Previous studies of a functional variant of the catechol-O-methyl transferase (COMT) gene, Val158Met, have provided inconsistent results with regard to blood pressure or hypertension. We examined the effect of this variant, the considering environmental factors of daily salt and energy intakes.
A total of 735 Japanese men (mean age, 47 years) were recruited from two separate occupational cohorts from Kanagawa and Kyoto prefectures. Participants were genotyped for the presence of COMT Val158Met (rs4680, G/A). Daily salt and energy intakes were evaluated by the food frequency questionnaire (FFQ).
Met/Met carriers had higher adjusted systolic blood pressure (SBP) (+4.79 mm Hg, P < 0.001) and diastolic blood pressure (DBP) (+2.33 mm Hg, P = 0.001) than Met/Val or Val/Val carriers. There was a significant association between being a Met/Met carrier and having a higher prevalence of hypertension (odds ratio = 2.448, 95% confidence interval = 1.426−4.205, P = 0.001). When salt and energy intakes were dichotomized, the effect of Val158Met on hypertension was observed only in the high-energy intake group, and was equivalent between low- and high-salt groups.
The Met allele of COMT Val158Met is associated with higher blood pressure and higher prevalence of hypertension in Japanese men, and energy intake may interact with this effect.
American Journal of Hypertension advance online publication 21 July 2011; doi:10.1038/ajh.2011.93
Catechol-O-methyl transferase (COMT) is a ubiquitous enzyme that catalyzes the transfer of a methyl group from S-adenoylmethionine to catechol neurotransmitters such as dopamine, epinephrine, and norepinephrine.1 Transfer of the methyl group leads to inactivation of these catecholamines, and is therefore thought to regulate dopaminergic tone in various tissues.
The human COMT gene, located on chromosome 22, contains a single-nucleotide polymorphism in the fourth exon, codon 158, which leads to a valine-to-methionine amino acid change. This variation has been shown to be functional in that Met/Met carriers display a three- to fourfold reduction in catecholamine-degrading enzymatic activity (which results in increased dopamine availability) compared with Val/Val carriers, whereas Met/Val heterozygotes display intermediate enzymatic activity.2
The discovery of a functional variant initially led to association studies for various neurogenic disorders including Parkinson's disease, schizophrenia, and bipolar disorder.3 More recently, prompted by studies of COMT function in animal models, the COMT Val158Met polymorphism has been examined with regard to blood pressure variation and hypertension prevalence in humans. A population-based cohort study from Norway reported that high systolic blood pressure (SBP) is associated with Val/Val carriers.4 In contrast, in a cohort of 51-year-old Swedish men, Met/Met carriers were associated with higher SBP (P = 0.003) and diastolic blood pressure (DBP) (P = 0.03), pointing to the opposite conclusion.5 In a Japanese cohort study, Met/Met carriers had a higher prevalence of hypertension but with only marginal statistical significance (P = 0.07).6 Therefore, results are still inconsistent and further studies are needed to quantify this polymorphism.
To explore to what extent this polymorphism may influence blood pressure, and also to investigate possible interactions with environmental factors such as salt and dietary energy intakes, we employed 735 apparently healthy Japanese men, recruited from two cohorts setup from occupational fields.
Subjects and measurements. The present study was undertaken in two occupational cohorts, which were organized to investigate the association of lifestyle factors with obesity according to different genetic factors in Japanese workers. The first cohort comprised 311 healthy Japanese men working for a company in Kanagawa Prefecture.7 The second cohort comprised 424 healthy Japanese men in Kyoto Prefecture working for a subsidiary of the first company. Information on the age, current smoking, drinking, and energy intake of all participants was obtained by means of a self-report questionnaire. The food frequency questionnaire (FFQ) was used for this purpose. Medical history was acquired by interview. Height and weight were measured, and body mass index (BMI) was calculated as weight in kg divided by height in m2. Blood pressure was measured using a posterior wall velocity/ankle-brachial index (PWV/ABI) device (Nippon Colin, Aichi, Japan), after individuals rested quietly for at least 10 min in a supine position. Blood pressure was measured twice and the average mean value was recorded. Hypertension was defined as SBP ≥140mmHg or DBP ≥90mmHg. The study was approved by the ethics review committee of the Medical Research Institute, Tokyo Medical and Dental University, and Keio University School of Medicine, and written informed consent was obtained from all participants.
Genotyping. A peripheral blood specimen was collected from each participant, and genomic DNA was extracted by conventional method. Genotyping for the COMT polymorphism was performed by PCR using the TaqMan genotyping assay (ABI) followed by allelic discrimination analysis using SDS software (Applied Biosystems, Foster City, CA).
