COX-2通过内源性EP信号系统调控线粒体功能的分子机制及其在肝脏缺血损伤中的作用

Hepatic ischemia/reperfusion (I/R) injury is a common complication during major hepatic surgeries such as hepatic transplantation and hepatectomy. It greatly slows postoperative recovery and is a burden to medical care. Although the specified mechanisms are not known, emerging evidences suggest a crucial role of mitochondrial dysfunction. Mitochondrial permeability transition pore (MPTP) opening is a crucial step during I/R induced mitochondrial dysfunction and following cell death. We formerly found that inhibition of COX-2 could alleviate hepatic I/R injury by inhibiting MPTP opening( Mol Med. 2012,18:1128 ). However, the mechanism that COX-2 modulates MPTP susceptibility remains unclear. PGE2, a major product of COX-2 activation, binds to 4 G protein-coupled receptors(EP1-EP4) to transduce COX-2 signaling. Our ongoing study find that AE1-329, an EP4 agonist, has a similar hepatoprotective effect as COX-2 inhibitors. Theses findings support the hypothesis that PGE2/EP signaling,which is also called as" endogenous EP signaling", might mediate COX-2 signaling in modulating the motochondrial function during I/R. We intend to use the in vitro and in vivo hepatic I/R model to further testify the hypothesis. By utilizing siRNA technique and using selective signaling inhibitors, we aim to identify the possible signaling pathways and responsible proteins in PGE2/EP-mediated COX-2 signaling. Furthermore, we plan to test the protective role of drugs that target PGE2/EP signaling during hepatic I/R injury. Understanding the mechanism of PGE2/EP-mediated COX-2 signaling in modulating mitochondrial function is helpful for effective hepatoprotection strategy searching and is of potential clinical significance.

缺血再灌注(I/R)损伤是肝部分切除及肝移植等常见的临床病理现象，可严重影响术后疗效。I/R损伤主要涉及ROS产生、固有免疫活化介导的炎症及细胞凋亡等过程，而线粒体功能障碍是关键。前期研究中我们发现抑制COX-2信号能抑制线粒体MPTP开放，减轻肝I/R损伤(Mol Med. 2012,18:1128)，但对COX-2如何调控MPTP开放的分子机制缺乏认识。预实验结果提示COX-2下游的EP受体信号参与了I/R时线粒体功能的调控。基于此,本课题拟进一步研究I/R时COX-2信号调节内源性EP受体信号对肝线粒体功能影响的分子机制及关键的信号分子；采用I/R模型，应用siRNA、靶点抑制剂等体内探究肝脏I/R后COX-2调控肝线粒体功能的关键信号分子靶点及其对肝脏I/R的作用，为深入认识I/R损伤分子机制、寻找有效的分子药物靶点防治肝脏I/R损伤提供理论依据，具有重要的科学意义和临床应用价值。

缺血再灌注(I/R)损伤是肝部分切除及肝移植等常见的临床病理现象，可严重影响术后疗效。资料显示I/R损伤主要涉及ROS产生、固有免疫活化介导的炎症及细胞凋亡等过程，而线粒体功能障碍是关键。前期研究中我们发现抑制COX-2信号能抑制线粒体MPTP开放，减轻肝I/R损伤，但对COX-2如何调控MPTP开放的分子机制缺乏认识。本课题通过（1）肝细胞缺氧/复氧模型和在体大鼠70%肝热缺血再灌注损伤模型，观察了肝I/R 时COX-2 的表达改变对内源性EP信号系统的影响，包括PGE2 水平和肝细胞特定EP 受体的表达，通过基因和蛋白层面筛选出COX-2调控线粒体功能的重要信号受体EP4；（2）对EP4调控肝I/R时线粒体功能的分子机制和信号级联进行了研究，结果发现EP4可通过下游cAMP-ERK1/2信号进一步影响MPTP调控相关的GSK-3β和JAK2-STAT3信号改变，从而调控MPTP通透性，影响ROS生成，调控肝I/R损伤；（3）并以上述研究为基础，进一步揭示了COX-2/EP4下游的重要信号分子GSK-3β在肝I/R时线粒体调控机制，发现使用药物Sal或GSK-3抑制剂SB21干预GSK-3β位点，能显著减轻再灌注时肝细胞坏死和凋亡。机制主要与调节线粒体能量代谢，诱发线粒体兴奋效应，从而抑制再灌注时MPTP开放，减轻线粒体相关的细胞坏死和凋亡有关。本研究拓展了我们对肝I/R损伤分子机制的认识，有助于肝I/R损伤机理的全面阐述，也为围术期肝I/R损伤的防治提供了新思路。