冬凌草甲素诱导人肾癌A-704细胞凋亡及机制研究

目的:研究冬凌草甲素( oridonin,Ori)对人肾癌细胞的生长抑制、诱导凋亡作用及其机制.方法:MTT法检测Ori对密度为1×104/mL的人肾癌A-704细胞的生长抑制作用;流式细胞仪Annexin V-FITC/PI双染色法检测Ori作用于人肾癌A-704细胞24 h的细胞凋亡率;实时监测聚合酶链反应(Real-time PCR)法检测32 μmol·L-1Ori作用人肾癌A-704细胞24h后的Bcl-2,Bax及Caspase-3基因表达水平的变化.结果:Ori可显著抑制人肾癌A-704细胞的生长,并呈明显的时间-效应关系和浓度-效应关系,64 μmol· L-1 Ori作用人肾癌A-704细胞60 h后抑制率为73％.随着Ori作用浓度增加,凋亡细胞数和坏死细胞数均增加,浓度越高,坏死细胞率增加越多,64 μmol·L-1Ori作用人肾癌A-704细胞24 h后,坏死细胞升至32.4％.随着32 μmol·L-1Ori作用时间延长,人肾癌A-704细胞Bax,Caspase-3基因的表达逐渐增强,Bcl-2基因的表达逐渐减弱(P均＜0.05).结论:Ori通过上调Bax基因和降低Bcl-2基因表达诱导细胞凋亡,从而抑制人肾癌A-704细胞的生长.

Oridonin Induced Apoptosis of Human Kidney Carcinoma Cells and its Mechanism

Objective:To investigate apoptosis induction effects of oridonin(Ori)on human kidney carcinoma cells and its mechanisms in vitro.Method:The MTT assay was used to measure the inhibitory effects of oridonin at the concentration of 1×104/mL on A-704 cells.The apoptosis rate of A-704 cells which were impacted by Ori for 24 hours was examined by Annexin V-FITC/PI staining of Flow cytometry with the increase of the concentration,the number of apoptotic cells and necrotic cells were increased,the higher the concentration,the more the ratio of necrotic cells increased expression of Bax,Caspase-3 and Bcl-2 of A-704 cells which were impacted by 32 μmol · L-1oridonin for 24 hours were evaluated by Real-time PCR.Result:Ori could inhibit the growth of A-704 cells significantly in a time-dependent and dose-dependent manner.The inhibition rate of A-704 cells which were impacted by 64 μmol · L-1oridonin for 24 hours could be reach 73%.With the concentration of Ori increased,the number of apoptotic and necrosis cells both increased.The higher concentration,the more rate of necrotic cells increased.The ratio of necrosis cells of A-704 cells which were impacted by 64 μmol · L-1Ori for 24 hours could be reach 32.4%.Along with the extention of time,Bax and Caspase-3 expressions were up-regulated,Bcl-2 expression was down-regulated(all P<0.05).Conclusion:Ori can inhibit A-704 cells growth by induction of apoptosis in cells via activation of Caspase-3 as well as up-regulation of Bax and down-regulation of Bcl-2 gene expression.