丙型肝炎病毒基因1b型F蛋白对人肝细胞癌细胞株HepG2凋亡的影响

摘要

目的 研究丙型肝炎病毒(HCV)基因1b型F蛋白对人肝细胞癌细胞株HepG2凋亡的影响.方法 构建真核表达重组质粒pcDNA3.0-F-EGFP,并利用Lipofectamine 2000转染人肝细胞癌细胞系HepG2,同时设pcDNA3.0-C-EGFP-HepG2阳性对照、pcDNA3.0-HcpG2为阴性对照及未转染HepG2为空白对照,以Act-D、TNFα诱导四组细胞系发生凋亡,Annexin V-FITC/PI双染检测肿瘤细胞凋亡率,并使用细胞凋亡DNALadder检测,进一步验证HCV 1b型F蛋白在肝细胞凋亡的生物学功能效应.结果 pcDNA3.0-F-EGFP-HepG2细胞系在发生凋亡的时间上比peDNA3.0-C-EGFP-HepG2快,但相对空白质粒转染组及未转染HepG2细胞系有明显延迟,凋亡的发生率也明显低于阴性对照及空白对照细胞系(P<0.001).结论 外源性基因HCV 1b型F的引入及表达,对HepG2细胞的凋亡有抑制作用.%Objective To investigate the effects of F protein of hepatitis C virus subtype lb on the apoptosis of human hepatocellular carcinom HepG2 cells. Methods HepG2 cells were transfected with recombinant plasmid pcDNA3.0-F-EGFP and pcDNA3.0-F-EGFP-HepG2 strain was exposed to Act-D and tumor necrosis factor a (TNFα) treatment in order to induce cell apoptosis with positive control pcDNA3.0-C-EGFP-HepG2, negative control pcDNA3.0-C-EGFP-HepG2 and blank control HepG2.Annexin V-FITC/PI of flow cytometry was performed to determine the number of apoptotic cells. DNA Ladder was used to observe the isolation of apoptotic DNA fragments in the apoptotie cells. Results pcDNA3.0-F-EGFP- HepG2 cell strain showed a much delayed apoptosis as well as obviously lowering the apoptotic rate when compared with the pcDNA3.0-HepG2 strain and HepG2 strain (P<0.001).Conclusion The introduction and expression of extraneous gene (the F gene of hepatitis C virus subtype 1b) could significantly inhibit the apoptosis of HepG2 cells.