Table_7_Maternal Smoking During Pregnancy Induces Persistent Epigenetic Changes Into Adolescence, Independent of Postnatal Smoke Exposure and Is Associated With Cardiometabolic Risk.xlsb

Background: Several studies have shown effects of current and maternal smoking during pregnancy on DNA methylation of CpG sites in newborns and later in life. Here, we hypothesized that there are long-term and persistent epigenetic effects following maternal smoking during pregnancy on adolescent offspring DNA methylation, independent of paternal and postnatal smoke exposure. Furthermore, we explored the association between DNA methylation and cardiometabolic risk factors at 17 years of age.
Materials and Methods: DNA methylation was measured using the Illumina HumanMethylation450K BeadChip in whole blood from 995 participants attending the 17-year follow-up of the Raine Study. Linear mixed effects models were used to identify differential methylated CpGs, adjusting for parental smoking during pregnancy, and paternal, passive, and adolescent smoke exposure. Additional models examined the association between DNA methylation and paternal, adolescent, and passive smoking over the life course. Offspring CpGs identified were analyzed against cardiometabolic risk factors (blood pressure, triacylglycerols (TG), high-density lipoproteins cholesterol (HDL-C), and body mass index).
Results: We identified 23 CpGs (genome-wide p level: 1.06 × 10−7) that were associated with maternal smoking during pregnancy, including associated genes AHRR (cancer development), FTO (obesity), CNTNAP2 (developmental processes), CYP1A1 (detoxification), MYO1G (cell signalling), and FRMD4A (nicotine dependence). A sensitivity analysis showed a dose-dependent relationship between maternal smoking and offspring methylation. These results changed little following adjustment for paternal, passive, or offspring smoking, and there were no CpGs identified that associated with these variables. Two of the 23 identified CpGs [cg00253568 (FTO) and cg00213123 (CYP1A1)] were associated with either TG (male and female), diastolic blood pressure (female only), or HDL-C (male only), after Bonferroni correction.
Discussion: This study demonstrates a critical timing of cigarette smoke exposure over the life course for establishing persistent changes in DNA methylation into adolescence in a dose-dependent manner. There were significant associations between offspring CpG methylation and adolescent cardiovascular risk factors, namely, TG, HDL-C, and diastolic blood pressure. Future studies on current smoking habits and DNA methylation should consider the importance of maternal smoking during pregnancy and explore how the persistent DNA methylation effects of in utero smoke exposure increase cardiometabolic risk.

研究背景：已有多项研究表明，当前吸烟及孕期母亲吸烟会对新生儿及日后生命过程中CpG位点（CpG site）的DNA甲基化产生影响。本研究假设，孕期母亲吸烟会对子代青少年的DNA甲基化产生长期且持续的表观遗传效应，且该效应不受父亲吸烟及产后烟雾暴露的影响。此外，本研究还探讨了17岁时DNA甲基化与心脏代谢风险因子之间的关联。
材料与方法：采用Illumina人类甲基化450K芯片（Illumina HumanMethylation450K BeadChip）对参与Raine研究17年随访的995名受试者的全血样本进行DNA甲基化检测。采用线性混合效应模型（Linear mixed effects models）识别差异甲基化CpG位点，校正孕期父母吸烟、父亲吸烟、被动吸烟及青少年时期吸烟的混杂因素。额外构建的模型则分析了全生命过程中父亲吸烟、青少年时期吸烟及被动吸烟与DNA甲基化的关联。对识别出的子代CpG位点，进一步分析其与心脏代谢风险因子的关联，包括血压、三酰甘油（triacylglycerols, TG）、高密度脂蛋白胆固醇（high-density lipoproteins cholesterol, HDL-C）及体质量指数。
研究结果：本研究共识别出23个与孕期母亲吸烟相关的CpG位点（全基因组显著性p值：1.06 × 10^−7），涉及的关联基因包括AHRR（癌症发生）、FTO（肥胖）、CNTNAP2（发育过程）、CYP1A1（解毒作用）、MYO1G（细胞信号转导）及FRMD4A（尼古丁依赖）。敏感性分析显示，母亲吸烟与子代甲基化水平之间存在剂量依赖关系。在校正父亲吸烟、被动吸烟或子代吸烟的影响后，上述结果未发生显著变化，且未识别出与这些吸烟暴露变量相关的CpG位点。经邦费罗尼校正（Bonferroni correction）后，23个识别出的CpG位点中的2个[cg00253568（关联FTO基因）及cg00213123（关联CYP1A1基因）]分别与三酰甘油（男女均相关）、舒张压（仅女性相关）或高密度脂蛋白胆固醇（仅男性相关）存在关联。
讨论：本研究证实，在全生命过程中，香烟烟雾暴露存在关键时间窗，可通过剂量依赖的方式使DNA甲基化产生持续改变并延续至青少年时期。子代CpG甲基化水平与青少年时期的心血管风险因子（即三酰甘油、高密度脂蛋白胆固醇及舒张压）之间存在显著关联。未来针对当前吸烟习惯与DNA甲基化的研究，应考虑孕期母亲吸烟的重要影响，并探究宫内烟雾暴露所导致的持续性DNA甲基化改变如何增加心脏代谢疾病风险。