【摘要】 目的探讨HBx蛋白对人肝细胞发生侵袭恶性表型的影响和作用机制。方法利用固相pH梯度双向凝胶电泳分离稳定表达HBx基因的CCL13-HBx细胞系及转染空质粒的CCL13-pcDNA3.1细胞系的总蛋白质,凝胶经蓝银显色后,采用Image Master 2-DE Elite 4.01图像分析软件进行比较分析、识别差异表达的蛋白质,MALDI-TOF-MS质谱仪得到相应的肽质指纹图谱后,搜索数据库鉴定部分差异蛋白质点。结果获得了背景清晰、分辨率高、重复性好的CCL13-HBx和CCL13-pcDNA3.1细胞总蛋白,图像分析识别差异蛋白质点共26个,通过质谱分析鉴定出14个非冗余的与细胞代谢、细胞周期及信号转导相关的差异表达蛋白质。结论建立了CCL13-HBx,CCL13-pcDNA3.1细胞系的2-DE图谱,鉴定了14个HBx诱发肝癌发生的相关蛋白质,为在蛋白质水平阐明HBx的诱发肝癌发生的机制提供了新线索。更多还原

【Abstract】 Objective To explore the effect of hepatitis B virus X protein on malignant transformation of hepatocytes and its mechanism.Methods The total protein content of CCL13-HBx and CCL13-pcDNA3.1 cells was extracted and separated by two-dimensional electrophoresis with immobilized pH gradients(2-DE).The silver-stained 2-DE was scanned with digital image scanner and analyzed with Image Master 2-DE Elite 4.01 software.To obtain peptide mass fingerprint(PMF) of differential protein spots,matrix assisted laser desorption/ionization time-of-flight mass spectrometry(MALDI-TOF-MS) was used.PMF was searched in SWISS-PROT database by Mascot software to identify differential expression proteins.Results The clear background,well-resolved and reproducible 2-DE maps were obtained.from CCL13-HBx and CCL13-pcDNA3.1 cells.Twenty-six protein spots were found as differentially expressed proteins,of which 14 protein spots were identified successfully.The identified proteins were correlated with cell metabolism,cell cycle and signal transduction.Conclusions The well-resolved,reproducible 2-DE maps of CCL13-HBx and CCL13 cells have been established and 14 differentially expressed proteins are identified.They provide new clues for the study of hepatocarcinogenesis at protein level.更多还原