Role of NF-κB activation and VEGF gene polymorphisms in VEGF up regulation in non-proliferative and proliferative diabetic retinopathy.

The present study was aimed to investigate the relation between nuclear factor kappa beta (NFκB) activation and downstream up-regulation of vascular endothelial growth factor (VEGF) in diabetic retinopathy (DR). Moreover the study was intended to evaluate the role of VEGF gene single nucleotide polymorphisms (SNPs) in DR occurrence and to investigate the functional relevance of VEGF gene SNPs in terms of VEGF expression in DR. Serum level of VEGF, VEGF R1 (receptor 1), VEGF R 2 (receptor 2) and NFκB (p50/65) activity was measured by enzyme linked immune sorbent assay. Genotyping and allelic composition of different SNPs i.e., rs2010963, rs3025039, rs1570360 and rs 2071559 were investigated by Taqman SNP genotyping assay. VEGF, NFκB p50/p65, and VEGF R1 & R2 gene expressions were quantified by real time quantitative polymerase chain reaction. Increased NFκB p50/p65 activity and expressions were observed in non proliferative diabetic retinopathy (NPDR) and proliferative diabetic retinopathy (PDR) subjects compared to type 2 diabetes mellitus without retinopathy (DNR) group. Significantly elevated levels of serum VEGF and highest VEGF expression were found among PDR subjects compared to DNR or NPDR subjects. CC genotype and C allele of rs2010963 and TT genotype and T allele of rs3025039 were significantly over represented among PDR subjects compared to DNR group. Increased activation of NFκß in NPDR and PDR subjects might involve increased up regulation of VEGF. VEGF SNPs i.e., rs2010963 C allele and rs3025039 T allele might be associated with PDR occurrence and in turn regulates VEGF expression among PDR subjects.

Role of hyperglycemia-mediated erythrocyte redox state alteration in the development of diabetic retinopathy.

PURPOSE: To evaluate erythrocyte redox state and its surrogates in patients with different stages of diabetic retinopathy and their association with cellular metabolic derangement developed in retinal microvascular cells. METHODS: Sixty type 2 diabetic patients with nonproliferative diabetic retinopathy (NPDR), 85 patients with proliferative diabetic retinopathy (PDR), and 70 patients with diabetes but without retinopathy were considered as diabetic control (DC) for the study. In addition, 65 normal individuals without diabetes were enrolled as healthy control in this study. Erythrocyte oxidized nicotinamide adenine dinucleotide phosphate / reduced nicotinamide adenine dinucleotide phosphate (NADP / NADPH), oxidized nicotinamide adenine dinucleotide / reduced nicotinamide adenine dinucleotide (NAD / NADH) glutathione, plasma and vitreous lactate, and pyruvate levels were determined by enzymatic reaction-based spectrophotometric assay for the patients and individuals. RESULT: Erythrocyte NADP+ to NADPH ratio to NADPH ratio was found to be significantly higher among NPDR and PDR patients compared with DC subjects (P < 0.0001). Erythrocyte-reduced glutathione was significantly decreased in patients of NPDR (P = 0.0004) and patients of PDR (P = 0.0157) compared to DC. Erythrocyte NAD to NADH ratio was also significantly decreased in patients of NPDR (P < 0.0001) and PDR (P < 0.0001) compared to DC subjects. Lactate to pyruvate ratio of plasma was elevated significantly in patients with NPDR compared with DC (P < 0.0001) and those having PDR (P = 0.0046). In the vitreous fluid, the lactate to pyruvate ratios were found to be significantly lower in normal individuals without diabetes compared with patients having PDR (P < 0.0001). CONCLUSION: Hyperglycemia-mediated erythrocyte redox state alterations might be a potential risk factor for the development of NPDR in poorly controlled diabetic subjects.

Association of vascular endothelial growth factor, transforming growth factor beta, and interferon gamma gene polymorphisms with proliferative diabetic retinopathy in patients with type 2 diabetes.

PURPOSE: Chronic hyperglycemia and hypoxemia are believed to be causal factors in the development of proliferative diabetic retinopathy (PDR) among individuals with type 2 diabetes. It is hypothesized that formation of new blood vessels in the retina due to prolonged hypoxia is associated with increased expression of several growth factors and angiogenic cytokines. In the present study, we investigated the association of genetic polymorphisms in vascular endothelial growth factor (VEGF), transforming growth factor beta (TGF-ß), and interferon γ (IFN-γ) genes, which may be responsible for the hypoxia-induced VEGF-mediated neovascularization pathway for the pathogenesis of PDR. METHODS: Our case-control association study composed of 493 ethnically matched volunteers (253 with PDR [cases] and 240 diabetic controls [DC]). Gene polymorphisms were determined with Taqman-based real-time PCR and amplification refractory mutation analysis system PCR. RESULTS: The VEGF-460C (rs833061C; p=0.0043) and IFN-γ +874T (rs2430561T; p=0.0011) alleles were significantly associated with PDR. CONCLUSIONS: Genetic variations at VEGF-460C and IFN-γ +874T might accelerate the pathogenesis of retinal neovascularization in PDR.

Impact of interleukin-6 promoter polymorphism and serum interleukin-6 level on the acute inflammation and neovascularization stages of patients with Eales' disease.

PURPOSE: To evaluate the role of interleukin-6 (IL-6) in the inflammatory and proliferative stages of Eales' disease (ED) and to determine the influence of IL-6-174G/C polymorphism in the IL-6 and IL-6-regulated protein expression, as well as the development of ED. METHODS: One hundred and twenty-one patients diagnosed with ED, 223 matched healthy controls, and 16 control patients with macular holes were recruited from the eastern Indian population. Serum and vitreous levels of IL-6 and vascular endothelial growth factors (VEGF) were measured by enzyme-linked immunosorbent assay. Serum levels of high-sensitivity C-reactive protein (hsCRP) were measured by enzyme immunoassay. Subjects were genotyped for the IL-6-174G/C polymorphism (rs1800795) by a custom TaqMan single-nucleotide polymorphism (SNP) Genotyping Assays system. RESULTS: Serum IL-6 (p<0.0001), hsCRP (p<0.0001), and VEGF (p=0.0031) levels were significantly higher in the inflammatory stage of ED than in healthy controls. Serum IL-6 also significantly correlated with hsCRP (Spearman's correlation coefficient; r=0.4992, p=0.0009), but not with VEGF in this stage in ED patients. At the proliferative stage of ED, significantly higher levels of vitreous IL-6 (p=<0.0001) and VEGF (p=<0.0001) were found compared with the vitreous of patients with macular holes. A significant correlation was observed between vitreous IL-6 and VEGF in ED patients (Spearman's correlation coefficient; r=0.5834, p=0.0087). A statistically significant association was found between the -174GG genotype (p=0.006) and occurrence of ED. Mean serum and vitreous concentrations of IL-6 were also higher in the subjects with the GG genotype than in those with the GC or CC genotype in this population. CONCLUSIONS: IL-6 expression, regulated by the allelic distribution of -174 loci and the enhanced level of IL-6, modulates CRP and VEGF concentration depending respectively on the acute inflammatory stimulation at the initial stage and angiogenic stimulation at the advanced stage of ED.