Preliminary analysis of the association between methylation of the ACE2 promoter and essential hypertension.

The aim of the present study was to investigate whether methylation of the angiotensin I converting enzyme 2 (ACE2) promoter increases the risk of essential hypertension (EH). A total of 96 patients with EH were recruited and 96 sex­ and age­matched healthy controls. Methylation of 5 CpG dinucleotides in the ACE2 promoter was quantified using bisulfite pyrosequencing. Logistic regression and multiple linear regression were used to adjust for confounding factors and the generalized multifactor dimensionality reduction (GMDR) method was applied to investigate high­order interactions. Methylation of CpG4 (adjusted P=0.020) and CpG5 (adjusted P=0.036) was significantly higher in patients with EH, with frequency 97.56±5.65% and 12.75±4.15% in EH individuals and 95.73±9.11% and 11.47±3.67% in healthy controls. GMDR detected significant interaction among the 5 CpG sites (odds ratio=7.33, adjusted P=0.01). Furthermore, receiver operating characteristic curves identified that CpG5 methylation was a significant predictor of EH. Notably, CpG2 methylation was significantly higher in males than in females (adjusted P=0.018). Conversely, CpG5 methylation was significantly lower in males (adjusted P=0.032). These results indicated that aberrant methylation of the ACE2 promoter may be associated with EH risk. In addition, sex may significantly influence ACE2 methylation.

Association of SCNN1B promoter methylation with essential hypertension.

The amiloride-sensitive sodium channel beta subunit (SCNN1B) gene encodes the beta subunit of the epithelial sodium channel, which is involved in blood pressure homeostasis. The aim of the present study was to investigate the association between SCNN1B gene promoter methylation and essential hypertension (EH), and to explore whether SCNN1B methylation was altered by antihypertensive therapy. The present study recruited 282 individuals: 94 controls, 94 incident cases and 94 prevalent cases. Subsequently, the methylation status of six CpG sites in the SCNN1B promoter region was measured using bisulfite pyrosequencing technology. Among the six CpG sites, a significant difference in CpG1 and CpG2 methylation levels were detected between controls and incident cases (CpG1: ß­standardized=0.17, adjusted P=0.015; CpG2: ß­standardized=­0.41, adjusted P=0.001). In addition, a significant difference was detected in CpG1 methylation levels between incident cases and prevalent cases (ß­standardized=­0.252, adjusted P=3.77E­04). The present study also demonstrated that CpG1 and CpG2 methylation levels were significantly lower in males compared with in females (CpG1: t=­3.180, P=0.002; CpG2: t=­2.148, P=0.033). CpG1 methylation was also shown to be positively correlated with age (controls: r=0.285, P=0.008; incident cases: r=0.401, P=0.0001; prevalent cases: r=0.367, P=0.001). These results indicated a significant association between EH and SCNN1B methylation, which was affected by age, gender and antihypertensive therapy.

Interactions between CYP11B2 Promoter Methylation and Smoking Increase Risk of Essential Hypertension.

Aldosterone synthase (CYP11B2) is closely linked to essential hypertension (EH). However, it remains unclear whether the methylation of the CYP11B2 promoter is involved in the development of EH in humans. Our study is aimed at evaluating the contribution of CYP11B2 promoter methylation to the risk of EH. Methylation levels were measured using pyrosequencing technology in 192 participants in a hospital-based case-control study. Logistic regression and multiple linear regression analyses were utilized to adjust for confounding factors and the GMDR method was applied to investigate high-order gene-environment interactions. Although no significant result was observed linking the four analyzed CpG sites to EH, GMDR detected significant interactions among CpG1, CpG3, CpG4, and smoking correlated with an increased risk of EH (OR = 4.62, adjusted P = 0.011). In addition, CpG2 (adjusted P = 0.013) and CpG3 (adjusted P = 0.039) methylation was significantly lower in healthy males than in healthy females. Likewise, after adjusting for confounding factors, CpG2 methylation (adjusted P = 0.007) still showed significant gender-specific differences among the participants of the study. CpG1 (P = 0.009) site was significantly positively correlated with age, and CpG3 (P = 0.007) and CpG4 (P = 0.006) were both inversely linked to smoking. Our findings suggest that gene-environment interactions are associated with the pathogenesis and progression of EH.

