Overexpression of endoplasmic reticulum-resident chaperone attenuates cardiomyocyte death induced by proteasome inhibition
Overexpression of endoplasmic reticulum-resident chaperone attenuates cardiomyocyte death induced by proteasome inhibition
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DOI：
10.1093/cvr/cvn128
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发表时间：
2008-09-01
影响因子：
10.8
通讯作者：
Kitakaze, Masafumi
中科院分区：
文献类型：
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作者：
Fu, Hai Ying;Minamino, Tetsuo;Kitakaze, Masafumi
目的蛋白酶体抑制剂是一类诱导肿瘤细胞生长的新型抗癌药物。通过内质网(ER)应激死亡。鉴于内质网应激参与心力衰竭的发生发展，我们研究了蛋白酶体抑制在内质网启动的心肌细胞死亡中的作用。用蛋白酶体酶底物检测蛋白酶体活性。3-(4，5-二甲基噻唑-2-基)-2，5-二苯基四氮唑溴化法检测细胞存活率，流式细胞仪检测细胞凋亡率。采用蛋白印迹分析、实时定量聚合酶链式反应和逆转录聚合酶链式反应检测蛋白质和信使核糖核酸的表达。共聚焦荧光显微镜观察高表达的葡萄糖调节蛋白(GRP)78的定位。蛋白酶体抑制可诱导心肌细胞死亡，激活内质网应激诱导的转录因子ATF6，但不能激活XBP1(X盒结合蛋白1)，而不上调内质网伴侣蛋白。胞嘧啶-胞嘧啶-腺嘌呤-腺嘌呤-胸腺嘧啶增强子结合蛋白(C/EBP)同源蛋白(CHOP)、c-Jun-N末端激酶(JNK)和caspase-12等内质网启动的细胞凋亡信号被蛋白酶体抑制激活。针对CHOP的短干扰RNA可减轻心肌细胞死亡，但不能阻断caspase-12或JNK途径。GRP78过表达可通过蛋白酶体抑制抑制CHOP的表达和心肌细胞的死亡。结论蛋白酶体抑制通过CHOP依赖的途径诱导ER启动的心肌细胞死亡，而不是ER伴侣的代偿性上调。补充和/或药物诱导GRP78可减轻蛋白酶体抑制所致的心肌损伤。
Aims Proteasome inhibitors are a novel class of anticancer agents that induce tumour cell. death via endoplasmic reticulum (ER) stress. Since ER stress is involved in the development of heart failure, we investigated the role of ER-initiated cardiomyocyte death by proteasome inhibition.Methods and results Rat neonatal cardiomyocytes were used in this study. Proteasome activity was assayed using proteasome peptidase substrates. Cell viability and apoptosis were measured by 3-(4,5-dimethylthiazol-2-yl)- 2,5-diphenol tetrazolium bromide and flow cytometry, respectively. Western blot analysis, real-time polymerase chain reaction (PCR) and reverse transcriptional PCR were used to detect the expression of protein and messenger ribonucleic acid (RNA). The location of overexpressed glucose-regulated protein (GRP) 78 was observed by confocal fluorescence microscopy. Proteasome inhibition induced cardiomyocyte death and activated ER stress-induced transcriptional factor ATF6, but not XBP1 (X-box binding protein 1), without up-regulating ER chaperones. ER-initiated apoptosis signalling, including cytosine-cytosine-adenine-adenine-thymine enhancer-binding protein (C/EBP) homologous protein (CHOP), c-Jun-N-terminal kinase (JNK), and caspase-12, was activated by proteasome inhibition. Short interference RNA targeting CHOP, but not the blockage of caspase-12 or JNK pathway, attenuated cardiomyocyte death. Overexpression of GRP78 suppressed both CHOP expression and cardiomyocyte death by proteasome inhibition.Conclusion These findings demonstrate that proteasome inhibition induces ER-initiated cardiomyocyte death via CHOP-dependent pathways without compensatory up-regulation of ER chaperones. Supplement and/or pharmacological induction of GRP78 can attenuate cardiac damage by proteasome inhibition.