage. fungal nstis are more frequent in patients on immunosuppressive treatments and are associated with significant in-hospital mortality.15 however, early recognition of the problem in a patient unresponsive to repeated serial debridement and broad-spectrum antibiotic cover, empiric treatment with antifungals should be given, particularly in those patients who are on immunosuppressive therapy, as it has shown to decrease mortality in admitted patients with fungal nstis.15,16,17 fig-2: clostridal nstis after an open cholecystectomy in diabetic patient. fig-3: 5 days after appendectomy patient is sepsis. wound opened to reveal extensive necrotizing myositis (clostridial) with near normal overlying skin 28 predictors of mortality in necrotizing soft tissue infections in adults – experience at a tertiary care hospital fig-4: fungal nsti in a patient after pancreatic necrosectomy. a recent landmark systematic review and metaanalysis by nawijn et al showed that although average mortality rates reported for nstis have remained constant (around 20%) over the past 2 decades or more, surgical debridement to fresh healthy bleeding margins at the earliest possible time after admission lowers the mortality rate for nsti by almost 50%. early surgical debridement of nstis within 12 hours is essential for reducing the mortality rate, while surgical intervention within 6 hours even further improve outcomes.13 like most other authors, we share similar findings. our records showed that 23.5% of the total mortalities were operated before 12 hours while 76.5% of the total mortalities were operated after 12 hours or beyond. therefore, timing of the first surgery is of the essence and there indeed does exist a “golden hour” for the management of nstis.11,13 like most other authors our data also shows a mean of 5 debridements for the survivors thus highlighting the importance of timely repeat serial debridements to arrest the spread of the nstis and to ensure improved survival.1-17 twenty three (32%) patients in our series were received in shock with raised white cell counts and evidence of impaired renal function. the mortality among these patients was 39% (9/23 patients) compared to 16.32% mortality in patients who were not received in shock (8/49 patients). so, it is obvious that the mortality was almost doubled in patients who had shock and evidence of organ dysfunction at the time of admission. although our data is lacking, it has been observed that a combination of multi organ dysfunction and comorbidities like heart disease, diabetes, obesity, immune compromise and nsti trunkal involvement. (fig-5a & 5b) either primarily or secondarily led to mortalities even in patients who were not received in shock. these are the findings of most other authors who have written about the subject1-5,7-9,11,13,19. delayed or suboptimal primary or subsequent serial debridements13 and the presence of clostridial,20 acinobacter,21,22 mrsa or fungi15,16,17 as the infecting organisms was related to a higher mortality. overwhelming sepsis was the cause of death in most of our patients except two who died due to major cardiac events during recovery in the hospital. fig-5a: nsti of the torso with superimposed fungal infection secondary to sc insulin injections in a diabetic. notice the blue areas of extension after the third debridement. fig-5b: same patient as in fig-5a. sharp excision to fresh healthy bleeding tissues. severe nstis of the perineum extending onto the limbs and trunk as the patient shown in fig-6a may require faecal diversion in addition to prompt serial debridements. survivors will require the help of the plastic surgeons for early reconstruction as the wounds get better (fig-6b). 29 predictors of mortality in necrotizing soft tissue infections in adults – experience at a tertiary care hospital fig-6a: severe nsti of the perineum involving a gas forming organism secondary to an im nsaid injection in a diabetic extending onto the trunk and right thigh at the second debridement. needed a diverting transverse loop colostomy. fig-6b: the same patient as in fig-6a after 2months and reconstruction. colostomy was reverted after 5 months. a study done at aga khan university hospital karachi showed that necrotizing infections of the head and neck region are fast spreading and are often not diagnosed properly. aggressive strategies for treatment, both surgical and medical are required for proper management but late diagnosis and extension beyond the neck region into the chest carries poor prognosis.26 a case report published in 2020, where a patient with no co-morbids presented at liaqat national hospital, karachi with an acute abdomen and was later diagnosed with retroperitoneal necrotizing fasciitis which is difficult to diagnose and the only option of treatment is prompt surgical intervention.27 another pakistani study suggested that in patients presenting with necrotizing fasciitis of the lower limb, sepsis was the commonest complication (70%), while the most common cause of death in these patients was myocardial infarction and the mortality rate was reported to be around 30%. diabetes mellitis was positive in the history of most patients.28 so a multidisciplinary approach in a tertiary care hospital is the best way to treat these patients. conclusion delay in diagnosis and surgical treatment, the presence of shock at admission accompanied by evidence of organ dysfunction, increasing comorbidity index and the presence of a nasty micro-organism (clostridia, fungi) all contribute to increased mortality in patients with nstis. diagnosing these infections early, followed by prompt resuscitation, a broad spectrum antibiotic cover and extensive surgical debridement are the cornerstones in improving the outcome in these desperately ill patients. increased time from admission to surgical management is associated with a higher mortality. therefore, timing of the first surgery is of the essence thus highlighting the importance of the “golden hour” for the management of nstis. references 1. gupta y, chhetry m, pathak kr, jha rk, ghimire n, mishra bn, karn nk, singh gk, bhagabati jn. risk factors for necrotizing fasciitis and its outcome at a tertiary care centre. j ayub med coll abbottabad. 2016 oct-dec; 28(4):680-682. pmid: 28586594. 2. bonne sl, kadri ss. evaluation and management of necrotizing soft tissue infections. infect dis clin north am. 2017 sep;31(3):497-511. 3. van sambeek chl, van stigt sf, brouwers l, bemelman m. necrotising fasciitis: a ticking time bomb? bmj case rep. 2017 oct 4; 2017:bcr2017221770. 4. hua c, bosc r, sbidian e, de prost n, hughes c, jabre p, chosidow o, le cleach l. interventions for necrotizing soft tissue infections in adults. cochrane database syst rev. 2018 may 31; 5(5):cd011680. 5. yilmazlar t, ozturk e, alsoy a, ozguc h. necrotizing soft tissue infections: apache ii score, dissemination, and survival. world j surg. 2007 sep; 31(9):1858-62. 6. hefny af, eid ho, al-hussona m, idris km, abuzidan fm. necrotizing fasciitis: a challenging diagnosis. eur j emerg med. 2007 feb; 14(1):50-2. 7. giuliano a, lewis f jr, hadley k, blaisdell fw. bacteriology of necrotizing fasciitis. am j surg. 1977 jul; 134(1):52-7. 8. chen kj, klingel m, mcleod s, mindra s, ng vk. presentation and outcomes of necrotizing soft tissue infections. int j gen med. 2017 jul 31; 10:215-220. 9. hedetoft m, madsen mb, madsen lb, hyldegaard o. incidence, comorbidity and mortality in patients 30 predictors of mortality in necrotizing soft tissue infections in adults – experience at a tertiary care hospital with necrotising soft-tissue infections, 2005-2018: a danish nationwide register-based cohort study. bmj open. 2020 oct 16; 10(10):e041302. 10. ramakrishnan k, salinas rc, agudelo higuita ni. skin and soft tissue infections. am fam physician. 2015 sep 15; 92(6):474-83. 11. khoury mk, pickett ml, cripps mw, park sy, nagaraj mb, hranjec t, hennessy sa. transfer is associated with a higher mortality rate in necrotizing soft tissue infections. surg infect (larchmt). 2020 mar; 21(2):136-142. 12. yiasemidou m, majumder s, basheer m. necrotising fasciitis after laparoscopic rectal cancer surgery. ann r coll surg engl. 2017 apr; 99(4):e113-e114. 13. nawijn f, smeeing dpj, houwert rm, leenen lph, hietbrink f. time is of the essence when treating necrotizing soft tissue infections: a systematic review and meta-analysis. world j emerg surg. 2020 jan 8; 15:4. 14. urschel jd. necrotizing soft tissue infections. postgrad med j. 1999 nov; 75(889):645-9. 15. khoury mk, heid ca, cripps mw, pickett ml, nagaraj mb, johns m, lee f, hennessy sa. antifungal therapy in fungal necrotizing soft tissue infections. j surg res. 2020dec; 256:187-192 16. horn cb, wesp bm, fiore nb, rasane rk, torres m, turnbull ir, ilahi on, punch lj, bochicchio gv. fungal infections increase the mortality rate threefold in necrotizing soft-tissue infections. surg infect (larchmt). 2017 oct; 18(7):793-798. 17. brook i. microbiology and management of soft tissue and muscle infections. int j surg. 2008 aug; 6(4):328-38. 18. meleney fl. bacterial synergism in disease processes: with a confirmation of the synergistic bacterial etiology of a certain type of progressive gangrene of the abdominal wall. ann surg. 1931 dec; 94(6):961-81. 19. bruun t, rath e, madsen mb, oppegaard o, nekludov m, arnell p, karlsson y, babbar a, bergey f, itzek a, hyldegaard o, norrby-teglund a, skrede s; infect study group. risk factors and predictors of mortality in streptococcal necrotizing soft-tissue infections: a multicenter prospective study. clin infect dis. 2021 jan 27; 72(2):293-300. 20. leiblein m, wagner n, adam eh, frank j, marzi i, nau c. clostridial gas gangrene a rare but deadly infection: case series and comparison to other necrotizing soft tissue infections. orthop surg. 2020 dec; 12(6):1733-1747. 21. guerrero dm, perez f, conger ng, solomkin js, adams md, rather pn, bonomo ra. acinetobacter baumannii-associated skin and soft tissue infections: recognizing a broadening spectrum of disease. surg infect (larchmt). 2010 feb; 11(1):49-57. 22. ali a, botha j, tiruvoipati r. fatal skin and soft tissue infection of multidrug resistant acinetobacter baumannii: a case report. int j surg case rep. 2014; 5(8):532-6. 23. berbudi a, rahmadika n, tjahjadi ai, ruslami r. type 2 diabetes and its impact on the immune system. curr diabetes rev. 2020; 16(5):442-449. 24. cancer-induced immunosuppression: il-18–elicited immunoablative nk cells magali terme, evelyn ullrich, laetitia aymeric, kathrin meinhardt, jérôme d. coudert, mélanie desbois, françois ghiringhelli, sophie viaud, bernard ryffel, hideo yagita, lieping chen, salaheddine mécheri, gilles kaplanski, armelle prévost-blondel, masashi kato, joachim l. schultze, eric tartour, guido kroemer, mariapia degli-esposti, nathalie chaput and laurence zitvogel cancer res june 1 2012; (72) (11) 2757-2767. 25. noor mt, manoria p. immune dysfunction in cirrhosis. j clin transl hepatol. 2017; 5(1):50-58. 26. ikram m, saeed o, ali ss, salahuddin i. necrotizing fasciitis of head & neck: the pakistani experience. asian j surg. 2002 jan; 25(1):13-7. 27. 1. beg a, qureshi o, siddiqui f, khurshaidi n. retroperitoneal necrotizing fasciitis in a 42-year-old male patient: a case report [internet]. 2020 [cited 3 may 2021]. available from: http://doi.org/10.47391/jpma.647 28. malik zi. necrotizing fasciitis of lower limb: a surgical emergency. jrmc.; 16(2):135-7. the authors: prof. haroon javaid majid head, dept. of general surgery & surgical oncology, shaikh zayed medical complex, lahore. prof. muhammad imran anwar head, department of surgery (unit-ii), shaikh zayed medical complex, lahore. dr. muhammad zaeem khalid trainee registrar, department of surgery (unit-i), shaikh zayed medical complex, lahore. dr. shafique-ur-rehman senior registrar, department of surgery (unit-i), shaikh zayed medical complex, lahore. dr. muhammad amir jameel senior registrar, department of surgery (unit-ii), shaikh zayed medical complex, lahore. corresponding author: prof. haroon javaid majid head, dept. of general surgery & surgical oncology, shaikh zayed medical complex, lahore. e-mail: hjmajid@gmail.com 69 proceedings s.z.m.c. vol: 35(3): pp. 69-74, 2021. pszmc-812-35-3-2021 fertility-related quality of life amongst married infertile females of lahore 1hira ashfaq, 2ahmad naseem, 3shabnum tariq, 4hina usman, 5tayyiba wasim, 4maria sharif, 6naheed humayun 1department of surgery, eme trust hospital, lahore 2department of medicine, jinnah hospital, lahore 3department of obstetrics and gynecology, lahore general hospital, lahore 4department of public health and community medicine, shaikh zayed medical complex, lahore 5department of obstetrics and gynecology, services institute of medical sciences, lahore 6department of community medicine, red crescent medical college, lahore abstract introduction: in today’s world of science and medicine, assessing ones’ quality of life has become a norm. it provides baseline information for future planning in order to elevate quality of life of individuals and communities. aims & objectives: to assess the quality of life of infertile women and identify factors influencing it. place and duration of study: data was collected from lahore general hospital and services institute of medical sciences, lahore during 2019. material & methods: a cross-sectional study was conducted on 260 married, infertile females attending infertility clinics of two tertiary care hospitals of lahore. 36-items fertiqol, an urdu validated version was used to assess the quality of life of females with infertility. it had six subscales and three total scores. data was analyzed through spss version 22. results: mean age of females was found to be 30+5.23 years and 85.8% were not working women. average duration of infertility was 6.7 years. 61.5% of females presented with primary infertility. 70.4% reported positive attitude of their husband towards this infertility while 56.2% of the participants reported positive attitude of their in-laws as well. 30.8% of the females rated their health as poor. 31.2% reported to be dissatisfied with their overall quality of life. in fertiqol, ‘emotional’ subscale showed mean 67±14 sd, ‘mind-body’ subscale mean 69±14 sd, ‘relational’ subscale mean 55±14.7 sd, ‘social’ subscale mean 66.2±14.5 sd, ‘treatment environment’ mean 70±17.5 sd and ‘treatment tolerability’ showed mean of 73±17.2 sd. ‘total scaled core score’ showed mean of 64.5±12 sd, ‘total scaled treatment score’ mean of 69.3±14 sd and ‘total scored fertiqol score’ showed mean of 66±12 sd. significant association between family income and mind body subscale; duration of marriage and treatment tolerability; time till infertility diagnosis and treatment environment was found. while overall fertiqol score and family income was found to be significantly associated. conclusion: overall quality of life of infertile females is not compromised as was thought of except for its association with family income. sub scales of mind-body, treatment tolerability and treatment environment were found to be associated with family income, duration of marriage and time till infertility diagnosis respectively. recognition of factors associated with poor quality of life will help in planning strategies to overcome them during infertility treatment. key words: fertility, infertility, quality of life, fertiqol introduction infertility by definition is failure to conceive after 1 year of unprotected intercourse amongst heterosexual couples.1 it is used synonymously with terms such as sterility, childlessness and subfertility. it can be devastating for those couples who want to have children and can affect both genders equally.2 as definition of infertility lacks clarity and uniformity, a systematic review was done in 2011 on defining demographic and epidemiological aspects of infertility.3 in a systematic review, 277 demographic and reproductive health surveys data showed infertility prevalence to be highest in south asia, sub-saharan africa, north africa/middle east, central/eastern europe and central asia.4 higher prevalence of sexual dysfunction which includes decreased sexual desire, orgasm and satisfaction was found to be 9.5 folds higher in secondary infertile women as compared to primary infertile.5 world health organization (who) defines quality of life (qol) as an 'individuals’ perceptions of their 70 fertility-related quality of life amongst married infertile females of lahore position in life in the context of the culture and value systems in which they live and in relation to their goals, expectations, standards and concern’.6 infertility influences quality of life of couples especially females. infertility-related perceptions and responses are huge and they influence qol of couples in different ways.7 polycystic ovarian syndrome (pcos) with growing incidence is one of the leading causes of infertility now. a significant amount of studies on pcos and other causes of infertility, showed concerns pointing towards infertility and its’ associated behavioral and quality of life issues in our societies.8,9 infertility related stress, anxiety, depression, psychological and emotional issues are influencing and determining factors of quality of life of couples.10-13 patientreported outcome measures were used to assess infertility-related quality of life in turkish, hungarian, tunisian, iranian and taiwanese couples.14-21 the impact of infertility on the psychological well-being, marital relationships, sexual relationships, and quality of life of couples was assessed in a systematic review of literature and found to be influenced by infertility.22 a comparison between infertile and fertile couples about quality of life, sexual satisfaction and marital satisfaction showed difference.23 a systematic review in 2013 was done on questionnaires measuring quality of life in infertile couples.24 cross-cultural differences were found in fertility specific quality of life in german, hungarian and jordanian couples attending a fertility center.25 a study done in pakistan in 2011 reported psychological consequences of infertility to be huge.26 multiple quality of life assessment tools were developed and validated in different languages over last few decades but a tool comprehensively addressing qol in infertile couples was still needed.27,28 european society of human reproduction and embryology (eshre) and the american society of reproductive medicine (asrm) jointly did efforts to create the fertility quality of life (fertiqol) instrument to measure qol in couples with fertility problems. fertiqol used same protocol as was used for the development of whoqol measure.6 fertiqol is a questionnaire internationally developed to measure fertilityspecific quality of life. it is considered to be a reliable tool and measures impact of fertility problems and their effect on qol. in total it consists of 36 items and has been translated into 20 different languages.29 limited literature is published from pakistan so there is a need to generate evidence on quality of life of infertile females.30,31,32 the objective of this study was to assess the quality of life of infertile females and identify the factors influencing it. material and methods a cross-sectional study was conducted in infertility clinics of two tertiary care teaching hospitals of lahore. administrative consent along with ethical clearance from departmental ethical committee was obtained. total 260 participants (married, infertile females) from 2 centers of lahore (lahore general hospital and services institute of medical sciences) were included in the study. sample size was calculated taking expected proportion of quality of life as 0.5, with 0.05 precision and 95% confidence level. females coming for infertility treatment were consecutively selected till the desired sample size was reached in both hospitals. verbal informed consent was obtained from each participant in presence of witness. fertiqol, which consists of 36 items that assess core (24 items) and treatment-related quality of life (qol) (10 items) and overall life and physical health (2 items) was used.29 the items in the fertiqol survey were rated on a scale of 0-4, a higher score means more favorable qol. it was translated in multiple languages and is available on (http://www.fertiqol.org/). forms were kept anonymous and their data privacy and confidentiality was maintained by the researchers. statistical analysis: statistical analysis was done on spss version 22.0. chi-square, t-test and anova were applied to identify association. results the mean age of the participants was found to be 30 years which shows that it is the average best-fitted value for the whole sample with a ±5.23 standard deviation (sd). duration of infertility of females had mean of 6.7±5.02 sd. total family income had mean of 25853.8 with ±25564.24 standard deviation and total family members had mean of 4.47±2.63 sd. results showed that out of 260 females, 106 (40.8%) had cousin marriages and majority of them (223, 85.8%) were housewives. husband’s occupation showed that most of them 132 (50.8%) were doing jobs. husband’s education revealed that 59 (22.7%) were illiterate, 30 (11.5%) had attained education till primary level, 90 (34.6%) had attained education till matriculation, 35 (13.5%) had done intermediate, 39 (15.0%) did graduation and only 7 (2.7%) had attained higher education. most of the women i.e, 160 (61.5%) had primary type of 71 fertility-related quality of life amongst married infertile females of lahore infertility. 77 (29.6%) of the females reported negative attitude of their husband while negative attitude of in-laws was reported by 114 (43.8%) of the respondents. 80 (30.8%) rated their health as poor and 81 (31.2%) reported to be dissatisfied with their quality of life (table-1). frequency (n=260) percent % cousin marriage yes 106 40.8% no 154 59.2% working women yes 37 14.2% no 223 85.8% husband occupation job 132 50.8% business 46 17.7% laborer 78 30.0% others 3 1.2% unemployed 1 0.4% husband education illiterate 59 22.7 primary 30 11.5 matric 90 34.6 intermediate 35 13.5 bachelors 39 15.0 higher education 7 2.7 infertility type primary 160 61.5% secondary 100 38.5% husband attitude positive 183 70.4% negative 77 29.6% in-laws attitude positive 146 56.2% negative 114 43.8% how would you rate your health? very poor 20 7.7% poor 80 30.8% neither good neither bad 72 27.7% good 70 26.9% very good 18 6.9% are you satisfied with your quality of life? very dissatisfied 2 0.8% dissatisfied 81 31.2% neither satisfied neither dissatisfied 59 22.7% satisfied 107 41.2% very satisfied 11 4.2% table-1: socio-demographic characteristics of married infertile females of lahore range along with mean and sd were calculated for total fertiqol and its subscales. emotional subscale showed a range of 37-95 with mean ± sd of 67±14, mind-body showed a range of 37-100 with mean ± sd of 69±14, relational subscale revealed a range of 20-87 with mean ± sd of 55±14.7, social subscale showed a range of 29-95 with mean ± sd of 66.2±14.5, environment revealed a range of 2992 with mean ± sd of 70±17.5, tolerability revealed a range of 25-100 with mean ± sd of 73±17.2. total scaled core score had range of 38.591 with mean ± sd of 64.5±12, total scaled treatment score had range of 37.5-95 with mean ± sd of 69.3±14 and total scored fertiqol score had range of 41-91 with mean ± sd of 66±12 (table-2). variables and subscales showed a significant association between family income and mind-body subscale, duration of marriage and treatment tolerability, time till infertility diagnosis and treatment environment (table-5). with fertiqol as dependent variable and age and family income as independent variables inferential analysis was performed. level of significance was set at <0.05. t-test was found to be significant (0.021) for fertiqol and family income (table-3). test of variance (anova) was applied for age, duration of marriage, family income and number of family members and was found to be significant only for family income (p-value= 0.019) (table-4). subscales & total fertiqol range (1-100) mean ± sd emotional 37-95 67 ± 14 mind-body 37-100 69 ± 14 relational 20-87 55 ± 14.7 social 29-95 66.2±14.5 treatment environment 29-92 70 ± 17.5 treatment tolerability 25-100 73 ± 17.2 *total scaled core score 38.5-91 64.5 ± 12 **total scaled treatment score 37.5-95 69.3 ± 14 ***total scaled fertiqolscore 41-91 66 ± 12 table-2: fertiqol and the subscales in married infertile females of lahore *total scaled core score= emotional+ mind-body+ relational+ social **total scaled treatment score= treatment environment+ treatment tolerability factors categories of factors mean t-test for equality of means tstatistic p-value age (years) < 25 65.26 0.268 0.789 > 25.1 65.81 family income (rupees) <25000 64.81 -2.322 0.021 >25001 68.45 table-3: t-test for age and family income variables t-test (anova) p-value age -1.320 0.188 duration of marriage 0.22 0.983 family income 2.354 0.019 family members 1.083 0.280 table-4: test of variance (anova) for age, duration of marriage, family income and number of family members 72 fertility-related quality of life amongst married infertile females of lahore variables emotional p value mind-body p value relational p value social p value treatment environment p value treatment tolerability p value total fertiqol p value p a g e 0.75 p a g e 0.40 p a g e 0.71 p a g e 0.44 p a g e 0.56 p a g e 0.00 p a g e 0.627 duration of marriage (years) 1-10 0 35 132 52 0 24 139 56 2 94 104 19 0 35 125 59 0 51 91 77 1 31 97 90 0 26 134 59 11-20 0 8 17 8 0 4 18 11 0 18 14 1 0 9 15 9 0 6 18 9 0 3 18 12 0 6 18 9 21-30 0 2 4 2 0 1 7 0 0 3 5 0 0 2 5 1 0 3 3 2 1 0 5 2 0 2 5 1 family income (rupees) 1-25000 0 34 109 37 0.15 0 22 124 34 0.00 2 82 83 13 0.69 0 32 107 41 0.31 0 48 71 61 0.08 1 27 87 65 0.18 0 25 113 42 0.211 26000250000 0 11 44 25 0 7 40 33 0 33 40 7 0 14 38 28 0 12 41 27 1 7 33 39 0 9 44 27 time till infertility diagnosis (years) 1-8 0 39 136 54 0.49 0 23 146 60 0.46 2 101 107 19 0.62 0 39 132 58 0.31 0 56 95 78 0.03 2 30 106 91 0.53 0 30 139 60 0.618 9-16 0 6 13 8 0 5 15 7 0 13 13 1 0 7 12 8 0 4 17 6 0 3 14 10 0 4 16 7 17-24 0 0 3 0 0 0 3 0 0 0 3 0 0 0 1 2 0 0 0 3 0 0 0 3 0 1 2 3 husband’s attitude positive 0 32 112 39 0.32 0 18 123 42 0.10 1 88 81 13 0.26 0 30 106 47 0.52 0 47 80 56 0.15 2 23 83 75 0.75 0 24 117 42 0.117 negative 0 13 41 23 0 11 41 25 1 27 42 7 0 16 39 22 0 13 32 32 0 11 37 29 0 10 40 27 table-5: association of socio-demographic variables with sub-scales & total fertiqol score discussion quality of life related to infertility and factors influencing it were identified in current research. the results of this study showed mean age of mothers at thirty years while majority was housewives. average duration of infertility was 6.7 years. so on average they were in early marriage phase, staying at home and facing infertility issue for a significant period of time. most of the women had primary type of infertility. these findings are close to a study findings done in karachi in 2016.33 the reason being females belonging to same socioeconomic class and an urban setting where there is better awareness and families are openminded. reproduction is an essential human desire and infertility can manifest itself as stress, anxiety, depression, low self-esteem and declined sexual satisfaction. in our study majority of them rated their health as poor because of infertility and most of them were dissatisfied with their quality of life. in a systematic review, infertility was found to affect couples showing infertility’s negative effect on sexual relationship and psychological well-being of couples, but the effect on quality of life and marital relationships was not found.22 in our study, total fertiqol measured inconclusive effect on quality of life but subscales showed a variable response. test of variance (anova) was applied for age, duration of marriage, family income and number of family members. it was found to be significant only for family income (p-value= 0.019). low income couples and less than 10 years of married life were found to face lower social subscale score in fertiqol in a recent turkish study.15 in our study, majority females reported positive attitude of their husband and in-laws towards their infertility issue. this was contradictory to findings from researches done in developing countries in previous decades but similar to latest research on infertility associated behaviors.34 a study from karachi reported a lower mean for all subscales of fertiqol as compared to our findings.33 our study’s total fertiqol score mean 66 was higher than this. majority (30.8%) of the women included in our study rated their health as poor. 31.2% were dissatisfied with their quality of life and relational subscale which refers to fertilityspecific issues experienced within the couple relationship had mean±sd of 55±14.7. in pakistan second marriage is considered if a woman does not bear child in early years of marriage this stress negatively effects their relationship with husband and his family and also effects physical and mental well-being of herself. treatment tolerability which effectively measures physical and mental well-being and disruption of daily routine due to infertility treatment was found to be significant in this study. treatment environment scale which assesses the positive environment provided by medical staff during infertility treatment showed a positive association with time till infertility. results of another study showed that the score of subscales was higher in women with secondary type of infertility meaning a better quality of life. duration of infertility was also found to be an important factor adversely affecting the quality of life. higher education status of females was found to be associated with better qol.24 it has been shown that lower income, worsened spousal relationship, infertility related perceptions, pressuring oneself or spouse due to infertility, and strong desire for children are significantly associated with a poor quality of life.7 our study also showed that family income adversely effects total fertiqol score but other factors did not show a conclusive influence. 73 fertility-related quality of life amongst married infertile females of lahore self-esteem scores were lower in the couples with longer infertility duration. women are found to be more affected by infertility as compared to men and twelve percent of the women seeking infertility treatment had poor quality of life. reason for this may be the natural desire of females to bear children. women reported about more depressive symptoms and poorer quality of life than men. both in men and women, the higher depression level correlated with lower level of quality of life. moreover, the presence of more depressive symptoms in women was related to men’s poorer quality of life. women are considered weak psychologically so these findings support this assumption.10-13 infertile couples have to face a wide range of psychological influences so there arises the need for psychological support and counseling of infertile couples.34 this was a cross-sectional study focusing on females due to convenience of their presence in infertility clinics as compared to men. a couple based approach may be better for future studies so both can give an insight into quality of life and its subscales. this study provides an comprehensive evaluation of fertility related quality of life in different aspects of infertile women. conclusion it is concluded that quality of life in married infertile women is compromised and is influenced by low family income on the whole but sub scales of mind-body, treatment tolerability and treatment environment are associated with family income, duration of marriage and time till infertility diagnosis respectively. these factors can be addressed to improve quality of life in these subscales/dimensions/ measures such as counseling or psychotherapy need to be incorporated in the conventional treatment so as to improve quality of life of infertile couples. references 1. tanha fd, mohseni m, ghajarzadeh m. sexual function in women with primary and secondary infertility in comparison with controls. international journal of impotence research. 2014; 26(4):132. 2. mascarenhas mn, cheung h, mathers cd, stevens ga. measuring infertility in populations: constructing a standard definition for use with demographic and reproductive health surveys. population health metrics. 2012; 10(1):17. 3. gurunath s, pandian z, anderson ra, bhattacharya s. defining infertility—a systematic review of prevalence studies. human reproduction update. 2011; 17(5):575-88. 4. mascarenhas mn, flaxman sr, boerma t, vanderpoel s, stevens ga. national, regional, and global trends in infertility prevalence since 1990: a systematic analysis of 277 health surveys. plos medicine. 2012; 9(12):e1001356. 5. keskin u, coksuer h, gungor s, ercan cm, karasahin ke, baser i. differences in prevalence of sexual dysfunction between primary and secondary infertile women. fertility and sterility. 2011; 96(5):1213-7. 6. group w. the world health organization quality of life assessment (whoqol): position paper from the world health organization. social science & medicine. 1995; 41(10):1403-9. 7. lau jt, wang q, cheng y, kim jh, yang x, tsui hy. infertility-related perceptions and responses and their associations with quality of life among rural chinese infertile couples. journal of sex & marital therapy. 2008; 34(3):248-67. 8. elsenbruch s, hahn s, kowalsky d, offner ah, schedlowski m, mann k, et al. quality of life, psychosocial well-being, and sexual satisfaction in women with polycystic ovary syndrome. the journal of clinical endocrinology and metabolism. 2003; 88(12):5801-7. 9. mccook jg, reame ne, thatcher ss. health-related quality of life issues in women with polycystic ovary syndrome. journal of obstetric, gynecologic, and neonatal nursing : jognn. 2005; 34(1):12-20. 10. newton cr, sherrard w, glavac i. the fertility problem inventory: measuring perceived infertilityrelated stress. fertil steril. 1999; 72(1):54-62. 11. nicoloro-santabarbara jm, lobel m, bocca s, stelling jr, pastore lm. psychological and emotional concomitants of infertility diagnosis in women with diminished ovarian reserve or anatomical cause of infertility. fertil steril. 2017; 108(1):161-7. 12. yusuf l. depression, anxiety and stress among female patients of infertility; a case control study. pakistan journal of medical sciences. 2016; 32(6):1340-3. 13. keramat a, masoomi sz, mousavi sa, poorolajal j, shobeiri f, hazavhei sm. quality of life and its related factors in infertile couples. journal of research in health sciences. 2014; 14(1):57-63. 14. kitchen h, aldhouse n, trigg a, palencia r, mitchell s. a review of patient-reported outcome measures to assess female infertility-related quality of life. health and quality of life outcomes. 2017; 15(1):86. 15. goker a, yanikkerem e, birge o, kuscu nk. quality of life in turkish infertile couples and related factors. human fertility (cambridge, england). 2017:1-9. 16. crawford nm, hoff hs, mersereau je. infertile women who screen positive for depression are less likely to initiate fertility treatments. human reproduction (oxford, england). 2017; 32(3):582-7. 74 fertility-related quality of life amongst married infertile females of lahore 17. aliyeh g, laya f. quality of life and its correlates among a group of infertile iranian women. medical science monitor : international medical journal of experimental and clinical research. 2007; 13(7):cr313-7. 18. cserepes re, bugan a. [impact of depressive symptomatology in hungarian infertile couples]. psychiatria hungarica : a magyar pszichiatriai tarsasag tudomanyos folyoirata. 2015; 30(1):50-9. 19. cserepes re, korosi t, bugan a. [characteristics of infertility-specific quality of life in hungarian couples]. orvosi hetilap. 2014; 155(20):783-8. 20. el kissi y, amamou b, hidar s, ayoubi idrissi k, khairi h, ali bb. quality of life of infertile tunisian couples and differences according to gender. international journal of gynaecology and obstetrics: the official organ of the international federation of gynaecology and obstetrics. 2014; 125(2):134-7. 21. hsu py, lin mw, hwang jl, lee ms, wu mh. the fertility quality of life (fertiqol) questionnaire in taiwanese infertile couples. taiwanese journal of obstetrics & gynecology. 2013; 52(2):204-9. 22. luk bh, loke ay. the impact of infertility on the psychological well-being, marital relationships, sexual relationships, and quality of life of couples: a systematic review. j sex marital ther. 2015; 41(6):610-25. 23. masoumi sz, garousian m, khani s, oliaei sr, shayan a. comparison of quality of life, sexual satisfaction and marital satisfaction between fertile and infertile couples. international journal of fertility & sterility. 2016; 10(3):290-6. 24. mousavi sa, masoumi sz, keramat a, pooralajal j, shobeiri f. assessment of questionnaires measuring quality of life in infertile couples: a systematic review. journal of reproduction & infertility. 2013; 14(3):110-9. 25. sexty re, hamadneh j, rosner s, strowitzki t, ditzen b, toth b, et al. cross-cultural comparison of fertility specific quality of life in german, hungarian and jordanian couples attending a fertility center. health and quality of life outcomes. 2016; 14:27. 26. sultan s, tahir a. psychological consequences of infertility. hellenic j psychol. 2011; 8:229-47. 27. boivin j, takefman j, braverman a. the fertility quality of life (fertiqol) tool: development and general psychometric properties. fertility and sterility. 2011; 96(2):409-15. e3. 28. mousavi sa, masoumi sz, keramat a, pooralajal j, shobeiri f. assessment of questionnaires measuring quality of life in infertile couples: a systematic review. journal of reproduction & infertility. 2013; 14(3):110. 29. boivin j, takefman j, braverman a. the fertility quality of life (fertiqol) tool: development and general psychometric properties. human reproduction. 2011; 26(8):2084-91. 30. abbasi s, kousar r. the fertility quality of life (fertiqol) questionnaire in pakistani infertile women. 2016. 31. hassan s-u-n, siddiqui s, friedman bd. health status and quality of life of women seeking infertility treatments in baluchistan, pakistan. the british journal of social work. 2019. 32. hi vartf, hi vartuk, jawad f, qureshi h, hameed k, mehmood k, et al. the journal of bahria university medical and dental college karachi, pakistan. 33. abbasi s, kousar r. the fertility quality of life (fertiqol) questionnaire in pakistani infertile women. journal of bahria university medical and dental college. 2016; 6(3):4-. 34. zarif golbar yazdi h, aghamohammadian sharbaf h, kareshki h, amirian m. psychosocial consequences of female infertility in iran: a metaanalysis. front psychiatry. 2020; 11:518961. the authors: dr. hira ashfaq house officer, department of surgery, eme trust hospital, lahore. dr. ahmad naseem house officer, department of medicine, jinnah hospital, lahore. dr. shabnum tariq associate professor, department of obstetrics and gynecology, lahore general hospital. dr. hina usman visiting faculty instructor, dept. of public health and community medicine, shaikh zayed medical complex, lahore. prof. tayyiba wasim department of obstetrics and gynecology, services institute of medical sciences, lahore. maria sharif biostatistician, dept. of public health and community medicine, shaikh zayed medical complex, lahore. prof. naheed humayun head, department of community medicine, red crescent medical college, lahore corresponding author: dr. ahmad naseem house officer, department of medicine, jinnah hospital, lahore. e-mail: ahmad.naseem3344@gmail.com 17 proceedings s.z.m.c. vol: 35(3): pp. 17-23, 2021. pszmc-803-35-3-2021 malus domestica peel extract counters carboplatin induced fall in red blood cell count and bone marrow erythroid cell percentage in mice 1abdul mudabbir rehan, 2fariha ahmad khan, 3qudsia umaira khan, 4rabia naseer khan, 5zoobia irum, 3sehrish zaffar, 6sadia chiragh 1department of pharmacology, d.g. khan medical college, dera ghazi khan 2department of pharmacology, akhtar saeed medical and dental college, lahore 3department of physiology, cmh medical college, lahore 4department of pathology, shahida islam medical and dental college, lodhran 5department of pharmacology, cmh institute of medical sciences, bahawalpur 6department of pharmacology, al-aleem medical college, lahore abstract introduction: the chemotherapy-induced anemia (cia) is a complex phenomenon impacted by many treatments and patient related variables. some variations within cia are type of chemotherapeutic agent, dose and schedule of administration of agent, type as well as stage of cancer, time of intervention, age, gender, nutritional status and renal function. the variety of patient presentation and symptomatology in cia is very thought-provoking while setting up widespread measures to oversee ideal administration of treatments. this study will be focused on protective role of apple peel extract (appe) on carboplatin induced fall in blood rbc count bone marrow erythroid cell percentage in mice. aims & objective: to evaluate the protective effect of malus domestica peel extract on carboplatin induced fall in red blood cell count and bone marrow erythroid cell percentage in mice. place and duration of study: the study was conducted at pharmacology department of postgraduate medical institute, lahore in june 2016. the total duration of study was 1 month. material & methods: forty adult male mice were randomly divided into 5 groups (n=8) and maintained on standard diet and water adlibitum. group a was negative control. group b was disease control wherein each animal received one dose of 125mg/kg carboplatin intraperitoneally. group c, d & e experimental groups were administered same dose carboplatin followed by the ethanolic apple peel extract in three different single oral doses of 25, 50 & 75mg/kg respectively for 7 days. blood & tissue sampling was done on the 8th day. data was analyzed using graf pad prism 7. results: acute carboplatin anaemia resulted in 4.01±0.851012/l rbc count & 21.6±1.06% bone marrow erythroid cell percentage in disease control group b. subsequent apple peel extract administration in a dose of 25, 50 and 75mg/kg countered carboplatin induced fall in groups c, d & e, simultaneously raising rbc counts & bone marrow erythroid cell percentage to (6.19±0.70×1012/l & 21.3±1.28%) (6.56±0.37×1012/l & 25.3±1.04%) and (6.26±0.74×1012/l & 23.5±1.41%.) respectively. the increase in rbc as well as erythroid cell counts was most significant with the dose of 50mg/kg and nearing that of healthy control levels (group a) conclusion: apple peel extract effectively prevented carboplatin induced fall in rbc count and erythroid cell percentage and produced most significant results in a dose of 50 mg/kg. key words: chemotherapy induced anemia (cia), carboplatin, malus domestica, erythroid cell percentage introduction the word anemia is taken from the “anaimia”, a greek word which means deficiency of blood. it also denotes the reduced oxygen carrying ability of rbcs in blood and it is associated very commonly with cancers.1 anemia is considered the most common and obstinate haematological issue in cancer patients.2 chemotherapy induced anemia (cia) occurs due to malignant invasion of healthy tissues which ultimately leads to loss of blood and marrow intrusion with a resultant disruption in erythropoiesis and functional deficiency of iron due to inflammation.3 anemia due to cancer chemotherapy is significant and very important because it may delay or limit treatment as well as cause serious fatigue and diminished life quality of cancer patients. the grading scale for anemia presented by the national cancer institute anemia scale is as follows:2 18 malus domestica peel extract counters carboplatin induced fall in red blood cell count… grade 0 – haemoglobin ranges from 14–18 g/dl for men and 12–16 g/dl for women grade 1 haemoglobin ranges from 10–14 g/dl for men and 10–12 g/dl for women grade 2 haemoglobin ranges from 8–10 g/dl grade 3 haemoglobin ranges form 6.5–8 g/dl grade 4 haemoglobin<6.5 g/dl grade 5 death the patients getting chemotherapy treatment usually have certain other comorbid conditions like insufficient renal function. approximately, 70% of cancer patients getting chemotherapy suffer from anemia.4 the adjunct care strategies like stem cell support and marrow transplant are crucial to alleviate the cytotoxic chemotherapeutic effects.2 the transfusion rates differ in cia patients, but approximately 50% of patients with terminal cancer need rbc transfusion at least once before finishing chemotherapy treatment.5 high dose chemotherapy produces higher response rate in cancer patients.2 the chemotherapy disturbs all cells which are rapidly dividing, including erythroid progenitor cells in a phenomenon called as eryptosis.6 the eryptosis is a process in which erythrocytes destroy and eliminate malfunctioning erythrocytes which prevents hemolysis & haemoglobin release.7 eryptosis and erythrocyte deficiency lead to anemia and it is worsened by insufficient compensatory erythropoiesis.7 lymphomas, gynecologic & genitourinary tumors and lung tumors have highest cia incidence with at least 50%–60% requiring blood or rbc transfusion.2 cia is precipitated by platinum based chemotherapies (cisplatin and carboplatin etc.).8 the development of platinum induced anemia is dependent upon many factors like an early decline in haemoglobin levels following the treatment, cumulative doses of platinum analogues, old age, therapeutic failure and high amount of outstanding platinum in following the administration.2 the mechanisms of cia due to platinum analogues comprises of direct suppression of erythroid progenitor cells within the bone marrow.9 carboplatin is an anticancer drug which belongs to alkylating class of cancer chemotherapeutic drugs. it is used effectively to treat many cancers. experimentally, carboplatin has been used to induce myelosuppression in mice.10 carboplatin induces myelosuppression by increasing overall oxidative stress and decrease in the glutathione content inside bone marrow.11 it also cross links the dna and generates oxidative stress products such as malondialdehyde.12 carboplatin suppresses the bone marrow production of red blood cells by interfering with redox reactions inside marrow cavity and shifts the redox equilibrium towards oxidative side, it can be concluded that toxicities of carboplatin are due to increase in the overall oxidative stress inside bone marrow and if this abrupt rise in oxidative stress could somehow be prevented then the serious untoward effects can be avoided. various antioxidant compounds like glutathione,13 squalene14 and l-carnitine11 have shown protective effects on carboplatin induced myelotoxicity in animal models. apples are the most widely consumed fruit worldwide and contain five major polyphenols. apple peel contains 3 to 6 times more flavonoids as compared to apple flesh that’s why; apple peel extracts have more antioxidant potential than apple flesh extracts.15 apple peel extract by virtue of its antioxidant property has potential to restore the redox equilibrium and improve the peripheral red rbc count and bone marrow erythroid cell percentage. we evaluated the protective effect of malus domestica peel extract on carboplatin induced fall in red blood cell count and bone marrow erythroid cell percentage in mice. material and methods materials used in this research were apples (red delicious variety), injection ketamine (indus pharma pvt. ltd), injection carpsol 150mg/15ml (pfizer pharma pvt. ltd), electronic balance (wuhan panscale hardware co. ltd, model no. dh-v300a), hematology analyzer (sysmex model no. kx-21, serial no. b 3483), light microscope (olympus cht, japan), edta blood vacutainer (bio-vac stars pakistan), whatman filter paper. no.1 (whatman international ltd, maidstone, uk), giemsa stain (diachem, china), hematoxylin & eosin stains (diachem, china), disposable syringes 3 & 5ml (bd syringe becton dickinson, pakistan), disposable examination gloves (max pluss-100, malaysia). it was an experimental study conducted at pharmacology department of post graduate medical institute (pgmi), lahore in june 2016 & approved by ethical committee for basic sciences of pgmi. forty healthy male adult swiss albino mice, 7-8 weeks of age weighing 25-35g were included in the study. apparently unhealthy-looking mice were excluded mice were purchased from the university of veterinary and animal sciences (uvas), lahore and kept in the animal house of pgmi under hygienic condition. these were randomly divided via simple lottery method into 5 19 malus domestica peel extract counters carboplatin induced fall in red blood cell count… groups (n= 8). the temperature was maintained in a range of 19-22°c, with a natural day and night cycle. before the onset of study, all mice were kept for a week to acclimatize and they were provided with diet and water ad libitum. a single intraperitoneal injection of carboplatin (injection carpsol 150mg/15ml by pfizer pharma) in a dose of 125 mg/kg was given to induce myelosuppression in mice.10 red delicious variety of local apples was selected because it contains the richest proportion of antioxidants among all the locally produced varieties.16 apples were washed with plain running water, air dried and peeled carefully so that the peel may not contain flesh. the collected peel was spread and allowed to dry in shade for 2 weeks. the partially dried peels were then put in hot air oven at a temperature of 60˚c for 3 hours. the completely dried peel was then coarsely ground with pestle and mortar. the powdered peel was soaked in 80% of ethanol (1:10, w/v) at room temperature for 3 days with daily shaking.16 the filtration of solution was done by filtering it through whatman filter paper no. 1 and was separated from the liquid extract. the excess of solvent was evaporated and gummy paste like extract was stored at 4°c.16,17 the five study groups were given drugs/each mouse as described in the table below. mice groups drug by intra peritoneal route (single dose on day 1 only) drug by oral route (once daily dose from day 1 to 7) a normal control sterile water 1.25 ml/kg distilled water 4 ml/kg b disease control carboplatin 125 mg/kg distilled water 4 ml/kg c experimental 25 mg/kg carboplatin 125 mg/kg apple peel extract (25mg/kg) 25 mg/4ml/kg d experimental 50 mg/kg carboplatin 125 mg/kg apple peel extract (50mg/kg) 50 mg/4ml/kg e experimental 75 mg/kg carboplatin 125 mg/kg apple peel extract (75 mg/kg) 75 mg/4ml/kg table-1: summary of drugs administered to all groups sampling technique & analyzed parameters 1. blood: twenty-four hours after the last dose, the mice were anesthetized with ketamine which was administered via single intraperitoneal injection in a dose of 100 mg/kg17,18 into the left lower quadrant of abdomen. the mice were dissected afterwards to expose the heart and 1.5ml blood was withdrawn directly from the right ventricle with the help of 23-gauge needle and 3 ml disposable syringe. the blood was collected in edta vacutainer.17 the red blood cell count was analyzed by haematology analyzer 2. bone marrow aspirate: the already dissected mice were euthanized by giving a single sharp cut at neck using surgical scalpel and further dissected to obtain the right femur bone. the contents of right femur were aspirated into 0.2ml of ice-cold phosphate buffer saline by using 23gauge needle and 10 ml syringe. the aspirate was then spread over slide and smear was prepared. once the smear was air dried it was dipped into methanol solution to fix the specimen over the slide. finally, the slide was stained with geimsa stain, washed with plain running water and cover slip was applied.17,19 differential cell count was done under oil immersion lens of light microscope. 200 cells were counted and relative percentage of red blood cell was entered in the proforma only.17,19 statistical analysis: the data was transcribed into graphpad prism version 7.0. as data was found to be normally distributed by shapiro wilk test, analysis of variance (anova) and post hoc tukey’s test were applied to see which group mean differs. the pvalue of this hypothesis was <0.05 (level of significance). results red blood cell count: the mean rbc count ± standard deviation of all the groups is illustrated in fig-1. the comparison of means of all groups by anova revealed a significant difference between the group means with a p value of <0.001. the post hoc tukey’s test was applied to analyze the rbc count difference between the groups. the disease control group had markedly lower rbc count as compared to normal control group, whereas all experimental groups had significant higher rbc count as compared to disease control group. all experimental groups had lower rbc count as compared to normal control and the difference amongst 3 doses of apple peel extract was statistically nonsignificant (table-2). erythroid cell percentage: the mean erythroid cell percentage ± standard deviation of all groups is illustrated in fig-2. the comparison of means of the all groups by anova revealed a significant difference between the group means with a p value of <0.001. 20 malus domestica peel extract counters carboplatin induced fall in red blood cell count… fig-1: effect of carboplatin and carboplatin plus 3 different doses of apple peel extract on red blood count (mean ±sd) in mice (n=8) ***=p value <0.001 (vs normal control), … = p value <0.001 (vs disease control), .. = p value <0.01 (vs disease control) post hoc tukey’s multiple comparison test mean difference significance group a normal control disease control 2.57 *** experimental (25 mg/kg) 0.40 ns experimental (50 mg/kg) 0.02 ns experimental (75 mg/kg) 0.35 ns group b disease control experimental (25 mg/kg) -2.17 *** experimental (50 mg/kg) -2.55 *** experimental (75 mg/kg) -2.22 *** group c experimental (25 mg/kg) experimental (50 mg/kg) -0.37 ns experimental (75 mg/kg) -0.05 ns group d experimental (50 mg/kg) experimental (75 mg/kg) 0.32 ns table-2: comparison of effect of apple peel extract, carboplatin and carboplatin plus 3 different doses of apple peel extract on red blood cell count in mice by post hoc tukey’s test (n=8) ns = not significant, * = p value < 0.05, ** = p value < 0.01, *** = p value <0.001 fig-2: effect of carboplatin and carboplatin plus 3 different doses of apple peel extract on erythroid cell percentage (mean ± sd) in bone marrow aspirate smear of mice (n=8) *** = p value <0.001 (vs normal control), ** = p value <0.01 (vs normal control), … = p value <0.001 (vs disease control) post hoc tukey’s multiple comparison test mean difference significance group a normal control disease control 8.25 *** experimental (25 mg/kg) 8.62 *** experimental (50 mg/kg) 4.62 *** experimental (75 mg/kg) 6.37 *** group b disease control experimental (25 mg/kg) 0.37 ns experimental (50 mg/kg) -3.62 *** experimental (75 mg/kg) -1.87 * group c experimental (25 mg/kg) experimental (50 mg/kg) -4.00 *** experimental (75 mg/kg) -2.25 * group d experimental (50 mg/kg) experimental (75 mg/kg) 1.75 ns table-3: comparison of effect of carboplatin, apple peel extract and carboplatin plus 3 different doses of apple peel extract on erythroid cell percentage in bone marrow aspirate smear of mice by post hoc tukey’s test (n=8) ns = not significant, * = p value < 0.05, ** = p value < 0.01, *** = p value <0.01 the post hoc tukey’s test was applied to analyze the erythroid cell percentage difference between the groups. the disease control group had markedly lower erythroid cell percentage as compared to 21 malus domestica peel extract counters carboplatin induced fall in red blood cell count… normal control, whereas experimental groups 50 & 75 mg/kg had significant higher erythroid cell percentage as compared to disease control group. experimental 50 mg/kg group and experimental 75 mg/kg group had significant higher erythroid cell percentage as compared to experimental 25 mg/kg group, whereas difference between experimental 50 & 75 mg/kg was statistically nonsignificant(table3). discussion cancer is the 2nd most common leading cause of death worldwide which causes severe morbidity and mortality in all age groups. chemotherapy is the prime option for initial medical management of various cancers, but it is the acute or cumulative toxicity of these agents which impairs treatment and ultimately leads to either dose reduction or treatment delay, thus reducing the drug efficacy and quality of life of cancer patients.20 the most common dose dependent adverse effects of various chemotherapeutic agents are alopecia, myelosuppression and gastrointestinal irritation, whereas myelosuppression is the only treatment limiting toxicity in almost all of the cancer patients.21 that is why adjuvants, neo-adjuvants and adjunct therapies are used along with the principal therapeutic strategy to abate cancers. the gm-csf has been used in cancer patients to improve peripheral neutrophils,22 erythropoietin to promote the production of erythrocytes,23 and thrombopoietin to treat thrombocytopenia.24 and blood transfusion to improve pancytopenia. however, the indications of the above regimens are limited because of their high costs and serious adverse effects like transfusion related immunomodulation.25,26 and risk of tumor growth and shortening of survival with the use of epoetin.2 the ultimate solution to chemotherapy induced myelosuppression lies in the natural compounds which can prevent the dose dependent chemotherapy induced myelosuppression by virtue of their antioxidant capacity. in current study, there was significant decrease in peripheral blood rbc count which was accompanied by a significant decrease in erythroid cell percentage in bone marrow aspirate smear of disease control group. the rbc count and erythroid cell percentage increased significantly in all experimental groups as compared to disease control group and the maximum increase was observed in 50 mg/kg group. apple peel extract significantly prevented carboplatin induced oxidative stress in experimental groups and there was rise in rbc count along with the rise in erythroid cells. in a similar study, significant rise in rbc count was observed in a compound lycium barbarum polysaccharide. lycium barbarum polysaccharide in a dose of 50, 100 and 200 mg/kg promoted the peripheral rbc count in carboplatin induced myelosuppressive mice.27 in another study conducted by shim and his colleagues, similar observations were made. ginsan, a traditional chinese herbal medicine improved the peripheral blood rbc count in chemotherapy treated mice.28 there was 1.5-fold increase in the blood cell counts in mice which were post-treated with the ginsan. ginsan increased the bone marrow cellularity also. the number of bone marrow cells in the mice treated with ginsan increased from 17.45 ± 0.07 in the chemotherapy treated group to 22.55 ± 0.19 x 106 cells (p value <0.05). the increase in the bone marrow cellularity is a remarkable effect which depicts the protective effect of ginsan just like the apple peel extract. no comparable studies on apple peel extract were found. in current study, three different doses of apple peel extract were used to find out the right therapeutic dose. the study results demonstrated that all three doses of apple peel extract caused an overall numerical increase in the peripheral rbc count and bone marrow cellularity of all experimental groups, but statistical analysis revealed that the apple peel extract in a dose of 50 mg/kg is more significant as compared to the rest of 2 doses. the significantly better results of 50 mg/kg dose as compared to 25 mg/kg dose demonstrates dose dependent preventive effect, while inferior effect of 75 mg/kg dose may be explained by the fact that antioxidants in a high dose act as pro-oxidants. adulteration and contamination of medicinal plants with transition metal ions such as copper and iron can aggravate the pro-oxidant effects of medicinal plants by catalyzing oxidation reactions. besides, polyphenols are metabolized by phase i and phase ii enzymes, and their metabolism can produce intermediate and final metabolites and reactive oxygen species (ros) with prooxidants properties. while cells exposed to medicinal extracts at short exposure times and low concentrations usually show increased cell viability, the powerful antioxidant extracts in high concentrations have been found to be cytotoxic by inducing severe oxidative stress29 conclusion apple peel extract is effective in preventing the fall in red blood cell count & erythroid cell percentage induced by the carboplatin driven oxidative stress. apple peel extract produced most significant results in a dose of 50 mg/kg. 22 malus domestica peel extract counters carboplatin induced fall in red blood cell count… acknowledgement: authors are thankful to staff of animal house and chemical pathology laboratory of postgraduate medical institute, lahore for their cooperation. references 1. radziwon p, krzakowski m, kalinka-warzocha e, zaucha r, wysocki p, kowalski d, et al. anaemia in cancer patients expert group recommendations. oncology in clinical practice. 2017; 13(5):202-10. 2. bryer e, henry d. chemotherapy-induced anemia: etiology, pathophysiology, and implications for contemporary practice. international journal of clinical transfusion medicine. 2018; 6:21. 3. gilreath ja, stenehjem dd, rodgers gm. diagnosis and treatment of cancer related anemia. american journal of hematology. 2014; 89(2):203-12. 4. barrett-lee pj, ludwig h, birgegård g, bokemeyer c, gascón p, kosmidis pa, et al. independent risk factors for anemia in cancer patients receiving chemotherapy: results from the european cancer anaemia survey. oncology. 2006; 70(1):34-48. 5. adamson jw. the anemia of inflammation/ malignancy: mechanisms and management. ash education program book. 2008; 2008(1):159-65. 6. taylor sj, duyvestyn jm, dagger sa, dishington ej, rinaldi ca, dovey om, et al. preventing chemotherapy-induced myelosuppression by repurposing the flt3 inhibitor quizartinib. science translational medicine. 2017; 9(402). 7. lang e, lang f. triggers, inhibitors, mechanisms, and significance of eryptosis: the suicidal erythrocyte death. biomed research international. 2015: 1-16. 8. bamias a, aravantinos g, kalofonos c, timotheadou n, siafaka v, vlahou i, et al. prevention of anemia in patients with solid tumors receiving platinum-based chemotherapy by 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2004; 40(11):1713-23. 13. pratibha r, sameer r, rataboli pv, bhiwgade da, dhume cy. enzymatic studies of cisplatin induced oxidative stress in hepatic tissue of rats. european journal of pharmacology. 2006; 532(3):290-3. 14. das b, yeger h, baruchel h, freedman m, koren g, baruchel s. in vitro cytoprotective activity of squalene on a bone marrow versus neuroblastoma model of cisplatin-induced toxicity: implications in cancer chemotherapy. european journal of cancer. 2003; 39(17):2556-65. 15. karaman ş, tütem e, başkan ks, apak r. comparison of antioxidant capacity and phenolic composition of peel and flesh of some apple varieties. journal of the science of food and agriculture. 2013; 93(4):867-75. 16. manzoor m, anwar f, saari n, ashraf m. variations of antioxidant characteristics and mineral contents in pulp and peel of different apple (malus domestica borkh.) cultivars from pakistan. molecules. 2012; 17(1):390-407. 17. rehan am,khan rn, zaffar s, malik m, chiragh s, hafeez a. protective effect of malus domestica (apple) peel extract on carboplatin induced fall in blood platelet count and bone marrow megakaryocyte percentage in mice. esculapio. 2020; 16(2):5. 18. xu q, ming z, dart am, du xj. optimizing dosage of ketamine and xylazine in murine echocardiography. clinical and experimental pharmacology and physiology. 2007;34(56):499-507 19. gibson fm, michael andrews c, diamanti p, rizzo s, macharia g, gordonsmith ec, et al. a new model of busulphan induced chronic bone marrow aplasia in the female balb/c mouse. international journal of experimental pathology. 2003; 84(1):31-48. 20. huitema a, spaander m, mathôt r, tibben m, holtkamp m, beijnen j, et al. relationship between exposure and toxicity in high-dose chemotherapy with cyclophosphamide, thiotepa and carboplatin. annals of oncology. 2002; 13(3):374-84. 21. wang y, probin v, zhou d. cancer therapy-induced residual bone marrow injury: mechanisms of induction and implication for therapy. current cancer therapy reviews. 2006; 2(3):271-9. 22. bennett cl, djulbegovic b, norris lb, armitage jo. colony-stimulating factors for febrile neutropenia during cancer therapy. new england journal of medicine. 2013; 368(12):1131-9. 23. debeljak n, solár p, sytkowski aj. erythropoietin and cancer: the unintended consequences of anemia correction. frontiers in immunology. 2014; 5:563. 24. kuter dj. managing thrombocytopenia associated with cancer chemotherapy.oncology.2015;29(4):282 25. blajchman m. transfusion immunomodulation or trim: what does it mean clinically? hematology. 2005; 10(sup1):208-14. 26. smith tj, bohlke k, lyman gh, carson kr, crawford j, cross sj, et al. recommendations for 23 malus domestica peel extract counters carboplatin induced fall in red blood cell count… the use of wbc growth factors: american society of clinical oncology clinical practice guideline update. journal of clinical oncology. 2015;33(28):3199-212 27. gong h, shen p, jin l, xing c, tang f. therapeutic effects of lycium barbarum polysaccharide (lbp) on irradiation or chemotherapy-induced myelosuppressive mice. cancer biotherapy & radiopharmaceuticals. 2005; 20(2):155-62. 28. shim j, han y, ahn j, yun y, song j. chemoprotective and adjuvant effects of immunomodulator ginsan in cyclophosphamidetreated normal and tumor bearing mice. international journal of immunopathology and pharmacology. 2007; 20(3):487-97. 29. nasri h, rafieian-kopaei m. medicinal plants and antioxidants: why they are not always beneficial? iranian journal of public health. 2014; 43(2):255. the authors: dr. abdul mudabbir rehan assistant professor, department of pharmacology, d. g. khan medical college, dera ghazi khan. dr. fariha ahmad khan assistant professor, department of pharmacology, akhtar saeed medical and dental college, lahore. dr. qudsia umaira khan assistant professor, department of physiology, cmh medical college, lahore. dr. rabia naseer khan assistant professor, department of pathology, shahida islam medical and dental college, lodhran dr. zoobia irum assistant professor, department of pharmacology, cmh institute of medical sciences, bahawalpur. dr. sehrish zaffar assistant professor, department of pharmacology, cmh medical college, lahore. prof. sadia chiragh department of pharmacology, al-aleem medical college, lahore. corresponding author: dr. abdul mudabbir rehan assistant professor, department of pharmacology, d. g. khan medical college, dera ghazi khan. e-mail: abdulmudabbir@yahoo.com 03 8 proceedings s.z.m.c. vol: 36(1): pp. 8-13, 2022. pszmc-826-36-1-2022 1department of pathology, king edward medical university, lahore 2institute of blood disease transfusion service (ibts) 3department of haematology, king edward medical university, lahore 9 frequency of leukoerythroblastic picture & hematological parameters in covid-19 patients… 10 frequency of leukoerythroblastic picture & hematological parameters in covid-19 patients… 11 frequency of leukoerythroblastic picture & hematological parameters in covid-19 patients… 12 frequency of leukoerythroblastic picture & hematological parameters in covid-19 patients… et al. 13 frequency of leukoerythroblastic picture & hematological parameters in covid-19 patients… 10 44 proceedings s.z.m.c. vol: 36(1): pp. 44-48, 2022. pszmc-833-36-1-2022 nigella sativa 1department of pharmacology, islam medical college, sialkot 2department of pharmacology, d.g. khan medical college, dera ghazi khan 3department of pharmacology, rawalpindi medical university, rawalpindi 4department of pharmacology, shahida islam medical college, lodhran 5department of pharmacology, islamabad medical and dental college, islamabad nigella sativa nigella sativa nigella sativa nigella sativa nigella sativa nigella sativa nigella sativa, 45 nigella sativa seeds protective ability in pyrazinamide induced hyperuricemia in mice cinnamomum cassia, artemisia vulgaris, onion , h. lantanaefolia, caesalpinia sappan, lycopus europaeus allium cepa nigella sativa nigella sativa nigella sativa n. sativa n. sativa nigella sativa ad libitum. nigella sativa nigella sativa ns (nigella sativa) 46 nigella sativa seeds protective ability in pyrazinamide induced hyperuricemia in mice nigella sativa nigella sativa nigella sativa nigella sativa post hoc . pp pppost hoc nigella sativa 47 nigella sativa seeds protective ability in pyrazinamide induced hyperuricemia in mice p nigella sativa p zingiber officinale cinnamomum osmopholieum p nigella sativa nigella sativa nigella sativa nigella sativa nigella sativa d 48 nigella sativa seeds protective ability in pyrazinamide induced hyperuricemia in mice 06 26 proceedings s.z.m.c. vol: 36(1): pp. 26-30, 2022. pszmc-829-36-1-2022 colebrookea oppositifolia 1department of pharmacology, post graduate medical institute, lahore 2department of pharmacology, king edward medical university, lahore 3department of pharmacology, fatima memorial hospital, lahore 4department of histopathology, jinnah hospital, lahore 5department of medicine, sir ganga ram hospital, lahore colebrookea oppositifolia (co) colebrookea oppositifolia c. oppositifolia colebrookea oppositifolia , colebrookea oppositifolia c0 27 colebrookea oppositifolia anti-arthritic potential vs methotrexate in pristane induced rat arthritis colebrookea oppositifolia c0 c0 c0 c0 28 colebrookea oppositifolia anti-arthritic potential vs methotrexate in pristane induced rat arthritis c0 co co 29 colebrookea oppositifolia anti-arthritic potential vs methotrexate in pristane induced rat arthritis c. oppositifolia c. oppositifolia c. oppositifolia co co co co co 30 colebrookea oppositifolia anti-arthritic potential vs methotrexate in pristane induced rat arthritis colebrookea oppositifolia sm. pergularia daemia moringa rivae untitled-2 6 proceedings s.z.m.c. vol: 35(4): pp. 6-12, 2021. pszmc-816-35-4-2021 1cmh lahore medical college, lahore 2department of community medicine, cmh lahore medical college, lahore 7 symptoms and associations of covid-19 in pakistan: a single centre, descriptive study 8 symptoms and associations of covid-19 in pakistan: a single centre, descriptive study 9 symptoms and associations of covid-19 in pakistan: a single centre, descriptive study 10 symptoms and associations of covid-19 in pakistan: a single centre, descriptive study 11 symptoms and associations of covid-19 in pakistan: a single centre, descriptive study 12 symptoms and associations of covid-19 in pakistan: a single centre, descriptive study file.indd 78 proceedings s.z.m.c. vol: 35(2): pp. 78-82, 2021. pszmc-798-35-2-2021 1department of pharmacology, al-aleem medical college, lahore 2allama iqbal medical college, lahore 3department of pharmacology, shalamar medical and dental college, lahore 4department of pharmacology, rahbar medical college, lahore 79 effect of atorvastatin alone and in combination with aspirin on uric acid handling of normal rats 80 effect of atorvastatin alone and in combination with aspirin on uric acid handling of normal rats p p p p p p p p p p 81 effect of atorvastatin alone and in combination with aspirin on uric acid handling of normal rats 82 effect of atorvastatin alone and in combination with aspirin on uric acid handling of normal rats file.indd 35 proceedings s.z.m.c. vol: 35(2): pp. 35-40, 2021. pszmc-790-35-2-2021 1department of plastic surgery, shaikh zayed medical complex, lahore 2department of ent, shaikh zayed medical complex, lahore 36 effectiveness of amnion on chronic wounds vs saline dressing 37 effectiveness of amnion on chronic wounds vs saline dressing 38 effectiveness of amnion on chronic wounds vs saline dressing 39 effectiveness of amnion on chronic wounds vs saline dressing wounds j dent res 40 effectiveness of amnion on chronic wounds vs saline dressing 12 56 proceedings s.z.m.c. vol: 36(1): pp. 56-61, 2022. pszmc-835-36-1-2022 1department of physiology, sharif medical & dental college, lahore 2department of anatomy, king edward medical university, lahore 3department of physiology, shaikh zayed medical complex, lahore 57 variations in symptomatology of migraine among local population of pakistan 58 variations in symptomatology of migraine among local population of pakistan 59 variations in symptomatology of migraine among local population of pakistan 60 variations in symptomatology of migraine among local population of pakistan 61 variations in symptomatology of migraine among local population of pakistan untitled-2 18 proceedings s.z.m.c. vol: 35(4): pp. 18-24, 2021. pszmc-818-35-4-2021 1university institute of diet and nutritional sciences, the university of lahore, lahore 2university institute of food sciences and technology, the university of lahore, lahore 19 food insecurity and its effect on mental health: a systematic review 20 food insecurity and its effect on mental health: a systematic review 21 food insecurity and its effect on mental health: a systematic review 22 food insecurity and its effect on mental health: a systematic review 23 food insecurity and its effect on mental health: a systematic review 24 food insecurity and its effect on mental health: a systematic review untitled-2 51 proceedings s.z.m.c. vol: 35(4): pp. 51-57, 2021. pszmc-824-35-4-2021 1department of oral pathology, dental section, bolan medical college, quetta 2health professional education and research department, faisalabad medical university, faisalabad 3consultant gastroenterologist, chughtai medical center, dha, lahore 52 an updated insight into learning approach of government sector dental students in balochistan 53 an updated insight into learning approach of government sector dental students in balochistan 54 an updated insight into learning approach of government sector dental students in balochistan 55 an updated insight into learning approach of government sector dental students in balochistan 56 an updated insight into learning approach of government sector dental students in balochistan 57 an updated insight into learning approach of government sector dental students in balochistan for web full book 7 proceedings s.z.m.c. vol: 36(2): pp. 7-13, 2022. pszmc-837-36-2-2022 1department of community medicine, shalamar medical and dental college, lahore. 2department of bioethics, shalamar medical and dental college, lahore. 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 8 reopening of universities for on-campus teaching in covid-19 pandemic: status of generalized… 9 reopening of universities for on-campus teaching in covid-19 pandemic: status of generalized… 10 reopening of universities for on-campus teaching in covid-19 pandemic: status of generalized… 11 reopening of universities for on-campus teaching in covid-19 pandemic: status of generalized… 12 reopening of universities for on-campus teaching in covid-19 pandemic: status of generalized… 13 reopening of universities for on-campus teaching in covid-19 pandemic: status of generalized… untitled-2 41 proceedings s.z.m.c. vol: 35(4): pp. 41-45, 2021. pszmc-822-35-4-2021 1department of forensic medicine, nawaz sharif medical college, gujrat 2department of forensic medicine, king edward medical university, lahore 42 determination of postmortem interval by estimating csf proteins concentration after death… 43 determination of postmortem interval by estimating csf proteins concentration after death… 44 determination of postmortem interval by estimating csf proteins concentration after death… 45 determination of postmortem interval by estimating csf proteins concentration after death… file.indd 68 proceedings s.z.m.c. vol: 35(2): pp. 68-72, 2021. pszmc-796-35-2-2021 1department of pulmonology, gulab devi chest hospital, lahore 2department of pulmonology, shaikh zayed medical complex, lahore 3department of ent, shaikh zayed medical complex, lahore 4department of cardiology, shaikh zayed medical complex, lahore 5department of plastic surgery, shaikh zayed medical complex, lahore 69 a comparative study between-small bore and large bore tubes for therapeutic pleural drainage 70 a comparative study between-small bore and large bore tubes for therapeutic pleural drainage 71 a comparative study between-small bore and large bore tubes for therapeutic pleural drainage 72 a comparative study between-small bore and large bore tubes for therapeutic pleural drainage file.indd 52 proceedings s.z.m.c. vol: 35(2): pp. 52-57, 2021. pszmc-793-35-2-2021 1department of anatomy, rashid latif medical college, lahore 2department of anatomy, shaikh zayed medical complex, lahore 3department of anatomy, ucmd, university of lahore 53 flax seed oil alleviate caffeinated energy drink induced metamorphosis in femur weight and relative tissue weight 54 flax seed oil alleviate caffeinated energy drink induced metamorphosis in femur weight and relative tissue weight 55 flax seed oil alleviate caffeinated energy drink induced metamorphosis in femur weight and relative tissue weight 56 flax seed oil alleviate caffeinated energy drink induced metamorphosis in femur weight and relative tissue weight 57 flax seed oil alleviate caffeinated energy drink induced metamorphosis in femur weight and relative tissue weight untitled-2 30 proceedings s.z.m.c. vol: 35(4): pp. 30-35, 2021. pszmc-820-35-4-2021 carica papaya 1department of pharmacology, fatima jinnah medical university, lahore 2department of pharmacology, services institute of medical sciences, lahore 3department of pharmacology, al-aleem medical college, lahore carica papaya carica papaya carica papaya carica papaya, carica papaya c.papaya carica papaya carica papaya carica papaya carica papaya carica papaya 31 effect of carica papaya leaf juice on blood cell count of busulfan-induced chronic bone marrow aplasia.. ad libitum carica papaya carica papaya carica papaya carica papaya carica papaya carica papaya carica papaya carica papaya post hoc carica papaya carica papaya p post hoc carica papaya 32 effect of carica papaya leaf juice on blood cell count of busulfan-induced chronic bone marrow aplasia.. carica papaya ** #9. 70 11 .2 4 10 .8 6 hb (1a) normal group busulfan group c.papaya+ busulfan group *** ###6. 5 0 2 51 0 .4 3 9 .5 4 rbc (1b) busulfan group c. papaya+ busulfan group normal group carica papaya carica papaya carica papaya carica papaya *** ### ** 45 16 .6 7 66 66 .6 7 81 16 .6 7 tlc (2a) normal group busulfan group c.papaya+ busulfan group *** ### 62 7. 83 88 1. 08 95 8. 92 plt (2b) normal group busulfan group c.papaya+ busulfan group carica papaya carica papaya 33 effect of carica papaya leaf juice on blood cell count of busulfan-induced chronic bone marrow aplasia.. neutrophils lymphocytes monocytes eosinophils ** ### 8 8 8 .7 5 3 0 9 7 .6 7 4 6 3 7 .4 1 3 4 6 8 .4 3 3 5 6 7 .6 3 3 3 9 3 .3 6 dlc normal group busulfan group c.papaya+ busulfan group carica papaya carica papaya carica papaya carica papaya carica papaya carica papaya carica papaya carica papaya carica papaya carica papaya carica papaya carica papaya carica papaya carica papaya carica papaya carica papaya p-value 34 effect of carica papaya leaf juice on blood cell count of busulfan-induced chronic bone marrow aplasia.. caricapapaya carica papaya carica papaya carica papaya carica papaya et al 35 effect of carica papaya leaf juice on blood cell count of busulfan-induced chronic bone marrow aplasia.. 01 dedicated to the researchers, clinicians & health care workers of pakistan bismillah hir rahman nir raheem. in the name of allah, the most beneficent and the most merciful message by the chairman shaikh zayed medical complex & chief editor of proceedings prof. mateen izhar prof. saadia s. alam (chairman & dean) (chief editor) our jan to march 2022 edition is being published as the 5th covid-19 wave with the omicron variant crests worldwide. rising infectivity, yet lesser mortality is its hallmark. we pray humanity is relieved of this deadly disease. we received hec y category for the 2nd year running alhumdolillah and saw the highest number of article submissions since its inception. a hundred and twelve articles on wide ranging topics were submitted and vetted as per hec guidelines. we have applied for adl and doaj indexation proceedings presents articles from medical, allied and dental health professionals and wide ranging basic and applied research. all articles have been judged on stringent international criteria of plagiarism and blind peer reviews. we have achieved expanded viewership, with researchers in diverse medical fields from different institutions publishing their research in proceedings our vision, building linkages with top journals and researchers both in the country and abroad “we venture forth” hec & pmc approved quarterly proceedings shaikh zayed medical complex vol. 36 (1) pcpb/24(94) p-121/3196 jan to mar 2022 editorial board patron in chief: prof. mateen izhar phd, mrcpath chairman & dean chief editor: prof. saadia shahzad alam phd external associate editor: dr. usman iqbal phd, associate editors: prof. ayesha humayun phd dr. adnan salim fcps members-national: prof. mamoon rashid frcs (sitara-e-eisaar) prof. nadira mamoon mrcpath prof. m. ashraf phd (tamgha-e-imtiaz) prof. m. arif nadeem (fcps) dr. sarah ghafoor phd members-international prof. renne koeffel phd (sui) prof.jane banaszak-holl phd (aus) prof. hisham al muhtaseb phd (ksa) dr. salma malik m.d (us) dr. sarwat shaheen m.d (us) dr. aliya asher mrcp (uk) dr. uzma nasim siddiqui fcps (aus) editors: dr. shahila jalil fcps dr. samira haque fcps dr. sadia maqsood mhpe dr. faraz bokhari m phil dr. noora hassan hezam alaqmer m phil mrs. saima mohsin msc consultant biostatician: mr. muhammad aasim m phil library & information sciences: mr. ihsan basit m phil dr.muhammad shahid soroya phd contents sibgha fatima, humaira gulnaz, syed hussain raza zaidi, saba saleem, zahra fatima, nabila kaukab medical students’ perception on learning anatomy online during covid-19 pandemic in an integrated modular system: comparison of online and face to face sgds & interactive lectures sindhu rehman, sobia ashraf, shahid mehmood, hafsa malik, rahat serferaz, saeed ahmed frequency of leukoerythroblastic picture & hematological parameters in covid-19 patients and association with disease severity almas iqbal, huma saleem, muhammad taqi efficacy of paediatric preinduction anxiety distraction techniques during oncologic procedures adila ashraf, shazia abid, m. b. jamil, naila mumtaz, syeda abida ahmed controlling post-partum hemorrhage using a novel technique of multiple sponge-holding-forceps applied along cervical canal maheen fatima, moneeb ashraf, urooj fatima, muhammad imran, amer hassan siddiqui, tafzeel fatima colebrookea oppositifolia anti arthritic potential vs methotrexate in pristane induced rat arthritis mavra fatima, ayesha younas, ayisha imran, asma nasir, nauman aslam malik, omar chughtai, akhtar sohail chughtai pseudobasophilia: a helpful screening tool in diagnosis of dengue hafiza hina pasha, uzma jabbar, ambreen anjum, aasia kanwal, chaman nasrullah, aisha bashir, hamid javed qureshi is coriandrum sativum hypolipidemic in alloxan induced diabetic rats? amna liaquat, mirza zeeshan sikandar, syed imran ali shah comparative study of déjà-vu and associated attributes among epileptics and non-epileptics amtul hafeez, abdul mudabbir rehan, zunera hakim, attiya munir, rabia naseer khan, aamna khokhar nigella sativa seeds protective ability in pyrazinamide induced hyperuricemia in mice nisar ahmed, amer hassan siddiqui, ambereen anwar, muhammad nauman shad, abdul karim therapeutic effect of berberine versus methotrexate on joint histopathology in a rat model of pristane-induced arthritis sana qanber abbasi, zahid bashir, shafeen zulfiqar, ghazal mansoor, qurat-ul-ain, sana javaid variations in symptomatology of migraine among local population of pakistan guidelines for authors page 1 8 14 19 26 31 35 39 44 49 56 62 the editorial board of proceedings of shaikh zayed medical complex, lahore is indebted to our honorable peer reviewers for their expert & timely reviews. advisoryboard basic & pre-clinical sciences prof. mateen izhar phd, mrcpath prof. sibgha zulfiqar m.phil prof. muhammad suhail m.phil prof. nasreen ehsan m.phil prof. tahira naseem m.phil dr. anwaar bashir m.phil dr. suleman dawood phd (uol) dr. nighat yasmeen phd (kemu) dr. saima chaudhary phd (uhs) medicine and allied prof. azeem taj fcps prof. abdul shakoor fcps prof. talha mahmud fcps prof. safoora aamir fcps dr. lubna riaz fcps dr. asfandyar fcps surgery and allied prof. ali rafique mirza fcps prof. muhammad ikram fcps prof. imran anwar frcs prof. shafqat mukhtar fcps dr. jamshaid rahim fcps dr. syed sajjad raza kazmi fcps diagnostic division prof. mona aziz fcps prof. saulat sarfraz fcps dr. amir khan frcr peer review board (national) prof. m. nauman ahmad ffarcs prof. naseem saud phd prof. m. ovais omer phd prof. shabbir bhatti phd prof. zahid niaz frcs prof. zamir ahmad phd prof. muhammad arif nadeem fcps prof. muhammad moin frcs prof. khalid mahmood phd prof. sadia chiragh m.phil prof. muhammad aslam fcps prof. abdul mannan m.s prof. naheed humayun fcps prof. farhat naz fcps prof. abdul hameed frcs prof. rafeea tafweez phd prof. inayat thaver phd prof. ashraf chaudhry fcps prof. kashif malik fcps prof. aftab turabi phd prof. uzma hussain fcps prof. muhammad pervaiz prof. fouzia shaukat fcps prof. maryam rashid phd prof. rabeia bilal phd prof. khwaja khursheed fcps prof. tanvir akhtar butt ms prof. naila akhtar fcps prof. sajida malik m.phil dr. farooq afzal frcs dr. saman shahid phd dr. muhammad shahzad phd dr. ali hussainy zaidi m.d dr. shahzad khuram akram phd dr. uzma malik fcps dr. ayesha mallick frcp dr. tehseen haider kazmi fcps dr. khalida ajmal m.phil dr. israr ahmed fcps dr. saima batool fcps dr. saleem muhammad rana phd dr. muhammad khurram habib fcps dr. saba riaz phd dr. raazia tasadduq phd dr. nageen hussain phd dr. saadia nosheen jan fcps dr. soumble zulfiqar phd dr. yasir abbas zaidi fcps dr. muhammad zeeshan sarwarmrcs dr. muhammad imran khokar mrcs dr. nabiha farasat m.phil peer review board (international) prof. renne koeffel phd (switzerland) prof. jane banaszak-holl phd (aus) prof. m. hisham al-muhtaseb phd (jordan) prof. mukhtiar baig phd (ksa) dr. sarwat shaheen m.d (usa) dr. zeeshan i. shaikh m.d (usa) dr. tanzeel qurat ijaz mrcp (uk) dr. hafiz sohail anjum fcps (ireland) dr. rafay azhar frcp (singapore) dr. mazhar nawaz m.d (usa) dr. ahmad azam malik phd (ksa) dr. ishtiaq ahmed fcps (ksa) dr. aamir omair phd (ksa) dr. sabina ahmed mir m.d (usa) dr. anita lamichhane m.d (nepal) dr. faiza durrani phd (uk) dr. waseem lodhi frcog (uk) dr. seerat zahra hammad m.s (ksa) dr. shafya shahid phd (u.s) dr. abdul waheed frcs (u.k) dr. humaira zareen fcps (ksa) dr. amira okud m.d (ksa) dr. asma ahmed mrcp (uk) dr. sabahat sabir mrcog (uk) dr. sarfaraz ahmad frcr (uk) file.indd dedicated to the researchers, clinicians & health care workers of pakistan hec & pmc approved bismillah hir rahman nir raheem. in the name of allah, the most beneficent and the most merciful message by the chairman shaikh zayed medical complex & chief editor of proceedings prof. mateen izhar prof. saadia s. alam (chairman & dean) (chief editor) we present our april to june 2021 edition in the month of ramadan 1442 h with prayers for the good fortune and wellbeing of all. with the covid 19 pandemic 3rd wave, humanity in general, and especially health workers are enduring an exhausting emotional and physical toll. it is a time to fortify our defenses against this deadly disease by all means possible including vaccinations. for proceedings 2020 was a year of achievements alhumdolillah with development of a new website at www.proceedings-szmc.org.pk of international standards supported by the ojs/pkp platform for online submission. during the year we got recognition from higher education commission of pakistan (hec). we thank them for reposing their confidence in us awarding “y” category, an honor indeed. we are now indexed by pmc, google scholar, crossref/doi pastic and pakmedinet. efforts are underway for international indexation. the april to june 2021 proceedings presents covid 19 community awareness data and wide ranging basic and applied research. all articles submitted for publication have been judged on stringent international criteria of plagiarism and blind peer reviews. we have achieved expanded viewership, with researchers in wide ranging medical fields from different institutions publishing their research in proceedings our vision, building linkages with top journals and researchers both in the country and abroad “we venture forth” hec & pmc approved proceedings shaikh zayed medical complex vol. 35 (2) pcpb/24(94) p-121/3196 april to june 2021 editorial board patron in chief: prof. mateen izhar chairman & dean chief editor: prof. saadia shahzad alam head, pharmacology & therapeutics external associate editor: dr. usman iqbal taipei medical university, taiwan associate editors: prof. ayesha humayun head, public health & community medicine prof. imran anwar head, surgery unit-ii dr. adnan salim assistant prof. gastroenterology sub editors: dr. uzma nasim siddiqui assistant prof. medicine dr. shahila jalil associate prof. histopathology prof. tahira naseem head, biochemistry & chemical pathology dr. samira haque associate prof. histopathology dr. lubna riaz assistant prof. pediatrics dr. sadia maqsood assistant prof. pharmacology dr. faraz bokhari associate prof. physiology dr.noora hassan hezam alaqmer physiology department mrs. saima mohsin incharge, nhrc consultant biostatician: dr. akram chaudhry mr. muhammad aasim library & information sciences: mr. ihsan basit dr. muhammad shahid soroya contents arif abdulmohsen al mousa, imtiaz ahmad qureshi, fahad anwar, nadeem shafique butt, marwan a bakarman,zohair jamil gazzaz,ahmad azam malik awareness and prevalence of risk factors of chronic kidney disease in rabigh, kingdom of saudi arabia: a community perspective mirza zeeshan sikandar, bilal husnain bajwa, hasan zahid, sobia ali, sabiha iqbal, syed imran ali shah knowledge, attitude, and practice trends towards covid-19 pandemic: comparison between individuals with medical and nonmedical backgrounds imran hanif, masooma ahmad, wardah anwar, maria ilyas, ambreen anjum, umair hanif, majid kaleem gender differences in the morphological severity of mitral stenosis in patients presenting in a tertiary care hospital qura-tul-ain, sidra mushtaq, naveeda manzoor, mahreen akhtar, anila errum, naseem saud ahmad comparison of xanthine oxidase inhibitory effect of swertia chirayita and febuxostat in vitro zoofishan qureshi, muhammad ashraf chaudhry, nayyer uzair, tariq mukhtar farani, uzair mumtaz, muhammad umer farooq high body mass index a risk factor of preeclampsia: a hospital based case-control study iram kamal, rizwan faisal, lubna amer, abdul mudabbir rehan, sadia chiragh effect of malus domestica (apple peel) extract on peripheral blood and bone marrow cells of prednisolone treated mice muhammad ali rafique mirza, ammara mazhar, waheed bhutto, abdul saeed khan effectiveness of amnion on chronic wounds vs saline dressing jawaria ghazanfar, aisha khalil, aniqua tahir, muhammad imran anwar, hina mahmood, ayesha humayun knowledge, attitude and practice of breast cancer screening in medical and non-medical females students of lahore, pakistan irfan sharif, athar mahmood, ghulam ghous, nabeel shafi, humayun saeed, muhammad shahzad anwar, muhammad farooq is upper pole puncture for percutaneous nephrolithotomy (pcnl) safe and effective? afifa waseem, muhammad suhail, tayyaba muzaffar, javed iqbal, hadia zulfiqar flax seed oil alleviates caffeinated energy drink induced metamorphosis in femur weight and relative tissue weight index of adult male albino rat alvia batool, maryam fatima, fozia farzana toxic effects of ribavirin on the testicular interstitium in albino rats anis farooq, haroon javaid, imran anwar a quality improvement project: development and implementation of a modified i-pass handover in the general surgery department of tertiary care hospital in pakistan muhammad ramzan, talha mahmud, muhammad saqib, abdul saeed khan, qazi abdul saboor, ali rafique mirza a comparative study between-small bore and large bore tubes for therapeutic pleural drainage muhammad ikram, amna javed, ayesha shahid, sara muneer, fariha tahir postpartum intrauterine contraceptive device (ppiucd) versus interval intrauterine contraceptive device (iucd) complications: which is better? usman aslam, muhammad ubaidullah khan, sadia chiragh, abdul karim, mushtaq ahmed effect of atorvastatin alone and in combination with aspirin on uric acid handling of normal rats tayyaba gul malik acute anterior uveitis (ocular manifestation of covid-19); a case report instructions for authors page 1 7 14 18 23 28 35 41 47 52 58 64 68 73 78 83 86 editorial advisory board basic & pre-clinical sciences prof. mateen izhar head, pathology division, szmc prof. sibgha zulfiqar head, physiology, szmc. prof. muhammad suhail head, anatomy, szmc. prof. nasreen ehsan head, forensic medicine, szmc dr. suleman dawood (uol) asst. prof. of allied health sciences dr. nighat yasmeen (kemu) asst. prof. of biomedical sciences dr. tania shakoori (kemu) asst. prof. of biomedical sciences dr. saima chaudhary (uhs) asst. prof. of oral pathology medicine and allied prof. azeem taj head, medicine, szmc. prof. abdul shakoor head, psychiatry, szmc prof. talha mahmud head, pulmonology, szmc prof. safoora aamir head, dermatology, szmc surgery and allied prof. mamoon rashid (sitara-e-eisaar) head, plastic surgery, shifa hospital, islamabad prof. ali rafique mirza professor of plastic surgery, szmc prof. muhammad ikram head, gynecology, szmc prof. shafqat mukhtar head, gynecology, szmc dr. jamshaid rahim associate professor urology, szmc dr. syed sajjad raza kazmi head, anesthesia & icu, szmc diagnostic division prof. nadira mamoon head, pathology, shifa hospital, islamabad prof. mona aziz head, hematology, szmc prof. saulat sarfraz head, radiology, szmc dr. amir khan asst. prof. radiology, szmc peer review board (international) prof. renne koeffel (switzerland) prof. m. hisham al-muhtaseb (jordan) prof. mukhtiar baig (ksa) dr. jane banaszak hall (usa) dr. dagmar stoiber sakaguchi (austria) dr. sarwat shaheen (usa) dr. zeeshan i. shaikh (usa) dr. aliya asher (uk) dr. tanzeel qurat ijaz (uk) dr. hafiz sohail anjum (ireland) dr. rafayazhar (singapore) dr. mazhar nawaz (usa) dr. ahmad azam malik (ksa) dr. ishtiaq ahmed (ksa) dr. aamir omair (ksa) dr. sabina ahmed mir (usa) dr. anita lamichhane (nepal) dr. faiza durrani (uk) dr. waseem lodhi (uk) dr. seerat zahra hammad (ksa) dr. shafya shahid (u.s) dr. madeeha afzal (u.k) dr. abdul waheed (u.k) dr. humaira zareen (ksa) dr. amira okud (ksa) dr. asma ahmed (uk) dr. sabahat sabir (uk) peer review board (national) prof. m. ashraf (tamgha-e-imtiaz) prof. arshad kamal butt prof. m. nauman ahmad prof. ziaullah prof. moeed iqbal qureshi prof. naseem saud prof. m. ovais omer prof. shabbir bhatti prof. zahid niaz prof. zamir ahmad prof. muhammad arif nadeem prof. muhammad moin prof. khalid mahmood prof. zujaja zaheer prof. muhammad aslam prof abdul mannan prof. naheed humayun prof. farhat naz prof. abdul hameed prof. rafeea tafweez prof. inayat thaver prof. ashraf chaudhry prof. kashif malik prof. aftab turabi prof. uzma hussain dr. maryam rashid dr. farooq afzal dr. saman shahid dr. sarah ghafoor dr. muhammad shahzad dr. ali hussainy zaidi dr. shahzad khuram akram dr. uzma malik dr. ayesha mallick dr. tehseen haider kazmi dr. khalida ajmal dr. israr ahmed dr. saima batool dr. saleem muhammad rana dr. muhammad khurram habib dr. paras khan dr. saba riaz dr. raazia tasadduq dr. nageen hussain dr. saadia jan dr. soumble zulfiqar dr. rabeia bilal dr. yasir abbas zaidi untitled-1 1 proceedings s.z.m.c. vol: 34(3): pp. 1-3, 2020. pszmc-753-34-3-2020 transfusion medicine practice during covid-19 pandemic: potential pitfalls and solutions for provision of blood products sarah tehseen saskatchewan health authority, canada ca. transfusion of blood components is an essential part of clinical medicine and can be significantly impacted by pandemics. therefore, hospital blood transfusion services must plan for the potential shortage of blood supply to continue the provision of this critical service in the face of emergencies.1 this article focusses on recommendations to ensure a safe and reliable supply of blood components. impact of a pandemic on the blood supply chain can be observed in multiple areas including the potential loss of donors, lack of reagent red cells or other materials needed for pre-transfusion testing, staff shortages due to illness and increased utilization by the hospital due to an increase in the number of sick patients.2 channels of communication: it is paramount that the blood transfusion services of each hospital review its inventory and communicate with the blood suppliers frequently to review the status of blood availability. frequent communication with patient care providers (anesthesia, oncology, medicine, etc.) to estimate the daily need for blood products is also critical. it is also crucial for each hospital to minimize blood utilization where possible and develop contingency plans for severe shortages.3 reducing hospital utilization: a large volume of high-quality data both in adults and pediatrics strongly favors restrictive transfusion thresholds in hospitalized patients in the absence of active hemorrhage, hemolysis or trauma.4,5,6 enforcement of these restrictive thresholds (table1a & 1b) will ensure a reduction in blood utilization and maintain blood inventory.7 this can be done by review of transfusion orders by the blood bank (table-2) as well as the widespread education of healthcare providers with a focus on addressing the cause of underlying anemia (table-1a). also, postponing elective procedures and surgeries will help reduce blood utilization during an active pandemic.3 other considerations include blood conservation strategies like normovolemic hemodilution and the use of cell savers during surgery. optimization of surgical control of bleeding and the use of antifibrinolytic agents such as tranexamic acid also plays an important role in reducing the need for blood transfusion.8,9 massive hemorrhage (>4.5 liters of blood loss in 30 minutes or 150 ml/min of blood loss or > anticipated need of 3 or more red cell units in 3 hours) requires fast delivery of multiple blood components and can deplete inventory quickly.18 it is recommended to initiate a conversation with the healthcare providers early in the situation of a massive hemorrhage to anticipate the need for blood components. switch to type-specific blood as soon as possible to protect the o negative inventory and consider the use of antifibrinolytics and coagulation factor concentrates such as fibrinogen and prothrombin concentrates if available.10 staff and testing supply shortages: shortage of staff personnel and testing supplies in the light of an ongoing pandemic must be considered. hospitals should have contingency plans for staff coverage should many technologists fall sick. remote working should be considered for medical directors whenever possible. at this time there are no specific recommendations for protective equipment for blood bank laboratory technologists beyond handwashing, social distancing, and universal precautions however in the light of the dynamic situation of the current pandemic, these may be reconsidered.11 hospitals should also have contingency plans in light of a shortage of testing reagents. for example, if testing reagents for pre-transfusion testing fall short, consider limiting to basic testing such as abo, rh and cross-matching for transfusions. specific issues in developing countries: pakistan lacks a centralized system or coordinated blood transfusion service leading to wide heterogeneity in terms of availability of blood 2 transfusion medicine practice during covid-19 pandemic components, the sufficiency of blood supply, presence and educational level of medical directors and technologists as well as infrastructure and other resources.12 while the blood supply for patients with hemoglobinopathies come from volunteer donors, most rural and small city clinics and hospital rely on whole blood donation from patients’ relatives and friends in cases of surgeries or other procedures which raises significant questions about the quality and safety of blood components. it is, therefore, critical that hospital systems providing care at various levels, collaborate to achieve the following: 1. sharing blood inventory details across different hospitals and providing components to centers in demand. 2. formulating and enforcing restrictive transfusion strategies and adequate hemorrhage control 3. contingency planning for shortages in staff and testing reagents with a delineated centralized hospital where specimens may be referred for advanced transfusion testing if needed blood collection perspective: given the anticipated increase in need of blood for patients, there should be national and provincial level initiatives for blood donations, especially to maintain the blood supply for patients with hemoglobinopathies. this is complicated in the light of social distancing recommendations which are crucial to prevent the spread of covid-19 infections. donor centers should consider the following measures for donor and staff protection 1. screen donors with questions regarding cough and fever before accepting them for donation to prevent droplet/airborne spread of the virus in collection centers. 2. avoiding crowding by maximizing donor appointments, staggering appointments, and offering donors the opportunity to wait in their cars or other convenient locations. 3. discouraging children & others from accompanying donors to blood drives. 4. using signs outside blood drives asking individuals at elevated risk for infection (due to travel, illness or contact with sick individuals) not to enter/donate. 5. donors and staff wear surgical masks.13 there is no current evidence that sars-cov2 is transmissible by blood however the measures mentioned above are crucial to prevent the spread of infection among donors and staff personnel performing blood collection. do not transfuse for anemia if other therapies or observation is effective limit transfusion to one unit at a time in a stable nonbleeding patient avoid plasma transfusion for a mildly elevated inr (<1.8) in a nonbleeding patient or before a procedure for inr correction do not order pre-transfusion testing for all preoperative patients unless significant bleeding is anticipated limit transfusion of o negative units to women of childbearing potential for emergencies and to patients who have o negative blood type table-1a: recommendations for red cell transfusion based on choosing wisely canada14,15,16 hemoglobin >9 g/dl mostly inappropriate unless special circumstances. hemoglobin 8-9 g/dl transfusion likely inappropriate unless symptomatic anemia (tachycardia, cns symptoms, hypotension). hemoglobin 7-8 g/dl one unit appropriate for transfusion in cardiac patients. hemoglobin <7 g/dl usually appropriate to transfuse 1 rbc or whole blood unit followed by checking hemoglobin unless: 1. the patient has chronic iron deficiency anemia (ida) where iv iron is a better alternative. 2. young/pediatric patient. hemoglobin <6g/dl transfusion appropriate unless chronic ida. table-1b: restrictive transfusion thresholds for nonbleeding stable patients17 >1 red cell/whole blood unit for orders outside of operating room (or) >4 red cell units requested / case for surgeries >1 platelet unit requested outside the or initiation of discussion between tm physician/blood bank and patient care provider in cases of massive hemorrhage to anticipate need for blood components table-2: criteria for prospective order review to conserve inventory by the transfusion medicine physician or lab technologists13 conclusion the covid-19 pandemic can have a significant impact on the ability of hospitals and blood systems to provide life-saving transfusions. in this light, preparation by enforcing restrictive transfusion thresholds, postponing elective procedures, physician education and contingency planning for staff and testing supply shortages can go a long way towards mitigating the adverse effects of blood shortages. the most crucial step in these measures is frequent communication between direct patient care providers and blood banks to review inventory and 3 transfusion medicine practice during covid-19 pandemic anticipate the need for blood products. this communication also needs to happen between hospitals in cities and provinces to address shortages in blood components. references 1. world health organization. maintaining a safe and adequate blood supply during pandemic influenza. available at: https://www.who.int/bloodsafety/publications/ who_guidelines_on_pandemic_influenza_and _blood_supply.pdf 2. zimrin, ann b., and john r. hess. "planning for pandemic influenza: effect of a pandemic on the supply and demand for blood products in the united states." transfusion 47.6 2007):1071-79 3. pagano, monica b., et al. "prepare to adapt: blood supply and transfusion support during the first 2 weeks of the 2019 novel coronavirus (covid-19) pandemic affecting washington state." transfusion (2020). 4. hébert, paul c., et al. "a multicenter, randomized, controlled clinical trial of transfusion requirements in critical care." new england journal of medicine 340.6 (1999):409-17 5. valentine, s. l., et al. "pediatric critical care transfusion and anemia expertise initiative (taxi)." pediatric critical care blood research network (blood net), and the pediatric acute lung injury and sepsis investigators (palisi) network pediatrcrit care med 19 (2018): 884. 6. holst, lars b., et al. "restrictive versus liberal transfusion strategy for red blood cell transfusion: a systematic review of randomized trials with meta-analysis and trial sequential analysis." bmj 350 (2015): h1354. 7. carson, jeffrey l., et al. "red blood cell transfusion: a clinical practice guideline from the aabb." annals of internal medicine 157.1 (2012): 49-58. 8. henry, david a., et al. "anti-fibrinolytic use for minimizing perioperative allogeneic blood transfusion." cochrane database of systematic reviews 3 (2011). 9. huët, charlotte, et al. "a meta-analysis of the effectiveness of cell salvage to minimize perioperative allogeneic blood transfusion in cardiac and orthopedic surgery." anesthesia & analgesia 89.4 (1999): 861. 10. dzik, walter h., et al. "clinical review: canadian national advisory committee on blood and blood products-massive transfusion consensus conference 2011: report of the panel." critical care 15.6 (2011): 242. 11. aabb townhall series: prepare to adapt. the washington state experience during covid-19 pandemic. pagano m. march 31st, 2020. 12. haddad, antoine, et al. "how to manage transfusion systems in developing countries: the experience of eastern and southern mediterranean countries." transfusion medicine (2020). 13. aabb townhall series: collection considerations during covid-19 pandemic. vassallo r, april 2nd, 2020 14. retter, andrew, et al. "guidelines on the management of anaemia and red cell transfusion in adult critically ill patients." british journal of haematology 160.4 (2013): 445-464. 15. carson, jeffrey l., et al. "transfusion thresholds and other strategies for guiding allogeneic red blood cell transfusion." cochrane database of systematic reviews 10 (2016). 16. hillis, christopher m., et al. "the canadian choosing wisely campaign: the canadian hematology society’s top five tests and treatments." annals of hematology 94.4 (2015): 541-545. 17. screening algorithm for red cell transfusions. saskatchewan health authority saskatoon canada. 18. savage, stephanie a., et al. "the new metric to define large-volume hemorrhage: results of a prospective study of the critical administration threshold." journal of trauma and acute care surgery 78.2 (2015): 224-230. the authors: dr. sarah tehseen saskatchewan health authority, canada ca. email: sarah.tehseen@gmail.com 08 35 proceedings s.z.m.c. vol: 36(1): pp. 35-38, 2022. pszmc-831-36-1-2022 coriandrum sativum 1department of physiology, shalamar medical & dental college, lahore 2department of biochemistry, fmh college of medicine & dentistry, lahore 3department of physiology, alaleem medical college, lahore 4department of physiology, ucmd, university of lahore 5department of physiology, akhter saeed medical & dental college, lahore coriandum sativum coriandrum sativum coriandrum sativum coriandrum sativum, coriandrum sativum l coriandrum sativum coriandrum sativum 36 is coriandrum sativum hypolipidemic in alloxan induced diabetic rats? oriandrum sativum coriandrum sativum coriandrum sativum coriandrum sativum oriandrum sativum coriandrum sativum 37 is coriandrum sativum hypolipidemic in alloxan induced diabetic rats? coriandrum sativum coriandrum sativum coriandrum sativum coriandrum sativum coriandrum sativum coriandrum sativum coriandrum sativum coriandrum sativum coriandrum sativum coriandrum sativum lanneaedulis coriandrum sativum coriandrum sativum 38 is coriandrum sativum hypolipidemic in alloxan induced diabetic rats? untitled-1 29 proceedings s.z.m.c. vol: 34(3): pp. 29-34, 2020. pszmc-758-34-3-2020 role of prophylactic dimenhydrinate versus ondansetron as an adjunct to dexamethasone on post-operative nausea and vomiting in laparoscopic abdominal procedures 1sadaf bukhari, 2aaifa khalid niazi, 1syed mahmood ali, 2maria arshad, 3muhammad taqi 1department of anesthesia, shaikh zayed medical complex, lahore 2department of anesthesia, jinnah hospital, lahore 3department of anesthesia, gulab devi hospital, lahore abstract introduction: post-operative nausea and vomiting (ponv) is a common complication in patients undergoing surgery. anti-emetic prophylaxis is recommended for all high risk patients. combination therapy is considered superior to single dose prophylaxis however its efficacy in varied laparoscopic abdominal surgical situations has not been investigated. aims & objectives: to compare the post-operative nausea and vomiting with dimenhydrinate plus dexamethasone versus ondansetron plus dexamethasone in patients following laparoscopic abdominal procedures. place and duration of study: operation theatre, shaikh zayed hospital lahore, for one year (1st april 2015 – 31st march 2016). material & methods: 464 patients were placed in two groups of 232 each. group a received prophylaxis with dexamethasone 8 mg i/v + dimenhydrinate 50mg i/v and group b received prophylaxis with dexamethasone 8 mg i/v + ondansetron 4 mg i/v. efficacy was judged with absence of nausea and vomiting in first 24 hours after surgery. data was analyzed in spss version 27. results: the frequency of nausea was 6% with dimenhydrinate while was 9% with ondansetron. similarly, vomiting was 18% vs. 24% respectively. the efficacy of dimenhydrinate was 88% while and ondansetron achieved efficacy in 65% cases, which was statistically significant (p<0.05). conclusion: the combination dexamethasone & dimenhydrinate was more efficacious in preventing ponv than the combination dexamethasone & ondansetron. key words: post-operative nausea and vomiting, antiemetic prophylaxis, dexamethasone, dimenhydrinate, ondansetron introduction post-operative nausea and vomiting (ponv) is the most disturbing and a very commonest problem often after surgery due to anesthesia. ponv may lead to severe patient’s anxiety, requirement of nursing care increases, delay in ambulation, also increases pain and in few severe cases, it also badly effects the outcome of the surgery as well as disturb the hemodynamics of patients.1,2 it is a troublesome complication and has been the subject of interest of researchers for the past many years.3 ponv is an unpleasant emotional experience that provokes both fear and anxiety in the patient. it might instill in the memory of patient for life time. in addition to the fear that it instills in patients, ponv can lead to serious medical complications.4 ponv occurs in 20-30% of patients undergoing surgery.5 in high risk patients who receive no prophylaxis the incidence of ponv goes up to 7080%.6,7,8 effective prevention of ponv improves patient satisfaction ratings. all these factors make the prevention and management of ponv a challenge for most anesthesiologists.9 the optimal control of ponv is based on three steps. firstly high risk patients at risk for ponv need to be identified. secondly such an anesthetic technique should be employed which uses low emitogenic agents and favors use of antiemetogenic agents. thirdly all patients at risk for ponv should receive antiemetic prophylaxis.6 antiemetic prophylaxis can be provided by pharmacological and non-pharmacological methods. pharmacological methods include a variety of drugs from different classes.10,11,12 combination therapy of more than one drug is considered far more efficacious as compared to single drug therapy. drugs with different mechanisms of action should be used in combination.13,14 this multi modal approach has 30 role of prophylactic dimenhydrinate versus ondansetron as an adjunct to dexamethasone shown better results when compared with single mode of action prophylaxis.14,15,16,17,18 the novelty of our study is this, that it's been done in the face of many risk factors of ponv. where laparoscopy itself is a risk factor for ponv and in the presence of other ponv risk factors (females, use of nitro oxide, use of opiods, use of volatiles). we have tried to demonstrate dexamethasone plus dimenhydrinate combinations' substantial effectiveness which is also very cost effective as per requirement in our country. other studies are done on tonsillectomy, breast surgeries, and laparoscopic cholecystectomies but not on all types of laparoscopic abdominal procedures. this being another attribute of our study as we have conducted this study in various abdominal laparoscopic surgeries, not just one. so keeping in mind ponv and its challenging outcomes, an effective drug combination is the need of the hour particularly in high risk cases. material and methods after taking irb permission no: 1371, informed consent of 464 patients, 18 years age and above, asa i & ii, undergoing abdominal laparoscopic surgery was obtained in pre-op area of shaikh zayed hospital lahore, over a period of one year. the patients were divided into two groups a & b by lottery method. i/v lines were secured: group a patients were given injection dexamethasone 8mg i/v and inj dimenhydrinate 50mg i/v. the patients in group b were given inj. dexamethasone 8mg and inj ondansetron 4mg i/v. in the operation theatre standard monitoring was done. injection ringer’s lactate(r/l) was given by 4,2,1 formula respective of their weights. all the patients were given standard general anesthesia. after pre-oxygenation using 100% oxygen, anesthesia was given along with injection fentanyl 1-2mcg/kg body weight followed by injection propofol 1-2mg/kg body weight. inj atracurium was given 0.5mg/kg body weight, patient was ventilated for 3 minutes and endotracheal tube was inserted in all patients. mechanical ventilation was done. anesthesia was sustained with 50% oxygen and 50% nitrous with 1% isoflurane. intraoperatively, the muscle relaxation was attained by adding incremental doses of atracurium i.e. 1mg/kg on sos basis. after surgery, anesthesia was stopped and remaining neuromuscular blockade was provoked by applying neostigmine injection along with glycopyrrolate. to minimize the effects of confounders standardized dose of opioid was used. also adequate, standardized hydration was done using ringer lactate with the 4,2,1 formula. inj fentanyl upto 50mcg i/v was permitted as need for management of post-op pain. after procedure patients were shifted to post anesthesia care unit (pacu) where they were monitored for 2 hours and then to ward where they were monitored for 24 hrs by pacu doctor and nurse on duty. statistical analysis: data was fed into spss version 17 and analyzed using this program. chi square test was used to compare the efficacy in both groups keeping pvalue 0.05 as significant. results the mean age was 45.65±9.37 years. in group a, mean age was 45±9 and in group b was 46±9 years. in group a, nausea was observed in 14 (6%) cases while in group b, nausea was present in 42 (18%) cases (p<0.05). within first 2 hours, nausea was observed in 14 (6%) cases in group a while in 27 (11%) cases in group b (p<0.05) while doing first 24 hours, nausea was observed in 0 (0%)cases in group a while in 15 (6%) cases in group b (p<0.05). similarly, in group a, vomiting was observed in 21 (9%) cases while in group b, vomiting was present in 55 (24%) cases (p<0.05). within first 2 hours, vomiting was observed in 14 (6%) cases in group a while in 25 (11%) cases in group b (p>0.05) while doing first 24 hours, vomiting was observed in 7 (3%) cases in group a while in 30 (12.9%) cases in group b (p<0.05). efficacy was achieved in 204 (88%) cases in group a while in 152 (65%) cases in group b. the difference between both group was significant (p<0.05). (table-1) dimenhydrinate ondansetron p-value age (years) 45±9 46±9 nausea yes 14(6%) 42(18%) <0.001 no 218 (94%) 190(82%) nausea at 2 hrs 14(6%) 27 (11%) 0.033 nausea at 224 hrs. 0(0%) 15(6%) 0.0001 vomiting yes 21(9%) 55(24%) <0.001 no 211 (91%) 177(76%) vomiting in first 2 hrs. 14(6%) 25(11%) 0.066 vomiting in 2-24 hrs. 7 (3%) 30 (12.9%) 0.0001 efficacy achieved yes 204(88%) 152(65%) <0.001 no 28(12%) 80(35%) table-1: characteristics of patients 31 role of prophylactic dimenhydrinate versus ondansetron as an adjunct to dexamethasone discussion ponv is one of the most common, heavy on the pocket and debilitating complication of general anesthesia. ponv occurs in 20-30% of patients undergoing surgery.19 in high risk patients for ponv who receive no prophylaxis the incidence of ponv goes up to 70-80%.4,8 although ponv is typically self-limiting, lasting less than 24 hours, the sequences are grave. it can lead to dehydration, electrolyte imbalance, metabolic alkalosis, gastric erosions and aspiration of gi contents with subsequent respiratory problems.20 it can also delay oral nutrition and oral drug intake. ponv, thereby delays convalescence and increases hospital stay and cost.21 the incidence of ponv depends upon the presence or absence of certain risk factors. factors that may play a role in development of ponv include sex, age, previous history of ponv, motion sickness and migraines, smoking status, type of surgery, duration of surgery, type of anesthesia, use of opioid analgesics and hydration during surgery.22,23 these factors can be broadly categorized to “patient factors” which includes females, people who do not smoke, past history of ponv/motion sickness, “anesthetic factors” like volatile anesthetic gases, nitrous oxide and use of perioperative opioid analgesics and lastly “surgical factors” that include type of surgery and duration of surgery.10 the optimal control of ponv is based on three steps. firstly high risk patients at risk for ponv need to be identified. secondly such an anesthetic technique should be employed which uses low emitogenic agents and favors use of antiemetogenic agents. thirdly all patients at risk for ponv should receive antiemetic prophylaxis.6 ponv must be controlled effectively especially in high risk patients. combination therapy of more than one drug is considered far more efficacious as compared to single drug therapy due to its multifactorial origin. therefore, drug combinations should be used which comprise of drugs that have different mechanisms of action.24,25 this multi modal approach has shown better results when compared with single mode of action prophylaxis. the combinations widely used include 5ht-3 receptor antagonist + dexamethasone, 5ht-3 receptor antagonist + droperidol, droperidol + dexamethasone, dimenhydrinate + dexamethasone.26,27,28 despite the presence of a plethora of publications on ponv only little is known on how to treat it. our study is another effort to know how to control this distressing ponv, best. in our study a clinically relevant question about the efficacy of two combination drug regimens was asked. with the knowledge that all these drugs (among them dexamethasone and ondanstron which are considered as gold standard for ponv and a relatively newer drug, dimenhydrinate, which is more widely available and very cost effective) reduce ponv with similar efficacy in patients at high risk, the better of two combinations was sought out. multimodal interventions are required to control ponv in high risk patients. dexamethasone is a good drug for combination therapy. it effectively reduces ponv of delayed onset that occurs after 12 hours. when combined with a short acting agent which reduces ponv in the short term (0-12 hrs.) the risk of ponv is reduced both in the short term (0-12 hrs.) and in the long term (12-24 hrs). the combination dexamethasone plus ondansetron is a popular one which has been studied extensively and has shown very good results. ondansetron is effective in early ponv while dexamethasone controls late ponv. however there are not many studies that have compared it with other combination. dimenhydrinate is a cheap and widely available drug. it has been shown to be effective in the management of ponv. it hasn’t gained as much popularity due to its side effects, sedation, and dry mouth. in theory it should make a good combination with dexamethasone for the same reasons as ondansetron. both of them are effective in prevention of early ponv. voight et al.,18 tried the combination and found results which are equivalent to the combination of dexamethasone plus ondansetron similar to our study. however no studies have proved superiority of one combination to another which our study hoped to do, in a ponv prone environment. all patients received a standardized anesthetic protocol. nitrous oxide was used as an analgesic. although nitrous oxide is a risk factor itself for ponv, it reduced the intraoperative usage of opioids which are a stronger risk factor for ponv. hydration and opioid doses were standardized to minimize confounding. aftab et al.,29 conducted a study of risk factor assessment for ponv in dow medical university karachi, pakistan. they concluded that ponv was significantly higher in females, patients with h/o ponv or motion sickness and patients undergoing laparoscopic and eye surgery. our study has addressed two of these risk factors among others 32 role of prophylactic dimenhydrinate versus ondansetron as an adjunct to dexamethasone and observed the effects of anti-emetics in the high risk population. in india, in year 2014, bendre and associates, in accordance to our study determined incidence nausea/vomiting, early and delayed with ondansetron and dexamethasone. their results demonstrated for early nausea 26.7% vs our study 11%. for delayed nausea their result was 23.3% vs our 6%. for early vomiting they calculated incident of 10% vs our 24% and for delayed vomiting their 6.7% vs our 18%.30 voigt and associates (2011) carried out a randomized, double blinded, placebo controlled trial. their results are almost same as of our study. one of their study group received dimenhydrinate and dexamethasone, half of this groups' patients received total intravenous anaesthesia (tiva) and half of them received volatile anesthesia. they recorded incidence of nausea, emesis or both, in early 0-2 hours and late 2-24 hours post operatively, similar to us. their result showed significant reduction of ponv 11.4% with volatiles. our results showed that the combination of dexamethasone and dimenhydrinate was significantly better in preventing nausea and vomiting than the ondansetron and dexamethasone combination. we observed that dexamethasone and dimenhydrinate showed an efficacy of 88% which is consistent to voight et al.18 like us dabbous and fellows (2010) did a randomized study on elective laparoscopic surgeries where they compared efficacy of the combination of dexamethason and ondanstron in prevention of ponv. during 24 hours post operatively dabbous et al observed that no patient had vomiting (100%) compared to our 97.6%.31 imam et al studied the effects of ondansetron 4 mg plus dexamethasone with ondansetron 4 mg alone and placebo in patients undergoing general anesthesia for cesarean section. episodes of nausea and vomiting were observed in the first 24 hours of extubation. in the combination group 5% of the patients had ponv. in ondansetron alone group 22.5% of the patients had ponv and in the placebo group 52.5% of patients required antiemetics. the study showed that ondansetron and dexamethasone is more effective in preventing ponv with minimal adverse effects,32 which is comparable to our study. bano and associates studied the effects of dexamethasone plus ondansetron with dexamethasone alone in patients undergoing laparoscopic cholecystectomy. they found that the combination of dexamethasone plus ondansetron is more effective in preventing postoperative nausea and vomiting (p 0.035), close to our <0.05.17 kothari at al., results are contrasting to ours. they conducted a randomized trial, in patients undergoing laparoscopic cholecystectomy. they observed anti-emetic efficacy of dimenhydrinate vs ondansetron. of the patients receiving ondansetron post-op vomiting occurred in 6% versus our 24% and nausea in 42% vs our 24% of the patients. while of the patients who received dimenhydrinate 29% vs our 9% of them vomited and 34% vs our 42% of them had nausea. concluding that ondansetron was superior in controlling vomiting than dimenhydrinate, in contrast to our results. the discrepancy probably due to our use of these drugs in combination with dexamethasone while kothari gave them alone.33 contrary to our results piwko c & associates showed that for vomiting the ondansetron group had an efficacy of 45.3% vs the dimenhydrinate group's 38.2% while ours was 75% for ondansetron vs a 91% in dimenhydrinate group. piwko evaluated prevention of vomiting only in children whereas our study population were adults. another factor leading to this dis-similarity in result could be due to the difference of surgical procedure, piwko chose strabismus surgery while our interest was in abdominal laparoscopic surgeries.34 conclusion our research suggests the combination of dexamethasone and dimenhydrinate is more efficacious and also more cost effective than the combination of dexamethason and ondansterone. this study can be done on a much larger scale including all kinds of surgeries and asa categories. in light of its promising results dexamethasone and dimenhydrinate combination seems to be an excellent arsenal for an anesthetic for better patient care against ponv. limitations: ponv was considered as equivalent condition regardless of the surgical procedure duration, or length of anesthesia. therefore, future studies can be done by standardizing the dose and checking its efficacy on different anesthesia lengths. a further limitation was that although we recorded side effects of the drugs used but we did not include them in our studies. in future we can do the same study but instead of ponv we can observe side effects of these drugs. 33 role of prophylactic dimenhydrinate versus ondansetron as an adjunct to dexamethasone references 1. cohen mm, duncan pg, deboer dp, tweed wa. the postoperative interview: assessing risk factors for nausea and vomiting. anesthesia and analgesia 1994; 78(1):7-16. 2. gan tj, diemunsch p, habib as, kovac a, kranke p, meyer ta, et al. consensus guidelines for the management of postoperative nausea and vomiting. anesthesia & analgesia. 2014; 118(1):85-113. 3. miller rd, eriksson li, fleisher la, wienerkronish jp, cohen nh, young wl. miller's anesthesia e-book: elsevier health sciences; 2014. 4. apfel cc, philip bk, cakmakkaya os, shilling a, shi y-y, leslie jb, et al. who is at risk for postdischarge nausea and vomiting after ambulatory surgery? anesthesiology: the journal of the american society of anesthesiologists. 2012; 117(3):475-86. 5. gustafsson u, scott m, hubner m, nygren j, demartines n, francis n, et al. guidelines for perioperative care in elective colorectal surgery: enhanced recovery after surgery (eras®) society recommendations: 2018. world journal of surgery. 2019; 43(3):659-95. 6. cao x, white pf, ma h. an update on the management of postoperative nausea and vomiting. journal of anesthesia 2017; 31(4):617-26. 7. d'souza n, swami m, bhagwat s. comparative study of dexamethasone and ondansetron for prophylaxis of postoperative nausea and vomiting in laparoscopic gynecologic surgery. international journal of gynecology & obstetrics. 2011; 113(2):124-7. 8. apfel cc, korttila k, abdalla m, kerger h, turan a, vedder i, et al. a factorial trial of six interventions for the prevention of postoperative nausea and vomiting. new england journal of medicine. 2004; 350(24):2441-51. 9. kranke p, eberhart lh. postoperative nausea and vomiting: rational algorithms for prevention and treatment based on current evidence. anasthesiologie, intensivmedizin, notfallmedizin, schmerztherapie: ains 2009; 44(4):286-94; quiz 95. 10. sbaraglia f, saviani m, timpano jm, rossi m. postoperative nausea and vomiting as a cause of tracheal injury: an underestimated lifethreatening adverse event? british journal of anaesthesia. 2019;123(3):e457-e8. 11. helms ra, quan dj. textbook of therapeutics: drug and disease management: lippincott williams & wilkins; 2006. 12. chisholm-burns ma, wells bg, schwinghammer tl. pharmacotherapy principles and practice: mcgraw-hill; 2016. 13. rother c. post-operative nausea & vomitinguse of anti-emetic agents in anaesthesia. scottish universities medical journal 2012;1(1). 14. smith hs, cox lr, smith ej. 5-ht3 receptor antagonists for the treatment of nausea/vomiting. ann palliat med 2012; 1(2):115-20. 15. mihara t, tojo k, uchimoto k, morita s, goto t. reevaluation of the effectiveness of ramosetron for preventing postoperative nausea and vomiting: a systematic review and metaanalysis. anesthesia & analgesia 2013; 117(2):329-39. 16. som a, bhattacharjee s, maitra s, arora mk, baidya dk. combination of 5-ht3 antagonist and dexamethasone is superior to 5-ht3 antagonist alone for ponv prophylaxis after laparoscopic surgeries: a meta-analysis. anesthesia & analgesia. 2016; 123(6):1418-26. 17. bano f, zafar s, aftab s, haider s. dexamethasone plus ondansetron for prevention of postoperative nausea and vomiting in patients undergoing laparoscopic cholecystectomy: a comparison with dexamethasone alone. j coll physicians surg pak. 2008; 18(5):265-9. 18. voigt m, fröhlich cw, waschke kf, lenz c, göbel u, kerger h. prophylaxis of postoperative nausea and vomiting in elective breast surgery. journal of clinical anesthesia. 2011; 23(6):461-8. 19. hung l-w, chou m-y, liang c-k, liu k, chou y-m. being older as a risk factor for vomiting in those undergoing spinal anesthesia. journal of clinical gerontology and geriatrics. 2012; 3(2):68-72. 20. strong m, merritt ccm. pharmacology and antibiotics. surgical critical care and emergency surgery: clinical questions and answers. 2018: 97-108. 21. feinleib j, kwan lh, yamani a, davidson a. postoperative nausea and vomiting. uptodate waltham (ma): uptodate 2018. 22. hijazi em, edwan h, al-zoubi n, radaideh h. incidence of nausea and vomiting after fasttrack anaesthesia for heart surgery. brazilian journal of cardiovascular surgery. 2018; 33(4):371-5. 23. gwinnutt m, gwinnutt cl. clinical anaesthesia: john wiley & sons; 2016. 34 role of prophylactic dimenhydrinate versus ondansetron as an adjunct to dexamethasone 24. bayter m, peña p, marquez m, cárdenascamarena l, macias a, rubio j, et al. incidence of postoperative nausea and vomiting when total intravenous anaesthesia is the primary anaesthetic in the ambulatory patient population. ambul surg. 2018; 24:8-11. 25. kranke p, wilhelm w, eberhart l. management of postoperative nausea and vomiting (ponv). enhanced recovery after surgery: a complete guide to optimizing outcomes. 2020:195-202. 26. tjia i, dalton m, watcha mf. management of postoperative nausea and vomiting in pediatric patients. postoperative nausea and vomiting: a practical guide. 2016:119-30. 27. chen x, qin y, li s, lei h, wu x, shu l. efficacy of 5-ht3 receptor antagonists (ondansetron) vs dopamine receptor antagonists (droperidol) for preventing postoperative nausea, vomiting and headache: a meta-analysis. pteridines. 2019; 30(1):146-52. 28. bang sr, jin kim h, ahn ej, choi hr, kim kw, ko mj, et al. effect of total intravenous anesthesia and prophylactic 5-ht3 receptor antagonist on postoperative nausea and vomiting after gynecologic laparoscopic surgery: a prospective, randomized controlled study. rawal medical journal 2017; 42(1):73-7. 29. aftab s, khan ab, raza g. assessment of risk factors for postoperative nausea and vomiting. j coll physicians surg pak 2008; 18(3):137-41. 30. bendre r, karthik s, potli s, madhusudhana r. a comparative study of ondansetron with dexamethasone and granisetron with dexamethasone for prevention of postoperative nausea and vomiting following abdominal surgeries under general anaesthesia. wjpps 2015; 4(3):605-20. 31. dabbous as, jabbour-khoury si, nasr vg, moussa aa, zbeidy ra, khouzam ne, et al. dexamethasone with either granisetron or ondansetron for postoperative nausea and vomiting in laparoscopic surgery. middle east j anesthesiol 2010; 20(4):565-70. 32. imam sm, imam sm, mufti tm, bhatti ah, faridi ma. efficacy of balanced antiemesis for prophylaxsis againt post operative nausea and vomiting [ponv]: comparative study of ondansetron and dexamethasone versus ondansetron alone, pak. armed forces med j 2004; 54(2):185-90. 33. kothari s, boyd w, bottcher m, lambert p. antiemetic efficacy of prophylactic dimenhydrinate (dramamine) vs ondansetron (zofran). surgical endoscopy.2000;14(10):926-9. 34. piwko c, lasry a, alanezi k, coyte pc, ungar wj. economic evaluation of ondansetron vs dimenhydrinate for prevention of postoperative vomiting in children undergoing strabismus surgery. pediatric anesthesia.2005;15(9):755-61 the authors: dr. sadaf bukhari senior registrar, department of anaesthesia, shaikh zayed hospital, lahore. dr. aaifa khalid, senior registrar, department of anaesthesia, jinnah hospital, lahore. dr. syed mahmood ali associate professor, department of anaesthesia, shaikh zayed hospital lahore maria arshad senior registrar, department of anaesthesia, jinnah hospital, lahore. dr. muhammad taqi senior registrar, department of anaesthesia, gulab devi hospital, lahore. corresponding author: dr. sadaf bukhari senior registrar, department of anaesthesia, shaikh zayed hospital, lahore. e-mail: dr.sadaf.bukhari@gmail.com untitled-1 18 proceedings s.z.m.c. vol: 34(3): pp. 18-23, 2020. pszmc-756-34-3-2020 comparison of salivary cotinine levels of neonates born to mothers exposed to varying degrees of passive cigarette smoke 1wardah anwar, 1ambreen anjum, 2nasreen akhtar, 3maria anwar, 4tabinda kazmi, 5sibgha zulfiqar, 6faiza khan 1department of physiology, al-aleem medical college, lahore 2department of forensic medicine & toxicology, al-aleem medical college, lahore 3department of physiology, independent medical college, faisalabad 4department of physiology, niazi medical and dental college, sargodha 5department of physiology, shaikh zayed medical complex, lahore 6department of pharmacology, al-aleem medical college, lahore abstract introduction: obstetrical data reveals that fetuses born to mothers who are exposed to passive smoking show adverse health outcomes. aims & objectives: to compare the cotinine levels in neonates born to mothers exposed to varying degrees of passive cigarette smoke versus the control group i.e., mothers who are not exposed to smoking. place and duration of study: this study was conducted in obstetrics and gynecology department of shaikh zayed hospital, lahore and in emergency labor room of ganga ram hospital, lahore from december 2015 to may 2016. material & methods: this was a cross-sectional comparative study in which neonatal salivary cotinine level were measured and its relationship with birth weight was observed in 120 subjects, divided in four groups: .i) neonates whose mothers were not-exposed to smoke, neonates whose mothers were exposed to, ii) 1-5 cigarette, iii) 6-10 cigarettes and iv) more than 10 cigarettes per day. results: salivary cotinine levels were raised in all four groups with maximum levels seen in group four but there was no statistical difference between groups. changes in the birth weight were also seen with increased exposure to passive smoking. conclusion: salivary cotinine concentration in neonates increases due to passive exposure to cigarette smoke in mothers because of disregard of smokers to the adverse effects of nicotine inside homes and almost no governmental enforcement of laws regarding cigarette smoking in public spaces. key words: passive smoking, salivary cotinine, neonate, non invasive introduction cotinine is derived from breakdown of nicotine. 70-80% of nicotine is converted to cotinine in humans. this conversion occurs in two steps. in the first step cytochrome p450 system produce nicotineδ1'(5’)-iminium and 5’-hydroxynicotine.1 in the second step cytoplasmic aldehyde oxidase acts as a catalyst to produce cotinine.2,3,4 cotinine is measured in blood, hair, urine and saliva.5,6,7,8 its measurement is mostly preferred because of their noninvasive nature as compared to blood-derived assays.9,10 out of aforementioned techniques, saliva from neonates within 24 hours of birth has not been explored among noninvasive method in pakistan. most studies have found increasing level of cotinine with increasing level of self-reported exposure to tobacco smoke.11 however, not enough focus has been given to second hand smoking (shs) exposure to the neonate health. our study combines the quantitative nicotine exposure through cotinine level with the questionnaire feedback. measuring cotinine is preferred to measuring nicotine because nicotine has half-life of 2 hours whereas cotinine remains in the body longer and is preferred due to its longer half-life of 17 hours.12,13,14 in humans birth weight is affected by the development during intrauterine period.15 birth weight is very important and significant as it is associated with likelihood of mortality and morbidity. according to a report of unicef, neonatal birth weight is good indicator of mother’s health but also indicates the chances of survival of the baby.16 19 comparison of salivary cotinine levels of neonates born to mothers exposed to varying degrees of passive cigarette smoke at term that is 37+ weeks, normal birth weight should be 2500 4200 grams (5 lbs. 8 ounces till 9 lbs. 4 ounces). 1) birth weight less than 2500g, is low birth weight. 2) birth weight less than 1500g, is known as very low birth weight. 