组蛋白去乙酰化酶抑制剂MS-275诱导骨髓瘤细胞株U266凋亡过程中Survivin的表达

MS-275, a histone deacetylase inhibitor, induces apoptosis and alters survivin gene expression in human myeloma cell line U266

背景与目的:组蛋白去乙酰化酶抑制剂(histone deacetylase inhibitors,HDACi)是一类具有抗肿瘤活性的新型药物,主要通过诱导细胞凋亡发挥作用。本实验研究HDACi(MS-275)对人骨髓瘤细胞系U266细胞增殖和凋亡的影响及其与Survivin表达的关系。方法:将不同浓度的MS-275作用于U266细胞不同时间后,用台盼蓝拒染法观察药物对细胞活力的影响;通过瑞氏-姬姆萨染色观察药物作用后细胞形态学的变化;用流式细胞仪分析细胞周期;用Western blot检测Survivin、P21和Cdk4等的蛋白表达,以及凋亡信号通路中Caspase-3活化及蛋白聚ADP核糖聚合酶[poly(ADP-ribose)polymerase,PARP]的裂解情况。结果:MS-275呈时间和剂量依赖性抑制U266细胞增殖,阻断细胞周期于G0/G1期。MS-275作用U266细胞48h时的IC50为1.39μmol/L;2μmol/L的MS-275作用U266细胞24h后,G0/G1期细胞占66.39%,36h后G0/G1期细胞占89.80%。瑞氏-姬姆萨染色显示细胞形态发生明显变化。Western blot检测结果显示,MS-275作用U266细胞后,Survivin和Cdk4表达下降,P21表达增加,Caspase3被裂解活化,其底物蛋白PARP发生剪切。结论:MS-275抑制人多发性骨髓瘤细胞系U266增殖并诱导细胞凋亡,可能与下调Survivin蛋白的表达有关。

Background and Objective： Histone deacetylase inhibitors （HDACi） exert antitumor effects through the induction of apoptosis. This study was to investigate the effects of HDACi on survivin gene expression, cell proliferation and apoptosis in human myeloma cell line U266. Methods：U266 cells were exposed to different concentrations of MS-275, a histone deacetylase inhibitor, at different time courses. Cell viability was measured using the trypan blue exclusion assay. Changes in cell morphology were observed with Wright-Giemsa staining. Cell cycle was analyzed using flow cytometry. Protein expressions of poly （ADP-ribose） polymerase （PARP）, caspase-3, survivin, P21 and CDK4 were detected by Western blot. Results： MS-275 inhibited the growth of U266 cells in a dose- and time- dependent manner. After exposure to 1.39 μmol/L MS-275 for 48 h, the cell cycle was arrested at the G0/G1 phase, and the cell viability was decreased by 50%. After the treatment with 2μmol/L MS-275 for 24 h and 36 h, the ratios of U266 cells at the G0/G1 phase were increased to 66.39% and 89.80% , respectively. Obvious changes in morphology of U266 were observed under microscopy. Cleaved PARP appeared, along with an increased protein level of P21 and a decrease of survivin and CDK4 levels in U266 cells treated with MS-275. Conclusion： MS-275 could suppress the proliferation, induce apoptosis and reduce the expression of survivin in human myeloma cell line U266, which may be associated with the downregulation of survivin.