胸苷激酶基因系统杀伤前列腺癌细胞的流式细胞术分析

Flow cytometric analysis of thymidine kinase gene system inducing cell death of prostatic cancer cells

目的 探讨单纯疱疹病毒胸苷激酶/更昔洛韦(HSV-TK/GCV)自杀基因系统诱导人-前列腺癌细胞死亡的机制。方法利用脂质体介导的基因转染技术将携带HSV-TK基因的EB病毒表达载体导入激素非依赖性人前列腺癌(HRPC)细胞系PC-3m并给予GCV,采用流式细胞术(FCM)观察PC-3m细胞周期变化以及细胞坏死和凋亡情况。结果HSV-TK/GCV系统可使PC-3m细胞周期S期比例升高,高浓度GCV组尤其明显,S期细胞比例由16.73％升至33.60％;10和100μmol/L GCV作用72 h后,PC-3m细胞可出现较为明显的细胞坏死和凋亡,比例分别为13.54％、12.45％、34.70％和10.11％。结论细胞坏死和凋亡均参与了HSV-TK/GCV系统诱导的PCa细胞死亡,以细胞坏死比例稍高;TK自杀基因系统杀伤前列腺癌细胞时可能不依赖于其p53表达背景,因而具有更广阔的应用前景。

Objective To investigate the mechanism of HSV-TK/GCV system inducing cell death of prostatic cancer cells. Methods A hormone-independent PCa cell line (PC-3m) was transfected with EBV-based expression vector containing HSV-TK gene by using liposome-mediated gene delivery technique. After administration of the GCV, the cell cycle distribution, apoptosis, and necrosis were analyzed by flow cytometry. Results There was a moderate increase in the relative number of TK-transfect-ed PC-3m cells at S phase of cell cycle following GCV treatment. The S phase blockage was more significant in the high dose group, in which the percents of the cells at S phase were elevated from 16. 73% to 33.60 % . Furthermore, analysis of apoptosis and necrosis displayed that 72 h administration of GCV triggered a rather high ratio of apoptosis and necrosis of TK gene-transduced PC-3m cells, and the magnitudes of those were 13. 54%, 12. 45 % , 34. 70 % and 10.11% respectively. Conclusion Both necrosis and apoptosis were involved in the cellular death of PC-3m cells induced by HSV-TK/GCV system. It seems that necrosis is of essential importance during the procedure. It is indicated that the killing effect of TK suicide gene approach on prostate cancer cells may independent of the background of p53 expression.