3) birth weight less than 1000g, is extremely low birth weight. out of these almost 2/3rd of the low birth weight infants are due to preterm birth and almost 1/3rd of these have growth restriction during intrauterine life.17 it has been well known that active smoking by mother is one of the environmental factor can decrease the birth weight of the infant markedly. there is an increase incidence of low birth weight infants born to smokers than to non-smokers.18,19 passive maternal smoking increases the likelihood of lowering the birth weight of new born. other environmental factors are a) lead poisoning b) combustion products of solid fuel c) carbon mono oxide d) mercury e) noise early morphological changes of the placenta are induced by cotinine, resulting in reduced volume of maternal intervillous space and also surface area of fetal capillaries. this leads to reduction in oxygen diffusion across the placenta.20the fetus suffers from chronic hypoxic stress as an outcome of smoking.21 these factors contribute to a reduced birth weight and length, and a smaller head circumference at birth.22,23 when normal placenta develops, its blood supply depends on the growth of maternal decidua and the fetal trophoblast. the trophoblastic cells undergo growth, maturation, migration and invade the decidual cells. if for any reason these migrations and penetration of cell is interrupted, it leads to less growth of fetus. it has been established that nicotine exposure has an endocrine like effect on the development of the placenta.24 exposure of nicotine obstructs the delivery of oxygen by causing vasoconstriction of all the blood vessels of body especially blood vessels of the umbilical cord. not only this but the red blood cells combine with car-boxyhemoglobin.25 the combination further reduces the oxygenation of tissues. as a result of decreased oxygenation of fetal tissues, the growth and development of baby is devastated. exposure of nicotine and its metabolites also cause direct damage to genetic material of fetus with impairment of lung development as well as increasing the heart rate and a decrease in breathing movement of fetus.26,27,28efforts are being made to avoid the incidence of lower than normal birth weight but these are not fruitful yet. there are other factors that increase the risk of infant mortality, the most important of which are socio-economic factors, medical risks before or during gestation and maternal lifestyles.29 keeping in view the potential damaging effects of nicotine exposure in mothers this study was conducted to compare the cotinine level of infants born to mothers who were divided in different groups. i) unexposed to cigarette, ii) exposed to 1-5 cigarettes, iii) 6-10 cigarettes and iv) more than 10 cigarettes per day. material and methods this study was conducted in obstetrics and gynecology department of shaikh zayed hospital, lahore and in emergency labor room of ganga ram hospital, lahore, after approval from the ethical committee and permission from the respective head of department. the study was conducted from december 2015 to may 2016. full term neonates, born to mothers aged (20 to 35 years), of same socioeconomic status and resident of lahore were included in the study. control group was neonates whose mothers were unexposed to passive smoking. other groups were of neonates born to mothers who were exposed to passive smoke. neonates of mothers with hypertension, respiratory diseases, cardiovascular disease, hiv infection and complicated pregnancy like anemia, poor weight gain were excluded. also the actively smoking mothers (cigarette, huqqa, cigar, etc.) or addicts to drugs, alcohol, pan and naswaar chewing were excluded. the sample size of 120 neonates divided into four equal groups (30 participant in each) was estimated by using 95% confidence level, 90% power of test with expected average cotinine levels (in nano grams per milliliter) of 0.077, 0.257, 0.466 and 1.013 for non-exposed, less than 05 cigarette per-day 6-10 per-day cigarette and more than 10 cigarette consumption respectively.30 a questionnaire was developed in order to collect information regarding the tobacco exposure in the study group. the questionnaire was selfadministered and answered by mothers at the time of saliva collection in the post-natal ward. personal data including name, age, and weight at first visit was recorded from their antenatal cards. detailed history and examination was performed and the birth weight was recorded on study performa used as study tool. 20 comparison of salivary cotinine levels of neonates born to mothers exposed to varying degrees of passive cigarette smoke saliva was taken from 120 neonates which were divided into four groups of 30 each according to the amount of maternal exposure to smoking i.e. i) nonexposed, ii) exposed to less than 5 cigarette per day, iii) exposed to 6-10 cigarette per day and iv) exposed to more than 10 cigarette per day each. the saliva of new born was collected within 24 hours of birth with sterile dropper. samples were collected at least 10 min before feeding and stored in eppendorf tubes. samples were kept cold in refrigerator and frozen at -20°c within 4 hours after collection, in order to avoid bacterial growth in the specimen. test was done by elisa technique in immunoassay laboratory, nhrc, lahore by calbiotech cotinine kit ,a solid phase competitive elisa. comparison between groups was made using one way anova followed by applying tukey's test where applicable. p-value of ≤0.05 for cotinine level and birth weight of neonate was considered significant statistically. statistical analysis: data was entered and analyzed by using spss version 20.0. results in this study, 120 pregnant females were enrolled and saliva from their neonates was tested. they were divided into four groups according to different levels of cigarette exposure. group 1 was considered as control group having neonates born to mothers without passive smoke exposure. the neonates in group 2 were of mothers who were exposed to less than 05 cigarettes per day. group 3 had neonates whose mothers were exposed to 05 to 10 cigarettes per day. while group 4 was of those neonates whose mothers had exposure to more than 10 cigarettes per day. fig-1: mean cotinine level (ng/ml) of all four groups with standard deviation. mean cotinine levels in group-1 was 1.0063 ± 0.70, in group-2 was 0.883 ± 0.68, in group-3 was 1.1537 ± 0.57 and in group-4 was 1.2520 ± 0.90. p-value of cotinine level was 0.216 which was not significant. mean birth weight (kg) in group-1 was 3.31 ± 0.50, in group-2 was 3.00 ± 0.28, in group-3 was 2.92 ± 0.48 and in group-4 was 2.66 ± 0.41. it was seen that birth weight was significantly different in all study groups as p-value was 0.001. mean birth weight of neonates showed inverse relation with shs exposure of the mothers. our results show that in group 1 and group 4 there was positive but insignificant pearson correlation between cotinine levels and birth weight that is (r=0.057, p-value= 0,767), (r=0.046, p-value=0.810) respectively. the pearson correlation in groups 2 and 3 showed insignificant correlation, having (r = 0.015, p-value=0.936) (r= -0.159, p-value=0.402) respectively. discussion mother and child are the two innocent members of our society, which suffer from the hazards of the passive/second hand smoke (shs). levels of shs during pregnancy can be measured by newborn salivary cotinine levels.31in our study we took the pregnant women of similar age and demographic characters, like all participants were residents of lahore for last one year. we made a questionnaire in which the data of mother and the baby was recorded specially the history of exposure to second hand smoke and the birth weight. we compared the maternal second hand exposure to cigarettes as reported in the questionnaire, with salivary cotinine levels and the neonatal birth weight. we also compared salivary cotinine levels of infants born to mothers who were not exposed to those who were exposed to indoor smoking during their pregnancy. it was seen that salivary cotinine level increased with the exposure to shs as noted in groups 3 and 4, as compared to groups 1 and 2. the relationship of group 1 and group 2 was unclear. group 1 cotinine levels did not show values aligned with study. the higher cotinine level detected in group 1 might be due to the fact that public smoking policy is not effectively implemented in pakistan. researches are also performed to assess the level of smoke in publically used areas like parks, transport and schools as these are the places where innocent victims get exposed to shs.32 as cotinine is not present in body normally and can only be detected due to exposure of nicotine. another study also reported cotinine to be detected in neonates and mothers who had no exposure of g-1: group 1 (control) g-2: group 2 (1-5 cigarettes) g-3: group 3 (6-10 cigarettes) g-4: group 4 (>10 cigarettes) 21 comparison of salivary cotinine levels of neonates born to mothers exposed to varying degrees of passive cigarette smoke cigarette at home hence they also infer that the exposure might be during visiting public places and they expressed it as the limitation of the study that exposure cannot be measured when visiting another house and in public areas.33 in our study indoor exposure of smoke during pregnancy was limited and evident from neonate weight but owing to traveling and recent exposure of shs, cotinine levels were overlapping with group 2. this group showed least exposure to indoor shs smoking, the similar findings were seen in another study in which cotinine was not detected in exposed neonates and it can be explained by difference in individual metabolism.34 cotinine levels in the saliva of neonates were effectively detected in all the groups and the birth weight showed decreasing trend with increasing exposure to shs. except group 1, the cotinine levels increased in rest of the studied groups according to the increase in shs exposure; 0.883 ng/ml, 1.153 ng/ml and 1.252 ng/ml in groups 2, 3 and 4 respectively. another fact about nicotine is that it remains on the surfaces of home, in the air and in the dust of smoking members and so these things get contaminated and the other members becomes passive smokers.35 in our study the mean birth weight (kg) in group-1 was 3.31 ± 0.50, in group-2 was 3.00 ± 0.28, in group-3 was 2.92 ± 0.48 and in group-4 was 2.66 ± 0.41. the birth weight showed inverse relation with the shs exposure, however while correlating with cotinine levels only groups 2 and group 3 showed inverse relationship with birth weight that was statistically insignificant. this might be due to small sample size. similar study to determine relationship between shs exposure and neonate’s birth weight was done in krakow using cord blood and it shows insignificant results when comparison was made with overall cord blood cotinine and birth weight but was significant when groups were compared.30 conclusion our study is one of the first in south asia whereby noninvasive technique of neonate saliva was used to determine correlation between maternal exposures to shs.maternal exposure to shs has inverse impact on neonate birth weight, which could have lasting effects on life of newborn baby. limitations: this study was a small scale study conducted on 120 cases. the results of cotinine levels are affected by many factors such as timing, the length of time since the last exposure as well as the exposure at public places and while travelling in public transport. the outdoor exposure is the one to which people are not aware, that they are being exposed and it is difficult to quantify it. recommendations: 1. large scale studies should be conducted particularly in south asian pregnant females exposed to shs so that more precise and comprehensive results regarding raised levels of cotinine and their inverse relationship with the birth outcome can be seen statistically. 2. awareness of shs hazards should be made public; especially pregnant mothers should be advised to take utmost care to avoid exposure not only to active smoking but also to shs. references 1. hecht ss, hochalter jb, villalta pw, murphy se. 2′-hydroxylation of nicotine by cytochrome p450 2a6 and human liver microsomes: formation of a lung carcinogen precursor. proceedings of the national academy of sciences of the united states of america. 2000; 97(23):12493-12497. 2. johnstone e, benowitz n, cargill a, jacob r, hinks l, day i, et al. determinants of the rate of nicotine metabolism and effects on smoking behavior. clinical pharmacology & therapeutics.2006; 80(4):319-330. 3. kandel db, hu mc, schaffran c, udry jr, benowitz nl. urine nicotine metabolites and smoking behavior in a multiracial/multiethnic national sample of young adults. american journal of epidemiology. 2007; 165(8):901-910 4. oscarson m. genetic polymorphisms in the cytochrome p450 2a6 (cyp2a6) gene: implications for inter individual differences in nicotine metabolism. drug metabolism and disposition. 2001; 29(2):91-95. 5. binnie v, mchugh s, macpherson l, borland b, moir k, malik k. the validation of self reported smoking status by analysing cotinine levels in stimulated and unstimulated saliva, serum and urine. oral diseases. 2004;10:287-93 6. figueiredo vc, szklo m, szklo as, benowitz n, lozana ja, casado l. determinants of salivary cotinine level: a population-based study in brazil. rev saudepublica. 2007;41(6):954-62 7. nishida n, yamamoto y, tanaka m, maeda k, kataoka k, nakayama k et al. association between passive smoching and salivary markers 22 comparison of salivary cotinine levels of neonates born to mothers exposed to varying degrees of passive cigarette smoke related to periodontitis. journal of clinical periodontology. 2006; 33(10):717-23. 8. nuca c, amariei c, badea v, zaharia ag, bucur l, nicolae c. salivary cotininebiomarker of tobacco consumption in the assessment of passive smoking prevalence. farmacia. 2012; 60(5):662-74. 9. srnt subcommittee on biochemical verification. biochemical verification of tobacco and cessation. nicotine and tobacco research. 2002; 4:149-159. 10. etter jf, vu due t, perneger tv. saliva cotinine levels in smokers and nonsmokers. american journal of epidemiology. 2000; 151(3):251-8. 11. bernert j.t. estimation of environmental tobacco smoke exposure during pregnancy using a single question on household smokers versus serum cotinine. journal of exposure analysis and environmental epidemiology. 2002; 12:1-10. 12. jarves m, feyerabend c, bryant a. passive smoking in the home. plasma cotinine concentration in non-smokers with smoking partners. tobacco control. 2001; 10:368-374. 13. figueiredo vc, szklo m, szklo as, benowitz n, lozana ja, casado l. determinants of salivary cotinine level: a population-based study in brazil. rev saudepublica.2007;41(6):954-62. 14. hawamdeh a, kasasbeh fa, ahmad ma. effects of passive smoking on children’s health: a review. eastern mediterranean health journal. 2003; 9(3):441-447. 15. melve k, skjaerven r .birth weight and perinatal mortality: paradoxes, social class and sibling dependencies. international journal of epidemiological association 2003; 32: 625-632. 16. https://www.unicef.org/specialsession/aboutpdf/15_lowbirthweight-d7341 17. tucker j, mcguire w. epidemiology of preterm birth. british medical journal. 2004;329:675-78. 18. vs knopik. maternal smoking during pregnancy and child outcomes: real of spurious effect? 2009; 34(1):1-36 19. salmasi g, grady r, jones j. environmental tobacco smoke exposure and perinatal outcomes: a systematic review and metaanalyses. acta obstetriciaetgynecologica scandinavica. 2010; 89(4):423-41. 20. bush pg, mayhew tm, abramovich dr, et al. a quantitative study on the effects of maternal smoking on placental morphology and cadmium concentration. placenta 2000; 21:247-56. 21. bush pg, mayhew tm, abramovich dr, et al. maternal cigarette smoking and oxygen diffusion across the placenta. placenta 2000; 21: 824-33. 22. hofhuis w, de jongste jc, merkus pj. adverse health effects of prenatal and postnatal tobacco smoke exposure on children. archives of disease in childhood. 2003; 88(12):1086-90. 23. robinson js, moore vm, owens ja, mcmillen ic. origins of fetal growth restriction. european journal of obstetrics & gynecology and reproductive biology. 2000; 92(1):13-9. 24. eric j, graham jb. morphological and biological effects of maternal exposure to tobacco smoke on the feto-placental unit. early human development. 2007; 83(11):699-706. 25. hukkanen j, gourlay sg, kenkare s, benowitz nl. influence of menstrual cycle on cytochrome p450 2a6 activity and cardiovascular effects of nicotine. clinical pharmacology & therapeutics. 2005; 77(3):159-69. 26. de la chica ra, ribas i, giraldo j, egozcue j, fuster c. chromosomal instability in amniocytes from fetuses of mothers who smoke. jama. 2005; 293(10):1212-22. 27. sherman j, young a, sherman mp, collazo c, bernert jt. prenatal smoking and alterations in newborn heart rate during transition. j obstetgynecol neonatal nurs. 2002; 31(6): 680-687. 28. sekhon hs, keller ja, benowitz nl, spindel er. prenatal nicotine exposure alters pulmonary function in newborn rhesus monkeys. american journal of respiratory and critical care medicine. 2001; 164(6):989-94. 29. de bernabé jv, soriano t, albaladejo r, juarranz m, calle me, martı́nez d, domı́nguezrojas v. risk factors for low birth weight: a review. eur j obstetgynaecolreprod biol. 2004; 116(1):3-15. 30. jedrychowski w, perera f, moroz e, edwards s, flak e, bernert jt. fetal exposure to secondhand tobacco smoke assesed by maternal self-reports and cord blood cotinine: prospective cohort study in karakow. matern child health.2009; 13:415-23. 31. wahabi ha, alzeidan ra, fayed aa, mandil a, al-shaikh g, esmaeil sa. effects of secondhand smoke on the birth weight of term infants and the demographic profile of saudi exposed women. bmc public health. 2013; 13(1):341. 32. ameer s, kandiah dm. detection of cotinine in passive smokers exposed to environmental tobacco smoke. biotech. 2017; 2(1):87-07 33. sachiyo k, kumiko a, keiko n, kaori k, sonomi o. effect of passive smoking using 23 comparison of salivary cotinine levels of neonates born to mothers exposed to varying degrees of passive cigarette smoke maternal and neonatal salivary cotinine measurements. nursing research.2012;61:140-4 34. nakajima m, fukami t, yamanaka h, higashi e, sakai h, yoshida r, yokoi t. comprehensive evaluation of variability in nicotine metabolism and cyp2a6 polymorphic alleles in four ethnic populations. clinical pharmacology and therapeutics. 2006;80:282-97 35. matt ge, quintana pje, hovell mf, et al. households contaminated by environmental tobacco smoke: sources of infant exposures. tob cont. 2004; 13(1):29-37. the authors: dr. wardah anwar assistant professor, department of physiology, al-aleem medical college, lahore. dr. ambreen anjum assistant professor department of physiology, al-aleem medical college, lahore. dr. nasreen akhtar assistant professor department of forensic and toxicology al-aleem medical college, lahore. dr. maria anwar assistant professor, department of physiology, independent medical college, faisalabad dr. tabinda kazmi assistant professor, department of physiology, niazi medical and dental college, sargodha. prof. sibgha zulfiqar head, department of physiology, shaikh zayed medical complex, lahore. dr. faiza khan assistant professor department of pharmacology, al-aleem medical college, lahore. corresponding author: dr. wardah anwar assistant professor, department of physiology, al-aleem medical college, lahore. e-mail: wardahtoseef@gmail.com 78 proceedings s.z.m.c. vol: 35(3): pp. 78-80, 2021. pszmc-814-35-3-2021 hereditary multiple exostoses with ulnar hemimelia syed wasif ali shah, muhammad abdul shakoor, usman amjad, haider ali, waleed imtiaz, shafqat raza department of orthopedic surgery, shaikh zayed medical complex, lahore abstract hereditary multiple exostoses is a skeletal dysplasia that is very rare and defined by formation of numerous cartilage capped benign tumours either pedunculated or sessile known as osteochondromas throughout skeleton especially around the growth plates of ribs, vertebrae, pelvis and long bones. rarely it can present forearm problems such bowing deformity of radius, ulnar shortening and radiocapitellar dislocation or subluxation. we are presenting a case of 20 year old female who presented with left distal ulnar exostosis resulting in ulnar shortening and radial bowing with restricted supination and pronation range of movement. other complaint was of multiple non tender bony hard lumps in both upper and lower limbs. excision of distal ulnar exostosis was done which resulted in marked improvement in pronation and supination range of movement. hereditary multiple exostoses with forearm deformities though very rare but can present and the treatment is conservative except if any bony swelling manifests any complications such as pain or associated deformity. key words: osteochondromas, distal ulnar exostosis, excision introduction diaphyseal achlasis also known as hereditary multiple exostoses is a genetic dysplastic condition having autosomal dominant mode of inheritance where several benign cartilage capped tumors originate from the perichondrium around the physis of bones. it has a prevalence of 1:50,000 in western countries.1 it usually involves vertebrae, scapula, pelvis, ribs and long bones. exostoses can result in reduction of skeletal growth, bony deformity, restricted joint movement, shortened stature, premature arthritis, and compression of peripheral nerves. it is very rare and can present with forearm deformities such as ulnar shortening, radial bowing and radiocapitellar subluxation.2 surgical excision of exostosis, limb lengthening and deformity correction are the available treatment options and are advised when osteochondromas become symptomatic or there is malignant transformation which rarely occurs.3 case report a 20 year old female presented to the outpatient orthopedic surgery clinic with bowing deformity and shortening of left forearm along with multiple swellings around knee and ankle joints. there as complaint of limitation of range of motion at wrist. problems were first noticed at age of 12 years. family history was positive for disproportionate dwarfism in a brother and an uncle. she was born by normal spontaneous vaginal delivery, with normal developmental milestones. she had no prior surgeries, was not taking any medications, and had no known drug allergies. physical examination revealed prominences around inferomedial aspect of bilateral knee and around both ankle joints. there was unilateral genu valgum deformity of right leg. limb length discrepancy of 2 cm noted in left leg with galeazzi sign positive for left femoral shortening. there was no contracture at hip or knee along with normal range of movement at hip, knee and ankle. there was varus deformity, shortening and dorso-radial bowing deformity of the left forearm with flexion contracture of little finger. there was 20° to 130° flexion and extension rom at left elbow with 10° supination and 0° pronation. the contra lateral (right) elbow had rom from 0° to 130° of flexion-extension with 60° supination and pronation. there were no other areas of pain or deformity in the upper extremities. antero-posterior and lateral radiographs of the left forearm were taken at first visit (fig-a). a distal ulnar exostosis was visualized projecting over the region of the interosseous membrane on both views. there were associated ulnar shortening and radial bowing. in addition, posterolateral radial head subluxation was seen. given the natural history of hme and the position of the distal ulnar exostosis, it was presumed that this lesion contributed to deformity of left forearm and resulted in restricted supination and pronation. complete skeletal survey was done which showed multiple exostosis (fig-b & c). 79 hereditary multiple exostoses with ulnar hemimelia fig-a: x-ray left radius/ulna ap and lat view showing distal ulnar exostosis, radial bowing and radiocapitellar subluxation fig-b:x-ray pelvis showing multiple osteochondromas fig-c: x-ray b/l tibia/fibula showing multiple exostoses the renal anomly scan and echocardiogram were normal. patient was operated and excision of exostosis of distal ulna was done (fig-d) with release of interosseous membrane. per operative assessment revealed improvement in supintion and pronation range of movement. postoperatively, patient was followed up in opd and at 2nd week follow up forearm range of movement was from 25° supination and 15° pronation. fig-d: per op pic showing distal ulnar exostosis discussion hereditary multiple exostoses is a rare skeletal dysplasia and mutations in exostosin genes are implicated in pathogens namely, ext1 (8q23-q24), ext2 (11p11-p13) and in rare cases, ext3 (on chromosome 19p). these mutations lead to dysregulation of cell surface heparin sulfate proteoglycans and consequently cartilage hyperproliferation.4 exostoses of upper extremities may result in forearm deformities such as shortened ulna with secondary disproportionate radius bone overgrowth been frequently noticed ultimately leading to radial bowing deformity.5 these lesions originate from perichondrium and flank the growth plate so these lesion tether the physis results in off balance growth of the bone. radiocapitellar subluxation or dislocation is common over time complication in the forearm deformities in context of hereditary multiple exostosis. dislocation of radial head has been associated with functional impairment such as loss of pronation and greater ulnar variance.6 masada et al.7 devised three types morphological classification for hereditary multiple exostoses linked forearm deformities. in type i, there is shortening of ulna secondary to distal ulnar exostosis and bowing of the radial bone. the radial head is not dislocated or subluxed. in type ii, there is radiocapitellar dislocation and shortened ulna. radial bone bowing is present but in milder form as compared to type i deformity. in type iia, the exostosis involves the proximal metaphyseal segment of radius as a result of which radial head is completely dislocated. in type iib, there is no exostosis involving proximal metaphysis of radius but still radial head dislocation happens. in type iii, there is disproportionate radial shortening relative to 80 hereditary multiple exostoses with ulnar hemimelia ulna due exostosis involving metaphysis of distal radius. he advocated to go for radial osteotomy simple exostosis excision and ulnar lengthening in type 1 deformity all thanks to his successful results in limited patients. controversy exists regarding the timing of surgical intervention. fogel et al.8 reported that the progression of forearm deformity comes to rest by simply excising exostosis in early age but it does not permanently provides complete correction. matsubara h et al.9 suggested that intervention should be performed older age after skeletal maturity as in this way a revision surgery could be evaded and satisfactory function could be obtained despite significant deformity. shin et al.10 assessed the clinical results of three different procedures sauvé kapandji, ulnar lengthening and simple excision for addressing forearm deformities manifested in patients of hereditary multiple exostoses. he reported that the mean supination improved by 11.8 by simple excision. no significant functional and clinical outcomes were attained from ulnar distraction lengthening procedure. results were consistent with previous studies showing that just going for simple excision was effective in improving forearm rotational range of movement. conclusion based on the results of this case presentation, we conclude that just performing simple exostosis excision can enhance the range of movement of the forearm in skeletally mature patients. references 1. g. a. schmale, e. u. conrad 3rd, and w. h. raskind, “the natural history of hereditary multiple exostoses”, the journal of bone & joint surgery, vol. 76, no. 7, pp. 986–992, 1994. 2. stieber, jonathan r. md; dormans, john p. md manifestations of hereditary multiple exostoses, journal of the american academy of orthopaedic surgeons: march 2005 volume 13 issue 2 p 110-120. 3. porter de, lonie l, fraser m, et al. severity of disease and risk of malignant change in hereditary multiple exostoses. a genotype phenotype study. j bone joint surg br. 2004; 86:1041e6. 4. alvarez cm, de vera ma, heslip tr, casey b. evaluation of the anatomic burden of patients with hereditary multiple exostoses. clin orthop. 2007; (462):73-79. 5. stieber jr, pierz ka, dormans jp. hereditary multiple exostosis: a current understanding of clinical and genetic advances. spring 2001;14:39-48. 6. 32. masada k, tsuyuguchi y, kawai h, kawabata h, noguchi k, ono k: operations for forearm deformity caused by multiple osteochondromas. j bone joint surg br 1989; 71:24-29. 7. jo ar, jung st, kim ms, oh cs, min bj. an evaluation of forearm deformities in hereditary multiple exostoses: factors associated with radial head dislocation and comprehensive classification. j hand surg am. 2017 apr; 42(4):292.e1-292.e8. 8. fogel gr, mcelfresh ec, peterson ha, wicklund pt: management of deformities of the forearm in multiple hereditary osteochondromas. j bone joint surg am 1984; 66:670-680. 9. matsubara h, tsuchiya h, sakurakichi k, yamashiro t, watanabe k, tomita k (2006). correction and lengthening for deformities of the forearm in multiple cartilaginous exostoses. j orthop sci. 11(5): 459-66. 10. shin ek, jones nf, lawrence jf (2006). treatment of multiple hereditary osteochondromas of the forearm in children: a study of surgical procedures. j bone joint surg br. 88(2): 255-60. the authors: dr. syed wasif ali shah associate professor and hod department of orthopedic surgery, shaikh zayed medical complex, lahore. dr. muhammad abdul shakoor trainee registrar department of orthopedic surgery, shaikh zayed medical complex, lahore. dr. usman amjad senior registrar department of orthopedic surgery, shaikh zayed medical complex, lahore. dr. haider ali trainee registrar department of orthopedic surgery, shaikh zayed medical complex, lahore. dr. waleed imtiaz trainee registrar department of orthopedic surgery, shaikh zayed medical complex, lahore. dr. shafqat raza trainee registrar, department of orthopedic surgery, shaikh zayed medical complex, lahore. corresponding author: dr. muhammad abdul shakoor trainee registrar, department of orthopedic surgery, shaikh zayed medical complex, lahore. e-mail: shakoorkhan00@gmail.com 75 proceedings s.z.m.c. vol: 35(3): pp. 75-77, 2021. pszmc-813-35-3-2021 idiopathic facial nerve paralysis & response to physiotherapy in pregnant woman; a case report 1wajida perveen, 2muhammad akhtar, 3riaz hashmi, 1misbah amanat ali, 4sahreen anwar, 5ayesha ghani 1department of physical therapy, sialkot college of physical therapy, amin welfare & teaching hospital, sialkot 2department of department of physical therapy, university of lahore 3department of physical therapy, syed medical complex, sialkot 4department of physical therapy, independent medical college, faisalabad 5department of gynecology & obstetrics, madina teaching hospital, faisalabad. abstract facial nerve palsy is a common condition affecting both genders and all age group. however, its incidence in pregnant females needs greater care. it may affect only lower half of the face or full one side of the face or in extreme condition bilateral sides. symptoms include deviation of angle of mouth towards sound side, flattening of nasolabial folds, loss of forehead wrinkles and inability to completely close the eyelid along with loss of taste on anterior two third of tongue. here we present a rare case report of facial nerve palsy in a pregnant woman in third trimester of pregnancy. she was offered an evidence based physiotherapy treatment regime along with pharmacological treatment and she recovered; however late complication of “ptosis” was reported. key words: facial palsy, bell’s palsy, pregnant women, ptosis, neurodynamic techniques, facial disability index (fdi), sunny brook facial grading system (sb fgs). introduction sir william gower defined facial paralysis as “the loss of motion of all the muscle supplied by 7th cranial nerve (facial nerve) on one side. most of the times it happens without ear infection or injury close to the nerve after it arises, it implies disease of the nerve as it passes through the bone.1 idiopathic bilateral loss of motion of facial muscles, albeit uncommon, seems to be more frequent during the last trimester of pregnancy and in the early puerperium. bilateral facial paralysis is less regularly idiopathic, and different etiologies had been found.2 onset is usually painful and acute, viral inflammation, immunosuppressant features of pregnancy and higher extracellular fluid content may be predisposing factor of the condition.3 facial paralysis in pregnancy is related with gestational and fetal difficulties, especially in the third trimester. sir charles bell in 1830 first identified a potential relationship of pregnancy with loss of movement of the muscles of facial expressions.4 in a case series reported, eight cases of pregnancy associated bell’s palsy (pabp) from 2006 to 2016, were identified out of them seven were found in third trimester and only one started two days postpartum. the recovery of these cases was satisfactory.5 but another study reported that, prognosis of the women with bell’s palsy in pregnancy is worse than that of the non-pregnant cases.6 case report a 32 years old female gravida 3 was in 36th week of her pregnancy. she woke up in the morning and found his mouth deviated to right side. her left eye lid was not closing completely, nasolabial folds were lost, and she was unable to produce wrinkles on left side of her forehead. the whole left half of the face was flat and expressionless. also there was loss of taste sensation on tongue, difficulty in drinking, chewing and swallowing. on the same day she visited her physician, who advised to take preventive and protective measures (especially eye care) only and referred to visit physiotherapist till delivery. she could not visit to physiotherapist but observed precautions at home. after seven days she gave birth to a normal healthy baby girl at 37th week of pregnancy who was 5.8 pounds. she was a known case of anxiety and depression since last five years and was also suffering from pregnancy induced hypertension. physiotherapy treatment was started on 3rd day of delivery after ten days of the 76 idiopathic facial nerve paralysis & response to physiotherapy in pregnant woman; a case report onset of symptoms. treatment included electric stimulation, support for the paralyzed facial muscles (provided by kinesiotaping) to prevent contracture on good side and overstretching of the facial muscles of affected side (left), exercises for facial muscles of left side three times a week. she was educated to self-administer home exercise programme according to condition of muscles. in the mean while neurophysician was consulted, who advised corticosteroids along with other pharmacological treatment. physiotherapy was continued for six weeks. her major symptoms were relieved except the bell sign which lasted partially even after eight weeks of onset. score of facial disability index7 before 1st physiotherapy treatment session and after last treatment session are expressed in table-1, and score of sunnybrook facial grading system8 as table-2. after six more weeks she noticed sudden drooping of eyelid on the affected side. this condition is although occasional but still present. physical functions social functions before 1st treatment session 33 76 after last treatment session 105 72 table-1: score of facial disability index. resting movements (compared to normal side) voluntary movements (degree of muscle excursion compared to normal side) synkinesis (degree of involuntary movements associated with each expression) composite score before 1st treatment session 20/20 24/100 12/15 12 after last treatment session zero/20 84/100 3/15 61 table-2: scores of sunnybrook facial grading system. discussion our study reported the case of left sided facial nerve paralysis in a woman in third trimester of pregnancy, but reported a rare incidence of bilateral facial palsy in pregnant woman in third trimester2 which was also reported in a recent case series.5 according to another case reported by n mathieu et al. in 2011; in a pregnant who had idiopathic facial palsy, pre-eclampsia and hellp syndrome, there was an association between idiopathic facial palsy which appeared in 3rd trimester and pre-eclampsia,9 but in our patient no such finding was observed. a 20 year old white lady was noted with a loss of voluntary movement of right half of upper lip and loss of hearing on same side during her last trimester of pregnancy. on assessment right sided facial paralysis was noted with obscure etiology,10 our patient also presented with facial palsy in last trimester. in 2008, al-husban hassan et al. conducted an investigation in which over a 10 years’ time from jan 1995 to jan 2005, 36 pregnant females with facial paralysis were found in the ent and obstetric gynecology facilities at king hussein medical center. the peak time of beginning of facial paralysis was thirty fourth week of gestation and was related with gestational and fetal complications. they reported causes of facial palsy in pregnancy from ten years record, but no information about any of the treatment given was reported, which we considered in our study. pre-eclampsia was diagnosed in 9 pregnant ladies (25%), 3 patients presented with gestational hypertension (8.3%), the frequency of c-section was 38.9% (14 patients), and 33.3% were induced and delivered vaginally (12 patients), 25% preterm birth (9 patients). fetal complications included 19.4% (7 youngsters) of low birth weight (less than 2500g), the rate of congenital anomalies 2.7% (1 infant), and twin births 5.6% (2 twins). they also inferred that facial paralysis in pregnancy is related with gestational and fetal complications, especially in third trimester.4 the strength of our study is that our patient was given an evidence based physiotherapy treatment and was found useful; however none of the previous studies reported any treatment and its outcomes. moreover, no further complication like ptosis which our patient suffered afterwards was observed in any of the previous studies. while limitation observed was her anxiety and depressive disorders which resulted in lack of adherence to the prescribed home programme. another case study reported, neurodynamics techniques of facial nerve were found effective in the treatment of facial palsy11 which is another novel method of treatment and may be considered in females with facial palsy during pregnancy as well. mazharbadsha et al conducted a randomized controlled trial to study the outcomes of steroids alone and in combination with facial exercise and electrical stimulation on 113 patients with bell’s palsy. they measured the outcomes on house brackmann facial nerve grading system and found the combination therapy was more useful12. results of our study are in line with the results of this trial. 77 idiopathic facial nerve paralysis & response to physiotherapy in pregnant woman; a case report conclusion facial palsy can affect any women during pregnancy especially during third trimester. physiotherapy treatment can help early recovery of the condition. references 1. cawthorne t, haynes d. facial palsy. british medical journal. 1956; 2(5003):1197. 2. kovo m, sagi y, lampl y, golan a. simultaneous bilateral bell's palsy during pregnancy. the journal of maternal-fetal & neonatal medicine. 2009; 22(12):1211-3. 3. cohen y, lavie o, granovsky-grisaru s, aboulafia y, diamant yz. bell palsy complicating pregnancy: a review. obstetrical & gynecological survey. 2000; 55(3):184-8. 4. al-husen h, abou-olim r, alzbone a, khreisat m, al rawashdeh b. relationship between the onset of facial palsy during pregnancy and the development of gestational complications.jrms.2008;15(2):19-22 5. leelawai s, sathirapanya p, suwanrath c. bell’s palsy in pregnancy: a case series. case reports in neurology. 2020; 12(3):452-9. 6. gillman gs, schaitkin bm, may m, klein sr. bell's palsy in pregnancy: a study of recovery outcomes. otolaryngology—head and neck surgery. 2002; 126(1):26-30. 7. vanswearingen jm, brach js. the facial disability index: reliability and validity of a disability assessment instrument for disorders of the facial neuromuscular system. physical therapy. 1996; 76(12):1288-98. 8. neely jg, cherian ng, dickerson cb, nedzelski jm. sunnybrook facial grading system: reliability and criteria for grading. the laryngoscope. 2010; 120(5):1038-45. 9. mathieu n, ledigabel j. pre-eclampsia and facial paralysis. gynecologie, obstetrique & fertilite. 2011; 39(2):e31-3. 10. story sr. conductive deafness, seventh nerve paralysis, and absence of vestibular response during pregnancy. ama archives of otolaryngology. 1958; 68(5):632-3. 11. kashoo fz, alqahtani m, ahmad m. neural mobilization in bell’s palsy: a case report. cranio®. 2019:1-4. 12. badshah m, umar m, naeem a, marryam m. a randomized control trial to review the effectiveness of combination therapy versus steroids alone, for the treatment of bells palsy. ann pak inst med sci. 2013; 9(3):118-21. the authors: wajida perveen associate professor, hod, programme director dpt, sialkot college of physical therapy, amin welfare & teaching hospital, sialkot. muhammad akhtar associate professor, department of physical therapy, university of lahore. riaz hashmi incharge, department of physical therapy, syed medical complex, sialkot. misbah amanat ali assistant professor, department of physical therapy, sialkot college of physical therapy, amin welfare & teaching hospital, sialkot. sahreen anwar assistant professor, department of physical therapy, independent medical college, faisalabad dr. ayesha ghani pg trainee, department of gynecology & obstetrics madina teaching hospital, faisalabad. corresponding author: wajida perveen associate professor, hod, programme director dpt, sialkot college of physical therapy, amin welfare & teaching hospital, sialkot. e-mail: wajopri@gmail.com numbering.indd 1 proceedings s.z.m.c. vol: 34(4): pp. 1-3, 2020. pszmc-762-34-4-2020 epidemic to pandemic: just a matter of time 1ayesha humayun, 2muhammad imran anwar, 3saadia shahzad alam, 4usman iqbal 1department of public health & community medicine, shaikh zayed medical complex,lahore 2department of surgery, shaikh zayed medical complex, lahore 3department of pharmacology & therapeutics, shaikh zayed medical complex, lahore 4global health & development, college of public health, taipei medical university, taiwan epidemic and pandemic potential of all respiratory diseases of zoonotic origin is well known. in recent history global public health threat was inflicted by zoonotic origin infectious diseases with pandemic potential including, ebola and marburg haemorrhagic fevers, severe acute respiratory syndrome (sars), influenza a (h5n1), middle east respiratory syndrome corona virus (merscov),1 and the recently discovered covid19.2change of epidemic to a pandemic is just a matter of time. a novel corona virus 2019 (covid-19) is appearing to be one of the rapidly transmitting diseases because of the social and economic linkages worldwide. emerging infectious diseases (eids) are not new but their incidence or geographical involvement is rapidly increasing.3 global temporal and spatial patterns of these emerging infections are studied well in literature concluding significant correlation of eids with socio-economic, environmental and ecological factors. it helps to identify regions as ‘hotspots’ for eids with a substantial risk of vector-borne and wildlife zoonotic emerging diseases.4 epidemic is “the occurrence in a community or region of cases of an illness . . . clearly in excess of normal expectancy”. pandemic is “an epidemic occurring over a very wide area, crossing international boundaries, and usually affecting a large number of people”.5 the decision of pandemic alert is influenced by the evidence evolving from virological, epidemiological, and clinical data.6 the phases of global pandemics are divided into four i.e, inter-pandemic, alert, pandemic, and transition.7 furthermore, the pandemic levels are classified into six phases where in phase 1 no virus circulation among animals is reported. in phase 2, a potential pandemic threat is there when domestic or wild animal circulation of virus found. phase 3 means human sporadic cases but usually no or limited manto-man transmission. in phase 4, also called risk of pandemic, when human-to-human transmission is verified. the phase 5 depicts that there is a global pandemic underway, if human-to-human transmission found in at least 2 countries of the same region classified by world health organization (who). phase 6 known as the pandemic phase, it keeps phase 5 criteria followed by post peak and post pandemic periods. from these phases, we can see that the last 3 phases show an actual threat and alert that concern stakeholders within countries and are required to take action with strong commitment by following instructions and warnings issued by who time to time accordingly with global situation.8 the pandemic risk level varies at different times among countries therefore, who advised to conduct their own national level risk assessments and plan management in coordination with its support.9 the international health regulations (2005) provide a global legal framework for public health potential threats to prevent, control, or respond to them to avoid emerging as public health emergencies of international concern (pheics). the risk assessment includes “preparedness,” “response,” and “recovery”, during pandemic.10 many developed countries took stringent initiatives during last decade to prepare for such a pandemic. despite the fact that these countries developed and tested pandemic preparedness plans after sars pandemic but it is unfortunate that covid pandemic has not been tackled well.11 recent covid-19 outbreak is a unique kind of respiratory infection as compared to sars (20022003) and mers (2012-ongoing). early in the outbreak of covid-19 in china, genetic analysis revealed similarity and difference from sars-cov and the corona virus isolated from bats found to be genetically similar.12 corona virus spread rapidly from wuhan, hubei, (end december 2019) only in a month within the mainland china. in wuhan city, travel ban was delayed which influenced epidemic progression by only 3 to 5 days in china but internationally 80% importations were not reduced until mid-february 2020.13 disease spread was rapid in covid-19 as compared to sars and mers, partly due to few factors triggering massive spread; epidemic focus in a metropolitan, wuhan, a very 2 epidemic to pandemic: just a matter of time economically active hub, well connected globally and within china; rapid connection through flights and massive rail transit; and chinese (lunar) new year celebration time. this rapid spread within china and globally showed pandemic illpreparedness of economic giant countries as well as the apathetic, cold and delayed response globally in european as well as other asian countries. inadequate risk assessment along with limited reporting within china contributed to the rapid spread of virus throughout china and other countries.14 this covid19 pandemic is creating a deleterious impact on human health and economics globally and dragging back countries’ economies to decades. preparedness plays a crucial role in protection of health and safety of employees and customers, limiting impact on businesses, community, economy and health system. a study of seven scenario’s of covid-19 epidemic shows that the impact on the global economy, even a contained outbreak could be significant in the short period.15 world should be learning lessons from countries that mitigated previous pandemics well such as taiwan, it has experienced sars in 2002.16 public health response mechanisms were well placed by taiwan (roc) after 2002, which enabled them to timely recognize this new covid-19 crisis and manage it through activated emergency management structures and processes. taiwanese government and society responded timely and quickly by taking basic measures such as wearing face masks, social distancing and quarantine along with other necessary measures.17 this editorial highlights the importance and need of identifying the global preemptive measures to thwart current and looming pandemics. in covid, pandemic alert phase should have been responded well to by the countries. there is a need for an extensive research on these novel emerging diseases especially corona viruses, focusing elucidation of route of transmission and pathogenesis along with development of drug and vaccines for effective prevention and control. moreover, we should emphasize on the technology utilization to get quality, reliable data from surveillance using advanced tools, epidemiologic field investigations, case series, cohort studies to identify burden of disease and attack rates in household and in special groups.18 vaccines and antiviral drugs are under trial but not clinically approved yet for public use. extensive research is needed to study natural history, appraise prevention strategies, develop therapies, draw conclusions and develop and evaluate impact of targeted public health interventions.19 developing and under-developed countries have enormous constraints, financial and social being the top most in the list. hence, a quick contextual needs assessment and smart objectives need to be devised to conceive and implement effective regional strategies. based on current scenario, previous knowledge and future predictions, we can try to find answers to these questions; did we learn from two prior epidemics of corona virus? were we well-prepared to deal with such highly contagious challenge? did we respond timely to covid? learning from previous experiences, timely and rapid information dissemination, prompt pandemic alerts and effective global collaborations can help us devise a way forward for public health threats. references 1. reusken cb, haagmans bl, müller ma, gutierrez c, godeke g-j, meyer b, et al. middle east respiratory syndrome corona virus neutralising serum antibodies in dromedary camels: a comparative serological study. the lancet infectious diseases. 2013;13(10):859-66. 2. wu z, mcgoogan jm. characteristics of and important lessons from the corona virus disease 2019 (covid-19) outbreak in china: summary of a report of 72 314 cases from the chinese center for disease control and prevention. jama. 2020 apr 7; 323(13):1239-1242. 3. morse ss. factors in the emergence of infectious diseases. plagues and politics: springer; 2001. p. 8-26. 4. jones ke, patel ng, levy ma, storeygard a, balk d, gittleman jl, et al. global trends in emerging infectious diseases. nature. 2008; 451(7181):990-3. 5. porta m. a dictionary of epidemiology: oxford university press; 2014. 6. doshi p. the elusive definition of pandemic influenza. bulletin of the world health organization. 2011; 89:532-8. 7. world health organization. pandemic influenza risk management: who interim guidance. world health organization, 2013: 6-7. 8. world health organization. pandemic influenza preparedness and response. a who guidance document 2009. available from url: https://apps.who.int/iris/bitstream/handle/10665/ 44123/9789241547680_eng.pdf;jsessionid=1f8 2125a878c00d08b9ac204eb320742?sequen ce=1. 3 epidemic to pandemic: just a matter of time 9. mccloskey b, dar o, zumla a, heymann dl. emerging infectious diseases and pandemic potential: status quo and reducing risk of global spread. the lancet infectious diseases. 2014; 14(10):1001-10. 10. katz r. use of revised international health regulations during influenza a (h1n1) epidemic, 2009. emerging infectious diseases. 2009; 15(8):1165. 11. holmberg m, lundgren b. framing postpandemic preparedness: comparing eight european plans. global public health. 2018; 13(1):99-114. 12. paraskevis d, kostaki eg, magiorkinis g, panayiotakopoulos g, sourvinos g, tsiodras s. full-genome evolutionary analysis of the novel corona virus (2019-ncov) rejects the hypothesis of emergence as a result of a recent recombination event. infection, genetics and evolution. 2020; 79:104212. 13. chinazzi m, davis jt, ajelli m, gioannini c, litvinova m, merler s, et al. the effect of travel restrictions on the spread of the 2019 novel coronavirus (covid-19) outbreak. science. 2020; 368(6489):395-400. 14. peeri nc, shrestha n, rahman ms, zaki r, tan z, bibi s, et al. the sars, mers and novel coronavirus (covid-19) epidemics, the newest and biggest global health threats: what lessons have we learned? int j epidemiol. 2020; 49(3):717-26. 15. mckibbin wj, fernando r. the global macroeconomic impacts of covid-19: seven scenarios. 2020. 16. yen my, chiu aw, schwartz j, king cc, lin ye, chang sc, et al. from sars in 2003 to h1n1 in 2009: lessons learned from taiwan in preparation for the next pandemic. j hosp infect. 2014; 87(4):185-93. 17. wang cj, ng cy, brook rh. response to covid-19 in taiwan: big data analytics, new technology, and proactive testing. jama. 2020; 323(14):1341-2. 18. cowling bj, leung gm. epidemiological research priorities for public health control of the ongoing global novel corona virus (2019ncov) outbreak. euro surveillance. 2020; 25(6). 19. li jy, you z, wang q, zhou zj, qiu y, luo r, et al. the epidemic of 2019-novel-corona virus (2019-ncov) pneumonia and insights for emerging infectious diseases in the future. microbes infect. 2020; 22(2):80-5. the authors: proceedings editorial team prof. ayesha humayun (associate editor) dept. of public health & community medicine, shaikh zayed medical complex, lahore. prof. muhammad imran anwar (associate editor) dept of surgery, shaikh zayed medical complex, lahore. prof. dr. saadia shahzad alam (chief editor) dept of pharmacology& therapeutics, shaikh zayed medical complex (szmc), lahore. dr. usman iqbal (external associate editor) global health and development, college of public health, taipei medical university, taiwan. 54 proceedings s.z.m.c. vol: 35(3): pp. 54-57, 2021. pszmc-809-35-3-2021 frequency and pattern of bronchiectasis in patients with chronic obstructive pulmonary disease presenting in a tertiary care hospital 1asifa karamat, 2huma batool, 3sohail anwar, 4shazia akram, 5atif masood, 5wajid ali rafai 1department of pulmonology, gulab devi hospital, lahore 2department of pulmonology, lahore general hospital, lahore 3department of pulmonology, university of lahore teaching hospital, lahore 4department of pulmonology, pakistan atomic energy commission, chashma 5department of medicine, university of lahore teaching hospital, lahore abstract introduction: chronic obstructive pulmonary disease (copd) is a high burden respiratory issue all over the world. it has high morbidity and mortality in the united states. bronchiectasis is associated with chronic obstructive pulmonary disease (copd) and is under diagnosed as diagnostic tools like hrct though easily available but an expensive test in developing countries like pakistan. we aim to establish the frequency and patterns of bronchiectasis in patients of copd. aims and objectives: to establish the frequency and pattern of bronchiectasis in copd in our population so that we can improve patient care and quality of life of these patients. place and duration of study: we did a cross sectional survey in department of pulmonology, gulab devi hospital, lahore. study was completed from 1st september 2017 to 28th february 2018. material & methods: after taking an informed consent 150 already diagnosed copd patients were included. bronchiectasis was seen on high resolution ct scan (hrct). data was collected on a structured proforma and analyzed on spss version 20. results: bronchiectasis was observed in 76 (50.6%) patients of copd while 74 patients had no bronchiectasis. out of 76, cylindrical bronchiectasis was seen in 82%. lower lobe and bilateral involvement was more common. means of age, gender, exacerbations of copd, and history of pulmonary tuberculosis were not related to bronchiectasis while pack years of smoking, duration of illness and modified medical research council (mmrc) dyspnea scale were significantly related to bronchiectasis. conclusion: this is observed that bronchiectasisis quite common (50.6%) in patients of copd in our population. key words: bronchiectasis, chronic obstructive pulmonary disease introduction chronic obstructive pulmonary disease (copd) is a high burden health issue globally. it is the 4th major cause of morbidity and mortality in the united states, and according to recent data, it is projected to cause heavy burden of disease globally.1,2,3 despite the diagnostic and therapeutic advancements the prevalence of respiratory diseases is increasing.1 the prevalence of copd is around 14%, 7%, and 3% in smokers, former smokers and non-smokers respectively.2 recent studies have projected that worldwide heavy budget will be spent on respiratory disease by 2030. in contrast, the per year will be the cost of smoking cessation program for controlling tobacco-related diseases is significantly low.4,7,8,9,10 patients with copd and bronchiectasis have more severe disease with poor outcome and higher frequency of exacerbations and complications.5 high resolution computed tomography (hrct) is a good diagnostic tool for determining the extent and severity of bronchiectasis.6,11,12 in a quite a few studies bronchiectasis was found in more than 50% of copd patients. cylindrical bronchiectasis was present in majority and cystic bronchiectasis in almost 20% of cases. commonly lower lobes were involved. mostly copd patients have bilateral involvement with no specific segmental predominance.7 various patients factors influence the frequency of bronchiectasis among copd patients, like mean smoking pack year, active smoking, duration of disease, number of exacerbations leading to hospital admission, mmrc score (modified medical 55 frequency and pattern of bronchiectasis in patients with chronic obstructive pulmonary disease research council dyspnea scale), past history of tuberculosis and degree of airway obstruction.7 we aim to establish the frequency and patterns of bronchiectasis in copd patients of our population. by doing so, we can improve the patient care and management of these patients thus improving their quality of life and exacerbation rates. material and methods it was a cross-sectional study conducted in gulab devi hospital, lahore. patients were labeled as having copd on the basis of both of the following two criteria based on values of pulmonary function test (pft’s) post bronchodilator fev1 less than or equal to 80% with reversibility less than 15% or less than 200ml. post bronchodilator fev1/ fvc, (forced expiratory volume in one second/forced vital capacity) less than 0.7. bronchiectasis was diagnosed on the hrct reported by consultant radiologist. the study was conducted in department of pulmonology, gulab devi hospital, lahore from 1st september 2017 to 28th february 2018.copd patients with age ≥ 40 & ≤ 80 years were included in this. people with 15 pack year smoking history were considered as chronic smoker.8 patient who were already diagnosed case of bronchiectasis were excluded from the study. sampling technique was non-probability consecutive purposive sampling. after taking informed consent from patient, data was collected. pulmonary function tests (pft’s) was done to measure fev1 and fvc in each patient (as per inclusion criterion). demographic characteristics included in this study were height, age, bmi, sex, current or ex-smoking history, history of huqqa smoking, pack year of smoking, number of copd exacerbations in previous one year and past history of anti-tuberculous treatment, and exposure to biomass fuel. mmrc dyspnea scale was used to assess functional status. hrct was performed as per protocol was reported by same consultant radiologist. presence of bronchiectasis as well as patterns of bronchiectasis was documented based on radiologist’s report. selection bias was addressed by using appropriate sample size and by using non-probability consecutive sampling. measurement bias was addressed by using uniform measurement scale/units for each patient. data was collected on a structured proforma (attached). statistical analysis: standard deviation and mean was taken of different variables i.e. bmi, age, no. of copd exacerbations, smoking pack years, fvc, mmrc score, and no. of copd exacerbations). frequency and percentages were taken for variables i.e. history of tb, gender, hukka smoker and biomass fuel exposure, current/ex-smoker, and presence and pattern of bronchiectasis. spss 20 version was used for analysis of this study. chi square and fisher exact tests were applied. results total 150 patients of mean age of 56.2± 5.9 years were included in the study. characteristics of studied population are shown in table 1. according to findings there were 71% and 29% males and females respectively. fifteen percent patients were previously treated for pulmonary tuberculosis. half of the sampled population was exposed to cigarette and huqqa. more than half of the patients (67%) have stopped smoking for past 1 year. in last one year average number of exacerbations is 3.2. 50% of our study population had bronchiectasis. 14 patients (18%) had cystic bronchiectasis while majority (82%) had cylindrical bronchiectasis on hrct. most of the patients having bronchiectasis in current study population have multiple lobe involvement (n = 65, 85%). lower lobe was the common site involved. majority (n=48, 64%) had bilateral involvement. (table-2) on comparison of groups with and without bronchiectasis there was non-significant difference of patients of bronchiectasis between ages more than and less than 60 years. similarly gender is also found not related with bronchiectasis in studied sample, p=0.14. mean number of exacerbations is same for both groups of patients, p= 0.24 there was non-significant difference of patients of bronchiectasis between exacerbations more than and less than 3 per year (p= 0.1). but in bronchiectasis group, smoking years (p=0.03); time elapse since in years (p=<0.001); and mmrc score (p=<0.001) were significantly higher. it is seen that patients who had previously received antituberculous treatment were equally distributed in bronchiectasis and non-bronchiectasis groups (p=0.2). similarly both ex-smokers and active were equally prone to develop bronchiectasis (p=0.58) as shown in table-3 56 frequency and pattern of bronchiectasis in patients with chronic obstructive pulmonary disease mean age 56.2 ±sd gender (%) male female 106/150(71) 43/150(29) smoking status cigarette smoker (%) huqqa smoker (%) cigarette + huqqa smoker (%) biomass fuel exposure (%) active smoker (%) ex-smoker (%) 33/150(22) 13/150(9) 69/150(46) 34/150(23) 45/150(33) 100/150(67) history of pulmonary tuberculosis (%) 22/150(15) mean copd exacerbation in last one year 3.2 ±1.7 mean duration of smoking(years) 41.3 ±18.9 mean mmrc score 2.8 ± 0.74 mean time elapsed since diagnosis (year) 7.09 ± 3 table-1: basic demographics of study population (n=150) bronchiectasis present (%) absent (%) 76/150 (50.6%) 74/150 (49.3) type of bronchiectasis cystic (%) cylindrical (%) 14/76 (18) 62/76 (82) extent of bronchiectasis one lobe more than one lobe upper lobe lower lobe right lung left lung bilateral 11/76(15) 65/76(85) 11/76(15) 65/76(85) 10/76(12) 18/76(24) 48/76(64) table-2: frequency and characteristics of bronchiectasis (n=150) parameter bronchiectasis present bronchiectasis absent p value age <60years >60years 48 28 50 24 0.7 gender male female 50 26 57 17 0.14 time elapsed since diagnosis <5 years <5 years 1 75 50 24 < 0.001 smoking status current smoker ex-smoker mean duration of smoking 23 53 45 26 48 38 0.2 0.03 mean mmrc 3 2.6 0.001 history of pulmonary tuberculosis 15 8 0.2 no. of exacerbation last year <3 >3 46 30 55 19 0.1 table-3: comparison of patient characteristics with and without bronchiectasis discussion in our study population copd was unequally distributed in males and females (71% in males & 29% in females) likely because smoking is more prevalent in males while in another study by aryals et all concluded copd to becoming more prevalent in females causing increase mortality.15 majority in our study though had quit smoking but were still having bronchiectasis (67%), showing contribution of smoking on permanent lung damage. biomass fuel exposure was common in females due to use of high carbon fuel in rural areas. more than half of the study population was having bronchiectasis which was comparable to observation of martínez-garcía má et al.7 although another study showed only 30% prevalence of bronchiectasis in copd patients.13 the pattern and distribution of bronchiectasis found in our study was cylindrical affecting bilateral lower lobes which was consistent with martínez-garcía má et al observation.7 bronchiectasis was significantly related to time elapsed since diagnosis, smoking pack years and mmrc score of severity of copd. while age, gender, history of pulmonary tuberculosis and recurrent exacerbations were not significantly related to bronchiectasis in patients with chronic obstructive pulmonary disease. jin, jianmin and yu, wenling et al. also observed the same factors which had significant impact on prevalence of bronchiectasis.14 conclusion it is concluded that the frequency of bronchiectasis is quite high (50.6%) in copd patients of our population and it needs to be ruled out. some copd patients are, more susceptible to develop this complication. references 1. decramer m, janssens w, miravitlles m. chronic obstructive pulmonary disease. lancet. 2012 apr 7; 379(9823):1341-51. 2. athanazio r. airway disease: similarities and differences between asthma, copd and bronchiectasis. clinics (sao paulo). 2012 nov; 67(11):1335-43. 3. the global strategy for the diagnosis, management and prevention of copd, global initiative for chronic obstructive lung disease (gold) 2011. available from: http://www.goldcopd.org/. 4. rosenbaum l, lamas d. facing a “slow-motion disaster” the un meeting on non-communicable diseases. n engl j med. 2011; 365(25):2345-48. 57 frequency and pattern of bronchiectasis in patients with chronic obstructive pulmonary disease 5. o'donnell ae. bronchiectasis in patients with copd: a distinct copd phenotype? chest. 2011 nov; 140(5):1107-8. 6. gupta pp, yadav r, verma m, agarwal d, kumar m. correlation between high-resolution computed tomography features and patients' characteristics in chronic obstructive pulmonary disease. ann thorac med. 2008 jul; 3(3):87-93. 7. martínez-garcía má, soler-cataluña jj, donatsanz y, catalán serra p, agramunt lerma m, ballestín vicente j, et al. factors associated with bronchiectasis in patients with copd. chest. 2011 nov; 140(5):1130-7. 8. yu liu, meng dai, yufang bi, min xu, active smoking, passive smoking, and risk of nonalcoholic fatty liver disease (nafld): a population-based study in china. j epidemiol. 2013; 23(2): 115-121. 9. launois c, barbe c, bertin e, nardi j, perotin jm, dury s, et al. the modified medical research council scale for the assessment of dyspnea in daily living in obesity: a pilot study. bmc pulm med. 2012 oct 1; 12:61. 10. hu g, zhou y, tian j, yao w, li j, li b, et al. risk of copd from exposure to biomass smoke: a metaanalysis. chest. 2010 jul; 138(1):20-31. 11. goeminne p, dupont l. non-cystic fibrosis bronchiectasis: diagnosis and management in 21st century. postgrad med j.2010aug;86(1018):493-501 12. javidan-nejad c, bhalla s. bronchiectasis. radiolclin north am. 2009 mar; 47(2):289-306. 13. o'brien c, guest pj, hill sl, et al. physiological and radiological characterization of patients diagnosed with chronic obstructive pulmonary disease in primary care. thorax 2000; 55:635-642. 14. jin, jianmin mda; yu, wenling mdb; li, shuling mmb; lu, lijinmda; liu, xiaofang mda; sun, yongchang mda, medicine: july 2016 volume 95 issue 29 p e4219. 15. aryal s, diaz-guzman e, mannino dm. influence of sex on chronic obstructive pulmonary disease risk and treatment outcomes. int j chron obstruct pulmon dis. 2014; 9:1145-54. the authors: dr. asifa karamat assistant professor, department of pulmonology, gulab devi hospital, lahore. dr. huma batool assistant professor, department of pulmonology, lahore general hospital, lahore. dr. sohail anwar assistant professor, department of pulmonology, university of lahore teaching hospital, lahore. dr. shazia akram assistant professor, department of pulmonology, pakistan atomic energy commission, chashma. dr. atif masood associate professor, department of medicine, university of lahore teaching hospital, lahore. dr. wajid ali rafai senior registrar, department of medicine, university of lahore teaching hospital, lahore. corresponding author: dr. sohail anwar assistant professor, department of pulmonology, university of lahore teaching hospital, lahore. e-mail: sohail.anwar@ucm.uol.edu.pk 46 proceedings s.z.m.c. vol: 34(3): pp. 46-50, 2020. pszmc-761-34-3-2020 clinical risk factors associated with mortality among burn patients admitted to jinnah burn and reconstructive surgery center, lahore fatima sakhawat, hamna sohail, fatima zafar, hassan bashir, hasan shafique, sana iftikhar department of community medicine, allama iqbal medical college, lahore abstract introduction: burn injuries being highly devastating in nature, are associated with extensive morbidity and mortality. although the incidence of burn injuries is less in developed countries but it is increasing in developing and low income countries. aims & objectives: the aim of this study was to determine clinical risk factors associated with mortality among burn patients. place and duration of study: jinnah burn and reconstructive surgery center, lahore from august 2018 till january 2019. material & methods: data was collected by researchers themselves by studying files of patients. data analysis was done by using spss and pearson chi square test was used to determine association of clinical risk factors with mortality among burn patients. results: out of 200 patients, 137 were males. the mortality rate was 12.50 %. cardiopulmonary arrest was the most common cause of death. sepsis was the most common complication among burn patients. higher mortality rates were observed among patients with arterial/venous and urinary catheterization done. higher survival rates were seen in those patients who were treated with debridement, grafting surgeries, colistin use and aided by mechanical ventilation. conclusion: aid through mechanical ventilation, grafting surgeries, wound debridement, use of colistin are associated with improved survival outcomes among burn patients. whereas inhalational injury, arterial/venous catheterization, urinary catheter and sepsis are seen as major predictors of mortality among burn patients. key words: mortality rate, burns, survival, clinical profiles introduction a burn is defined as an injury to skin or other organic tissue primarily caused by heat radiation, electricity or contact with chemicals.1 burns are the most common destructive form of injury. improved outcomes after burn injuries are seen due to advances in field of medicine in terms of fluid replacement therapies, nutritional care pulmonary care, burn wound care and infection control practices.2 much improvements in treatment strategies are still needed to further decrease the mortality associated with burn injury. according to facts and figures by who, burn injuries cause death of nearly 180,000 people per annum.3 fire related burns is one of the major causes of disability adjusted life years (dalys) in countries with average or below average income.4 who statistics show that nearly 75% of burn injuries are common in south east asia, eastern mediterranean and african subcontinent5 the incidence of burn injuries is estimated to be 243 per 100,000 people per year with mortality rate equal to 11.6 % in low income countries of south east asia in comparison to 1 death per 100,000 people in high income countries.6 a study of research paper from 14 african countries showed that malawi has the highest mortality rate due to burn injury.7 although there is no proper record keeping system functional in pakistan, however, it is estimated that mortality due to burn injuries reaches up to 36% which is still 5 times higher than that in high income countries. people with severe burn injuries have increased risk of nosocomial infections which is considered to be the most common cause of death after burn.8 patients who survive the burn injuries recover gradually, however recovery is incomplete in a large number of cases. moreover it also has a remarkable effect on the psyche of patients as the patient has to undergo multiple reconstructive surgeries.9 despite of the due importance, the epidemiological studies of burn injury are under-researched in pakistan.10 by studying the co relation of outcome of burn injury with various clinical factors inpatients’ profiles, this 47 clinical risk factors associated with mortality among burn patients study may help in revealing predictors of mortality in burn patients. material and methods lahore is a city of punjab, pakistan with a total population of 11.13 million. jinnah burn and reconstructive surgery center is a part of jinnah hospital, lahore which is the second largest tertiary referral teaching hospital of the province of punjab. the cross sectional study was conducted in jinnah burn and reconstructive surgery center, lahore over a period of 6 months (from august 2018 to january 2019).the permission for this study was obtained by ethical committee of the institute. informed consent was taken from patients. confidentiality of the data of patients was maintained. sample size was calculated using open epi software at 95% confidence interval with margin of error 5% and taking frequency of anticipated factor (mortality among burn patients) as 11.6%. calculated sample size was 137. we recruited 200 patients in this study (total number of patients who were admitted during six months study period). patients with contractures were excluded. non probability purposive sampling was done to recruit patients. a questionnaire was designed and pretested to collect the data about clinical profile of patients. files of the patients were studied in detail by the researchers to obtain the dataon patients age, gender, inhalational injury, grafting, debridement surgeries, complications of burn injury, its outcomes, culture reports, venous catheterization, use of urinary catheter, support on mechanical ventilation and use of colistin. statistical analysis: data analysis was done by using statistical package for the social sciences (spss) version 24. cross tabulation of data was done between outcome of burn injury and clinical profiles of the patients. we applied pearson's chi square test to determine statistical significance of clinical risk factors with mortality among burn patients. p value <0.05 was considered statistically significant. results out of 200, 68.5% patients were males. male to female ratio is 2.17:1. majority (35%) of the patients belonged to the age group of 16-25 years (n=70). majority (52.5%) of the patients suffered from mixed thickness type of burn injury. (graph-1) the mortality among burn patients was found to be 12.5%. the most common complication of the burn injury was sepsis 42.6% (n=84), second most common complication seen was cardiopulmonary arrest 11.7% (n=23). the most common cause of mortality seen among burn patients was cardiopulmonary arrest (92%). inhalation injury was present in 21.5% of the patients and mortality rate among them was 32.5%, (p=0.00). amputation of limb was done in only 5 percent of the patient. wound culture was done in about 61% patients, about 21.5 % of culture reports were negative. among positive culture report the most common organism found was pseudomonas 18.85% followed by coinfection of pseudomonas and klebsiella 13.1% (fig-1).survival rate was statistically better among those patients in whom wound culture was done(p=0.00).arterial/venous catheter was passed in about 79% of the patients. mortality rate was higher among those in which arterial/venous catheter was passed (96%, p=0.026). urinary catheter was passed in about 64.5% patients. mortality rate among patients who had urinary catheter passed was 88%, higher than that of others who had not got urinary catheter passed (p=0.009).only 16% patients required mechanical ventilation, survival rate was higher in those who were provided mechanical ventilation aid (68.75%) as compared to those who weren’t (p=0.00). colistin was given in 29.5% patients and it improved survival rate (p=0.012) .grafting surgeries were done in 36.5% patients and out of them only 2.73% patients deceased. so, grafting was found to improve the outcome of burn injury (p=0.007).debridement was done in 35% patients. out of 70 patients who were treated with debridement, 97.1% patients survived. so, like grafting debridement was also found to have increased the survival rates among the patients (p=0.008). (table-1) graph-1: degree of burn injury 48 clinical risk factors associated with mortality among burn patients figure 1: wound culture report outcome of burn injury p-value patient deceased patient survived inhalational injury yes 14 29 0.00 no 11 146 number of debridements done 0 23 107 0.008 1-2 1 59 >2 1 9 number of grafting surgeries done 0 23 104 0.007 1-2 2 69 >2 0 2 venous catheter passed yes 24 134 0.026 no 1 41 urinary catheter passed yes 22 107 0.009 no 3 68 aid through mechanical ventilation yes 10 22 0.00 no 15 153 use of colistin yes 2 57 0.01 no 23 118 complications of burn injury sepsis 1 83 0.00 cardiopulmonary arrest 23 3 both sepsis and cardiopulmonary arrest 0 2 ectropion 0 1 shock 0 1 dry gangrene 0 1 respiratory arrest 0 3 both sepsis and respiratory distress 0 3 no complications 1 78 table-1: relation of outcome of burn injury with variables discussion finding out the outcome of burn injury and its association with various risk factors is important to improve not only management and care plan for our patients but to make better preventive schemes at individual and social levels in this era as well. the goal of the study is to guide health care professionals in improving approaches regarding treatment of the patient. this study has determined the mortality rate as 12.5% among burn patients which is far less than that of other studies done. another study from karachi pakistan had shown mortality rates of 36.12% in that region due to the same cause.11 a malaysian study had shown the mortality rates of 12.2% among burn patients which is closer to our study.12 a multi institutional study form united states had shown mortality rate of 3.3% due to burn injury13 which can be attributed to good infection control practices over there. despite the excel in disease management and patient care, pakistan is still facing higher mortality rates due to burn injury in comparison to developed countries in the western world which implies that much is needed to be done practically to lower the mortality in our patients. the various predictors of mortality in burn patients include inhalational injury, arterial/venous catheterization, urinary catheterization, complications such as sepsis and cardiopulmonary arrest, aid through mechanical ventilation, grafting surgeries and debridement of the wound. a generalized improved impact of use of colistin on outcome of burn injury was also observed. it has been observed that in all the cases of mortality, the cause was cardiopulmonary arrest, except two cases; out of them, the cause of death of one patient was declared as septic shock and the cause of death of the other patient was unknown. burn injury causes surge of catecholamines which causes cardiac stress after burn injury. this is because of increased myocardial oxygen utilization which causes myocardial hypoxia and cardiac death.14 most common complication among those who have survived was nosocomial infections/sepsis. about 47.51% patients suffered from such complications in our hospital settings. sepsis triggers inflammatory cascades which causes multi organ damage and failure leading to death of the patient.15 according to our study, most of the patients who suffered from inhalational injury, died. inhalational injury is associated with exposure of respiratory tract to heat leading to thermal burn inside and 49 clinical risk factors associated with mortality among burn patients damage due to constitutional toxins of inhaled smoke.16 survival rates were found to be high among those who were aided by mechanical ventilation than those who were not. this is because of the reason that burn patients suffer from respiratory distress, especially when it comes to diagnose the cases of inhalational injury associated with burn trauma .in most of the cases, the culture report was positive for pseudomonas and second most frequent organism found infecting was klebsiella. survival rates was high in patients who had negative microbe culture report. out of the patients who died, most had urinary foley catheter and iv line passed. although iv lines are very important in administering medicines such as saline and ringer’s lactate and benefits of foleycathetar can also never be denied when it comes to drain the urine and treat exaggerated urinary continence but both of them are associated with increased risk of infections due to iatrogenic causes. this may be due to flora at periuretheral area and skin and also the biofilm formation. this bacterial infection when spreads through the blood is a cause of multi organ dysfuntion and consequently death. yet, the primary factors are always the total body surface area burnt, degree and site of burn injury.17 use of iv colistin is highly protective in patients of burn injury according to our study. it is a polymyxin e antibiotic very effective against multidrug resistant strains of pseudomonas and aceintobacter species.18 debridement of a wound was clearly found as a beneficial factor to achieve the health and well being in a patient. burn patients lack primary protective layers of the body such as skin and fascia which expose underlying tissue to microorganisms in the environment and surrounding thus increasing the risk of infections. however, some time these infections being destructive, damage the tissue to such an extent that it cannot be spared by using antibiotics and the only preventive treatment left to prevent the spread of infections is to do debridement and grafting. in severe cases amputation of the limb may be the option to save the life from vicious septic cycles and multi organ failures.19 this study highlights the importance of certain factors towards reducing the burden of mortality due to burn injury, however, a lot of research with larger sample sizes is still needed to support our findings. moreover, mortality rate in our study is low in comparison to other studies in pakistan, this may be due to the reason that this study was conducted at tertiary care hospital. therefore, to get a more generalized result, such studies should also be promoted at level of district and tehsil headquarter hospital settings so that a significant addition can be made in terms of exploring the causes and risks of mortality among burn patients. attempt should be made to find out the ways which can help in further reduction of the mortality rates taking into account the impact of associated risk factors and to develop the strategies for improvement of patient management and health care services accordingly. conclusion aid through mechanical ventilation, grafting surgeries, wound debridement, use of colistin are associated with improved survival outcomes among burn patients. whereas inhalational injury, arterial/venous catheterization, urinary catheter and complications such as cardiopulmonary arrest and sepsis are seen as major predictors of mortality among burn patients. references 1. mathers c, fat dm, boerma j. the global burden of disease: 2004 update. bmc res notes, 6 (2013), p. 545 geneva: world health organization; 2008. 2. hussain a, choukairi f, dunn k. predicting survival in thermal injury: a systematic review of methodology of composite prediction models. burns. 2013 feb 2; epub ahead of print. [pubmed] 3. global burden of disease. the institute for health metrics and evaluation. geneva: world health organization; 2008. 4. peck md. epidemiology of burns throughout the world. part i: distribution and risk factors. burns. 2011; 37(7):1087-1100. [pubmed] 5. who fact sheets; http://www.who.int/mediacentre/factsheets/fs36 5/en/ 6. global burden of disease. the institute for health metrics and evaluation. http://www.healthdata.org/gbd 7. a. golshan, c. patel, a.a. hyder a systematic review of the epidemiology of unintentional burn injuries in south asia j public health (oxf), 35 (3) (2013), pp. 384-396. 8. o'keefe ge, hunt jl, purdue gf. an evaluation of risk factors for mortality after burn trauma and the identification of genderdependent differences in outcomes. j am coll surg. 2001; 192(2):153-60. [pubmed] 9. rafla k. tredget ee. infection control in burn unit. burn 200; 37:5-15. 50 clinical risk factors associated with mortality among burn patients 10. batra ak. burn mortality: recent trends and socio cultural determinants in rural india burn. pubmed. 2003; 29:270-5. 11. ibran et al. mortality associated with burn injuryacross sectional study from karachi, pakistan. bmc research notes 2013; 6:545. 12. tan chor lip, tan, thomas, imran, azmah tuan mat: survival analysis and mortality predictors of hospitalized severe burn victims in a malaysian burns intensive care unit. 2019 jan. 13. zavlin, chegireddy, boukovalas, nia, branski, friedman, echo. multi-institutional analysis of independent predictors for burn mortality in the united states: 2018 aug 22; 6:24. 14. felicia n williams, md; david n herndon, md;oscar e suman, phd, ; jong o lee, md; william b norbury, md; ludwik k branski, md; ronald p mlcak, phd; and marc g jeschke, md, phd1: changes in cardiac physiology after severe burn injury: j burn care res. 2011 mar-apr. 15. bharwana, s., farid, m., ali, s., rizwan, m., nadeem, m., haider, m., tauqeer, h., sallahud-din, r., & ahmad, r. (2016). mortality rate associated with hospital acquired infections among burn patients. biomedical research and therapy, 3(9), 790-799. 16. vittorio pavoni, lara gianesello, laura paparella, laura tadini buoninsegni, and elisabetta barboni: outcome predictors and quality of life of severe burn patients admitted to intensive care unit: 2010 apr 27. 17. dr. barbara w. trautner, md and dr. rabih o. darouiche, md catheter-associated infections pathogenesis affects prevention. 18. ludwik k. branski, ahmed al-mousawi, haidy rivero, marc g. jeschke, arthur p. sanford, and david n. herndon emerging infections in burns: surg infect (larchmt). 2009 oct; 10(5): 389-397. 19. bowo sa, soedjana h. effectiveness of early excisional debridement in burn injuries to sepsis incidence and mortality rate at burn unit of hasan sadikin hospital. j plast rekons. 2017, jan 24; 3(2):40-4 the authors: fatima sakhawat 5th year m.b.b.s. student, department of community medicine, allama iqbal medical college, lahore. hamnasohail 5th year m.b.b.s. student, department of community medicine, allama iqbal medical college, lahore. fatima zafar 5th year m.b.b.s. student, department of community medicine, allama iqbal medical college, lahore. hassan bashir 5th year m.b.b.s. student, department of community medicine, allama iqbal medical college, lahore hasan shafique 5th year m.b.b.s. student, department of community medicine, allama iqbal medical college, lahore. dr. sana iftikhar assistant professor, department of community medicine, allama iqbal medical college, lahore. corresponding author: fatima sakhawat 5th year m.b.b.s. student, department of community medicine, allama iqbal medical college, lahore. e-mail: fatimasakhawat1@gmail.com 64 proceedings s.z.m.c. vol: 35(3): pp. 64-68, 2021. pszmc-811-35-3-2021 prevalence of right ventricular dysfunction & pulmonary hypertension and their relationship to the number of hemodialysis sessions in patients of end-stage renal disease (esrd) 1rakesh bahadur adhikari, 2qazi abdul saboor, 3mateen akram, 4saira fayyaz, 2husnain bashir, 3muhammad rizwan-ul-haque 1department of cardiology, shahid gangalal national heart centre, kathmandu, nepal 2department of cardiology, shaikh zayed medical complex, lahore 3department of nephrology, shaikh zayed medical complex, lahore 4department of cardiology, city care hospital, khanewal abstract introduction: chronic hemodialysis (hd) ends up with right ventricular (rv) dysfunction and increased pulmonary hypertension (phtn). left to right shunt in dialysis patients due to arterio-venous fistula (avf) causes chronic volume overload, independent of rise in body water leading to worsening rv overload and rv dysfunction (rvd). aims & objectives: to determine the prevalence of rv dysfunction & pulmonary hypertension and its relationship to the number of dialysis sessions in patients of esrd. place and duration of study: department of cardiology & nephrology, sheikh zayed hospital, lahore for one year from march 2016 march 2017. material & methods: this cross-sectional analytical hospital based study enrolled 145 patients of end-stage renal disease (esrd) on regular 4-hours hd sessions at two or more times per week for at least 3 months. echocardiography (echo) with 2-d, m (motion) mode & doppler studies were done. rv dysfunction by tapse value less than 15mm & phtn by systolic pulmonary artery pressure >35 mm hg or tricuspid regurgitation velocity (vtr) ≥2m/s at rest were noted. data was analyzed on spss version 20. results: rv dysfunction was seen in 40.7% (59) of patients and the frequency rose across the 4 dialysis session groups (13.8%, 37.3%, 51.7% & 100%, p<0.001). phtn was observed in 44.1% of the patients and the prevalence progressively increased across the groups (17.2%, 45.1%, 53.3% & 80.01%, p<0.003). there was significant association between rvd and phtn (p=0.011). conclusion: we observed positive correlation between rv dysfunction and phtn with total number of hemodialysis sessions. early detection of sub-clinical rv dysfunction may improve mortality and morbidity by optimizing treatment options. key words: esrd patients on hd, echo, rv dysfunction, phtn. introduction cardiovascular diseases are leading cause of mortality among dialysis patients accounting for 50% of deaths,1 due to congestive cardiac failure (ccf) was found to be most common with poor prognosis.2 but right ventricle (rv) is the least studied topic. chronic hemodialysis ends up with rv dysfunction and increased pulmonary pressure. left to right shunt in dialysis patients due to arteriovenous fistula (avf) causes chronic volume overload, independent of rise in body water leading to worsening rv overload.3 clinical manifestations of cardiac disease at the start of esrd therapy are high.4 and 50% of patients starting a dialysis program have some type of preexisting cardiovascular disease.5,6 available data on rv dysfunction is scarce but there is significant evidence of pulmonary hypertension in patients on chronic hemodialysis and survival of patients has been related to function of heart rather than values of pulmonary pressure.7 also rv dysfunction may affect left ventricular filling through interventricular interactions.8 previous studies focused on the relation between dialysis and pulmonary hypertension gave more emphasis to the tissue doppler imaging (tdi) indices of left ventricle (lv) function and showed that diastolic dysfunction is more prevalent in these patients.9,11 in 2010 a retrospective study by paneni et al showed that in presence of brachial avf, risk of rv dysfunction increased among hemodialysis patients10, and showed to predict mortality.12 65 prevalence of right ventricular dysfunction & pulmonary hypertension and their relationship… on echocardiography, tricuspid annular plane systolic excursion (tapse) is a reliable parameter to evaluate rv function. it is a good prognostic marker for cardiovascular risk stratification and is unaffected by heart rate and rhythm.13 and 20% of hemodialysis patients with av fistula showed abnormal tapse values (<15 mm).14 another recent study showed that the reduction in rv dysfunction was independent of left ventricular hypertrophy (lvh), diastolic dysfunction of the lv, or pulmonary hypertension.15 but this has not been described in pakistani literature up to present. our objective is to determine the prevalence of pulmonary hypertension and rv dysfunction & their relationship to the number of dialysis sessions in patients esrd on maintenance hemodialysis. material and methods this is an analytical cross-sectional research study done in department of cardiology & nephrology, shaikh zayed hospital, lahore during one year, march 2016 march 2017. 145 patients met the inclusion criteria and were enrolled in the study. the sample size of 145 was estimated by using 95% confidence level, 8% margin of error with expected frequency of rv dysfunction 60% among hemodialysis cases.14 rv dysfunction (rvd) is declared when on transthoracic echocardiography tapse value less than 15mm14 and chronic kidney disease (ckd) is declared when reduced renal function that is glomerular filtration rate less than 60ml/min/1.73m2 for at least 3 months irrespective of cause.17 pulmonary hypertension is said significant when systolic pulmonary artery pressure >35 mm hg or tricuspid regurgitation velocity (vtr) ≥2m/s at rest.18 inclusion criteria: 1) adults above the age of 18 years. 2) patients on regular 4 hours hd sessions at two or more times per week for at least 3 months using bicarbonate-buffered dialysate. exclusion criteria: 1) echocardiographic or clinical evidence of ischemic heart disease, lv dysfunction, valvular heart disease or previous renal transplantation. 2) predisposing clinical conditions to pulmonary hypertension, coad, interstitial lung diseases, primary pulmonary hypertension, congenital left to right shunt, chronic thromboembolic disease and connective tissue disorders. hd sessions were calculated as total hd sessions = frequency per week × 4 × number of months of hd. all of them underwent transthoracic echocardiography including 2d. m-mode and doppler studies of the rv. echocardiography was performed within 1 hour after completing hemodialysis in clinically stable patients with optimal dry weight to avoid any overestimation due to volume overload. images were obtained using ge vivid i portable ultrasound machine. statistical analysis: collected data was entered and analyzed using spss 20.0. data for age and tapse was described using mean ±sd if normally distributed and median (iqr) otherwise. data for rvd, pulmonary hypertension, smoking, dm, htn and total number of hd sessions was described by using frequency and percentages. furthermore, stratification of the study population was done based on the total number of hd sessions into 4 groups as: <200, 201-300, 301-400 &>400. comparison of tapse between the hd session groups was made using kruskal wallis test. comparison of rvd between the groups was made by using chi-square test. also the relationship between rvd and pulmonary hypertension was tested by the chi-square test. a p-value ≤ 0.05 was considered significant. results out of 145 patients 82(56.55%) were male while 63 (43.45%) were female. the average age of the patient was 53.46± 7.75 although the minimum age of patient was 20 years and maximum age was 78 years. mean duration of hd was 26.22 month with sd 9.0. the minimum duration of hd was 10 months and maximum duration of hd was 30 months. out of 145 patients, 50 (34.48%) patients were receiving twice-weekly hemodialysis while, 95 (65.52%) patients were on thrice-weekly hemodialysis sessions. as regard to risk factors, diabetes mellitus was found in 48 (33.10%) patients while 97 (66.90%) were non-diabetics. out of 145 patients, hypertension was found in 39(26.9%) patients, while 106 (73.1%) were non-hypertensive. fifty three patients were smokers while ninety two patients were nonsmokers. further sub analysis is mentioned in tables given below. the distribution with respect to hemodialysis sessions shown in table-1, rv dysfunction with respect to hemodialysis shown in table-2 and pulmonary hypertension is shown in table-3. comparison of tapse value in different groups are shown in table-4 & 5. 66 prevalence of right ventricular dysfunction & pulmonary hypertension and their relationship… hd sessions n % ≤ 200 29 20.0 201 300 51 35.2 301 400 60 41.4 401+ 5 3.4 total 145 100.0 table-1: distribution with respect to hemodialysis sessions hd sessions right ventricular dysfunction yes no total n % n % n % ≤ 200 4 13.8 25 86.2 29 100.0 201 300 19 37.3 32 62.7 51 100.0 301 400 31 51.7 29 48.3 60 100.0 401+ 5 100.0 0 0.0 5 100.0 total 59 40.7 86 59.3 145 100.0 table-2: stratification of rvd with respect to hd session cross tabulation hd sessions pulmonary hypertension yes no total n % n % n % ≤ 200 5 17.2 24 82.8 29 100.0 201 300 23 45.1 28 54.9 51 100.0 301 400 32 53.3 28 46.7 60 100.0 401+ 4 80.0 1 20.0 5 100.0 total 64 44.1 81 55.9 145 100.0 table-3: stratification of pulmonary hypertension with respect to hd session cross tabulation hd sessions tapse mean sd q1 median q3 ≤ 200 19.2 3.9 18.0 19.0 22.0 201 – 300 17.6 5.1 13.0 19.0 22.0 301 – 400 15.4 4.2 12.0 15.0 19.0 401+ 8.8 .8 8.0 9.0 9.0 total 16.7 4.9 12.0 18.0 20.0 table-4: tapse among 4 hd session groups hd sessions(i) hd sessions(j) mean difference (i) – (j) z p-value ≤ 200 201 300 1.67 -1.19 0.233 301 400 3.89* -3.85 < 0.001 401+ 10.44* -3.35 0.001 201 300 301 400 2.22 * -2.50 0.012 401+ 8.77* -3.39 0.001 301 400 401+ 6.55* -3.46 0.001 table-5: group wise comparison of tapse among 4 hd session groups discussion patients on long-term hemodialysis (hd) therapy at some stage develop rv dysfunction and pulmonary hypertension and it is more common in hd patients as compared to the patients on peritoneal dialysis (pd).10 we did this study with a view to determine the prevalence of rv dysfunction and pulmonary hypertension and their relation to the total number of dialysis sessions as local data are lacking in this regard. although there are some international studies, they showed inconsistent results. rv dysfunction was seen in 59 (40.7%) of the patients in our study. the frequency increased with increase in total hd sessions. the patients who had less than 200 sessions in total had rvd prevalent among 13.8%, while, 37.3% among cases with sessions 201-300 and 51.7% among cases with 301400 sessions. there were 5 cases with more than 400 sessions and all had rvd. the total sessions of hd had a significant association with p-value <0.001 (table-2). paneni et al. showed 71.3% prevalence of rvd in hd patients and found further that the prevalence was more in hd than pd and brachial than radial avf10. another study showed that rv dysfunction is increased in hd patients in which they compared the hd patients with the controls.15 however these studies did not compare the effects of the total number of dialysis sessions on the rv dysfunction. regarding phtn, yigla m et al in their study reported its prevalence of 40% in hd patients.19 our study has similar results with 44.1% prevalence in the hd patients. like rv dysfunction (rvd) the pulmonary hypertension (phtn) was also prevalent in increasing order with the increase of hd sessions. the minimum prevalence was 17.2% among group with total sessions less than 200 and 80.0% among those with more than 400 sessions (table-3). it was observed that there were 34 (23.4%) cases that had rvd as well as phtn while in 56 (38.6%) both were absent. there were 25 cases that had rvd but no phtn and similarly there were 30 cases that had phtn but no rvd. still the association between rvd and phtn was statistically significant with pvalue 0.011. present study reported that the rvd parameter viz. tricuspid plane systolic excursion (tapse) value decreased significantly with increasing duration of hd sessions. the mean tapse value for the group with hd sessions less than 200 was found to be 19.2±3.9mm with median of 19.0(18.0-22.0). the median (iqr) for other three groups were 19.0 (13.0-22.0), 15.0 67 prevalence of right ventricular dysfunction & pulmonary hypertension and their relationship… (12.0-19.0) and 9.0 (8.0-9.0) respectively. when compared among groups by using kruskal wallis test it was noted that the difference of tapse among four groups by hd sessions was highly significant (p-value <0.001). thus, it is evident that once the dialysis crosses 400 sessions the tapse value starts falling in the rv dysfunction range (table-4). mohamed momtaz et al. reported a statistically significant decrease in tricuspid plane systolic excursion (tapse; 2.2±0.94 vs. 2.1±0.63 cm) in hd group as compared to the control group.15 but unlike this study it was not correlated with the number of hd sessions. in our study majority (65.52%) of patients was on thrice weekly hd and the rest were on twice weekly hd. the proportion of men was greater than that of the women (56.55% vs. 43.45% respectively) similar to the study by momtaz m et al. in which they studied the rv dysfunction in hd patients in which men were 62% and women were 38%.15 so our results are comparable with other studies but the effect of number of hd sessions on rv function and phtn was significant to make this study different from other studies. limitations: there is no control group, the number of patients were less and that it is a cross-sectional study. the relationship of frequency, dose, length and the duration of dialysis on the rv dysfunction and pulmonary hypertension needs to be addressed in larger prospective studies. conclusion this study confirmed the higher prevalence of rv dysfunction and pulmonary hypertension among esrd patients with higher sessions of hd. so, periodic echocardiography should be considered for early detection of cardiac abnormalities and prioritize such patients for early renal transplant. references 1. mohiud din k, bali hk, banerjee s, sakhuja v, jha v. silent myocardial ischemia and high-grade ventricular arrhythmias in patients on maintenance hemodialysis. ren fail 2005; 27:171-75. 2. trespalacios fc, taylor aj, agodoa ly, bakris gl, abbott kc. heart failure as a cause for hospitalization in chronic dialysis patients. am j kidney dis 2003; 41:1267-77. 3. foley rn, parfrey ps, sarnak mj. clinical epidemiology of cardiovascular disease in chronic renal disease. am j kidney dis 1998; 32 (suppl 3): s112–19. 4. collins aj, ma jz, umen a, keshaviah p. urea index and other predictors of hemodialysis patient survival. am j kidney dis 1994; 23:272-82. 5. foley rn, herzog ca, collins aj. smoking and cardiovascular outcomes in dialysis patients: the united states renal data system wave 2 study. kidney int 2003; 63:1462-67. 6. curtis bm, parfrey ps. congestive heart failure in chronic kidney disease: disease-specific mechanisms of systolic and diastolic heart failure and management. cardiol clin. 2005; 23:275-84. 7. d’alonzo ge, barst rj, ayres sm, bergofsky eh, brundage bh, detre km, et al. survival in patients with primary pulmonary hypertension. results from a national prospective registry. ann intern med 1991; 115: 343-49. 8. piazza g, goldhaber sz: the acutely decompensated right ventricle: pathways for diagnosis and management. chest 2005; 128: 1836-52. 9. gulel o, soylu k, yuksel s, karaoglanoglu m, cengiz k, dilek m, et al. evidence of left ventricular systolic and diastolic dysfunction by color tissue doppler imaging despite normal ejection fraction in patients on chronic hemodialysis program. echocardiography. 2008; 25:569-74. 10. paneni f, gregori m, ciavarella gm, sciarretta s, de biase l, marino l, et al. right ventricular dysfunction in patients with end-stage renal disease. am jnephrol. 2010; 32:432-38. 11. kittipovanonth m, bellavia d, chandrasekaran k, villarraga hr, abraham tp, pellikka pa: doppler myocardial imaging for early detection of right ventricular dysfunction in patients with pulmonary hypertension. j am soc echocardiogr 2008; 21:1035-41. 12. wang ay, wang m, lam cw, chan ih, zhang y, sanderson je: left ventricular filling pressure by doppler echocardiography in patients with endstagerenal disease. hypertension 2008; 52: 107-14. 13. di lullo l, floccari f, rivera r, granata a, d’amelio a, logias f, et al. the nephrologist and the role of ultrasound imaging in the diagnosis of cardiorenal syndrome. g ital nefrol. 2012;29:321-27 14. di lullo l, floccari f, polito p. right ventricular diastolic function in dialysis patients could be affected by vascular access. nephron clinical practice. 2011; 118(3):c257-61. 15. momtaz m, fishawy ha, aljarhi um, al-ansi rz, megid ma, khaled m. right ventricular dysfunction in patients with end-stage renal disease on regular hemodialysis. eur j int med. 2013; 25:127-32. 16. rashid aa, soleiman ar, nikoueinejad h. the evaluation of increase in hemodialysis frequency on c-reactive protein level and nutritional status. acta medical ranica. 2013; 51(2):119-24. 17. kdigo 2012 clinical practice guideline for the evaluation and management of chronic kidney disease. kdigo. 2013; 3:18-34. 68 prevalence of right ventricular dysfunction & pulmonary hypertension and their relationship… 18. nakhoul f, yigla m, gilman r, reisner sa, abassi z. the pathogenesis of pulmonary hypertension in haemodialysis patients via arterio-venous access. nephroldial transplant. 2005; 20:1686-92. 19. yigla m, nakhoul f, sabag a, tov n, gorevich b, abassi z, et al. pulmonary hypertension in patients with end-stage renal disease. chest journal. 2003 may1; 123(5):1577-82. the authors: dr. rakesh bahadur adhikari clinical & interventional cardiologist, shahid gangalal national heart centre (sgnhc), kathmandu, nepal. prof. qazi abdul saboor head, department of cardiology, shaikh zayed medical complex, lahore. dr. mateen akram assistant professor, department of nephrology, shaikh zayed medical complex, lahore. dr. saira fayyaz consultant cardiologist, city care hospital, khanewal dr. husnain bashir medical officer, department of cardiology, shaikh zayed medical complex, lahore. dr. muhammad rizwan-ul-haque associate professor, department of nephrology, shaikh zayed medical complex, lahore. corresponding author prof. qazi abdul saboor head, department of cardiology, shaikh zayed medical complex, lahore. email: drsaboor04@gmail.com untitled-1 41 proceedings s.z.m.c. vol: 34(3): pp. 41-45, 2020. pszmc-760-34-3-2020 an association of vitamin d insufficiency with elevated serum uric acid levels among postmenopausal women 1mariya ali, 2tahira naseem, 3farooq abdul hakim bajwa, 4saira zafar, 4asma akram, 4hirra ghaffar 1department of chemical pathology, continental medical college, lahore 2department of biochemistry & chemical pathology, shaikh zayed medical complex, lahore 3department of radiology, lahore general hospital, lahore 4department of microbiology, continental medical college, lahore abstract introduction: deficiency of vitamin d and raised levels of serum uric acid are considered independent risk factors for causing cardiovascular diseases. aims & objectives: this study was carried out to study serum uric acid and vitamin d levels in postmeopausal women to find association of hypovitaminosis d with hyperuricemia. place and duration of study: it was a cross-sectional analytical study. postmenopausal women more than 55 years of age with bmi between 25-30 kg/m2 were included in the study. the study was conducted at the department of biochemistry and chemical pathology, shaikh zayed federal postgraduate medical institute, lahore over a period of six months. material & methods: eighty subjects were included in this study. they were divided into two groups i-e, normouricemic group and hyperuricemic group based onserum uric acid level.vitamin d levels of both groups were then measured. results: the hyperuricemic group had markedly lower vitamin dlevels as compared to normouricemic group. moreover, vitamin d had a negative correlation with uric acid in the hyperuricemic group i-e, higher the uric acid level, lower was vitamin d. multiple regression analysis was also performed for studying the relation of vitamin d with uric acid levels in the two groups overall. it was observed that one mg/dl increase in uric acid caused an average decrease of 2.43 ng/ml in vitamin d. conclusion: it may be concluded from this study that association of hypovitaminosis d with hyperuricemia in postmenopausal women can be used for planning an early intervention to prevent cardiovascular diseases in them. key words: hyperuricemia, hypovitaminosis d, cardiovascular diseases introduction hyperuricemia is considered to be a predictor for cardiovascular diseases1 and at the same time deficiency of vitamin d is also considered an independent risk factor for the development of cardiovascular diseases.2 hypovitamnosis d and hyperuricemia are being considered major public health concerns worldwide and the prevalence has been increasing in both developed and developing countries.3 vitamin d deficiency is defined as the serum level of 25-hydroxy vitamin d less than 20ng/ml while a serum level of 25 hydroxy vitamin d in the range 20-29.9ng/ml falls into the category of vitamin d insufficiency.4 approximately 1 billion people throughout the world are suffering from vitamin d deficiency.5 a study conducted in pakistan on about 200 postmenopausal women showed that 59% of these women were vitamin d deficient.6 it has been documented that risk of myocardial infraction doubles in patients having vitamin d levels <34ng/ml.7 studies show that serum vitamin d of more than 30ng/ml is implicated in preventing cardiovascular diseases.8 uric acid (2,6,8-trihydroxypurine) is a naturally occurring nitrogenous waste product in the body.9 the reference interval in plasma or serum for females is 2.6-6.0 mg/dl and in the twenty four hour urine of adult it is 250-750 mg/dl.10 hyperuricemia is a metabolic disorder characterized by either raised uric acid synthesis or decreased renal excretion.11 women are considered to have hyperuricemia if their serum uric acid level is ≥ 6.0 mg / dl.15 in pakistan about 27% of adult females have been found to be hyperuricemic.12 hyperuricemia is found to be more prevalent in postmenopausal 42 an association of vitamin d insufficiency with elevated serum uric acid levels among postmenopausal women women, some ethnic groups like african americans and patients with renal diseases.13 raised serum uric acid (sua) in postmenopausal women is linked to the lack of estrogen which is believed to be involved in the excretion of uric acid.14 excess uric acid has been related to the risk of hypertension, atherosclerosis and cardiovascular conditions.15 relationship of raised sua level with vitamin d insufficiency is a significant metabolic disorder that is associated with cardiovascular diseases.16 low levels of vitamin d cause the release of parathyroid hormone through the activation of parathyroid gland.17 some previous studies showed that high levels of parathyroid hormone increase the occurrence of hyperuricemia.18 it is believed that there are some absorptive and secretory transporters in renal tissue at the basolateral and apical membranes which are involved in the transportation of uric acid.19 parathyroid hormone affects some of the transporters of proximal tubules of kidney, for example, its inhibition of sodium/hydrogen ion exchange type 3 isoform is known.20 renal secretion of uric acid is reduced among hyperuricemic patients having high levels of parathyroid hormone, the severity of which relies on the change in the rate of absorption.21 the current study has been designed to study serum uric acid and vitamin d levels and to find out the relationship of low levels of vitamin d with high serum uric acid levels among postmenopausal women in our population. due to an increasing trend of hyperuricemia and hypovitaminosis d in the middle aged and elderly population, the present study might have significant public health implications and it may also help out in planning an appropriate intervention to prevent cardiac diseases in these patients. improved exposure to the sun, better diet, supplementation of vitamin d and measures to reduce serum uric acid may therefore be utilized to improve general health, normalize serum uric acid and prevent cardiovascular diseases. material and methods it was a “cross-sectional analytical study”.the samples were collected from a medical camp in bajwa clinic, kahna, ferozpure road, lahore. sample size was 80 which was estimated for each group by using 95% confidence level.90% power of test with expected mean vitamin d levels of 43±12 and 35±10 in uric acid normal and uric acid elevated group respectively.22 this was calculated by using power and precision 3.0 software. the sampling technique was non probability convenient sampling. postmenopausal women above 55 years of age and having bmi between 25-30 were included in the study. patients with self-reported gout or arthritis, self-stated parathyroid or thyroid disease, having used uric acid lowering medicine in last 14 days, deranged liver function tests, history of chronic renal disease, tumours and cardiac disease were excluded from the study. after approval from institutional review board of fpgmi (letter no.f38/nhrc/admin/irb/226), consent forms were filled and signed by the participants of study. serum uric acid assay was done by uricase method on fully automated chemistry autoanalyzer au480 by beckmann coulter and subjects were divided into two groups based on their uric acid level i.e, normouricemic and hyperuricemic. each group comprised of 40 postmenopausal women. vitamin d levels of both groups were then measured by chemiluminescent immunoassay on fully automated analyzer access 2 by beckman coulter. statistical analysis: data analysis was performed using latest version of spss 22.0. mean ± sd and median of the two parameters was obtained. statistical significance was calculated by student’s t-test and mann whitney u test. p<0.05 was considered significant. step wise regression analysis was done to determine the effect of sua on vitamin d. results the study included 80 postmenopausal women, 40 having normal uric acid level and 40 hyperuricemic. the mean age for normouricemic group was 59.4±4.4 years and that of hyperuricemic group was 61.1±3.5 years. on comparing the vitamin d levels of both groups it was found that hyperuricemic group had relatively close range of vitamin d levels while for normouricemic the range was quite wide with a significant p-value of 0.006. in normouricemic group there were 12 cases of vitamin d deficiency while 19 cases were of vitamin d insufficiency. on the other hand in hyperuricemic group there were 19 cases of vitamin d deficiency and 19 cases fell into vitamin d insufficiency. pvalue of 0.049was significant on comparison of both groups. 43 an association of vitamin d insufficiency with elevated serum uric acid levels among postmenopausal women group normouricemic hyperuricemic vitamin d (ng/ml) mean 24.9 19.5 sd 12.3 5.0 q1 18.2 17.3 median 23.7 20.1 q3 28.2 22.2 mann whitney = 516.5 p-value* = 0.006 vitamin d status < 20.0 (deficient) 12 19 20.0 29.9 (insufficient) 19 19 30.0+ (sufficient) 9 2 chi-square = 6.04 p-value = 0.049 table-1: vitamin d levels of study subjects of both groups based on sua level. p * means that p value is < 0.05 (significant) fig-1: boxplot presenting distribution for vitamin d levels amongst both group of study subjects. the correlation analysis was performed to see the association of vitamin d with uric acid. this analysis was carried out within each group and overall. it was noted that overall uric acid had significant negative correlation with vitamin d with r= -0.471 with p-values < 0.001. when the correlation analysis was performed in hyperuricemic group it was noted that the uric acid level had a good negative correlation of -0.577 with vitamin d showing that higher the uric acid levels lower the vitamin d levels. the multiple linear regression analysis was done to see the effect of uric acid on vitamin d. result found at 17th step showed that the final significant model was decided. it was noted that the uric acid had a significant impact on vitamin d level. when all 80 cases were included, the average vitamin d level was 35.53ng/ml and one unit increase in uric acid caused an average decrease of 2.43ng/ml in vitamin d. model unstandardized coefficients t p-value b std. error 17 (constant) 35.53 3.52 10.11 0.000* uric acid (mg/dl) -2.43 0.61 -3.94 0.000* table-2: multiple linear regression analysis for studying relation of vitamin d with uric acid by using backward method in both groups combined (r2 = 0.166 adj r2= 0.156) dependent variable: vitamin d (ng/ml) the same model was applied on hyperuricemic group and it was noted that in this group serum uric acid had highly significant and negative effect on vitamin d with p-values <0.050. this model was obtained in 12th step with adjusted r2 0.555 showing reasonably good fit model. an increase of one mg/dl in uric acid level in this group caused an average decrease of 2.02 ng/ml in vitamin d. discussion eighty postmenopausal women were included in this study. they were divided into two groups: normouricemic and hyperuricemic groups. the difference between the ages of hyperuricemic and normouricemic groups was insignificant. on comparing vitamin d levels of both groups it came into notice that mean vitamin d level for hyperuricemic group was lower as compared to normouricemic subjects. the difference was significant with p-value of 0.006.the results are similar to a study by takir m et al in which hyperuricemic group had lower values of vitamin d as compared to normouricemic subjects. there was a negative and important relationship between sua and vitamin d.23 similar findings were also seen in another study done in turkey. in this research vitamin d and uric acid levels of the patients were obtained. their findings suggested that subjects having increased sua levels had low levels of vitamin d. they concluded that low level of vitamin d were correlated with high serum uric acid levels.24 correlation analysis was performed to see the relationship of vitamin d with uric acid within each group and overall. when multiple regression analysis was performed it was found that uric acid had a significant impact on vitamin d. these results are in agreement with a study done in china in which multiple regression analysis was performed and it was found that there was an independent and significant relationship between vitamin d insufficiency and high sua levels among the postmenopausal women. the subjects with vitamin 44 an association of vitamin d insufficiency with elevated serum uric acid levels among postmenopausal women d insufficiency were more likely to have raised sua compared with those without vitamin d insufficiency among postmenopausal women. their findings suggested that vitamin d status is negatively associated with elevated serum uric acid. they also summarized that exposure to the sunlight and vitamin d supplementation are easy as well as an inexpensive ways for preventing low levels of vitamin d and other health issues.22 another study conducted in saudi arabia concluded that vitamin d was inversely related with serum uric acid level.25 a study from egypt also concluded that low level of vitamin d was inversely correlated with high serum uric acid level.26 a study carried out in turkey suggested that patients who have elevated serum uric acid had lower vitamin d levels.27 another research carried out on 100 patients for finding out the possible effect of vitamin d on uric acid level concluded that there was an inverse relationship between vitamin d and serum uric acid.28 a systemic review was conducted in 2019 which showed that patients with vitamin d deficiency and insufficiency had a significantly higher level of serum uric acid compared with individuals with normal vitamin d.29 conclusion it may be concluded from our study that postmenopausal women with higher serum uric acid level had lower level of vitamin d as compared to those who had normal serum uric acid levels. a negative correlation was observed between serum uric acid and vitamin d level in the group with hyperuricemia. low vitamin d and raised serum uric acid are important risk factors for the development of cardiovascular diseases. by improving vitamin d level, serum uric acid can be normalized. association of hypovitaminosis d with hyperuricemia in postmenopausal women can be used in planning an early intervention to prevent cardiac diseases in such subjects. limitations of study: it was a single center study with a small sample size which may not be sufficient to correlate low levels of vitamin d with elevated sua. a study should be carried out with larger sample size and in different socio economic strata to get more accurate information. references 1. borghi c, cicero af. serum uric acid and acute coronary syndrome: is there a role for functional markers of residual cardiovascular risk?. int. j. cardiol. 2018; 250:62-3. 2. kim sh, oh je, song dw, cho cy, hong sh, cho yj, yoo bw, shin ks, joe h, shin hs, son dy. the factors associated with vitamin d deficiency in community dwelling elderly in korea. nutr res pract. 2018; 12(5):387-95. 3. liu h, zhang x m, wang y. l, liu b-c. prevalence of hyperuricemia among chinese adults: a national cross-sectional survey using multistage, stratified sampling. jnephrol. 2014; 27(6):653-8. 4. pfotenhauer km, shubrook jh. vitamin d deficiency, its role in health and disease, and current supplementation recommendations. j am osteopath assoc. 2017; 117(5):301-5. 5. ilson lr, tripkovic l, hart kh, lanham-new sa. vitamin d deficiency as a public health issue: using vitamin d 2 or vitamin d 3 in future fortification strategies. brit j nutr. 2017; 76(3):392-9. 6. akhtar s, jan r. prevalence of vitamin d inadequacy among postmenopausal women. med forum mon j spec. 2019; 24(11):58-9. 7. kheiri b, abdalla a, osman m, ahmed s, hassan m, bachuwa g. vitamin d deficiency and risk of cardiovascular diseases: a narrative review. j. clin. hypertens. 2018; 24(1):1-9. 8. mozos i, marginea o. links between vitamin d deficiency and cardiovascular diseases. biomed res. intern. 2015; (12). 9. hafez rm, abdel-rahman tm, naguib rm. uric acid in plants and microorganisms: biological applications and genetics-a review. j. adv. res. 2017; 8(5):475-86. 10. maiuolo j, oppedisano f, gratteri s, muscoli c, mollace v. regulation of uric acid metabolism and excretion. int. j. cardiol. 2016; 213:8-14. 11. yokokawa h, fukuda h, suzuki a, fujibayashi k, naito t, ueharo a, et al. association between serum uric acid levels /hyperuricemia and hypertension among 85, 286 japenese workers. j clin hypertens. 2016; 18(1):53-9. 12. raja s, kumar a, ahooja r, et al. frequency of hyperuricemia and its risk factors in adult population. cureus. 2019; 11(3):4198-4202. 13. cho sk, winkler ca, lee sj, chang y, ryu s. the prevalence of hyperuricemia sharply increases from the late menopausal transition stage in middle-aged women. j. clin. med. 2019; 8(3):296. 14. naseem r, zafar sm, jawed s, mukhtar s, ijaz f, aftab rk. influence of serum estradiol on serum uric acid level in pre and postmenopausal women. prof. med. j. 2019; 26(09):1587-91. 45 an association of vitamin d insufficiency with elevated serum uric acid levels among postmenopausal women 15. muiesan ml, agabiti-rosei c, paini a, salvetti m. uric acid and cardiovascular disease: an update. eur cardiol. 2016; 11(1):54. 16. thakkinstian a, anothaisintawee t, chailurkit l, ratanachaiwong w, yamwong s, sritara p, ongphiphadhanakul b. potential causal associations between vitamin d and uric acid: bidirectional mediation analysis. scientific reports. 2015; 5:14528. 17. martins js, palhares md, teixeira oc, gontijo ramos m. vitamin d status and its association with parathyroid hormone concentration in brazilians. nutr metab. jan 1 2017. 18. sugimoto r, watanabe h, ikegami k, enoki y, imafuku t, sakaguchi y, murata m, nishida k, miyamura s, ishima y, tanaka m. downregulation of abcg2, a urate exporter, by parathyroid hormone enhances urate accumulation in secondary hyperparathyroidism. kidney int. 2017; 91(3):658-70. 19. girardi c, titan s, malnic g, reboucas n. chronic effect of parathyroid hormone on nhe3 expression in renal proximal tubules. kidney int .2000; 58(4):1623-31. 20. hisatome i, ishimura m, sasaki n. renal handling of urate in two patients with hyperuricemia and primary hyperparathyroidism. int med j. 1992; 31(6):807-11. 21. wen c, david c, chan h, tasai m, chung w. is a high serum uric acid a risk marker or target for treatment. am j nephrol. 2010;56(2):273-88 22. peng h, li h, li c, chao x, zhang y. association between vitamin d insufficiency and elevated serum uric acid among middle aged and elderly chinese han women. pubmed. 2013; 8(4):1-9. 23. takir m, solak y, erek a, kostek o, oral a, elcioglu o, et al. association between serum uric acid and vitamin d insufficiency among middle aged and elderly population. turk neph dial transpl. 2016; 25(2):182-6. 24. takir m, tprok e, elcioglu c, bakan a, kostek a, cakhh t, et al. is there a link between vitamin d and uric acid?. clin chem lab med. 2014; 52(1):22-6. 25. gehan et al. relationship between hyoeruricemia and vitamin d deficiency among adults in al kharj, saudia arabia. intr j adv res. 2015; 3(1):269-273. 26. khalid al syed, magdy elasyed, reem sabry. relationship between serum uric acid and vitamin d among elderly egyptians. j am sci. 2014; 9(12):984-988. 27. yandi y, geenra f, kilauz b, bakin a, sahim h, billi m et al. cyanocobalamin and 25 hydroxy vitamin d levels in gout; an overlooked issue. ojra an acadm publsh.2016:6(4):96-101 28. aakay e, korakaelci f, gurosay g, karabag y, yilidiz m, erodgu h, et al .the possible effect of vitamin d on uric acid in diabetic patients. world clin j med sci. 2017; 1(2):77-83. 29. charoenngam n, ponvilawan b, ungprasert p. vitamin d insufficiency and deficiency are associated with a higher level of serum uric acid: a systematic review and meta-analysis. modern rheumatology. 2019 mar4. the authors: dr. mariya ali, demonstrator, department of chemical pathology, continental medical college, lahore. dr. tahira naseem hod, biochemistry and chemical pathology, shaikh zayed medical complex, lahore. dr. farooq abdul hakim bajwa, medical officer, department of radiology, lahore general hospital, lahore. dr. saira zafar, assistant professor, department of microbiology, continental medical college, lahore. dr. asma akram, demonstrator, department of pathology, continental medical college, lahore. hirra ghaffar demonstrator, department of pathology, continental medical college, lahore. corresponding author: dr. mariya ali, demonstrator, department of chemical pathology, continental medical college, lahore. e-mail: drmariafarooq@hotmail.com for web full book 14 proceedings s.z.m.c. vol: 36(2): pp. 14-18, 2022. pszmc-838-36-2-2022 1department of anatomy, fmh college of medicine & dentistry, lahore 2 department of neurology, fmh college of medicine & dentistry, lahore 3department of physiology, fmh college of medicine & dentistry, lahore 4department of anatomy, azra naheed medical college, lahore 5department of anatomy, rahbar medical college, lahore 6department of anatomy, akhtar saeed medical and dental college, lahore 7department of pediatric medicine, fmh college of medicine & dentistry, lahore 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 15 morphometry of foramen magnum: an anatomical study 16 morphometry of foramen magnum: an anatomical study 17 morphometry of foramen magnum: an anatomical study 18 morphometry of foramen magnum: an anatomical study for web full book 29 proceedings s.z.m.c. vol: 36(2): pp. 29-34, 2022. pszmc-841-36-2-2022 1department of pharmacology, shifa college of medicine, shifa tameer-e-millat university, islamabad 2department of pathology, shifa college of medicine, shifa tameer-e-millat university, islamabad 3department of biochemistry, shifa college of medicine, shifa tameer-e-millat university, islamabad 4shifa college of medicine, shifa tameer-e-millat university, islamabad 5department ofphysiology, shifa college of medicine, shifa tameer-e-millat university, islamabad r= r= 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 30 correlation between post exercise heart rate recovery and body composition in healthy female adults 31 correlation between post exercise heart rate recovery and body composition in healthy female adults p r p p 32 correlation between post exercise heart rate recovery and body composition in healthy female adults 33 correlation between post exercise heart rate recovery and body composition in healthy female adults 34 correlation between post exercise heart rate recovery and body composition in healthy female adults numbering.indd 64 proceedings s.z.m.c. vol: 34(4): pp. 64-69, 2020. pszmc-773-34-4-2020 secondary heamorrhage after cold steel tonsillectomy: an audit performed in shalamar hospital 1arshadullah afridi, 2sana tariq, 3muhammad dawood, 4sarwat iqbal, 5usna athar, 1aisha hanif 1department of ent, shalamar hospital, lahore 2department of bioethics, shalamar medical & dental college, lahore 3department of forensic medicine, shalamar hospital, lahore 4department of medicine, shalamar hospital, lahore 5department of community medicine, shalamar hospital, lahore abstract introduction: tonsillectomy is a common surgical procedure in which post tonsillectomy heamorrhage remains the most serious complication. the cold steel dissection method is the oldest and safest method used to perform tonsillectomies. aims & objectives: this study was done to see the frequency of posttonsillectomy heamorrhage with cold steel dissection method. it was a clinical audit performed retrospectively to compare our results with international standards. place and duration of study: our clinical audit was conducted at otolaryngology department in shalamar hospital, lahore, pakistan. this study was conducted in a period of 6 months. material & methods: the charts of patients (n=344) that underwent tonsillectomy at shalamar hospital from january 2016 till december 2019 were retrospectively viewed, in a cross-sectional, pilot study, using convenience sampling technique. all the tonsillectomy procedures were performed using the cold steel dissection method. frequencies were recorded and chi-square test was applied. results: the complication of secondary heamorrhage occurred in 7 patients and the frequency was calculated as 2%. a significant difference (p=0.01) of age was found in patients suffering from secondary heamorrhage. the incidence was lower in patients below 18 years of age (0.7%) as compared to patients above 18 years of age (7.14%). the rate of heamorrhage was found to be slightly higher in males than in females (2.6% and 1.31% respectively). conclusion: we conclude that the rate of secondary heamorrhage in our institution is up to the rate reported in literature. different hot techniques for tonsillectomy should also be performed in clinical settings and compared with the standard cold steel dissection tonsillectomy. key words: tonsillectomy, adeno-tonsillectomy, secondary heamorrhage, cold steel method introduction tonsillectomy is one of the most common and frequently performed surgical procedures in otolaryngology practices in which both palatine tonsils are removed from the throat. in some cases, the adenoid may also be removed which is then termed as "adeno-tonsillectomy".1 it is often categorized as low risk operation with quite a few chances of complications whilst the post tonsillectomy heamorrhages remains the most serious complication.2 post-tonsillectomy heamorrhages are fairly divided into two main categories: primary heamorrhage in which postoperative bleeding occurs within 24 hours after the surgery while the patient is in hospital, and the secondary heamorrhage which occurs up to 2 weeks post operatively.3 to reduce both the intra and post-operative compilations, and to minimize the surgery duration and morbidity, the tonsillectomy techniques are evolving constantly.4 tonsillectomy can be performed by various methods which are divided into two major categories including ‘cold’ and ‘hot’ methods. dissection and snare, guillotine method, intra-capsular, harmonic scalpel, plasma mediated ablation technique and cryosurgical techniques are the cold methods to perform tonsillectomy.5 the hot methods include; electrocautery, laser tonsillectomy using (co2 or ktp), colbation tonsillectomy and radio frequency.6 however, it cannot be clearly stated that which of these techniques shows ascendancy over the other.4 65 secondary heamorrhage after cold steel tonsillectomy: an audit performed in shalamar hospital primarily, the main indications for tonsillectomy are based on obstructive and infectious causes. a surgery is required when there are periodic episodes of throat infections whose record is maintained through proper documentation. the physician might recommend tonsillectomy if the frequency of throat infection is of seven episodes per year, five episodes per 2 years or at least three episodes per 3 year, including the history ofpfapa syndrome ( periodic fever, aphthous stomatitis, pharyngitis and cervical adenitis) , peri-tonsillar abscesses multiple antibiotic allergy and drug intolerance.1 other indicators such as obstructive sleep apnoea syndrome, dysphonia, dysphagia, suspected malignancy, chronic tonsillitis, halitosis, tonsilloliths are also the reasons for considering tonsillectomy.1,7 the cold steel dissection method was introduced about 100 years ago, in which the blunt dissection is done involving a metal instrument and the subsequent bleeding is controlled with the help of gauze.8 the cold steel method is often considered as the gold standard because every other innovative techniques for tonsillectomy are compared with it.9 it is also suggested that the young ent trainees must be taught to perform tonsillectomy with cold method first.10 the primary rationale behind this institutional review and analysis is to determine the frequency of secondary heamorrhages over the period of four years in shalamar medical and dental college and shalamar hospital. this will help in ascertaining the risk factors associated with cold steel dissection tonsillectomy. this clinical audit may prove beneficial as a scaffold for comparing the results with different studies in order to monitor and modify our practices. this would help in improving patients’ clinical outcomes. clinical audits and reviews are essential in health care services. as clinical practices and guidelines are changing with time and the advent of technology, surgeons must put their work under a microscope and modify their techniques accordingly for a state-of-the-art health care provision. material and methods a retrospective, cross sectional study was conducted in ear, nose and throat department (ent) of shalamar hospital. this study can be classified as a pilot audit conducted by the authors. ethical approval was taken from the institutional review board (irb) at shalamar institute of health sciences (sihs). after the irb approval, the study was completed within 6 months. since this is a clinical audit, so it needs no specific sample size calculation. convenience sampling was used; the study was conducted in duration of 6 months. the medical charts of the patients that underwent the tonsillectomy and/or adenotonsillectomy were prepared dating from january 2016 till december 2019. the data of total of 344 cases was obtained from the otolaryngology (ent) department of smdc which included both adults and children. the inclusion criteria was determined as patients with a history of recurrent attack of recurrent attacks of tonsillitis accompanied by clinical examination findings of high grade fever, palpable jugulodigastric lymph nodes, exudates on tonsil and absence of cough. all patients aged 3 years and above, who underwent the ‘cold steel dissection’ method for tonsillectomy were included. patients having suspicion of malignancy or undergoing tonsillectomy as a part of uvulopalatoplasty were excluded. according to the set pattern of surgery, operations were performed on each patient and the surgical technique followed for tonsillectomy was ‘cold steel dissection’ specifically. the tonsils were removed using tonsillar dissector and haemostasis was secured by ligation and bipolar electro-cautery. heamorrhage was monitored qualitatively in the post-operative phase during their stay in the hospital. the oropharyngeal cavity including the surgical site was examined every six hours using a tongue depressor and a light source to check for any possible bleeding. the patients were asked to report back to the hospital in case of any signs of bleeding, after discharge. statistical analysis: the data of tonsillectomy cases was analyzed by using spss version 20. frequency tables were formed. chi-square test was applied for analytical comparison and p-value was set as 0.05 with 95% confidence interval. results a total of 344 patients underwent tonsillectomy and adeno-tonsillectomy by the cold steel method only during january 2016 till december 2019. demographic features and frequency of secondary heamorrhage were studied. chi-square test was applied with the level of significance set at 0.05. figure 1 shows the frequency of operations done according to the age of the patients. 66 secondary heamorrhage after cold steel tonsillectomy: an audit performed in shalamar hospital fig-1: frequency of tonsillectomy cases against the age of the patients. table-1 illustrates other demographic features as well as the results. out of the total number of tonsillectomy cases, 337 cases did not experience secondary heamorrhage. only 7 cases reported postoperative heamorrhage so the rate of secondary heamorrhage recorded was 2%. there were zero cases reported in our hospital for mortality as a result of tonsillectomy. age (in years) mean 13.66 median 10 mode 7 std. deviation 10.11 range 52 minimum 3 maximum 55 sex male 192 (55.8%) female 152 (44.2%) type of surgery: adenotonsillectomy 137 (39.8%) tonsillectomy 207 (60.2%) rate of secondary haemorrhage haemorrhage 7(2%) table-1: characteristics of study participants undergoing tonsillectomy (n=344) shalamar institute of health sciences is a trust hospital where people from different financial backgrounds are treated. fig-2 represents the statistics of 4 consecutive years starting from 2016, where the mode of payment for the number of different patients is categorized into 3 categories accordingly. fig-2: comparison of ‘general, private & company’ patients of tonsillectomy over past 4 years. the data of the patients were divided into two groups on the basis of their age. our study shows that only 0.72% of patients under the years of 18 experienced secondary heamorrhage whereas for the patients older than 18 years of age the percentage was 7.14%. the rate of secondary heamorrhage was higher for the patient with age more than 18 years and this result was statistically significant (p value=0.01) in our study, the rate of secondary heamorrhage in males is 2.6% and for females it is 1.31%. there is no significant difference between male and female regarding the rate of secondary heamorrhage (p value=0.401) in our study, no statistically significant difference was recorded between adeno-tonsillectomy and tonsillectomy regarding the rate of secondary heamorrhage (p value=0.869). discussion the rate of secondary heamorrhage in our study is 2% which is higher than the rates reported by o'leary and vorrath (1.85%) but lower than described by brick et al. 4.23%.4,11 harju and numminen reported higher secondary heamorrhage rate (12%) as 208 patients out of total 1734 patients who underwent tonsillectomy experienced secondary haemorrhage.12 when looking at the two groups divided on the basis of age, 0ur study showed similar results to the one done by walker p, in which post-tonsillectomy heamorrhage for 18 years old patients was 10.1% and age was defined as a highly statistically significant (p value=0.001) risk factor for posttonsillectomy haemorrhage.13 the cold steel dissection method has been preferred in older children, adolescents and adults.9 0 5 10 15 20 25 30 35 40 3 6 9 12 15 18 21 24 27 30 33 37 40 45 53 fr eq ue nc y age 16 18 13 13 45 25 28 16 26 8 10 9 0 5 10 15 20 25 30 35 40 45 50 2019 2018 2017 2016 general private company 67 secondary heamorrhage after cold steel tonsillectomy: an audit performed in shalamar hospital in contrast to our findings, ali et al. shows a high rate of post tonsillectomy bleeding (6.68%) in and it was indicted that the bleeding rate was higher in males as compared to the females (7.66 and 5.9% respectively).14 in another comparative study, the group of patients (n=40) subjected to the cold steel dissection tonsillectomy showed 2.5% of secondary heamorrhage rate. it was also indicated that the patients who underwent tonsillectomy with the cold steel method showed quick healing with lesser amount of pain as compared to the other group patients subjected to bipolar diathermy.15 preforming tonsillectomy through the cold steel method is quite a common practice even in the united states of america as it is characterized as cheaper and safer.16 as the tissue healing is better in cold steel method, it is reported as less painful technique in the literature.17 cold steel dissection method may have certain advantages but surgeons should be trained in several other techniques of tonsillectomy and should be permitted to use different methods for individual patients as well. extreme precautions are required for cold dissection in order to avoid the vascular damage. regardless of the operative technique, the frequency of post-operative bleeding also depends upon the skills and experiences individual surgeon.18it has been seen in a study that the complication of post-tonsillectomy heamorrhageis reportedly higher when young resident perform the coblation technique. hence, when younger surgeons are to perform this specific technique, supervision and surveillance are extremely vital.19 to further emphasize this, a research described that the rate of post-tonsillectomy heamorrhage was not dependent on the method used specially when the consultants performed the surgery.20 post-operative administration of dexamethasone was also seen to raise the frequency of heamorrhage after tonsillectomy.21 male gender and age were also studied as some of the risk factors for increased post-tonsillectomy heamorrhage rates and returns to the operating table.22 the complication of post-tonsillectomy heamorrhage has high mortality and thus must be evaluated with priority. some patients with this particular complication may be managed conservatively, but most of them need an intervention ranging from cautery to external carotid artery ligation.23 rising rates of secondary heamorrhage after tonsillectomy can be reduced by employing quality improvement projects like the one used in sweden which included hands-on workshops and supervision.24 various quality assurance programs targeting the local hospitals and clinics can improve the clinical outcomes, which is an indicator of health care provision in a country.25 an important component of such quality improvement programs is clinical audit, much similar to the one performed by us. a regular audit and maintenance of a registry can provide clinicians and surgeons with evidence to modify practice guidelines. conclusion there are many studies published showing the frequency of secondary heamorrhage after tonsillectomies but no study has been published containing data from shalamar institute of health sciences. it was seen in our pilot study that the frequency of secondary heamorrhagic complication was similar to what was reported in other peerreviewed publication. this in turn exhibits that standard techniques are being applied by our surgeons performing tonsillectomies. the introduction of electronic surgical performa along with the manual notes has resulted in improving the quality of data maintenance for routinely performed tonsillectomies. as evident, there is a low amount of data available right now, over the years the reauditing for different factors will become systematic and easy. limitations: there is a lot of literature available on this topic with the with sample size ranging in thousands. since before 4 years the electronic data system did not exist, our study only comprises of records of the past 4 years hence the sample size is low. in other studies, the comparison of different tonsillectomy technique is analyzed. however, in shalamar medical and dental college tonsillectomy is only performed with cold steel dissection method. the proper documentation of the initial indicators for performing tonsillectomy is also lacking. references 1. mitchell rb, archer sm, ishman sl, rosenfeld rm, coles s, finestone sa, et al. clinical practice guideline: tonsillectomy in children (update). 2019;(4)s1-s42. 2. patel a, foden n, rachmanidou a. is weekend surgery a risk factor for post-tonsillectomy haemorrhage? j laryngol otol. 2016; 130(8): 763-7. 3. lechner m, chandrasekharan d, vithlani r, sutton l, grandidge c, elmiyeh b. evaluation 68 secondary heamorrhage after cold steel tonsillectomy: an audit performed in shalamar hospital of a newly introduced tonsillectomy operation record for the analysis of regional posttonsillectomy bleed data: a quality improvement project at the london north west healthcare nhs trust. bmj open qual.2017;6(2):e000055 4. brkic f, mujic m, umihanic s, hrncic n, goga a, goretic e. haemorrhage rates after two commonly used tonsillectomy methods: a multicenter study. med arch (sarajevo, bosnia herzegovina). 2017; 71(2):119-21. 5. walker ra, syed za. harmonic scalpel tonsillectomy versus electrocautery tonsillectomy: a comparative pilot study. otolaryngol head neck surg. 2001; 125(5): 449-55. 6. verma r, verma rr, verma rr. tonsillectomy-comparative study of various techniques and changing trend. indian j otolaryngol head neck surg.2017;69(4):549-58. 7. moroco ae, saadi ra, wilson mn. posttonsillectomy respiratory complications in children with sleep disordered breathing. int j pediatr otorhinolaryngol. 2020; 131(october 2019):109852. 8. bao-hns, rcs. national prospective tonsillectomy audit. royal college of surgeons of england. 2005. 1-50 p. 9. blanchford h, lowe d. cold versus hot tonsillectomy: state of the art and recommendations. orl. 2013; 75(3):136-41. 10. söderman ach, odhagen e, ericsson e, hemlin c, hultcrantz e, sunnergren o, et al. post-tonsillectomy haemorrhage rates are related to technique for dissection and for haemostasis. an analysis of 15734 patients in the national tonsil surgery register in sweden. clin otolaryngol. 2015; 40(3):248-54. 11. o’leary s, vorrath j. postoperative bleeding after diathermy and dissection tonsillectomy. laryngoscope. 2005; 115(4):591-4. 12. harju t, numminen j. risk factors for secondary post-tonsillectomy haemorrhage following tonsillectomy with bipolar scissors: four-year retrospective cohort study. j laryngol otol. 2017; 131(2):155-61. 13. walker p, gillies d. post-tonsillectomy hemorrhage rates: are they techniquedependent? otolaryngol head neck surg. 2007; 136(4 suppl.). 14. ali rb, smyth d, kane r, donnelly m. posttonsillectomy bleeding: a regional hospital experience. ir j med sci. 2008; 177(4):297-301. 15. chughtai a, haq au, ullah s. is the cold steel dissection method still the most effective method of tonsillectomy? 2016; 66(5). 16. leinbach rf, markwell sj, colliver ja, lin sy. hot versus cold tonsillectomy: a systematic review of the literature. otolaryngol head neck surg. 2003; 129(4):360-4. 17. aydin s, taskin u, altas b, erdil m, senturk t, celebi s, et al. post-tonsillectomy morbidities: randomised, prospective controlled clinical trial of cold dissection versus thermal welding tonsillectomy. j laryngol otol.2014;128(2):163-5 18. motta s, testa d, ferrillo b, massimilla ea, varriale r, motta g, et al. surgical techniques and post-tonsillectomy haemorrhage. 2017; 21(4):559-66. 19. heidemann ch, wallén m, aakesson m, skov p, kjeldsen ad, godballe c. post-tonsillectomy hemorrhage: assessment of risk factors with special attention to introduction of coblation technique. european archives of oto-rhinolaryngology. 2009 jul 1; 266(7):1011. 