转染TNF-α及其突变体的成纤维细胞的杀瘤效应  

Antitumor effects of the fibroblastes transfected TNF-α gene and its mutants

作　　者：李清芬[1] 李莉[1] 李卓娅[1] 龚非力[1] 冯玮[1] 姜晓丹[1] 徐勇[1] 熊平[1] 

机构地区：[1]华中科技大学同济医学院免疫学系,武汉430030

出　　处：《中国免疫学杂志》2003年第1期21-25,共5页Chinese Journal of Immunology

基　　金：国家自然科学基金重点项目资助 (批准号:392 70 314 )

摘　　要：目的 :比较跨膜型和分泌型TNF α在体内的抗瘤效应。方法 :将 3种TNF α基因 (分泌型TNF α突变体 ,S TN Fm、跨膜型TNF α突变体 ,TM TNFm和野生型TNF α,Wt TNF)通过逆转录病毒载体转染成纤维细胞NIH/3T3 ;并用Southern印迹、RT PCR、流式细胞仪和生物活性检测等方法 ,从基因的整合、RNA转录、蛋白的表达及其生物学活性 4个方面对转染细胞进行检测鉴定。在小鼠接种肿瘤细胞的第 3天 ,于接种部位分别皮下注射表达TNF α及其突变体的NIH3T3细胞 ,效 /靶比为5∶1或 1∶1 ,观察这 3种TNF α的体内抗瘤效应。结果 :NIH3T3转染细胞可表达高水平的TNFα及其突变体。在体外 ,NIH3T3/TM TNFm的固定细胞、NIH3T3/S TNFm的上清、NIH3T3/Wt TNF的固定细胞和上清均可有效杀伤肿瘤细胞H2 2。在体内 ,当效 /靶比为 5∶1时 ,注射NIH3T3/TM TNFm的抑瘤效果最强 ;而当效 /靶比为 1∶1时 ,则NIH3T3/S TNFm的疗效最好。此外 ,在过继治疗期间未见明显副作用。结论 :上述结果提示跨膜型和分泌型TNF α均可在体内有效杀瘤 ;但如效 /靶比适当 ,TM TNF α显示的杀瘤效应较S TNFObjective:To compare the antitumor effects of transmembrane TNF α(TM TNFα)and secreted TNF α(S TNF α) in vivo.Methods:Mouse fibroblasts NIH3T3 were transfected separately with three types of retrovirus containing wild type TNF α (Wt TNF);transmembrane TNF α mutant(TM TNFm); secreted TNF α mutant(S TNFm). The cells were further identified for TNF gene integration, transcription, translation and its biologic activity by Southern blot, RT PCR, FACS and bioassay. On the third day after the H22 tumor cell challenge, fibroblasts NIH3T3 expressing TNF α and its mutants were separately injected into the mice at the sites of tumor cell inoculation according to 5∶1 or 1∶1 effector/target cell ratio, respectively, to observe the antitumor effects of the three types of TNF α in vivo. Results:The transfected NIH/3T3 could express high levers of TNF α and its mutants. It was found that the fixed cell of NIH3T3/TM TNFm, the supematant of NIH3T3/S TNFm, both fixed cell and supernatant of NIH3T3/Wt TNF could effectively kill tumor cell H22 in vitro. At the effector/target cell ratio=5∶l, the injection of NIH3T3/TM TNFm was found to result in the highest tumor regression, while NIH3T3/S TNFm was shown to exert the strongest tumor depressing effect at the effector/target cell ratio=1∶1 in vivo. No obvious side effects were noted throughout the course of treatment. Conclusion:The results suggest that tansmembrane and secreted TNF α could cause tumor regression. The antitumor effect of TM TNF α would be more powerful than that of S TNF α at the proper effector/target cell ratio.

关 键 词：基因治疗 跨膜型TNF-Α 分泌型TNF-Α 逆转录病毒载体 抗瘤效应 

分 类 号：R730.51[医药卫生—肿瘤]