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目的 研究含单纯疱疹病毒Ⅱ型胸苷激酶基因 (HSV ⅡTK)对人原发性肝细胞癌SMMC 772 1的治疗作用。方法 用已构建的真核表达载体 pcDNA3/TK ,通过阳离子脂质体Lipo fectin介导 ,体外转染人原发性肝细胞癌SMMC 772 1。加入GCV ,细胞培养。用RT PCR ,光镜 ,MTT ,流式细胞仪等方法检测HSV ⅡTK在SMMC 772 1细胞中的表达及其对该细胞的杀伤作用。结果 实验组细胞通过RT PCR获得与目的基因 (HSV ⅡTK)大小一致 (10 0 0bp)的特异性条带 ,而对照组细胞则无任何条带出现 ;MTT法检测显示 ,实验组的OD值明显低于对照组 ,且有显著意义(P <0 0 1) ;流式细胞仪检测显示 ,实验组可发现明显的凋亡峰 ,而对照组则无凋亡峰出现。结论 通过阳离子脂质体Lipofectin介导 ,HSV ⅡTK在SMMC 772 1细胞中能够获得表达 ,其表达产物结合GCV可明显抑制人原发性肝细胞癌细胞的增殖。
Objective To investigate the therapeutic effect of herpes simplex virus type 2 thymidine kinase gene (HSV ⅡTK) on human primary hepatocellular carcinoma SMMC 772 1. Methods The constructed eukaryotic expression vector pcDNA3 / TK was transfected into human primary hepatocellular carcinoma SMMC 772 1 by Lipofectin, a cationic liposome. Join GCV, cell culture. The expression of HSV Ⅱ TK in SMMC 772 1 cells was detected by RT PCR, light microscopy, MTT and flow cytometry. Results The specific bands of the target gene (HSV ⅡTK) in the experimental group were identical with those of the target gene (HSV ⅡTK) in the experimental group, while the control group did not show any bands. The MTT assay showed that the OD value of the experimental group was significantly (P <0.01). The results of flow cytometry showed that the apoptotic peak was found in the experimental group and no apoptotic peak appeared in the control group. Conclusion HSV ⅡTK can be expressed in SMMC 772 1 cells mediated by cationic liposomes, and its expression product combined with GCV can significantly inhibit the proliferation of human primary hepatocellular carcinoma cells.