摘要
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.
作者
娄晓丽
侯彦强
Lou Xiaoli;Hou Yanqiang(Department of Central Laboratory,Shanghai Songjiang District Central Hospital,Shanghai 201600,China)
出处
《中华实验和临床病毒学杂志》
CAS
CSCD
2019年第4期394-399,共6页
Chinese Journal of Experimental and Clinical Virology
基金
国家自然科学基金青年项目(81702729)。