Shubhangi Arora, Nibhriti Das, Kamna Srivastava
Background: Currently hypertension grips around 25% of the entire world population. More than 90% of the hypertensive patients suffer from essential hypertension. In Asian Indians hypertension is the predominant risk factor for Coronary Artery Disease among others. Nitric Oxide (NO) is synonymous with endothelial derived relaxation factor. Acting via cGMP (cyclic guanosine monophosphate) it causes smooth muscle relaxation, prevents platelet aggregation and acts as an anti-inflammatory agent. iNOS (inducible Nitric oxide synthase), nNOS(neuronal nitric oxide synthase) and eNOS (endothelial nitric oxide synthase) are the three enzymes producing the gas nitric oxide in the human body. eNOS is the main source of NO under physiological conditions. It is known to have a number of polymorphisms. The most well known ones being the G to T polymorphism in exon 7, the T to C polymorphism in the promoter region and the a/b polymorphism in the intron 4. While the G to T polymorphism has been associated with hypertension in many races including the north Indian population, the association of other polymorphisms has been more of a controversy. Not much study has been done on the Asians especially those in India regarding these polymorphisms.
Aims: To elucidate the association between the intron4a/b polymorphism in the eNOS gene and nitric oxide levels and essential hypertension.
Objectives: 1. To determine the genotype frequencies of the above mentioned polymorphism in patients and controls 2. To study the levels of NO in the plasma of the patients and controls 3. To find out correlation if any between this polymorphism and plasma NO levels 4. To find a correlation if any between this polymorphism and essential hypertension
Materials and Methods: The study design was a case control study. 10 ml of venous blood was taken from 45 patients (selected from the department of Cardiology All India Institute of Medical Sciences, ages between 25 to 55 yrs and not on any antihypertensive medications) and controls (healthy volunteers with normal blood pressure, ages between 25 to 55 yrs, not on antihypertensive treatment, not having any diabetes mellitus or endocrine disorder or any acute illness and responding to a publicized call. It was separated into plasma and packed cells. The alleles were identified by method of Wang et al with slight modification using PCR and Agarose Gel electrophoresis with 2.7% agarose. The nitric oxide was measured using levels of the surrogate marker, nitrite by the simple and economical method of Ding et al using Griess reagent. The Data was analyzed using SPSS software. The level of significance was fixed at p<0.05.
Results/Findings: The patient and control populations were found to be in Hardy Weinberg equilibrium [p value (patient)= 0.05, p value(control) >0.05] . The mean level of nitric oxide in the patients was found to be 42% less than that of the controls. The majority of patients (76%) had plasma nitrite levels below 5uM. On the other hand a majority of controls (64%) had plasma nitrite levels more than or equal to 5uM. The fraction of patients with nitrite levels below 5uM (76%) was more than double that of the controls (34%). The patients were equally distributed between the lower two groups. The controls however showed two peaks the higher one being that of greater than 7uM. There was no significant difference (p value= 0.79) in the genotype distribution between patients and controls. There was also no significant difference between in the levels of nitrite between ab and bb genotypes in both patients and controls. Conclusion: The levels of nitrite were found to differ significantly between patients and controls. Although no statistically significant correlation could be found in the present study a trend towards higher frequency of a allele could be envisaged among the patients of essential hypertension. A detailed study with a larger sample size is needed to establish or refute the role of this polymorphism in essential hypertension in the Indian population