20. hinton-bayre ad, noonan k, ling s, vijayasekaran s. experience is more important than technology in paediatric post-tonsillectomy bleeding. the journal of laryngology & otology. 2017 jul; 131(s2):s35-40. 21. ordemann ag, hartzog aj, seals sr, spankovich c, stringer sp. is weight a predictive risk factor of postoperative tonsillectomy bleed? laryngoscope investigative otolaryngology.2018jun;3(3):238-43 22. hinton-bayre ad, noonan k, ling s, vijayasekaran s. experience is more important than technology in paediatric post-tonsillectomy bleeding. the journal of laryngology & otology. cambridge university press; 2017; 131(s2):s35-s40. 23. abdel rahman aa, al-morsy ma, hussein ms. incidence, prevention and management of post tonsillectomy hemorrhage. the egyptian journal of hospital medicine. 2019 jan 1; 74(5):1123-7. 24. odhagen e, sunnergren o, söderman ac, thor j, stalfors j. reducing post-tonsillectomy haemorrhage rates through a quality improvement project using a swedish national quality register: a case study. european archives of oto-rhino-laryngology. 2018 jun 1; 275(6):1631-9. 25. kaplan hc, provost lp, froehle cm, margolis pa. the model for understanding success in quality (musiq): building a theory of context in healthcare quality improvement. bmj qual saf. 2012; 21(1):13-20. 69 secondary heamorrhage after cold steel tonsillectomy: an audit performed in shalamar hospital the authors: dr. arshadullah afridi assistant professor, department of ent, shalamar hospital, lahore. dr. sana tariq research associate, department of bioethics, shalamar medical & dental college, lahore. dr. muhammad dawood demonstrator, department of forensic medicine, shalamar hospital, lahore. dr. sarwat iqbal assistant professor, department of medicine, shalamar hospital, lahore. dr. usna athar demonstrator, department of community medicine, shalamar hospital, lahore. dr. aisha hanif medical officer, department of ent, shalamar hospital, lahore. corresponding author: dr. arshad ullah afridi assistant professor, department of ent, shalamar hospital, lahore. e-mail: arshadullah@hotmail.com 12 proceedings s.z.m.c. vol: 35(3): pp. 12-16, 2021. pszmc-802-35-3-2021 community acquired methicillin resistant staphylococcus aureus (ca-mrsa) infection in paediatric subcutaneous abscesses in pakistan 1muhammad ali sheikh, 1amna idrees, 1jamil akhter munir ahmad, 1kaneez fatima, 2lubna riaz, 1amna ikram 1department of pediatric surgery, shaikh zayed medical complex, lahore 2department of pediatric medicine, shaikh zayed medical complex, lahore abstract introduction: staphylococcus aureus is one of the common pathogens of the human body. previously a change in its sensitivity pattern was observed in which it became methicillin resistant but this strain was usually seen in hospital settings or in immunocompromised patients. recently a changing trend has been observed in which a new variant of methicillin resistant staphylococcus aureus (mrsa) is being isolated specially in pediatric patients, who do not have any risk factors. this strain is labeled as community acquired mrsa. aims & objectives: to find the frequency of community acquired methicillin-resistant staphylococcus aureus (camrsa) in paediatric sub-cutaneous abscesses. place and duration of study: a cross-sectional prospective study was conducted in department of paediatric surgery shaikh zayed medical complex, lahore from january 2017 till december 2020. material & methods: all the patients from 7 days to 14 years of age from both genders having subcutaneous abscesses were included. neonates having hospital stay of more than 24 hours at birth were excluded. patients having abscess secondary to any surgical intervention, recent hospitalization or trauma were also excluded. patients were divided into four groups based on age. pus samples were taken for culture & sensitivity. data analysis was performed using spss version 20. results: there were 143 patients and 75 (52.4%) of them were males. patient were divided in groups on the basis of age. the most common location of abscess was head & neck (43.4%) followed by lower limbs and buttocks (30.1%). the most common organism to grow was staph. aureus (75%) and out of these 107 cases 70 (65.4%) had mrsa. no growth was obtained in 30 (21%) patients. other less common organisms were seen in 6 (4%) cases. in less than 1 month of age of patients 76% had mrsa which was significantly higher than in other age groups (pvalue 0.047). conclusion: the frequency of ca-mrsa in paediatric subcutaneous abscesses is on the rise across the globe and more cases of invasive infections are being reported. so, population-based studies are required to assess the prevalence of this bacteria amongst paediatric population of pakistan. key words: community acquired methicillin resistant staphylococcus aureus, pediatric, subcutaneous abscess introduction staphylococcus aureus (s. aureus) is one of the common pathogen of human body and the frequent cause of skin and soft tissue infections. some of the strains of s. aureus are resistant to all beta lactam antibiotics and are labelled as methicillin resistant s. aureus.1 methicillin-resistant staphylococcus aureus (mrsa) was initially isolated in 1960s and since that time it has been spreading increasingly in the recent years 2. simultaneously the increase in community-acquired soft tissue abscesses in the pediatric population has been noted across the globe in last few years. previously, mrsa infections were primarily hospital or institution acquired infections in patients with underlying risk factors. several studies have shown that mrsa infections of otherwise healthy patients in the community are on the increase, with a reported incidence ranging from 33% to 69%.3 soft tissue infections due to community-acquired mrsa (ca-mrsa) are becoming increasingly prevalent in the pediatric population even in western population.4 data from 43 children's hospitals represented in the pediatric health information system was analyzed by, gerber et al. and they reported an increase in mrsa soft tissue infections from 0.2% of admissions in 2002 to 1.1% of admissions in 2007. likewise, an analysis of discharge records in the united states national center for health statistics demonstrated an increasing rate of community 13 community acquired methicillin resistant staphylococcus aureus (ca-mrsa) infection in paediatric acquired mrsa from 1996 to 2006, reaching an incidence of 25.5 cases per 100,000 children in 2006.1,5 doctors around the globe have been dealing with the complexity of management of mrsa for quite some time now. a few years back, the term mrsa was almost always associated with an infection that was hospital acquired. but since a decade or so the term community acquired mrsa (ca-mrsa) has emerged, which has brought along a whole new chapter of treatment and etiologies.6 the centers for disease control and prevention (cdc) active bacterial core surveillance program defined a ca-mrsa case as a patient with an mrsa infection with no history of surgery, hospitalization, or residence in a medical facility within the year before infection, presence of a percutaneous device or indwelling catheter, dialysis within the previous year, hospitalization more than 48 h before mrsa culture, or previous mrsa infection or colonization.7 although there have been research publications internationally on ca-mrsa, still there is insufficient data and information available on role of ca-mrsa in paediatric population of pakistan, which led us to undertake the current research. especially, there is lack of studies on ca-mrsa on local population except five cases reported by agha khan university hospital karachi.8 the aim of the study was to find the frequency of community acquired methicillin-resistant staphylococcus aureus (ca-mrsa) in paediatric sub-cutaneous abscesses. material and methods a cross-sectional prospective study was conducted in department of paediatric surgery shaikh zayed medical complex, lahore from january 2017 till december 2020. all the patients from 7 days to 14 years of age from both genders having subcutaneous abscesses were included. neonates having hospital stay of more than 24 hours at birth were excluded. patients having abscess secondary to any surgical intervention, recent hospitalization for any other disease in last 2 years or trauma were also excluded. patients were divided into four groups based on age. pus samples of the all the patients were obtained for culture and sensitivity at the time of surgery. most of the cases were managed on outpatient basis. patients were initially started on empirical antibiotics and later changed according to sensitivity report. age of the patient, gender, area involved, temperature, hemoglobin levels, tlc count, weight of the patient and type of organism were recorded. statistical analysis: data was analyzed with spss version 20. qualitative variables were analyzed with chi square and p values were calculated. results during the study period 143 patients met the inclusion criteria. there were 75 (52.4%) male patients and 57 (40%) were less than 1 year of age. the age and gender distribution are shown in table1. the most common location of abscess was head & neck (43%), followed by lower limb & buttocks (30%). the details of location of abscess are shown in table-2. the most common bacteria to grow in pus was mrsa in 70 (49%) patients, followed by methicillin sensitive staphylococcus aureus (mssa) in 37 (26%) patients and 30 (21%) patients did not grow any bacteria. other less common organisms were seen in 6 (4%) cases. in less than 1 month of age patients 76% had mrsa which was significantly (p=0.047) higher than in other age groups. the details are shown in table-3. as the age of the patients increased the number of cases infected with mssa increased. the most common location of abscesses was head & neck and in these cases 50% had mrsa but the difference was not statistically significant. comparison was also made between type of growth and tlc but no significant difference was observed, except that 50 % of cases of mrsa had less than 11,000 tlc. growth of mrsa was common in female patients i.e., 41(58.6 %) out of total 70 cases of mrsa and it was statistically significant with p value of = 0.033. mssa was slightly more common in male patients 29 out of 37 (56.75%). fever was present in 71 patients, 33 (46.5%) with mrsa and 23 (32.4%) with mssa had fever. remaining 72 patients remained a febrile. (p value = 0.140). only 5 (3.5%) of the patients had 2 or more abscesses, the remaining patients had a single abscess. there was no significance of hemoglobin levels as compared to the type of growth. hemoglobin levels of 30 (50%) patients with mrsa were between 10.4-12.5 (p value =0.942). no significance of weight of the patient was found in relation to either location, age or type of growth. 14 community acquired methicillin resistant staphylococcus aureus (ca-mrsa) infection in paediatric age group male female total percentage <1 month 12 17 29 20.3 2 month12 months 14 14 28 19.6 1-5 years 24 22 46 32.2 >5 years 25 15 40 28.0 total 75 68 143 table-1: age & gender distribution of patients (n=143) area involved no of cases percentage head & neck 62 43.4% upper limb 18 12.6% lower limb & buttocks 43 30.1% chest wall 9 6.3% abdominal wall 5 3.5% back 6 4.2% table-2: distribution according to location of abscess mssa mrsa no growth others p value n % n % n % n % age groups <1 month (n=29) 3 10.3 22 75.9 4 13.8 0 0.0 0.047 <1 year (n=28) 9 32.1 15 53.6 3 10.7 1 3.6 1-5 years (n=46) 13 28.3 17 37.0 13 28.3 3 6.5 >5 years (n=40) 12 30.0 16 40.0 10 25.0 2 5.0 gender male (n=75) 21 28.0 29 38.7 20 26.7 5 6.7 0.033 female (n=68) 16 23.5 41 60.3 10 14.7 1 1.5 location of abscess head& neck (n=62) 14 22.6 31 50.0 15 24.2 2 3.2 0.350 upper limb (n=18) 2 11.1 10 55.6 6 33.3 0 0.0 lower limb & buttocks(n=43) 15 34.9 18 41.9 6 14.0 4 9.3 chest wall (n=9) 3 33.3 5 55.6 1 11.1 0 0.0 abdominal wall (n=5) 1 20.0 2 40.0 2 40.0 0 0.0 back (n=6) 2 33.3 4 66.7 0 0.0 0 0.0 total leukocyte count × 1000 <= 11.0 (n=44) 10 22.7 22 50.0 11 25.0 1 2.3 0.571 11.1 15.0 (n=45) 16 35.6 19 42.2 8 17.8 2 4.4 >15.1 (n=53) 10 18.9 29 54.7 11 20.8 3 5.7 table-3: comparison of patient parameters with type of growth discussion methicillin resistant staph aureus (mrsa) was initially identified among hospitalized patients and remained predominantly a nosocomial infection till late 1980s.1 a new strain of mrsa started to appear in 1990s, which started to cause infections in healthy people and was labeled as community acquired mrsa (ca-mrsa).1,2 ca-mrsa is becoming a world-wide threat with studies reporting the increasing prevalence of nasal carriers of ca-mrsa. the proportion of ca mrsa isolates among all mrsa clinical isolates increased from 19.7% in 2007 to 36.4% in 2011 in canada.4 in spain, ca-mrsa rates increased over time while comparing 2004-2007 period (0.43%) with 2008-2012 period (5.44%).4 in brazil, a study has reported ca-mrsa nasal colonization of 0.9% among healthy people living in the community, whereas 8.7% was found among adults with chronic wounds.4 camrsa does not follow the usual risk factors that were previously associated with mrsa infection, like repeated antibiotic therapy, prolonged hospitalization, exposure to people already infected with mrsa.9 risk factors for ca-mrsa infections have been identified in some studies, such as age, family history of staphylococcal infections, high body weight, previous use of antibiotics, aboriginal ethnicity, stay in day care settings and presentation during the spring.4 but we did not encounter similar risks in our study. the most common organism cultured from subcutaneous abscesses in our study was staphylococcus aureus which was noted in 107 cases (75%). out of these 107 cases 70 (65.4%) had mrsa. this was comparable with other international studies; olesevich et al. reported 66.3% and rossini et al reported 74.75% infection with staphylococcus aureus in their patients. in these patients ca-mrsa was detected in majority of cases 49.2% by olesevich and 66 % by rossini et al.2,7 this shows the trend that ca-mrsa is increasing in the general population. in current study ca-mrsa was significantly (p=0.033) more common in females (58.6%), similar finding was noted by olesevich et al7 but the difference was not statistically significant in their study. a recent study from china observed similar results as our study with ca-mrsa being slightly more common (52%) in females.10 no clear reason for this female preponderance in our patients could be found. the patients were divided into groups based on their age and ca-mrsa was significantly (p =0.047) more common in less than 1 month of age patients. 15 community acquired methicillin resistant staphylococcus aureus (ca-mrsa) infection in paediatric similar findings were reported by salazar et al.3 this could be due to the immature immune system in this age group which allows easy colonization by the organism and rapid spread of infection. in our study 40% of patients were less than 1 year of age which was comparable with the study by salazar (41.2%)3 and wang et al (42%).10 this finding shows that ca-mrsa causes more invasive infections in younger age groups and is amongst the leading cause of skin and subcutaneous tissue infections in this population. the most common location of abscess in our study was head & neck which was seen in 43.4% of patients. the large number of cases in this area were on the occipital region. as per practice of our population the baby’s head is rested against a firm surface to shape it. it results in pressure sore and infection on dependent occipital area. rossini et al. reported buttocks to be the most common location (41.5%) in their study. the second most common location in our study was buttocks and lower limbs (30.1%). buttocks are more vulnerable because of commonly occurring diaper rash resulting in breach of skin barrier and bacteria could easily enter in the subcutaneous tissue and cause infection. we correlated presence of fever, hemoglobin levels, and weight of the children with type of growth and did not find any significant relation. in patients with ca-mrsa total leukocyte count was less than 11,000 in 50% of cases which show poor response from immune system against the bacteria, specially in less than 1 year age group. there are some studies from asian countries which have highlighted the menace of rising trend of camrsa in this part of the world as well. immergluck et al. conducted an epidemiological study and found that ca-mrsa rates increased at higher rates compared to non-resistant forms (mssa), p = 0.01. children with no public health insurance had higher odds of ca-mrsa infection. black children were almost 1.5 times as likely as white children to have ca-mrsa infections (or 95% ci 1.44,1.75, p < 0.0001).11 alaklobi et al. studied children between 1 month to 13 years for nasal carriage of ca-mrsa and found that 20% of their patients were carrying staph aureus and 23% among them were having mrsa.12 a study by rodriguez et al. reported that the prevalence of ca-mrsa increased from 21% between 2009-2010 to 54% between 2012-2013 and most of them (74%) caused skin & soft tissue infections in children.13 alzomar et al. reported bones and joint infections, deep seated abscesses, necrotizing pneumonia and infective endocarditis that were caused by camrsa in children.14 they concluded that camrsa infections could become invasive and infect previously healthy children with no known risk factors making it one of the virulent organisms. invasive diseases may comprise of bacteremia related with metastatic areas of infection like endocarditis, bone & joint infections and necrotizing pneumonia. incidence of ca-mrsa seems to be variable, but with these types of invasive infections, the knowledge of the wide range of ca-mrsa disease among the treating physicians is important because of early identification and start of effective empirical antibiotics is important. the current study confirms that ca-mrsa is an emerging pathogen not only in paediatric population of pakistan but also in other parts of the world, accounting for approximately one third of all s.aureus strains isolated from children with severe community-acquired infections. moreover, we noticed a trend of increasing incidence density over the four-year period of the study. a large population-based study should be conducted to know the carrier state of mrsa. conclusion the frequency of ca-mrsa in paediatric subcutaneous abscesses is on the rise across the globe and more cases of invasive infections are being reported. the choice of empirical antibiotics should be based on common organisms found in a particular population. so, population-based studies are required to assess the prevalence of this bacteria amongst paediatric population of pakistan. references 1. dukic v, lauderdale d, wilder j, daum r, david m. epidemics of community-associated methicillinresistant staphylococcus aureusin the united states: a meta-analysis. plos one. 2013; 8 (1) e52722. 2. rossini c, moriarty k, tashjian d, garb j, wait r. geographic distribution of community–acquired methicillin–resistant staphylococcus aureus soft tissue infections. j pediatr surg 2011; 46: 1089-92. 3. salazar-ospina l, jiménez jn. high frequency of methicillin-susceptible and methicillin-resistant staphylococcus aureus in children under 1 year old with skin and soft tissue infections. j pediatr (rio j). 2018; 94(4):380-389. 4. paternina-dela-ossa r, prado si, cervi mc, lima dafs, martinez r, bellissimo-rodrigues f. is community associated methicillin resistant staphylococcus aureus an emerging pathogen among children in brazil? braz jour infec dis. 2018; 22(5):371–376. 5. gerber js, coffin se, smathers sa, et al. trends in the incidence of methicillin-resistant staphylococcus 16 community acquired methicillin resistant staphylococcus aureus (ca-mrsa) infection in paediatric aureus infection in children's hospitals in the united states. clin infect dis 2009; 49:65-7. 6. frei cr, miller ml, lewis js, lawson ka, hunter jm, oramsionwu cu et al. trimethoprim sulfamethoxazole or clindamycin for communityassociated mrsa (ca-mrsa) skin infections. j am board fam med. 2010; 23:714-19. 7. olesevich m, kennedy a. emergence of community acquired methicillin resistant staphylococcus aureus soft tissue infections. j pediatr surg 2007; 42:765768. 8. khalid m, junejo s, mir f. invasive community acquired methicillin resistant staphylococcal aureus infections in children. j coll physicians surg pak 2018; 28 (special supplement):174-77. 9. frei cr, makos br, daniels kr, oramsionwu cu. emergence of community-acquired methicillinresistant staphylococcus aureus skin and soft tissue infections as a common cause of hospitalization in united states children. j pediatr surg 2010; 45: 1967-1974. 10. wang x, shen y, huang w, zhou y. characterization of community acquired staphylococcus aureus causing skin and soft tissue infections in a children’s hospital in shanghai, china. epidemiology and infection 147, 2019, e323, 1–6. 11. immergluck lc, leong t, malhotra k, parker tc, ali f, jerris rc et al. geographic surveillance of community associated mrsa infections in children using electronic health record data. bmc infect dis 2019; 19:170 12. alaklobi f, aljobair f, alrashod a, alhabibi r, alshamrani m, alamin w et al. the prevalence of community associated methicillin resistant staphylococcus aureus among outpatient children in a tertiary hospital: a prospective observational study in riyadh, saudi arabia. int j pediatr adolesc med 2015; 2:136-140. 13. rodríguez f, salinas c, fernandez s, haim s, mollerach m, basualdo w et al. communityassociated methicillin-resistant staphylococcus aureus (ca-mrsa) clones from paraguayan children. j infect dev ctries 2020; 14(3):290-297. 14. alzomar o, alfawaz t, alshahrani d. invasive community acquired methicillin resistant staphylococcus aureus (ca-mrsa) infection in children: case series and literature review. int j pediatr adolesc med. 2017; 4:119-123. the authors: prof. muhammad ali sheikh head, department of pediatric surgery, shaikh zayed medical complex, lahore. dr. amna idrees p.g. trainee, department of pediatric surgery, shaikh zayed medical complex, lahore. dr. jamil akhter munir ahmad senior registrar, department of pediatric surgery, shaikh zayed medical complex, lahore. dr. kaneez fatima medical officer, department of pediatric surgery, shaikh zayed medical complex, lahore. dr. lubna riaz assistant professor, department of pediatric medicine, shaikh zayed medical complex, lahore. dr. amna ikram p.g. trainee, department of pediatric surgery, shaikh zayed medical complex, lahore. corresponding author: prof. muhammad ali sheikh head, department of pediatric surgery, shaikh zayed medical complex, lahore. e-mail: drali444@yahoo.com for web full book 40 proceedings s.z.m.c. vol: 36(2): pp. 40-45, 2022. pszmc-843-36-2-2022 1department of pathology, university college of medicine and dentistry, lahore 2department of pathology, jinnah hospital, lahore 3department of pathology, fmh college of medicine & dentistry, lahore 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 41 adverse blood transfusion reaction in tertiary care hospitals an initiative towards improvement: a… 42 adverse blood transfusion reaction in tertiary care hospitals an initiative towards improvement: a… 43 adverse blood transfusion reaction in tertiary care hospitals an initiative towards improvement: a… 44 adverse blood transfusion reaction in tertiary care hospitals an initiative towards improvement: a… 45 adverse blood transfusion reaction in tertiary care hospitals an initiative towards improvement: a… 1 proceedings s.z.m.c. vol: 35(3): pp. 1-5, 2021. pszmc-800-35-3-2021 histological subtypes & staging of post-chemotherapy wilms tumor according to siop 2001 protocol: study at the children’s hospital, lahore 1fariha sahrish, 2abeera assad rasool, 2zonaira rathore, 2mahvish hussain, 2fizza waqar, 2samina zaman 1department of histopathology, azra naheed medical college, lahore 2department of histopathology, children’s hospital, lahore abstract introduction: pediatric renal tumors constitute 7 to 8% of pediatric solid malignancies and most common is wilms tumor. it usually presents as unilateral mass with sporadic and familial associations. it is currently treated by nwts and siop protocols worldwide. in our hospital setup we follow siop 2001 protocol to subcategorize different histological subtypes and staging of wilms tumor after completing four cycles of chemotherapy. aims & objectives: to determine the frequency of histological subtypes of wilms tumor (wt) in post chemotherapy nephrectomy specimens place and duration of study: histopathology section of pathology department, children’s hospital and institute of child health lahore from january 2015 to june 2018. material & methods: ninety-three radical nephrectomy specimens of different histological subtypes i.e. wilms tumor (wt) consisting of blastemal predominant (bp), epithelial predominant (ep), stromal (st), mixed (mt), regressive (rp), completely necrotic (cn) and diffuse anaplastic (da) were analyzed. risk categorization and staging proposed by siop 2001 was appraised. data analysis was done using spss version 23. results: a total of 93 cases were included in the present study. mean age of children recorded was 42 months with male predominance. mean tumor size after chemotherapy was 7.25 cm. majority cases were observed on the right side. regarding the histological subtype there was a predominant group of mt while least observed was da. based on subtypes and necrosis, majority of wt were of intermediate risk (ir) and stage i tumors. conclusion: majority of pretreated cases (n=80, 85.1%) were stage 1 tumors. commonest histological subtype is mt followed by rp, cn, ep, bp, st and da. key words: wilms tumor (wt), (siop staging) internal society of pediatric oncology) national wilms tumor study group (nwts), post chemotherapy wt. introduction pediatric renal tumors comprise 7 to 8 % pediatric solid malignancies in first 15 years of life. most common renal tumor is wilms tumor (wt), also known as nephroblastoma (n) which accounts for 85% of cases. it usually presents as palpable abdominal mass noticed by mother.1,2 median age of children is 3-4 years.3 wt mostly presents as unilateral renal mass but 5-10 % cases are bilateral.4 wt1 gene located on chromosome 11p13 is associated with both familial and sporadic cases. vast majority of wt are sporadic and only 10% are familial. familial wt are associated with beckwith wiedemann syndrome, denys drash syndrome and wagr syndrome. no specific risk factor contributing towards wt has been identified uptil now.1,3,4 grossly most of the cases of wt are unicentric, whereas 5% wt are multicentric.5 undifferentiated blastemal element, epithelial component and stromal component constitute classic wt. the potential precursor of wt are nephrogenic rests which are embryonic stem cells and are of perilobar and intralobar types.3,5 wt is managed according to two protocols, national wilms tumor study group (nwts) and international society of pediatric oncology (siop) which are established over years of multicenter trials. these groups have enormous contributions towards the management and excellent prognosis of patients with wt.3,4,6,7,8 the nwts, established in 1969, completed 5 sequential trials from nwts-1 to nwts-5. it endorses the diagnosis of wt on radiological examination followed by core biopsy and prompt radical nephrectomy.2 currently, nwts-5 is 2 histological subtype & staging of post-chemotherapy wilms tumour according to siop 2001 protocol analyzing genetic prognostic factors such as loss of heterozygosity on chromosome 1p and 16q.9 shortly after nwts, siop trials were started in 1971 and evolved from the siop 1 to siop 9 (19712001). it advocates prophylactic chemotherapy after initial diagnosis made on core biopsy. after cessation of chemotherapy course, it endorses radical nephrectomy in unilateral cases and partial nephrectomy in bilateral cases.10,11 the histological classification of wt differs in nwts and siop 2001 due to preoperative chemotherapy. accurate histological subtyping and staging of wt is very important in further management and prognosis of patient.1,2 according to nwts, histology groups are divided on the basis of presence and absence of anaplasia in three groups. the favorable histology wt is cystic partially differentiated nephroblastoma (cpdn), standard risk (sr) histology group includes nonanaplastic wt and its variants and high risk (hr) diffuse anaplasia (da). according to the siop wt 2001 classification the subtypes are classified on the basis of percentage of necrosis, presence of viable tumor, presence and absence of regressive changes and presence of focal and diffuse anaplasia. low risk (lr) include completely necrotic (cn), intermediate risk (ir) include epithelial predominant (ep), stromal type (st), mixed type (mt), regressive predominant (rp), focal anaplasia (fa) and high risk (hr) tumors include blastemal predominant (bp) and diffuse anaplasia (da).3,10,11,12 in our centre we are following siop 2001 protocol. the present study is carried out to determine the frequency and morphological features of wt according to siop 2001 protocol as our centre is following their protocol. material and methods the present study is a cross-sectional study approved by ethical review board vide its letter no. 02/173/17 dated 01/02/2017 and carried out at the histopathology department of children’s hospital and institute of child health lahore from january 2015 to june 2018. ninety-three radical nephrectomy specimens of children between 0-15 years of age in both genders of biopsy proven cases of wt tumor were included. partial nephrectomy specimens and blocks received for review were excluded. different histological subtypes i.e. wt consisting of blastemal, epithelial, stromal, mixed, regressive, completely necrotic and anaplastic were analyzed. risk categorization and staging proposed by siop 2001 was appraised. radical nephrectomy specimens were meticulously sectioned as per protocol of american college of pathologists. specimen size, areas of suspected tumor rupture, capsular and perinephric invasion, sectioning of distal end of ureter, renal vein and artery for tumor metastasis and tumor thrombus was noted prior to opening of specimen at the department. sectioning of tumor at one centimeter was done and assessment of necrotic areas, viable tumor areas and its relation to the normal kidney, capsule, renal hilum and renal sinus was done grossly. additional sections from the tumor and normal kidney were taken to assess nephrogenic rests. tissue slices were processed in the automatic tissue processor for 16 hours overnight dehydrated with ethyl alcohol, cleared by xylene, impregnated with paraffin wax and later on tissue blocks were made. hematoxylin and eosin stains were done. statistical analysis: data analysis was done on spss (statistical packages of social sciences) version 23. qualitative variables like age, gender, laterality, histological subtypes and staging were summarized as frequencies and percentages. no statistical association is determined between any variables. results a total of 93 cases were included in this study. the main clinical features of the study are presented in table-1. age at diagnosis mean 3.6 years minimum age 6 months maximum age 15 years number (n) percentage (%) 0 to 3 years 52 55.9 4 to 6 years 30 32.3 7 to 9 years 8 8.6 10 to 12 years 2 2.2 more than 12 years 1 1.1 gender male 52 55.9 female 41 44.1 laterality right 52 55.9 left 37 39.8 tumor size in cm mean tumor size 7.25 minimum tumor size 1.5 maximum tumor size 16 table-1: principal clinicopathological features of wilms tumor 3 histological subtype & staging of post-chemotherapy wilms tumour according to siop 2001 protocol majority of the patients were seen in first 3 years of age with a male predominance n=52(55.9%). most cases were in right kidney (n=52, 55.9%) with mean tumor size 7.25cm. however, largest tumor size observed was 16cm after chemotherapy (fig-1). regarding the histological subtype the commonest was mt (n=41, 44.1%), followed by rp (n=16, 17.2%). there were equal number of ep and cn (n=12, 12.9%). in addition, 9 cases (9.7%) were of bp, 2 cases (2.2%) of st whereas only one case (1.1%) was of da. considering necrosis, majority of the tumors (n=65, 69.9%) showed less than 66% necrosis and 28 cases (30.1%) showed more than 66% necrosis. on the basis of histological subtypes and necrosis, (76.3%) cases belonged to ir, whereas 12.9% cases were of lr and 10.8% cases were of hr according to siop staging protocol. in the present study majority of the cases were limited to kidney, only in 7 cases hilum was involved. capsule was involved in 2 cases and similar number of cases showed tumor thrombi in renal vein whereas in a single case inferior vena cava was involved by tumor thrombus clinically. distal end of ureter, perinephric fat infiltration and nephrogenic rests were seen in one case each. we received lymph nodes in 28 cases, which were all negative for metastatic disease. 80 cases (86%) had stage i, 11 cases (11.8%) had stage ii and 2 cases (2.2%) had stage iii. the pathological features concerning histological subtypes, staging and largest tumor size is presented in table-2. histological subtypes stage i n (%) stage ii n (%) stage iii n (%) largest tumor size(cm) mixed type 34 (36.5) 6 (6.4) 1 (1.07) 16 regressive predominant 14 (15) 2 (2.1) 0 (0) 10 completely necrotic 11 (11.8) 1 (1.07) 0 (0) 14 epithelial predominant 10 (10.7) 2 (2.1) 0 (0) 11 blastemal predominant 8 (8.6) 0 (0) 1 (1.07) 14 stromal predominant 2 (2.1) 0 (0) 0 (0) 6 anaplasia – diffuse 1 (1.07) 0 (0) 0 (0) 10 total no. 80 (79) 11(11.8) 2 (2.1) 81 table-2: histological subtypes, staging and largest tumor size according to siop 2001. concerning the stage distribution as shown in table-2 the greater proportion of cases were of stage i with predominant histological type mt. fig-1:a post chemotherapy bivalved kidney shows a well circumscribed tumor with mahogany cut surface (down & right arrow), and periphery shows thin rim of normal renal parenchyma (up arrow) fig-2: photomicrograph of h&e staining showing neoplastic tubules (arrows) in residual wilms tumor x 20 fig-3: photomicrograph of h&e staining showing a renal hilum (right arrow), involved by wilms tumor (arrow) adjacent normal kidney parenchyma (elbow arrow) x 20 4 histological subtype & staging of post-chemotherapy wilms tumour according to siop 2001 protocol discussion the prevalence of malignant pediatric tumors is remarkably higher in the developing countries as compared to the developed nations. in developing countries, pediatric malignancies prevalence is reported as 4.38% to 12.6% as compared to 2% in developed countries.13 it affects approximately one child in every 10,000 children, worldwide before the age of 15 years.4 according to world health organization (who), international incidence of childhood cancer, the incidence-age specific rate per million in pakistan and india is 3.6 and 4.4 respectively.14 according to annual cancer registry report by shaukat khanum cancer hospital there were 27 cases of wt, nos in 2018 in pakistani population.15 we received 93 cases of post chemotherapy wt in approximately 3.5 years with the mean age of 3.5 years (table-1). similarly, guruprasad et al. observed the mean age of 3.3 years in the duration of 6 years in indian population.16 mazumder et al. a peak age of approximately 4 years in a 20 patients cross sectional study,17 anwar et al. had the observation of mean age of wt at the age of 3 years18 whereas onuigbo noticed the peak age of wt lies between 2 years to approximately 5 years.19 regarding sex, we had majority of the male patients (table-1). this observation was similar to gruparasad et al,16 mazumder et al,17 anwar et al,18 onuigbo et al,19 weirich et al11 and sayed et al21 whereas shende sa showed female preponderance.22 there was a predominately right sided renal involvement in our index study (table-1). similar to mazumder et al,17 anwar et al,18 onuigbo et al19 and basu et al.20 however, weirich et al11 and sayed et al21 observed predominantly left renal involvement. bilateral wt was reported by pianezza et al,23 gruparasad et al,16 anwar et al18 and sayed et al21 whereas bilateral wt was excluded in our study. regarding the histological subtype, there was a marked predominance of mt in our study and only 12% of the total cases were cn (table-2). similar to mazumder et al who also observed predominantly tumors of mt in their study.17 sayed et al. had the observation that most of the cases were of nonanaplastic type and had the mt histological picture followed by ep type.21 however, vujanic et al. showed that in 258 of the total pretreated nephrectomy specimens, majority of the cases were of rp followed by mt.24 bocon-gibond et al. showed majority as cn in their study.12 in contrast, vujanic et al studied 195 pre treated nephrectomy specimens and he noticed a major chunk of mt and cn which was seen in 17% of the cases.25 in our study majority were of stage i tumors (table2) similar to sayed et al,21 vujanic et al24 and hung ij et al.27 in contrast, faranoush et al. observed slight predominance of stage ii tumors26 where as, guruprasad et al16 and anwar et al18 observed majority of the patients were in stage iii. verschuur ac et al showed ep and st showed excellent clinical outcomes which was beyond the scope of our case study.28 conclusion following siop 2001 protocol the commonest histological type is mixed subtype which has intermediate risk. majority of our tumors were of stage i tumors. references 1. vujanić gm and sandstedt b. the pathology of wilms’ tumour (wt): the international society of paediatric oncology approach. j clinpathol 2010; 63(2):102-9. 2. davidoff am. wilms tumor. advpediatr. 2012; 59(1):247-67. 3. varan a. wilms’ tumor in children: an overview. nephron clinpract 2008; 108:83-90. 4. charlton j, irtan s, bergeron c, pritchard-jones k. bilateral wilms tumour: a review of clinical and molecular features. expert rev mol med.2017;19:e8 5. friedman ad. wilms tumorpediatrics in review 2013; 34(7):328-30. 6. bhatnagar s. management of wilms tumor: nwts vs siop. journal of indian association of pediatric surgeons. 2009; 14(1):6-14. 7. lopes ri, lorenzo a. recent advances in the management of wilms' tumor. f1000res.2017;6:670 8. brok j, lopez-yurda m, tinteren hv, treger td, furtwängler r, graf n, bergeron c, heuvel-eibrink mmvd, pritchard-jones k olsen oe, camargo b, verschuur a, spreafico f. relapse of wilms' tumour and detection methods: a retrospective analysis of the 2001 renal tumour study group-international society of paediatric oncology wilms' tumour protocol database. lancet oncol.2018;19(8):1072-81 9. grundy pe, breslow ne, li s, perlman e, beckwith jb, ritchey ml, shamberger rc, haase gm, d'angio gj, donaldson m, coppes mj, malogolowkin m, shearer p, thomas pr, macklis r, tomlinson g, huff v, green dm; national wilms tumor study group. loss of heterozygosity for chromosomes 1p and 16q is an adverse prognostic factor in favorable-histology wilms tumor: a report from the national wilms tumor study group. j clinoncol. 2005; 10;23(29):7312-21. 10. metzger m and dome j. current therapy for wilms tumor. pediatr. oncol. 2005; 10:815-26. 5 histological subtype & staging of post-chemotherapy wilms tumour according to siop 2001 protocol 11. weirich a, leuschner i, harms d, vujanic gm, troger j, abel u, graf n, schmidt d, ludwig r and voute pa. clinical impact of histologic subtypes in localized non-anaplastic nephroblastoma treated according to the trial and study siop-9/gpoh. annals of oncology. 2001; 12:311-9. 12. boccon-gibod l, rey a, sandstedt b, delemarre j,harms d, vujanic g, de kraker j, weirich a, and tournade mf. complete necrosis induced by preoperative chemotherapy in wilms tumor as an indicator of low risk: report of the international society of paediatric oncology (siop) nephroblastoma. medical and pediatric oncology 2000; 34:183-190. 13. shah sh, soomro in, siddiqui ms, pervez s, and hassan sh. immunohistochemical evaluation of small round cell tumors of childhood. j pak med assoc. 1999; 49(4). 87-9. 14. cunningham me, klug td, nuchtern jg, chintagumpala mm, venkatramani r, lubega j, mathuria bj. global disparities in wilms tumor. j surg res. 2019; 247(832):34-51. 15. yousaf a, mahmood s, faraz r, quader a, asif h, atif a, nadeem l, parveen n, tanveer r, hussain a, badar f.annual cancer registry report-2018, of the shaukatkhanum memorial cancer hospital & research center, pakistan. 2016, 1-21. 16. guruprasad b, rohan b, kavitha s, madhumathi ds, lokanath d and appaji l. wilms’tumor: single centre retrospective study from south india indian j surgoncol 2013; 4(3):301-304. 17. mazumder m, islam a, farooq n, zaman m. clinicopathological profile of wilms’ tumour in children. j banagladeshcollphyssurg 2014; 32:5-8 18. anwar s, faizan m, khan s, tallat n saleem m, khalid a, qaiser, qureshi a, ali as, rathore a. a five year experience of wilms tumor at a tertiary care centre, where we stand, a developing country perspective. p j m h s 2017; 11(4):1263-6. 19. onuigbo w. comparative approach to the epidemiology of wilm's tumor: archives in cancer research. 2016; 4.10.21767/2254-6081.100048. 20. basu k, chattopadhyay s, das s, basu n, chatterjee s, das s. role of fnac in wilms’ tumor aninterim report, j assocpediatr, 2001;6:130-3. 21. sayed har, ali am, hamza hm, and abdalla ma. long-term follow-up of infantile wilms tumor treated according to international society of pediatric oncology protocol: seven years’ followup. urology 2011; 77(2):446-5. 22. shende a, sathe pa. clinicopathologic analysis of wilms’ tumor – a retrospective study of 35 cases over 10 years. ann pathol lab med. 2018; 5(2):a151-157. 23. pianezza ml, rubin s, bass j, chou s, pike jg, leonard mp. wilms tumor at the children’s hospital of eastern ontario. can j urol. 2004, 11: 2151-6. 24. vujanić gm, d'hooghe e, popov sd, sebire nj, kelsey a. the effect of preoperative chemotherapy on histological subtyping and staging of wilms tumors: the united kingdom children's cancer study group (ukccsg) wilms tumor trial 3 (ukw3) experience. pediatr blood cancer. 2019; 66(3):27549. 25. vujanić gm, kelsey a, mitchell c, shannon rs, and gornall p, “the role of biopsy in the diagnosis of renal tumors of childhood: results of the ukccsg wilms tumor study 3. med pediatroncol 2003; 40(1):18-22. 26. faranoush m, bahoush gr, mehrvar a, hejazi s, vossough p, hedayatiasl aa, rahiminejad ms, seighali f, ghorbani r and ehsani ma. wilm’s tumor: epidemiology and survival. research journal of biological sciences 2009; 4 (1): 86-89. 27. hung ij, yang cp et al, epidemiology, clinical feature and treatment outcome of wilms’ tumor in taiwan: a report from taiwan pediatric oncology group. j formos med assoc, 2004; 103:104-11. 28. verschuur ac, vujanic gm, van tinteren h, jones kp, de kraker j, sandstedt b. stromal and epithelial predominant wilms tumours have an excellent outcome: the siop 93 01 experience. pediatr blood cancer. 2010 aug; 55(2):233-8. the authors: dr. fariha sahrish senior demonstrator, department of histopathology, azra naheed medical college lahore. dr. abeera assad rasool women medical officer, department of histopathology, children’s hospital, lahore. dr. zonaira rathore assistant professor, department of histopathology, children’s hospital, lahore. dr. mahvish hussain assistant professor, department of histopathology, children’s hospital, lahore. dr. fizza waqar women medical officer, department of histopathology, children’s hospital, lahore. prof. samina zaman department of histopathology, children’s hospital, lahore. corresponding author: dr. fariha sahrish senior demonstrator, azra naheed medical college, lahore. e-mail: awan.fariha44@gmail.com full book 11 proceedings s.z.m.c. vol: 36(4): pp. 11-25, 2022. pszmc-859-36-4-2022 1department school of health sciences, university of management and technology, lahore, pakistan 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 12 impact of food literacy on consumer's food purchasing habits and dietary intake a systematic review 13 impact of food literacy on consumer's food purchasing habits and dietary intake a systematic review 14 impact of food literacy on consumer's food purchasing habits and dietary intake a systematic review 15 impact of food literacy on consumer's food purchasing habits and dietary intake a systematic review 16 impact of food literacy on consumer's food purchasing habits and dietary intake a systematic review 17 impact of food literacy on consumer's food purchasing habits and dietary intake a systematic review 18 impact of food literacy on consumer's food purchasing habits and dietary intake a systematic review 19 impact of food literacy on consumer's food purchasing habits and dietary intake a systematic review 20 impact of food literacy on consumer's food purchasing habits and dietary intake a systematic review 21 impact of food literacy on consumer's food purchasing habits and dietary intake a systematic review 22 impact of food literacy on consumer's food purchasing habits and dietary intake a systematic review 23 impact of food literacy on consumer's food purchasing habits and dietary intake a systematic review 24 impact of food literacy on consumer's food purchasing habits and dietary intake a systematic review 25 impact of food literacy on consumer's food purchasing habits and dietary intake a systematic review numbering.indd 70 proceedings s.z.m.c. vol: 34(4): pp. 70-75, 2020. pszmc-774-34-4-2020 topical lignocaine with diltiazem or glyceryltrinitrate for paediatric acute anal fissure: a randomized clinical trial 1mukhtiar ahmed, 2jamil akhter munir ahmad, 2muhammad ali sheikh, 3tariq latif, 4abdul qayyum 1department of paediatric surgery, abbottabad international medical college, abbottabad. 2department of paediatric surgery, shaikh zayed hospital, lahore 3department of paediatric surgery, shaukat khanum memorial cancer hospital & research centre, lahore 4department of paediatric surgery, fatima memorial hospital, lahore abstract introduction: anal fissure is a common problem in children, the exact etiology of which is unknown and it mostly presents with painful defecation and bleeding per rectum. the standard treatment of anal fissure is lateral internal sphincterotomy but due to risk of fecal incontinence chemical sphincterotomy is used as alternative to surgical sphincterotomy. aims & objectives: to compare the effectiveness of topical diltiazem and lignocaine with glyceryl trinitrate and lignocaine in relieving of symptoms and healing of acute anal fissure in children. place and duration of study: this study was conducted in the department of paediatric surgery, shaikh zayed hospital, lahore & department of paediatric surgery, fatima memorial hospital, lahore from september 2017 to september 2018. material & methods: total 228 children were enrolled in the study and randomly divided in group a and b, 114 children in each group. group a received topical 2% diltiazem cream and 2% lignocaine gel, while group b received topical 0.2% glyceryl trinitrate and 2% lignocaine gel, applied locally, twice daily. results: there were 78(68.4%) male children in group a and 66(57.9%) in group b. all patients completed 6 week treatment course. the symptoms and condition of the anal fissure were evaluated before start of treatment and at subsequent follow up periods. in group a 55 (48.2%) cases completely healed by second week, while in group b, 33 (28.9%) cases healed. the number of completely healed cases at 4 weeks follow up in group a and group b were 91 (79.8%) and 69(60.5%) respectively, while at week 6 follow up this rate was 95 (83.3%) and 73 (64.0%) respectively. symptomatic relief in painful defecation observed in group a and group b was 74(64.9%) and 55(48.2%) at week 2, 95(83.3%) and 74(64.9%) at week 4 while 95(83.3%) and 77(67.5%) at week 6 in two groups respectively. conclusion: use of combination of topical diltiazem and lignocaine for the treatment of acute anal fissure in paediatric population is preferred over combination of glyceryl trinitrate and lignocaine. key words: anal fissure, children, diltiazem, glyceryl trinitrate. introduction anal fissure (af) is a longitudinal tear in the mucosa of distal anal canal extending into the anal verge. the most common site of anal fissure is posterior midline, though it may occur anywhere in anal circumference. in females the most common location is anterior midline.1,2 the anal fissure commonly presents with painful defecation and bleeding per rectum (pr). it is confirmed by observing a tear in the anal mucosa.3 the exact etiology of anal fissure is unknown. however, it is suggested that trauma to the anal canal causes tear which fails to heal due to internal anal sphincter (ias) spasm, creating high pressure in the anal canal leading to local ischemia of the anal mucosa.4 in the management of anal fissure, the first line of treatment is to relieve constipation (causing trauma) by increase in fluid intake, high fiber diet and stool softeners. application of local anesthetics and warm sitz bath, also help in healing of anal fissures.5 different pharmacological treatment options are available which include topical glyceryl trinitrate, diltiazem, botulinum toxin, bethanechol, indoramin, and nifedipine. different studies have reported healing of anal fissure by 32% to 64% with lignocaine, 74% to 88.46% with glyceryl trinitrate and with diltiazem upto 96%.3,6,7,8,9 klin et al used 71 topical lignocaine with diltiazem or glyceryltrinitrate for paediatric acute anal fissure nifedipine (which is a calcium channel blocker) with lignocaine and anal fissure completely healed in 92.9% of their patients.10 surgical treatment options include dilatation of internal anal sphincter, fissurectomy and lateral internal sphincterotomy (lis), but these may cause incontinence of stool .5 it was observed by richard et al that internal sphincter spasm was the common finding in anal fissure. in order to overcome spasm of internal sphincter, surgery was the common option but due to fear of fecal incontinence interest in non-surgical treatment has developed.11 the current study was conducted to compare the effectiveness of combination of topical diltiazem and lignocaine with combination of glyceryl trinitrate and lignocaine in relieving of symptoms and healing of acute anal fissure in children. material and methods the study was a randomized clinical trial and was conducted in the department of paediatric surgery, shaikh zayed hospital, lahore & department of paediatric surgery, fatima memorial hospital, lahore september 2017 to september 2018 after the approval from institution review board. the estimated sample size of 228 (divided into two groups of 114 each) was calculated by using 95% confidence level, 80% power with expected healing rate of 93% and 82% in a and b group respectively. treatments were allocated to patient at random by using lottery method. group a topical 2% diltiazem cream and 2% lignocaine gel. group b topical 0.2% glyceryl trinitrate cream and 2% lignocaine gel. all preparations were prepared by a local pharmacy. inclusion criteria: patients of both genders from 1 year to 14 years of age with acute anal fissure of less than 6-week duration, presenting in paediatric surgery out patient department and paediatric emergency. exclusion criteria patients having: inflammatory bowel disease perianal fistula and abscess hemorrhoids hirschsprung’s disease congenital heart disease previously treated anal fissure severe headache (migraine) previous anal surgery & anorectal malformation. longitudinal tear in the mucosa of distal anal canal extending to the anal verge noticed on visual inspection was labelled as anal fissure. partial healing was labelled when granulation tissue was noted at the site of anal fissure on follow up. complete healing was labelled when complete epithelialization of the mucosa at the site of anal fissure was noted on follow up. after taking written consent, the patients were divided at random by lottery method into group a and b for treatment. group a received combination of topical 2% diltiazem and 2% lignocaine while group b received combination of topical 0.2% glyceryl trinitrate and 2% lignocaine. individuals in each group were advised to apply the given creams of pea-sized quantity at anal margins twice daily with 4 hours interval for 6 weeks period. follow up were made after 2, 4 and 6 weeks of treatment. along with local application it was advised to increase fluid intake, stool softeners and sitz baths in both the groups to treat and prevent constipation. constipation was treated aggressively in patients of both groups so that it did not affect our results. success indicator for both groups was the complete healing of anal fissure and relief of symptoms. statistical analysis: data was analyzed by using spss 20.0. quantitative data like age, duration of painful defecation, duration of bleeding frequency were described by using mean & sd for two groups. comparison between two groups for these variables was made by using independent sample t-test. qualitative data like gender, location of fissure, painful defecation, bleeding per rectum, bleeding frequency per week at enrolment were all described by using frequency and percentages for two groups, and comparisons between two groups were made by using chi-square test. p-value ≤0.05 was considered statistically significant. results the study included 228 children with anal fissure (af). these children were divided into two groups of 114 each. the group a was treated with combination of topical 2% diltiazem and 2% lignocaine and group b with combination of topical 0.2% glyceryl trinitrate and 2% lignocaine. the results were observed after 2, 4 and 6 week time. there were 78 (68.4%) male children in group a and 66 (57.9%) in group b with p value of 0.099, (table-1). the mean age for both groups was almost same with p value of 0.890. the number of children with af located posteriorly in group a were 94 72 topical lignocaine with diltiazem or glyceryltrinitrate for paediatric acute anal fissure (82.5%) wile in group b were 71 (62.3%). the children in group b had significantly higher number of fissures in anterior location (37.7%) vs (17.5%) as compared to group a with p value of 0.001 (table-1). painful defection was also significantly higher in children in treatment group b with p-value 0.016. (table-1) the duration for painful defecation in group a was 14.34 ± 7.74 days while in group b was 12.36 ± 7.17 days, with p-value 0.054. bleeding per rectum was found similar in both groups. (table-1) the average duration of bleeding at presentation in group a was 11.83 ± 8.41 days while 8.06 ± 5.61 days in group b, and the difference was statistically significant. symptomatic relief in painful defecation observed in group a and group b was 74 (64.9%) and 55 (48.2%) at week 2, 95 (83.3%) and 74 (64.9%) at week 4 while 95(83.3%) and 77 (67.5%) at week 6 in two groups respectively. the difference in relief of pain while defecation between two groups was statistically significant with p-values 0.011, 0.001 and 0.006 at 2, 4 and 6 weeks follow up times respectively. (table-2) when changes in painful defecation compared between follow up times in each group, the change between 2 and 4 weeks was significant for both groups with p-value <0.001. the change in group b was insignificant between 4 and 6 weeks with pvalue 0.375, while in group a there was no symptomatic improvement in painful defecation between weeks 4 and 6. at start of study in group a 61 (53.5%) cases had no bleeding per rectum while in group b there were 65 (58.0%) cases without bleeding per rectum. at 2 weeks follow up the numbers increased to 86 (75.4%) and 81 (71.1%) in group a and group b respectively. at 4 weeks follow up these numbers increased to 98 (86%) and 94 (82.4%) and at 6 weeks follow up increased to 100 (87.7%) and 101 (88.6%) in group a and group b respectively. the difference between two groups was insignificant with p-values 0.209 and 0.245 at 4 and 6 weeks respectively, while at week 2 the p-value was 0.058. when comparison of bleeding frequency was made between follow up times within each group, the change in group a was found significant between 2 and 4 weeks with p-value 0.001. the mcnemar was not measureable for group b at both times while for group a at week 6. in group a 55 (48.2%) cases completely healed by second week, while in group b, 33 (28.9%) cases healed. the number of completely healed cases at 4 weeks follow up in group a and group b were 91 (79.8%) and 69 (60.5%) respectively, while at week 6 follow up this rate was 95 (83.3%) and 73 (64.0%) respectively. (table-3) within group a, the healing of af between 2 and 4 weeks’ time was significant with p-value <0.001 and same was the case for group b. then between 4 and 6 weeks follow up the change in group b was significant with p-value 0.003, while in group a only four partially healed fissures converted to fully healed and no change in unhealed fissures, mcnemar was not measureable for group a. at 6 weeks follow up 83.3% cases in group a and 67.5% patients in group b were free of symptoms and none of them had constipation. none of the patient developed headache or itching in both the groups. group pvalue group a n = 114 group b n = 114 n % n % gender male 78 68.4 66 57.9 0.099 female 36 31.6 48 42.1 location of af anterior 20 17.5 43 37.7 0.001 posterior 94 82.5 71 62.3 painful defecation yes 102 89.5 111 97.4 0.016 no 12 10.5 3 2.6 bleeding pr yes 53 46.5 47 42.0 0.493 no 61 53.5 65 58.0 frequency of bleeding per week nil 61 53.5 67 58.8 0.147 1.00 3 2.6 9 7.9 2.00 25 21.9 20 17.5 3.00 15 13.2 10 8.8 4.00 3 2.6 4 3.5 5.00 1 0.9 2 1.8 6.00 0 0.0 1 0.9 7.00 6 5.3 1 0.9 table-1: basic features of patients at the time of enrolment in both groups symptomatic relief in painful defecation group pvalue group a n = 114 group b n= 114 n % n % week 2 yes 74 64.9 55 48.2 0.011 no 40 35.1 59 51.8 week 4 yes 95 83.3 74 64.9 0.001 no 19 16.7 40 35.1 week 6 yes 95 83.3 77 67.5 0.006 no 19 16.7 37 32.5 table-2: symptomatic relief in painful defecation observed at three follow up times in both groups 73 topical lignocaine with diltiazem or glyceryltrinitrate for paediatric acute anal fissure condition of af group pvalue group a n=114 group b n=114 n % n % week 2 healed 55 48.2 33 28.9 0.001 partially healed 39 34.2 38 33.3 not healed 20 17.5 43 37.7 week 4 healed 91 79.8 69 60.5 0.006 partially healed 4 3.5 10 8.8 not healed 19 16.7 35 30.7 week 6 healed 95 83.3 73 64.0 0.001 partially healed 0 0.0 4 3.5 not healed 19 16.7 37 32.5 table-3: condition and status of fissure at three follow up times in both groups discussion the exact etiology of anal fissure is unknown. however, it is suggested that anal fissure may be caused by trauma to the anal canal by passage of hard stool or bouts of loose stool. increased tone of internal anal sphincter and presence of ischemia leads to non-healing and persistence of anal fissure.15 anorectal manometry showed fewer internal sphincter relaxations in chronic anal fissure.16 the passage of hard stool causes tear in anal mucosa that produces painful defecation and bleeding per rectum. next time the child voluntarily holds stool to avoid another painful defecation which results in exacerbation of constipation that leads to harder stool and thus produces vicious cycle. the main stay of treatment is directed to break this vicious cycle, by blocking spasm of internal anal sphincter and relieving constipation.3,5,17 in children functional constipation due to behavioral changes has also been reported in early age group. mother with high psychological distress and low level of education were reported as a risk factor for constipation in children.12 different therapeutic options are available for the treatment of acute anal fissure (aaf) including chemical (pharmacological) and surgical sphincterotomy. the standard treatment is lateral internal sphincterotomy but it is invasive, requires hospitalization, spinal or general anesthesia and may cause incontinence. so chemical sphincterotomy is considered as the first option in the treatment of acute anal fissure in children.18 half of fissures heal by increasing fluid intake, increase in fiber diet, use of stool softeners, application of local anesthetics and warm sitz baths.5. many studies have shown that topical glyceryl trinitrate (gtn) treatment heals af in 40.4 68% cases and it is superior to topical lignocaine.11,19 demirbag et al. reported 83.87% healing rate of anal fissure with 8 weeks of gtn therapy and transient fecal incontinence was reported in 1 patient.20 side effects of topical gtn like headache, perianal dermatitis, hypotension and incontinence limits its use.19,21 another drawback of gtn treatment of af is recurrence of the fissure at the rate of 7.9% to 50%.23 diltiazem (dtz) is more effective in treating af than gtn and relieves symptoms quickly.3 dtz has fewer side effects as compared to gtn.3,18 patients with af who do not respond or who develop side effects by the use of topical nitroglycerine can be treated with dtz, which heals af between 50% – 92% cases.5 in other studies healing rate of af with dtz was observed between 67% to 89.4%.13,23,24 a large number of studies were conducted in adults comparing diltiazem and gtn, while in children most studies described the role of topical gtn, diltiazem and lignocaine individually in the treatment of acute anal fissure. in a study by klin et al combination of 0.2% nifedipine with lidocaine was effective in the treatment for anal fissures in children which also showed good results of healing with no side effects.14 one pilot study was conducted in israel in which combination of calcium channel blocker (nifedipine) with lignocaine was used to treat anal fissure in children.10 in our study the incidence of aaf was higher in male children (63%) than female (37%) in both groups. male to female ratio was 1.7: 1. in other studies females were slightly more than males.3,10 in the study by cevik et al. the reported incidence of af more in female was 52.7%.3 mean age of children in our study was 4.72 ± 2.48 years in group a and 4.67 ±2.89 years in group b, while in a study conducted by cevik at el the mean age was 2.5 years in diltiazem group and 2 years in gtn group.3 in study conducted by demirbag at el mean age was 3.1 year in gtn group.22 in comparison to cevik et al. study the mean age in our study population was almost double which shows that our patients seek medical advice late. in our study posterior anal fissures (72.37%) were more common than anterior fissures (27.63%). in group a 82.5% fissures were located posteriorly and 17.5% anteriorly, while in group b posterior and anterior fissures were 82.3% and 37.7% respectively. the difference in location of af in our study was statistically significant (p 0.001). this was confirmed by other studies in which posterior anal fissure were more common than anterior fissure.3,9,10 klin et al. reported 90% posterior anal 74 topical lignocaine with diltiazem or glyceryltrinitrate for paediatric acute anal fissure fissure and around 10% anterior fissure, and giridhar et al. reported 93.3% posterior fissure.9,10 in our study 93.4% children with aaf presented with painful defecation. in a study conducted by cevik et al. the incidence of painful defecation in children was reported in 94.6%, while in study conducted by klin et al. painful defecation was reported in 87% cases.3,10 our results were comparable with results reported in literature. in our study 37.7% of children had bleeding per rectum, cevik et al. reported bleeding per rectum in 68.4% cases3, while klin et al. reported 84% cases with bleeding per rectum.10 in our study a smaller number of children presented with bleeding per rectum as compared to cevik at el and klin et al. studies.3,10 in our study symptomatic relief in painful defecation was observed in 83.3% and 67.5% in group a and b respectively. klin et al. and cevik et al. reported 83.9% and 92.9% symptomatic relief respectively.3,10 in our study relief in pain between group a and b was statistically significant (p 0.006). in a study conducted by pardan at el reported 76%% good to excellent symptomatic relief with dtz and 52% with gtn.8 in our study no bleeding was observed in 87.7% (group a) and 88.6% cases (group b) after completion of 6 weeks’ treatment. the relief in bleeding per rectum was statistically insignificant (p 0.245). cevik et al. reported 92.9% symptomatic relief with dtz and 82.1% with gtn therapy.3 complete healing of af was observed in 96 (83.3%) with dtz and 73 (64.0%) with gtn in our study. the difference was statistically significant (p 001). cevik et al. reported a healing rate of 92.9% and 82.1% after 8 weeks of therapy with dtz and gtn respectively.3,10 no untoward effects were observed in both of our groups after 6 weeks of treatment. cevik et al. reported perineal dermatitis in one patient in dtz group and one in gtn group, while no headache was observed in both groups.3 joda et al , reported showed that gtn ointment was effective treatment for anal fissure in children, with good healing, early relief of symptoms and less side effects.22 demirbag at el reported one case of transient fecal incontinence with gtn therapy and no headache was noted in any of the child.20 newman et al also reported that topical diltiazem hydrochloride (2%) or topical nifedipine (0.2–0.5%) both proved to be effective alternatives with less side effects when compared with gtn.25 bansal et al reported that 2% diltiazem ointment and .02% gtn in the treatment of chronic anal fissure, were equally effective in relieving pain, healing and recurrence, but patients treated with gtn ointment had headache. so 2% diltiazem ointment may be preferred in the medical management of chronic anal fissure as first line of treatment.26 may be due to less duration (6 week) of treatment and as parents were also instructed not to apply gtn inside the anus to decrease systemic absorption, no side effect were observed in our study population. the limitation of our study was that both of these creams were not available as readymade in combination with lignocaine, so there were dispensed by the pharmacy on our request. conclusion use of combination of topical diltiazem and lignocaine for the treatment of acute anal fissure in paediatric population is preferred over combination of glyceryl trinitrate and lignocaine or any of these drugs alone. topical diltiazem with lignocaine can be used as 1st line treatment in the management of acute anal fissure in children. references 1. zaghiyan kn, fleshner p. anal fissure. clin colon rectal surg. 2011; 24(01):022-30. 2. ayantunde aa, debrah sa. current concepts in anal fissures. world j surg.2006;30(12):2246-60 3. cevik m, boleken me, koruk i, ocal s, balcioglu me, aydinoglu a, et al. a prospective, randomized, double-blind study comparing the efficacy of diltiazem, glyceryl trinitrate, and lidocaine for the treatment of anal fissure in children. pediatr surg int. 2012; 28(4):411-6. 4. katsinelos p, kountouras j, paroutoglou g, beltsis a, chatzimavroudis g, zavos c, et al. aggressive treatment of acute anal fissure with 0.5% nifedipine ointment prevents its evolution to chronicity. world j gastroenterol. 2006; 12(38):6203-6. 5. rakinic j. anal fissure.clin colon rectal surg. 2007; 20(02):133-7. 6. motie mr, hashemi p. chronic anal fissure: a comparative study of medical treatment versus surgical sphincterotomy. acta med iran. 2016; 54(7):437-40. 7. ahmad j, andrabi s, rathore m. comparison of topical glyceryl trinitrate with lignocaine ointment for treatment of anal fissure: a randomised controlled trial.int j surg. 2007; 5(6):429-32. 8. pardhan a, azami r, mazahir s, murtaza g. diltiazem vs. glyceryl tri-nitrate for symptomatic relief in anal fissure: a randomised 75 topical lignocaine with diltiazem or glyceryltrinitrate for paediatric acute anal fissure clnical study. j pak med assoc. 2014; 64(5):510-3. 9. giridhar c, babu p, rao ks. a comparative study of lateral sphincterotomy and 2% diltiazem gel local application in the treatment of chronic fissure in ano. j clin diagn res. 2014; 8(10):nc01-2. 10. klin b, abu-kishk i, efrati y, lotan g. nifedipine gel with lidocaine in the treatment of anal fissure in children a pilot study and review of the literature. complementary pediatrics. 2012; 4: 53-71. 11. nelson r. a systematic review of medical therapy for anal fissure. dis colon rectum. 2004; 47(4):422-31. 12. kilincaslan h, abali o, demirkaya s k, bilci m. clinical, psychological and maternal characteristics in early functional constipation. pediatr int 2014; 56: 588–93. 13. carapeti ea, kamm ma, phillips rk. topical diltiazem and bethanechol decrease anal sphincter pressure and heal anal fissures without side effects. dis colon rectum. 2000; 43(10):1359-62. 14. klin b, efrati y, berkovitch m, et al. anal fissure in children: a 10-year clinical experience with nifedipine gel with lidocaine. minerva pediatr 2016; 68: 196–200. 15. jonas m, scholefield jh. anal fissure. gastroenterol clin north am.2001;30(1):167-81 16. farouk r, duthie g, macgregor a, bartolo d. sustained internal sphincter hypertonia in patients with chronic anal fissure. dis colon rectum. 1994; 37(5):424-9. 17. poh a, tan k-y, seow-choen f. innovations in chronic anal fissure treatment: a systematic review. world j gastrointest surg. 2010; 2(7):231-41. 18. hashmi f, siddiqui fg. diltiazem (2%) versus glyceryl trinitrate cream (0.2%) in the management of chronic anal fissure. j coll physicians surg pak. 2009; 19(12):750-3. 19. knight j, birks m, farouk r. topical diltiazem ointment in the treatment of chronic anal fissure. br j surg. 2001; 88(4):553-6. 20. demirbag s, tander b, atabek c, surer i, öztürk h, cetinkursun s. long-term results of topical glyceryl trinitrate ointment in children with anal fissure. ann trop paediatr. 2005; 25(2):135-7. 21. tankova l, yoncheva k, kovatchki d, doytchinova i. topical anal fissure treatment: placebo-controlled study of mononitrate and trinitrate therapies.int j colorectal dis. 2009; 24(4):461-4. 22. joda ae, al-mayoof af. efficacy of nitroglycerine ointment in the treatment of pediatric anal fissure. j pediatr surg 2017; 52:1782-6. 23. kocher h, steward m, leather a, cullen p. randomized clinical trial assessing the side-effects of glyceryl trinitrate and diltiazem hydrochloride in the treatment of chronic anal fissure. br j surg. 2002; 89(4):413-7. 24. jonas m, speake w, scholefield jh. diltiazem heals glyceryl trinitrate–resistant chronic anal fissures. dis colon rectum. 2002;45(8):1091-5. 25. newman m, collie m. anal fissure: diagnosis, management, and referral in primary care. br j gen pract. 2019; 69(685): 409-410. 26. bansal a. r., pawan kumar yadav, rajesh godara, noresh pal, rathindra tripura, jaikaran. comparative evaluation of 0.2% glyceryl trinitrate vs. 2% diltiazem ointment in treatment of chronic anal fissure: a case trial. hellenic journal of surgery 2016; 88: 25-30. the authors: dr. mukhtiar ahmed senior registrar, department of paediatric surgery, abbottabad international medical college, abbottabad. dr. jamil akhter munir ahmad senior registrar, department of paediatric surgery, shaikh zayed hospital, lahore. prof. muhammad ali sheikh head, department of paediatric surgery, shaikh zayed hospital, lahore. prof. tariq latif consultant paediatric surgeon, department of paediatric surgery, shaukat khanum memorial cancer hospital, lahore. dr. abdul qayyum senior registrar, department of paediatric surgery, fatima memorial hospital, lahore. corresponding author: dr. jamil akhter munir ahmad senior registrar, department of paediatric surgery, shaikh zayed hospital, lahore. e-mail: doctorjamil@yahoo.com 37 proceedings s.z.m.c. vol: 35(3): pp. 37-43, 2021. pszmc-806-35-3-2021 tamarix dioica (ghaz) protective potential in the carbon tetrachloride-induced hepatotoxicity animal model 1sumra komal, 2aqna malik, 3naheed akhtar, 4syed asif jahanzeb kazmi, 4fayyaz anjum, 4ayesha rida 1department of pharmacology, school of basic medical sciences, zhengzhou university, zhengzhou, china 2department of pharmacy, the university of lahore, gujrat 3department of medicine, cmh institute of medical sciences, bahawalpur 4department of pharmacology and therapeutics, cmh institute of medical sciences, bahawalpur abstract introduction: hepatic diseases remain the leading cause of death worldwide. despite overall advancements in health care, mortality due to hepatic diseases is constantly growing. more than 2 million people globally are estimated to die each year from liver diseases, and current treatment offers little for its management. thus, it is essential to find more effective and less toxic pharmaceutical alternatives for the treatment of liver diseases. aims & objectives: tamarix dioica, a shrub broadly used in herbal medicine for the treatment and prevention of various diseases. the current study was designed to analyze the hepatoprotective effect of t. dioica in balb⁄cmice against ccl4-induced acute liver damage. place and duration of study: the study was conducted in nih, islamabad, pakistan, for six months in 2016-2017. material & methods: for in vivo evaluation, the animals (n= 42) were randomly divided into seven groups (n=6), three control (i.e. group, i or normal control, group ii or induction control received 0.9% normal saline orally, and group iii or positive control received silymarin 100 mg/kg per oral), and four treatment groups (i.e. iv, v,vi and vii were treated with oral t.dioica 200 mg/kg/day, 300mg/kg/day methanol extract, 200mg/kg/day and 300mg/kg/day of aqueous extracts respectively for six days, followed by intraperitoneal administration of ccl4 on the seventh day. the blood samples were collected for analysis of lfts, and hepatic tissue was taken for histological analysis. data was analyzed using spss version 16, one-way anova with duncan’s multiple range test (dmrt). results: ccl4 induction in group 2 resulted in severe hepatic derangement manifested as highly elevated mean lfts (alt 7245.56, ast 3292.11, alp 340.09 u/l, bilirubin 4.64 mg/dl) as compared to healthy controls (alt 38.97, ast 50.20, alp 57.17 u/l, bilirubin 1.25 mg/dl: (group 1) levels p<0.001. pretreatment with different extracts of t.dioica for 6 days before ccl4 administration produced varying degrees of hepatoprotection. 300mg/kg aqueous extract t.dioica (group7) prevented damage with maximal hepatoprotection, reduced lfts (alt: 339.95 , ast: 242.90 , alp: 116.86 u/l, bilirubin: 1.38 mg/dl) and normalized liver histology as compared to group 2 and standard drug silymarin 100mg/kg, (alt: 6483.23, ast: 2567.69, alp: 272.19 u/l, bilirubin: 2.84 mg/dl: group 3) p<0.001. lesser hepatoprotection was provided by t.dioica aqueous extract 200mg/kg (alt: 439.93, ast: 367.87, alp: 180.62 u/l bilirubin: 1.53 mg/dl: group vi) and least by 300mg/kg & 200mg/kg methanolic extracts groups v & iv (alt: 6338.06, 6443.91, ast: 2800.81, 3012.34, alp: 242, 248 u/l & bilirubin: 2.82 & 3.62 mg/dl) respectively. further, no drug-induced toxicity symptoms were observed 24 hours after administration of the high dose oral t. dioica 2000 mg/kg/body weight aqueous and methanolic extracts were administered. conclusion: pretreatment with t. dioica extracts especially 300mg/kg aqueous extract reduced acute ccl4-mediated liver damage, ameliorated histopathological as well as biochemical parameters and was free of toxicity in 2000mg/kg /body weight dose in the mice experimental model. t. dioica has potential in hepatoprotective drug research. key words: tamarix dioica, hepatoprotective, silymarin, carbon tetrachloride introduction liver regulates the majority of body functions, including detoxification, metabolism, antimicrobial defenses, and immune response whereas, any disturbance in its normal functioning may lead toward serious consequences.1 the extensive metabolic capacity of the liver makes it more susceptible to chemicals includes liver injuries.2 carbon tetrachloride (ccl4) is identified as a possible human carcinogen. in animals, it is rapidly absorbed into the systemic circulation via the gastrointestinal tract.3,4ccl4 administration by inhalation, gastric infusion, and oral bolus gave its peak concentration in fats, liver, kidney, brain, and lung, therefore extensively used to induce experimental hepatopathy.5,6 in experimental murine 38 tamarix dioica (ghaz) protective potential in the carbon tetrachloride-induced hepatotoxicity animal model models, the cytochrome p450 2e1 is predominantly involved in the bio activation of ccl4.7 accidental exposure to ccl4 can cause acute hepatic and renal failure followed by dialysis when exposed to 200 ppm ccl4 for less than 3 hours.8 furthermore, hepatic tumors were observed at higher ccl4 doses followed by loss of cellular ca+2 sequestration and disruption of ca+2 homeostasis with subsequent cell damage.9ccl4 also stimulates fibrosis by activating nitric oxide (no), tumor necrosis factor α (tnf-α), and transforming growth factor (tgf)-α/β in the cell and pushes the cell toward apoptosis.10 hepatitis cirrhosis, alcoholic liver disease, hemochromatosis, and hepatomas are some examples of liver diseases and current medical treatments for such hepatic diseases are either too expensive or associated with adverse effects.11 medicinal plants have been used worldwide for their therapeutic purposes since ancient times. extracts of several medicinal plants show permissible results against various diseases without any critical side effects.12 multiple studies validated the usage of traditional medicine in the management of various diseases.13 tamarix dioica (t. dioica) is a small shrub that belongs to the family tamaricaceae.14 the plant is well-known in ancient and present herbal remedies for its antidiabetic, anti-fungal, anti-infective and antidermatosis, carminative, anti-inflammatory, and diuretic properties.15,16 nonetheless, there are still few clinical studies for the therapeutic effectiveness of t. dioica. keeping in view these facts, the present study is designed to evaluate the hepatoprotective potential of t. dioica plant aqueous and methanolic extracts against ccl4-induced toxicity into balb/c mice. material and methods plant material leaves of t. dioica were obtained and identified by the department of botany, pmas-arid agricultural university rawalpindi, pakistan. chemicals and reagents carbon tetrachloride, silymarin, olive oil, methanol, distilled water, serum bilirubin and diagnostic kits (huma star 600) of pharmacopoeia grade were obtained from national institute of health (nih), islamabad, pakistan. animal selection and maintenance balb⁄c mice body weights 35-45g on standard laboratory diet were obtained from nih islamabad, pakistan, and were kept at 25±1ºc temperature, the relative humidity of 10% with normal light and dark schedule. the research proposal was evaluated and approved by research ethical committee and a lot reference number 10-m.ph/lcwu-17981. extract preparation tamarix dioica aqueous extract dried t. dioica aqueous extraction were prepared by grinding followed by soaking in distilled water (20g/200ml) for overnight and then boiled at 100oc. the extract was cooled at room temperature and supernatant was then filtered through whatman® grade 42 filter paper.17 tamarix dioica methanolic extract dried plant material was ground and extracted with 80% methanol (20g/200ml) to obtain the crude methanolic extract. the extract was concentrated by rotary evaporation and stored in the refrigerator at 4oc for future study and dose preparation.17 biological study drug administration protocol and animal grouping animals were divided into seven groups (n=6). all groups received standard food orally throughout the study period. group, i (normal control or nc) and group ii (induction control) were administered 0.9% normal saline orally for six days. group iii (positive control) received silymarin 100 mg/kg per oral. two treatment groups (iv and v) were treated with methanol extract (me), and the other two groups (vi and vii) received aqueous extracts (ae) orally at the dose of 200 and 300 mg/kg respectively, for six days. all groups, except normal control, received an intraperitoneal injection (i.p.) of ccl4 in olive oil (1:1) at the dose of 0.5 mg/kg for hepatotoxicity induction on the seventh day of study. acute toxicity studies male mice were selected for the acute toxicity testing according to oecd guidelines-423.18 t.dioica aqueous and methanolic extracts at the dose of 2000 mg/kg body weight were administered p.o while distilled water was given to the control group. blood and liver sample collection the experimental animals were sacrificed for blood and liver collection. blood was collected directly from the aorta followed by centrifugation for five minutes at 12000 rpm/min. the supernatant was collected and preserved at -20oc for further analysis. livers samples were preserved in 10% formalin solution and stored for subsequent analysis.19 liver biochemical assessment liver function tests (lfts) including aspartate aminotransferase (ast), alkaline phosphatase (alp), alanine transaminase (alt), and total bilirubin were determined via commercially available diagnostic kits (humastar 600). histopathological observation animal liver samples were obtained for histopathological examination and preserved in 39 tamarix dioica (ghaz) protective potential in the carbon tetrachloride-induced hepatotoxicity animal model formalin (10% v/v) solution followed by paraffin embedding and cutting into 5μm thick sections with a microtome and were stained subsequently with hematoxylin and eosin (h&e) dye and were observed under the light microscope (im-910 irmeco gmbh & co. germany). statistical analysis: statistical analysis was done by using spss software version 16.0, one-way analysis of variance (anova) with duncan's multiple range test (dmrt). all results were expressed as mean ± se and 5% level of significance (p ≤ 0.05). results effect of t. dioica extracts on bilirubin levels against ccl4-induced acute liver injury in balb/c mice: the results depicted that the bilirubin levels in the induction group (ii) significantly increased, p< 0.05, (4.64 ± 0.029) as compared to the control group (1.25 ± 0.018). it shows that ccl4 induces hepatotoxicity. the groups that received the pretreatment with me and ae have statistically significant lower levels of bilirubin (p<0.05), as shown in table-1 and fig-1. effect of t. dioica on liver parameters a significant increase level of ast and alt (p≤ 0.05) was observed in the carbon-tetrachloride treated (ii) group as compared to the normal control group (table-1). the me and ae pretreatment and standard drug treatment have significantly (p<0.05) reduced the upregulated serum liver markers (ast and alt), which showed the hepatoprotective effects of t. dioica. as shown in table-2 and fig-2. furthermore, among various treatment groups, maximum protection was observed in ae treated group against ccl4-induced acute liver injury. as shown in table-3 and fig-3. tamarix dioica protects mice from ccl4-induced acute liver injury liver histopathological examination showed significant changes on liver section exposed with ccl4 (b) when compared to the control (nc) group (a) shown in fig-4. the changes in ccl4 exposed liver section revealed abnormal morphological characteristics, vacuolated hepatocytes, fat accumulation, mitotic figures and the severity of hepatic damage. the histopathological examination of liver sections of standard group (c) showed to have significantly reduced mitotic figures, vacuolated hepatocytes and no fat accumulation. the pretreated mice groups with ae (d: 200mg/kg & e: 300mg/kg) and me (f: 200mg/kg & g: 300mg/kg) found to have significantly organized liver tissues, highly significant lower fat accumulation and vacuolated hepatic cells as compared to ccl4 exposed (b) group. the animal group treated with ae 300mg/kg showed to have maximum liver protection. furthermore, no toxicity symptoms were observed after 24 hours when a high dose of 2000 mg/kg body weight of t. dioica aqueous and methanolic extracts was administered p.o. compare to the control group. serum parameters (u/l) alp alt ast total bilirubin (mg/dl) nc (group i) 57.17 ± 0.731 38.97 ± 0.54 50.20 ± 0.69 1.25 ± 0.018 induction control (group ii) 340.09 ± 5.709 7245.56 ± 84.40 3292.11 ± 38.51 4.64 ± 0.029 positive control (group iii) 272.19 ± 3.386 6483.23 ± 28.24 2567.69 ± 28.29 2.84 ± 0.13 treatmentme 200 (group iv) 248 ± 3.396 6443.91 ± 36.74 3012.34 ± 36.40 3.62 ± 0.094 treatment me 300 (group v) 242 ± 3.715 6338.06 ± 44.64 2800.81 ± 29.58 2.82 ± 0.133 treatmentae 200 (group vi) 180.62 ± 5.349 439.93 ± 0.86 367.87 ± 7.54 1.53 ± 0.039 treatmentae 300 (group vii) 116.86 ± 2.681 339.95 ± 2.91 242.90 ± 3.49 1.38 ± 0.109 table-1: effect of silymarin and t. dioica pretreatment at seventh day on liver function tests of ccl4intoxicated mice serum parameters (u/l) positive control treatmentme 200 treatmentme 300 treatmentae 200 treatmentae 300 alp 20.40% 40.13% 43.47% 18.98% 93.13% alt 98.33% 85.62% 82.21% 99.07% 100% ast 96.29% 93.34% 91.98% 94.21% 98.89% total bilirubin (mg/dl) 100% 89.65% 68.96% 93.1% 75.86% table-2: percentage protection after t. dioica extracts pretreatment compares to the standard treatment group treatment groups percentage protection silymarin (100mg/kg) + ccl4 78.75 % methanolic extract (200mg/kg) + ccl4 77.2 % methanolic extract (300mg/kg) + ccl4 71.65 % aqueous extract (200mg/kg) + ccl4 76.34 % aqueous extract (300mg/kg) + ccl4 91.97 % table-3: percentage protection of silymarin and t. dioica leaves extracts against ccl4-induced liver injury 40 tamarix dioica (ghaz) protective potential in the carbon tetrachloride-induced hepatotoxicity animal model fig-1: t. dioica protects experimental animals from ccl4-induced liver damage and significantly alters the level of serum biomarkers in treatment groups. pretreatment with ae of t. dioica (300 mg/kg/day p.o) for six days prior to ccl4 exposure (0.5 mg/kg) on seventh day significantly protects hepatocytes in balb/c mice (n = 6) from liver injury due to the ccl4 as assessed by measuring (a) serum alt, (b) ast, (c) alp, and (d) total bilirubin activity. each value represents mean ± s.e.m. (n=6) at p<0.001. ap<0.001 as compared to group i (normal control). bp<0.001 as compared to group ii (induction control). cp<0.001 as compared to group iii (positive control). fig-2: percentage protection of various biochemical parameters against ccl4-induced hepatic injury. percentage protection of treatment groups, when compared to that of induction control group, revealed that aqueous extract (300mg/kg) produced maximum hepatocellular protection as depicted by values of all serum biomarkers. fig-3: effects of t. dioica extract on hepatic cells protection t. dioica aqueous extract (300mg/kg) exhibited the maximum overall protection to hepatic cells (91.97 %) when compared to the positive control group (silymarin 100mg/kg) and methanolic extracts (me200 and me-300) respectively. fig-4 shows; (a) normal cells, (b) ccl4-induced cells, (c) standard or silymarin-induced cells (200 mg/kg), (d) me-200 + ccl4 treated cell, (e) me300 + ccl4 treated cell, (f) ae-200+ ccl4 treated cell, (g) ae-300+ ccl4 treated cell. these results show that pretreatment with t. dioica methanolic (d& e) and aqueous (f & g) extracts significantly protects the hepatic cells from injury compared to silymarin (c) and ccl4-induce (b) groups. 41 tamarix dioica (ghaz) protective potential in the carbon tetrachloride-induced hepatotoxicity animal model fig-4: histopathological examination of liver samples collected from balb/c mice. discussion medicinal plants are the most important source of traditional medicine for the majority of the world’s population. natural products have attracted more attention in the last 20 years as potential sources of new antibacterial, antiviral, and phytotoxic chemicals.20 t. dioica requires significant attention due to its biological activities. previous studies on t. dioica phytochemical screening showed that steroids and phlobatannins are present in all parts of the plant, while phenols, flavonoids, and tannins, are present in leaves, flowers, and roots; saponins and terpenoids are present in flowers, leaves, and stems.21 studies revealed that plants that contains the highest contents of phenols and flavonoids, are enriched in antioxidants and exhibits their hepatoprotection action against ccl4-induced liver injuries by inhibiting nf-кb, tnf-α, and tgf-α/β signaling pathways.22,23 previous studies on phytochemicals screening of t. dioica have revealed its strong antiallergic, anti-inflammatory, anticarcinogenic, antiviral, antithrombotic, and hepatoprotective, as well as antioxidant potential.24. however, its antioxidant and anti-inflammatory potential supports our findings.25 the present study on liver-protective action showed that t. dioica leaves extracts, both aqueous and methanolic have given promising results against ccl4-induced livers injury in mice and shown relatively great improvement than silymarin. the antihepatotoxic effects of silybum marianum have been studied several times.26 t. dioica aqueous and methanolic extracts have found non-toxic when administered orally to the animals.27,28 animals pretreatment with trichosantes dioica aqueous and methanolic extracts exhibit marked improvement against ccl4-induced hepatotoxicity and also shows significant hepatoprotective effect against ccl4 toxicity in contrast with silymarin.29 a significantly raised (p<0.005) serum level of ast, alp, alt, and total bilirubin were observed in ccl4 treated animals (group ii) as compared to the group treated with standard drug or silymarin (group iii).30 in our study the animal receiving aqueous and methanolic extracts of t. dioica (group iv and vii) show a significant reduction in serum markers levels as a comparison to diseased control. the study further confirms that the administration of t. dioica extracts at the dose of 200mg/kg, 300 mg/ kg revert the level of serum enzymes towards normal. t. dioica aqueous extracts at the dose of 300 mg/kg show efficient therapeutic effects that are comparable to the standard drug silymarin. no comparative data to support or refute our findings regarding t. dioica efficacy against ccl4 were found. moreover, no drug-induced toxicity symptoms were observed when a high dose of 2000 mg/kg body weight of t. dioica aqueous and methanolic extracts were administered p.o. hence, the study revealed that t. dioica extracts are safe and have the potential for future development of advanced hepatoprotective drugs. 31 conclusion t. dioica extracts reduced acute ccl4-mediated liver damage in vivo and ameliorated the histopathological as well as biochemical parameters in mice. the t. dioica has the potential to alleviate the severity of liver damage caused by carbon tetrachloride. hence, it is concluded that t. dioica can be used as supportive therapy for the treatment of drug-induced and other oxidative stress-mediated hepatoxicity in the future. acknowledgement: we pay our sincere gratitude to research supervisor prof. dr. hafeez ikram and co-supervisor prof. dr. mobasher ahmad butt. we extend endless thanks to our family, colleagues and friends for provision of consistent courage and support. references 1. you sl, il jc, joo wk, min kl, sae kk, jae sc, hae j l. hepatoprotective effects of blue 42 tamarix dioica (ghaz) protective potential in the carbon tetrachloride-induced hepatotoxicity animal model honeysuckle on ccl4-induced acute liver damaged mice. food scinutr. 2018; 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9:3689-3695. 20. samejo mq, sumbul a, shah s, memon sb, chundrigar s. phytochemical screening of tamarix dioica roxb. ex roch. j. pharm. res. 2013; 7:181-3. 21. saijou e, enomoto y, matsuda m, yuet yin kok c, akira s, tanaka m, and miyajima a. neutrophils alleviate fibrosis in the ccl4 induced mouse chronic liver injury model. hepatol commun. 2018;2:703-17 22. saijou e, enomoto y, matsuda m, yuet yin kok c, akira s, tanaka m, and miyajima a. neutrophils alleviate fibrosis in the ccl4 induced mouse chronic liver injury model. hepatol commun. 2018;2:703-17 23. nipanikar su, chitlange ss, nagore d. pharmacological evaluation of hepatoprotective activity of ahpl/aytab/0613 tablet in carbon tetrachloride-, ethanol-, and paracetamol-induced hepatotoxicity models in wistar albino rats. pharmacognosy res. 2017; (suppl 1):s41-s47. 24. rohini mv, padmini e. preliminary phytochemical screening of selected medicinal plants of polyherbal formulation. j. pharmacogn. phytochem.2016;5:277. 25. iqbal a, din su, bakht j, khan iu, shah z. antiinflammatory, anti-pyretic and analgesic activities of tamarix dioica. pak. j. pharm. sci. 2019;32. 26. madani h, talebol hosseini m, asgary s and naderi gh. hepatoprotective activity of silybum marianum and cichorium intybus against thioacetamide in rat. pakistan j nutr 2008; 7: 172-176. 27. ezzat mi, okba mm, ahmed sh, el-banna ha, prince a, mohamed so, ezzat sm. in-depth hepatoprotective mechanistic study of phyllanthus niruri: in vitro and in vivo studies and its chemical characterization. plos one. 2020; 15:e0226185. 28. dutta s, chakraborty ak, dey p, kar p, guha p, sen s, kumar a, sen a, chaudhuri tk. amelioration of ccl4 induced liver injury in swiss albino mice by antioxidant rich leaf extract of croton bon plandianus baill. plos one. 2018; 13:e0196411. 43 tamarix dioica (ghaz) protective potential in the carbon tetrachloride-induced hepatotoxicity animal model 29. shivhare y, singh p, rajak h, patil uk, pawar rs. antioxidant potential of trichosanthes dioica roxb. (fruits) phcog j. 2010; 2:107-111. 30. clichici s, olteanu d, nagy al, oros a, filip a, mircea pa. silymarin inhibits the progression of fibrosis in the early stages of liver injury in ccl4treated rats. j. med. food. 2015; 18:290-298. 31. khan s, ullah f, mahmood t. in vitro antimicrobial and cytotoxic activity of tamarix dioica roxb. leaves. turk. j. biol. 2013; 37:329-35. the authors: sumra komal researcher, department of pharmacology, school of basic medical sciences, zhengzhou university, zhengzhou, china. dr. aqna malik, assistant professor, department of pharmacology, the university of lahore, gujrat campus. dr. naheed akhtar assistant professor, department of medicine, cmh institute of medical sciences, bahawalpur. prof. syed asif jahanzeb kazmi hod, pharmacology and therapeutics, cmh institute of medical sciences, bahawalpur. fayyaz anjum lecturer, department of pharmacology and therapeutics, cmh institute of medical sciences, bahawalpur. ayesha rida lecturer, department of pharmacology and therapeutics, cmh institute of medical sciences, bahawalpur. corresponding author: prof. syed asif jahanzeb kazmi hod, pharmacology and therapeutics, cmh institute of medical sciences, bahawalpur. e-mail: drsajk@hotmail.com untitled-1.indd 1 proceedings s.z.m.c. vol: 34(2): pp. 1-3, 2020. pszmc-742-34-2-2020 an approach to screen covid-19 patients using diagnostic kits: with special reference to pakistan 1faiza gul durrani, 2farhat zaheer 1consultant, science shines ltd., london 2retired chief scientific officer (cso), institute of nuclear medicine and oncology, lahore abstract the world health organisation declared covid-19 as a pandemic on 11th march 2020. the main approach to tackle covid-19 worldwide is to screen and provide supportive care to patients. for screening purpose rtqpcrbased test are used as an initial detections assay. the test is expensive, time consuming and not suitable for mass scale screening/ confirmation requirement. a recent advancement is development of immunoassay procedures (liquid phase tests or bed side 10-20 minute strip test). in order to help and accelerate bringing life to normal after lock down, pakistan is in dire need to develop and adopt the immunoassay procedures for mass scale screening and confirmation of covid-19 infection. it is cheap and easy to perform without a lab requirement. introduction an outbreak of novel corona virus infection (covid 19) in wuhan (china), has rapidly spread throughout the world. a current estimate indicates total number of incidences of infected persons in the world is 1,452,378, total deaths 83,614 and recovered persons are 282,147 (8th april, 2020).1 in pakistan, the first patient was reported on 26th february 2020 and number of incidences reported on 8th april 2020 was 4,196 cases, deaths 60 (1.42%) and the number of recovered cases, 467(11.1%) according to a ministry of health report, the projected number of incidences is 50,000 by the end of april.2 background: since 1960s, corona viruses have been recognized as one of the infection sources responsible for common cold in humans. six species of human corona viruses have been identified to date(hku1 ,229e , nl63 ,mers and sarscov).3 the pandemic virus has been designated as a sister to sars-covs, hence the name sars-cov-2 by a corona virus study group of the international committee on taxonomy of viruses. the corona viruses are enveloped viruses with a positive-sense, single stranded rna. the first and complete genome sequence of covid-19 (genbank: mn908947.3) and the structure of the virus’s main protease to the protein data bank (doi: 10.2210/pdb6lu7/pdb) have been reported4. its genome constitutes 29891 nucleotides, encoding for 9860 amino acids. the virus has 70 percent similarity in genetic sequence with sars. the x-ray crystallographic protein data bank structure explained the glycosylated crown shape covering (hence give the name “corona”, latin for crown). these spike proteins facilitate the virus to bind to the receptors on the cells. the covid-19 has 2 s domains s1 and s2. the computational analysis of s1 and s2 has shown the importance of the s1 domain which has many unique residues to not only bind to cd26 receptor cells but also ace2 cells5. it has been reported that this particular virus can infect human respiratory epithelial cells 100-1000 times more as compared to previous corona strains because of its unique binding sites for ace2 receptors. in severe cases, it can cause organ failure and even death6. for any viral infection/medical illness there is a set route of diagnosis and treatment. the strategy advised by who is based on screening and providing supportive system to the patients in need. throughout the world, screening of the infected individuals is essentially based upon: thermal monitoring of the body temperature, breathing problem, and dry cough. it is noteworthy to mention here that clinical symptoms after infection would appear after a period of 3-5 days of infection. these may be delayed for two weeks, besides the fact that some of the infected subjects may remain asymptomatic7. infected individuals identified by screening are isolated at least for 14 days (quarantine) as per who guidelines. 2 an approach to screen covid-19 patients using diagnostic kits: with special reference to pakistan diagnostic tests for covid-19 with special reference to pakistan in pakistan, at present in order to confirm the covid-19 infection, only molecular based rtqpcr is allowed. rt-pcr technique is very well established in all the research institutes of the country.17 in particular case of covid-19, it is performed with the kits available to the govt. (imported purchase or donation). most of the kits contain 3 assays in which the targets are orf1 gene (human rna polymerase protein) e-gene (envelope protein) and the n-gene (the nucleocapsid protein)8 so if the virus does mutate the chance of all three targets mutation is low, however if the results of 1 or 2 of these essays is not positive then the result needs to be revised. a possibility of a false negative and a false positive cannot be excluded and in order to confirm the results it needs to be repeated for verification. bearing in mind that the procedure is lengthy, requires bsl-2 lab facility and it is an expensive test especially in view of repeat required9. in patients with confirmed covid-19 diagnosis, the laboratory evaluation should be repeated to evaluate for viral clearance prior to being released from observation. the possibility of a local rt-q pcr kit production is being explored or it is in the process of validation in pakistan. however, the primers are not synthesized in pakistan. though, at present the requirement of tests is being managed by the country adequately as far as possible however it is still limited to a single test performance, with hardly required repeat tests as mentioned above. also, in view of the rising number of infected patients being identified, the country will reach an exhaustive stage on how to manage the required number of tests. it will not be possible to manage the required lab workload; the country requires to be prepared for that situation. the diagnostic test in particular for covid-19, approved by who in its briefing (early march, 2020) was by rt-qpcr. however, who also emphasized the importance of immunoassay serological tests, which can be used as bedside tests and will be a big breakthrough for testing service at mass level. these immunoassay-based tests reagents/kits have been manufactured by several countries including usa, china, south korea, ireland etc.12,13 who is working on its validation. as soon as the validation is completed, these tests will be recommended because of their specificity for detection of antibody igm, igg in patients who are in recovering phase or have had been previously infected, 10. moreover, the immunoassay test can also be applied for the detection of antigen (virus), during the first week of infection. the detection of antibodies, (igm and igg) in blood by immunochromatography procedure is simple and straightforward. these two substances are produced in blood after infection, usually by day 7 and 14 respectively. it requires a nitrocellulose strip with a sample pad, and coated anti igm and igg pads. to the sample pad a drop of sample (blood) is added by a finger-prick followed by buffer. the sample and buffer move laterally towards first igmantibody pad band where igm, if present in the sample will bind to antibody and a colour band is produced. similarly, the next pad with anti-igg coated will identify the presence of igg produced in blood as a result of infection. both igm and igg identify the status of infection present or past respectively. a quality control colour band will also be produced at its respective position on the strip to confirm that the test procedure is working accurately.11 the immunoassay technique reagents are available both for enzyme linked immunosorbent assay (elisa) procedure in liquid phase and lateral flow immunoassay (lfia) for immunochromatographic method. bulk reagents, viz. antibody, active site fragments of viral protein antigen, enzyme linked tracer for elisa are available besides the availability of nanoparticle linked, fluorescent probes etc. the immunoassay kits for the detection of viral antigen and antibody are available internationally.12,13 as described above several countries, including china, south korea, uk, europe, and the usa are in process of producing immunoassay kits for covid-19 antigen/antibody detection. the main consideration in hand in western countries is the validation of the assay which is essential for quality control. it is anticipated that assays validation will be achieved in a few weeks’ time if not days. once it is achieved, there is no reason why immunoassay technique will not become a front-line technique and priority of choice to be applied for mass scale screening and for confirmation. a comparison of the two techniques, rt-qpcr and the immunoassay techniques are clearly indicative that immunoassay procedure is less expensive, easy to perform and results are available within 10-20 minutes14. conclusion it is concluded, that pakistan should adapt and validate the immunoassay procedures. this would be first step forward towards normalization after lockdown and quarantine situation. it will be helpful 3 an approach to screen covid-19 patients using diagnostic kits: with special reference to pakistan to bring the healthy recovered persons back to work, hence normalize routines14 furthermore, pakistan has the capability and expertise to produce immunoassay-based kits as previously developed by inmol lahore (paec) for hormones and tumour marker assays.15,16 pakistan should invest to prepare the antibody-based immunoassay kit because of its high specificity, low in cost and easy to use methodology. references 1. ”corona virus resource center” coronavirus.jhu.edu, viewed on 8th april 2020, <https://coronavirus.jhu.edu/> 2. ashfaq, a “corona virus: more than 50,000 covid-19 patients expected in pakistan by end of april”, gulfnews.com, 5th april 2020 <https://gulfnews.com/world/asia/pakistan/coro navirus-more-than-50000-covid-19-patientsexpected-in-pakistan-by-end-of-april1.70815880> 3. xu x, chen p, wang j, et al. 2020 “evolution of the novel corona virus from the ongoing wuhan outbreak and modelling of its spike protein for risk of human transmission” science china life sciences; 2020;63(3):457-60 4. vankadari n, wilce j, 2020 “emerging wuhan (covid-19) corona virus: glycan shield and structure prediction of spike glycoprotein and its interaction with human cd26” emerging microbes & infections, 2020 9:1, 601-604. 5. lu h, stratton c, tang y, 2020 “outbreak of pneumonia of unknown etiology in wuhan china: themystery and the miracle” journal of medical virology 2020; 92(4):401-2. 6. huang c, wang y, li x, et al. 2020 “clinical features of patients infected with 2019 novel corona virus in wuhan, china” lancet 2020; 395(10223):497-506. 7. xiaolong cai. 2020. “an insight of comparison between covid-19 (2019-ncov disease) and sars in pathology and pathogenesis”. 10.31219/osf.io/hw34x. 8. broughton j, deng x, yu g, fasching c, singh et al. 2020. “rapid detection of 2019 novel corona virus sars-cov-2 using a crisprbased detectr lateral flow assay” https://doi.org/10.1101/2020.03.06.20032334 9. souf s, 2016 “recent advances in diagnostic testing for viral infections”, bioscience horizons: the international journal of student research, volume 9, 2016, hzw010, https://doi.org/10.1093/biohorizons/hzw010 10. “corona virus disease (covid-19) pandemic”, who.int , viewed on 1-8 april 2020, <https://www.who.int/emergencies/diseases/nov el-coronavirus-2019> 11. li z, yiy, luo x, xiong n, liu yet al. 2020. “development and clinical application of a rapid igm-igg combined antibody test for sars-cov-2 infection diagnosis” journal of medical virology. 10.1002/jmv.25727. 12. “corona virus elisa kits/biocompare”, viewed on 1-8 april 2020 <www.biocompare.com> 13. “corona virus covid-19 igm elisa assay kit-eagle bioscience” viewed on 1-8 april 2020 <www.eaglebio.com> 14. “expert reactions to corona virus”, sciencemediacentre.org, viewed on 1-8 april 2020<https://www.sciencemediacentre.org/tag/c ovid-19/> 15. zaheer, f. 1992 “behaviour of 1-125 labelled monoclonal anti tsh, and cellulose linked (solid phase) antibody in a supersensitive tsh irma “iaea-sm-324/69: 73-81. proc. on dev. ria and related procedures vienna austria. 16. zaheer, f. 1992 “new perspectives for radioimmunoassay in developing countries “iaea-sm-324/71: 579-583. proc. on dev. ria and related procedures vienna austria. 17. durrani fg, gul r, sadaf s, akhtar mw.2015 “expression and rapid purification of recombinant biologically active ovine growth hormone with dsba targeting to escherichia coli inner membrane”. applied microbiology and biotechnology. 99(16):6791-6801. doi: 10.1007/s00253-015-6751-6. the authors: dr. faiza gul durrani consultant, science shines ltd., london. dr. farhat zaheer retired chief scientific officer (cso), institute of nuclear medicine and oncology lahore (inmolpaec). corresponding author: dr. faiza gul durrani consultant, science shines ltd., london. e-mail: faizaguldurrani@gmail.com 4 proceedings s.z.m.c. vol: 34(3): pp. 4-12, 2020. pszmc-754-34-3-2020 use of point-of-care cholesterol testing in population based non-communicable disease surveillance: caveats and challenges 1ruhina akbar, 2khadija irfan khawaja, 2sara mahmood, 3ian y. goon, 3john campbell chambers, 1saman sarwar, 1ayesha shahid, 4mahina iftikhar baloch 1department of chemical pathology, services institute of medical sciences, lahore 2department of endocrinology &metabolism, services institute of medical sciences, lahore 3department of epidemiology and biostatistics, school of public health, imperial college, london 4department of hematology, jinnah hospital, lahore abstract introduction: point of care testing (poct) for total cholesterol (tc) is invaluable in non-communicable disease (ncd) surveillance programs, as it may permit rapid risk stratification for efficient channeling of limited finances in resource constrained settings. nevertheless, one needs to be aware of some caveats to the dependability of poct results for tc in high load situations. aims & objectives: to evaluate the analytical performance of poct for tc in a population-based ncd surveillance study, by comparing its results with a laboratory assay, and to identify sources of error. place and duration of study: mangamandi, lahore (sampling); services institute of medical sciences, lahore (laboratory), from december 2019 to march 2020. material & methods: poct for tc was done as part of cvd risk stratification in a large ncd surveillance project. lower than expected readings of tc on poct were flagged during routine data quality checking, and this prospective study was designed to determine accuracy of poct readings by testing the same sample in a laboratory. mean ±sd of two methods were compared in overall sample and in subgroups. linear regression analysis was done to determine correlation between the two methods. after a significant disparity was confirmed, poct process was scrutinized to identify its cause, and re-testing after its correction confirmed the source of interference. results: mean tc level in overall sample (n= 699) by poct was significantly lower than that of laboratory method: 2.80 (±0.30 sd) mmol/l vs. 5.28 (±1.27 sd) mmol/l (p <0.0001) r2 0.085. this trend persisted in subgroup analysis. a significant difference between the two methods was seen in a bland altman plot. poct process evaluation identified optical window interference as a possible cause of the discrepancy, and after this was corrected, poct results started showing a higher trend and became comparable with laboratory: 4.67 (±1.50 sd) vs. 5.45 (±1.89 sd) mmol/l, r2 0.9157.conclusion: even though the utility of poct for cvd risk stratification in ncd surveillance programmes is undeniable, some caveats and challenges remain. non-compliance with device maintenance protocols in high throughput situations encountered in field testing may contribute to inaccurate results. cholesterol poct requires careful operator training, technical support and strong quality assurance backup. key words: point-of-care cholesterol testing, optical window interference, non-communicable disease screening introduction the 21st century has witnessed a paradigm shift in the global burden of disease from communicable to non-communicable diseases (ncd). according to the 2017 global burden disease (gbd) study of who, 73.4 % of all deaths in 2017 were due to ncds; an increase of almost 22.7% over the preceding decade.1recognizing this fact, who has included ncd control in its sustainable development goals for 2030.2a substantial number of these deaths (17.8 million in the 2017 survey) occur due to premature chd in lower middle income countries (lmic) like pakistan, where ncd programmes are still in infancy.1,3 in resource constrained settings, it is cost-effective to focus on primary prevention, using a two-pronged approach of disease surveillance and population based low cost interventions like the who package of 5 use of point-of-care cholesterol testing in population based non-communicable disease surveillance essential non communicable disease interventions (who pen), delivered though frontline workers.4 the global health research unit (ghru) on diabetes and cardiovascular disease in south asia is an international collaborative project between uk and four south asian countries including pakistan, funded by national institute for health research (nihr), uk. details of the collaboration and current projects can be found on the project website: (https://fundingawards.nihr.ac.uk/award/16/136/68). under the umbrella of this project, a ncd surveillance study was started in the punjab province in pakistan in 2019, with the aim of ascertaining the true prevalence of ncd in the province. blood cholesterol level, as an important marker of chd, was among the various parameters being recorded in this population based surveillance study. point of care testing (poct) for total cholesterol was done to help in risk stratification using cvd risk prediction tools, for inclusion in the participant health assessment report. based upon risk stratification, participants could then be channeled towards ensuing ncd control projects. while poct for estimation of blood glucose and glycosylated hemoglobin has been a part of clinical management of diabetes for a long time,5 poct for cholesterol in whole blood has become widely available relatively recently.6,7,8 the advantage of including poct for tc in ncd surveillance programmes lies in that it may be used for early detection of elevated cholesterol allowing risk stratification and timely institution of risk mitigation measures like statin therapy.6 however, this is a relatively new technology, and while it offers advantage of speed and convenience, the caveat lies in the fact that in a population-based screening setting, poct has a high usage load and is mainly performed by front-line workers with little technical background, who are usually not well-versed in the technical details of the device and testing process.9 this may be an even greater challenge in low resource countries like pakistan where availability of technical and quality control support is minimal particularly in remote locations. indeed, concerns have been raised about reliability of poct results compared to the conventional laboratory testing, which may be ascribable to operator dependent issues rather than a limitation of poct per se, as the reliable operation of the device requires an understanding of the principle of colorimetric detection, and ability to service the device optical window regularly. proper operator training, and continuous quality control (qc) checks are a mandatory requirement for dependable poct testing in high throughput settings.10 in the ghru surveillance project, poct for tc was performed for immediate chd risk assessment, while blood was collected for complete biochemistry as a batch in a central lab at a later date. to select a suitable tc poct device for use in the project, a range of available devices were compared on the basis of reliability of results, live data capture and stability of performance in high ambient temperatures likely to be encountered in field testing in south asian countries including pakistan. the aina poct device (jana care inc, usa) was selected by the central steering committee, as it fulfilled above mentioned criteria.11 a pilot study was done to compare the device results with laboratory findings, and showed comparable results (unpublished data, available on request). frontline community health workers were trained in aina device usage as per documented standard operating protocols of the surveillance project, and a detailed operations manual was made available to each of them. for quality assurance, the central project team was running continuous quality control (qc) checks on the collected data of the aina poct device. the device continued to perform well for several months into the surveillance project, however, after performance of almost 2200 tests on the aina device, routine qc checks identified that measured tc levels were showing a consistent drift towards low values. the rationale of present study was to identify practical issues in the implementation of a new smart phone linked cholesterol poct device as a cost effective tool for population based surveillance studies in limited resource settings like pakistan. tc results of poct were evaluated by comparing its analytical performance with tc measured in a clinical laboratory, in the backdrop of lower than expected tc levels, in the first instance, to confirm this finding, and if confirmed, to critically evaluate the poct process to detect the source of any disparity, institute remedial measures, followed by re-testing to verify improvement in performance. the objective of this exercise was to identify the challenges and to suggest practical ways to overcome these difficulties. material and methods as mentioned previously, the present study was conducted on a subset of the participants in the ongoing ghru surveillance study which aims to screen 150000 adults for ncd in four south asian 6 use of point-of-care cholesterol testing in population based non-communicable disease surveillance countries, out of which 30000 would be from pakistan. the project has been approved by national bioethics committee (ref: no.4-87/nbc347/19/1506 dated 01.31.2019) and the hospital institutional review board (ref: irb/2018/ 461/sims dated 09.24.2018). equipment used in the study included aina poct lipid device (jana care inc, usa) for cholesterol poct and cobas c311 analyzer (roche diagnostics gmbh, germany) for laboratory assays as a standard for comparison. the aina poct device has a reported clinical accuracy of 100% samples within 20% bias and a good correlation (r2= 0.973) with dimension rxl max analyzer (siemens, usa) and a measuring range of 2.59 to 10.34 mmol/l for tc working at 10 to 40o c with test time of 2 minutes. in addition, it has an advanced feature of cloud readiness i.e. safe transfer of data to central databases.11 cobas c311 is an automatic clinical laboratory analyzer which was based in an iso 15189 accredited laboratory with internal and external quality controls. participants were enrolled and given a translated information sheet one day before the surveillance activity, and fasting venous samples were collected next day after their written informed consent. while serum was stored for a wide range of biochemical tests to be run as a batch at a later date, poct for blood glucose and tc were done to be included in the participant health assessment report. in december 2019, the continuous qc checks identified that the poc cholesterol results had drifted well below expectations over a four week period immediately preceding this study (october, 2019: range 2.59-5.67 mmol/l, mean 3.23 (±0.54 sd) mmol/l vs november/december, 2019: range 1.71-4.68 mmol/l, mean 2.92 (±0.35 sd) mmol/l. to determine if this inconsistency was a chance occurrence or was due to a malfunction of the poct, a cross-sectional, prospective study was conducted to compare the poct results with laboratory analysis results. sampling was done in the area of mangamandi, in suburbs of lahore, over a four week period from mid-december 2019 to january 2020. poct was performed in a mobile health unit (mhu) specially designed for ncds surveillance, with a mini-laboratory set up for poc testing, sample processing and storage. the average ambient daytime temperature during this time ranged between 16.1°c-19.5°c with 75% humidity.12,13 a total of 699 participants were included in the study, with 455 females (65.1%) and 244 males (34.9%). venous blood sample, of participants fasting for 8-14 hours, was collected from a single venipuncture into 2 ml edta and 3.5 ml serum gel tubes through multi-sample needles. for poct, 15 l whole blood was pipetted onto the cholesterol test strip of aina device. for laboratory testing, serum was separated within 4 hours, and was evaluated in clinical laboratory of services institute of medical sciences, lahore. tc was measured by an enzymatic colorimetric method using cobas c311 analyzer (roche diagnostics gmbh, germany). the results of the two methods were compared overall, as well as in subgroups based on gender, age and laboratory cholesterol level. further course of action was to be determined by the results of the first phase. it was planned that if on initial testing, poct cholesterol readings differed significantly from the laboratory results, poct process would be evaluated to identify the source of the error, and testing would be repeated after the fault had been eliminated, to confirm that this was the cause of the anomaly. statistical analysis: data analysis was performed using microsoft excel (2013). mean ± sd of two methods was obtained, and % bias calculated. bland-altman plot was used to assess agreement between two methods. statistical significance calculated by student’s t-test, was defined at p< 0.05. linear regression analysis was done to determine the existence of correlation between two methods. results 1. initial comparison of poct cholesterol results with laboratory testing: in the overall population, mean (± sd) tc by laboratory method was 5.28 (±1.27) mmol/l and by poct device method it was 2.81 (±0.30) mmol/l. overall bias and % bias was -2.47 and -46.78 respectively while r2 was 0.085. bland altman plot showed significant negative bias (fig-1). fig-1: bland altman plot for the difference between the poct device and the laboratory for tc -12 -10 -8 -6 -4 -2 0 2 4 6 0 1 2 3 4 5 6 7 8 9 10 d iff er en ce b et w ee n d ev ic e tc a nd la b tc m et ho d (m m ol /l ) mean (in mmol/l) of tc by lab method and by poct device method +2sd (agreement limit) mean -2sd (agreement limit) 7 use of point-of-care cholesterol testing in population based non-communicable disease surveillance on gender specific analysis, tc mean (±sd) in females was 5.32 (±1.23) mmol/l, and in males was 5.19 (±1.33)mmol/l by laboratory method, while by poct, it was 2.83 (±0.31) mmol/l in females and 2.74 (±0.28) mmol/l in males. the % bias, p-value and correlation between poct and laboratory method based on gender is given in (table-1) and (fig-2) sex n % % bias p-value female 455 65.1 0.00103 <0.0001 male 244 34.9 0.00194 <0.0001 note. n=number of participants table-1: analysis of groups by gender fig-2: correlation between device method and laboratory method according to gender analysis of subgroups based on age for tc by poct method vs cholesterol by laboratory method can be seen in table-2 and fig-3. age group (years) n total cholesterol device (mmol/l) mean ±sd total cholesterol by automation (mmol/l) mean ±sd % bias p-value i. 18-35 202 2.77±0.28 4.86±1.24 0.00213 <0.0001 ii. 36-59 463 2.82±0.31 5.45± 1.23 0.00104 <0.0001 iii. >60 32 2.76±0.32 5.39± 1.51 0.01525 <0.0001 note. n=number of participants. sd=standard deviation table-2: total cholesterol by device versus laboratory estimation according to age groups fig-3:correlation between device method and laboratory method according to age table-3 and fig-4 details analysis of tc measured by poct and laboratory method based on cholesterol levels by laboratory method. cholesterol level (mmol/l) n tc device (mmol/l) mean±sd tc by automation (mmol/l) mean±sd % bias p-value a) < 5.147 333 2.73±0.27 4.28±0.69 0.00109 <0.0001 b) 5.1476.18 226 2.79±0.25 5.63±0.30 0.00222 <0.0001 c) ≥6.19 138 2.98±0.30 7.09±0.94 0.00421 <0.0001 grand total 699 2.80±0.30 5.28±1.27 note. n=number of participants. tc=total cholesterol. sd=standard deviation. table-3: comparison of device versus laboratory estimation according to cholesterol level fig-4: correlation between device method and laboratory method according to cholesterol level 2. identification of the source of error as the anomaly identified on qc was confirmed by the result of the first phase of testing, a biomedical engineer scrutinized the poct process and found that the sample collection and processing operating r² = 0.0858 r² = 0.0605 r² = 0.1408 0 50 100 150 200 0 100 200 300 400 500 total female male linear (total) linear (female) linear (male) r² = 0.1123 r² = 0.0712 r² = 0.0625 0 50 100 150 200 250 0 200 400 600 group i: 18-35yrs group ii: 36-59yrs group iii: ≥60yrs linear (group i: 18-35yrs) linear (group ii: 36-59yrs) linear (group iii: ≥60yrs) 8 use of point-of-care cholesterol testing in population based non-communicable disease surveillance protocols were being followed correctly. however, it was identified that while the device was cleaned externally with isopropyl alcohol on a daily basis, as per protocol, the optical window was not being serviced as this needed to be exposed by opening the device body. once the optical window was exposed by opening the device body (fig-5), it was cleaned with 70% isopropyl alcohol, and device performance was retested after cleaning. fig-5: optical window exposed after sliding strip adaptor from the device body retesting after optical window servicing: after the device optical window was cleaned, a series of duplicate tests were run on randomly selected samples over a three week period (february-march 2020). it was immediately apparent that the range of poct results had increased beyond the low values seen in the preceding period (range: 2.59-7.42 mmol/l, mean 3.99 (±0.81 sd) mmol/l. the results of the poct compared to the laboratory testing are given in (table-4) and (fig-6). this confirmed that the device performance issue had been due to optical window interference. cholesterol level n device method (mmol/l) mean lab method (mmol/l) mean correlation r2 pvalue a) < 5.147 11 3.26± 0.80 3.72±0.6 0.785 0.61632 0.1429 b) 5.147 6.18 2 5.21± 0.55 5.59±0.5 5 1 1 0.3845 c) ≥ 6.19 10 5.96± 0.68 7.33±0.8 6 0.904 0.8173 0.00098 overall 23 4.67±1.5 5.45±1.89 0.9569 0.9157 0.099 note. n =number of participants. sd=standard deviation. r=coefficient of determination. table-4: correlation between device method and laboratory method according to cholesterol level after optical window cleaning. (overall r2=0.9157) fig-6: correlation between device method and lab method according to cholesterol level after optical window cleaning (overall r2=0.9157) 3. retraining and remediation: as remedial measures, the operating technicians were retrained in the device maintenance procedure, and the operations manual was expanded to include a section on routine device maintenance, emphasizing the technique of opening and cleaning the optical window. discussion knowledge of the patient’s cholesterol levels is invaluable in risk stratification for focusing preventive measures and directing risk mitigation programmes like the who pen intervention, deliverable by trained frontline workers. poct for tc offers an attractive solution, whereby patients can be triaged and assigned to a particular intervention in a single encounter, minimizing loss to follow-up. however, in order to be a useful part of clinical assessment, poct systems should yield results which are accurate and comparable with laboratory analysis.14 cholesterol poct technology became generally available around the year 2000, but devices were initially large and cumbersome. over time, devices have become more portable and compact and capable of integration with data servers.15 the aina poct device offers the advantage of both being very compact and directly pluggable into a smartphone for live data capture by the central server11 the present study was the first time it had been used in population based screening in pakistan. the device showed accurate results compared with laboratory assays in the pilot study, and in the first three months of the surveillance project, as confirmed by regular quality assurance checks. however, in the period immediately preceding this study, lower than expected readings had been 9 use of point-of-care cholesterol testing in population based non-communicable disease surveillance flagged by these checks. during the initial phase of this study, meant to confirm the low readings, it was seen that the poct cholesterol results were significantly lower than those from the laboratory cholesterol assay (p <0.001), with very low correlation between the two (r2= 0.085). this was of great concern to us, as it was likely to have an impact on the validity of the health report given to the participants. even more worrying was the fact that this was at odds not only with the pilot study, but also the device performance in the initial three months. the second part of the study focused on a step by step analysis of the testing process to pinpoint the source of the disparity, which identified the device optical window to be responsible, and this was confirmed when poct result correlation with laboratory assay increased significantly after its correction (r2= 0.916). as the availability of poct for cholesterol is a relatively new development, there is a paucity of external validation studies, especially for the newest generation of smartphone compatible devices. some of these studies have raised concerns about lack of accuracy of cholesterol poct devices, with a bias towards an underestimation of tc.16 similar trend was quantified in the initial phase of our study; the negative bias of -2.46 showed that poc device values were less than those of the gold standard laboratory method indicating accuracy issues, however the bias was eliminated after the removal of the source of interference. the difference in the overall sample mean of tc by the two methods in our study was almost 2.47 mmol/l (95.5mg/dl). this difference was plotted in the bland altman plot (fig-1) showing several readings lying outside + 2sd limits of agreement which seemed to indicate significant measurement bias between the two methods.17,18 in fact such a difference had been previously reported by park et al in a poct tc validation study with -15.9% bias14 whereas standard for accuracy set by ncep guidelines is ≤ ± 5%.19 in another study of 111 cases, xavier et al reported moderate correlation between poct device and clinical laboratory for tc values (r2 = 0.796).20 whitehead et al, reported that the poct analyzer showed a negative bias for tc of 17.6±13.4% when compared to the laboratory method.21 in a study by matteucci et al. poct also underestimated tc (bias 6.5%).22 other studies have, however, shown satisfactory correlation between poct and laboratory values of tc. ferreira et al, reported good correlation between poct and laboratory method for tc: (r2 0.879, average bias 4.0 %) in a study on 516 participants.23 indeed, the differences in the experiences of different groups of researchers suggest that something beyond a limitation of the device technology is involved here. clearly, it is a matter for concern that, while the device performance in device validation studies usually correlates highly with laboratory data, when testing in field conditions, a significant difference between cholesterol results on poc testing and laboratory readings has been noted by many authors.8,20,21 our study highlights the fact that falsely low cholesterol readings may be due to operational factors, reflecting the difficulty in running simple but sensitive technology in field conditions, where the operators are often nontechnical personnel,21,24 and technical support may not be immediately available. such user dependent challenges were identified by o’kane and colleagues as a major source of quality errors in poct.25 furthermore, it highlights the need for rigorous quality control measures, to detect any deviation from expected trends.8,9,22,23 extrapolating from our results, we can speculate that the accuracy issues observed in other poc cholesterol devices could be attributed to a similar issue. inaccuracy in poct results can be an operator dependant issue due to a failure to follow device maintenance protocols, rather than a limitation of device per se.22,23 this was also noted by whitehead et al in the field setting of the outreach nhs health screening clinics in england.19 it is important to raise this issue because in actual field testing, with a large number of samples being handled in circumstances, these best practices are frequently ignored.10,19,23 although cholesterol poct devices are simple to use, they utilize a sensitive technology that requires careful and regular device maintenance by trained operators. these devices, including the one used in our study, are based on optical bio-sensing technique, which uses enzyme catalyzed color reaction24 for cholesterol identification and reflectance photometry for changing the chemical signal into an optical signal.25 cholesterol concentration is then quantified through photometric detection.28 the reaction area, where test strip/cassette is inserted, is a removable part of the device and lies directly above the optical system which is a non-removable intrinsic part of the device body. the optical window is made from optical material with specific qualities tailored for reflected light transmission into the optical system.29,30,31 changes in the optical properties of this window due to contamination (dust, dirt, blood etc) or abrasion can cause distortions in reflected light or interference with its transmission leading to erroneous results.32 the optical window needs to be 10 use of point-of-care cholesterol testing in population based non-communicable disease surveillance exposed by opening the device body, to be cleaned regularly. although this is not a complicated step per se, it may be omitted by the field operators who are usually not from a technical background, and may be unable to understand the technical requirements of the system.21,25,28 operator factors were recently reported as a reason of resistance towards poct system acceptance in workflow by a large primary care cvd risk assessment program in new zealand and it was suggested that continuous training and support could help in achieving the recognized benefits of poct.10 indeed, technical support may not always be available in field screening, and such remediable sources of error may go unnoticed. this highlights the importance of careful training and re-training of field operators, and indeed their supervisors, who may themselves be unaware of the technical requirements of the system.24,25 furthermore, the importance of quality control checks cannot be overemphasized, as these are able to flag potential sources of error, and indicate the need for remedial measures.24,25,28 in our study, we were fortunate to have both technical support and a rigorous quality assurance system in place, because of which we were able to quickly identify and correct the issue. in low resource countries like pakistan, in particular where cholesterol poct is being used in population screening in remote locations, this issue may become very relevant. it is noteworthy that interference due to dirty window can be an issue in all optical biosensors, including those in glucose poct devices. however, in contrast to cholesterol poct devices, the newer generation glucose poct devices have resolved this issue by using electrochemical bio-sensing.33 technology for cholesterol poct devices is in emerging state, with optical biosensors being the most cost effective solution for the time being.34 if cholesterol poct is to be successfully incorporated into population based screening programs, devices which are less dependent on technical maintenance need to be developed. furthermore, thorough operator training, robust technical support as well as rigorous quality assurance with periodic evaluation of results against laboratory cholesterol assays are essential to maintain validity of results. conclusion in conclusion, operator training in technical aspects of poct device maintenance is an essential part of the internal quality control (iqc) protocols. it is important to highlight this issue because in field testing for population screening, where a large number of samples are handled in less than ideal circumstances, these best practices may not be followed due to inadequate operator training or the high workload. clearly, while the ease of use and speed of results for poct is undeniable, unless these devices are operated with careful adherence to operating and maintenance protocols, with appropriate technical support, device accuracy may become compromised. limitations: as this was not a formal validation study, it was not possible to control for every factor that might have an impact on the results. the study was designed in the context of an ongoing surveillance project with narrow focus on the study’s own poct accuracy concerns raised by routine qc. however, we wanted to share our experience so that public health researchers especially in low resource countries may be aware of difficulties in poct device usage in high load settings and can take suitable measures to avoid these for a reliable and cost effective data output. conflict of interest: the authors have no conflict of interest to declare. acknowledgement: this study is funded by the uk national institute for health research (nihr) [global health research unit (award id 16/168/68) / department of health and social care] https://fundingawards.nihr.ac.uk/award/16/136/68 references 1. roth ga, abate d, abate kh, abay sm, abbafati c, abbasi n, et al. global, regional, and national age-sex-specific mortality for 282 causes of death in 195 countries and territories, 1980-2017: a systematic analysis for the global burden of disease study 2017. lancet. 2018; 392(10159):1736-88. 2. united nations o. transforming our world: the 2030 agenda for sustainable development: united nations; 2015 [12-07-20]. 3. bennett je, stevens ga, mathers cd, bonita r, rehm j, kruk me, et al. ncd countdown 2030: worldwide trends in non-communicable disease mortality and progress towards sustainable development goal target 3.4. lancet. 2018; 392(10152):1072-88. 4. world health o. package of essential non 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[accessed 29 june 2020]. available at: <https://www.timeanddate.com/weather/pakista n/lahore/historic?month=1&year=2020 > 14. park p h, chege p, hagedorn i c, kwena a, bloomfield g s, pastakia s d. assessing the accuracy of point-of-care analyzer for hyperlipidemia in western kenya. trop med int health. 2016; 21(3):437-44 15. plüddemann a, thompson m, price cp, wolstenholme j, heneghan c. point-of-care testing for the analysis of lipid panels: primary care diagnostic technology update. br j gen pract. 2012; 62(596):e224-e226. 16. haggerty l, tran d. cholesterol point-of-care testing for community pharmacies: a review of the current literature. j pharm pract. 2017; 30(4):451-458. 17. hanneman sk. design, analysis and interpretation of method-comparison studies. aacn adv crit care [internet]. 2008 apr–jun; 19(2):223–234 doi: 10.1097/01.aacn.0000318125.41512.a3 18. bland jm, altman dg. agreement between methods of measurement with multiple observations per individual. j biopharm stat. 2007;17(4):571-582, doi: 10.1080/10543400701329422 19. national cholesterol education program. recommendations on lipoprotein measurement. u.s. department of health and human services: washington, dc, 1995. 20. xavier h t, ruiz r m, kencis l, melone g, costa w, fraga rf et al. clinical correlation between the point-of-care testing method and the traditional clinical laboratory diagnosis in the measure of the lipid profile in patients seen in medical offices. j bras patol med lab 2016; 52(6): 387-390. 21. whitehead sj, ford c, gama r. a combined laboratory and field evaluation of cholestech ldx and cardio chek pa point-of-care testing lipid and glucose analyzers. ann clin biochem 2014; 51(pt 1):54-67. 22. matteucci e, bartola dl, rossi l, pellegrini g, giampietro o. improving cardio check pa analytical performance: three-year study. clin chem lab med 2014; 52(9): 1291-6. 23. ferrira ces, franca cn, correr cj, zucker ml. clinical correlation between a point-of-care testing system and laboratory automation for lipid profile.clinicachimicaacta.2015;446: 263-6 24. warade jp. challenges in poct. medico research chronicles. 2014; 1(1):49-55. 25. o'kane mj, mcmanus p, mcgowan n, lynch pm. quality error rates in point-of-care testing. clinical chemistry.2011;57(9):1267-71 26. li hl, dutkiewicz ep, huang yc, zhou hb, hsu cc. analytical methods for cholesterol quantification. j food drug analy. 2019; 27(2): 375-386. 27. bhalla n, jolly p, formisano n, estrela p. introduction to biosensors. essays biochem. 2016; 60(1):1-8. 28. rifai n, horvath ar, wittwer ct. tietz fundamentals of clinical chemistry and molecular diagnostics 8 e; south asia edition; e-book [internet]. google books. 2019 [cited 29 june 2020]. 12 use of point-of-care cholesterol testing in population based non-communicable disease surveillance 29. polymer technology systems, inc. cardio check pa system: user guide. [internet] 2018. indianapolis, usa. 30. alere group of comapnies. cholestech ldx: system user manual. [internet] 2013. san diego, usa. 31. roche diagnostics. accutrends plus: user’s manual. [internet]. 2012. indianapolis, usa. 32. rajkumar, singh r, pandya kb, kumar a. effects of pressure gradients on laser beam propagation through an optical window for tokamak plasma diagnostics. fus scien tech. 2012. [61. 51. 10.13182/fst12-a13338] 33. pezzuto f, scarano a, marini c, rossi g, stocchi r, cerbo ad et al. assessing reliability of commercially available point of care in various clinical fields. open public health j 2019. [12. 10.2174/1874944501912010342]. 