IBN AL- HAITHAM J. FOR PURE & APPL. S CI. VOL.23 (2) 2010 De termination of Adenosine De aminase Activity in type 1 and type 2 Diabetes Mellitus H. Mossa and Z. Hussein Departme nt of Biology, Collage of Education Ibn-Al-Haitham Unive rsity of Baghdad Abstract Serum adenosine deaminase (ADA) activity was determined in 30 blood sample of type 1 diabetic individuals 30 blood sample for the ty p e 2 and 15 normal children as a control for ty p e 1 15 normal adults as control for ty p e 2. The mean ADA activity and sp ecific activity in ty p e 1 was (8.85± 5.55 U/mg of p rotein) which is comp ared with control (32.11± 1.54 U/mg of p rotein) while in ty p e 2 was (48.46±11.91 U/mg of p rotein) is comp ared with control (5.18± 2.27 U/mg of p rotein ). We conclude that t he altered blood level of ADA activity may help in p redicting immunological dy sfunction in diabetic individuals and also has a p rognost ic value. Introduction Diabetes mellitus is a group of devast ating metabolic disease caused by insufficient insulin sy nthesis, increased insulin destruction or in effective insulin action. All of its metabolic effect result when the body 's cells fail to acquire glucose from the blood. The metabolic imbalances that occur have serious, but not life-threatening, consequences (Figure 1). In insulin dependent diabetes mellitus (IDDM ), also called ty p e 1 diabetes, inadequate amount of insulin are secreted because the B-cells of the p ancreas were destroy ed. Because IDDM usually occurs before the age of 20, it has (until recently ) been referred to as juvenile- onset diabetes. Non insulin dependent diabetes mellitus (NIDDM ), also called ty p e 2 or adult- onset, is caused by the insensitivity of target tissues to insulin. Although these forms of diabetes share some features, t hey differ significantly in other. The most obvious sy mptom of diabetes in hy p erglycemia ( high blood glucose levels), is caused by adequate cellular up take of glucose. Because the kidney s cap acity to reabsorb glucose from the urinary filtrate is limited, glucose app ears in the urine (glucosuria). Glucosuria results in osmotic diuresis, a p rocess in which an excessive loss of water and electrolytes (Na‾, K, and Cl‾) is caused by the p resence of solute in the filtrate. With out insulin to regulate level metabolism, its three p rincipal target t issues (liver, adip ose tissue, and muscle) fail to absord nut rient app rop riately . Inst ead, there tissues function as if the body were undergoing st arvation. Insul in- Dependent diabetes In most cases of insulin-dependent diabetes the insulin p roduced B-cells have been destroy ed by the immune sy st em. Although the sy mptom of IDDM often manifest themselves abruptly, it now app ears that B-cell destruction is caused by an inflammatory p rocess over several y ears. The sy mptom are not obvious until virtually all insulin p roducing cap acity is destroy ed. As in other inflammatory and autoimmune processes, B-cell destruction is initiated when an antibody bind to cell surface antigen. Aut o antibodies to insulin and the ty rosine p hosp hotase. IA-2 have been detected. The most acute sy mptom of ty p e 1 diabetes is ketoacidosis. Elevated concentration of ketones in the blood and low blood PH a long with hy p erglycemia cause excessive water losses, ketoacidoses and dehydration, it left untreated, can lead to coma and death. Certain HLA antigens are found in a large majority of ty p e 1 diabetes. IHJPAS IBN AL- HAITHAM J. FOR PURE & APPL. S CI. VOL.23 (2) 2010 Non-insul in-dependent diabete s Non-insulin-dep endent diabetes is a milder disease them the insulin dependent form. Its onest is slow, often occurring after the age of 40. Individual with ty p e 2 diabetes have normal or often eveated blood levels of insulin. Ty p e 2 p atients are resistant to insulin. The most common cause of insulin resistance is the down-regulation of insulin receptor. Ap p roximately 85% of ty p e 2 diabetics are obese. Treatment of NIDDM usually consists of diet controland exercise. In same cause oral hy p oglycemic drugs are used. When the failure of ty p e 2 diabetic p atients to control hy p erglycemia is accomp atied by other medical condition ( e.g infection) a serious metabolic st ate referred to as hy p erosmolar hy p erglycemic nonketosis (HHNK) can result. Because of the additional metabolic st ress, insulin resistance in exacerbated, and