放射诱导启动子介导CDglyTK基因治疗Tca8113细胞的实验研究

目的 观察人工放射诱导启动子介导CDglyTK双自杀融合基因治疗Tca8113细胞的疗效，为舌鳞癌治疗探索新的途径。方法 构建人工放射诱导启动子调控双融合自杀基因的真核表达质粒pcDNA3.1（+）/E-CDglyTK，用脂质体为载体转染Tca8113细胞后予以3 Gy诱导放疗，描绘细胞生长曲线，RT-PCR检测CDglyTK的表达，流式细胞仪检测细胞的凋亡和增殖。结果 诱导放疗可显著增强双自杀基因CDglyTK对Tca8113细胞的毒性作用；RT-PCR半定量分析诱导放疗增强了CDglyTK基因的mRNA表达；流式细胞检测发现治疗组细胞的凋亡指数明显高于对照组，而增殖指数则低于对照组，诱导放疗显著提升了这种差距。结论 人工合成放射诱导启动子可作为基因治疗中的分子开关调节CDglyTK基因在Tca8113细胞中靶向表达。低剂量放射性照射可显著提高放射诱导启动子调控的CDglyTK基因治疗Tca8113细胞的疗效。

Synthetic Radiation-inducible Promoter Mediated CDglyTK Gene in Treatment of Tca8113 Cells

Objective To observe the therapeutic effect of CDglyTK gene mediated by synthetic radiation-inducible promoter in the treatment of Tca8113 cells.Methods CDglyTK gene in pCEA-CDglyTK was subcloned into pcDNA3.1( ) to construct plasmid pcDNA3.1( )-CDglyTK,and then the synthetic radiation-inducible promoter in pMD18-T-E was inserted into pcDNA3.1( )-CDglyTK to construct plasmid pcDNA3.1( )/E-CDglyTK.The recombinant plasmid was transfected into Tca8113 cells by lipofectamine,and then exposed to 3 Gy irradiation.Cytotoxicity was evaluated by MTT.The expression of CDglyTK gene was detected by RT-PCR.The apoptosis and proliferation were examined by flow cytomtery.Results The plasmid pcDNA3.1( )/E-CDglyTK was constructed successfully.The comparative survival rate of Tca8113 cells was markedly decreased by induction irradiation.Up-regulation of CDglyTK expression was found in Tca8113 cells exposed to irradiation.The apoptosis index(AI) of Tca8113 cells exposed to irradiation was higher than that of Tca8113 cells without irradiation,the other way round,the proliferation index(PI) of Tca8113 cells exposed to irradiation was lower than that of Tca8113 cells without irradiation.Conclusion The synthetic radiation-inducible promoter can be served as a molecular switch to improve the expression of CDglyTK gene in Tca8113 cells,and low dose induction radiation can significantly improve the therapeutic efficiency.