Maternal smoking during pregnancy (MSDP) affects development of multiple organ systems including the placenta, lung, brain, and vasculature. In particular, children exposed to MSDP show lifelong deficits in pulmonary function and increased risk of asthma and wheeze. Our laboratory has previously shown that vitamin C supplementation during pregnancy prevents some of the adverse effects of MSDP on offspring respiratory outcomes. Epigenetic modifications, including DNA methylation (DNAm), are a likely link between in utero exposures and adverse health outcomes, and MSDP has previously been associated with DNAm changes in blood, placenta, and buccal epithelium. Analysis of placental DNAm may reveal critical targets of MSDP and vitamin C relevant to respiratory health outcomes. DNAm was measured in placentas obtained from 72 smokers enrolled in the VCSIP RCT: NCT03203603 (37 supplemented with vitamin C, 35 with placebo) and 24 never-smokers for reference. Methylation at one CpG, cg20790161, reached Bonferroni significance and was hypomethylated in vitamin C supplemented smokers versus placebo. Analysis of spatially related CpGs identified 93 candidate differentially methylated regions (DMRs) between treatment groups, including loci known to be associated with lung function, oxidative stress, fetal development and growth, and angiogenesis. Overlap of nominally significant differentially methylated CpGs (DMCs) in never-smokers versus placebo with nominally significant DMCs in vitamin C versus placebo identified 9059 candidate “restored CpGs” for association with placental transcript expression and respiratory outcomes. Methylation at 274 restored candidate CpG sites was associated with expression of 259 genes (FDR < 0.05). We further identified candidate CpGs associated with infant lung function (34 CpGs) and composite wheeze (1 CpG) at 12 months of age (FDR < 0.05). Increased methylation in the DIP2C, APOH/PRKCA, and additional candidate gene regions was associated with improved lung function and decreased wheeze in offspring of vitamin C-treated smokers. Vitamin C supplementation to pregnant smokers ameliorates changes associated with maternal smoking in placental DNA methylation and gene expression in pathways potentially linked to improved placental function and offspring respiratory health. Further work is necessary to validate candidate loci and elucidate the causal pathway between placental methylation changes and outcomes of offspring exposed to MSDP. Clinical trial registration ClinicalTrials.gov, NCT01723696. Registered November 6, 2012. https://clinicaltrials.gov/ct2/show/record/NCT01723696 .

中文翻译：

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补充维生素 C 对与母亲吸烟相关的胎盘 DNA 甲基化变化的影响：与基因表达和呼吸结果的关联


母亲怀孕期间吸烟 (MSDP) 会影响多个器官系统的发育，包括胎盘、肺、大脑和脉管系统。特别是，接触 MSDP 的儿童会出现终生肺功能缺陷，并增加患哮喘和喘息的风险。我们的实验室此前已表明，怀孕期间补充维生素 C 可预防 MSDP 对后代呼吸系统结局的一些不利影响。表观遗传修饰，包括 DNA 甲基化 (DNAm)，可能是子宫内暴露与不良健康结果之间的联系，而 MSDP 此前曾被认为与血液、胎盘和口腔上皮中的 DNAm 变化有关。胎盘 DNAm 分析可能揭示与呼吸系统健康结果相关的 MSDP 和维生素 C 的关键靶点。对参加 VCSIP RCT：NCT03203603 的 72 名吸烟者（其中 37 名补充维生素 C，35 名补充安慰剂）和 24 名从不吸烟者的胎盘进行了 DNAm 测量，以供参考。与安慰剂相比，补充维生素 C 的吸烟者中一个 CpG（cg20790161）的甲基化达到了 Bonferroni 显着性，并且甲基化程度较低。对空间相关 CpG 的分析确定了治疗组之间的 93 个候选差异甲基化区域 (DMR)，包括已知与肺功能、氧化应激、胎儿发育和生长以及血管生成相关的位点。从不吸烟者与安慰剂中名义显着差异甲基化 CpG (DMC) 与维生素 C 与安慰剂中名义显着差异甲基化 CpG (DMC) 的重叠确定了 9059 个与胎盘转录本表达和呼吸结果相关的候选“恢复 CpG”。 274 个恢复的候选 CpG 位点的甲基化与 259 个基因的表达相关 (FDR < 0.05)。 我们进一步确定了与婴儿肺功能 (34 个 CpG) 和 12 个月龄复合喘息 (1 CpG) 相关的候选 CpG (FDR < 0.05)。 DIP2C、APOH/PRKCA 和其他候选基因区域甲基化的增加与接受维生素 C 治疗的吸烟者后代的肺功能改善和喘息减少相关。怀孕吸烟者补充维生素 C 可改善与母亲吸烟相关的胎盘 DNA 甲基化和基因表达的变化，这些变化可能与改善胎盘功能和后代呼吸健康有关。需要进一步的工作来验证候选基因座并阐明胎盘甲基化变化与暴露于 MSDP 的后代结果之间的因果途径。临床试验注册 ClinicalTrials.gov，NCT01723696。 2012 年 11 月 6 日注册。 https://clinicaltrials.gov/ct2/show/record/NCT01723696 。