高水平乙型肝炎病毒复制诱导肝细胞QSG-7701凋亡  

High replicated hepatitis B virus induces apoptosis of hepatocytes

作　　者：潘孝本[1] 韩进超[1] 高燕[1] 魏来[1] 

机构地区：[1]北京大学人民医院北京大学肝病研究所,100044

出　　处：《中华医学杂志》2008年第12期840-843,共4页National Medical Journal of China

基　　金：国家自然科学基金资助项目（30700697）;国家高技术研究发展计划（863）基金资助项目（2006AA02A410）

摘　　要：目的观察高水平的HBV复制对QSG-7701细胞可能产生的致病效应。方法采用磷酸钙沉淀方法转染质粒pUC18-HBV1．2（实验组）和空质粒pUC18（对照组）至QSG-7701细胞，细胞计数观察细胞生长曲线。转染后4d，采用荧光实时定量PCR方法检测培养上清中HBV DNA水平；免疫荧光细胞化学染色检测细胞内的HBsAg表达；电子显微镜和末端脱氧核苷酸转移酶介导duTP缺口末端标记法（TUNEL）检测细胞凋亡；Oliga信号传导基因芯片检测实验组和对照组的基因差异表达。结果对照组细胞转染6d后细胞数量增加（8．3±1．2）倍，凋亡细胞极少见。实验组在转染后6d细胞数量仅增加（1．1±0．2）倍，HBsAg阳性细胞35．4％±6．7％，细胞凋亡占15．2％±4．3％。基因差异表达谱分析显示与细胞生长和凋亡相关的部分基因如CASP3（2．7981）、CASP7（2．2643）、3．Apr（3．5013）、CDC2（0．4380）、MAPK6（0．4447）和MAP3K2（0．2785）等表达水平发生显著改变。结论高水平的HBV复制明显抑制QSG-7701细胞生长，并诱导部分细胞发生凋亡。Objective To investigate the effects of high level hepatitis B virus （HBV） replication on the hepatocytes. QSG-7701 cells. Methods Human hepatocytes of the line QSG-7701 were cultured and transfected with the plasmid pUC18-HBV1.2 or pUC18 containing 1.2 full length HBV DNA by the standard calcium phosphate precipitation method. Other QSG-7701 cells were transfected with the plasmid pUC18 as controls. Cell growth curves were drawn for 7 days after transfection. Four 4 days after transfection, HBV DNA in the culture medium was detected by using fluorescence quantitative real-time PCR. Cell apoptosis was detected by using terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling and electronic microscopy. Differential expressed genes were analyzed by using Oliga signal pathway micro-array. Results The curves of cell growth showed that the amount of control QSG-7701 cells increased by （8.3 ± 1.2） times, significantly faster than the pUC18-HBV1. 2 transfected QSG-7701 cells that increased only by （1.1 ±0.2） times （P 〈0.01）. Four days after transfection, the HBsAg positive rate of the pUC18- HBV1.2 transfected cells was 35.4% ±6.7%, and the apoptotic rate was 15.2% ±4.3%. The HBV DNA level in the culture supernatant peaked 4 days adder transfection with the maximum value of （5.8 ± 2.6） × 106 copies/ml. Genes related to cell growth and apoptosis, such as CASP3 （2. 7981 ） ,CASP7 （2. 2643 ）, 3-Apr （3.5013）, CDC2 （0.4380）, MAPK6 （0.4447）, and MAP3K2 （0. 2785）, were differentially expressed. Conclusion High replicated HBV markedly inhibits the growth of hepatocytes and induces cell apoptosis.

关 键 词：肝炎病毒 乙型 肝细胞 凋亡 

分 类 号：R686[医药卫生—骨科学]