摘要
目的 探讨在乳腺癌细胞系MCF-7中下调S期激酶相关蛋白（Skp2）表达诱导细胞凋亡及其机制.方法 应用RNAi方法在体外下调乳腺癌细胞系MCF-7中Skp2的表达水平,48小时后表阿霉素处理细胞,TUNEL和Hoechst 33258染色检测凋亡,Western Blot检测细胞周期调控相关因子及凋亡相关蛋白表达情况,研究其机制.结果 下调MCF-7中Skp2表达水平后,乳腺癌细胞凋亡增加.下调Skp2使p27、p21和CyclinE蛋白表达水平升高.表阿霉素处理MCF-7细胞后,Skp2蛋白水平下调.Skp2 siRNA与表阿霉素有协同诱导凋亡的作用,p53蛋白水平升高.结论 p27、p21和CyclinE在通过下调Skp2诱导的凋亡中发挥作用.Skp2 siRNA和表阿霉素协同诱导细胞凋亡,与p53依赖的凋亡途径有关.Skp2可能是乳腺癌治疗的靶点.
Objective The F - box protein S - phase kinase - associated protein 2 （Skp2） is one of the positive regulators of the cell cycle that promotes abiquitin mediated proteolysis of some cyclin - dependent kinase inhibitors. In this study, we investigated the mechanism of apoptosis induced by down - regulation Skp2 expression using siRNA in MCF - 7 human breast cancer cell lines. Methods We downregulated Skp2 in MCF - 7 cells using siRNA （ small interfering RNA ）. After 48h, cells were treated with epirnbicin. TUNEL assay and Hoeehst33258 stain were used to detected apoptosis. We also investigated the mechanism of apoptosis by detecting the expression of some factors associated with cell cycle using Western blot analysis. Results Knockdown of Skp2 by RNAi increased cellular apoptosis in vitro. Down regulation of Skp2 increased the level of p27, p21 and CyclinE protein. Epirubicin also decreased Skp2 protein levels in MCF - 7 cells. Treatment with Skp2 siRNA fol- lowed by treatment with epirnbicin not only increased apoptosis synergistically, but also activated p53 in MCF -7 cells. Conclusion In vitro, Skp2 siRNA induced apoptosis in breast cancer cells, p27, p21 and CyclinE regulated apoptosis by downregulation of Skp2 in breast cancer cells. Skp2 RNAi sensitized MCF -7 cells to apoptosis induced by epirubicin potentially through p53 - dependent way. Skp2 could be a therapeutic target in breast cancer.
出处
《实用肿瘤学杂志》
CAS
2012年第2期109-113,共5页
Practical Oncology Journal
基金
黑龙江省科技厅青年基金（QC08C07）
哈尔滨市科技创新人才项目（2009RFXXS017）
黑龙江省卫生厅重点项目（2006-189）
