Antioxidative enzyme gene polymorphism and serum magnesium, ferritin and iron among Sudanese diabetic retinopathy patients

Abstract

Abstract Background: Diabetic retinopathy the commonest micro-vascular complication of diabetes mellitus. It is the main cause of blindness among young adults world widely. Poor glycemic control in addition to longer diabetic duration are the main risk factors for diabetic retinopathy. Inter-racial variation in clinical severity and incidence of diabetic retinopathy has been observed. This provide the rational to genetic factor involvement. Many genes has been postulated as candidate genes for diabetic retinopathy. Little is known about anti-oxidative enzyme gene polymorphism and its association with diabetic retinopathy, mainly for catalase enzyme and mangenes superoxide dismutase genes. Therefore the aim of the current study to determine the association of CAT-21A/T and MnSOD-47 C/T gene polymorphisms with diabetic retinopathy, in addition to determine the association of magnesium, ferritin and iron in the pathogenesis of diabetic retinopathy. Methodology: This is a case-control hospital based study carried out in Makkah Eye Complex during the period of March — December 2013. Cases were 50 diabetic patients complicated with diabetic retinopathy while control were 50 diabetic patients without diabetic retinopathy. All cases and control were selected randomly and informed consent were obtained from all participants. All patients underwent retinal examination by slit-lamp with assisted lenses, after mydriasis by using Tropicamide (Mydriasis Alcon) eye drop. Diabetic retinopathy was graded according to the American Academy of Ophthalmology scale. DNA was extracted from patients’ blood using Blood DNA Extraction kits, and then polymerase chain reaction (PCR) was done for the selected gene target. PCR product for CAT gene segment digested by Hinfl enzyme to determine the genotype by (PCR-RFLP). For MnSOD the PCR product underwent Sanger DNA sequencing to determine its genotyping. Serum was obtained to determine the concentration of magnesium and iron by using atomic absorption spectrophotometry. Serum ferritin was measured by immunofiuorescent assay. Result: In this study, the overall male to female ratio was 1 : 1 .9. The majority of the patients were 98(98%) type II diabetes and only 2 patients were type I diabetes mellitus. The mean

age i SD of the cases group versus controls was [59 :k 11.2 vs. 56:1: 10.5 years; P=0.3l7]. The mean duration of diabetes in years was higher among cases group than in controls [l6.5:I:7.5 vs. l0.7i6.3; P=0.544]. The median (25"‘ -75"‘ quartile) of serum magnesium was significantly lower in cases l.48(0.75-1.64) than control group 1.92 (1.4-2.3) mg/dl; P= 0.022. While there was no significant difference between the cases and control in serum ferritin and iron, The TT genotype frequency of the CAT-2 lA/T was significantly higher in cases (P=0.043), and showed a risk effect in diabetic retinopathy patients (OR=2.56, 95% CI=l.0l-6.45). While the AA genotype showed a protective effect (P= 0.011, OR=0.33, 95% CI= 0.13- 0.79) among controls. The allele frequency of the variant T allele was significantly different in the cases group (P=0.003, OR=2.35, 95% CI=l.33— 4.16). The genotyping for MnSOD 47C/T showed that, the fiequency of genotype CC was significantly lower in cases compared with control and OR (95% CI) = 0.088 (0.034-0.224), P=<0.00l. While the frequency of the CT heterozygote genotype was significantly higher in cases group compare with control, the OR= 3.76(l.4l-10.05); P=0.006. While frequency of the TT genotype were significantly higher in cases than controls and OR= 5.31(1.9l-14.75), P=0.001. The C allele is observed in 81% of the control while the T allele — risky allele- observed in 61% of the cases, OR= O.l50(0.079-0.285), P=<0.00l. Conclusion: In this setting, longer duration of diabetes is significantly associated with diabetic retinopathy. Hypomagnesaemia was associated with diabetic retinopathy while no association was observed in case of serum ferritin and iron. The TT genotype in both genes CAT -21A/T and MnSOD-47C/T were significantly associated with diabetic retinopathy. These finding consolidate the involvement of genetic factors in pathogenesis of diabetic retinopathy beside other known risk factors. Although hypomagnesaemia is significantly associated with diabetic retinopathy, we did not detemiine the cause of hypomagnesaemia whether it is dietary deficiency or urinary loss. This is one of the limitation of our study. However, further study is needed.