【作者】 张维红； 任秀宝； 于津浦； 李慧； 任宝柱；

【Author】 Zhang Wei-Hong Ren Xiu-Bao Yu Jin-Pu Li Hui Ren Bao-Zhu Department of Immunology, Tianjin Medical University Affiliated Cancer Hospital, Tianjin 300060, China

【机构】 天津医科大学附属肿瘤医院免疫学研究室；

【摘要】 目的:本研究旨在探讨CD40L在CD4+CIK细胞诱导肿瘤细胞凋亡中的生物学作用。方法:构建含有sCD40L 基因的质粒pIRES2-EGFP-sCD40L,大量扩增后转染CHO 细胞,同时用pIRES2-EGFP转染CHO细胞作为空对照。瞬时表达后,应用ELISA法检测其上清中sCD40L的分泌量; 并将转染质粒pIRES2-EGFP-sCD40L的CHO细胞(CHO- sCD40L)与乳腺癌细胞T47D共孵育,同时设置空质粒转染组作为对照,流式细胞法比较24h后T47D表面分子Fas的表达变化;加入Fas激活性抗体(CH-11)后观察24h后T47D细胞的凋亡。结果:成功构建含有sCD40L基因的质粒pIRES2- EGFP-sCD40L;成功转染CHO细胞后,培养上清中sCD40L 的分泌量较空白对照明显增加(P<0．05)。与T47D共培养后, T47D表面分子Fas可被诱导升高,与空白对照有显著性差异 (P<0．05);加入CH-11后24h后可明显观察到实验组T47D 细胞的凋亡。结论:CD40／CD40L的相瓦作用可提升T47D 细胞表面功能性Fas的表达,从而使通过Fas／FasL途径诱导的T47D细胞的凋亡增加。更多还原

【Abstract】 Objective: To investigate the effect of CD40L in apoptosis induced by CD4+CIK. Methods: To construct plasmid carried sCD40L gene pIRES2-EGFP-sCD40L. CHO cells were transfected with plasmid pIRES2-EGFP-sCD40L(CHO-sCD40L) and control plasmid pIRES2-EGFP(CHO-pIRES2) separately. After transfected, sCD40L secreted by CHO-sCD40L was evaluated by ELISA. Then they were cocultured with T47D cells to observe the expression of Fas on T47D.The apoptotic rates of T47D cell ware observed after cocultured with the anti-Fas activating antibody(CH11). Results: We constructed plasmid carried sCD40L successfully. CHO cells was transfected with plasmid pIRES2-EGFP-sCD40L successfully and the transfected rate reached about 20%. SCD40L secreted by CHO-sCD40L increased significantly .After cocultured with CHO-sCD40L, the expression of Fas on T47D increased significantly. The apoptotic rates of T47D cell increased cocultured with the anti-Fas activating antibody(CH-11). Conclusion: The interaction of CD40/ CD40L can increase the expression of functional Fas and induce apoptosis on tumor cell.更多还原