乙肝病毒x蛋白通过Wnt／β-catenin信号通路调控Gankyrin表达  

Regulation on expression of Gankyrin by hepatitis B virus X protein through Wnt/β-catenin signaling

作　　者：刘锐[1] 魏续福[1] 沈艾[1] 吴忠均[1] 

机构地区：[1]重庆医科大学附属第一医院肝胆外科,重庆400016

出　　处：《重庆医科大学学报》2014年第7期923-928,共6页Journal of Chongqing Medical University

基　　金：国家自然科学基金资助项目(编号:81171562)

摘　　要：目的:研究乙肝病毒X蛋白(hepatitis B virus X protein,HBX)对Gankyrin表达水平的影响以及Wnt/β-catenin信号通路在HBX基因调控Gankyrin表达中的作用。方法:HBX腺病毒表达载体(Ad-HBX-GFP)感染转染2种人肝癌细胞系HepG2、SMMC-7721细胞,Si-HBX分子(Si-HBX oligo)用LipofectamineTM 2000转染入HepG2.2.15细胞,用RT-PCR和Western blot法检测HBX与Gankyrin的表达情况;β-catenin腺病毒载体转染HepG2、SMMC-7721细胞,用Western blot法检测β-catenin与Gankyrin的表达情况;在沉默β-catenin表达的HepG2、SMMC-7721细胞中过表达HBX基因,Western blot法检测β-catenin与Gankyrin的表达情况。结果:①HepG2.2.15细胞中Gankyrin mRNA与蛋白表达水平均高于HepG2细胞(P<0.05)。②HepG2与SMMC-7721细胞系中Ad-HBX-GFP组Gankyrin mRNA与蛋白表达水平均高于绿色荧光蛋白腺病毒表达载体(Ad-GFP)组(P<0.01)。③HepG2.2.15细胞中Si-HBX组Gankyrin mRNA与蛋白表达水平均低于Si-阴性对照(Si-NC)组(P<0.01)。④在HepG2与SMMC-7721细胞系中过表达HBX后,Ad-HBX-GFP组β-catenin的蛋白表达水平均高于Ad-GFP组(P<0.01)。⑤在HepG2与SMMC-7721细胞系中过表达β-catenin后,β-catenin腺病毒表达载体(Ad-β-catenin-GFP)组Gankyrin的蛋白表达水平均高于Ad-GFP组(P<0.01)。⑥在HepG2与SMMC-7721细胞系中沉默β-catenin后,过表达HBX基因,Si-β-catenin腺病毒表达载体(Ad-si-β-catenin-RFP)组与红色荧光蛋白腺病毒表达载体(Ad-RFP)组相比,Gankyrin的蛋白表达水平未见升高(P<0.01)。结论:HBX-Wnt/β-catenin-Gankyrin信号转导通路存在与肝癌细胞中,抑制β-catenin的功能可阻遏HBX对Gankyrin的上调作用,本结果将可能为肝癌的治疗提供新的思路。Objectlve：To investigate the effects of HBX on Gankyrin expression and role of Wnt/β-catenin signaling pathway in the regulation of HBX on Gankyrin. Methods： HBX adenovirus vector Ad-HBX-GFP was transfected into human hepatocellular carcinoma cell line HepG2, SMMC-7721 cells and Si-HBX molecules was transfected into HepG2 2.15 ceils with LipofectamineTM 2000. Expres- sions of HBX and Gankyrin were detected by RT-PCR and Western blot. β-catenin adenovirus vector was transfected into HepG2 and SMMC-7721 cells, and expressions of β-catenin and Gankyrin were detected by Western blot. HBX was overexpressed in β-catenin silenced HepG2 and SMMC-7721 cells,and expressions of β-catenin and Gankyrin were detected by Western blot. Results：①Expres- sions of Gankyrin were higher in HepG2.2.15 cells than in HepG2 cells（P〈0.05）. ②mRNA and protein expressions of Gankyrin were increased significantly in Ad-HBX-GFP group than in Ad-GFP group in HepG2 and SMMC-7721 cell line（P〈0.01）. ③mRNA and protein expressions of Gankyrin were decreased significantly in Si-HBX group than in Si-NC group in HepG2.2.15 cells（P〈0.05）. ④Protein expressions of β-catenin were increased significantly in Ad-HBX-GFP group than in Ad-GFP group after overexpressing HBX protein in HepG2 and SMMC-7721 cell line（P〈0.01）. ⑤Protein expressions of Gankyrin were increased significantly in Ad- HBX-GFP group than in Ad-GFP group after overexpressing β-cateninin HepG2 and SMMC-7721 cell line（P〈0.01）. ⑥Protein ex- pressions of Gankyrin were not increased in Ad-si-β-catenin-RFP group than in Ad-RFP group after silencing β-catenin in HepG2 and SMMC-7721 cell line and overexpressing HBX gene（P〈0.01 ）. Conclusion：HBX-Wnt/β-catenin-Gankyrin signal transduc-tion pathways are existed in hepatocarcinogenesis. Inhibiting the function of β-catenin can interrupt the up-regulation of Gankyrin by HBX,which may provide a new treatment method for hepa- tocellular carcinoma.

关 键 词：肝癌 乙肝病毒X蛋白 Gankyrin Β-CATENIN 

分 类 号：R735.7[医药卫生—肿瘤]