米托蒽醌对人骨髓瘤细胞RPMI8226增殖与凋亡的影响

本研究探讨米托蒽醌对人骨髓瘤细胞RPMI8226的增殖抑制作用及其分子机制。将米托蒽醌作用于骨髓瘤细胞,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BCL-2、BAX及caspase-3 mRNA表达量的变化,Western blot检测BCL-2、BAX和caspase-3 mRNA蛋白表达量变化。结果表明,米托蒽醌可抑制RPMI8226细胞增殖,抑制率呈时间（r=0.924）和浓度（r=0.947）依赖性增强。米托蒽醌作用24 h后在光学显微镜下可见米托蒽醌组细胞数量减少,排列紊乱,细胞形态变得不规则,可见凋亡细胞;在48 h后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,在米托蒽醌组RPMI8226细胞凋亡率明显增高（P〈0.05）;米托蒽醌作用48 h后,在低浓度1.0μg/ml组RPMI8226细胞阻滞于G2/M期（P〈0.05）,在高浓度2.0μg/ml组RPM I8226细胞阻滞于S期（P〈0.05）;米托蒽醌作用48 h,BCL-2 mRNA与BCL-2蛋白表达量减少（P〈0.05）,BAX、caspase-3mRNA及蛋白表达量增加（P〈0.05）。结论：米托蒽醌可诱导RPM I8226细胞凋亡,此细胞凋亡可能与细胞周期阻滞,激活细胞内、外源性凋亡的通路有关。

Effect of Mitoxantrone on Proliferation and Apoptosis of Human Multiple Myeloma Cell RPMI8226

This study was aimed to investigate the effects of mitoxantrone on proliferation and apoptosis of human multiple myeloma cell line RPMI-8226 and its mechanism.The inhibitory rate of RPMI8226 cells proliferation was assayed by MTTthe morphological changes of RPMI-8226 cells were observed by inverted flurescent microscopy and transmission electron microscopythe apoptosis rate and the cell cycle distribution of RPMI-8226 cells were detected by flow cytometry.The effects of mitoxantrone on the expression of BCL-2BAXcaspase-3 mRNA were detected by RTPCRthe BCL-2BAXcaspase-3 protein expression of RPMI-8226 cells was analyzed by Western blot.The results showed that mitoxantrone could inhibit the proliferation of RPMI-8226 cells in time-and dose-dependent manners.Light microscopy showed that the cell number in mitoxantrone group was significantly less than that in control group and the cell growth arrangement was irregularapoptotic cells could be seen.Under electron microscopetypical apoptotic morphological and ultrastructural changes could be observedthese results confirmed that the mitoxantrone could induce apoptosis of RPMI-8226 cellsthe difference have statistical significance（P〈0.05）.The 1.0 μ/ml low concentration of mitoxantrone mainly arrested RPMI-8226 cells in the G2 /M phase（P〈0.05）,and the 2.0 μ/ml high concentration of mitoxantrone mainly arrested RPMI-8226 cells in the S phase（P〈0.05）.The expression of BCL-2 mRNA decreased（ P〈0.05）,while the expression of BAXcaspase-3 mRNA increased（ P〈0.05）.Western blot indicated that BCL-2protein expression also decreased（P〈0.05） and BAXcaspase-3 protein expression increased.It is concluded that the mitoxantrone can inhibit the proliferation of RPMI-8226 cells by inducing cell apoptosis.Activation of the mitochondrial and death receptor pathways of apoptosis may be involved in the mitoxantrone-induced apoptosisthe cell cycle arrest may also play an important role in the apoptosis mechanism.