blood glucose level would rise[1]. Adenosine deaminase (ADA),as an enzy me that is involved in nucleic acid metabolism [2]. Its main biological activity is defected in T lymphocyte function[3]. So it was considered as good marker of cell mediated immunity [4], and it has acrucial role in lymphocyte p roliferation and differentiation [5]. It has been reported that adenosine deaminase is a good marker for insulin function [6,7]. But its connection with immune sy st em was not y et established in diabetic subject. Even through there are some reports available on ADA levels in diabetic subject there are all inconclusive and controversial [8]. We have undertaken apreliminary st udy to determine its blood activity and to highlight its role in ty p e 1 and ty p e 2 diabets mellitus. Material and Methods There were 30 blood samp les for adult p atients form (both sex) who had a history of not less than six y ears of diabetes mellitus (sample were collected from AL-Yarmuk hosp ital). They were aged 20 to 50 y ears. All of them were in the category of ty p e 2 diabetes mellitus. None of subject have a hist ory of infection or other factor (like drugs) at the time of the st udy . And t here were 30 blood samples of p atients with ty p e 1 diabetes mellitus and there were in range of 5 to 16 y ears (samplewere collected from AL-Yarmuk hosp ital).Non of the subjects have ahist ory of infection or other element at the time of st udy . A group of 15 healthy adult individuals were served as control and 15 healthy children served as control for ty p e 1. ADA activity was determinded according to the Giusti method (9). T he tot al activity was defined as the amount of enzy me required to release 1 mol of ammonia p er minute from adenosine at st andard assay condition and it was exp ressed as U/ml. The sp ecific activity was exp ressed as U/mg of p rotein. The st atist ical analysis was p erformed by using T - test to comp are the mean value of ADA in p atients with control. Re sults Adenosine deaminase activity was significantly (p<0.01) decreased in p atients with ty p e 1 sy mptom, it was (8.85± 5.55 U/mg of p rotein) as comp ared with control (32.11 ± 1.54 U/mg of p rotein). While sp ecific activity in patients (2.01± 1.25 U/mg of p rotein) as comp ared with control (9.44 ± 0.78 U/mg of p rotein). The results in p atient with ty p e 2 show a significant increased level in ADA activity , it was (48.46 ± 16.91 U/mg of p rotein) in p atient as comp ared with control ( 5.18 ± 2.27 U/mg of p rotein). While the sp ecific activity levels in p atients were (2.01± 1.26 U/mg of p rotein) as comp ared with control which were (9.44 ± 0.78 U/mg of p rotein). The activity and sp ecific activity of p atients and control for ty p e 1 and ty pe 2 were shown in table 1. Discussion Enzy me is useful in modern medical p ractice for several reasons. Enzy me assay s p rovide imp ortant information concerning the p resence and severity of disease. In addition, enzy me often provide a means of monitoring p atients resp onse to therap y . Genetic IHJPAS IBN AL- HAITHAM J. FOR PURE & APPL. S CI. VOL.23 (2) 2010 p redisp osition to a certain disease may also be determined by measuring sp ecific enzy me activities [1]. Adenosine deaminase is an enzy me necessary for the normal catabolism of p urine. ADA cataly ses the conversion of adenosine and deoxy adenosine to inosine and deoxy inosin. Exp erimental evidence indicates that adenosine, in increased amount, may result in increased cAM P activity . which is known to be associated with the inhibition of ly mphocyte function so immunodeficiency may result as consequenes of ADA defect [10]. In this st udy we observe that t here were significant decreases in the ADA levels in ty p e 1 individuals while there were elevated levels of ADA activity in ty p e 2 individuals. In regard to ty p e 2 our finding is similar to result rep orted by [6,11]. The decreased level of ADA activity in ty p e 1 may result from the detect in the action of insulin that is required for the function of ly mphocyte. It is also thought that in diabetic individuales, insulin may be a good target for killing by antibody dependent cellular cytotoxicity resp onse [12], that has control over T-lymphocyte function[8]. The increased level of ADA in ty p e 2 may releated to t he elevated level of insulin in blood, insulin has a modulating action on immune resp onse [13]. So according to this st udy we can conclude that t here were immunological disturbances associated with this disease, and the altered level of ADA may help in est ablishing this enzy me as a good marker for assessing CM I in diabetes individuals. However, this st udy has a few limitation, further studies of ADA in lymphocyte DM individuals is required. Re frences 1- Trudy M ckee,and Tames R. M ckee,(2003) Biochemist ry . 