Abstract 089: A Genetic Variant of Human Aldosterone Synthase Gene Causes Salt-Dependent High Blood Pressure in Transgenic Mice

Abstract

Aldosterone, synthesized by the enzyme CYP11B2, induces positive sodium-balance and predisposes to hypertension. Various investigators, using genomic DNA analyses, have linked -344T polymorphism in the hCYP11B2 gene to human hypertension. We have identified three SNPs, in linkage disequilibrium, in the hCYP11B2 gene: T/A at -663, T/C at -470 and C/T at -344. Variants ACT occur together and form the haplotype I (Hap I) while variants TTC constitute haplotype II (Hap II). We hypothesize that these SNPs, when present together, will lead to haplotype-dependent transcription of the hCYP11B2 gene, differentially increase aldosterone and affect blood pressure. To this end, novel transgenic (TG) mice with the hCYP11B2 gene, targeted to the mHPRT locus, with either haplotype II or I variant are used in the study. ChIP assay, using anti-RNA pol II antibody, shows increased Pol II binding to the chromatin from Hap I TG mice in adrenal (2.8 fold higher, p<0.05) and renal tissues (1.3 fold higher, p<0.05) as compared to chromatin extracts from Hap II TG mice. Immunoblot analysis shows upregulation of the hCYP11B2 in adrenal (2.7 fold higher, p<0.05) and renal tissues (1.35 fold higher, p<0.05) of Hap I vs. Hap II-TG mice; no significant difference was observed in mCYP11B2 between the two haplotypes. Complementary ELISA shows higher circulating levels (p<0.05) of aldosterone in Hap I mice (1504±48.7 pg/mL) as compared to both, Hap II (778±142.8 pg/mL) and C57 mice (740±28.9 pg/mL). Importantly, we observed increased baseline blood pressure in Hap I TG mice (Hap I- 117±2.5 vs. Hap II- 109±1.9 mm Hg, p<0.05), an effect accentuated by high-salt diet (Hap I- 135±2.6 vs. Hap II- 122±2.2 mm Hg, p<0.05). Elevated aldosterone was accompanied by up-regulation (p<0.05) of proinflammatory markers in renal tissues from Hap I TG mice (IL1β, MCP1, ICAM). Thus, this study identifies -344T as a reporter polymorphism for Hap I of the hCYP11B2 gene. SNPs in Hap I promote increased transcription and expression of the gene, in multiple tissues, with resultant elevation of plasma aldosterone levels. Pathophysiological impact of this haplotype-dependent transcriptional regulation of the hCYP11B2 is highlighted by increased inflammation and blood pressure in TG mice with the Hap I of this transgene.