目的 观察缺氧后处理与氧自由基清除剂超氧化物歧化酶(SOD)联合过氧化氢酶(CAT)预处理,对心肌细胞膜和线粒体Bcl-2、Bax蛋白表达的影响,探讨两者调控心肌细胞凋亡的机制.方法 建立乳鼠心肌细胞缺氧/复氧损伤模型,分对照组、缺氧/复氧组、缺氧后处理组及缺氧/复氧+SOD+CAT组.应用荧光探针测定心肌细胞线粒体活性氧水平,Hoechst染色及流式细胞仪检测心肌细胞凋亡,Western blot法检测心肌细胞膜和线粒体Bcl-2、Bax蛋白表达.结果 与对照组比较,缺氧/复氧组、缺氧后处理组、缺氧/复氧+SOD+CAT组细胞线粒体活性氧及细胞凋亡率明显升高(P＜0.01)；与缺氧/复氧组比较,缺氧后处理组、缺氧/复氧+ SOD+CAT组细胞线粒体活性氧及细胞凋亡率明显降低(P＜0.01)；缺氧后处理组及缺氧/复氧+ SOD+ CAT组细胞膜和线粒体Bcl-2蛋白明显上调,Bax蛋白明显下调.结论 缺氧后处理及氧自由基清除剂预处理抑制线粒体活性氧爆发,减轻缺氧/复氧再灌注心肌细胞凋亡,其抗凋亡可能机制与细胞膜和线粒体Bcl-2蛋白表达上调及Bax蛋白表达下调有关.%Objective To observe the effect of postconditioning and free radical scavengers,super-oxide dismutase(SOD)and catalase(CAT) ,on expression of Bcl-2 and Bax proteins in cardiomyo-cytes and mitochondria,and study the machinism of Bcl-2 and Bax underlying apoptosis of cardio-myocytes. Methods A hypoxia(H)/reoxygenation(R) model of suckling mouse cardiomyocytes was established. The suckling mouse cardiomyocytes were divided into control group, H/R group, postconditioning (PC) and H/R+SOD+CAT group. Mitochondrial reactive oxygen species(ROS) in cardiomyocytes and mitochondria were detected with fluorescence probes. Apoptosis of cardio-myocyte was assayed by flow cytometry with Hoechst 33342 staining. Expressions of Bcl-2 and Bax proteins in cardiomyocytes and mitochondria were detected by Western blot. Results The number of mitochondrial ROS and apoptotic cardiomyocytes was significantly less in PC group and H/R+SOD+CAT group than in H/R group[28. 59±2. 93 and 28. 43±1. 76 vs 58. 19± 3.29,(26.99 + 3.35)% and (26.45 + 5.08)% vs (45. 86 + 3. 29)%,P<0. 01]. The Bcl-2 expression level was higher while the Bax expression level was lower in PC group and H/R + SOD + CAT group than in H/R group. Conclusion Postconditioning and free radical scavengers reduce hypoxia while reoxygenation induces apoptosis of cardiomyocytes by upregulating the Bcl-2 expression and down-regulating the Bax expression in cardiomyocytes and mitochondria.
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