34. wang x, hu l. review-enzymatic strips for detection of serum total cholesterol with point-of-care testing (poct) devices: current status and future prospect. j electro soci. 2020 [10.1149/1945-7111/ab64bb]. the authors: dr. ruhina akbar head, department of chemical pathology, services institute of medical sciences, lahore. dr. khadija irfan khawaja head, dept. of endocrinology & metabolism, services institute of medical sciences, lahore. dr. sara mahmood research project manager, imperial college london research projects/ department of endocrinology & metabolism, services institute of medical sciences, lahore. dr. ian y. goon department of epidemiology and biostatistics, school of public health, imperial college, london. prof. john campbell chambers department of epidemiology and biostatistics, school of public health, imperial college, london. dr. saman sarwar pg trainee, department of pathology, services institute of medical sciences, lahore. dr. ayesha shahid demonstrator, department of pathology, services institute of medical sciences, lahore. dr. mahina iftikhar baloch pg trainee, department of hematology, allama iqbal medical college, lahore corresponding author: dr. ruhina akbar head, department of chemical pathology, services institute of medical sciences, lahore. e-mail: ruhina_akbar@yahoo.com for web full book 44 proceedings s.z.m.c. vol: 37(3): pp. 44-49, 2023. pszmc-896-37-3-2023 comparison of learning styles used by clinical faculty of hospital and general practitioners for their professional development muhammad atique, department of pathology, pakistan kidney & liver institute & research centre, lahore. abstract aims & objectives: to compare the learning styles used by hospital clinical faculty and general practitioners for their professional development/ continued medical education. place and duration of study: this was a comparative cross-sectional study carried out at pakistan kidney and liver institute and research canter lahore from october 2019 to november 2020. material & methods: total number of seventy-six medical professionals comprising thirty-eight members of clinical faculty and similar number of general practitioners were included in the study. amongst these, 45 were males and 31 females’ learning style questionnaire adopted from honey and mumford was distributed to all the participants. according to the learning styles they were grouped into activists, reflectors, theorists and pragmatists with preference categorization of very strong, strong, moderate, low and very low. data was entered and analyzed usin spss version 22 chi-square test was applied to see the significant difference in two groups and p value was calculated and value of less than 0.05 was considered significant. results: there were 45 males and 31 females with ratio of 1.45:1 .in general practitioners the strongest learning style was reflector, followed by theorist, activist and pragmatists.while in the clinical faculty the strongest preference was again for reflectors, followed by activists then theorists and finally pragmatists, the significant difference statistically was only seen in the moderate preference in activist group which was 0.038. conclusion: reflector type of learning style based on pondering, experiencing and observing different perspectives was strongly observed in both genders of consultants and general practitioners. however, a moderate degree of activist type of learning style influenced by doing and feeling was also noted in the general practitioners. further planning instructional strategy and assessment based on these learning styles could benefit the career growth of these two groups of health professionals. keywords: learning style, general practitioners, clinical faculty, health professionals introduction human beings have similar biopsychological and social characteristics however their ability to grasp information and transferring it into logical meaning is different. this difference is termed learning style1. it is also defined as the characteristic cognitive, affective, and psychological behaviors that serve as relatively stable indicators of how learners perceive, interact with, an d respond to the learning environment2. it is also defined as a way of thinking comprehending and processing information3 and to kolb it is a method of personal choice to perceive or process information4. curry in 1987 proposed that overall learning depends on three factors and the model was stated as onion model. the central most part was personality variable, the middle one was of learning style and the outer most layer of onion depended upon instructional format preference5.there are about thirteen major ways that learning styles are classified6. however, the commonest ones are of four types one was by flemings who classified as vak-visual 29%, auditory34% and kinesthetic 37%7. this style has been used to assess the achievements of medical professionals8.the second one was given by kolb who classified as (accommodators), divergers, assimilators and convergers)8. honey and mumford classified as activists, reflectors, theorists and pragmatists. this classification was merged with that form kolb as activists (accommodators), reflectors (divergers), theorists (assimilator) and pragmatists (convergers)9. the honey and mumford learning styles 9are plotted around two axes (fig 1). the horizontal axis is called the processing continuum and refers to how we approach a task, that is, preference to learn by watching or doing. whereas the vertical axis is termed the perception continuum and refers to our emotional response to the task, or how we think and feel about it. reflectors learn best when learning activity provides them opportunity to think and reflect. they want to understand things thoroughly before they try them 6 1department of public health, health services academy, islamabad. 2department of management sciences, riphah int university, islamabad. 3department of medicine, ihitc, islamabad. 4department of rheumatology, pims, islamabad. 5department of medicine, dow university of health sciences, karachi. 70 1department of orthopedics, thq, khanewal 2department of orthopedics, lgh, lahore 3department of orthopedics, thq, ferozwala 4department of orthopedics, cmh, lahore 5department of hand & upper limb surgery, cmh, lahore 6department of orthopedics, al-rehmat benevolent trust 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 45 comparison of learning styles used by clinical faculty of hospital and general practitioners for….. out. the activist learns in an environment of new, varied and continued activity. they don’t want to hear what they should be doing; they want to dive in head-first and have a go. fig-1: honey and mumford learning style. the theorist learns best when they have time to analyze and synthesize. theorists like to understand how the new learning fits into their ‘framework’ and into previous theories. they’re likely to be uncomfortable with things that don’t fit with what they already know. the pragmatist learns through practical and concrete issues of learning. pragmatists care about what works in the real world. they aren’t interested in abstract concepts; they just want to know if it works10.many individuals may not use single learning style and work in a continuum using more than one type11. a learner must understand how to identify their learning goals integrate learning style apply a properskill and be self-regulated to achieve the best results from learning12. multiple studies examining the learning styles of health professionals were noted during literature search. some of the studies were comparative, whereby the comparison was done amongst various groups of health professionals a) specialists of different medical fields i.e surgeons and medical specialist 13. b) residents and faculty members14, c) undergraduate medical students with postgraduate medical students15, medical students from various cities i.e. islamabad, gujranwala and from a particular university16,17. the other studies were confined to one type of professionals i.e. pathologists17, nurses and general practitioners. however, in the literature search no study was found which compared learning styles of general practitioners and consultants. the progress in medical profession could be dependent on the learning styles and different medical professionals have particular learning styles19.the clinical faculty and general practitioner follow entirely different approaches towards career. the consultants after doing graduation enter a postgraduate programme, while the general practitioners go into practice usually without getting into a postgraduate degree/diploma program. the rationale of this present study was to see how the learning styles differ/or are similar among these two categories of medical professionals. if the two groups are following a particular learning style and are successful in their careers, then further planning instructional strategy and assessment would be easy and would benefit the growth of these two groups of health professionals. material and methods the study design was comparative cross-sectional. settings and participants: all members of clinical faculty (thirty-eight) of different specialities from pakistan liver and kidney institute and research centre were included in the study excluding those on leave during datacollection. similarly, thirtyeight general practitioners were inducted from various locations of lahore. data collection instrument: quantitative data was collected through self-administered honey and mumford learning style questionnaire containing 80 questions. this was handed over to the participants and collected subsequently. the questionnaire comprised three sections, first section dealt with demographic data, second section -comprised 80 questions and the third section dealt with scoring and categorization. the scores were calculated according to question number which were assigned to a particular learning style and participants were categorized into activists, theorist, pragmatist, and reflectors according to very strong, strong, moderate low and very low preferences. basic demographic data was also collected including age, gender, experience and speciality. completed form in all aspects were included and incomplete /poorly filled forms were excluded from the data. statistical analysis the data was analysed using spss version 22. mode was calculated for the continuous variables like age. frequency and percentage were calculated for the categorical variables like learning styles, gender and speciality. chi square test was used to compare the frequency of different styles among the two categories. the p value of <0.05 was considered significant. 46 comparison of learning styles used by clinical faculty of hospital and general practitioners for….. results there were forty-five males and thirty-one females with ratio of 1.45:1. (fig 2), the age range was between 25 and 75 years with mode of 25-35 years of age (fig, 3). the results were compiled for two groups, clinical faculty and general practitioners according to the learning styles with the preference categorized as very strong, strong, moderate, low and very low. general practitioners: in the general practitioner group, the very strong preferred learning style was reflector (11), followed by theorist (8), pragmatist (7) and activist (4) in the descending order. the females were also in the reflector group in the very strong and strong preferences (table-1). clinical faculty (consultants): in the consultants the most preferred learning style was reflector (8) followed by activist (5), pragmatists (4) and theorists (3) in descending order .the females also followed the same pattern with majority in reflector group (table-2).also seen in the comparative table of learning styles of consultants and general practitioners (table-3). however, the statistical analysis showed the significant p value was obtained in activists groups in moderate preference category (table-4). in this category general practitioners are more than clinical faculty. fig-2: gender distribution of all participants in the study on learning style –n=76 fig-3: age distribution of all participants in the study on learning style –n=76 general practitioners total category gender very strong strong moderat low very low activist m 1(3%) 4(12%) 7(14 %) 4(16 %) 1(6%) 17 f 3(10%) 2(6%) 11(22 %) 4(16 %) 2(12%) 22 reflector m 6(20%) 3(9%) 5(10 %) 2(8 %) 1(6%) 17 f 5(16%) 8(25%) 6(12 %) 2(8 %) 1(6%) 22 theorist m 4(13%) 4(12%) 6(12 %) 1(4 %) 2(12%) 17 f 4(13%) 6(19%) 5(10 %) 4(16 %) 3(18%) 22 pragmatist m 5(16%) 1(3%) 4(8%) 5(18 %) 2(12%) 17 f 2(6%) 3(9%) 7(14 %) 5(18 %) 5(30%) 22 total 30 31 50 27 17 table-1: learning styles with preference category n=38 general practitioners.*+1gp data clinical faculty (consultants) total category gender very strong strong moderate low very low activist m 4(20%) 2(7 %) 7(16 %) 12(3 3%) 4(25%) 29 f 1(5%) 3(11 %) 2(4%) 3(8. 5%) 0 9 reflector m 6(30%) 8(29 %) 9(19 %) 5(13 .5%) 1(6%) 29 f 2(10%) 3(11 %) 2(4%) 1(3 %) 1(6%) 9 theorist m 3(15%) 5(18 %) 14(29 %) 7(19 %) 2(12.5% ) 31 f 0 3(11%) 2(4%) 1(3 %) 1(6%) 7 pragmatist m 4(20%) 3(11 %) 6(12 %) 7(19 %) 6(37.5% ) 26 f 0 1(3%) 6(12 %) 1(3 %) 1(6%) 9 total 20 28 48 37 16 table-2: learning style with preference category n=38clinical faculty (consultants). 47 comparison of learning styles used by clinical faculty of hospital and general practitioners for….. combined general practitioners &clinical faculty (consultants) tota l % categ ory gende r very strong stro ng mode -rate low very low activis t m 5(10%) 6(10%) 14(13 %) 16(2 5%) 5(14%) 46 14 f 4-8% 5-8% 1312% 710% 2-6% 31 10 reflect or m 12-24% 118% 1413% 710% 2-6% 46 14 f 7-14% 11-8% 8-8% 34% 2-6% 31 10 theori st m 7-14% 9-15% 2019% 812% 5-14% 49 15 f 4-8% 9-15% 9-9% 57% 4-11% 31 10 pragmatist m 9-18% 4-6% 1010% 1218% 8-22% 43 13 f 2-4% 4-6% 1312% 69% 6-17% 31 10 total 50 59 101 64 34 308 table-3: combined general practitioners &clinical faculty (consultants) learning style n=76 learning style clinical faculty g.p p-value very strong preference activist 5 4 0.126 reflector 8 11 0.426 theorist 3 8 0.103 pragmatist 4 7 0.328 strong preference activist 5 6 0.744 reflector 11 11 1 theorist 8 10 0.589 pragmatist 4 4 1 moderate preference activist 9 18 0.03(significant) reflector 11 11 1 theorist 18 11 0.098 pragmatist 12 11 0.802 low preference activist 19 11 0.06 reflector 8 6 0.553 theorist 12 10 0.255 pragmatist 15 17 0.642 very low preference activist 4 3 0.307 reflector 2 2 0.867 theorist 4 5 0.872 pragmatist 7 7 0.414 table-4: comparison of learning styles between clinical faculty and general practitionerspvalue n=76. discussion this study compared the learning styles of consultants and general practitioners. these two groups follow different approaches in their carrier progression. the consultants were from clinical faculty of a hospital which in this study was (pkli&rc) lahore. this hospital is a tertiary care hospital mostly dealing with liver/kidney transplant and related surgeries along with their medical management. the general practitioners were selected on the basis that they did not follow any specialized training to be in a clinical faculty. these were recruited from the general practitioner group practicing in various localities of lahore. on literature search, the learning style preference was variable in different groups of consultants. in the earlier studies medical specialists were found to be pragmatist13,14. however, in the later studies on full time consultants the learning style preference was for reflectors14,15, this was true for the postgraduate students also15 a study comparing learning styles of residents and faculty of internal medicine showed that both groups were mostly (theorist)20. in another study comparing learning styles among post graduate residents and full-time faculty showed consultants to be mostly reflector/ theorist14. the learning style also differed among postgraduate and undergraduate medical students. the postgraduate students were reflectors and undergraduate students were mostly activist14.in two other studies medical students were mostly found to be reflectors16,17. among the pathologists the dominating learning style was reflectors followed by theorists18. a study conducted in amman comparing learning style of surgeons to their medical colleagues showed mostly a mixed pattern, with most having combination of two learning styles. eighty to ninety percent of both surgeons and physicians showed combination of reflector and theorist learning styles21. another study on implication of surgical education from alberta school canada showed that most of the medical students had a assimilating (theorist) learning style and the faculty was (pragmatist) converger22. the undergraduate and practitioner nurses were found to be reflectors23,24 .a comparative study from a dentistry school in latin america showed that both students and professors had a preference for reflector learning style followed by theorist25. learning styles were assessed in the participants of continued medical education and found that the most preferred learning style was assimilating (theorist ) followed by diverger (reflector)26. the preferred learning styles also shows variation over time as seen in the studies in a same set up two decades apart. which has shown a change from theorist learning style in 1993 to reflectors learning style in 201026. adult learners who were inducted for diploma course showed preference for three learning styles 48 comparison of learning styles used by clinical faculty of hospital and general practitioners for….. activists, reflector and theorists. the pragmatist style was missing in this group27,28 . in all of the previous studies in different categories of medical professionals around the world the preferred learning style was reflectors in most of the consultant groups, in post graduate residents, and in nursing groups. in our study also the most strongly preferred learning style was reflector among the clinical faculty, so this showed a similarity between most of the previous studies. considering the characteristics of this learning style the clinical faculty like to collect data and think about it. they make use of a concrete experience in perceiving and reflective observation in organizing, their strength is creativity, understanding others and developing a large perspective. it is likely that if they follow this learning style would likely help in their professional development. general practitioners: different studies on the learning styles in general practitioners were also evaluated. in one study it was found that out of 274 general practitioners about 43.8% were theorists, followed by reflectors 21%, and then pragmatists 18 % and finally activists 16%29. in another study from hong kong a preassessment followed by post assessment showed that most of them were reflector in both assessments30. in another study on general practitioner registrar suggested that the most prevalent learning style was reflector/theorist31. in our study the general practitioners were mainly reflectors followed by theorists, and then activists followed by pragmatists. although there was some difference in the results, however in the previous studies a significant percentage of general practitioners were reflectors. in both the groups under study the preferred learning style was reflector so they both may follow similar learning strategies in their professional development. they work as an integral part of health team as observers. they develop creative solutions, self-critique and analysis. the only statistically significant difference was seen in the moderate preference category of activist learning style. that means that a significant number of general practitioners like to have new experiences. they like acting based on their feelings rather than mental analyses. their strong point was practicality and they love to learn by doing. if activists follow the strategy of practical approach towards the professional development they are likely to succeed more. best learning strategy for them would be case based discussion, and interactive learning. they must have a hands-on experience for producing best results15. conclusion reflector type of learning style based on pondering, experiencing and observing different perspectives was strongly observed in both genders of consultants and general practitioners. however a moderate degree of activist type of learning style nfluenced by doing and feeling was also noted in the general practitioners. continued medical education (cme), instructional strategy and assessment based on these learning styles could enhance further academic career growth of these two groups of health professionals. references 1. shuell, t. j. cognitive conceptions of learning. review of educational research 1986, 56, 411-436. 2. mil d, milosevic and al .adaptiveleaning by using scos metadata .inter disciplinary journal of knowledge and learning objects2007 ,vol 3,1631744-3-jensen, e. . 3. -pashler, h., mcdaniel, m., rohrer, d., & bjork, r. (2008). learning styles: concepts and evidence. psychological science in the public interest, 9(3),105–19. https://doi.org/10.1111/j.15396053.2009.01038.x 4. kolb, d. a. experiential learning: experience as the source of learning and development.. englewood cliffs: prentice hall; 1984 5. curry, l. integrating concepts of cognitive or learning style: a review with attention to psychometric standards. ottawa, canada: canadian college of health service executives.1987 6. alaya nb. elferchichi a, towards the elaboration of a meta model unifying learning styles .conference flic 2007; sausse tunisia 7. fleming nd. teaching and learning styles: vark strategiesn.d. fleming, christchurch, new zealand (2001) 8. mozaffari, hr, janatolmakan m, sharifi r, ghandinejad,f andayeshgar b & alireza khatony a.. the relationship between the vark learning styles and academic achievement in dental students,: advances in medical education and practice,2020: 11:, 15-19, doi: 10.2147/amep.s235002 9. kolb, d. a., boyatzis, r., &mainemelis, c. (2001). experiential learning theory: previous research & new directions. in r. sternberg, & l. zhang (eds.), perspectives on cognitive learning & thinking styles (2001) (pp. 228-247). mahwah, nj: lawrence erlbaum associates. 10. honey p, mumford a. the manual of learning styles .peter honey publications, maidenhead, england (1992) 49 comparison of learning styles used by clinical faculty of hospital and general practitioners for….. 11. ehrman, m. second language learning difficulties: looking beneath the surface. thousand oaks, ca: sage.1996. 12. paris, s. g., &winograd, p. promoting metacognition & motivation of exceptional children. remedial & special education.1990 11, 7-15 13. plovnick, m. s. primary care career choices and medical student learning styles. journal of medical education1975 50(9), 849-855. 14. mansoor sn, yousaf o, rahman ss. comparison of learning styles among post graduate residents and full timespeciality clinicians pursuing higher education degree. pak armed forces med j2019; 69(4):887-93 15. shukr i, zainab r, rana mh. learning styles of postgraduate and under graduate medical students .jcpsp.2013,vol23(1):25-30 16. fatima sh, hussain a, naseem s, fatimasr.different and dominant preferences of learning styles among undergraduate medical students from islamabad and gujranwala cities, pakistan.2022.gjms;20:4,172-177 17. guraya ss, guraya sy, habib fa, khoshhal ki. learning styles of medical students at taibah university: trends and implications. j res med sci. 2014 dec; 19(12):1155-62. doi: 10.4103/17351995.150455. pmid: 25709657; pmcid: pmc4333524. 18. ahmed a, wojcik em, ananthanarayananv, mulder l ,mirza km. learning style in pathology :a comparative analysis and implications of learner centred education .academic pathology;6:2019 19. gokalp m-the effect of student’s learning styles to their academic success-success creative education 2013; 46:728-737 20. adesunloyeba , aladesanm oi, henriques m forsythe, ivonye c the preferred learning style among residents and faculty members of an internal medicine residency program j natl med assoc.2008 feb;100(2):172-5. 21. uraiqat a, rubeihat h, alsudi m, amireh a. surgeons ‘learning style in comparison to their medical colleagues .jrms december2015 ;22(4):2430. 22. engels pt, gara cd. learning styles of medical students, general surgeryresidents and general surgeons:implication for surgical education .bmcmedical education 2010,10:15. 23. rasool gh, rawaf s, .learning style preferences of under graduate nursing students .nursing standard 2007 ;24:21(32) . 24. austin f, clos sj, hughes n. the self-reported leaning style preferences of females, macmillan clinical nurse specialist .nurse edu today 2006;26(6):475-8319. 25. ardila cm, goméz-restrepo am, echeverrirestrepo c. learning styles of students and professors of a dental school in latin america: a cross-sectional study. global journal of health science. 2019 nov 20; 11(13):104 26. scott collins s, nanda s, brian a. palmer, arya b. mohabbat, cathy d. schleck, jayawant n. mandrekar, saswati mahapatra, thomas j. beckman & christopher m. wittich (2019) a cross-sectional study of learning styles among continuing medical education participants, medical teacher, 41:3, 318324, doi: 10.1080/0142159x.2018.1464134. 27. amponsah, s. exploring the dominant learning styles of adult learners in higher education. int rev educ 66, 531–550 (2020). https://doi.org/10.1007/s11159-020-09845-yeubank tf, pitts j.a comparison of learning styles across the decades .optometric education 2011.vol 36.2/winter/spring 28. eubank tf, pitts j.a comparison of learning styles across the decades .optometric education 2011.vol 36.2/winter/springlewis ap, bolden kj. general practitioners and their learning styles. journal of royal college of general practitioners, 1989, 39:187189. 29. lewis ap, bolden kj. general practitioners and their learning styles. journal of royal college of general practitioners, 1989, 39:187-189.mccall l, klien b, piterman l, lam tp. learning style preferences for hongkong gps recruited in distance learning course.hk pract2005;27 :261-267. 30. mccall l, klien b, piterman l, lam tp. learning style preferences for hong kong gps recruited in distance learning course.hk pract2005;27 :261-31. 31. .lesmes-anel j, robinson g moody s learning preferences and learning styles: a study of wessex general practice registrars. british journal of generalpractice 2001; 51 (468): 559-564. corresponding author: prof (brig r). dr. muhammad atiquesi(m), chairman /professor of pathology, department of pathology, pakistan kidney & liver institute & research centre (pkli), lahore. email: muhammad.atique@pkli.org.pk untitled-1.indd 7 proceedings s.z.m.c. vol: 34(2): pp. 7-10, 2020. pszmc-744-34-2-2020 the potential role of acei and arbs in covid-19; a perspective ahmad hammad hassan primary and secondary healthcare department, punjab, pakistan abstract covid-19 pandemic has caused significant morbidity and mortality around the world. the disease severity ranges from mild upper respiratory infection to severe lower respiratory and cardiac illness. acute respiratory distress syndrome (ards) is the most serious complication and results in diffuse inflammatory alveolar damage, respiratory failure, and death. components of the renin-angiotensin-aldosterone-system (raas) are involved in an inflammatory reaction in the lungs. various studies have shown that blocking raas peptides in the lungs especially angiotensin-converting enzyme (ace) and type-1 angiotensin receptor (atr1) reduces lung injury, improves respiratory function and is associated with better clinical outcomes in the covid-19 patients. we suggest that angiotensin-converting enzyme inhibitors (acei) and angiotensin receptor blockers (arbs) – drugs that block raas peptides – be considered for a repurposed use in covid19 induced lung injury. introduction corona virus disease 2019 (covid-19) emerged as a respiratory illness of unknown infectious origin in china. it then spread all around the world and has caused substantial mortality. the disease-causing virus, sars-cov2 (later identified), is closely related to other beta corona viruses, sars-cov (2003) and mers-cov (2012), that have caused severe respiratory disease pandemics in the past two decades. the disease follows a variable pattern, ranging from asymptomatic or mild upper respiratory or gastrointestinal infection1 to a severe lower respiratory tract (pneumonia) or cardiac (myocarditis) illness, especially in patients with other co-morbidities.2 in very severe cases, a hyperinflammatory response causes cytokine-storm leading to acute respiratory distress syndrome (ards), respiratory failure, sepsis, and death.3 the exact mechanism of this variable disease expression is not yet understood. the pathogenicity and tissue tropism of apathogenis determined by the type of interaction with the host receptors and the tissue distribution of interacting receptors in the host.4 sars-cov2 is genetically 72% similar to sars-cov and likewise interacts with angiotensin-converting enzyme 2 (ace2)as receptor through its spike glycoprotein (s) to bind, fuse and enter into the host cells.5,6,7 ace2 is a component of the renin-angiotensin-aldosteronesystem (raas) and is expressed abundantly in airway epithelia, lung parenchyma, heart, intestines, esophagus, bladder, and kidney.5 the raas regulates fluid balance and blood pressure through several peptides, including renin, angiotensin 2 (at2) and aldosterone. the precursor angiotensinogen is released from the liver and is cleaved by renin into angiotensin 1 which is, in turn, converted into angiotensin 2 by the angiotensinconverting enzyme 1 (ace) in the lungs. angiotensin 2 (at2) maintains vascular tone, blood pressure and fluid volume through its vasoconstrictor effect, the release of aldosterone and vasopressin, and direct catecholaminergic effects on cardiac myocytes. at2 is then converted into angiotensin (1-7) by angiotensin-converting enzyme 2 (ace2). angiotensin (1-7) mitigates the effects of at2 and thus a balance exists between ace and ace2 for appropriate homeostasis.8 angiotensin-converting enzyme inhibitors (acei) and angiotensin receptor blockers (arbs) are drugs that act on the raas and are used in hypertension, cardiomyopathies and heart failure. much controversy exists on the effects of these drugs on the respiratory and cardiac pathology of covid-19.9 long term use of acei can upregulate ace25 due to the absence of its natural ligand at2. the long-term use of arbs has been also been associated with the upregulation of ace2.10 this may cause an increased number of ace2 receptors made available to the virus for infecting host cells and consequently enhanced pathogenicity. indeed, hypertension and cardiovascular diseases are the top two co-morbidities associated with the worst outcome in covid-192 and that may be attributed to the long term use of aceis or arbs. 8 the potential role of acei and arbs in covid-19; a perspective role of raas components in covid-19 lung injury: in addition to their respective roles in raas, ace, ace2, and at2 are also involved in the pathogenesis of acute lung injury. interestingly, an imbalance between ace and ace2 activity is found in various disease models of ards (the most common complication of covid-19). increased ace activity and decreased ace2 activity in the lungs is associated with diffuse alveolar edema and lung injury.11 at2 has been found to have proinflammatory and pro-fibrotic effects (in addition to raas effects)12 and lung levels of at2 were found to be elevated in mice model of sars-cov13 induced ards and in covid-19 patients.14 ace2 has been shown to protect mice from severe lung damage induced by acid aspiration.13 ace2 also hydrolyzes and inactivates des-bradykinin, a proinflammatory peptide,15 that can potentially worsen inflammation-induced lung damage. blockade of angiotensin receptor 1 (atr1) with losartan attenuates acute lung injury in mouse models of ards.13 the lung-protective effects of acei and arbs against lung injury in ards have been described in various clinical studies as well.16,17 accordingly, increased ace2 activity and atr1 antagonism have a protective role in acute lung injury and ards.13,18 a hypothesized mechanism of covid-19 lung injury (based on studies on previous sars-cov) is that the sars-cov2 binds ace2 in the lungs through the spike glycoprotein (s). s is cleaved at a polybasic site by trans membrane protease (tmprss2) into s1 and s2,6 the virion fuses with the epithelial cell membrane and is internalized along with ace2 by ph-dependent endocytosis.19 internalization results in the down regulation of ace2,20 thus tipping the balance towards relative excess of ace. this results in more angiotensin 2 and that may worsen lung injury and ards (fig a). increased number of ace2 receptors can cause increased viral load that causes more lung damage. this presents a conflicting paradox to the lungprotective effects of ace2 and atr1 blockade. it can be partly explained by the timing of expression5,21 of ace2 receptors on airway epithelia and lung parenchyma. the preexisting increased numbers of ace2 (due to long-term acei or arb use) can provide a good homing ground to the sars-cov2 and may cause increased pathogenicity. also, after interaction with the virus, the ace2 would internalize, thus decreasing the number of available ace2 at the time of lung injury that comes late after virus internalization. contrarily, inducing ace2 expression, or blocking atr1 at the time of lung-injury may enhance the protective effects of these mechanisms. these mechanisms are not fully characterized and we suggest further experimental studies to determine the exact underlying mechanisms. fig-a:hypothesized mechanism of sars-cov2 cell-infectivity and subsequent lung-damage with focus on the balance between ace (yellow) and ace2 (black). sars-cov2 (red spiked spheres) via its spike glycoprotein s bind with ace2 on lung epithelial cells. s is cleaved to s1 and s2 (not shown) by tmprss (blue stripe) and internalized. this also leads to the internalization of ace2 and thus decreased numbers of ace2 on the epithelium. decreased ace2 and relative increased ace disturbs the balance. this subsequently results in more at2 compared to at1 (as shown in the second part of the figure). at2 binds its receptor atr1 and can potentially contribute towards diffuse alveolar edema and inflammatory acute lung injury as shown by edema and infiltrates in the second part of the figure. red spiked spheres = sars-cov2. yellow chevron arrows = ace. black chevron arrows = ace2. blue strips = tmprss. light blue box = angiotensin 2 (at2). green box = angiotensin 1-7 (at1). purple triangle = angiotensin 2 receptor 1(atr1). 9 the potential role of acei and arbs in covid-19; a perspective fig-b:schematic role of raas components (ace, ace2, at2, and atr1) in acute lung injury and sites where raas antagonists (race2, acei, and arbs) block the inflammatory pathway and attenuate inflammatory lung injury. s (yellow triangle) = spike glycoprotein of sars-cov2. ace2 counter balance ace and subsequent pathway by deactivating at2. race2 = recombinant ace2, can potentiate activity of ace2. the potential lung-protective roleof raas antagonistsin covid-19: based on the fact that angiotensin 2 (at2) has proinflammatory activity in the lungs and the inflammatory basis of acute lung injury and ards in covid-19 patients, the following strategies may protect against the fatal lung damage in covid-19. (a) therapeutically inducing ace2 activity. (b) blocking lung angiotensin receptor 1 (atr1). (c) decreasing lung angiotensin 2 (at2) levels. (fig. b). recombinant ace2 receptor can both block the virus from binding natural ace2 and also counterbalance the proinflammatory effects of ace. it has shown promising results in experimental studies22 but clinical trials are yet to be completed. acei and arbs are available drugs and their pharmacokinetics, pharmacodynamics, and safety profile are well established. these drugs can work by decreasing the at2 levels and blocking atr1 in the lungs, both effects rescue acute lung injury. a recent clinical study (n=417) has demonstrated the beneficial effects of local renin-angiotensin antagonism against inflammatory lung injury in covid-19 patients. arb and acei use was shown to be associated with less inflammatory lung injury, enhanced acquired immunity, decreased viral load, and better clinical outcomes in covid-19 patients.23 we, therefore suggest that these drugs can be repurposed to be used in covid-19 lower respiratory tract injury8 after appropriate testing, dose adjustments and optimization. it is worth mentioning here, that, although there has been an association between the use of acei/arbs and the worst cardio respiratory outcome in covid-19, there is no adequate experimental or clinical evidence in humans yet that long-term use of acei or arbs worsen the cardio respiratory illness associated with sars viruses.4,24 some further clinical trials are going on to ascertain and confirm the exact effects of renin-angiotensin antagonism on covid-19 associated lung-injury and results are awaited25 (nct04330300). current recommendations, however, are to continue the use of acei and arbs for their renal and cardiac protective benefits24. conclusion the proinflammatory role of raas components, ace and at2 in acute lung injury coupled with the inflammatory nature of lung damage in covid-19, justifies that raas antagonists (arbs and acei) can be considered as a therapeutic modality to protect against acute lung injury and respiratory failure in covid-19. as these drugs are already in use, and their kinetics and dynamics are well known, they can be readily readopted for their lungprotective benefits. the dose and timing of administration of these drugs will need to be optimized. references 1. gu j, han b, wang j. covid-19: gastrointestinal manifestations and potential fecal-oral transmission. gastroenterology [internet]. 2020; (april):118-9. available from: https://doi.org/10.1053/j.gastro.2020.02.054 2. zhou f, yu t, du r, fan g, liu y, liu z, et al. clinical course and risk factors for mortality of adult in patients with covid-19 in wuhan, china : a retrospective cohort study. lancet [internet]. 2020; 395(10229):1054-62. 3. mehta p, mcauley df, brown m, sanchez e, tattersall rs, manson jj, et al. covid-19: consider cytokine storm syndromes and immunosuppression. lancet [internet]. 2020; 395(10229):1033-4. 4. maginnis ms. virus-receptor interactions: the key to cellular invasion. j mol biol [internet]. 2018/06/18. 2018 aug 17; 430(17):2590-611. 5. 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system blockade in patients with acute respiratory distress syndrome : a retrospective casecontrol study. 2017; 29(04):154-63. 17. ruthman ca, festic e. emerging therapies for the prevention of acute respiratory distress syndrome. ther adv respir dis [internet]. 2015/05/22. 2015 aug; 9(4):173-87. 18. imai y, kuba k, rao s, huan y, guo f, guan b, et al. angiotensin-converting enzyme 2 protects from severe acute lung failure. 2005; 436(july). 19. yang n, shen h. targeting the endocytic pathway and autophagy process as a novel therapeutic strategy in covid-19. 2020; 16. 20. nl c, glowacka i, bertram s, herzog p, pfefferle s, steffen i, et al. differential down regulation of ace2 by the spike proteins of severe acute respiratory syndrome corona virus and human. 2010; 84(2):1198-205. 21. rockx b, baas t, zornetzer ga, haagmans b, sheahan t, frieman m, et al. early upregulation of acute respiratory distress syndrome-associated cytokines promotes lethal disease in an aged-mouse model of severe acute respiratory syndrome corona virus infection. 2009; 83(14):7062-74. 22. monteil v, kwon h, prado p, hagelkrüys a, wimmer ra. inhibition of sars-cov-2 infections in engineered human tissues using clinical-grade soluble human ace2. 2020; 23. meng j, xiao g, zhang j, he x, ou m, bi j, et al. renin-angiotensin system inhibitors improve the clinical outcomes of covid-19 patients with hypertension. 2020; 1751. 24. angiotensinpi. cardiovascular disease : a viewpoint on the enzyme inhibitors/ angiotensin receptor. 2020; 2019:1-5. 25. clinicaltrials.gov [internet]. john william mcevoy (mbbch, mhs): national library of medicine (us). 2020 apr 01 -. identifier nct04330300, corona virus acei/arb investigation (coronacion); 2020 apr 01. availablefrom:https://clinicaltrials.gov/ct2/show /nct04330300 corresponding author: dr. ahmad hammad hassan, medical officer, primary & secondary health care department, punjab. e-mail: ahmadhammadhassan@gmail.com full book for web 25 proceedings s.z.m.c. vol: 36(3): pp. 25-29, 2022. pszmc-848-36-3-2022 department of pathology, king edward medical university, lahore. 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 26 haematological and clinical findings especially third space fluid accumulation observed in major… 27 haematological and clinical findings especially third space fluid accumulation observed in major… 28 haematological and clinical findings especially third space fluid accumulation observed in major… 29 haematological and clinical findings especially third space fluid accumulation observed in major… full book for web 55 proceedings s.z.m.c. vol: 36(3): pp. 55-59, 2022. pszmc-853-36-3-2022 1department of pediatrics, cmh medical college, lahore. 2department of community medicine, cmh medical college, lahore. 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 56 preterm neonatal need for mechanical ventilation in prophylactic continuous positive air pressure… 57 preterm neonatal need for mechanical ventilation in prophylactic continuous positive air pressure… 58 preterm neonatal need for mechanical ventilation in prophylactic continuous positive air pressure… 59 preterm neonatal need for mechanical ventilation in prophylactic continuous positive air pressure… full book for web 60 proceedings s.z.m.c. vol: 36(3): pp. 60-64, 2022. pszmc-854-36-3-2022 department of physiology, pgmi, ameer-ud-din medical college & general hospital, lahore p 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 61 oxidative stress in non-obese & obese young healthy adults p p ppp ppp 62 oxidative stress in non-obese & obese young healthy adults ppp p p p p p 63 oxidative stress in non-obese & obese young healthy adults 64 oxidative stress in non-obese & obese young healthy adults full book for web 36 proceedings s.z.m.c. vol: 36(3): pp. 36-42, 2022. pszmc-850-36-3-2022 1department of medicine, cmh lahore medical college & institute of dentistry, lahore. 2department of medicine, fatima memorial hospital, lahore. 3department of pediatrics, cmh lahore medical college & institute of dentistry, lahore 4department of nephrology, fatima memorial hospital lahore. 5department of physiology, cmh lahore medical college & institute of dentistry, lahore. 6department of medicine, pkli hospital, lahore. 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 37 perception and status of vaccination in individuals aged 60 years and above: an urban lahore analysis 38 perception and status of vaccination in individuals aged 60 years and above: an urban lahore analysis 39 perception and status of vaccination in individuals aged 60 years and above: an urban lahore analysis 40 perception and status of vaccination in individuals aged 60 years and above: an urban lahore analysis 41 perception and status of vaccination in individuals aged 60 years and above: an urban lahore analysis 42 perception and status of vaccination in individuals aged 60 years and above: an urban lahore analysis full book for web 1 proceedings s.z.m.c. vol: 36(3): pp. 01-12, 2022. pszmc-845-36-3-2022 tlr4 1department of zoology, government college university, lahore. 2department of biological sciences, national university of medical sciences, rawalpindi. 3director, science shines ltd. london, u.k. 4department of zoology, university of gujrat. tlr4 gene 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 2 family based genetic association study of tlr4 gene with myocardial infarction in pakistan 3 family based genetic association study of tlr4 gene with myocardial infarction in pakistan 4 family based genetic association study of tlr4 gene with myocardial infarction in pakistan 5 family based genetic association study of tlr4 gene with myocardial infarction in pakistan 6 family based genetic association study of tlr4 gene with myocardial infarction in pakistan tlr4 7 family based genetic association study of tlr4 gene with myocardial infarction in pakistan 8 family based genetic association study of tlr4 gene with myocardial infarction in pakistan 9 family based genetic association study of tlr4 gene with myocardial infarction in pakistan p fisher p 10 family based genetic association study of tlr4 gene with myocardial infarction in pakistan tlr4tlr4 tlr4 11 family based genetic association study of tlr4 gene with myocardial infarction in pakistan 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 12 family based genetic association study of tlr4 gene with myocardial infarction in pakistan full book 45 proceedings s.z.m.c. vol: 36(4): pp. 45-50, 2022. pszmc-863-36-4-2022 1department of anatomy, ucmd, university of lahore 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 46 faculty perception about e-learning and teaching anatomy: an experience of covid pandemic 47 faculty perception about e-learning and teaching anatomy: an experience of covid pandemic 48 faculty perception about e-learning and teaching anatomy: an experience of covid pandemic 49 faculty perception about e-learning and teaching anatomy: an experience of covid pandemic 50 faculty perception about e-learning and teaching anatomy: an experience of covid pandemic 404 not found full book 6 proceedings s.z.m.c. vol: 36(4): pp. 06-10, 2022. pszmc-858-36-4-2022 1department of pharmacology, fmh college of medicine and dentistry, lahore 2department of pharmacology, king edward medical university, lahore 3department of pharmacology, post graduate medical institute, lahore 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 7 does short term atorvastatin treatment improve symptomatic control in patients with mild to moderate… 8 does short term atorvastatin treatment improve symptomatic control in patients with mild to moderate… ppp9 does short term atorvastatin treatment improve symptomatic control in patients with mild to moderate… 10 does short term atorvastatin treatment improve symptomatic control in patients with mild to moderate… full book for web 70 proceedings s.z.m.c. vol: 36(3): pp. 70-73, 2022. pszmc-856-36-3-2022 department of orthopedic surgery, lahore general hospital, lahore, pakistan. 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 71 obturator type of hip dislocation with ipsilateral non-displaced neck of femur fracture: a case report 72 obturator type of hip dislocation with ipsilateral non-displaced neck of femur fracture: a case report 73 obturator type of hip dislocation with ipsilateral non-displaced neck of femur fracture: a case report full book for web 11 proceedings s.z.m.c. vol: 37(1): pp. 11-17, 2023. pszmc-870-37-1-2023 1public health department, uhs, university of health sciences, lahore. 2primary & secondary health care department, lahore. 3department of anesthesia, sir ganga ram hospital, lahore. 4department of community medicine, uol, lahore. 70 1department of orthopedics, thq, khanewal 2department of orthopedics, lgh, lahore 3department of orthopedics, thq, ferozwala 4department of orthopedics, cmh, lahore 5department of hand & upper limb surgery, cmh, lahore 6department of orthopedics, al-rehmat benevolent trust 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 12 attitude, awareness and recommendations regarding fortified food usage amongst health care….. 13 attitude, awareness and recommendations regarding fortified food usage amongst health care….. 14 attitude, awareness and recommendations regarding fortified food usage amongst health care….. 15 attitude, awareness and recommendations regarding fortified food usage amongst health care….. 16 attitude, awareness and recommendations regarding fortified food usage amongst health care….. 17 attitude, awareness and recommendations regarding fortified food usage amongst health care….. full book for web 1 proceedings s.z.m.c. vol: 37(1): pp. 01-05, 2023. pszmc-868-37-1-2023 ,2 ,2 ,3 ,1 1department of radiology, pakistan kidney and liver institute and research center (pkli &rc), lahore. 2department of interventional radiology, pakistan kidney and liver institute and research center (pkli &rc), lahore. 3department of oncology, shaukat khanum memorial cancer hospital and research centre (skmch &rc), lahore. 70 1department of orthopedics, thq, khanewal 2department of orthopedics, lgh, lahore 3department of orthopedics, thq, ferozwala 4department of orthopedics, cmh, lahore 5department of hand & upper limb surgery, cmh, lahore 6department of orthopedics, al-rehmat benevolent trust 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 2 technical considerations and complication of percutaneous transhepatic biliary drainage: a single….. 3 technical considerations and complication of percutaneous transhepatic biliary drainage: a single….. 4 technical considerations and complication of percutaneous transhepatic biliary drainage: a single….. 5 technical considerations and complication of percutaneous transhepatic biliary drainage: a single….. full book 37 proceedings s.z.m.c. vol: 36(4): pp. 37-44, 2022. pszmc-862-36-4-2022 department of community medicine, shalamar medical and dental college, lahore 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 38 dual informants reporting: do we observe behavioral problems in primary school children? 39 dual informants reporting: do we observe behavioral problems in primary school children? 40 dual informants reporting: do we observe behavioral problems in primary school children? 41 dual informants reporting: do we observe behavioral problems in primary school children? 42 dual informants reporting: do we observe behavioral problems in primary school children? 43 dual informants reporting: do we observe behavioral problems in primary school children? 44 dual informants reporting: do we observe behavioral problems in primary school children? full book for web 41 proceedings s.z.m.c. vol: 37(1): pp. 41-46, 2023. pszmc-875-37-1-2023 1department of pulmonology, shaikh zayed hospital, federal postgraduate medical institute, lahore pakistan. 2department of surgery, combined military hospital, sialkot pakistan. 3department of thoracic surgery, combined military hospital, multan pakistan. 4department of medicine, university college of medicine and dentistry lahore pakistan. 5centre of excellence in molecular biology, university of punjab, lahore. 70 1department of orthopedics, thq, khanewal 2department of orthopedics, lgh, lahore 3department of orthopedics, thq, ferozwala 4department of orthopedics, cmh, lahore 5department of hand & upper limb surgery, cmh, lahore 6department of orthopedics, al-rehmat benevolent trust 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 42 a 5 year tertiary centre experience of frequency & risk factors association with primary and secondary... 43 a 5 year tertiary centre experience of frequency & risk factors association with primary and secondary... 44 a 5 year tertiary centre experience of frequency & risk factors association with primary and secondary... 45 a 5 year tertiary centre experience of frequency & risk factors association with primary and secondary... . 46 a 5 year tertiary centre experience of frequency & risk factors association with primary and secondary... full book for web 52 proceedings s.z.m.c. vol: 37(1): pp. 52-55, 2023. pszmc-877-37-1-2023 1department of orthopedic surgery, lahore general hospital, lahore, pakistan. 70 1department of orthopedics, thq, khanewal 2department of orthopedics, lgh, lahore 3department of orthopedics, thq, ferozwala 4department of orthopedics, cmh, lahore 5department of hand & upper limb surgery, cmh, lahore 6department of orthopedics, al-rehmat benevolent trust 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 53 case report: neglected locked anterior shoulder dislocation with upper brachial plexus injury 54 case report: neglected locked anterior shoulder dislocation with upper brachial plexus injury 55 case report: neglected locked anterior shoulder dislocation with upper brachial plexus injury full book for web 18 proceedings s.z.m.c. vol: 37(1): pp. 18-22, 2023. pszmc-871-37-1-2023 1department of ophthalmology, shaikh zayed hospital, lahore. 2department of ophthalmology, continental medical college, lahore. 70 1department of orthopedics, thq, khanewal 2department of orthopedics, lgh, lahore 3department of orthopedics, thq, ferozwala 4department of orthopedics, cmh, lahore 5department of hand & upper limb surgery, cmh, lahore 6department of orthopedics, al-rehmat benevolent trust 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 19 hba1c levels an indicator of glycemic control in diabetics with and without retinopathy 20 hba1c levels an indicator of glycemic control in diabetics with and without retinopathy 21 hba1c levels an indicator of glycemic control in diabetics with and without retinopathy 22 hba1c levels an indicator of glycemic control in diabetics with and without retinopathy full book foer web 19 proceedings s.z.m.c. vol: 36(2): pp. 19-22, 2022. pszmc-839-36-2-2022 1department of anatomy, ucmd, university of lahore, lahore 2department ofmedical education, pak red crescent medical and dental college, lahore 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 20 perception of mbbs students about effectiveness of modified unobserved anatomy ospe… 21 perception of mbbs students about effectiveness of modified unobserved anatomy ospe… 22 perception of mbbs students about effectiveness of modified unobserved anatomy ospe… for web full book 35 proceedings s.z.m.c. vol: 36(2): pp. 35-39, 2022. pszmc-842-36-2-2022 1department of cardiology, wapda hospital, lahore 2department of cardiology, shaikh zayed medical complex, lahore 3department of cardiology, social security hospital, lahore 4department of pulmonology, shaikh zayed medical complex, lahore 5department of neurology, shaikh zayed medical complex, lahore 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 36 association of left ventricular hypertrophy on echocardiography with hemorrhagic… 37 association of left ventricular hypertrophy on echocardiography with hemorrhagic… 38 association of left ventricular hypertrophy on echocardiography with hemorrhagic… 39 association of left ventricular hypertrophy on echocardiography with hemorrhagic… for web full book 46 proceedings s.z.m.c. vol: 36(2): pp. 46-51, 2022. pszmc-844-36-2-2022 department of general surgery, sir ganga ram hospital, lahore. 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 47 association between depressed mood and morbid obesity: experience from a bariatric surgery program 48 association between depressed mood and morbid obesity: experience from a bariatric surgery program 49 association between depressed mood and morbid obesity: experience from a bariatric surgery program 50 association between depressed mood and morbid obesity: experience from a bariatric surgery program 51 association between depressed mood and morbid obesity: experience from a bariatric surgery program for web full book 23 proceedings s.z.m.c. vol: 36(2): pp. 23-28, 2022. pszmc-840-36-2-2022 1department of biochemistry, azra naheed medical college, lahore 2department of biochemistry, lahore medical and dental college, lahore 3department of physiology, azra naheed medical college, lahore 4department of pathology, azra naheed medicalcollege, lahore 5department of pathology, al-aleem medical college, lahore 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 24 assessment of lipid profile in females suffering from endometriosis 25 assessment of lipid profile in females suffering from endometriosis 26 assessment of lipid profile in females suffering from endometriosis p< 27 assessment of lipid profile in females suffering from endometriosis 28 assessment of lipid profile in females suffering from endometriosis full book for web 31 proceedings s.z.m.c. vol: 37(1): pp. 31-35, 2023. pszmc-873-37-1-2023 1department of surgery, chaudhry mohammad akram hospital, lahore. 70 1department of orthopedics, thq, khanewal 2department of orthopedics, lgh, lahore 3department of orthopedics, thq, ferozwala 4department of orthopedics, cmh, lahore 5department of hand & upper limb surgery, cmh, lahore 6department of orthopedics, al-rehmat benevolent trust 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 32 early postoperative pain in video assisted anal fistula treatment versus open fistulectomy 33 early postoperative pain in video assisted anal fistula treatment versus open fistulectomy 34 early postoperative pain in video assisted anal fistula treatment versus open fistulectomy 35 early postoperative pain in video assisted anal fistula treatment versus open fistulectomy full book for printing 15 proceedings s.z.m.c. vol: 37(2): pp. 15-18, 2023. pszmc-880-37-2-2023 1department of anatomy, university college of medicine & dentistry, university of lahore, lahore. 2department of anatomy, abu umara medical and dental college, lahore. 3department ofanatomy, akhtarsaeed medical and dental college, bahria town, lahore. p 70 1department of orthopedics, thq, khanewal 2department of orthopedics, lgh, lahore 3department of orthopedics, thq, ferozwala 4department of orthopedics, cmh, lahore 5department of hand & upper limb surgery, cmh, lahore 6department of orthopedics, al-rehmat benevolent trust 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 16 local sexual dimorphism based on chilotic index of human dried hip bone: a morphometric study p 17 local sexual dimorphism based on chilotic index of human dried hip bone: a morphometric study p 18 local sexual dimorphism based on chilotic index of human dried hip bone: a morphometric study full book for printing 1 proceedings s.z.m.c. vol: 37(2): pp. 01-08, 2023. pszmc-878-37-2-2023 1department of clinical chemistry and immunology, chughtai institute of pathology. 2department of histopathology, chughtai institute of pathology. 3research director, shifa tameer-e-millat university, pitras bukhari road, islamabad, pakistan. 70 1department of orthopedics, thq, khanewal 2department of orthopedics, lgh, lahore 3department of orthopedics, thq, ferozwala 4department of orthopedics, cmh, lahore 5department of hand & upper limb surgery, cmh, lahore 6department of orthopedics, al-rehmat benevolent trust 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 2 characterization of mannose binding lectin (mbl) levels in type-2 diabetes mellitus patients amongst … 3 characterization of mannose binding lectin (mbl) levels in type-2 diabetes mellitus patients amongst … 4 characterization of mannose binding lectin (mbl) levels in type-2 diabetes mellitus patients amongst … 5 characterization of mannose binding lectin (mbl) levels in type-2 diabetes mellitus patients amongst … 6 characterization of mannose binding lectin (mbl) levels in type-2 diabetes mellitus patients amongst … journalofimmunology scandinavian journal of 7 characterization of mannose binding lectin (mbl) levels in type-2 diabetes mellitus patients amongst … immunology diabetes 8 characterization of mannose binding lectin (mbl) levels in type-2 diabetes mellitus patients amongst … for web full book 20 proceedings s.z.m.c. vol: 37(3): pp. 20-23, 2023. pszmc-891-37-3-2023 case report: isolated trochlear fracture of the humerus 1shehzad anwer, 1shuja uddin,1hasnain khalid,1rizwan ul haq,1adeel hamid,1mubashir farhan,1zia uddin,1mian muhammad haneef 1department of orthopedics, general hospital, lahore. abstract the fracture of the distal part of the humerus usually involves the capitellum and variable part of the trochlea. the deep position of the trochlea within the elbow joint makes it a rare occurrence and protects it from direct damage. a 23-yearold local man named mumtaz presented with an intra-articular half-moon-shaped fragment that had traveled and forward, leading us to assume a capitellar fracture. that was fixed through the medial approach of distal humerus. postoperatively, patient a had good pain-free range of motion. keywords: fracture, trochlea, humerus introduction the first description of an isolated trochlear fracture of the humerus was in 1853 by laugier. thus, the trochlea fracture is also sometimes acknowledged as laugier’s fracture. the fracture of the distal part of humerus usually involves capitellum and variable part of the trochlea. the deep position of the trochlea within the elbow joint makes it a rare occurrence and protects it from direct damage. these fractures are known to result in elbow post-traumatic arthritis and instability. therefore, management of these types of fractures requires a systematic approach, obtaining an articular anatomic reduction and providing a painless, stable and mobile elbow joint3. the laugier's fracture, also known as an isolated fracture of the trochlea, is uncommon and frequently accompanied by elbow dislocation, ligamentous injuries, capitellar fractures, radial head fractures, and/or olecranon fractures2,9. regarding its management, there is no agreement, which is based on trans articular distal humeral fractures. posterior splint immobilization for 3 weeks is recommended if the fracture is undisplaced5. if there are small unamenable osteochondral fragments, excision followed by early elbow movement is the treatment of choice2,10. displaced fractures should be anatomically reduced to restore articular congruency, using headless herbert screws or bioabsorbable screws2, 11, 12. case report a 23-year old local man named mumtaz presented with trauma to the right elbow sustained due a to fall from a height. radiographs revealed an isolated trochlear fracture of the humerus. a lateral radiograph revealed an intra-articular half-moon-shaped fragment that had traveled forward, leading us to assume a capitellar fracture. anteroposterior radiographs merely revealed an abnormality of his medial joint area (fig-1). a computed tomography (ct) scan confirmed an isolated front-line fracture of his trochlea, without the participation of the capitellum (fig-2). open reduction and internal fixation were planned for our patient. the joint was opened through a medial approach, passing between the triceps brachii in the back and brachialis in the front. his ulnar nerve was dissected and protected. accessing the joint capsule required de-insertion of the humeral part of his pronator teres, without interrupting the medial collateral ligament, which was intact. his trochlea was fractured across the front line, with the persistence of the posterior wall. the fragment had moved up and forward without any compaction or loss of cartilage substance5. fig-1: isolated fracture of trochlea and cranial migration of fragment. 6 1department of public health, health services academy, islamabad. 2department of management sciences, riphah int university, islamabad. 3department of medicine, ihitc, islamabad. 4department of rheumatology, pims, islamabad. 5department of medicine, dow university of health sciences, karachi. 70 1department of orthopedics, thq, khanewal 2department of orthopedics, lgh, lahore 3department of orthopedics, thq, ferozwala 4department of orthopedics, cmh, lahore 5department of hand & upper limb surgery, cmh, lahore 6department of orthopedics, al-rehmat benevolent trust 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 21 case report: isolated trochlear fracture of the humerus fig-2: ct scan of elbow localizing the fracture fragment. fig-3: 3d ct scan of elbow showing the isolated trochlea fracture. an orif(open reduction internal fixation) was done through medial approach and fracture was fixed with herbert screws. (fig-4, fig-5) post operatively early range of motion exercises of the elbow was commenced. patient regained nearly full elbow range of movements. fig-4: radiagraph ap-view shows fracture fixation via headless screw. fig-5: radiograph lateral view shows fracture fixation. this case report describes an isolated fracture of the humeral trochlea treated with open reduction internal fixation. radiographic union was present at 13 weeks, and at 20 weeks post-op, the patient had regained full elbow range of motion minus 5 degrees of terminal flexion. open reduction and internal fixation can be performed with success if the trochlear fragment is large enough4. discussion the trochlea is an articular surface of the elbow joint and acts as a pulley on which hinged motion of the ulno-humeral joint occurs. non-operative treatment or excision of coronal shear fractures leads to stiffness or instability of the elbow joint. fracture of the trochlea is usually associated with elbow dislocation and capitellar or medial condylar fracture6. isolated fracture of the humeral trochlea is very rare. this is explained by its position deep within the trochlear notch cavity and the absence of any muscular or ligamentous attachments at this level, which protects it against direct and indirect trauma7. isolated trochlear fractures occur as a result of axial loading when the elbow is in extension, such as a fall on an outstretched hand, and the force transmits through the palm and along the ulna.13 a role of varus stress with axial loading in isolated trochlear fractures is also suggested12. furthermore, the ulno-humeral joint is subjected to very light compressive and shear forces compared to those experienced by the radio-humeral joint, which explains the high frequency of capitellar fractures compared to trochlear fractures8. evaluation of radiographs in the anteroposterior view may show an irregularity at the ulno-humeral joint3, but the image can be interpreted ‘normal’5. in a lateral view, the appearance of an articular 22 case report: isolated trochlear fracture of the humerus half-moon-shaped fragment moved up and forward could suggest a capitellar fracture. for this reason, diagnosis is based on the results of a ct scan3,4. it is unclear what caused the trochlear rather than the entire articular capitello-trochlearblock to fracture. as the anterior part of the trochlea is subjected to the force from the coronoid process of the ulna in a flexed-elbow position, it selectively causes the coronal shear fracture of the trochlea14. the medial approach enables good visualization of the anteromedial aspect of the distal humerus for fixation of trochlear and/or medial epicondylar fractures15. it avoids the need for the more extensive posterior approach combined with olecranon osteotomy. the lateral approach has limited exposure and cannot fix fractures extending into the capitellum. therefore, preoperative computed tomography is essential. in the reported cases, the cancellous screws were passed from the non-articular area. their direction was oblique, from front to back and from medial to lateral, fixing the trochlea to the capitulum. alternatively, herbert screws were inserted into the articular surface buried beneath the cartilage; their direction was perpendicular to the fracture line, securing the fragment of the trochlea to the posterior wall with maximum compression. we opted for this type of osteosynthesis because it is more stable from a biomechanical point of view but the prognosis of coronal trochlear fractures is poor when there is articular cartilage damage (particularly small coronal osteochondral fractures)11. the trochlear fractures can be fixed using the olecranon osteotomy, which many surgeons may be familiar with. the medial approach enables direct access to the trochlear fracture (particularly anterior coronal fractures), preserves the blood supply, and avoids additional surgical morbidity and complications from olecranon osteotomy. the vascular supply to the trochlea is through its posterior surface. thus, the posterior approach may disrupt these vessels and increase the risk of avascular necrosis. in a cadaveric study, trochlear excision leads to considerable instability of the elbow joint12. both the capitellum and trochlea have an important role in stability; excision of the trochlea results in multiplanar instability of the ulno-humeral and radio-capitellar joints. the choice of fixation depends on the size of fragment and the comminution17. in this case we managed the displaced trochlear fracture using single headless herbert screw. herbert and fischer in 1984 introduced the double-threaded design of screws that allows them to sink below the articular cartilage level and provide necessary compression for early mobilization18,19.using the kirschner wires and compression screws with the combination early range of motion exercise, have also borne successful result. conclusion isolated trochlear fractures are uncommon and only appear infrequently in the literature. we advocate open reduction and internal fixation for displaced fractures where prior studies have indicated that patients had a positive outcome after orif. successful outcomes have been recorded in our case report of solitary trochlear fractures with good rom and dash(disability of arm, shoulder, hand) scores. references 1. hotchkiss rn, green dp. fractures and dislocations of the elbow. in: rockwood ca, green dp, bucholz rw, editors. rockwood and green’s fractures in adults. 3rd ed. philadelphia: lippincott; 1991. p. 739–841. 2. dubberley jh, faber kj, macdermid jc, patterson sd, king gj. outcome after open reduction and internal fixation of capitellar and trochlear fractures. j bone joint surg am 2006;88:46–54. 3. isolated fracture of trochlea: a rare case entity, dr.gourabbose international journal of scientific research volume 9 | issue 8 | august – 2020 4. foulk da, robertson pa, timmerman la. fracture of the trochlea. j orthop trauma. 1995;9(6):530-2. doi: 10.1097/00005131199509060-00013. pmid: 8592269. 5. abbassi, n., abdeljaouad, n., daoudi, a. et al. isolated fracture of the humeral trochlea: a case report and review of the literature. j med case reports 9,121(2015).https://doi.org/10.1186/s1325 6-015-0564-1 6. grant ir, miller jh. osteochondral fracture of the trochlea associated with fracture-dislocation of the elbow. injury. 1975 feb;6(3):257-60. doi: 10.1016/0020-1383(75)90118-7. pmid: 1126752. 7. nauth, aaron md, frcsc1; mckee, michael d. md, frcsc1; ristevski, bill md, frcsc1; hall, jeremy md, frcsc1; schemitsch, emil h. md, frcsc1. distal humeral fractures in adults. the journal of bone & joint surgery 93(7):p 686-700, april 6, 2011. | doi: 10.2106/jbjs.j.00845 8. chamseddine a, hamdan h, obeid b, zein h. [articular coronal fractures of the distal humerus]. chirurgie de la main. 2009 dec;28(6):352-362. doi: 10.1016/j.main.2009.08.009. pmid: 19811942 23 case report: isolated trochlear fracture of the humerus 9. bryan rs. fractures about the elbow in adults. instr course lect1981;30:200–23. 10. ring d, jupiter jb, gulotta l. articular fractures of the distal part of the humerus. j bone joint surg am 2003;85:232–8 11. ashwood n, verma m, hamlet m, garlapati a, fogg q. transarticular shear fractures of the distal humerus. j shoulder elbow surg 2010;19:46–52. 12. nakatani t, sawamura s, imaizumiy, sakurai a, fujioka h, tomioka m, et al. isolated fracture of the trochlea: a case report. j shoulder elbow surg 2005;14:340–3. 13. worrell rv. isolated, displaced fracture of the trochlea. n y state j med 1971;71:2314–5. 14. kwan mk, khoo eh, chuayp, mansor a. isolated displaced fracture of humeral trochlea: a report of two rare cases. inj extra 2007;28:461–5. 15. kaushal r, bhanot a, gupta pn, bahadur r. isolated shear fracture of humeral trochlea. inj extra 2005;36:210–1. 16. sabo mt, fay k, mcdonald cp, ferreira lm, johnson ja, king gj. effect of coronal shear fractures of the distal humerus on elbow kinematics and stability. j shoulder elbow surg 2010;19:670– 80 17. w. osman, m. braiki, z. alaya, n. naouar, m. ben ayeche, combined isolated laugier’s fracture and distal radial fracture: management and literature review on the mechanism of injury, case rep. orthop. 2016 (figure 2) (2016) 1–6. 18. l.j. zimmerman, j.j. jauregui, c.e. aarons, isolated shear fracture of the humeral trochlea in an adolescent: a case report and literature review, j.pediatr.orthop.b 24 (5) (2015) 412–417. 19. a.p. singh, i.k. dhammi, a.k. jain, s. jain, neglected isolated fracture of the trochlea humeri, chin.j.traumatol.engl.ed. 13 (4) (2010) 247–249, https://doi.org/10.3760/cma.j.issn.10081275.2010.0 4.011 the authors: dr. shehzad anwer, consultant orthopedic surgeon, department of orthopedic surgery, general hospital, lahore, pakistan. dr. shuja uddin, consultant orthopedic surgery, department of orthopedic surgery, general hospital, lahore, pakistan. dr. hasnain khalid, registrar orthopedic surgery, department of orthopedic surgery, general hospital, lahore, pakistan. dr. rizwan ul haq, consultant orthopedic surgery, department of orthopedic surgery, general hospital, lahore, pakistan. dr. adeel hamid, consultant orthopedic surgery, department of orthopedic surgery, general hospital, lahore, pakistan. mubashir farhan, consultant orthopedic surgery, department of orthopedic surgery, general hospital, lahore, pakistan. dr. zia uddin, consultant orthopedic surgery, department of orthopedic surgery, general hospital, lahore, pakistan. dr. mian muhammad haneef, professor oforthopedics, department of orthopedic surgery, general hospital, lahore, pakistan. corresponding author: dr. shuja uddin, consultant orthopedic surgery, department of orthopedic surgery, general hospital, lahore, pakistan. email:drshujauddin111@gmail.com numbering.indd 4 proceedings s.z.m.c. vol: 34(4): pp. 4-9, 2020. pszmc-763-34-4-2020 detection of helicobacter pylori through histochemistry & immunofluorescent staining in biopsies of patients with chronic gastritis asia jahanzeb, abdul hannan nagi department of morbid anatomy & histopathology, university of health sciences, lahore abstract introduction: helicobacter pylori (h.pylori) is a helix shaped gram negative rod which is usually associated with chronic gastritis and also a major cause of other gastroduodenal diseases as well. aims & objectives: the present study used histochemical and immunofluorescent stains on formalin fixed paraffin embedded human gastric biopsies for detection of h.pylori. comparison was also done to evaluate best staining method. place and duration of study: this study was accomplished in about one year. sampling of gastric biopsies and rapid urease test were executed at the endoscopy suite of lahore general hospital whereas the histopathological examination and immunofluorescent staining were done in university of health sciences, lahore. material & methods: thirty patients (n=30cases) were included in the study following inclusion criteria. diagnostic upper gi endoscopy was carried out in all cases. five gastric biopsies were taken from each patient/case (total n=150 biopsies) according to the updated sydney system. rapid urease test was performed at the site of endoscopy. biopsies fixed in 10% formalin were brought to the concerned department where they were assigned a specific laboratory number then processed and stained. results: endoscopic examination revealed chronic gastritis and rapid urease tests were positive. all cases (n=30) were positive for h.pylori on histopathology. the calculated sensitivity and specificity of h&e, giemsa, modified mcmullen’s stain and immunofluorescent method in present study were 71% and 100%,83% and 100%, 82% and 100%, 90% and 100% respectively. conclusion: special stains makes h.pylori identification easier in tissue sections. however, immunofluorescent test is the most sensitive and specific method as compared to histochemical stains. key words: helicobacter pylori, chronic gastritis, rapid urease test, immunofluorescent stain. introduction in 1982, h.pylori was first discovered by drs. barry marshall and robin warrens in gastric mucosa of patients having gastritis and ulcers.1 h.pylori causes chronic gastritis and has been related with other gastroduodenal diseases including peptic ulcer diseases (pud), gastric and duodenal ulcers, mucosa associated lymphoid tissue (malt) lymphoma and even gastric carcinoma.2 more than 50% of patients reveals h.pylori upon endoscopy as humans are the principle host and it predominantly inhabits human gastric mucosa that leads to inflammation and other pathological change.3 numerous studies have reported different incidence rates of h.pylori infection that varies broadly due to topographical area, age, race, culture and living standards. infections are more common in developing countries due to poor living and sanitary conditions.4 so penurious living conditions, poor hygiene, overpopulation and genetic susceptibility are the possible risk factors of h.pylori infection. presence of h.pylori dna in saliva, vomitus, gastric juices, feces and water shows its diverse transmission sources.4 presence of virulence genes of h.pylori in different samples of drinking water is also reported from pakistan.5 in pakistan, the prevalence of h.pylori infection is also high like other developing countries and reported in the range of 50–90%.6 several h.pylori virulence factors like enzymes, toxins and genetic factors are known that are involved in the pathogenesis of diseases.7 cytotoxin associated gene (caga) of h.pylori is potentially considered as a carcinogen.8 virulent h.pylori strains elicits more powerful inflammatory response and are related with increased risk of gastric carcinoma.9 5 detection of helicobacter pylori through histochemistry & immunofluorescent staining in biopsies of patients various invasive and non-invasive tests are available for h.pylori diagnosis.10 endoscopy is the most frequently practiced invasive procedure today and histological examination provides additional information ranging from inflammatory to malignant conditions.11 so, histochemical stains and immunofluorescent (anti h.pylori) method were used in study for detection of helicobacter pylori. statistical analysis was done to evaluate the most sensitive and specific method. material and methods after approval by ethical review committee uhs/education letter no. 126-17/863. this diagnostic study was conducted in uhs during period from june 2017 to july 2018. convenient sampling technique was used. data, history and gastric biopsy of adult dyspeptic patients admitted to lahore general hospital were taken after obtaining written consent. exclusion criteria: patients with co-morbid conditions (e.g. hepatitis b, c), patients undergoing antibiotic or proton pump inhibitor (ppi) treatment were excluded. inclusion criteria: thirty patients (n=30) of both gender and age range of 18-45 years reporting for the primary diagnosis of the disease were enrolled in present study. endoscopy: it was done in lahore general hospital by gastroenterologist using olympus cv-190 video endoscope. five biopsies (two each from antrum and corpus and one from incisura angularis) were taken from 30 patients (total n=150 biopsies).11 in present study two biopsies (one each from antral and corpus (greater curvature) were used for the rapid urease test in each patient and other biopsies were fixed in 10% formalin solution for histopathological examination. the biopsy sections were stained with h&e, giemsa and modified mcmullen’s stain.12,13 grading of gastritis was done following updated sydney classification. histopathological variables like activity (neutrophil infiltration), chronic inflammation (mononuclear cell infiltration), atrophy (loss of normal glands from antrum or corpus) and gastric intestinal metaplasia (replacement of gastric epithelium by intestinal type) were also documented that also defined the prognosis of gastric infection in study population.14 immunofluorescent staining: polyclonal anti h.pylori antibody conjugated with fitc (fluorescein iso-thiocyanate conjugate) was used on formalin fixed paraffin embedded gastric biopsies.15 fluorescence microscope (olympus bx51), model u-lh1oohg was used for microscopy and images were captured. fitc usually gives apple green fluorescence that locates the antigen in tissue.16 statistical analysis: the data was analyzed using spss (20.0). mean and standard deviation were considered for quantitative variables in addition to the frequencies along with percentages for qualitative variables. categorical variables were investigated by using pearson’s chisquare test. a p-value less than 0.05 (p<_0.05) was taken significant. comparison of staining methods was done and statistical sensitivity, specificity, negative predictive value, positive predictive value and diagnostic accuracy were also calculated.17 results mean age of the population was 34±8. the gender distribution showed that females were predominant that is n=17 (56.7%) and n=13 (43.3%) were males. the female to male ratio was 1.3:1. the mean age of males and females were 29.92±7.6 and 36.12±7.8 years respectively. regarding localization and distribution of gastritis, the most common gastritis on histopathology was antral gastritis n=16 (53.3%), antral predominant pan gastritis and corpus predominant pan gastritis was seen in n=11 (36.7%) and n=3 (10%) of the cases respectively.(table 1) clinic endoscopic features n %age age 18-30years 9 30% 31-45 years 21 70% hyperemia 26 86.7% erosion 1 3.3% ulceration 1 3.3% nodularity 0 0 co-exiting duodenitis 2 6.7% location of endoscopic gastritis -antral gastritis 16 53.3% -corpus predominant 3 10% -antral predom pangastritis 11 36.7% normal gastric mucosa 0 0% table-1: distribution of cases with respect to clinical and endoscopic data 6 detection of helicobacter pylori through histochemistry & immunofluorescent staining in biopsies of patients morphological variables g0 (nil) grades g1 (mild) g2 (moderate) g3 (marked) h. pylori density n = 0 none n = 4 1-3 bacteria n=11 layer of bacteria n=15n clusters activity (neutrophil infiltrate ) n =18 60% n = 12 40% n=o n=0 chronic inflammation (lymphocytic infiltrate ) n = 0 n = 4 13.3% n=11 37% n=15 50% glandular atrophy n=27 (90%) n=0 n=1 n=2 6.6% gastric intestinal metaplasia n=30 (100%) n=0 n=0 n=0 table-2: grading of histopathological variables of antral and corporal mucosa according to the updated sydney system. go=no inflammation, g1=mild inflammation, g2=moderate inflammation, g3 marked inflammation. h.pylori density = mild, moderate and marked number of h.pylori overlying epithelium. density was observed in all n=30 cases of chronic gastritis. cases of g1 and g2 grades of chronic gastritis showed low density of bacteria while n=15 cases of g3 showed high density. g3 i.e. marked chronic gastritis was the most commonly observed grade of chronic inflammation. the grade of activity was mild for n=12 (40%) cases. however no activity was observed in n=18 (60%) of the cases. the finding of chronic inflammation was observed in all n=30 (100%) cases of which n=15 (50%) cases had marked chronic inflammation, n=11 (36.7%) cases had moderate chronic inflammation and n=4 (13.3%) cases had mild chronic inflammation. histologically n=2 (6.6%) cases of marked chronic inflammation revealed atrophy but no gastric intestinal metaplasia was observed (table-2). staining sensitivity a/a+c x100 specificity d/b+dx 100 ppv a/a+b x100 npv d/d+c x100 accuracy a+d/a+b +c+dx100 h&e 71% 80 % 98 20 66% giemsa 83% 100% 100 14.2 84% modified mcmullen’s 82% 100% 100 16.6 83% immunoflu orescent 90% 100% 100 0.00 90% table-3: statistical analysis fig-1: rapid urease test, strong positive result, test paper turns from yellow to cherry red fig-2: h&e shows h.pylori in gastric pits fig-3: h&e chronic mild antral gastritis 7 detection of helicobacter pylori through histochemistry & immunofluorescent staining in biopsies of patients fig-4: h&e chronic moderate antral gastritis fig-5: h&e chronic marked antral gastritis fig-6: h.pylori on giemsa staining 40x fig-7: h.pylori in gastric pits on modified mcmullen’s stain 40x fig-8: immunofluorescent stain (apple green fluorescence) discussion h.pylori induced gastritis is the most common gastric infection that fluctuates in diverse areas of world depending on several factors that effects its acquisition, transmission and pathogenesis. the prevalence in our study came out to be 100% that was relatively higher as compared to the previous study reported.6 this was probably due to small sample size of our study in which we had included patients whose severe clinical symptoms were suggestive of chronic gastritis. females were predominant in present study that was contrary to previous studies that showed the predominance of males. this can be related to low educational status of females in pakistan and increased exposure to risk factors like domestic hygiene, large number of siblings with crowded living conditions and poor sanitation. zoonotic and vector borne transmission 8 detection of helicobacter pylori through histochemistry & immunofluorescent staining in biopsies of patients could be a source but considered the lowest risk factor.4,6 the mean age in our study population was 34±8.4 that was less compared to previous studies.18 this was due the inclusion criteria of our study i.e. the age range was 18-45 years and patients under 18 years and above 50 years of age were excluded. however, the relation of infection with age cannot be explained and justified in present study because pediatric and elderly patients were not incorporated in this study. the clinical symptoms presented by patients were in accordance to the previous studies reported both locally and internationally.19 rapid urease test was performed immediately after endoscopy using commercially available kits (lituo biotech) and observed to be positive in all cases n=30 (100%). these findings were consistent with both local and international studies.20 cases were also evaluated on basis of yield of h.pylori at various anatomical sites and it was found that probability of finding h.pylori was highest at the antrum. in present study g3 i.e. marked chronic gastritis was the most commonly observed grade of chronic inflammation probably because patients had the symptoms of gastritis since 8-9 years. activity (active inflammation) was seen in n=12 (40%) cases, a finding which is accordant to both local and international studies.6 gastric intestinal metaplasia (gim) cases were not found in present study that represented very low risk of developing gastric carcinoma in our study population due to small sample size of our study but a large number of population would be needed to assess the risk of gastric cancer. the sensitivity and specificity of h&e staining were found to be 71% and 80 % respectively that were comparable to previous studies.11 (table3). the sensitivity and specificity of giemsa staining were ā83% and 100% respectively and compatible with the previous studies.11 the sensitivity and specificity of modified mcmullen’s stain were 82% and 100 % respectively and compatible with previously done researches.12,13 statistical analysis in present study showed that immunofluorescent test was the most sensitive and specific test for h.pylori identification as the p value was less than 0.05,x2 (0.05)=1.16. (table3) the calculated sensitivity and specificity of immunofluorescent test in our study were 90% and 100% respectively probably due to strongly positive cases of h.pylori, but compatible with previous study.15 the antibody (polyclonal anti h. pylori antibody) used in immunofluorescent assay did not react with the closely matched bacteria present in gastric mucosa thus detected even low concentration of an antigen in tissue as in corpus biopsies where the yield of bacteria was usually low. in this study no false positive cases were revealed due to strongly positive cases of h.pylori induced chronic gastritis. conclusion h.pylori associated chronic gastritis is an appalling issue in our country which needs to be noticed seriously. modified mcmullen’s stain can be used in pathological laboratories like giemsa stain for detection of h.pylori due to its better contrast. immunofluorescent staining is the most sensitive and specific method but it is not being used in pathological laboratories commonly due to its high cost and complexity. it should be restricted to research based studies. every pathological laboratory desires to use cheap and simple method in routine practice with much emphasis on time/ labor cost. so, giemsa stain is still the most economical and convenient staining method that would help as good as immunofluorescent method in our setting where resources are limited. limitations of the study: disposable biopsy forceps, which is not routinely practiced in our hospitals, were expensive. immunofluorescent assay was also an expensive and time consuming procedure that’s why due to budget and time we were restricted to smaller sample size. so, a project on higher scale would be more persuasive and statistically important to access the efficacy of endoscopic biopsies, histochemical stains and immunofluorescent assay as well as its diagnostic and prognostic role in gastric infections. acknowledgement: we are thankful to dr. nadia naseem, head department of morbid anatomy and histopathology, uhs lahore. references 1. marshall bj, warren jr. unidentified curved bacilli in the stomach of patients with gastritis and peptic ulceration. lancet (london, england). 1984; 1(8390):1311-5. 2. konturek jw. discovery by jaworski of helicobacter pylori and its pathogenetic role in peptic ulcer, gastritis and gastric cancer. journal of physiology and pharmacology : an official journal of the polish physiological society. 2003; 54 suppl 3:23-41. 9 detection of helicobacter pylori through histochemistry & immunofluorescent staining in biopsies of patients 3. mcnulty ca. campylobacter pyloridisassociated gastritis. the journal of infection. 1986; 13(2):107-13. 4. brown lm. helicobacter pylori : epidemiology and routes of transmission. epidemiologic reviews. 2000; 22(2):283-97. 5. samra zq, javaid u, ghafoor s, batool a, dar n, athar ma. pcr assay targeting virulence genes of helicobacter pylori isolated from drinking water and clinical samples in lahore metropolitan, pakistan. journal of water and health. 2011; 9(1):208-16. 6. mehmood k, awan aa, muhammad n, hasan f, nadir a. helicobacter pylori prevalence and histopathological findings in dyspeptic patients. journal of ayub medical college, abbottabad : jamc. 2014; 26(2):182-5. 7. kusters jg, van vliet ah, kuipers ej. pathogenesis of helicobacter pylori infection. clinical microbiology reviews.2006;19(3):449-90 8. hatakeyama m. helicobacter pylori caga and gastric cancer: a paradigm for hit-and-run carcinogenesis. cell host & microbe. 2014; 15(3):306-16. 9. wroblewski le, peek rm, jr., wilson kt. helicobacter pylori and gastric cancer: factors that modulate disease risk. clinical microbiology reviews. 2010; 23(4):713-39. 10. best lm, takwoingi y, siddique s, selladurai a, gandhi a, low b, et al. non-invasive diagnostic tests for helicobacter pylori infection. the cochrane database of systematic reviews. 2018; 3:cd012080. 11. lee jy, kim n. diagnosis of helicobacter pylori by invasive test: histology. annals of translational medicine. 2015; 3(1):10. 12. rotimi o, cairns a, gray s, moayyedi p, dixon mf. histological identification of helicobacter pylori: comparison of staining methods. journal of clinical pathology. 2000; 53(10):756-9. 13. lwaki h, sugiyama t, asaka m. a modified mcmullen's staining for helicobacter pylori: a high-contrast, visibly prominent method. helicobacter. 1998; 3(1):45-8. 14. stolte m, meining a. the updated sydney system: classification and grading of gastritis as the basis of diagnosis and treatment. canadian journal of gastroenterology = journal canadien de gastroenterologie. 2001; 15(9):591-8. 15. rivera e, lópez-vidal y, luqueño v, ruizpalacios gm. indirect immunofluorescence assay for detection of helicobacter pylori in human gastric mucosal biopsies. j clin microbiol. 1991; 29(8):1748-51. 16. makki js. diagnostic implication and clinical relevance of ancillary techniques in clinical pathology practice. clin med insights pathol. 2016; 9:5-11. 17. baratloo a, hosseini m, negida a, el ashal g. part 1: simple definition and calculation of accuracy, sensitivity and specificity. emerg (tehran). 2015; 3(2):48-9. 18. yakoob j, abbas z, jafri w, usman mw, jafri f, awan s. comparison of the virulence markers of helicobacter pylori and their associated diseases in patients from pakistan and afghanistan. saudi j gastroenterol. 2013; 19(5):211-8. 19. suzuki h, moayyedi p. helicobacter pylori infection in functional dyspepsia. nature reviews gastroenterology & hepatology. 2013; 10(3):168-74. 20. uotani t, graham dy. diagnosis of helicobacter pylori using the rapid urease test. annals of translational medicine. 2015; 3(1):9. the authors: asia jahanzeb dept. of morbid anatomy & histopathology, university of health sciences, lahore. prof. abdul hannan nagi former head, dept. of morbid anatomy & histopathology, university of health sciences, lahore. corresponding author: asia jahanzeb dept. of morbid anatomy & histopathology, university of health sciences, lahore. e-mail: asaiarao1000@gmail.com full book for web 18 proceedings s.z.m.c. vol: 36(3): pp. 18-24, 2022. pszmc-847-36-3-2022 department of general surgery, sir ganga ram hospital, lahore. 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 19 shock index in patients undergoing elective and emergency general surgical procedures… 20 shock index in patients undergoing elective and emergency general surgical procedures… 21 shock index in patients undergoing elective and emergency general surgical procedures… 22 shock index in patients undergoing elective and emergency general surgical procedures… 23 shock index in patients undergoing elective and emergency general surgical procedures… 24 shock index in patients undergoing elective and emergency general surgical procedures… full book 31 proceedings s.z.m.c. vol: 36(4): pp. 31-36, 2022. pszmc-861-36-4-2022 1department of physiology, shalamar medical and dental college, lahore 2department of physiology, fatima jinnah medical university, lahore 3fatima jinnah medical university, lahore 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 32 frequency of overweight and obesity and its associated factors among school children in lahore pakistan 33 frequency of overweight and obesity and its associated factors among school children in lahore pakistan 34 frequency of overweight and obesity and its associated factors among school children in lahore pakistan 35 frequency of overweight and obesity and its associated factors among school children in lahore pakistan 36 frequency of overweight and obesity and its associated factors among school children in lahore pakistan full book for web 65 proceedings s.z.m.c. vol: 36(3): pp. 65-69, 2022. pszmc-855-36-3-2022 s.z.m.c. vol: 36(3): pp. 65-69, 2022. pszmc-855-36-3-2022 department of surgery, ayub teaching hospital, abbottabad. department of medicine, women medical college, dhq teaching hospital, abbottabad. institute of kidney diseases, peshawar. dhq hospital haripur. 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 66 comparison between lateral internal sphincterotomy & lord’s operation for chronic anal fissure 67 comparison between lateral internal sphincterotomy & lord’s operation for chronic anal fissure 68 comparison between lateral internal sphincterotomy & lord’s operation for chronic anal fissure 69 comparison between lateral internal sphincterotomy & lord’s operation for chronic anal fissure 05 26 proceedings s.z.m.c. vol: 36(4): pp. 26-30, 2022. pszmc-860-36-4-2022 1department of bds, islamabad medical and dental college, islamabad 26 proceedings s.z.m.c. vol: 36(4): pp. 26-30, 2022. pszmc-860-36-4-2022 1department of bds, islamabad medical and dental college, islamabad department of oral and maxillofacial surgery, 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 27 perception, attitude & barriers amongst dentists using digital and electronic technology 26 proceedings s.z.m.c. vol: 36(4): pp. 26-30, 2022. pszmc-860-36-4-2022 1department of bds, islamabad medical and dental college, islamabad 28 perception, attitude & barriers amongst dentists using digital and electronic technology 29 perception, attitude & barriers amongst dentists using digital and electronic technology department of oral and maxillofacial surgery 30 perception, attitude & barriers amongst dentists using digital and electronic technology department of oral and maxillofacial surgery department of oral and maxillofacial surgery department of oral and maxillofacial surgery department of oral and maxillofacial surgery department of oral and maxillofacial surgery department of oral and maxillofacial surgery full book for printing 35 proceedings s.z.m.c. vol: 37(2): pp. 35-41,2023. pszmc-884-37-2-2023 1department of radiology, dow university of health sciences. 2department of radiology, gambat institute of medical sciences, gambat. 3department of general medicine, northwest general hospital and research centre, peshawar. 4department of clinical investigation, maroof international hospital. 70 1department of orthopedics, thq, khanewal 2department of orthopedics, lgh, lahore 3department of orthopedics, thq, ferozwala 4department of orthopedics, cmh, lahore 5department of hand & upper limb surgery, cmh, lahore 6department of orthopedics, al-rehmat benevolent trust 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 36 efficacy of pigtails for the management of refractory malignant ascites or effusion: a systematic….. 37 efficacy of pigtails for the management of refractory malignant ascites or effusion: a systematic….. 38 efficacy of pigtails for the management of refractory malignant ascites or effusion: a systematic….. 39 efficacy of pigtails for the management of refractory malignant ascites or effusion: a systematic….. 40 efficacy of pigtails for the management of refractory malignant ascites or effusion: a systematic….. am j emerg med 41 efficacy of pigtails for the management of refractory malignant ascites or effusion: a systematic….. full book for web 30 proceedings s.z.m.c. vol: 36(3): pp. 30-35, 2022. pszmc-849-36-3-2022 1department of radiology, shaikh zayed hospital, lahore. 2department of obstetrics and gynecology, shaikh zayed hospital, lahore. 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 31 value of transvaginal sonography in antenatal management of ectopic pregnancy 32 value of transvaginal sonography in antenatal management of ectopic pregnancy 33 value of transvaginal sonography in antenatal management of ectopic pregnancy 34 value of transvaginal sonography in antenatal management of ectopic pregnancy 35 value of transvaginal sonography in antenatal management of ectopic pregnancy full book for web 13 proceedings s.z.m.c. vol: 36(3): pp. 13-17, 2022. pszmc-846-36-3-2022 1department of radiology, holy family hospital, rawalpindi. 2department of radiology, rawalpindi institute of cardiology, rawalpindi. 3department of radiology, advanced international hospital, islamabad. 4 maroof international hospital, islamabad. 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 14 comparative diagnostic accuracy of sonographic strain elastography and fnac in breast lesions 15 comparative diagnostic accuracy of sonographic strain elastography and fnac in breast lesions 16 comparative diagnostic accuracy of sonographic strain elastography and fnac in breast lesions 17 comparative diagnostic accuracy of sonographic strain elastography and fnac in breast lesions full book for web 50 proceedings s.z.m.c. vol: 36(3): pp. 50-54, 2022. pszmc-852-36-3-2022 haidan hospital, saadah, yemen. 2al-sabeen hospital, sana’a, yemen. al-thawrah hospital, sana’a, yemen. 4al-alya’a hospital, sana’a, yemen. 5doctors without borders, amran, yemen. 6faculty of medicine sana’a university, yemen. 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 51 quality of care for pediatric leukemia patients at a hospital in sana’a as perceived by patients’ attendants 52 quality of care for pediatric leukemia patients at a hospital in sana’a as perceived by patients’ attendants 53 quality of care for pediatric leukemia patients at a hospital in sana’a as perceived by patients’ attendants 54 quality of care for pediatric leukemia patients at a hospital in sana’a as perceived by patients’ attendants full book for printing 23 proceedings s.z.m.c. vol: 37(2): pp. 23-28, 2023. pszmc-882-37-2-2023 1department of medical education, university college of medicine & dentistry, lahore. 2department of school of health professions' education, medical college and institute of dentistry cmh, lahore. 3department of physiology, university college of medicine & dentistry, lahore. 70 1department of orthopedics, thq, khanewal 2department of orthopedics, lgh, lahore 3department of orthopedics, thq, ferozwala 4department of orthopedics, cmh, lahore 5department of hand & upper limb surgery, cmh, lahore 6department of orthopedics, al-rehmat benevolent trust 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 24 qualities of pbl leader: the students’ perspective 25 qualities of pbl leader: the students’ perspective 26 qualities of pbl leader: the students’ perspective 27 qualities of pbl leader: the students’ perspective "it is said that 2 heads are better than one". team is a group of individuals working together in order to attain a common goal and to achieve their desired objectives11. emotional intelligence is the ability to observe between the one's own and others' feelings and emotions, to discriminate among them and use this information to guide one's thinking and actions12. “what goes around comes around” “leader’s behavior of asserting strong authority and control over subordinates and demanding unquestioned loyalty, submission and obedience from them15. 28 qualities of pbl leader: the students’ perspective for web full book 24 proceedings s.z.m.c. vol: 37(3): pp. 24-27 , 2023. pszmc-892-37-3-2023 an indigenous study on the palmaris longus muscle anatomical variations 1alvia batool, 2faizia batool, 1afifa waseem, 1saira munawar, 1maryam fatima 1department of anatomy, fmh college of medicine and dentistry, lahore. 2department of dermatology, arif memorial hospital, lahore. abstract introduction: the palmaris longus muscle is one of the superficial flexor muscles of the forearm that flexes the hand on the wrist and makes the palmar aponeurosis tense for gripping. it is variable both in number and form like inverted, doubled, split, three-headed or absent palmaris longus. these variabilities may cause clinical features due to abnormal pressure on the median and ulnar nerves. its tendon can be used as a graft in various transplant surgeries. aims & objectives: this study was conducted to observethe number of cadavers showing anatomical variations of palmaris longus muscle in the pakistani population. place and duration of study: this study was conducted in the anatomy department of fmhcm&d, lahore. the duration of the study was january 2019 january 2022. material & methods: cadaveric dissected upper limb specimens with normal palmaris longus were listed in group: a, upper limbs having reversed palmaris longus were listed in group: b, while upper limbs with absent palmaris longus were listed in group: c. the rpl (reversed palmaris longus) muscle is the structure that is tendinous proximally and muscular distally (opposite of normal palmaris longus).results were tabulated and percentages calculated. results: it was observed that out of 80 dissected cadaveric upper limbs,77 (96.25%) showed normal palmaris longus while a right cadaveric upper limb (1.25%) showed anomalous palmaris longus with the reversed tendon. palmaris longus muscle was found absent in two cadaveric left upper limbs (2.5%). conclusion: our study highlighted the percentage of different anatomical variations of the palmaris longus muscle in terms of its presence with reversed tendons as well as its absence in a few human cadaveric upper limbs. this information may help anatomists, radiologists, physiotherapists and plastic surgeons. keywords: palmaris longus, anatomical variations, reversed tendon introduction palmaris longus is a thin spindle-shaped muscle present midway between the flexor carpi radialis and flexor carpi ulnaris muscles in the superficial muscle group of the anterior fascial compartment of the forearm.1its nerve supply is the median nerve (c7, c8). it has a common origin from the anterior aspect of the medial epicondyle of the humerus. it has a small muscle belly and an elongated strandlike tendon that passes just in front of the flexor retinaculum and joins it and the tip of the palmar aponeurosis.1,2,3 in the palm, it breaks into lengthwise running fibers of the palmar aponeurosis. the palmaris longus tendon is helpful in finding the median nerve at the wrist. here it is located a little bit medial or in front of the median nerve. it is a weak flexor of the hand at the wrist and makes the palmar aponeurosis tense. it also attaches the skin and fascia of the hand during clasping. its tendon is useful as a graft in transplant surgeries.1,2normally, the tendon can be noticed and explored through touch when the wrist is flexed and the tips of the thumb and little finger are in opposition1,2,3. the palmaris longus muscle shows remarkable anatomical variabilities in contrast with other muscles of the upper limb. the most commonly noticeable variability is its deficiency. in 10-14% of individuals, it might not be present on one or both sides, most commonly on the left1,2,3. the muscle belly of palmaris longus might be present midway or distant in some individuals instead of a proximate one that is normal. the muscle is comparatively fragile that’s why its non-existence does not create any dysfunction1. it varies both in quantity and appearance namely duplicated, bifid, three-headed, reversed, or hypertrophied palmaris longus tendon. these anomalies may cause median and ulnar nerve compression with inflammation of the anterior side of the wrist4,5. the rpl reversed palmaris longus muscle is the structure that is tendinous proximally and muscular distally (opposite of normal palmaris longus).palmaris longus is used as a transplant in the surgical relocation of a tendon for correcting 6 1department of public health, health services academy, islamabad. 2department of management sciences, riphah int university, islamabad. 3department of medicine, ihitc, islamabad. 4department of rheumatology, pims, islamabad. 5department of medicine, dow university of health sciences, karachi. 70 1department of orthopedics, thq, khanewal 2department of orthopedics, lgh, lahore 3department of orthopedics, thq, ferozwala 4department of orthopedics, cmh, lahore 5department of hand & upper limb surgery, cmh, lahore 6department of orthopedics, al-rehmat benevolent trust 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 25 an indigenous study on the palmaris longus muscle anatomical variations droopy eyelids while doing the cosmetic procedure to increase lip fullness and in plastic surgeries because of its adequate dimensions and convenient picking without causing any harm to the benefactor’s area6,7,8. the aim of this study is to highlight various anatomical variations of the palmaris longus muscle in the pakistani population. this knowledge would be essential not only for anatomists, radiologists, and physiotherapists but for plastic surgeons as well. material and methods a study on cadaveric upper limbs fixed in 10% formalin was conducted in the anatomy department offmh college of medicine & dentistry, lahore, from january 2019 to january 2022 to investigate the presence of normal palmaris longus and palmaris longus with anatomical variations. after getting approval of the institutional review board vide ref letter no. fmh-12-2020-irb-841-m. cadavers of both sexes with the age group ranging from 40 to 60 years were included in this study. the upper limbs that were noted with visible trauma were excluded. this study was designed after an accidental finding of palmaris longus muscle with an inverted tendon in the right upper limb of a male cadaver during routine dissection of its flexor compartment of the forearm using standard procedure.9his occupational and medical records were unknown. median and ulnar nerves were found normal in thickness. the right and left upper limbs of all cadavers were observed for the presence of palmaris longus muscle with anatomical variabilities. the upper limbs that were found to have palmaris longus muscles with normal tendons were noted in group a, while the upper limbs having palmaris longus with inverted tendons were listed in group b and the upper limbs with absent palmaris longus muscles were noted in group c.the groups, laterality, numbers and their percentages are presented in table-1. results out of 80 dissected cadaveric upper limb specimens (40 right and 40 left upper limbs), 77(96.25%) were showing palmaris longus with the normal tendon of insertion fig-1. 01(1.25%) right upper limb of a cadaver out of 80 dissected cadaveric upper limb specimens was having reversed palmaris longus. the origin of reversed palmaris longus was found tendinous from the medial epicondyle of the humerus and then the musculotendinous junction was seen and the muscular belly was inserted into the ventral aspect of the wrist fig-2. 02(2.5%) left upper limbs of two cadavers out of 80 dissected cadaveric upper limb specimens were deficient in palmaris longus muscle fig-3. results were tabulated, percentages were calculated and represented in the form of table-1. groups side number of upper limb specimens percentage of variations right left a (upper limb specimens with normal palmaris longus) 39 38 77 96.25% b (upper limb specimen with reversed palmaris longus rpl) 01 01 1.25% c (upper limb specimens with absent palmaris longus) 02 02 2.5% total 40 40 80 100% table-1: showing percentage of upper limb specimens with normal and anatomically variable palmaris longus muscle fig-1: showing normal palmaris longus muscle in the cadaveric left upper limb specimen. fig-2: showing reversed palmaris longus rpl in the cadaveric right upper limb specimen. 26 an indigenous study on the palmaris longus muscle anatomical variations fig-3: showing absence of palmaris longus in the cadaveric left upper limb specimen. discussion palmaris longus exhibits different variabilities in its number, morphology and location. in the present study of variations of palmaris longus muscle in cadaveric upper limb specimens, it was found normal in 77(96.25%) of cases, right-sided palmaris longus muscle with reversed tendon was noted in1(1.25%) and in 2(2.5%) of the left-sided upper limbs it was found absent. reiman et al.,10studied the upper limbs of 1600 cadavers for anomalies of the palmaris longus muscle for the first time. he noticed that the occurrence of its absence was 12.9% and the incidence of other anomalies was 9% such as the muscle belly of palmaris longus may be present in the middle or distally placed or there may be two bellies10.these findings support the present study results. rajesh s et al.,11studied cadaveric upper limbs and observed that palmaris longus was normal in 45 (90%) upper limbs, reversed was found in two upper limbs (4%), and its agenesis was noted in two (4%) upper limbs. the results of this study are more or less close to the present study results as both variations like reversal and absence of palmaris longus were noted in the same percentage. this information about this muscle is not only important for radiologists to diagnose wrist swellings with involvement of forearm nerves but for surgeons in grafting procedures of limbs like tendon transfer for first dorsal interosseous muscle, lumbrical replacement, reconstruction of tendo-calcaneus and glottis11. sunitha r et al.,12observed that palmaris longus was absent in one male cadaver on both sides and its agenesis was found in one female cadaver on the right side. total absence of palmaris longus was 5%. right-sided absence is 3.3% and left-sided absence is 1.6%. reversed belly of the right-sided palmaris longus in a male cadaver was also noted. the results of this study are consistent with the present study findings. the presence of reversed palmaris longus may produce the most likely clinical features of carpal tunnel syndrome or guyon’s canal syndrome due to the involvement of the median nerve or ulnar nerve respectively. the knowledge of the variability of palmaris longus is essential for surgeons, clinicians, orthopedic surgeons, anatomists, researchers and interventional radiologists12. gune ar et al.,13 found a case with bilateral reversed palmaris longus muscle with insertion on three different sites, first in fascia covering the thenar muscles, second into palmar aponeurosis and third with abductor digiti minimi. reversed palmaris longus can compress the median nerve or ulnar nerve and may cause compartment syndrome leading to swelling and pain in the wrist. using this knowledge, a surgeon can use it as a graft while doing procedures on the wrist region and hands13. lalit m et al.,14 while teaching undergraduate students, noticed the right forearm of a 54 years old male cadaver having a reversed palmaris longus with the tendinous origin and muscular belly with two slips near insertion. this muscle is of great interest to surgeons because of its pivotal role in repairing collateral ligaments and in the treatment of facial paralysis and other nerve palsies14. hashem et al noted a patient with right-sided distal forearm painful swelling on the ventral side due to rpl affecting the median nerve that produced clinical features of median nerve involvement. ultrasonography was done to see the painful wrist mass and then magnetic resonance imaging was done to know the details of this mass. he was treated with anti-inflammatory medicines; wrist supports and physiotherapy15. demir ci et al.,16 observed a case of reversed palmaris longus with clinical features of median nerve compression. a female patient of 16 years of age was reported for the presence of swelling in front of the wrist due to rpl muscle that was causing compression of the median nerve leading to the feeling of pins and needles in the first three fingers of the left hand and an increase in severity of the feeling was noted while doing some hand activity. the clinical tests showed the presence of reversed palmaris longus muscle. ultrasonography and then mri highlighted the rpl muscle along with its attachments. while making the diagnosis of various swellings at the wrist, the rpl muscle may be one of the causes that should also be kept in consideration16. waghray net al.,17the palmaris longus was found absent in left upper limb of a 58 years old cadaver. it was concluded that the information about palmaris longus is significant for surgeons who use 27 an indigenous study on the palmaris longus muscle anatomical variations the tendon of this particular muscle as a graft in various replacement surgeries17. conclusion the knowledge of anatomical variations of palmaris longus muscle is important for anatomists for the awareness of their presence and the related clinical significance. this information is useful for radiologists as well for diagnosing patients who come with wrist swellings and pains. these details related to the variations of the palmaris longus muscle are decisive for physiotherapists in order to treat patients with clinical features of the median nerve or ulnar nerve compression. these findings are also necessary for plastic surgeons who are planning to use the palmaris longus tendon as a graft for reconstructive surgeries in human bodies. references 1. richard s. snell, clinical anatomy by regions. 10th ed. lippincott williams & wilkins, wolters kluwer (philadelphia) pvt ltd, philadelphia. 2019. pages: 127-128, 130 2. keith l. moore, arthur f. dalley, anne m.r. agur. moore clinically oriented anatomy, 8th ed. wolters kluwer (india) pvt. ltd. new delhi. 2018. page: 218-220. 3. chummy s. sinnatamby, last’s anatomy, regional and applied. 12th ed. churchill livingstone. elsevier ltd. great britain. 2011. page: 66 4. georgiev gp et al. palmaris longus muscle variations: clinical significance and proposal of new classifications.folia medica. 2017;59(3): 289 97. 5. longhurst g, stone d, mahony n. review bilateral reversed palmaris longus muscle: a case report and systematic literature review surgical and radiologic anatomy (2020) 42:289-95. 6. mathew aj,sukumarantt,joseph s. versatile but temperamental: a morphological study of palmaris longus in the cadaver. journal of clinical and diagnostic research. 2015, 9(2): 1-3. 7. quadros ls, punja r, nayak vs, yadav js, nayak n, d souza as.palmaris longus muscle in the south indian population a cadaveric study. online j health allied scs. 2017; 16(2)9:1-3. 8. murabita,gnarram,mohamed a. reversed palmaris longus muscle: anatomical variant – case report and literature review. can j plast surg. 2013;21(1):5556. 9. koshi r. cunningham’s manual of practical anatomy vol 1 upper and lower limbs. 16th ed. india. oxford university press, india. chapter 8: the forearm and hand. 2012: 94-5. 10. reimann af, daseler eh, anson bj, beaton le. the palmaris longus muscle and tendon. a study of 1600 extremities. the anatomical record. 1944 ;89(4):495-505. 11. rajesh s, singaram vijayakumar, anju balaji more. a study on morphologyof palmaris longus. int.j.bioassays, 2014, 3(05): 2087-9. 12. sunitha r, prathap kumar j. a study of palmaris longus muscle: its anatomic variations with embryological significance and clinical importance. int j anat res.2018;6(2.2):5222-7. 13. gune ar, pote aj, patil ad, patil ps, nikram vr. bilateral reversed palmaris longus muscle with trifid insertion, a rare variation. int j res med sci. 2014; 2(20): 741-3. 14. lalit m, singla rk, piplani s. bifid inverted palmaris longus muscle – a case report. eur. j. anat. 2014; 18(4): 341–3. 15. hashem m, alatassi r, narinder k and emran f. hypertrophied reversed palmaris longus muscle (pseudotumor) of the forearm causing median nerve compression: a case report. journal of medical case reports. 2020;14(60): 1-4. 16. demir ci, ya ar ek, dursun b and alagoz ms. reversed palmaris longus muscle with median nerve compression symptoms a case report and review of the literature. annals of plastic surgery. 2021; 86(2): 237-41. 17. waghray n and jyothia. unilateral absence of palmaris longus muscle – a case report. apollo medicine. 2014; 11(4): 297-99. the authors: dr. alvia batool, associate professor, department of anatomy, fmh college of medicine and dentistry, lahore. dr. faizia batool, postgraduate resident mcps, department of dermatology, arif memorial hospital, lahore. dr. afifa waseem, assistant professor, department of anatomy, central park medical college, lahore. dr. saira munawar, associate professor, department of anatomy, fatima jinnah medical university, lahore. dr. maryam fatima, medical officer, dermics, gulberg 3, lahore. corresponding author: dr. alvia batool, associate professor, department of anatomy, fmh college of medicine and dentistry, lahore. email: alvia.batool@gmail.com for web full book 15 proceedings s.z.m.c. vol: 37(3): pp. 15-19, 2023. pszmc-890-37-3-2023 postoperative complications in carcinoma gall bladder: a tertiary care hospital experience 1qaiser naveed haral, 1ammad ud din nasir, 1ahmed hassan malik, 1muhammad qasim butt, 1rashid zahid ali, 1arwah mansoor, 2hassan mumtaz 1department of general surgery, nums, rawalpindi. 2department of clinical investigation, maroof international hospital, islamabad. abstract introduction: gallbladder cancer is more common than any other part of the biliary system. accounting between 2% and 3% of all cholecystectomies. aims &objectives: to analyze different postoperative complications in various stages of operated gall bladder carcinoma. place and duration of study: army liver transplant unit (a.l.t.u.) pak emirates military hospital rawalpindi (p.e.m.h.), from november 2020 to april 2022. material & methods: this cross-sectional descriptive study looked at all patients diagnosed with gallbladder cancer before surgery or who were subsequently found to have benign gallbladder cancer presenting with intra-luminal lesions and localized or diffuse wall thickening amenable to surgical treatment. a sequential sampling method was used to select the patients. a proforma was developed to streamline the data collection process. the patient's gallbladder cancer was staged using the a.j.c.c.'s eighth edition staging system, and the degree of complications was classified using the clavien-dindo system. data was entered and analysed using spss version 23. results: a total of 27 patients were included in the study, with male to female ratio of 1:3; and a mean age of 50.1 ± 4.7 years (range 25-70 years). six patients were in stage-1 of whom only 1 (3.7%) developed grade – i complication; 9 patients were in stage-2, out of which 2 (7.4%) developed grade – ii complications; 6 patients were in stage-3, out of which 2 (7.4%) patients developed grade – iii and 1 (4.7%) patient developed grade-ii postoperative complications. six (22.2%) were in stage-4 and underwent curative resection, where no patient developed postoperative complications. conclusion: surgery remains a solitary curative option for gall bladder carcinoma, and when patients are selected carefully, postoperative complications after radical surgery are not very high. moreover, as the stage of the disease increases, so does the complication grade. keywords: gall bladder carcinoma, surgical resection, t.n.m. staging. introduction gallbladder cancer is more common than any other part of the biliary system. gallbladder cancer accounts for between 2% and 3% of all cholecystectomies. its incidence has grown in the last two decades1. the increased incidence of the disease is thought to be attributed to two factors: the general practice of reporting histological findings for all gallbladders removed and the rising popularity of laparoscopic cholecystectomies2. people with non-accidental gallbladder cancer are rarely diagnosed with the disease immediately. surprisingly, even when the severity of the disease is considered, the outcomes are worse than when diseases are discovered by accident. the incidence of gallbladder cancer varies greatly by location3. approximately 22 out of every 100,000 women in northern india are affected. the incidence rate is 11 per 100,000 indians and pakistanis in north america4. there are few reported cases in europe (between 0 and 4 per 100,000)5. the prognosis for gallbladder carcinoma is poor, with a less than 5% 5-year survival rate. with early detection and surgery based on the disease’s stage, 75% of patients diagnosed with cancer can survive for five years6.criteria for stage-adjusted therapy have been published in some countries, but there is a significant deal of diversity in the literature, the standards, and the degree to which people adhere to the guidelines. patients with pt1 cancer and clean margins after cholecystectomy may not need further treatment. patients with pt2 or pt3 incidental cancer should have a drastic re-operation to increase their chances of survival. major surgery, including the removal of the liver and lymph nodes, is an appropriate treatment for cancer7 for the treatment of gallbladder cancer; the s3 guidelines suggest either a wedge resection of the gallbladder bed or resection of liver segments 4b and 510 . both of these 6 1department of public health, health services academy, islamabad. 2department of management sciences, riphah int university, islamabad. 3department of medicine, ihitc, islamabad. 4department of rheumatology, pims, islamabad. 5department of medicine, dow university of health sciences, karachi. 70 1department of orthopedics, thq, khanewal 2department of orthopedics, lgh, lahore 3department of orthopedics, thq, ferozwala 4department of orthopedics, cmh, lahore 5department of hand & upper limb surgery, cmh, lahore 6department of orthopedics, al-rehmat benevolent trust 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 16 postoperative complications in carcinoma gall bladder:a tertiary care hospital experience operations necessitate removing a segment of the gallbladder's bed. gallbladder cancer patients at stages t2 and t3 commonly receive hepatoduodenal ligament lymphadenectomy after undergoing this treatment8. according to the criteria provided by the national comprehensive cancer network, this specific method should be used for carcinomas at stages t1b and above9. this study aims to examine data from patients with gallbladder cancer at various stages of the disease and following surgery at our institution. material and methods this descriptive cross-sectional study was conducted from november 2020 to april 2022, at the department of general surgery, army liver transplant unit (a.l.t.u.) pak emirates military hospital rawalpindi. the hospital's board of directors gave the ethics review clearance vide reference number a/28/ec/450/2022. patients diagnosed with gallbladder carcinoma either before surgery or after surgery for a benign condition were included in the study." patients whose diagnoses included metastatic disease were not included in the research. the study included 27 participants. sequential sampling was used to select them. surgeons, gastroenterologists, diagnostic and interventional radiologists, oncologists, and anesthesiologists worked together to treat each patient. patient demographic information, surgical details, and post-op complications were recorded on a proforma. gallbladder cancer in patients were staged using the a.j.c.c. 8th edition7. clinical outcomes and case-based compensation following surgery strongly correlated with the c.d.c. grading system. as a result, it is a reliable method for contrasting different surgical procedures10. statistical analysis the data was analyzed using s.p.s.s. 23. descriptive statistics calculate values like the mean and standard deviation, while qualitative analysis uses metrics like frequency and percentages. results there was a total of 27 patients who were diagnosed with gallbladder cancer, and the ratio of males to females was 1:3. table-1 illustrates a variety of characteristics of the group under consideration. six of 27 patients were found to be in t1 disease on histopathology report after routine cholecystectomy. among these six patients, three were in the t1a category (tumor had invaded lamina propria) for which no further treatment was required, and 3 were in the t1b category (tumor had invaded muscular layer). out of the patients with t1b disease, three patients had clear cystic duct margin on histopathology report for which radical cholecystectomy plus portal lymph node dissection was done, and one had cystic duct margin involvement on histopathology report for which radical cholecystectomy plus bile duct excision and portal lymphadenectomy was done. parameters n (%) gender male 9 (33.33%) female 18 (66.66%) age (years) mean 50.1 ± 4.7years range (min-max) 25-70 years table-1: characteristics of study population (n=27) nine patients were found with t2 disease (tumor enters peri-muscular connective tissue without extension into the serosa). six individuals had t3 disease (tumor punctures serosa or directly affects the liver or adjacent organs). five underwent radical cholecystectomy. two patients' cystic duct margins required bile duct resection. right hemi hepatectomy was done for hepatic duct involvement. six individuals with t4 illness exhibited peritoneal metastases on diagnostic laparoscopy and one during laparotomy. table-2 shows a.j.c.c. 8th edition stage-wise patient distribution. out of these 27 patients studied, 6 patients developed postoperative complications. according to clavien dindo's classification of different stages of the carcinoma gall bladder, postoperative complications were studied (table-3). among stage i patients, one developed grade–complication (superficial surgical wound infection). among stage ii patients, two patients developed grade – ii complications. one developed basal atelectasis, and the other had a minor bile leak. among stage 3 patients, two developed grade–iii complications and one developed grade–ii complications. one patient had a biloma, which needed image-guided drainage. and one patient developed a postoperative 17 postoperative complications in carcinoma gall bladder:a tertiary care hospital experience hemorrhage, which required laparotomy. grade – ii complication patient had developed pneumonia. ajcc 8 th edition total % age stage i 6 (22.2%) t1a, n0, m0 3 (11.1%) t1b, n0, m0 3 (11.1%) stage ii 9 (33.3%) t2a, n0, m0 2 (7.4%) t2b, n0, m0 7 (25.9%) stage iii 6 (22.2%) t3, n0, m0 6 (22.2%) stage iv(a&b) (t4n0 m0) (anyt, n2, m0) (anyt, anyn, m1) 6 (22.2%) table-2: stage-wise distribution of patients according to a.j.c.c. 8th edition. stages grade-i grade -ii grade -iii grade -iv grade -v total ( %) stage1 1 0 0 0 0 1 (3.7%) stage2 0 2 0 0 0 2 (7.4%) stage3 0 1 2 0 0 3 (11.1%) stage4 0 0 0 0 0 0 (0%) total 1 3 2 0 0 6 (22.2%) table-3: postoperative complications according to clavien dindo classification in different stages of carcinoma (n=6). *where the grade is according to claviendindo classification and stage according to a.j.c.c. 8th edition. discussion only surgical resection, which removes the affected portion of the gall bladder, can cure gall bladder cancer 1. because it is a disease that is so aggressive and potentially fatal, making an early diagnosis is of the utmost significance. numerous studies have shown that the t.n.m. staging system may give information on survival benefits after surgical procedures11. in our research, patients were staged using the t.n.m. classification of gallbladder carcinoma according to the eighth edition of the a.j.c.c. guidelines. table 4 presents the t.n.m. classification according to the a.j.c.c.'s eighth edition. simple cholecystectomy is sufficient for t1a tumors, but t1b requires more aggressive treatment12. extended cholecystectomy with regional lymph node dissection and en bloc hepatectomy is recommended for t2 patients13. in t3 disease, r0 resection (negative margins and lymph node dissection one level beyond microscopically involved lymph nodes) is associated with improved survival14. our study followed suit. r0 resection for t4 disease is not possible, and 5-year survival is low, so extended radical surgeries were not attempted. such patients receive adjuvant chemoradiotherapy. in our study, claviendindo classification was used to classify postoperative complications. primary tumor tx cannot be evaluated t0 no indication of a primary tumor this carcinoma in situ t1a tumor enters lamina propria t1b tumor enters the muscular layer t2 tumor enters peri-muscular connective tissue without extension into the serosa. t3 tumor pierces the serosa (visceral peritoneum) and directly enters the liver or a nearby structure. t4 a tumor involves the main portal vein or hepatic artery or enters 2 or more organs. regional lymph nodes nx cannot be evaluated n0 no regional lymph node involved n1 metastases to 1-3 regional lymph nodes n2 metastases to 4 regional lymph nodes distant metastasis m0 no distant metastasis m1 distant metastasis table-4: a.j.c.c. 8th edition t.n.m. staging system for gallbladder cancer7. complications after surgery include hemorrhage, surgical site infection, pneumonia, and bile leakage. a study on 106 patients operated for gallbladder carcinoma demonstrated surgical site infection in 20.8% of patients, pneumonia in 7.5% and bile leakage in 6.6% of patients15. one patient in our study developed a surgical site infection; one developed basal atelectasis, and one developed pneumonia. bile leaks also occurred in two patients. no in-hospital mortality occurred in our study. our results are comparable to those found in literature16. 18 postoperative complications in carcinoma gall bladder:a tertiary care hospital experience hospital mortality of up to 7.5% after surgery for gallbladder carcinoma has been reported in a study, with failure to rescue from complications as an important cause of mortality17. rapid diagnosis and prompt management of postoperative complications after intricate surgeries for carcinoma gallbladder is paramount to ensure a speedy recovery and discharge from the hospital18. compared to any other surgery, surgery on the liver is complex, with risks such as prolonged duration of surgery, intra-operative hemorrhage, massive fluid shifts, and hypotension, along with postoperative complications such as coagulopathy or bleeding, pulmonary complications, bile leaks, liver or renal failure19. all of these present challenges to the operating team. enhanced recovery after surgery (eras) program has been found to be safe and effective in reducing postoperative complications in patients undergoing hepatobiliary surgery20. it has been found to enhance patient recovery and reduce hospital stays without increasing treatment costs or complications21. eras program involves various pre-operative, intra-operative and postoperative elements22. preoperative elements include minimal fasting, no mechanical bowel preparation, pre-operative carbohydrate load and venous thromboembolism prophylaxis. intra-operative elements include antimicrobial prophylaxis, skin preparation, use of balanced crystalloids, normothermia and glycemic control. postoperative elements include multimodal analgesia, early mobilization and early oral intake23. eras program should be implemented in true letter and spirit in our setup to reduce postoperative complications, as it has been proven its utility in multiple studies23. grades definition grade i any nonconformity from usual postoperative recovery without requiring pharmacological, surgical, endoscopic or radiological interventions. anti-emetics, antipyretics, analgesics, diuretics, electrolytes, and physiotherapy may be utilized. wound infections opened at the bedside are also included. grade ii pharmacological treatment with drugs other than those mentioned for grade i complications. grade iii requiring surgical, endoscopic or radiological intervention iiia intervention not under general anesthesia iiib intervention under general anesthesia grade iv life-threatening complication requiring i.c./icu-management iva single organ dysfunction (including dialysis) ivb multi-organ dysfunction grade v death of a patient table-5: claviendindo classification for postoperative complications13. conclusion surgery remains a solitary curative option for carcinoma gallbladder, and when patients are selected carefully, postoperative complications after radical surgery are not very high. moreover, as the stage of the disease increases, so does the complication grade. references 1. gourgiotis s, kocher hm, solaini l, yarollahi a, tsiambas e, salemis ns. gallbladder cancer. am j surg. 2008 aug;196(2):252–64. 2. toyonaga t, chijiiwa k, nakano k, noshiro h, yamaguchi k, sada m, et al. completion of radical surgery after cholecystectomy for accidentally undiagnosed gallbladder carcinoma. world j surg. 2003 mar 4;27(3):266–71. 3. schmidt ma, marcano-bonilla l, roberts lr. gallbladder cancer: epidemiology and genetic risk associations. chin clinoncol. 2019 aug;8(4):31. doi: 10.21037/cco.2019.08.13. p.m.i.d.: 31484487. 4. pawlik tm, gleisner al, vigano l, kooby da, bauer tw, frilling a, et al. incidence of finding residual disease for incidental gallbladder carcinoma: implications for re-resection.j gastrointest surg. 2007 nov 1;11(11):1478–87. 5. key statistics for gallbladder cancer [internet]. www.cancer.org.availablefrom:https://www.cancer. org/cancer/gallbladder-cancer/about/key statistics.html 6. goetze to. gallbladder carcinoma: prognostic factors and therapeutic options. world j gastroenterol. 2015 nov 21;21(43):12211-7. doi: 10.3748/wjg.v21.i43.12211. p.m.i.d.: 26604631; p.m.c.i.d.: pmc4649107. 7. adjusted a.j.c.c. 6th ed t,n,m, and stage-seer documentation [internet].seer.[cited 2023 jan 16].availablefrom:https//seer.cancer.gov/seerstat/var iables/seer/ajcc-stage/6th 8. goetze to, paolucci v. influence of highand lowvolume liver surgery in gallbladder carcinoma. world j gastroenterol wjg. 2014 dec 28;20(48):18445–51. 19 postoperative complications in carcinoma gall bladder:a tertiary care hospital experience 9. pluchino la, d'amico ta. national comprehensive cancer network guidelines: who makes them? what are they? why are they important? ann thorac surg. 2020 dec;110(6):1789-1795. doi: 10.1016/j.athoracsur.2020.03.022. epub 2020 apr 13. p.m.i.d.: 32298647. 10. téoule p, bartel f, birgin e, rückert f, wilhelm tj. the clavien-dindo classification in pancreatic surgery: a clinical and economic validation. j invest surg. 2019 jun;32(4):314-320. doi 10.1080/08941939.2017.1420837. epub 2018 jan 16. p.m.i.d.: 29336625. 11. benson ab, abrams ta, ben-josef e, bloomston pm, botha jf, clary bm, et al. n.c.c.n. clinical practice guidelines in oncology: hepatobiliary cancers. j natlcomprcancnetw. 2009 apr 1;7(4):350–91. 12. liao x, zhang d. the 8th edition american joint committee on cancer staging for hepato-pancreatobiliary cancer: a review and update. arch pathol lab med. 2021 may 1;145(5):543–53. 13. dindo d, demartines n, clavien p-a. classification of surgical complications: a new proposal with evaluation in a cohort of 6336 patients and results of a survey. ann surg. 2004 aug;240(2):205–13. 14. yuza k, sakata j, hirose y, miura k, ando t, katada t, et al. outcome of radical surgery for gallbladder carcinoma according to t.n.m. stage: implications for adjuvant therapeutic strategies. langenbecks arch surg. 2021 may;406(3):801–11. 15. mekeel kl, hemming aw. surgical management of gallbladder carcinoma: a review. j gastrointest surg. 2007 aug 22;11(9):1188–93. 16. park tj, ahn ks, kim yh, kim t-s, hong jh, kang kj. the optimal surgical resection approach for t2 gallbladder carcinoma: evaluating the role of surgical extent according to the tumour location. ann surg treat res. 2018 feb 28;94(3):135–41. 17. hong ek, kim kk, lee jn, lee wk, chung m, kim ys, et al. surgical outcome and prognostic factors in patients with gallbladder carcinoma. korean j hepato-biliary-pancreat surg. 2014 nov 30;18(4):129–37. 18. wu xs, zhu yd, jin yp, li ml, gong w, liu yb. [diagnosis and treatment for unexpected gallbladder carcinoma(a retrospective study of 45 cases)]. zhonghuawaikezazhi. 2019 apr 1;57(4):265–70. 19. filmann n, walter d, schadde e, bruns c, keck t, lang h, et al. mortality after liver surgery in germany. br j surg. 2019 oct;106(11):1523–9. 20. agarwal v, divatia jv. enhanced recovery after surgery in liver resection: current concepts and controversies. korean j anesthesiol. 2019 apr;72(2):119–29. 21. utsumi m, aoki h, nishimura s, une y, kashima h, kimura y, et al. safety of surgical treatment for elderly patients with gallbladder carcinoma. acta med okayama. 2019;73(3):6. 22. wu x, li b.l., sun j, zheng cj, he xd, liu w, et al. [application of enhanced recovery after surgery in the perioperative management of patients with gallbladder carcinoma]. zhonghuawaikezazhi. 2022 apr 1;60(4):373–8. 23. melloul e, hübner m, scott m, snowden c, prentis j, dejong chc, et al. guidelines for perioperative care for liver surgery: enhanced recovery after surgery (eras) society recommendations. world j surg. 2016 oct 1;40(10):2425–40. the authors: dr. qaiser naveed haral postgraduate resident department of general surgery nums dr. ammad uddin nasir, classified gen surgeon, department of general surgery, nums. dr. ahmed hassan malik, postgraduate resident, department of general surgery, nums. dr. muhammad qasim butt, classified gen surgeon, department of general surgery, nums. dr. rashid zahid ali, postgraduate resident, department of general surgery, nums. dr. arwah mansoor, postgraduate resident, department of general surgery, nums. dr. hassan mumtaz, senior clinical research associate, department of clinical investigation, maroof international hospital. corresponding author: dr. hassan mumtaz, senior clinical research associate, department of clinical investigation, maroof international hospital. email: hassanmumtaz.dr@gmail.com full book 1 proceedings s.z.m.c. vol: 36(4): pp. 01-05, 2022. pszmc-857-36-4-2022 al-sabeen maternal and child hospital, sana’a, yemen 2 haidan hospital, saadah, yemen university of sciences and technology hospital, sana’a, yemen 4 al-noor medical center, shabwah, yemen 5 jiah government medical center, hajjah, yemen 6 faculty of medicine sana’a university, sana'a, yemen 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 2 anemia and the factors associated with it in pregnant women attending al-sabeen maternal and child… 3 anemia and the factors associated with it in pregnant women attending al-sabeen maternal and child… 4 anemia and the factors associated with it in pregnant women attending al-sabeen maternal and child… 5 anemia and the factors associated with it in pregnant women attending al-sabeen maternal and child… proceedings full book dedicated to the valiant health care warriors, medical instructors, future doctors & researchers of pakistan bismillah hir rehman nir raheem. in the name of allah, the most beneficent and the most merciful message by the chairperson shaikh zayed medical complex & chief editor of proceedings prof. sibgha zulfiqar prof. saadia s. alam (chairperson& dean) (chief editor) we have achieved 3 years of hec y category indexation. our ojs supported proceedings website of international standards connects us to the world. the 6th covid 19 pandemic wave has emerged with increasing cases of the omicron variants globally. the lesser-known monkey pox virus cases are also on the rise across europe and the u.s. while in pakistan dengue has once again 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patron in chief: prof. sibgha zulfiqar chairperson & dean chief editor: prof. saadia shahzad alam phd editor: prof. ayesha humayun phd associate editors: prof. m. imran anwar frcs dr.adnan salim mrcp external associate editor: dr. usman iqbal phd members-national: prof. mamoon rashid -frcs (sitara-e-eisaar) prof. nadira mamoon mrc path prof. m. ashraf phd (tamgha-e-imtiaz) prof.m.arif nadeem (fcps) prof.khalid mahmood (phd) dr.sarah ghafoor phd members-international prof.renne koeffel phd (sui) prof.jane banaszak-holl phd(aus) prof. hisham al muhtasebphd (ksa) dr. salma malik m.d (us) dr. sarwat shaheen m.d (us) dr. aliya asher mrcp (uk) dr. uzma nasim siddiqui fcps (aus) section editors: dr. shahilajalil fcps dr. samira haque fcps dr. lubna riaz fcps dr. sadiamaqsood mhpe dr. faraz bokhari m phil dr. noora hassan hezamal aqmer m phil mrs.saimamohsin msc consultant biostatician: mr. muhammad aasim m phil library & information sciences: mr. ihsan basit m phil dr. muhammad shahid soroyaphd technical support: m.saad ullah sheikh (bs.engr) contents riffat iqbal, muhammad shehzad, shaista aslam, faiza gul durrani, kiran aftab, iram liaqat family based genetic association study of tlr4 gene with myocardial infarction in pakistan isma shabbir, aniqua saleem, misbah durrani, umme kalsoom, asma qasim, hassan mumtaz comparative diagnostic accuracy of sonographic strain elastography and fnac in breast lesions vishal farid raza, absar nazir, muhammad shabbir chohan, muhammad yousaf, iqrawaris, khalid javeed khan shock index in patients undergoing elective and emergency general surgical procedures: a predictor of morbidity risk and outcomes hira babar, zunairah mughal, sobia ashraf, sindhu rehman, arsala rashid, samina qamar haematological and clinical findings especially third space fluid accumulation observed in major dengue outbreaks of pakistan saulat sarfaraz, khadija arif, abdullah hussain, shoaib iftikhar, muhammad ikram, amna javed value of transvaginal sonography in antenatal management of ectopic pregnancy rizwana kitchlew, afaq shamim, spenta kakalia, arous khaqan, sonia latif, faizan farooq perception and vaccination status of elderly individuals aged 60 years and above: an urban lahore analysis javeria faridi, ahmed ahson khan, ayesha faridi, sabeen afshan, umair jan bugti, hassan mumtaz frequency of her2/neu immuno histochemical expression in bladder urothelial carcinoma: a study at cmh quetta sulaiman al-sharabi, reham hassan hezam al-aqmer, ghada abdullatef mahmood al-rajami, nader fatehi al-kubati, ahmed al-mukhanjef, amany omer yousef taib, simon hassan dammag, enas ali hussein al-shoaebi, amera mohammed hussein al-okaishi, wedad qaid abdoh hamdan, omar alasali quality of care for paediatric leukemia patients at a hospital in sana’a as perceived by patients’ attendants nida zeeshan, tooba karim, farid adnan, m. ashraf chaudhry preterm neonatal need for mechanical ventilation in prophylactic continuous positive air pressure (cpap) versus therapeutic cpap isma meer, muniza saeed, sana rasheed chaudhry, komal iqbal, muneeza taseer, samita yasmin oxidative stress in non-obese and obese young healthy adults rabeeha bashir, mir jalal-ud-din, shawana asad, sher ali khan, bilawal gul, shabeena naz comparison between lateral internal sphincterotomy and lord’s operation for chronic anal fissure shuja uddin, adeel hameed, mian haneef, usman nazeer gill obturator type of hip dislocation with ipsilateral non-displaced neck of femur fracture: a case report instructions for authors page 1 13 18 25 30 36 43 50 55 60 65 70 74 advisory board basic & pre-clinical sciences prof. mateen izhar phd, mrcpath prof. sibgha zulfiqar m.phil prof. muhammad suhail m.phil prof. nasreen ehsan m.phil prof. tahira naseem m.phil dr. anwaar bashir m.phil dr. suleman dawood phd (uol) dr. nighat yasmeen phd (kemu) dr. saima chaudhary phd (uhs) medicine and allied prof. azeemtaj fcps prof. abdul shakoor fcps prof. talha mahmud fcps prof. 