摘要
目的:通过研究雷公藤内酯醇(TP)对COC1/DDP体外活性的影响,初步探讨雷公藤内酯醇促进COC1/DDP细胞凋亡的机制,为TP成为治疗晚期或耐顺铂卵巢癌药物提供实验依据。方法:采用MTT法检测顺铂(DDP)、TP及两者联用对COC1/DDP的生长抑制作用;用透射电镜观察TP作用24h后细胞超微结构的变化;采用流式细胞术分析不同浓度TP对COC1/DDP细胞周期和细胞凋亡的影响;用DNA电泳分析用药后细胞基因组DNA断裂状况;以免疫组化分析TP对Caspase-7蛋白表达的影响。结果:DDP在24h、48h和72h三个时间段对COC1/DDP细胞的半数抑制量(50%concentration of inhibi-tion,IC50)分别为5.567μg/ml、2.866μg/ml和1.161μg/ml,其对COC1/DDP细胞增殖表现出浓度-时间依赖性的抑制作用(P<0.05);TP可抑制COC1/DDP细胞增殖,其抑制率呈浓度-时间依赖性(P<0.05);TP作用细胞24h后,电镜下可见凋亡小体形成;流式细胞术分析表明,各浓度TP组G1期细胞明显升高,细胞凋亡率呈浓度依赖性(P<0.05);DNA电泳分析可见细胞凋亡特有的DNA断裂形成的"梯形"条带;免疫组化表明Caspase-7参与了TP诱导COC1/DDP细胞凋亡过程。结论:TP对COC1/DDP具有明显的杀伤和促凋亡作用,凋亡机制与Caspase-7蛋白表达上调有关。
Objective：To study the effect and underlying mechanism of triptolide on COC1/DDP in vitro.Methods：MTT assay was used to analyze the growth inhibition effect of DDP and TP.Twenty-four hours after the treatment of TP,transmission electron microscopy was used to observe COC1/DDP ultrastructure,and flow cytometry was used to observe the change of cell apoptosis rate and cell cycle in different concentration of TP.The apoptosis fracture zone was investigated with DNA electrophoresis.The difference of caspsase-7 protein expression was monitored by immunohistochemistry.Results：The results showed the inhibitory effect of DDP on the COC1/DDP cells in a dose and time dependent manner（P0.05）.The IC50 after 24h was 5.567μg/ml,2.866μg/ml after 48h and 1.161μg/ml after 72h.TP could inhibit COC1/DDP in a time-and dose-dependent manner.After treatment of TP by 24 hours,apoptotic body formation could be observed under transmission electron microscopy.Apoptosis rate was positively correlated with TP dose.Apoptosis cycle was arrested at G1 phase.Different lengths of apoptosis specific DNA break were presented as specific ＂ladder＂ change observed by agarose gel electrophoresis.Immunohistochemistry showed that caspase-7 protein was involved in COC1/DDP cell apoptosis induced by TP.Conclusions：TP can inhibit the growth and promote the apoptosis of COC1/DDP.Apoptosis mechanism appears to be related to caspase-7 protein up-regulation.
出处
《现代妇产科进展》
CSCD
北大核心
2011年第11期846-850,共5页
Progress in Obstetrics and Gynecology
基金
江西省科技支撑计划和江西省教育厅科技计划项目(No:GJJ08074)
