摘要
探讨过表达IDO的大鼠骨髓间充质干细胞(bone marrow mesenchymal stem cell,BMSC)免疫抑制调节作用机制。通过慢病毒转染体系构建过表达IDO大鼠BMSC并加入基因开启剂。采用体外共培养方式共分12组,检测CD40、CD86、CD80、MHCⅡ、CD274、CD45RA、CD45RA+CD45RB、OX62、Treg、CD4、CD8表达;利用液相芯片检测IL-1α、IL-4、IL-1β、IL-2、IL-10、IFN-γ、IL-18、TGF-β1、TGF-β2、TGF-β3的变化情况。结果显示各组与DC单独培养48h后,共培养组中过表达IDO-BMSC+DC培养组中DC表达CD40、CD86、CD80、MHCII、CD45RA、CD45RA+CD45RB、OX62是降低的,而CD274的表达是升高的,说明过表达IDO提高了BMSC对DC的免疫抑制作用;各组与T细胞单独培养48h后,共培养组中过表达IDO-BMSC+T细胞培养组中T细胞中Treg数量是升高的,CD4+T细胞的变化无明显规律性,但CD8+T细胞是降低的,说明过表达IDO-BMSC可调节CD8+T细胞并可以促进Treg的表达;当将各组BMSC+DC+T细胞培养48h后,可见:共培养组中悬浮细胞表达CD40、CD86、CD80、MHCⅡ、CD45RA、CD45RA+CD45RB、OX62是降低的,而CD274是升高的,再次说明过表达IDO提高了BMSC对DC的免疫抑制作用,而Treg数量增加,CD4+T细胞的变化无明显规律性,但CD8+T细胞是降低的,再次说明IDO能调节CD8+T细胞并可促进Treg的表达;各组培养48h后的上清液进行液相芯片检测,可见IDO-BMSC+DC+T细胞培养组上清中IL-1α、IL-4、IL-1β、IL-2、IFN-γ、IL-18减低,IL-10、TGF-β1、TGF-β2、TGF-β3表达量升高。综上,过表达IDO的BMSC可以通过有效调节DC、T细胞水平及细胞因子分泌,从多免疫途径进行免疫抑制调节。
We aimed to explore the mechanism of immunosuppressive regulation of rat bone-marrow mesenchymal stem cells(BMSCs)with overexpressed IDO(IDO-BMSC).IDO-BMSCs were constructed by using a lentivirus transfection system and by adding gene promoters.The IDO-BMSCs were divided into 3 groups which were co-cultured for 48 hours with DC(group1),T cells(group 2)and DC plus T cells(group 3)respectively.The expression levels of CD40,CD86,CD80,MHCII,CD274,CD45 RA,CD45 RA+CD45 RB,OX62,Treg,CD4 and CD8 were measured.Liquid-phase chips were used to measure changes in IL-1α,IL-4,IL-1β,IL-2,IL-10,IFN-γ,IL-18,TGF-β1,TGF-β2 and TGF-β3.The results showed that at the end of culture,the DC in group 1 downregulated the expression of CD40,CD86,CD80,MHCII,CD45 RA,CD45 RA+CD45 RB,and OX62;conversely,CD274 expression increased.This finding indicated that IDO overexpression enhanced the immunosuppressive effect of BMSCs on DC.In group 2,the number of Treg of T cells in the overexpressed IDO-BMSC+T cell culture group increased.The CD4+T cells showed no obvious changes while CD8+T cells decreased.These results suggested that IDO overexpression in BMSCs can regulate CD8+T cells and promote Treg expression.In group 3,the suspension cells showed decreased expression of CD40,CD86,CD80,MHCII,CD45 RA,CD45 RA+CD45 RB,and OX62,but increased expression of CD274.Thus,IDO overexpression indeed enhanced the immunosuppressive effect of BMSCs on DC.Although the number of Treg cells increased,the change of CD4+T cells was not clear.However,CD8+T cells decreased,further indicating that IDO can regulate CD8+T cells and promote the expression of Treg cells.Liquid-phase chips were subsequently used to detect the supernatant amount of cytokines in each group after 48 hof cultivation.We found that in the supernatant of group 3,IL-1α,IL-4,IL-1β,IL-2,IFN-γand IL-18 decreased,whereas the expression of IL-10,TGF-β1,TGF-β2 and TGF-β3 increased.In summary,BMSCs with overexpressed IDO can regulate immune suppression through multiple immune levels by effectively regulating the levels of DC and T cells,as well as cytokine secretion.
作者
李贝贝
韩金秀
严丹
撒亚莲
李宏远
贺继刚
LI Bei-bei;Han Jin-xiu;Yan Dan;SA Ya-lian;Li Hong-yuan;He Ji-gang(Cardiovascular Surgery1 , Basic Research Institute2, the Affiliated Hospital of Kunming University of Science and Technology, the First People's Hospital of Yunnan Province, Kunming 650032, China)
出处
《现代免疫学》
CAS
CSCD
北大核心
2018年第3期191-196,共6页
Current Immunology
基金
国家自然科学基金(81460073
31460298)
云南省科技厅-昆明医科大学应用基础研究联合专项(2014FB089)
云南省教育厅科学研究基金(2015Z051)
中国博士后科学基金(2015M582764XB)
成都医学院2015年度科研项目(CYZ15-18)