HCV核心蛋白与NS4B对HepG2细胞增殖的影响

目的 探讨HCV核心蛋白与非结构蛋白4B（NS4B）对HepG2细胞增殖的影响及可能机制.方法 重组质粒pcDNA3.1（-）Core与pcDNA3.1（-）NS4B单独和联合转染HepG2细胞,同时以转染空载体和未转染HepG2细胞作为对照.RT-PCR及Western Blot检测各组细胞中HCVCore、NS4B 、Wnt1、β-catenin 、c-myc及CyclinD1表达；MTT法,平板克隆形成试验检测HCV核心蛋白与NS4B对HepG2细胞增殖的影响；流式细胞术检测细胞周期.结果 ①pcDNA3.1（-）Core与pcDNA3.1（-）NS4B单独和联合转染HepG2细胞,成功表达HCV Core或/（和）NS4B mRNA和蛋白.②pcDNA3.1（-）Core和pcDNA3.1（-）NS4B单独转染和联合转染的HepG2细胞Wnt1、β-catenin、c-myc、CyclinD1 mRNA与蛋白的相对表达量均高于HepG2/pcDNA3.1（-）组和HepG2组（P＜0.01）.③与HepG2/pcDNA3.1（-）组和HepG2组比较,pcDNA3.1（-）Core和pcDNA3.1（-） NS4B单独转染和联合转染的HepG2细胞活力和克隆形成能力增强,S期和G2/M期细胞比例升高（P＜0.01）.结论 HCV核心蛋白与NS4B能加速HepG2细胞周期进程,促进细胞增殖,这种效应可能与其增强Wnt1、β-catenin、c-myc及CyclinD1的表达相关.

Effect of hepatitis C virus core protein and NS4B on the proliferation of HepG2 cells

Objective To investigate the effect of hepatitis C virus （HCV） core protein and nonstructural protein 4B（NS4B） on the proliferation of HepG2 cells and its possible mechanism.Methods The two recombinant plasmid pcDNA3.1 （-）Core and pcDNA3.1 （-） NS4B were transiently transfected respectively and co-transfected into HepG2 cells by lipofectamine, simultaneously HepG2 cells transfected with pcDNA3.1 （-） and untransfected HepG2 cells were used as control.The expression of mRNA and protein of HCV Core,NS4B,Wnt1,β-catenin,c-myc and CyclinDl in the cells of each group were detected by RT-PCR and Western blot respectively.Cell proliferation changes were measured by MTT assay and plate colony formation assay.The cell cycle distribution was tested by flow cytometry.Results ①HCV Core or/and NS4B mRNA and protein were expressed successfully in the HepG2 cells transfected with pcDNA3.1 （-）Core,pcDNA3.1 （-）NS4B alone or in combination.② The relative expression levels of mRNA and protein of Wnt1,β-catenin,c-myc and CyclinD1 were higher in the cells transfected with pcDNA3.1 （-） Core,pcDNA3.1 （-） NS4B alone or in combination than those in the cells transfected with pcDNA3.1 （-） and untransfected HepG2 cells（P 〈0.01）.③Compared with the HepG2 cells transfected with pcDNA3.1 （-）and untransfected,cell viability,cloning efficiency and the percentage of cells at S and G2/M phases were significantly increased in the cells transfected with pcDNA3.1 （-） Core,pcDNA3.1 （-） NS4B alone or in combination （P 〈 0.01）.Conclusion HCV core protein and NS4B can accelerate cell cycle progression and promote cell proliferation of HepG2 cells probably through enhancement of Wntl,β-catenin,c-myc and CyclinD1 expression.