For each sample, 10 ng of purified DNA was mixed with 0.25µl of 20× single-nucleotide polymorphism genotyping assay mix containing forward and reverse primers, VIC & FAM probes, 2.5µl of TaqMan universal PCR master mix and 2.25µl of DNase-free water. PCR amplification was performed by an Applied Biosystems 9700 Thermal Cycler with a holding temperature of 90°C for 10 min, 40 cycles of denaturation at 92°C for 15 s and annealing at 60°C for 1 min, and a final cooling step at 4°C.
Following PCR, allelic discrimination was performed using a sequence detection system (ABI PRISM 7900HT; SDS software package version 2.2.1; Applied Biosystems) with a successful genotype call rate of 99.98%.
Statistical analysis. The allele frequency was determined by direct counting. Deviation of the genotype distribution from Hardy–Weinberg equilibrium was confirmed by χ2-test. Differences in the mean values of age, clinical characteristics, current smoking, drinking, and energy intake levels between the genotype groups were compared by analysis of variance or χ2-test. Logistic regression analysis was performed to examine the relationship of genotype and hypertension adjusted for age, current smoking, and drinking. The participants were classified into two groups according to salt and energy intake based on their replies to the FFQ (1st = 1, 2nd = 2). Multiple logistic regression analysis was used to analyze whether interactions between the COMT genotypes and energy and salt intake levels were associated with hypertension. All analyses were carried out using Statistical Package for Social Science (SPSS) for Windows version 11.0 (SPSS, Chicago, IL). P values <0.05 were considered statistically significant.
The baseline characteristics of the population are shown in Table 1. Among the total of 735 participants, 474 were normotensive and 261 were hypertensive according to the criteria of SBP ≥140mmHg or DBP ≥90mmHg. The prevalence of hypertension was 35.6% in this cohort. Age, BMI, SBP, and DBP were significantly higher for the hypertensive group. As for the serum lipid profile, total cholesterol and serum triglyceride were higher and serum high-density lipoprotein cholesterol were lower in the hypertensive group. There were no differences in total energy intake, salt intake, or percentage of current smokers and drinkers between the normotensive and hypertensive groups. The Met-allele frequency (the minor allele: A) was 28% in the normotensive group and 36% in the hypertensive group (P = 0.003), indicating that this is likely to be the risk allele. The genotype distribution obeyed the Hardy–Weinberg Equilibrium in both groups.
Characteristics of Met/Met, Met/Val, and Val/Val carriers
Table 2 shows clinical characteristics and lifestyle factors according to the Val158Met genotypes. SBP and DBP were highest in patients who were Met/Met homozygotes and lowest in patients who were Val/Val homozygotes. The prevalence of hypertension was 52.6% among Met/Met homozygotes, 34.7% in Met/Val heterozygotes, and 32.5% Val/Val homozygotes. There were no differences among age, BMI, daily energy intake, daily salt intake, or frequencies of smokers or drinkers among the genotype groups. There was a difference in total serum cholesterol level among Met/Met and Met/Val + Val/Val carriers (P = 0.013), there were no differences in serum triglyceride and serum high-density lipoprotein levels among the different genotypes.
Because Met/Met carriers showed a prominent difference in SBP and DBP levels compared with Met/Val and Val/Val carriers, we combined the latter two genotypes and logistic regression analysis was performed adjusting for age, BMI, serum total cholesterol, daily salt intake, and daily energy intake (Table 3). The result showed that the Met/Met genotype was associated with hypertension after these adjustments (OR = 2.448, 95% confidence interval: 1.426–4.205, P = 0.001). In a linear regression analysis, the Met/Met genotype showed significant association with higher adjusted SBP (+4.79mmHg, P < 0.001) and DBP (+2.33mmHg, P = 0.001) than Met/Val or Val/Val genotypes.
Interaction with daily energy and salt intakes
To study potential interactions with environmental factors, we divided daily salt intake and energy intake into high and low intake groups at the median (NaCl: 9.06 g and energy intake: 1,800 kcal), and the prevalence of hypertension in each group was examined by logistic regression analysis (Table 4). In high- and low-energy intake groups, the Val158Met variant was associated with hypertension only in the high-energy intake group (P < 0.001), and not in the low-energy intake group (P = 0.264). Moreover, there is significant interaction (P = 0.037) between daily energy intake and the Val158Met variant on the prevalence of hypertension. However, in high- and low-salt intake groups the prevalence of hypertension was equally associated with the variant (P < 0.02).
In the present study, we found that the Met allele of the COMT gene Val158Met variant is associated with blood pressure and hypertension in apparently healthy Japanese men. Whereas this genetic effect was observed equally in both high- and low-salt intake groups, the effect was observed only in the high-energy intake group.
