甘露糖结合凝集素对CD11c^+髓样树突状细胞表型和功能的影响

Impacts of mannan-binding lectin on phenotype and function of CD11c-positive human peripheral blood myeloid dendritic cells

目的探讨甘露糖结合凝集素(MBL)对人外周血CD11c+髓样树突状细胞(CD11c+m DC)表型和功能的影响。方法磁珠分选技术获得健康志愿者CD11c+m DC和CD4+T淋巴细胞。在CD11c+髓样树突状细胞中加入不同浓度的MBL(5、10、20μg/ml)刺激,以不加MBL的细胞作为对照,应用ELISA法检测细胞培养上清液中的白细胞介素(IL)12水平,流式细胞仪检测细胞表面分子CD40、CD80、CD86及HLA-DR的表达。用MTT法测定CD11c+m DC刺激CD4+T淋巴细胞的增殖能力。ELISA法检测细胞培养液中IL-4和干扰素(IFN)γ水平。计量资料多组间比较采用单因素方差分析,进一步两两比较采用LSD-t检验。结果与未加MBL刺激组相比,3种浓度MBL均显著增强CD11c+m DC表面分子CD40、CD80、CD86、HLA-DR表达及IL-12分泌(F值分别为44.34、27.35、15.57、48.38、38.27,P值均<0.001),其中IL-12分泌呈MBL浓度依赖性;MBL刺激组细胞增殖能力显著高于正常对照组和未刺激组(F=23.43,P<0.001);MBL刺激组细胞的IFNγ水平显著高于正常对照组和未刺激组(F=28.25,P<0.001);而MBL刺激组细胞的IL-4水平明显低于正常对照组和未刺激组(F=40.03,P<0.001)。结论 MBL能够有效刺激CD11c+m DC的活化,诱导CD4+T淋巴细胞向辅助性T淋巴细胞1分化,提示其可能通过调节CD11c+m DC的表型和功能参与HBV的控制和清除。

Objective To investigate the impacts of mannan- binding lectin（ MBL） on the phenotype and function of CD11c- positive human peripheral blood myeloid dendritic cells（ CD11c＋m DC）. Methods CD11c＋m DC and CD4＋T lymphocytes from healthy human volunteers were isolated by magnetic bead sorting and were stimulated by different concentrations of MBL（ 5,10,20 μg/ml）. Compared with the non- MBL stimulation group,the levels of interleukin- 12（ IL- 12） in the supernatant of culture medium in different MBL stimulation groups were determined by enzyme- linked immunosorbent assay（ ELISA）. The expression of CD40,CD80,CD86,and HLA- DR on CD11c＋m DC surface was measured by flow cytometry. CD11c＋m DC- stimulated proliferation abilities of CD4＋T lymphocytes were determined by MTT assay. The levels of interleukin- 4（ IL- 4） and interferon- gamma（ IFNγ） in the coculture medium were measured by ELISA. Comparison of the means between multiple groups was made by one- way ANOVA and pairwise comparison between any two groups was made by LSD t- test. Results Compared with the non- MBL stimulation group,the MBL stimulation groups（ 5,10,20 μg/ml） had significantly higher expression of CD40,CD80,CD86,and HLA- DR on CD11c＋m DC surface and significantly increased IL- 12 secretion（ F = 44. 34,P〈0. 001; F = 27. 35,P〈0. 001; F = 15. 57,P〈0. 001; F = 48. 38,P〈0. 001; F = 38. 27,P〈0. 001）. The IL- 12 secretion was MBL concentration- dependent. The proliferation ability of CD4＋T lymphocytes was significantly higher in the MBL stimulation group than in the non- MBL stimulation group and the control group（ F = 23. 43,P〈0. 001）. The MBL group had a significantly higher IFNγ level but a significantly lower IL- 4 level compared with the non- MBL group and the control group（ F = 28. 25,P〈0. 001; F =40. 03,P〈0. 001）. Conclusion MBL can effectively stimulate the activation of CD11c＋m DC and induce the differentiation from CD4＋T lymphocytes to type 1 helper T cells. Therefore,MBL is possibly involved in the control and clearance of hepatitis B virus by regulating the phenotype and function of CD11c＋m DC.