硼替佐米对胰腺癌细胞BxPC3、SW1990生长抑制作用的体外研究

目的 观察硼替佐米对胰腺癌细胞BxPC3、SW1990增殖、凋亡的影响,探讨硼替佐米杀伤癌细胞的可能机制.方法 应用1、10、50、100、500 nmol/L及1、10 μmol/L的硼替佐米干预BxPC3、SW1990细胞,以硼替佐米未干预的细胞作为对照组.采用四甲基偶氮唑蓝（MTT）法检测细胞的增殖;流式细胞仪检测细胞凋亡;RT-PCR法检测Bak、Bax、Bcl-2、Bcl-xl、survivin mRNA表达;蛋白质印迹法检测pro-caspase-3、cleaved-caspase-3、Bax、Bcl-2、surviving蛋白表达.结果 大于50 nmol/L的硼替佐米干预胰腺癌BxPC3、SW1990细胞时,呈浓度及时间依赖性抑制细胞的增殖,其中硼替佐米对BxPC3细胞的生长抑制作用显著大于对SW1990细胞的作用,两者差异有统计学意义（P值均＜0.05）.100 nmol/L硼替佐米干预组BxPC3和SW1990细胞凋亡率分别为（22.56±4.23）％和（（12.71±2.23）％,显著高于对照组的（2.15±0.47）％和（2.32±0.54）％（P值均＜0.05）,且BxPC3细胞的凋亡率显著高于SW1990细胞（P＜0.05）.100 nmol/L硼替佐米干预48 h后BxPC3和SW1990细胞的Bak mRNA表达无显著变化,Bax mRNA及蛋白表达显著增加（P值均＜0.05）,Bcl-2 mRNA和蛋白表达及Bcl-xl mRNA表达均减少（P值均＜0.05）.survivin mRNA和蛋白表达在BxPC3细胞中均减少,而在SW1990细胞中均增加（P值均＜0.05）.2株细胞的pro-caspase-3蛋白表达减少,而cleaved-caspase-3蛋白表达增加（P值均＜0.05）.结论 硼替佐米可抑制胰腺癌细胞BxPC3、SW1990的增殖、诱导凋亡,对BxPC3细胞的作用高于对SW1990细胞,其机制可能与激活线粒体内源性凋亡途径及survivin参与的肿瘤耐药相关.

Effects of bortezomib on growth inhibition of pancreatic cancer cells BxPC3 and SW1990 in vitro

Objective To investigate the effect of bortezomib on proliferation and apoptosis of pancreatic cancer cell lines BxPC3,SW1990 and explore possible mechanisms of bortezomib＇s killing effect on cancer cells.Methods BxPC3,SW1990 cells were treated by using 1,10,50,100,500 nmol/L and 1,10 μmol/L of bortezomib,and cells without bortezomib treatment were considered as control group.The cell proliferation was determined by MTF assay,and apoptosis was determined by flow cytometry.Bak,Bax,Bcl2,Bcl-xl,survivin mRNA expressions were measured by RT-PCR,and Western blot was applied to determine the expressions of pro-caspase-3,cleaved-caspase-3,Bax,Bcl-2,surviving protein.Results When bortezomib concentration was higher than 50 nmol/L,it inhibited the proliferation of two cell lines in a dose and time-dependent manner.And with the same treatment the rate of proliferation inhibition of BxPC3 cells by bortezomib was greater than that of SW1990 cells,and the difference between the two cell lines was statistically significant （P ＜0.05）.Apoptosis rates in the groups of BxPC-3 and SW1990 cells treated by 100 nmol/L bortezomib were （22.56 ± 4.23） ％ and （12.71 ± 2.23） ％,which were significantly higher than those in control group （2.15 ± 0.47） ％ and （2.32 ± 0.54） ％,P ＜ 0.05）.In addition,apoptosis rate of BxPC3 cells was significantly higher than that of SW1990 cells （P＜0.05）.Bak mRNA expression of BxPC3 and SW1990 cells after 100 nmol/L bortezomib treatment were not significantly changed,but the expression of Bax mRNA and protein was significantly increased （P ＜0.05）.Bcl-2 mRNA and protein,as well as Bcl-xl mRNA expressions was significantly decreased （P ＜0.05）.The expression of survivin mRNA and protein in BxPC3 cells were decreased,but were increased in SW1990 cells（P ＜0.05）.The expression of pro-caspase-3 protein in the two cell lines was decreased,while the expression of cleaved-caspase-3 protein was increased （P ＜0.05）.Conclusions Bortezomib can inhibit the pr