RevTet-Off系统调控凋亡素基因在人肝癌细胞表达的研究  

Study on expression of apoptin gene in human heptoma cells controlled by RevTet-Off gene expression system

作　　者：顾红祥[1] 朱惠明 刘玉杰[1] 黄友明[1] 

机构地区：[1]中山大学附属东华医院消化内科,广东东莞523110 [2]深圳市人民医院消化内科,广东深圳518020

出　　处：《世界肿瘤杂志》2006年第1期38-40,48,共4页Tumour Journal of the World

摘　　要：目的 应用RevTet-Off系统，于体外观察四环素调节下凋亡素基因的表达及其对人肝癌细胞系HepG2的作用。方法 将RevTet-Off基因调控系统的调节质粒pRevTet-Off和含凋亡素基因的重组表达质粒载体pRevTRE-VP3分别转染PT67细胞，包装出RevTet-Off和RevTRE-VP3重组逆转录病毒，依次将此两种病毒感染HepG2细胞，建立整合了RevTet-Off和RevTRE-VP3重组逆转录病毒的阳性细胞株HepG2／Off-VP3。通过四环素调节凋亡素基因在HepG2／Off-VP3中的表达，Annexin V-FITC／PI双染色后，以流式细胞仪检测细胞凋亡率。结果 经RT-PCR证实凋亡素基因在HepG2／Off-VP3-8细胞得到了表达；HepG2细胞在无四环素环境下的细胞凋亡率明显高于1μg／mL四环素环境下的凋亡率。结论 凋亡素基因经RevTet-Off系统导入HepG2细胞后，可在四环素调控下表达产生凋亡素并诱导细胞凋亡。Objective To investigate the effect of apoptin gene expression in vitro on HepG2, a human heptoma cell line in the control of tetracycline with RevTet-Off system. Methods Retroviral regulator vector pRevTet-Off and response vector pRevTRE-VP3 were transfected into packaging cell PT67 respectively. The RevTet-Off virus and recombinant retroviral RevTRE-VP3 were collected and were, in turn, used to infect HepG2 cells. After antibiotic selection, the resulting cell lines HepG2/Off-VP3, which were integrated by RevTet-Off virus and RevTRE-VP3 virus, were established. Apoptin gene expression in HepG2/Off-VP3 cells was controlled by the presence or the absence of tetracycline in medium. After Annexin V-FITC/PI staining, the apoptosis rate in HepG2/Off-VP3 cells was analysed with flow cytometry. Results The expression of apoptin gene in HepG2/Off-VP3-8 was confirmed by RT-PCR. Apoptosis rate in the HepG2/Off-VP3-8 cells incubated in medium free of tetracycline was significant higher than that in the HepG2/Off-VP3-8 cells incubated in medium with 1μg/mL tetracycline. Conclusions Apoptin gene could express apoptin and induce apoptosis in HepG2 cells in the control of tetracycline after apoptin gene is transduced into HepG2 with RevTet system.

关 键 词：凋亡素 四环素调控 逆转录病毒载体 细胞凋亡 肝癌细胞 

分 类 号：R730.21[医药卫生—肿瘤]