Medical Journal of Babylon Vol. 12- No. 3: 625-631, 2015 http://www.medicaljb.com ISSN 2312-6760©2015 University of Babylon

Original Research Article Evaluation The Total in Patients with Diabetes Mellitus (Type I and Type II) and Study Genetic Level of Glutathione-S- Transferaseµ 1

Nisreen Jalal Mohammed* Moaed Emran AL-Gazally Maysam Ali Ameen Awadh College of Medicine , University of Babylon , Hilla , IRAQ

* E-mail : [email protected]

Accepted 1 June , 2015 Abstract Diabetes mellitus is a group of metabolic diseasesdue to defect in insulin secretion or action or both. in diabetes creates free radicals .These free radicals produce oxidative stress and thus debilitate the endogenous antioxidant defense system.If the amount of insulin available is insufficient, cells response badly to the effects of insulin or if the insulin itself is abnormal. The net effect is persistently elevating levels of blood glucose, low protein synthesis. The current study evaluate the biochemical changes in diabetes mellitus patients in using different medications and investigate the glutathione S-transeferase M1 gene deletion in different treatments. The present study was conducted on (75) diabetic patients, (25) of them were treated with insulin, (25) were using insulin and metformin and the last (25) were on metformin and glibinclimide .The study also included (25) apparently healthy subjects were taken as control group. The blood sampleswere collected from Merjan Teaching Hospital from November 2013 to April 2014.The total serum protein was measured by colorimetric method described by Gornall. LSD test showed that there was significant difference between insulin & metformin group and metformin &glibinclimide in total serum protein (p ≤0.05). Metformin &glibinclimide group was less mean difference with the control group. In the gene level of the study, genotyping of glutathione S-transferase mu 1gene by PCR were defined as GSTM1 and GSTM0 or deletion association to the present and absences of the guanine nucleotide in the gene sequence. There was statistically significant difference in the genotyping distribution and the frequency of GSTM0 among study groups were 44% for insulin and metformin group, 68% for metformin and glibinclimide, 44% insulin and 28% for control healthy group. there was significant decrease in serum total protein in diabetic patients. Genetic polymorphism of glutathione S- transferase mu 1gene may be considered as risk factor for both types of diabetes mellitus.

Key words:Diabetes mellitus,GST,GSTM1,Serum total protein. اﻟﺧﻼﺻﺔ داء اﻟﺳﻛري ﻫو ﻣﺟﻣوﻋﺔ ﻣن اﻷﻣ ارض اﻻﺳﺗﻘﻼﺑﯾﺔ ﺑﺳﺑب ﺧﻠل ﻓﻲ إﻓ ارز اﻷﻧﺳوﻟﯾن أواﻹﺟ ارءات أوﻛﻠﯾﻬﻣﺎ.ارﺗﻔﺎع اﻟﺳﻛر ﻓﻲ اﻟدم ﻓﻲ ﻣرض اﻟﺳﻛري ﯾﺧﻠق اﻟﺟذور اﻟﺣرة.ﻫذﻩ اﻟﺟذور اﻟﺣرة ﺗﻧﺗﺞ اﻻﻛﺳدة ﺑﺎﻟﺗﺎﻟﻲ إﺿﻌﺎف ﻧظﺎم اﻟدﻓﺎع اﻟﻣﺿﺎد ﻟﻸﻛﺳدة اﻟذاﺗﯾﺔ.إذا ﻛﺎﻧت ﻛﻣﯾﺔ اﻷﻧﺳوﻟﯾن اﻟﻣﺗﺎﺣﺔ ﻏﯾر ﻛﺎﻓﯾﺔ،ﺑﺷدة اﻟﺧﻼﯾﺎ اﺳﺗﺟﺎﺑﺔ اﻟﺧﻼﯾﺎ ﯾﻛون ﺳﯾﺎ ﻵﺛﺎر اﻷﻧﺳوﻟﯾن أو إذا ﻛﺎن اﻷﻧﺳوﻟﯾن ﻓﻲ ﺣد ذاﺗﻪ ﻫو ﻏﯾر طﺑﯾﻌﻲ. اﻷﺛر اﻟﺻﺎﻓﻲ ﻫو ارﺗﻔﺎع ﻣﺳﺗوﯾﺎت اﻟﺟﻠوﻛوز ﻓﻲ اﻟدم،ﺗﺧﻠﯾق اﻟﺑروﺗﯾن ﺑﺻورﻩ ﻣﻧﺧﻔﺿﺔ. إن اﻟد ارﺳﺔ اﻟﺣﺎﻟﯾﺔ ﺗﻘﯾﯾم اﻟﺗﻐﯾ ارت ﺑﺎﻟﻛﯾﻣﯾﺎء اﻟﺣﯾوﯾﺔ ﻓﻲ اﻟﻣرﺿﻰ اﻟذﯾن ﯾﻌﺎﻧون ﻣرض اﻟﺳﻛري ﻓﻲ اﺳﺗﺧدام اﻷدوﯾﺔ اﻟﻣﺧﺗﻠﻔﺔ،وﺑﺎﻟﺗﺣﻘﯾق ﻓﻲ اﻟﺣذف اﻟﺣﺎﺻل ﺑﺎﻟﺟﻠوﺗﺎﺛﯾون اس ﺗ ارﻧﺳﻔرﯾز ﺑﺎﻟﻌﻼﺟﺎت اﻟﻣﺧﺗﻠﻔﺔ.

