IBN AL- HAITHAM J. FOR PURE & APPL. S CI. VOL. 24 (3) 2011 Evaluation Interleukin ─ 8 and High Sensitivity C - Reactive Protein in Sera of Control Group and Human Female Patients with Osteoarthritis Nodal in Iraqi's Patients B. H. AL-Wihaly Departme nt of Chemistry ,College of Education Ibn – Alhaitham, Unive rsity of Baghdad Received in : 13 March 2011 Accepte d in : 30 May 2011 Abstract The p resent st udy conducted on 30 female p atients with ost eoarthritis OA a att ending Baghdad teaching hosp ital, in addition to 30 healthy females, all subjects were (35-65) y ear's old .Parameters were measured in the sera of p atients group and healthy group s were interleukin -8 (IL-8) and high sensitivity C-reactive p rotein (hs-CRP).The results revealed a higher significant in all p arameters of p atients sera comp ared to healthy group s. Conclusion was obtained that IL-8 and hs- CRP were a markers or p lay er inflammatory disease like ost eoarthritis nodal Key words: Interleukin -8(IL-8),High sensitivity CRP(hs-CRP) ,Ost eoarthritis Introduction Osteoarthritis (OA) is a slow p rogressive disorder of sy novial joints .T his joints disorder is characterized by a loss of balance between sy nthesis and degradation of the articular cartilage constituents leading to subsequent erosion of joint cartilage remodeling of the underly ing bone oseop hy te formation and variable degree of sy nop itis [1].When clinical characterist ics of OA (p ain loss of mobility and radiographic narrowing of the joint sp ace ) manifest , the actual changes in articular cartilage and subchondrla ,bone have st arted long ago[2]. Interleukins are group of cytokines [3] because they regulate in formation transfer among different ty p es of leukocytes during various st age of immune or inflammatory resp onse [4] secreted Prot eins/signaling molecules/ that were first seen to be exp ressed by white blood cells (leukocytes). The term interleukin derives from (inter-) as a means of communication '' and (-leukin)'' deriving from the fact that many of these proteins are p roduced by leukocytes and act on leukocytes, it has since been found that interleukins are p roduced by a wide variety of body cells . The function of the immune sy st em depends in a large p art on interleukins and rare deficiencies of number of them have been described, all featuring autoimmune diseases or immune deficiency , the majority of interleukins are sy nthesized by help er CD4+T lymphocytes as well as through monocots macrophages, and endothelial cells. They p romote the development and differentia on of T, B, and hematop oietic cells. Interleukin – 8 one of these interleukins secreted from macrop hages, lymphocytes, epithelial cells and endothelial IBN AL- HAITHAM J. FOR PURE & APPL. S CI. VOL. 24 (3) 2011 cells, CXCR1/IL8RA, CXCR2/IL8RB/CD128 were the target receptors, Neutrop hiles, lymphocytes were target cell [3] so as IL-8 was chemotaxis for Neutrop hils [5]. C-Reactive protein (CRP) is member of the class of acute-phase reactant as its levels rise dramatically during inflammatory p rocesses occurring in the body [6, 7][atherosclerotic p laques][9] .CRP binds to p hosp ocholine on microbes, it is thought to assist in comp lement binding to foreign and damaged cells and enhances p hagocytosis by macrophages (op sonin mediated p hagocytosis) ,which exp ress a receptor for CRP[10]. It is also believed to p lay another imp ortant role in innate immunity as an early defense sy st em against infections [11] so is used as a clinical marker of a cute sy st emic inflammation and infection [9] CRP a rise up to 50,000 fold in acute inflammation such as infection .It rises above normal limits within 6 hours, and p eaks at 48 