miR-1及其DNA甲基化在三氯乙烯致先天性心脏病的作用机制研究

In China, congenital heart diseases are recognized as the most common type of birth malformations. In addition to genetic mutations, epigenetic changes caused by environmental factors also contribute to the disease development. Epidemiological studies and animal models indicate that trichloroethylene, a widespread environmental organic pollutant, can induce congenital heart disease, but the underlining molecular mechanism is largely unknown, particularly for human being. In consideration of little mutagenic effects of trichloroethylene and its main metabolites, epigenetic mechanisms might play an important role in the cardiac developmental toxicity. Our preliminary data showed that trichloroethylene inhibited the myocardial differentiation of cardiovascular progenitor cells. Trichloroethylene also down-regulated the expression level of miR-1, which was accompanied by the DNA hypermethylation in miR-1 modulation area and upregulation of target genes. We hypothesize that trichloroethylene modulates the expression of miR-1 by DNA methylation, which could disturb the expression of target genes, leading to the unbalance of proliferation and differentiation and aberrant endothelial/cardiac lineage specification. In this project, we will use human embryonic stem cells and rats as in vitro and in vivo models to clarify the effect of DNA methylation in regulating the expression of miR-1 and the role of miR-1 along with its target genes in trichloroethylene induced congenital cardiac defects. The study will expand our understanding of the molecular mechanism of congenital heart diseases induced by pregnancy exposure of trichloroethylene, and has potential benefits in providing molecular targets for the prevention and treatment of the diseases.

我国先天性心脏病（先心病）位居出生缺陷疾病首位，除遗传因素外，环境因素所导致的表观遗传改变也是主要诱因。三氯乙烯是分布广泛的环境有机污染物，流行病学和动物实验研究均显示与先心病发生有关，但其对人心脏发育毒性的分子机制尚不明确。由于三氯乙烯及其主要代谢物无显著遗传毒性，提示表观遗传机制可能在其心脏发育毒性中起重要作用。我们的前期实验显示，三氯乙烯可抑制心血管前体细胞的心肌定向分化，引起miR-1低表达并伴随DNA甲基化升高及miR-1靶基因上调，推测三氯乙烯可能通过DNA甲基化影响miR-1，进而干扰增殖调控和心肌/内皮细胞谱系分化。本项目除大鼠饮用水染毒模型外引入人胚胎干细胞模型，研究DNA甲基化在三氯乙烯调控miR-1表达中的机制，探讨miR-1及其下游靶基因在三氯乙烯致先心病中的作用。该项目有助于阐明孕期三氯乙烯暴露致先心病的分子机制，并为保护和干预提供分子靶标。

先天性心脏病是最常见的出生缺陷，非感染原因死亡率第一位。三氯乙烯是全球最重要的有机污染物之一，广泛存在于大气、土壤和地下水中。流行病学研究显示孕期三氯乙烯暴露与先天性心脏病相关，但机制尚不明确。DNA甲基化和微小RNA是最重要的表观遗传调控机制， miRNAs可与DNA甲基化相互作用，形成调控网络，影响心脏发育。该研究采用人胚胎干细胞心肌定向分化模型结合模式生物斑马鱼染毒模型，围绕成簇存在心肌特异性基因miR-1/133开展研究，从细胞增殖，分化，基因表达，氧化应激等方面探讨三氯乙烯对心脏发育的毒性作用机制。该研究发现：①三氯乙烯染毒导致心肌分化异常。②三氯乙烯影响细胞分化和增殖；③三氯乙烯染毒引起miR-1/133簇的表达变化；④miR-1/133簇可直接或间接通过刺激线粒体ROS升高影响细胞增殖；⑤三氯乙烯暴露可改变DNA甲基化水平，miR-1/133簇的表达可通过改变甲基化相关酶的表达参与DNA甲基化调控。该项目从表观遗传层面阐述孕期三氯乙烯暴露致先心病的分子机制，并有望为保护干预提供分子靶标。