625

Mohammed et al. MJB-2015

ﻗد أﺟرﯾت ﻫذﻩ اﻟد ارﺳﺔ ب (٧٥) ﻣن ﻣرﺿﻰ اﻟﺳﻛري، (٢٥) ﻣن ﻫم ﯾﻌﺎﻟﺟون ﺑﺎﻷﻧﺳوﻟﯾن, (٢٥) ﻛﺎﻧوا ﯾﺳﺗﺧدﻣون اﻷﻧﺳوﻟﯾن واﻟﻣﯾﺗﻔورﻣﯾن وآﺧرون (٢٥) ﻛﺎﻧوا ﻋﻠﻰ ﻣﯾﺗﻔورﻣﯾن وﺟﻠﺑﯾﻧﻛﻠﯾﻣﺎﯾد .ﺷﻣﻠت اﻟد ارﺳﺔ (٢٥) ﻋﻠﻰ ﻣﺎﯾﺑدو ﻛﺎﻧوا أﺻﺣﺎء ﺗؤﺧذ ﻛﻣﺟﻣوﻋﺔ ﺿﺎﺑطﺔ. ﻋﯾﻧﺎت اﻟدم ﺗم ﺟﻣﻌﻬﺎ ﻣن ﻣﺳﺗﺷﻔﻰ ﻣرﺟﺎن اﻟﺗﻌﻠﯾﻣﻲ ﻣن ﻧوﻓﻣﺑر ٢٠١٣ إﻟﻰ أﺑرﯾل ٢٠١٤.اﻟﺑروﺗﯾن اﻟﻛﻠﻲ ﻓﻲ اﻟدم ﻛﺎن ﯾﻘﺎس ﺑﺎﻟطرﯾﻘﺔ اﻟﻠوﻧﯾﺔ اﻟﺗﻲ وﺻﻔﻬﺎ Gornall. أظﻬر اﺧﺗﺑﺎر LSD اﻟﻔﺎرق اﻟواﺿﺢ ﺑﯾن ﻣﺟﻣوﻋﺗﻲ اﻷﻧﺳوﻟﯾن & اﻟﻣﯾﺗﻔورﻣﯾن و اﻟﻣﯾﺗﻔورﻣﯾن واﻟﺟﻠﺑﻧﺟﻠﯾﻣﺎﯾد ﻓﻲ ﻣﺳﺗوى اﻟﺑروﺗﯾن اﻟﻛﻠﻲ ﻓﻲ ﻣﺻل اﻟدم (ع ≤٠.٠٥). ﻣﺟﻣوﻋﺔ اﻟﻣﯾﺗﻔورﻣﯾن واﻟﺟﻠﺑﯾﻧﺟﻠﯾﻣﺎﯾد ﻛﺎﻧت ﻓرق اﻟﻣﺗوﺳط أﻗل ﻣﻊ اﻟﻣﺟﻣوﻋﺔ اﻟﺿﺎﺑطﺔ. ﻓﻲ اﻟﻣﺳﺗوى اﻟﺟﯾﻧﻲ ﻟﻠد ارﺳﺔ،اﻟﺗﻧﻣﯾط اﻟﺟﯾﻧﻲ ﻣن اﻟﺟﻠوﺗﺎﺛﯾون S-ﺗ ارﻧﺳﻔﯾ ارزﻣو١ﺑواﺳطﺔ PCR ﺗم ﺗﻌرﯾﻔﻬﺎGSTM1 اﻟﺣذف GSTM0 ﻣﺗﺻﺎﺣﺑﻪ ﺑوﺟود او ﻏﯾﺎب اﻟﻧوﻛﻠﯾوﺗﯾداﺗﺟواﻧﯾن ﻓﻲ اﻟﺗﺳﻠﺳل اﻟﺟﯾﻧﻲ. ﺣﯾث ﻛﺎن ﻫﻧﺎك إﺣﺻﺎﺋﻲ ااﻟﻔرق اﻟﺗﻧﻣﯾط اﻟﺟﯾﻧﻲ ﻓﻲ ﺗوزﯾﻊ وﻛﺎﻧت وﺗﯾرةGSTM0ﺑﯾن ﻣﺟﻣوﻋﺎت اﻟد ارﺳﺔ ٤٤٪ ﻟﻸﻧﺳوﻟﯾن وﻣﺟﻣوﻋﺔ اﻟﻣﯾﺗﻔورﻣﯾن، ٦٨٪ ﻟﻠﻣﯾﺗﻔورﻣﯾن واﻟﺟﻠﺑﻧﺟﻠﯾﻣﺎﯾد، و ٤٤٪ أﻧﺳوﻟﯾن و ٢٨٪ ﻟﻠﻣﺟﻣوﻋﺔ اﻟﻣ ارﻗﺑﺔ اﻟﺻﺣﯾﺔ. ﻫﻧﺎك اﻧﺧﻔﺎض ﻛﺑﯾر ﻓﻲ اﻟﺑروﺗﯾن اﻟﻛﻠﻲ ﻓﻲ اﻟدم ﻓﻲ ﻣرﺿﻰ اﻟﺳﻛري. وﯾﻣﻛن اﻋﺗﺑﺎر ﺗﻌدد اﻷﺷﻛﺎل اﻟﺟﯾﻧﻲ ﻟﻼﻟﺟﻠوﺗﺎﺛﯾون S-ﺗ ارﻧﺳﻔﯾ ارزﻣو ١ geneﻛﻌﺎﻣل ﺧطر ﻟﻛﻼ اﻟﻧوﻋﯾن ﻣن ﻣرض اﻟﺳﻛري. ــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــ Introduction group are dispersed around 20 kilo base iabetes mellitus (DM) is a group of pairs (kb) separator, and orchestrated as 5*- metabolic diseases characterized GSTM4–GSTM2–GSTM1– GSTM5-3*. Dby hyperglycemia resulting from This map has been utilized to confine the defects in insulin secretion, insulin action, end purposes of the polymorphic GSTM1 or both. The chronic hyperglycemia of cancellation [5]. diabetes is associated with long-term Aim of this study was toevaluate the total damage, dysfunction, and failure of serum protein changes in diabetes mellitus different organs, especially the eyes, patients in using different medications and kidneys, nerves, heart, and