巨噬细胞外泌体PPARγ调控腹膜炎性损伤的机制研究

Peritoneal dialysis related peritonitis(PDRP) caused by gram negative bacteria which pathogenicity is an endotoxin known as Lipopolysaccharide(LPS) has pool clinical outcome, but their mechanisms remain unclear. Exosome is one kind of membrane vesicles which consist of a limiting lipid bilayer, transmembrane proteins and a hydrophilic corecontaining proteins, mRNAs and microRNAs. They are transported in tissue fluid and protected by lipid bilayers and act as excellular vehicles through the delivery of their functional cargo to recipient cells, with many important biological, physiological and pathological functions. In fact, exosomes and microvesicles are difficult to distinguish and so we all called them as exosomes. Peroxisomeproliferator-activated receptorgamma(PPARγ),which is recognized as a component of the exosomes circulating in plasma, is expressed in the rat peritoneal mesothelial cells (PMCs). Our previous studies has proved LPS is a factor that regulates the expression of PPAR-γ. We also confirmed that exosomes may derived from LPS-treated macrophage. The mechanism study of macrophage-derived exosomal PPARγ modulate peritoneal inflammation has not elucidated yet. Based on this reports and our results, we plan to do intense research in the molecular mechanisms of PDRP in vivo and in vitro using technical methods of molecular biology, biochemistry, cell culture, animal model, et al. And we will prove whether macrophage-derived exosomal PPARγ involves in the process of peritoneal inflammation induced by LPS or not. On the one hand, this study will provide the experimental and theoretical evidence for molecular mechanisms of PDRP. On the other hand, this study will provide new ideas and new targets for prevention and treatment of peritoneal dialysis related peritonitis.

革兰氏阴性菌（其脂多糖LPS）感染致腹膜透析相关腹膜炎临床结局差。外泌体是一种细胞自身分泌的囊泡，因其能够携带多种物质在体内进行远距离传输而成为细胞间信息传递的优良载体并发挥着重要作用。腹腔巨噬细胞（MФ）可产生外泌体，除了可转运胞浆物质，外泌体还可转运核受体PPARγ。然而，外泌体PPARγ调控腹膜炎症的机制尚无报道。我们前期研究发现：PPARγ磷酸化参与腹膜间皮细胞（PMCs）炎症调节，因此，我们提出假说：MФ的外泌体转运PPARγ及其磷酸化相关蛋白Cdk5/ERK至受体细胞——PMCs参与调控腹膜炎性损伤。本项目拟从分子、细胞、组织及动物模型等多层次阐释外泌体在炎症状态下对MФ与PMCs间生物信息传递的作用，探讨外泌体转运PPARγ的机制，揭示PPARγ磷酸化保护腹膜免受炎性损伤的机制。本研究将从外泌体转运核受体新观点探讨腹膜炎发生的新机制，为腹膜透析相关腹膜炎防治提供新思路。

革兰氏阴性菌（脂多糖LPS是其主要致病因子）感染致腹膜透析相关腹膜炎临床预后差。本项目主要研究了LPS所致巨噬细胞炎症过程中外泌体PPARγ相关调控机制，LPS所致腹膜间皮细胞炎性损伤过程中PPARγ通过与TRIM21相互作用调节CD14的表达进而影响炎症过程，LPS所致腹膜间皮细胞炎性损伤过程中PPARγ及其磷酸化体系ERK1/2-CDK5的调控机制，腹膜炎模型中PPARγ通过调控GLUT1的表达进而调节腹膜纤维化的分子机制。重要结果及关键数据主要为1)揭示了LPS诱导细胞炎症状态下巨噬细胞外泌体蛋白质组学的变化特征；2）鉴定了PPARγ对LPS诱导炎症状态下巨噬细胞外泌体蛋白质组学变化的影响；3）创新实现巨噬细胞中PPARγ蛋白进入其外泌体，进而进入腹膜间皮细胞的追踪观察，并发现过表达PPARγ的Raw264.7细胞上清可以抑制腹膜间皮细胞炎性因子的表达；4）首次揭示了腹膜间皮细胞中LPS刺激下PPARγ转录因子结合的下游基因变化特征，并证明TRIM21与PPARγ相互作用，二者通过正反调节CD14的表达参与腹膜间皮细胞的炎性损伤；5）揭示了ERK1/2-CDK5-PPARγ信号轴参与腹膜间皮细胞炎性损伤的作用机制；6）首次揭示PPARγ通过正调节GLUT1的表达介导调控腹膜纤维化发生发展的分子机制。本项目的研究进一步明确了巨噬细胞外泌体转运PPARγ至腹膜间皮细胞的新观点及其对腹膜间皮细胞炎性因子表达的影响，进一步明确了PPARγ参与腹膜间皮细胞炎性反应的调控机制及其调控腹膜纤维化的作用机制，将为腹膜透析相关腹膜炎防治提供新思路。