抑制Kupffer细胞RIP140表达诱导内毒素耐受减轻肝移植缺血再灌注损伤的实验研究

Our researches have identified that one of the most important reasons for ischemia / reperfusion injury (I/RI) after liver transplantation was the activation of Kupffer cells (KCs) triggered by LPS during reperfusion phase, and I/RI could be significantly alleviated after endotoxin tolerance (ET) induced in KCs. Macrophages activated by LPS through many sophisticated signal transduction pathways.Recent studies have demonstrated that changes on chromatin of specific LPS signal transduction proteins caused by nuclear receptors provide the major regulatory mechanisms for ET induction. NF-kB was allimportant nuclear receptors to activate macrophages caused by LPS, and receptor-interacting protein 140 (RIP140) mainly expressed in liver was one of the most important co-activator for NF-κB of macrophages. RIP140 degradation promotes ET in macrophages. As 80-90% macrophages are known as KCs located in hepatic sinusoids, we deduced that RIP140 could also play a significant role in KCs activation by LPS, and future therapeutic approaches may be targeted to supression of RIP140 gene expression to establish ET in KCs during reperfusion phase to alleviate I/RI after liver transplantation.

我们前期研究发现内毒素(LPS)触发的再灌注期Kupffer细胞(KCs)过度激活是导致肝移植缺血再灌注损害(I/RI)的重要原因之一，诱导KCs对内毒素耐受(ET)后可显著减轻I/RI。最近研究表明：LPS必须经系列的信号传递才能激活单核/巨噬细胞，调控核受体活性使众多LPS相关传导因子染色质发生改变是诱导ET建立的主要原因。NF-kB是LPS活化单核/巨噬细胞的最重要核受体之一，而富集表达于肝脏的RIP140又是调控单核/巨噬细胞NF-kB转录活性的关键性辅激活因子，抑制RIP140表达即可诱导ET。 鉴于体内80%-90%的单核/巨噬细胞是定植于肝血窦的KCs，我们提出，如能证实RIP140也是LPS诱导KCs活化的关键调控因子，那抑制RIP140表达将从调控核受体活性角度有效阻断LPS对再灌注期KCs的激活并诱导建立ET，从而为防治肝移植I/RI的基因治疗提供适宜的新靶点。

项目组前期研究证实内毒素(LPS)触发的再灌注期 Kupffer 细胞(KCs)过度激活是启动肝移植缺血再灌注损伤(I/RI)进程的关键事件，诱导 KCs 对内毒素耐受(ET)可通过抑制KC激活并减轻I/RI，但其确切机制并不清楚。在基金资助下，项目组通过相关实验研究获得了以下重要信息：（1）通过对不同细胞转染方式的对比，找到了KCs慢病毒干预转染新的理想途径，使得对KCs的基因干预过程能在最低细胞毒性下获得最佳的转染效率，同时实验条件可控、效果稳定，为后续KCs相关研究打下良好基础；（2）通过肝移植I/RI动物模型及相关细胞水平研究，找到了调控KC激活及内毒素耐受的关键分子RIP140，并证实了它在肝移植I/RI进程中的重要作用，提示其作为治疗靶点的可行性和有效性，为临床防治肝移植 I/RI提供了潜在的新策略；（3）通过对细胞间信息传递机制的相关研究，发现了肝星状细胞（HSCs）通过释放携炎症信号的外泌体以激活KCs并推进肝移植I/RI这一新机制，并证实了miR-9作为炎症调控的关键分子在肝移植I/RI中发挥的重要作用及其作为监测/调控靶点的潜在可行性。以上研究结果将从KCs相关基础研究、肝移植I/RI临床防治靶点选择以及相关新机制探索等三个层面为解决临床肝移植I/RI问题提供理论思路和研究基础。