blood vessels investigate the glutathione S-transeferase [1]. M1 gene deletion in different treatments. If the amount of insulin available is insufficient, cells response badly to the Material and Methods effects of insulin (insulin insensitivity or The diabetic gropes who subjected in this insulin resistance) or if the insulin itself is study were divided into (3) gropes each abnormal, then glucose will not be group had (25) patients: First group had absorbed by the body cells properly that treated with insulin, their age ranging from need it, and it will not be stored in the liver (14-77) years, second group had treated and muscles normally. The net effect is with insulin accompanied with metformin, persistently elevating levels of blood their ages ranging from (18-64) years, third glucose, low protein synthesis, and other group would treat with combination of oral metabolic derangements, such as acidosis hypoglycemic drugs metformin and [2]. glibinclimide. All the patients had suffered Glutathione S-transferase (GST) is group of from DM for about 5-15 years, they should enzymes that assumes a vital part in be not smokers or pregnant, they had no detoxification of xenobiotics. GST other chronic disease like hypertension and catalyzes attachment of the thiol of they had taken medications of DM glutathione to electrophiles[3]. regularly. Cytosolic GSTs are classified into 13 1. Serum Total Protein classes according to their structures: alpha, The total serum protein was measured by beta, delta, epsilon, zeta, theta, mu, nu, pi, colorimetric method described by Gornall. sigma, tau, phi, and omega [4]. The peptide bonds of react with A physical map has been developed of Cu+2 in alkaline solution to form a coloured the human class Mu GST genes on complex which absorbance, proportional to chromosome 1p13.3. The GST genes in this the concentration of total protein in the

