Liu Ling, Yang Cheng, Herzog Christian, Seth Rohit, Kaushal Gur P
University of Arkansas for Medical Sciences, Department of Medicine, Little Rock, Arkansas 72205, United States.
Biochem Pharmacol. 2010 Jan 15;79(2):137-46. doi: 10.1016/j.bcp.2009.08.015. Epub 2009 Aug 20.
We demonstrate the effect of proteasome inhibitors in mitochondrial release of apoptosis-inducing factor (AIF) in cisplatin-exposed renal tubular epithelial cells (LLC-PK(1) cells) and in a model of cisplatin nephrotoxicity. Immunofluorescence and subcellular fractionation studies revealed cisplatin-induced translocation of AIF from the mitochondria to nucleus. Mcl-1, a pro-survival member of the Bcl-2 family, is rapidly eliminated on exposure of renal cells to cisplatin. Proteasome inhibitors PS-341 and MG-132 blocked cisplatin-induced Mcl-1 depletion and markedly prevented mitochondrial release of AIF. PS-341 and MG132 also blocked cisplatin-induced activation of executioner caspases and apoptosis. These studies suggest that proteasome inhibitors prevent cisplatin-induced caspase-dependent and -independent pathways. Overexpression of Mcl-1 was effective in blocking cisplatin-induced cytochrome c and AIF release from the mitochondria. Downregulation of Mcl-1 by small interfering RNA promoted Bax activation and cytochrome c and AIF release, suggesting that cisplatin-induced Mcl-1 depletion and associated Bax activation are involved in the release of AIF. Expression of AIF protein in the mouse was highest in the kidney compared to the heart, brain, intestine, liver, lung, muscle, and spleen. In an in vivo model of cisplatin nephrotoxicity, proteasome inhibitor MG-132 prevented mitochondrial release of AIF and markedly attenuated acute kidney injury as assessed by renal function and histology. These studies provide evidence for the first time that the proteasome inhibitors prevent cisplatin-induced mitochondrial release of AIF, provide cellular protection, and markedly ameliorate cisplatin-induced acute kidney injury. Thus, AIF is an important therapeutic target in cisplatin nephrotoxicity and cisplatin-induced depletion of Mcl-1 is an important pathway involved in AIF release.
我们证明了蛋白酶体抑制剂在顺铂处理的肾小管上皮细胞（LLC-PK(1)细胞）以及顺铂肾毒性模型中对凋亡诱导因子（AIF）线粒体释放的影响。免疫荧光和亚细胞分级分离研究显示，顺铂诱导AIF从线粒体转位至细胞核。Mcl-1是Bcl-2家族的一个促生存成员，肾细胞暴露于顺铂时会迅速被清除。蛋白酶体抑制剂PS-341和MG-132可阻断顺铂诱导的Mcl-1耗竭，并显著防止AIF从线粒体释放。PS-341和MG132还可阻断顺铂诱导的执行性半胱天冬酶激活和凋亡。这些研究表明，蛋白酶体抑制剂可预防顺铂诱导的半胱天冬酶依赖性和非依赖性途径。Mcl-1的过表达可有效阻断顺铂诱导的细胞色素c和AIF从线粒体释放。小干扰RNA下调Mcl-1可促进Bax激活以及细胞色素c和AIF释放，表明顺铂诱导的Mcl-1耗竭及相关的Bax激活参与了AIF的释放。与心脏、脑、肠、肝、肺、肌肉和脾脏相比，AIF蛋白在小鼠肾脏中的表达最高。在顺铂肾毒性的体内模型中，蛋白酶体抑制剂MG-132可防止AIF从线粒体释放，并通过肾功能和组织学评估显著减轻急性肾损伤。这些研究首次提供证据表明，蛋白酶体抑制剂可预防顺铂诱导的AIF从线粒体释放，提供细胞保护，并显著改善顺铂诱导的急性肾损伤。因此，AIF是顺铂肾毒性中的一个重要治疗靶点，顺铂诱导的Mcl-1耗竭是AIF释放所涉及的一个重要途径。
