雪胆素乙通过破坏微丝结构和促进p21Cip1表达抑制前列腺癌PC-3细胞的增殖  

Inhibitory effect of CucurbitacinⅡb on proliferation of human prostate cancer PC-3 cells through disruption of microfilaments and upregulation of p21^(Cip1) expression

作　　者：任帅[1] 徐丽慧[1,2] 曾龙辉[1] 欧阳东云[1] 何贤辉[1] 

机构地区：[1]暨南大学生命科学技术学院、免疫生物学系 [2]细胞生物学系,广东广州510632

出　　处：《中国药理学与毒理学杂志》2012年第6期835-841,共7页Chinese Journal of Pharmacology and Toxicology

基　　金：国家自然科学基金(81173604);中央高校基本科研业务费专项资金(21609403)~~

摘　　要：目的分析雪胆素乙(CuⅡb)对人前列腺癌PC-3细胞增殖及细胞周期的影响,探讨其作用机制。方法 MTS法检测CuⅡb 0.064～200μmol·L-1作用48 h后PC-3细胞增殖;CuⅡb 2和20μmol·L-1作用24 h,相差显微镜观察细胞形态;CuⅡb 2和20μmol·L-1作用48 h,流式细胞术分析细胞周期分布;免疫荧光染色分析CuⅡb 20μmol·L-1分别作用1,4和24 h后微丝和微管细胞骨架变化;免疫印迹法测定CuⅡb20μmol·L-1分别作用1,4和24 h后G肌动蛋白、F肌动蛋白、p21Cip1及细胞周期蛋白A,B1,E和D1的表达。结果 CuⅡb以浓度依赖方式抑制PC-3细胞的增殖(r=0.9817,P<0.05)。CuⅡb 20μmol·L-1作用48 h,使细胞周期阻滞于G2/M期,从溶剂对照组的(27.7±1.5)%上升为(45.3±1.8)%(P<0.01),相差显微镜见细胞发生明显收缩。CuⅡb 20μmol·L-1作用24 h,使G肌动蛋白水平显著下降,F肌动蛋白发生严重聚集(P<0.05),而对微管只有轻微影响。与溶剂对照组相比,CuⅡb 20μmol·L-1作用24 h后,细胞p21Cip1表达明显升高,细胞周期蛋白A表达显著下调,其他细胞周期蛋白表达上调(P<0.05)。结论 CuⅡb能明显抑制人前列腺癌PC-3细胞的增殖,其机制可能是通过诱导肌动蛋白聚集,破坏微丝骨架,促进抑癌因子p21Cip1表达,阻滞细胞周期的进程。OBJECTIVE To explore effects of cucurbitacin Ⅱb （ Cu Ⅱ b） on human prostate cancer PC-3 cells and its underlying mechanism. METHODS The proliferation of cells was detec- ted by MTS assay after PC-3 cells were treated with CuⅡb 0. 064 - 200 μmol· L-1 for 48 h. After treated with Cu Ⅱ b 2 and 20 μmol·L-1 for 24 h, cell morphologic changes were observed under phase contrast microscopy. After exposed with Cu Ⅱb 2 and 20 p,mol· L- 1 for 48 h, cell cycle distri- bution was measured by flow cytometry using propidium iodide （PI） staining. When cultured with CuⅡ b 20 μmol· L- 1 for 1, 4 and 24 h dividedly, the changes of microfilament and microtubule structures were assessed by immunofluorescence staining. After separately treated with Cu Ⅱb 20 μmol·L-1 for 1,4 and 24 h, the protein expression levels of F-actin, G-actin, p21cipl , cyclin A, cyclin B1, cyclin D1 and cyclin E were detected by western blotting （P 〈 0.05 ）. RESULTS Cu Ⅱb inhibited the proliferation of PC-3 cells in a concentration-dependent manner. Cu Ⅱ b 20 μmol＇L-t increased the cell rates of G2/M phase （tetraploid） to （45.3 ± 1.8） % from （27.7 ± 1.5 ） % in control group （ P 〈 0.01 ）, indicating an arrest of cell cycle progression. The cells became shrunk after Cu Ⅱ b treatment. Meanwhile, Cu Ⅱ b 20 μmol· L-1 decreased G-actin levels and induced severe F-actin aggregation （P 〈 0.05 ）, but had minimal effect on the microtubules. In addition, CuⅡb 20 μmol-L-1 also elevated p21cipl expression and downregulated cyclin A level in PC-3 cells, whereas the other cyclins were slightly upregulated （P 〈 0.05 ）. CONCLUSION Cu 11 b can significantly inhibit the proliferation of human prostate cancer PC-3 cells probably through induction of actin aggregation, disruption of microfilaments, upregulation of tumor suppressor p21cipl expression and arrest of cell cycle progression.

关 键 词：雪胆素乙 前列腺癌细胞 肌动蛋白聚集 细胞周期阻滞 

分 类 号：R737.25[医药卫生—肿瘤]