【摘要】 目的研究锌离子对脂多糖(LPS)诱导的大鼠腹膜间皮细胞(RPMC)凋亡的影响及机制的探索,从而为锌离子在腹膜透析中的应用提供实验基础。方法胰蛋白酶消化法培养原代大鼠腹膜间皮细胞、传代、经鉴定后分组:①对照组;②LPS组;③ZnSO4组;④ZnSO4/LPS组。应用AnnexinV-FITC凋亡检测试剂盒及流式细胞仪检测RPMC凋亡率,并采用Westen Blot方法检测细胞外信号调节激酶(PERK),BAX,凋亡诱导因子(AIF),天冬氨酸特异性半胧氨酸蛋白酶3(caspase3),天冬氨酸特异性半胧氨酸蛋白酶9(caspase9)蛋白表达。应用流式细胞术,采用活性氧自由基(ROS)检测试剂盒检测各组别的ROS蛋白表达情况。结果与对照组相比,LPS组RPMC凋亡细胞数量和相关凋亡蛋白(BAX,AIF,caspase3,caspase9)表达升高(P<0.01)。与LPS组相比,ZnSO4作用组RPMC凋亡细胞数量和相关凋亡蛋白表达明显降低(P<0.01)。LPS组ROS显著高于对照组而ZnSO4作用组ROS显著低于LPS组(P<0.01,P<0.01)。与LPS组相比,ZnSO4作用组P-ERK的表达明显升高(P<0.01)。结论 ZnSO4可能可以逆转LPS所致的RPMC凋亡,对腹膜透析相关腹膜炎过程中所导致的RPMC损伤具有保护作用,其作用机制可能是通过ERK通路而发挥作用。更多还原

【Abstract】 Objective To explore whether Zn+2 can be used in peritoneal dialysis, we investigated the effect of Zn + 2 on lipopolysaccharide(LPS)- induced cell apoptosis in rat peritoneal mesothelial cells(RPMCs) and its underlying mechanism. Methods RPMCs were isolated by enzyme disaggregation, cultured, and then identified by phase contrast microscopy, transmission electron microscopy, and immunocytochemistry methods. RPMCs were incubated with 100 μg/ml LPS for 24 hours, or stimulated by 100 μg/ml LPS after incubation with 50 μM ZnSO4for 24 hours. RPMCs incubated in regular medium were used as the controls. The expressions of p-ERK, BAX, AIF, caspase 3, and caspase 9 were assayed by western blot. Reactive oxygen species was measured by a reactive oxygen species assay kit. Results ZnSO4significantly inhibited the LPS- induced RPMC apoptosis by attenuating ROS production, inhibiting LPS- induced BAX, AIF, and the activation of caspase 3 and caspase 9. Further studies revealed that ZnSO4treatment facilitated cell survival through the activation of ERK signaling pathway. Conclusion These results indicate that Zn+2 inhibits LPS-induced apoptosis in RPMCs by attenuating ROS production and inhibiting BAX, AIF, and the activation of caspase 3 and caspase 9. The ERK signaling pathway may take part in these changes.更多还原