乙型肝炎病毒X基因促TRAIL诱导Huh7细胞凋亡的机制

目的探讨乙型肝炎病毒(hepatitis B virus,HBV)X基因(HBX)促肿瘤坏死因子相关凋亡诱导配体(TNF relatedapoptosis inducing ligand,TRAIL)蛋白诱导人肝癌细胞Huh7凋亡的机制。方法应用脂质体转染法将pcDNA3.1-HBX、pcDNA3.1分别转染Huh7细胞,建立稳定表达HBX的细胞模型Huh7-HBX和空载体对照细胞模型Huh7-3.1;运用TRAIL蛋白分别作用Huh7-HBX细胞、Huh7-3.1及Huh7细胞后,采用CCK8、流式细胞术检测细胞增殖和凋亡情况;Western blot检测死亡受体4(deathreceptor 4,DR4)和死亡受体5(death receptor 5,DR5)的表达情况;分光光度法检测细胞caspase8和caspase3活性。结果 RT-PCR及Western blot证实成功构建细胞株Huh7-HBX;CCK8和流式细胞术检测结果均显示Huh7-HBX细胞的凋亡率明显高于Huh7-3.1及Huh7(P<0.05);Western blot显示Huh7-HBX细胞的DR4、DR5的表达量明显高于Huh7-3.1及Huh7细胞(P<0.05);caspase活性检测结果显示,Huh7-HBX细胞caspase8及caspase3的活性明显高于Huh7-3.1及Huh7(P<0.05)。结论 HBX能够通过上调Huh-7细胞表面死亡受体DR4、DR5的表达,进而促进TRAIL蛋白诱导的细胞凋亡,为进一步研究HBX的促凋亡机制提供实验依据。

Hepatitis B virus X gene enhances TRAIL-induced apoptosis in Huh7 cells

As is known to all,HBX can induce apoptosis of HepG2 cells by promoting the expression of FasL.In this study,we aimed to explore the effects of HBX on TRAIL-induced apoptosis of Huh7 human hepatoma cell line.Firstly,Huh7-HBX cell was established through lipid-mediated transfection.Then the cells proliferation and apoptosis were detected through CCK8 and FCM.Western blot was used to detect the expressions of death receptor 4 and death receptor 5.We also detected the activities of caspase8 and caspase3.We found that the apoptosis ratio of Huh7-HBX cell was higher than that of Huh7-3.1 and Huh7 cells;death receptor 4 and death receptor 5 on the surface of the Huh7-HBX cell were richer than that on Huh7-3.1 and Huh7 cells;the activities of caspase8 and caspase3 in Huh7-HBX cell were higher than that of Huh7-3.1 and Huh7 cells.Thus,we arrived at a conclusion that HBX can enhance TRAIL-induced apoptosis in Huh7 cells by promoting the expression of death receptor 4 and death receptor 5.