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prof. khwaja khursheed fcps prof. tanvir akhtar butt ms prof. naila akhtar fcps prof. abidashaheenphd prof. sajida malik m.phil prof. rumina hasan phd prof. khalid javeed khan frcs dr. farooq afzal frcs dr. samanshahid phd dr. muhammad shahzad phd dr. ali hussainy zaidi m.d dr. shahzad khuram akram phd dr. fahad azam phd dr. khadija irfan fcps dr. uzma malik fcps dr. ayesha mallick frcp dr. tehseen haider kazmi fcps dr. khalidaajmal m.phil dr. israr ahmed fcps dr. saimabatool fcps dr. saleem muhammad rana phd dr. muhammad khurram habib fcps dr. saba riaz phd dr. raaziatasadduq phd dr. nageen hussain phd dr. saadia nosheen jan fcps dr. soumble zulfiqar phd dr. yasir abbas zaidi fcps dr. muhammad zeeshan sarwar mrcs dr. muhammad imran khokar mrcs dr. nabihafarasat m.phil dr. tayyaba muzaffar m.phil dr. zaeem khalid fcps dr. nafeesah fatimah fcps peer review board (international) prof. khalid javeed khan frcs prof. renne koeffel phd (switzerland) prof. jane banaszak-holl phd (aus) prof. m. hisham al-muhtaseb phd (jordan) prof. mukhtiar baig phd (ksa) dr. sarwat shaheen m.d (usa) dr. zeeshan i. shaikh m.d (usa) dr. tanzeelquratijaz mrcp (uk) dr. hafiz sohail anjum fcps (ireland) dr. rafayazharfrcp (singapore) dr. mazhar nawaz m.d (usa) dr. ahmad azam malik phd (ksa) dr. ishtiaq ahmed fcps (ksa) dr. aamiromairphd (ksa) dr. sabina ahmed mir m.d (usa) dr. anita lamichhane m.d (nepal) dr. faizadurrani phd (uk) dr. waseemlodhi frcog (uk) dr. seerat zahra hammad m.s (ksa) dr. shafya shahid phd (u.s) dr. abdul waheed frcs (u.k) dr. humaira zareen fcps (ksa) dr. amira okud m.d (ksa) dr. asma ahmed mrcp (uk) dr. sabahat sabir mrcog (uk) dr. sarfaraz ahmad frcr (uk) dr. zouina sarfaraz m.d (uk) untitled-1.indd 4 proceedings s.z.m.c. vol: 34(2): pp. 4-6, 2020. pszmc-743-34-2-2020 transfusion medicine practice in a pandemic; use of convalescent plasma in the management of covid-19 sarah tehseen saskatchewan health authority, canada ca. covid-19 pandemic has now infected over 1 million people and caused 62,000 deaths around the world as of april 5th, 2020.1 it is caused by a virus of coronaviridae family now named sars-cov (2) which leads to severe viral pneumonia and acute respiratory distress syndrome (ards).2 the number of cases continue to rise globally with a growing increase in mortality. at this time, no specific treatment is found to be curative for covid-19 however multiple experimental treatments are being explored.3 current therapy for covid-19 is mainly supportive with a focus on prevention by reducing and containing the spread of the virus.3the experimental therapies being investigated include antivirals like lopinavir-ritonavir, interferon-1β, the rna polymerase inhibitor remdesivir and chloroquine.4 the use of plasma from convalescent covid-19 patients is an attractive option since it provides passive immunity by neutralizing antibodies and has precedence in the management of viral pneumonia caused by other viruses of coronoviridae family as well. though the quality of evidence of available literature is moderate to low, the studies showed a consistent reduction in mortality among patients with influenza or sars-cov (1).5 blood supplier systems in canada and the us are currently working towards developing clinical trials to investigate the effect of convalescent plasma on morbidity and mortality from covid-19.6,7 fda has approved its use for critically ill patients following expedited approval on a case by case basis.12 the pathophysiological basis for use of convalescent plasma: both sars-cov(1) and the novel sars-cov-2 are highly pathogenic coroviruses with a strong affinity for lungs. these viruses enter the target cells by (s) spike glycoproteins through the ace-2 receptors. it is postulated that the neutralizing antibodies against these viruses inhibit the s spike mediated entry into target cells.8 multiple animal models have also shown that these antibodies prevent re-infection and viral replication both sars cov (1) and cov (2) viruses.9,10 the source of these neutralizing antibodies can be human convalescent plasma or specific preparations in animal hosts. the human convalescent plasma can then be fractionated and treated to form hyperimmune globulin. hyper-immune globulin and animal-based antibody sources both require multiple processing steps which will be ultimately beneficial but cannot provide a rapid source of antibodies for the current epidemic. hence, currently, straight administration of convalescent covid-19 plasma seems to be the most rational step in the management of critically ill patients.13,14 as mentioned above, plasma from convalescent patients will act through neutralizing antibodies that prevent viral entrance into target cells and potential replication. other mechanisms such as antibodymediated cellular cytotoxicity and phagocytosis may also be possible.14 a recent report of five critically ill individuals with ards and sars-cov (2) infection indicated resolution of ards in 2 patients and an overall improvement in respiratory status with no mortality in all 5 patients. all patients demonstrated a significant reduction in viral load. these patients received plasma from five donors who had recovered from sars cov (2) infection with high titers of neutralizing antibody.11 this case series in human subjects suggests a beneficial effect of the use of convalescent plasma, however, it is not possible to determine the extent of benefit from convalescent plasma as these patients were receiving multiple therapies at the same time including antivirals and anti-inflammatory agents. potential complications of plasma administration: human plasma administration is not risk-free and can lead to mild as well as severe complications such as transfusion reaction associated lung injury (trali) and anaphylaxis. however, most studies do not indicate any significant side effects of plasma transfusion.5 there were two reported cases of possible trali after the use of convalescent plasma for an ebola and a middle eastern respiratory syndrome (mers) patient.15,16 another consideration is the pro-inflammatory characteristics of plasma and antibodies which may 5 transfusion medicine practice in a pandemic; use of convalescent plasma in the management of covid-19 exacerbate the ards contracted by covid-19 patients. although the previous reports on the use of convalescent plasma as well as the current case series of 5 patients report no such adverse effects, this risk should be contemplated when deciding on plasma administration. dose and timing of plasma administration: there is no recommended dose of plasma for covid-19 patients. the patients treated during the sars-cov2 epidemic received two consecutive transfusions of 200-250 ml of abo compatible plasma collected from donors on the same day11 patients with ebola virus were also treated with a similar dose.17 clinical trials being designed to study the impact of convalescent plasma are using 2 doses of 200-250ml of plasma over 1-2 days.23 data obtained from the use of convalescent plasma for viral illnesses has shown its maximum benefit when administered early during disease. it can be hypothesized that the rapid reduction of high viral inoculum with neutralizing antibodies leads to the blunting of pro-inflammatory response and ards compared to late administration when the inflammatory response has been established.18 in the reported case series on the use of human plasma for covid-19 patients, it was given between days 1022 of hospital admission.11 among patients infected with sars cov-1, the plasma was administered on day 14 of admission according to observational studies18 the use of convalescent plasma is only investigated in patients with severe respiratory illness needing significant support.7,11 donor perspectives: donors should be considered for plasma collection after complete resolution of symptoms for at least 28 days before donation or resolution of symptoms for 14 days with a negative test for sars-cov2.7 they can be contacted through the hospital registries after discharge or through widespread educational and marketing campaigns catered to both public and healthcare providers. screening the population for titers of igg antibodies against sars-cov2 as potentials for plasma donation is another option but has significant budgetary constraints.21 these donors should obey the local guidelines for donor eligibility. at the very least infectious disease testing for transfusion-transmitted disease must be performed before accepting the plasma donation. it is recommended that plasma should be collected from men and women with no prior history of pregnancy to mitigate the risk of trali.7 the titers of neutralizing antibodies in the plasma of convalescent sars-cov(1) patients reached a peak at 4 months and continued to decline afterward.19 the patterns of antibody rise and decline in sarscov (2) convalescent patients are still undetermined but it is likely beneficial to consider plasma donation in the first 4 months after diagnosis of covid-19 for maximal benefit. it is also interesting that individuals with severe illness due to sarscov1 were more likely to have high titers of neutralizing antibodies in their plasma. whether this holds for sars-cov2 as well needs to be elucidated.20 ideally, the collected plasma should be tested for titers of neutralizing antibodies against sarscov2. current clinical trials in the us are using a minimum titer of 1:64 as a criterion for plasma collection based on prior research on sars-cov1.22 however, the determination of antibody titers may prove to be difficult in resource-limited settings. at a minimum, potential donors should be tested for the presence of neutralizing antibodies using elisa or similar techniques.21 plasma collection and storage: plasma collection can be undertaken using pheresis machines or following a whole blood donation. one benefit of pheresis is that it will allow the donor to donate plasma every 2 weeks whereas whole blood donations can only be performed every 12 weeks. whole blood donation and subsequent separation and storage of plasma may be more applicable to resource-limited settings and should be considered. after collection from convalescent donors, plasma can be donated immediately or frozen at -18 degrees celsius for up to 12 months for later use or pooled to form hyperimmune globulin. pathogen reduction technique for viral inactivation of plasma, if available, should be performed before plasma administration.17 conclusion there are no proven therapeutic modalities for the cure of covid-19. while significant research is underway to develop medical therapies and vaccines, none are available in a time-sensitive fashion. the use of convalescent plasma as a source of passive immunity against the virus is a feasible and timely solution for critically ill patients. this option can be utilized in resource-limited settings as well with appropriate modifications to plasma collection, donor testing, and administration based on locally developed protocols. as plasma administration has some significant risks, its use must be carefully weighed against the benefits when considering 6 transfusion medicine practice in a pandemic; use of convalescent plasma in the management of covid-19 treatment. ongoing clinical trials will help determine the optimal dose, timing and the patient cohort most likely to benefit from this treatment modality. references 1. https://www.who.int/docs/default-source/ coronaviruse/situation-reports/20200405-sitrep76-covid-19.pdf?sfvrsn=6ecf0977_4 2. lu, roujian, et al. "genomic characterization and epidemiology of 2019 novel corona virus: implications for virus origins and receptor binding." the lancet 395.10224. 2020;565-574. 3. fauci, anthony s., h. clifford lane, and robert r. redfield. "covid-19 navigating the uncharted." 2020; 1268-1269. 4. world health organization. who r&d blueprint: informal consultation on prioritization of candidate therapeutic agents for use in novel corona virus 2019 infection, geneva, switzerland, 24 january 2020. no. who/heo/r&d blueprint (ncov)/2020.1. world health organization, 2020. 5. mair-jenkins, john, et al. "the effectiveness of convalescent plasma and hyperimmune immunoglobulin for the treatment of severe acute respiratory infections of viral etiology: a systematic review and exploratory metaanalysis." the journal of infectious diseases 211.1. 2015; 80-90. 6. levy, prokopchuk-gauc et al. nebmc statement on convalescent plasma and ivig for treatment of covid-19 infection.march31st,2020 7. https://www.fda.gov/vaccines-blood-biologics/ investigational-new-drug-ind-or-deviceexemption-ide-process-cber/investigationalcovid-19-convalescent-plasma-emergency-inds 8. walls, alexandra c., et al. "structure, function, and antigenicity of the sars-cov-2 spike glycoprotein." cell (2020). 9. subbarao, kanta, et al. "prior infection and passive transfer of neutralizing antibody prevent replication of severe acute respiratory syndrome corona virus in the respiratory tract of mice." journal of virology 78.7. 2004; 3572-7. 10. bao l, deng w et al. re-infection could not occur in sars-cov (2) infected rhesus macaques. boprxiv. 2020 0313 (not yet peerreviewed) 11. shen, chenguang, et al. "treatment of 5 critically ill patients with covid-19 with convalescent plasma." jama. 2020. 12. tanne, janice hopkins. "covid-19: fda approves use of convalescent plasma to treat critically ill patients." 2020. 13. beigel, john h, et al. "safety and tolerability of a novel, polyclonal human anti-mers corona virus antibody produced from transchromosomic cattle: a phase 1 randomized, double-blind, single-dose-escalation study." the lancet infectious diseases 18.4. 2018; 410-18. 14. casadevall, arturo, and liise-anne pirofski. "the convalescent sera option for containing covid-19." the journal of clinical investigation 130.4 (2020). 15. mora-rillo, marta, et al. "acute respiratory distress syndrome after convalescent plasma use: treatment of a patient with ebola virus disease contracted in madrid, spain." the lancet respiratory medicine 3.7. 2015; 554-62. 16. chun, sejong, et al. "possible transfusionrelated acute lung injury following convalescent plasma transfusion in a patient with the middle east respiratory syndrome." annals of laboratory medicine 36.4. 2016; 393-5. 17. tiberghien, pierre, et al. "collecting and evaluating convalescent plasma for covid-19 treatment: why and how." vox sanguinis. 2020. 18. momattin, hisham, et al. "therapeutic options for middle east respiratory syndrome coronavirus (mers-cov)–possible lessons from a systematic review of sars-cov therapy." international journal of infectious diseases 17.10 .2013; 792-8. 19. cao, wu-chun, et al. "disappearance of antibodies to sars-associated corona virus after recovery." new england journal of medicine 357.11. 2007; 1162-3. 20. hung, ivan fn, et al. "effect of clinical and virological parameters on the level of neutralizing antibody against pandemic influenza a virus h1n1 2009." clinical infectious diseases 51.3. 2010; 274-9. 21. focosi, daniele, et al. "convalescent blood product therapies for covid-19: a systematic review."2020. 22. https://clinicaltrials.gov/ct2/show/study/nct04 325672 23. https:/clinicaltrials.gov/ct2/show/nct04332835 the authors: dr. sarah tehseen saskatchewan health authority, canada ca. email: sarah.tehseen@gmail.com for web full book 39 proceedings s.z.m.c. vol: 37(3): pp. 39-43, 2023. pszmc-895-37-3-2023 perception of e-learning in medical students of fatima jinnah medical university 1shazo sana, 2hafiz muhammad umair, 1nagmana lateef, 3tahir ayub, 1javeria zafar, 1hina pervaiz 1department of physiology, fjmu, lahore. 2department of hematology, fjmu, lahore. 3department of library science, uet, lahore. abstract introduction: in the modern era, advancing information technology is leaving an impact on the educational system as well wherein classroom learning is being replaced by e-learning. this type of learning is a part of formal educational system in many institutions around the globe. hec is emphasizing the implementation of elearning in formal education, thus perception of students about e learning need to be known. aims & objectives: to determine the perception of medical students of fjmu regarding e-learning. all institutions throughout world had to shift to online learning during covid19 pandemic. this new online system is challenging for many students. the current study was planned to know the perception of medical students of fjmu about elearning. place and duration of study: this was a descriptive cross-sectional study that took place in fjmu, lahore and duration of study was 2 weeks in august 2021. material & methods: the study was conducted on 377 medical students of 1st and 2nd year mbbs. nonprobability purposive sampling technique was employed. a predesigned and pretested questionnaire was administered to the students in august 2021. data was analyzed by spss 23 and frequency % was calculated for quantitative variables as perceived usefulness, perceived ease of use and e learning stressors were calculated. results: mean age of students were 19.4 years, with range of 18-22 years and all were females. majority (85%) used mobile phones for e-learning. 72.6% of the students preferred face to face learning over e-learning. major stressors included inadequate it skills, lack of motivation and poor internet connections. conclusion: medical students of fjmu didn’t prefer elearning over face to face learning. keywords: e-learning, covid 19, medical students introduction learning is the process of obtaining knowledge and skills through study or other means for a specific purpose. as time is passing, the trends and modes of learning are changing day by day. the majority students are using mobile phones, laptops, tabs and tools like that for acquiring knowledge. the traditional ways of learning are gradually being replaced by new systems involving use of information technology (it)1. thus, the modes of learning are classroom model, online or e-learning and blended learning. classroom model demands the physical presence of teacher and students in a room and learning is by their direct communication to each other. e-learning is totally virtual by means of use of computer and internet technologies in which there is no physical communication amongst teachers and students, while blended model is the mixture of both forms2. e-learning system is a part of formal education in many universities across the world and many learners prefer this system due to being easy, flexible, and controllable in terms of time and place3,4,5. a study conducted in syiah kuala university, indonesia showed a positive response of students towards online learning and their students enjoy this learning6. but other learners prefer face to face learning due to lack of student teacher interaction and complexity of the process as students of air-langga university, also in indonesia rejected the elearning system due to being a complex process7. similarly, another study on university students in oman highlighted barriers to online learning and preferred face to face system8, while a study conducted in university of sharjah in united arab emirates showed a positive attitude of students towards online learning9. in comparison to the world, in pakistan elearning is not a part of our routine educational system but in year 2020, this system was used in many institutions. due to the covid-19 pandemic, almost 6 1department of public health, health services academy, islamabad. 2department of management sciences, riphah int university, islamabad. 3department of medicine, ihitc, islamabad. 4department of rheumatology, pims, islamabad. 5department of medicine, dow university of health sciences, karachi. 70 1department of orthopedics, thq, khanewal 2department of orthopedics, lgh, lahore 3department of orthopedics, thq, ferozwala 4department of orthopedics, cmh, lahore 5department of hand & upper limb surgery, cmh, lahore 6department of orthopedics, al-rehmat benevolent trust 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 40 perception of e-learning in medical students of fatima jinnah medical university all educational institutions were closed around the globe. to prevent the academic loss, all institutions including medical schools have tried to facilitate the students by providing online learning using google classrooms, webinars, online video meetings etc. this has raised multiple challenges for the students10,11. thus most of our faculty members and students are in process of getting used to new online teaching system, as reflected by the study conducted in liaquat medical & dental college in karachi showed negative impact of online learning in medical students during the current situation22. higher education system of pakistan is emphasizing the implementation of e-learning system in formal education throughout the country13. that’s why it is necessary to know the students point of view regarding the acceptability of elearning system. so, a study is planned to determine the perception of medical students of fjmu regarding e-learning in the current situation. material and methods ethical approval of study was taken from institutional/ ethical review board of fjmu with number 12/cierb dated 09-08-2021. a descriptive cross sectional study was conducted at fjmu, lahore for a period of 2 weeks in august 2021. using the non probability purposive sampling 377 1st and 2nd year mbbs were inducted in the study as calculated by rao soft14) a predesigned and pretested questionnaire obtained from a study conducted on university students in ghana with slight modification was used15. the questionnaire had 21 questions divided into 6 subdivisions. 1st one was the general information. the remaining 4 sections were based on the technology acceptance model (tam) and covered the following: perceived usefulness (pu) of elearning perceived ease of use (pe) attitude towards e-learning (at) intention to use (iu) e-learning in future. and final subdivision included elearning stressors. all items were assessed on 5 point likert scale, from strongly disagree 1, disagree 2, no response 3, agree 4 and strongly agree 5. at the end, participants were thanked for their cooperation. data was collected by administering the questionnaire to the students during tutorial classes. data analysis: data was entered and analyzed in spss-23. descriptive statistics was applied for getting the frequency percentage of the data. results the current study has been conducted on 377, 1st and 2nd year medical students of fjmu. mean age of students was 19.4years with range of 18-22 years. all students were females and majority used mobile phones (85%) for the online learning. most of the students liked zoom app for learning (74.2%). the general characteristics of study participants are given in table-1. sr.no frequency percentage (%) 1 gender male 0 0 female 377 100 2 year of study 1st year 158 41.9 2nd year 219 58 3 choice of gadget/device mobile phones 321 85 laptop 56 14.8 4 choice of platform zoom app 280 74.2 google 27 7.1 classroom microsoft team 70 18.5 table-1:general characteristics of study population perceived usefulness of elearning(pu): majority of the medical students thought that online learning improves the access to the learning materials. students strongly disagreed to the better engagement of students in learning. students thought that this online learning is much time consuming and less interactive among students and teachers. overall this learning system impacted the respondents less as compared to face to face learning as shown in table-2. perceived ease of use: majority of students found online learning more difficult than consulting library. listening to lectures online was troublesome for majority of students, moreover students also faced difficulty in expressing their thoughts during composing and attempting online elearning assignments and submissions. most students disagreed with the statement that they were quick while using web for e learning as depicted in table-3. 41 perception of e-learning in medical students of fatima jinnah medical university sr. no questions strongly disagree disagree no response agree strongly agree 1 e-learning develops admittance to learning material 52.8% 36.2% 3.45% 5.3% 2.22% 2 e-learning increase learners commitmentin learning 55.6% 36.1% 0.6% 2.2% 5.8% 3 elearning better develops educator’s and learners’ connection 48.7% 33.3% 4.2% 9.2% 5.2% 4 e-learning is excessively tedious to utilize 12.3% 8.9% 7.4% 31.5% 41.4% 5 e-learning has little influence on me 5.8% 12.1% 10.1% 34.2% 38.4% 6 e-learning help to support my insight 40.2% 33.9% 9.2% 13.2% 4.2% 7 my research abilities increased several fold by e learning 33.1% 32.1% 22.5% 9.8% 2.5% table-2: showing percentage for perceived usefulness sr. no questions stron gly disagr ee disagre e no respons e agree strongly agree 1 utilizing e learning is more simple than utilizing library 34.8 % 36.3 % 20.1 % 2.5 % 6.3 % 2 i can peruse the lecture notes through web without any problem 45.4 % 39.2 % 9.1 % 6.4 % 3 it is easy to expressmy viewpoint by composing through e learning 33.4 % 18.8 % 27.2 % 10.8 % 9.8 % 4 i am quick while using the web for e learning 36.4 % 33.3 % 10.1 % 11.4 % 8.8 % 5 i find e learning clear and understandable 37.5 % 45.3 % 12.4 % 4.8 % table-3: showing percentage for perceived ease of use. attitude towards elearning (at): 49.7% students strongly disagreed with the idea of online learning as shown in the fig-1 and 40% suggested that online learning should be discouraged as given in fig-2 revealing our medical students to have negative attitude toward online learning. fig-1: showing likeness of e-learning fig-2: students view about encouraging e-learning e-learning stressor: the reasons behind this negative attitude towards online learning included inadequate it skills and inadequate training on using online learning, poor motivation and expectation from part of students and major stressor included the slow internet connections as given in table-4. intention to use in future: majority of our students (72.5%) preferred face to face learning over online learning, 20.1% also choose hybrid method and only 8.9% students preferred online learning shown in fig-3. 42 perception of e-learning in medical students of fatima jinnah medical university stressors strongly disagree disagree no response agree strongly agree my inadequate computer /mobile skills stresses me. 6.8 20.8 29.0 43.4 inadequate training on using e learning stresses me. 23.5 7.3 39.4 29.8 poor motivation and expectation from me regarding e-learning stresses me. 9.8 18.8 4.5 39.9 27.0 limited resources as weak internet connections stresses me. 10.8 20.6 1.3 31.9 35.4 table-4: stressors of e-learning fig-3: showing intention of students to use elearning discussion the current study was planned to assess the perception of students of fjmu about elearning during covid-19 pandemic. our study showed that the majority of our students used mobile phones for e-learning and preferred zoom platform for learning followed by microsoft teams. many studies also showed preference of mobile phone for online learning in students16,17,18, the reason being the easy interaction at mobile and easy to carry anywhere. our students were forced to e-learn during covid 19 pandemic as many institutions throughout the world but their overall perception about e-learning was not positive. majority of students had negative views like e-learning has little impact, is time consuming and less engaging, this is in line with many studies round the world like poland19, saudi arabia20 and india21 etc. one study conducted in liaquat college of medicine and dentistry in our country also highlighted that 77% of medical students had negative perception for e-learning during covid 19 pandemic12. another study conducted by ansar et.al. in 10 different medical, engineering and arts institutions also concluded that 81% of students didn’t want to continue with elearning22. in our country, online learning is going on in many institutions even before covid 19, students had different opinions regarding elearning, some preferred while others rejected this system. a study conducted in nursing students highlighted that nursing students preferred to adopt online learning23, while in our study, 72% of our students preferred face to face learning. thus the majority of students around the globe are not in favor of online learning. our study has also highlighted the major stressors that might be the reason for rejecting online learning. these included inadequate training, lack of information technology skills and poor internet connection, the same stressors have also been noted in study conducted in india24. by overcoming these stresses, online learning acceptability may be increased and this online system can be adopted in a better way, if its need arises again in future. conclusion e-learning system was not digestible for medical student of fjmu during covid19 pandemic lock down period. the university should take measures to overcome the stressors of e-learning well in time so that students learning isn’t affected if a similar situation arises. references 1. ali m, hossain khalid, ahmed t (2018). effectiveness of e-learning for university students: evidence from bangladesh. asian j emp res. 2018; 8(10), 352-60 2. woods m, rosenberg me. educational tools: thinking outside the box. clin j am soc nephrol. 2016;11(3):51826. doi:10.2215/cjn.02570315 3. govindasamy t. successful implementation of e-learning: pedagogical considerations. internet high educ. 2001;4(3).287-99. 4. blas tm, fernandez as. the role of new technologies in the learning process: moodle as a teaching tool in physics. comput educ. 2009;52(1):35-44. 5. bentley y, selassie h, shegunshi a. “design and evaluation of student-focused elearning” elect j elearn.2012;1(10):1-12. 43 perception of e-learning in medical students of fatima jinnah medical university 6. vitoria l, mislinawati m, nurmasyitah n. students’ perceptions on the implementation of e-learning: helpful or unhelpful? j physics. 2018;1088 7. kwary da, fauzie s. students’ achievement and opinions on the implementation of e-learning for phonetics and phonology lectures at airlangga university. educ pesqui. 2018;44. 8. al-senaidi os, lin l, poirot j. barriers to adopting technology for teaching and learning in oman. comp educ. 2009: 53;575-90 9. elshami w, taha mh, abuzaid m, saravanan c, al kawas s, abdalla me. satisfaction with online learning in the new normal: perspective of students and faculty at medical and health sciences colleges. med edu online. 2021: 26; 1-10 10. kow a, chong y, lau t, samarasekera dd, li d, goh m, et al. response and lessons learnt managing the covid19 crisis by school of medicine, national university of singapore. mededpublish. 2020; 9(1): 1–12. 11. aggarwal a, comyn p, fonseca pm. discussion: continuing online learning and skills development in times of the covid-19 crisis. 27 march 17 april. availableonline:https://www.skillsforemployment.org/ks p/en/ discussions/edmsp1_256625. 12. abbasi s, ayoob t, malik a, memon si. perceptions of students regarding e-learning during covid-19 at a private medical college. pak j med sci. 2020;36(covid19s4):covid19-s57-s61. 13. higher education commission. hec covid-19 policy papers. 2020 [cited 2020apr22].https://www.hec.gov.pk/english/hecannoun cements/documents/ncovirus/covid-19-policy-guidanceno.5-online readiness.pdf 14. raosoft. sample size calculator. 2004. available online. http://www.raosoft.com/samplesize.html 15. mamattah rs. student perception of e-learning. [master’s thesis]. ghana: linköping university;2016. 16. yilmaz o. e-learning: students input for using mobile devices in science instructional settings. edu learn. 2016;5:182. doi: 10.5539/jel.v5n3p182 17. roberts n, rees m. student use of mobile devices in university lectures. australas j educ technol. 2014;30:4. doi:10.14742/ajet.589 18. martinez ig, sanchiz dc, batanero jmf, rosa aldl. using mobile devices for improving learning outcomes and teachers’ professionalization. sustainability. 2019;11:6917. doi: 10.3390/su11246917 19. micha b, b czek z, monika s, andrzej j, beata kw. students’ perception of online learning during the covid19 pandemic. medicine. 2021;100(7): e24821 doi: 10.1097/md.0000000000024821 20. al zahrani, em, al naam ya, alrabeeah sm et al. e learning experience of the medical profession’s college students during covid-19 pandemic in saudi arabia. bmc med educ, 2021; 443 https://doi.org/10.1186/s12909-021-02860-z 21. gupta s, dabas a, swarnim s, mishra d. medical education during covid-19 associated lockdown: faculty and students’ perspective. med j armed forces india 2021;77(suppl 1):s79-s84. https://doi.org/10.1016/j.mjafi.2020.12.008 22. ansar f, ali w, khattak a, naveed h, zeb s. undergraduate students’ perception and satisfaction regarding online learning system amidst covid-19 pandemic in pakistan. j ayub med coll abbottabad 2020;32(suppl. 1):644–50 23. ali n, jamil b, sethi a, ali s. attitude of nursing students towards elearning. adv health prof educ. 2016;2(1):2429 24. chakraborty m, reddy ya, ghoshal ja, amudharaj d, tripathi m. preparedness of medical students towards elearning conducted during covid-19 lockdown: a crosssectional descriptive study. j edu health promot2021;10:302 doi: 10.4103/jehp.jehp_1125_20 the authors: dr. shazo sana, demonstrator, department of physiology, fjmu, lahore. dr. hafiz muhammad umair, senior demonstrator, department of hematology, fjmu, lahore. dr. nagmana lateef, associate professor, department of physiology, fjmu, lahore. tahir ayub, internee, department of library science, uet, lahore. dr. javeria zafar, demonstrator, department of physiology, fjmu, lahore. dr. hina pervaiz, demonstrator, department of physiology, fjmu, lahore. corresponding author: dr. shazo sana, demonstrator, department of physiology, fjmu, lahore. email: shazirose2011@hotmail.com untitled-1.indd 11 proceedings s.z.m.c. vol: 34(2): pp. 11-15, 2020. pszmc-745-34-2-2020 comparison of different doses of petroleum ether extracts of sphaeranthus indicus linn against dutasteride on testosterone and psa in murine model of benign prostatic hyperplasia 1sabiha erum khan, 2bisma fatima aslam, 1saadia shahzad alam 1department of pharmacology, shaikh zayed medical complex, lahore 2department of pharmacology, azranaheed medical college, lahore abstract introduction: benign prostatic hyperplasia (bph) is the most common health problem of male elderly population resulting in lower urinary tract symptoms (luts), characterized by frequency, urgency, hesitancy, nocturia, dysuria and incomplete voiding. aims & objectives: to compare the different doses of petroleum ether extracts of sphaeranthus indicus linn (sip) against dutasteride, on hormonal parameters (serum testosterone and psa) in testosterone induced bph in albino mice. place and duration of study: this experimental study was conducted at the animal house of pgmi, lahore for 6 months. material & methods: thirty-six healthy adult male mice, divided into 6 groups, were administered testosterone, different doses of petroleum ether extract sil extracts (sip) and dutasteride. animals in group 1, taken as control, were given 0.1ml corn oil daily subcutaneously for 28 days. the animals in the remaining 5 groups (# 2-6) were given 3 mg/kg body weight testosterone dissolved in corn oil subcutaneously for 28 days, to induce bph. the animals in group 3 were given 20 mg/kg/day dutasteride orally for 28 days. animals in groups 4-6 were given 25,50mg and 75mg/ kg body weight petroleum ether extract (sip) orally for 28 days. blood samples were drawn on day 0, 14 and 28 for the estimation of serum testosterone and serum psa, by elisa technique. data was analyzed using spss version 20.0. comparison (between groups at baseline, day14 and day 28)was done by using one-way anova and post hoc tukey’s test results: decrease in psa level and increase in serum testosterone supports the alpha reductase inhibiting activity of sip, in a dose dependent manner, in testosterone induced bph. si petroleum ether extract 75mg/kg exhibited efficacy similar to that of dutasteride. conclusion: sphaeranthus indicus linn (sip) has shown efficacy equal to that of dutasteride in attenuating the testosterone induced bph, in albino mice. key words: benign prostatic hyperplasia, testosterone, prostate specific antigen, dutasteride, sphaeranthus indicus linn (sil), petroleum ether extract of sphaeranthus indicus linn (sip). introduction “i do not need you to remind me of my age. i have a bladder to do that for me”. (stephen fry—author of “the fry chronicles”) benign prostatic hyperplasia (bph) is the most common, although benign, bothersome malady causing significant urinary symptoms in the male elderly population.1 based on histology, the term refers to proliferation of smooth muscles and epithelial cells situated in the transition zone of the gland.2 bph could progress unchecked, if not managed rationally, leading to lower urinary tract symptoms (luts) manifested by weak urinary stream, hesitancy, incomplete voiding and irritative symptoms including frequency, urgency, nocturia and dysuria.3 incomplete voiding results in stasis of residual urine with an increased risk of recurrent urinary infections. chronic urinary retention may result in renal failure in some patients.4 generally , it is not a life-threatening condition.5 however, it is well known to have significant detrimental effect on a patient's quality of life (qol).6 whereas the incidence is on increase due to an increase in average life span, the threat of bph is hanging over most of elderly men as the “sword of damocles”.7 it affects 50 percent men between 51 and 60 years and about 80 percent in their eighties.8 12 comparison of different doses of petroleum etherextracts of sphaeranthus indicus linn against dutasteride the etiology is not clearly understood.9 however, the development of bph is associated with the presence of testicular androgens during prostate development and growing age.10 the major portion of testosterone, in men, is produced by leydig cells of the testes but also, to a lesser extent, by adrenal cortex. in the body tissue, the testosterone is converted into dihydrotestosterone (dht) which is more potent and plays an important role in the development of prostate. however, the presence of dht, in adulthood, could be injurious as it would result in unchecked growth of the gland.11 the physiologically well characterized 5ar substrate, the androgen testosterone and its more potent metabolite dht are essential hormones responsible for male phenotypic sexual differentiation and maturation through their actions at the androgen receptor”.12 psa is chemically a serine protease, produced primarily by the glands in the transition zone and is androgen dependent. the highest amount of psa is found in these minal fluid; some amount of psa escapes the prostate and could be detected in the serum. the rising levels of psa in serum are associated with prostate cancer, however the levels may also rise in bph. the normal value for total psa in human blood is 4ng/ml. when between 410, it indicates possibility of bph or cancer. the normal value of psa levels in mice is 0.140.86ng/ml.14 alpha blockers and 5alpha reductase inhibitors are the two fda approved treatment options. administration of a 5alpha reductase inhibitor (dutasteride and finasteride), offers rapid and sustained inhibition of dht and is therefore the major treatment for bph, at the present. the patients are at risk of acquiring drug related problems (drps), notably dizziness and orthostatic hypotension in case of alpha blockers and sexual dysfunction and gynecomastia by 5alpha reductase inhibitors.15 sphaeranthus indicus linn (gorakmundi) which literally means round flower, in greek, is a medicinal plant fascinating healthcare provider since centuries. it is an indigenous herb of south asians being in use in ayurvedic pharmacopeia, for many ailments. it grows frequently both in cultivated and un-cultivated lands, from sea level to 1200 m altitude, in indian sub-continent, malaysia, china, africa and australia.16 it is a spreading aromatic plant (becoming odorless on long standing) with spreading glandular stem and branches with purple or pink flowers.17 the phytochemical investigations of the plant revealed the presence of17: ● an alkaloid "spaeranthine" ● a novel "flavonoid c-glycoside ● stigmasterol and b-sitosterol ● bicyclic sesquiterpene from petroleum extract although the efficacy of this medicinal plant in bph was postulated earlier, the first published study, on its potential use in bph, was conducted in india, in 2011, by nahata and vinod kumar dixit. they induced prostatic hyperplasia, in albino rats, by giving them testosterone. the administration of sil was found to attenuate the action of testosterone.18 to the best awareness of authors, no authentic published work is available on its potential use in bph, in pakistan. whereas nahata and dixit used finasteride as positive control, we opted dutasteride instead. it is already known that finasteride (selective 5α reductase inhibitor) achieves 70% suppression which in case of dutasteride (from non-selective group) reaches up to 94.8%.19 material and methods animals: 36 adult male mice of balb c species, weighing 2030 gm, were purchased and housed in university of health sciences, lahore. they were kept in standard polyprolene cages, at controlled room temperature of 25+10˚c and relative humidity 6070%. six groups of 6 animals each, were used for the experiment. duration of study was total of 28 days. testosterone propionate (tesvot 250 mg) injections were purchased from local pharmacy. all groups (except the control one) received 3mg/kg body weight testosterone propionate subcutaneously for 28 days, to induce bph.20 dutasteride (avodart 0.5 mg) tablets were purchased from the local pharmacy. animals in group 3 were given 20 mg/kg/day dutasteride orally for 28 days. mouse testosterone elisa kits and mouse psa elisa kits were purchased from glory science limited usa. petroleum ether was purchased from merck pharmaceuticals. plant material: flower heads of sil were collected from lahore area. they were identified and authenticated from the herbarium maintained by the department of botany of university of the punjab lahore. they were kept at normal room temperature. petroleum ether was prepared in the chemistry department of pcsir laboratory lahore. 13 comparison of different doses of petroleum etherextracts of sphaeranthus indicus linn against dutasteride preparation of petroleum ether extract: 500gms of powdered flower heads of sil were dipped in 2l petroleum ether, kept for 48 hrs with periodic shaking and left for half an hour to settle down. it was then filtered in a big flask using whatman’s filter paper and glass funnel. the filtrate evaporated in rotary evaporator until all the petroleum ether was removed and dark greenish yellow waxy mass was obtained.18 from 500g herb, the extract obtained was 14gm (2.8% yield). animal grouping: animals in group 1 (control group) were given 0.1ml corn oil daily subcutaneously for 28 days. the animals in the remaining 5 groups (2-5 groups) were given 3 mg/kg body weight of testosterone (containing 0.075 mg testosterone) dissolved in 0.1ml of corn oil subcutaneously for 28 days. animals in group 3 were given 20 mg/kg/day dutasteride orally for 28 days. those in group 4, 5 and 6 received 25 mg, 50 mg and 75 mg/kg body weight sip extract orally for 28 days. statistical analysis: data was analyzed by spss version 20.0. the data for serum testosterone and serum psa levels obtained by using mean±sd. comparison (between groups at baseline, day14 and day 28)was done by using one-way anova and post hoc tukey’s test. lettering was assigned on the basis of tukey’s test. different letters in superscript represented significant difference from each other and those having common letters were non significant with pvalue 0.05. cluster bar diagrams were made to give visual difference between groups at each reading time and error bars were used to show the standard deviations. p-value ≤ 0.05 was considered statistically significant. results the mean serum testosterone levels at baseline were 1.55 to 1.59 and had no significant difference (pvalue 0.271). at day 14 the group 6 had highest level, significantly different from all other groups, followed by group 3, 5 and 4 respectively. the group 1 and 2 had no significant difference from each other on day 14 but had significantly lower mean testosterone levels as compared to all other groups. on day 28 the testosterone levels increased in all groups as compared to day 14 and group 3 had the highest mean level of 10.48 ng/ml, non significantly different from group 6 but significantly higher than all other groups. the group 2 also had significantly higher level as compared to group 1, but significantly lower than group 4 and 5. (fig-1) fig-1: cluster bar diagram presenting mean serum testosterone levels for six study groups (error bars presenting standard deviation) the mean serum psa levels, ranging between 0.22 and 0.25ng/ml at base line with standard deviations ranging between 0.02 and 0.04, were insignificantly different with p-value 0.651. on day 14 the mean levels in all groups (except group1) rose significantly, the highest being in group 2. groups 3, 5 and 6 had no significant difference from each other, either on day 14 or day 28. group 4 had significantly higher level in comparison to other groups but lower than group 2. the same was the difference on day 28, though the mean levels in all groups were higher as compared to day 14. (table2, fig-2) fig-2: cluster bar diagram presenting mean serum psa levels for six study groups (error bars presenting standard deviation) 0 2 4 6 8 10 12 group 1 group 2 group 3 group 4 group 5 group 6m ea n se ru m t es to st er on el ev el o f an im al s + sd (n g/ m l) study groups day 0 day 14 day28 0 0.2 0.4 0.6 0.8 1 1.2 1.4 1.6 group 1 group 2 group 3 group 4 group 5 group 6 m ea n se ru m p sa le ve l o f a ni m al s + sd (n g/ m l) study groups day 0 day 14 day28 14 comparison of different doses of petroleum etherextracts of sphaeranthus indicus linn against dutasteride discussion bph is a progressive age-related health problem of elderly men. it is clinically characterized by prostatic enlargement and resultant luts causing significant detrimental effects on qol of the life of the patients.1,6 it is not a premalignant condition and has anetiology different from prostatic cancer. a variety of growth factors and inflammatory process could contribute to the causation.9 although many medical and surgical treatment options are available, the problems of the patients remain unresolved in a significant number of cases. the medical therapy does not work for everyone. the drug related problems have a serious impact. the surgical intervention may, in some cases, worsen the condition. the immediate post-operative complications include bacterial infection (up to 15%), impotence (upto 17%) and retrograde ejaculation (upto 77%). however, the incidence of actual post-operative complications, as concluded by stefanie petrou binder, is higher than that reported in literature.22 phytotherapy, as a new treatment option for bph, is gaining popularity on global level. sil is in use in ayurvedic system of traditional medicine, for many ailments.23 it is an ingredient of “prostabliss” which is said to have beneficial effects on bph.14 the present study was conducted to assess the efficacy of various doses of petroleum ether extracts of this herb in comparison to dutasteride in testosterone induced prostatic hyperplasia in albino mice. testosterone levels were measured in this study on day 0, 14 and 28 as a marker of 5 alpha reductase inhibition. all the groups (except the control one) showed an elevation of testosterone levels because of exogenous testosterone treatment. the effect on group 3 (testosterone+dutasteride) was quite similar to that in group 6 (testosterone+ sip) manifesting an increment of 572.5% in serum testosterone level. results show that group 3 had significantly higher serum testosterone level than all the groups except 6 with a p-values <0.05. these findings support the alpha reductase inhibitory effect of sil.25 at day 28, the average mean serum psa level of group 1 was 0.25ng/ml and that for group 2 was 1.37ng/ml. the comparatively highest level in group 2 (testosterone alone) could be explained because of the resultant prostatic hyperplasia. group 1 had significantly lower levels as compared to all other groups while group 2 showed significantly higher psa levels as compared to all other groups with pvalues <0.05. it shows that the testosterone opposing effect of dutasteride was comparable with sip 50mg/kg and sip 75mg/kg body weight. the only available comparable study (nahata et al) also showed a decrease in serum psa by beta sitosterol of sil.18 a large multi-centered study is required to further evaluate the alpha reductase inhibitory effect of the fascinating herb. the results of the present study, if confirmed on a large scale, would, most hopefully, bring a revolutionary change in the management of bph and the patients would be saved from the injurious untoward effects of the presently available therapeutic regime. this joint venture of the pharmacologists, urologists and pharmacists is the need of the day. conclusion in conclusion, this fascinating herb has shown efficacy equal to that of dutasteride in attenuating the testosterone induced bph in albino mice. the results suggest that petroleum ether extracts of sil prevented prostatic hyperplasia significantly in a dose dependent manner. the best activity was exhibited by sip 75 mg /kg body weight. references 1. auffenberg gb, helfand b, mc vary kt. established medical therapy for benign prostatic hyperplasia-urolclin north am. 2009; 36(4):443-459. 2. craft j, gordon c, adriana p et al. understanding pathophysiology. anz adaptation-elsvier health sciences april 2013. 3. wang jy, liu m, zhang yg. relationship between lower urinary tract symptoms and objective measures of benign prostatic hyperplasia: a chinese survey-china med j. 2008; 121(20):2042-2045. 4. timothy jw. benign prostatic hyperplasia. bmj. 2008; 336: 146. 5. dereijke tm, klarskov p. comparative efficacy of two a1-adrenoceptor antagonists, doxazocin and alfuzocin in patients with lower urinary tract symptoms from benign prostatic enlargement. bju int. 2004; 93:757-762. 6. bruskewitz rc. quality of life and sexual function in patients with benign prostatic hyperplasia. rev urol. 2003; 5(2):72-80. 7. vision vip. en. international people groupbiologically active food suppl -prostate 8. napalkov p, maisonneuve p, boyle p. worldwide patterns of prevalence and mortality from benign prostatic hyperplasia. urology. 1995; 46(3):41-46. 15 comparison of different doses of petroleum etherextracts of sphaeranthus indicus linn against dutasteride 9. lepor h. pathophysiology, epidemiology and natural history of benign prostatic hyperplasia. rev urol. 2004; 6(suppl 9):s3-s10. 10. briganti a, capitanio u, suardi n. et al. benign prostatic hyperplasia and its aetiologies. eururolsuppl 8. 2009; 865-871. 11. carson c, rittmaster r. the role of dihydrotestosterone in benign prostate hyperplasia. urology. 2003; 61(4 suppl):2-7. 12. nef s, parada lf. hormones in male sexual development. genes dev. 2000; 14:3075-3086. 13. balk sp, ko y-j, bubley gt. biology of prostate specific antigen. j clinoncol. 2003; 21:383-391. 14. kindblom j. role of prolactin in the prostate gland: studies in transgenic mouse model. phd thesis goteborg university. goteborg2003 15. huri hz, sulaiman cz. drug related problems in patients with benign prostatic hyperplasia: a cross-sectional retrospective study. plos/one. 2014; 9(1). 16. ramachandran s. review on sphaeranthus indicus (kottaikkarantai). pharmacogonosy review. 2013; jul-dec 7(14):157-169. 17. chakraborty ak, behera m, roy h, rambhade s. sphaerathus indicus linn-a phytophamacological review. int j drug farmulation research. 2010; vol 1(3):113-133. 18. nahata a, dixit vk. sphaeranthus indicus attenuates testosterone induced prostatic hypertrophy in albino rats. phytotherapy research. john willey and sons ltd. 2011. 19. nickel jc. comparison of clinical trials with finestride and dutasteride. reviews in urology. 2004; 6 suppl9: s31-s39. 20. gleave me, hsieh jt, wu hc et al. serum prostate specific antigen levels in mice bearing human prostate lncap tumors are determined by tumors volume and endocrine and growth factors. cancer res.1992;1598-1605 21. urology care foundation. bph: surgical management. urology a-z. july 2013. 22. binder sp. bph surgery complications underestimated in clinical trials. urology times june 01, 2012. 23. suter a, saller r, riedi e et al. improving bph symptoms and sexual dysfunctions with a saw palmetto preparation? results from a pilot trial. phytother res. 2013; 27(2):218-226. 24. mhfw (ministry of health and family welfare). the ayurvedic pharmacopea of india -1st edition part 1 vol 3. the controller of publications, new delhi 2001. 25. nahata a, sexena a, suri n dixit vk. sphaeranthus indicus induces apoptosis through mitochondrial dependent pathway in hl-60 cells and exerts cytotoxic potential on several human cancer cell lines. integ cancer therapy. august 22, 2012. the authors: dr. sabiha erum khan p.g. trainee, department of pharmacology& therapeutics, shaikh zayed medical complex, lahore. dr. bisma fatima aslam demonstrator, department of pharmacology & therapeutics, azra naheed medical college, lahore. prof. saadia shahzad alam hod, pharmacology & therapeutics, shaikh zayed medical complex, lahore. corresponding author: dr. sabiha erum khan p.g. trainee, department of pharmacology& therapeutics, shaikh zayed medical complex, lahore. e-mail: drsabihaerum@gmail.com full book 59 proceedings s.z.m.c. vol: 36(4): pp. 59-64, 2022. pszmc-865-36-4-2022 1department of orthopedics, general hospital, lahore 2department of hand & upper limb surgery, cmh, lahore 3department of orthopedics, thq, ferozwala 4department of orthopedics, thq, khanewal 5department of orthopedics, al-rehmat benevolent trust, pasroor 6department of orthopedics, general hospital, lahore 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 60 comparison of external fixation with illizarov method versus open reduction with locking plate in… 61 comparison of external fixation with illizarov method versus open reduction with locking plate in… 62 comparison of external fixation with illizarov method versus open reduction with locking plate in… 63 comparison of external fixation with illizarov method versus open reduction with locking plate in… 64 comparison of external fixation with illizarov method versus open reduction with locking plate in… full book for web 47 proceedings s.z.m.c. vol: 37(1): pp. 47-51, 2023. pszmc-876-37-1-2023 1department of surgery, azra naheed medical college, chaudhry mohammad akram hospital, lahore. , , 70 1department of orthopedics, thq, khanewal 2department of orthopedics, lgh, lahore 3department of orthopedics, thq, ferozwala 4department of orthopedics, cmh, lahore 5department of hand & upper limb surgery, cmh, lahore 6department of orthopedics, al-rehmat benevolent trust 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 48 combination of botulinum toxin and topical diltiazem versus lateral sphincterotomy in treatment of… 49 combination of botulinum toxin and topical diltiazem versus lateral sphincterotomy in treatment of… 50 combination of botulinum toxin and topical diltiazem versus lateral sphincterotomy in treatment of… 51 combination of botulinum toxin and topical diltiazem versus lateral sphincterotomy in treatment of… 02 6 proceedings s.z.m.c. vol: 37(1): pp. 06-10, 2023. pszmc-869-37-1-2023 2 1department of public health, health services academy, islamabad. 2department of management sciences, riphah int university, islamabad. 3department of medicine, ihitc, islamabad. 4department of rheumatology, pims, islamabad. 5department of medicine, dow university of health sciences, karachi. 70 1department of orthopedics, thq, khanewal 2department of orthopedics, lgh, lahore 3department of orthopedics, thq, ferozwala 4department of orthopedics, cmh, lahore 5department of hand & upper limb surgery, cmh, lahore 6department of orthopedics, al-rehmat benevolent trust 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 6 proceedings s.z.m.c. vol: 37(1): pp. 06-10, 2023. pszmc-869-37-1-2023 2 1department of public health, health services academy, islamabad. 2department of management sciences, riphah int university, islamabad. 3department of medicine, ihitc, islamabad. 4department of rheumatology, pims, islamabad. 5department of medicine, dow university of health sciences, karachi. 7 is covid vaccine hesitancy unfounded? a local comparative study of different covid vaccinations for….. 6 proceedings s.z.m.c. vol: 37(1): pp. 06-10, 2023. pszmc-869-37-1-2023 2 1department of public health, health services academy, islamabad. 2department of management sciences, riphah int university, islamabad. 3department of medicine, ihitc, islamabad. 4department of rheumatology, pims, islamabad. 5department of medicine, dow university of health sciences, karachi. 8 is covid vaccine hesitancy unfounded? a local comparative study of different covid vaccinations for….. 9 is covid vaccine hesitancy unfounded? a local comparative study of different covid vaccinations for….. 10 is covid vaccine hesitancy unfounded? a local comparative study of different covid vaccinations for….. untitled-1.indd 16 proceedings s.z.m.c. vol: 34(2): pp. 16-20, 2020. pszmc-746-34-2-2020 diagnostic significance of serum total bile acid levels in suspected cases of obstetric cholestasis 1hijab batool, 1muhammad dilawar khan, 2omar rasheed chughtai, 1shakeel ashraf 1department of clinical chemistry and immunology,chughtai institute of pathology, lahore 2director operations, chughtai institute of pathology, lahore abstract introduction: intrahepatic cholestasis of pregnancy (icp) is significantly associated with adverse maternal and fetal outcome and therefore accurate diagnosis and timely initiation of therapy is of utmost importance. aims & objectives: to find out the diagnostic significance of serum total bile acid measurement in suspected cases of obstetric cholestasis. place and duration of study: it was a cross sectional study conducted in the department of clinical chemistry and immunology, chughtai institute of pathology from july 2018 to february 2019. material & methods: two hundred pregnant females whose serum total bile acid (tba) was determined were included in the study. pregnant females with cholestasis due to medical reasons other than icp were excluded from the study. the patients were contacted for relevant clinical details which were recorded on a pre-designed proforma. data was analyzed using spss 23.0. univariate analysis was done to find out percentages and frequencies. results: mean age of 200 pregnant females was 27 years having mean tba value of 32.8 μmol/l. 63% females were primiparous. 197 (98.5%) patients gave the history of pruritus while 193 (96.5%) patients presented with dark urine. only 30 patients (15%) gave history of using progesterone during pregnancy. total bile acid value was >10 μmol/l for 84 primiparous and 48 multiparous cases (n=132). there was no significant difference in the mean tba value on the basis of parity, medication, mode of delivery or presence and absence of itch. conclusion: icp is a multi factorial reversible condition and bile acid estimation plays a major role in correct clinical assessment of patients with obstetric cholestasis. key words: total bile acid, obstetric cholestasis, pruritus, jaundice introduction bile acids are synthesized in the liver from cholesterol as precursor. primary bile acids are cholic and chenodeoxycholic acid which are metabolized in the intestinal lumen by action of bacterial enzyme 7-α-dehydroxylase. as a result of this metabolism secondary bile acids (deoxycholic and lithocholicacid) and a fraction of tertiary bile acid (ursodeoxycholicacid) are formed. glycine and taurine react with the bile acids to form conjugated bile acids. the term “total bile acids” refer to primary, secondary and tertiary bile acids along with their conjugates.1 bile is secreted by the liver and stored in the gallbladder of which bile acids are a major constituent. more than 90% of the bile acids which pass through git are reabsorbed from small intestine via enterohepatic circulation, processed in the liver and re-secreted into the gall bladder. impairment of enterohepatic circulation leads to increased level of bile acids in the serum and urine.2 metabolically bile acids have several important roles. these are the primary metabolic pathway for cholesterol catabolism, aid in digestion specifically lipid and lipid soluble vitamins and play a role in glucose and lipoprotein metabolism.3 bile acids are increased after food ingestion therefore a fasting sample is recommended.4 within a period of 24 hours, liver removes about 20 grams of bile salts from the circulation and abnormal bile acid levels indicate presence of liver disease.4,5 there can be many potential sites for the primary or secondary disturbances of bile acid levels. hepatocyte dysfunction can result in decreased level of primary bile acid with a decreased primary to secondary bile acid ratio. in the non-fasting state, there is decreased extraction from plasma leading to increased concentration of bile acids. on the other hand, in cholestatic disorders, there is increased concentration of total bile acids with increased 17 diagnostic significance of serum total bile acid levels in suspected cases of obstetric cholestasis primary to secondary bile acid ratio. in intestinal disorders, there is low concentration of both primary and secondary bile acids due to increased fecal loss.1 moreover, increase in total bile acid level can also be seen in hepatic coma, alcoholic liver disease, chemical and drug induced liver damage, liver cirrhosis and cystic fibrosis.6 some of the studies in animals have shown that bile acids may damage pancreatic cells and can lead to pancreatitis.7 the prevalence of icp is higher in south asian and south american populations as compared to european countries.8 according to a survey conducted in 2014 on 1000 obstetric patients in pakistan, the frequency of patients having icp was 3.1%.9 bile acid estimation is used as a marker for hepatic integrity and aids in the diagnosis of obstetric cholestasis which is one of the commonest causes of liver disease during gestational period. factors such as advanced maternal age, previous history of icp, positive family history, multiple pregnancies and use of oral contraceptive pills are found to be associated with increased risk of icp.10, 11the genetic predisposition becomes clear by the evidence of family clustering, ethnic variations and discovery of mutations in the genes encoding for hepatobiliary transport proteins.12 intrahepatic cholestasis of pregnancy (icp) is characterized by pruritus (unexplained) with increased level of serum bile acids and/or abnormal liver function tests in the second or third trimester of pregnancy. abnormal laboratory picture and pruritus occurs in the absence of other liver diseases and there is resolution of all the signs and symptoms within 2 to 3 weeks after delivery of the neonate.13,14 icp is significantly associated with adverse maternal and fetal outcome and therefore accurate diagnosis and timely initiation of therapy is of utmost importance. bile acid estimation plays a major role in correct clinical assessment of patients with obstetric cholestasis.5 non availability of serum bile acid analysis is a major limitation in majority of the studies and in many clinical settings the diagnosis of icp is made by exclusion of other medical conditions and clinical features. assessment of the diagnostic accuracy of tba may help the clinicians improve the approach towards the disease directly benefit outcome of pregnant females and their newborns. the range of values used to assess pregnant females is same as that of non pregnant population in most of the clinical laboratories i.e. 0-10 μmol/l.15 material and methods it was a cross sectional study conducted on pregnant females whose total bile acid test was performed. after obtaining permission from institutional ethical review board, pregnant females fulfilling the inclusion criteria whose serum tba were performed from july 2018 (since the test was included in the panel) to february 2019 were included in the study (n=200) the patients were contacted for history and any relevant clinical details. history along with mode of delivery and neonatal outcome (only if patient had delivered at the time of study) were recorded on a pre-designed proforma. serum total bile acids were analyzed on architect ci8200 by isoenzymatic cycling colorimetric method where bile acids in the sample are converted to their corresponding oxosteroids in the presence of excess nadh (nicotinamide adenine dinucleotide) and thio-nad + (thionicotinamide adenine dinucleotide). for quality assessment controls were run on a daily basis. statistical analysis: the results were analyzed statistically using spss 23.0. a univariate analysis was done using percentages, frequencies and means. mann-whitney test was used to find out difference between mean serum tba among different groups. a p value of <0.05 was considered significant. results mean age of patients was 27 ± 4 years (minimum age 18 years maximum age 39 years). mean tba value was 32.8 μmol/l (iqr 23.88, minimum 2.10 μmol/l-maximum 346.80 μmol/l). out of all the 200 pregnant females, 126 (63%) were primiparous and 74 (37%) females were multiparous. in our study, 197 (98.5%) females gave the history of pruritus whereas only 3 (1.5%) females did not have any pruritus (tba value was less than 10μmol/l) and they only requested the test because they were advised by their consultants. 193 (96.5%) females presented with dark urine whereas only 7 (3.5%) females did not give history of dark colored urine. 30 females (15%) gave history of using progesterone during their pregnancy as prescribed by their gynecologist and stated that they developed pruritus right after starting the medicine. 44(22%) females gave birth to the neonate via spontaneous vaginal delivery(svd). 127 (63.5%) females had caesarean section (c-section) while 29 (14.5%) females were still pregnant at the time of study. among the multiparous females, 12 females who gave birth vaginally in their previous pregnancies had to undergo c-section due to threatened complications. total bile acid value was more than 18 diagnostic significance of serum total bile acid levels in suspected cases of obstetric cholestasis 10μmol/l for 84 primiparous and 48 multiparous females (n=132). among the 30 females who took progesterone in their pregnancy, 17 had tba value more than 10 μmol/l. there was no significant difference between the tba value among primiparous and multiparous women. similarly, the women who took progesterone during their pregnancy had mean tba value of 24.1 μmol/l, which was not significantly different from the women who did not take progesterone. mean tba value for women who underwent c-section was 32.8μmol/l and 40.4μmol/l for women who had a normal vaginal delivery. there was no significant difference in mean serum total bile acid levels in various groups of patients (p-value >0.05 mannwhitney test) (table-1). groups mean tba value (μmol/l) p value primiparous females 35.6 0.340 multiparous females 32.4 women who took progesterone 24.1 0.341 women who did not take progesterone 34.2 woman who delivered via spontaneous vaginal delivery 40.4 0.395 woman who delivered via c section 32.8 woman presented with itching 32.7 0.786 woman with no history of itching 27.4 table-1: serum total bile acid levels in various groups of patients (n=200) discussion icp appear as a hepatic disorder in pregnancy and is the second most common cause of jaundice in pregnancy is icp following viral hepatitis.16,17 a combination of multiple factors is responsible for the development of obstetric cholestasis including environmental and hormonal factors acting in conjugation with genetic tendency.18 a multiethnic study conducted in united kingdom revealed that the incidence of icp in multiethnic population was 0.7% where as in indian and pakistani population this incidence was 1.2 to 1.5%.19 in an indian study the diagnosis of icp was made only on the basis of liver function tests and clinical features of patients and the authors concluded that in the absence of liver function tests, aminotransferases can aid in the diagnosis of icp.20 more than 90% of the females in our study gave history of pruritus. a pakistani study showed similar results in which pruritus was present in 85% of the patients.9 majority of the patients (63%) had delivered via c section in our study and the results are similar to the above stated study in which 58% of the patients had c-section.9 a systematic review published in 2019 stated that a cut off value of 10umol/l for tba had an overall sensitivity ranging from 72% to 98% and specificity ranging from 81% to 97%.21a retrospective study conducted in china revealed that pregnant females had increased fasting serum bile acid levels even without the presence of symptoms and these asymptomatic pregnant females had similar or a higher risk of still birth than patients with icp.22 demir along with his colleagues while establishing relationship between serum bile acid levels and maternal and fetal outcomes found that mean tba level in preterm birth group was significantly higher as compared to the term birth group.23 in another study determining the role of bile acid ratio of cholic acid to chenodeoxycholic acid in diagnosis of icp stated that the ratio contributed to the diagnosis of icp and use of liver function tests in the diagnosis of icp with bile acid ratios decreased the percentage of positive results.24 yannik bacq in 2014 concluded in his study that serum total bile acid levels can be used to diagnose cases of icp and can also be used as a followup investigation.25 some studies have shown that tba level may rise up to 25 fold in suspected cases of obstetric cholestasis. also there can be a marked rise in the ratio of cholic and chenodeoxycholic acid in icp. a triad of increased tba, elevated cholic acid level and a decreased glycine to taurine ratio can be used to exclude other causes and aid in differential diagnosis. elevated alt, ast and mild hyperbilirubinemia (rarely >6mg/dl) may further help to reach a final diagnosis.8 other investigations which might help in timely diagnosis are plasma glucose levels and clotting profile of the patient.17 in the absence of bile acids, obstetric cholestasis may be diagnosed with the help of clinical findings and elevated transaminases, provided both of these resolve after delivery.26 pregnancy specific reference ranges are necessary for proper interpretation of results. egan along with his colleagues conducted a study in which he proposed reference ranges of total bile acid in pregnancy in relation with gestational age. the authors found that value of serum bile acids ranged from 0.3 to 9.8 μmol/l, which is considerably different from the range given for non-pregnant patients.2 another chinese study found the mean tba value raised from 24th gestational week (range 19 diagnostic significance of serum total bile acid levels in suspected cases of obstetric cholestasis 0.3-8.6 μmol/l) and reached its maximum at 36th gestational week with a mean of 2.97 μmol/l (range 0.5-12.1μmol/l).27 lee et al conducted a study to find out bile acid concentration reference ranges in latin pregnant females and found a range of <8.5 μmol/l.28 non availability of an established reference range of tba in pregnancy is a major limitation in accurate diagnosis of icp. it is very important to create clinical awareness among the general population regarding icp. early diagnosis with timely initiation of therapy is important for improved fetal prognosis in icp. availability of bile acid testing in clinical settings can be very helpful in this regard. due to nonavailability of this test in many clinical settings of pakistan, the diagnosis of icp is based on exclusion. further a reference range of tba according to gestational age should be established. this pregnancy related tba reference range will be helpful to physicians for correct diagnosis of icp. conclusion in our study serum total bile acid measurement provided valuable clinical information while diagnosing icp. serum total bile acid was raised in cases of intra hepatic cholestasis of pregnancy. conflict of interest: in accordance with our ethical obligation as researchers we are declaring that omar rasheed chughtai is director operations at chughtai institute of pathology. limitations: our study is a single centered study with limited sample size. further supporting studies from multiple clinical settings with larger sample size are suggested to provide more clinical evidence relating to diagnostic accuracy of serum total bile acids. our desire was to share the findings to create a parameter which can be replicated in other studies. references 1. burtis ca, ashwood er, bruns de. tietz textbook of clinical chemistry and molecular diagnostics. 5th ed. st. louis, mo: elsevier saunders. 2012; 1641-43. 2. egan n, bartels a, khashan a, et al. reference standard for serum bile acids in pregnancy. bjog. 2012; 119:493-8. 3. deaguiarvallim tq, tarling ej, edwards pa, pleiotropic roles of bile acids in metabolism. cell metab. 2013; 17:657-69. 4. mcpherson ra, pincus mr. henry's clinical diagnosis and management by laboratory methods. 21sted. philadelphia, pa: elsevier; 2007; 266-7. 5. hofmann af, hagey lr. key discoveries in bile acid chemistry and biology and their clinical applications: history of the last eight decades. j lipid res. 2014; 55:1553-95. 6. maleszka a, dumnicka p, matuszyk a, pędziwiatr m, mazur-laskowska m, sporek m et al. the diagnostic usefulness of serum total bile acid concentrations in the early phase of acute pancreatitis of varied etiologies. international journal of molecular sciences. 2017; 18(1):106. 7. venglovecz v, rakoczay z, hegyi p. the effects of bile acids on pancreatic ductal cells. panceapedia. 2012. 8. ozkan s, ceylan y, ozkan o, yildirim s. review of a challenging clinical issue: intrahepatic cholestasis of pregnancy. world journal of gastroenterology.2015;21(23):7134-41 9. hafeez m, ansari a, parveen s, salamat a, aijaz a. frequency of intrahepatic cholestasis of pregnancy in punjab pakistan: a single centre study. jpak med assoc.2016;66(2):203-6 10. lee nm, brady cw .liver disease in pregnancy. world j gastroenterol. 2009 feb 28; 15(8):897-906. 11. williamson c, geenes v. intrahepatic cholestasis of pregnancy. obstet gynecol. 2014 jul; 124(1):120-33. 12. anzivino c, odoardi mr, meschiari e, baldelli e, facchinetti f, neri i et al. abcb4 and abcb11 mutations in intrahepatic cholestasis of pregnancy in an italian population. dig liver dis. 2013 mar; 45(3):226-32. 13. lammert f, marschall hu, glantz a, matern s. intrahepatic cholestasis of pregnancy: molecular pathogenesis, diagnosis and management. j hepatol.2000; 33:1012-21. 14. beuers u, pusl t. intrahepatic cholestasis of pregnancy-a heterogeneous group of pregnancy related disorders. hepatology. 2006; 43:647-9. 15. geenes v, williamson c. intrahepatic cholestasis of pregnancy. world j gastroenterol 2009; 15:2049-66. 16. diken z, usta im, nassar ah. a clinical approach to intrahepatic cholestasis of pregnancy. am j perinatol. 2014 jan; 31(1):1-8. 17. saleh mm, abdo kr j. intrahepatic cholestasis of pregnancy: review of the literature and evaluation of current evidence. women’s health (larchmt). 2007; 16(6):833-41. 20 diagnostic significance of serum total bile acid levels in suspected cases of obstetric cholestasis 18. than nn, neuberger j. liver abnormalities in pregnancy. best pract res clingastroenterol. 2013; 27(4):565-75. 19. madazli r, yuksel ma, oncul m, tuten a, guralp o, aydin b. pregnancy outcomes and prognostic factors in patients with intrahepatic cholestasis of pregnancy. j obstetgynaecol 2015; 35:358-61. 20. singh g, sidhu k. cholestasis of pregnancy: a prospective study. medical journal armed forces india. 2008; 64(4):343-345. 21. manzotti c, casazza g, stimac t, nikolova d, gluud c. total serum bile acids or serum bile acid profile, or both, for the diagnosis of intrahepatic cholestasis of pregnancy. cochrane database of systematic reviews 2019, issue 7. 22. feng d, he w. asymptomatic elevated total serum bile acids representing an unusual form of intrahepatic cholestasis of pregnancy. international journal of gynecology & obstetrics. 2016; 134(3):343-344. 23. demir b, güneş e, atalay m. intrahepatic cholestasis of pregnancy: relationship between bile acid levels and maternal and fetal complications. journal of turkish society of obstetric and gynecology. 2014;11(3):148-152. 24. huang w, gowda m, donnelly j. bile acid ratio in diagnosis of intrahepatic cholestasis of pregnancy. american journal of perinatology. 2008; 26(04):291-294. 25. bacq y, sentilhes l. intrahepatic cholestasis of pregnancy: diagnosis and management. clinical liver disease. 2014; 4(3):58-61. 26. geenes v, williamson c, chappell l. intrahepatic cholestasis of pregnancy. the obstetrician &gynaecologist.2016;18(4):273-81 27. chen l, wen h, zhu b, qian y, he j. reference standard of total bile acid concentration of chinese pregnant women: analyses of 11022 chinese pregnant women. int j clinexp med. 2018; 11(7):7312-7318. 28. lee hr, ouzounian j, goodwin t, incerpi m, miller d, zhang k, caulfield m et al. bile acid concentration reference ranges in a pregnant latina population. amer j perinatol 2013; 30(05): 389-394. the authors: dr. hijab batool resident pathologist, department of clinical chemistry and immunology, chughtai institute of pathology, lahore prof. muhammad dilawar khan professor of chemical pathology, department of clinical chemistry and immunology, chughtai institute of pathology, lahore dr. omar rasheed chughtai assistant professor of histopathology, director operations, chughtai institute of pathology, lahore shakeel ashraf technical supervisor, department of clinical chemistry and immunology, chughtai institute of pathology, lahore corresponding author: dr. hijab batool resident pathologist, department of clinical chemistry and immunology, chughtai institute of pathology, lahore e-mail: batool.hijab@gmail.com 08 34 proceedings s.z.m.c. vol: 37(2): pp. 34-38, 2023. pszmc-894-37-2-2023 mental health impact on patients of esrd on renal replacement therapy: a cross sectional survey using beck and deck inventory 1rashid ali, 1sardar sabeel, 2danial zahoor, 1bushra arif, 1adnan khan, 1saqib malik 1department of medicine, ayub teaching hospital, abbottabad. 2deprtment of surgery, jpmc, karachi. abstract introduction: chronic kidney disease (ckd) affects approximately 8 to 16% of population worldwide. with increasing incidence of diabetes mellitus and an aging population, ckd is putting an enormous burden on health care resources. the prevalence of ckd in pakistan has been estimated to vary between 5%-12.5%. depression can be found in 25% to 42% of hemodialysis patients. there are no previous studies done in pakistan to know the mental health impact in endstage renal disease (esrd)patients undergoing dialysis. aims &objectives: to assess the mental health impact on patients of esrd on renal replacement therapy using beck and deck inventory in our population. place and duration of study: department of medicine, ayub teaching hospital, abbottabad from february 2019 to july 2019. material & methods: 97 patients were enrolled using non-probability, consecutive sampling. patients having ckd and on dialysis for 3 months from both genders from ages 15 to 60 years were included in the study. patients were assessed using dsm-iv/bdi criteria for presence of depression. data was entered and analyzed using spss version 20 results: of the 97 patients included in the study 57 (58.8%) were male and 40 (41.2%) were females. the mean age of patients was 45.00 ±14.82 years. the mean length of time since the patients were on dialysis was 20.47 ±22.81 months. hypertension was the commonest cause of ckd (75.3%). the mean bdi score was 15.25 ±7.46. the number of patients who had no or minimal depression was 40 (41.2%). the number of patients having mild, moderate and severe depre ssion was 57 (58.8%) patients. single patients were more likely to be normal (77.8%) compared to married individuals (37.5%). of the patients who were on dialysis for more than 48 months, 75% were normal with no element of depression. of the patients who were on dialysis for less than a year, 59.6% had depression. this was statistically significant. conclusion: depression is more prevalent in esrd patients on hemodialysis. it is more prevalent than found in other major illnesses and in the general population. it should be recognized earlier so that these patients can be treated, and they can have a better quality of life. keywords: depression, esrd, dialysis, chronic kidney failure. introduction chronic kidney disease (ckd) is rapidly becoming a global health concern. there has been a steep rise in the number of patients reaching endstage renal disease (esrd). ckd affects an approximately 8 to 16% of population worldwide1. ckd, as a cause of death, has climbed from 27thto 18th position in two decades2.this has translated to 82.3% rise in the number of deaths caused by ckd in the last twenty years. this is the third largest increase among the top 25 causes of deaths, paralleled only by aids and diabetes3.ckd is an alarming public health priority also because of the fact that the number of esrd patients on renal replacement therapy is approximately 1.4 million and this number is increasing by 8% annually4. with increasing incidence of diabetes mellitus and an aging population, ckd is putting an enormous burden on health care resources5. in western countries, the prevalence of ckd varies between 5.8% (poland) to 14.8% (united states) in the adult population6. however, prevalence of ckd increases in diabetic people, in whom it varies between 34.7 and 45.4% according to different populations7. the prevalence of ckd in pakistan has been studied very sparsely. it has been estimated to vary between 5% to 12.5% in a review published by imtiaz s. et al.in 20188. most of the esrd patients undergo renal replacement therapy (hemodialysis) in a hospital setting. although this has increased life expectancy in ckd patients, many studies suggest that these patients often suffer from poor quality of life9,10. 8 intraoperative peritoneal lavage in peritonitis: normal saline vs metronidazole 1hira aslam, 2muhammad khurram jameel, 2pir muneeb, 1aneeqa nasir qureshi, 2fawad hameed, 3muhammad ammar, 2maham qazi, 4abrar ul hassan pirzada 1department of surgery, mayo hospital, kemu, lahore 2department of surgery, anmc / chaudhary muhammad akram hospital, lahore 3department of urology, chaudhary muhammad akram hospital, lahore 4department of plastic surgery, anmc / chaudhary muhammad akram hospital, lahore, pakistan abstract introduction: globally, acute generalized peritonitis ranks among the top surgical emergencies. different studies have been conducted to show the amplitude of peritonitis worldwide eliciting a huge impact on overall patient morbidity and mortality. largely peritonitis is caused by a gastrointestinal perforation or anastomotic leak. in peritonitis, anaerobes & gram-negative organisms are mostly responsible for sepsis and morbidity due to the overactive inflammatory cascade by endotoxins which is amenable to timely intervention. aims & objectives: the study's aim was to evaluate whether using normal saline or metronidazole solution during intraoperative peritoneal lavage (iopl), results in a lower rate of postoperative wound infection. place and duration of study: this study was undertaken at the south surgical ward, mayo hospital lahore for 6 months from february 2nd, 2021, to august 1st, 2021. material & methods: consecutive sampling strategy followed by a randomized controlled trial were used to induct and provide intervention to 90 patients aged 15-65 years with peritonitis caused by hollow viscus perforation. the patients were subdivided into 2 groups a& b(n=45 each).two liters of normal saline were used for peritoneal lavage in group a, while two liters of normal saline were combined with 200 ml of metronidazole solution and administered to group b. intraoperatively.baseline physiological parameters such as age, sex, bmi , intra operative surgical parameters as duration of operation and post-operative course were recorded till discharge. on 10th pod, patients returned to opd for further monitoring. an infection was diagnosed if the patient had post-operative symptoms such as a high temperature, increased tlc, wound discharge, redness, or pain. data was entered and analyzed using spss version 23. results: a majority (54.44%) of the patients were young adults. mean age of 37.33 ± 10.53 years of patients in the metronidazole group was comparable to mean age 40.04 ± 11.96 years in the saline group, difference was not significant (p=0.067). male/female ratio in metronidazole and saline groups were 17/25 and 10/18, respectively. patients who received intraperitoneal lavage with normal saline were more likely to develop wound infections (17/45) (37.78%), while only 3/45) (6.67% of those who received metronidazole solution did so (p 0.0001). conclusion: based on the results of this experiment, using metronidazole solution for intraoperative peritoneal lavage instead of normal saline reduces the occurrence of postoperative wound infection. keywords: peritonitis, postoperative wound infection, intraoperative peritoneal lavage introduction globally, acute generalized peritonitis ranks among the top surgical emergencies1. it is more common in third world nations. the prevalence of perforation is low (0.6% 4.9%) in developed nations but high (33% 63%) in west africa2. 554 persons were discovered to have peritonitis in a study that took place over three years in india3. researchers in pakistan have conducted studies with similar methods, with one study reporting 650 cases in a just 9 months4. most cases of peritonitis are caused by a gastrointestinal perforation or anastomotic leak5. in the case of peritonitis, anaerobes and gramnegative organisms are mostly responsible for sepsis and morbidity due to the overactive inflammatory cascade brought on by the release of endotoxins5.clinical evidence is used to identify peritonitis. diagnosis can be achieved via upright plain x-ray of the abdomen, usg, or ct scan. this is often done through diagnostic laparoscopy nowadays6.resuscitation, diagnosis, prompt exploration, treatment of the underlying cause, and extensive surgical peritoneal lavage have always been the cornerstones of peritonitis therapy regimens (iopl)7,8. regular iopl is performed to lessen bacterial contamination and burden. even though large volumes of normal saline are used in iopl, the rates of sepsis, wound infection, and mortality remain alarmingly high. another method 6 1department of public health, health services academy, islamabad. 2department of management sciences, riphah int university, islamabad. 3department of medicine, ihitc, islamabad. 4department of rheumatology, pims, islamabad. 5department of medicine, dow university of health sciences, karachi. 70 1department of orthopedics, thq, khanewal 2department of orthopedics, lgh, lahore 3department of orthopedics, thq, ferozwala 4department of orthopedics, cmh, lahore 5department of hand & upper limb surgery, cmh, lahore 6department of orthopedics, al-rehmat benevolent trust 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 1dr. adnan, 1saqib malik 35 mental health impact on patients of esrd on renal replacement therapy:a cross sectional survey….. depression is frequently associated with hemodialysis in ckd patients11. according to the american psychological association's diagnostic and statistical manual-v, depression is characterized by low mood, a decreased interest or pleasure in activity, and at least three of seven other co-occurring symptoms present for a period of more than two weeks12. depression can be found in patients in 25% to 42% of hemodialysis patients13,14. further, depression frequently goes unrecognized in ckd patients on dialysis and it significantly affects the quality of life in such patients15. there are no previous studies done in pakistan to know the mental health impact in esrd patients undergoing dialysis. therefore, this study was planned to assess the problem in our population. material and methods this cross-sectional study was conducted in the department of medicine, ayub teaching hospital, abbottabad from february 2019 to july 2019. irb clearance was received vide letter number (884-1) sample size was calculated to be 97 cases with 95% confidence level, 4% margin of error and taking expected frequency depression in ckd patients on dialysis as 7.8%16.non-probability, consecutive sampling was used. all patients having esrd who were on dialysis for 3 months from both genders from ages 15 to 60 years were included in the study. patients having previous history of depressive illness or mood disorders, antidepressant or antipsychotic use, history of stroke, bed ridden patients or having any malignancy were excluded from the study. after taking consent, patients were assessed using dsm-iv/beck and deck inventory (bdi) criteria for presence of depression. demographic data was also collected on a proforma. data was analysed by using spss-20. results a total of 97 patients were included in the study of which 57 (58.8%) were male and 40 (41.2%) were females. the mean age of the patient’s was 45.00 ±14.82 years and the age ranged between 11 years and 80 years. the mean length of time since the patients were on dialysis was 20.47 ± 20.0 months. the mean time since the patients were diagnosed as having esrd was 24.20 ± 24.08 months. hypertension was the commonest cause of ckd (75.3%). most of the patients, 93 (96%), were on twice weekly dialysis. the average time the patients travelled to the hospital for dialysis was 2.35 ±1.91 hours. only 2% of patients were either hepatitis b or c positive before the initiation of dialysis. 44.3% patients contracted hepatitis c after being put on dialysis. no patient contracted hepatitis b after starting dialysis. 85.6% patients did not associate any adverse outcomes with previous dialysis. the mean bdi score was 15.25 ±7.46. the number of patients who had no or minimal depression was 40 (41.2%). the number of patients having mild, moderate and severe depression was 57 (58.8%) patients. (table-1). results show significant association between adverse outcome during previous dialysis and the level of depression. patients who did not experience any adverse outcome in previous sessions of dialysis were more likely to be normal (30%) compared to those that had an adverse outcome (7%). marital status was also associated with a significant difference in the level of depression. single patients were more likely to be normal (77.8%) compared to married individuals (37.5%). mean bdi scores were 9.5 in single individuals and 15.8 in married individuals. this association was statistically significant, indicating that married individuals were more likely to be depressed. there was no significant difference in mean bdi scores in educated and uneducated individuals. results showed significant association between total duration the patient was on dialysis and the level of depression. of the patients who were on dialysis for more than 48 months, 75% were normal with no element of depression. of the patients who were on dialysis for less than a year, 59.6% had depression. bdi categories number of participants (%) minimal depression (0-13) 40 (41.2) mild depression (14-19) 29 (29.9) moderate depression (20-28) 23 (23.7) severe depression (29-63) 5 (5.2) table-1: bdi categories and number of participants discussion the most common psychiatric disorder in esrd patients is depression17. it has been associated with an increased risk of cardiovascular events, hospitalization and death18. psychological stress and disturbing physical symptoms lead to reduced quality of life (qol) which shows the way to depression19. many studies have shown a significant association between depression and mortality in esrd patients on dialysis20. 8 intraoperative peritoneal lavage in peritonitis: normal saline vs metronidazole 1hira aslam, 2muhammad khurram jameel, 2pir muneeb, 1aneeqa nasir qureshi, 2fawad hameed, 3muhammad ammar, 2maham qazi, 4abrar ul hassan pirzada 1department of surgery, mayo hospital, kemu, lahore 2department of surgery, anmc / chaudhary muhammad akram hospital, lahore 3department of urology, chaudhary muhammad akram hospital, lahore 4department of plastic surgery, anmc / chaudhary muhammad akram hospital, lahore, pakistan abstract introduction: globally, acute generalized peritonitis ranks among the top surgical emergencies. different studies have been conducted to show the amplitude of peritonitis worldwide eliciting a huge impact on overall patient morbidity and mortality. largely peritonitis is caused by a gastrointestinal perforation or anastomotic leak. in peritonitis, anaerobes & gram-negative organisms are mostly responsible for sepsis and morbidity due to the overactive inflammatory cascade by endotoxins which is amenable to timely intervention. aims & objectives: the study's aim was to evaluate whether using normal saline or metronidazole solution during intraoperative peritoneal lavage (iopl), results in a lower rate of postoperative wound infection. place and duration of study: this study was undertaken at the south surgical ward, mayo hospital lahore for 6 months from february 2nd, 2021, to august 1st, 2021. material & methods: consecutive sampling strategy followed by a randomized controlled trial were used to induct and provide intervention to 90 patients aged 15-65 years with peritonitis caused by hollow viscus perforation. the patients were subdivided into 2 groups a& b(n=45 each).two liters of normal saline were used for peritoneal lavage in group a, while two liters of normal saline were combined with 200 ml of metronidazole solution and administered to group b. intraoperatively.baseline physiological parameters such as age, sex, bmi , intra operative surgical parameters as duration of operation and post-operative course were recorded till discharge. on 10th pod, patients returned to opd for further monitoring. an infection was diagnosed if the patient had post-operative symptoms such as a high temperature, increased tlc, wound discharge, redness, or pain. data was entered and analyzed using spss version 23. results: a majority (54.44%) of the patients were young adults. mean age of 37.33 ± 10.53 years of patients in the metronidazole group was comparable to mean age 40.04 ± 11.96 years in the saline group, difference was not significant (p=0.067). male/female ratio in metronidazole and saline groups were 17/25 and 10/18, respectively. patients who received intraperitoneal lavage with normal saline were more likely to develop wound infections (17/45) (37.78%), while only 3/45) (6.67% of those who received metronidazole solution did so (p 0.0001). conclusion: based on the results of this experiment, using metronidazole solution for intraoperative peritoneal lavage instead of normal saline reduces the occurrence of postoperative wound infection. keywords: peritonitis, postoperative wound infection, intraoperative peritoneal lavage introduction globally, acute generalized peritonitis ranks among the top surgical emergencies1. it is more common in third world nations. the prevalence of perforation is low (0.6% 4.9%) in developed nations but high (33% 63%) in west africa2. 554 persons were discovered to have peritonitis in a study that took place over three years in india3. researchers in pakistan have conducted studies with similar methods, with one study reporting 650 cases in a just 9 months4. most cases of peritonitis are caused by a gastrointestinal perforation or anastomotic leak5. in the case of peritonitis, anaerobes and gramnegative organisms are mostly responsible for sepsis and morbidity due to the overactive inflammatory cascade brought on by the release of endotoxins5.clinical evidence is used to identify peritonitis. diagnosis can be achieved via upright plain x-ray of the abdomen, usg, or ct scan. this is often done through diagnostic laparoscopy nowadays6.resuscitation, diagnosis, prompt exploration, treatment of the underlying cause, and extensive surgical peritoneal lavage have always been the cornerstones of peritonitis therapy regimens (iopl)7,8. regular iopl is performed to lessen bacterial contamination and burden. even though large volumes of normal saline are used in iopl, the rates of sepsis, wound infection, and mortality remain alarmingly high. another method 6 1department of public health, health services academy, islamabad. 2department of management sciences, riphah int university, islamabad. 3department of medicine, ihitc, islamabad. 4department of rheumatology, pims, islamabad. 5department of medicine, dow university of health sciences, karachi. 70 1department of orthopedics, thq, khanewal 2department of orthopedics, lgh, lahore 3department of orthopedics, thq, ferozwala 4department of orthopedics, cmh, lahore 5department of hand & upper limb surgery, cmh, lahore 6department of orthopedics, al-rehmat benevolent trust 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 36 mental health impact on patients of esrd on renal replacement therapy:a cross sectional survey….. a systemic review and meta-analysis which included 249 populations and 55,982 participants showed that prevalence of depression to be 39.3% in esrd patients21. in a study done by chi-ken chen et al, on 200 patients on hemodialysis, depression was present in 35.0% of patients22. however, the estimates of depression in esrd on hemodialysis patients have varied between 5% to 58%23. the prevalence of depression in our study is 58.8%. this rate is higher than that reported in many western populations. however, it corresponds to the levels of depression in indian sub-continent. a study done in western rajasthan, india showed the prevalence of depression in hemodialysis patients to be 61%24. similarly, kumar et al. showed a prevalence of depression to be 61.3% in ckd patients on hemodialysis25. in our study, married people were likely to be depressed that single individuals. this is in contrast to some other studies which found that married people were less likely to be depressed and had better qol than widowed/divorced individuals26,27. however, amjad khan et al., found that married people were more likely to be depressed than single individuals. they found 85.6% of married people to be depressed28. in our study, 62.5% married individuals were depressed. the difference could be explained by the fact that the first two studies compared married to widowed/divorced individuals, whereas amjad khan et al. and our study compared married to single unmarried individuals. a study done in pakistan by anees m et al. also showed that married individuals are more likely to be depressed than single individuals29. in our social setup, a married individual is usually a guardian of 4-8 dependents. as the disease process affects their employment status, this leads to increased mental stress as there is no social support for the family. this could cause higher levels of depression. in our study, the duration of dialysis was associated with the level of depression in an individual. of the patients who were on dialysis for less than a year, 59.6% were found to have depression, whereas patients who were on dialysis for more than 4 years, 75% of them had no element of depression. this corresponds to findings of some other studies which propose that the commencement of dialysis puts a significant emotional and social stress on an individual, and as the duration increases the patient gradually adapts to the burden of the disease30,31. although treatment can be challenging, it is very important to identify depression early in esrd patients. pharmacological and cognitive behavioral therapy (cbt) can lead to improved quality of life in these patients32,33. conclusion depression is more prevalent in esrd patients on hemodialysis. it is more prevalent than found in other major illnesses and in the general population29,31. it should be recognized earlier so that these patients can be treated and they can have a better quality of life. references 1. jha v, garcia-garcia g, iseki k, li z, naicker s, plattner b, et al. chronic kidney disease: global dimension and perspectives. lancet 2013;382(9888):260-72. 2. lozano r, naghavi m, foreman k, lim s, shibuya k, aboyans v. global and regional mortality from 235 causes of death for 20 age groups in 1990 and 2010: a systematic analysis for the global burden of disease study2010. lancet 2012;380(9859):2095128. 3. radhakrishnan j, remuzzi g, saran r, williams de, rios-burrows n, powe n. taming the chronic kidney disease epidemic: a global view of surveillance efforts. kidney int2014;86(2):246-50. 4. white sl, chadban sj, jan s, chapman jr, cass a. how can we achieve global equity in provision of renal replacement therapy? bull world health organ 2008;86(3):229-37. 5. couser wg, remuzzi g, mendis s, tonelli m.the contribution of chronic kidney disease to the global burden of major noncommunicable diseases. kidney int2011;80(12):1258-70. 6. de nicola l, zoccali c. chronic kidney disease prevalence in the general population: heterogeneity and concerns. nephrol dial transplant 2016;31(3):331-5. 7. wu b, bell k, stanford a, kern dm, tunceli o, vupputuri s, et al. understanding ckd among patients with t2dm: prevalence, temporal trends, and treatment patterns-nhanes 2007-2012.bmj open diabetes res care 2016;4(1):e000154. 8. imtiaz s, salman b, qureshi r, drohlia mf, ahmad a. a review of the epidemiology of chronic kidney disease in pakistan: a global and regional perspective. saudi j kidney dis transpl 2018;29(6):1441-1451. 9. merkus mp, jager kj, dekker fw, boeschoten ew, stevens p, krediet rt. quality of life in patients on chronic dialysis: self-assessment 3 months after the start of treatment. the necosad study group.am j kidney dis 1997;29(4):584-92. 10. kang gw, lee ih, ahn ks, lee j, ji y, woo j. clinical and psychosocial factors predicting healthrelated quality of life in hemodialysis patients. hemodial int 2015;19(3):439-46. 37 mental health impact on patients of esrd on renal replacement therapy:a cross sectional survey….. 11. king-wing ma t, kam-tao li p.depression in dialysis patients.nephrology (carlton) 2016;21(8):639-46. 12. miller db, o'callaghan jp. personalized medicine in major depressive disorder -opportunities and pitfalls. metabolism 2013;62suppl 1:s34-9. 13. son yj, choi ks, park yr, bae js, lee jb. depression, symptoms and the quality of life in patients on hemodialysis for end-stage renal disease. am j nephrol 2009;29(1):36-42. 14. teles f, azevedo vf, miranda ct, miranda mp, teixeira mdo c, elias rm. depression in hemodialysis patients: the role of dialysis shift. clinics (sao paulo) 2014;69(3):198-202. 15. farragher jf, polatajko hj, jassal sv. the relationship between fatigue and depression in adults with end-stage renal disease on chronic inhospital hemodialysis: a scoping review. j pain symptom manage 2017;53(4):783-803.e1. 16. hedayati ss, bosworth hb, briley lp, sloane rj, pieper cf, kimmel pl, et al. death or hospitalization of patients on chronic hemodialysis is associated with a physician-based diagnosis of depression. kidney international 2008;74(7):930–6. 17. kimmel pl, cukor d, cohen sd, peterson ra. depression in end-stage renal disease patients: a critical review. adv chronic kidney dis 2007;14(4):328-34. 18. ibrahim s, el salamony o. depression, quality of life and malnutrition-inflammation scores in hemodialysis patients. am j nephrol 2008;28(5):784-91. 19. kimmel pl, patel ss. quality of life in patients with chronic kidney disease: focus on end-stage renal disease treated with hemodialysis. semin. nephrol2006;26:68-79. 20. farrokhi f, abedi n, beyene j, kurdyak p, jassal sv. association between depression and mortality in patients receiving long-term dialysis: a systematic review and meta-analysis. am j kidney dis 2014;63(4):623-35. 21. palmer s, vecchio m, craig jc, tonelli m, johnson dw, nicolucci a, pellegrini f, saglimbene v, logroscino g, fishbane s, strippoli gf. prevalence of depression in chronic kidney disease: systematic review and meta-analysis of observational studies. kidney int. 2013;84(1):179-91. 22. chen ck, tsai yc, hsu hj, wu iw, sun cy, chou cc, et al. depression and suicide risk in hemodialysis patients with chronic renal failure. psychosomatics 2010;51(6):528-528.e6. 23. murtagh fe, addington-hall j, higginson ij. the prevalence of symptoms in end-stage renal disease: a systematic review. adv chronic kidney dis 2007;14(1):82-99. 24. gadia p, awasthi a, jain s, koolwal gd. depression and anxiety in patients of chronic kidney disease undergoing haemodialysis: a study from western rajasthan. j family med prim care 2020;9(8):4282-4286. 25. kumar v, khandelia v, garg a. depression and anxiety in patients with chronic kidney disease undergoing hemodialysis. ann indian psychiatry 2018;2:115–9. 26. theofilou p. depression and anxiety in patients with chronic renal failure: the effect of sociodemographic characteristics. int j nephrol ;2011:514070. 27. chiang ck, peng ys, chiang ss, yang cs, he yh, hung ky, et al. health-related quality of life of hemodialysis patients in taiwan: a multicenter study. blood purif 2004;22(6):490-8. 28. khan a, khan ah, adnan as, sulaiman sas, mushtaq s. prevalence and predictors of depression among hemodialysis patients: a prospective followup study. bmc public health. 2019;19(1):531. 29. anees m, barki h, masood m, mumtaz a, kausar t. depression in hemodialysis patients. pak j med sci 2008;24(4):560-5. 30. elkheir hk, wagaella as, badi s, khalil a, elzubair th, khalil a, et al. prevalence and risk factors of depressive symptoms among dialysis patients with end-stage renal disease (esrd) in khartoum, sudan: a cross-sectional study. j family med prim care 2020;9(7):3639-3643. 31. dziubek w, kowalska j, kusztal m, rogowski , go biowski t, nikifur m, et al. the level of anxiety and depression in dialysis patients undertaking regular physical exercise training--a preliminary study. kidney blood press res. 2016;41(1):86-98. 32. gregg lp, hedayati ss. pharmacologic and psychological interventions for depression treatment in patients with kidney disease. curropin nephrol hypertens 2020;29(5):457-464. 33. lerma a, perez-grovas h, bermudez l, peraltapedrero ml, robles-garcía r, lerma c. brief cognitive behavioural intervention for depression and anxiety symptoms improves quality of life in chronic haemodialysis patients. psychol psychother 2017;90(1):105-123. the authors: dr. rashid ali, assistant professor, department of medicine ayub teaching hospital,abbottabad dr. sardar sabeel, post graduate trainee, department of medicine, ayub teaching hospital, abbottabad dr. danial zahoor, post graduate trainee, department of surgery, jpmc, karachi 38 mental health impact on patients of esrd on renal replacement therapy:a cross sectional survey….. dr. bushra arif, post graduate trainee, pulmonology unit, ayub teaching hospital, abbottabad dr. adnan khan, assistant professor, department of medicine ayubteaching hospital, abbottabad dr. saqib malik, professor, department of medicine ayub teaching hospital,abbottabad corresponding author: dr. adnan khan, assistant professor, department of medicine, ayub medical college, abbottabad email:adnanjrmcw1983@gmail.com dr. adnan, dr. adnan, full book for printing 50 abstract real time pcr or rt-qpcr (including reverse transcriptase pcr) is a gold standard in diagnostic testing for pathogens such as bacteria and viruses that contain rna or dna for amplification. as a validated molecular assay technique, rt qpcr has been instrumental in microbial detection because of its high throughput, quantification of dna or rna samples as well as quick turnaround especially when compared with culturing methods that can take multiple days to generate result. while the advancement of pcr has had significant impact for diagnosis of diseases such as covid 19through its high sensitivity and specificity, the test also brings with it its own challenges of false positive and false negative results. this is normally attributed to pcr's susceptibility to contamination or inhibitors. however, there are other conditions like dna extraction techniques and protocols in pcr methodology that can impact the accuracy of the pcr results as well. this paper, therefore, aims to synthesize factors that contribute to false positive and negatives in pcr results, improvements to protocols that includes the use of dna intercalating dyes such as propidium monoazide (pma) in pcr assay to improve dna extraction and discuss the importance of an optimized pcr assay for diagnostic testing efficacy from a social and economic standpoint. review article rt-qpcr protocol optimization assessment with propidium monoazide for viable and non-viable pathogen detection 1mariam sajid ,1mohamed askar, 1roger bayston 1department of academic orthopaedics, queen's medical centre, university of nottingham, c floor, west block, derby road, nottingham, ng7 2uh, uk. introduction the covid-19 pandemic provided a lot of lessons for epidemiologists and the scientific community at large on how to manage a pandemic. one of the keys determining factors in controlling the spread and impact of the disease is in diagnostic testing1,2. the quicker one is able to accurately diagnose a person with the infection, can determine how quickly the spread of the infection can be contained in a particular population through treatment and disease management protocols3. during the pandemic many lateral flow devices such as rapid antigen test kits4 and antibody serology tests were optimized for population wide screening. these types of tests offer shorter run time of around 10-15 minutes and are useful in differentiating those with active infections compared to those who have recovered5. however, their low sensitivity and specificity leads the confirmatory diagnostics such as a real time pcr (rt-qpcr) to remain the gold standard. polymerase chain reaction (pcr) is a molecular assay that aims to detect specific genetic material from pathogens such as bacteria or virus through dna detection. using the process of amplification, pcr replicates the small sample or fragments found of the test sample (virus or bacteria) and amplifies to a level that it becomes detectable as a signal. the overall testing accuracy is determined by how sensitive and specific the assay is. sensitivity refers to the ability of the test to accurately detect infected individual while specificity is whether the uninfected test samples come up negative. for a laboratory performance, a pcr test is usually 95% sensitive and specific. however, in reality this performance does drop down because of external factors that include how the assay is handled, potential risk of contamination and other varying conditions in pcr protocols, all of which can determine the likelihood of false positive and false negative results appearing6. false negative and false positive results: false negative results, if we take the example of covid-19 test samples, would indicate that a person who has viral infection is showing a negative result on the pcr instead of a true positive7. with an assumed 95% sensitivity, there is a 5% chance of false negative results appearing. this could be attributed to not just errors in handling of testing, cross contamination but also proceedings s.z.m.c. vol: 37(2): pp. 50-57, 2023. pszmc-886-37-2-2023 rt-qpcr protocol optimization assessment with propidium monoazide for viable and non-viable….. 51 low viral load that isnot enough for the pcr to amplify and detect. one study has suggested that up to 58% covid-19 patients may have false negative results on their initial rt-pcr7because of pcr testing protocols that include how early a person in their infection period is tested. on the other hand, a false positive8,9 indicates that a person does not have an active viral infection while the pcr result shows up as positive rather than a true negative. false positives are less common compared to false negatives and yet both types of results can potentially cause harm in high and low prevalence population setting, of a disease state in question. prevalence indicates how common a disease, for example covid-19, is in a specific population at a specific point in time. and this prevalence (or disease burden) plays a role on the positive predictive value (ppv) and negative predictive value (npv) of a diagnostic test. a ppv is a probability that the testing samples showing up positive are truly positive, while an npv is the probability that samples testing negative are truly negative. in a high prevalence area, the ppv will increase but the npv will decrease indicating that in a high prevalent risk population, false negatives will become more of an issue. while in a low prevalence setting with a decreased ppv and an increased npv, false positives will be a concern10,11. impact of pcr testing errors: the adverse effects linked with false positive and false negative results, in disease infections, cannot be underestimated. in usa, centers for disease control and prevention (cdc) states that roughly 1 million healthcare associated infections and 90,000 deaths occur each year. among the hospital acquired infections staphylococcus aureus and methicillin resistant s. aureus (mrsa) are the prevailing pathogens with approximately 5 10% prevalence in hospitals compared to 2-6% in general population. adding to this, a study states that mrsa related false negative results can go from 6 to 30%, and false positive results around 20% through rt-pcr testing12. looking at the existing burden of hospital infections on hospital resources and cost, the additional impact that diagnostic testing error is causing has to be given importance. with the rt-pcr already an established standard in testing, the optimization of its testing protocols to reduce its error rate is the next step. outside of contamination, poor techniques, low quality sampling and reagents and general handling of the test13(table-1), looking into pcr methodology can also aim to reduce errors14. one of the causes for false positive or negatives tests can be attributed to pcr machine being unable to differentiate between live and dead pathogen dna. given that dna from dead bacteria can remain in clinical samples for months15 even after a person has been cured of infection, therefore can tamper with the pcr results. table-1: pcr testing error factors the main principle of pcr is to detect dna in the test sample and begin to amplify it through various rounds of thermal cycling16, generating enough dna sample that the fluorescent dye in the pcr is able to bring up a signal determining whether the pathogen dna is present (positive) or not (negative). quantification of the dna is also done by measuring how many cycles it takes for the pcr to amplify the dna at the threshold level. lower the number of cycles, higher the number of genetic material present in initial sample which thereby indicates the severity of the infection.17 however, if dna from dead bacteria or virus is still present in the sample from previous infections, that dna can also be amplified as the pcr is unable to discriminate between live and dead dna thus overestimating the presence of active infection in a sample i.e., false positive18. propidium monoazide dye: give then importance of viable and non-viable nucleic acid distinction in samples whether in a clinical setting or environmental, standardized protocols have still not been fully developed. several viability assessments have been considered to distinguish live vs dead bacterial samples such as cultivation where live samples are detected using population growth methods such as gram positive/ gram negative bacterial cultures, flow cytometry and more. this process is limited rt-qpcr protocol optimization assessment with propidium monoazide for viable and non-viable….. 52 by the cultivatable species as well as time constraints. another assessment can be done through a metabolism-based approach such as molecular viability testing. the principal behind this technique looks at metabolic activities carried out by viable (live) samples such as changes in cellular energy (atp production), respiratory measurements and more. limitation of this approach is linked with risks of false positives due to presence of active extracellular enzymes that can show up in non-viable samples19. the third assessment is using a membrane-based method that looks at the integrity of membranes in live vs dead samples using different stains, dyes and treatments. for this approach, the viability pcr20 uses ethidium monoazide (ema) or propidium monoazide (pma) as a membrane binding dye. while both ema and pma follow the same process flow, some studies have shown that ema is not as effective in distinguishing samples by penetrating both dead and live nucleic acid samples21,22. propidium monoazide (pma) is a dna and rna binding photo reactive dye that is membrane impermeant, which means that the dye is not able to penetrate a cell membrane allowing them to only bind with dead cells. once they enter the dead cell, through the broken membrane, they bind to the dna or rna covalently once made reactive by light. this binding mechanism modifies the nucleic acid thus preventing it from being amplified during the amplification process in a pcr. the remaining pma that is unbound and free-floating becomes inactivated upon light exposure as well therefore eliminating the risk of it attaching to dna during the dna extraction phase for example (fig-1). fig-1: pma flow chart for pma to work in optimal conditions there are several factors that need to be considered such as the pma concentration being used, time periods for its dark incubation and the time period for photo activation process during which it binds with dead cells. all these parameters need to be tested to find the conditions necessary to give accurate results. askar et al. (2019), in their study looked at these parameters when assessing affects of pma on pcr performance for detection of periprosthetic joint infections23 (staphylococcus aureus) using tissue samples. looking at cycle threshold difference(δct), between viable and nonviable bacterial suspensions, δct viable and δct dead were calculated. δct viable being the difference in ct value of viable sample with and without pma dye and same for ∆ct dead. fig-2: optimization of pma: amplification plot of live (viable) and dead (non-viable) samples with and without pma dye.askar etal.(2019). the ct difference in live(viable) samples with and without pma showed to be less than one cycle difference. while the ct difference in the dead(non-viable) samples with and without pma were significantly different, showcasing that the inclusion of pma dye on non-viable samples increased the cycle numbers before reaching cycle threshold. in another paper (pany.et al), the quantification of viable listeria monocytogenes in food products was tested using pma conditions. in that paper both ema and pma conditions were tested where ema was found to be toxic to viable cells compared to pma regardless of the varying conditions provided, further validating the shift to pma from ema dyes in dna extraction24. further, the paper lists out pma concentrations and incubation temperatures as key conditions that impact the ct values of pma-pcr assay. similarly (vesper, 2008) found that optimizing the pma conditions during dna extraction for their fungal infection samples contributed to viable sample distinction and also suggested altering the pma conditions for different species to get best outcomes25. this was discovered when the optimized conditions for pma treatment which led rt-qpcr protocol optimization assessment with propidium monoazide for viable and non-viable….. 53 to effective distinction of viable samples in cyanobacterial species did not provide the same distinction when same conditions were applied in diatom species analyses26. pma rt-qpcr results in laboratory and clinical settings: in a laboratory setting with bacterial suspensions, pma dye shows good discrimination in live and dead bacteria. however, the question arises on whether similar results can be seen in clinical setting where samples are more complex in nature through presence of other material that can hinder pma efficacy. when looking at tissue suspension samples (ex vivo) the ct difference of viable cells in tissue suspension compared with viable cells in bacterial suspension showed a delay in amplification. this could indicate pcr inhibition by other tissue materials in the sample. when ct difference in viable tissue suspension samples was measured with pma dye for amplification, the delay in amplification was seen to be less in pma samples compared to the previous example, indicating that pma could also be playing a role in reacting with various organic materials in the tissue sample that inhibit pcr amplification. while there are other factors that also need optimization such as improved techniques for dna extraction that can reduce presence of human dna and other homogenates that inhibit the pcr activity; the presence of pma as a pre treatment pcr technique does work to improve pcr sensitivity as well as specificity. as indicated in the study pcr efficacy increased from 71.7 to 89.1% for viable pathogen detection. the value of the pma technique is dependent on its versatility in different microbial diagnostic testing. outside of staphylococcus aureus, it is important to consider how pma interacts with other bacteria and viruses. mycobacterium tuberculosis 27 or pseudomonas aeruginosa that come under the category of viable but not culturable bacteria (vbnc), present issues with public health providers as culturing is not only time-consuming (taking an average of 5-7 days to generate results) but such vbnc pathogens are difficult to culture altogether due to their low metabolic activity. in environmental sampling, detection of such bacteria in open bodies of water such as swimming pools is time sensitive, which is where a rt-qpcr with a pma dye will not only speed up the time taken for detection of sample but can also differentiate whether sample contains viable or non-viable bacteria. one study28 showcased that using pma in water samples for detection of bacteria helped to reduce false positives in rt-pcr29 testing due to the presence of damaged bacteria in the samples while not impacting the viable dna from being amplified, or being affected by environmental materials that could otherwise hinder efficacy. pma rt-qpcr testing protocol modifications: one study looked at the effect of pma rt-qpcr for uti infection detection in urine samples. in most cases, with a prolonged infection, clinical symptoms of a uti infection can persist even if bacterial cultures turn up negative. this can be attributed to long antibiotic courses that are able to still give false negatives in cultures. while the pma dye inclusion in rt-qpcr can bring the distinction between live and dead bacteria and improving accuracy over a conventional pcr, the efficacy of rt-qpcr when reacting with urine components is impacted as pma efficiency in binding with live vs dead cells is reduced. this again highlights the need to augment the pma rtqpcr protocols depending on the external conditions of the samples provided. in this study (zeng, d. et al.,2016). pma efficiency increased once the bacterial samples were centrifuged, sediments collected and suspended in phosphate buffered saline (pbs). similar to modification of pma rt-qpcr by suspending or removing organic material in target sample, there are other modifications to pma for different pathogens that have yielded improved results. such as dithiothreitol cotreatment in bacillus subtilis spores that helped pma penetration in inactivated spores compared to viable spores31. other strategies include the already discussed parameters such as incubation time for light and dark periods, concentration but also temperature for incubation that can help to increase dye penetration in non-viable cells. as pma can work with any organism that has its nucleic acid enclosed in a pma impermeable membrane or structure, it should also function on viral organisms. an effective method to detect infectious viral particle is not only important from a diagnostic clinical perspective but also from assessing disinfection treatments. to that effect, viral culturing or virus induced plaques are the gold standard methods in determining viral infectivity. in virus culturing, a virus is injected in laboratory cell lines to observe whether cell damage and death occurs or not and if the virus rt-qpcr protocol optimization assessment with propidium monoazide for viable and non-viable….. 54 reproduces. while this is effective, culturing is time consuming, has a long turn-around time for results, needs biosafety level protocols and is expensive. a pcr approach is still preferable but the distinction of the virus being active vs inactive remains crucial as every pcr result cannot be sent for a viral culture confirmation afterwards. the sars-cov-2 is an enveloped protein virus that upon damage to its membrane can allow pma to penetrate and bind. one study showed that the addition of a surfactant sodium dodecyl sulfate (sds) helped the pma dye penetration32 in damaged cells as it is a membrane destabilizing agent. the study found that that sds-pma assisted q-rtpcr led to a quicker distinction between the viable and non-viable samples. this is because pma is able to react to the free rna released or inside damaged virions but is unable to penetrate active (live) virus and bind with its rna, when used with sds (0.0005%). the rna that does bind to pma therefore inhibits the pcr amplification, allowing for only live virions to be detected33. the addition of sds is an important augmentation as simply heat-treating the virus (in moderate conditions) does not damage its viral envelope. if a higher temperature (95°c) is used it would damage the envelope enough for pma to bind but such conditions, as are not present in the environment, are not optimal to include as part of protocols. pma rt-qpcr in viral testing: pma pre-treatment for viral infections is still not completely validated since pma is limited by its requirement of a damaged membrane for penetration. not all inactivated viruses have a damaged capsid34, for example, there are some viruses that can lose receptors that allow them to remain infectious while still having their capsid intact. in this scenario, the virus is inactive but pma is unable to penetrate and block it from amplification in rt-qpcr thus retaining the risk of a false positive35.thus highlighting a critical note to membrane-based method for viable sample detection as the integrity of a membrane cannot fully determine the viability or active nature of a cell. one study36 looked at t4 bacteriophage to determine the conditions suitable for effective pma pre-treatment to distinguish between viable and non-viable (inactive) viruses. it determined that moderate temperatures and protease treatments might not be effective as it did not damage the capsid that protected the viral nucleic acid. therefore, other additions to the pma parameters are needed for further optimization in viral testing. for water treatments or general assessment of disinfectant efficacy, using extreme conditions of heat, chlorination can help to inactivate enteric viruses such as poliovirusand hepatitis a37and damage their capsid. in that case, using a pma rt-qpcr would be effective in determining whether such conditions worked in inactivating viruses from a particular environment38. however, further research into which type of conditional changes (for example susceptibility of hepatitis a, e and other non culturable viruses to temperate, surfactants etc39,40) are optimal for pma dyes to penetrate and perform, is critical for the success of this pcr approach. while false negatives are often reported and investigated, false positive results though less common still need to be understood. high number of cases of false positives could be attributed to the pcr sensitivity in picking up dead viral dna from previous infections. from a covid-19 contagion standpoint, a false positive can have a psychological impact on a person by triggering panic and anxiety. moreover, as part of contact tracing and isolation protocols that have been put in place, such a false positive report can create more harm whereby the person could be quarantined with other covid-19 positive patients thus increasing likelihood of a getting an actual infection. in such scenarios, the dna assays need to be able to assess pathogenic risk instead of the conventional pcr approach where a signal is detected from infectious or non-infectious viral dna. with continuous research and development, the pcr methodology continues to be optimized and expanded allowing for changes in the protocols to accommodate for pcr efficacy. conclusion using intercalating dna binding dye has been on an evolutionary journey for many years, where the previous dye ethidium monoazide (ema) has now been replaced with propidium monoazide (pma) because pma is a more impermeant membrane than ema. however, there are still limitations41 to the pma pre-treatment that needs to be further reviewed. the additions of surfactants or alterations in temperature, concentrations and other parameters can be customized to pathogen specific testing protocols. however, one limitation that does not have a clear-cut solution is the dyes inability to detect inactivation of pathogen that does not damage the rt-qpcr protocol optimization assessment with propidium monoazide for viable and non-viable….. 55 membrane. in that scenario, the only alternative approach that can be considered is a molecular viability test (mvt)42,43. mvt uses a pre ribosomal rna as a detection method to differentiate between viable and non viable bacteria. in viable bacterial cells, pre rrnas are intermediates in ribosomal rna synthesis that are found abundantly in growing bacterial cells. given a nutritional stimulus these pre-rrna intermediates (pools) rapidly replenishes which can be observed as fluctuations that can be detected. this can help improve the sensitivity of pcr detection for bacterial viability discrimination since mvt does not require the membrane to be damaged for detection. however, the limitation with this approach is its being relatively new with little data and research behind it. it can therefore be considered as an alternative for scenarios where pma with its alterations to protocol are not providing efficient rt-qpcr results, with culturing techniques as a secondary approach. overall, pma is an elegant addition to pathogen detection through rt-pcr44,45. with population expansion and climate change, the spread of various 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(2022) “review of covid-19 testing and diagnostic methods,” talanta, 244, p. 123409.available at:https://doi.org/10.1016/j.talanta.2022.123409. 2. vengesai, a. et al. (2021) “a systematic and meta analysis review on the diagnostic accuracy of antibodies in the serological diagnosis of covid 19,” systematic revie mytton, o.t. et al. (2021) interpreting a lateral flow sars-cov-2 antigen test, the bmj. british medical journal publishing group. availableat:https://www.bmj.com/content/373/bmj. n1411 (accessed: december 10, 2022). ws, 10(1). available at: https://doi.org/10.1186/s13643-021 01689-3. 3. paltiel, a.d., zheng, a. and sax, p.e. (2021) “clinical and economic impact of widespread rapid testing to decrease sars-cov-2 transmission.” availableat:https://doi.org/10.1101/2021.02.06.212 51270. 4. mytton, o.t. et al. (2021) interpreting a lateral flow sars-cov-2 antigen test, the bmj. british medical journal publishing group. availableat:https://www.bmj.com/content/373/bmj. n1411 (accessed: december 10, 2022). 5. kwok, h.f. (2022) “the significance of advanced covid-19 diagnostic testing in pandemic control measures,” international journal of biological sciences,18(12),pp.46104617.availableat:https://d oi.org/10.7150/ijbs.72837. 6. braunstein, g.d. et al. (2021) false positive results with sars-cov-2 rt-pcr tests and how to evaluate a rt-pcr-positive test for the possibility of a false positive result, journal of occupational and environmental medicine. u.s.nationallibraryofmedicine.availableat:https:/ /www.ncbi.nlm.nih.gov/pmc/articles/pmc7934325 /(accessed: december 10, 2022). 7. kanji, j.n. et al. (2021) “false negative rate of covid-19 pcr testing: a discordant testing analysis,” virology journal, 18(1). available at: https://doi.org/10.1186/s12985-021-01489-0. 8. kong, x.-q. et al. (2022) false-positive results of sars-cov-2 rt-pcr in oropharyngeal swabs from vaccinators, frontiers. frontiers. available at: https://www.frontiersin.org/articles/10.3389/fmed. 2022.847407/full (accessed: december 10, 2022). 9. karel, t., mazouch, p. and fischer, j. (1970) the economic impact of false positivity of covid-19 pcr testing in the czech republic, international journal of economic sciences. available at: https://www.eurrec.org/ijoes-article-117019 (accessed: december 10, 2022). 10. pecoraro, v. et al. (2021) “estimate false-negative rt-pcr rates for sars-cov-2. a systematic review and meta-analysis,” european journal of clinical investigation, 52(2). available at: https://doi.org/10.1111/eci.13706. 11. alsheikh, s.h., ibrahim, k. and alfaraj, d. (2021) the impact of false positive covid-19 result, cureus. u.s. national library of medicine.availableat:https://www.ncbi.nlm.nih.go v/pmc/articles/pmc8752407/ (accessed: december 10, 2022). 12. the impact of incorrect mrsa diagnoses medical laboratory observer (no date). available at: https://www.mlo online.com/home/article/13004269/the-impact-of incorrect-mrsa-diagnoses (accessed: december 10, 2022). 13. bergeret, m. and profile, f. (no date) crispr: the future of molecular diagnostics?,today's clinical lab.availableat:https://www.clinicallab.com/trend s/molecular-diagnostics/crispr-the-future-of molecular-diagnostics-25264 (accessed: december 10, 2022). 14. kralik, p. and ricchi, m. (2017) a basic guide to real time pcr in microbial diagnostics: definitions, parameters, and everything, frontiers. frontiers.availableat:https://www.frontiersin.org/a rticles/10.3389/fmicb.2017.00108/full (accessed: december 10, 2022). http://www.bmj.com/content/373/bmj http://www.bmj.com/content/373/bmj http://www.bmj.com/content/373/bmj http://www.bmj.com/content/373/bmj http://www.ncbi.nlm.nih.gov/pmc/articles/pmc7934325 http://www.ncbi.nlm.nih.gov/pmc/articles/pmc7934325 http://www.frontiersin.org/articles/10.3389/fmed http://www.frontiersin.org/articles/10.3389/fmed http://www.frontiersin.org/articles/10.3389/fmed http://www.eurrec.org/ijoes-article-117019 http://www.eurrec.org/ijoes-article-117019 http://www.ncbi.nlm.nih.go/ http://www.ncbi.nlm.nih.go/ http://www.clinicallab.com/trend http://www.frontiersin.org/a rt-qpcr protocol optimization assessment with propidium monoazide for viable and non-viable….. 56 15. guo, j. et al. (2022) “a new pma-qpcr method for rapid and accurate detection of viable bacteria and spores of marine-derived bacillus velezensis b 9987,” journal of microbiological methods, 199, p. 106537.availableat:https://doi.org/10.1016/j.mimet .2022.106537. 16. understanding covid-19 pcr testing (no date) genome.gov.availableat:https://www.genome.gov/ about-genomics/fact-sheets/understandingcovid 19-pcr-testing (accessed: december 10, 2022). 17. how is the covid-19 virus detected using real time rt-pcr? (2020) iaea. iaea. (accessed: december 10, 2022). 18. askar, m. et al. (2019) propidium monoazide– polymerase chain reaction for detection of residual periprosthetic joint infection in two-stage revision molecular biology reports, springerlink. springer netherlands.availableat:https://link.springer.com/a rticle/10.1007/s11033-019-05092-z (accessed: december 10, 2022). 19. bullington, j. (no date) digging into dna: evaluating propidium monoazide as a tool to remove nonviable microbial dna from deep-sea sedimentcores,mbari.org.availableat:https://www. mbari.org/wp content/uploads/2019/11/bullington.pdf (accessed: march 28, 2023). 20. viability pcr (2022) biotium. available at: https://biotium.com/technology/pma-for-viability pcr/ (accessed: december 10, 2022). 21. nocker, a., cheung, c.-y. and camper, a.k. (2006) “comparison of propidium monoazide with ethidium monoazide for differentiation of live vs. dead bacteria by selective removal of dna from dead cells,” journal of microbiological methods, 67(2),pp.310320.availableat:https://doi.org/10.101 6/j.mimet.2006.04.015. 22. flekna, g. et al. (2007) “insufficient differentiation of live and dead campylobacter jejuni and listeria monocytogenes cells by ethidium monoazide (ema) compromises ema/real-time pcr,” research in microbiology, 158(5), pp. 405–412. availableat:https://doi.org/10.1016/j.resmic.2007.0 2.008. 23. yamamoto, y. (2002) pcr in diagnosis of infection: detection of bacteria in cerebrospinal fluids, clinical and diagnostic laboratory immunology. u.s. national library of medicine. availableat:https://www.ncbi.nlm.nih.gov/pmc/arti cles/pmc119969/ (accessed: december 10, 2022). 24. pan, y. and breidt, f. (2007) “enumeration of viable listeria monocytogenes cells by real-time pcr with propidium monoazide and ethidium monoazide in the presence of dead cells,” applied and environmental microbiology, 73(24), pp. 80288031.availableat:https://doi.org/10.1128/aem. 01198-07. 25. vesper, s. et al. (2008) “quantifying fungal viability in air and water samples using quantitative pcr after treatment with propidium monoazide (pma),”journalofmicrobiologicalmethods,72(2),p p.180184.availableat:https://doi.org/10.1016/j.mim et.2007.11.017. 26. joo, s., park, p. and park, s. (2019) applicability of propidium monoazide (pma) for discrimination between living and dead phytoplankton cells, plos one. u.s. national library of medicine. available at:https://www.ncbi.nlm.nih.gov/pmc/articles/pmc 6592542/ (accessed: march 28, 2023). 27. zeng, d. et al. (2016) advances and challenges in viability detection of foodborne pathogens, frontiers in microbiology. u.s. national library of medicine.availableat:https://www.ncbi.nlm.nih.go v/pmc/articles/pmc5118415 (accessed: december 10, 2022). 28. golpayegania;douraghim;rezaeif;alimohammad im;nodehi rn; (no date) propidium monoazide quantitative polymerase chain reaction (pma qpcr) assay for rapid detection of viable and viable but non-culturable (vbnc) pseudomonas aeruginosa in swimming pools, journal of environmental health science & engineering. u.s. national library of medicine. available at: https://pubmed.ncbi.nlm.nih.gov/31297217/ (accessed: december 10, 2022). 29. kibbee, r.j. and örmeci, b. (2017) “development of a sensitive and false-positive free pma-qpcr viability assay to quantify vbnc escherichia coli and evaluate disinfection performance in wastewater effluent,” journal of microbiological methods,132,pp.139147.available:https://doi.org/1 0.1016/j.mimet.2016.12.004. 30. lee, a.s. et al. (2022) a novel propidium monoazide-based pcr assay can measure viable uropathogenic e. coli in vitro and in vivo, frontiers.frontiers.availableat:https://www.frontie rsin.org/articles/10.3389/fcimb.2022.794323/full (accessed: december 10, 2022). 31. rawsthorne h, dock cn, jaykus la. 2009. pcr based method using propidium monoazide to distinguish viable from nonviable bacillus subtilis spores. appl. environ. microbiol. 75:2936–2939. 10.1128/aem.02524-08 32. hong, w. et al. (2021) “rapid determination of infectious sars-cov-2 in pcr-positive samples by sds-pma assisted rt-qpcr,” science of the total environment, 797, p. 149085. available at: https://doi.org/10.1016/j.scitotenv.2021.149085. 33. chung, y.-s. et al. (2021) validation of real-time rt-pcr for detection of sars-cov-2 in the early stages of the covid-19 outbreak in the republic of korea, nature news. nature publishing group. availableat:https://www.nature.com/articles/s4159 8-021-94196-3 (accessed: december 10, 2022). 34. leifels, m. et al. (2021) “capsid integrity quantitative pcr to determine virus infectivity in environmental and food applications – a systematic review,” water research x, 11, p. 100080.availableat:https://doi.org/10.1016/j.wroa. 2020.100080. http://www.genome.gov/ http://www/ http://www.ncbi.nlm.nih.gov/pmc/arti http://www.ncbi.nlm.nih.gov/pmc/arti http://www.ncbi.nlm.nih.gov/pmc/arti http://www.ncbi.nlm.nih.gov/pmc/articles/pmc http://www.ncbi.nlm.nih.gov/pmc/articles/pmc http://www.ncbi.nlm.nih.go/ http://www.ncbi.nlm.nih.go/ http://www.nature.com/articles/s4159 rt-qpcr protocol optimization assessment with propidium monoazide for viable and non-viable….. 57 35. braunstein, g.d. et al. (2021) “false positive results with sars-cov-2 rt-pcr tests and how to evaluate a rt-pcr-positive test for the possibility of a false positive result,” journal of occupational & environmental medicine, 63(3).availableat:https://doi.org/10.1097/jom.0000 000000002138. 36. gs;, p.s.l.i.f. (no date) use of propidium monoazide in reverse transcriptase pcr to distinguish between infectious and noninfectious enteric viruses in water samples, applied and environmental microbiology. u.s. national library ofmedicineavailableat:https://pubmed.ncbi.nlm.ni h.gov/20472736/ (accessed: december 10, 2022). 37. joo, s., park, p. and park, s. (2019) applicability of propidium monoazide (pma) for discrimination between living and dead phytoplankton cells, plos one. u.s. national library of medicine. available at:https://www.ncbi.nlm.nih.gov/pmc/articles/pmc 6592542/ (accessed: december 10, 2022). 38. molecular viability testing (mvt) (no date) environmental & occupational health sciences. availableat:https://deohs.washington.edu/cangelosi lab/molecular-viability-testing-mvt(accessed: december 10, 2022). 39. zi, c. et al. (2018) an improved assay for rapid detection of viable staphylococcus aureus cells by incorporating surfactant and pma treatments in qpcr bmc microbiology, biomed central. biomedcentral.availableat:https://bmcmicrobiol.b iomedcentral.com/articles/10.1186/s12866-018 1273-x (accessed: december 10, 2022). 40. soejima t, minami j, iwatsuki k. rapid propidium monoazide pcr assay for the exclusive detection of viable enterobacteriaceae cells in pasteurized milk. journal of dairy science. 2012;95(7):3634– 42. 10.3168/jds.2012-5360 41. limitations of using propidium monoazide with qpcr to discriminate ... (no date). available at: https://journals.sagepub.com/doi/10.4137/mbi.s17 723 (accessed: december 10, 2022). 42. molecular viability testing (mvt) (no date) environmental & occupational health sciences.availableat:https://deohs.washington.edu/ cangelosilab/molecular-viability-testing mvt(accesseddecember 10, 2022). 43. cangelosi, g.a. and meschke, j.s. (2014) dead or alive: molecular assessment of microbial viability, applied and environmental microbiology. u.s. national library of medicine. available at: https://www.ncbi.nlm.nih.gov/pmc/articles/pmc41 78667/#b48 (accessed: december 10, 2022). 