SOX7介导心内膜EndMT调控心脏发育及先天性心脏病发生的作用机制研究

Congenital heart defects (CHD) due to abnormal heart development ranks top in birth defects. As a key part of heart development, aberrations of endocardial endothelial-to-mesenchymal transition (EndMT) is related to atrioventricular canal (AVC) and out flow tract (OFT) malformation. The role of the important embryonic development-related transcription factor, SOX7, during heart development, has not been revealed. Preliminary research of our group showed SOX7 is expressed in a subset of endothelial cells as well as mesenchymal cells of the AVC and OFT in the embryonic human heart, and mutations of SOX7 was found in 3 cases of patients with OFT malformation, wherein SOX7 mutants influenced candidate gene targets VE-cadherin and β-catenin, suggesting SOX7 can affect heart development via regulating endocardial EndMT with VE-cadherin and β-catenin. This research aims to elucidate the underlying mechanisms of SOX7-mediated endocardial EndMT by using SOX7-knockout mice experiments, AVC/OFT explant assays and analysis of cell functions. We’ll also screen for SOX7 mutations in CHD and analyze the functions of SOX7 mutants and their roles in CHD. This research will deepen our understandings of the mechanisms of the development of the heart and CHD, and provide the basis for early diagnosis and genetic counseling of CHD.

在出生缺陷中，心脏发育异常引起的先天性心脏病居于首位。心内膜内皮间质转化（EndMT）是心脏发育的重要环节，其异常与房室通道、流出道畸形发生相关。胚胎发育重要转录因子SOX7在心脏发育中的作用机制尚未阐明。本课题组前期研究发现SOX7表达于人胚心脏房室通道、流出道的心内膜及间质细胞。在3例流出道异常患儿中，SOX7基因存在突变，而且突变蛋白对潜在靶基因VE-cadherin、β-catenin的调控作用发生改变，提示SOX7可能通过VE-cadherin、β-catenin介导心内膜EndMT，影响心脏发育。为阐明该机制，本课题拟利用SOX7敲除小鼠、房室管/流出道移植物、细胞功能分析等技术手段，研究SOX7介导心内膜EndMT的机制，并筛查先心病中SOX7突变，分析突变体蛋白功能及其在先心病发生中的作用。本项目将拓宽对心脏发育和先心病致病机制的认识，为先心病的早期诊断、遗传咨询提供基础。

心内膜内皮间质转化（endothelial-to-mesenchymal transition, EndMT）是流出道（outflow tract, OFT）发育的重要环节，其异常可导致圆锥动脉干畸形 （conotruncal defects, CTD）。我们课题组于2013年报道在一个流出道异常先心病家系中发现1例患儿SOX7基因拷贝数增加，并进一步在同类型散发病例中得到证实，但SOX7在OFT发育中的作用机制尚未阐明。.为了阐明相关机制，免疫组化显示SOX7主要表达在OFT启动EndMT的心内膜内皮细胞和间充质细胞。OFT体外移植物模型表明，SOX7通过抑制心内膜细胞迁移、减少间充质细胞来负向调控EndMT。在内皮细胞中过表达SOX7可增加粘附蛋白VE-cadherin的表达，并抑制内皮细胞迁移。此外，干扰VE-cadherin的表达可部分挽救SOX7介导的细胞迁移抑制。萤光素酶报告基因和电泳迁移率 （electrophoretic mobility shift assays, EMSA）分析表明，SOX7在内皮细胞中可直接与VE-cadherin的启动子结合并上调VE-cadherin的表达。对536名散发CTD患者和300名健康对照中的SOX7基因的编码区和剪接区域进行测序，发现了4个SOX7杂合突变，其中2个突变对VE-cadherin启动子的反式激活作用减弱。.上述结果提示SOX7通过直接上调粘附分子VE-cadherin的表达，维持内皮细胞紧密联系，抑制OFT发育期间的EndMT，猜测SOX7突变通过调节VE-cadherin蛋白导致EndM受损，可能与CTD发病机理相关。本课题将拓宽对心脏发育和先心病致病机制的认识，为先心病的早期诊断、遗传咨询提供基础。.