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目的:通过研究乙型肝炎病毒X蛋白(hepatitis B virus X protein,HBx)对人肝细胞株L02细胞增殖、细胞周期以及细胞中糖原合成酶激酶3β(glycogen synthase kinase 3β,GSK3β)表达的影响,探讨乙型肝炎病毒(hepatitis B virus,HBV)相关性原发性肝细胞癌(hepatoccellular carcinoma,HCC)的发生机制。方法:用HBx腺病毒(Ad-HBx)感染人肝细胞株L02细胞后,RT-PCR法检测L02细胞中HBx和GSK3β mRNA表达情况;MTT法检测L02细胞的增殖率变化;FCM法检测细胞周期中各时相所占比例;蛋白质印迹法检测HBx、总GSK3β(total-GSK3β,t-GSK3β)、磷酸化GSK3β(phospho-GSK3β,p-GSK3β)、β-连环素(β-catenin)以及细胞用期蛋白(cyclinD1)等蛋白的表达水平。结果:Ad-HBx感染L02细胞后,HBx的mRNA和蛋白均出现表达,细胞增殖率随着时间的延长而增加;G1期细胞所占比例较对照组减少,S期和G2期细胞比例较对照组增加(P<0.05)。感染Ad-HBx后,t-GSK3β在mRNA和蛋白水平上均无明显变化,而p-GSK3β、β-catenin以及cyclinD1蛋白的表达量增加(P<0.05)。结论:HBx可能通过促进人肝细胞株L02细胞中GSK3β的磷酸化,激活Wnt/β-catenin下游信号通路,从而促进细胞的增殖。
AIM: To investigate the effects of hepatitis B virus X protein (HBx) on the proliferation, cell cycle and the expression of glycogen synthase kinase 3β (GSK3β) in human hepatocellular line L02 cells To investigate the pathogenesis of hepatitis B virus (HBV) -related hepatocellular carcinoma (HCC). Methods: The expression of HBx and GSK3β mRNA in L02 cells was detected by RT-PCR after infected with HBx adenovirus (Ad-HBx). The proliferation rate of L02 cells was detected by MTT assay. The cell cycle , Total GSK3β (t-GSK3β), phospho-GSK3β (p-GSK3β), β-catenin and cell were detected by Western blotting, Expression of proteins such as cyclinD1. Results: The expression of HBx mRNA and protein were all found in L02 cells infected with Ad-HBx, and the cell proliferation rate increased with time; the proportion of cells in G1 phase decreased compared with that in control group, and the proportion of cells in S phase and G2 phase Group increased (P <0.05). After infection with Ad-HBx, t-GSK3β mRNA and protein levels did not change significantly, while p-GSK3β, β-catenin and cyclinD1 protein expression increased (P <0.05). Conclusion: HBx can promote the proliferation of human hepatocyte L02 cells by promoting the phosphorylation of GSK3β and activating the downstream Wnt / β-catenin signaling pathway.