过度自噬介导绒毛外滋养细胞免疫损伤：子痫前期发病新机制？

Preeclampsia (PE) is a serious pregnant complication. Its specific molecular mechanism is unclear, and declined invasion of extravillous trophoblast (EVT) is key point. Our preliminary study found that there was excessive autophagy in placental trophoblast and CD8+ cytotoxic T cells specific cellular immunity. Based on excessive autophagy activated CD8 + T cells through endoplasmic reticulum related pathways, we also found that there was excessive autophagy in EVT cell model of PE, and the function of EVT declined in further preliminary experiments.There was endoplasmic reticulum different proteins between normal EVT and EVT cell model of PE. So we provide a hypothesis: EVT excessive autophagy is induced by multiple factors stimulation. Antigenic peptides generated from intra-cellular materials of patrilineal by the action of the proteasome in autophagy are transported into the endoplasmic reticulum (ER) lumen by TAP, where they can undergo additional trimming and bind with HLA-C. Antigenic peptide-HLA-C complexes reach the cell surface through the secretory pathway. So allotransplants patrilineal is presented to CD8+T lymphocytes by unprofessional APCs , then induce EVT immunolesion and preeclampsia. We will test all pathways by the technology about molecular biology, subcellular organelle proteomics, cytobiology and immunology in EVT, normal/PE placenta tissue and mice PE model. And verify the the hypothesis from "cell - placental tissue - whole" level. We wish the results can provide some news of PE immunological pathogenesis and therapeutic targets.

子痫前期（PE）是严重的妊娠并发症，绒毛外滋养细胞（EVT）侵袭力下降是发病关键点，具体分子机制不清。课题组前期研究发现患者胎盘滋养细胞存在过度自噬，且母胎界面存在以CD8+T细胞为主的特异性细胞免疫应答。基于过度自噬需通过内质网相关通路激活CD8+T细胞，我们进一步预实验发现：EVT的PE细胞模型上存在过度自噬且功能下降，与正常EVT相较，存在内质网差异蛋白。因而推测"多因素刺激EVT发生过度自噬，将其内的父源成分降解成抗原片段，然后进入内质网与HLA-C结合并转位至细胞膜上，并将这一父系的同种异体移植抗原递呈给CD8+T细胞，诱发特异性细胞杀伤效应，造成EVT的免疫损伤，参与PE发病"。项目拟在EVT、正常/PE胎盘组织及小鼠模型上应用分子生物学、亚细胞器蛋白质组学、细胞生物学及免疫学等方法，从"细胞-胎盘组织-整体"验证上述假说，将为深入研究PE免疫学发病机制提供新思路和治疗新靶点。

本课题以过度自噬为切入点，重点针对缺血缺氧和氧化应激导致的过度自噬通过免疫呈递损害绒毛外滋养细胞正常生理功能进行研究，揭示并深入探讨了通过过度自噬导致胚胎父系遗传物质的抗原性暴露，最终招致母亲免疫系统的强烈攻击。研究发现：虽然自噬和凋亡调节通路上存在交叉点，例如Bcl-2、PARP1，同种组织凋亡和自噬可以同时发生，也可相互诱导先后出现，在不同条件、细胞不同反应下，以其中一种途径占主导地位。为了排除凋亡对实验结果的影响，用Western-blot检测凋亡蛋白cleaved-caspase3蛋白表达，结果提示在该细胞模型中以自噬占主导地位。验证细胞模型存在过度自噬后，进一步证实了过度自噬对细胞功能的影响， Transwell侵袭实验检测HTR8/SVneo细胞侵袭力，实验结果显示在子痫前期细胞模型中细胞侵袭力明显下降，而抑制自噬能部分恢复细胞侵袭力。从而推测，早发型子痫前期病例，胎盘细胞处于持续氧化应激的微环境，引起滋养细胞自噬活性增加，细胞发生过度自噬，过度降解自身基础细胞器，细胞功能受损，甚至自噬性死亡增加。最终导致滋养细胞侵袭不足，子宫螺旋动脉重铸不全、胎盘浅着床，胎盘出现缺血、缺氧等病理生理改变，诱发一系列的子痫前期临床表现。证实了早发型子痫前期胎盘滋养细胞发生过度自噬，过度自噬可能是早发型子痫前期胎盘滋养细胞功能紊乱另一重要发生机制。