子痫前期胎盘外泌体的miR-199b-5p促进微血管内皮细胞缺氧损伤的机制研究

The mechanism of systemic vascular endothelial cell injury induced by regional abnormalities of placental trophoblast in PE patients remains unknown. Exosome became a research focus due to the ability of intercellular transmission of genetic contents such as miRNAs. Our previous research showed that the quantity of exosomes derived for PE placenta significantly increased, and miR-199b-5p contained in the exosomes from PE placenta was significantly higher than that from control group by miRNA microarray screening. Therefore, the scientific hypothesis was proposed: miR-199b-5p is transmitted to endothelial cells by placental exosomes and then promotes endothelial cell injury for hypoxia. This current project aims to study the changes of PE placental exosome and its effect on endothelial cell injury for hypoxia using a case - control study. Both immunofluorescence and assay of heterogeneous nuclear RNA and mRNA will be used to confirm the transmission ability in vitro. The biological function of endothelial cells will be studied to evaluate the affection of miR-199b-5p. We aim to clarify the mechanism of injury promotion of miR-199b-5p in microvascular endothelial cells undergoing hypoxia, and to identify the target gene and signaling pathway of miR-199b-5p. The results will explain the mechanism of endothelial cells' injury induced by miR-199b-5p, which is transmitted by exosomes, and provide a chance of preventing and treating PE during the period of placenta hypoxia and vascular endothelial cell injury.

子痫前期（PE）胎盘局部滋养细胞异常导致全身血管内皮细胞损伤的机制尚不清楚，外泌体因能介导不同细胞间miRNA等遗传物质的传递而受到关注。课题组前期发现PE胎盘外泌体数量增加且内含的miR-199b-5p显著高于对照组。基于此，提出假说：PE胎盘外泌体介导miR-199b-5p传递至内皮细胞，促进内皮细胞缺氧损伤。本项目拟开展病例对照研究探讨PE胎盘外泌体数量变化及其对内皮细胞缺氧损伤的影响；细胞实验中运用荧光标记示踪、前体和成熟体表达鉴定等明确外泌体可介导miR-199b-5p传递至内皮细胞，并通过细胞生物学功能检测确认miR-199b-5p促进内皮细胞缺氧损伤；筛选并验证miR-199b-5p靶基因，揭示其促进内皮细胞缺氧损伤的分子机制和下游通路。研究结果将阐释外泌体介导的miR-199b-5p在内皮细胞缺氧损伤中的调控机制，从而在胎盘低灌注诱发内皮细胞损伤的关键环节进行干预和治疗。

子痫前期（PE）是妊娠期特有的多系统疾病，是影响孕产妇和围生儿发病率及病死率的主要因素。胎盘灌注减少所致胎盘缺血缺氧，从而引起孕妇全身血管内皮细胞的广泛激活和损伤，导致多系统器官不同程度受累是PE特征性的病理改变，但胎盘灌注不足诱发血管内皮细胞受损的机制并不明确。本项目聚焦于外泌体介导细胞间遗传物质的传递作用，在前期研究结果的基础上，主要研究了PE胎盘来源的外泌体在内皮细胞缺氧损伤中的作用和机制。主要包括以下内容：(1) 通过比较PE胎盘和正常胎盘来源的外泌体对内皮细胞生长和功能的不同影响，确认PE胎盘来源的外泌体能加剧内皮细胞的缺氧损伤。(2)通过PE胎盘来源外泌体中miRNA的芯片筛查，明确PE胎盘来源外泌体中的miRNA表达谱；(3) 通过细胞水平的表达干预，确定miR-199b-5p的过表达具有缓解内皮细胞缺氧损伤的作用；(4)通过靶基因筛选及验证，明确了ARHGAP12是miR-199b-5p的靶基因，miR-199b-5p通过抑制ARHGAP12表达，参与缓解内皮细胞的缺氧损伤。本项目执行期间共发表SCI收录论文1篇，授权实用新型专利2项，参编论著1部。参加国内学术交流会议3人次（口头报告1次，壁报交流2次）。协助培养硕士研究生1名，博士研究生1名。