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目的利用不同种类树突状细胞(dendritic cells,DC)体外扩增获得表型和功能稳定的CD4+CD25+Foxp3+调节性T细胞(Treg)。方法免疫磁珠法(MACS)分离Balb/c小鼠CD4+CD25+调节性T细胞,利用与调节性T细胞同基因或异基因成熟DC、未成熟DC和调节性DC刺激其扩增,流式细胞术测定其纯度和表型。以CD4+CD25-T细胞作为反应细胞,验证扩增前后Treg细胞的免疫抑制功能。结果MACS分离的CD4+CD25+调节性T细胞纯度达到(95.38±1.82)%,同基因和异基因DC都能有效刺激Treg细胞体外扩增,其中同基因成熟DC扩增效果最为明显。而且同基因成熟DC扩增后CD4+CD25+调节性T细胞纯度达到(94.16±1.88)%,而且高表达Foxp3分子。当CD4+CD25+调节性T细胞与效应T细胞比例为1∶1时,能够有效的抑制效应T细胞的增殖,而且,同基因成熟DC扩增的CD4+CD25+调节性T细胞的抑制效果比新分离的Treg效果更好。结论同基因成熟DC能够体外扩增表型和功能稳定的Treg细胞。
OBJECTIVE: To clone and characterize CD4 + CD25 + Foxp3 + regulatory T cells (Tregs) with stable dendritic cells (DCs) in vitro. Methods CD4 + CD25 + regulatory T cells were isolated from Balb / c mice by immunomagnetic beads method (MACS), and stimulated by syngeneic or allogeneic maturation DCs, immature DCs and regulatory DCs with regulatory T cells. Flow cytometry Cytometry measures its purity and phenotype. CD4 + CD25-T cells were used as reactive cells to verify the immunosuppressive function of Treg cells before and after expansion. Results The purity of CD4 + CD25 + regulatory T cells isolated by MACS was (95.38 ± 1.82)%. Both syngeneic and allogeneic DCs could effectively stimulate Treg cells to proliferate in vitro. The purity of CD4 + CD25 + regulatory T cells reached 94.16 ± 1.88% after DC maturation, and Foxp3 was overexpressed. When the ratio of CD4 + CD25 + regulatory T cells to effector T cells was 1: 1, the proliferation of effector T cells was effectively inhibited, and the inhibitory effect of CD4 + CD25 + regulatory T-cells expanded with the same gene maturation was more pronounced Separated Treg better. Conclusions DC with the same gene can expand phenotype and stable Treg cells in vitro.