MOLECULAR GENETIC ANALYSIS OF SUSCEPTIBLE DIABETIC NEPHROPATHY GENES IN TYPE 2 DIABETICS

Abstract

Diabetic Nephropathy (DN) is a major complication of both type 1 and type 2 diabetes and a leading cause of mortality due to End Stage Renal Disease (ESRD) worldwide. This overwhelming clinical problem is also affecting a number of diabetic patients in Pakistan. Several molecular pathways could lead to the development of DN. In this thesis, the role of genes in one classical pathway, the Renin Angiotensin Aldosterone System (RAAS) along with fatty acid metabolism, and cardiovascular related pathways were investigated in type 2 diabetics, nephropathy and ESRD cohorts. The aim was to see the allele and genotype frequency differences of Pakistani, French and UK population. The study is divided into three parts for Pakistani, French and UK cohorts. For Pakistani Punjabi population, 206 subjects mainly from the Faisalabad district were included. The study design comprised of four groups which were analyzed for biochemical and molecular characterization. PCR and PCR-RFLP based genotyping assays followed by Real Time PCR technique (TaqMan Assay) were carried out for the ACE, AGT, CYP11B2 and ACACB genes to identify the genotypes of each group individually, and calculate and compare their genotype frequencies among four groups as well with individual non-genetic parameters in each group. The association of genotypes was analyzed for their correlation with all of the biochemical parameters, as well as with hypertension, family history of diabetes and nephropathy status. The results showed the association of CYP11B2 gene polymorphism (T/C) with diabetes with OR (95%CI, P value) of 3.20 (1.15-8.91, 0.02), and a significant association of ACACB gene polymorphism (G/A) with obesity/BMI (P =0.009) and type 2 diabetes (P=0.02). For French cohorts, one of the genes of fatty acid metabolism pathway i.e. ACACB (G/A) was analyzed in five different cohorts (n=7438) of diabetic patients who were at various stages of nephropathy. The genotypes were compared for ethnic variations as well as association for different parameters. Results showed a significant association (P=0.02) with ESRD. For UK population, the association study of cardiovascular and hypertension related SNPs in LPA, DAB2IP and TLL1 genes with baseline characteristics or disease outcomes in four UK cohorts (n=5602) was performed. Results showed LPA SNPs association with serum Lp(a) levels in one of the cohorts (NPHS-II group), whereas for DAB2IP SNP (rs7025486), the significant association with High Density Lipoprotein levels (P=0.01) and Diastolic Blood Pressure (P=0.03) was observed. From this study, it is suggested that if similar studies are carried out on large number of samples (replication and meta-analysis), it will be helpful in identifying genetic biomarkers for DN on the basis of ethnic variations and devising pharmacogenomics based interventions.