Hypertension is a multifactorial complex disease that is a result of gene-environment interactions. Interplay between global DNA methylation and MTHFR C677T gene polymorphism seems to be important in hypertension. The results of the present study indicate that global DNA hypomethylation is seen in hypertension (irrespective of the treatment) and is not associated with folate, vitamin B-12 and homocysteine. The global DNA methylation levels seem to be significantly dropped in individuals with both SBP-DBP high who are not on medication. Intervention with medication for treatment of hypertension seems to improve methylation levels.
Global DNA hypomethylation has been reported as a consequence of age in numerous studies involving both human and mice [23, 24]. Global DNA methylation has also been reported to be higher among males [25, 26]. In the present study, all the selected demographic (age and sex) and lifestyle (smoking, alcohol consumption and diet) confounders for hypertension were found to be significantly higher among the cases, whereas the biochemical (folate, vitamin B-12 and homocysteine) and genetic (MTHFR C677T gene polymorphism) variables were found to be distributed similarly in both the groups. Global DNA hypomethylation was seen among cases even after adjustments for all the confounders for hypertension, albeit with no statistical significance, suggesting that global DNA hypomethylation may be associated with hypertension irrespective of the confounders. Similar findings that associate hypertension with global hypomethylation were reported in previous studies [9, 19, 20, 27].
Global DNA methylation levels were seen to be higher among cases that were on medication compared to cases that were not on medication, or in other words, it may be inferred that global DNA methylation seems to improve after medication among cases. No published literature is available on the influence of antihypertensive drugs on global DNA methylation, but the results of the present study suggest an improvement in methylation levels due to intervention with antihypertensive drugs. The effect of this global DNA hypomethylation on the genes influencing the blood pressure control among hypertensives as also proposed by Smolarek et al., 2010 cannot be overlooked . Though Smolarek et al., 2010 have discussed about the pleiotropic effects of antihypertensive drugs on global DNA methylation, but no correlation was observed between the treatment and global DNA methylation in their study. Being a rural population, the hypertensive individuals in the present study usually visit the local government hospitals where common antihypertensive drugs such as calcium channel blockers like amlodipine or angiotensin receptor blockers (ARBs) like telmisartan are prescribed. These drugs seem to be acting on the blood pressure controlling genes too, as also suggested by Smolarek et al., 2010 . In the present study also it has been observed that anti-hypertensive drugs seem to alter global DNA methylation levels.
Since hypertension with both high SBP-DBP is a severe form of hypertension that predicts cardiovascular risk , when individuals with both SBP-DBP hypertension (not on medication) were considered, they seem to be having significantly reduced global DNA methylation compared to their respective controls (Fig. 2). Similar observations were seen after adjusting for the effect of confounders (r2-0.859, p-value – 0.01). Global DNA hypomethylation which is an effect of hypertension in turn may lead to other metabolic adversities (such as cardiovascular adversities)  due to genomic instability caused by global DNA hypomethylation , but may be reduced or reversed through medication. Interestingly, the trend of 5mC% among cases (SBP-DBP high) seemed similar to that of controls whereby global DNA methylation levels were improved among cases on medication. This suggests that medication may improve the global DNA methylation status.
Further, global DNA methylation is also reported to be affected by mutations in the candidate genes specific to ‘one-carbon’ metabolic pathway like that of MTHFR. MTHFR gene affects the release of free methyl groups in the homocysteine metabolic pathway during the irreversible reduction of 5,10-methylenetetrahydrofolate to 5-methyltetrahydrofolate . MTHFR C677T gene polymorphism can reduce the enzymatic activity of MTHFR, resulting in decreased 5-methyl-THF concentrations, increased homocysteine concentrations, and reduced methylation capacity . Thus, T allele carrying individuals of MTHFR C677T gene polymorphism are likely to exhibit hypomethylation owing to enhanced methyl group acceptance capacity (higher among TT genotype carrying individuals vs CT and CC individuals) . In the present study, when an attempt was made to understand this phenomenon among cases and controls, similar trend of decreased methylation with an increase in T allele dose (TT < CT < CC) among cases (not on medication) was observed. However, such trend was not found among controls and cases (on medication). Global DNA methylation levels were found to be lower among CT genotype carrying individuals compared to CC genotype but an increase in global DNA methylation level was observed with respect to TT genotype in both controls and cases on medication. A study conducted by Arruda et al., 2013 among healthy individuals has also reported global DNA methylation levels in CT genotype carrying individuals to be lower compared to CC and TT genotype carrying individuals . However, the reason for such a trend among CT genotype carrying individuals with respect to global DNA hypomethylation needs to be explored. Further, the variation of global DNA methylation with respect to genotypes seems to be different only among cases (on medication and not on medication) whereas no such difference was observed among controls.
Intervention with medication seems to play an important role in improvement of DNA methylation status. This could probably be due to two mechanisms. First, medication may directly improve methylation levels and the flux of methyl groups may be diverted to the blood pressure controlling genes, thereby activating them to control blood pressure. Second, the drug may directly act on blood pressure control and this may result in the improvement of methylation status.
Further, the intervention with medication is given irrespective of the MTHFR genotype status of an individual but there seems to be an improvement in the methylation status only among homozygote individuals on medication with CC and TT genotypes. An abrupt drop in methylation levels among cases on medication with CT genotype is indicative of poor response of these individuals to antihypertensive drugs. Intervention with medication is supposed to normalize the elevated blood pressure levels and therefore the cases on medication seem to be imitating the 5mC% pattern among controls, as also seen in the present study but the exact mechanisms involved in such a phenomenon are yet to be explored. No comprehensive studies are available that confirm the role of MTHFR genotype with respect to the effect of drugs and global DNA methylation.
The sample size in the present study is not robust enough to come to major conclusions on the complex interplay between global DNA methylation, MTHFR C677T gene polymorphism and hypertension. However, the results of the present study are indicative of an improvement in global DNA methylation levels due to intervention with antihypertensive drugs, which is more pronounced among individuals with CC and TT genotypes. The findings of the present study and previous studies [9, 11] thus seem to build evidence on the association of global DNA hypomethylation with disturbances in cardiovascular system. Further, the mere fact that medication is improving global DNA methylation among cases is a welcoming observation which can be a primer to the interventional strategies adopted in blood pressure regulation.