小鼠骨髓来源未成熟树突状细胞诱导同种幼稚T淋巴细胞分化的研究

Using mice immature dendritic cells to induce the differentiation of allogenic naive T cells in vitro

目的研究小鼠骨髓来源未成熟树突状细胞（imDC）诱导同种幼稚T淋巴细胞分化为调节性T淋巴细胞（Treg细胞）及辅助性T淋巴细胞（TH）1、TH2和TH17的能力。方法培养Balb／c小鼠骨髓来源树突状细胞（DC），加入脂多糖刺激DC成熟。分别将inff）C与成熟DC（mDC）与同种C57BL／6小鼠幼稚T淋巴细胞共同培养，用酶联免疫斑点法检测Tn1类细胞因子[γ干扰素（IFN-γ）和白细胞介素（IL）-2]、TH2类细胞因子（IL-4和IL-10）、TH17类细胞因子（IL-17）的分泌情况。imDC和miX；分别与同种幼稚T淋巴细胞在加入IL-2及转化生长因子B，的条件下共同培养，用流式细胞术检测T淋巴细胞中Treg细胞比例。结果分别与imDC或mDC共同培养后，T淋巴细胞中分泌IFN-γ细胞的数量分别为（11．67±2．18）／2×105个细胞和（182．00±23．71）／2×105个细胞（P〈0．01），分泌．IL-2细胞的数量分别为（26．67±2．96）／2×105个细胞和（318．30±18．62）／2×105个细胞（P〈0．01），分泌IL-4细胞的数量分别为（17．00±3．78）／2×105个细胞和（45．33±3．48）／2×105个细胞（P〈0．01），分泌IL-10细胞的数量分别为（7．00±1．00）／2×105个细胞和（158．70±10．90）／2×105个细胞和（P〈0．001），分泌IL-17细胞的数量为（0．66±0．33）／2×105个细胞和（238．30±24．39）／2×105个细胞（P〈0．001）。imDC诱导产生的Treg细胞占（22．70±1．53）％，高于mDC诱导产生的（5．42±1．27）％（P〈O．001）。结论imDC诱导同种幼稚T淋巴细胞较多分化为Treg细胞，而较少分化为TH1、TH2和TH17细胞。

Objective To explore the differentiation of allogeneic naive T cells to regulatory T cells （Tregs） and T helper （TH） 1/2/17 cells by coculture with bone marrow-derived,immature dendritic cells （imDC）. Method Bone marrow-derived imDC were cultivated from Balb/c mice. Lipopolysaccharide-stimulated DC were harvested as mature dendritic cells （mDC） and unstimulated cells were collected as imDC. Then imDC or mDC were cocultured with allogeniec naive T cells, respectively. TH1 cytokines [-interferon-γ （IFN-γ） and interleukin （IL）-2], TH2 cytokines （IL-4 and IL-10）, and TH 17 cytokine （IL-17） of co-cultured cells were detected by enzyme linked immunospot assay. CD4＋ Forkhead box p3 （FoxP3） ＋ Treg proportion in CD4＋ cells in the co-cultured system with IL-2 and transforming growth factor-β1 （TGF-β1） was analyzed by flow cytometry. Result As compared with mDC, naive T cells cocultured with imDC secreted much less IFN-γ （11. 67± 2. 18 vs. 182. 00 ± 23. 71 ,P〈O. 01）, IL-2 （26. 67± 2. 96 vs. 318. 30 ± 18. 62, P〈0. 01）, IL-4 （17. 00 ± 3. 78 vs. 45. 33 ± 3. 48, P〈0. 01）, IL-10 （7. 00 ± 1.00 vs. 158. 70 ±10. 90, P〈0. 001） and IL-17 （0. 66± 0. 33 vs. 238. 30± 24. 39, P〈0. 001）. Furthermore, imDC induced more CD4＋ FoxP3＋ Tregs than mDC after adding IL-2 and TGF-131 in the coculture system for 7 days （22. 70 ± 1.53% vs. 5. 42 ± 1.27%, P〈0. 01）. Conclusion imDC are more effective to induce nave T cells to Tregs, but not differentiate to TH1/TH 2/TH 17 cells. These findings provide in vitro experimental evidence for induction of transplant tolerance by adoptive transfer of imDC.