626

Mohammed et al. MJB-2015 specimen, is measured at 550 nm. The volume to 100µl with deionized distilled biuret reagent contains sodium potassium H2O. Reactions were heated for 2 min at tartrate to complex cupric ions and 94°C, 1 min at 55°C and 1.5 min at 72°C maintains their solubility in alkaline for 35 cycles in a Thermal Cycler. PCR solution. products 273bp were electrophoresed on an 2. Gene Level 8% acryl amide gel. In this experiment, Primers were designed based on the cDNA sequence for human Results and Discussions GSTµ.The primers hybridize to the 5' 1. Serum Total Protein region of exon 4 (5'- There was a highly significant mean CTGCCCTACTTGATTGATGGG-3') and difference of serum total protein (p value ≤ the 3' region of exon 5 (5' 0.01) when compare three treated groups CTGGATTGTAGCAGATCATGC- 3') of with control group using ANOVA test GSTµ1 [6]. (Table 1). Poly chain reactions had been occurred The results showed significant means by mixing 12.5µl of master mix GoTaq® difference within treated diabetic groups, Green Master Mix (Promega-Green Master by using LSD test which showed that there Mix), 2µl of primer mix ,4µl of DNA is significant differences between extraction (200 ng) and 6.5µl of deionized Metformin &Glibinclimide and other distille.H2O. Net volume is 25µl. Primer treated groups;p= 0.044(≤0.05). mix consisted of 10µl of first primer and 10µl of second primer and complete

Table 1 :Mean difference of total protein by study groups

Variable Groups Mean ± SD P value

Control 52.67±1. 75 g/L Insulin&MetforminMetform 39.45± 6.58g/L* <0.01 $ Total Protein in&Glibinclimide 44.13± 6.78g/L* * Insulin 41.87± 9.40g/L*

*p value ≤ 0.05 is significant in compeer with control $p value ≤ 0.05 is significant within treated groups

The diabetic patients suffer from of circullating and intrarenal proteins, significant decrease in serum TP as a result hypertension, and anomalous intrarenal of chronically elevated blood glucose leads hemodynamics[8]. to the formation of advanced glycosylated In the glomerulus of diabetes, protein products with subsequent hyperfiltration (a passes across the endothelial cells of the potential increase of 5%-10% in GFR) and glomerular capillaries, across the basement glomerular hypertrophy [7] and that is membrane and through the slit pores of the agree with Evans and Capell who glomerular epithelial cells (podocytes) into demonstrated that pathophysiologic Bowman’s space [9]. components of diabetic nephropathy are not The metformin &glibinclimide group entirely seen yet incorporate glycosylation was the lowest mean difference from

