子痫前期中滋养细胞凋亡坏死与TNF-α异常甲基化的关系研究

Our previous studies and epidemiologic studies showed that hypoxia and TNF-α are possibly etiological factor of preeclampsia. We propose that hypoxia may cause abnormal DNA methylation of TNF-α, leading to increased necrotic trophoblasts/necroptotic trophoblasts and preeclampsia accompanied with increased TNF-α. This study will focus on the following 3 parts: Firstly, we want to find out the correlation between increased TNF-α and the change of death characteristic of trophoblast; Secondly, the effect of hypoxia on DNA methylation and histone will be repectively explored by bisulfite sequencing, MassARRAY; Thirdly, in trophoblast cell lines, bioinformatics technology, PCR, plasmid construction, plasmid transfection and reporter gene assay will be used to explain the role of DNA methylation in the pathogenesis of preeclampsia. ELISA, histochemistry, Western blotting and qRT-PCR will be applied for detecting TNF-α expression, bisulfite sequencing and MassARRAY for DNA methylation. The study will provide more epigenetic information for the etiology of preeclampsia.

前期研究和流行病学发现缺氧及TNF-α与子痫前期有关，但机制不明。我们提出“缺氧导致子痫前期的表观遗传假说”：缺氧可能造成滋养细胞DNA 甲基化修饰异常，TNF-α异常表达增加，导致坏死／坏死性凋亡滋养细胞异常增多，最终导致子痫前期发生”。本项目拟从临床、细胞、发病机制三个层面进行验证:1)应用病例对照研究，分析TNF-α异常表达增加同滋养细胞死亡特性改变的相关性；2)体外细胞培养分析缺氧对TNF-α DNA甲基化修饰状态的影响；3)通过生物信息学、PCR、质粒构建、质粒转染、报告基因活性测定、凝胶阻滞实验等技术，从发病机制及分子生物学的角度进一步探究DNA甲基化调控在TNF-α异常表达增加进而导致子痫前期发病中的作用机制。本项目为子痫前期发病机制的表观遗传研究提供新思路。