慢病毒介导的融合基因SP-TAT-Apoptin对HepG2细胞的作用  

The effect of the fused gene of SP-TAT-Apoptin transfected by lentivirus on HepG2 cells

作　　者：韩苏夏[1] 赵晶[1] 马瑾璐[1] 黄辰[2] 吕毅[1] 欧玮[1] 贾茜[1] 

机构地区：[1]西安交通大学医学院第一附属医院肿瘤中心,陕西西安710061 [2]西安交通大学医学院中心实验室,陕西西安710061

出　　处：《细胞与分子免疫学杂志》2010年第4期310-312,共3页Chinese Journal of Cellular and Molecular Immunology

基　　金：国家自然科学基金资助项目(30672069)

摘　　要：目的:将已构建的SP-TAT-Apoptin融合基因真核表达载体包装成感染性慢病毒颗粒,并用病毒颗粒感染肝癌HepG2细胞,测定其诱导凋亡的效率。方法:通过脂质体Li-pofectamine TM2000将SP-TAT-Apoptin融合基因真核表达载体与其他包装质粒共同转导入293FT细胞,包装并收集感染性病毒颗粒,用实时定量PCR法测定病毒滴度,免疫荧光法检测重组慢病毒感染的293FT细胞中SP-TAT-Apoptin融合基因的表达,同时通过流式细胞术(FCM)测定慢病毒感染后HepG2细胞的凋亡率。结果:SP-TAT-Apoptin重组慢病毒感染293FT细胞后,用V5抗原单克隆抗体(mAb)进行免疫荧光化学检测,示SP-TAT-Apoptin融合基因可成功表达于293FT细胞;通过Anexin-VPI法检测示SP-TAT-Apoptin融合基因慢病毒感染肝癌HepG2细胞后可引起细胞凋亡,其凋亡效率明显高于单纯脂质体转染组。结论:成功包装出可表达SP-TAT-Apoptin融合基因且具感染性的慢病毒颗粒,感染HepG2肝癌细胞后可引起其凋亡,为进一步研究融合基因SP-TAT-Apoptin体内治疗效果极其在临床中的应用奠定了基础。AIM：To Package the recombinant lentivirus containing the fused gene of SP-TAT-Apoptin to infect HepG2 cell and measure the efficiency of apoptosis. METHODS：The eukaryotic expression vector of SP-TAT-Apoptin fused gene and other packaging plasmids were transfected into 293FT cells by Lipofectamine^TM2000 reagent. The supernatant of the cultured 293FT cells was harvested and virus titration was determined by real time PCR. The expression of the fused gene of SP-TAT-Apoptin in 293FT cells infected by the recombinant lentivirus was examed by immunofluorescence histochemistry method. At the same time,the apoptosis rates of the HepG2 cells infected by the recombinant lentivirus were determined by using flow cytometer. RESULTS：The 293FT cells infected by the recombinant lentivirus could express the fused protein SP-TAT-Apoptin. Anexin-V PI assay showed that SP-TAT-Apoptin carried by the recombinant lentivirus could cause the HepG2 cell apoptosis,and its apoptosis rate was significantly more than paired control group and SP-TAT-Apoptin carried by liposomes only. CONCLUSION：The recombinant lentivirus of SP-TAT-Apoptin is successfully packaged and it can induce HepG2 cells to apoptosis.

关 键 词：APOPTIN 融合基因 慢病毒 细胞凋亡 

分 类 号：R730.5[医药卫生—肿瘤]