627

Mohammed et al. MJB-2015 control group as a result of many [6].Frequencies of GSTM1 homozygous populations NIDDM develops much later null genotype (GSTM1*0) were 42%-60% protein urea and less decrease in serum in Caucasians and 16%-36% in Africans total protein and the risk of death from non- [14]. renal causes is much higher than in patients In this study, result shows that the largest with IDDM of similar duration [10]. percentage of deletion occur in Albumin execration typically increased metformin&glibinclimide group 68% who in a continuous manner over several years, represented patients with type ΙΙ diabetes the average increase in albumin execration mellitus that mean GSTM1*0 represent one rate and decrease in TP range from 10 to of risk factor for type ΙΙ diabetes mellitus 30% per year until over nephropathy and that is reaching agreement with develops [11]. Bhandari who proved that the GSTT1 and The results of this study disagree with GSTM1 genes, alone or combined, have an Venkataramana group who showed that all influence on the risk of having type 2 patients in their study were diabetes mellitus [15] and Bidet al who normoalbuminuric and without detectable demonstrated that the relationship of a micro- and macro vascular complications. joined impact of GSTM1, T1 and P1 In this study plasma total protein levels genotypes in accomplice of Indian patients were statistically significantly with T2DM. Since noteworthy affiliation increased[12]. was seen in GSTM1 invalid and GSTP1 3. Genomic level and numerous relationship in GSTM1 Genotyping of the GSTM1 gene is invalid, T present and P1 (I/I), these GSTM1*A/A, GSTM1*B/B or polymorphisms can be screened in the GSTM1*A/B genotypes. But due to populace to focus the diabetic danger [16]. presence of a deletion in this gene, the The result of this thesis is disagreeing possible genotypes would be GSTM1*A/A, with Guoyinget al who showed that there GSTM1*A/0, GSTM1*B/B, GSTM1*B/0, was no association between either GSTM1 GSTM1*A/B or GSTM1*0/0. PCR polymorphism and risk of T2 DM [17].The readings cannot differentiate between test displays that GSTM1*0 in insulin and GSTM1*A/A, GSTM1*A/0 genotype, as insulin & metformin groups was the same both appear as positive band at the specific 44% which were significant increase than locus, are considered to be GSTM1*A wild present of control group and these groups genotype, while GSTM1*B/B and represent type Ι DM that mean GSTM1*0 GSTM1*B/0 are considered as GSTM1*B represent as risk factor for type Ι DM and mutant genotype [13]. that is agree with the result of Lynn, et al; In this study the result showed that the who recommend that the GSTM1 invalid differentiation between GSTM1*A/A, genotype is connected with T1D assurance GSTM1*A/0, GSTM1*B/B, GSTM1*B/0, and T1D age-at-onset and that helplessness GSTM1*A/B (represent positive results) in to T1D may include GST conjugation [18], the hand and GSTM1*0/0(negative results) similarly Vojtkováet al;who stated that which called null or deletions by PCR. genotype combination GST T1 null/M1 Percentage ofcontrol group was 28%in wild was significantly more prevalent in Babylon community and this agreed with subjects with diabetes and represented 2.9- Seidgardet al who showed that total of 20 fold risk for T1D developing [19]. to 50% of persons had gene deletion, is called the GSTM1*0, or null allele

628

Mohammed et al. MJB-2015

Figure shows the product of PCR in polyacryl amide gel 8% under UV light (1, 3,4,5,7,8,10 and 11) are normal GSTµ1. (2, 9, and 12) are deleted GSTµ1 6 is 100 b.p. DNA ladder.

Table 3:Association of Study Groups with GSTµ1 deletion

Groups Variable Control Insulin & Metformin&G Insulin P (%) Metformin libinclimide( (%) value (%) %)

GSTµ1 Deletion 7 (28.0) 11 (44.0) 17 (68.0) 11 (44.0) 0.042 Normal 18 (72.0) 14 (56.0) 8 (32.0) 14 (56.0) *

*p value ≤ 0.05 is significant.

629

Mohammed et al. MJB-2015 Deletion in the GSTM1 gene had a 4. Oakley A., (2011).Glutathione defensive part for diabetic retinopathy. A transferases: a structural perspective. Drug substantial number of studies on GSTM1-0 Metab. Rev. 43 (2): 138–51. GSTT1-0 invalid genotypes report an 5. Shi-jieXu, Ying-ping Wang, Bruce Roe expanded danger for improvement and and William R. Pearson (1998). movement of rheumatoid joint Characterization of the Human Class Mu inflammation and asthma [20]. Glutathione S-Transferase Gene Cluster A large-scale cohort study in Egyptian and the GSTM1Deletion, J. Biol. Chem. population may confirm the role of 273(6):3517-3527. GSTM1, T1 and P1 gene polymorphisms in 6. Seidgard, J., Vorachek, W.R., Pero, the pathogenesis of T2DM and its related R.W. and Pearson, W.R. (1988). Hereditary complications [21]. differences in the expression of the human Significant correlation between GSTM1 glutathione S-transferase activity on trans- null genotype and retinopathy could stilbene oxide are due to a gene deletion. indicate this fact that impairs cellular Proc. Natl. Acad. Sci.USA 85: 7293-7297. result in increased free radicals 7. Radbill B., Murphy B. and LeRoith D., and oxidative pressure. Therefore, GST (2008).Rationale and strategies for early null genotypes may result in decrease detection and management of diabetic antioxidant capacity which causes side kidney disease. Mayo Clin Proc.; 83(12): effects of diabetes [22]. 1373–1381. 8. Evans T. and Capell P., (2000). Diabetic Conclusions Nephropathy.CLINICAL DIABETES; Diabetic patient suffer from decrease in 18(1). serum total protein may be due to increase 9. Marshall M. S. ,(2004) . Recent advances excretion due to nephropathy or decrease in in diabetic nephropathy .Clin Med;4:277– production due to defect in insulin secretion 82. or action or both. From the result of 10.Parving H., Gall M., Skøtt P., Jørgensen glutathione S- transeferase µ 1 deletion H., Løkkegaard H., Jørgensen F., Nielsen may represent a highly risk factor for both B., and Larsen S. (1992). Prevalence and types of diabetes mellitus. causes of albuminuria in non-insulin- dependent diabetic patients. Kidney Int References 41:758-62. 1. American Diabetic Association (2013), 11.Sacks D., Bruns D., Goldstein D., Diagnosis and Classification of Diabetes Maclaren N., Donald J. and Parrott Mellitus. Diabetes Care; January (36). M.,(2002). Guidelines and recommendation 2.David G and Shoback D. (2011). for laboratory analysis in the diagnosis and Greenspan's basic & clinical management of diabetes endocrinology, 9th Ed. New York: mellitus.ClinChem; 48:436-72. McGraw-Hill Medical. 12. Venkataramana G. , Indira P and 3.Udomsinprasert R., Pongjaroenkit S., D.V.M. Rao, (2013). Changes of Plasma Wongsantichon J., Oakley AJ, Total proteins, Albumin and Fibrinogen in PrapanthadaraLA,Wilce MC and Type 2 Diabetes mellitus- A Pilot study. Ketterman AJ.,(2005). "Identification, Original article;7(2): 679-685. characterization and structure of a new 13. Alshagga M., Mohamed N.,Suhid A., Delta class glutathione Ibrahim A., and Zakaria S., (2011). transferaseisoenzyme".Biochem. J. 388 (Pt Frequencies of glutathione s-transferase 3): 763–71. (GSTM1, GSTM3 AND GSTT1)

