Evaluation of endothelial nitric oxide synthase gene expression in mice genetically heterozygous in Cx43 (Cx43 + / - mice)

Abstract

Gap junctions are proteins made of connexins which are involved in the regulation of vascular function. Deletion of connexins 43 (Cx43) modifies expression of genes known to be involved in the regulation of the vasculature, differentiation and function of vascular cells. Interestingly, mutant mice lacking endothelial nitric oxide synthase (eNOS) gene have been shown to be hypertensive, suggesting that nitric oxide (NO) plays a role in the physiological control of blood pressure. It was therefore hypothesised that the endothelial deletion of Cx43 in the pulmonary vasculature induces endothelial dysfunction and causes eNOS impairment thereby reducing NO biosynthesis, thus leading to vasoconstriction and vascular remodelling which subsequently leads to the development of pulmonary arterial hypertension (PAH). This project was aimed at evaluating eNOS gene expression in mice genetically heterozygous (HET) in Cx43 (Cx43 +/-mice). This was achieved by using lung tissues from four groups of wild type (W/T) and Cx43 +/-(male and female) mice. Ribonucleic acid (RNA) was isolated from the lung tissues using RNA II isolation system and was reverse transcribed to complementary deoxyribonucleic acid (cDNA). End -point polymerase chain reaction (PCR) and real time PCR were used to measure the expression of eNOS gene. eNOS gene expression levels were found to be the same in all four groups of mice tested, with no significant difference. The result therefore suggests that eNOS gene is expressed in mice genetically heterozygous in Cx43 (Cx43 + / -).Recent data reveal that in PAH patients, the mean resting pulmonary artery pressure rises above 25 mmHg [6] and tends to increase further to 30 mmHg during exercise [8]. Elevated arterial blood pressure is a key risk factor leading to cardiovascular diseases such as right heart failure which is the main cause of mortality In PAH patients [9,10]. There is poor prognosis and rate of survival from the disease has been reported as only 58% after 3 years on therapy [11]. The disease which occurs more frequently in women than in men presents a remodelled and obliterated pulmonary vasculature, advancing to right ventricular (RV) dysfunction and death [10,12].