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目的 :观察反转录病毒介导的双自杀基因对 Raji淋巴瘤细胞的杀伤增强作用 ,探讨淋巴瘤的基因治疗方法。方法 :通过脂质体将含有双自杀基因的反转录病毒载体 p WZL neo CDglytk导入病毒包装细胞 PA317,经 G4 18筛选后大量培养产病毒的阳性克隆 PA317/ CD+ tk细胞株 ,收集病毒上清 ,浓缩后转染 Raji淋巴瘤细胞 ,再次经 G4 18筛选 ,获得稳定表达双自杀基因的 Raji/ CD+ tk细胞株。用RT- PCR检测双自杀基因的表达。给予前体药物 5 -氟胞嘧啶 (5 - flourocytosine,5 - FC)和 /或无环鸟苷(Ganciciovir,GCV)后 ,MTT法测定细胞的存活率及 CDglytk双自杀基因对 Raji细胞的杀伤作用。结果 :双自杀基因在 Raji细胞中可稳定表达 ,联合使用 5 - FC和 GCV时 Raji细胞的存活率 (13.83% )明显低于单独使用 GCV (5 0 .6 5 % )或 5 - FC(5 7.6 8% )时 ,各组比差异具有显著性 (P<0 .0 1)。结论 :反转录病毒介导双自杀基因对 Raji淋巴瘤的杀伤作用明显增强
Objective: To observe the effect of retrovirus-mediated double suicide gene on the enhancement of Raji lymphoma cells and to explore the gene therapy of lymphoma. Methods: The retroviral vector pWZL neo-CDglytk containing double suicide gene was introduced into PA317 virus packaging cell by lipofectamine. After selection by G418, the positive clone PA317 / CD + tk cell line was obtained and the virus supernatant . Raji lymphoma cells were transfected into the Raji lymphoma cells. After screening by G418, Raji / CD + tk cells stably expressing double suicide genes were obtained. Double suicide gene expression was detected by RT-PCR. After the prodrug 5 - flourocytosine (5 - FC) and / or guanciciovir (GCV) were given, the viability of cells and the killing effect of CDglytk double suicide gene on Raji cells were determined by MTT assay . Results: The double suicide gene was stably expressed in Raji cells. The survival rate of Raji cells (13.83%) in combination with 5 - FC and GCV was significantly lower than that of Raji cells treated with GCV alone (5 0.56%) or 5 - FC 7.6 8%), the differences of each group were significant (P <0.01). Conclusion: The killing effect of retrovirus-mediated double suicide gene on Raji lymphoma was significantly enhanced