3 rd ed., : 550-554. 2- Amja, P.O.,Ekanem, E.A., Short Term.(2005) Glucose load and serum Adenosine Deaminase Activity in Diabetic Nigerians. Journal of Medical Lab. Oratory science; 14:1. 3- M ishra, O.P.; Gup ta, B.L.; Ali, Z.; Nath, G.and Chandra, L. (1994) Adenosine deaminase Activity in ty p hoid fever. TNDIAN PEDIAT RICS: 31:1380-1383. 4- Bayhanha, M .F.; Pego, A. Lima, M .A.etal.(1990) Serum and p leural adenosine deaminase correlation with ly mphocyte pop ulation. Chest ; 87: 605-610. 5- Sullivan, J.L.; Osborne, W.R.A.;Wedgwood, R.J.(1977) Adenosine deaminase activity in lymphocytes. Br. J. Hematol. 37:157-158. 6- Kurt ul, N.; Penu, S.; Akarsu, E.; Kocogla, U.; Akosoy , Y.and Akosoy , H.(2005) Adenosine deaminase activity in the serum of ty p e 2 diabetic p atients, Acta. Medic (Hradec Kralove), 48(1):63. 7- Hoshino, T.; Yamada, K.and M asuoka, K. et al. (1994) Elevated adenosine deaminase activity in serum of p atients with diabetes mellitus. Diabetes Res Clin Pract; 25: 97-102. 8- Frankie, B. and Abass, E. (2003) Activated T-ly mphocyte in ty p e 2 diabetes: imp lications from in vitro studies. Curr. Drug Targets, 4:493-503. 9- Giust i, G. Adenosine deaminase. In: Bergmey er, H. U. (Ed.) M ethods of Enzy me Analysis.(1981). 2 nd ed. Academic Press., 2:1092-1099. 10- Daniel, D.;Stites,John D.;Stobo, J.,Vivian Wells (1987) Basic and clinicl immunology , 6 th ed.,:339-340. 11- Casanueva, V.; Ximena, C.;Cavicchioli, G.;Oelkev, M .; Cofre, J.and Chiang, M .I.(1992) Serum adenosine deaminase in the early diagnosis of ty p hoid fever. Pediatr. Intec. Dis. J.,11:828-830. 12- Foveny , J.J., K., T otp al, E., Thassz, ,T, Garan (1984). 13- Ivan, M ., Roitt and Peter, J., Delves (2004) 10 th ed.: 214-215. IHJPAS IBN AL- HAITHAM J. FOR PURE & APPL. S CI. VOL.23 (2) 2010 Table (1): Adenosine deaminase activity in serum of diabetes patie nts type 1 and type 2. Sp ecific activity U/mg of p rotein ADA activity U/mg of p rotein Number Age (both sex) 9.44 ± 0.78 32.11 ± 1.54 15 control 5-13 2.01 ± 1.26* 8.85 ± 5.55* 30 p atients 5-13 7.66 ± 0.48 5.18 ± 2.27 15 control 20-50 46.19 ± 16.55* 48.46 ± 11.91* 30 p atients 20-50 *S igni ficant (p<0.01) Insulin deficiency decreased secretion resistance, or both Increased hep atic ketogensis Increased hepatic gluconeogensis Decreased ketone utilization Decreased glucose Ut ilization Increased lip olysis Increased hep atic glycogenoly sis Hy p erketonemia Hy p erglycemia Osmotic Diuresis Excessive loss Of H2O, K, Na and Cl Ketoacidosis Dehy dration Fig .(1) The me tabolic consequences of insulin deficie ncy or resistance IHJPAS IBN AL- HAITHAM J. FOR PURE & APPL. S CI. VOL.23 (2) 2010 Co ntrol Patients Co ntrol Patients 0 5 10 15 20 25 30 35 40 45 50 Activity of Enz yme (U/mg) Co ntrol P atients Fig.(2) Activity and specific activity for patients and controls IHJPAS 2010) 2( 32المجلد مجلة ابن الهیثم للعلوم الصرفة والتطبیقیة یم ادینوسین دي امینیز في مرضى السكري من النوع الثانيظقیاس فعالیة ان حازمة موسى وزهراء حسین جامعة بغداد، كلیة التربیة ابن الهیثم ،قسم علوم الحیاة الخالصة مرضــى الســـكر النـــوع االول فـــي (ADA)دینوســـین المزیــل لمجموعـــة االمـــین مــن االیم ظـــناالة فعالیـــ تسیـقــ )IDDM ( والنوع الثاني)NIDDM ( بـالنوع الثـاني آبـالنوع االول وثالثـین مصـاب آالثین مصابـثوذلك بأخذ عینات دم من وقـد اظهـرت النتـائج وجـود ارتفـاع كبیـر فـي فعالیـة االنظـیم . وقورنت هذه العینات مع عینات االشخاص السـلیمین لكـل نـوع . في حین وجد انخفاض معنوي في عینات النوع االول مقارنة بالسیطرة ،في مرضى النوع الثاني مقارنة بالسیطرة فـي انقسـام وتمـایز الخالیـا دور رئـیس ADAا االنظـیم لـدى مرضـى السـكر حیـث ان لـلهدفت الدراسة الى تحدید فعالیـة هـذ كمـا ان االنسـولین یعـد محـور للفعالیـة المناعیــة لـذا حـددنا هـذا النـوع مـن االمــراض . التائیـة التـي هـي اسـاس المناعـة الخلویــة . ADAلدراسة فعالیة انظیم آكهدف ین االشخاص المصابین بالنوع كل من في ADAمستوى فعالیة ناك اختالفآ في ه ومن خالل النتائج یمكننا االستنتاج ان .لدى هؤالء المرضى لحصول اظطراب مناعيفعاآل آمؤشر قد یعد ان هذا االختالف . مقارنة بالسیطرة الثاني األول و IHJPAS