雷帕霉素对人CD4^+CD25^+调节性T细胞体外扩增的影响  

Effect of rapamycin on the expansion of human CD4^+CD25^+ regulatory T cells in vitro

作　　者：郝俊[1,2] 曹国番[1,2] 杨洁[2] 杨懿铭[2] 李振华[1,2] 杨燕[2,3] 范华骅[2] 

机构地区：[1]华东师范大学生命科学学院生物医学系(上海市血液中心联合培养),上海200062 [2]上海市血液中心血液工程研究室,上海200052 [3]复旦大学附属华山医院临床检验诊断系,上海200032

出　　处：《现代免疫学》2012年第5期404-411,共8页Current Immunology

基　　金：上海市科委项目(11140902902)

摘　　要：探讨有效的体外扩增人CD4^+CD25^+调节性T细胞(Treg)的方法以及雷帕霉素(rapamycin,RAPA)对人CD4^+CD25^+Treg细胞体外扩增的影响。采用免疫磁珠分离人外周血的CD4^+CD25^+Treg,并将实验分为3组:第1组在CD4^+CD25^+Treg培养体系中加入200 U/ml IL-2和包被CD3/CD28单克隆抗体的磁珠;第2组培养体系中除加入100 nmol/LRAPA以外,其余与第1组相同;第3组是CD4^+CD25^+Treg培养第12天后,将细胞进一步分选获取CD4^+CD25^+CD127^- Treg再继续培养,培养体系与第2组相同。实验结果显示,加RAPA培养的CD4^+CD25^+Treg细胞扩增效率比不加RAPA的低,但细胞的纯度明显提高(92%vs 65%)。而加RAPA培养的第2、3组CD4^+CD25^+Treg细胞在纯度上没有明显的差异。加RAPA培养的CD4^+CD25^+Treg细胞扩增后保持其无能状态。体外抑制实验显示体外扩增的CD4^+CD25^+Treg细胞对CD4^+效应T细胞增殖均有明显的抑制作用,而且加RAPA扩增的Treg细胞表现出更强的抑制功能。另外,与第1组体外扩增的Treg细胞相比,加RAPA扩增的Treg细胞表达低水平的IL-2和IFN-γ。实验证明雷帕霉素抑制了CD4^+ CD25^+Treg细胞中混杂的CD4^+效应T细胞的扩增,使用雷帕霉素可以提高体外扩增CD4^+CD25^+Treg细胞的纯度及其免疫抑制功能。This research was designed to develop an effective method for amplifying CD4＋ CD25＋ Treg in vitro, and to study the effect of rapamycin on the expansion of human CD4＋ CD25＋ regulatory T cells in vitro. CD4＋ CD25＋ regulatory T cells were purified from adult peripheral blood with immunomagnetic beads （MACS）, and we set up three groups： in group one, CD4＋ CD25＋ Tregs were cultured with 200 U/ml IL2 and CD3/CD28 antibodycoated beads~ in group two, all culture condition was the same as group one except the addition of 100 nmol/L rapamycin; in group three, CD4＋ CD25＋ Tregs were further sorted into CD4＋ CD25＋ CD127Treg on Day 12, and then were cultured for the second cycle, and the culture condition was the same as in group two. As a result, all cells in the three groups amplified significantly. Compare with the first group, the expansion ratio of the latter two groups is lower （1.0〉（ 10avs 1.8X 103）, but the cell purity increased significantly（92% vs 65%）. The expanded CD4＋ CD25＋ Tregs of the latter two groups still retain the cell anergy. Although the latter two groups showed a stronger inhibiting activity than was the first group, there were no obvious differences between the latter two groups. Moreo ver, compare with the first group, the latter two groups expressed lower levels of IL2 and IFNy. The results of this study show that rapamycin inhibited the expansion of CD4＋ effective T cells which were contaminated in CD4＋ CD25＋ Tregs popula tion therefore the use of rapamycin may effectively improve the cell purity of in vitro expanded CD4＋ CD25＋ Tregs and their immunosuppressive function.

关 键 词：人调节性T细胞 体外扩增 雷帕霉素 

分 类 号：R392.12[医药卫生—免疫学]