【摘要】 目的以蛋白激酶C-ε(PKC-ε)转位为切入点,探讨2类多巴胺受体(DR2)在心肌缺血后适应抑制细胞凋亡中的作用及可能机制。方法复制原代培养乳鼠心肌细胞缺氧/复氧和缺血后适应模型。MTT检测心肌细胞的存活率;Hoechst 33342染色观察细胞凋亡;Western blotting检测Bcl-2、caspase-3、caspase-9、PKC-ε蛋白的表达和细胞色素C(Cyt c)的释放;免疫共沉淀检测PKC-ε和DR2的相互作用。结果与正常组比较,缺氧/复氧组细胞存活率降低,细胞凋亡增加,促凋亡因子(Cyt c、caspase-3、caspase-9)表达增加,抑凋亡因子(Bcl-2)表达亦增加,PKC-ε没有转位到细胞膜,PKC-ε和DR2不存在相互作用。与缺氧/复氧组比较,缺血后适应明显升高细胞存活率,降低细胞凋亡,抑制促凋亡因子(Cyt c、caspase-3、caspase-9)表达,促进抑凋亡因子(Bcl-2)表达,PKC-ε转位到细胞膜,PKC-ε和DR2存在相互作用。与缺血后适应组比较,DR2激动剂进一步增加缺血后适应的保护作用,而DR2抑制剂则取消了DR2激动剂的作用。结论DR2参与心肌缺血后适应抑制细胞凋亡,其机制与DR2促进PKC-ε转位及DR2和PKC-ε存在相互作用有关。更多还原

【Abstract】 Objective lo investigate the eflect and possible mechanism of dopamine receptor-2(DR2) activation in ischemic postconditioning(PC) inhibited cardiomyocytes apoptosis through translocation of PKC-ε.Method The hypoxia/reoxygenation(H/R) injury and PC models was established using primarily cultured neonatal rat cardiomyocytes.The cell survival rate was detected by MTT.The cell apoptosis was observed using Hoechst 33342 starling.The protein expression of Bcl-2,caspase-3,caspase-9,the release of cytochrome c(Cyt c) and translocation of PKC-e were analyzed using Western blotting.The interaction of DR2 and PKC-e was tested by co-immunoprecipitation.Result Compared with Control group,the cell survival rate was decreased,cardiomyocytes apoptosis was heightened,the expression of Bcl-2,caspase-3,caspase-9 and the release of Cyt c were increased,PKC-e were not translocated to the cell membrane and the interaction of DR2 and PKC-e was not exist in H/R group.Compared with H/R group,PC increased the cell survival rate,decreased cardiomyocytes apoptosis,down-regulated the expression of caspase-3,caspase-9 and the release of Cyt c,up-regulated the expression of Bcl-2,promoted translocation of PKC-e and the interaction of DR2 and PKC-ε.Compared with PC group,the agonist of DR2 further increased cardioprotection of PC,however,the antagonist of DR2 canceled this effect of DR2 agonist.Conclusion DR2 activation is involved in PC inhibited apoptosis by promoting translocation of PKC-ε.更多还原