IFN-γ刺激培养未成熟树突状细胞免疫耐受效应机制研究  

Study on mechanism of immune tolerance effect of IFN-γ stimulated cultured immature dendritic cells

作　　者：吴银平[1] 袁发焕[1] 侯卫平[1] 

机构地区：[1]第三军医大学新桥医院肾病科,重庆400037

出　　处：《重庆医学》2010年第21期2852-2854,2857,共4页Chongqing medicine

基　　金：国家自然科学基金资助项目(30700371)

摘　　要：目的通过γ-干扰素(IFN-γ)刺激培养未成熟树突状细胞,研究在体外IFN-γ诱导未成熟树突状细胞免疫耐受的效应机制。方法采用经典的GM-CSF+IL-4诱导方案,以获得细胞表面缺乏共刺激分子的大鼠骨髓来源的未成熟树突状细胞,经IFN-γ刺激培养后,通过外源性抗原T细胞表位pCol(28-40)多肽培养,流式细胞仪检测表面MHCⅡ类分子及共刺激分子CD80、CD86,DC表面相对特异性标记OX62的情况,将脾脏分离淋巴细胞分为3组,即对照组、imDC组和imDCIFN-γ组,采用MTT法检测各组淋巴细胞增殖情况,Anneix-V-Flous检测各组淋巴细胞凋亡,ELISA法检测体外培养脾细胞上清液Th1/Th2型细胞因子,观察体外CD4+T细胞Th亚群的分化情况。结果 IFN-γ刺激培养的未成熟树突细胞通过外源性抗原T细胞表位pCol(28-40)多肽培养后低表达表面MHCⅡ类分子(14.01%)及共刺激分子CD80(4.78%)、CD86(19.79%),同样低表达DC表面相对特异性标记OX62(15.31%,P<0.05,n=4);MTT法检测共培养的脾淋巴细胞imDC组i、mDCIFN-γ组与对照组增殖差异有统计学意义(P<0.05,n=5);但imDCIFN-γ组凋亡率为(12.9±0.02)%,与对照组和imDC组相比均明显增高(P<0.05,n=5);Th1型细胞因子明显向Th2型细胞因子偏移。结论 IFN-γ刺激培养的未成熟树突状细胞对外源性抗原T细胞表位pCol(28-40)多肽摄取及提呈能力明显下降;抑制T细胞增殖,促进T细胞凋亡;并且促使Th1型细胞因子向Th2型细胞因子漂移,诱导形成免疫耐受。Objective To study the immune tolerance mechanisms in vitro of the IFN-γ stimulated cultured immature dendritic cells.Methods The classical method of GM-CSF ＋ IL-4 was adopted to obtain the lack of cell surface costimulatory molecules in rat bone marrow-derived.Immature dendritic cells were stimulated culture by IFN-γ,then through the exogenous antigen T cell epitope pCol（28-40） peptide culture,flow cytometry detection of surface MHC Ⅱ molecules and costimulatory molecules CD80,CD86,DC-specific surface sign OX62.Separated lymphocytes in the spleen were divided into three groups： control group,imDC group and imDCIFN-γ group.MTT was used to detect lymphocyte proliferation in each group;Anneix-V-Flous to detect lymphocyte apoptosis in each group;ELISA assay was used to culture spleen cell supernatant Th1/Th2 type cytokines,CD4＋ T-cell differentiation of Th subsets in vitr was observed.Results IFN-γ stimulated cultured immature dendritic cells through the exogenous antigen T cell epitope pCol（28-40） peptide cultured had low expression of surface MHC Ⅱ molecules（14.01%）,and co-stimulatory molecule CD80（4.78%）,CD86（19.79%）,the same low expression of DC surface,a relatively specific sign OX62（15.31%,P〈0.05,n=4）,MTT assay co-cultured spleen lymphocytes imDC group,imDCIFN-γ group and the control group had no significant proliferation（P〈0.05,n=5）;but the apoptosisrate of imDCIFN-γ group（12.98±0.02）% was significantly higher than that of control group and imDC group（P〈0.05,n=5）;Th1 type cytokines apparenlty drifted to Th2 type cytokines Conclusion IFN-γ stimulated cultured immature dendritic cells,then through exogenous antigen T cell epitope pCol（28-40） peptide culture show dendritic cells geting antigen and antigen-presenting ability significantly decreases,inhibits T cell proliferation and promotes T-cell apoptosis;the experiments also show that IFN-γ stimulated cultured immature dendritic cells promote Th1 type cytokines drifting to Th2 type cytokines,indu

关 键 词：Γ-干扰素 未成熟树突状细胞 免疫耐受 T细胞 

分 类 号：R392.32[医药卫生—免疫学]