POS-804 Donor vascular endothelial growth factor gene polymorphism association with acute allograft rejection in live related renal transplant recipient patients

Abstract

Renal allograft rejection risk associated with donor’s vascular endothelial growth factor (VEGF) gene polymorphism remain unelucidated till now. Although, studies have shown, an association of recipient’s VEGF polymorphism with the end-stage renal disease and early acute rejection. VEGF has pleiotropic function, which regulates vasculogenesis, endothelial cell survival signaling. Endothelial cell regulates tonicity, the permeability of blood vessels and egression of allo-stimulated inflammatory cell in intragraft compartments, thus regulate the events of rejection. In the current study, we aimed to investigate the distribution of VEGF -634C>G, -1154 G>A, -1190G>A, -1455T>C, -1499 C>T, -2578 C>A, -2549 18bp Insertion/Deletion, +405 C>G and +936 C>T SNPs among donors and recipients and to evaluate the VEGF mRNA and protein expression in intragraft tissue and in plasma. We analyzed VEGF gene polymorphism in 160 Kidney donor and 320 Kidney allograft recipients (n=160, rejecters and n=160, non-rejecters patients). One hours prior to patient admission in kidney transplant unit, a 5 ml of blood was collected in K2EDTA vials for plasma, DNA and RNA isolation from donor and recipients. After centrifugation at 1500RPM for 5 minutes, plasma supernatants were separated and 1ml cell pellet was used for DNA isolation and 1 ml pellet was dissolved in Trizole solution for RNA isolation. A core of allograft biopsy specimen was collected from 50 patients for allograft recipient for VEGF gene and protein expression in intragraft tissue by the RT-PCR and Immunohistochemistry techniques respectively and soluble plasma VEGF level was analyzed by the ELISA. Genotyping of VEGF SNPs were done through PCR- RFLP technique. Difference between donor and recipient group was calculated by the Fisher exact test. P value <0.05 was considered statistically significant. The odd ratio was calculated to measure the difference in association between genotypes. On comparison between donors and recipients genotypes of VEGF +936 C>T [CT (OR=7.16; 95% CI=4.33- 11.84; P=0.00) and TT (OR=49.30; 95% CI=11.84-205.29; P=0.00)], -1154 G>A [AG (OR=2.22; 95% CI=1.40-3.50; P=0.00)], -1190G>A [GG (OR=2.21; 95% CI=1.22-4.01; P=0.00)], -634C>G [GG (OR=2.34; 95% CI=1.34-4.10; P=0.00)], -2549 18bp Insertion/Deletion [ID (OR=1.58; 95% CI=1.01- 2.46; P=0.04) were significantly associated with risk of rejection. Genotypic frequencies of +936 C>T [TT (OR=2.43; 95% CI=1.33-4.44; P=0.004)], -1154 G>A [GG (OR=1.94; 95% CI=1.03-3.67; P=0.04)], -1190G>A [GA (OR=1.83; 95% CI=1.07-3.13; P=0.02)], -2549 18bp Insertion/Deletion [ID (OR=2.35; 95% CI=1.38- 3.99; P=0.002) and -1455T>C [TT (OR=3.13; 95% CI=1.07-9.10; P=0.03)] shown risk of allograft rejection whereas mutant genotypes of -2578 C>A [ CA (OR=0.45; 95% CI=0.26-0.79; P=0.005) and CC (OR=0.23; 95% CI=0.11-0.46; P=0.000)] and +405 C>G [GG (OR=0.43; 95% CI=0.20-0.91; P=0.02)] have shown protective association with rejection. The intragraft VEGF mRNA and protein expression and plasma soluble VEGF level was significantly higher in allograft recipients compared to healthy donor, which was further higher in rejecters compared to non-rejecters. Mutant genotypes of VEGF +936 C>T, -1190G>A, -2549 18bp Insertion/Deletion and -1455T>C SNPs was associated with increased risk for allograft rejection