ManLAM对树突状细胞成熟及下游免疫的影响

Effects of ManLAM on the maturation of dendritic cells and downstream immunity

目的:研究结核分枝杆菌(MTB)成分带甘露聚糖帽的脂阿拉伯甘露聚糖(mannose-capped lipoarabinomannan,ManLAM)对树突状细胞(dendriticcells,DCs)成熟及下游免疫的影响。方法:分离健康人外周血单个核细胞,用GM-CSF和IL-4诱导DCs生长,培养至第五天换液加细胞因子,第六天分三组,按实验设计加ManLAM和LPS。第七天收集细胞送流式检测DC-SIGN、HLA-DR、CD86、CD83表达水平;ELISA检测DCs培养上清液中IL-12和IL-10水平;混合淋巴细胞反应检测DCs诱导CD4+T淋巴细胞增值能力;DCs与初始CD4+CD45RA+T细胞共培养48小时后,ELISA检测培养上清液中IFN-γ水平。结果:(1)ManLAM组DCs表达CD86、CD83等成熟标志物较成熟DCs组下降,差异有统计学意义(P<0.05);(2)ManLAM组DCs培养上清液中IL-10水平较成熟组升高,IL-12水平较成熟组下降,差异有统计学意义(P<0.05);ManLAM组DCs刺激淋巴细胞增殖能力及刺激初始CD4+CD45RA+T细胞向Th1细胞分化能力较成熟组下调,差异有统计学意义(P<0.05)。结论:ManLAM干扰DCs成熟,下调DCs诱导的淋巴细胞增殖能力和激活初始CD4+CD45RA+T细胞向Th1细胞分化能力。

Objective：To observe the effects of mycobacterium tuberculosis components （mannose-capped lipoarabinomannan,ManLAM） on the maturation of dendritic cells and downstream immunity.Methods：The peripheral blood mononuclear cells were obtained from healthy individuals,and then were cultured in medium with GM-CSF and IL-4.Five days later the medium was changed and cytokine was added to.On the sixth day,the cells were divided into three groups.Meanwhile ManLAM and LPS were added to stimulate the DCs according to the experimental design.Seven days later,the cells were collected and the level of DC-SIGN,HLA-DR,CD86 and CD83 were detected by flow cytometry.The levels of IL-12 and IL-10 in the supernatant were measured by enzyme-linked immunosorbent assay （ELISA）.Proliferative induction of DCs to CD4＋T lymphocytes was measured by mixed lymphocyte reaction（MLR）.The naive CD4＋CD45RA＋ T cells and allogeneic DCs were co-cultured for 48 hours.Subsequently,the supernatant IFN-γ level in each group was measured by ELISA.Results：（1）The expression of CD86 and CD83 on DCs in ManLAM group was lower than those of the mature group （P0.05）.（2）In ManLAM group,the IL-10 level in the supernatant was higher and the IL-12 level was lower than those in mature group （P0.05）.（3）The potency of proliferative induction of DCs to CD4＋T cells and the activation of DCs from naive CD4＋ CD45RA＋T cells to Th1 cells in ManLAM group were lower than those in mature group（P0.05）.Conclusion：The maturation of DCs could be restrained by ManLAM,so are the proliferation of T cells induced by DCs too and the activation of nave CD4＋ CD45RA＋T cells into Th1 cells.