【摘要】 目的:探讨原癌基因Pim-3对抗心肌急性损伤的作用是否与线粒体信号转导途径介导的细胞色素C(CytC)的释放有关。方法:采用大鼠乳鼠心肌细胞,随机分为4组,对照组;缺氧复氧(A/R)组,建立A/R损伤模型;pcDNA3.1+A/R组,将pcDNA3.1转染入心肌细胞,24h后进行A/R损伤;pcDNA3.1/Pim-3+A/R组,将Pim-3重组质粒(pcDNA3.1/Pim-3)转染入心肌细胞,24h后进行A/R损伤。实验结束后,检测培养液中乳酸脱氢酶(LDH)活性,四唑盐(MTT)比色试验测定细胞存活率,TUNEL法检测细胞凋亡,蛋白印迹检测Pim-3蛋白表达水平,线粒体、胞浆CytC动态变化和caspase-3活性。结果:与对照组比较,A/R组细胞存活率明显下降,LDH值和凋亡指数明显升高,差异有统计学意义(P<0.01);与A/R组比较,pcDNA3.1/Pim-3+A/R组细胞存活率明显升高,LDH值和凋亡指数明显下降,并且CytC由线粒体向胞浆的释放明显被抑制,同时caspase-3活性也下降,差异有统计学意义(P<0.01)。结论:Pim-3对抗心肌急性损伤的机制是通过抑制CytC易位释放入胞浆激活caspase-3而致。更多还原

【Abstract】 Objective:To investigate the protective effect of proto-oncogene Pim-3 on cardiomyocytes against anoxia-reoxygenation (A/R) injury and the relationship with the cytochrome C release mediated by mitochondrial signal transduction pathways thereof. Methods:The primary neonatal rat ventricular cardiomyocytes were randomly divided into 4 groups:control group; A/R group:the A/R models of primary cultured neonatal rat myocytes; pcDNA3.1+A/R group:pcDNA3.1 was transfected into the cardiomyocytes,then A/R models were established 24 h after transfection; pcDNA3.1/Pim-3+A/R group:pcDNA3. 1/Pim-3 were transfected into the cardiomyocytes,then A/R models were established 24 h after transfection. The level of lactate dehydrogenase (LDH) activity,MTT and TUNEL were detected after treatment. The expressions of Pim-3 and caspase-3 and the release of cytochrome C from mitochondria to cytoplasm were detected by Western blotting. Results:In the A/R group,the cellular survival was significantly decreased,while the LDH activity and the apoptotic index of the cells were markedly increased compared with those of the control group (P < 0.01). In the pcDNA3.1/Pim-3+A/R group,the cellular survival was significantly increased while the LDH activity and the apoptotic index of the cells were markedly decreased compared with that of the A/R group. Furthermore,the release of cytochrome C from mitochondria to cytoplasm was inhibited and the expression of caspase-3 was significantly decreased in the A/R group(P < 0.01). Conclusion:The protective mechanism of Pim-3 should be the inhibition of the release of cytochrome C from mitochondria to cytoplasm,which leads a suppression of caspase-3 activity.更多还原