hours [12] . A high – sensitivity CRP (hs-CRP) test measures low levels of CRP using laser nephelomoetry the test gives results in 30 minutes with sensitivity down t o 0.04 mg\ L. Normal concentration in healthy human serum is usually lower than l0 mg \L, Slightly increasing with ageing, higher levels are found in late p regnant women, mild inflammation and viral infection (10-40 mg \L), severe bacterial in fections and burns 200 mg \L [13] Aim of the S tudy The aim of this st udy is to evaluate some biochemical p arameters such as interleukin - 8, (IL-8), hs-CRP in sera of OA female p atients which is used as a clinical marker of an acute inflammation and infection. Experimentaly S ubjects The p resent st udy was p erformed on a group of 30 human females aged (35-65) y ears with on diagnosed by examining the p atients by in the (Baghdad teaching) hosp ital and by x- ray examination on. In addition, a group of 30 healthy females were enrolled in the st udy as control group . S ampling Blood samples of 5ml were drawn all from subjects enrolled in this st udy , and kept in p lain tubes 1eft t o clot at room temp erature for /5 min. then centrifuged at 3500 g for 10 min to sep arate the serum. - Inte rle ukin – 8 (IL-8) determinations Interleukin 8(IL-8) has been estimated by using enzy me immunoassay (ELISA) technique. The procedure was done according to the manufacturer instruction as sup p lied with kit from Biosource Europe S.A. Comp any, Belgium. - High se nsitivity C - reactive protei n (hs CRP) determination - High sensitivity C - reactive p rotein (hs CRP) enzy me immunoassay for the quantitative determination in human serum was used. DRG international. Inc. USA. T his is done by ELASA test. S tatisti cal analysi s The results were exp ressed as mean± SD of mean, using st udent-t est, significant Variation is considered significant when p -values are ≤ 0.05. Re sults and Discussion Results obtained in the present study st ate that t he serum level of IL – 8 was significantly higher in females p atients with OA than in healthy control. (Table1),figure(1). The elevated IL-8 level found in the serum of females' p atients with OA of the p resent st udy is in agreement as in pervious studies [14, 15] who found increase in the level of IL-8 in serum of OA p atients comp ared with control and found that serum levels of IL-6 in arthritis p atients [like fibro myologi a sy ndrome (FM S) p atients] was t he same as in controls. IBN AL- HAITHAM J. FOR PURE & APPL. S CI. VOL. 24 (3) 2011 IL -8 is t he major chemoattractant for neutrop hils; it mediates changes in neutrop hil shap e. Transendothelial migration, induces exocytosis of granule enzy mes such as elastase and myelop eroxidase and a resp iratory burst where hy drogen p eroxide and sup er oxide radical are p roduced [15] IL-8 was up regulated by p roinflammatory signals such as IL-1 and IL-17 and inhibited by anti -inflammatory cytokine (IL-4, IL-10) and glucocorticoids [16] furthermore IL-8 has been defined as a mediator of sy mpathetic p ain [17]. C - reactive protei n Results obtained showed that the serum level of CRP was significantly higher in p atients with OA than in controls (T able1),figure(2). These results are in agreement with other st udies reported that CRP level is regularly used in the clinic, which is useful to discuss it in terms of a diagnost ic tool and therefore as indicator of an underly ing disease in acute infections.