AFP增强子驱动的HSV-TK/GCV自杀基因系统对肝癌细胞杀伤的实验研究

Expression of AFP Enhancer Driven HSV-TK/GCV Suicide Gene in Hepatocellular Carcinoma Cells

目的探讨人AFP增强子驱动的单纯疱疹病毒胸苷激酶/丙氧鸟苷(HSV-TK/GCV)自杀基因系统体外靶向杀伤肝癌细胞效应。方法构建人AFP增强子驱动的pAFP-CDNA3.1-TK自杀基因真核表达质粒,脂质体转染肝癌细胞,检测TK mRNA和蛋白表达,MTT法检测GCV对肝癌细胞的杀伤作用。结果成功构建pAFP-CDNA3.1-TK自杀基因真核表达质粒,在AFP阳性HepG2细胞中检测到TK mRNA和蛋白表达,添加GCV可特异性地杀伤HepG2细胞,而AFP阴性的SMMC7721细胞生长不受影响。结论 AFP增强子驱动的TK/GCV自杀基因系统可以靶向杀伤AFP阳性肝癌细胞。

Objective To investigate the effect of expression of AFP driven thymidine kinase suicide gene on the in vitro growth of hepatocellular carcinoma cells. Methods To construct hepatocellular carcinoma-specific eukaryotic expression vector carrying a suicide gene driven by AFP enhancer. HepG2 （AFP positive） and SMMC7721 （AFP negative） hepatocellular carcinoma cells were transfected with the plasmid using liposome. Expression of mRNA and protein was detected by RT-PCR and Western blot,respectively. Cytotoxicity of ganciclovir （GCV） on the transfected cells was determined by MTT method. Results Selectively expression of the suicide gene in AFP positive HepG2 cells was verified by RT-PCR and Western blot. GCV dose-dependently inhibited the proliferation and growth of AFP positive HepG2 cells. GCV had no effect on the AFP negative SMMC7721 cells. Conclusion AFP enhancer driven suicide gene transfected hepatocellular carcinoma cells are compromised with GCV-mediated cytotoxicity.