Association of AGTR1 Promoter Methylation Levels with Essential Hypertension Risk: A Matched Case-Control Study.

UNLABELLED: The purpose of the present study was to investigate whether methylation of the angiotensin II type 1 receptor (AGTR1) promoter contributed to the risk of essential hypertension (EH). A total of 96 EH cases and 96 gender- and age-matched healthy controls were recruited. Methylation of 8 CpG dinucleotides (CpG1-8) in the AGTR1 promoter was examined using the bisulphite pyrosequencing technology. Three CpG dinucleotides (CpG6-8) could not be well sequenced, therefore only the remaining 5 CpG sites were analysed. A significantly lower CpG1 methylation level was identified in EH cases than in controls (cases vs. CONTROLS: 6.74 ± 4.32% vs. 9.66 ± 5.45%, p = 0.007), and no significant association was observed in the remaining analyses. In addition, significantly lower CpG1 (p = 0.028) and higher CpG2 (p = 0.032) methylation levels were observed in males than in females. In the breakdown association test by gender, a higher CpG1 methylation level was also identified in EH in both males (p = 0.034) and females (p = 0.020). Receiver operating characteristic curves showed that CpG1 methylation was a significant predictor of EH. Furthermore, CpG1 methylation was inversely correlated with uric acid levels in controls. The present study suggests that CpG1 hypomethylation in the AGTR1 promoter is likely associated with the risk of EH in the population assessed.

Aberrant methylation of the GCK gene body is associated with the risk of essential hypertension.

Essential hypertension (EH) is commonly accompanied by a dysfunction of glucose metabolism. Glucokinase (GCK) is a key enzyme involved in glucose metabolism. The aim of the present study was to investigate whether GCK gene-body methylation contributed to the risk of EH. A total of 47 patients with EH and 47 age-matched controls were recruited for methylation research in the current study. GCK gene-body methylation was measured using bisulphite pyrosequencing technology. DNA methylation levels were closely correlated among CpG1, CpG2 and CpG3 (r>0.70; P<0.001), in contrast with a weaker correlation between CpG4 and the preceding three CpGs (r<0.3 or r=1; P>0.05). Significantly lower CpG13 methylation (cases vs. controls, 49.13 ± 5.72 vs. 53.49 ± 7.53%; adjusted P=0.006) and significantly higher CpG4 methylation (cases vs. controls, 46.34 ± 6.48 vs. 34.74 ± 12.73%; adjusted P=0.002) were observed in patients with EH. The present study indicated that aberrant methylation of the GCK gene body was significantly associated with the risk of EH in the population assessed. The discrepancies between CpG1­3 and CpG4 methylation may suggest distinct roles for each of them in the determination of the risk of EH.

Lower ADD1 gene promoter DNA methylation increases the risk of essential hypertension.

The goal of our study is to investigate the contribution of promoter DNA methylation of α-adducin (ADD1) gene to the risk of essential hypertension (EH). Using the bisulphite pyrosequencing technology, DNA methylation levels of five CpG dinucleotides on ADD1 promoter were measured among 33 EH cases and 28 healthy controls. Significantly higher ADD1 DNA methylation levels were observed in the females than in the males (CpG1: P = 0.016; CpG2-5: P = 0.021). A breakdown analysis by gender showed that lower CpG1 methylation was associated with an increased risk of EH in females (adjusted P = 0.042). A much more significant association between lower CpG2-5 methylation levels and the increased risk of EH was found in males (adjusted P = 0.008). CpG1 methylation was inversely correlated with age in females (r = -0.407, P = 0.019) but not in males. ADD1 CpG1 and CpG2-5 methylation levels were significantly lower in post-menopausal (>50 years) women than pre-menopausal (≤50 years) women (CpG1: P = 0.006; CpG2-5: P = 0.034). A significant interaction between CpG1 methylation and age was found in females (CpG1*age: P = 0.029). CpG2-5 methylation was shown as a significant predictor of EH in males [area under curve (AUC) = 0.855, P = 0.001], in contrast that CpG1 methylation was a trend toward indicator in females (AUC = 0.699, P = 0.054). In addition, significant differences were observed between males and females for alanine aminotransferase (ALT, P = 0.001), aspartate aminotransferase (AST, P = 0.005) and uric acid (P<0.001). The concentration of AST was inversely correlated with ADD1 CpG2-5 methylation levels in female controls (r = -0.644, P = 0.024). These observations may bring new hints to elaborate the pathogenesis of EH.

