Dong L, Chen X, Shao H, Bai L, Li X, Zhang X
Tianjin Medical University Eye Hospital, Eye Institute & School of Optometry and Ophthalmology, Tianjin 300384, China.
Department of Ophthalmology and Visual Sciences, Kentucky Lions Eye Center, University of Louisville, Louisville, KY 40202-1594, United States.
Curr Mol Med. 2018 Mar 9;17(7):478-487. doi: 10.2174/1566524018666180207155614.
We have previously reported that MSCs inhibited experimental autoimmune uveitis (EAU) in rodent models induced by either uveitogenic antigens or antigen-specific T cells. In this study, we explored the inhibitory mechanisms of MSCs on dendritic cells (DCs) in EAU.
We collected the DCs from the lymph nodes of MSC treated or untreated EAU rats, as well as bone marrow derived DCs cultured in vitro with or without MSC treatment. The levels of costimulatory molecules of CD80, CD86, CD40, OX40L and suppressors of cytokine signaling (SOCS1, SOCS2, and SOCS3) on these DCs were analyzed by flow Cytometry. The expression of CCR-7 and MMP-9 was examined by real time PCR and western blots. Total proteins of STAT1 and STAT6 signaling molecules and their phosphorylation were examined by western blots. ShRNA of STAT1 and STAT6 were respectively employed to explore the influence of STAT1 and STAT6 knockdown on DCs.
MSC treatment down-regulated the expression of CD80, CD86, CD40, and OX40L, as well as CCR-7 and MMP-9, but increased the levels of SOCS1, SOCS 2, and SOCS3 on DCs. STAT1 phosphorylation was reduced while STAT6 phosphorylation was enhanced in MSC treated DCs. Moreover, MSC treatment and STAT1 shRNA equally reduced CCR-7 and MMP-9 levels in DCs, and inhibited the proliferation of R16-specific T cells. In contrast, knockdown of STAT6 in DCs by STAT6 shRNA increased the expression of CD80 and CD86 and accelerated the proliferation of R16-specific T cells.
MSCs inhibit DC maturation by regulating Stat1 and Stat6 phosphorylation in EAU.
我们之前报道过，间充质干细胞（MSCs）可抑制由葡萄膜炎抗原或抗原特异性T细胞诱导的啮齿动物模型中的实验性自身免疫性葡萄膜炎（EAU）。在本研究中，我们探讨了MSCs对EAU中树突状细胞（DCs）的抑制机制。
我们从接受或未接受MSCs治疗的EAU大鼠的淋巴结中收集DCs，以及在体外接受或未接受MSCs治疗培养的骨髓来源的DCs。通过流式细胞术分析这些DCs上共刺激分子CD80、CD86、CD40、OX40L以及细胞因子信号抑制因子（SOCS1、SOCS2和SOCS3）的水平。通过实时PCR和蛋白质印迹法检测CCR-7和MMP-9的表达。通过蛋白质印迹法检测信号转导和转录激活因子1（STAT1）和信号转导和转录激活因子6（STAT6）信号分子的总蛋白及其磷酸化水平。分别使用STAT1和STAT6的短发夹RNA（shRNA）来探讨STAT1和STAT6基因敲低对DCs的影响。
MSCs治疗下调了DCs上CD80、CD86、CD40和OX40L以及CCR-7和MMP-9的表达，但增加了DCs上SOCS1、SOCS2和SOCS3的水平。在接受MSCs治疗的DCs中，STAT1磷酸化水平降低而STAT6磷酸化水平增强。此外，MSCs治疗和STAT1 shRNA同样降低了DCs中CCR-7和MMP-9的水平，并抑制了R16特异性T细胞的增殖。相反，通过STAT6 shRNA敲低DCs中的STAT6会增加CD80和CD86的表达并加速R16特异性T细胞的增殖。
在EAU中，MSCs通过调节Stat1和Stat6磷酸化来抑制DC成熟。
