乙肝病毒HBx基因稳转细胞系的建立及其诱导的内质网应激  

Establishment of stable cell line expressing HBx and induction of endoplasmic reticulum stress by HBx

作　　者：陆鹏[1,2] 姜同翠[1,2] 陈露[1,2] 沈玉君[1,2] 沈玉先[1,2] 方圣云[1,2] 

机构地区：[1]安徽医科大学基础医学院,合肥230032 [2]安徽医科大学生物药物研究所,合肥230032

出　　处：《安徽医科大学学报》2014年第3期283-287,共5页Acta Universitatis Medicinalis Anhui

基　　金：国家自然科学基金(编号:81173074;91129729;81372576)

摘　　要：目的建立乙肝病毒(HBV)HBx基因稳转细胞系,并观察该细胞系的增殖活性及内质网应激相关基因的表达情况。方法采用RT-PCR法从HepG2.2.15细胞株中克隆HBx基因编码区片段,构建pcDNA3.1-HBx真核表达质粒。利用脂质体将pcDNA3.1-HBx转染至人HepG2细胞系,通过G418筛选后,经Western blot法对HBx的表达进行验证。并用MTT法检测细胞的增殖活性,同时观察过表达HBx蛋白对内质网应激相关基因表达的影响。结果成功构建了HBx的真核表达质粒;建立了HBV HBx基因稳转细胞系;MTT法检测显示该细胞系增殖活性明显上升;同时也发现HBx基因的过表达可上调内质网应激相关基因BiP、PERK、ATF6、XBP1、MANF的表达,而CHOP的表达降低。结论 HBx基因过表达可诱导内质网应激,并促进HepG2细胞增殖。Objective To establish the cell line stably expressing hepatitis B virus X protein （ HBx） by transfection of HBx to HepG2 cells and to investigate the effect of HBx on ER stress and proliferation. Methods HBx gene en-coding fragment cloned from HepG2 . 2 . 15 cells by RT-PCR was inserted to pcDNA3 . 1 vector to construct the pcD-NA3. 1-HBx eukaryotic expression plasmid and established the stable cell line after G418 screening. Cell prolifera-tion was determined by MTT method. The expressions of UPR genes were detected by PCR method. Results The HepG2s cell line tably expressing HBx was obtained. Overexpression of HBx up-regulated BiP, PERK, ATF6, XBP1, and MANF and down-regulated CHOP. The cell proliferation was enhanced in the HepG2s cells tably ex-pressing HBx. Conclusion Stably expressing HBx induces ER stress by differentially regulating UPR genes and promotes the cell proliferation.

关 键 词：细胞增殖 内质网应激 

分 类 号：R34[医药卫生—基础医学]