Positive correlation between variants of lipid metabolism­related genes and coronary heart disease.

Four gene variants related to lipid metabolism (including the rs562338 and rs503662 variants of the APOB gene, the rs7767084 variant of the LPA gene and the rs2246942 variant of the LIPA gene) have been shown to be associated with coronary heart disease (CHD). The aim of the present study was to assess their association with CHD in the Han Chinese population and to assess the contribution of these gene variants to CHD. Using the standardized coronary angiography method, we enrolled 290 CHD patients and 193 non-CHD patients as non-CHD controls from Lihuili Hospital (Ningbo, China). In addition, we recruited 330 unrelated healthy volunteers as healthy controls from the Xi Men Community (Ningbo, China). Our results demonstrated that the rs503662 and rs562338 variants of the APOB gene were extremely rare in the Han Chinese population (minor allele frequency <1%). Genotype rs2246942-GG of the LIPA gene was associated with an increased risk of CHD [CHD cases versus healthy controls: P=0.04; odds ratio (OR)=1.63; 95% confidence interval (CI)=1.02-2.60). Genotype rs7767084-CC of the LPA gene was identified as a protective factor against CHD in females (CHD cases versus non-CHD controls: P=0.04, OR=0.21; CHD cases versus healthy controls: P=0.02, OR=0.21). The results of our meta-analysis indicated that rs7767084 was not associated with a high risk of CHD (P=0.83; combined OR=0.93; 95% CI=0.47-1.85). In the present study, two single nucleotide polymorphisms (SNPs) of genes involved in lipid metabolism (rs2246942 and rs7767084) were identified to be significantly associated with CHD in the Han Chinese population. Specifically, rs2246942-GG of the LIPA gene was a risk factor for CHD, while rs7767084-CC of the LPA gene was a protective factor against CHD in females. However, our meta-analysis indicated that rs7767084 is not associated with a higher risk of CHD.

Association between PCSK9 and LDLR gene polymorphisms with coronary heart disease: case-control study and meta-analysis.

OBJECTIVE: To explore the association of rs11206510 (PCSK9 gene) and rs1122608 (LDLR gene) polymorphisms with coronary heart disease (CHD) in Han Chinese. METHODS: A total of 813 participants (290 CHD cases, 193 non-CHD controls and 330 healthy controls) were recruited in the case-control study. DNA genotyping was performed on the SEQUENOM® Mass-ARRAY iPLEX® platform. χ(2)-test was used to compare the genotype distribution and allele frequencies. Two meta-analyses were performed to establish the association between the two polymorphisms with CHD. RESULTS: No significant associations between the two SNPs and the risk of CHD were observed in the present study. The meta-analysis of rs11206510 of PCSK9 gene comprises 11 case-control studies with a total of 69,054 participants. Significant heterogeneity was observed in Caucasian population in subgroup analysis of the association studies of rs11206510 with CHD (P=0.003, I(2)=67.2%). The meta-analysis of LDLR gene rs1122608 polymorphism comprises 7 case-control studies with a total of 20,456 participants and the heterogeneity of seven studies was minimal (P=0.148, I(2)=36.7%). CONCLUSION: The results of the meta-analyses indicated that both SNPs were associated with CHD in Caucasians (P<0.05) but not in Asians. The results from our case-control study and meta-analyses might be explained by genetic heterogeneity in the susceptibility of CHD and ethnic differences between Asians and Caucasians.