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目的观察CDglyTK双自杀基因对C6胶质瘤细胞的杀伤作用及旁观者效应。方法利用逆转录病毒介导的胞嘧啶脱氨酶(CD)、单纯疱疹病毒胸苷激酶(HSV-tk)融合基因转染C6胶质瘤细胞,通过细胞集落形成试验、细胞生长抑制率(GIR)测定(MTT法),检测和分析CD/5-氟胞嘧啶(5-FC)、HSV-tk/更昔洛韦(GCV)双自杀基因系统对肿瘤细胞的杀伤作用和旁观者效应。结果RT-PCR检测分析融合基因的表达,显示逆转录病毒介导的双自杀基因在C6胶质瘤细胞中表达。不加5-FC和GCV时,转染组和对照组肿瘤细胞集落形成和倍增时间分别为94h、96h和25.7h、26.6h,组间差异比较无显著意义(P>0.05)。在5-FC(80.0mg/L)和GCV(10-1mg/L)浓度下,GIR分别为83.36%、7.08%,差异比较有显著意义(P<0.01)。转染C6胶质瘤细胞在混育细胞中比例占5%时,即可获得明显的旁观者效应,细胞生长抑制率可达38.48%。结论CDglyTK融合基因联合双前药治疗能取得显著的抗胶质瘤作用。
Objective To observe the killing effect and bystander effect of CDglyTK double suicide gene on C6 glioma cells. Methods C6 glioma cells were transfected with retrovirus-mediated cytosine deaminase (CD) and herpes simplex virus thymidine kinase (HSV-tk) fusion gene. The cell growth inhibition rate (GIR (MTT) assay was used to detect and analyze the cytotoxicity and bystander effect of CD / 5-FC and HSV-tk / ganciclovir (GCV) dual suicide gene on tumor cells. Results The expression of fusion gene was detected by RT-PCR. The result showed that retrovirus-mediated double suicide gene was expressed in C6 glioma cells. In the absence of 5-FC and GCV, the colony formation and doubling time of tumor cells in transfected and control groups were 94h, 96h and 25.7h, 26.6h, respectively. There was no significant difference between the two groups (P> 0.05). The GIRs were 83.36% and 7.08% respectively at the concentrations of 5-FC (80.0 mg / L) and GCV (10-1 mg / L), with significant difference (P <0.01). Transfection of C6 glioma cells in the proportion of mixed cells accounted for 5%, you can get a significant bystander effect, cell growth inhibition rate up to 38.48%. Conclusion CDglyTK fusion gene combined with dual-prodrug treatment can achieve significant anti-glioma effect.