630

Mohammed et al. MJB-2015 polymorphisms in a Malaysian population, polymorphisms and associations with type Arch Med Sci.; 7(4): 572–578. 1 diabetes age-at-onset.Informa Healthcare; 14.Garte S, Gaspari L, Alexandrie AK. 38(8): 567-575. (2001).Metabolic gene polymorphism 19.Vojtková J., Ďurdík P., Čiljaková1 M., frequencies in control populations. Cancer Michnová1 Z.,Turčan T. and Babušíková Epidemiol Biomarkers Preven.; 10:1239– E., (2013). The association between gene 48. polymorphisms of glutathione s-transferase 15.Bhandari M., (2014) .Association of the t1/m1 and type 1 diabetes in slovak Glutathione s-transferase (GST) gene children and adolescents.; 21 (2): 88–91. polymorphisms with type 2 diabetes 20. Tamer L, Calikoglu M, Ates NA, mellitus risk factors in north Indian Yildirim H, Ercan B, Saritas E, Unlu A, population. Scholarly Journal of Scientific Atik U. (2004).Glutathione-S-transferase Research and Essay (SJSRE); 3(6): 77-82. gene polymorphisms (GSTT1, GSTM1, 16. Bid H., Konwar R., Saxena M., GSTP1) as increased risk factors for Chaudhari P., Agrawal C. and Banerjee asthma. Respirology;9:493– 498. M.,(2010). Association of glutathione S- 21. Zaki M., Moghazy T. ,El-Deeb M. , transferase (GSTM1, T1 and P1) gene Mohamed A., and Mohamed polymorphisms with type 2 diabetes N.(2014).Glutathione S-transferase M1, T1 mellitus in north Indian population. J and P1 gene polymorphisms and the risk of Postgrad Med. 2; 56(3):176-81. developing type 2 diabetes mellitus in 17. Guoying W., Lu Zhang and Qiongfang Egyptian diabetic patients with and without Li., (2006). Genetic polymorphisms of diabetic vascular complications. Alexandria GSTT1, GSTM1, and NQO1 genes and Journal of Medicine;51: 71-83. diabetes mellitus risk in Chinese population 22. Dadbinpour A., Sheikhha M.,Darbouy , Biochemical and Biophysical Research M. and Afkhami-Ardekani M. (2013) . Communications; 341(2):310–313. Investigating GSTT1 and GSTM1 null 18.Lynn M., Cindy S., Morgan P., Jana H., genotype as the risk factor of diabetes type Brian V., Elizabeth R., Federico F., 2 retinopathy. J. Diabetes and Metabolic Terrance J., and Ake L. (2005). Disorders, 12:48. Glutathione-s-transferase M1 and T1

631