The regulation of blood pressure is highly complex and hypertension is a multifactorial disease involving various genetic and environmental factors. The role of the COMT gene in hypertension has been studied in experimental animals. Expression of the COMT gene has been shown to decrease after salt-loading in the kidney of Dahl-S rats, which are prone to hypertension.8,9COMT gene-deficient mice have reduced natriuresis due to loss of dopaminergic reaction in the kidney.10 Because Met/Met carriers have three- to fourfold lower catalytic activities compared to Val/Val carriers, it would be reasonable to speculate that the Met allele may be a risk allele for hypertension. Our result was consistent with this hypothesis in that blood pressures were highest and the prevalence of hypertension was greatest among Met/Met homozygotes, and that blood pressures were lowest and the prevalence of hypertension was least among Val/Val homozygotes. This result is in keeping with findings from a study conducted by Annerbrink et al., who reported an association between blood pressure and abdominal obesity in a group (n = 240) of 51-year-old Swedish men5 And, the COMBINE study of alcohol dependent individuals (n = 839) conducted by Stewart et al., also showed that Met/Met genotype was associated with higher adjusted baseline blood pressure compared to Val/Val genotype.11 In a large cohort study in Japan, a marginally statistically significant association of COMT and hypertension was detected only in men (P = 0.07), where the Met allele pointed to the risk.7 In contrast, a cohort study reported by Hagen et al. showed an association toward the opposite direction.4 The reason for this contradictory result may be due to the “flip–flop phenomenon” of the COMT Val158Met polymorphism. The “flip–flop” of risk alleles may have interlocus correlation with a causal variant at another locus through linkage disequilibrium. Alternatively, the contradictory result may be due to the heterogeneous effect of this Val158Met variant among ethnic groups as a result of differences in genetic background or environmental factors.12 However, in-depth study for this genomic context is required to verify this different association of COMT polymorphism among population groups.
Because nutrition has a strong influence on blood pressure and hypertension,13 we studied whether salt intake or energy intake affects the genetic effect of COMT Val158Met. We expected that there might be an interaction between COMT genotype and salt intake, because COMT enzyme activity is involved in sodium retention.14 However, we did not detect differences in the effect of COMT Val158Met between high- and low-salt intake, indicating that the interaction is unlikely in our study. It may be worthwhile to note that, although COMT has been considered as a candidate gene for salt-sensitive hypertension, a recent paper using quantitative trait locus analysis of Dahl-S rats demonstrated that COMT was excluded from the candidate genes contributing to salt-sensitive hypertension in these animals.15 Therefore, the COMT genotype might affect hypertension through physiological actions other than salt regulation. The enzyme activity of COMT is not restricted to the kidney; the brain also has abundant COMT activity.16 Indeed, Hirano et al. demonstrated that in high-salt loaded Dahl salt-sensitive rats, COMT activities in the cerebral cortex were significantly reduced, causing norepinephrine release in the cortex and affecting blood pressure by the activation of sympathetic outflow to tissue.17 However, whether or not COMT genotype affects blood pressure regulation through an extrarenal mechanism warrants further study.
Unexpectedly, in our study, an association between COMT Val158Met and hypertension was observed in the high-energy intake group but not in low-energy intake group, suggesting a potential interaction. There was no difference in BMI between the low- and high-energy intake groups (P = 0.295, Student's t-test), and the association remained positive after adjustment for BMI. Therefore, the energy intake per se was likely to interact with the genetic effect. High-energy intake is considered to be a risk factor of hypertension, usually through causing obesity;18 however, it might also be a risk factor for hypertension independent of obesity through number of mechanisms.13 Whether or not specific nutrients are involved in this interaction should be studied in a more thorough analysis.
One of the limitations to our study is the accuracy of the FFQ. Our FFQ has been validated to have a significant correlation coefficient of r = 0.47 and 0.43 with a 7-day dietary record of salt and energy intakes, respectively.19 Nevertheless, the abbreviated nature of the FFQ may have limited our ability to accurately measure dietary salt and energy intakes. Such limited accuracy may produce a potential for random measurement error in false negative associations. Thus, if these two factors are considered as confounders for the association between the genotype and blood pressure, we cannot exclude the possibility of residual confounding effects.
In conclusion, the Met allele of the COMT Val158Met variant is associated with hypertension in Japanese men. This genetic effect may have an interaction with energy intake.
We acknowledge the funding from Grant-in-Aid for Scientific Research (C) (22590547) from Ministry of Education, Culture, Sports, Science and Technology (to M.M.) and Grant-in-Aid for Scientific Research (C) (19390173) from Ministry of Education, Culture, Sports, Science and Technology (K.M.).
The authors declared no conflict of interest.