类风湿关节炎小鼠模型骨髓间充质干细胞体外的免疫调节功能

In vitro immunomodulatory function of bone marrow mesenchymal stem cells in mouse model with rheumatoid arthritis

目的 探索类风湿关节炎(RA)模型小鼠自体骨髓间充质干细胞(MSC)体外的免疫调节功能。方法 构建胶原诱导关节炎(CIA)小鼠模型,分离其骨髓MSC体外培养,鉴定细胞表型;同时分离培养正常小鼠的骨髓MSC作为对照。建立小鼠骨髓MSC与小鼠脾细胞共培养体系,流式细胞术检测脾细胞增殖情况。建立小鼠骨髓MSC与小鼠Na6ve CD4^(+)T细胞共培养体系,分别诱导Na6ve CD4^(+)T细胞向Th17细胞或调节性T细胞(Treg细胞)分化,流式细胞术检测Th17细胞或Treg细胞比例,ELISA法检测细胞上清液IL-17A的水平。结果 体外分离培养的骨髓MSC表达干细胞抗原1(Sca-1)、CD105、CD44和CD73,不表达CD11b、CD34、CD45和CD31。共培养实验结果显示正常小鼠与CIA小鼠骨髓MSC在抑制脾细胞增殖方面无统计学差异(P>0.05)。与正常小鼠骨髓MSC相比,CIA小鼠骨髓MSC同样可以抑制Th17细胞分化,减少IL-17A的分泌(P>0.05)。体外实验进一步表明,正常小鼠与CIA小鼠骨髓MSC在促进Treg细胞分化方面无统计学差异(P>0.05)。结论 RA动物模型自体骨髓MSC在体外抑制淋巴细胞增殖、调节Th17细胞或Treg细胞分化等方面无明显异常,骨髓MSC的免疫调节功能可能与RA体内的炎症环境密切相关。

Objective To explore the immune regulatory function in vitro of autologous bone marrow mesenchymal stem cells(MSCs) in the mouse model with rheumatoid arthritis(RA).Methods The mouse model with collagen-induced arthritis(CIA) was constructed.Bone marrow MSCs were isolated and cultured in vitro,and their cell phenotypes were identified.At the same time, the bone marrow MSCs from normal mice were used as the controls.The co-culture system of mouse bone marrow MSCs and mouse spleen cells was established, and the proliferation of spleen cells was detected by flow cytometry.The co-culture system of mouse bone marrow MSCs and mouse Na6 ve CD4^(+)T cells was established to induce T cells to differentiate into Th17 or Treg cells, respectively.The proportion of Th17 or Treg cells was detected by flow cytometry, and the level of IL-17 A in cell supernatant was detected by ELISA.Results Bone marrow MSCs expressed stem cell antigen-1,CD105,CD44 and CD73,but did not express CD11 b, CD34,CD45 and CD31.There was no significant difference in the inhibition of splenocyte proliferation between normal MSCs and CIA MSCs(P>0.05).Compared with normal MSCs, CIA MSCs could also inhibit the differentiation of Th17 cells and reduce the secretion of IL-17 A(P>0.05).In vitro experiments further showed that there was no significant difference in promoting Treg differentiation between normal MSCs and CIA MSCs(P>0.05).Conclusion CIA autologous bone marrow MSCs have no significant abnormalities in inhibiting lymphocyte proliferation and regulating Th17 or Treg differentiation in vitro.The immunomodulatory function of bone marrow MSCs may be closely related to the inflammatory environment in vivo in RA.