Abstract P377: Effect of Salt Diet on Haplotype Specific Expression of Human Angiotensinogen Gene

Human Angiotensinogen gene (hAGT) is an important component of the Renin- Angiotensin System. The 2.5 Kb promoter of the hAGT gene has ten polymorphisms that form two haplotype (Hap) blocks. Hap-I containing -6A, -20A, -217A, -532T, -793A, -1074T, -1178G, -1561T, -1562C, and -1670A is associated with increased blood pressure whereas Hap-II containing -6G, -20A, -217G, -532C, -793G, -1074G, -1178A, -1561G, -1562G, and -1670G is associated with normal blood pressure in human subjects. In order to understand the role of these SNPs in the transcriptional regulation, we have generated novel transgenic mice with the hAGT gene, targeted to the mHPRT locus with either Hap I or II variants. In the present study, we have examined the effect of high salt diet on the hAGT gene expression and transcriptional regulation in the 8 weeks old male transgenic mice containing these two haplotypes. Quantitative real time PCR analysis revealed an increase in hAGT expression in the liver of Hap-I as compared to Hap-II in basal condition (4.09 fold), Hap-I high salt vs Hap-I basal (2.15 fold), Hap-II high salt vs Hap-II basal (1.9 fold) and Hap-I high salt vs Hap-II high salt (2.47 fold) with p<0.05 in these transgenic mice. Immunoblot analysis also indicated the upregulation of the hAGT in plasma in all the study groups, viz; Hap-I compared to that of Hap-II (2.5 fold), Hap-I high salt vs Hap-I basal (1.9 fold) and Hap-II high salt vs Hap-II basal (1.7 fold) with p<0.05. Our studies show that the transgenic mice fed high salt diet have increased mRNA levels in the liver for the transcription factors HNF1, FOXO1, FOXA1 and MR (4, 3.13, 3.93 and 5.4 fold increase respectively; p<0.05). CHIP assay also complements the above observations. Our results strongly suggest that SNPs in Hap-I promote increased transcription and expression of the hAGT gene in multiple tissues, with resultant elevation of plasma hAGT levels. Our studies would provide new insights into the susceptibility of individuals to salt induced hypertension and explain the novel paradigm in salt induced transcriptional regulation of human angiotensinogen gene.