REDD1对妊娠期肝内胆汁淤积症滋养细胞低氧应激反应中糖代谢的调控机制研究

Intrahepatic cholestasis of pregnancy (ICP) is one of the most common liver disorders in pregnancy. It can result in premature delivery, fetal distress and fetal loss. It is well known that ICP increases the risk of morbidity and mortality for the mother and the fetus. However, the mechanism of ICP is not yet clearly known（remains unclear）. Considering the fetus parasitize on the metal by the placenta, placental hypoxia stress maybe play an important role in the fetal distress. Our previous research found that HIF-1α increased sensitively to acute hypoxia in the placenta of ICP. As a downstream gene of HIF-1α and a key regulator of mTOR in response to hypoxia, our recent study further comfirmed that REDD1(Regulated in Development and DNA Damage responses) protein was located in the cytoplasm of trophoblasts. Moreover, it also showed a similar expression trend just like HIF-1α during the whole pregnancy. All these evidences indicate that HIF-1α maybe regulate REDD1 in response to hypoxia in human placenta and REDD1 may participate in the process of trophoblast implantation during early pregnancy. In order to better understand the pathophysiological mechanism of placental hypoxic stress responses in ICP, this project aims to study innovativly how HIF-1α- REDD1- mTOR pathway regulates glucose transportation and glycolysis by such aspects as Bewo cells in hypoxia, animal models and placental samples with or without ICP.

妊娠期肝内胆汁淤积症（ICP）是一种妊娠特发性疾病，可引起突发性胎儿宫内窘迫、死胎、死产，对胎儿危害极大。由于胎儿通过胎盘寄生于母体，胎盘低氧应激可能是引起胎儿缺氧的重要原因之一。课题组前期实验已经证实HIF-1α是ICP胎盘组织急性缺氧的敏感指标。发育及DNA损伤反应调节基因REDD1(Regulated in Development and DNA Damage responses)作为HIF-1α的下游基因，是mTOR通路对缺氧做出快速反应的关键因子。课题组近期实验进一步证实了REDD1随孕周增大呈下降趋势，与HIF-1α的表达呈现一致性。本项目将通过缺氧Bewo细胞、ICP动物模型及临床标本的研究，创新性地从多角度阐明HIF-.1α-REDD1-mTOR信号通路对ICP滋养细胞糖转运及糖酵解的影响，以期揭示ICP胎儿缺氧时胎盘低氧应激的病理生理机制。

妊娠期肝内胆汁淤积症（ICP）是一种妊娠特发性疾病，可引起突发性胎儿宫内窘迫、死胎、死产，对胎儿危害极大。本课题组通过缺氧Bewo细胞、ICP动物模型及临床标本的研究，创新性地从多角度阐明HIF-1α-REDD1-mTOR信号通路对ICP滋养细胞糖转运及糖酵解的影响，对揭示ICP胎儿缺氧时胎盘低氧应激的病理生理机制起了一定作用。.第一部分实验对比研究了人类正常胎盘组织(n=20)及ICP晚孕胎盘组织(n=20) HIF-1α, REDD1, mTOR, GLUT1, PGK1, LDHA的变化，结果发现除mTOR，其它所有的基因其蛋白表达水平ICP组均高于正常对照组。在低氧及氧化应激损伤刺激下，ICP胎盘组织HIF-1α, REDD1的应激性升高，进一步刺激了糖转运及糖酵解的增加。因此，HIF-1α, REDD1,mTOR可能对于ICP胎盘组织的低氧及氧化应激反应，调节糖代谢起着很重要的作用。.第二部分动物实验中，分为EE-IR，EE-sham，control-IR，control-sham 组，对比研究了4组实验动物胎盘组织上述6个基因的变化。结果发现EE-sham，control-IR组HIF-1α, REDD1, GLUT1 和PGK1表达显著增加，而mTOR 和LDHA表达水平却呈现下降趋势。EE-IR组REDD1和mTOR没有发生变化，HIF-1α, GLUT1和 PGK1表达显著增加，而 LDHA表达水平却呈现下降趋势。各组的所有基因mRNA水平没有差异。说明雌激素诱导的妊娠期肝内胆汁淤积动物模型中，缺氧诱导HIF-1α,REDD1的功能发生了紊乱，削弱了HIF-1α下游基因的低氧应激反应，可能导致ICP胎儿不良结局的发生。.第三部分转染实验中，检测Bewo细胞转染前后，常氧和低氧培养的不同时间点REDD1 mRNA和蛋白的表达变化，分析HIF-1α,REDD1及mTOR蛋白的相互关系。结果显示缺氧4h和8h，缺氧组HIF-1α 表达明显上升,也高于此两个时间点上 HIF-1α转染缺氧组的表达；缺氧8h时，REDD1 转染组REDD1蛋白的表达明显低于常氧组；缺氧2h,4h,8h, REDD1转染缺氧组REDD1的表达高于常氧组。证实了缺氧均能够刺激Bewo细胞启动HIF-1α-REDD1-mTOR通路。