环氧合酶抑制剂NS-398增强放射诱导的肝癌细胞凋亡

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-3mRNA 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。

In vitro enhancement of radiation-induced apoptosis in human hepatoma cell line HepG2 by a selective inhibitor of cyclooxygenase-2 enzyme NS-398

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.