PINK1/PARK2介导的线粒体自噬在对比剂致急性肾损伤中的作用及机制

Reactive oxygen species (ROS) overproduction and renal tubular epithelial cell (TEC) apoptosis are key mechanisms of contrast media-induced acute kidney injury (CI-AKI), but the underlying molecular mechanisms that contrast media induce ROS overproduction remain unclear. It is rarely reported that whether mitochondria injury and mitochondrial autophagy (mitophagy) are involved in the contrast media-induced ROS overproduction and renal tubular cell apoptosis. Our previous studies have demonstrated that the exposure of contrast media could activate PINK1/PARK2 signal pathway and induce mitophagy in cultured renal tubular epithelial cell, and that enhancing autophagy could alleviate contrast media-induced ROS overproduction and decrease TEC apoptosis. In the present program, the role and molecular mechanism of PINK1/PARK2-mediated mitophagy in CI-AKI will be explored: The effects of inhibiting or enhancing autophagy on contrast media-induced changes in ROS production, mitochondrial injury, mitophagy and TEC injury will be observed in CI-AKI rats and in cultured TECs; The signal transduction pathway of contrast media-induced mitophagy and the relationship between mitophagy and ROS will be explored in cultured TECs by PARK2 gene silencing; And the role and molecular mechanism of PINK1/PARK2-mediated mitophagy in the CI-AKI will be further investigated in PARK2 gene knockout rats. In summary, the new therapy target points against CI-AKI and associated theory basis are proposed in the present program.

活性氧(ROS)过度产生及肾小管上皮细胞(TEC)凋亡是对比剂致急性肾损伤（CI-AKI）的关键机制。但对比剂诱导ROS过度产生的分子机制仍不清楚。线粒体损伤及线粒体自噬改变是否影响对比剂诱导的ROS增加及TEC凋亡鲜见报道。我们前期研究发现对比剂可激活PINK1/PARK2信号通路，诱导TEC线粒体自噬；增加自噬可降低ROS、减少TEC凋亡。本项目拟进一步研究PINK1/PARK2介导的线粒体自噬改变在CI-AKI中的作用及分子机制：通过体外实验和CI-AKI动物模型探讨增强和抑制自噬对TEC线粒体损伤、线粒体自噬、ROS生成、细胞损伤等的影响；通过PARK2基因沉默探讨对比剂诱导线粒体自噬发生的信号传导通路及其与ROS的关系；通过PARK2基因敲除进一步研究PINK1/PARK2介导的线粒体自噬在CI-AKI中的作用及分子机制。本研究将为CI-AKI的防治提供新的治疗靶点及理论依据。

肾小管上皮细胞(Renal tubular epithelial cell, RTEC)异常凋亡和氧化应激是对比剂致急性肾损伤（CI-AKI）的重要原因，但其调节机制仍不清楚。本项目组前期研究发现对比剂可激活PINK1/PARK2介导的线粒体自噬,增强线粒体自噬可减轻ROS产生和RTEC凋亡。本项目旨在探讨PINK1/PARK2介导的线粒体自噬在对比剂致急性肾损伤中的作用和分子机制。本研究发现对比剂在体外培养RTEC及体内动物模型中均能诱导RTEC线粒体损伤、氧化应激激活及细胞凋亡，且氧化应激激活剂ox-LDL可模拟对比剂对肾小管上皮细胞的损伤效应，两者共处理可加重急性肾损伤。对比剂刺激可诱导自噬相关蛋白LC3Ⅱ/Ⅰ、Beclin-1和Pink1的表达，自噬增强剂雷帕霉素预处理可减轻对比剂诱导的肾小管上皮细胞凋亡及氧化应激。利用Parkin siRNA转染抑制HK2细胞线粒体自噬可进一步加重了对比剂诱导的细胞凋亡和氧化应激。结论：PINK1/PARK2信号通路介导的线粒体自噬参与RTEC氧化应激和细胞凋亡的调节。在对比剂刺激或ox-LDL刺激等病理条件下，RTEC的PINK1/PARK信号通路激活，线粒体自噬增强，但其不足以拮抗病理状态下的RTEC损伤。进一步增强PINK1/PARK信号诱导线粒体自噬可减少ROS产生和细胞凋亡，减轻对比剂引起的肾脏损伤。

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