乙型肝炎病毒x蛋白通过激活p38和JNK信号通路促进肝癌细胞增殖

Hepatitis B virus x protein promote human hepatocellular carcinoma cells proliferation by activating p38 and JNK signalling pathway

目的探讨乙型肝炎病毒x(HBx)蛋白对人肝癌细胞增殖的分子作用机制。方法构建HBx蛋白的绿色荧光蛋白真核表达载体HBx-pEGFP-C1,利用脂质体转染技术将质粒HBx-pEGFP-C1转染人肝癌细胞系HepG2细胞株,荧光显微镜检测报告基因表达产物EGFP,RT-PCR和Western blot方法检测X基因的表达情况。采用生长曲线和TUNEL染色检测HBx蛋白对HepG2细胞增殖和凋亡的作用。采用Western blot方法检测caspase3、p-p38、p-JNK、p-Akt的表达变化。结果成功构建HBx-pEGFP-C1真核表达载体并转染HepG2细胞,转染HBx蛋白的HepG2细胞与对照组相比增殖率明显升高(t=-0.8999,P=0.012),但细胞凋亡率和caspase3表达水平无明显变化。HBx组p-p38(24 h)(t=-11.058,P=0.0004)、p-JNK(48 h)(t=-15.022,P=0.0001)与对照组相比表达升高明显。结论HBx蛋白通过激活p38和JNK信号通路促进人肝癌细胞HepG2增殖。

Objective To investigate the effects and mechanism of hepatitis B virus x protein(HBx)on human hepatocellular carcinoma cells proliferation.Methods Eukaryotic expression vector HBx-pEGFP-C1 was constructed.HepG2 cells were transfected transiently using Lipofectamine 2000.HBx expression in transfected cells were measured by RT-PCR and Western blot.Cells proliferation and apoptosis were detected by using growth curves and TUNEL staining.The protein levels of caspase-3,p-p38,p-Akt and p-JNK were measured by Western blot.Results HBx was successfully expressed in HepG2 cells.Growth curve result showed that HBx promoted cell proliferation(t=-0.8999,P=0.012).Compared with control group,the levels of p-p38(24 h)(t=-11.058,P=0.0004)and p-JNK(48 h)(t=-15.022,P=0.0001)in HBx-pEGFP-C1 group were increased significantly.There is no significant difference between the two groups′apoptosis.Conclusions Transient overexpression of HBx promoted human hepatic carcinoma cells proliferation and activated the p38 and JNK signalling pathway.