组蛋白去乙酰化酶抑制剂联合吉西他滨对骨肉瘤MG-63细胞抑制作用的研究  

Experimental study of MGCD0103 in combination with gemcitabine inhibiting the growth of human osteosarcoma MG-63 cells in vitro

作　　者：胡旭昌[1] 王栓科[1] 康学文[1] 汪静[1] 安丽萍[1] 夏亚一[1] 狄天宁[1] 马靖琳[1] 王翠芳[1] 胡杰亮[1] 

机构地区：[1]兰州大学第二医院骨科,甘肃省骨关节疾病研究重点实验室,甘肃兰州730030

出　　处：《肿瘤》2013年第6期520-525,共6页Tumor

基　　金：甘肃省自然科学基金资助项目(编号:2009GS03170)

摘　　要：目的:研究组蛋白去乙酰化酶(histone deacetylase,HDAC)抑制剂MGCD0103和吉西他滨(gemcitabine)单药或联合作用对体外培养的人骨肉瘤细胞株MG-63生长的影响,并探讨其可能的作用机制。方法:体外传代培养人骨肉瘤MG-63细胞,将不同浓度的MGCD0103(1、10和100μmol/L)和吉西他滨(0.1、0.2和0.4mg/L)分别单独或联合作用于MG-63细胞;采用MTT法检测2种药物单药或联合作用对MG-63细胞的生长抑制作用,并观察其有效的作用浓度;在荧光显微镜下观察细胞的凋亡情况;实时荧光定量-PCR(real-time fluorogenic quantitative-PCR,RFQ-PCR)检测MG-63细胞中转移相关基因1(metastasis-associated gene1,MTA1)mRNA表达的变化,蛋白质印迹法检测p21、Bax及血管内皮生长因子(vascular endothelial growth factor,VEGF)表达水平的变化。结果:MGCD0103和吉西他滨单药或联合作用都能明显抑制MG-63细胞的生长(P<0.05),并呈剂量依赖性,MGCD0103和吉西他滨具有协同作用。荧光显微镜下观察结果显示,MGCD0103和吉西他滨单药或联合作用于MG-63细胞后,大量具有凋亡形态特征的细胞出现,凋亡细胞数明显增多;RFQ-PCR和蛋白质印迹法检测结果分别显示,MTA1 mRNA及VEGF蛋白的表达量明显下调,而p21和Bax蛋白的表达量明显上调。结论:MGCD0103和吉西他滨通过诱导细胞凋亡及抑制细胞增殖而发挥体外抗骨肉瘤作用,其作用机制涉及细胞周期和凋亡相关基因表达的调控。MGCD0103和吉西他滨具有协同作用,有望成为治疗骨肉瘤治疗的一种新手段。To investigate the inhibitory effect of HDAC （histone deacetylase） inhibitor MGCD0103 and gemcitabine on the proliferation of human osteosarcoma MG-63 cells in vitro, and to explore its possible mechanism. Methods： The cell proliferation inhibitory rates of MG-63 cells treated with different concentrations of MGCD0103 （1, 10 and 100 ~mol/L） or gemcitabine （0.1, 0.2 and 0.4 mg/L） alone and in combination were detected by MTT assay. The change of the apoptosis of MG-63 cells was observed by fluorescence microscope. The expression level of MTA1 （metastasis-associated gene 1） mRNA after MGCD0103 and gemcitabine treatment was measured by RFQ-PCR （real-time fluorogenic quantitative-PCR）. The expressions levels of Bax, p21 and VEGF （vascular endothelial growth factor） proteins were detected by Western blotting. Results： Different concentrations of MGCD0103 or gemcitabine alone or in combination could obviously inhibit the proliferation of MG-63 cells in a dose- dependent manner （P 〈 0.05）. MGCD0103 in combination with gemcitabine had a synergistic effect. After MG-63 cells were treated with MGCD0103 and gemcitabine alone or in combination, the apoptotic cells presenting typical morphologic change could be observed, and the apoptotic ratio was also increased. The expression levels of MTA1 mRNA and VEGF protein were down-regulated, but the expression levels of Bax and p21 were up-regulated. Conclusion： MGCD0103 and gemcitabine have antitumor effects on osteosarcoma in vitro through inducing apoptosis and inhibiting the cellular proliferation, which might be related to the regulation of the expressions of genes involved. MGCDO103 and gemcitabine may be used as new antitumor agents for treatment of osteosarcoma.

关 键 词：骨肉瘤 组蛋白脱乙酰化酶类 吉西他滨 抗肿瘤联合化疗方案 MG-63细胞 

分 类 号：R733.3[医药卫生—肿瘤]