索拉非尼对人多发性骨髓瘤细胞RPMI8226增殖与凋亡的影响

Effects of Sorafenib on Proliferation and Apoptosis of Human Multiple Myeloma Cell RPMI 8226

本研究旨在探索索拉非尼对人骨髓瘤细胞RPMI 8226增殖与凋亡的影响及其作用的分子机制。采用MTT法检测索拉非尼对人多发性骨髓瘤细胞的抑制率,透射电子显微镜观察索拉非尼干预骨髓瘤细胞后其超微结构的变化,流式细胞术检测索拉非尼诱导骨髓瘤细胞凋亡的情况及对细胞周期分布的影响,半定量RT-PCR和Western-blot法分别检测索拉非尼作用前后骨髓瘤细胞caspase-3、BCL-2和MCL-1 mRNA和蛋白的表达变化。结果显示,索拉非尼(0-10.0μmol/L)可抑制RPMI 8226细胞增殖,抑制率呈时间-浓度依赖性;48 h后在透射电子显微镜下可见到凋亡小体等典型凋亡改变;索拉非尼可明显诱导骨髓瘤细胞凋亡(P<0.05),主要将细胞阻滞于G1期(P<0.05);索拉非尼作用48 h后,与对照组相比细胞BCL-2及MCL-1 mRNA与蛋白表达量均减少(P<0.05),caspase-3表达量变化不大,而蛋白水平的活化型caspase-3表达增加(P<0.05)。结论:索拉非尼通过诱导细胞凋亡有效抑制骨髓瘤增殖,其机制可能与细胞周期阻滞和激活死亡受体途径有关。

This study was aimed to investigate the effects of sorafenib on proliferation and apoptosis of MM cell line RPMI-8226,and to explore the its potential anti-tumor mechanism.The inhibitory rate of multiple myeloma cell proliferation was tested by MTT.Transmission electron microscopy was used to observe morphological and ultrastructural changes of RPMI-8226 cells treated with sorafenib.The effects of sorafenib on the apoptosis and cell cycle of RPMI-8226 cells was detected by flow cytometry.The effects of sorafenib on the expression of caspase-3、BCL-2 and MCL-1 mRNA and protein were assayed by RT-PCR and Western blot respectively.The results showed that sorafenib （0-10.0 μmol/L） could obviously inhibit the proliferation of RPMI-8226 cells in time and dose-dependent manner.Flow cytometry results showed that sorafenib could induce apoptosis of RPMI-8226 cells,the difference was statistical significance （P 〈 0.05）.Sorafenib mainly arrested RPMI-8226 cells in the G1 phase （P 〈 0.05）.Typical apoptotic morphological and ultrastructural changes of MM cells could be observed under transmission electron microscope,Examination of cellular signaling pathways showed that sorafenib induced upregulation of cleaved-caspase-3 expression,and simultaneous downregulation of BCL-2 and MCL-1 expression.It is concluded that sorafenib displays anti-myeloma activity.Activating the death receptor pathway and arresting cell cycle may be two of the relatated mechanisms.