【摘要】 目的:探讨环氧合酶-2(COX-2)选择性抑制剂NS-398对放射诱导的人肝癌细胞HepG2凋亡的影响及可能作用机制。方法:应用MTT法检测NS-398对细胞的抑制率;透射电子显微镜观察细胞凋亡的形态学变化,流式细胞术(FCM)定量检测细胞凋亡;实时荧光定量PCR检测凋亡相关基因bcl-2、bax及caspase-3 mRNA表达;Western blotting检测Bc1-2、Bax蛋白表达;比色法分析caspase-3酶活性变化。结果:NS-398对HepG2细胞的生长抑制作用呈时间与剂量依赖性;电镜下处理组细胞呈现典型的凋亡形态学变化,NS-398显著增加放射诱导的细胞凋亡,上调bax mRNA、Bax蛋白及caspase-3 mRNA表达,并增强caspase-3酶活性,而Bcl-2表达无明显变化(P>0.05)。结论:NS-398能增加放射诱导的HepG2细胞凋亡,其机制可能与上调Bax、caspase-3表达,上升Bax/Bcl-2比例,激活线粒体凋亡通路,活化caspase-3,最终诱导细胞凋亡相关。更多还原

【Abstract】 AIM: To investigate the possible role of NS-398,a selective inhibitor of cyclooxygenase-2 enzyme,in radiation-induced apoptosis of human hepatoma cell line HepG2 in vitro.METHODS: Hepatoma cell line HepG2 was treated with various concentrations(25,50,100,200 μmol/L) of NS-398 before MTT assay was used to evaluate the cytotoxicity of NS-398.Transmission electron microscopy(TEM) was used to observe the changes of apoptosis in morphology.FCM was performed to quantify the apoptotic percentage.Real-time PCR was used to detect the expression of bcl-2,bax and caspase-3 mRNA,Western blotting was used to measure the expression of Bcl-2 and bax protein,and colorimetric method was provided to analyze the change of caspase-3 activity.RESULTS: The cytotoxicity of NS-398 increased in time-dependent and dose-dependent manners.NS-398 significantly enhanced radiation-induced apoptosis(P<0.01),increased the expression of bax mRNA,Bax protein,caspase-3 mRNA and enhanced caspase-3 activity,whereas no significant change in Bcl-2 expression was found(P>0.05).CONCLUSION: NS-398 enhances radiation-induced apoptosis in hepatoma cell line HepG2.The mechanism may be associated with the up-regulation of the expression of Bax,caspase-3 and enhancement of the activity of caspase-3,which ultimately induce apoptosis in HepG2.更多还原