转化生长因子-β1联合白细胞介素-2体外诱导非肥胖糖尿病小鼠调节性T细胞的产生

In vitro conversion of CD4^＋ CD25^ - Naive T cells to CD4^＋ CD25 ^＋ Foxp3^＋ regulatory T cells from non-obese diabetic mice by TGF-β1 combined with IL-2

目的体外诱导非肥胖糖尿病（NOD）小鼠CD4^＋CD25^＋Foxp3^＋调节性T细胞（Treg）的产生并检测其免疫抑制功能。探讨外源性白细胞介素（IL）-2在诱导方案中的作用。方法分选NOD小鼠童贞T细胞（NmveT），利用Anti—CD3、Anti—CD28刺激，同时给予转化生长因子-β1（TGF—B1）和白细胞介素-2（IL-2），共同培养5d，收获诱导性Treg（iTreg），经流式细胞仪检测其表型。利用体外T细胞增殖体系，对比NOD小鼠天然Treg（nTreg），评价iTreg的免疫抑制能力。将诱导方案中的IL-2撤除以观察其作用。结果TGF-β1联合IL-2能诱导NOD小鼠NaiveT转化为iTreg，较对照组有统计学意义[（41．33±3．21）％比（8．00±3．00）％，P〈0．05]。iTreg可有效抑制T细胞增殖，其能力与nTreg的差异无统计学意义[（40．33±1．03）％比（38．33±3．06），P〉0．05]。外源性IL-2有利于iTreg的产生[（41．33±3．21）％比（15．00±1．00）％，P〈0．05]。结论TGF-β1联合IL-2可在体外诱导Naive T转化为具有免疫抑制功能的Treg。

Objective To investigate the roles of TGF-β1 and IL-2 to induce the differentiation of CD^＋ CD25^＋Foxp3 ^＋regulatory T cells from non-obese diabetic （NOD） mice and the mechanisms. Methods （1） The Naive T cells from NOD mice were stimulated by Anti-CD3 and Anti-CD28 with TGF-β1 and IL-2 for a 5-day culture. Four groups were established：control group （0 μg/L TGF-β1 ） ,low concentration group （ 1 μg/L） ,middle concentration group （5 μg/L） and high concentration group （ 10 μg/L）. At the end-point of culture ,the ratio of CD4^＋CD25^＋Foxp3^＋T cells were measured by FACS individually. （2） The CD90^＋cells from NOD mice were separated by MACS and stimulated by Anti-CD3 and Anti- CD28 for establishing the proliferation system of T cells. CD4^＋CD25^＋T cells were isolated by MACS and added into the proliferation system to measure the immunosuppressive activity of these cells. Meanwhile the role of IL-2 in this protocol was studied. Results TGF-β1 combined with IL-2 could convert Naive T cells from NOD mice to iTreg which could suppress the proliferation of T cells [ （41.33 ± 3.21 ） % vs （ 8.00 ± 3.00）% ,P 〈 0.05]. 5 μg/L TGF-β1 was an effective dose which could increase the ratio of CD4^＋CD25^＋Foxp3^＋Treg cells. IL-2 was essential in this protocol [ （41.33 ± 3.21 ） % vs （ 15.00 ± 1.00） %, P〈0.05].Conclusion TGF-β1 combined with IL-2 induced CD4^＋CD25^＋Foxp3^＋Treg from Naive T cells,which could suppress the proliferation of T cells.