同种异型抗原冲击不成熟CD8α^+ DCs抑制T细胞增殖的体外实验  

Allogeneic antigen-pulsed murine immature CD8α^+ dendritic cells prevent T-cells proliferation in vitro

作　　者：陈康[1] 曹开源[2] 徐霖[3] 吴耀良[1] 袁广卿[2] 张甜[2] 傅强[4] 

机构地区：[1]中山大学第一附属医院黄埔院区检验科,广东广州510700 [2]中山大学临床检验标准化研究中心,广东广州510080 [3]中山大学中山医学院免疫学教研室,广东广州510080 [4]中山大学中山医学院微生物教研室,广东广州510080

出　　处：《中国病理生理杂志》2010年第11期2202-2206,共5页Chinese Journal of Pathophysiology

基　　金：广东省自然科学基金资助项目(No.8451008901000213);中山大学医科青年教师启动基金资助项目(No.2007003)

摘　　要：目的:探索同种异基因抗原体外冲击后小鼠优势不成熟CD8α+树突状细胞(DCs)体外抑制T细胞增殖的最佳条件。方法:取SPF级C57BL/6(H-2b)小鼠的骨髓细胞,GM-CSF+IL-4+SCF+Flt3L诱导制备优势不成熟CD8α+DCs,于培养最后1d加入梯度蛋白浓度(0、2.5、5、10和20mg/L)同种异基因(H-2d)小鼠脾细胞抗原冲击DCs。经抗原冲击后的DCs分别与同系T细胞按1∶1、2∶1、4∶1比例共培养,MTT法观察T细胞增殖情况。ELISA法检测上清中细胞因子(IFN-γ和IL-10)的含量。以GM-CSF+IL-4+TNF-α诱导的成熟DCs作为对照。结果:优势不成熟CD8α+DCs与T细胞1∶1共培养时不能有效刺激T细胞增殖,与2∶1及4∶1组相比刺激能力最弱(P<0.05);上清中IFN-γ的分泌明显低于成熟DCs对照组,且经小剂量抗原(<2.5mg/L)冲击时IL-10的分泌高于对照组。结论:经小剂量(2.5mg/L)同种异基因抗原冲击后的优势不成熟CD8α+DCs,与T细胞1∶1混合,是抑制T细胞增殖的最佳条件。AIM： To explore the optimal condition of predominant immature CD8α＋ dendritic cells（predo-iDCs） preventing T-cell proliferation pulsed by allogeneic antigen. METHODS： Predo-iDCs, induced with GM-CSF ＋IL-4 ＋SCF ＋Flt3L from SPF healthy C57BL/6 murine bone marrow cells, were pulsed by different doses （0, 2.5, 5, 10 and 20 mg/L）of allogeneic murine splenocyte antigen. Syngeneic T-cells were co-incubated with Ag-pulsed DCs （DCs/T=1∶ 1, 2∶ 1, 4∶ 1） and the T-cell proliferation was measured by MTT. The secretion of cytokines （IFN-γ and IL-10） in the co-incubated supernatants was detected by ELISA. The effect of prevention of T-cell proliferation generated by murine predo-iDCs pulsed by allogeneic antigen was detected. The control derived from mature dendritic cells（mDCs）induced by GM-CSF ＋IL-4 ＋TNF-α. RESULTS： The effect of Ag-pulsed predo-iDCs for stimulating T-cell proliferation was the slightest in predo-iDCs/T 1∶ 1 group, compared with that in predo-iDCs/T 2∶ 1 and 4∶ 1 group （P〈0.05）. The secretion of IFN-γ in mixed lymphocyte reaction（MLR） was significantly lower than the one in mDCs control group, while the secretion of IL-10 was higher than that in control group when low dose of antigen （〈2.5 mg/L） was added into MLR. CONCLUSION： Predominant iDCs pulsed by low dose of allogeneic antigen （2.5 mg/L） mixed 1∶ 1 with T-cells is the optimal condition for the prevention of T-cells proliferation.

关 键 词：树突状细胞 移植物抗宿主病 T淋巴细胞 

分 类 号：R318.06[医药卫生—生物医学工程]