CDglyTK双自杀基因杀伤C6胶质瘤细胞的体内实验研究

目的 观察CDgtyTK双自杀基因对C6实体胶质瘤生长的抑制作用. 方法 利用逆转录病毒介导的大肠杆菌胞嘧啶脱氨酶(CD)和单纯疱疹病毒胸苷激酶(HSV-TK)融合基因转染小鼠C6实体胶质瘤,RT-PCR检测分析融合基因的表达,并观察对照组和治疗组肿瘤体积、重量、抑瘤牢及生存期的变化,同时观察其对C6胶质瘤细胞凋亡的影响,分析CDglyTK双自杀基因系统对肿瘤的抑制作用. 结果 RT-PCR检测显示逆转录病毒介导的COgtyTK双自杀基因在C6胶质瘤中有表达.对照组第4周肿瘤已长至20 mmx30 mm大小.并出现小鼠死亡现象,至第6周末全部死亡;治疗组肿瘤体积未增长,约5 mm大小.部分肿瘤消失.肉眼观察可见对照组肿瘤体积大,色红,血供丰富,治疗组肿瘤体积减小或消失.21 d后处死取瘤称重,对照组[(2.51±0.58)g]与治疗组肿瘤质量[(0.35±0.26)g]比较差异有统计学意义(P<0.05),治疗组抑瘤牢为86.1%.流式细胞仪检测可见治疗组细胞凋亡率(34.41%±5.20%)明显高于对照组(2.92%±1.30%),差异有统计学意义(P<0.05).电镜观察可见治疗组肿瘤细胞凋亡小体形成. 结论 效 CDglyTK双自杀基因联合双前药治疗能取得显著的抗胶质瘤作用.

In vivo antitumor effect of CDglyTK double suicide gene on C6 glioma cells

Objective To observe the inhibitory effect of CDglyTK double suicide gene on the growth of C6 glioma in vivo. Methods Thirty 4- to 6-week-old male Balb/c nude mice with subcutaneous injection of C6 glioma cells were randomized into two groups for injections with 5x105 CFU of CDglyTK (treatment group) or 50 μL PBS (control group) for 5 consecutive days. RT-PCR was performed to examine the mRNA expression of the CDglyTK fusion gene in the tumor cells. All the mice received intraperitoneal injections of 5-FC (500 mg/kg) and GCV (60 mg/kg) for 7 days, and the tumor volume, weight and mice survival time were observed. The effect of CDglyTK gene transfer on the apoptosis of the C6 glioma cells was evaluated using flow cytometry. Results RT-PCR demonstrated effective expression of the fusion gene in C6 glioma cells. In the fourth week of tumor cell inoculation, the tumor in the control group grew to the volume of 20 mmx30 mm with occasional death of the mice, and in week 6, all the control mice died. In the treatment group, the tumors showed no obvious growth with a volume of around 5 mm, and some of tumors even disappeared; no death of the mice occurred at the end of the third month. By gross observation, the tumors in the control mice were large and red with rich blood supply, but those in the treatment group showed reduced volume. On day 21 following tumor cell inoculation, the weight of the tumors was significantly greater in the control group than in the treatment group (2.51±0.58 vs 0.35±0.26 g, P<0.05), and the growth inhibition rate was 86.1% in the latter group. Flow cytometry showed significantly higher apoptotic rate of the tumor cells in the treatment group than in the control group (34.41%±5.2% vs 2.92%±1.3%, P<0.05), and electron microscope demonstrated the formation of apoptotic bodies in the tumor cells in the former group. Conclusion Significant antitumor effects can be obtained with the CKglyTK fusion gene in combination with the two prodrugs.