人卵巢癌细胞SKOV3对硼替佐米和紫杉醇联合应用的药物敏感性及可能机制  

The role of GSK-3βin apoptosis induced by proteasome inhibitors combined with paclitaxel in ovarian cancer cells and its possible mechanism

作　　者：翁丹卉[1] 宋晓红[1] 孔繁飞[1] 范良生[1] 栗妍[1] 邢辉[2] 马丁[1] 王世宣[1] 

机构地区：[1]华中科技大学同济医学院附属同济医院妇产科,湖北武汉430030 [2]华中科技大学同济医学院附属襄樊中心医院妇产科,湖北襄樊441021

出　　处：《中国癌症杂志》2008年第6期401-404,共4页China Oncology

基　　金：国家自然科学基金支助项目(No．30571950；No．30528012);国家973重点基础科学基金(No．2002CB513107)。

摘　　要：背景与目的:蛋白酶体抑制剂硼替佐米是新型抗癌药物,体外对多种恶性肿瘤细胞均具有良好的疗效。本实验研究硼替佐米与紫杉醇单独及联合应用对人卵巢癌细胞株SKOV3的存活率和凋亡率的影响,并对糖原合成酶激酶-3β(GSK-3β)和髓样白血病-1基因(Mcl-1)在其诱导细胞凋亡中的作用机制进行探讨,为临床治疗卵巢癌提供理论依据。方法:用50 nmol/L硼替佐米、90 nmol/L紫衫醇,或50 nmol/L硼替佐米联合90 nmol/L几紫衫醇分别作用于SKOV3细胞12、24、36、48及72 h后,四甲基偶氮唑蓝(MTT)比色法测定细胞增殖活性,并计算细胞存活率,Annexin-V/PI法流式细胞仪检测细胞凋亡率,蛋白质免疫印迹法(Western blot)检测相关蛋白表达水平。结果:50 nmol/L硼替佐米和90 nmol/L紫衫醇联合作用于SKOV3细胞,12、24、36、48及72 h时间点细胞存活率分别为(65.2±5.8)%,(58.3±14.4)%,(35.3±5.0)%,(19.2±1.5)%和(11.4±2.5)%,与紫杉醇单用组比较,细胞生长抑制增强,差异有显著性(P<0.05)。药物单独或联合作用细胞24 h后,紫杉醇单用组、硼替佐米单用组和两者联用组细胞凋亡率分别为(14.7±0.5)%、(15.1±0.8)%和(20.5±0.7)%,硼替佐米与紫杉醇联用组凋亡率显著增高(P<0.05)。药物处理细胞后,经免疫印迹法检测p-GSK-3β和Mcl-1蛋白表达水平,硼替佐米与紫杉醇联用组p-GSK-3β和Mcl-1表达水平降低最为明显,分别为正常细胞组表达水平的(19.3±0.4)%和(31.6±3.1)%,差异均有显著性(P<0.05)。硼替佐米和紫杉醇单用组中p-GSK-3β表达水平有所降低,分别为正常细胞组的(78.7±1.2)%和(85.1±1.6)%,Mcl-1表达水平分别为对照组的(68.2±4.5)%和(57.0±4.1)%,差异均具显著性(P<0.05)。结论:硼替佐米与紫杉醇联用能增强人卵巢癌细胞对紫杉醇的药物敏感性,并促进GSK-3β磷酸化Mcl-1,使其降解,诱导细胞,GSK-3β/Mcl-1信号通路可能在硼替佐米与紫杉醇联用诱导凋亡中发挥了重要作用�Background and purpose： Proteasome inhibitors such as bortezomib, represent an interesting new class of potential anticancer drugs. In the present study,we explored the sensitivity of ovarian cancer cell line SKOV3 to paclitaxel, proteasome inhibitors and their combination, and also studied the involvement of GSK-3β/Mcl-1 signaling pathway in the regulation of apoptosis induced by those agent. Methods： Methyl thiazolyl tetrazolium （MTT） assay was applied to examine the cell viability, Annexin-V/PI apoptosis detection kit was used to determine the apoptosis rate of different groups, and western blot assay was introduced to evaluate the expression levels of phosphorylated GSK-3β and Mcl-1. Results： In the MTT assay, the cell viability ratios of combination group at serial time points from 12 to 72 hr were （65.2±5.8）%, （58.3±14.4）%, （35.3±5.0）%, （19.2±1.5）% and （11.4±2.5）%, and there were significant differences as compared to the treatment of paclitaxel alone （P〈0.05）. After drug treatments, apoptosis rates of paclitaxel group, bortezomib group and combination group were （14.7±0.5）%, （15.1±0.8）% and （20.5±0.7）%,respectively. The rate of combination group was much higher than that in the other two groups （P〈0.05）. Western blot assay showed that after the treatment of paclitaxel and bortezomib combination, phosphorylated GSK- 3β and Mcl-1 were significantly down-regulated （P〈0.05）. Conclusions： The cytostatic effect of paclitaxel could be increased significantly by the combination of proteasome inhibitors like bortezomib and paclitaxel. And the GSK- 3β/Mcl-1 signaling pathway may play an important role in the molecular mechanism of apoptosis induced by the combination treatment.

关 键 词：蛋白酶体抑制剂 硼替佐米 糖原合成酶激酶-3Β 紫杉醇 化疗药物敏感性 细胞凋亡 

分 类 号：R73-361[医药卫生—肿瘤]