苦参碱联合Akt信号通路抑制剂影响肾癌细胞的活力与凋亡

Effect of matrine combined with an Akt signaling pathway inhibitor on viability and apoptosis in renal carcinoma cells in vitro

目的本研究探讨苦参碱联合蛋白激酶B(Akt)信号通路抑制剂对肾癌细胞增殖活力及凋亡的影响及机制。方法以肾癌细胞为研究对象,用不同浓度的苦参碱作用后,噻唑蓝(MTT)检测细胞增殖活力,计算半数抑制浓度。用半数抑制浓度的苦参碱和Akt信号通路抑制剂分别处理细胞记为苦参碱组和抑制剂组,同时以半数抑制浓度的苦参碱和Akt信号通路抑制剂共同处理后的细胞为联合组,以正常培养的细胞为对照组,MTT检测细胞增殖活力,流式细胞术检测细胞凋亡,激光扫描共聚焦显微镜检测细胞中活性氧(reactive oxygen species,ROS)水平,蛋白质印迹法检测细胞中p38、磷酸化的p38(p-p38)、活化的含半胱氨酸的天冬氨酸蛋白水解酶3(cleaved cysteinyl aspartate specific proteinase 3,cleaved caspase-3)表达。结果苦参碱能够呈浓度依赖的抑制肾癌细胞增殖活力。与对照组比较,苦参碱组、抑制剂组、联合组细胞存活率降低,凋亡率升高,ROS水平升高,p-p38水平、cleaved caspase-3蛋白水平升高。与苦参碱组、抑制剂组比较,联合组胞存活率降低,凋亡率升高,ROS水平升高,p-p38水平、cleaved caspase-3蛋白水平升高。结论苦参碱联合Akt信号通路抑制剂能够抑制肾癌细胞增殖活力,促进肾癌细胞凋亡,ROS和p38信号通路可能是其作用机制之一。

Objective To explore the effect of matrine combined with an Akt signaling pathway inhibitor on the proliferation and apoptosis in renal carcinoma cells in vitro.Methods Renal cell carcinoma ACHN and GRC-1 cells were treated with different concentrations of matrine,and cell proliferation activity was detected by MTT assay to calculate the half inhibitory concentration.Cells treated with a half inhibitory concentration of matrine or an Akt pathway inhibitor were named the matrine group and inhibitor group,respectively;cells treated with both were the combined group,and untreated cells were used as controls.Cell proliferation was detected by MTT,apoptosis was measured by flow cytometry,levels of reactive oxygen species(ROS)were examined by laser scanning confocal microscopy,and the expression of p38,p-p38,and cleaved caspase-3 was investigated by western blotting.Results Matrine can inhibit the proliferation of renal cancer cells in a concentration-dependent manner.Compared with the control group,the cell survival rate decreased,apoptosis rate,ROS level,p-p38 level and cleaved caspase-3 protein level increased in the matrine,inhibitor and combination groups.Compared with the matrine group and inhibitor group,the combined group had a lower survival rate,higher apoptosis rate,higher ROS level,higher p-p38 level and cleaved caspase-3 protein level.Conclusions Matrine combined with an Akt signaling pathway inhibitor inhibits the proliferation of renal carcinoma cells,and promotes their apoptosis.ROS and p38 signaling pathways may be one of the mechanisms of action of this effect.