[18] Less in known on the CRP level in chronic disorders t hat have an Inflammatory disease comp onent [19] High sensitivity C-Reactive p rotein activates the comp lementy sy st em with a p rimary function to bind and detoxify endogenous toxic substance p roduced as a result of tissue damage [20] CRP is not only a marker of, but a p lay er in, cardiovascular disease (CVD)., CRP may directly p romote atherosclerosis and endothelial inflammation by att enuating the release of NO, a key molecule in the endothelium that p lay s a pivotal role in the maintenance of vascular tone [21] In addition, besides the p otent vasodilator effects of NO, it meditates many p rotective functions in the endothelium, for examp le inhibiting the of p ro-inflammatory cytokines, chemokines and leukocyte adhesion molecules, there by limiting vascular recruitment of leucocy tes and p latelets [22].NO also inhibits vascular smooth muscle cell p roliferation, an early sign of atherosclerosis. Therefore CRP may have an imp ortant deleterious role in the atherosclerotic p rocess. i .e .by inhibiting NO formations however the involvement of CRP as a functional p lay er in the endothelium has recently been questioned as it has been suggested that the p ro-inflammatory effects by CRP might be due to contaminating artifact in the commercial CRP p reparation [23]. Over the last few y ears t here have been a lot of p romising clinical markers p rop osed to link inflammation and atherosclerosis. One of these markers, hs – CRP [24].And these st udies are in agreement with this st udy . From the present study , a conclusion could be st ated that IL-8 increase concentration in sera of OA p atients is due to interleukins that, regulate information transfer among different ty p es of leukocytes during various st age of immune or inflammatory resp onse, which is confirmed by increase in hs –CRP concentration. This finding may indicate that inflammatory activity might influence cytokine p roduction in these p atients [25] [26] Re ferences 1-Ana M .Valdes and Timothy , D. Specttro (2009) The contribution of Genes to osteoarthritis 66-(1) :4593J.M edical Clinical of North America 2-Robert, B.Wallace; Johan, M . Last , et al(2008) "Public Health & Preventive M edicine " 50 th Edition ,M cGraw .Hill Companies ,Inc(U.S.A) p .1128-1129 3- Lip p incott Williams and Wilkins (2007) :Lipp incott's Illustrated Reviews Immunology . 3 th Edition Lipp incott Williams Wilkins ,U.S.A Pa ge 68. 4- Devlin M .Thomas: cytokine Recep tors (2006): text book of Biochemist ry with clinical correlations, 6 th Ed. A John Wiley &Sons, Inc. , USA, p .507. 5- Karen M .Henkelsa., et, al. (2011). IL-8-induced neutrop hil chemotaxis is mediated by Janus kinase 3 (JAK3). Edited by Beat Imhof 585(1): 159-166. 6- Danesh ,J. ;wheeler ,JG.; Hirschfield ,G.M . et al (2004) C-reactive p rotein and other circulating markers of inflammation in the prediction of coronary heart disease .N. En gl 1397 -: 1387 350.J. M ed. (3): 74 30reactive p rotein, J. Australian p rescriber -R. (2007) C,Gleen -7 8- Harp er ' s (2006) Harp ers lustrated Biochemist ry 27 th ed. Lange med ical Books M cGraw- Hill M edical publishin g Division NewYork p 590- 591. 9- Päiväniemi O.E. paulak, et al (2009) : Local C-Reactive p rotein Exp ression in obliterative lesions and the Bronch ial wall in post transp lant obliterative Bronchiolitis Volume 2009(2009), Art icle ID 510254, 7 p ages doi : 10. 1155/2009/510254. 10- Pepy s M.B. Hawkins p .n., et al. (2005) :p roinflammatory effects of bacterial recombin ant human C-reactive protein are caused by contamination with bacterial p roducts , 103.-:97 97reactive protein itself .Circ.Res. -not by C 11- Pickup J.C, (2004): Inflammation and activated innate immunity in the p athogenesis 823.-:81327of type 2 diabetes .Diabetes care 12- Ron Kennedy.M .D.(2011) C-reactive Prot ein (CRP), Cardiac C-reactive Prot ein, (Hi gh Sensitivity C-reactive Potein) .The Doctors' M edical Library . 