Brief Report

Association of a Polymorphism of the Endothelial Constitutive Nitric Oxide Synthase Gene With Myocardial Infarction in the Japanese Population

Cardiovascular diseases, particularly coronary artery disease (CAD) and myocardial infarction, are the leading cause of death among people worldwide. CAD is exceedingly complex in its interplay of environment and genetics, with numerous genetic loci contributing to its heritability. Here, we aim at looking into the effects of the NOS3 c.894G>T and 27-bp VNTR polymorphisms on susceptibility to CAD in a population of Turkish Cypriots, at seeing whether these effects correlate with plasma lipid levels and at predicting the functional consequences of each polymorphism tested. A total of 50 subjects with CAD and 100 otherwise healthy subjects were included in the present case–control study. Genomic DNA was extracted from peripheral blood samples, and the two NOS3 polymorphisms were determined by restriction endonuclease analysis of PCR amplicons. Complementary methods of statistical analysis and computational modeling were employed accordingly to achieve the aims above. Our findings show that the 27-bp VNTR polymorphic locus, but not the c.894G>T polymorphic locus, is associated with CAD and that it may regulate NOS3 pre-mRNA splicing in a length-dependent manner. Overall, along with additional, yet-to-be ascertained susceptibility markers the 27-bp VNTR 4a/4b marker may be employed in risk stratification in community-level screening for CAD among Turkish Cypriots.

Endothelium nitric oxide (eNO) deficiency may lead to premature coronary artery disease (pCAD). This defect could be due to the effect of some eNOS gene variants on its gene expression.

The aim of this study is to investigate the association between four eNOS gene variants, independently and as four-locus haplotypes, with pCAD in the southern population of Iran.

One hundred fifty pCAD patients and 150 age and sex-matched controls were enrolled in this study. Polymerase chain reaction and PCR restriction fragment length polymorphism methods were used for 4a/4b variable number tandem repeat and “–922A/G, –786T/C, 894G/T” single nucleotide polymorphisms, respectively.

The results indicated that genotype frequencies of four studied variants between case and control groups were different significantly (p < .05) except for GG (−922A/G), CC (−786 T/C), GT and TT (894G/T) genotypes (p > .05). Likewise, all of the studied four mutant alleles (−922G, 786C, 894 T, and 4a) were associated with increased risk of pCAD (p < .05, OR = 1.966, OR = 3.107, OR = 2.21, OR = 1.650, respectively). H4 (bGCT) and H7 (aGCG) haplotypes were significantly associated with pCAD which introduced susceptible haplotypes (p < .01, p < .0001, respectively). However, H1 (bATG) as a protective haplotype, was in a significant negative association with pCAD (P < .0001).

In conclusion, this study suggested three novel four-locus haplotypes of eNOS gene variants that could be in association with pCAD in the southern population of Iran.

In the past ten years, several chromosomal loci have been identified by genome-wide association studies to influence the risk of coronary artery disease (CAD) and its risk factors. The GUCY1A3 gene encoding the α₁ subunit of the soluble guanylyl cyclase (sGC) resides at one of these loci and has been strongly associated with blood pressure and CAD risk. More recently, further genes in the pathway encoding the endothelial nitric oxide synthase, the phosphodiesterases 3A and 5A, and the inositol 1,4,5-trisphosphate receptor I-associated protein (IRAG), i.e., NOS3, PDE3A, PDE5A, and MRVI1, respectively, were likewise identified as CAD risk genes. In this review, we highlight the genetic findings linking variants in NO-cGMP signaling and cardiovascular disease, discuss the potential underlying mechanisms which might propagate the development of atherosclerosis, and speculate about therapeutic implications.

It is widely accepted that the classic risk factors for atherosclerosis only partly explain the incidence of coronary artery disease and the development of acute coronary syndromes. Therefore, genetic factors that vary among human populations seem to be involved in the clinical manifestations of such patients. Substantial data suggest that a significant proportion of genetic polymorphisms involved in endothelial function, inflammation, lipid metabolism, and thrombosis/fibrinolysis are often present in patients with acute coronary syndromes. Despite the well-established contribution of candidate gene studies to complex cardiovascular diseases such as acute coronary syndromes, the clinical impact and the pathophysiological implications of polymorphisms are elusive. With the evolution of genomic technologies, the necessary genetic tools may be soon available to introduce novel diagnostic and therapeutic strategies that will alter cardiovascular morbidity and mortality.

Tobacco smoking, with its wide variety of constituents, has a multitude of effects on the cardiovascular system, ranging from endothelial dysfunction, increased thrombosis, increased atherosclerosis, damage to coronary artery vessels, arrhythmias, coronary artery spasm, and heart failure. Smoking causes endothelial dysfunction through reduction in the nitric oxide level and damage to the endothelium. Oxygen radicals are released by cigarette smoking and cause increased atherosclerosis through a number of pathways. Smoking causes increased levels of prothrombotic factors, such as tissue factor (TF) and von Willebrand factor (VWF), which cause increased platelet adhesion and increased thrombotic effects through a number of pathway modulations. It is well known that smoking causes dyslipidemia. There are significantly increased rates of atherosclerosis in patients who smoke due to a number of mechanisms such as smoking’s effect on inflammatory factors and macrophages. It also causes increased angiogenesis, increased red cell mass, and higher levels of leucocytes. Smoking causes increased arterial stiffness and subsequent hypertension. The main risk factor for coronary artery spasm is also smoking. In addition, it predisposes individuals to increased left ventricular dysfunction and higher rates of ventricular tachycardia in patients with left ventricular dysfunction. Also, smoking increases mortality as a result of impaired left ventricular function. Other forms of tobacco exposure, such as smokeless or chewing tobacco, have been used and found to have varying levels of cardiovascular risk. In addition, passive smoking is considered to be a significant risk factor for coronary artery disease. There are a number of methods that can be adopted for smoking cessation such as the use of nicotine replacement therapy, bupropion, patches, and counseling sessions.

Controversial results regarding the association of eNOS gene (NOS3) polymorphisms with myocardial infarction (MI) have been reported. This study investigated the relationship of the −786T>C (rs2070744), 894G>T (rs1799983) and 4a4b polymorphisms of the NOS3 gene with the presence of MI in the Tunisian population. In addition, we also examined the association of NOS3 gene haplotypes with MI in Tunisian subjects.

A total of 303 patients with MI and 225 controls were included in the study. The 894G>T and −786T>C single nucleotide polymorphisms were analyzed by PCR-RFLP, and 4a4b polymorphism just for PCR.

There was significant linkage disequilibrium between the three NOS3 polymorphisms (p < 0.0001). The genotype distribution and allele frequency of NOS3 4a4b, but not −786T>C and 894G>T, polymorphism was significantly different between MI patients and controls. The univariate logistic regression analysis showed a significant association of the 4a4b polymorphism and MI according to co-dominant, dominant and recessive models (co-dominant model OR: 4.38, 95%CI: 1.24–15.41; p = 0.021, dominant model OR: 1.66, 95%CI: 1.14–2.42); p = 0.007, and recessive model OR: 3.85, 95%CI: 1.10–13.47; p = 0.035). The multivariate analysis, adjusted for traditional cardiovascular risk factors, revealed that the NOS3 4a4a genotype was an independent predisposing factor to MI, according to the models considered. In addition, a haplotype 7 (C-T-4a), (OR = 12.05, p = 0.010) was a risk factor of MI after controlling for classical risk factors.

These finding suggest that the 4a4b polymorphism of the NOS3 gene was associated with MI in Tunisian patients.