乙肝病毒X蛋白对肝星状细胞的增殖及其表达TGFβ1和CTGF的影响  

Effects of Hepatitis B Virus X Protein on Proliferation of Hepatic Stellate Cells and mRNA Expression of TGFβ1 and CTGF

作　　者：陈红英[1] 陈治新[1] 王小众[1] 

机构地区：[1]福建医科大学 附属协和医院消化科,福州350001

出　　处：《福建医科大学学报》2013年第3期127-132,共6页Journal of Fujian Medical University

基　　金：教育部高校博士点专项基金(20093518110003);福建省科技人才基金(2008F3043);福建省卫生厅青年基金立项医院基金(XH200801);福建省临床医学重点专科资助项目

摘　　要：目的研究乙肝病毒X蛋白(HBx)对肝星状细胞(HSC)增殖的影响及其可能的分子机制。方法运用分子生物学方法构建稳定表达HBV X基因的HL-7702肝细胞株(L02/x)。RT-PCR、Western blot鉴定L02/x细胞HBV X基因的稳定表达。将HSC细胞分别与L02/x、转染空质粒和未转染的肝细胞共培养36h,分别命名为HSC/x、HSC/ctr和HSC/NC。CCK-8法检测各组HSC增殖,Real-time PCR法检测在HSC增殖中起重要作用的TGFβ1和CTGF基因在各组HSC中的mRNA表达。结果 RT-PCR和Western blot结果显示,L02/x细胞中有HBV X基因mRNA和蛋白的表达。CCK-8法检测显示,HSC/x的增殖(0.737 7±0.014 2)显著高于HSC/ctr(0.497 0±0.008 5)和HSC/NC(0.491 3±0.018 6),差别具有统计学意义(P<0.01)。Real-time PCR显示,HSC/x细胞中的TGFβ1和CTGF mRNA相对表达量(3.146 1±0.070 5,2.982 9±0.031 5)均显著高于HSC/ctr(1.004 6±0.004 0,1.022 2±0.062 7)和HSC/NC(1.000 0±0.000 0,1.000 0±0.000 0),差别具有统计学意义(P均<0.01)。结论肝细胞中表达的HBV X基因可以上调HSC细胞TGFβ1和CTGF基因的表达,促进共培养的HSC增殖。Objective To investigate the effects of hepatitis B virus X protein (HBx) on the pro-liferation of human hepatic stellate cells (HSCs) and to discuss possible mechanism (s) in the pathway . Methods The eukaryotic expression vector of HBV X gene (p HBV-X-IRES2-EGFP) was transfected in-to human liver cell HL-7702 and selected by G418 to establish a cell line L02/x ,which was confirmed to stably express H BV X gene by RT-PCR and Western blot analysis . HSCs were co-cultured with L02/x cells ,empty vector-transfected cells and non-transfected cells for 36 h ,and were named HSC/x ,HSC/ctr and HSC/NC respectively . Then the proliferation of HSCs in these three groups were detected by CCK8 method and mRNA expressions of TGFβ1 and CTGF in HSCs were analyzed by Real-time PCR . Results RT-PCR and Western blot analysis showed L02/x cells expressed HBV X gene steadily . The prolifera-tions of HSC/x (0 .737 7 ± 0 .014 2)were significantly higher than those of HSC/ctr(0 .497 0 ± 0 .008 5) and HSC/NC(0 .491 3 ± 0 .018 6) ,P<0 .01 . Real-time PCR showed mRNA levels of TGFβ1 and CTGF in HSC/x (3 .146 1 ± 0 .070 5 ,2 .982 9 ± 0 .031 5) were significantly higher than those in HSC/ctr(1 .004 6 ± 0 .004 0 ,1 .022 2 ± 0 .062 7)and HSC/NC (1 .000 0 ± 0 .000 0 ,1 .000 0 ± 0 .000 0) ,both P< 0 .01 . Conclusion The expression of HBV X gene in HL-7702 cells can promote the proliferation of co-cultured HSCs through up-regulating mRNA expressions of TG Fβ1 and CTG F .

关 键 词：病毒蛋白质类 肝炎病毒 乙型 转化生长因子β 肝细胞 星形细胞 即早蛋白质类 

分 类 号：R735.7[医药卫生—肿瘤] R392.11[医药卫生—临床医学]