细胞因子诱导的人脐血CD4 T细胞Th17极化态的建立

目的建立一个合理的CD4T细胞极化的体外细胞培养方法,研究细胞因子刺激的人脐血CD4T细胞极化时产生IL-17的效应/记忆T细胞（Th17细胞）的表达。方法用TGF-β、抗IL-4、抗IFN-γ、IL-6、IL-1β活化幼稚脐血CD4T细胞,促成Th17细胞表型,IL-23具有维护功能。用流式细胞术分析在Th17极化态和Th0非极化态时CD4T细胞上IL-17和IFN-γ的表达,同时用酶联免疫吸附试验（ELISA）测定细胞培养上清液中IL-17和IFN-γ的水平。结果IL-17＋IFN-γ-（Th17）细胞比例在Th17极化态时为（28±2.3）%,非极化态时为（3.2±0.4）%;培养上清液中分泌的IL-17浓度在Th17极化态时为（7350±1530）pg/ml,非极化态时为（218±32）pg/ml,两项指标在Th17极化态时都显著高于非极化态时,差异均有高度统计学意义（P〈0.001）。结论多种细胞因子刺激的CD4T细胞极化时促进了IL-17产物的表达,体外Th17极化态的建立为研究IL-17相关的免疫性疾病提供了一个平台。

Building of Th17-polarizing Condition on Induced Human Cord Blood CD4 T Cells by Cytokines

Objective To build a appropriate cell culture method of CD4-polarized T cells in vitro and to investigate expression of interleukin 17 （IL-17）-producing cells （Th17 cells） when human cord blood CD4 T cells was polarized by cytokines. Methods Activation of native cord blood CD4 T cells with TGF-β, anti-IL-4, anti-IFN-γ, IL-6, IL-1βresulted in the development of a Th17 phenotype, and maintained the Th17 phenotype with IL -23. IL -17 and IFN -γ phenotype were detected under Th17 polarizing and Th0 non-polarizing condition in CD4 T cells by flow cytometry. Meanwhile, IL-17 and IFN-γ protein levels were measured in cell culture supernatants using enzyme-linked immunosorbent assay（ELISA）. Results The proportion of IL-17＋IFN-γ -（Th17） cells was increased significantly in Th17- polarized cultures（28.0±2.3）% compared with the nonpolarized cultures（3.2±0.4）% （P〈0.001）. Analysis of secreted cytokines within cell culture supernatants revealed a significant concentration of IL-17 in Th17-polarized cuhures（7 350±1 530）pg/ml compared with nonpolarized cultures（218±32）pg/ml（P〈 0.001）. Conclusion Expression of IL-17-production is enhanced when human cord blood CDg T cells is polarized by cytokines. In vitro Th17-polarizing condition builded provide a plateform for investigating IL-17 related immune disease.