过敏性哮喘小鼠树突状细胞负荷Der p2后改变及与T细胞增殖分化的关系研究

Effects of Der p2 on DC and the Relationship between DC and T Cell Proliferation

目的:探讨哮喘小鼠与正常小鼠骨髓源性树突状细胞(DC)负荷Der p2抗原后表达表面分子(CD11c、CD86)和细胞因子(IL-10、IL-12p70)的差异及其对Th1和Th2型细胞因子平衡的影响,进一步研究过敏性哮喘发生中DC的可能作用。方法:分别从哮喘组和对照组提取骨髓培养DC,第五天负荷Der p2,24小时后吹打收集细胞,观察DC形态,用流式细胞仪检测孵育后细胞表面CD11c、CD86表达。并留取负荷Der f2前后培养上清,ELISA法检测IL-10及IL-12p70含量。同时以DC:反应细胞比例为1:10混合培养,72 h后ELISA法检测混合培养上清中IL-4、IL-5、IFN-γ的水平。结果:1负荷Der p2后,哮喘组CD86、CD11c表达比对照组高,分别为(t=11,P<0.05)(t=4.9,P<0.05),差异有统计学意义;2在细胞因子分泌方面,Der p2负荷前,两组DC均能分泌IL-10与IL-12p70,IL-10水平哮喘组高(t=9.5,P<0.05),而IL-12p70水平对照组高(P<0.05);负荷Der p2后,对照组IL-10、IL-12p7分泌量比负荷前明显增加(P<0.05),哮喘组无明显差异(P>0.05);3在DC刺激同种T细胞因子分泌方面,负荷Der p2后哮喘组DC刺激T细胞分泌IL-4、IL-5分泌能力明显增强(P<0.05),而刺激INF-γ能力降低(P<0.05)。结论:DC在过敏性哮喘中起着重要作用,异常DC通过增加CD86、CD11c的表达和减少IL-10及IL-12的合成,致使T细胞向Th2细胞优势分化。

Objective： To investigate the differences of the expression of CD11 c, CD86 and the secretion of IL-10 and IL-12p70 in the dendritic cells（DCs） from allergic asthmatics and healthy donors, which loaded the Der p2 antigen, and further study the possible role of DCs in the occurrence of allergic asthma. Methods： Marrow-derived dendritic cells（DC） were extracted and cultured, and the cells were collected after the Derp2 load 24 hours at the 5th day, the cells morphology was observed, and the CD11 c, CD86 expression was tested with flow cytometry cell. After and before the Derp2 loaded, the culture supernatants was collected to test the IL-10 and IL-12P70 levels with ELISA, then co-culture the DC and T cells with a proportion of 1：10 to test the levels of IL-4, IL-5 and INF-γ in culture supernatant with ELISA. Results： 1 After loading Der p2, compared with control group, the expression of CD86, CD11 c in asthma group was increased with statistical significance, respectively（t=11, P〈0.05）（t=4.9, P〈0.05）; 2 In the aspect of secretion of the cytokines, both groups could secrete IL-10 and IL-12P70 before Der p2 loaded, and the level of IL-10 in the asthma group was higher（t=9.5, P〈0.05）, while the level of IL-12p70 in the control group was higher（t=6.56, P〈0.05）; after Der p2 load, the level of IL-10 and IL-12p70 of the control group was increased significantly（t=19.4, P〈0.05）, while asthma group had no significant difference（P〈0.05）. 3In the aspect of DC stimulation on secretion of T cells, after Der p2 loaded, for the asthma group, the levels of IL-4 and IL-5 secreted by T cells by DC stimulation was significantly enhanced（P〈0.05）, while the level of INF-γ was decreased（P〈0.05）. Conclusion： These results identify abnormal DC can increase CD86 and CD11 c expression and reduce the synthesis of IL-10 and IL-12 which are necessary and sufficient for the T cells differentiation into Th2 cells.