Abstract 3751: A Promoter Polymorphism of the Endothelial Nitric Oxide Synthase Gene Reduces Endothelial Nitric Oxide Synthase Expression in Patients With Heart Failure
A thymidine to cytosine (T786-C) transition polymorphism of the promoter region of the gene encoding endothelial nitric oxide synthase (eNOS) has been associated with spontaneous coronary vasospasm and greater autonomic imbalance in patients with heart failure. It has been presumed that these findings are the result of reduced eNOS expression, yet this has not been confirmed by in vivo humans studies. To test whether this polymorphism determines eNOS expression in vivo, we performed quantitative polymerase chain reaction (QPCR) for eNOS mRNA expression in samples of myocardial tissue obtained from the explanted hearts of patients undergoing heart transplantation. Tissue from 39 explanted hearts was submitted to QPCR after isolation of mRNA and reverse transcription to cDNA. Genomic DNA was isolated from each sample and genotyped using PCR amplification, restriction digestion with Msp1 enzyme and electrophoresis for identification of long (homozygous wild type), short (homozygous polymorphism), and of long and short fragments (heterozygotes). Twenty five samples were found to be heterozygous or homozygous for the T to C polymorphism. Samples containing the eNOS promoter polymorphism were found to have a significant 50% decrease (p = 0.02) in mRNA expression compared to samples that were homozygous for the wild type gene ( 0.23 versus 0.42 reciprocal expression units relative to beta actin house keeping gene). These data are the first in vivo demonstration in human subjects that the promoter polymorphism of the eNOS gene is associated with reduced gene expression. The results provide a mechanistic basis for greater vascular and autonomic dysfunction in patients with heart failure who possess this polymorphism and thus identify a subgroup of patients at greater risk for disease progression and arrhythmogenic sudden death.