44. rogers, g.b. et al. (2010) “the exclusion of dead bacterial cells is essential for accurate molecular analysis of clinical samples,” clinical microbiology and infection, 16(11), pp. 1656– 1658. available at: https://doi.org/10.1111/j.1469 0691.2010.03189.x. 45. zhou, l. and pollard, a.j. (2012) a novel method of selective removal of human dna improves pcr sensitivity for detection of salmonella typhi in blood samples bmc infectious diseases, biomed central.biomedcentral.availableat:https://bmcinfe ctdis.biomedcentral.com/articles/10.1186/1471 2334-12-164 (accessed: december 10, 2022). the authors: miss. mariam sajid, msc. molecular genetics and diagnostics, department of academic orthopaedics, institute of queen's medical centre, university of nottingham,uk. dr. mohamed askar, academic and clinical fellow, department of academic orthopaedics, institute of queen's medical centre, university of nottingham,uk. email: mohamed.askar@nhs.net dr. roger bayston, professor emeritus surgical infection (school of medicine), department of academic orthopaedics, institute of queen's medical centre, university of nottingham,uk. email: roger.bayston@nottingham.ac.uk corresponding author: miss. mariam sajid, msc. molecular genetics and diagnostics, department of academic orthopaedics, institute of queen's medical centre, university of nottingham, uk. email: mariamsajid25@gmail.com http://www.ncbi.nlm.nih.gov/pmc/articles/pmc http://www.ncbi.nlm.nih.gov/pmc/articles/pmc http://www.ncbi.nlm.nih.gov/pmc/articles/pmc41 http://www.ncbi.nlm.nih.gov/pmc/articles/pmc41 http://www.ncbi.nlm.nih.gov/pmc/articles/pmc41 file.indd 14 proceedings s.z.m.c. vol: 35(2): pp. 14-17, 2021. pszmc-786-35-2-2021 1department of cardiology, al-aleem medical college, lahore 2department of anatomy, al-aleem medical college, lahore 3department of physiology, al-aleem medical college, lahore 4advance centre of research, al-aleem medical college, lahore 15 gender differences in the morphological severity of mitral stenosis in patients presenting in a tertiary care hospital p 16 gender differences in the morphological severity of mitral stenosis in patients presenting in a tertiary care hospital 17 gender differences in the morphological severity of mitral stenosis in patients presenting in a tertiary care hospital file.indd 47 proceedings s.z.m.c. vol: 35(2): pp. 47-51, 2021. pszmc-792-35-2-2021 department of urology, sims/services hospital, lahore 48 is upper pole puncture for percutaneous nephrolithotomy (pcnl) safe and effective? 49 is upper pole puncture for percutaneous nephrolithotomy (pcnl) safe and effective? 50 is upper pole puncture for percutaneous nephrolithotomy (pcnl) safe and effective? 51 is upper pole puncture for percutaneous nephrolithotomy (pcnl) safe and effective? full book for printing 19 proceedings s.z.m.c. vol: 37(2): pp. 19-22, 2023. pszmc-881-37-2-2023 1department of , allama iqbal medical college, lahore. 2department of anatomy, bakhtawar amin medical and dental college, multan. 3department of anatomy, king edward medical university, lahore. 70 1department of orthopedics, thq, khanewal 2department of orthopedics, lgh, lahore 3department of orthopedics, thq, ferozwala 4department of orthopedics, cmh, lahore 5department of hand & upper limb surgery, cmh, lahore 6department of orthopedics, al-rehmat benevolent trust 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 20 indigenous cadaveric variations in lung fissures and lobes pattern 21 indigenous cadaveric variations in lung fissures and lobes pattern 22 indigenous cadaveric variations in lung fissures and lobes pattern full book for printing 9 proceedings s.z.m.c. vol: 37(2): pp. 09-14, 2023. pszmc-879-37-2-2023 1department of community medicine, rlku medical college lahore. 2department of biochemistry, rlku medical college lahore. 3department of physiology, rlku medical college lahore. 70 1department of orthopedics, thq, khanewal 2department of orthopedics, lgh, lahore 3department of orthopedics, thq, ferozwala 4department of orthopedics, cmh, lahore 5department of hand & upper limb surgery, cmh, lahore 6department of orthopedics, al-rehmat benevolent trust 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 10 diet in pregnancy & preeclampsia: is there an association? a hospital based matched case control study 11 diet in pregnancy & preeclampsia: is there an association? a hospital based matched case control study 12 diet in pregnancy & preeclampsia: is there an association? a hospital based matched case control study 13 diet in pregnancy & preeclampsia: is there an association? a hospital based matched case control study 14 diet in pregnancy & preeclampsia: is there an association? a hospital based matched case control study full book for web 36 proceedings s.z.m.c. vol: 37(1): pp. 36-40, 2023. pszmc-874-37-1-2023 1department of anatomy, govt. khawaja muhammad safdar medical college, sialkot. 2 department of anatomy, faisalabad medical university, faisalabad. 3department of anatomy, akhtar saeed medical & dental college, lahore. 4 department of anatomy, bakhtawar amin medical and dental college, multan. 5department of anatomy, fatima jinnah medical university, lahore. 6department of anatomy, amna inayat medical college, lahore. 70 1department of orthopedics, thq, khanewal 2department of orthopedics, lgh, lahore 3department of orthopedics, thq, ferozwala 4department of orthopedics, cmh, lahore 5department of hand & upper limb surgery, cmh, lahore 6department of orthopedics, al-rehmat benevolent trust 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 37 embryo-hepatotoxic potential of spearmint aqueous extracts: a histopathological study 38 embryo-hepatotoxic potential of spearmint aqueous extracts: a histopathological study 39 embryo-hepatotoxic potential of spearmint aqueous extracts: a histopathological study 40 embryo-hepatotoxic potential of spearmint aqueous extracts: a histopathological study full book for printing 29 proceedings s.z.m.c. vol: 37(2): pp. 29-34, 2023. pszmc-883-37-2-2023 1department of biochemistry, gujranwala medical college, gujranwala. 2department of pharmacology and therapeutics, fatima jinnah medical university, lahore. 3department of ibb, university of lahore, lahore. 4department of biochemistry, fmh medical and dental college lahore, lahore. 70 1department of orthopedics, thq, khanewal 2department of orthopedics, lgh, lahore 3department of orthopedics, thq, ferozwala 4department of orthopedics, cmh, lahore 5department of hand & upper limb surgery, cmh, lahore 6department of orthopedics, al-rehmat benevolent trust 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 30 heavy metal ion and haptoglobin association with career duration in pharmaceutical industries: a….. 31 heavy metal ion and haptoglobin association with career duration in pharmaceutical industries: a….. 32 heavy metal ion and haptoglobin association with career duration in pharmaceutical industries: a….. . exs. . 33 heavy metal ion and haptoglobin association with career duration in pharmaceutical industries: a….. . environ. health 34 heavy metal ion and haptoglobin association with career duration in pharmaceutical industries: a….. full book for web 23 proceedings s.z.m.c. vol: 37(1): pp. 23-30, 2023. pszmc-872-37-1-2023 15th year mbbs, cmh lmc & iod, lahore. 2department of community medicine, cmh, lmc & iod, lahore. 70 1department of orthopedics, thq, khanewal 2department of orthopedics, lgh, lahore 3department of orthopedics, thq, ferozwala 4department of orthopedics, cmh, lahore 5department of hand & upper limb surgery, cmh, lahore 6department of orthopedics, al-rehmat benevolent trust 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 24 medical students’ perception regarding community medicine as a subject and a career option amongst….. 25 medical students’ perception regarding community medicine as a subject and a career option amongst….. 26 medical students’ perception regarding community medicine as a subject and a career option amongst….. 27 medical students’ perception regarding community medicine as a subject and a career option amongst….. p 28 medical students’ perception regarding community medicine as a subject and a career option amongst….. 29 medical students’ perception regarding community medicine as a subject and a career option amongst….. 30 medical students’ perception regarding community medicine as a subject and a career option amongst….. full book for printing 58 proceedings s.z.m.c. vol: 37(2): pp. 58-61, 2023. pszmc-887-37-2-2023 case report 1department of orthopedic surgery, lahore general hospital, lahore, pakistan. 70 1department of orthopedics, thq, khanewal 2department of orthopedics, lgh, lahore 3department of orthopedics, thq, ferozwala 4department of orthopedics, cmh, lahore 5department of hand & upper limb surgery, cmh, lahore 6department of orthopedics, al-rehmat benevolent trust 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 59 case report: a case of glomus tumor; missed diagnosis as enchondroma for previous 13 years 60 case report: a case of glomus tumor; missed diagnosis as enchondroma for previous 13 years 61 case report: a case of glomus tumor; missed diagnosis as enchondroma for previous 13 years file.indd 7 proceedings s.z.m.c. vol: 35(2): pp. 7-13, 2021. pszmc-785-35-2-2021 1department of community medicine, central park medical college, lahore 2department of cardiology, allama iqbal memorial teaching hospital sialkot 2department of physiology, central park medical college, lahore 3department of biochemistry, university of hafr al-batin, saudia arabia 8 knowledge, attitude, and practice trends towards covid-19 pandemic: comparison between individuals 9 knowledge, attitude, and practice trends towards covid-19 pandemic: comparison between individuals 10 knowledge, attitude, and practice trends towards covid-19 pandemic: comparison between individuals 11 knowledge, attitude, and practice trends towards covid-19 pandemic: comparison between individuals 12 knowledge, attitude, and practice trends towards covid-19 pandemic: comparison between individuals 13 knowledge, attitude, and practice trends towards covid-19 pandemic: comparison between individuals full book for web dedicated to prof.dr. sibgha zulfiqar the first lady chairperson & dean of sheikh zayed medical complex lahore bismillah hir rehman nir raheem. in the name of allah, the most beneficent and the most merciful message by the chairperson shaikh zayed medical complex & chief editor of proceedings prof. sibgha zulfiqar prof. saadia s. alam (chairperson& dean) (chief editor) we bid farewell to prof. sibgha zulfiqar our patron in chief proceedings and honorable chairperson & dean as she attains super annuation after 3 decades of service at szmc lahore. her impeccable and whole hearted commitment to the entire complex and especially to proceedings facilitated us to achieve international recognition in the directory of open access journals (doaj), list of cpsp recognized journals, over 3 years of hec y category indexation, all the while maintaining our ojs supported proceedings website of international standards which connects us to the world. we will truly miss her and wish her the best of health and allah’s blessings in her future endeavours. the current covid 19 pandemic wave has re-struck china as it suddenly relaxed it zero covid policy allowing unrestricted intermingling of the populace causing indeterminable number of covid infections and tens of thousands of fatalities reported by who. global warming , hunger and war have impacted global health significantly.in these rapidly changing scenarios, we steadfastly publish a diverse january to march 2023 proceedings articles on varied aspects of medical, surgical, and allied health disciplines. all articles submitted for publication have been judged on stringent international standards of plagiarism and blind peer reviews. our editions have achieved expanded viewership, with researchers in wide ranging medical fields from different institutions publishing their research in proceedings. our nexus of undergraduate and postgraduate researchers, supervisors, academicians, and reviewers remains imperative to achieving rigorous standards of research, writing and evaluation in proceedings. our desire is to build linkages with top journals and researchers both in the country and abroad in sha allah. our motto “we venture forth” recognised by pmc, hec, cpsp & doaj proceedings shaikh zayed medical complex vol. 37 (1) pcpb/24(94) p-121/3196 jan to mar 2023 editorial board patron in chief: prof. sibgha zulfiqar chairperson & dean chief editor: prof. saadia shahzad alamphd editor: prof. ayesha humayunphd associate editors: prof. m. imran anwar frcs dr. adnan salimmrcp external associate editor: dr. usman iqbalphd members-national: prof. mamoon rashid-frcs (sitara-e-eisaar) prof. nadiramamoonmrcpath prof. m. ashraf phd (tamgha-e-imtiaz) prof. m. arif nadeem (fcps) prof.khalidmahmood(phd) dr. sarah ghafoorphd members-international prof.rennekoeffelphd(sui) prof.jane banaszak-holl phd(aus) prof. hisham al muhtasebphd (ksa) dr. salma malik m.d (us) dr. sarwat shaheenm.d(us) dr. aliya asher mrcp(uk) dr. uzma nasim siddiqui fcps(aus) section editors: dr. shahila jalil fcps dr. samira haque fcps dr. lubna riaz fcps dr. sadia maqsood mhpe dr. faraz bokhari m phil dr. noora hassan hezam al aqmer m phil mrs.saimamohsinmsc consultant biostatician: mr. muhammad aasimm phil library & information sciences: mr. ihsan basit m phil dr. muhammad shahid soroyaphd technical support: m.saad ullah sheikh (bs.engr) contents muhammad tayyab zia, ahmed zia ud din, abdullah saeed, sadia abdullah, muhammad junaid tahir technical considerations and complication of percutaneous transhepatic biliary drainage: a single center retrospective study nimrah komal, ali mujtaba, muhammad wajid munir, aftab ahmed, syed abdul ahad, hassan mumtaz, syed abdul rehman shah is covid vaccine hesitancy unfounded? a local comparative study on short-term side effects of different covid vaccinations wardah nisar, rabbiya sarwar, shahid nabi, zainab bilal malik, shamaila zamir attitude, awareness and recommendations regarding fortified food usage amongst health care professionals of punjab atif mansoor ahmed, irum abbas, sohail ahmed siddiqui hba1c levels an indicator of glycemic control in diabetics with and without retinopathy hamna azhar, izzah baig, fareha azam, jannat ehsan, danyal faisal, zainab omer medical students’ perception regarding community medicine as a subject and a career option amongst medical colleges of lahore city maham qazi, danyal anwar shiraz, ch. muhammad aqeel, pir muneeb ur rehman, umer farooq, mohammad aslam early postoperative pain in video assisted anal fistula treatment versus open fistulectomy asma zulfiqar, uzma ali, asma siddique, saqib mansoor, gul afshan, muhammad suhail embryo-hepatotoxic potential of spearmint aqueous extracts: a histopathological study talha mahmud, muhammad saqib, yasser saeed khan, adeel wyne, faisal rahman, muhammad muntazir mehdi, rabbia abbas, shazia rafique, muhammad idrees a 5 year tertiary centre experience of frequency & risk factors association with primary and secondary lung cancers ch. muhammad aqeel, humaira alam, pir muneeb rehman, aqsa jmail, maham qazi, mohammad aslam combination of botulinum toxin and topical diltiazem versus lateral sphincterotomy in treatment of chronic anal fissure shuja uddin, ejaz ahmad, husnain khalid, hafiz m. umar, ahmed umair asad khan, mian m. haneef case report: neglected locked anterior shoulder dislocation with upper brachial plexus injury instructions for authors_____________________________________ page 01 06 11 18 23 31 36 41 47 52 56 advisoryboard basic & pre-clinical sciences prof. mateen izharphd, mrcpath prof. sibgha zulfiqarm.phil prof. muhammad suhailm.phil prof. nasreen ehsanm.phil prof. tahira naseemm.phil dr. anwaar bashir m.phil dr. suleman dawood phd (uol) dr. nighat yasmeen phd (kemu) dr. saima chaudhary phd (uhs) medicine and allied prof.azeemtajfcps prof. abdul shakoorfcps prof. talha mahmudfcps prof. safooraaamirfcps dr. lubna riaz fcps dr. asfand tariq fcps surgery and allied prof. ali rafiquemirzafcps prof. muhammad ikram fcps prof. imran anwarfrcs prof. shafqat mukhtarfcps dr. jamshaid rahimfcps dr. syed sajjad raza kazmifcps diagnostic division prof. mona azizfcps prof. saulatsarfrazfcps dr. amir khanfrcr peer review board (national) prof. m. nauman ahmadffarcs prof. naseem saudphd prof. m. ovais omerphd prof. shabbir bhattiphd prof. zahidniazfrcs prof. zamir ahmadphd prof. muhammad arif nadeem fcps prof. muhammad moinfrcs prof. khalid mahmood phd prof. sadia chiraghm.phil prof. muhammad aslamfcps prof. abdul mannan m.s prof. naheedhumayunfcps prof. farhat naz fcps prof. abdul hameed frcs prof. rafeea tafweez phd prof. inayat thaverphd peer review board (national) prof. ashraf chaudhryfcps prof. kashifmalikfcps prof. aftabturabiphd prof. uzma hussainfcps prof. muhammad pervaizfrcs prof. fouzia shaukat fcps prof. maryam rashidphd prof. rabeia bilal phd prof. khwaja khursheedfcps prof. tanvir akhtar butt ms prof. naila akhtar fcps prof. abidashaheenphd prof. sajida malik m.phil prof. rumina hasan phd prof.khalid javeed khan frcs dr. farooq afzal frcs dr. saman shahidphd dr. muhammad shahzadphd dr. ali hussainy zaidi m.d dr. shahzad khuram akramphd dr. fahad azam phd dr. khadija irfan fcps dr. uzma malikfcps dr. ayesha mallick frcp dr. tehseenhaiderkazmifcps dr. khalida ajmal m.phil dr. israr ahmedfcps dr. saima batoolfcps dr. saleem muhammad ranaphd dr. muhammad khurram habibfcps dr. saba riaz phd dr. raaziatasadduqphd dr. nageen hussain phd dr. saadia nosheen jan fcps dr. soumble zulfiqar phd dr. yasir abbas zaidi fcps dr. muhammad zeeshan sarwarmrcs dr. muhammad imran khokarmrcs dr. nabiha farasatm.phil dr. tayyaba muzaffar m.phil dr. zaeem khalid fcps dr. nafeesah fatimah fcps peer review board (international) prof. khalid javeed khan frcs prof. renne koeffel phd (switzerland) prof. jane banaszak-holl phd (aus) prof. m. hisham al-muhtaseb phd (jordan) prof. mukhtiar baig phd (ksa) dr. sarwat shaheenm.d(usa) dr. zeeshan i. shaikh m.d (usa) dr. tanzeel qurat ijazmrcp (uk) dr. hafiz sohail anjum fcps (ireland) dr. rafayazharfrcp (singapore) dr. mazhar nawaz m.d (usa) dr. ahmad azam malik phd (ksa) dr. ishtiaq ahmed fcps (ksa) dr. aamiromairphd (ksa) dr. sabina ahmed mir m.d (usa) dr. anita lamichhanem.d (nepal) dr. faiza durraniphd (uk) dr. waseem lodhifrcog (uk) dr. seerat zahra hammadm.s(ksa) dr. shafya shahid phd (u.s) dr. abdul waheedfrcs (u.k) dr. humaira zareenfcps (ksa) dr. amira okud m.d (ksa) dr. asma ahmed mrcp (uk) dr. sabahat sabir mrcog (uk) dr. sarfaraz ahmad frcr (uk) dr. zouina sarfaraz m.d (uk) dr. humaira zaman fcps (uk) dedicated to our most respected prof. dr. zia ullah 20-3-1960 to 30-11-2020 former head of department of medicine, shaikh zayed medical complex, lahore a shaheed with a legacy of service to the nation bismillah hir rahman nir raheem. in the name of allah, the most beneficent and the most merciful message by the chairman shaikh zayed medical complex & chief editor of proceedings prof. mateen izhar prof. saadia s. alam (chairman & dean) (chief editor) we present our jan to march 2021 edition with prayers for the good fortune and wellbeing of all. as the covid 19 pandemic rages, it is a moment to capitulate the personal, emotional and physical toll endured by humanity in general and especially frontline workers during the previous year. the sudden loss of our very own prof. zia ullah to this deadly disease has left us an indelible memory. for proceedings 2020 was a year of achievements alhumdolillah, with development of a new website of international standards supported by the ojs/pkp platform for online submission www.proceedings-szmc.org.pk. during the year we got recognition and funding from higher education commission of pakistan (hec). we thank them for reposing their confidence in us awarding “y” category, an honor indeed. we are now indexed by pmc, google scholar, crossref/doi pastic and pakmedinet. efforts are underway for international indexation. the jan to march 2021 proceedings presents research on covid 19, health care, basic sciences, therapeutics and diagnostics. all articles submitted for publication have been judged on stringent international standards of plagiarism and blind peer reviews. as our editions reflect we have achieved expanded viewership, with researchers in wide ranging medical fields from different institutions publishing their research in proceedings. our vision is to create an r&d department at szmc in collaboration with the national health research complex (nhrc) and build linkages with top journals and researchers both in the country and abroad “we venture forth” http://www.proceedings-szmc.org.pk/ hec & pmc recognized proceedings shaikh zayed medical complex vol. 35 (1) pcpb/24(94) p-121/3196 jan to mar 2021 editorial board patron in chief: prof. mateen izhar chairman & dean chief editor: prof. saadia shahzad alam head, pharmacology & therapeutics external associate editor: dr. usman iqbal taipei medical university, taiwan associate editors: prof. ayesha humayun head, public health & community medicine prof. imran anwar head, surgery unit-ii dr. adnan salim assistant prof. gastroenterology sub editors: dr. uzma nasim siddiqui assistant prof. medicine dr. shahila jalil associate prof. histopathology prof. tahira naseem head, biochemistry & chemical pathology dr. samira haque associate prof. histopathology dr. lubna riaz assistant prof. pediatrics dr. sadia maqsood assistant prof. pharmacology dr. faraz bokhari associate prof. physiology dr.noora hassan hezam al-aqmer physiology department mrs. saima mohsin incharge, nhrc consultant biostatician: dr. akram chaudhry mr. muhammad aasim library & information sciences: mr. ihsan basit dr. muhammad shahid soroya contents munaza javed, muhammad atif qureshi, muhammad naeem akhtar, saadat ullah, osama habib, muhammad saqib correlation of qsofa with severity of covid-19 infection assessed on admission at tertiary care hospital in lahore khansa saleem, muhammad ilyas, saad razzaq, fahad maqbool, qaiser abbas, aneem abdullah, abu bakar shahid improvement in survival outcomes of acute lymphoblastic leukemia patients treated with berlin-frankfurt-munster 76/79 protocol muhammad saqib, talha mahmud, shanawer qaiser, muhammad ramzan, muhammad asim bronchoscopic microbial diagnosis in non-responding & non-resolving pneumonia naheed humayun sheikh, ayesha humayun, shakila zaman contraceptive and abortion-related practice among females of lower income group in district lahore shanawer qaiser, muhammad saqib, talha mahmud frequency of raised total serum ig e, sputum eosinophilia and blood eosinophil count in patients with severe persistent asthma maleeha tahir butt, mukarma sajjad, tahira naseem relation of vitamin d deficiency with breast cancer staging in patients from inmol cancer hospital saba saleem, muhammad suhail, saba amjad, faiza irshad protective effects of ocimum basilicum against atrophic changes in graafian follicles in cyclophosphamide induced ovarian toxicity nadeem yaqoob, mufakhara fatimah, farwa naqvi, javeria sarfraz, sidra mushtaq, sadia chiragh comparative evaluation of aloe vera and diclofenac on body weight, blood pressure and renal function of hypertensive rats zoofishan qureshi, muhammad ashraf chaudhry, seema imdad, tariq mukhtar farani, ayesha humayun association of maternal hyperhomocysteinemia with preeclampsia: a hospital based case-control study instructions for authors page 1 7 12 16 23 27 33 39 45 51 editorial advisory board basic & pre-clinical sciences prof. mateen izhar head, pathology division, szmc prof. sibgha zulfiqar head, physiology, szmc. prof. muhammad suhail head, anatomy, szmc. prof. nasreen ehsan head, forensic medicine, szmc dr. suleman dawood (uol) assistant prof. of allied health sciences dr. nighat yasmeen (kemu) assistant prof. of biomedical sciences dr. tania shakoori (kemu) assistant prof. of biomedical sciences dr. saima chaudhary (uhs) assistant prof. of oral pathology medicine and allied prof azeem taj head, medicine, szmc. prof. abdul shakoor head, psychiatry, szmc prof. talha mahmud head, pulmonology, szmc prof. safoora aamir head, dermatology, szmc surgery and allied prof. mamoon rashid (sitara-e-eisaar) head, plastic surgery, shifa hospital, islamabad prof. ali rafique mirza professor of plastic surgery, szmc prof. muhammad ikram head, gynecology, szmc prof. shafqat mukhtar head, gynecology, szmc dr. jamshaid rahim associate professor urology, szmc diagnostic division prof. nadira mamoon head, pathology, shifa hospital, islamabad prof. mona aziz head, hematology, szmc prof. saulat sarfraz head, radiology, szmc dr. amir khan assistant prof. radiology, szmc peer review board (international) prof. renne koeffel (switzerland) prof. mohammad hisham al-muhtaseb (jordan) prof. mukhtiar baig (ksa) dr. jane banaszak hall (usa) dr. dagmar stoiber sakaguchi (austria) dr. sarwat shaheen (usa) dr. zeeshan i. shaikh (usa) dr. aliya asher (uk) dr. tanzeel qurat ijaz (uk) dr. hafiz sohail anjum (ireland) dr. rafay azhar (singapore) dr. mazhar nawaz (usa) dr. ahmad azam malik (ksa) dr. ishtiaq ahmed (ksa) dr. aamir omair (ksa) dr. sabina ahmed mir (usa) dr. anita lamichhane (nepal) dr. faiza durrani (uk) dr. waseem lodhi (uk) dr. seerat zahra hammad (ksa) dr. shafya shahid (u.s) dr. madeeha afzal (u.k) dr. abdul waheed (u.k) dr. humaira zareen (ksa) dr. amira okud (ksa) peer review board (national) prof. m. ashraf (tamgha-e-imtiaz) prof. arshad kamal butt prof. m. nauman ahmad prof. ziaullah prof. moeed iqbal qureshi prof. naseem saud prof. m. ovais omer prof. shabbir bhatti prof. zahid niaz prof. zamir ahmad prof. muhammad arif nadeem prof. muhammad moin prof. khalid mahmood prof. zujaja zaheer prof. muhammad aslam prof abdul mannan prof. naheed humayun prof. farhat naz prof. abdul hameed prof. rafeea tafweez prof. inayat thaver prof. ashraf chaudhry prof. kashif malik prof. aftab turabi dr. maryam rashid dr. farooq afzal dr. saman shahid dr. sarah ghafoor dr. muhammad shahzad dr. ali hussainy zaidi dr. shahzad khuram akram dr. uzma malik dr. ayesha mallick dr. tehseen haider kazmi dr. khalida ajmal dr. israr ahmed dr. saima batool dr. saleem muhammad rana dr. muhammad khurram habib dr. paras khan dr. saba riaz dr. raazia tasadduq dr. nageen hussain dr. saadia jan dr. soumble zulfiqar dr. rabeia bilal full book for printing dedicated to the memory of our very dear professor azeem taj head of medicine szmc lahore 18th july 1973 to 14th march 2023 inna lillahe wa inna elaihai rajeoon bismillah hir rehman nir raheem. in the name of allah, the most beneficent and the most merciful chief editor of proceedings prof. saadia s. alam (chief editor) the april to june 2023 edition begins with a stark reminder of the mortality of human life. the sudden, shocking, and untimely passing of our very own professor azeem taj head of medicine shaikh zayed medical complex lahore has left us all introspecting on the truth of being mortal and finite. prof azeem was a good soul, a great academician and splendid physician. may allah bless his journey to jannah. on the proceedings front we have applied for continued hec y category indexation. our ojs supported proceedings website of international standards connects us to the world. continuing covid 19 infections despite robust vaccinations globally and in pakistan tell us of its undeniable persistence and presence on our infectious disease panel. dengue and monkey pox have once again reared their heads. in these rapidly changing scenarios, we steadfastly publish a diverse april to june 2023 proceedings articles on medical, surgical and diagnostic disciplines. all articles submitted for publication have been judged on stringent international standards of plagiarism and blind peer reviews. our editions have achieved expanded viewership, with researchers in wide ranging medical fields from different institutions publishing their research in proceedings. our nexus of undergraduate and postgraduate researchers, supervisors, academicians, and reviewers remains imperative to achieving rigorous standards of research, writing and evaluation in proceedings. our desire is to build linkages with top journals and researchers both in the country and abroad in sha allah. our motto “we venture forth” recognised by pmc, hec, cpsp& doaj proceedings shaikh zayed medical complex vol. 37 (2) pcpb/24(94) p-121/3196 apr to jun 2023 editorial board chief editor: prof. saadia shahzad alamphd editor: prof. ayesha humayunphd associate editors: prof. m. imran anwar frcs dr. adnan salim mrcp external associate editor: dr. usman iqbalphd members-national: prof. mamoon rashid-frcs (sitara-e-eisaar) prof. nadiramamoonmrcpath prof. m. ashraf phd (tamgha-e-imtiaz) prof. m. arif nadeem(fcps) prof. khalid mahmood(phd) dr. sarahghafoorphd members-international prof. renne koeffel phd(sui) prof.jane banaszak-holl phd(aus) prof. hisham al muhtasebphd (ksa) dr. salma malik m.d (us) dr. sarwat shaheenm.d(us) dr. aliya asher mrcp(uk) dr. uzma nasim siddiqui fcps(aus) section editors: dr. shahila jalilfcps dr. samira haquefcps dr. lubna riaz fcps dr. sadia maqsood mhpe dr. faraz bokhari m phil dr. noora hassan hezamal aqmer m phil mrs.saima mohsinmsc consultant biostatician: mr. muhammad aasimm phil library & information sciences: mr. ihsan basit m phil dr. muhammad shahid soroyaphd technical support: m.saad ullah sheikh (bs.engr) contents hijab batool, muhammad dilawar khan, omar rasheed chughtai, sana rafaqat, fouzia sadiq, hafiz muhammad bilal characterization of mannose binding lectin (mbl) levels in type-2 diabetes mellitus patients amongst pakistani,population zoofishan imran, umer saeed ansari, shamim akram, shehla javaid, hifza noor lodhi diet in pregnancy and preeclampsia: is there an association? a hospital based matched case control study sibgha fatima, saba saleem, noor ijaz, nabila kaukab, fauzia qureshi, asma siddique local sexual dimorphism based on chilotic index of human dried hip bone: a morphometric study shazia tufail, maria ilyas, saqib mansoor, syeda shabeeh rubab, ayesha afzal indigenous cadaveric variations in lung fissures and lobes tayyaba azhar, junaid sarfraz, maimoona nasreen qualities of pbl leader: the students’ perspective uzma jabbar, muniza qayyum, rukhshan khurshid, maira mahmood heavy metal ion and haptoglobin association with career duration in pharmaceutical industries: a potential occupational hazard? faryal farooq, nasreen naz, maryam asghar, amjad sattar, nauman al qamari, wiqar ahmad, parvez ahmed, hassan mumtaz efficacy of pigtails for the management of refractory malignant ascites or effusion: a systematic retrospective chart review of an institution abdul rasheed qureshi, muhammad irfan, huma bilal, muhammad sajid, zeeshan ashraf ultrasound guided complete thoracentesis, precluding residual pleural thickening in pleural tuberculosis: a case-control study mariam sajid, mohamed askar, roger bayston review article rt-qpcr protocol optimization assessment with propidium monoazide for viable and non-viable pathogen detection shuja uddin, zia uddin, mubashir farhan, adeel hamid, rizwan ul haq, hasnain khalid, yousaf gull, mian muhammad haneef case report a case of glomus tumor; missed diagnosis as enchondroma for previous 13 years instructions for authors ________________________________________________________________________________________________ page 01 09 15 19 23 29 35 42 50 58 62 advisoryboard basic & pre-clinical sciences prof. nasreen ehsanm.phil prof. tahira naseemm.phil dr. anwaar bashir m.phil dr. suleman dawood phd(uol) dr. nighat yasmeen phd(kemu) dr. saima chaudhary phd(uhs) medicine and allied prof. talha mahmudfcps prof. safoora aamirfcps dr. lubna riazfcps dr. asfand tariqfcps dr. nabeel ibad fcps surgery and allied prof. ali rafique mirzafcps prof. muhammad ikram fcps prof. imran anwarfrcs dr. jamshaid rahimfcps dr. syed sajjad raza kazmifcps diagnostic division prof. mona azizfcps prof. saulat sarfrazfcps dr. amir khanfrcr peer review board (national) prof. m. nauman ahmadffarcs prof. naseem saudphd prof. m. ovais omerphd prof. shabbir bhattiphd prof. zahid niazfrcs prof. zamir ahmadphd prof. muhammad arif nadeem fcps prof. muhammad moinfrcs prof. khalid mahmood phd prof. sadia chiraghm.phil prof. muhammad aslamfcps prof. abdul mannan m.s prof. naheed humayunfcps prof. farhat naz fcps prof. abdul hameed frcs prof. rafeea tafweezphd prof. inayat thaverphd peer review board (national) prof. mateen izharphd, mrcpath prof. sibgha zulfiqarm.phil prof. abdul shakoorfcps prof. ashraf chaudhryfcps prof. kashif malikfcps prof. aftabturabiphd prof. khalid javeed khan frcs prof. uzma hussainfcps prof. muhammad pervaizfrcs prof. fouzia shaukat fcps prof. maryam rashidphd prof. rabeia bilal phd prof. khwaja khursheedfcps prof. tanvir akhtar butt ms prof. naila akhtar fcps prof. abida shaheenphd prof. sajida malik m.phil prof. shafqat mukhtarfcps prof. rumina hasan phd prof. khalid javeed khan frcs dr. farooq afzal frcs dr. samanshahidphd dr. muhammad shahzadphd dr. ali hussainy zaidim.d dr. shahzad khuram akramphd dr. fahad azam phd dr. khadija irfan fcps dr. uzma malikfcps dr. ayesha mallick frcp dr. tehseen haider kazmifcps dr. khalida ajmalm.phil dr. israr ahmedfcps dr. saima batool fcps dr. saleem muhammad ranaphd dr. muhammad khurram habibfcps dr. saba riaz phd dr. raaziatasadduqphd dr. nageen hussain phd dr. saadia nosheen jan fcps dr. soumble zulfiqar phd dr. yasir abbas zaidi fcps dr. muhammad zeeshan sarwarmrcs dr. muhammad imran khokarmrcs dr. nabiha farasat m.phil dr. tayyaba muzaffar m.phil dr. zaeem khalid fcps dr. nafeesah fatimah fcps dr. athar mahmood fcps dr. fariha sahrish fcps peer review board (international) prof. renne koeffel phd(switzerland) prof. jane banaszak-holl phd(aus) prof. m. hisham al-muhtaseb phd (jordan) prof. mukhtiar baig phd(ksa) dr. sarwat shaheenm.d(usa) dr. zeeshan i. shaikh m.d (usa) dr. tanzeel qurat ijaz mrcp (uk) dr. hafiz sohail anjum fcps(ireland) dr. rafay azharfrcp (singapore) dr. mazhar nawaz m.d (usa) dr. ahmad azam malik phd (ksa) dr. ishtiaq ahmed fcps (ksa) dr. aamir omairphd (ksa) dr. sabina ahmed mir m.d (usa) dr. anita lamichhanem.d (nepal) dr. faiza durraniphd (uk) dr. waseem lodhifrcog (uk) dr. seerat zahra hammadm.s(ksa) dr. shafya shahid phd(u.s) dr. abdul waheedfrcs (u.k) dr. humaira zareenfcps (ksa) dr. amira okudm.d (ksa) dr. asma ahmed mrcp (uk) dr. sabahat sabir mrcog (uk) dr. sarfaraz ahmad frcr (uk) dr. zouina sarfaraz m.d (uk) dr. humaira zaman fcps (uk) untitled-1 12 proceedings s.z.m.c. vol: 35(1): pp. 12-15, 2021. pszmc-777-35-1-2021 bronchoscopic microbial diagnosis in non-responding & non-resolving pneumonia 1muhammad saqib, 1talha mahmud, 1shanawer qaiser, 2muhammad ramzan, 3muhammad asim 1department of pulmonology, shaikh zayed medical complex, lahore 2department of pulmonology, gulab devi chest hospital, lahore 3nhrc, shaikh zayed medical complex, lahore abstract introduction: pneumonia is a common and potentially serious illness associated with considerable morbidity and mortality, especially in the elderly and those with notable comorbidities. more than 100 microbes (bacteria, viruses, fungi, and parasites) can cause pneumonia. in hospitalized community-acquired pneumonia (cap), 40% are non-responding cap. the reliability of using bronchoscopic procedures when compared with sputum cultures, routinely processed bronchoscopic specimens demonstrate improved sensitivity. aims & objectives: isolation of a responsible microbe causing respiratory infection in non-responding & nonresolving pneumonia. place and duration of study: this study was conducted at the department of pulmonology shaikh zayed medical complex, lahore from 01-01-2018 to 31-12-2019. material & methods: retrospective analysis of 109 bronchoscopic procedures conducted in patients with nonresponding & non-resolving pneumonia with a fiberoptic bronchoscope over two years (01-01-2018 to 3112-2019). results: one hundred nine (109) procedures were done in patients with non-responding and resolving pneumonia. bronchial washing microbiology yielded results in 71 (65.1%) patients. pseudomonas aeruginosa was the most frequent organism and was isolated in 30 (42.3%) patients; klebsiella pneumoniae was found in 23 (32.4%) patients. other organisms included staphylococcus aureus (22.5%), acinetobacter baumanni (14.1%), escherichia coli (11.3%), streptococcus pneumoniae (11.3%), stenotrophomonas maltophilia (4.2%), haemophilus influenzae (2.8%), enterococcus (1.4%) and mycobacterium tuberculosis (4.2%). conclusion: pseudomonas aeruginosa and klebsiella pneumoniae are the most common isolated organisms in patients with non-responding and non-resolving pneumonia. bronchoscopy is an extremely useful tool for sample collection among non-responding & non-resolving pneumonia and making the correct microbiological diagnosis. key words: non-responding and resolving pneumonia, bronchoscopy, retrospective. introduction pneumonia is a universal and potentially critical disease, and it is associated with substantial morbidity and mortality, particularly in the elderly and those with significant comorbidities.1 the nonresolving or slowly resolving pneumonia is a common clinical term. it has been used reciprocally to mention the persistence of radiographic abnormalities past the expected time limit2. more than 100 microbes (bacteria, viruses, fungi, and parasites) can cause pneumonia.3 in hospitalized cap, 40% are non-responding cap. non-resolving or slowly resolving pneumonia accounts for 10% 15% of nosocomial pneumonias.4,5 delay in diagnosis and treatment may cause fatality by 3-5% in both community-acquired and nosocomial pneumonia. incorrect diagnosis, impaired host defense, atypical organisms, inadequate antibiotic therapy, resistant pathogens, tuberculosis, etc. are the common etiologies of non-resolving or slowly resolving pneumonia.6 slow or incomplete pneumonia resolution, despite treatment, requires a more rigorous evaluation. microbiological, histopathological, and cytological tests of the specimens can be done for diagnosing the underlying cause. the sputum culture has a low diagnostic yield,7 but the reliability of using bronchoscopic procedures compared with sputum cultures demonstrate improved sensitivity. the sensitivity of bronchoscopic specimens is around 40% in the context of non-responding cap and 70% in nosocomial icu pneumonia.6 in this study, we intend to establish the characteristics of microorganisms isolated from 13 bronchoscopic microbial diagnosis in non-responding & non-resolving pneumonia bronchial washing specimens of patients with nonresolving pneumonia or non-responding pneumonia. material and methods the study was a descriptive case series, conducted at the department of pulmonology shaikh zayed fgmi, lahore from 01-01-2018 to 31-12-2019. all cases of non-responding and resolving pneumonia underwent bronchoscopy during the period were included. the selected cases were 109. statistical analysis: the data were recorded in spss 20.0. data were described by using frequencies and percentages and presented in tables, pi-charts, and bar charts. results the total number of patients registered in the study was 109, having ages between 15 and 85 years; there were 71 (65.1%) males & 38 (34.9%) females. ninety-five (87.2%) patients were admitted to the pulmonology ward while 7 (6.4%) patients were admitted to the intensive care unit, and 7 (6.4%) patients were referred from other hospitals (table-1). bronchoscopy yielded result in 65.1% patients, and the most common pathogen isolated was pseudomonas aeruginosa (42.3%) followed by klebsiella pneumoniae (32.4%), staphylococcus aureus (22.5%), acinetobacter baumanni (14.1%), escherichia coli (11.3%), streptococcus pneumoniae (11.3%) and haemophilus influenzae (2.8%). (figure 1) in 46 (64.8%) patients, only one organism was isolated, while in 20 (28.2%) cases, two organisms were detected in 5 (7%) patients, three organisms were found (table-1). in 64.8% of patients, single organism was isolated in bal culture, while 28.2% of patients had two organisms, and 7% revealed three organisms. diabetes was the most common comorbidity in this study, observed among 39.4% of all the cases; other comorbidities include hypertension (22.5%), chronic obstructive pulmonary disease (copd) (11.4%), asthma (5.6%), end-stage renal disease (esrd) (5.6%), while nine patients had a history of organ transplant (fig-2). thirty-one (28.4%) patients had a history of symptoms for one week, 29 (26.6%) patients for two weeks, 12 (11.4%) for three weeks, 23 (21.1%) patients had the disease for four weeks & 14 patients (12.8%) had a presentation with symptoms more than 4 weeks (table-1). n % gender male 71 65.1 female 38 34.8 age ˂ 40 years 14 12.8 40 to 60 years 57 52.2 ˃ 60 years 38 34.9 culture result positive 71 65.1 negative 38 34.8 admission place ward 95 87.2 icu 7 6.4 referred from other hospital 7 6.4 duration of symptoms 1 week 31 28.4 2 weeks 29 26.6 3 weeks 12 11.0 4 weeks 23 21.1 ˃4 weeks 14 12.8 table-1: various characteristics of study cases fig-1: organisms isolated fig-2: co-morbidities pseudomonas aeruginosa, 30 klebsiella pneumoniae, 23 staphylococcus aureus, 16 acinetobacter baumanni, 10 escherichia coli, 8 streptococcus pneumoniae, 8 stenotrophomonas maltophilia, 3 haemophilus influenzae, 2 enterococcus, 1 0 5 10 15 20 25 30 14 bronchoscopic microbial diagnosis in non-responding & non-resolving pneumonia n % number of organisms single 46 64.8 two 20 28.2 three 5 7.0 table-2: number of organisms isolated on bronchoalveolar lavage (bal) culture discussion non-resolving or non-responding pneumonia is not a rare clinical entity to pulmonologists, and simultaneously, it can be a cause of concern in everyday clinical practice. amberson first described the term "unresolved organizing or protracted pneumonia" in 1943. 8 most patients with cap show clinical improvement within 72 hours of initial antibiotic treatment. it has been determined that 6 to 15 percent of hospitalized patients with cap do not respond within this time, and the failure rate for those admitted to icu may be as high as 40 percent. causes of non-responding & non-resolving pneumonia may include atypical organisms, inadequate antibiotic therapy, resistant pathogens, tuberculosis, etc.6 the radiographic and clinical improvement in pneumonia varies with age, comorbidities & with the particular infectious agent causing pneumonia. in general, the resolution is more rapid with nonbacteremic streptococcus pneumoniae, mycoplasma pneumoniae, chlamydophila species, and moraxella catarrhalis than with other organisms.9 in this study, 85% of patients were above the age of 40, and 34.9% were over the age of 60. in a prospective cohort study, it was found that age alone has the most remarkable impact on the resolution of pneumonia.10 broncho-alveolar lavage (bal) fluid was collected from all patients who underwent bronchoscopy during this study, and it remained diagnostic in 65.1% cases, whereas during a study conducted in india, bal culture-confirmed diagnostic yield was 71.4%.6 in a study conducted by silver et al., bronchoscopy was diagnostic in 86% of cases.11 balamugesh et al. have also observed bronchoscopy a very useful tool in evaluating non-resolving pneumonia.12 a study conducted in barcelona, spain revealed that 13% of patients with pneumonia had mixed infection.13 in our research, the mixed infection was seen in 35.2% of cases. we also observed that all five patients where three organisms were isolated on bal culture were admitted to icu and had multiple comorbidities. gram-negative bacteria were seen to be the predominant pathogens in our study, and among them, pseudomonas (42.3 %), klebsiella (32.4%), and acinetobacter (14.1%) were the common pathogens. chaudhuri et al. & fein also shared similar observations of an increased occurrence of gram-negative etiology of pneumonia in patients with non-resolving pneumonia or non-responding pneumonia.6,14 diabetes mellitus was the most frequent comorbidity in our study and was seen in 39.4% of cases and while the most common organisms in the diabetic patients were klebsiella (39.3%) & pseudomonas (35.7%). avijgan has also reported that diabetes mellitus was significantly associated with the delayed resolution of pneumonia.15 klebsiella pneumoniae and mycobacterium tuberculosis were the most common etiologies in diabetics and were detected in 33.3% and 25% cases, respectively. begamy has also published an increased appearance of klebsiella pneumoniae in thoracic infections in diabetic patients.16 in our study, we had nine patients with post organ transplant history, and among these, the most common organisms were also klebsiella (44.4%) & pseudomonas (44.4%) reflecting their ability to cause non-responding & non-resolving pneumonia. conclusion non-responding & non-resolving pneumonia is usually an area of clinical dilemma. the microbiological profile varies from that of commonly known organism for communityacquired respiratory tract infection as most common pathogens isolated in our study were pseudomonas aeruginosa & klebsiella pneumoniae. bronchoscopy is an extremely useful tool for sample collection among non-responding & non-resolving pneumonia and making the correct microbiological diagnosis, this can help in timely modification of antimicrobial therapy. references 1. mandell la, wunderink rg, anzueto a, et al. infectious diseases society of america/ american thoracic society consensus guidelines on the management of community-acquired pneumonia in adults. clin infect dis 2007; 44 suppl 2:s27. 2. weyers cm, leeper kv. non resolving pneumonia. clin chest med. 2005; 26:143-58. 15 bronchoscopic microbial diagnosis in non-responding & non-resolving pneumonia 3. johansson n, kalin m, tiveljung-lindell a, et al. etiology of community-acquired pneumonia: increased microbiological yield with new diagnostic methods. clin infect dis 2010; 50:202. 4. gotway mb, leung jw, dawn sk, hill a. nonresolving pneumonia in an otherwise healthy patient. clin pulm med 2004; 11:198-200. 5. menendez r, perpina m, torres a. evaluation of non-resolving and progressive pneumonia. semin respir infect. 2003; 18:103-11. 6. chaudhuri ad, mukherjee s, nandi s, bhuniya s, tapadar sr, saha m. a study on nonresolving pneumonia with special reference to role of fiberoptic bronchoscopy. lung india 2013; 30:27-32. 7. sputum cultures: low yield in communityacquired pneumonia bukata, w. richard md author information emergency medicine news: december 2002 volume 24 issue 12 p31-34. 8. amberson jb. significance of unresolved organizing or protracted pneumonia. j mich state med soc 1943;42: 599-603. 9. fein am, feinsilver sh, niederman ms, et al. "when the pneumonia doesn't get better". clin chest med 1987; 8:529. 10. el solh aa, aquilina at, gunen h, ramadan f. radiographic resolution of community-acquired bacterial pneumonia in the elderly. j am geriatr soc 2004; 52:224-9. 11. feinsilver sh, fein am, niederman ms, schult de, faegenburg dh. utility of fiberoptic bronchoscopy in non-resolving pneumonia. chest 1990; 98:1322-6. 12. balamugesh t, aggarwal an, gupta d, behera d, jindal sk. profile of repeat fiberoptic bronchoscopy. indian j chest dis allied sci 2005; 47:181-5. 13. de roux a, ewig s, garcía e, et al. mixed community-acquired pneumonia in hospitalized patients. eur respir j 2006; 27:795. 14. fein am. pneumonia in the elderly: overview of diagnostic and therapeutic approaches. clin infect dis 1999; 28:726-9. 15. avijgan m. specificity and sensitivity of clinical diagnosis for chronic pneumonia. east mediterr health j 2005; 11:1029-37. 16. begamy t. thoracic empyema. is its microbiology changing? pul rev com. 2000; 5:10-2. the authors: dr. muhammad saqib assistant professor, department of pulmonology, shaikh zayed medical complex, lahore. prof. talha mahmud head, department of pulmonology, shaikh zayed medical complex, lahore. dr. shanawer qaiser registrar, department of pulmonology, shaikh zayed medical complex, lahore. dr. muhammad ramzan senior registrar, department of pulmonology, gulab devi chest hospital, lahore. muhammad asim statistical officer, nhrc, shaikh zayed medical complex, lahore. corresponding author: dr. muhammad saqib assistant professor, department of pulmonology, shaikh zayed medical complex, lahore. e-mail: dr.saqib.ch@gmail.com full book dedicated to the valiant health care warriors, humanitarians, social workers medical instructors, future doctors & researchers of pakistan bismillah hir rehman nir raheem. in the name of allah, the most beneficent and the most merciful message by the chairperson shaikh zayed medical complex & chief editor of proceedings prof. sibgha zulfiqar prof. saadia s. alam (chairperson & dean) (chief editor) our oct to dec 2022 proceedings heralds our induction into the doaj a premier global repository of open access journals. along with 3 years of hec y category indexation and renewed pmdc recognition it is a culmination of continuous efforts. our ojs supported proceedings website of international standards connects us to the world. catastrophic floods impacting one third of pakistan in september have led to multiple public health crises with outbreaks of malaria, dengue , gastroenteritis & cholera in the rain swamped areas. public, governmental, armed forces and global humanitarian responses are ongoing. meanwhile covid 19 cases have declined. our current edition publishes articles on pediatrics and maternal health, obstetrics, medical, dental, orthopedics and basic health research. all articles submitted for publication have been judged on stringent international standards of plagiarism and blind peer reviews. our 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nadira mamoon mrcpath prof. m. ashraf phd (tamgha-e-imtiaz) prof. m. arif nadeem fcps prof.khalid mahmood phd dr. sarah ghafoor phd members-international prof.renne koeffel phd (sui) prof.jane banaszak-holl phd(aus) prof. hisham al muhtasebphd (ksa) dr. salma malik m.d (us) dr. sarwat shaheen m.d (us) dr. aliya asher mrcp (uk) dr. uzma nasim siddiqui fcps (aus) section editors: dr. shahilajalil fcps dr. samira haque fcps dr. lubna riaz fcps dr. sadiamaqsood mhpe dr. faraz bokhari m phil dr. noora hassan al-aqmer phd mrs.saimamohsin msc consultant biostatician: mr. muhammad aasim m phil library & information sciences: mr. ihsan basit m phil dr. muhammad shahid soroyaphd technical support: mr.m.saad ullah sheikh (bs.engr) contents reham hassan hezam al-aqmer, sulaiman al-sharabi, asmaa abduljabar alnoor, marwa jamal nasir babreek, ruqaih esmail hussain hanash, najwa yahya ahmed al-ghomari, nusaibah abdulqawi hael al-shameri anemia and the factors associated with it in pregnant women attending al-sabeen maternal and child hospital in sana’a, yemen urooj fatima, moneeb ashraf, maheen fatima, muhammad irfan malik, amer hassan siddiqui, sadaf humayun khan does short term atorvastatin treatment improve symptomatic control in patients with mild to moderate and uncontrolled asthma? amber yaseen, saira amir, rimsha arshad, laiba imran, mehar nigar, muteeba irshad, kainat ashraf, dr umar bacha, afifa tanweer. impact of food literacy on consumer's food purchasing habits and dietary intake a systematic review muhammad jamal, hania abouzer, urooj sajjad, manahil rahat, ishtiaq ahmad, jamal ahad perception, attitude & barriers amongst dentists using digital and electronic technology adeela shahid, muhammad shahid saeed, aasia kanwal, sophia shahid frequency of overweight and obesity and its associated factors amongst school children in lahore pakistan saadia shahzad dual informants reporting: do we observe behavioral problems in primary school children? saba amjad, zahra fatima, nabila kaukab, hadia zulfiqar, saba saleem, bushra ijaz faculty perception about e-learning and teaching anatomy: an experience of covid 19 pandemic saba batool, sheikh maria qammar, tooba malik, naseem saud ahmad, bushra shaheen, javaria fatima anti-ulcerogenic evaluation of torilis leptophylla plant extract on indomethacin induced gastric ulcer in mice muhammad umair abbas, shuja uddin, safdar iqbal, zaigham habib, hafiz m. umar, muhammad naeem hassan, ahmed umair asad khan comparison of external fixation with illizarov method versus open reduction with locking plate in comminuted distal radius fractures saira niaz, shazia abid, adila ashraf, naila mumtaz efficacy of oxytocin in reducing blood loss during abdominal myomectomy hafiz m. umar, shuja uddin, zaigham habib, muhammad umair abbas, safdar iqbal, muhammad naeem hassan, muhammad umair khan, mian muhammad haneef outcome of longer lever arm plates with minimally-invasive-plateosteosynthesis in ao type c distal femoral fractures: a descriptive case series instructions for authors_____________________________________ page 1 6 11 26 31 37 45 51 59 65 70 76 recognised by pmc, hec, cpsp & doaj proceedings shaikh zayed medical complex vol. 36 (4) pcpb/24(94) p-121/3196 oct to dec 2022 editorial board patron in chief: prof. sibgha zulfiqar chairperson & dean chief editor: prof. saadia shahzad alam phd editor: prof. ayesha humayun phd associate editors: prof. m. imran anwar frcs dr. adnan salim mrcp external associate editor: dr. usman iqbal phd members-national: prof. mamoon rashid frcs (sitara-e-eisaar) prof. nadira mamoon mrcpath prof. m. ashraf phd (tamgha-e-imtiaz) prof. m. arif nadeem fcps prof.khalid mahmood phd dr. sarah ghafoor phd members-international prof.renne koeffel phd (sui) prof.jane banaszak-holl phd(aus) prof. hisham al muhtasebphd (ksa) dr. salma malik m.d (us) dr. sarwat shaheen m.d (us) dr. aliya asher mrcp (uk) dr. uzma nasim siddiqui fcps (aus) section editors: dr. shahilajalil fcps dr. samira haque fcps dr. lubna riaz fcps dr. sadiamaqsood mhpe dr. faraz bokhari m phil dr. noora hassan al-aqmer phd mrs.saimamohsin msc consultant biostatician: mr. muhammad aasim m phil library & information sciences: mr. ihsan basit m phil dr. muhammad shahid soroyaphd technical support: mr.m.saad ullah sheikh (bs.engr) contents reham hassan hezam al-aqmer, sulaiman al-sharabi, asmaa abduljabar alnoor, marwa jamal nasir babreek, ruqaih esmail hussain hanash, najwa yahya ahmed al-ghomari, nusaibah abdulqawi hael al-shameri anemia and the factors associated with it in pregnant women attending al-sabeen maternal and child hospital in sana’a, yemen urooj fatima, moneeb ashraf, maheen fatima, muhammad irfan malik, amer hassan siddiqui, sadaf humayun khan does short term atorvastatin treatment improve symptomatic control in patients with mild to moderate and uncontrolled asthma? amber yaseen, saira amir, rimsha arshad, laiba imran, mehar nigar, muteeba irshad, kainat ashraf, dr umar bacha, afifa tanweer. impact of food literacy on consumer's food purchasing habits and dietary intake a systematic review muhammad jamal, hania abouzer, urooj sajjad, manahil rahat, ishtiaq ahmad, jamal ahad perception, attitude & barriers amongst dentists using digital and electronic technology adeela shahid, muhammad shahid saeed, aasia kanwal, sophia shahid frequency of overweight and obesity and its associated factors amongst school children in lahore pakistan saadia shahzad dual informants reporting: do we observe behavioral problems in primary school children? saba amjad, zahra fatima, nabila kaukab, hadia zulfiqar, saba saleem, bushra ijaz faculty perception about e-learning and teaching anatomy: an experience of covid 19 pandemic saba batool, sheikh maria qammar, tooba malik, naseem saud ahmad, bushra shaheen, javaria fatima anti-ulcerogenic evaluation of torilis leptophylla plant extract on indomethacin induced gastric ulcer in mice muhammad umair abbas, shuja uddin, safdar iqbal, zaigham habib, hafiz m. umar, muhammad naeem hassan, ahmed umair asad khan comparison of external fixation with illizarov method versus open reduction with locking plate in comminuted distal radius fractures saira niaz, shazia abid, adila ashraf, naila mumtaz efficacy of oxytocin in reducing blood loss during abdominal myomectomy hafiz m. umar, shuja uddin, zaigham habib, muhammad umair abbas, safdar iqbal, muhammad naeem hassan, muhammad umair khan, mian muhammad haneef outcome of longer lever arm plates with minimally-invasive-plateosteosynthesis in ao type c distal femoral fractures: a descriptive case series instructions for authors_____________________________________ page 1 6 11 26 31 37 45 51 59 65 70 76 recognised by pmc, hec, cpsp & doaj proceedings shaikh zayed medical complex vol. 36 (4) pcpb/24(94) p-121/3196 oct to dec 2022 editorial board patron in chief: prof. sibgha zulfiqar chairperson & dean chief editor: prof. saadia shahzad alam phd editor: prof. ayesha humayun phd associate editors: prof. m. imran anwar frcs dr. adnan salim mrcp external associate editor: dr. usman iqbal phd members-national: prof. mamoon rashid frcs (sitara-e-eisaar) prof. nadira mamoon mrcpath prof. m. ashraf phd (tamgha-e-imtiaz) prof. m. arif nadeem fcps prof.khalid mahmood phd dr. sarah ghafoor phd members-international prof.renne koeffel phd (sui) prof.jane banaszak-holl phd(aus) prof. hisham al muhtasebphd (ksa) dr. salma malik m.d (us) dr. sarwat shaheen m.d (us) dr. aliya asher mrcp (uk) dr. uzma nasim siddiqui fcps (aus) section editors: dr. shahilajalil fcps dr. samira haque fcps dr. lubna riaz fcps dr. sadiamaqsood mhpe dr. faraz bokhari m phil dr. noora hassan al-aqmer phd mrs.saimamohsin msc consultant biostatician: mr. muhammad aasim m phil library & information sciences: mr. ihsan basit m phil dr. muhammad shahid soroyaphd technical support: mr.m.saad ullah sheikh (bs.engr) contents reham hassan hezam al-aqmer, sulaiman al-sharabi, asmaa abduljabar alnoor, marwa jamal nasir babreek, ruqaih esmail hussain hanash, najwa yahya ahmed al-ghomari, nusaibah abdulqawi hael al-shameri anemia and the factors associated with it in pregnant women attending al-sabeen maternal and child hospital in sana’a, yemen urooj fatima, moneeb ashraf, maheen fatima, muhammad irfan malik, amer hassan siddiqui, sadaf humayun khan does short term atorvastatin treatment improve symptomatic control in patients with mild to moderate and uncontrolled asthma? amber yaseen, saira amir, rimsha arshad, laiba imran, mehar nigar, muteeba irshad, kainat ashraf, dr umar bacha, afifa tanweer. impact of food literacy on consumer's food purchasing habits and dietary intake a systematic review muhammad jamal, hania abouzer, urooj sajjad, manahil rahat, ishtiaq ahmad, jamal ahad perception, attitude & barriers amongst dentists using digital and electronic technology adeela shahid, muhammad shahid saeed, aasia kanwal, sophia shahid frequency of overweight and obesity and its associated factors amongst school children in lahore pakistan saadia shahzad dual informants reporting: do we observe behavioral problems in primary school children? saba amjad, zahra fatima, nabila kaukab, hadia zulfiqar, saba saleem, bushra ijaz faculty perception about e-learning and teaching anatomy: an experience of covid 19 pandemic saba batool, sheikh maria qammar, tooba malik, naseem saud ahmad, bushra shaheen, javaria fatima anti-ulcerogenic evaluation of torilis leptophylla plant extract on indomethacin induced gastric ulcer in mice muhammad umair abbas, shuja uddin, safdar iqbal, zaigham habib, hafiz m. umar, muhammad naeem hassan, ahmed umair asad khan comparison of external fixation with illizarov method versus open reduction with locking plate in comminuted distal radius fractures saira niaz, shazia abid, adila ashraf, naila mumtaz efficacy of oxytocin in reducing blood loss during abdominal myomectomy hafiz m. umar, shuja uddin, zaigham habib, muhammad umair abbas, safdar iqbal, muhammad naeem hassan, muhammad umair khan, mian muhammad haneef outcome of longer lever arm plates with minimally-invasive-plateosteosynthesis in ao type c distal femoral fractures: a descriptive case series instructions for authors_____________________________________ page 1 6 11 26 31 37 45 51 59 65 70 76 advisory board basic & pre-clinical sciences prof. mateen izhar phd, mrcpath prof. sibgha zulfiqar m.phil prof. muhammad suhail m.phil prof. nasreen ehsan m.phil prof. tahira naseem m.phil dr. anwaar bashir m.phil dr. suleman dawood phd (uol) dr. nighat yasmeen phd (kemu) dr. saima chaudhary phd (uhs) medicine and allied prof. azeemtaj fcps prof. abdul shakoor fcps prof. talha mahmud fcps prof. safooraaamir fcps dr. lubna riaz fcps dr. asfandyar fcps surgery and allied prof. ali rafique mirza fcps prof. muhammad ikram fcps prof. imran anwar frcs prof. shafqat mukhtar fcps dr. jamshaid rahim fcps dr. syed sajjad raza kazmi fcps diagnostic division prof. mona aziz fcps prof. saulat sarfraz fcps dr. amir khan frcr peer review board (national) prof. m. nauman ahmad ffarcs prof. naseem saud phd prof. m. ovais omer phd prof. shabbir bhatti phd prof. zahid niaz frcs prof. zamir ahmad phd prof. muhammad arif nadeem fcps prof. muhammad moin frcs prof. khalid mahmood phd prof. sadia chiragh m.phil prof. muhammad aslam fcps prof. abdul mannan m.s prof. naheedhumayun fcps prof. farhat naz fcps prof. abdul hameed frcs prof. rafeeatafweez phd prof. inayat thaver phd peer review board (national) prof. ashraf chaudhry fcps prof. kashif malik fcps prof. aftabturabi phd prof. uzma hussain fcps prof. muhammad pervaiz frcs prof. fouzia shaukat fcps prof. maryam rashid phd prof. rabeia bilal phd prof. khwaja khursheed fcps prof. tanvir akhtar butt ms prof. naila akhtar fcps prof. abida shaheenphd prof. sajida malik m.phil prof. rumina hasan phd prof. khalid javeed khan frcs dr. farooq afzal frcs dr. saman shahid phd dr. muhammad shahzad phd dr. ali hussainy zaidi m.d dr. shahzad khuram akram phd dr. fahad azam phd dr. khadija irfan fcps dr. uzma malik fcps dr. ayesha mallick frcp dr. tehseen haider kazmi fcps dr. khalida ajmal m.phil dr. israr ahmed fcps dr. saima batool fcps dr. saleem muhammad rana phd dr. muhammad khurram habib fcps dr. saba riaz phd dr. raaziatasadduq phd dr. nageen hussain phd dr. saadia nosheen jan fcps dr. soumble zulfiqar phd dr. yasir abbas zaidi fcps dr. muhammad zeeshan sarwar mrcs dr. muhammad imran khokar mrcs dr. nabiha farasat m.phil dr. tayyaba muzaffar m.phil dr. zaeem khalid fcps dr. nafeesah fatimah fcps dr. rabbiya sarwar m.phil peer review board (international) prof. khalid javeed khan frcs prof. renne koeffel phd (switzerland) prof. jane banaszak-holl phd (aus) prof. m. hisham al-muhtaseb phd (jordan) prof. mukhtiar baig phd (ksa) dr. sarwat shaheen m.d (usa) dr. zeeshan i. shaikh m.d (usa) dr. tanzeel qurat ijaz mrcp (uk) dr. hafiz sohail anjum fcps (ireland) dr. rafayazhar frcp (singapore) dr. mazhar nawaz m.d (usa) dr. ahmad azam malik phd (ksa) dr. ishtiaq ahmed fcps (ksa) dr. aamir omairphd (ksa) dr. sabina ahmed mir m.d (usa) dr. anita lamichhane m.d (nepal) dr. faiza durrani phd (uk) dr. waseem lodhi frcog (uk) dr. seerat zahra hammad m.s (ksa) dr. shafya shahid phd (u.s) dr. abdul waheed frcs (u.k) dr. humaira zareen fcps (ksa) dr. amira okud m.d (ksa) dr. asma ahmed mrcp (uk) dr. sabahat sabir mrcog (uk) dr. sarfaraz ahmad frcr (uk) dr. zouina sarfaraz m.d (uk) dr. humaira zaman fcps (uk) inner.indd 1 proceedings s.z.m.c. vol: 34(1): pp. 1-5, 2020. pszmc-734-34-1-2020 problem based learning: learning from the experiences of medical students 1asma rasheed, 2ahsan sethi, 3muhammad afaaq agha, 1sana tariq, 1lubna humayun, 4uzma shaheen 1department of pathology, ucmd, university of lahore 2department of medical education, khyber medical university, peshawar 3department of forensic medicine, shaikh zayed medical complex, lahore 4department of pathology, dera ghazi khan medical college, dg khan abstract introduction: problem based learning (pbl) is student centered learning approach that has been implemented in many medical colleges. since the literature has controversial takes on the utility of pbl, exploring student perspectives might share insights on the contextual merits and demerits of pbl approach. aims & objectives: to evaluate experience of medical students regarding pbl in hybrid integrated curriculum. place and duration of study: may to june 2018 at two medical colleges of lahore (shalamar medical college & university college of medicine and dentistry. material & methods: descriptive cross sectional study conducted in may to june 2018 at two medical colleges. sample size was 188 students of 1st and 2nd year mbbs of institute 1 and 110 students of 1st and 2nd year mbbs of institute 2. pre validated questionnaire was distributed and students were asked to record their experience about pbl using a 5-point’s likert scale. data was analyzed by using non-parametric statistics. results: institute 1 (188 participants), institute 2 (110 participants), females being 205 (68.79%). mean score <3 indicates bad experience while >3 indicates good experience about pbl. results showed that students of both institutes found several key benefits of pbl acquiring critical thinking, problem solving, communication skills and team work. pbl was perceived as better learning approach than lectures. moreover students of institute 1 mean score <3.0 showed dissatisfaction regarding tutor performance in pbl facilitation which was statistically significant (0.048). conclusion: it is recommended that tutor and student training should be mandatory before introducing pbl. pbl session marks can be included in internal assessment. only senior faculty and volunteering to facilitate should conduct pbl sessions. key words: problem based learning, students experiences, pbl tutoring, skill development, pbl critical thinking, teamwork. introduction the change of medical education from traditional to integrated curriculum has been adopted by different medical schools worldwide.1 traditional curriculum is teacher centered while integrated curriculum is more inclined towards student centeredness. one of these approaches is problem based learning (pbl).2 problem based learning is implement in medical colleges either as pure or hybrid models. in pure problem based learning model, it is the main teaching and learning strategy adopted throughout the curriculum. students learning in hybrid problem based learning model is supported by previous knowledge of students through formal lecturing, small group discussion, skill labs before the pbl session.1 few medical schools placed 1st session of pbl at the start of module and students gained knowledge for 2nd session by didactic lectures, small group discussions and by self-directed learning.2 problem based learning is a constructivist pedagogy which has many advantages. it helps students to explore the concepts, build on prior knowledge3 so that they become selfdirected learners, critical thinkers and problem solvers. student become able to integrate basic subjects to clinical subjects more better.4 in addition, pbl also contributes in the development of communication, interpersonal and presentation skills.5 the tutor has important role in pbl sessions as facilitator, to guide students how they keep themselves focused on their objectives.6 that’s why, 2 problem based learning: learning from the experiences of medical students skilled and trained facilitator plays an important part in the accomplishment of pbl session.7 although most studies tend to report an optimistic and positive impact of pbl on students learning, there are some controversial finding as well. for instance few studies reported that pbl is a time consuming process and it may not have a significant impact on knowledge acquisition. two studies in this regard had conflicting findings. chang j revealed pbl as a positive teaching and learning strategy which enhances student’s knowledge and interpersonal skills with potential areas of improvement and continued fine-tuning.8 on the other hand, khan ia suggested that pbl was time taking, resource demanding approach, majority students were unable to identify the benefits of pbl.9 these controversial findings demand further exploration of student perceptions. this study aimed to investigate the experience of medical students in different educational settings to understand the expected pbl outcomes (skill development, studying in pbl small groups, learning preference and tutoring practices).10 material and methods this descriptive cross sectional study was conducted from may 2018 to june 2018 at two different medical colleges, the university college of medicine and dentistry, university of lahore and shalamar medical and dental college lahore, the targeted population was 1st year and 2nd year mbbs students. a total of 188 students of 1st year and 2nd year mbbs of the university college of medicine and dentistry and 110 students of 1st and 2nd year of shalamar medical and dental college were targeted. inclusion criteria: 1st year and 2nd year undergraduate mbbs students of either gender from the academic year 2017-18. exclusion criteria: students of bds, post graduate students, incomplete questionnaires were excluded, those students not willing to participate. data collection technique: this study was based on self-administered and pre-validated questionnaire on experience of medical students regarding pbl. the questionnaire was devised by extensive literature review using the search engines medline, pubmed and google scholar. the key words were problem based learning experience of medical students, pbl in integrated curriculum, prevalidated questionnaire on pbl, controversial studies on pbl. questionnaire was reviewed by current research supervisor and piloted on twenty third year medical students. the questionnaire was divided into four subscales and had total 16 items. the first subscale consisted of 5 questions, which measured the participants experience related to the benefits of problem based learning sessions regarding skill development. second subscale consisted of 4 questions, which measured the student’s experience of working and study in groups (group dynamics) during and in between pbl session. the third subscale consist of 2 questions which measured students learning preference pbl or lecture. the fourth subscale consisted of 4 questions, which measured the students experience about tutor practice during pbl session. last item was open ended question, if someone liked to add anything else regarding pbl. these questions were answered on a five point’s likert scale. strongly disagree (1), disagree (2), neutral (3), agree (4) and strongly agree (5) ethical approval: the research ethical committee, university college of medicine and dentistry, university of lahore approved the study. participants were informed about purpose of the study, information about items in the prevalidated questionnaire were explained and study subjects individuality was assured by giving them a code before data analysis. statistical analysis: data were coded and entered into microsoft excel software and analyzed by using spss version 21.0. data were analyzed and presented as mean+ sd. data was not normally distributed, shapiro wilk test were applied (p= 0.00) therefore non parameter test such as mann-whitney –u test were used to identify the significance difference (p <0.05) among the subscale and groups (institute 1 and 2). results fig-1: number of participant fig-2: gender distribution of institute wise participants 0 50 100 150 200 institute 1 institute 2 188 110 female 69% male 31% 3 problem based learning: learning from the experiences of medical students s. no item institue 1 n=188 mean+ sd institue 2 n=110 mean+sd p value institue 1 and 2 1 i gained critical thinking skills through pbl tutorials 3.27+ 1.34 3.31+ 1.08 0.484 2 i gained problemsolving skills in pbl tutorials 3.47+ 1.23 3.29+ 1.02 0.016 3 i gained the ability to think laterally in solving problems in pbl tutorials 3.26+ 1.18 3.32+ 1.12 0.472 4 i have gained skills in making diagnosis in pbl 3.43 + 1.23 3.47 + 1.05 0.859 5 pbl enhanced my communication skills development 3.67 + 1.26 3.56 + 1.16 0.12 sum (mean score) 3.42+ 0.16 3.39+ 0.11 0.32 6 pbl tutorials promote team work 3.59 + 1.29 3.59 + 1.17 0.11 7 participants in pbl group contributes actively in the discussion 3.17 + 1.33 3.07 + 1.24 0.987 8 small groups in pbl session encourage the students to share their ideas in group 3.49 + 1.28 3.63 + 1.06 0.179 9 students like to share their knowledge with group members in pbl session 3.69 + 1.20 3.55 + 1.15 0.08 sum (mean score) 3.48+ 0.22 3.4+ 0.22 0.12 10 students prefer pbl classes on didactic lecturing 2.93 + 1.32 3.17 + 1.18 0.13 11 students understand difficult concepts much better in pbl rather than teach in a lecture 3.25 + 1.34 3.45 + 1.18 0.06 sum (mean score) 3.09+ 1.33 3.3+ 1.18 0.16 12 tutor provides a relax atmosphere during pbl session 3.19 + 1.38 3.46 + 1.12 0.007 13 tutor asks frequent questions during session about the problem being solved 2.65 + 1.26 3.22 + 1.05 0.197 14 tutor uses questions to keep the students on track 2.45 + 1.29 3.41+ 1.06 0.009 15 tutor actively resolves the conflicts among group participants 2.80 + 1.33 3.50 + 1.10 0.02 sum (mean score) 2.7+ 0.32 3.2 +0.4 0.048 table-1: evaluation of experience of medical students regarding pbl in two different institutes. discussion according to this study, the students of both institutes found pbl a better learning methodology in acquiring critical thinking, problem solving, communication skills and studying in small groups. pertaining to tutor performance, the institute 1 mean score <3.0 showed student’s dissatisfaction towards their tutor performance in pbl conduction, p value (0.048) is statistically significant. in imparting skill development subscale (item 1-5) the mean score of institute 1 was 3.42, which is higher than institute 2(3.39), that was good response (table-1). this shows that the pbl system of the institute 1 is more effective in imparting skill development competencies in its students as compared to the institute 2. regarding group process subscale (item 6-9) the mean score was 3.4 for both institute which is a concurrent finding. the pbl system is equally effective in both institutes when it comes to the outcome of group process. pertaining to learning preference subscale (item 1011) the mean score of institute 1 and institute 2 is 3.09 and 3.4 respectively. both institutes showed adequate response but the institute 2 is lead in generating learning preferences about pbl system. regarding tutor practice subscale (item 12-15) the mean score of institute 1 was 2.7 which showed their dissatisfaction towards tutor practice in pbl conduction while the mean score of institute 2 was 3.2 which was superior response towards their tutor practice in pbl conduction. the finding was similar to those from turan etal, in which students are not satisfied with their tutor performance in pbl process. this may be because of unpreparedness of tutor for pbl (tutor didn’t go through facilitator guide for pbl), lack of training of faculty who were involved in pbl conduction. on the other hand students of institute 1 devalued their tutors because of other factors, as students may have had issues with a specific tutor or vice versa. it is suggested that proper and frequent training sessions should be conducted for faculty, hiring of more qualified, experienced faculty and, assigning pbl to well versed, passionate faculty members and voluntary participants so they follow all steps of pbl in true spirit. this finding is supported by another study conducted by el aziz el naggar11 in which he evaluated the educational effectiveness of implementing a facilitator training workshop. results of his study showed that facilitator training workshop was effective in improving tutor facilitation skills in the areas of constructive active learning, collaborative learning, self-directed 4 problem based learning: learning from the experiences of medical students learning, and increased the educational outcomes of the pbl sessions from students. it was observed in open ended question of this study survey, most of the students complained that pbl marks were not included in internal evaluation, therefore students were not taking pbl and tutor sincerely. it was deduced that pbl marks should be included in internal evaluation.12 results of this study showed that students found pbl a better learning approach with successful outcome. that pbl should be commenced by more medical colleges gradually, to extract and spread advantages of pbl. the study also revealed the importance of evaluation of effectiveness and weaknesses of existing pbl model which would eventually help in improvement through feedback of actual product of this system. this cross-sectional study was based on selfreported information provided by the students and on convenient sampling method. due to limited time frame, this was a pilot study of small sample size in between two institutes only. similar studies should be conducted in other medical colleges, upon introduction of pbl as one of the methods of active learning in curriculum. their faculty and administrative staff involvement is essential for data regarding logistics, feedback, concerns and hindrances pertaining to pbl. conclusion this study reveals that medical students of both institutes consider pbl as valuable learning methodology. it also emphasizes the important role of pbl in curriculum which enables self reflection in students, regarding enhancement in their different learning skills and helps students assess their tutor performance during pbl conduction. the investigator strongly recommends tutor and student training be made mandatory before introducing pbl. references 1. al-drees aa, khalil ms, irshad m, abdulghani hm. students’ perception towards the problem based learning tutorial session in a system-based hybrid curriculum. saudi med j.2015;36(3):341-8 2. hande s, mohammed ca, komattil r. acquisition of knowledge , generic skills and attitudes through problem-based learning : student perspectives in a hybrid curriculum. j taibah univ med sci. 2015; 10(1):21-5. 3. jawawi r, matzin r, shahrill m, jaidin jh. the benefits of adopting a problem-based learning approach on students. learning developments in secondary geography lessons. 2016; 1(2): 25-27. 4. silva a, bispo a, rodriguez d, vasquez f. problem-based learning, revista de gestão. 2018; 25(2):160-177. 5. pagander l, read j. is problem-based learning (pbl) an effective teaching method ? a study based on existing reserch. seminariedatum. 2014; 581(83):1-45. 6. yew ehj, goh k. problem-based learning: an overview of its process and impact on learning. heal prof educ. 2016; 2(2):75-9. 7. wijnen m, loyens smm, smeets g, kroeze m, van der molen h. comparing problem-based learning students to students in a lecture-based curriculum: learning strategies and the relation with self-study time. eur j psychol educ. 2017; 32(3):431-47. 8. chang jb. problem-based learning in medical school: a student's perspective. annals of medicine and surgery. 2016; 12:88-89 9. khan ai, al-swailmi fk. perceptions of faculty and students regarding problem based learning: a mixed methods study. j pak med assoc. 2015; 65(12):1334-38. 10. azu oo. a survey of problem-based learning and traditional methods of teaching anatomy to 200 level pharmacy students of the university of lagos , nigeria. humanity & social sciences journal. 2014; 5(2):105-111. 11. el aziz, el naggar. effectiveness of implementing a tutor training workshop for problem based learning class tutors at the faculty of medicine, suez canal university. intel prop rights. 2013; 1(1):1-10. 12. cavanagh a, vanstone m, ritz s. problems of problem-based learning:towards transformative critical pedagogy in medical education. perspect med educ. 2019; 8(1):38-42. 13. mahmood su, syed f, khan nr, batool z, rehman r.comparison of problem based with case based learning. pak j physiol. 2017; 13(4):52-6. the authors: dr. asma rasheed assistant professor, department of pathology, ucmd, the university of lahore. dr. ahsan sethi assistant professor, department of medical education, institute of health professions education and research, khyber medical university, peshawar 5 problem based learning: learning from the experiences of medical students dr. muhammad afaaq agha assistant professor department of forensic medicine, shaikh zayed medical complex, lahore. dr. sana tariq demonstrator, department of pathology, ucmd, the university of lahore. dr. lubna humayun assistant professor, department of pathology, ucmd, the university of lahore. dr. uzma shaheen assistant professor, department of pathology, dera ghazi khan medical college, dg khan. corresponding author: dr. asma rasheed assistant professor, department of pathology, ucmd, the university of lahore. e-mail: asma.rasheed@ucm.uol.edu.pk file.indd 1 proceedings s.z.m.c. vol: 35(2): pp. 1-6, 2021. pszmc-784-35-2-2021 1dammam medical complex, dammam, saudi arabia 2department of pathology, king abdulaziz university, jeddah, saudi arabia 3department of family and community medicine, king abdulaziz university, jeddah, saudi arabia 4department of medicine, king abdulaziz university, jeddah, saudi arabia 5university institute of public health, the university of lahore, pakistan 2 awareness and prevalence of risk factors of chronic kidney disease in rabigh, kingdom of saudi arabia 3 awareness and prevalence of risk factors of chronic kidney disease in rabigh, kingdom of saudi arabia 4 awareness and prevalence of risk factors of chronic kidney disease in rabigh, kingdom of saudi arabia 5 awareness and prevalence of risk factors of chronic kidney disease in rabigh, kingdom of saudi arabia 6 awareness and prevalence of risk factors of chronic kidney disease in rabigh, kingdom of saudi arabia numbering.indd 16 proceedings s.z.m.c. vol: 34(4): pp. 16-20, 2020. pszmc-765-34-4-2020 localization of trochlear spine & fovea in human skulls 1iram atta, 1maria nouman, 2raafea tafweez 1department of anatomy, king edward medical university, lahore 2department of anatomy and histology, king edward medical university, lahore abstract introduction: many otolaryngologist and the orbital surgeons have been devoted to finding a better way to achieve optimal outcome in fields of the paranasal sinuses and surgery on medial orbital wall. this could be achieved when the surgeon gives sufficient consideration to orbital anatomy. the cartilaginous pulley of superior oblique muscle is vulnerable during the operations on frontoethmoidal sinus. this pulley passes through an anatomical landmark called trochlear fovea and spine aims & objectives: to describe proper location of trochlear fovea and spine by using the measurements of frontal and sagittal plane. place and duration of study: the study was conducted at king edward medical university, lahore in 2020 on 61 dry skulls. material & methods: four measurements were taken in frontal plane using two lines passing at right angle through supra orbital notch and fronto zygomatic suture while in sagittal plane the distance of fovea/spine was measured from orbital margin anteriorly and optic canal posteriorly. results: the distance of fovea and spine from the lines along supra orbital notch and fronto zygomatic suture was 7.22 ± 0.93mm and 6.14 ± 0.83mm respectively. it was 3.77 ± 0.73mm behind the margin and 38.22 ± 2.98mm in front of optic canal. conclusion: to prevent unwanted injury to superior oblique pulley surgeons should know the topographic location and variation of fovea and spine so that the postoperative diplopia could be prevented while approaching the paranasal sinuses and surgery on medial orbital wall. key words: trochlear spine, trochlear fovea, orbital cavity introduction with the advancement of surgical techniques in the field of ophthalmology, more studies on the relations of various orbital structures is needed so that the postoperative complications of these procedures could be avoided.1 the eyeball is moved by the extra ocular muscles. the strap shaped four recti arise from the common tendinous ring located at the orbital apex. as these muscles pass forward from the apex they broaden out and form cone of muscles around the eye.2 the orientation of recti is in such a way that lateral rectus is parallel to the lateral wall of the orbit, medial rectus is along a sagittal plane, the superior and inferior recti following the orbital axis roughly forms an angle of 23° with the medial and lateral walls of the orbit.3 on the other hand, the two oblique muscles approach the eyeball in a front to back and medial to lateral direction. inferior oblique, a thin, narrow muscle arises from floor of the orbit and is inserted into the lateral part of the sclera behind the equator of the eyeball. whereas the superior oblique, a fusiform muscle arises from the body of the sphenoid superomedial to the optic canal and runs forwards to end in a round tendon that passes through a fibrocartilaginous loop or pulley, the trochlea, which attached to the trochlear fovea of the frontal bone and the tendon then gets attached to the sclera in the superolateral part of the posterior quadrant behind the equator.4 the trochlear fovea is a 3-5 mm shallow depression on the frontal bone at the anteromedial aspect of the orbital roof. bony spicula called as trochlear spine sometimes arises from this fovea. their location is at the superomedial angle of orbit being 4-5 mm behind the orbital opening5. localization of trochlear fovea and spine is of great importance for the ophthalmologist while dealing with the frontal sinus surgeries as many cases of post-operative diplopia have been reported due to injury to trochlea during the surgical procedure.6,7 moreover repair of fractures at the orbital roof could also result in disruption of the trochlea leading to visual disparity.8 therefore, to provide ophthalmologist with some useful references on measurements of human dry skulls which were used to document 17 localization of trochlear spine & fovea in human skulls incidence and variation in the anatomical location of trochlear fovea and spine in pakistani population. material and methods the study was conducted on 61 dry adult human skulls stored in the department of anatomy, king edward medical university, lahore, pakistan. overall 122 orbits were observed. out of these the data could be collected from 120 orbits as 2 were spoiled. all the skulls were examined to determine the gender, incidence and exact location of trochlear fovea or spine. the sex of the skull was determined by the morphological discriminant method.9 in order to locate the trochlear fovea or spine two lines were drawn in the frontal plane from the supraorbital notch and fronto zygomatic suture respectively. these two significant anatomical landmarks were used to measure the interval of trochlear spine and fovea from the roof, medial wall, supraorbital notch and fronto zygomatic suture. a vertical line was drawn from the supraorbital notch (l1) and a horizontal line was drawn through fronto zygomatic suture in the frontal plane (l2). (l3) and (l4) were drawn through the trochlear fovea and spine base transversely and vertically respectively. distances measured between the base of the spine and center of the fovea and l1 and l2 were recorded as d1 and d3 respectively (fig-1), gap between l3 and the orbital roof measured along l1is shown as d2. interval between l4 and the medial wall of the orbit measured along l2 is represented as d4 (fig-2). further two measurements were taken in the sagittal plane to localize the trochlear fovea and spine. d5 was measured from the anterior orbital opening and the base of the trochlear spine and center of the fovea so d6 was recorded from the anterior margin of the optic canal and the base of the trochlear spine and center of the fovea (fig-3). all the readings were recorded with the vernier calipers and were taken in millimeters. fig-1: (l1) and (l2) are shown passing through the supraorbital notch and fronto zygomatic suture respectively. (d1) indicates the interspace between trochlear fovea/spine and (l1) while (d3) indicates the interspace from (l2). fig-2: (l3) and (l4) are drawn perpendicular to each other through the trochlear fovea/spine. along l1, d2 was recorded between orbital roof and l3. moreover, d4 was taken along l2 between the medial wall of the orbit and l4. fig-3: d5 is shown along the sagittal plane from the anterior orbital opening and trochlear fovea or spine. d6 represents the distance of spine or fovea from the optic canal. statistical analysis: data was entered and analyzed by using spss-21-. student t-test was applied to authenticate the difference between male and female position of trochlear fovea and spine. significance between the incidence of the trochlear fovea and spine in male and female skulls was tested by z-test. p<0.05 was taken as significant. 18 localization of trochlear spine & fovea in human skulls results 120 orbits were studied in which trochlear fovea was present in 101 orbits and trochlear spine was observed in 20 orbits. trochlear fovea trochlear spine side number % side number % right 55 54.45 right 15 75 left 46 45.54 left 5 25 total 101 84.17 total 20 16.67 table-1: incidence of trochlear fovea and spine the incidence of trochlear fovea was found to be 84.17% with more prevalence on right side. whereas the incidence of spine was 16.67 % with the percentage on right being more than on left. moreover t-test showed that there was no sex related preference of trochlear fovea and spine as the frequency of fovea in male (84.2%) and female (80%) showed no statistical difference. similarly, percentage of trochlear spine in male (16.7%) and female (19.8%) did not show any significant difference statistically. distance measured mean (mm) maximum (mm) minimum (mm) d1 7.22 ± 0.93 8.64 5.35 d2 4.86 ± 0.93 6.98 3.04 d3 6.14 ± 0.83 8.13 4.43 d4 0.98 ± 0.48 1.92 0.10 table-2: measurements of trochlear spine and fovea in frontal plane where d1 is the distance measured from fovea/ spine till supraorbital notch (l1). d2 is the distance measured from fovea/ spine till orbital roof along l3. d3 is the distance measured from fovea/ spine till fronto zygomatic suture (l2). d4 is the distance measured from fovea/ spine till medial wall of orbit along l4. on applying ttest values of d1 and d4 showed no statistical difference in male and female whereas d2 and d3 were significantly different in male and female as depicted in table-3 d1 mean (mm) d2 mean (mm) d3 mean (mm) d4 mean (mm) male 7.20 ± 0.90 4.59 ± 0.75 6.64 ± 0.75 0.98 ± 0.4 female 7.29 ± 0.96 5.81 ± 0.85 5.75 ± 0.69 0.96 ± 0.3 p-value 0.49 0.01 0.02 0.43 p-value less than 0.05 is taken as significant table-3: mean values of distances measured in male and female distance measured mean (mm) maximum (mm) minimum (mm) d5 3.77 ± 0.73 5.71 2.4 d6 38.22 ± 2.98 44.35 29.18 table 4: measurements of trochlear spine and fovea in sagittal plane where d5 is the distance measured between fovea/spine and anterior orbital opening d6 is the distance measured between fovea/spine and optic canal like the measurements taken in frontal plane, d5 values also didn’t show any statistically significant difference between male (3.81± 0.77) and female (3.62 ± 0.73). however there was statistically significant difference among values of d6 in male orbits (38.77 ± 3.1) and female orbits (35.25 ± 2.7) discussion the surgeries for frontal sinus or even ethmoidal sinus involving an internal approach with a rigid endoscope and external approach via skin crease the upper eyelid, is followed sometimes postoperatively by diplopia.10 the diplopia results from damage to the pulley of superior oblique. the anatomical landmark which marks the location of this cartilaginous pulley is the trochlear fovea and spine which in turn is located at the superomedial wall of orbit.11 surgical repair of medial blowout fractures of orbit could also result in damage to the muscle pulley.12 therefore the current study was attempted to document about the incidence, location and variation of fovea and spine in pakistani population to prevent the unwanted damage to cartilaginous pulley postoperatively. in this study the incidence of trochlear fovea was found to be 84.17% while that of trochlear spine was 16.67%. this incidence can be compared with italian population only since there is no data available on any other population. the frequency of fovea in italian populace was 73.39% and that of spine was 15.32%.3 where frequency of fovea was a bit higher in pakistani population, spine frequency was almost the same. further it was observed in the study that prevalence of spine was more on the right side than left. moreover, it was seen that there was no sex related preference of the occurrence of fovea and spine in female and male orbits. fovea was seen in 84.2% of male orbits and 80% of female orbits. likewise spine frequency in male (16.7%) and female orbits (19.8%) was not statistically significant. aglianó m et al (2018) had reported the same finding in their study. 19 localization of trochlear spine & fovea in human skulls the position of trochlea was defined by taking two measurements (d5 and d6) in sagittal plane. it was found to be 3.77 ± 0.73mm behind the orbital margin and 38.22 ± 2.98mm in front of optic canal. but the surgeon should keep in mind the variation that it could be found anywhere between 2.4 to 5.7 mm behind the orbital opening. more explanation on the location of trochlea four measurements (d1, d2, d3 and d4) were taken in frontal plane. in this regard two perpendicular lines were drawn from supra orbital notch and fronto zygomatic suture. the distance of trochlea from supra orbital notch (d1) was found to be 7.22 ± 0.93mm and that from orbital roof (d2) was found to be 4.86 ± 0.93mm. trochlea was found to be at 6.14 ± 0.83mm from fronto zygomatic suture (d3) and at 0.98 ± 0.48mm from medial wall of orbit (d4). the distance (d3) from the horizontal line (l2) ranges between 8.1 and 4.4 mm while on the contrary the distance (d1) from vertical line (l1) ranges between 8.6mm to 5.4mm. the measurements which used in this study were quite comparable to the italian population.3 but when we analyzed the measurements in male and female orbits, there was no significant difference statistically in the values of d1, d4 and d5 but d2, d3 and d6 showed significant difference (table-3). this finding was contrary to the italian study which reported the significant difference only in the measurement of d6 between male and female orbits. this disparity might be due to the difference in ethnic groups. this difference in d6 between male (38.77 ± 3.1) and female (35.25 ± 2.7) should be expected as there is a substantial difference in the average size of male and female orbits.13 the present study revealed the precise position of trochlear fovea and spine in pakistani population. we noticed that the position of fovea from orbital roof and fronto zygomatic suture differed in male and female pakistani skull, a finding contrary to italian populace. the reason for this disparity might be race related but still enough data on different ethnic groups is not available. so further research should be done to find out the cause of variation in its location in different populations. conclusion the knowledge of topographic location and variation of trochlear fovea and spine is of extreme significance not only to the ophthalmologists but also to the ent surgeons for they could prevent postoperative diplopia by sparing the pulley of the superior oblique muscle. references 1. haladaj r. normal anatomy and anomalies of the rectus extraocular muscles in human: a review of the recent data and findings. biomed res int. 2019. 2. sinnatamby cs. last’s anatomy regional and applied. 12th ed. edinberg: elsevier; 2011. chapter 6, head and neck and spine; p.399-402 3. aglianó m, franci d, volpi n, orsini d, messina m, lorenzoni p. osteologic topography of the trochlear spine and fovea as landmarks to locate the superior oblique trochlea. italian journal of anatomy and embryology. 2018; 123:304-11. 4. standring s. gray’s anatomy. 41st ed. united kingdom: elsevier; 2015. chapter 41, orbit and accessory visual apparatus; p. 670-75. 5. lang j. clinical anatomy of head. berlin: springer-verlag; 2012. chapter 3, orbit and contents; p. 62-3. 6. locker p, plitt m, papagiannopoulos p, smith r, tajudeen ba. anatomic relationship of the first olfactory neuron and trochlea: cadaveric study with surgical implications. int forum allergy rhinol. 2017; 7:1085-88. 7. lin js, liu tt, manes rp, galvin ja. superior oblique palsy: a complication of endoscopic sinus surgery. jaapos. 2015; 19:180-1. 8. choi j, lorenz hp, spain da. review of facial trauma management. j trauma acute care surg. 2020; 88:124-30. 9. standring s. gray’s anatomy. 41st ed. united kingdom: elsevier; 2015. chapter 27, external skull; p. 416-28. 10. beigi b, vayalambrone d, kashkouli mb, prinsley p, saada j. combined external and endonasal approach to fronto-ethmoidal mucocele involving the orbit. j curr ophthalmol. 2016; 28:37-42. 11. alekseenko s, karpischenko s. comparative analysis of the outcome of external and endoscopic frontal sinus surgery in children. acta otolaryngol. 2020; 140:687-92. 12. ji sy, yoo jh, ha w, lee jw, yang ws. three cases of acquired simulated brown syndrome after blowout fracture operations. arch plast surg. 2015; 42:346. 13. avelar le, cardoso ma, bordoni ls, de miranda avelar l, de miranda avelar jv. aging and sexual differences of the human skull. plast reconstr surg glob open. 2017; 5. 20 localization of trochlear spine & fovea in human skulls the authors: dr. iram atta demonstrator, department of anatomy, king edward medical university, lahore. dr. maria nouman demonstrator, department of anatomy, king edward medical university, lahore. prof. raafea tafweez professor and chairperson, anatomy and histology department, king edward medical university, lahore. corresponding author: dr. maria nouman demonstrator, department of anatomy, king edward medical university, lahore. e-mail: marriahnouman@gmail.com numbering.indd 10 proceedings s.z.m.c. vol: 34(4): pp. 10-15, 2020. pszmc-764-34-4-2020 frequency of occult hepatitis b in pregnant women attending antenatal care unit of a tertiary care hospital 1sameen bint ali, 2mateen izhar, 2chetan lal, 3asma yaqoob, 2hadiqa tul hafsa, 4asma akram 1department of pathology, services institute of medical sciences, lahore 2department of microbiology and virology, shaikh zayed medical complex, lahore 3department of pathology, fatima memorial medical & dental college, lahore 4department of pathology, continental medical college, lahore abstract introduction: occult infections (obis) caused by hepatitis b virus (hbv) are detected by the presence of hbv dna without surface antigens. the prevalence of hbsag in general population of pakistan is around 3%, but there is no data regarding the occurrence of obis. aims & objectives: the goal of this research was to assess the frequency of occult hepatitis b infection in pregnant females presenting to antenatal care unit of shaikh zayed hospital, lahore. place and duration of study: these samples were handled at the microbiology department of shaikh zayed hospital, lahore. the duration of study was one year after the approval of synopsis. material & methods: four hundred and sixty three pregnant women were included in this research. serum was separated from their blood samples. hbsag, anti-hbcand anti-hbs tests were performed using the elisa technique on all the samples. real time pcr was performed to find hbv dna. results: the frequency of occult hepatitis b was < 0.25 in this research. among the 463 women tested during pregnancy, hepatitis b surface antigen was positive in 2 (0.4%) women. these two were excluded from the study. out of 461 samples, hepatitis b surface antibody was positive in 61 (13.2%) samples and they were negative for hepatitis b core antibody. these 61 samples were also omitted from the research. 25 (5%) samples were positive for both anti-hbc and anti-hbs. 11 samples (2%) were anti-hbc positive and antihbs negative. in 364 (79%) samples, both anti-hbs and anti-hbc were absent. polymerase chain reaction was performed on 400 samples. conclusion: the frequency of occult hepatitis b is very low < 0.25% in pregnant women. it is not recommended to routinely screen pregnant women for hepatitis b virus dna. key words: hbv dna, occult hepatitis, pregnant women introduction occult hepatitis b (obi) is defined when hepatitis b virus dna (hbv dna) is present in the liver or serum of individuals who are negative for hbv surface antigen (hbsag).1 in cases of true obi the amount of viral dna in the serum is typically very low that is less than 200 iu/ml. since it is not always practical or possible to test liver tissue, serum hbv dna and viral marker tests are often used to diagnose obi.2 as the endemic level of hbv is different in different parts of the globe so it makes the prevalence of obi quite variable. around the world it is stated to fluctuate from 1% to 95%.3 this pervasiveness rate relies upon geographic contrasts (endemicity) and diverse patient qualities. obi has important clinical significance because hbv can be transmitted through transfusion, organ transplantation and perinatal route.4 in patients with low immunity or those who are having chemotherapy occult hbv infection can reactivate and lead to acute hepatitis which may develop hepatic fibrosis.3,5,6 by its proto-oncogenic effect it is also a risk factor that can develop hcc.7 obi is related in some cases to virus genetic variants. a modified surface antigen may be produced by the virus that is not detected by the diagnostic assays. according to different studies it may be due to host immune system which strongly suppress replication-competent viruses in their replication and transcriptional activities.8,9 in obi, absence of hbsag may also be because of the screening done during very initial stage of acute infection before the development of antibodies and in the serum hbsag is also not present. this also 11 frequency of occult hepatitis b in pregnant women attending antenatal care unit of a tertiary care hospital happens during chronic hbv infection when hbsag is declines to a very low level that cannot be detected.10 on the basis of the hepatitis b virus antibody profile, obi can be distinguished into seropositiveobi and seronegative-obi. in seropositive-obi anti hbc is positive irrelevant with the positivity of anti hbs. in seronegative-obi antibodies of both anti hbc and anti hbs are negative.11 approximately 10% of the patients have shown occult hbv infection (obi) in whom hbsag is negative and anti hbc is positive, in which serum hbv dna is measurable. detection of hbsag alone is not sufficient for identifying all chronic hbv infection. because it may effect patient hbv screening, blood donation, follow-up of patients who have cleared the infection.1,12 various clinical contexts are seen in which occult hepatitis b virus may occur including patients with immunosuppresion from endemic areas of hbv. pregnancy is a state related with immune suppression. cellular immunity is disturbed during pregnancy. in normal pregnancy, to evade elimination of the fetus, the maternal th 1 immune response may be suppressed. as a result of this, for the continuation of pregnancy th2 phenotype becomes predominant. cytokines such as interleukin (il)-4 and (il)-10 which are anti-inflammatory and are present during this time may lead to increased risk of infection. it has significant clinical implication not only to the mother’s wellbeing but also because there is a chance of transmission of occult hbv from mother to fetus. one research demonstrates that hbv can be transferred from mother to child even if the mother is negative for hbsag and positive for anti-hbc.13 to examine infants at danger for vertical transmission of hbv infection, serologic screening of pregnant women includes test for hbsag. it is not enough being unable to perceive occult hepatitis b virus infection. recently it was manifested that individuals with occult hbv infection can infect others, even with low viremic level. in highly endemic areas, mother to child (perinatal transmission) spread of hepatitis b is most common.14 approximately 90% of infants born with hbv infection always lead to a chronic carrier state. moreover, vertical transmission of hbv leads infected infants to liver cirrhosis and carcinoma of liver in young adulthood. so it is very important to prevent vertical transmission of hbv.15,16 viral hepatitis during pregnancy is related with a great possibility of maternal problems and it has been reported as the important reason of maternal demise.17 according to a study conducted in district kohat, prevalence of hepatitis b was found to be 6% among pregnant women.18 in another study conducted in rawalpindi military hospital, the prevalence was 4.69% in pregnant women.19 whereas, in normal population hbsag positivity was 2.5% according to the national survey done by the pakistan medical research council (pmrc).20 in all these studies, patients were tested for hbsag but they were not investigated for hbv dna by pcr. as pregnancy itself is a state of immune suppression we think it is under detected and we may be missing occult hepatitis b infection. moreover, there is the risk of vertical transmission of hbv infection it is important to diagnose occult hepatitis b virus infection. this study is planned to detect hbv dna by pcr in pregnant women screened negative by hbsag surface marker. material and methods it was a cross sectional descriptive study. 369 samples of blood were collected based on international data.21 informed consent and history was taken. hbsag negative pregnant women of all ages and parity were included in this examination visiting shaikh zayed hospital from august to october 2017. pregnant women with positive antihbs and negative anti-hbcigg antibody were excluded from this research. these samples were processed for serum at the department of microbiology of shaikh zayed hospital, lahore. hbsag and anti-hbc was estimated using elisa kit manufactured by roche diagnostics usa (cobas e 411). further, the real time polymerase chain reaction was performed to detect hepatitis b virus dna to diagnose occult hepatitis b by roche cobastaqman 48 kit usa. statistical analysis: data was entered and analyzed using spss version 23. p value <0.05 considered significant. results four hundred and sixty three pregnant women were registered in this research. their mean age with standard deviation (mean + sd) was 27.26 + 4.264. the minimum and maximum age of the patient was 17 and 40 years respectively. the highest proportion of participants (43%) was in the age group 26-30 years, followed by 33.5% in the group for 21-25 years. (table-1) the mean standard deviation of the duration of pregnancy was 25.14 weeks + 6.926. the minimum 12 frequency of occult hepatitis b in pregnant women attending antenatal care unit of a tertiary care hospital duration of pregnancy was 4 weeks and maximum duration of pregnancy was 37 weeks. (fig-1) in our study 7.1% of the patients were of high socioeconomic status. 74.1% belonged to middle socioeconomic class and 18.8% of the patients were of low socioeconomic status. (fig-2) out of 463 patients 2 (0.4%) patients were positive for hbsag serological marker and rest were negative (99.6%). and these 2 were excluded from the study. (table-2) out of 461 samples, hepatitis b surface antibody was positive in 61 (13.2%) samples and they were negative for hepatitis b core antibody. these 61 samples were also omitted from the research. 25 (5%) samples were positive for both anti-hbc and anti-hbs. 11 samples (2%) were antihbc positive and anti-hbs negative. in 364 (79%) samples both anti-hbs and anti-hbc were absent. and they were included in the study. (table-3) polymerase chain reaction was done in all patients to identify hbv dna in their serum samples. 400 study samples were run on roche cobastaqman 48 kit usa. hbv dna was not detected in any of the study sample. therefore, according to this study the frequency of occult hepatitis b ranges from 0% to 0.24%. age groups frequency percentage (%) under 20 18 3.9 21-25 155 33.5 26-30 199 43.0 31-35 78 16.8 36 & above 13 2.8 total 463 100.0 table-1: age groups of the patients hbsag frequency percentage (%) negative 461 99.6 positive 2 0.4 total 463 100.0 table 2: frequency and percentage of hbsag in study participants serological markers number (%) isolated anti-hbs positive (anti-hbs positive but anti-hbc negative) 61 13% isolated anti-hbc positive (anti-hbc positive but anti-hbs negative) 11 2% both anti-hbs and anti-hbc positive 25 5% both anti-hbs and anti-hbc negative 364 79% table 3: serological markers of study subjects fig-1: gestational age groups of study participants(weeks) fig-2: socioeconomic status discussion viral hepatitis b is endemicaly present in pakistan.22 it is a major health problem not only in pakistan but worldwide as well.18 in pakistan a study was done in 2016 at rawalpindi medical college to find the prevalance of hepatitis b in pregnant women in pakistan. the study inculded 519 pregnant females. out of 519 pregnant females, 7 were posistive for hbsag. the prevalance of hepatits b was found to be 1.3%.23 in another study conducted in district kohat, khyber pakhtunkhaw 100 pregnant females were tested from 8 december 2016 to 12 june 2017. out of these 100 females 6 women were seropositive against hbv infection. the prevalance of hepatitis b was found to be 6%.18 unidentified occult hepatitis b in pregnant women is a possible cause of infection from mother to fetus. the spread of hepatitis b virus from the pregnant women to her fetus can be stopped by a diagnosis of occult hepatitis b. the prevalance of occult hepatitis b has not been investigated in pregnant women in pakistan. however, a study has been conducted in pakistan in blood donors. 966 blood donors were selected from armed forces institute of transfusion, rawalpindi. eighteen (1.86%) donors were positive for hbsag. out of the remaining 948 13 frequency of occult hepatitis b in pregnant women attending antenatal care unit of a tertiary care hospital hbsag negative blood donors, hbv dna was detected in 5 (0.53%) blood donors.24 prevalance rate of obi among hemophalia patients in pakistan was 1.7% as reported by borhany and colleagues.25 a study conducted in india showed 413 (19.8%) blood donors were positive for hepatitis b core antibody and negative for hepatitis b surface antigen. of these 7.5% were positive for hbv dna.26 our results showed that the mean age of the study participants was 27.26 years and age did not seem to be an issue for hbv infection as there was no change in ages of women with hbv versus those without hbv. patients were selected randomly in our study. their past history of hepatitis b and the hepatitis b core antibody and hepatitis b surface antibody levels were unknown. in our study 61 (13.3%) were positive for hepatitis b surface antibody but negative for hepatitis b core antibody. this indicated that they were self vaccinated. out of these 61 ladies only 11 (18.03%) pregnant women gave history of vaccination rest 50 (81.96) were unaware of their vaccination status. in a study conducted in karachi in 2015, 17% of the females had received hbv vaccination during childhood.22 routine hbv vaccination has been recommended by the world health organization for all children.21 in pakistan, hbv vaccination was included in epi since 2009. because of this prevalance rate of hepatitis b surface antigen has declined and the positive rate of hepatitis b surface antibody has been on the rise. the prevalance rate of occult hepatitis b in pregnant women was 6.6% in botswana. six hundered and twenty two hbsag negative samples were tested for hbv dna. 41 were positive for obi.27 in another study conducted in korea, 4% pregnant women were positive for hbv dna with the taqman pcr assay. 202 healthy pregnant women were enrolled in this study and hbv dna was detected in eight of 202 individuals with taqman pcr assay.21 in our study 25 patients were positive for both antihbc and anti-hbs. this meant that these patients had recovered from past infection and were immune. eleven individuals were positive for antihbc and negative for anti-hbs. serological analysis of occult hepatitis b infection patients showed that they may b either seropositive (antihbc alone or together with anti-hbs positive) or seronegative (anti-hbc and anti-hbs negative). the subjects who are hepatitis b core antibody positive and hepatitis b surface antibody negative have the highest rate of detection of hbv dna.28 so the actual rate of hbv infection in these women is 8.02%. out of which 0.43% were carriers of hbv infection and 7.7% had some past infection. a study conducted at ayub teaching hospital abbotabad from december 2015 to may 2016. through non probablity consecutive sampling 174 pregnant females were registered in the study. elisa was the technique used to screen blood samples for hepatitis b surface antigen. six patients were positive for hbsag. so the prevalance rate was 3.4%.29 3.4% were carriers in comparison to 0.43% in our study. the prevalance of occult hepatitis b infection varies from 1% to 87% from diverse areas of the sphere. in some geographical areas with little hbv endemicity, obi was still reported. a number of factors are responsible for these variations. these include sensitivity of hepatitis b virus dna detection assay, the sample size, and the detection of of hepatitis b virus dna in liver tissue or serum sample.30 the actual prevalance of obi is underestimated by detecting hepatitis b virus dna in the serum. hbv dna detection in liver biopsy is the best way for diagnosing occult hepatitis b infection. but the liver specimen are not easily available as liver biopsy is required and it is a procedure which require invasiveness. moreover for finding hbv dna in the liver tissue, fda has not permitted to use any standardized and valid assay.28 in our study 79% of these women are negative for the antibodies of both hbsag and hbcag and they are the true negatives. and these women can get benefit from vaccination. as carriage rate of hepatitis b is very low in our study population that is why there was no occult hepatitis b detected in these pregnant women as endemicity of hbv infection associates with occurrence of obi.31 conclusion the frequency of occult hepatitis b is <0.25% in pregnant women of shaikh zayed hospital. it was not even present in anti-hbc positive women or those who had no serelogical marker. occult hepatitis b can be transmitted from mother to fetus but routine testing for hbv dna in pregnant women is not recommended at this time from our study. limitations of the study: • the study participants from a single antenatal care centre of shaikh zayed hospital were included in our study population. so the frequency of occult hepatitis b in the pregnant women from all over the country cannot be predicted from our research. 14 frequency of occult hepatitis b in pregnant women attending antenatal care unit of a tertiary care hospital • the true frequency of obi may have been under detected because hepatitis b virus dna was detected on the serum samples and not in the pbmcs and liver. recommendations: we recommend before testing hbsag, anti-hbc screening in patients should be done. if anti-hbc is negative, immunize the patient according to convention pursued by post immunization antihbs levels. if anti-hbc is positive, hbsag should be tested further. if hbsag is positive the patients should be evaluted for establishment of the stage of disease and treatment. if hbsag is negative no further vaccination is required in patients as they had past hepatitis b virus infection. the study participants must be selected from different hospitals units. along with the serum samples, the peripheral blood mononuclear cells and liver specimens (if possible) should be tested for detecting hbv dna. references 1. portilho mm, nabuco lc, villela-nogueira ca, brandão-mello ce, pilotto jh, flores gl, et al. detection of occult hepatitis b in serum and oral fluid samples. memórias do instituto oswaldo cruz. 2018; 113(1):62-5. 2. seo dh, whang dh, song ey, han ks. occult hepatitis b virus infection and blood transfusion. world journal of hepatology. 2015; 7(3):600. 3. kwak m-s, kim yj. occult hepatitis b virus infection. world journal of hepatology. 2014; 6(12):860. 4. rendon jc, cortes-mancera f, restrepogutierrez jc, hoyos s, navas m-c. molecular characterization of occult hepatitis b virus infection in patients with end-stage liver disease in colombia. plos one. 2017; 12(7):e0180447. 5. bajjou t, tagajdid mr, elkochri s, abi r, touil n, rahoui 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diagnosis and clinical significance. world journal of hepatology. 2015; 7(2):253. 11. said zn, el sayed mh, salama ii, aboel-magd ek, mahmoud mh, el setouhy m, et al. occult hepatitis b virus infection among egyptian blood donors. world journal of hepatology. 2013; 5(2):64. 12. romero m, madejón a, fernández-rodríguez c, garcía-samaniego j. clinical significance of occult hepatitis b virus infection. world journal of gastroenterology: wjg. 2011; 17(12):1549. 13. chang p, tu j, chesterman a, kim r, robertson p, rawlinson wd, et al. development of occult hepatitis b viral infection in pregnancy: implications for antenatal screening in women from endemic areas. obstetric medicine. 2010; 3(3):115-8. 14. walz a, wirth s, hucke j, gerner p. vertical transmission of hepatitis b virus (hbv) from mothers negative for hbv surface antigen and positive for antibody to hbv core antigen. the journal of infectious diseases. 2009; 200(8):1227-31. 15. zenebe y, mulu w, yimer m, abera b. seroprevalence and risk factors of hepatitis b virus and human immunodeficiency virus infection among pregnant women in bahir dar city, northwest ethiopia: a cross sectional study. bmc infectious diseases. 2014; 14(1):118. 16. khamduang w, ngo-giang-huong n, gaudygraffin c, jourdain g, suwankornsakul w, jarupanich t, et al. prevalence, risk factors, and impact of isolated antibody to hepatitis b core antigen and occult hepatitis b virus infection in hiv-1–infected pregnant women. clinical infectious diseases.2013;56(12):1704-12 17. banks t, kang j, watts i, tyrosvoutis meg, min am, tun nw, et al. high hepatitis b seroprevalence and risk factors for infection in pregnant women on the thailand-myanmar border. the journal of infection in developing countries. 2016; 10(04):384-8. 15 frequency of occult hepatitis b in pregnant women attending antenatal care unit of a tertiary care hospital 18. hussain r, mansoor h, iqbal t, askar r, shaheen u, rehman hu. prevalence of hepatitis b (hbv) infection in pregnant women in district kohat, khyber pakhtunkhwa, pakistan. 2018. 19. anwar r, razzaq k, imran a. frequency of hepatitis b virus among pregnant women attending military hospital rawalpindi. pakistan armed forces medical journal. 2016; 66(6). 20. ahmad i. prevalence of hepatitis b and c viral infection among pregnant women in peshawar, pakistan. hepatitis monthly. 2016; 16(6). 21. kwon ci, hwang sg, shin sj, chang sw, kim sy, ko kh, et al. occult hepatitis b virus infection in pregnant woman and its clinical implication.liver international.2008;28(5):667-74 22. shakeel s, iffat w, rehman h, jamil n, nazeer f, naveed s, et al. assessing the level of awareness and vaccination status of hepatitis b among young female community of pakistan. bangladesh journal of medical science. 2015; 14(4):376-83. 23. farooq a, rukhsar a, talib n, mahmood s, rukhsar n, asad m. frequency of hepatitis b and hepatitis c in pregnant females of rawalpindi. group.15:25. 24. bhatti fa, ullah z, salamat n, ayub m, ghani e. anti–hepatits b core antigen testing, viral markers, and occult hepatitis b virus infection in pakistani blood donors: implications for transfusion practice. transfusion.2007;47(1):74-9 25. arababadi mk, ahmadabadi bn, daredor hy, kennedy d. epidemiology of occult hepatitis b infection among thalassemic, hemophilia, and hemodialysis patients. hepatitis monthly. 2012; 12(5):315. 26. asim m, ali r, khan la, husain s, singla r, kar p. significance of anti-hbc screening of blood donors & its association with occult hepatitis b virus infection: implications for blood transfusion. indian j med res. 2010; 132(312):7. 27. mbangiwa t, kasvosve i, anderson m, thami p, choga w, needleman a, et al. chronic and occult hepatitis b virus infection in pregnant women in botswana. genes. 2018; 9(5):259. 28. said zna. an overview of occult hepatitis b virus infection. world journal of gastroenterology: wjg. 2011; 17(15):1927. 29. jadoon sm, adeel m, aslam s, rasool a. hepatitis b and hepatitis c virus in women with first pregnancy. journal of ayub medical college abbottabad. 2017; 29(4):614-8. 30. makvandi m. update on occult hepatitis b virus infection. world journal of gastroenterology. 2016; 22(39):8720. 31. raimondo g, caccamo g, filomia r, pollicino t, editors. occult hbv infection. seminars in immunopathology; 2013: springer. the authors: dr. sameen bint ali demonstrator, department of pathology, services institute of medical sciences, lahore. prof. mateen izhar head, department of microbiology and virology, shaikh zayed medical complex, lahore dr. chetan lal assistant professor, department of microbiology and virology, shaikh zayed medical complex, lahore. dr. asma yaqoob senior demonstrator, department of pathology, fatima memorial medical & dental college, lahore. dr. hadiqa tul hafsa demonstrator, department of microbiology and virology, shaikh zayed medical complex, lahore. dr. asma akram senior demonstrator, department of pathology, continental medical college, lahore corresponding author: dr. sameen bint ali demonstrator, department of pathology, services institute of medical sciences, lahore. e-mail: sameenbinteali@gmail.com untitled-1 45 proceedings s.z.m.c. vol: 35(1): pp. 45-50, 2021. pszmc-783-35-1-2021 association of maternal hyperhomocysteinemia with preeclampsia: a hospital based case-control study 1zoofishan qureshi, 1muhammad ashraf chaudhry, 2seema imdad, 3tariq mukhtar farani, 3ayesha humayun 1department of community medicine, cmh lahore medical college, lahore 2department of nutrition and dietetics, institute of public health, lahore 3department of surgery, cmh, lahore 2department of community medicine, shaikh zayed medical complex, lahore abstract introduction: preeclampsia is among the leading causes of feto-maternal morbidity and mortality throughout the world, especially in developing countries like pakistan. the exact pathophysiology of preeclampsia is still unclear but recent research on homocysteine shows its important role. aims & objectives: the aim of this study was to determine association of high serum homocysteine level with preeclampsia among antenatal women. place and duration of study: tertiary care hospitals of lahore (services, lahore general hospital and sir ganga ram hospitals) from 1st january 2019 to 30th june 2019. material & methods: a matched case-control design was employed. sixty-six diagnosed pre-eclamptic antenatal women were selected as cases and they were age and gestational weeks matched with sixty-six normotensive antenatal women as control, with a case to control ratio of 1:1. serum homocysteine level in fasting sample was estimated by enzyme linked immuno assay. data was entered and analyzed through spss version 20. results: the mean plasma homocysteine was significantly higher in cases (16.05 ± 2.25) as compared to controls (9.44±2.83) with a p value <0.001. highly significant statistical association was found between hyperhomocysteinemia and preeclampsia (p-value < 0.001 with adjusted odds ratio of 4.72). conclusion: homocysteine levels a high in preeclamptic women as compared to normotensives showing that hyperhomocysteinemia is significantly associated with preeclampsia in antenatal women. key words: hyperhomocystenemia, preeclampsia, case-control study introduction preeclampsia is among major causes of fetomaternal morbidities and mortalities all over the world. preeclampsia is a hypertensive disorder which is found in 3–10% of pregnancies worldwide.1 prevalence ranges from 1.8% to 16.7% in developing countries.2,3,4 preeclampsia ranks among priority public health issues in both advanced and less developed countries. it contributes significantly to maternal and perinatal morbidities and mortalities. preeclampsia is a pregnancy specific disorder defined as the combination of high blood pressure, (hypertension), swelling (edema) and protein in the urine (albuminuria/proteinuria), developing after the 20th week of pregnancy.5 preeclampsia has also been recognized as significant leading cause of maternal morbidity along with perinatal mortality and these women are more prone to have cardiovascular disease in later life.6 reduced blood flow to placenta causes ischemia/hypoxia which is proposed to trigger the release of a various placental factors that have marked effects on arterial pressure regulation and blood circulation.7 preexisting hypertension, renal disease, strong family history of pre-eclampsia/eclampsia, systemic lupus erythematous and poor antenatal care are among predisposing/risk factors.8,9 it has also been proposed that the vasculature of pregnant mothers may show increased sensitivity to plasma level of homocysteine.10 homocysteine (hhcy) is a non-essential amino acid that contains sulphur and it is derived from demethylation of methionine. methionine is an essential amino acid and it needs folate, vitamin b12 and b6 as coenzymes for its metabolism. women developing severe preeclampsia tend to have higher plasma homocysteine levels as compared to women who 46 association of maternal hyperhomocysteinemia with preeclampsia: a hospital based case-control study have normal blood pressure throughout pregnancy.11 the vascular damage is accompanied by multi organ dysfunction with varying clinical features in advanced disease.12 women with higher concentration of plasma homocysteine (hyperhomocysteinemia) levels during early pregnancy were found to have greater chance of developing preeclampsia and intrauterine growth restriction (iugr).7,13,14,15 recent literature based on large cohorts and many case-control studies reports significant association between homocysteine concentrations in maternal serum during early pregnancy and adverse pregnancy outcome and changes in placental vasculature.16,17 current study was planned and conducted to identify the association as well as the strength of association between homocysteine levels and preeclampsia in local antenatal women of lahore city. material and methods it was a matched casecontrol study conducted in gynecology/obstetric outpatient departments of services hospital lahore, lahore general hospital and sir ganga ram hospitals from 1st january 2019 to 15th july 2019. a total of 132 pregnant women were recruited comprising of 66 cases and 66 controls (keeping the cases to control ratio 1:1) through non–probability purposive sampling technique based on predetermined inclusion and exclusion criteria. matching was done for age and period of gestation. the sample size was calculated using the standard deviation (sd) of first group as 4, sd of second group 5, the mean homocysteine level among normotensive women taken as 17, while in preeclamptic women as 13.50.18 the effect size in 0.77 while the probability of α error was set at 0.05. the power of the study at 0.95 and the allocation ratio for cases and controls is taken as 1. including non-response, the minimum total sample size was estimated to be 100, 50 in each minimum but we increased sample size to 66 in each group so as to enhance sample size a bit. definition, source and selection of cases: pregnant women aged between 20 to 35 years at ≥ 20 weeks of gestation having confirmed preeclampsia by a gynecologist (with systolic blood pressure≥140mmhg and diastolic blood pressure≥90mmhg along with presence of albuminuria i-e1+ or more by dipstick method) were included in the study. these females were recruited through purposive sampling after taking written informed consent from antenatal opds of selected tertiary care hospitals of lahore. definition, source and selection of controls: pregnant women aged between 20 to 35 years at ≥ 20 weeks of gestation, normotensive with systolic blood pressure≤120mmhg and diastolic blood pressure ≤80mmhg and no albumin in urine were enrolled as controls. these females were recruited through purposive sampling after taking written informed consent from antenatal opds of the same hospitals. females with following known conditions were excluded from the study; eclampsia, cardiovascular disorder, diabetes mellitus, renal failure, liver disease, multiple gestation, polyhydramnios and urinary tract infections. fasting blood samples were collected, centrifuged and stored for serum analysis. five ml of blood was taken from median cubital vein under complete aseptic conditions. edta-plasma was used for estimation of homocysteine enzyme immunoassay (eia). samples were put in the ice packs before centrifugation. edta plasma samples were centrifuged or put on ice immediately after drawing. samples were kept on ice for up to six hours prior to separation by centrifugation. serum was separated by centrifuging at 3000 rpm for 15 minutes and then stored at 2-8º c till further analysis. hyperhomocystenemia is an independent variable in this study while pre-eclampsia is a dependant variable. hyperhomocystenemia is defined as plasma level of homocysteine ≥ 15 micro moles/l through elisa kit. age and gestational age were found to be confounding the relationship so they were matched among cases and control through pair matching. statistical analysis: data was collected through questionnaire, which was entered in to computer using spss 20.0. for quantitative variables mean and standard deviation were calculated and for qualitative variables frequency and percentages were calculated. independent sample t test was used to find out mean difference of homocysteine levels in cases and controls. chi-square test of significance was applied to find out association between categorical variables. the pvalue, 95% confidence interval and odds ratio was calculated to measure strength of association. the study was carried out as per helsinki declaration of human rights protection. scientific and ethical review of this research was done by department of community medicine cmh lahore medical college. 47 association of maternal hyperhomocysteinemia with preeclampsia: a hospital based case-control study results the mean age in cases and controls was 26.76 ± 5.82 years and 25.39 ± 4.69 years respectively. there was no statistically significant difference in mean age in both groups which showed that the age matching was appropriately done. the mean gestational period was 32.20 ± 4.12 weeks in cases and 30.76 ± 4.96 weeks in controls with no significant difference again due to appropriate matching of gestational age among cases and controls. study groups pvalue cases controls educational status illiterate 11(16.7%) 10(15.2%) 0.019 primary 34(51.5%) 19(28.8%) matric 18(27.3%) 26(39.4%) intermediate 3(4.5%) 6(9.1%) graduation and above 0(0%) 5(7.6%) monthly income (rs) <15000 51(77.3%) 46(69.7%) 0.324 ≥ 15000 15(22.7%) 20(30.3%) occupation working 1(1.5%) 4(6.1%) 0.171 house wife 65(98.5%) 62(93.9%) table-1: comparison of educational status, income and occupation among study population (n=132) among cases 51.5% had primary education, and among controls 39.4% females were matriculated. significant difference was seen in cases and controls as regards education level with low education level in cases (p=0.019). a total of 77.3% cases had monthly of income <15000 rupees while among controls 69.7% females had a monthly income of <15000 rupees. majority of cases (98.5%) and controls 93.9% were house wives. no significant difference was observed among cases and controls neither with regard to income nor with occupation status, (p-value = 0.324 and 0.171 respectively), (table-1). study groups mean s.d min. max. pvalue plasma homocysteine level case (n=66) 16.05 5.25 7.90 28.00 <0.001 control (n=66) 9.44 2.83 6.00 17.90 total (n=132) 12.74 5.35 6.00 28.00 table-2: comparison of mean plasma homocysteine level in cases and controls (n = 132) the mean plasma homocysteine was significantly higher in cases (16.05 ± 5.25) as compared to controls (9.44 ± 2.83), p-value = 0.001) (table-2). *hhcy cases (preeclamptic) n=66 controls (normotensive) n=66 chisquare value pvalue or 95 % c.i yes 34 (52%) 9 (14%) 21.56 0.001 6.72 2.86_ 15.79 no 32 (48%) 57 (86%) (*hhcy= hyperhomocysteinemia) table-3: association between preeclampsia and hyperhomocysteinemia out of 66 cases 52% had hhcy and 48% had normal plasma homocysteine level while 14% controls had hhcy and 86% were with normal plasma homocysteine level and this difference was statistically highly significant (p = 0.001, odds ratio = 6.72.) (table-3). the odds of presence of elevated homocysteine levels are 6.72 times higher in preeclamptic female cases as compared to normotensive female controls. variable category study group p-value or β p value aor c-i cases controls 0.001 6.72 1.55 0.001 4.73 1.86-11.85 *hhcy yes 34(79%) 9(20%) no 32(36% 57(64%) age(y) ≥30 15(68%) 7(32%) 0.062 2.47 0.001 0.999 0.99 0.32-3.20 <30 51(46%) 59(54%) bmi ≥25 46(63%) 27(37%) 0.001 3.32 0.92 0.021 2.66 1.16-6-10 <25 20(34%) 39(66%) *edu. status illiterate 11(52%) 10(48%) 0.812 1.12 0.12 0.820 1.13 0.38-03.31 literate 55(49%) 56(51%) table-4: logistic regression model to determine association between hyperhomocysteinemia and preeclampsia (n =132). *hhcy=hyperhomocysteinemia. *edu.status= education status after controlling for age, education level and bmi, risk of developing preeclampsia is found to be 4.73 times higher in the subjects with hyperhomocysteinemia as compared to controls (pvalue= 0.001) (table-4), which shows that hyperhomocysteinemia is independent risk factor for preeclampsia. 48 association of maternal hyperhomocysteinemia with preeclampsia: a hospital based case-control study discussion this research provides insight into strong association between hyperhomocysteinemia and syndrome of preeclampsia as seen in the current study. in this study, there was no difference in mean age and gestational age as they were matched and known hypertensives were excluded because evidence suggests that plasma homocysteine is positively associated with age as well as severe systolic hypertension.19 results showed mean plasma homocysteine level significantly higher in cases (16.05 ± 2.25) as compared to controls (9.44 ± 2.83) with an unadjusted odds ratio of 6.72 and adjusted or 4.73. the strength of association in our study is 4.73 times more in cases than in controls, which is quite high than found in previous literature. similar relationship was found in many recent and old case control studies15,18 evaluating correlation and association of homocysteine levels in preeclamptics comparing with normotensives. in another study, raised total homocysteine serum levels were seen in women with severe preeclampsia as compared to mild preeclamptic females (17.40±2.7 vs 11.49±1.19 μmol/l, p=0.001) and normal group (17.40±2.7 vs 6.38±0.3 μmol/l. (11.49±1.19 vs 6.38±0.3).2 another case-control study, explored the association of hcy in 3 groups of normotensive, pre-eclamptic and eclamptic women showing hcy levels to be significantly increased in both case groups as compared to controls.11 another netherland based case-control study concluded threefold increased risk of developing preeclampsia with raised hcy.20 yet another study showed significantly higher serum homocysteine levels in women with severe preeclampsia than those with moderate and mild stage (p = 0.025).14 the higher is homocysteine blood level more is the severity of preeclampsia.15 supplementation of pregnant women with vitamin b6 and folic acid lowered homocysteine level and also helped to control preeclampsia and improve fetal growth suggested in an interventional study.21 another study conducted in south india found out significant percentage of eclamptic patients had hyperhomocystenemia.22 very strong association of higher homocysteine levels was shown to exist in etiology of preeclampsia as well as statistically significant association with severity of preeclampsia.13,14 recent retrospective cohort study also supports the evidence that high homocysteine concentration in the first trimester serves as an independent risk factor for development of severe preeclampsia but not a good marker for the gestational hypertension and mild preeclampsia.23 a systematic review published in 2005, based on previous researches concluded that there is lack of consistency of association, dose-response effect and biological plausibility of homocysteine in etiology of preeclampsia.24 the major cause of it is largely lack of consistency in data, particularly in our local literature.21 now a recent meta analysis published in 2019 to identify causal association between homocysteine and hypertension of 40 173 individuals using mendelian randomization concluded that there is a clear evidence on causal association between homocysteine concentration and the risk of developing hypertension.25 it is well evident from above discussion that many cited case control, cohort and experimental studies are supporting the link between hcy and preeclampsia and the relationship between the preeclampsia but still more quality research is needed focusing on doseresponse relationship and biological plausibility to suggest causal link. a number of diagnostic tests for biomarkers of preeclampsia are under exploration and many show restricted sensitivity and specificity. there is no single test which can accurately predict preeclampsia. most of these tests have poor positive predictive values so a predictive model by identifying a number of factors and their effect can be helpful.14 raised plasma hcy levels are linked to an unhealthy lifestyle and faulty diet. hcy levels may be improved by modification in lifestyle and dietary habits. the best approach is by educating people about the risks of raised hcy levels, and empowering them to have a healthy lifestyle and diet. a healthy dietary pattern not only will lower hcy levels but, will more notably boost health. current study has the limitation of not considering other potential links/ associations like with folate and b12 and the fact that case control studies can’t rule out the effect of residual confounding. in south asian context, specifically in pakistan, we can suggest more studies to see the racial differences in homocysteine levels among general population as well as antenatal women. conclusion study results conclude that higher antenatal blood concentration of homocysteine (≥ micromole/l) is found to be five times higher in pre-eclamptic women as compared to normotensive pregnant women. 49 association of maternal hyperhomocysteinemia with preeclampsia: a hospital based case-control study references 1. organization wh. the world health report: 2005: make every mother and child count: world health organization; 2005. 2. osungbade ko, ige ok. public health perspectives of preeclampsia in developing countries: implication for health system strengthening. journal of pregnancy. 2011;2011. 3. bilano vl, ota e, ganchimeg t, mori r, souza jp. risk factors of pre-eclampsia/eclampsia and its adverse outcomes in low-and middle-income countries: a who secondary analysis. plos one. 2014;9(3):e91198. 4. shah n, khan nh. third delay of maternal mortality in a tertiary hospital. rawal med j. 2007; 32(supplement 2):163-7. 5. phipps e, prasanna d, brima w, jim b. preeclampsia: updates in pathogenesis, definitions, and guidelines. clinical journal of the american society of nephrology. 2016; 11(6):1102-13. 6. white wm, turner st, bailey kr, mosley jr th, kardia sl, wiste hj, et al. hypertension in pregnancy is associated with elevated homocysteine levels later in life. american journal of obstetrics and gynecology. 2013; 209(5):454. e1-e7. 7. laskowska m, oleszczuk j. homocysteine in pregnancies complicated by preeclampsia with and without iugr: a comparison with normotensive pregnant women with isolated iugr and healthy pregnant women. open journal of obstetrics and gynecology. 2011; 1(4):191-6. 8. shamsi u, saleem s, nishter n, ameen a. epidemiology and risk factors of preeclampsia; an overview of observational studies. al ameen j med sci. 2013; 6(4):292-300. 9. ahmad s, nazli r, lutfullah g. frequency of eclampsia and maternal complications in a tertiary care facility of peshawar. pak j med res. 2008; 47(4):79-82. 10. noto r, neri s, noto z, cilio d, abate g, noto p, et al. hyperhomocysteinemia in preeclampsia is associated to higher risk pressure profiles. european review for medical and pharmacological sciences. 2003; 7:81-7. 11. hasanzadeh m, ayatollahi h, farzadnia m, ayati s, khoob mk. elevated plasma total homocysteine in preeclampsia. saudi medical journal. 2008; 29(6):875. 12. palei ac, spradley ft, warrington jp, george em, granger jp. pathophysiology of hypertension in pre‐eclampsia: a lesson in integrative physiology. acta physiologica. 2013; 208(3):224-33. 13. miglani s, nautiyal r, prakash a. hyperhomocysteinemia in pre-eclampsia: is routine screening rational? international journal of reproduction, contraception, obstetrics and gynecology. 2017; 6(4):1272. 14. townsend r, o’brien p, khalil a. current best practice in the management of hypertensive disorders in pregnancy. integrated blood pressure control. 2016; 9:79. 15. serrano nc, quintero-lesmes dc, becerrabayona s, guio e, beltran m, paez mc, et al. association of pre-eclampsia risk with maternal levels of folate, homocysteine and vitamin b12 in colombia: a case-control study. plos one. 2018; 13(12):e0208137. 16. maged am, saad h, meshaal h, salah e, abdelaziz s, omran e, et al. maternal serum homocysteine and uterine artery doppler as predictors of preeclampsia and poor placentation. arch gynecol obstet. 2017; 296(3):475-82. 17. nwogu cm, okunade ks, adenekan ma, sekumade ai, john-olabode s, oluwole aa. association between maternal serum homocysteine concentrations in early pregnancy and adverse pregnancy outcomes. ann afr med. 2020; 19(2):113-8. 18. ghike s, jain s, kumare b, gupta m, shembekar c. a study of serum homocysteine levels during normal pregnancy and preeclampsia. jsafog. 2011; 3(2):71-4. 19. onyemelukwe ou, maiha bb. prevalence of hyperhomocysteinaemia, selected determinants and relation to hypertension severity in northern-nigerian hypertensives: the abu homocysteine survey. ghana med j. 2020; 54(1):17-29. 20. steegers-theunissen rp, van iersel ca, peer pg, nelen wl, steegers ea. hyperhomocysteinemia, pregnancy complications, and the timing of investigation. obstetrics & gynecology. 2004; 104(2):336-43. 21. qureshi sb, ahmad m, qureshi pma, memon a, qazi ra. hyperhomocysteinaemia, vascular related pregnancy complications and the response to vitamin supplementation in pregnant women of pakistan. jpma the journal of the pakistan medical association. 2010; 60(9):741. 22. arun m, gopinath m, nirmala c. prevalence of hyperhomocysteinemia among preeclampsia patients. j med sci clin res.2017;5(04):21063-9 23. sun f, qian w, zhang c, fan j-x, huang h-f. correlation of maternal serum homocysteine in 50 association of maternal hyperhomocysteinemia with preeclampsia: a hospital based case-control study the first trimester with the development of gestational hypertension and preeclampsia. medical science monitor: international medical journal of experimental and clinical research. 2017; 23:5396. 24. mignini le, latthe pm, villar j, kilby md, carroli g, khan ks. mapping the theories of preeclampsia: the role of homocysteine. obstetrics & gynecology. 2005; 105(2):411-25. 