Abstract 626: Differential Regulation of Blood Pressure in Transgenic Mice Containing two Haplotypes of Human Angiotensin Receptor Type 1

Abstract

The renin angiotensin system (RAS) is the main regulator of blood pressure and fluid homeostasis. Most of the known physiological and pathophysiological action of RAS are mediated through the angiotensin receptor type 1 (AT 1 R). Dose dependent increase in AT 1 R gene increases blood pressure in transgenic mice. Human AT 1 R gene promoter has four single nucleotide polymorphisms (SNPs) ( T/A at -777, T/G at -680, A/C at -214, and A/G at -119). Our studies have shown that variants -777T, -680T, -214A, and -119A almost always occur together in Caucasian subjects, creating haplotype I (TTAA) and II (AGCG) respectively. We have analyzed the genomic DNA from 388 normotensive and 374 hypertensive subjects and shown that haplotype I of the h AT 1 R gene is associated with hypertension in Caucasian subjects (p=< 0.001, χ 2 = 12.26). Our transient transfection studies in adrenal cortical (H295R) and vascular smooth muscle (VSMC) cells show that haplotype I of h AT 1 R gene has increased basal and induced promoter activity as compared to haplotype II. Our gel shift assays have shown that transcription factor USF-2, C/EBP beta and delta bind strongly to an oligonucleotide containing nucleoside A at -214 (present in haplotype-I). In order to understand the physiological effect of these haplotypes in an in vivo situation, we have generated transgenic mice containing 160Kb long BAC DNA (containing 67Kb of the 5’-flanking region, all five exons and four introns, and 44 Kb of the 3’-UTR of hAT 1 R gene) and differ only in 1.2Kb promoter containing either haplotype I or haplotype II. Our studies show that 8 weeks old male transgenic mice containing haplotype I have increased h AT 1 R mRNA level in the kidney (5.5 fold) and heart (3.7 fold) as compared to haplotype II. We also show that transgenic mice containing haplotype-I have increased systolic blood pressure (7 mmHg) as compared to transgenic mice containing haplotype II. Our results suggest that increased expression of haplotype I of the hAT 1 R gene may be a contributing factor for hypertension in Caucasian subjects.