依托泊苷对人骨髓瘤细胞 RPMI8226增殖与凋亡的影响

摘要

Objective:To investigate the effects of Etoposide on human multiple myeloma cell line RPMI8226 and its mechanism.Methods:The proliferation inhibitory rate of RPMI8226 cells was assayed by MTT,the morphological changes of RPMI8226 cells were observed by inverted flurescent microscopy and transmission electron microscopy,the apoptosis rate and the cell cycle distribution of RPMI8226 cells were detected by flow cytometry.The effects of Etopo-side on the expression of Bax,Caspase -3 mRNA were tested by RT -PCR.Bax,Caspase -3 protein expression of RPMI8226 cells was analyzed by Western blot.Results:Etoposide could inhibit the proliferation of RPMI8226 cells in time and dose -dependent manner.Cell number in Etoposide group was significantly less than that in control group under light microscope,and the growth arrangement was irregular,apoptotic cells could be seen.Under electron micro-scope,typical apoptotic morphological and ultrastructural changes could be observed.Flow cytometry results showed that Etoposide could induce apoptosis of RPMI8226 cells,the difference had statistical significance(P <0.05).The Etoposide mainly arrested RPMI8226 cells in the S phase(P <0.05).The expression of Bax,Caspase -3 mRNA in-creased(P <0.05).Western blot indicated that Bax,Caspase -3 protein expression increased.It is concluded that the Etoposide can inhibit the proliferation of RPMI8226 cells by inducing cell apoptosis.Conclusion:Activation of the mi-tochondrial and death receptor pathways of apoptosis may be involved in the Etoposide -induced apoptosis,cell cycle arrest may also play an important role in the apoptosis mechanism.%目的：探讨依托泊苷对人骨髓瘤细胞 RPMI8226的增殖抑制作用及其分子机制。方法：将依托泊苷作用于骨髓瘤细胞，用 MTT 法检测细胞增殖抑制率，光学显微镜和电子显微镜观察细胞形态及超微结构变化，流式细胞术检测细胞凋亡率和细胞周期分布，半定量 RT －PCR 检测 Bax 及 Caspase －3 mRNA 表达量的变化，Western blot 检测 Bax 及 Caspase －3蛋白表达量变化。结果：依托泊苷可抑制 RPMI8226细胞增殖，抑制率呈时间（r ＝0．926）和浓度（r ＝0．938）依赖性增强。24h 后在光学显微镜下可见依托泊苷组细胞数量减少，排列紊乱，细胞形态变得不规则，可见凋亡细胞及坏死细胞。48h 电子显微镜下可见细胞典型凋亡改变，凋亡小体形成。流式细胞术检测结果显示，依托泊苷组 RPMI8226细胞凋亡率明显增高（P ＜0．05）；依托泊苷作用48h 后将 RPMI8226细胞阻滞于 S 期（P ＜0．05）；依托泊苷作用48h，Bax、Caspase －3 mRNA 及蛋白表达量增加（P ＜0．05）。结论：依托泊苷可诱导 RPMI8226细胞凋亡，可能与细胞周期阻滞，激活细胞内、外源性凋亡通路有关。