25. fu l, li yn, luo d, deng s, wu b, hu yq. evidence on the causal link between homocysteine and hypertension from a metaanalysis of 40 173 individuals implementing mendelian randomization.2019;21(12):1879-94. the authors: dr. zoofishan qureshi assistant professor, department of community medicine, cmh lahore medical college, lahore. prof. muhammad ashraf chaudhry head, department of community medicine, cmh lahore medical college, lahore. prof. seema imdad department of nutrition and dietetics, institute of public health, lahore. dr. tariq mukhtar farani assistant professor, department of surgery, cmh, lahore. prof. ayesha humayun hod public health & community medicine, shaikh zayed medical complex, lahore. corresponding author: dr. zoofishan qureshi assistant professor, department of community medicine, cmh lahore medical college, lahore. e-mail: zimran195@gmail.com untitled-1.indd 36 proceedings s.z.m.c. vol: 34(2): pp. 36-40, 2020. pszmc-750-34-2-2020 small group discussion; medical students perspective 1lubna humayun, 1asma rasheed, 2muhammad afaaq agha 1department of pathology, university college of medicine & dentistry, university of lahore 2department of forensic medicine & toxicology, shaikh zayed medical complex, lahore abstract introduction: small group discussion teaching is a key instructional strategy being incorporated in medical education and is aimed at enhancing the personal, social and cognitive skills of students. learning in small groups helps the students to enhance the acquisition, processing and retention of the medical knowledge. aims & objectives: to assess medical student’s perception about small group discussion at a medical college. place and duration of study: september 2019 at the university college of medicine and dentistry, university of lahore. material & methods: this crosssectional survey was conducted during september 2019 at university college of medicine and dentistry, university of lahore. convenient sampling technique was used to obtain a sample of 322 students from the first three years of mbbs. a pre validated questionnaire was distributed and the students were asked to record their experience about small group discussion using a 5point likert scale. data was analyzed by using spss22 and reported as percentage distribution and mean± s.d. results: regarding quality of these small group discussion sessions, 25.8% of the students appeared satisfied, 24.8% of the students were neutral, whereas, 49.4% of the students remained unsatisfied. when asked about being clear of their role in a sgd session, 55.9% of the students agreed, 22.7% were unclear and 21.4% neutral. as regards the benefits from small group sessions, majority (35.1%) agreed that activities taught lifelong learning, 8.7% strongly agreed. 27% remained neutral with 16.5% disagreeing and 12.75% strongly disagreeing. with reference to assessment of tutors, majority of students found sgd facilitators enthusiastic about process with 13.4% strongly agreeing and 36.3% agreeing, 28% stayed neutral while 9.6% disagreed and 12.75% strongly disagreed. majority of students, (35.1% agreed, 13.4% strongly agreed) to that tutors just gave mini lecture, 27.6% responded neutral to statement while 11.2% disagreeing and 12.7% strongly disagreeing to the enquiry. conclusion: the study reveals that majority of the students are clear about their role in small group discussion sessions and majority (35.1%) also agreed that activities taught them lifelong learning. tutors are helpful and enthusiastic, however, they provide plenty of information and need retraining for conducting a small group learning session. key words: small group discussion, students’ perspective, teaching, learning. introduction traditionally, the undergraduate students of mbbs are divided into small groups to carry out the practical, tutorial and the ward clerkship learning. these small groups function either individually or under the supervision of a facilitator, and usually consist of 8-12 members. in small-group discussion the teacher announces a topic or idea for group discussion among the participants and they conclude the discussion usually in that session.1 in the recent years, a number of inadequacies have been recognized in the traditional teaching methods and this has led to major reforms and innovations in the medical education both in the fields of curriculum development as well as teaching strategies. medical educationists have come to a broad consensus for a need of a shift from the traditional didactic lecturing to the more studentcentered delivery of curriculum.2 research has demonstrated that small group discussion results in greater synthesis and retention of learning materials and the sgd sessions have changed the role of a teacher from a lecturer to a facilitator responsible for the active learning by the students.3,4,5 small group teaching method develops the intellectual skills of the learners such as reasoning, problem solving, the development of attitudes and the acquisition of interpersonal skills such as listening, speaking, arguing and group leadership.6 sgds can take on a variety of different forms such as problem-based learning, case-based learning, role play, discussions, brainstorming sessions and debate sessions.6 pbl is a variation of small group discussion strategy where the students define the problem and identify the action steps to create a solution for that problem. it extends over two 37 small group discussion; medical students perspective sessions.7 despite the increased use of small group discussion in medical education, relatively little is known about how our students feel about this strategy.8 although, small group teaching holds various advantages but it also poses certain limitations. teaching in sgds can be costly because it requires a higher teacher-student ratio. some teachers find sgd teaching relatively difficult as compared to delivering a long lecture.9 in the recent years there has been a shift of curriculum in most of the medical universities from the ‘traditional’ towards the ‘integrated curriculum’ and various new modes of information transfer to students have been introduced. out of these, the small group discussion is the fundamental teaching strategy being incorporated in these medical universities in various ways such as pbl, tbl and cbl sessions.10 this is recommended strategy to develop competencies of the students.11 with this increase in the use of small group discussion, there is a growing need for research on this strategy. this study is significant as it provides information about the student’s perception on the small group discussion sessions. material and methods this crosssectional survey was conducted during september 2019 at university college of medicine and dentistry, university of lahore. the targeted population was the 1st year, 2nd year and 3rd year students of mbbs. the students of the first three years of mbbs who are mainly using sgd as the learning strategy were targeted and the total number of students was 322. inclusion criteria: 1st year, 2nd year and 3rd year students of mbbs students of either gender from the academic year 2018-19. exclusion criteria: students of bds, post graduate students, those students not willing to participate, incomplete questionnaires were excluded. ethical approval: the study was approved by the ethical review board (erb) university college of medicine & dentistry, under the registration number ref: erc/09/19/04. it was conducted among a total of 322 students of the first three years of mbbs who are mainly using sgd as the learning strategy. data collection technique: this study was based on pre-validated questionnaire on experience of medical students regarding sgl. the questionnaire that was used contained 19closed ended questions that addressed the issues related to small group sessions in the integrated curriculum. this questionnaire is an established tool designed by the medical education department at the king saud medical school, and this proforma is available online for general use. questionnaire was reviewed by the current research supervisor and was piloted on twenty 2nd year mbbs students. the questionnaire was divided into four subscales and had total 19 items that included closed ended questions that addressed the issues of concern regarding the small group sessions. the questions assessed the structure of the small group learning sessions, the degree of student satisfaction during the process of learning, the assessment of tutors, and the usefulness and the quality of the small group sessions. students’ responses were quantitatively measured in relation to statements on the questionnaire using a five point likert scale. strongly disagree (1), disagree (2), neutral (3), agree (4) and strongly agree (5) the questionnaire was distributed among the participants after filling a consent form and sufficient time was given to each student to properly comprehend and fill it. the students were not required to disclose their names and roll numbers and were assured about the confidentiality of the information they provided in the questionnaire. statistical analysis: the data was collected by the 19-item questionnaire and the analysis was done on spss 22. data was reported as percentage distribution and standard deviation. results the results obtained from the study are compiled in table-1. when asked whether sgl session objectives were made clear, majority of students (65.2%) agreed to that, out of this total percentage 26.4% strongly agreed while 38.8% agreed. 20.8% students remained neutral while only 13.9% disagreed. when asked, if role was made clear to students in sgl session, 42.9% agreeing while 13% strongly agreeing to statement. 21.4% remained neutral while 12.1% disagreed and 10.6% strongly disagreeing. when asked about suitability of location and facilities to conduct sgl session, 26.4% remained neutral with 12.7% disagreeing and 13.4% strongly disagreeing. major percentage (30.75%) of students agreeing while 16.8% strongly agreeing to the statement. when enquired about their satisfaction levels of small group sessions’ progress, 34.5% students agreed that sessions were thought provoking, 16.1% strongly agreed while 27.6% remained neutral. 13% students disagreed while only 8.7% strongly 38 small group discussion; medical students perspective disagreed to this inquiry. 32.9% students agreed that sessions led them to active learning, 18.9% strongly agreed while 25.8% preferred to remain neutral. 13.4% thought sessions did not lead to active learning while only 9% strongly disagreed to this enquiry. when asked if sessions led to deep learning, 31.4% remained neutral, 25.5% agreed and 14.9% strongly agreed while 18.9% disagreed and only 9.3% strongly disagreed. regarding enquiry about sessions helping students to identifying learning needs, only 9% strongly disagreeing and 14.6% just disagreeing while 24.2% preferred to stay neutral. majority agreed to the statement with 17.4% strongly agreeing while 34.8% just agreeing to it. for 31.4% students agreed, that the small group sessions were easy to look for learning needs while 15.5% strongly agreed. 11.8% strongly disagreed, 11.5% just disagreeing while 29.8% stayed neutral in this regard. with regard to assessment of tutors, majority of students found sgd facilitators enthusiastic about process with 13.4% strongly agreeing and 36.3% agreeing. 28% stayed neutral while 9.6% disagreed and 12.75% strongly disagreed. clear majority (33.9%) remained neutral when asked if tutors provided lots of information while 14% thought they did not provide such information and 10.9% strongly thought against it. 13.7% students strongly agreed that tutors provided lots of information while 27.65 agreed to it. majority of students, (35.1% agreed, 13.4% strongly agreed to that tutors just gave mini lecture, 27.6% responded neutral to statement while 11.2% disagreeing and 12.7% strongly disagreeing to the enquiry. when asked whether tutors talked a lot in some sessions, 10.9% disagreed absolutely, 14% just disagreed while 28.3% remained neutral. 22.7% agreed to statement while 16.8% strongly agreed to the statement. when asked about benefits from small group sessions, majority (35.1%) agreed that activities taught them lifelong learning and 8.7% strongly agreed. 27% remained neutral with 16.5% disagreeing while 12.75% strongly disagreeing to this query. 32% students found sessions important and thought they will be of use in future and 16.8% strongly agreed. small percentage i.e., 9.95% disagreed, 10.2% strongly disagreed to it while significant 31.1% preferred to comment neutral. 37% agreed and 11.8% strongly agreed that these activities helped them to improve ability to think and solve problems rather than just memorizing information, where as 13.7% disagreed and 8.1% disagreed with 29.5% staying neutral about this question. 41.6% students agreed, while 12.1% strongly agreed that these activities helped them to develop skills in working as member of a team. 11.2% disagreed, 9.3% strongly disagreed while 25.8% students commented neutral in this respect. majority of students i.e., 36.5% agreeing, 16.1% strongly agreeing to statement that these activities improved their abilities to communicate effectively, 26.1% stayed neutral while 13% disagreed and 8.7% strongly disagreed. when enquired about sessions improving leadership skills, 16.8% disagreed, 10.2% strongly disagreed and 28% opted to stay neutral. 28.9% agreed while 16.1% strongly agreed that sessions improved their leadership skills. 36% students agreed while 13.4% strongly agreed that they were satisfied with quality of these sessions with 12.1% disagreeing, 13.7% strongly disagreeing to this statement. 24.8% students stayed neutral in this regard. items strongly disagree disagree neutral agree strongly agree structure of small group discussion sessions 1. the sgd session objectives were made clear to me 22 (6.8%) 23 (7.1%) 67 (20.8%) 125 (38.8%) 85 (26.4%) 2. my role was made clear to me in the sgd session 34 (10.6%) 39 (12.1%) 69 (21.4%) 138 (42.9%) 42 (13%) 3. the location and facilities were suitable for the conduction of sessions 43 (13.4%) 41 (12.7%) 85 (26.4%) 99 (30.75% 54 (16.8%) satisfaction of small group discussion sessions’ progress 4. the sessions were thought provoking 28 (8.7%) 42 (13%) 89 (27.6%) 111 (34.5%) 52 (16.1%) 5. the sessions led me to active learning 29 (9%) 43 (13.4%) 83 (25.8%) 106 (32.9%) 61 (18.9%) 6. the sessions led me to deep learning process 30 (9.3%) 61 (18.9%) 101 (31.4%) 82 (25.5%) 48 (14.9%) 7. the sessions led me to identify my learning needs 29 (9%) 47 (14.6%) 78 (24.2%) 112 (34.8%) 56 (17.4%) 8. it was easy to look for the learning needs 38 (11.8%) 37 (11.5%) 96 (29.8%) 101 (31.4%) 50 (15.5%) assessment of tutors 9. sgd facilitators were enthusiastic about the process 41 (12.75%) 31 (9.6%) 90 (28%) 117 (36.3%) 43 (13.4%) 10. the tutors provided us lots of information 35 (10.9%) 45 (14%) 109 (33.9%) 89 (27.6%) 44 (13.7%) 11. the tutors gave us a minilecture 41 (12.7%) 36 (11.2%) 89 (27.6%) 113 (35.1%) 43 (13.4%) 12. the tutors were talking a lot in some of the sessions 35 (10.9%) 69 (21.4%) 91 (28.3%) 73 (22.7%) 54 (16.8%) benefits from small group discussion sessions 13. the activities taught me lifelong learning 41 (12.75%) 53 (16.5%) 87 (27%) 113 (35.1%) 28 (8.7%) 14. what i learned in these sessions was important and will be of use to me in the future 33 (10.2%) 32 (9.95%) 100 (31.1%) 103 (32%) 54 (16.8%) 15. these activities helped me improve my ability to think and solve problems rather than just memorizing information 26 (8.1%) 44 (13.7%) 95 (29.5%) 119 (37%) 38 (11.8%) 16. these activities helped me to develop skills in working as a member of a team 30 (9.3%) 36 (11.2%) 83 (25.8%) 134 (41.6%) 39 (12.1%) 17. these activities improved my ability to communicate effectively 28 (8.7%) 42 (13%) 84 (26.1%) 116 (36.5%) 52 (16.1%) 39 small group discussion; medical students perspective 18. these sessions improve my leadership skills 33 (10.2%) 54 (16.8%) 90 (28%) 93 (28.9%) 52 (16.1%) 19. overall, i was satisfied with the quality of these sessions 44 (13.7%) 39 (12.1%) 80 (24.8%) 116 (36%) 43 (13.4%) mean 33.68 42.84 87.68 102.63 49.37 standard deviation 6.27 10.75 10.34 26.59 11.63 table-1: undergraduate medical student’s perception about small group discussion sessions at the university college of medicine, university of lahore. discussion according to this study the medical students found sgd a better learning methodology and are generally satisfied with the small group discussion sessions in the integrated curriculum. students have difficulty in remaining attentive during lectures and usually cannot retain the knowledge effectively in long lectures. this study found that small group discussions give a chance of active participation to all the learners and they feel more in control of the learning process. the findings are similar to those from study by pradeep et al12 in which students showed interest in the small group discussion methodology as it enhanced their intellectual skills such as reasoning, problem-solving, and critical thinking. similar studies carried out in shifa college of medicine islamabad,13 malaysia14 and the university of sharjah15 all have agreed that the students find the sgds to be the most effective strategy for learning and they are satisfied with the teacher’s role in a sgd. in contrast, the present study found 35.1% of the medical students are dissatisfied with the teachers’ role in sgd and believed that tutors gave mini-lectures in these sessions. the teachers actively participate in teaching the topic to the students and forget their passive role of a facilitator who just has to guide the learners if they go astray from the topic. it is suggested that proper and frequent training sessions and workshops should be arranged for the faculty members so that they conduct the sgd sessions in true spirit. this research will effectively contribute towards the implementation of strategies that would improve the conduction of sgd sessions. the results of this study will help to understand the student attitudes towards small group work, effectiveness of group dynamics and assessing the role of tutors in this teaching strategy. for future research, small group discussion sessions should also be assessed during all phases of medical education, from undergraduate clinical training to postgraduate residency education, it would be worthwhile to also measure the perceptions of teachers regarding small group teaching and compare the perceptions of teachers and students to gain a better understanding. moreover, it would also be valuable to measure the effectiveness of sgd in a more rigorous fashion, assessing the effects it has on a student’s educational progress and professional development.16,17 it will be better if we could repeat the study and identify the trend of student feedback about the sgds over years. conclusion the study reveals that the students found these sessions to be interactive and friendly and they bridged the gap between the teacher and student. the negative perception of the students is about the role of teachers during the sessions as they are providing too much information during the sgd. the tutors should leave it up to the learners to attain the learning objectives. the investigator strongly recommends that at institutional level this mode of learning should be used as routine method at appropriate intervals for teaching medical students. references 1. edmunds s, brown g. effective small group learning: amee guide no. 48. med teach [internet]. 2010 sep 26; 32(9):715-26. available from:http://www.tandfonline.com/doi/full/10.31 09/0142159x.2010.505454 2. wijnen m, loyens smm, smeets g, kroeze m, van der molen h. comparing problem-based learning students to students in a lecture-based curriculum: learning strategies and the relation with self-study time. eur j psychol educ. 2017 jul 1; 32(3):431-47. 3. hamann k, pollock ph, wilson bm. assessing student perceptions of the benefits of discussions in small-group, large-class, and online learning contexts. college teaching, 2012 v60 n2 p65-75. 4. silva ab da, bispo ack de a, rodriguez dg, vasquez fif. problem-based learning: a proposal for structuring pbl and its implications for learning among students in an undergraduate management degree program. rev gestão. 2018; 25(2):160-77. 5. chang bj. problem-based learning in medical school: a student’s perspective. vol. 12, annals of medicine and surgery. elsevier ltd; 2016. p. 88-9. 40 small group discussion; medical students perspective 6. annamalai n, manivel r, palanisamy r. small group discussion: students perspectives. int j appl basic med res. 2015; 5, suppl s1:18-20. 7. elaine h.j. yew, karen goh. problembased learning: an overview of its process and impact on learning. health professions education. 2016; 2(2): 75-79. 8. hameed, s., khalid, t., aslam, s., ahmad, m., farhan, f., batool, z., & hamid, s. (2017). team based learning in pathology effect on test scores and student satisfaction. pakistan armed forces medical journal, 67(3), 423-28. 9. mirza aub, khan ra, zia s, iqbal y. medical students’ perspective about factors motivating participation in small group discussions. jpma 64: 1339; 2014 10. yew ehj, goh k. problem-based learning: an overview of its process and impact on learning. heal prof educ. 2016 dec 1;2(2):75-9 11. raut dss, shreechakradhar u dm, more dsr, rathod dvs, gujar dvm, nardele d v., et al. developing competencies of medical students using group discussion as tl method. iosr j dent med sci. volume 13, issue 1 ver. iv (jan. 2014), pp 24-27. 12. sahu p, nayak s, & rodrigues v. medical students’ perceptions of small group teaching effectiveness in hybrid curriculum. journal of education and health promotion. 2018; 7(1),30. 13. ahmad rn, bhatti mm, khan ah, ghayur s, rafi s, naseem s, et al. small group discussion as a learning strategy for medical undergraduates. rawal med j. 2014; 39(3):344-348. 14. aziz n, nasir r, salam a. students’ perception of small group teaching: a cross sectional study. middle east j fam med. 2008;6(5) 37-39 15. mousa abu ghoush, mohammed abdul qadir, zaharaa al-lami, safa al-abdullah, nihar dash. undergraduate medical students’ perception about learning in small group at university of sharjah. journal of health science. 4 (2016); 207-214. 16. steinert y. student perceptions of effective small group teaching. med educ [internet]. 2004 mar; 38(3):286-93. 17. soliman mm, alnassar sa. student perception of small group teaching in first year mbbs at king saud university medical college saudi arabia. pak j med sci 2011; 27(5):963-966. the authors: dr. lubna humayun assistant professor, department of pathology, university college of medicine and dentistry, the university of lahore. dr. asma rasheed assistant professor, department of pathology, university college of medicine and dentistry, the university of lahore. dr. muhammad afaaq agha assistant professor, department of forensic medicine &toxicology, shaikh zayed medical complex, lahore. corresponding author: dr. lubna humayun assistant professor, department of pathology, university college of medicine and dentistry, the university of lahore. email: lubnahumayun155@gmail.com for web full book 8 proceedings s.z.m.c. vol: 37(3): pp. 8-14, 2023. pszmc-889-37-3-2023 intraoperative peritoneal lavage in peritonitis: normal saline vs metronidazole 1hira aslam, 2muhammad khurram jameel, 2pir muneeb, 1aneeqa nasir qureshi, 2fawad hameed, 3muhammad ammar, 2maham qazi, 4abrar ul hassan pirzada 1department of surgery, mayo hospital, kemu, lahore 2department of surgery, anmc / chaudhary muhammad akram hospital, lahore 3department of urology, chaudhary muhammad akram hospital, lahore 4department of plastic surgery, anmc / chaudhary muhammad akram hospital, lahore, pakistan abstract introduction: globally, acute generalized peritonitis ranks among the top surgical emergencies. different studies have been conducted to show the amplitude of peritonitis worldwide eliciting a huge impact on overall patient morbidity and mortality. largely peritonitis is caused by a gastrointestinal perforation or anastomotic leak. in peritonitis, anaerobes & gram-negative organisms are mostly responsible for sepsis and morbidity due to the overactive inflammatory cascade by endotoxins which is amenable to timely intervention. aims & objectives: the study's aim was to evaluate whether using normal saline or metronidazole solution during intraoperative peritoneal lavage (iopl), results in a lower rate of postoperative wound infection. place and duration of study: this study was undertaken at the south surgical ward, mayo hospital lahore for 6 months from february 2nd, 2021, to august 1st, 2021. material & methods: consecutive sampling strategy followed by a randomized controlled trial were used to induct and provide intervention to 90 patients aged 15-65 years with peritonitis caused by hollow viscus perforation. the patients were subdivided into 2 groups a& b(n=45 each).two liters of normal saline were used for peritoneal lavage in group a, while two liters of normal saline were combined with 200 ml of metronidazole solution and administered to group b. intraoperatively.baseline physiological parameters such as age, sex, bmi , intra operative surgical parameters as duration of operation and post-operative course were recorded till discharge. on 10th pod, patients returned to opd for further monitoring. an infection was diagnosed if the patient had post-operative symptoms such as a high temperature, increased tlc, wound discharge, redness, or pain. data was entered and analyzed using spss version 23. results: a majority (54.44%) of the patients were young adults. mean age of 37.33 ± 10.53 years of patients in the metronidazole group was comparable to mean age 40.04 ± 11.96 years in the saline group, difference was not significant (p=0.067). male/female ratio in metronidazole and saline groups were 17/25 and 10/18, respectively. patients who received intraperitoneal lavage with normal saline were more likely to develop wound infections (17/45) (37.78%), while only 3/45) (6.67% of those who received metronidazole solution did so (p 0.0001). conclusion: based on the results of this experiment, using metronidazole solution for intraoperative peritoneal lavage instead of normal saline reduces the occurrence of postoperative wound infection. keywords: peritonitis, postoperative wound infection, intraoperative peritoneal lavage introduction globally, acute generalized peritonitis ranks among the top surgical emergencies1. it is more common in third world nations. the prevalence of perforation is low (0.6% 4.9%) in developed nations but high (33% 63%) in west africa2. 554 persons were discovered to have peritonitis in a study that took place over three years in india3. researchers in pakistan have conducted studies with similar methods, with one study reporting 650 cases in a just 9 months4. most cases of peritonitis are caused by a gastrointestinal perforation or anastomotic leak5. in the case of peritonitis, anaerobes and gramnegative organisms are mostly responsible for sepsis and morbidity due to the overactive inflammatory cascade brought on by the release of endotoxins5.clinical evidence is used to identify peritonitis. diagnosis can be achieved via upright plain x-ray of the abdomen, usg, or ct scan. this is often done through diagnostic laparoscopy nowadays6.resuscitation, diagnosis, prompt exploration, treatment of the underlying cause, and extensive surgical peritoneal lavage have always been the cornerstones of peritonitis therapy regimens (iopl)7,8. regular iopl is performed to lessen bacterial contamination and burden. even though large volumes of normal saline are used in iopl, the rates of sepsis, wound infection, and mortality remain alarmingly high. another method 6 1department of public health, health services academy, islamabad. 2department of management sciences, riphah int university, islamabad. 3department of medicine, ihitc, islamabad. 4department of rheumatology, pims, islamabad. 5department of medicine, dow university of health sciences, karachi. 70 1department of orthopedics, thq, khanewal 2department of orthopedics, lgh, lahore 3department of orthopedics, thq, ferozwala 4department of orthopedics, cmh, lahore 5department of hand & upper limb surgery, cmh, lahore 6department of orthopedics, al-rehmat benevolent trust 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 9 intraoperative peritoneal lavage in peritonitis: normal saline vs metronidazole of reducing bacterial count and the associated risk of bacterial contamination and surgical site infection is intra-abdominal antibiotic lavage9.antibiotic solutions like ampicillin, doxycycline, lincomycin, gentamycin, and many others have all been employed before for this function10. metronidazole a synthetic antimicrobial drug has been infrequently used. working through diffusion of its nitro group into bacterial and protozoal cells it destabilizes dna and electron transport chains to produce a bactericidal and antiprotozoal effect. this mechanism of action sets it apart as an effective weapon against anaerobes11. metronidazole and sterile water solution is used in iopl. this will specifically target anaerobes which alone normal saline cannot do. indian research into postoperative wound infections showed a significantly reduction in these employing metronidazole solution as iopl. lavage with normal saline resulted in a 21% infection rate, while metronidazole solution resulted in a 15% infection rate7. another study indicated that whereas 42% of wounds became infected after being lavaged with regular saline, only 30% did so after being treated with a metronidazole solution. also, the difference was not statistically significant (p>0.05)7. even though normal saline has been used for iopl for a long time, post-operative wound infection is still a problem .the use of antimicrbial and antibiotic lavage, has thus shown conflicting scientific data regarding reduction of postoperative wound infection rates. patients presenting within 72 hours of developing peritonitis will be the focus of this study, as opposed to all peritonitis patients in previous research. as a result of this research, our community will have local data for the use of metronidazole solution as iopl, which will decrease chances of sepsis and wound infection and the length of time patients spend in the hospital in our setup. material and methods the study's goal was to evaluate the efficacy of intraoperative peritoneal lavage (iopl) with either normal saline or metronidazole solution in reducing wound infection. the investigation employed a randomized controlled trial using consecutive sampling as its study strategy. this study was executed in the south surgical ward of the surgery department of mayo hospital in lahore. the time frame for this study was from february 2nd, 2021, to august 1st, 2021. the sample size of this study was determined by calculating the expected proportion of wound infection in the normal saline group at 6.53% and in the metronidazole group at 26.3%, using a significance threshold of 5% and 80% power of test, respectively. this yielded a total of 90 patients (45 in each group). patients between the ages of 15 and 65 years of both sexes, and those diagnosed with peritonitis related to hollow viscous perforation (from trauma or any other cause) met the inclusion criteria used in the sample strategy. patients with primary or spontaneous peritonitis (based on clinical examination and history), and patients with peritonitis related to solid organ injury (on clinical examination) were not eligible for this study. ninety (90) patients meeting the aforementioned criteria were enrolled after obtaining ethics board approval vide 756 /rc/kemu dated 4/1/2021 for the data collection process. before the operation, everyone gave their informed consent. characteristics such as age, gender, body mass index, and time since peritonitis diagnosis were considered. patients were equally divided into two equal groups by random lottery method. two (2) liters of normal saline were used for peritoneal lavage in group a, while two liters of normal saline were combined with 200 ml of metronidazole solution for group b. using general anesthesia; a single surgical team performed all procedures with the help of the assisting researcher. all the time spent during operation was recorded. patients were monitored in the operating rooms and released from the facility after a period of 72 hours following surgery. after 10 days, patients returned to opd for further monitoring. patients were diagnosed with wound infection if they experienced post-operative complications such as fever, increased tlc, wound discharge, redness, and pain (as per operational definition). proformas were used to record all of this data. infections that developed in patients' surgical wounds after surgery were treated according to accepted medical practice. statistical analysis: all the information was entered into spss 23. age, body mass index, length of peritonitis, and surgical time were some of the quantitative factors provided as means +/standard deviations. qualitative factors such as gender and postoperative wound infection were given in the form of frequency and percentage. the chi-square test was used to examine the difference in rates of infection after surgery between the two groups. the cutoff for significance was set at a 0.05 probability level. the information was separated into groups based on variables such as age, gender, body mass index, length of peritonitis, and length of surgery. after the groups were divided into strata, the chi-square test was used to compare them based on the presence of postoperative wound 10 intraoperative peritoneal lavage in peritonitis: normal saline vs metronidazole infection. the cutoff for significance was a p value of 0.05. results patients included in the study were between 15 to 65 years with overall mean age of 38.67 ± 12.31 years.the mean age of group a was 40.04 ± 11.96 years and 37.33 ± 10.53 years in group b. out of 90 patients 32 (35.56%) were males and 58 (64.44%) were females with male to female ratio of 1:1.8. mean duration of peritonitis was 42.24 ± 12.01 hours. in table-1 mean bmi was 28.48 ± 2.92 kg/m2. mean operative time being 62.34 ± 11.56 minutes. frequency of postoperative wound infection with normal saline was found in 17/45 (37.78%) and in metronidazole solution in intraoperative peritoneal lavage was found to be 03/45 (6.67%) patients (pvalue = 0.0001). age di str i b uti on for both gr oup s (n=90) age group a (n=45) group b (n=45) total (n=90) no. of patie nts %age no. of patient s %age no. of patie nts %age 15-40 24 53.33 25 55.56 49 54.44 41-65 21 46.67 20 44.44 41 45.56 mea n ± sd 40.04 ± 11.96 37.33 ± 10. 53 38.67 ± 12. 31 di str i b uti on of pati e nt s ac c or di ng to dur ati o n of pe r i toni ti s durat ion (hr’s) group a (n=45) group b (n=45) total (n=90) no. of pati ent s %age no. of patient s %a ge no. of pati ents %age 36 19 42.22 18 40.0 37 41.11 >36 26 57.78 27 60.0 53 58.89 mean ± sd 41.40 ± 12. 21 43.44 ± 11. 28 42.24 ± 12. 01 di str i b uti on of pati e nt s ac c or di ng to bm i bmi (kg/ m 2) group a (n=45) group b (n=45) total (n=90) no. of pati ent s %age no. of patie nts %age no. of pati ents %age 27 15 33.33 17 37.78 32 35.56 >27 30 66.67 28 62.22 58 64.44 mean ± sd 28.98 ± 3.21 28.04 ± 2.67 28.48 ± 2.92 di str i b uti on of pati e nt s ac c or di ng to ope r ati ve ti me opera tion (min) group a (n=45) group b (n=45) total (n=90) no. of patie %age no. of patient s %age no. of patie %age nts nts 60 16 35.56 18 40.0 34 37.78 >60 29 64.44 27 60.0 56 62.22 mean ± sd 63.84 ± 11. 94 61.89 ± 11. 39 62.34 ± 11. 56 table-1: different parameters of patients and their results table-2 shows the ratio of wound infection with respect to various parameters. the stratification of postoperative wound infection with respect to age groups has significant p value (0.002) in group a patients with younger population between 15-40 years.similarly statistically significant difference was found in postoperative wound infection among both genders having female predominance in group a. postoperative wound infection according to duration of peritonitis is more common among 36 hours of group a, while it is nil in group b.the impact of higher bmi ( 27) on postoperative wound infection in group a is statistically important with significant p value (0.0001), while it is nil in metronidazole group (b). lastly, the postoperative wound infection again was significantly reduced in patients of metronidazole group irrespective of duration of surgery with p value (0.005& 0.018). in fig-1 comparison of the frequency of postoperative wound infection with normal saline versus metronidazole solution in intraoperative peritoneal lavage is shown. fig-1: comparison of the frequency of postoperative wound infection with normalsaline (a) versus metronidazole solution (b) in iopl 0 5 10 15 20 25 30 35 40 45 group a group b 17 3 28 42 po st -o pe ra tiv e w ou nd in fe ct io n 11 intraoperative peritoneal lavage in peritonitis: normal saline vs metronidazole ound infection with res pect to age age of patient (yrs) group a (n=45) group b (n=45) p-value postoperative wound infection postoperative wound infection yes no yes no 15-40 12 13 02 22 0.002 41-65 05 15 01 20 0.067 wound infection with res pect to gender gender group a (n=45) group b (n=45) p-value postoperative wound infection postoperative wound infection yes no yes no male 05 10 00 17 0.010 fem ale 12 18 03 25 0.011 wound infection with res pect to duration duration (hrs ) group a (n=45) group b (n=45) p-value postoperative wound infection postoperative wound infection yes no yes no 36 04 14 03 16 0.618 >36 13 14 00 26 0.022 wound infection with res pect to bmi bmi (kg/m 2) group a (n=45) group b (n=45) p-value postoperative wound infection postoperative wound infection yes no yes no 27 04 13 03 12 0.810 >27 13 15 00 30 0.0001 wound infection with res pect to operative tim e operativ e tim e (min) group a (n=45) group b (n=45) p-value postoperative wound infection postoperative wound infection yes no yes no 60 07 11 00 16 0.005 >60 10 17 03 26 0.018 table-2: ratio of wound infection with respect to various stratified parameters of patients. discussion acute widespread peritonitis is a surgical emergency that poses significant treatment challenges. it is crucial to identify and treat sepsis as soon as possible so that the underlying cause can be addressed14.in the past, since laparotomy on purpose had not yet been achieved in the operating room; early treatment of peritonitis had to rely on medical interventions. rest, purgatives (particularly magnesium sulphate), fasting, lemon juice, cold therapy, and very limited opium use were all recommended for treating peritonitis15. in a later presentation, mikulicz argued in favor of abdominal opening. he also popularized the use of drainage tubes and a 2% thymol solution for "toilet of the peritoneum," in which the unclean intestines are sponged clean. in another study of tait, the solution of blood and warm water was introduced in the abdomen and all internal organs washed until the water ran clear. amniotic fluid, 25% glucose, water, saline, and different antibiotics were some of the fluids utilized in peritoneal lavage and aspiration in the past16. peritoneal lavage can lessen the number of germs in the body by dilution, which in turn decreases the risk of infection and septic shock at the wound site. saline lavage significantly decreases the numbers of aerobic and anaerobic bacteria in peritoneal fluid and provides an estimate of the debris load in the wash fluid17. this research was carried out to evaluate the efficacy of intraoperative peritoneal lavage with either normal saline or metronidazole (nitroimidazoles) solution on the incidence of wound infection following perforated hollow viscus peritonitis surgery. to compare, our study found that 17/45 (37.78%) patients who had intraoperative peritoneal lavage with normal saline developed postoperative wound infections, while only 3/45 (6.67%) received intraoperative peritoneal lavage with metronidazole solution (p 0.0001). in contrast, a trial indicated that the metronidazole lavage group had a lower rate of wound infection than the saline lavage group, the difference was statistically significant (p0.05)12. a further investigation found a 26.3 percent infection rate in the metronidazole lavage group compared to 6.5 percent in the saline lavage group (p0.05)13. harpreet singh, malika agrawal, naveen singh18 reported that incidence of 50% (21 out of 42) of wound infection when no local drug was used as compared to 30% when metronidazole was used. dalvi et al19 reported 20% reduction in incidence of wound infection with use of intraoperative topical metronidazole. similarly, khan et al20 reported 20% reduction in incidence of wound infection, when superoxide solution was used for iopl. on contrary, schein et al did not find any difference in incidence of wound infection when chloramphenicol was used for iopl21. the etiology of postoperative wound infection is complicated by the heterogeneous nature of these 12 intraoperative peritoneal lavage in peritonitis: normal saline vs metronidazole parameters which include both patient factors (age, gender, bmi) and procedural factors (duration of surgery, prior infection due to long peritonitis timing)29.reviewing published data, shows that most researchers have not reported different parameters of patients like age, gender, bmi, time of peritonitis and duration of surgery in developing postoperative wound infection. in our study however parameters like higher bmi and increasing duration of surgery had statistically proportionate p values 0.0001, .018, 0.005 respectively in metronidazole group (b).which may infer that increasing operative time and higher bmi are possibly factors for greater peritoneal infection which responds significantly better to metronidazole than normal saline. moreover, metronidazole was significantly more effective in reducing peritonitis linked postoperative wound infection between the ages of 15-40 and in all genders. increasing age above 40 however did not respond significantly better to metronidazole versus normal saline (p.067) probably due to the presence of likely comorbidities such as diabetes. an important point noted was that metronidazole was substantially more effective in eliminating prolonged peritonitis linked wound infection than normal saline (p value 0.0001) because of its specific antigram-ve bacterial action11 peritoneal lavage was found to be therapeutically effective in a study conducted by nathens and colleagues22.similarly positive findings were seen in a trial using peritoneal irrigation and local antibiotics by hau t et al. and colleagues. lavage with antibiotics for irrigation has been found in experimental tests to significantly reduce mortality in cases of peritonitis23. some studies have failed to show any advantage of peritoneal lavage over simple systemic antibiotics24. it has been proposed that peritoneal dialysis fluid is an appropriate lavage agent since it is gentler on the mesothelial cells than regular saline (0.9% sodium chloride)25. metronidazole, gentamicin, cephalosporin, lincomycin, ampicillin, kanamycin and doxycycline and the antiseptics povidine iodine and chlorhexidine have all been studied with conflicting results.26in a review of 15 studies by josie chundamala j et al, and james g et al. wright showed a definite reduction of surgical site infection (ssi) with povidine iodine when compared to saline or no irrigation26. in all above studies, we can conclude that surgical wound infections after laparotomy of peritonitis are the leading cause of morbidity, poor quality of life and increase length of hospital stay. thus creating the physically, financially and mentally impact on patient’s family27. this necessitates thinking the ways to reduce ssi after laparotomy not only in infected abdomen but also in other laparotomy for any reason. metronidazole is thus an effective drug to use in ssi against gram-positive and gramnegative anaerobic bacteria, as well as grampositive anaerobic bacteria that produce spores . it has been used both alone and in conjunction with other antibiotics in peritoneal lavage, and both methods have been shown to be safe and effective28. limitations this was a single center study having a relatively small sample size and mostly 3,4 surgeons performed all operations. future larger clinical trials at more centers are necessary for comprehensive information on the use of metronidazole for iopl in peritonitis. conclusion this study concludes that the frequency of postoperative wound infection is less using metronidazole solution in intraoperative peritoneal lavage as compared to normal saline use. so, we recommended that metronidazole solution should be used routinely in intraoperative peritoneal lavage in order to prevent the post-operative wound infection. references 1. steinbach cl, töpper c, adam t, kees mg. spectrum adequacy of antibiotic regimens for secondary peritonitis: a retrospective analysis in intermediate and intensive care unit patients. ann clin microbiolantimicrob 2015; 14(1):48. 2. shanker m, nahid m, prajwal s. a clinical study of generalised peritonitis and its management in a rural setup. int surg j 2018;5(11):3496-504. 3. ghosh ps, mukherjee r, sarkar s, halder sk, dhar d. epidemiology of secondary peritonitis: analysis of 545 cases. int j sci stud 2016;3(12):83-8. 4. khalid s, bhatti aa. audit of surgical emergency at lahore general hospital. j ayub med coll abbottabad 2015;27(1):74-7. 5. brown d, alvarado jac. sepsis (septic), peritonitis. statpearls [internet]: statpearls publishing; 2018. 6. masud m, khan a, adil m, gondal zi, aquil a, jahangeer mh, et al. etiological spectrum of perforation peritonitis. pak armed forces med j 2016;66(5):756-60. 7. singh h, agrawal m, singh nk, kaul rk, ilahi i, ahuja r. intraoperative lavage in peritonitis: comparison between saline and metronidazole. int j sci stud 2016;4(3):131-4. 8. ahmad mm, wani m, dar hm, thakur sa, wani ha, mir in. spectrum of perforation peritonitis in kashmir: a prospective study at our tertiary care centre. int surg j 2016;2(3):381-4. 9. abd elsisy a, hagag m, ewida m. the effect of peritoneal lavage with a mixture of lincomycin– gentamicin on postoperative infection in cases of 13 intraoperative peritoneal lavage in peritonitis: normal saline vs metronidazole colorectal cancer surgery. menoufia med j 2017;30(2):393-. 10. santhosh c, singh ac, shetty kk. efficacy of imipenem lavage versus saline lavage in perforation peritonitis. int surg j 2018;5(6):2148-53. 11. ceruelos ah, romero-quezada l, ledezma jr, contreras ll. therapeutic uses of metronidazole and its side effects: an update. eur rev med pharmacol sci 2019;23(1):397-401. 12. sulli d, rao ms. comparative study of saline versus metronidazole peritoneal lavage in operated peritonitis cases. j evid based med healthc 2016;3(31):1446-8. 13. gangwal m. intraoperative lavage in patients undergoing emergency laparotomy for prevention of wound infection: a comparitive study between normal saline and metronidazole. jmscr 2017;5(3):18552-6. 14. petersen s, deder a, prause a, et al. transverse vs. median laparotomy in peritonitis and staged lavage: a single center case series. ger med sci. 2020; 18:doc07. published 2020 sep 14. doi:10.3205/000283 15. sarada b, bhargavi g. sobha rani b, et al. comparative study of povidone iodine versus metronidazole in normal saline in peritoneal lavage in cases of peritonitis. j evid based med healthc 2020; 7(34), 1798-1803 16. paolo gg, antonino b, antonino g. which surgical strategy in sepsis from retro peritoneal colic perforation. biomedical journal of scientific & technical research. 2019; 21(2):15765-72. 17. ramachandran ml. age, sex incidence with signs and symptoms of peritonitis. int j res med sci 2014;2(3):916-919. 18. singh h, agrawal m, singh nk, kaul rk, ilahi i, ahuja r. intraoperative lavage in peritonitis: comparison between saline and metronidazole. int j sci stud 2016; 4(3):131-134. 19. dalvi an, gondhalekar ra, upadhye as. postoperative irrigation in the management of amoebic peritonitis. j postgrad med 1987; 33:61-4. 20. sheeraz m, verma ak, shahid m, manal mk, abrar a. evaluation of pre-operative peritoneal lavage by superoxidized solution in peritonitis. middle east j intern med 2009;2,3:15-33. 21. kreis be, de mol van otterloo aj, kreis rw. open abdomen management: a review of its history and a proposed management algorithm. med sci monit. 2013;19:524-533. published 2013 jul 3. doi:10.12659/msm.883966 22. hesami ma, alipour h, nikoupourdaylami h, alipour b, bazargan-hejazi s, ahmadi a. irrigation of abdomen with imipenem solution decreases surgical site infections in patients with perforated appendicitis: a randomized clinical trial. iran red crescent med j. 2014;16(4):e12732. doi:10.5812/ircmj.12732 23. kirkpatrick aw, coccolini f, tolonen m, et al. the unrestricted global effort to complete the cool trial. world j emerg surg. 2023;18(1):33. published 2023 may 11. doi:10.1186/s13017-023-00500-z 24. hamilton al, kamm ma, ng sc, morrison m. proteus spp. as putative gastrointestinal pathogens. clin microbiol rev. 2018;31(3):e0008517. published 2018 jun 13. doi:10.1128/cmr.00085-17 25. singh i, singh g, singh h, singh m. comparative study of outcome of peritoneal lavage using saline, metronidazole and betadine solution in peritonitis. int j acad med pharm. 2023; 5(2):1688-91.. 26. chundamala j, wright jg. the efficacy and risks of using povidone-iodine irrigation to prevent surgical site infection: an evidence-based review. can j surg. 2007;50(6):473-81. 27. agi m mcfarland and others, impact of surgical-site infection on health utility values: a metaanalysis, british journal of surgery, volume 110, issue 8, august 2023, pages 942–949. 28. zabaglo m, sharman t. postoperative wound infection. [updated 2022 sep 19]. in: statpearls [internet]. treasure island (fl): statpearls publishing; 2023 jan-. the authors: dr. hira aslam, senior registrar, department of surgery, mayo hospital kemu, lahore. dr. muhammad khurram jameel, assistant professor, department of surgery, anmc/cma hospital, lahore. dr. pir muneeb, assistant professor, department of surgery, anmc/cma hospital, lahore. dr. aneeqa nasir qureshi, senior registrar, department of surgery, mayo hospital kemu, lahore. dr. fawad hameed, associate professor, department of surgery, anmc/cma hospital, lahore. dr. muhammad ammar, senior registrar, department of urology, anmc/cma hospital, lahore. 14 intraoperative peritoneal lavage in peritonitis: normal saline vs metronidazole dr. maham qazi, senior registrar, department of surgery, anmc/cma hospital, lahore. dr. abrar ul hassan pirzada, associate professor, department of plastic surgery, anmc/cma hospital, lahore. corresponding author: dr. muhammad khurram jameel, assistant professor, department of surgery, anmc/cma hospital, lahore. email: khurramjameel999@gmail.com for web full book 1 proceedings s.z.m.c. vol: 37(3): pp. 1-7, 2023. pszmc888-37-3-2023 trans-arterial chemoembolization for the treatment of hepatocellular carcinoma: a single tertiary care institute experience 1parvez ahmed, 1nasreen naz, 1amjad sattar, 1nauman al qamari, 2imtiaz ali, 1faryal farooq, 1muhammad asadullah, 3rakesh kumar 1department of radiology, dow university of health sciences, karachi. 2department of radiology, liaquat national hospital, karachi. 3department of radiology, jpmc, karachi. abstract introduction: trans-arterial chemoembolization (tace) is a mainstay therapeutic option indicated in early-stage nonresectable hepatocellular carcinoma (hcc) and has been shown to be associated with survival improvements. this study aimed to evaluate the radiological and clinical response of those who underwent tace. aims & objectives: we aimed to evaluate the radiological and clinical response in patients who underwent transarterial chemoembolization. place and duration of study: this study was conducted in the vascular interventional radiology department, dow institute of radiology, dow university of health sciences, ojha campus karachi for a period of 18 months between january 2021 and june 2022. material & methods: hcc patients (n=181) who underwent tace as their primary treatment at dow hospital ojha campus karachi between january 1st, 2021, and june 30th, 2022 were included. inclusion criteria followed the barcelona clinic liver cancer (bclc) and child-pugh staging systems. tumor response was evaluated using “modified response evaluation criteria in solid tumors (mrecist)”, and patients were categorized into complete response (cr) or partial response (pr) groups. the study compared background, clinico-laboratory, and radiological features between these groups, including hcc sizes and ct scan findings before and after tace. the retrieved data was entered and analyzed using spss version.21. results: of the total 118 patients, 51.70% showed cr to the tace, while pr was noted in 48.30%. age, sex, viral hepatitis, and co-morbids showed no intergroup differences. however, child-pugh stage and bclc were significantly associated with tumor response. similarly, laboratory parameters revealed significant mean differences between the two groups (p<0.001), except international normalized ratio and alanine transaminase. those who had achieved cr had a mean number of tumor less than that of pr. similarly, tumor size significantly decreased post-tace (p<0.0001). moreover, arterial-phase enhancement and portal venous and delayed phases washout tumors before tace were reported in 76.30%. over half of the patients had no residual tumor tissue after tace. tumors with arterial-phase enhancement and portal venous and delayed phases washout were highly susceptible to tace.furthermore, child-pugh classa had a greater estimated mean survival than class b (p<0.001). conclusion: over half of the patients showed complete response to the tace. bclc staging (b), child-pugh (stage a), and small tumors had a favourable effect on the radiological and clinical response in the early-stages of hcc. tumors with an arterial-phase enhancement and portal venous and delayed phases washout were more susceptible to tace. keywords: hepatocellular carcinoma; trans-arterial chemo-embolization; chronic liver disease; tace; radiological response introduction hepatocellular carcinoma (hcc) is a serious and a major global health issue with about 500,000 new cases identified yearly, making it the most prevalent and the 5th most frequent primary liver cancer1,2. it represents between 70%-90% of all cases of primary liver cancer3. in asian countries, the prevalence ranges between 0.3% and 1.6%4. chronic viral hepatitis or cirrhosis, environmental contaminants, secondary viral infections such as hepatitis b(hbv) and c (hcv) and alcoholic or fatty liver conditions, lifestyle aspects including smoking, alcohol intake, and dietary habits, metabolic conditions like diabetes and high body mass index, and genetic and hereditary issues are the most common causes3,5. despite diagnostic, and therapeutic advancements, hcc continues to pose significant diagnostic and treatment limitations5,6. 16 1department of public health, health services academy, islamabad. 2department of management sciences, riphah int university, islamabad. 3department of medicine, ihitc, islamabad. 4department of rheumatology, pims, islamabad. 5department of medicine, dow university of health sciences, karachi. 70 1department of orthopedics, thq, khanewal 2department of orthopedics, lgh, lahore 3department of orthopedics, thq, ferozwala 4department of orthopedics, cmh, lahore 5department of hand & upper limb surgery, cmh, lahore 6department of orthopedics, al-rehmat benevolent trust 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 2 trans-arterial chemoembolization for the treatment of hepatocellular carcinoma: a single….. “the barcelona clinic liver cancer (bclc) staging system” considers tumor characteristics, performance status and liver function for evidencebased treatment selections7. hepatic resection, radio-frequency ablation (rfa) and liver transplantation are recommended for early stage hcc bclc (stage a). nevertheless, due to specific situations, these modalities cannot always be utilized for all hcc patients. loco-regional treatment (lrt) is a bridging technique for patients on the transplant queue8,9. trans-arterial chemo embolization (tace) is a therapeutic approach that involves the selective delivery of chemotherapeutic agents and embolic materials directly into the tumor-feeding arteries, leading to the dual effect of tumor necrosis and arterial occlusion. it is the most frequently used treatment in patients with bclc stage b, which includes multi-nodular tumors with child-pugh (a or b) stage and good performance status and is not susceptible to resection. arterial neoangiogenesis is the hallmark of hcc or hepatoma. these aggressive liver tumors rely heavily on the development of an extensive blood supply to sustain their rapid growth and progression. tace blocks tumor blood flow, inhibits tumor growth and produces significant findings in terms of tumor response, which is about 50%9,10. this means that about half of the patients treated with tace experience a significant reduction in tumor size or stabilization of the disease, leading to improved clinical outcomes. these findings highlight the considerable impact of tace in managing hcc and its potential as a curative or palliative treatment modality. the radiological and clinical response of tace in the treatment of hcc has been a subject of significant interest and research. assessing tumor response to tace is crucial for determining treatment efficacy and future therapy.11 to assess response to lrt, a “modified response evaluation criterion in solid tumours (mrecist) are a set of published rules used to assess tumor burden in order to provide an objective assessment of response to therapy with targeted agents for hepatocellular carcinoma (hcc). it considers the extent of viable contrast-enhancing regions within the tumor9,10,12. although there is a validated relationship between hcc enhancement patterns and image results, these findings are closely associated with tumor differentiation, which plays a crucial role in understanding the aggressiveness and prognosis of hcc11,13,14however there is a data scarcity of this region and the therapy outcomes remain diverse. considering this, the present study is conducted to determine the radiological and clinical response of hcc patients who underwent tace. material and methods this retrospective cross-sectional study analysed hcc patients from january 1st, 2021, to june 30th, 2022, at the department of radiology, dow hospital ojha campus, karachi. the study protocol was agreed by the dow university of health sciences ethical review committee vide number irb-2790/dums/exemption/2023/31 dated 19th nov 2022. all patients who have undergone tace for multicentre hcc during the study were included, without calculating formal sample size. 118 patients completed this study.the criteria for inclusion were diagnosed case of chronic liver disease (cld) with unresectable multi-centric hcc (those with partial portal vein thrombosis and those with multi-centric hcc not amenable to percutaneous ablation).exclusion criteria included diagnosed cld with complete portal vein thrombosis, childpugh stage c , local or distant metastases, and received prior treatment from another institute, or loss of follow-up during treatment. background characteristics such as age and sex, clinical and laboratory characteristics, including viral hepatitis, co-morbids, child-pugh stage, bclc, total albumin and bilirubin, alanine aminotransferase (alt), alpha-fetoprotein (afp), inr, and tumor characteristics such as number of tumors, appearance, size, and location were recorded. tace technique: prior to the tace procedure, written informed consent was obtained. once the aseptic measures were taken, a local anaesthesia was administered and the common right femoral artery was punctured. a 6fr vascular sheath was then placed, and the celiac and superior mesenteric arteries were surveyed angiographically. the next step was to perform a common hepatic angiography to determine the tumor's blood supply. after that, a microcatheter was cautiously and selectively inserted into the artery that was feeding the tumor. doxorubicin 50mg, the chemotherapy agent mixed with lipiodol was injected. additional embolisation was performed with pva particles (100–350 microns). the interventional radiologist determined the correct amount of the chemotherapeutic and embolic agents based on the several factors like size and number of tumors, the tumor arterial blood supply, the degree of liver impairment, and renal function. for those who had 3 trans-arterial chemoembolization for the treatment of hepatocellular carcinoma: a single….. adequate hepatic function reserve (child-pugh a or child-pugh b < 8) and a residual viable tumor, repeat tace treatments were planned at 6 to 8weeks intervals after the initial treatment. however, those patients who showed no evidence of residual viable disease (i.e., with cr according to mrecist), imaging follow-up was recommended every 2 to 3 months. radiological characteristics: the preand post-tace triphasic abdominal ct scans were performed, and tumor characteristics were recorded as reported by experienced radiologists. the hcc attenuation in each phase was classified as arterially enhancing with washout in porto-venous and delayed phases tumor and heterogeneously enhancing with washout in portal venous and delayed phases. the pattern of enhancement is classified as residual tumor enhancement or no residual tumor enhancement. tumor size was compared preand post tace. post tace tumor response: patients were evaluated at 6weeks of post tace period by contrast ct scan of abdomen with triphasic protocol and then followed up every 2 to 3months, depended on individual tumor response until they reached the endpoints, which included death or survival. the tumor response was measured based on the modified recist (mrecist) standards12 with two categories of response: complete response (cr) and partial response (pr) obtained at first follow-up visit. statistical analysis: the retrieved data was analysed spss version 21. quantitative variables were presented as mean and standard deviation, medians and interquartile ranges, or both. numbers and percentages were used for categorical variables. the chi square test or fisher exact was used to determine the association between categorical variables, and numerical data differences were calculated using the student independent t test. a paired sample t-test was used to see the preand post-mean differences. survival analysis was done using the kaplan-meier curve. statistical significance was set at 5% with a 95% confidence interval. p value 0.05 was considered significant, results the details of the study patients (n=118) are summarised in table-1. the mean age of the study patients was 49.42±10.74 years [median (iqr) 50.00 (16.25)]. nearly two-thirds of the study participants were male. viral hepatitis caused by hcv was observed in 81 (68.64%) patients and hbv in 37 (31.36%) patients. those who had coexisting illnesses were 53 (44.92), where hypertension was documented in 41 (34.75%), and diabetes mellitus in 38 (32.10%). majority patients were grouped into child-pugh stage a, whereas 75.42% of the patients were in stage b based on "the barcelona clinic liver cancer (bclc) staging system." all patients had multiple tumors. the hcc diameter was 3.99±0.90 cm, with 83 (70.30%) having a tumor size >3 cm. tumors were located in more than half of the patients in right lobe of the liver. on the other hand, one-fourth of the patients had portal vein tumor thrombus. the study patients were divided into two tumor response categories. of the total 118 patients, 51.70% showed complete response to tace, while 48.3% had partial response (pr). patient background, laboratory, and tumor characteristics, as well as the outcome of both tumor response categories, are presented abridged in table-1. age, sex, viral hepatitis infection, and co-morbid conditions showed no intergroup differences. childpugh stage and bclc were significantly associated with tumor response. laboratory parameters including total bilirubin, albumin, and afp showed a significant mean inter group differences (p<0.001). those who responded completely had a mean tumor (3.00±1.00 vs.5.00±1.00 p<0.001) compared to the partial responders. those with smaller ( 3cm) tumor sizes had a higher rate of complete response than those with larger (>3cm) ones did not [(26(42.60%) vs. 9(15.80%)], and the association was statistically significant (p<=0.001). patients were followed, and at the endpoint, no mortality was noted in those who responded completely. tumor size was assessed using a ct scan of the abdomen with a triphasic protocol, and the results were compared preand post-6-weeks tace as shown in fig-1. a paired sample t-test was run to ascertain whether there was a statistically significant mean difference before and after tace. there was a significant mean difference in tumor size (3.27±0.90 vs. 3.99±1.02, p<0.0001). the paired mean difference was 0.72 (95% ci [0.62-0.82]). arterially enhancing with washout in portovenous and delayed phases tumor appearance before tace was documented in 90 (76.30%), followed by heterogeneously enhancing with washout in portal venous and delayed phases in 28 (23.70%). posttace, no residual enhancement was documented in more than half of the patients. a high proportion of patients showed no residual enhancement of tumor in arterial phase, as given in fig-2 and fig-3. 4 trans-arterial chemoembolization for the treatment of hepatocellular carcinoma: a single….. the average duration of patient survival was 12.55 months (95% confidence interval: 11.90–13.19), with a median survival of 14.00 months (95% confidence interval: 12.36–15.64). tumor response yielded no statistics because all cases were censored in cr. the computed mean [(12.68, 95% 11.9913.37 vs. 11.63, 95% 10.42-12.83)] duration for the absence of ppvt was greater compared to the presence (p=0.414). overall, the estimated mean duration for tumor size 3 was greater than that for tumour size >3 (12.42 vs.12.22) but the difference was statistically nonsignificant (p=0.138). likewise, bclc (stage b) had an estimated mean survival time greater than that of bclc c, and the difference was statistically nonsignificant (p=0.414). on the contrary, child-pugh score a had an estimated mean survival time greater than that of score b, and the difference was statistically significant (p<0.001) as shown in fig-4. fig-1: tumor size before and 6 weeks after tace fig-2: tumor appearance before and 6 weeks after tace. fig-3: the pre-tace ct scan of the patient's abdomen with contrast (triphasic protocol), followed by a post-tace ct with contrast, revealing a remarkable complete response of the hcc. fig-4: overall survival comparing (a) partial portal vein thrombosis (b) tumor size (c) bclc (d) child-pugh score 5 trans-arterial chemoembolization for the treatment of hepatocellular carcinoma: a single….. characteristics total tumor response p-value cr n=61 pr n=57 age(years) mean±sd 49.4± 10.7 48.2± 11.9 50.7± 9.2 0.201 sex, n (%) female 42 (35.6) 17 (27.9) 25 (43.9) 0.070 male 76 (64.4) 44 (72.2) 32 (56.2) viral hepatitis, n (%) hbv 37 (31.4) 19 (31.2) 18 (31.6) 0.960 hcv 81 (68.7) 42 (68.9) 39 (68.4) presence of comorbid, n (%) no 65 (55.1) 34 (57.7) 31 (54.4) 0.883 yes 53 (44.9) 27 (44.3) 26 (45.6) diabetes mellitus, n (%) no 80 (67.8) 42 (68.9) 38 (66.7) 0.800 yes 38 (32.2) 19 (31.2) 19 (33.3) hyperten sion, n (%) no 77 (65.3) 40 (65.6) 37 (64.9) 0.940 yes 41 (34.6) 21 (34.4) 20 (35.1) childpugh score, n (%) a 72 (61.02) 53 (86.89) 19 (33.33) <0.001 b 46 (38.9) 08 (13.1) 38 (66.7) bclc staging b 89 (75.4) 54 (88.5) 35 (61.4) 0.001 c 29 (24.6) 07 (11.5) 22 (38.6) total bilirubin (mg/dl) 1.86± 0.29 1.76± 0.28 1.96± 0.27 <0.001 albumin (g/dl) 3.56± 0.26 3.67± 0.23 3.45± 0.25 <0.001 international normalised ratio (inr) 1.23± 0.14 1.22± 0.13 1.24± 0.15 0.300 alphafetoprotei n (ng/ml) 1588.9 ± 1315.0 1170.1 ± 1009.6 2037.2 ± 1458.3 <0.001 alaninetr ansamina( alt) u/l 53.4± 5.9 53.52± 6.00 53.33± 6.01 0.863 number of tumor 4.19± 1.39 3.00± 1.00 5.00± 1.00 <0.001 tumor location in liver both lobe 39 (33.1) 05 (8.2) 34 (59.7) <0.001 left lobe 15 (12.7) 12 (19.7) 03 (5.3) right lobe 64 (54.3) 44 (72.2) 20 (35.1) ppvt no 89 (75.4) 54 (88.5) 35 (61.4) 0.001 yes 29 (24.6) 7 (11.5) 22 (38.6) tumor size 3 35 (29.7) 26 (42.6) 9 (15.8) 0.001 >3 83 (70.3) 35 (57.4) 48 (84.2) outcome alive 102 (86.4) 61 (100.0) 41 (71.9) <0.001 expire 16 (13.6) 0 16 (28.1) table-1: background, biochemical, tumor related characteristics and intergroup differences. discussion tace is the preferred loco-regional treatment for intermediate-stage hcc, according to the clinical guidelines. multimodal therapies based on tace have been shown to be more efficient than conservative management. to evaluate the efficacy, it is crucial to perform post-tace radiological response analysis, utilizing the modified recist principle, which recommends one-dimensional evaluation of the longest remaining tumor dimension and the recist numerical criteria of response12,15. the measurement of residual viable tumor tissue after tace to determine personal responses is a proxy indication of the survival rate. our findings revealed that a cr was achieved in 51.70% of patients. a study conducted by lee, s. w., et al9 reported radiological cr in 60% of the study patients. a similar study in italy with a single hcc undertaking tace reported a 64% cr rate and 26% pr16.the variation could be attributed to the tumour size, inclusion criteria, selectivity of technique, and expertise of the treating consultant. most clinico-laboratory variables had no effect on the cr. however, a smaller child-pugh grade was found to have significant impact on tace response in the study. this association has been shown frequently in studies10,17,18, and it may be associated to the aggressive therapy regimen that patients with a better functional liver status were able to tolerate as compared to that utilised in the wake of advanced disease.patient with larger tumors ( 3 cm) had a significantly higher percentage of cr compared to those with smaller tumors (<3 cm) (42.60% vs 57.40%).tace is the treatment of choice for large tumors and neoadjuvant chemoembolization to reduce the size of the tumor for liver transplant or resection19. previous studies suggest that small tumors are more likely to achieve cr, unlike our study findings16,20. the plausible explanation for these differences, where larger tumors had a higher percentage of cr, could be attributed to differences in the patient population, tumor characteristics, or therapy protocols. it is also possible that the sample size or methodology of the earlier studies was not robust enough to detect the effect of tumor size on tace response. in our study, those with no partial portal vein tumor thrombus (pvtt) had a high radiological complete response. tace paired with radiation has demonstrated greater efficacy in patients with hcc and pvtt by maintaining portal blood flow, preventing the loss of liver function, and preventing intra-vascular tumour progression19. the presence of pvtt poses a therapeutic challenge, and its 6 trans-arterial chemoembolization for the treatment of hepatocellular carcinoma: a single….. presence may compromise the candidature for tace due to the risk of liver function deterioration and hepatic infarction. however, selective, or superselective tace procedures can still be performed safely in some cases. to further understand what factors, influence hcc responsiveness, the study divided hcc lesions into two groups based on radiological appearance and found that 76.30% of tumors were arterially enhancing with washout in the porto-venous and delayed phases, while 23.70% were heterogeneously enhancing with washout in the portal venous and delayed phases. more than half of the patients had no residual enhancement after tace. in arterial phase, a large number of patients had no residual tumor enhancement as found by zhang wie et al10, found that lesions with strong arterial phase enhancement had a higher probability of nearcomplete necrosis compared to those with mild to moderate enhancement (37%).considering enhancement and margin factors may help determine tace outcomes, and examining both variables before treatment may be beneficial.the findings imply that the application of enhancement and margin factors may aid in determining the probability of tace outcome, and henceforth it may be beneficial to examine both variables concurrently before subjecting patients to surgical procedures. the absence of ppvt and tumor size 3 had a longer mean duration than the presence and larger size. bclc (b) had a greater estimated mean survival time than bclc (c), while child-pugh score a had a greater estimated mean survival time than score b, with a significant difference. cr had no statistics for tumor response, but tace-induced radiological tumor response predicted overall survival, and no mortality was observed in those who achieved cr. the study's findings are consistent with those of other studies regarding smaller tumors, child-pugh score, and bclc staging on cumulative survival9,16,20. tace is an effective therapy for unresectable hcc, including bclc-c and child-pugh-b stages, and can improve survival and responsiveness without affecting liver function19,21. the study has limitations such as a retrospective design, selection and reporting biases, small sample size, grouping of radiological response into two categories only, and lack of characterization of tumor margin shape. prospective studies with more cases and covariates are needed to improve the findings. conclusion based on this study's findings, it can be concluded that over 50% of the patients achieved a complete response to tace. factors such as bclc b stage, child-pugh class a, and small tumor size positively influenced the response to tace during the early stages of hcc. additionally, tumors exhibiting arterial enhancement and washout in the portovenous and delayed phases were found to be more susceptible to tace treatment. the use of ct imaging could aid in refining the criteria for selecting tace as a treatment option, thereby enhancing overall outcomes and patient survival. it is important to consider an individual's risk profile, co-morbid conditions, and potential benefits when deciding to pursue tace as a therapeutic approach, with the goal of improving survival rates while minimizing adverse events. references 1. yoo gs, yu ji, park hc, hyun d, jeong wk, lim hy, et al. do biliary complications after proton beam therapy for perihilar hepatocellular carcinoma matter? cancers (basel). 2020;12(9):2395 2. dhanasekaran r, nault j-c, roberts lr, zucmanrossi j. genomic medicine and implications for hepatocellular carcinoma prevention and therapy. gastroenterology. 2019;156(2):492-509 3. baecker a, liu x, la vecchia c, zhang z-f. worldwide incident hepatocellular carcinoma cases attributable to major risk factors. eur j cancer prev. 2018;27(3):205-212 4. naseer l, sabir s, azeem mk, zaman s, mahmood au, mastoor m. profile of hepatocellular carcinoma: an experience from a tertiary care hospital. pakistan j medical health sci. 2022;16(10):399-399 5. rawla p, sunkara t, muralidharan p, raj jp. update in global trends and aetiology of hepatocellular carcinoma. contemp oncol. 2018;22(3):141-150 6. rawla p, sunkara t, barsouk a. epidemiology of colorectal cancer: incidence, mortality, survival, and risk factors. prz gastroenterol. 2019;14(2):89-103 7. han k, kim jh. transarterial chemoembolization in hepatocellular carcinoma treatment: barcelona clinic liver cancer staging system. world j gastroenterol. 2015;21(36):10327-10335 8. european association for the study of the liver. easl clinical practice guidelines: management of hepatocellular carcinoma. j hepatol. 2018;69(1):182-236 9. lee sw, lee ty, cheng yc, yen cl, yang ss. the correlation with tumor radiological characteristics and prognosis of patients with early-stage hepatocellular carcinoma receiving transcatheter 7 trans-arterial chemoembolization for the treatment of hepatocellular carcinoma: a single….. arterial chemoembolization. medicine (baltimore). 2021;100(26):e26414 10. zhang w, xu a-h, wang w, wu y-h, sun q-l, shu c. radiological appearance of hepatocellular carcinoma predicts the response to trans-arterial chemoembolization in patients undergoing liver transplantation. bmc cancer. 2019;19(1):1041 11. zhou w, lv y, hu x, luo y, li j, zhu h, et al. study on the changes of ct texture parameters before and after hcc treatment in the efficacy evaluation and survival predication of patients with hcc. front oncol. 2022;12:957737 12. bargellini i, bozzi e, campani d, carrai p, de simone p, pollina l, et al. modified recist to assess tumor response after transarterial chemoembolization of hepatocellular carcinoma: ct–pathologic correlation in 178 liver explants. eur j radiol. 2013;82(5):e212-e218 13. lee jh, lee jm, kim sj, baek jh, yun sh, kim kw, et al. enhancement patterns of hepatocellular carcinomas on multiphasic multidetector row ct: comparison with pathological differentiation. br j radiol. 2012;85(1017):e573-e583 14. nishie a, yoshimitsu k, okamoto d, tajima t, asayama y, ishigami k, et al. ct prediction of histological grade of hypervascular hepatocellular carcinoma: utility of the portal phase. jpn j radiol. 2013;31:89-98 15. lencioni r, llovet jm. modified recist (mrecist) assessment for hepatocellular carcinoma. semin liver dis. 2010;30(1):52-60 16. terzi e, piscaglia f, forlani l, mosconi c, renzulli m, bolondi l, et al. tace performed in patients with a single nodule of hepatocellular carcinoma. bmc cancer. 2014;14:601 17. grieco a, marcoccia s, miele l, marmiroli l, caminiti g, ragazzoni e, et al. transarterial chemoembolization (tace) for unresectable hepatocellular carcinoma in cirrhotics: functional hepatic reserve and survival. hepatogastroenterology. 2003;50(49):207-212 18. brown db, fundakowski ce, lisker-melman m, crippin js, pilgram tk, chapman w, et al. comparison of meld and child-pugh scores to predict survival after chemoembolization for hepatocellular carcinoma. j vasc interv radiol. 2004;15(11):1209-1218 19. manjunatha n, ganduri v, rajasekaran k, duraiyarasan s, adefuye m. transarterial chemoembolization and unresectable hepatocellular carcinoma: a narrative review. cureus. 2022;14(8):e28439 20. baek my, yoo jj, jeong sw, jang jy, kim yk, jeong so, et al. clinical outcomes of patients with a single hepatocellular carcinoma less than 5 cm treated with transarterial chemoembolization. korean j intern med. 2019;34(6):1223-1232 21. piscaglia f, ogasawara s. patient selection for transarterial chemoembolization in hepatocellular carcinoma: importance of benefit/risk assessment. liver cancer. 2018;7(1):104-119 the authors: dr. parvez ahmed, fellow, department of vascular interventional radiology, dow university of health sciences, karachi. dr. nasreen naz, head of department, department of radiology, dow university of health sciences, karachi. dr. amjad sattar, associate professor, department of radiology, dow university of health sciences, karachi. dr. nauman al qamari, assistant professor, department of radiology, dow university of health sciences, karachi. dr. imtiaz ali senior registrar, department of radiology, liaquat national hospital, karachi dr. faryal farooq fellow, department of vascular interventional radiology, dow university of health sciences, karachi. dr. muhammad asadullah fellow, department of vascular interventional radiology, dow university of health sciences, karachi. dr. rakesh kumar consultant department of radiology, jpmc, karachi corresponding author: dr. parvez ahmed, fellow, department of vascular interventional radiology, dow university of health sciences, karachi. email:drshaikhparvez786@gmail.com full book 42 proceedings s.z.m.c. vol: 37(2): pp. 42-49, 2023. pszmc-885-37-2-2023 abstract introduction: pleural effusion is the commonest extra-pulmonary manifestation of tuberculosis in endemic populations. this is managed in routine, by diagnostic aspiration and treated with anti-tb medicines. a significant number of treated patients remain visiting pulmonology and thoracic surgery departments, for chest pain, abnormal chest x-ray, and shortness of breath due to residual pleural thickening (rpt). aims &objectives: the study was conducted to determine the usefulness of complete thoracentesis under ultrasound guidance over diagnostic aspiration in preventing residual pleural thickening. place and duration of study: this case-control study was conducted from 01-07-21 to 30-06-2022 at pulmonology opd of gulab devi teaching hospital lahore-pakistan. material & methods: after approval from irb and informed consent, 300 consecutive patients with age 14-85 years, diagnosed tb-pleural effusion, willing for intervention and participation in study were included in the study while pediatric patients, non-tb effusions and unwilling for aspiration or participation were excluded. two groups of 150 each were allocated through odd and even numbers. therapeutic aspiration under ultrasound guidance for interventional group and diagnostic aspiration of 50-100 ml for control group was performed. all patients were treated with same standard and followed up according to the principles of dots. pleural thickness was measured utilizing ultrasonography at day 0, 60 and 180. outcome was compared in both groups. spss-24, software was utilized for analysis. p ≤ 0.05 was taken as significant. results: only 31/150 patients (20.66%) from interventional group displayed residual pleural thickening while148/150 cases (98.66%) with residual pleural thickening were noted in control group. the p-value was 0.00001 conclusion: complete thoracentesis under ultrasound guidance is a tremendous tool for precluding residual pleural thickening in tubercular pleural effusions. keywords: effusion, pleural, residual pleural thickening (rpt), tuberculous, thoracentesis, therapeutic ultrasound guided complete thoracentesis, precluding residual pleural thickening in pleural tuberculosis: a case-control study 1abdul rasheed qureshi,1muhammad irfan,1huma bilal,1muhammad sajid,2zeeshan ashraf 1 department of pulmonology, gulab devi teaching hospital lahore, pakistan 2 department of statistics, gulab devi institute of social sciences lahore, pakistan introduction tuberculosis (tb) remains a challenging public health issue in resource limited countries1. about 1/3rd of the globe is infected with mycobacterium and tb is the leading infectious cause of death worldwide2. according to national institute of health pakistan study 2022, pleura is the commonest extra-pulmonary site involved by tuberculosis and disease is manifested as pleural effusion. when a sub-pleural tubercular focus ruptures into pleural space, inflammation is triggered by mycobacterium tuberculosis and its metabolites and evoked by hypersensitivity phenomenon. the pleural inflammation starts with neutrophilic response which is followed by macrophage influx and later on t-helper lymphocyte driven immune reaction occurs, resulting in granuloma formation3. inflammation evolves an increased capillary permeability, influx of proteins and fluid into pleural space, resulting in accumulation of protein rich fluid in pleural space. if patient is not treated timely with appropriate strategy, fibrin protein deposition occurs on pleural surfaces. the lymphatic stomata in parietal pleura are blocked, resulting in a decreased rate of fluid absorption. in this way, the dynamic equilibrium between fluid formation and absorption is lost emerging in effusion accumulation, pleural4 thickening, septae and adhesions formation and in some cases even trap lung can be the outcome. vorster mj et al (2015) communicated that residual pleural thickening is a well-recognized complication of tubercular pleural effusion (tbpe) having prevalence, varying between 5-55%5. it is estimated that 3-25 % of tb-patients develop pleural effusion in endemic populations. in a ultrasound guided complete thoracentesis, precluding residual pleural thickening in pleural t.b….. 43 series of 833 chinese patients, 40.1% were confirmed tb on medical thoracoscopy while chen rl et al (2017) reported a percentage of 23.3%6,7. in india, prabhu vg and co-workers (2012) revealed 23.5% tb pleural effusion8. diacon ah (2003) disclosed 84.2% in south africa9. depew zs and associates (2014) divulged 0.0% tubercular pleural effusions in usa10. the incidence of tb pleural effusion is increasingly recognized, even in developed countries following hiv pandemic11 gopi a et al. (2006) exhibited that even after successful treatment with standard anti-tb drugs, pleural thickening and calcification can occur in 10–72% of patients12. kumar a et al described pleural thickening as organizing phase after granuloma formation in tubercular pleural effusions13. meghji j et al manifested that post treatment pleural thickening of more than 10 mm was seen in 20–46% patients14. a tb-pleural effusion presents usually, as an acute illness by chest pain, shortness of breath, fever and dry cough, interfering with daily activities8,9. about 50-100ml pleural fluid is aspirated for diagnostic purpose in routine and remaining fluid is left inside to be absorbed by medical treatment. around 50% patients show variable degree of pleural thickening at the conclusion of treatment which appears as a homogenous opacity obscuring costo-phrenic angle. while the patient experiences chest pain & shortness of breath and remains upset for abnormal x-ray even after completing full course of treatment. patient visits pulmonology or thoracic surgery department for the rectification of these issues. each following physician considers such cases as pleural effusion but no fluid is found on attempted pleural aspiration. patients are subjected to sophisticated costly procedures in search of diagnosis an ultrasound and or ct-scan usually uncovers residual pleural thickening. literature points out that complete thoracentesis may reduce residual pleural thickening15,16. since we encounter a significant number of such cases in our department daily, we conducted this research to determine the effect of complete thoracentesis on post treatment pleural thickening. and its objective is to determine the usefulness of complete thoracentesis over diagnostic aspiration in tb-pleural effusions, for precluding residual pleural thickening. material and methods this prospective, case-control study was conducted at pulmonology-opd of gulab devi teaching hospital lahore-pakistan. the research ensued on 01-07-2021 and continued till 30-06 2022. ethical approval was obtained from irb of the hospital vide no admin/gdec/257/19.. an interventional case was defined as a patient for which thoracentesis was done under ultrasound guidance till complete evacuation. while a control case was defined as a patient for which only 50 100ml fluid was drawn for diagnostic purposes only, while remaining fluid was left behind, in pleural space. three hundred adult patients of both genders with unilateral pleural effusion, having volume 500ml-1000ml measured by trans-thoracic ultrasound, willing for intervention and participation in study were included via out-patient department. while patients with age < 14 years, small pleural effusion, bilateral pleural effusion, massive effusion, suspected non-tb effusion, pregnancy, hematopoietic malignancy, known hepatic disease, not willing for intervention, ada estimation and participation in study were excluded. 300 consecutive patients were divided into two groups interventional (group-a) and control (group-b), containing 150 patients each with approximately similar demography, by odd and even numbers. all patients were investigated with thorough history, physical examination, radiography, ultrasonography, cbc, esr and pertinent biochemical tests. the group-a patients underwent complete thoracentesis, but diagnostic aspiration was performed for g±±roup-b patients. fluid aspiration was carried out under ultrasound guidance and complete evacuation was obtained under real-time visualization, using sonovista fx(siemen)machine. pleural fluid biochemistry, cytology and ada estimation was done. a diagnosis of tubercular etiology was made by clinical scenario, exudative lymphocytic pleural fluid combined with pleural fluid adenosine deaminase (ada) level. a cut-off value of 40 iu/l or more was considered diagnostic for tb. all patients were treated with anti-tb medicines according to dots (directly observed treatment short course) protocol. patients were followed up for 6-9 months. chest radiography and ultrasonography were performed at day 0, 60 and 180. pleural thickening was measured by trans thoracic ultrasound. a pleural thickness < 5mm were considered mild, 5-10mm moderate, while > 1.0cm were labeled as gross pleural thickening. all findings were recorded on a preformed case report form. findings were summarized, tabulated and conclusions were drawn by statistical analysis. spss-24, software, was used for statistical analysis. categorical variables were represented as ultrasound guided complete thoracentesis, precluding residual pleural thickening in pleural t.b….. 44 frequency; quantitative data was computed as mean with ± sd and fisher exact test was utilized for comparison. odds ratio was calculated to determine the level of association between complete thoracentesis and residual pleural thickening (rpt). sensitivity, specificity, ppv, npv and accuracy of therapeutic thoracentesis was calculated, considering “complete clearance” as benchmark. 95 % confidence interval was observed for defining precision. a value of p < .05 was considered to be statistically significant. results the study population included 210 (70%) male and 90 (30%) female patients. male to female ratio was 2.3:1. the age range was 14-85 years. mean age was 32.94 ± 17.38 years for group (a) and 33.45 ± 17.15 years for (b) respectively with a p value of 0.19. patients presented with general and respiratory symptoms (table-1). trans-thoracic ultrasonography showed 96/150 cases (64.0%) with complex septated pleural effusions and 54/150 patients (36.0%) with non-septated pleural effusions in interventional group. the control group showed 89/150 cases (59.33%) with complex septated pleural effusions & 61/150 patients (40.66) with complex non-septated pleural effusions at the initiation of the study. the p-value was 0.4762. the result was not significant at p<0.05. transthoracic findings were recorded in (table-2). no. study groups pleural thickening cases n(freq) day-0 pleural thickening cases n(freq) day-60 pleural thickening cases n(freq) day-180 1. interventional group a (n=150) mild = 37(24.66%) mild = 54(36.0%) mild = 19(12.66%) moderate = 94(62.66%) moderate = 22(14.66%) moderate = 12(8.0%) gross = 19(12.66%) gross = nil(0.0%) gross = nil(0.0%) no pleural thickening (complete clearance)= 119 (79.33%) 2. control group b (n=150) mild = 45(30.0%) mild = 37(24.66%) mild = 12(8.0%) moderate = 87(58.0%) moderate = 99(66.0%) moderate = 25(16.66%) gross = 18(12.0%) gross = 14(9.33%) gross = 111(74.0%) no pleural thickening (complete clearance)= 02 (1.33%) table-2: ultrasonographic findings n=300 *mild pleural thickening:< 0.5cm, moderate pleural thickening: 0.5-1.0cm, gross pleural thickening:> 1.0cm, n: number. the interventional group showed mild to moderate pleural thickening in 31 cases while complete clearance was noted in 119 cases. this group displayed no cases with gross pleural thickening. the control group demonstrated variable pleural thickening in 148 cases while complete clearance table-1: frequency of patients clinical features in 300 was found only in 02cases. fisher exact test statistic value was <0.00001, significant at p< .05. shown in fig-1 and fig-2. the comparative analysis is shown in table-3. odds ratio was calculated using table-4. considering, sonographic clearance as positive outcome, the odds of a patient experiencing a positive outcome, with complete thoracentesis was calculated as: odds = p(positive) / 1 – p(positive) = (119/150) / 1-(119/150) = (0.793) / (0.207) = 3.83. similarly, the odds of a patient experiencing a positive outcome with existing treatment, was calculated as: odds = p(positive) / 1 – p(positive) = (2/150) / 1-(2/150) = (0.013) / (0.987) = 0.013. odds ratio = 3.83 / 0.013 = 294.61 therapeutic efficacy of complete evacuation was tabulated table-5. no. clinical presentation observed cases frequency 1. chest pain. 280 93.33% 2. shortness of breath 230 76.66% 3. fever 225 75.00% 4. decreased appetite 219 73.00% 5. weight loss 240 80.00% 6. cough 168 56.00% 7. contact history 54 18.00% 8. h/o night sweats 221 73.66% ultrasound guided complete thoracentesis, precluding residual pleural thickening in pleural t.b….. 45 therapeutic outcome positive outcome negative outcome effect with new treatment (intervention) group-a 119 31 effect with existing treatment (control) group-b 02 148 fig-1: sonographic grading of pleural thickening. a:mild pleural thickening. b:moderate pleural thickening. c:gross pleural thickening. fig-2: radiographical appearance before & after treatment. a: cxr-before treatment. b: cxr-after treatment. c: sonogramresidual pleural thickening. table-3: comparative analysis between groups table-4: odds ratio calculation for complete clearance. statistic value 95% confidence interval sensitivity 98.35% 94.16% to 99.80% specificity 82.68% 76.33% to 87.92% positive predictive value 79.33% 73.58% to 84.11% negative predictive value 98.67% 94.92% to 99.66% accuracy 89.00% 84.90% to 92.31% table-5: efficacy of complete thoracentesis for complete sonographic clearance. discussion the aim of this study was to find ways for reducing the frequency of rpt in tubercular pleural effusions and in turn the miseries of the patients. any patient once treated should be able to lead a normal life rather than repeated visits to pulmonology or thoracic surgery departments. people had been using corticosteroids, intra pleural fibrinolytics and intercostal drainage by catheter for preventing rpt but results are variable17,19. barbas cs et al (1991) concluded their research by commenting that approximately 50% of patients of pleural tuberculosis will develop residual pleural thickening at treatment completion but which patient is going to develop is quite uncertain20. in this study, patients of interventional and control groups were between 15 to 38 years of age. a p-value of 0.19, not significant at p<.05, indicated that both groups were comparable demographically to each other at the baseline. a mean age 32.94 years for group a and 33.45 years for group b, reflected that tubercular pleural effusions are common in younger age group. this finding is in agreement with the reports of bhuniya s. et al (2012) and singla r. and colleagues (1995)21,22. developing residual pleural thickening at a young age and having a good quality of life ahead is not possible and is of major concern . sonographic appearence group a patient group b patient fisher exact test statistic value p-value significant at p<.05 mild pleural thickening 19 12 0.261 not significant moderate pleural thickening 12 25 0.0546 not significant gross pleural thickening 00 111 <0.00001 highly significant total pleural thickening cases 31 148 <0.00001 highly significant complete clearance 119 02 <0.00001 highly significant ultrasound guided complete thoracentesis, precluding residual pleural thickening in pleural t.b….. 46 male to female ratio 2.3:1 indicated male predominance in tb-pleural effusions, which may be owing to socioeconomic responsibilities imposed on the male gender. males being responsible in our setup especially for supporting the family, remain working outside and are at greater risk of tb exposure and of contracting the disease. likewise, the disease is more common in males of lower socio-economic group where being hard pressed to earn, they go for medical consultation only when their problems interfere with the work-place activities. this delayed health seeking attitude may be responsible for more involvement of male gender in rpt. additionally the high prevalence of cigarette smoking in male may be another factor for promoting pleural thickening23. this finding is incongruous with the report of naseem khan r et al (2019) who found a female preponderance24. while kumar rai d et al (2021) reported male gender with increased risk of rpt25. because, male are more vulnerable to rpt and being the dominant earning member of the family, they should remain fit for fulfilling their responsibilities. rpt interfering with normal living can be a source of uneasiness for their family also, therefore it requires real rectification. this study demonstrated that most of the patients had symptoms of fever, chest pain, dyspnea and weight loss. these features are similar to those reported by nasim khan r et al (2019), basu, a et al (2012), valdés, l and colleagues (2010)24,26,27. in this study, group-a patients, who were managed by complete aspiration, displayed immediate relief in shortness of breath, chest pain and cough just after complete thoracentesis while the patients of group-b had a gradual relief from symptoms. this subjective improvement may possibly be due to altered nerve impulses arising from the lung or the chest wall after complete evacuation. according to estenne m and associates (1983), the relief of dyspnea after complete evacuation results primarily from reduction in size of the thoracic cage which improves the efficiency of inspiratory muscles to work on a more advantageous portion of their length–tension curve28. similarly, these patients returned to their daily routine activities immediately after complete evacuation while the control patients took far more time to rejoin their workplace because of delayed improvement. even with regular anti-tb treatment, the incidence of rpt still ranges from 10-72%. these patients develop rpt and remain anxious for abnormal chest x-ray even after full course of treatment and consult a number of physicians and even quakes desperately, with a hope of rectification. some of these patients are subjected to decortication but they have to face the complications of thoracotomy as well11. ana maria et al. (2011) displayed that therapeutic aspiration provides relief from dyspnea not only at rest but also on exertion and allows quick and better re-adaptation of patients to routine activities29. the exact mechanism of pleural thickening in tubercular pleural effusion is not clearly understood. however, circumstantial evidence from literature provides ample clues in this context. in tubercular effusion, exudate contains high levels of protein “fibrin” which is deposited on pleural surface, lymphatic vessels are clogged, thus hindering the absorption of fluid, accumulating effusion and resulting in adhesions formation leading to pleural thickening. this may be due to disordered fibrin turn-over. fibrin formation is up-regulated while fibrin dissolution is down-regulated disrupting equilibrium. philip joët f et al (1995) reported decreased level of fibrinolytic activity in tb-pleural effusion as compared to malignancy30. furthermore, tgf-β and tnf-a present in pleural fluid, promote fibrin matrix formation. hua cc et al (1999) reported that tb of pleura may enhance the release of pro inflammatory cytokines like tnf-alpha, which may increase plasminogen activator inhibitor-1 (pai-1) and decrease tissue plasminogen activator (tpa) in pleural fluid. this imbalance of pai-1 and tpa may lead to fibrin deposition and pleural thickening31.chen wl (2015) and co workers reported about the possible role of tnf-a in the pathogenesis of rpt.32 extensive thickening may results in severe restrictive ventilatory disorder33. kunter e et al (2002) reported that high levels of glucose, alpha-1 acid glycoprotein (aag) and creactive protein in pleural fluid, might be responsible for pleural thickening.34hsieh cy et al (2019) disclosed that thrombin upregulates plasminogen activator inhibitor-1 (pa1-1) and mesothelial-mesenchymal transition through protease-activated receptor-1 and cause pleural fibrosis in tubercular pleural effusion35. gopi et al (2007) revealed that development of pleural thickening is closely related to time for diagnosis and treatment, pleural fluid protein content and timely thoracentesis.36 it means if pleural fluid is allowed to stay for a longer time in the cavity, protein keeps on depositing on pleural surface, resulting in pleural thickening. establishing tb ultrasound guided complete thoracentesis, precluding residual pleural thickening in pleural t.b….. 47 diagnosis in pleural effusion is a time-consuming process because ultimate diagnosis is usually made on biopsy, histopathology or culture for acid fast bacillus. therefore, this malady can be controlled by finding rapid ways of diagnosis. kwon js and co-workers (2008) described symptom duration as predictors of rpt37. therefore, early diagnosis and immediate complete aspiration can be employed for prevention of rpt. it is very logical to expect if protein-rich fluid is removed from the cavity, there is nothing behind to be deposited on pleural surface, eventually no pleural thickening can occur. furthermore, there is no contraindication for complete thoracentesis in tubercular pleural effusions. so complete thoracentesis can be a very simple tool for preventing rpt. doosoo jeon (2014) communicated that the prevention of pleural fibrosis should be included in the treatment goals for tubercular pleural effusions38. feller kopman and associates suggested that pleural space should be drained completely in order to avoid rpt39. lai et al communicated that complete drainage had no beneficial effect on rpt reduction40.but balkissou ad and associates (2016) conveyed that rpt patients can be benefitted by appropriate management41. the objective of this study was to determine the usefulness of complete evacuation of pleural space for precluding residual pleural thickening. in this case-control study, an odds ratio of 294.61 (table iv) indicated tremendous association between complete evacuation and complete clearance. this result demonstrated that rpt can be prevented effectively by early diagnosis and complete thoracentesis. wyseret al, and chung cl et al reported that standard anti-tb medicines and early complete aspiration are adequate for the management of tubercular pleural effusion42,43. bhuniya et al also demonstrated a lower rate of rpt with therapeutic drainage21. considering “complete sonographic clearance” (no residual pleural thickening) as reference, the sensitivity 98.35% (95% ci: 94.16% to 99.80%), specificity 82.68% (95% ci: 76.33% to 87.92%) and an accuracy of 89.00% (95% ci: 84.90% to 92.31%) was observed, displaying remarkable efficacy. the main limitation of this study is that it is a single centered study performed on limited number of patients. a short follow-up period was another limitation. we encountered some cases with longer follow up showing decreased pleural thickness three to nine months after treatment completion. a multi-centered study with larger sample size and longer follow up can explore the subject more effectively and more generalizable results can be obtained. since, pleural thickening was estimated by trans-thoracic ultrasound which is a new opted method depending upon the expertise of the operator, can create a source of bias by inter-observer variation. the rpt cases, encountered even in interventional group, were perhaps those who reported late for the management. they spent significant time for getting treatment from local physicians and developed some pleural fibrosis before reaching the hospital. this patient-dependent factor may be confounding. we obtained promising results by complete evacuation under ultrasound guidance. a sensitivity of 98.35% for sonographic clearance is a remarkable outcome. the frequency of rpt was very low because this procedure precluded residual pleural thickening. the results of current study are very important because by utilizing this technique, rpt and post treatment afflictions of patients can be minimized effectively. although minimally invasive, but procedure is simple, rapid, safe and can be performed at day care clinics, even in peripheral areas of the country. additionally, by opting for this tool, we can not only reduce the suffering of patients, but rather lessen the burden on radiology, pulmonology and thoracic surgery departments by reducing rpt. in the light of this discussion, we recommend without any hesitation that we should not allow protein rich pleural fluid, stay in pleural space for longer time. complete thoracentesis under ultrasound guidance and the earliest institution of anti tb medicine can be the best strategy for preventing residual pleural thickening. conclusion complete thoracentesis of tubercular pleural effusion under ultrasound guidance, combined with anti tb medicines, effectively precludes residual pleural thickening. references 1. gbd tuberculosis collaborators. the global burden of tuberculosis: results from the global burden of diseasestudy2015. lancetinfectdis.2018;18(3):26 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[pubmed] [google scholar] 40. lai, yf, chao, ty, wang, yh, lin, as. pigtail drainage in the treatment of tuberculous pleural effusions:a randomised study. thorax. 2003;58(2):149–151.google scholar | crossref | medline 41. balkissou ad, pefura-yone ew, netonggamgne m, et al. opacitépleuralerésiduelleen fin de traitementpour tuberculosepleurale à yaoundé [residual pleural opacity in patients treated for pleural tuberculosisin yaounde]. rev pneumolclin.2016;72(2):115121.doi:10.1016/j.pne umo.2015.09.004 42. wyser, c, walzl, g, smedema, jp, swart, f, van schalkwyk, em, van de wal, bw. corticosteroids in the treatment of tuberculous pleurisy: a double blind,placebocontrolledrandomizedstudy.chest.199 6;110(2):333–338.google scholar | crossref | medline 43. chung cl, chen ch, yeh cy, sheu jr, chang sc. early effective drainage in the treatment of loculated tuberculous pleurisy. eurrespir j. 2008;31(6):12611267.doi:10.1183/09031936.0012 2207. the authors: dr. abdul rasheed qureshi, head of opd, department of pulmonology, gulab devi teaching hospital, lahore. dr. muhammad irfan, incharge chest emergency, department of pulmonology gulab devi teaching hospital, lahore. dr. huma bilal, women medical officer, department of pulmonology gulab devi teaching hospital, lahore. dr. muhammad sajid, medical officer, department of pulmonology gulab devi teaching hospital, lahore. dr. zeeshan ashraf, lecturer, department of statistics, gulab devi institute of social sciences lahore. corresponding author: dr. muhammad irfan, incharge chest emergency, department of pulmonology gulab devi teaching hospital, lahore. email: irfan16d2@gmail.com mailto:irfan16d2@gmail.com numbering.indd 46 proceedings s.z.m.c. vol: 34(4): pp. 46-51, 2020. pszmc-770-34-4-2020 caffeine consumption & perception of its effects amongst university students 1umer maqsood, 1roop zahra, 2muhammad zahid latif, 3hena athar, 3gul muhammad shaikh, 3syed bilal hassan 1department of physical therapy, azra naheed medical college, lahore 2department of community medicine & medical education, azra naheed medical college, lahore 3department of public health, university institute of public health, the university of lahore abstract introduction: a variety of research studies have shown that caffeine usage is highly prevalent among university students and many of them have different perception of its effect on their mental and physical health. these perceptions can influence the intake of caffeine which may lead to its addiction, resulting in serious public health consequences. aims & objectives: the objective of the study is to determine the frequency of caffeine consumption and the perception of its effects among university students. place and duration of study: this study was conducted within 6 months duration, from may 2018 to october 2019 among students of three private universities in lahore, pakistan. material & methods: this was a descriptive cross-sectional study. a sample size of 670 university students was taken and the data were collected from three private universities of lahore through convenience sampling technique. the perception of effects of caffeine was determined through a pre-validated questionnaire, caffeine expectancy questionnaire (caffeq) from consumers and nonconsumers of caffeine. results: a total of 670 university students participated in the study, out of which, 278 (41.5%) were males and 392 (58.5%) were females. 506 (75.5%) participants were consumer of caffeine, while 164 (24.5%) were non consumer. the occurrence of caffeine consumption was more in males (79.9%) than in females (72.4%). participants reported that they consume caffeine because it makes them feel more alert (83.4%), they experience caffeine withdrawal without caffeine (83%) and caffeine makes them feel more energetic (80.6%). the perceptions due to which participants don't consume caffeine were that they expect caffeine makes their heartbeat irregular (81.7%), they don't like the way caffeine makes them feel (80.5%) and caffeine makes them irritable (76.8%). conclusion: the study concludes that male students had more tendencies towards caffeine consumption than females. perceptions for consuming caffeine were feeling of alertness, experiencing caffeine withdrawal symptoms, feeling of energy and ability to work over long periods of time after having caffeine. the perceptions for not consuming caffeine were irregular heartbeat, bad feelings, irritability, and sleep disturbances. key words: caffeine, perception, caffeine consumption, university students introduction caffeine is believed to be the most widely used psychoactive substance globally and easily accessible in pakistani society.1 it is commonly used as an additive in drinks manufactured for weight reduction, sports enhancement and medications.2 caffeine is the constituent of variety of food sources including tea, coffee, drinks and chocolates. the amount ranges from 40 to 180 mg/150 ml for coffee, 24 to 50 mg/150 ml for tea, 15 to 29 mg/180 ml for cola, 2 to 7 mg/150 ml for cocoa, and 1 to 36 mg/28g for chocolate.3 the common use of the caffeinated products is considered to be connected with the food related social behaviors, individual habits and hereditary traits.4 although the moderate use, 400 mg/day is considered safe but children especially girls in the reproductive age are at higher risk of adverse issues due to the altering quantity of caffeine intake.3 the consumption of caffeine among youth is remarkably high worldwide and variety of caffeinated products are easily available.5 the regular use of caffeine has been reported among 92% of college students in united states.6 another study conducted among dutch university students concluded that 87.8% participants ingested caffeine 47 caffeine consumption & perception of its effects amongst university students products during the last 24 hours.7 the consumption of caffeine products among canadian youth adults was found to be 73.6%. similarly, high (94%) proportion of university students in pakistan consumes caffeinated products.8 the intake of caffeine reported is 97% in india and 98.5% in united arab emirates.9,10 the literature describes multiple positive and negative factors related with regular caffeine exposure. caffeine may enhance multiple cognitive and behavioral processes which are associated with effective sports, exercise and cognitive performance like concentration, energy levels, and feelings of fatigue. the moderate intake of caffeine may enhance alertness, decrease tiredness and reduces the risk of certain diseases.11 on the other hand, the negative effects of caffeine consumption include hunger suppression, mood swings, sleep problems and anxiety.12 the abrupt withdrawal in chronic caffeine users may lead to severe headache, muscle pain, stiffness, vomiting and nausea.13 the excessive consumption of caffeine is associated with multiple health related adverse effects like cardiovascular problems, decrease in bone density and calcium deposition.4,8,9,14,15 caffeine is also used as an analgesic due to its pain relieving effects.16 although it reduces fatigue, but the excessive quantity may lead to intoxication and result in insomnia, anxiety and restlessness. the above mentioned health concerns are challenging after consumption of pharmacologically active caffeine. however, the psychological variations such as determination, motivation, mood states and beliefs are associated with perception of oral caffeine consumption the use of caffeine in youngsters especially students in the form of coffee, tea, energy drinks and chocolates is an established phenomenon. the excessive intake may lead to addiction, resulting in serious public health consequences. based on the review of the literature, a gap was detected regarding caffeine consumption and its perception among university students.17 this study aims to determine the frequency of caffeine consumption and the perception of its effects among university students. the findings will help to overcome the issue through awareness, educational programs and also improve the quality of life of university students. material and methods a descriptive cross-sectional study was conducted during a period of six months, from may 2018 to october 2019 after taking approval from the ethical review board of azra naheed medical college. the data was collected from the students of three private universities of lahore, pakistan. the sample size was calculated by taking z1-α/2= 1.96 (standard normal variate at 5% type i error), p= 0.68 (expected proportion in population)and d= 0.05(absolute error).8 the required sample was 335, however to improve the power and accuracy of study, it was increased to 670. non-probability convenience sampling technique was adopted and three private universities in lahore were selected. students were approached in common areas of universities including cafeterias, grounds, mosques etc. students of age group 18-25 years (both male and female) were included in the study. drug addicts and pregnant female students were excluded. the perception of effects of caffeine was determined through a pre-validated questionnaire, caffeine expectancy questionnaire (caffeq) from consumers and nonconsumers of caffeine.18 a written informed consent was taken from 698 students, out of which 21 refused to take part in the study and seven were not fulfilling the inclusion criteria. a total of 670 students filled the questionnaire. the questionnaire comprises of 7 factors, out of which 5 factors were describing perception that led to consumption, and 2 factors were describing perception that led to nonconsumption of caffeine. consumers and nonconsumers were asked about the perception for consuming and not consuming the caffeine respectively. these seven factors include 47 questions, each of which was individually validated in a previous study. the validity of individual questions ranges from 0.36-0.98 and reliability of questionnaire ranges from 0.80 to 0.94.18 the data was analyzed through spss version 24. the data was analyzed by using the chi-square test and a pvalue of ≤0.05 was considered as significant. statistical analysis: data was entered and analyzed by using spss 24. the quantitative variables e.g. age were presented as mean ± s.d. the qualitative variables e.g. gender and perception of effects of caffeine was presented as frequencies and percentages. results there were 670 study participants with a mean age of 21.64±1.91 sd (range= 18 to 25 years). out of 48 caffeine consumption & perception of its effects amongst university students the total subjects, 278 (41.5%) were males and 392 (58.5%) were females, 506 (75.5%) participants were consumer of caffeine, while 164 (24.5%) were non consumers. caffeine consumption was more in males (79.9%) than in females (72.4%). the p value (0.03) calculated through chi square test showed that gender was associated with caffeine intake and caffeine consumption among males was significantly higher than females. the most common form of caffeine intake was tea (54.1%) followed by coffee (22.9%), chocolates (13.4%) and energy drinks (9.5%) respectively. (table-1) perception of effects of caffeine in five categories were asked from consumers of caffeine. these categories include appetite suppression, social mood enhancement, energy/work enhancement, withdrawal/dependence, and physical performance enhancement. in the category of appetite suppression, largest proportion (77.1%) of participants perceived that caffeine suppresses their hunger. regarding social mood enhancement, the majority of participants perceived (79.8%) that caffeine makes them feel happy. in questions related to energy/work enhancement, the largest proportion (83.4%) perceived that caffeine makes them feel more alert. about questions related to withdrawal/ dependence, the majority of students (75.5%) perceived that they will experience headaches without caffeine. in the category of physical performance enhancement, the largest proportion of participants perceived that caffeine improves their athletic performance. (table-2) variable male n=278 female n=392 total n=670 pvalue caffeine consumer yes 222 (79.9%) 284 (72.4%) 506 (75.5%) 0.03* no 56 (20.1%) 108 (27.6%) 164 (24.5%) caffeine intake form coffee 55 (24.8%) 61 (21.5%) 116 (22.9%) 0.716 tea 115 (51.8%) 159 (56.0%) 274 (54.2%) chocolate 29 (13.1%) 39 (13.7%) 68 (13.4%) energy drinks 23 (10.4%) 25 (8.8%) 48 (9.5%) p value≤0.05 was considered significant table-1: socio-demographic characteristics perception of effects of caffeine in consumers frequency % appetite suppression (n=506) suppress feeling of hunger 390 77.1 decrease appetite 342 67.6 allows to skip meals 310 61.3 helps to avoid eating more than usual 298 58.9 helps to control weight 316 62.5 social/mood enhancement (n=506) caffeine makes friendlier 354 70 conversations are better with caffeine 318 62.8 feel more sociable after having caffeine 354 70 feel more confident after having caffeine 352 69.6 caffeine makes feel happy 404 79.8 caffeine improves mood 344 68 energy/work enhancement (n=506) caffeine makes feel more alert 422 83.4 caffeine picks up when feeling tired 384 75.9 caffeine makes feel more energetic 408 80.6 feel less sleepy after having caffeine 320 63.2 caffeine helps to work over long periods of time 404 79.8 caffeine increases motivation to work 372 73.5 caffeine improves concentration 394 77.9 caffeine improves attention 390 77.1 withdrawal /dependence (n=506) experience caffeine withdrawal without caffeine 420 83 need caffeine everyday 372 73.5 get headache if went without caffeine 382 75.5 strong desire without usual amount 370 73.1 feel miserable without usual caffeine 328 64.8 difficulty starting day without caffeine 324 64 feel sick if went without caffeine 326 64.4 trouble giving up caffeine 366 72.3 often crave caffeine 360 71.1 get anxious if abstained from caffeine 380 75.1 unable to function without caffeine 374 73.9 physical performance enhancement (n=506) workouts are better after having caffeine 336 66.4 exercise longer if have caffeine 286 56.5 improves athletic performance 370 73.1 table-2: perception of effects of caffeine in consumers perception of effects of caffeine in two categories were asked from non-consumers of caffeine. these categories were anxiety/negative physical effects and sleep disturbance. in the category of 49 caffeine consumption & perception of its effects amongst university students anxiety/negative physical effects, majority of participants (81.7%) reported that they don’t take caffeine because it makes their heart beat irregular and in the category of sleep disturbance, the largest proportion (74.4%) reported that they don’t consume caffeine because they feel difficulty in sleeping after caffeine. (table-3) perception of effects of caffeine in non-consumers frequency % anxiety/ negative physical effects (n=164) caffeine makes feel nervous 110 67.1 easily stressed after having caffeine 106 64.6 caffeine makes jittery 114 69.5 even small amount of caffeine makes anxious 100 61 don’t like the way caffeine makes feel 132 80.5 caffeine makes irritable 126 76.8 caffeine makes heartbeat irregular 134 81.7 caffeine makes to sweat 118 72 caffeine upsets stomach 104 63.4 sleep disturbance (n=164) using caffeine late in day disrupts sleep 120 73.2 caffeine late in day gives insomnia 102 62.2 difficulty sleeping after having caffeine 122 74.4 caffeine at any time of day throws off sleep 96 58.5 table-3: perception of effects of caffeine in nonconsumers discussion the aim of current study was to determine the frequency of caffeine consumption and perception of its effects among university students and we found that high proportion (75.5%) of the participants was caffeine user. these findings are consistent with the results of different studies describing a high consumption of caffeine among youngsters worldwide.4,7,8,9,10 the present study concluded that among all the caffeinated products, the use of tea was found to be most common (51.8%). these results also corroborates with the findings of a similar research which shows a high consumption of tea among pakistan population.19 this high utilization of tea may be associated with the societal dietary patterns as it is considered as the mandatory component of the breakfast. it has also been found that the consumption of caffeine was significantly high (p=0.03) among males as compared to female participants of the study. these findings are also aligned with the evidence available in the relevant global literature describing that the caffeinated products are more commonly used in males.20 it is worth mentioning that (77%) of the participants reported to use the caffeinated products for the suppression of appetite. however, this finding is not supported by the literature which describes that caffeine is not an effective appetite suppressant.21 the participants reported that they consume caffeine because it enhances their mood. this perception was supported by another study which concludes that caffeine improves mood and enhances cognitive performance.22 participants were using caffeine because they think that it enhances their energy. this perception was also supported by literature.23 another reason for consuming caffeine was that perception of experiencing withdrawal effects. this finding is consistent with the findings of a previous study.24 there is evidence that this caffeine dependence can be reduced by education about caffeine sources and its ability to produce physical dependence.24 the non-consumer of caffeine reported that they experience anxiety and sleep disturbances after consuming caffeine. these perceptions are also supported in literature because caffeine consumption is found to be associated with anxiety related behavior and insomnia.25 the availability of caffeinated products and foods has been rising over the past decade. due to this increased availability and usage people generally link certain perceptions with caffeine consumption. the optimal amount of caffeine usage varies in literature and depends on various factors like age, gender and presence of any disease. a considerable amount of literature has been published on the benefits and harmful effects of caffeine consumption.4,7,8,9,10,26,27 the majority of the literature suggests that mild to moderate caffeine consumption is relatively safe, but that higher doses of caffeine consumption (>400 mg) can cause physiological, psychological, and behavioral harm.28 the current article addresses an important public health issue and contributes to providing better understanding of perceptions that are linked with the effects of caffeine. the findings of this study are beneficial and can lead to implementing changes to 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c, o’brien e, temple jl. caffeine transiently affects food intake at breakfast. j acad nutr diet. 2018; 118(10):1832-43. 22. irwin c, mccartney d, khalesi s, desbrow b. caffeine content and perceived sensory characteristics of pod coffee: effects on mood and cognitive performance. curr res nutr food sci jour. 2018; 6(2):329-45. 23. diaz-lara fj, del coso j, portillo j, areces f, garcía jm, abián-vicén j. enhancement of high-intensity actions and physical performance during a simulated brazilian jiu-jitsu competition with a moderate dose of caffeine. int j sport physiol. 2016; 11(7):861-7. 24. juliano lm, kardel pg, harrell pt, muench c, edwards kc. investigating the role of expectancy in caffeine withdrawal using the balanced placebo design. human psychopharmacology: clinical and experimental. 2019; 34(2):e2692. 25. o’neill ce, newsom rj, stafford j, scott t, archuleta s, levis sc, et al. adolescent caffeine consumption increases adulthood anxiety-related behavior and modifies neuroendocrine signaling. psychoneuroendocrinology. 2016; 67:40-50. 26. rutherfurd-markwick k, ali a. caffeine use in the 21st century: considerations for public health. jj food nutri. 2016; 3(1). 27. reid jl, mccrory c, white cm, martineau c, vanderkooy p, fenton n, et al. consumption of caffeinated energy drinks among youth and young adults in canada. preventive medicine reports. 2017; 5:65-70. 28. temple jl. review: trends, safety, and recommendations for caffeine use in children and adolescents. journal of the american academy of child and adolescent psychiatry. 2019; 58(1):36-45. the authors: dr. umer maqsood assistant professor department of physical therapy, azra naheed medical college, lahore. dr. roop zahra lecturer department of physical therapy, azra naheed medical college, lahore prof. muhammad zahid latif director, department of community medicine & medical education, azra naheed medical college, lahore. dr. hena athar lecturer department of public health, university institute of public health, the university of lahore. dr. gul muhammad shaikh phd scholar department of public health, university institute of public health, the university of lahore. syed bilal hassan lecturer, department of public health, university institute of public health, the university of lahore. corresponding author: dr. umer maqsood assistant professor, department of physical therapy, azra naheed medical college, lahore. e-mail: umer.maqsood@gmail.com numbering.indd 40 proceedings s.z.m.c. vol: 34(4): pp. 40-45, 2020. pszmc-769-34-4-2020 antenatal domestic violence & fetal outcome: a cross-sectional study 1sohaib ashraf, 2muhammad ahmad imran, 3hina mahmood, 4khawar nawaz, 5tayyab mughal, 6umair hafeez sheikh, 7hassan mujtaba cheema, 3ayesha humayun 1 wellman centre of photomedicine, massachusetts general hospital, boston, usa 2department of microbiology, shaikh zayed medical complex, lahore 3department of public health & community medicine, shaikh zayed medical complex, lahore 4department of paediatrics, university hospital of brooklyn sunny downstate medical centre newyork,usa 5department of radiology, shaikh zayed medical complex, lahore 6department of ent, shaikh zayed medical complex, lahore 7amna inayat medical and educational complex, lahore abstract introduction: antenatal domestic violence is a global public health and human rights concern. it increases the risk of maternal and fetal morbidity and mortality. aims & objectives: to assess the frequency of antenatal domestic violence and associated factors and also to determine effect of domestic violence on fetal outcome. place and duration of study: this cross sectional study was conducted from january to june 2015 in post-natal wards of department of gynecology & obstetrics of six tertiary care hospitals of lahore. material & methods: a total of 255 females admitted in post-natal wards of department of gynecology & obstetrics enrolled after proper verbal informed consent. a structured, self-constructed questionnaire was administered on females enrolled consecutively till the desired sample size was complete. data on domestic violence (verbal or physical), its associated factors, gestational age at birth, mode of delivery and birth weight of the neonate was analyzed on spss version 21.0. results: 22% (55/255) females experienced antenatal domestic violence during current pregnancy and out of 55, 72% faced verbal abuse while 27% experienced physical violence. in total 65/255(26%) w low birth weight neonates were born and out of them 58.18% (32/55) were born to mothers facing violence during pregnancy. husband was mostly responsible for violence in 49% of cases. antenatal domestic violence was significantly associated with low birth weight (p=<0.001). conclusion: antenatal domestic violence is associated with low birth weight babies. antenatal domestic violence prevention needs implementation of legislation and changing behaviors of violence in communities so to avoid health implications. key words: abuse, antenatal care, prenatal care, birth weight, domestic violence. introduction one of the common forms of violence against women is intimate partner violence (ipv) which includes physical, sexual or emotional violence. it even includes controlling behaviors by the intimate partner. ipv is not only restricted to developing or underdeveloped countries but is reported from all settings. unfortunately, this global burden of ipv is endured mostly by women. in rare cases women can also be violent but mostly they are the victims by male partners. types of ipv are; 1) acts of physical violence which includes slapping, hitting, beating or any other form of abusing physically, 2) sexual violence which includes assault, abuse etc., 3) emotional or psychological abuse which is subjecting the other person to such a behavior that can cause depression, anxiety, sleep disorders, feeling of worthlessness, regression etc. this may lead to suicidal attempts by the victim, 4) controlling behaviors which includes dominating the partner in unhealthy manner, wanting everything in a way they want, monitoring the partner continuously etc.1 domestic violence is defined “as the threat or exercise of physical, psychological, and/or emotional violence; i.e., any type of force against another person with the intent of inflicting harm or exercising power and control over them by a perpetrator belonging to the victim’s “domestic 41 antenatal domestic violence & fetal outcome: a cross-sectional study environment”. that can be an intimate partner, husband, former intimate partner, family member, friend or by someone who has some connection or familiarity. whether or not the victim actually lives in the same household as the perpetrator is not crucial for the definition of domestic violence.2 the term “intimate terrorism” is used interchangeably with domestic violence as well.3 factors associated with ipv are low levels of education, poor employment status of the husband, physical and verbal abuse during one’s childhood, absence of parental support, and history of violence in family of origin.4 it has been documented that almost seventy-five percent of violence against women is committed by their domestic partners.5 in countries like pakistan where joint or extended family system is prevalent6 women are not only treated poorly by their husbands but also by their family members.7 potential perpetrators, including spouses and partners, parents, other family members, neighbors, and men in positions of power or influence.8 psychological violence is found to be the most common type of violence in rural women residing in pakistan.9 a systematic review conducted in pakistan showed that men are the perpetrators of ipv and women are the victims.10 a systematic review done in ethiopia showed that a significant number of women experienced violence while they were pregnant.11 ipv/domestic violence is also found to be associated with infertility or sub fertility.12 living in an abusive environment leads to chronic stress which may affect birth weight of the newborn.13 a study conducted in karachi pakistan correctly pointed out the scarcity of data on antenatal domestic violence in muslim countries.14 therefore, the aim of this study was to assess the frequency of antenatal domestic violence during pregnancy, factors associated with it and its association with fetal outcomes in lahore. the aim of this study is to assess the frequency of antenatal domestic violence during pregnancy, its associated factors and to determine its effect on fetal outcome in females availing obstetric services in tertiary care hospitals of lahore. material and methods this cross sectional study was conducted for duration of 6 months from january 2015 to june 2015. data was collected from the department of gynecology and obstetrics of government hospitals i.e.shaikh zayed hospital, jinnah hospital, general hospital, mayo hospital, lady wallington and sir ganga ram hospital. formal ethical approval was taken from ethical review board of shaikh zayed hospital. a total of 255 participants were recruited in this study using consecutive sampling technique. sample size was calculated by keeping desired precision at 0.05, confidence level of 95%, for a population size of 750 and estimated proportion 0.53 of domestic violence.15 all women who had delivered babies within last 2 days in obstetric units of selected tertiary care hospitals were included in the study. a self-constructed, structured questionnaire was administered and filled after verbal informed consent of the participant in front of two witnesses. questionnaire was piloted and cronbach’s alpha was run for measuring internal consistency of the tool which came out to be 0.7. the study approval was taken from irb, shaikh zayed hospital lahore. during this study anonymity and confidentiality of the participant was ensured and no monetary benefit was given. operational definitions: antenatal domestic violence in the study was taken as physical and verbal abuse/violence; antenatal physical violence means slapping, pushing, shoving, hitting, kicking, dragging, beating or throwing something at her or was punched or slapped in the abdomen while she was pregnant by her husband or from someone from the immediate family. antenatal verbal abuse means insulting her, humiliating her, belittling her, scaring her etc. during pregnancy.16 foetal outcome in current study included birth weight (low, normal, over weight), gestational age at birth (pre-term, term, post-term) and still birth. neonates <2.5 kg birth weight are low, while between 2.5 to 4 kg are normal and above 4 kg over weight. before 37 completed gestational weeks are pre-term, 37 completed weeks till 40 completed weeks is term and more than that is post-term. domestic violence is taken as an independent variable while foetal outcome as dependent. all the concerned socio-demographic variables are given in table-1. statistical analysis: data was collected and entered in spss version 22.0. frequencies and percentages of various variables were determined. chi-square test was applied to determine association of antenatal domestic violence with various socio-demographic variables and fetal outcomes. p value of <0.05 was considered statistically significant. 42 antenatal domestic violence & fetal outcome: a cross-sectional study results 68% (173/255) of participants belonged to age group of 20 to 30 years while 32% (82/255) belonged to age group of 31 to 40 years. 78% (200/255) were from rural area (as most of the patients that visited our government sector hospitals belonged to peripheral areas, therefore those patients were considered in rural setting). 60% (155/255) were illiterate. 58% (150/255) had selfowned houses while 42% lived in rented houses. 18% (45/255) of the females underwent svd while 82% had caesarian section. 22% (55/255) of females experienced antenatal domestic violence during current pregnancy while 78% did not. as fetal outcome was in terms of birth weight, it was observed that 74% (190/255) of the babies had normal birth weight while 26% had low birth weight (table-1). 22% (55/255) females experienced antenatal domestic violence during current pregnancy and out of 55, 72% faced verbal abuse while 27% experienced physical violence. in total 65/255 (26%) were low birth weight neonates and out of them 58.18% (32/55) were born to mothers facing violence during pregnancy. verbal and physical abuse by husband were found to be most common with 49% (27/55) and 18% (10/55) respectively (table-3). association of age, residence, education, type of housing and type of delivery were assessed with antenatal violence which was non-significant (table-2), while a p-value of 0.001 showed a significant association of antenatal domestic violence with birth weight of the neonate (table-4). socio-demographic variables frequency (n=255) % age of mother 20-30 years 173 68% 31-40 years 82 32% locality rural 200 78% urban 55 22% educational level illiterate 155 60% matric and above 100 40% type of housing own 150 58% rent 100 42% antenatal domestic violence faced during married life yes 80 32% no 175 68% antenatal domestic violence faced during current pregnancy yes 55 22% no 200 78% type of delivery svd 45 18% cs 210 82% birth weight of the baby lbw (<2.5 kg) 65 26% normal weight (>2.5 kg) 190 74% table-1: antenatal domestic violence according to sociodemographic variables variables categories antenatal domestic violence chi’s square test. present not present p-value age 20-30 years 37 136 0.918 31-40 years 18 64 residence rural 43 157 0.959 urban 12 43 education uneducated 35 120 0.990 educated 22 78 housing own 33 122 0.913 rent 22 78 type of delivery svd 10 35 0.906 table-2: association of socio-demographic variables with antenatal domestic violence type of abuse frequency (n=55) % physical abuse by husband 10 18% by husbands’ family 5 9% verbal abuse by husband 27 49% by husbands’ family 13 23% table-3: type of abuse during pregnancy domestic violence birth weight of the baby p-value < 2.5 kg > 2.5kg yes 32 158 0.001 no 23 42 table-4: association of antenatal domestic violence with birth weight of the baby discussion the results of this study showed that twenty-two percent of women were victims of antenatal domestic violence during their pregnancy. results of a study conducted in turkey showed that 3.2% of females faced antenatal domestic violence during pregnancy.17 another study conducted in ethiopia showed that approximately 23% of women experienced ipv during their pregnancy. association of ipv with low birth weight was also found to be significant.18 results of a systematic review and meta-analysis conducted showed that about 23 % of women experienced domestic violence including ipv during pregnancy. the results also indicated an association of ipv with lbw.19 global incidence of ipv is 30%, in some countries it can be as high as 38%.13 thus, it can be said that women from different regions of the world faced violence during their pregnancies and the frequency is also approximately similar in developing countries. results of another national 43 antenatal domestic violence & fetal outcome: a cross-sectional study cross sectional study conducted in pakistan showed that 37.9% of participants reported experiencing ipv, a significant association was found between ipv, unintended pregnancy and pregnancy loss.20 the frequency of women who have suffered from violence is even greater than the frequency reported in our study. this variation in frequency might be due to the fact that it was a national level study as compared to this hospital based study. an analysis conducted on 19 countries concluded that ipv during pregnancy is commonly encountered and is more common than other maternal health conditions which are screened during antenatal. in our study verbal and physical abuse by husband were found to be most common 49% and 18% respectively, whereas physical and verbal abuse by husband family was 9% and 23% respectively. domestic violence including ipv during pregnancy is associated with various negative consequences such as decreased infant birth weight and increased risk of premature births. studies conducted in 2002, 2004 and 2008 signified a strong association of low birth weight of the neonate with antenatal domestic violence (ipv).21,22,23 these results are consistent with the findings of our study. our study found domestic violence association with low birth weight of the new born with a pvalue of <0.001. low birth weight infants and preterm births are main causes of neonatal morbidity and mortality. gestational age of the new-born and birth weight at delivery are considered to be the strongest predictor variables of health outcomes of the child. in extremely premature and low birth weight infants cognitive defects, delayed motor development, behavioral and psychological problems are very common24. domestic violence including ipv was found to be significantly associated with lbw and preterm birth.25 it is said that violence against women is perhaps one of the most shameful human rights violation. millennium development goals (mdg) 3,4,5& 6 could not be achieved without empowering women. antenatal domestic violence can be labelled as a security issue. in spite of recognition of this issue, preventive strategies still remain inadequate.26 a study conducted in pakistan highlights the magnitude of violence and its association with reproductive health of women.27 results of a study conducted in iran showed that the women with a history of violence during antenatal period had 1.9 fold higher risk of rupture of membranes and 2.9 fold higher risk of low birth weight as compared to women who did not face any violence.28 incidence of psychological symptoms such as depression, anxiety are significantly associated with antenatal domestic violence.29 however, in 2011, a study conducted in karachi, pakistan significantly reports high prevalence (56 to 57%) of physical violence14 which has now considerably been reduced as shown by other studies (but somewhat still present) and probable reasons may be better education and awareness toward women rights and empowerment. ipv as a part of domestic violence is considered to be a strongly associated with abortion in different settings. pregnancy terminated through unsafe method often leads to death of the mother or development of serious complications. therefore, reducing incidence of ipv can significantly reduce the worst outcomes related to maternal health.30 ipv as a part of domestic violence is found to be significantly associated with unintended pregnancies and unfavorable maternal outcomes such as depression, distress, poor antenatal care, bleeding from vagina, abortion etc. and in infants it can lead to premature birth, low birth weight and development of other complications etc.31 as birth weight is a reliable variable of measuring health of the new-born therefore, this study focuses mainly on birth weight of the new-born. our study is limited by the fact that six hospitals were selected based on administrative and geographical convenience. in future a community based survey or a representative multicentre hospital based survey could be helpful in getting a better insight into the burden of problem. another limitation is that the females were approached in hospitals after delivery so we missed abortions as fetal outcome. this is only a tip of the iceberg of domestic violence which we see in tertiary care hospitals. for a true picture we must not miss the females facing severe domestic violence and delivering at home or small unregistered centres. a community based survey can overcome these limitations in future. the antenatal domestic violence reported in this study requires serious attention of policymakers, political allies, and health professionals. global initiatives which focus on reducing maternal mortality and improving health of the mother should give more attention to violence during pregnancy. therefore, it can be suggested that history of antenatal domestic violence should be part of normal antenatal check-up. legal actions are required to outlaw violence against women. focus should be given on male and female education. personnel working in healthcare facilities need training on identification, counseling, management, and prevention of violence against women. involvement of mass media is necessary to 44 antenatal domestic violence & fetal outcome: a cross-sectional study create a debate on women’s empowerment. most importantly, there is a need to return to the islamic notion of the husband as a caretaker, sympathetic head of the household. conclusion antenatal domestic violence is found to be associated with low birth weight of the neonate. antenatal domestic violence is a neglected domain in maternal & child care and implementation of legislation is required to provide complete protection to mothers acknowledgement: we would like to thank dr. shahroze arshad, dr. abdul basit abbasi, dr. qurrat ul ain iqbal for proof reading this article. we also thank dr. muhammad hassan riaz, dr. abeer bin awais and muhammad kiwan akram for their intellectual and statistical input in writing this article. references 1. organization wh. understanding and addressing violence against women: intimate partner violence. who, 2012. 