Common Genetic Variation at the Endothelial Nitric Oxide Synthase Locus and Relations to Brachial Artery Vasodilator Function in the Community

Abstract

Background-Sequence variants at the endothelial nitric oxide synthase (NOS3) locus have been associated with endothelial function measures, but replication has been limited. Methods and Results-In reference pedigrees, we characterized linkage disequilibrium structure at the NOS3 locus using 33 common single nucleotide polymorphisms (SNPs). Eighteen SNPs that capture underlying common variation were genotyped in unrelated Framingham Heart Study participants (49.5% women; mean age, 62 years) with measured brachial artery flow-mediated dilation (nϭ1446) or hyperemic flow velocity (nϭ1043). Within 3 defined blocks of strong linkage disequilibrium that spanned NOS3, 11 SNPs captured Ͼ80% of common haplotypic variation. Among men, there were nominally significant associations between 8 NOS3 SNPs (minimum Pϭ0.002) and between haplotypes (minimum Pϭ0.002) and either flow-mediated dilation or hyperemic flow velocity. In women, we did not observe significant associations between NOS3 SNPs or haplotypes and endothelial function measures. To correct for multiple testing, we constructed 1000 bootstrapped null data sets and found that empirical probability values exceeded 0.05 for both phenotypes. Conclusions-A parsimonious set of SNPs captures common genetic variation at the NOS3 locus. A conservative interpretation of our results is that, accounting for multiple testing, we did not observe statistically significant relations between NOS3 sequence variants and endothelial function measures in either sex. The nominal associations of select NOS3 variants with endothelial function in men (unadjusted for multiple testing) should be viewed as hypothesisgenerating observations and may merit testing in other cohorts and experimental designs.

Endothelium-dependent vasodilation in conduit and resistance vessels in relation to the endothelial nitric oxide synthase gene

“…Recently, the Framingham Heart Study published an investigation relating 18 SNPs covering 480% of the common variation in the NOS3 gene to FMD in a large sample. 17 Although they found significant associations between FMD and eight of the examined SNPs, these relations were not significant after adjustment for multiple testing, necessitating validation studies. Also, they did not assess endothelium-dependent vasodilation in resistance vessels, as they did not use invasive forearm techniques.…”

Polymorphisms in the endothelial nitric oxide synthase gene and bone density/ultrasound and geometry in humans

“…Table 1 displays the eighteen NOS3 SNPs evaluated in the current analyses. All SNPs in this study were in Hardy-Weinberg equilibrium [28]. Minor allele frequencies of SNPs ranged from 6% to 46%, with no significant differences in the genotype frequencies by sex.…”

“…The 18 NOS3 SNP variants were selected from a previous study conducted by Kathiresan and colleagues [28] in which 33 SNPs were first selected on the basis of the underlying common variation using the LD patterns in the Centre d'Etude du Polymorphisme Humain (CEPH) panel (Coriell Institute for Medical Research, Camden, NJ) [27]. Of the 33 SNPs, 11 SNPs were selected as tag SNPs with customized software (http://www.broad.mit.edu/mpg/tagger/), which represented the minimum subset of SNPs required to predict all common haplotypes (≥5% frequency) within each block, with r 2 ≥0.9.…”

“…There were three "haplotype blocks" ( Table 1) with evidence of strong LD defined among the 18 SNPs capturing 97%, 83%, and 98% of common haplotypic variation, respectively [28]. Several common haplotypes with the haplotype frequency exceeding 5% were observed in the sample: 4, 6 and 3 common haplotypes in LD regions 1, 2 and 3, respectively, SNPs 1-5 made up LD region 1, but the four common haplotypes were composed by SNPs 1-4 and thus only these were considered in the analysis.…”

“…Using the haplotype block (LD region) definitions of Kathiresan et al [28], we summarized selected results from haplotype analysis in Tables 4 and 5. Table 4 displays selected haplotype analysis results for bone density/ultrasound.…”