【摘要】 目的:研究大鼠CD4+CD25+T调节细胞(Tr)的分离培养,并对其功能进行初步分析。方法:无菌条件下切取大鼠脾脏分离脾淋巴细胞。用免疫磁珠细胞分离系统(MACS)分选CD4+CD25+T细胞,并以流式细胞术检测其纯度后,对其进行扩增。采用混合淋巴细胞反应研究CD4+CD25+Tr细胞对CD4+CD25-T细胞的免疫抑制作用。用ELISA法检测培养上清中IL-2、IFN-γ及IL-10水平的差异。结果:MACS分离的CD4+CD25+T细胞的纯度达86%~93%。该细胞与CD4+CD25-T细胞相比能特异性地表达Foxp3基因。体外培养中能明显抑制效应T细胞增殖及其分泌IFN-γ、IL-2,但其自身能分泌Th2型细胞因子IL-10。结论:采用MACS系统阴性加阳性分选,可高效快速的获得理想纯度和免疫抑制功能的大鼠CD4+CD25+T调节细胞,该细胞对CD4+CD25-T细胞具有明显的免疫抑制作用,并能特异性的表达Foxp3基因。更多还原

【Abstract】 AIM: To investigate the isolation method and to analyze the function of rat CD4~+ CD25~+ regulatory T cells. METHODS: Lymphocytes were isolated from the rat spleens and then CD4~+ CD25~+ T cells were sorted by magnetic bead cell sorting (MACS) system. The purity and Foxp3 expression of CD4~+ CD25~+ T cells were analyzed by flow cytometry(FCM) and RT-PCR, respectively. The suppressive effect of CD4~+ CD25~+ T cells on the proliferation of CD4~+ CD25~- T cells was analyzed by mixed lymphocyte reaction. IL-2, IFN-γ and IL-10 levels in culture supernatant were detected by ELISA. RESULTS: The purity of CD4~+ CD25~+ T cells sorted by MACS was 86%-93%. The CD4~+ CD25~+ T cells could specifically express the Foxp3 gene as compared with CD4~+ CD25~- T cells. In vitro CD4~+ CD25~+ T cells could suppress the proliferation of CD4~+ CD25~- T cells and IFN-γ, IL-2 production, but they themselves could secrete IL-10. CONCLUSION: We established an effective procedure for enrichment of CD4~+ CD25~+ regulatory T cells by MACS with satisfactory cell purity, viability and function. CD4~+ CD25~+ T cells can suppress CD4~+ CD25~- T cells and specifically express Foxp3 gene.更多还原