2. flury m, nyberg e. antenatal domestic violence against women: definitions, epidemiology, risk factors and consequences. swiss med wkly. 2010; 140(3536). 3. johnson mp. a typology of domestic violence: intimate terrorism, violent resistance, and situational couple violence: upne; 2010. 4. hassan s, malik aa. factors for intimate partner violence (ipv) in urban pakistani families. pak. j. clin. psychol. 2011; 10(1). 5. aizer a. the gender wage gap and domestic violence. am.econ.rev. 2010; 100(4):1847-59. 6. 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physical abuse during pregnancy and preterm delivery. american journal of obstetrics and gynecology. 2008; 198(2):171. e1-e6. 24. bailey ba. partner violence during pregnancy: prevalence, effects, screening, and management. int. j. women's health. 2010; 2:183. 25. hill a, pallitto c, mccleary-sills j, garcia-moreno c. a systematic review and meta-analysis of intimate partner violence during pregnancy and selected birth outcomes. int. j. gynecol. obstet. 2016; 133(3):269-76. 26. garcia-moreno c, watts c. violence against women: an urgent public health priority. bull world health organ. 2011; 89:2-. 27. zakar r, zakar mz, mikolajczyk r, krämer a. intimate partner violence and its association with women's reproductive health in pakistan. int. j. gynecol. obstet. 2012; 117(1):10-4. 28. abdollahi f, abhari fr, delavar ma, charati jy. physical violence against pregnant women by an intimate partner, and adverse pregnancy outcomes in mazandaran province, iran. j family community medicine. 2015; 22(1):13. 29. howard lm, oram s, galley h, trevillion k, feder g. antenatal domestic violence and perinatal mental disorders: a systematic review and meta-analysis. plos med. 2013; 10(5). 30. pallitto cc, garcía-moreno c, jansen ha, heise l, ellsberg m, watts c, et al. intimate partner violence, abortion, and unintended pregnancy: results from the who multi-country study on women's health and domestic violence. international journal of gynecology & obstetrics. 2013; 120(1):3-9. 31. han a, stewart de. maternal and fetal outcomes of intimate partner violence associated with pregnancy in the latin american and caribbean region. international journal of gynecology & obstetrics. 2014; 124(1):6-11. the authors: dr. sohaib ashraf research associate, wellman centre of photomedicine, massachusetts general hospital, harvard medical school, boston, usa. dr. muhammad ahmad imran senior house officer, department of microbiology, shaikh zayed medical complex, lahore. dr. hina mahmood visiting faculty, dept. of public health & community medicine, shaikh zayed medical complex, lahore. dr. khawar nawaz resident pgr-1, department of paediatrics, university hospital of brooklyn sunny downstate medical centre, new york. dr. tayyab mughal post graduate resident, department of radiology, shaikh zayed medical complex, lahore. dr. umair hafeez sheikh post graduate resident, department of ent, shaikh zayed medical complex, lahore dr. hassan mujtaba cheema chairman, amna inayat medical and educational complex, lahore. prof. ayesha humayun hod, public health & community medicine, shaikh zayed medical complex, lahore. corresponding author: dr. sohaib ashraf research associate, wellman centre of photomedicine, massachusetts general hospital, harvard medical school, boston, usa. e-mail: sohaib-ashraf@outlook.com numbering.indd 30 proceedings s.z.m.c. vol: 34(4): pp. 30-34, 2020. pszmc-767-34-4-2020 nephroprotective evaluation of moringa oleifera & beta vulgaris leaves in murine methotrexate induced toxicity 1shumaila kanwel, 1bushra suhail, 2mudassara saqib, 2saadia shahzad alam 1department of pharmacology, university college of medicine & dentistry, lahore 2department of pharmacology, shaikh zayed medical complex, lahore abstract introduction: methotrexate has been used for treatment of multiple diseases. chronic use of this drug can result in oxidative damage resulting in nephrotoxicity. beta vulgaris and moringa oleifera are well known herbs for their antioxidant properties, so they can have potential nephroprotective effect against mtx induced renal damage. aims & objectives: the study was designed to evaluate the therapeutic role of moringa oleifera and beta vulgaris leaves against mtx induced nephrotoxicity in rats. place and duration of study: time duration of our research project was 6 months. it was carried out in pgmi lahore and fpgmi, shaikh zayed medical complex, lahore. material & methods: 5 groups of 9 rats each were made. group1 were maintained as healthy controls while group ii as diseased control groups iii, iv and v were administered daily p.o for 24 days 200mg /kg of ethanolic extract of beta vulgaris( eebv),and 800 mg/kg ethanolic extract of moringa oleifera (eemo) and combination of extracts (100mg/kg eebv+400mg/kg eemo respectively. nephrotoxicity was produced on 21st day via single i/p route injection of 20mg/kg mtx in all groups except group 1. rfts were performed on days 1, 21 and 24th day. nephroprotection was determined by measuring serum urea, creatinine along with histopathological study of kidneys at study end. results: eebv provided significant neproprotection in group 3 which had significantly lower levels of creatinine as compared to group 2 with p-value <0.001. the groups 4 and 5 had nonsignificant difference from group 2 with p-values 0.975 and 0.252. further confirmation by histopathological examination of kidneys revealed group 3 to have improved status of kidney( normal kidney epithelium, normal tubules, mild congestion, mild necrosis) as compared to other groups with (distorted kidney epithelium ,dilated tubules ,moderate to severe congestion and necrosis). conclusion: nephroprotective effect was shown by beta vulgaris plant while moringa oleifera plant extracts remained unable to show their protection against mtx induced nephrotoxicity. key words: ethanolic extract of beta vulgaris (eebv), methotrexate, ethanolic extract of moringa oleifera (eemo), renal function tests (rfts). introduction the kidneys are important organs with many functions in the body including metabolism, body water and salts regulation, acid base regulation and waste products excretion along with excretion of hormones. therefore, they are often at risk to damage caused by infections, harmful substances, prescribed and otc drugs.1 many drugs are responsible for acute nephrotoxicity. 20 % of all acute renal failure cases are caused by drugs.2 most drugs cause nephrotoxicity through different pathogenic mechanisms including, tubular cell damage, inflammation, crystal nephropathy. commonly responsible drugs are nsaids, aminoglycosides, ace inhibitors and amphotericin b.3 methotrexate is a folic acid antagonist. it has multiple uses like blood dyscrasias, inflammatory diseases of joints, skin disorder and inflammatory bowel disease. toxic dose and prolonged use of methotrexate can lead to nephrotoxicity. despite all advantages with mtx, their extensive use is still reduced due to their adverse effects. leucovorin, being antidote for methotrexate toxicities can save rapidly dividing cells only like gastrointestinal and hematopoietic cells. however, it is still unable to give protection against other adverse effects of mtx.4 oxidative stress is responsible for mtx induced toxicities within kidney. the main reason behind this toxicity is low nadph, which is essential for glutathione synthesis. ultimately cells are more exposed to ros (reactive oxygen species).5 31 nephroprotective evaluation of moringa oleifera & beta vulgaris leaves in murine methotrexate induced toxicity different plants have shown protective effect against different toxicities. swiss chard (variety of bv) has shown nephroprotective effect in diabetic rats.6 moringa oleifera has proven to have nephroprotective activity against dmba (7,12-dimethylbenz-anthracene) induced renal carcinogenesis.7 taking into account the protecting effect of different herbs, moringa oleifera and beta vulgaris were chosen. beta vulgaris leaves are useful as tonic, diuretic, anti-inflammatory.8 moringa oleifera is another herb, which has multiple uses like git diseases, diabetes mellitus and heart disorders.9 initial phytochemical analysis has shown that leaf extracts of moringa oleifera and beta vulgaris contain various phytochemicals like tannins, flavonoids, alkaloids.8,9 being known antioxidants, these active principles may have protective role in management of mtx induced toxicities. so, the purpose of our study was to evaluate protective effect of these plants. material and methods plant materials: authenticity of leaves of moringa oleifera and beta vulgaris was done by botany department of punjab university lahore. ethanolic extracts of these herbs were made in pcsir (pakistan council of scientific & industrial research) lahore. the powdered form of beta vulgaris leaves were extracted with 90% ethanol using soxhlet apparatus. the material left after extraction was macerated with distilled water for 24 hours. solvents were distilled off under reduced pressure and temperature to get greenish paste of eebv.8 absolute alcohol was mixed with powder form of moringa oleifera leaves extract at room temperature. after evaporation of concentrated form, greenish paste was obtained.10 animals: 45 rats weighing 150-200g were kept in iron cages under controlled room temperature (25±10 ◦c). rats were provided with typical lab diet. settings: animal house & research laboratory of pgmi lahore. grouping: rats were randomly divided into groups of 9 animals. extracts of moringa oleifera, beta vulgaris or their combination were administered daily orally throughout entire study period of 24 days as per group designation. on 21st day single injection of methotrexate was given through i/p route in all groups except group 1 to induce toxicity. . group 1 (negative, healthy control): the animals in this group were administered water only. group 2 (positive control): single injection of mtx 20mg/kg i/p was administered in rats of this group on 21st day to induce nephrotoxicity. group 3 (test): beta vulgaris leaf ethanolic extract of 200 mg/kg/p/o group 4 (test): ethanolic extract of moringa oleifera’s leaves in dose of 800mg/kg p/o. group 5 (test): combined eebv 400 mg/kg and eebv 100 mg/kg po. biochemical parameters: on day 1 and 21, blood for analysis of biochemical parameters was collected from tail of rats. 3 cc syringes were used for collection of blood by intracardiac puncture on 24th day, rfts were analyzed on chemistry analyzer in biochemistry department of sheikh zayed hospital, lahore. histopathology of kidneys: on 24th day animals were sacrificed and kidney samples was sent for histopathological examination kidneys tissues samples were isolated after dissection, stained with haematoxylin and eosin .slides were prepared and examined under microscope. statistical analysis: data was evaluated by using spss (statistical package for social sciences) 20.0. renal parameters like urea and creatinine were elaborated by using mean ±s.d. anova was applied for comparison among groups. p value<0.001 was considered significant for all parameters. results the mean urea level for group 1 was 21.6±6.0 mg/dl, the group 2 had levels 91.6±14.9mg/dl and group 3, 4 and 5 had their mean urea levels at 98.1±13.0mg/dl, 81.1±18.6mg/dl and 91.3±15.3mg/dl respectively here the group, 2, 3, 4 and 5 all had significantly higher levels as compared to group1 with p-values <0.001. the group 2 had nonsignificant difference from group 3, 4 and 5 with p-values 0.863, 0.530 and 1.000 respectively. the group 3 and 4 had an nonsignificant difference with p-value 0.102 and other differences between experimental groups were also nonsignificant (fig-a). group 1 had mean creatinine level of 0.39±0.11mg/dl which was lowest and group 5 had 0.87±0.05mg/dl which was highest. the group 2, 3 and 4 had creatinine levels of 0.79±.08mg/dl, 0.50±0.07mg/dl and 0.81±0.08 mg/dl. group 1 had significantly lower creatinine levels as compared to all other groups with p-values <0.001. 32 nephroprotective evaluation of moringa oleifera & beta vulgaris leaves in murine methotrexate induced toxicity the group 4 and 5 had nonsignificant difference from group 2 with p-values 0.975 and 0.252 but group 3 had significantly lower levels as compared to group 2 with p-value <0.001. the group 3 also had significantly lower levels as compared to group 4 and 5 with p-values <0.001. the difference between group 4 and 5 was nonsignificant with pvalue 0.581 (fig-b). histopathological study of kidneys in group 2 showed detached epithelium, dilated tubules, congestion and necrosis ,showing damage of kidney by mtx (fig 2),as compared to group 1(normal group) similar damage was observed in groups 4 and 5 (fig-4,5) while group 3 showed improved kidney status as compared to other groups (fig-3) figa: mean urea levels of five study groups at 24th day fig-b: mean creatinine level in five study groups at 24th day fig-1: showing normal histology of kidney in group 1 (negative control) a. normal kidney epithelium b. no congestion, c. no tubular dilatation d. no necrosis fig-2: showing kidney damage by mtx in group 2 a. distorted kidney epithelium b. dilated tubules, c. congestion d. necrosis fig-3: showing protective effect on kidney tissue in group 3 received extract of beta vulgaris showing a. normal kidney epithelium b. normal tubules c. mild congestion d. mild necrosis 33 nephroprotective evaluation of moringa oleifera & beta vulgaris leaves in murine methotrexate induced toxicity fig-4: showing damage of kidney in group 4 received extract of mo a. distorted kidney epithelium b. dilated tubules, c. moderate to severe congestion d. necrosis fig-5: showing damage of kidney in group 5 received combined extract of bv and mo a. distorted epithelium b. dilated tubules c. moderate to severe congestion d. necrosis discussion drugs like mtx can affect multiple organs, it is employed for management of various diseases like inflammatory diseases of joints, git, blood cancers and skin diseases. short and long-term use of drug has caused many adverse effects, which have been observed in many studies.4 mtx can affect multiple organs e.g bone marrow, git mucosal cells and hair follicles. leucovorin being mtx antidote cannot rescue all adverse effects.11 phytotherapy is attaining increasing consideration. moringa oleifera and beta vulgaris herbs exhibited variety of active principles like flavonoids, alkaloids, saponins, tannins. these plants were used for the treatment of multiple diseases.8,9 main purpose of our research was to appraise these plants for their nephroprotective activity against mtx in rats. at days 1 and 21 renal function test revealed values within normal range in comparison to group 1. this reflected safety of these herbal drugs if used as future potential medicines. in our project, we have found that group 2 (mtx) has shown (405% & 133% increase of serum urea and creatinine in rats, reflecting lethal effects of mtx on kidney. results were as expected.12 histopathological examination of kidney samples from group 2 showed (kidney architecture damage, congestion, necrosis, tubular dilatation) in comparison to group 1 (fig-2). previous studies also demonstrated same results.13 at end of our project, we have observed that group 2 (mtx) showed (333% increase in urea level in comparison to group 1) and when we compared it with urea levels of groups 3, 4 and 5, we found out that there was no significance difference from urea levels (366%, 285%, 333%) observed in groups 3, 4 and 5, showing our plant extracts remained unsuccessful in reverting nephrotoxicity induced by mtx. however urea is a non-specific test renal functions.15 its level can also be raised because of other causes like dehydration16, heart failure17, git bleeding 18 so it is not a confirmatory indicator of failure of nephroprotection of our plant extracts. at the end of study, mean creatinine level of group 1 was lowest 0.39±0.11 and group 5 had highest level 0.87±0.05. nephrotoxicity was not reversed in groups 4 and 5 in terms of increase of creatinine levels (166% and 167%) as compared to group 2(mtx) which has shown 133% increase in creatinine level in comparison to group 1. however group 3 showed 66% elevation of creatinine level which was less as compared to group 2 (133%), reflecting nephroprotective effect of this extract. nephroprotective effect of beet root extract against gentamicin induced nephrotoxicity seen in this study also favors our results14. as we detected in our project, nephrotoxicity was not reversed in two groups, 4 (eemo) and 5 (eebv+eemo), rather it came out to be worst, studies in past exposed that when mtx is co administered with certain drugs, their interactions might increase toxicity of mtx by competing with mtx for renal tubular secretion.19 this mechanism might elucidate failure of nephrotoxicity reversal in our study. histological evaluation further supported our result, where status of histopathological changes (kidney epithelium, tubules, congestion, necrosis) was better in group 3 (eebv) as compared to groups 4 and 5 (fig-3,4,5) with (distorted kidney epithelium, dilated tubules,moderate to severe congestion and necrosis). 34 nephroprotective evaluation of moringa oleifera & beta vulgaris leaves in murine methotrexate induced toxicity conclusion nephroprotective effect was shown by beta vulgaris plant while mo plant extracts remained unable to show their protection against mtx induced nephrotoxicity. this parameter should be further evaluated. references 1. kumar v, abbas a.k, fausto n, mitchell rn. robbins basic pathology: 10thed. delhi: elsevier 2017. 2. shahrbaf fg, assadi f. drug induced renal disorder.2015; 4(3):57-60. 3. noilin td, himmelfarb j. mechanism of drug induced nephrotoxicity. handb exp pharmacol 2010; (196):111-30. 4. harvey ra, champe pc and william l. lippincott illustrated reviews pharmacology. 6th ed delhi: wolters kuwer 2015; 592-4. 5. sener g, demiralpe e, cetiner m, erean f, yagen b. beta glucan ameliorates mtx induced oxidative organ injury via its antioxidants and immunomodulatory effect. european j of pharma 2006; 542(13):170-8. 6. bolkent s, yanardaq r, saren o bulan k. effect of chard on kidney tissue, serum urea and creatinine levels of diabatic rat.2002;16(8):758-61 7. paliwal r, sharma v, pracheta, sharma s, yadev s. anti nephrotoxic effect of administration of dmba induced renal carcinogenesis in swiss albino mice. bio medicine. 2011; 3(2):27-33. 8. jain n, singhai ak. protective role of beta vulgaris leaves extract and fractions on ethanol mediated hepatotoxicity. acta poloniae pharma drug res 2012; (69):945-50. 9. mehta j, shukla a, bukhariya v, charde r. the magic remedy of moringa oleifera: an overview. international j bio and advanr.2011;2(6):215-27 10. yousaf fm, khattab hr, sindhi ha. effectiveness of moringa oleifera leaves extract against methotrexate induced acute hepatotoxicity in male rats. international j pharm. 2018; (14):1029-1037. 11. b g ketzung, susan b, anthony j: 14th ed. new dehli 2017; 957-58. 12. kintzel pe, anticancer drug induced kidney disorder 2010; 24(1):19-38. 13. asci h, ozer m.k, calapoglu m, savran m, once m. effect of misoprostol on methotrexate induced hepatic and renal damage. j bio life sci 2011; 2(1):32-7. 14. elgamal a, alsaid m, raish, sohaibani ma, h. beet root ameliorates gentamicin induced nephrotoxicity associated oxidative stress, inflammation and apoptosis in rodent model 2014:p12. 15. higgins c. urea and the clinical value of measuring blood urea concentration.2016:p1-6 16. witting md. ed predictors of upper gastrointestinal tract bleeding in patients without hematemesis. am j emerg med 2006; 24(3):280-85. 17. aronson d. elevated blood urea nitrogen level as a predictor of mortality in patients admitted for decompensated heart failure. the am j med 2004; 116(7): 466-73. 18. mehta ar. why does the plasma urea concentration increase in acute dehydration? advances in physiology education. 2008; 32(4):336. 19. brigitte c, widermann p, c adamson. understanding and managing methotrexate induced nephrotoxicities 2006; 11(6):694-703. the authors: dr. shumaila kanwel assistant professor, department of pharmacology, ucmd, university of lahore. dr. bushra suhail assistant professor, department of pharmacology, ucmd, university of lahore. dr. mudassara saqib associate professor, department of pharmacology & therapeutics, shaikh zayed medical complex, lahore. prof. saadia shahzad alam head,department of pharmacology & therapeutics, shaikh zayed medical complex, lahore. corresponding author: dr. shumaila kanwel assistant professor, department of pharmacology, ucmd, university of lahore. e-mail: dr.shumaila8@gmail.com proceeding.indd dedicated to the valiant health care warriors who gave their lives battling the covid 19 pandemic bismillah hir rehman nir raheem. in the name of allah, the most beneficent and the most merciful message by the chairman shaikh zayed medical complex & chief editor of proceedings prof. mateen izhar prof. saadia s. alam (chairman & dean) (chief editor) as the covid 19 pandemic rages on, we steadfastly publish a diverse april to june 2020 proceedings with international and indigenous, basic and applied research on varied aspects of health care, therapeutics and diagnostics. all articles submitted for publication have been judged on stringent 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proceedings shaikh zayed medical complex vol. 34 (3) pcpb/24(94) p-121/3196 jul to sep 2020 editorial board patron in chief: prof. mateen izhar chairman & dean chief editor: prof. saadia shahzad alam head, pharmacology & therapeutics external associate editor: dr. usman iqbal taipei medical university, taiwan associate editors: prof. ayesha humayun head, public health & community medicine prof. imran anwar head, surgery unit-ii dr. adnan salim assistant prof. gastroenterology sub editors: dr. uzma nasim siddiqui assistant prof. medicine dr. shahila jalil associate prof. histopathology prof. tahira naseem head, biochemistry & chemical pathology dr. samira haque associate prof. histopathology dr. lubna riaz assistant prof. pediatrics dr. sadia maqsood assistant prof. pharmacology dr. faraz bokhari associate prof. physiology dr.noora hassan hezam al-aqmer physiology department mrs. saima mohsin incharge, nhrc consultant biostatician: dr. akram chaudhry mr. muhammad aasim library & 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postoperative complications in carcinoma gall bladder:a tertiary care hospital experience shehzad anwer, shuja uddin, hasnain khalid, rizwan ul haq, adeel hamid, mubashir farhan, zia uddin, mian muhammad haneef case report: isolated trochlear fracture of the humerus alvia batool, faizia batool, afifa waseem, saira munawar, maryam fatima an indigenous study on thepalmaris longus muscle anatomical variations hadia zulfiqar, muhammad suhail, amna rehman, afifa waseem, faeza rauf, hafiza sadiaahmed emblica officinalis reduces copper mediated inflammation and preserves liver morphology in the murine model rashid ali, sardar sabeel, danial zahoor, bushra arif, adnan khan, saqib malik mental health impact on patients of esrd on renal replacement therapy: a cross sectional survey using beck and deck inventory shazo sana, hafiz muhammad umair, nagmana lateef, tahir ayub, javeria zafar, hina pervaiz perception of e-learning in medical students of fatima jinnah medical university muhammad atique 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(national) prof. m. nauman ahmadffarcs prof. naseem saudphd prof. m. ovais omer phd prof. shabbir bhatti phd prof. zahidniazfrcs prof. zamir ahmadphd prof. muhammad arif nadeem fcps prof. muhammad moinfrcs prof. khalid mahmood phd prof. sadia chiraghm.phil prof. muhammad aslamfcps prof. abdul mannan m.s prof. naheedhumayunfcps prof. farhat naz fcps prof. abdul hameed frcs prof. rafeeatafweezphd prof. inayatthaverphd peer review board (national) prof. ashraf chaudhryfcps prof. kashifmalikfcps prof. aftabturabiphd prof. uzmahussainfcps prof. muhammad pervaizfrcs prof. fouzia shaukat fcps prof. maryam rashidphd prof. rabeia bilal phd prof. khwaja khursheedfcps prof. tanvir akhtar butt ms prof. naila akhtar fcps prof. abidashaheenphd prof. sajida malik m.phil prof. rumina hasan phd prof.khalid javeed khan frcs dr. farooq afzal frcs dr. samanshahidphd dr. muhammad shahzadphd dr. ali hussainyzaidim.d dr. shahzad khuramakramphd dr.fahad azam phd dr. khadija irfan fcps dr. uzmamalikfcps dr. ayesha mallick frcp dr. tehseenhaiderkazmifcps dr. khalidaajmalm.phil dr. israr ahmedfcps dr. saimabatoolfcps dr. saleem muhammad ranaphd dr. muhammad khurram habibfcps dr. saba riaz phd dr. raaziatasadduqphd dr. nageen hussain phd dr. saadia nosheen jan fcps dr. soumble zulfiqar phd dr. yasir abbas zaidi fcps dr. muhammad zeeshan sarwarmrcs dr. muhammad imran khokarmrcs dr. nabihafarasatm.phil dr. tayyaba muzaffar m.phil dr. zaeem khalid fcps dr. nafeesah fatimah fcps dr. athar mahmood fcps dr. fariha sahrish fcps dr. tehreemzulfiqar m.phil dr,fatima chaudhry m.phil dr. saifullah khan niazi fcps dr. bilal saeed fcps dr. chetan lal m.phil dr. amna mudassarfcps dr. riffat saeed fcps peer review board (international) prof. renne koeffel phd(switzerland) prof. jane banaszak-holl phd(aus) prof. m. hisham al-muhtaseb phd (jordan) prof. mukhtiar baig phd(ksa) dr. sarwatshaheenm.d(usa) dr. zeeshan i. shaikh m.d (usa) dr. tanzeelquratijazmrcp (uk) dr. hafiz sohail anjum fcps(ireland) dr. rafayazharfrcp (singapore) dr. mazhar nawaz m.d (usa) dr. ahmad azam malik phd (ksa) dr. ishtiaq ahmed fcps (ksa) dr. aamir omairphd (ksa) dr. sabina ahmed mir m.d (usa) dr. anita lamichhanem.d (nepal) dr. faiza durraniphd (uk) dr. waseemlodhifrcog (uk) dr. seerat zahra hammadm.s(ksa) dr. shafya shahid phd (u.s) dr. abdul waheedfrcs (u.k) dr. humaira zareenfcps (ksa) dr. amira okudm.d (ksa) dr. asma ahmed mrcp (uk) dr. sabahat sabir mrcog (uk) dr. sarfaraz ahmad frcr (uk) dr. zouina sarfaraz m.d (uk) dr. humaira zaman fcps (uk) numbering.indd 35 proceedings s.z.m.c. vol: 34(4): pp. 35-39, 2020. pszmc-768-34-4-2020 perceived stress of pakistani parents having normal children aged under ten visiting a tertiary care hospital in lahore, pakistan 1saadia shahzad, 2rosheen zahid, 3bushra rehman, 4tahseen haider kazmi 1department of community medicine, shalamar medical and dental college, lahore 2department of surgery, shalamar medical and dental college, lahore 3department of ent, shalamar medical and dental college, lahore 4department of community medicine, shalamar medical and dental college, lahore abstract introduction: having children changes the dynamics of family functioning and parenting role can become a source of stress for parents, even if their child is following a normal growth pattern without any unusual problem. a maximal number of studies in our setup have been conducted to assess stress levels in parents who have differently abled (special needs) children. this study, in contrast, will assess the perceived stress level in parents who have normal children following a normal growth pattern. aims & objectives: 1) to determine perceived stress levels in parents having normal children ≤10 years of age. 2) to identify the association of perceived stress levels in parents with socio-demographic factors. place and duration of study: shalamar hospital, dec 18, 2019-mar 19, 2020. material & methods: a cross-sectional study was conducted in one tertiary level hospital in lahore. the study subjects were parents with normal children ≤10 years of age. purposive sampling technique was applied to recruit the subjects. study variables were: a) sociodemographic b) variables of perceived stress among parents. data was collected upon the perceived stress scale (pss), and a structured socio-demographic performa. data was analyzed using spss 20, with perceived stress as the outcome. descriptive statistics were calculated, while the independent samples t-test was used to test differences between pss scores and characteristics of the family. multivariable logistic regression model was applied for studying associations of perceived stress with socio-demographic characteristics. results: from a total of 390 respondents, the mean parental perceived stress score was 41.59±8.27. among these, 32.6% were categorized in low stress, 50.8% moderate, and 16.6% in high stress category. a significant association was found between perceived stress and the working status of the mother (pvalue=0.011), and with the father’s monthly income (p-value=0.000). the independent sample t-test showed significant differences between pss total score and structural types of the family (p-value=0.001). conclusion: the development of a child with parents’ positive mental health is important, and parental stress compromises the healthy growth of children. the current results warrant further study at a community level with a representative sample to have a more in-depth analysis of the application of some intervention at an early stage. this will assist in minimizing parental stress and improve parenting quality. keywords: perceived parental stress, ≤10-year-old, typical children introduction typical/ normal by the dictionary means; being of the same sort or kind that is conventionally expected as usual. typical growth of a child or typical child thus implies that he/she is showing all important traits of a type, class, or group. similarly, the normal growth of a child or a normal child implies that he/she lacks deviation from the usual/ expected conventional norms.1 parenting stress according to literature refers to the intensity and relative magnitude of stresses that exist in the parent-child relationship.2 literature search has revealed the fact that having children naturally changes the dynamics of family function. thus parental role could be a source of stress for many parents.3 many studies have revealed that the demands and responsibilities that are associated with raising and caring for children are stressful for both parents.4 studies also reveal that both mothers and fathers have reported perceiving stress differently.5,6 36 perceived stress of pakistani parents having normal children aged under ten visiting a tertiary care hospital literature shows that techniques of coping and social support could help people to face stress. however, it was found that gender affected how people evaluate, perceive, and respond to stressors.7,8 literature shows that mothers as compared to fathers perceive more parenting stress and depression. although over the period the rate of father’s involvement in parenting has increased; mothers still have the major part of parenting thus feel more stressed out. especially mothers with fiveyear-old children get distressed upon their child’s school adjustment shortly .2 most reviewed research articles that investigated parental stress when parents have children with some congenital disorder e.g. autism, developmental delays, cerebral palsy, etc.9,10,11 however, very few studies have examined the stress level in parents having normal children. parental stress prevalence in parents having children with autism was found to be 47.4% in fathers, and 52.6% in mothers.12 another study conducted in jordan to assess parental stress level among parents with normal children have found a significant difference in stress levels between fathers and mothers; with mothers having significantly higher scores. the mean stress score for mothers was 17.94 ±6.41, compared with 15.13 ±7.12 for fathers.13 in a study various factors were reported to be predictors of parental stress like disability in a child, gender of a parent, socioeconomic status, number of children, years married, age of parents and age of the child, and parent’s social support network.14 researchers believe that parental stress is assumed to be at a higher side in developing countries like pakistan as compared to developed countries. this phenomenon is probably due to various influencing factors that play a key role in this situation like social set up, cultural values, societal norms, family system, socioeconomic status, lack of resources, low literacy rate, lack of awareness regarding good parenting techniques, and overt use of social media by the children. a literature search could not find any such study conducted in pakistan to date. there is very limited information in our setup about the magnitude of parental stress among parents having healthy children <10 years of age. likewise, awareness regarding coping techniques for parental stress in a positive manner is also grossly lacking among parents in our setup. social issues must be looked into in detail to have a better quality of life. moreover, this study is very significant as to the maximum knowledge of the researcher this study is the first of its kind conducted in our pakistani society; as most of the studies had been done to assess stress among parents with physically challenged children. the fact that stress among parents who are raising normal children must be taken into account as it affects their mental health as well as their parenting quality. objectives of the study are 1) to determine perceived stress levels in parents having typical children ≤10 years of age 2) to identify the association of perceived stress levels in parents with socio-demographic factors. material and methods it was a cross-sectional study conducted in the pediatric out-patient department of the shalamar hospital, lahore; in three months (dec 18, 2019mar 19, 2020). the sample size of 390 was calculated using open epi software with an assumed prevalence of 50%, &a margin of error of 5%. our study subjects were parents visiting the outpatient department; &purposive sampling technique was applied to select the study participants. the inclusion criteria were: 1) willingness of the respondent parents 2) parents having normal children ≤10 years of age. the exclusion criteria: 1) parents having a child with any congenital abnormality. study variables were: a) sociodemographic variables of parents b) variables of parental stress. data was collected upon the questionnaire after taking informed consent from the respondent. to measure parental stress, the perceived stress scale (pss) from the state of new hampshire employee assistance program was used; which is available in free access at info@mindgarden.com. cut-offs for the pss were 0-13 considered as a lowstress category, 14-26 as moderate stress, and 27-40 considered as a high-stress category. the total score for perceived stress was taken as an outcome. all ethical considerations were duly considered and observed during the study; and participants were assured of the confidentiality of the data. institutional review board certification was acquired from the ethical board of the shalamar institute of health sciences, lahore, before the start of the study. statistical analysis: descriptive stats, mean± sd, frequency trends, & percentages for categorical factors were calculated. multivariable logistic regression was applied to determine the association of perceived stress with sociodemographic factors, comparing moderate to high stress against the low-stress category. independent samples t-test was used to test 37 perceived stress of pakistani parents having normal children aged under ten visiting a tertiary care hospital differences between pss scores and characteristics of the family; & p-value of<0.05 was considered significant. results data of 390 parents were entered and analyzed via spss 20. there were 59 (15.1%) fathers and 331 (84.9%) mothers in the research. the mean age was 28.99 ± 4.55. the mean stress score for respondents was 18.03 ±8.8. table-1 shows the socio-demographic profile of the recruited subjects. the majority of the respondents were educated, with about 40% having an education of intermediate and above. most respondents reported having less than 4 children in their household. more than 70% of subjects had their family income less than pkr 50,000. s. no variable n (%) 1. father education class 1-10 intermediate and above 240(61.5%) 150(38.4%) 2. mother education class 1-10 intermediate and above 233(59.8%) 157(40.2%) 3. number of children 1-3 >3 343(88%) 47(12%) 4. type of family structure nuclear joint 102(26.2%) 288(73.8%) 5. house status rented own 86(22.1%) 304(77.9%) 6. monthly income of the father up to pkr 50000 > pkr 50000 285(73.07%) 97(24.87%) table-1: socio-demographic profile of the respondents (n=390) fig-1: categorization of perceived stress among both parents (n=390) fig-1 shows the categorization of stress scores among responding parents. the figure shows that 49% of respondents had moderate stress levels, followed by 33% in the low category. high-stress scores were obtained from among 18% of the subjects. table-2 shows the logistic regression analysis of stress with family characteristics. the working status of the mother and the family’s monthly income was significantly associated with perceived stress in the final adjusted model; the working status of mothers increases stress by 4 times [or 4.12 (95% ci: 1.39-12.25)]. variables univariate analysis or(95% ci) p-value multivariate analysis or(95%ci) p-value total number of children 1.250 (1.019-1.533) 0.033 * 1.218 (0.978-1.516) 0.078 house type own: rented: 1 1.730 (1.003-2.984) 0.049* 1.380 (0.777-2.451) 0.272 status of mother house wife: working: 1 4.018 (1.537-10.500) 0.005* 4.120 (1.386-12.248) 0.011 * frequent job changes by father no: yes: 1 3.072 (1.039-9.079) 0.042* 1.578(0.501-4.974) 0.436 monthly income 0.626 (0.505-0.777) 0.000 * 0.653 (0.524-0.815) <.001 * age of mother 0.977 (0.933-1.023) 0.326 age of father 0.991 (0.954-1.030) 0.651 gender male: female: 1 1.456 (0.824-2.570) 0.196 familytype joint: nuclear: 1 0.736 (0.449-1.205) 0.223 table-2: regression analysis of variables related to perceived stress among parents (n=390) table-3 shows the analysis of the differences of pss scores with family structure type and gender. this shows that scores for joint family structures were found to significantly high compared to nuclear families (p-value 0.001). 33% 49% 18% low stress moderete stress high stress 38 perceived stress of pakistani parents having normal children aged under ten visiting a tertiary care hospital variable pss mean score ±sd p-value type of family nuclear: joint: 40.287 43.187 7.75 8.63 0.001* gender male: female: 42.58 41.21 8.28 8.25 0.151 p-value<.05 significant table-3: differences across pss total score with type of family and gender discussion in the past, research defines parental stress as a mismatch between perceptions of environmental demands and personal resources.6 furthermore, previous studies have found that mothers and fathers report perceiving stress differently.5 our study found a higher frequency trend of perceived stress total score among both parents, the maximum range is around 40, and its mean score being 18.03±8.8. categorical breakdown of the perceived stress total score revealed 33% in lowstress category, 49% in moderate, and 18% as high category stress level. this finding is strongly supported by another study that reported mothers had higher stress as compared to fathers.13 literature search revealed that very few numbers studies had been done to study perceived stress among parents having normal children and the majority of studies were conducted to study stress levels among parents having children with special needs. the frequency of engagement with children was found to be related to the mother’s parenting stress as compared to the father’s parenting stress.15 our study also suggests that a greater number of mothers and fathers (40.2% and 38.4% respectively) had a decent education level (intermediate and above); yet they perceived stress scores were high suggesting that their difficulty in coping with demands and resources. the working status of mothers and family income might be the main influencing factors of their incapability towards coping. the total number of children in the family may be another important factor in this context for higher parental stress. this finding is corroborated in other studies as well.16 compromised economic status might contribute further to parental stress and our study findings show that majority of respondents had an income of less than pkr 50,000. this finding is strongly supported by another study that has linked accessibility to socio-economic resources and economic strain to an appraisal of stress.17 contemporary norms of fatherhood emphasize the dual demands of bread-winning plus daily involvement with children. the results of recent qualitative research show that it is difficult for working-class fathers to meet the demands of quality parenting. furthermore, on account of job instability and workplace inflexibility, this may increase.15 additionally, joint family structures can further aggravate the perceived stress among individuals. our study indicates scores to be higher among joint families, however, the multivariable model showed this to be non-significant. there are limitations to this research. foremost, the study being cross-sectional, we cannot suggest causality of the above-mentioned factors with perceived stress among parents. second, the sample size drawn from the target population might not be representative as the subjects were conveniently selected from a single hospital in lahore. but the study does add an important finding that stress is ubiquitous among most families and a study from the community level with an expanded sample size might provide more in-depth knowledge for stress scores among parents. moreover, culturally, the role of grandparents might be another important factor to influence stress among parents that was not explored in this research. literature suggests that grandparents are found to be a likely source of support for many parents, providing emotional, financial, and instrumental assistance.18 future studies could explore that relationship in detail. conclusion the study suggests that parents of normal children are afflicted with moderate to a high category of stress. the working status of mothers and family’s income are significant influencers of perceived stress among parents. to raise children with positive mental health, parents themselves must have a positive and stress-free mental state, and measures are needed to assist such families as much as possible. community counseling is one of the many measures that can be opted for a level of the community, assisting to minimize stress so that parents may provide good quality parenting for their children. references 1. definition of typical/ normal. available at: http://www.merriam-webster.com/dictionary visited on 23.6.19. 2. kim ke, choi jh. sources of parenting stress for mothers and fathers of young children. ijst 2015; 8(35):1-8. 39 perceived stress of pakistani parents having normal children aged under ten visiting a tertiary care hospital 3. soltanifar a, akbarzadeh f, moharreri f, ebrahimi a, mokhber n, et al. comparison of parental stress among mothers and fathers of children with an autistic spectrum disorder in iran. iran j nurs midwifery res 2015; 20(1): 93-98. 4. cohen s, janicki dd, miller ge. psychological stress and disease. jama.2007;298(14):1685-87 5. masa’deh r, collier j, hall c, alhalaiqa f. predictors of the stress of parents of a child with cancer: a jordanian perspective. glob j health sci. 2013; 5(6):81. 6. pelchat d, lefebvre h, levert mj. gender differences and similarities in the experience of parenting a child with a health problem: current state of knowledge. j child health care 2007; 11(2):112-131. 7. lazarus rs. stress and emotion: a new synthesis: new york: springer publishing company. 2006; 342. 8. eaton rj, bradley g. the role of gender and negative affectivity in stressor appraisal and coping selection. int j stress manag 2008; 15(1):94-115. 9. sipal rf, schuengel c, voorman jm, van eck j, becher g. course of behavioral problems of children with cerebral palsy: the role of parental stress and support. child care health dev. 2010; 36(1):74-84. 10. moh ta, magiati i. factors associated with parental stress and satisfaction during the process of diagnosis of children with autism spectrum disorders. res in autism spectrdisord. 2012; 6(1):293-303. 11. neece cl. mindfulness-based stress reduction for parents of young children with developmental delays: implications for parental mental health and child behavior problems. j appl res intellect disabil. 2014; 27(2):174-186. 12. al farsi oa, al farsi ym, al sharbatti mm, al adawi s. stress, anxiety, and depression among parents of children with an autism spectrum disorder in oman: a case-control study. neuropsychiatr dis treat 2016; 12:1943. 13. masa-deh r, bawadi h, saifan a, aburuz m. perceived stress of jordanian parents: a comparative study between mothers and fathers. j nursedupract. 2015; 5(11): 89-95. 14. abiding rr. parenting stress index. charlottesville, va: pediatric psychology press 1990. 15. nomaguchi k, johnson w. parenting stress among low-income and working-class fathers: the role of employment. j fam issues 2016; 37(11): 1535-1557. 16. kins jr, marcenko mo. parenting stress among child welfare involved families: differences by child placement. child youth serv rev 2014; 46:19-27. 17. lewis m, weinraub m. the father’s role in child development. ny: john wiley & sons. 2010: 157-184. 18. barth r. social support networks: informal helping in human services. social support networks in services for adolescents and their families. 1983: 299-330. the authors: dr. saadia shahzad assistant professor, department of community medicine, shalamar medical and dental college, lahore. dr. rosheen zahid house officer, department of surgery, shalamar medical and dental college, lahore. dr. bushra rehman house officer, department of ent, shalamar medical and dental college, lahore. prof. tahseen haider kazmi department of community medicine, shalamar medical and dental college, lahore. corresponding author: dr. saadia shahzad assistant professor, department of community medicine, shalamar medical and dental college, lahore. e-mail: saadiazahur@live.com inner.indd 11 proceedings s.z.m.c. vol: 34(1): pp. 11-16, 2020. pszmc-736-34-1-2020 culture-positivity and sensitivity in post-surgical patients in a tertiary care hospital 1ehsaan ahmed, 2muhammad imran anwar, 1ruqaya idrees, 1sheza arif toor, 1asad mahmood, 1ayesha humayun 1department of public health & community medicine, skzmc, shaikh zayed medical complex, lahore 2department of surgery, shaikh zayed medical complex, lahore abstract introduction: hospital-acquired infection (hai) is a serious health hazard across the world, being a major contributor to the rate of morbidity and mortality. inadequate evidence is available on the magnitude of hospital-acquired infections in tertiary care hospitals of pakistan, thus resulting in a poor focus on this aspect of quality of care. aims & objectives: the study aims to measure the frequency of post-op culture-positivity, sensitivity, and its associated attributes in a tertiary care hospital in lahore, pakistan. place and duration of study: tertiary care teaching hospital in lahore, punjab from april to july, 2016. material & methods: a cross-sectional study was conducted in a public sector, tertiary care, teaching hospital in lahore using secondary data. all discharged patients’ files from surgical and allied specialties, operated from 1st oct, 2015 to 31st dec 2015, having ages between 18 to 60 years and of both gender, were included consecutively in the study. data was collected on a structured, self-constructed form and analyzed through spss version 21. results: out of the total 420 patient files, 47 (11.2%) had culture positive reports, out of which 74.5% had utis, 17% tissue/wound infection, 4.3% fluid infections and 4.3% had other site infections. 46% of the females and 32% of the males were found to be culture positive out of the suspects of hai. the highest infection rate was found among patients of urology (71%), while there was no positive culture found for neurosurgery patients. the most frequently found organisms in cultures were coliform species (38%), pseudomonas aeruginosa (13%), followed by escherichia coli, staphylococcus aureus and candida albican. mixed growth was found in 26% of the cultures. the antibiotic sensitivity of the cultures was to amikacin (38.3%), imipenem (29.8%), tazocin (21.3%) and gentamycin (14.9%). the antibiotic resistance among the cultures was to; ciproxin (44.7%), amoxicillin (34.0%), cefotaxime (31.9%) and ampicillin (23.4%). conclusion: the infection rate among suspects of hai, was found to be high in this study which calls for an urgent need of robust infection control policy and strategies. key words: hospital acquired infections, nosocomial infections, culture-positivity, culture sensitivity introduction hospital acquired infection (hai) is a serious health hazard across the world. world health organization (who) described it as one of the major threats posing huge economic impact. nosocomial infections, hospital acquired infections and health-care associated infections are terms used interchangeably. despite much advances in the control and prevention of hospital acquired infections, still continue to remain major contributor to the rate of morbidity, mortality and cost of care.1,2 nosocomial infections can be defined as those occurring within 48 hours of hospital admission, 3 days of discharge or 30 days of a surgery. postoperative nosocomial infection is a surgical site or blood stream infection occurring within 48 hours of surgery until the time of discharge from hospital with clinical signs and symptoms and laboratory confirmation.3 the signs of infection include pain, tenderness, warmth, erythema, swelling and discharge.4 prevalence of hai is 9.4 percent in the developed countries. in a meta analysis and systematic review on health-care-associated infections reported icu pooled densities at least three times higher than from usa. surgical-site infection was leading with proportions higher than reported in developed countries. gram-negative bacilli are commonly identified with methicillin resistant staphylococcus aureus.5 surgical services constitute a large portion of hospital care with wide 12 culture-positivity and sensitivity in post-surgical patients in a tertiary care hospital range of major and minor operative procedures being conducted at a tertiary care setup. postoperative period is of great concern while looking at surgical services as a whole. the primary objective of postoperative care is to minimize patient’s discomfort, assist restitution of health and prevent infections.6,7 infection of surgical wounds resulting in pyrexia, discomfort, serious complications and subsequent prolonged use of antimicrobials are quite common in our part of the world. the source of these infections can be exogenous or endogenous bacteria. this has a grave impact on patient’s overall health and finances and a burden on hospitals. conventionally prophylactic antibiotics are prescribed to the patient both before and after the surgery. duration and type of prophylactic antibiotics received by the patient matters a lot in the development of postoperative infections along with patient factors as well.8 the increasing poly antimicrobial resistance amongst hospital pathogens also poses a great threat.9 usually wound, blood, sputum, fluid, stool and urine samples are sent for culture testing to identify microbial presence, growth as well as sensitivity and resistance to specific antibiotics.10,11 postoperative infections result in prolonged duration of hospital stay and pose economic, physical and psychological burden on the patient and the entire family. this raises serious questions about hospital’s performance, standards, hygiene, maintenance and economics.12 postoperative infections are good indicators for hospital associated infections (nosocomial infections) and quality of hospital care. multiple factors influence infection control behaviors and practices in surgery therefore interventions are needed to ensure shared ownership of all clinical outcomes.13 quality assurance and control activities are the backbone of hospital services so evidence-informed guidelines are greatly needed to provide new and updated evidence-based recommendations for the prevention of ssi and should be incorporated into comprehensive surgical quality improvement programs to improve patient safety.14 healthcare expenditure in organization for economic cooperation and development (oecd) countries is increasing due to increased pharmaceutical expenditure, hospital care and the high prevalence of chronic diseases and their financial consequence.15,16,17 this has created a need to implement systems to improve the efficiency and quality of health care18 which is the reason behind this study. an iranian study showed 60.6% antimicrobial resistance in nosocomial infection cases.19 in pakistan the frequency of nosocomial infection in icu of a tertiary care hospital was reported 29.13% in 200820 and surgical site infection in post surgical patients was 7.3% in another hospital in 2011.21 a multi-centered study from pakistan showed a acinetobacter baumannii, the most prominent antibiotic-resistance phenotype.22 alarmingly, we are moving towards pre-antibiotic era very fast and hospitals are a source of antimicrobial resistance.23 this study was urgently much needed to highlight the issue so the findings can be taken up to develop contextualized, evidence-informed hospital policies. recent quality literature from pakistan on the magnitude of nosocomial infections in surgical and allied services of tertiary care hospitals is limited. current study will guide administrative policies guidelines and plans for infection control in surgical settings and services. current study is aimed at measuring the frequency of postoperative culture positivity and sensitivity at one of the largest tertiary care hospitals in lahore. material and methods a descriptive cross-sectional study was conducted at a tertiary care teaching hospital in lahore, punjab from april to july, 2016 on secondary data of patients who were operated upon during 1st october to 31st december, 2015. this hospital houses more than 1000 beds. eligibility criteria: all patient files, all those operated upon from 1st october 2015 to 31st december 2015 in general surgery or the allied specialties except gynecology and obstetrics were our study population. researchers had constraints of time so they only conducted this study on 3 months record therefore this project was limited by time convenience. patients who had been re-operated within 30 days of first operation, had developed a complication or were declared immune-compromised were not included in the study. tampered files and files with inadequate data were also not included. the data was collected from the medical record room of the hospital by the research team maintaining anonymity and confidentiality of data. the patients’ records were used to fill the selfconstructed questionnaire designed specifically to acquire information about culture indicated, culture positivity and sensitivity to the available antibiotics. for this purpose, the microbiology reports of the specimens sent (including but not restricted to: 13 culture-positivity and sensitivity in post-surgical patients in a tertiary care hospital blood, sputum, urine, stool, wound/tissue and fluids) to the microbiology laboratory of the hospital were used. current study assessed the frequency of culture positivity and sensitivity according to specialty, type of organisms identified, antibiotics sensitivity and resistance along with gender-wise distribution of these variables. statistical analysis: the data was entered and analyzed in the ibm spss version 21 software. results total patients discharged during the study period were 420, out of which 41.9% were females and 58.1% were males. out of 420, 58.4% patients were operated in general surgery and 41.6% patients in allied surgical specialties (mentioned in inclusion criteria). average duration of stay of 420 patients was 9.92 days with sd+8.37 and range of 1 to 55 days. overall 268/420 (63.8%) patients had prolonged stay (>5 days) and 152/420 (36.2%) had 5 days or less post-op stay at the hospital. the usual practice is to give post operative antibiotic cover and those patients with symptoms and signs of hai are investigated by sending samples for culture sensitivity. 124/420 (29.5%) were suspected to have acquired infection during hospital stay (as per their record files) so, samples were sent of 40.3% females and 59.7% males for culture sensitivity. out of total 420 patients operated, 47(11.19%) were culture positive during hospital stay, for acquired infection. out of 124 cultures indicated, 47 (37.9%) were found to be positive while 77 (62.1%) were culture negative. 51.1% of our culture positives were male patients and 48.9% females. 64.9% of the total culture negatives were males and 35.1% females. there was no significant difference (p=>0.05). out of culture positives (47/124), urinary tract infections (utis) was leading the list with 35 out of 47 (74.5%) and tissue/ wound infection was second with 8 out of 47 (17.0 %), blood/ fluid infection was 2 /47 (4.3%) and other site infections were 2 (4.3%). distribution of culture indicated/sent and culture positivity according to type of surgical specialty showed that; urology cultures sent were 93/124 (75%) out of which 33 were positive. in general surgery, 18/124 cultures were sent showing 8 positive. in orthopedics, 9/124 (7.26%) cultures were sent with 4 positive. in cts, 4/124 (3.22%) cultures were sent out of which 50% were positive. in neuro-surgery, 1/124 (0.81%) culture was sent during the period of study which came back negative. department of urology was the highest and neuro-surgery being the lowest among them. antibiotic sensitivity results showed that 38.9% (18/47) of positive cultures were sensitive to amikacin, 29.8% (14/47) of positive cultures were sensitive to imipenem, 21.3% (10/47) of positive cultures were sensitive to tazocin and 14.9% (7/47) of positive cultures were sensitive to gentamycin. antibiotic resistance results showed that 44.7% (21/47) of positive cultures were resistant to ciproxin, 34.0% (16/47) of positive cultures were sensitive to amoxicillin, 31.9% (15/47) of positive cultures were resistant to cefotaxime and 23.4% (11/47) of positive cultures were resistant to ampicillin. the most frequently found organisms in cultures were coliform species (38%), pseudomonas aeruginosa (13%), escherichia coli (10%), staphylococcus aureus (8%) and candida albicans (5%). mixed growth was found in 26% of the cultures (above data in table-1). variables frequency percentage gender (n=47) males 24 51.1% females 23 48.9% type of infection (n=47) uti 35 74.5% tissue/wound 8 17% blood/fluid 2 4.3% other sites 2 4.3% specialties (n=47) urology 33 71% general surgery 8 17% orthopedics 4 8.5% cardiothoracic 2 4.2% neurosurgery 0 0% antibiotic sensitivity (n=47) amikacin 18 38.9% imipenem 14 29.8% tazocin 10 21.3% gentamycin 7 14.9% antibiotic resistance (n=47) ciproxin 21 44.7% amoxicillin 16 34% cefotaxime 15 31.9% ampicillin 11 23.4% microorganism growth (n=47) coliform 18 38% pseudomonas a. 6 13% escherichia coli 5 10% staphlococcus a. 4 8% candida a. 2 5% mixed growth 12 26% table-1: culture sensitivity and resistance pattern among post-surgical culture positive samples in a tertiary care hospital 14 culture-positivity and sensitivity in post-surgical patients in a tertiary care hospital discussion our results provide evidence that nosocomial infections are a serious problem in tertiary care hospitals in pakistan, since the rates found are high (11.19%). in a systematic review conducted on healthcare associated infections in africa showed a hospital-wide prevalence of 2.5% to 14.8% in surgical wards, surgical site infection(ssi) was the main focus.24 rates of current study are within this range and near to the maximum limit found in literature from africa, while it is more than another systematic literature review and meta-analysis of 41 studies, on the burden of healthcare-associated infections (hais) in southeast asia, showing a pooled prevalence of overall hais as 9.0%.25 in literature from pakistan, the nosocomial infection rate of icu was reported to be 29.13% in 200820 while surgical site infection (ssi) rate was 7.3% in 2011.21 nosocomial infection surveillance and its epidemiology are found to be well studied in literature over last decade in different operative patients.26,27,28 in current study, average duration of stay was 9.92 days with sd+8.37 and range of 1 to 55 days. 63.8% patients had prolonged stay (>5 days). this is different from the findings of a study, where ssi rate reported was 9.85%, the median time to ssi was the 7th postoperative day and the average length of stay was 4.2 ± 3.4 days.29 in present study findings, amongst the post-op positive culture reports, urine culture was 74.5% followed by tissue/ wound infection/ssi 17.0 %. in a study of 2019, from a hospital in pakistan, done on post-operative patients, reported ssi rate as 33.68% which is very high as compared to our study findings.30 in current study resistant strains were mostly sensitive to amikacin followed by imipenem, tazocin and and gentamycin while antibiotic resistance was found mainly to ciproxin, followed by amoxicillin and cefotaxime. in our study, coliform species were most frequently found (31.9%) followed by pseudomonas aeruginosa, escherichia coli, staphylococcus aureus and candida albicans. mixed growth was found in 26% of the cultures. in a study in india, escherichia coli was most commonly isolated microbe followed by klebsiella pneumoniae, pseudomonas aeruginosa, enterococcus faecalis, staphlococcus aureus and coagulase negative staphylococci.31 klebsiella spp. was the most frequently (27%) isolated bacterium in another study in 2016.11 in a systematic review and meta-analysis published in 2019 on 66 studies, escherichia coli, klebsiella pneumoniae and staphylococcus spp were reported in majority of studies.32 in the age of antibiotics, nosocomial infections are still uncontrollable. implementation of infection control practices through continuous training and monitoring of health care providers could help in reducing nosocomial infection rates. irrational use of antibiotics is an important cause of drug-resistant organisms. nosocomial surveillance in hospital needs to be stringent followed by adequate measures to address deficiencies in tertiary care health setups in countries like pakistan. limitations in current research include the data extraction from manual record keeping system of patient files. surgical and medical specialties and different hospitals could be compared to get a larger evidence for development and revision of uniform infections control policies in hospitals of pakistan. qualitative case studies can give in-depth insight into the problem and solutions of hai. conclusion our study found high frequency of culture positivity in all suspects of nosocomial infections in different surgical specialties of a tertiary care hospital. antimicrobial resistance was high and this is found to be consistent with local and international literature. references 1. ahmed mi. prevalence of nosocomial wound infection among postoperative patients and antibiotics patterns at teaching hospital in sudan. north american journal of medical sciences. 2012; 4(1):29. 2. jarvis wr. selected aspects of the socioeconomic impact of nosocomial infections: morbidity, mortality, cost, and prevention. infection control & hospital epidemiology. 1996; 17(08):552-7. 3. mulu w, kibru g, beyene g, damtie m. postoperative nosocomial infections and antimicrobial resistance pattern of bacteria isolates among patients admitted at felege hiwot referral hospital, bahirdar, ethiopia. ethiopian journal of health sciences. 2012; 22(1):7-18. 4. cutting kf, white r. defined and refined: criteria for identifying wound infection revisited. british journal of community nursing. 2004; 9(3):s6-15. 5. allegranzi b, bagheri nejad s, combescure c, graafmans w, attar h, donaldson l, et al. 15 culture-positivity and sensitivity in post-surgical patients in a tertiary care hospital burden of endemic health-care-associated infection in developing countries: systematic review and meta-analysis. lancet (london, england). 2011; 377(9761):228-41. 6. stulberg jj, delaney cp, neuhauser dv, aron dc, fu p, koroukian sm. adherence to surgical care improvement project measures and the association with postoperative infections. jama. 2010; 303(24):2479-85. 7. awad ss. adherence to surgical care improvement project measures and postoperative surgical site infections. surgical infections. 2012; 13(4):234-7. 8. flores-mireles a, hreha tn, hunstad da. pathophysiology, treatment, and prevention of catheter-associated urinary tract infection. top spinal cord inj rehabil. 2019;25(3):228-40 9. huang ch, chiu ch, chen iw, hung sy, lin cw, hsu br, et al. antimicrobial resistance and outcomes of community-onset bacterial bloodstream infections in patients with type 2 diabetes. j glob antimicrob resist. 2018; 15:271-6. 10. satti mz, hamza m, sajid z, asif o, ahmed h, zaidi smj, et al. compliance rate of surgical antimicrobial prophylaxis and its association with knowledge of guidelines among surgical residents in a tertiary care public hospital of a developing country. cureus. 2019; 11(5):e4776 11. nouetchognou js, ateudjieu j, jemea b, mesumbe en, mbanya d. surveillance of nosocomial infections in the yaounde university teaching hospital, cameroon. bmc res notes. 2016; 9(1):505. 12. zimlichman e, henderson d, tamir o, franz c, song p, yamin ck, et al. health care-associated infections: a meta-analysis of costs and financial impact on the us health care system. jama intern med. 2013; 173(22):2039-46. 13. troughton r, mariano v, campbell a, hettiaratchy s, holmes a, birgand g. understanding determinants of infection control practices in surgery: the role of shared ownership and team hierarchy. antimicrob resist infect control. 2019; 8:116. 14. berríos-torres si, umscheid ca, bratzler dw, leas b, stone ec, kelz rr, et al. centers for disease control and prevention guideline for the prevention of surgical site infection. jama surgery. 2017; 152(8):784-91. 15. armstrong bk, gillespie ja, leeder sr, rubin gl, russell lm. challenges in health and health care for australia. the medical journal of australia. 2007; 187(9):485-9. 16. lorenzoni l, belloni a, sassi f. health-care expenditure and health policy in the usa versus other high-spending oecd countries. the lancet. 2014; 384(9937):83-92. 17. squires da. explaining high health care spending in the united states: an international comparison of supply, utilization, prices, and quality. issue brief (commonwealth fund). 2012; 10:1-14. 18. hoque dm, kumari v, ruseckaite r, romero l, evans sm. impact of clinical registries on quality of patient care and health outcomes: protocol for a systematic review. bmj open. 2016; 6(4):e010654. 19. rajabi m, abdar me, rafiei h, aflatoonia mr, abdar ze. nosocomial infections and epidemiology of antibiotic resistance in teaching hospitals in south east of iran. glob j health sci. 2015; 8(2):190-7. 20. shaikh jm, devrajani br, shah s, akhund t, bibi i. frequency, pattern and etiology of nosocomial infection in intensive care unit: an experience at a tertiary care hospital. j ayub med coll abbottabad. 2008; 20(4):37-40. 21. bibi s, channa ga, siddiqui tr, ahmed w. frequency and risk factors of surgical site infections in general surgery ward of a tertiary care hospital of karachi, pakistan. int j infect control. 2011; 7(3):5. 22. hasan b, perveen k, olsen b, zahra r. emergence of carbapenem-resistant acinetobacter baumannii in hospitals in pakistan. j med microbiol. 2014; 63(pt 1):50-5. 23. michael ca, dominey-howes d, labbate m. the antimicrobial resistance crisis: causes, consequences, and management. frontiers in public health. 2014; 2:145. 24. nejad sb, allegranzi b, syed sb, ellis b, pittet d. health-care-associated infection in africa: a systematic review. bulletin of the world health organization. 2011; 89:757-65. 25. ling ml, apisarnthanarak a, madriaga g. the burden of healthcare-associated infections in southeast asia: a systematic literature review and meta-analysis. clin infect dis. 2015; 60(11):1690-9. 26. iordanou s, middleton n, papathanassoglou e, raftopoulos v. surveillance of device associated infections and mortality in a major intensive care unit in the republic of cyprus. bmc infect dis. 2017; 17(1):607. 27. gomez-lopez r, barge-caballero e, fernandez-ugidos p, paniagua-martin mj, barge-caballero g, couto-mallon d, et al. inhospital post-operative infection after heart 16 culture-positivity and sensitivity in post-surgical patients in a tertiary care hospital transplantation: epidemiology, clinical management, and outcome. surg infect (larchmt). 2019. 28. curcio d, cane a, fernandez f, correa j. surgical site infection in elective clean and clean-contaminated surgeries in developing countries. int j infect dis. 2019; 80:34-45. 29. zejnullahu va, isjanovska r, sejfija z, zejnullahu va. surgical site infections after cesarean sections at the university clinical center of kosovo: rates, microbiological profile and risk factors. bmc infect dis.2019;19(1):752 30. sattar f, sattar z, zaman m, akbar s. frequency of post-operative surgical site infections in a tertiary care hospital in abbottabad, pakistan. cureus. 2019; 11(3). 31. ginawi i, saleem m, sigh m, vaish ak, ahmad i, srivastava vk, et al. hospital acquired infections among patients admitted in the medical and surgical wards of a nonteaching secondary care hospital in northern india. j clin diagn res. 2014; 8(2):81-3. 32. mouiche mmm, moffo f, akoachere jtk, okah-nnane nh, mapiefou np, ndze vn, et al. antimicrobial resistance from a one health perspective in cameroon: a systematic review and meta-analysis. bmc public health. 2019; 19(1):1135. the authors: dr. ehsaan ahmed research assistant (honorary), department of public health & community medicine, shaikh khalifa bin zayed al-nahyan medical college, shaikh zayed medical complex, lahore. prof. muhammad imran anwar department of surgery, shaikh zayed medical complex, lahore. dr. ruqaya idrees research assistant (honorary), department of public health & community medicine, shaikh khalifa bin zayed al-nahyan medical college, shaikh zayed medical complex, lahore. dr. sheza arif toor research assistant (honorary), department of public health & community medicine, shaikh khalifa bin zayed al-nahyan medical college, shaikh zayed medical complex, lahore. dr. asad mahmood research assistant (honorary), department of public health & community medicine, shaikh khalifa bin zayed al-nahyan medical college, shaikh zayed medical complex, lahore. prof. ayesha humayun department of public health & community medicine, shaikh khalifa bin zayed al-nahyan medical college, shaikh zayed medical complex, lahore. corresponding author: prof. ayesha humayun department of public health & community medicine, shaikh khalifa bin zayed al-nahyan medical college, shaikh zayed medical complex, lahore. e-mail: drayeshah@gmail.com untitled-1 33 proceedings s.z.m.c. vol: 35(1): pp. 33-38, 2021. pszmc-781-35-1-2021 protective effects of ocimum basilicum against atrophic changes in graafian follicles in cyclophosphamide induced ovarian toxicity 1saba saleem, 2muhammad suhail, 1saba amjad, 3faiza irshad 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, shaikh zayed medical complex, lahore 3department of anatomy, m islam medical & dental college, gujranwala abstract introduction: cyclophosphamide is one of the alkylating chemotherapeutic drug used in cancer patients that has antifertility effects on female gonads. ocimum basilicum is a natural herb rich in polyphenols and is known to improve fertility. aims & objectives: the study was designed to evaluate the role of natural herb, ocimum basilicum extract, as a preventive agent against ovarian follicular toxicity induced by cyclophosphamide. place and duration of study: this experimental study was performed in the department of anatomy, shaikh zayed postgraduate medical institute, lahore. the duration of study was 8 months. material & methods: 45 female albino rats were divided equally in control group a, experimental group b and group c each contained 15 rats. group a rats received single dose of 150 mg/kg normal saline intraperitoneally on 8th day of experiment, while group b was given single intraperitoneal dose of 150 mg/kg cyclophosphamide at day 8 of experiment. group c rats were pretreated with methanolic basil (ocimum basilicum) seeds extract for 7 days followed by single intraperitoneal dose of 150 mg/kg cyclophosphamide at day 8 of experiment. all the rats were dissected 48 hours after the last dose. results: graafian follicles were atrophied showing atretic granulosa cells in group b when compared with control group a with p value <0.001. however, significant improvement in status of graafian follicles was observed in group c, when compared with group b with p value <0.025. conclusion: this study depicts that basil seeds extract can prevent the cellular toxicity in graafian follicles caused by cyclophosphamide treatment. so the use of basil seeds during chemotherapy can significantly limit its toxic effects on graafian follicles. key words: cyclophosphamide, oxidative stress, follicular atrophy, basil. introduction cyclophosphamide is a commonly used effective anticancer alkaloid as well as an effective immunosuppressive drug. cyclophosphamide gave the best desired result when tested against 33 tumors in comparison with 1000 other agents.1 it is a useful alkylating drug that belongs to group oxazophosphorine and was first introduced by bourseaux, arnold and brock in 1958.2 this alkylating drug causes cytotoxicity by shifting its alkyl group to various constituents of tumor cells especially their dna which causes cell death. alkylation of guanine is common which results in abnormal pairing of guanine with thymine causing formation of abnormal dna strands.3 cyclophosphamide is being used to treat malignant breast carcinoma, hodgkin’s lymphoma and autoimmune disorders like systemic lupus erythematosus and glomerulonephritis.4,5,6 cyclophosphamide, like other chemotherapeutic drugs, has adverse effects on different organ systems of body. severity of adverse effects showed direct relation with dose of drug. most common side effects are immunosuppression, nausea, vomiting, myelosuppression, weight loss and loss of appetite.7 it is known to cause increased susceptibility to infections, haemorrhagic cystitis, nausea, vomiting, hair loss and permanent infertility in humans even at therapeutic doses.8 nowadays, increasing use of chemotherapeutic drugs for treatment of lupus nephritis and breast cancer in young females is causing premature menopause and secondary infertility due to post treatment ovarian failure.9,10 the mechanism of ovarian toxicity with cyclophosphamide involves decreased levels of antioxidant enzymes i.e. superoxide dismutase and catalase, resulting in excess of free radicals causing oxidative tissue damage.11 a study on female rats showed the dose dependent relationship between different doses of cyclophosphamide and antral follicular atresia.12 34 protective effects of ocimum basilicum against atrophic changes in graafian follicles ocimum basilicum (basil) plant belongs to the family lamiacaea, species ocimum basilicum and genus ocimum.13 basil is called “king of herbs” because of its wide usage and advantages.14 high content of phenolics in extract of various parts of this basil give it significant antioxidant potential in various in-vitro studies.15 when basil was used along with other medicinal herbs for treating oligospermia in males, it showed significant improvement in semen catalase levels and semen parameters proving that reduction in oxidative stress by increasing antioxidant enzymes in gonads can improve chances of fertility because free radicles are a big factor in infertility.16 an experiment on female rats also proved antioxidant potential of its methanolic extract given at dose of 1.5g/kg in combination with exposure to electromagnetic field (emf) and results showed significant improvement in various histological parameters of ovaries including granulosa cell apoptosis, fibrosis and venous congestion, when compared with other group that received only emf exposure.17 cyclophosphamide is being frequently used in recent past years to treat malignancies. major adverse effect of cyclophosphamide, when used in young females having ca breast, is secondary infertility due to its direct effects on primordial and growing follicles resulting in oxidative damage.18 it is obvious from past studies that natural antioxidants have protective effect when used in combination with cyclophosphamide.11 present study is therefore designed to see the antioxidant potential of basil seeds extract against cyclophosphamide induced ovarian toxicity. material and methods this experimental study was performed in the department of anatomy, shaikh zayed postgraduate medical institute, lahore. the sample size was estimated by using power and precision 3.0 software with 0.48 effect using and 2.28 as error sd. based on this a total of 45 adult healthy female albino rats (3-4 months old), average female weight 190-240 gm were used in this study. all these animals were kept in cages in the animal house of the department of anatomy, punjab pgmi, lahore. the animals were allowed free access to food and water. a commercial brand of chick feed no. 1 was provided to rats, (appendix-i). in every 5 kg of this feed, wheat flour 2.5 kg, molasses 1 kg, fish meal 100 grams and water was added. a 12:12 light: dark cycle was maintained. temperature was maintained between 22-25 ℃. basil seeds were extracted with methanol. nonvolatile compounds were extracted by solvent extraction method. weighted samples were taken in a flask and it was filled with solvent. these samples were regularly shaken for 48 hours with 3 hours interval. then sample was filtered with filter paper and subjected to rotatory evaporation in order to remove solvent and then by air evaporation. sample was stored in freezer to avoid loss of antioxidant compounds.19 dosage solution for oral administration was prepared by dissolving extract in normal saline.20 cyclophosphamide was purchased from pharmacy and dose of drug as 150mg/kg body weight was calculated for each rat. all rats were divided in three groups a, b and c. each group contained 15 rats that were further named as a1-a15, b1-b15 and c1-c15 by lottery method. the rats were assigned that number and marked with permanent marker and placed in the specific group cage. the weight of each rat was carefully recorded, with the help of weighing machine, in a proforma. the dose schedule was as follows: group a (control): the rats of this group were not given any extract or toxic drug. these rats were provided with routine oral food daily and only given the single dose of normal saline 150 mg/kg body weight intraperitoneally on day 8 of experiment.20 group b (experimental): the rats of this group received routine oral food with other rats for 7 days followed by only a single dose of 150 mg/kg cyclophosphamide intraperitoneally at day 8 of experiment.11 group c (experimental): this experimental group received basil seed extract as dose of 1.5 g/kg/day through gastric intubation for 7 days followed by single 150 mg/kg intraperitoneal dose of cyclophosphamide at day 8 of experiment. the extract was given on the same fixed time daily. all the animals of group a, b and c were properly given analgesia at the end of experiment, 48 hours after giving the last toxic dose, by using 50mg/kg ketamine followed by 50mg/kg xylazine intraperitoneally. the animals were dissected and ovaries were removed. hematoxylin and eosin stained slides were prepared for both right and left ovaries of each rat having 5 serial tissue sections in each slide. graafian follicles were observed for atretic changes in their follicular cells. the follicle was considered tertiary when oocyte was seen surrounded by corona radiata cells and there was only a single large cavity between compacted layers of granulosa cells and corona radiata cells.12 granulosa cells were considered atretic when having pyknotic nuclei, 35 protective effects of ocimum basilicum against atrophic changes in graafian follicles irregular arrangement around oocyte and detachment from their basement membrane.11 statistical analysis: qualitative data for graafian follicles was analyzed by using frequency and percentage for each group. then comparison was made among the groups by using chi-square test. p value was considered significant when <0.05. quantitative data was reported by using mean ± sd. comparison among the groups was made by using anova. results graafian follicles were observed normal in 13 rats (86.75) of control group a. graafian follicles in only 2 rats of control group were atretic and deshaped. atresia was seen in 12 rats (80%) of group b and 6 rats (40%) of group b, (table-1). group graafian follicle normal atretic total n % n % n % group a 13 86.7 2 13.3 15 100.0 group b 3 20.0 12 80.0 15 100.0 group c 9 60.0 6 40.0 15 100.0 table-1: status of graafian follicles observed in control group a, experimental groups b and c. diameters of graafian follicles showed significant decrease in diameters in toxic group b, when compared with control group a with p value <0.001. there was improvement in diameters of follicles in experimental group c when compared with toxic group b with p value 0.005, (fig-1). fig-1: diameter of graafian follicle measured for control group a, experimental group b and c (error bar represents standard deviation). fig-2: photomicrograph graafian follicle of control group showing well arranged granulosa cells (gc), follicular antrum (a), cumulous oophorus (co) and corona radiate (cr), (h & e, 20x). the follicles were labeled atretic because granulosa cells were pulled apart from their basement membrane and from each other having pyknotic nuclei. granulosa cell layer was also irregular and with decreased thickness when compared with control group graafian follicle, (table-1, fig-2,3). remaining rats in group b and c had normal rounded mature follicles with well defined theca interna and theca externa. fig-3: photomicrograph of graafian follicle in ovarian cortex of experimental group b showing irregular granulose cell layer (igc), central oocyte (o), pyknotic granulosa cells pulled away from basement membrane (red arrows) and epithelium (e), (h & e, 20x). groups a & b <0.001** groups a & c 0.062 groups b & c 0.005* 36 protective effects of ocimum basilicum against atrophic changes in graafian follicles fig-4: photomicrograph of ovary of experimental group c having graafian follicle showing follicular antrum (a), well arranged granulosa cell layer (gc), corona radiate (cr) and cumulous oophorus (co), (h & e, 20x). when group wise comparison was made between control group a and experimental group b, the results were highly significant with p-value <0.001. the difference observed between control group and experimental group c was not significant with pvalue 0.093. comparison between experimental groups b and c showed significant difference with p-value 0.025, (table-2). (i) group (j) group p-value group a group b <0.001** group c 0.093+ group b group c 0.025* table-2: group wise comparison for status of graafian follicles in control group a, experimental groups b and c. discussion in the present study, cyclophosphamide affected graafian follicles causing cell death and apoptosis of granulosa cells resulting atrophy of follicles. the results showed that there is significant improvement in status of graafian follicles and its size in experimental group c receiving pretreatment with basil seeds extract, in comparison with toxic group b receiving only cyclophosphamide. these changes observed in graafian follicles in toxic group b are due to decreased levels of antioxidant enzymes and overproduction of free radicles caused by toxic effects of cyclophosphamide resulting in initiation of cell damage and apoptosis of damaged cells.11,21 another important mechanism that explains the cell death and apoptosis of granulosa cells causing atrophy of graafian follicles, is the cytotoxic action of phosphoramide mustard, one of its active metabolic compound formed by liver.22 this phosphoramide mustard caused alkylation of guanine base of dna and resulted in formation of abnormal dna causing cell cycle arrest and apoptosis in rapidly dividing cells of gonads.3 the significant improvement in status and size of graafian follicles was observed in group c, given basil seed extract and cyclophosphamide, (fig1,2,3,4). this can be explained by the antioxidant and free radical scavenging property of seed extract of ocimum basilicum that can be responsible for prevention of oxidative damage in gonads.23-28 the antioxidant mechanism of action of ocimum basilicum has also been proved in the study performed by khaki a. in this study ocimum basilicum extract protected the rat tissue against oxidative damage induced by electromagnetic field.29 conclusion the results of the present study proved that basil seed extract can protect graafian follicles when given as a preventive treatment along with cyclophosphamide by improving the atrophic changes in granulosa cells of mature follicles induced by cyclophosphamide. the results also suggest the clinical use of basil seeds extract for prevention of infertility induced by cyclophosphamide. references 1. emadi a, jones r.j., brodsky ra. cyclophosphamide and cancer: golden anniversary. nature reviews clinical oncology. 2009; 6 (11):638-647. 2. chighizola c, ong vh, denton cp. cyclophosphamide as disease-modifying therapy for scleroderma: pros and cons. international journal of clinical rheumatology. 2011; 6(2):219-227. 3. edward c. alan c. sartorelli. cancer chemotherapy, in: bertram g. katzung, susan b. basic and clinical pharmacology 12th edition. new york, usa: mc graw hill companies; 2012. p. 953. 4. edward c. alan c. sartorelli. cancer chemotherapy, in: bertram g. katzung, susan b. masters (eds.). basic and clinical pharmacology 12th edition. new york, usa: mc graw hill companies; 2012. p. 953. 37 protective effects of ocimum basilicum against atrophic changes in graafian follicles 5. hudson mm, schwartz c, constine ls. treatment of pediatric hodgkin lymphoma. in pediatric lymphomas 2007: 35-48. 6. houssiau fa, vasconcelos c, d'cruz d, sebastiani gd, garrido ed, danieli mg, abramovicz d, blockmans d, mathieu a, direskeneli h, galeazzi m. immunosuppressive therapy in lupus nephritis: the euro‐lupus nephritis trial, a randomized trial of low‐dose versus high‐dose intravenous cyclophosphamide. arthritis & rheumatism. 2002; 46(8):2121-31. 7. zhong s, huang m, yang x, liang l, wang y, romkes m et al. relationship of glutathione s‐transferase genotypes with side‐effects of pulsed cyclophosphamide therapy in patients with systemic lupus erythematosus. british journal of clinical pharmacology. 2006; 62(4): 457-72. 8. teles ka, medeiros-souza p, lima fa, araújo bg, lima ra. cyclophosphamide administration routine in autoimmune rheumatic diseases: a review. revista brasileira de reumatologia. 2017; 57(6):596-604. 9. wetzels jf. cyclophosphamide-induced gonadal toxicity: a treatment dilemma in patients with lupus nephritis. netherlands journal of medicine. 2004; 62(10):347-52. 10. meirow d, nugent d. the effects of radiotherapy and chemotherapy on female reproduction. human reproduction update. 2001; 7(6):535-43. 11. yener na, sinanoglu o, ilter e, celik a, sezgin g, midi a et al. effects of spirulina on cyclophosphamide-induced ovarian toxicity in rats: biochemical and histomorphometric evaluation of the ovary. biochemistry research international. 2013; 764262. 12. kamarzaman s, shaban m, abdul rahman s. the prophylactic effect of nigella sativa against cyclophosphamide in the ovarian follicles of matured adult mice: a preliminary study. j anim plant sci. 2014 jan 1; 24:81-85. 13. stanojković-sebić a, dinić z, iličić r, pivić r, jošić d. effect of indigenous pseudomonas chlororaphis strains on morphological and main chemical growth parameters of basil (ocimum basilicum l.). ratarstvo i povrtarstvo. 2017; 54(2):42-7. 14. michele meyers. basil: an herb society of america guide. the herb society of america 9019 kirtland chardon rd. kirtland.2003.p.1-7. 15. juliani hr, simon je. antioxidant activity of basil. trends in new crops and new uses. 2002;575 16. alizadeh h, khaki a, farzadi l, nouri m, ahmadi-asrbadr y, seyed-ghiasi g et al. the therapeutic effects of a medicinal plant mixture in capsule form on catalase levels in the semen of men with oligospermia. crescent journal of medical and biological sciences. 2015;2(1):1-4. 17. khaki a, khaki aa, ezzatzadeh a, hamidreza a. effect of ocimum basilicum on ovary tissue histopathology after exposure to electromagnetic fields (emf) in rats. african journal of pharmacy and pharmacology. 2013; 7(25): 1703-1706. 18. amoura z, duhaut p, sbai a, costedoat n, wechsler b, piette jc. risk of ovarian failure and fertility after intravenous cyclophosphamide. a study in 84 patients. the journal of rheumatology. 2002; 29(12):2571-6. 19. aydemir t, becerik s. phenolic content and antioxidant activity of different extracts from ocimum basilicum, apium graveolens and lepidium sativum seeds. journal of food biochemistry. 2011; 35(1):62-79. 20. rasekh hr, hosseinzadeh l, mehri s, kamlinejad m, aslani m, tanbakoosazan f. safety assessment of ocimum basilicum hydroalcoholic extract in wistar rats: acute and subchronic toxicity studies. iranian journal of basic medical sciences. 2012; 15(1):645. 21. devine pj, perreault sd, luderer u. roles of reactive oxygen species and antioxidants in ovarian toxicity. biology of reproduction. 2012; 86(2):27, 1-10. 22. chouhan rs, ahmad b, chauhan r, shrivastava vk. ameliorative role of ascorbic acid against hepatotoxicity induced by chemotherapeutic drug cyclophosphamide in male albino rat. international journal of biological & pharmaceutical research. 2014; 5(11):830-42. 23. kaurinovic b, popovic m, vlaisavljevic s, trivic s. antioxidant capacity of ocimum basilicum l. and origanum vulgare l. extracts. molecules. 2011; 16(9):7401-14. 24. gajendiran a, thangaraman v, thangamani s, ravi d, abraham j. antimicrobial, antioxidant and anticancer screening of ocimum basilicum seeds. bulletin of pharmaceutical research. 2016; 6(3):114-9. 25. fekri n, khayami m, heidari r, jamee r. chemical analysis of flaxseed, sweet basil, dragon head and quince seed mucilages. research journal of biological sciences. 2008; 3(2):166-70. 26. stevanović zd, radanović d. proceedings of the seventh conference on medicinal and 38 protective effects of ocimum basilicum against atrophic changes in graafian follicles aromatic plants of southeast european countries, (proceedings of the 7th cmapseec). institute for medicinal plant research and association for medicinal and aromatic plants of southeast european countries (amapseec). subotica, serbia, 2012: 267-273. 27. hussain ai, anwar f, sherazi st, przybylski r. chemical composition, antioxidant and antimicrobial activities of basil (ocimum basilicum) essential oils depends on seasonal variations. food chemistry.2008;108(3):986-95. 28. shirazi mt, gholami h, kavoosi g, rowshan v, tafsiry a. chemical composition, antioxidant, antimicrobial and cytotoxic activities of t agetesminuta and ocimum basilicum essential oils. food science & nutrition. 2014; 2(2):146-55. 29. khaki a. protective effect of ocimum basilicum on brain cells exposed to oxidative damage by electromagnetic field in rat: ultrastructural study by transmission electron microscopy. crescent journal of medical and biological sciences. 2016; 3(1):1-7. the authors: dr. saba saleem, assistant professor department of anatomy, ucmd, university of lahore. prof. muhammad suhail head, department of anatomy, shaikh zayed medical complex, lahore. dr. saba amjad, assistant professor, department of anatomy, ucmd, university of lahore. dr. faiza irshad, associate professor, department of anatomy, m islam medical and dental college, gujranwala. corresponding author: dr. saba saleem, assistant professor, department of anatomy, ucmd, university of lahore. e-mail: saba_saleem786@hotmail.com for web full book 28 proceedings s.z.m.c. vol: 37(3): pp. 28-33, 2023. pszmc-893-37-3-2023 emblica officinalis reduces copper mediated inflammation and preserves liver morphology inthe murine model 1hadia zulfiqar, 2muhammad suhail, 3amna rehman,3afifa waseem,4faeza rauf,1hafiza sadia ahmed 1department of anatomy, ucmd, university of lahore, lahore. 2department of anatomy, amna inayat medical college, lahore. 3department of anatomy, central park medical college, lahore. 4department of anatomy, rashid latif khan university medical and dental college, lahore. abstract introduction: copper (cu) is a metal widely used in agriculture and in industries. its hepatotoxicity is established in literature. emblica officinalis, locally known as amla, has many beneficial health effects and is scientifically reported to be a powerful antioxidant. aim & objective: to determine the role of emblica officinalis fruit in reducing cu induced inflammation and distortion of hepatic lobules of adult albino rats. place and duration of study: the study was done in the department of anatomy, shaikh zayed medical college, lahore for a period of 4 weeks. material & methods: 36 adult male albino rats were equally divided into three groups (a, b, c) by randomization and acclimatized for 1 week. 1.5 ml of normal saline was given to rats of group a (control) and cuso4 (200mg/kg b.w./day) was given to those in group b (cu treated) once daily via orogastric tube. rats in group c (cu+eo treated) were given both cuso4 (200mg/kg bw) and emblica officinalis fruit extract (300mg/kg bw) once daily by orogastric intubation. the animals were sacrificed after 28 days and livers were dissected out for histological study. data was entered and analyzed using spss version 22. results: cu treated rats developed significant distortion of liver architecture and inflammation while emblica officinalis co-treatment lead to decreased inflammation with preservation of shape of hepatic lobule. these findings were further confirmed by difference in diameters of hepatic lobules between control, cu-, cu+eotreated groups. conclusion: emblica officinalis, by limiting oxidative damage, ameliorates hepatic inflammation and preserves liver architecture. therefore, its use as a hepatoprotective agent should be encouraged. keywords: emblica officinalis, copper, inflammation, liver fibrosis, hepatoprotective. introduction recently liver disease has become major cause of morbidity and mortality worldwide. who survey claims liver cirrhosis to be the fifteenth major cause of death in adults worldwide with a prevalence of 1.5billion cases worldwide. in south-east asian region, liver cirrhosis stands as tenth major cause of death1.there are a variety of factors leading to liver toxicity and cirrhosis. these include hereditary factors, environmental toxins and micro-organisms. exposure to oxidant species such as copper (cu) is one such factor. increased serum cu levels have been reported in chronic cases of viral hepatitis, wilson’s and non-wilson’s copper toxicosis in multiple national and international studies2. considered to be the first metal used by man, today cu is one of the most commonly used commercial metals3and has found extensive application in agriculture, cosmetics, architecture, plumbing, electric and telecommunication industries4. these advances in commercial and industrial usage of copper have led to contamination of water sources with copper. increased amount of copper is found in drinking water coming from copper pipes or from water treated with algaecides5. discharge of industrial effluents into water streams also increases copper in water6. multiple studies have revealed increased copper content in vegetables grown on sewerage water and in those where copper-based pesticides and insecticides have been used. the highest concentration is found in green leafy vegetables and liver meat7. copper bioaccumulation has been reported in several fish species as well8. several international studies have found that copper concentration in multivitamins exceeds the permissible limit with few samples containing as much as 10 times the allowed limit of copper9. it is also a documented contaminant of herbal medicines and medicinal plants and has been found in slimming products as well10. copper is a known hepatotoxic agent and exerts its effect by oxidative damage to cells. local and international studies on human populations have linked increased amount of serum copper to hepatitis and chronic liver disease11. excess copper leads to damage of membrane lipids by formation 6 1department of public health, health services academy, islamabad. 2department of management sciences, riphah int university, islamabad. 3department of medicine, ihitc, islamabad. 4department of rheumatology, pims, islamabad. 5department of medicine, dow university of health sciences, karachi. 70 1department of orthopedics, thq, khanewal 2department of orthopedics, lgh, lahore 3department of orthopedics, thq, ferozwala 4department of orthopedics, cmh, lahore 5department of hand & upper limb surgery, cmh, lahore 6department of orthopedics, al-rehmat benevolent trust 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 29 emblica officinalis reduces copper mediated inflammation and preserves liver morphology in murine…... of per-oxy radicals and also causes peroxidation of the hepatic lysosomal membranes12.it decreases gsh levels and diminishes the activity of cytochrome c oxidase and catalase and impairs liver mitochondrial respiration13. emblica officinalis is commonly known as amla or indian gooseberry. it is grown in tropical countries of the asian continent including china, india, indonesia, sri lanka and pakistan. it is claimed to be the best source of vitamin c and contains 30 times the amount of ascorbic acid found in oranges. it has found extensive use in ayurveda, unani, arabic and tibetan medicine and is used since ancient times as a liver, skin and hair tonic14. it is also used locally in the form of powder, pickles, chutneys and murabba (jams). emblica officinalis is a rich source of many antioxidants. the tannins (emblicanin a and emblicanin b) and ascorbic acid present in it act as potent anti-oxidants and prevent tissue damage by inhibiting lipid peroxidation. they also augment antioxidant system and scavenge free radicals, ros and no in a dose dependent manner15. amla extract increases naturally occurring anti-oxidant enzymes including superoxide dismutase, gsh, catalase, gsh transferase and gsh reductase16.the ascorbic acid, phenols and flavonoids present in emblica officinalis also act as metal chelators. the protective effects of emblica officinalis against metals like lead, cadmium and mercury have been reported in literature17. emblica officinalis is claimed to restrict inflammation by preventing immune cells infiltration at the site of inflammation and by reducing the formation of several chemokines and no. it also promotes the formation of immunoprotective cytokines to speed the repair process 18. a study conducted on murine model showed that concurrent administration of emblica officinalis extract with arsenic decreased the levels of tnf, il-1 and il-6 in serum with recovery in b and t lymphocyte populations. there was also reduced arsenic burden in lymphoid organs like spleen and thymus which can be attributed to metal chelation properties of emblica officinalis19. it is also reported to decrease acute and chronic inflammation by decreasing edema and limiting the formation of granulomas in mice models20. emblica officinalis is reported to protect human liver from different environmental toxins such as heavy metals and alcohol by antagonizing apoptosis and lipid peroxidation21. it was found to be effective against ethanol induced liver damage and nonalcoholic steatohepatitis (nash) in murine models22. its fruit extract when given in a dose of 200mg/ kg/ day for 1 week to rats was found to be protective against carbon tetrachloride induced liver damage. it prevented necrosis, vacuolization of hepatocytes and fatty changes in liver. it also attenuated derangement in levels of alt, ast, total protein and albumin-globulin ratio23. similar hepatoprotective effects of emblica officinalis were reported in sulphur dioxide24, sodium arsenite25 and iron induced hepatic injury in rats26.since there is lack of hepatoprotective drugs, therefore, this study is designed to establish role of emblica officinalis as a hepatoprotective agent both in copper induced liver toxicity and in general toxic insults to liver. to determine the role of emblica officinalis fruit in reducing cu induced inflammation and distortion of hepatic lobules of adult albino rats. material and methods this study was a randomized animal trial done in department of anatomy, shaikh zayed medical college vide irb f-39 /185/acad/1417 dated 9-82016. initial sample size of 4 rats in each group was estimated by using 95% confidence level and 95% power using power & precision 3.0 software. the sample size was increased to 12 rats per group so that the parameters under observation could be compared with good power of study. 36 adult male albino rats (2-4 months of age) with average weight of 250-300g were marked and divided in three groups at random after 1 week of acclimatization. they were kept in three separate cages labelled a, b and c. rats showing signs of any ailment (lethargy, decrease in appetite and sleep time, sneezing, nasal or eye discharge, breathing problems and unexplained bleeding) were excluded from the study. extract from fresh fruits of emblica officinalis was made in pcsir laboratory, lahore. 20g of this extract was dissolved in 500 ml of normal saline to make extract solution. group a (control): the rats of this group were fed 1.5ml saline once daily by orogastric tube for 4 weeks. group b (cu treated): animals of this group were fed 1.5ml of copper sulphate stock solution once daily by orogastric tube for 4 weeks. group c (cu and emblica officinalis treated): rats of this group were fed by orogastric tube with 1.5ml of copper sulphate solution once daily. after 8 hours a single dose of 1.5 ml emblica officinalis solution was given via orogastric tube.the animals were fed with cu and emblica officinalis extract for 4 weeks. 24-hours after the last dose, they were euthanized. a midline incision was made from chin to groin. this incision was extended laterally to 30 emblica officinalis reduces copper mediated inflammation and preserves liver morphology in murine…... expose the liver. liver was taken out by dissecting the falciform ligament and hepatic vessels. livers were washed with saline after isolation and were placed in labelled jars containing 10% formaldehyde solution for fixation. after tissue processing, paraffin blocks of liver were made. each block was tagged with the name of respective animal. 5 m thick sections were cut by a rotatory microtome and stained with h & e stain for detailed histological evaluation. presence of multiple mononuclear cells like neutrophils and lymphocytes within the liver parenchyma and portal triad was labelled as inflammation. statistical analysis: spss 22 software was used to analyze the obtained data. the data for qualitative parameter such as shape of hepatic lobule and presence or absence of inflammation is stated by using frequency and percentages in each group. comparison of diameter of hepatic lobules among groups was made by using one way anova and post-hoc tukey test. p-value of 0.05 was considered significant. results 1. shape of hepatic lobule: shape of hepatic lobules of all rats in group a (control) was normal. the lobules were polygonal in shape with a central centrilobular vein. 5 to 6 portal triads were visible in periphery of each lobule. in group b (cu treated), all rats had hypertrophy and distortion of hepatic lobules whereas in group c (cu+eo treated), 10 (83.3%) rats had normal and 2 (16.7%) had enlarged and distorted hepatic lobules (fig-1,fig-2,fig-3,fig-5, fig-6). hypertrophy of lobules was assessed by calculating number of hepatic lobules/ central veins in one hpf and was later confirmed by micrometry. fisher’s exact test showed that there was an association between shape of hepatic lobule and groups.(table-1) shape of hepatic lobule total normal hypertrophy group a 12 (100.0%) 0 (0.0%) 12 (100.0%) group b 0 (0.0%) 12 (100.0%) 12 (100.0%) group c 10 (83.3%) 2 (16.7%) 12 (100.0%) p-value < 0.001* table-1: distribution of shape of hepatic lobule in control and experimental groups. fig-1: photomicrograph of the liver of adult albino rat from control group showing normal polygonal hepatic lobule. capsule (cap), portal triad (pt), central vein (red arrow) are also visible (h & e stain, 40x). fig-2: photomicrograph of liver of adult albino rat of group b (cu treated) showing disrupted margins of hepatic lobule, dilated central vein (red arrow) and portal triad (pt). (h&e stain, 40x). fig-3: photomicrograph of liver of adult albino rat of group c (cu+eo treated) showing normal polygonal lobule architecture. portal triad (pt) and central vein (red arrow) are also visible. (h&e stain, 40x). 31 emblica officinalis reduces copper mediated inflammation and preserves liver morphology in murine…... 2. inflammation in portal triad: presence of inflammation in portal triad was confirmed by presence of mononuclear infiltrate. fig-4 shows the percentage of presence or absence of inflammation in group a, b and c. fig-4: bar chart showing inflammation in portal triad among groups. fisher’s exact test showed that there was an association between inflammation in portal triadand groups with p-value <0.001 (table-2). inflammation in portal triad total present absent group a 0 (0.0%) 12 (100.0%) 12 (100.0%) group b 11 (91.7%) 1 (8.3%) 12 (100.0%) group c 2 (16.7 %) 10 (83.3%) 12 (100.0%) p-value < 0.001* table-2: inflammation in portal triad in control and experimental groups. *statistically significant difference (p-value < 0.05) fig-5: photomicrograph of adult albino rat of group b showing lymphocytes (red arrow) and fibrosis (yellow arrow) in portal triad. inset shows lymphocytic infiltration in portal triad (h&e stain, 400x). fig-6: photomicrograph of adult albino rat of group c showing normal portal triad (pt). hepatocytes with rounded central nuclei (red arrow), hepatic sinusoids (blue arrow) and centrilobular vein (cv) are also visible. (h&e stain, 400x). 3. diameter of hepatic lobule ( m): the diameter of hepatic lobule in all groups was measured using ocular micrometer. the mean diameter of hepatic lobule in each group are given in table-3 group diameter of hepatic lobule ( m) mean ± sd minimum maximum a 688.8 ± 22.80 662 726 b 957.5 ± 26.71 918 1012 c 691.5 ± 9.568 676 710 table-3: diameter of hepatic lobule ( m) in control and experimental groups. one-way anova test was used to compare the mean diameter of hepatic lobule among groups which shows significant p value of <0.001. for multiple comparisons, post hoc tukey test was used which showed that mean diameter of hepatic lobule in group b was significantly higher as compared to group a and c. however, no significant difference was found in the mean diameter of hepatic lobule between groups a and c (table-4). multiple comparison s. no. gro ups gro ups mean differe nce (i-j) std. error p-value diameter of hepatic lobule ( m) 1 a b -268.8 8.580 < 0.001* c -2.750 8.580 0.945+ 2 b c 266.1 8.580 < 0.001* table-4: pair wise comparison of diameter of hepatic lobule ( m) among groups. + statistically insignificant difference (p value > 0.05) *statistically significant difference (p value < 0.05) 32 emblica officinalis reduces copper mediated inflammation and preserves liver morphology in murine…... discussion the present study was designed to assess the ameliorative role of emblica officinalisextract on copper induced hepatotoxicity in adult albino male rats. liver was selected as the organ of study as it is the main site of metabolization for both cu27 and emblica officinalis. in previous studies the protective role of emblica officinalis on various hepatotoxic agents including heavy metals, alcohol and insecticides has been studied, but no research has yet been done on its protective role against cu toxicity. the hepatic lobules of rats treated with cu (group b) appeared grossly enlarged and their normal polygonal structure was distorted. this can be attributed to hyperemia due to inflammation12 as congestion was noted in hepatic sinusoids, central veins and in portal triads of rats in group b. lymphocytic infiltration was also observed in portal triads of cu treated rats (fig-5). el-hak et al reported similar finding due to cu toxicity which they attributed to cu induced oxidative stress that leads to disturbance of normal liver architecture, congestion in vessels and hepatic sinusoids as well as inflammation and ultimately fibrosis in hepatic lobules27.these findings were absent in animals of control group and cu+eo treated group (group a and group c). amelioration of cu toxicity by emblica officinalis can be explained by the fact that it prevents oxidative stress. restoration and protection of liver parenchyma by emblica officinalis against carbon tetrachloride, ethanol and iron induced hepatotoxicity has been reported25,26. micrometric studies of the hepatic lobules of rats showed statistically significant difference among groups (p-value <0.001, table-4). this gross difference in diameter between groups can be explained by hypertrophy of hepatocytes coupled with congestion in sinusoids and mononuclear infiltration in cu treated group as was reported by ghinomiet al28. limitations: this study is based on the murine model and may not be wholly applied to humans. the study could be of longer duration to ascertain chronic aspects of cu toxicity. conclusion the current research establishes the ameliorative role of emblica officinalis against copper induced hepatotoxicity in rats. its use may also be recommended in patients suffering from liver inflammation as it is cheap, easily available and is a safe dietary supplement. references 1. cheemerla s, balakrishnan m. global epidemiology of chronic liver disease. clin liver dis. 2021 may 1;17(5):365–70. 2. barber rg, grenier za, burkhead jl, cuajungco p, linder mc, tolmasky me. copper toxicity is not just oxidative damage: zinc systems and insight from wilson disease. biomedicines2021;9:316. available from: https://doi.org/10.3390/biomedicines9030316. 3. nørgaard hw. metalcraft within the nordic bronze age : combined metallographic and superficial imaging reveals the technical repertoire in crafting bronze ornaments. j archaeol sci. 2015;64:110-28. 4. mudd gm, jowitt sm. growing global copper resources, reserves and production: discovery is not the only control on supply. econ geol. 2018;113(6):1235-67. 5. manne r., kumaradoss mmrm, iska rsr. et al. water quality and risk assessment of copper content in drinking water stored in copper container. appl water sci. 2022; 12:27. https://doi.org/10.1007/s13201-021-01542 6. memon ah, ghanghro ab, jahangir tm, khand aa, muneer g, yuan q. health risk assessment of trace metals during preand post-monsoon seasons in drinking water of jamshoro, sindh. int j scieng res. 2017;8(4):1431-49. 7. bost m, houdart s, oberli m, kalonji e, huneau jfo, margaritis i. dietary copper and human health: current evidence and unresolved issues. j trace elem med biol [internet]. 2016;35:107–15. available from: http://dx.doi.org/10.1016/j.jtemb.2016.02.006 8. singh r, singh a, bhadouria r, yadav hk. impact and assessment of heavy metal toxicity on water quality, edible fishes and sediments in lakes : a review. trends biosci. 2017;10(8):1551-60. 9. andrews kw, roseland jm, gusev pa, palachuvattil j, dang pt, savarala s, et al. analytical ingredient content and variability of adult multivitamin/mineral products: national estimates for the dietary supplement ingredient database. am j clinnutr. 2017;105(2):526-39. 10. zin nm, chit ym, abubakar nf. commercial herbal slimming products: concern for the presence of heavy metals and bacteria. pakistan j biol sci. 2014;17(3):356-63. 11. khokhar zu, naveed m, ilyas m. estimation of serum copper level in hepatitis-b patients by using spectrophotometer. sci int. 2017;29(3):723-8. 12. gaetke lm, chow-johnson hs, chow ck. copper: toxicological relevance and mechanisms. arch toxicol. 2014;88(11):1929-38. 13. musacco-sebio r, saporito-magriñá c, acosta jm, boveris a, repetto mg. iron and copper toxicity in rat liver: a kinetic and holistic overview. liver res – open j. 2017;2(1):9-13. 14. lanka s. a review on pharmacological, medicinal 33 emblica officinalis reduces copper mediated inflammation and preserves liver morphology in murine…... and ethnobotanical important plant: phyllanthus. world j pharm res. 2018;7(04):380-96. 15. mandal a, reddy jm. a review on phytochemical, pharmacological and potential therapeutic uses of phyllanthusemblica. world j pharm res. 2017;6(7):817-30. 16. rao p. antioxidant effect of triphala critical review. j ayurveda integr med sci. 2017;2(1):2139. 17. wang y, zhou h, che y, wan x, ding x, zheng s, et al. emblicaofficinalis mitigates intestinal toxicity of mice by modulating gut microbiota in lead exposure. ecotoxicology and environmental safety. 2023; 15:253:114648. 18. nisar mf, he j, ahmed a, yang y, li m, wan c. chemical components and biological activities of the genus phyllanthus: a review of the recent literature. molecules. 2018;23:1-25. 19. singh mk, yadav ss, yadav rs, chauhan a, katiyar d, khattri s. protective effect of emblica-officinalis in arsenic induced biochemical alteration and inflammation in mice. springer plus. 2015;4(438):1-8. 20. golechha m, sarangal v, ojha s, bhatia j, arya ds. anti-inflammatory effect of emblicaofficinalis in rodent models of acute and chronic inflammation: involvement of possible mechanisms. int j inflam. 2014;1-6. 21. bhatti s, ranga d, meena d, agarwal m, chakrawarti a, purohit r. ameliorative effect of emblicaofficinalis in radiation and cadmium induced alteration in mice liver. world j pharm res. 2014;3(10):846-63. 22. tung y, huang c, lin j, yen g. effect of phyllanthusemblica l. fruit on methionine and choline-deficiency diet-induced nonalcoholic steatohepatitis. j food drug anal. 2018;26(4):124552. 23. deori c, das s, bordoloi sk. role of emblicaofficinalis (amla) in the prophylaxis of hepatic injury by carbon tetrachloride (ccl4) in albino rats. int j basic clinpharmacol. 2017;6(8):1992-5. 24. yadav m. change in liver histoarchitecture by sulphur dioxide induced toxicity and its vitalization by emblicaofficinalis in albino rat. j adv lab res biol. 2018;9(3):71-6. 25. kumar a, bala r, bano f, kumar r, ali m. emblicaofficinalis protects against sodium arsenite induced hepatotoxicity & nephrotoxicity in rats. int j sci. 2015;4(3):47-55. 26. sarkar r, hazra b, brain n. amelioration of iron overload-induced liver toxicity by a potent antioxidant and iron chelator, emblicaofficinalisgaertn. toxicolind health. 2013;1(2):1-14. 27. linder mc. copper homeostasis in mammals, with emphasis on secretion and excretion. a review. international journal of molecular sciences. 2020; 21(14):4932. https://doi.org/10.3390/ijms2114493 28. ghonimi wam, alferah maz, dahran n, el-shetry es. hepatic and renal toxicity following the injection of copper oxide nanoparticles (cuo nps) in mature male westar rats: histochemical and caspase 3 immunohistochemicalreactivities. environ scipollut res [internet]. 2022 nov;29(54):81923– 37.availablefom:https://link.springer.com/article/10. 1007/s11356-022-21521-2. the authors: dr. hadia zulfiqar, assistant professor, department of anatomy, ucmd, lahore. dr. muhammad suhail, professor department of anatomy, amnainayat medical college, lahore. dr. amna rehman, senior demonstrator department of anatomy, central park medical college, lahore. dr. afifa waseem, assistant professor department of anatomy, central park medical college, lahore. dr. faeza rauf, assistant professor department of anatomy, rashid latif khan university medical and dental college, lahore. dr. hafiza sadia ahmed, assistant professor department of anatomy, ucmd, lahore. corresponding author: dr. hadia zulfiqar, assistant professor, department of anatomy, ucmd, lahore. email:hadia2137@gmail.com for web full book 50 proceedings s.z.m.c. vol: 37(3): pp. 50-56, 2023. pszmc-897-37-3-2023 concept paper: it’s time to make inclusivity and positive rehabilitation the basic right of children with down syndrome: a pakistani perspective saadia shahzad department of community medicine, shalamar medical and dental college, lahore. abstract social inclusion and positive rehabilitation of children with down syndrome (ds) is the need of the hour in our society; which can be achieved by following the right principles and strategies. the current status regarding this issue at the local level has been analyzed and compared with the global status, and gaps have been identified. the author of this paper has put forward the argument that inclusivity of persons with ds ought to be the norm of this society, and early intervention onset and support mechanism of a comprehensive nature can develop these children into self-supporting independent persons. priorities and a way forward are suggested through a very comprehensive model for the local setup in pakistan. keywords: down syndrome, social inclusion, local situation, global situation, gaps, model introduction in the developed world progressive research and development in rehabilitation and social inclusion of children with down syndrome (ds) has played a phenomenal role in the upbringing of these children to the level of becoming independent and productive members of society. whereas the situation is almost in contrast in developing countries and especially in pakistan. three decades back the term ds never warranted any attention from the special education and rehabilitation personnel; hence no formal/ informal rehabilitation program for them. then there was some change in the scene and special needs schools started mushrooming in abundance in our local setup; still with no actual awareness of how to rehabilitate ds children to their optimum capacity, thus making them constructive members of the society. for the past, twenty years enhanced awareness regarding ds started infiltrating the minds of society to some extent and efforts were put in practically to incorporate these children into the special education system, to give them skill training, but rarely in an integrated educational plan. and this started happening when parents themselves came to the front with positive rehab of their child and making him/ her a success story. but these individual efforts required herculean efforts by the parents to get their child to this level. this concept paper is an effort to accumulate the experiential facts on the ground, compare them with the global situation, and come up with a first-of-its kind working model for the positive rehabilitation and social inclusion of ds children to the utmost; benefit all the stakeholders i.e. ds child/ person, parents/ immediate family, and society at large. concept paper preamble: the un sustainable development goals (sdgs) were based upon one principle that stated "equitable and universal access to quality education at all levels, health care and social protection; where physical, mental, and social wellbeing are assured"1. in a nutshell, this vision of a healthy society included differently abled and challenged children too. the un convention on the rights of the child (crc) further imposed that no child with any disability must be left out from mandatory primary and secondary education on account of their physical or mental challenge2. successful inclusion in schools and society can follow the principles given by salend3. these principles are: first: learner shares equal access, defining it as placing all the learners together in general education classrooms regardless of their learning ability, economic status, gender, learning style, ethnicity, religion, cultural background, family structure, and sexual orientation3. second: focusing on individual strengths, needs, and diversity. where diversity is defined as an inclusive classroom where all students are valued as individuals capable of learning and contributing to society. they are taught to appreciate diversity, to 6 1department of public health, health services academy, islamabad. 2department of management sciences, riphah int university, islamabad. 3department of medicine, ihitc, islamabad. 4department of rheumatology, pims, islamabad. 5department of medicine, dow university of health sciences, karachi. 70 1department of orthopedics, thq, khanewal 2department of orthopedics, lgh, lahore 3department of orthopedics, thq, ferozwala 4department of orthopedics, cmh, lahore 5department of hand & upper limb surgery, cmh, lahore 6department of orthopedics, al-rehmat benevolent trust 1 1department of anatomy, ucmd, university of lahore, lahore 2department of anatomy, punjab medical college, faisalabad medical university, faisalabad 3department of medical education, pak red crescent medical and dental college, lahore p 51 it’s time to make inclusivity and positive rehabilitation the basic right of children with down….. value and learn from each other's similarities and differences3. third: effective inclusion requires adopting reflective practices by the educators and practicing differentiation in their assessment techniques, differentiation in teaching methods, and classroom management. teachers must see each child with their specific needs and strengths3. fourth: successful inclusion needs collaboration from all stakeholders: educators, parents, families, society, government, and agencies3. teachers feel apprehensive by the presence of a special needs child in their regular class as if it is a test of their capabilities4. although developed countries like usa and uk are leading in adopting and effectively implementing a positive approach towards the inclusion of such children in the mainstream education system5 in pakistan collaboration between special education and mainstream education department is grossly lacking; gravely impacting the societal view of the special gifted children. the last principle by salend3 shows that the involvement of all the stakeholders is the key driver in this chain for the successful rehabilitation of special needs children in a community. parents/ families/ society's attitude is critical and plays an important role in the process of getting the child included in a mainstream regular school6. the past four decades of exploratory and applied research in developed countries in the sphere of positive rehabilitation and social inclusion of children with down syndrome (ds), had achieved fruitful results. globally it is estimated that 70% of children with disability including intellectual challenges of mild to moderate degree, are capable of attending regular schools; provided the institution has a supportive, encouraging/ conducive, and customized environment for these students; showing that they are willing to accommodate the children7.unicef strongly urges that the ultimate goal should be to enable all such children to become independent and productive members of society and take part in the development of their community.to meet this daunting goal, all structures and stakeholders need to collaborate and provide community-based contemporary services; accessible to all without discrimination7. current status regarding the issue: local level: history reveals that in pakistan teaching and training of special needs children/ adults was in the domain of religious bodies, which were in a minimal number at the time of independence. after independence, a few private organizations also came into the field for this purpose7. in 1959 the national commission on education for the first time placed the agenda of provision of vocational education for children/ adults with mental challenges, as well as training of teachers for this specific purpose7. the education policy of 1972 allocated funds for special education, for the first time7. in the decades of 80s and 90s, the un asked its member states to focus on the challenges/ problems of special needs persons (though there was no categorization of different disabilities as such); after publishing their report in this regard7. hence government of pakistan at that time increased its allocated budget to many special education institutions, and as a result, more than 200 institutions came into being. as per personal experience, i feel that the findings of this report and its recommendations are very much valid for the vulnerable population in question; and more importantly, these are still unmet in our setup since 2003. in pakistan, the scope of this problem includes: 1. scant research on down syndrome (ds) and their rehabilitation: regarding perceptions of parents and immediate family, problems faced by parents and families, perceptions of teachers in mainstream schools, the approach of teachers towards ds children and their rehabilitation, perceptions of the community regarding this concept and fact, scope and impact of awareness campaigns in this regard. 2. social perception and social fabric: one of the biggest hindrances to progress in this area is the social perception and social fabric of our country. till the last decade least interest and value were given to a child with a disability at the family, as well as societal level. 3. understanding of the ds child status:there is a gross lack of understanding of the proven fact that children with mild to moderate deficiency (iq of 40-70) will benefit more with constructive social inclusion at all levels; enhancing their capabilities. 4. lack of social awareness: additionally, the problem is overshadowed by the lack of awareness in society at large and the socioeconomic status of a developing country. 5. lack of contemporary professional expertise: capacity building of the professional staff is required at all levels for inclusive rehabilitation. all these factors contribute towards the minimal creation of opportunities for children with ds. against this background, this paper argues that: since the ethical principle of respect for the person 52 it’s time to make inclusivity and positive rehabilitation the basic right of children with down….. and the best interest of the child (and no harm/ less harm to the child) are imperatives, inclusivity of persons with ds in society ought to be the norm. for this social inclusivity to happen it is important to know (and overcome) the hindrances. inclusion in educational institutions and thus social inclusion of ds persons depends on some factors that can be separated into the following domains: 1. family: parental literacy, socio-economic status, family size (number of children in the family), philosophy of life in a household, attitudes of parents and family members, and family influence in a society like ours. above all the will and determination of parents as groups is the key driver behind the achievement of this change in a community. though many parents are working at the individual level in our country coordination and collaboration among all of them is missing; as a result, unified influence is not achieved at the required levels in the society. parents' attitude in all respect within the home and outside is the foundation stone for making a positive outcome-based road map of rehabilitation. research has shown that the key points driving towards choosing or not choosing inclusive education might depend upon basic socio-demographic factors. 2. societal: a) social attitudes: cultural and social norms, and lack of correct societal awareness towards this concept and its desired outcome. b) social awareness: at the policy level, expert human resource for inclusive education, intervention services' availability, and accessibility by all (from the very start), education experts who are well aware of the idea of inclusiveness of ds children, planning and implementation of the right policies for these children at the state level (since there are none at the moment), and inter-sectoral collaboration. furthermore, the ethical concept of the best interest of the child and no harm/ less harm to the child is applicable in this situation. it needs to be practiced both by the parents/ family and the society at large8. research has shown that the key points driving towards choosing or not choosing inclusive education might depend upon basic sociodemographic factors9. inclusive education in pakistan was started in 1994 by the forceful push of unicef; which started the system of enrolling children with disabilities in every school10. it was reported in a previous study that mainstream schools are a much better opportunity for children with disabilities; if the problems associated with teaching these children in mainstream schools are tackled appropriately by expert staff11. in a study conducted by unicef, the focus was on the policies and practices in inclusive education for children with disabilities in pakistan7. the few inclusive schools that demonstrate good practices are limited to bigger cities and in the private sector; neither accessible nor affordable to all7. there is a lack of knowledge as to what capability the differently abled children can have superimposed by the lack of educational facilities, this puts them in a very low profile.7 at the local level for ds children there is a gross lack of awareness for rehabilitation; inclusive education; comprehensive social inclusion; support, counseling/ mentor system for parents; coordination between the ministry of education, social welfare, and special education; and early intervention system. even at the federal level in pakistan, the implications for inclusive education for ds children are recognized in principle onlyand not practiced at a larger scale7.my argument is: that early intervention onset and support mechanisms of a comprehensive nature can develop these children into self-supporting independent persons. making a child with ds an independent member of society is the desired outcome when all the rehabilitation programs are working. thus ds persons are having a much fuller life in the west as compared to developing countries. locally, to date, the following has been done. 1. there is an increased number of special needs schools in both the public and private sectors that take in some number of ds children as per their vacancies. 2. special olympics pakistan is giving opportunities to children with ds in sports. 3. mainstream inclusive education is available in a scant number of schools, but the whole package is unaffordable for the majority. furthermore, teachers in mainstream schools are not specifically trained for this purpose. 4. some parent support groups have evolved in the past decade that are trying to give support to parents; again in bigger cities and among affluent populations. a few groups of affected and educated parents joined hands to make organizations like karachi down syndrome program (kdsp) for rehabilitation purposes, providing good services to the community. 5. few organizations in the private sector are providing structured programs for job placement for children with disability (physical); but not for ds children on account of their mental challenges. 6. mcdonald's (an international franchise) is offering a structured job placement program for 53 it’s time to make inclusivity and positive rehabilitation the basic right of children with down….. well-rehabilitated ds children/ persons too, for the past ten years or so. global level globally in the past 5-6 decades, vast exploratory and applied research has been conducted in the area of ds. studies have been conducted to search the impact of having a child with ds on family12,support system13,community values14, structured rehabilitation program, independent development, job customization and placement, and many other areas of life. west has identified as many as twenty research gaps still, in the area of rehabilitation of ds and its impact; to name a few better characterization of the natural history of cognition, language, and behavior, identification of mental health comorbidities, and risk and protective factors for their development, understanding the effects of educational and social services and supports.globally current state of knowledge regarding ds encompasses the areas of epidemiology, physical and mental health, cognitive and linguistic profiles, aging, impact on family, and current strategies for educating professionals, families, and the general population at large; has been studied15. local global scant research in ds rehab; unable to make use of the global research evidence in this regard optimum applied research in the area of ds rehabilitation. minimal awareness in the majority local population regarding ds rehabilitation and inclusion social awareness of ds inclusion across cultures is better16.. early intervention programs grossly missing for the child after birth early intervention is available from day one of birth and its effectiveness is determined17. no formal support services are available to parents and families (counseling, guidance, social support, a mentor system, etc) early intervention is available for mothers18. no affordable structured occupational therapy services available to all contemporary methods are available to all. capacity building of specialized human resources for ds inclusionand rehabilitation is missing a large number of ds rehabilitation experts are available. registration as a citizen with the issuance of scnic through disability certificate issuance; has started recently19. the implementation of the benefits (health coverage, transport/ fare coverage, etc) of this is yet to be seen for ds children/ persons all basic facilities and services are covered for ds children a formal policy for social inclusion at the school level/ customized skill training/ job placement/ entrepreneurship/ social unicef's newer revised guidelines clearly state the inclusion21. clubs' services is in place, but its effective implementation is not there at all for ds persons20. child protection bill does not state the term down syndrome specifically22.so things are left to the perception of the reader. the national down syndrome society (usa) advocacy & public policy team works directly with legislators and administrators to make sure their work is increasingly inclusive of the down syndrome community and progressively beneficial. no authentic and specialized central record maintenance of ds person record maintenance is there lack of collaboration between the stakeholders (mainstream education, special education, legal, social/ welfare services, health care, and families); for inclusive rehabilitation of ds children unicef's newer revised guidelines clearly state the significance of collaboration between the stakeholders21. lack of collaboration between current resources working for ds children; in both the public and private sector collaboration is there as many such organizations are working across the globe e.g. ndss in the usa, down syndrome association uk, ds association scotland. table-1: gaps in the handling of this problem locally and comparative global situation. few parents and special institutions understand the philosophy behind the rehabilitation of ds children duly supported with scientific evidence. as a result, there is a gross difference in the results for ds children in pakistan and globally. priorities and suggested way forward in pakistan: as a public health specialist, i propose the following model for the rehabilitation of children with ds. it is important to understand that ds children/ persons have different needs as compared to other physical as well as mental disabilities; and the disability act, 2020 by the government of pakistan does not take into account this significant difference. as a result, things have not worked out well for children with ds uptil now. i have tried to take into account all the required organizations and levels of society for the successful rehabilitation of ds children; with the outcome of making them productive members of society and independent and confident individuals. still, in my opinion, this model will not be practically successful unless and 54 it’s time to make inclusivity and positive rehabilitation the basic right of children with down….. until maximal sensitization upon it is done for all stakeholders and state/ society to join hands in it. as the result of my experiential learning, and as a public health specialist i have tried to develop a working model for this purpose. i propose the following working model for the rehabilitation and social inclusion of children with ds: fig-1: model for the positive rehabilitation of children with down syndrome. 1. parent support groups should be developed to build networking among the parents and support system development. 2. society awareness needs to be raised through various sustained long-term strategies like the use of electronic/ print media, seminars, talks, and symposia; on a war footing. 3. the mainstream integrated educational system can be effectively pulled up through capacity building of the staff for this specific purpose; individual educational plan development; and pulled-back programs. 4. a reliable and trustworthy peer and buddy system needs to be introduced in society at all levels for these children, starting from the household level. these children have a great tendency and capability of learning by imitating others and the buddy system encourages them to a great extent. 5. social inclusion and networking can be achieved through membership in social clubs/ sports clubs/ public events and household events etc. 6. subsidized medical care is a need that can only be provided through central policy implementation. 7. collaboration of government and private sector is necessary for the creation of customized skill training opportunities; customized job creation where needed and sustenance of these activities. 8. legal safeguard of ds children through the right policy making, implementation, and monitoring and evaluation (m & e). 9. working organizational bodies are a combination of policy makers + learned parents who have personal experience with ds child rehabilitation. 10. central policy-making specifically for ds children/ persons is needed as all types of disabilities have a different scope of rehabilitation 11. implementation and then m & e of the policies is very important to meet the objectives. 12. feedback should be disseminated to the stakeholders at individual and collective levels 13. there is a dire need for authentic record maintenance of ds children and persons with details of the level of challenge. 14. central ds regulatory bodies are required at the provincial as well as central levels, to manage the system of rehabilitation for the ds children/ persons. conclusion educated and experienced parents who have worked with their such children need to get together in greater numbers to build up their resources that work for the improvement of this situation in the suggested model, supported by philanthropists, special education experts, educationists, health care professionals and things can be set in motion at some pace. there is a dire need for policy making upon social inclusion for ds children that takes into account all the required levels of positive rehabilitation, with the clear and precise outcome of making these children confident, positive, and productive human beings. they must not become a liability for themselves, family, and society; when they have all the capability of becoming constructive members in all respects. stakeholders need to work hand in hand, and drivers would definitely be these parents themselves, there is no other option or solution to it locally. i as a mother of a son with ds and a public health specialist, reinforce the arguments given above with my practical life example and stand by my arguments that acceptance, inclusiveness, social 55 it’s time to make inclusivity and positive rehabilitation the basic right of children with down….. support, the best interest of the child, no harm/ less harm to the child, result in oriented efforts to achieve the objectives; all are contributive towards the development of a ds child into an independent and productive member of the family and society. my son is working in the mcdonald's chain on half time salaried job since 2017 and recently got promoted to training manager in sep 2021. i support my argument given above that parents need to become unified and start helping each other and society to understand the problem and work in a result-oriented manner. the practical example is there to support it too as i am the founder of a parent support group myself and another has been mentioned above. i can vouch with all my conviction that these children are better human beings than many of us on account of their innocent character, love, friendliness, caring attitude, and untiring willingness to learn with due encouragement and guidance. we need to learn from them the art of being better human beings. they bring with them sunshine of happiness and love and smiles. statements of declaration funding: the author declares that no fund, grant, or other support was received during the preparation of this manuscript. competing interests: the author has no relevant financial or non-financial interests to disclose. this article was presented by the author at the 14th world conference on bioethics, medical ethics, and health law in porto, portugal in march 2022. author contribution: i am the sole author of this manuscript conception, working model design, and write-up. ethics approval: ethics approval was not required as this was a thought paper and no human subject was involved in any data collection, so consent to participate was not required. consent for publication: the author consents to the publication of the manuscript with the condition that the model developed and given in the manuscript shall not be published separately by another name and the copyrights of the given model will stay with the author. strengths locally scant research work has been done in this specific area. hence, this concept paper will be a comprehensive addition of experiential knowledge and ground realities combined. it can be taken as the foundation and varied avenues for future research can open from it. for the reader's interest and benefit of the researchers, the author has built up the concept as a story; 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2020. 21. united nations children’s fund, disability inclusion policy and strategy (dipas) 2022–2030, unicef, new york, december 2022. 22. the gazette of pakistan 2018. child protection act; 2018. corresponding author: dr.saadia shahzad, associate professor, community medicine, shalamar medical and dental college, lahore. email: saadiazahur@live.com