Integrative analysis identifies the association between CASZ1 methylation and ischemic stroke

Abstract

Objective: To highlight potential epigenetic risk factors for blood pressure (BP) and ischemic stroke (IS) in loci identified by genome-wide association studies (GWASs).

Methods: We detected DNA methylation for BP (317,756 individuals from UK Biobank) and IS (521,612 individuals from MEGASTROKE) in Europeans by using the summary data-based mendelian randomization (SMR) method. We selected the most relevant gene to validate the association in 1,207 patients with hypertensive IS and 1,269 controls from the Chinese populations.

Results: We first identified 173 CpG sites in 90 genes, 337 CpG sites in 142 genes, and 9 CpG sites in 7 genes that were significantly associated with systolic, diastolic BP, and IS, respectively. The methylation level of cg12760995 in CASZ1 was associated with systolic (P _SMR = 1.74 × 10^-12), diastolic BP (P _SMR = 2.48 × 10^-10), and IS (odds ratio [OR] = 0.92 [95% confidence interval [CI]: 0.91-0.94]; P _SMR = 2.28 × 10^-8) in Europeans. The methylation levels of 17 sites in the promoter of CASZ1 were measured in the Chinese individuals, and 10 of them were significantly associated with IS. The higher methylation level of CASZ1 was associated with a lower risk of IS (adjusted OR = 0.97 [95% CI: 0.96-0.99]). CASZ1 seemed to be hypomethylated in hypertensive cases, and the level was negatively correlated with BP. Systolic and diastolic BP mediated approximately 61.2% (p = 3.49 × 10^-6) and 45.0% (p = 0.0029) of the association between CASZ1 methylation and IS, respectively.

Conclusions: This study identified DNA methylations that were associated with BP and IS. CASZ1 was hypomethylated in Chinese patients with hypertensive IS.

Figure 1. Results for the association between DNA methylations and IS

The x-axis represents the chromosome positions. The y-axis shows the –log10 p values for the association between levels of DNA methylation sites and BP (UK Biobank) and IS (MEGASTROKE) in European populations. Among the CpG sites analyzed for (A) SBP, (B) DBP, and (C) IS, 430, 581, and 28 passed the suggestive significance threshold of 1.0 × 10-5 (blue line), respectively. Only the top 20 genes were annotated for SBP and DBP. Other CpG sites associated with SBP and DBP are presented in tables e-1 and e-2 (links.lww.com/NXG/A313), respectively. BP = blood pressure; DBP = diastolic BP; IS = ischemic stroke; SBP = systolic BP.

Figure 2. The association between CASZ1 methylation and BP and IS

The 3 panels present the associations between CASZ1 methylation and SBP (A), DBP (B), and IS (C) in European populations. Each panel consists of 2 parts. The x-axis represents the genomic position (GRCh37.p13). The lower part of each panel shows the results of mQTL. The y-axis represents –log10(P mQTL). In this part, we can see that CASZ1 SNPs were strongly associated with DNA methylation levels (mQTL). The upper part of each panel shows the results of GWAS and SMR analysis. The y-axis represents –log10(P GWAS or SMR). In this part, we can see that CASZ1 SNPs were strongly associated with SBP (A), DBP (B), and IS (C). According to the mendelian randomization analysis, a DNA methylation level of cg12760995 in the CASZ1 gene was significantly associated with SBP (A), DBP (B), and IS (C). DBP = diastolic BP; SBP = systolic BP; GWAS = genome-wide association study; IS = ischemic stroke; SMR = summary data–based mendelian randomization.

Figure 3. The relationship of mean CASZ1 methylation levels with BP and IS among Chinese

The mean methylation levels for CASZ1 for each individual were calculated using methylation levels of all measured CpG sites within the genomic fragment sequenced. The bar plots depict the means, and the error bars in the figures represent SDs; the differences were assessed by t tests. (A) This gene was significantly hypomethylated in the 1,207 patients with hypertensive IS compared with the 1,269 controls. (B) This target was hypomethylated in the hypertensive cases (1,207 IS hypertensives plus 381 non-IS hypertensives or only 381 non-IS hypertensives) compared with the 888 controls. The 2 scatter plots present the relationship between the mean methylation level of CASZ1 and SBP (C) and DBP (D). The blue lines represent linear regression lines, and the gray areas represent the corresponding 95% confidence intervals. DBP = diastolic BP; IS = ischemic stroke; SBP = systolic BP.

Figure 4. The indirect effect of CASZ1 methylation on IS

Proportion mediated: the proportion of the association between methylation and ischemic stroke that was explained by the mediator. The analysis was performed on the 1,207 patients with hypertensive IS from the CATIS and 1,269 controls. ACME = average causal mediation effect; ADE = average direct effect; DBP = diastolic blood pressure; SBP = systolic blood pressure.