Alterations in the placental methylome with maternal obesity and evidence for metabolic regulation

The inflammatory and metabolic derangements of obesity in pregnant women generate an adverse intrauterine environment, increase pregnancy complications and adverse fetal outcomes and program the fetus for obesity and metabolic syndrome in later life. We hypothesized that epigenetic modifications in placenta including altered DNA methylation/hydroxymethylation may mediate these effects. Term placental villous tissue was collected following cesarean section from lean (prepregnancy BMI30) women. Genomic DNA was isolated, methylated and hydroxymethylated DNA immunoprecipitated and hybridized to the NimbleGen 2.1M human DNA methylation array. Intermediate metabolites in placental tissues were measured by HPLC-ESI-MS, ascorbate levels by reverse phase HPLC and gene expression by RT-PCR. Differentially methylated and hydroxymethylated regions occurred across the genome, with a 21% increase in methylated but a 31% decrease in hydroxymethylated regions in obese vs lean groups. Whereas increased methylation and decreased methylation was evident around transcription start sites of multiple genes in the GH/CSH and PSG gene clusters on chromosomes 17 and 19 in other areas there was no relationship. Increased methylation was associated with decreased expression only for some genes in these clusters. Biological pathway analysis revealed the 262 genes which showed reciprocal differential methylation/ hydroxymethylation were enriched for pregnancy, immune response and cell adhesion-linked processes. We found a negative relationship for maternal BMI but a positive relationship for ascorbate with α-ketoglutarate a metabolite that regulates ten eleven translocase (TET) which mediates DNA methylation. We provide evidence for the obese maternal metabolic milieu being linked to an altered DNA methylome that may affect placental gene expression in relation to adverse outcomes.

孕妇肥胖所伴随的炎症与代谢紊乱，可构建不利的宫内微环境，增加妊娠并发症与不良胎儿结局风险，并使胎儿在成年后罹患肥胖与代谢综合征的表型被程序化设定。本研究假设，胎盘的表观遗传修饰（epigenetic modifications）——包括DNA甲基化（DNA methylation）与羟甲基化（hydroxymethylation）水平改变——或可介导上述效应。研究收集了剖宫产术后的足月胎盘绒毛组织，供体为孕前BMI<18.5的体质量偏轻者与孕前BMI≥30的肥胖女性。提取基因组DNA后，通过甲基化DNA免疫沉淀与羟甲基化DNA免疫沉淀技术富集目标片段，随后将其与NimbleGen 2.1M人类DNA甲基化芯片进行杂交。采用高效液相色谱-电喷雾电离质谱（HPLC-ESI-MS）检测胎盘组织中的中间代谢产物，通过反相高效液相色谱法测定抗坏血酸水平，并利用实时荧光定量聚合酶链反应（RT-PCR）检测基因表达水平。全基因组范围内均存在差异甲基化区域与差异羟甲基化区域；与体质量偏轻组相比，肥胖组的甲基化区域增加21%，而羟甲基化区域减少31%。在17号与19号染色体上的生长激素/绒毛膜生长催乳激素（GH/CSH）及妊娠特异性糖蛋白（PSG）基因簇的多个基因转录起始位点周围，甲基化水平升高与降低现象显著，但在其他区域则无此类关联。仅该基因簇中的部分基因，其甲基化水平升高与基因表达下调存在相关性。生物学通路分析显示，262个呈现互异性差异甲基化/羟甲基化的基因，显著富集于妊娠、免疫应答及细胞黏附相关的生物学过程。本研究发现，母亲孕前BMI与α-酮戊二酸呈负相关，而抗坏血酸与α-酮戊二酸呈正相关；α-酮戊二酸作为一种调控性代谢物，可调控介导DNA甲基化的十十一易位酶（ten eleven translocase, TET）。本研究证实，孕妇肥胖相关的代谢微环境与DNA甲基化组改变存在关联，而该改变或可通过影响胎盘基因表达，参与不良妊娠结局的发生发展。