13- Castoldi G., Galimberti S., et al .(2007): Association between serum values of C-reactive p rotein and cy tokine p roduction in whole blood of p atients with type 2 diabetes .Clin . Sci. 108 -: 103113 14- Gur A, and Karakoc M .(2002) Cytokines and depression in cases with 61. -(2): 35829fibromy algia .J. Rheumatol. 15- Ransohoff RM , and Benvenist c EN.2005 Cy tokines and the CNS .2 th ed .London, New York: Tay lor &Francis Boca Raton ; pp 217. 16- Uceyler N, Valenza R, and Stock M .2006. Reduced levels of anti- inflammatory cytokines in patiente with chronic widesp read p ain : Arthritis Rheum ; 54 : 2656- 2664 . 17- M ey er TD , Hautninger M , Allgmine . (2003) .Depressions Skala Normuerun g an M inderjahrigen and Erweiterung zur Erfassung manischer sy mptom .Blood 365.-:360132mononuclear cells. Clin EXP Immunol. 18-Salzman B, . E. (2007). A p rimary care ap p roach to the use and interp retation of common rheumatolo gic tests. Clin Fam Pract, 335-358. 19- M arnell L,M old C, DU Closs T W .(2005) C-reactive p rotein: ligands, recpcors and role 11.-:104117 in inflammation .Clin Immuno l; 20- Devaraj S, and Du closs TW , Jialal I.(2005): Bind ing and internalization of C- Reactive protein by FC gamma recep tors on human aortic endothelial cells 3.136-:135925mediates biolo gical effects . Arerioscler thromb vasc Biol. 21- Verma, S., Wan g, C, H., Li, et al. (2002). Aself - fulfilling p rop hecy : C-reactive p rotein att enuates nitric oxide p roduction and inhibits an giogenesis. Circulation 919.-,913106 22- Verma , S., Kul iszewski , M . A., Li, S.H. et al (2004)C- reactive protein att enuates endothelial p rogenitor cell survival , differentiation and function : further evidence of a mechan istic link between C- reactive p rotein and cardiovascular 2067.-,2058109isease .Circu lation d 23-p epy s, M .B., Hirshfield,G.M .,Tannent ,G.A.Et. al. (2006) Targeting 1221 -: 1217440reactive protein for the treatmeent of cardiovascular d isease. Nature -C 24- Szmitko, P.E., Wan g, C.H., Weise l, R.D., de Almeida, J.R., Anderson ,T .J. and Verma , S.(2003) New markers of inf lammation and endothelial cell activation I. 1923. -,1917108Circulation -:79113Clin ical .Science . reactive protein: amarker or play er? -Thomas N. (2007): C -25 81. 26- Gaede, P., Ved , P., L arsen, N., Jensen, G.V., Parvin g, H.H. and Pedersen , O. (2003)M ultifactorial intervention and cardiovascu lar disease in p atients with type 2 393. -:383348diabetes N. Engl .J. M ed . IBN AL- HAITHAM J. FOR PURE & APPL. S CI. VOL. 24 (3) 2011 IBN AL- HAITHAM J. FOR PURE & APPL. S CI. VOL. 24 (3) 2011 Table (1): IL-8 and hs –CRP concentration i n sera of OA patients and control P value Control N=30 OA p atients n= 30 p <0.05 84.91±14.58 120.66±37.42 IL-8 Pg/ml M ean ±SD p <0.05 0.583±0.141 1.211±0.524 Hs-CRP mg/dl M ean± SD Fig .(1) : IL-8 concentration in sera of OA patients and control. Fig .(2): ) hs –CRP concentration in sera of OA patients and control. 2011) 3( 24المجلد مجلة ابن الھیثم للعلوم الصرفة والتطبیقیة في مصل الحساسیةتقدیر االنترلیوكین الثامن وبروتین سي الفعال العالي مریضات بالتهاب المفاصل العقدي في العراقمجموعة نساء أصحاء يالو حیلبشرى حمید جامعة بغداد ،كلیة التربیة ابن الهیثم ،قسم الكیمیاء 2011آذار 13:استلم البحث في 2011آیار 30:حث في قبل الب الخالصة مـن 30 آل عـنضـفمن مستشفى بغـداد التعلیمـي أخذتمریضة بالتهاب المفاصل العقدي 30تضمنت هذه الدراسة ،الـدم للمجمـوعتین وقیسـت المتغیـرات الحیویـة فـي مصـل، ةسـن 35-65المجمـوعتین بـین أعمـاروتراوحت .األصحاءالنساء إحصـائي معنـوي محسـوس مرتفـع فـرق میـع النتـائج جعكست . )الحساسیةوبروتین سي الفعال العالي ،ثامناالنترلیوكین ال( مثـل ،االلتهابیـة اإلمـراضلمعرفـة اهذین المتغیرین مقیاس المعنستنتج من ذلك انه یمكن است .المجموعتین في مصل نساء التهاب المفاصل العقدي