Investigation for immunomodulatory and anti-inflammatory mechanisms of mesenchymal stem cells on T cells and dendritic cells in CIA mice

의과대학/박사Background Mesenchymal stem cells (MSCs) have profound immunomodulatory properties. Using their properties, MSCs-based therapies have been applied in several inflammatory diseases. The immune modulation of MSCs is related to inhibition of immune cell proliferation and activation. Dendritic cells (DCs) play pivotal roles in initiating immune response as DCs are important antigen-presenting cells. DCs are able to mature into inflammatory DCs, sustaining a continuous activation of adaptive immune system. T cells critical effector immune cells in affecting and regulating immune response and can differentiate into one of several subtypes, including TH1, TH2, TH17 or regulatory T cells (Tregs). As the effects of MSCs on DCs and T cells have not been fully understood, the immunomodulatory mechanisms of MSCs on DCs and T cells were investigated. Methods Bone marrow-derived CD11c+ mononuclear cells of DBA/1 mice were differentiated cultured with GM-CSF and IL-4, to immature DCs (iDCs) for 7 days. CD4+ T cells were isolated from spleen of DBA/1 mice by negative selection using CD4+ T cell isolation kit. The iDCs were maturated using lipopolysaccharides (LPS). We co-cultured human bone marrow-derived MSCs (BM-MSCs) and mouse DCs and CD4+ T cells directly with different ratios. The effect of BM-MSCs on DC maturation and T cell differentiation was assessed by maturation markers by flow cytometry, and supernatants for induced production of cytokines. Gene expressions were analyzed by quantitative RT-PCR and microarray. In addition, immunohistochemistry was performed in inflammatory tissues in collagen-induced arthritis (CIA) mice. Results Human BM-MSCs significantly inhibited the DCs maturation by decreasing CD86 and major histocompatibility complex (MHC)Ⅱ expressions in both 1:1 and 1:10 co-culture ratio. And, human BM-MSCs decreased the levels of interleukin (IL)-12p70, tumor necrosis factor (TNF)-α, and IL-6 effectively, and increased the levels of IL-10 and transforming growth factor (TGF)-β. Also, the relative gene expressions of indoleamine 2,3-dioxygenase (IDO) and heme oxygenase (HO)-1 were increased in 1:1 co-culture ratio at 6 hr. The expression of CD4+CD25+FoxP3+ Tregs were highly induced in co-culture condition with hBM-MSCs. Human BM-MSCs significantly induced the Tregs by increasing programmed death-1 (PD-1) and neuropilin-1 (Nrp-1) expressions in different co-culture ratio. And, human BM-MSCs increased the levels of IL-10 and TGF-β1. Moreover, immunohistochemical analysis of inflamed tissues in BM-MSC treated CIA mice showed significant immunopositive staining for PD-1 on CD4+ T cells. Conclusion The results demonstrated that human BM-MSCs inhibited the maturation of DCs and induced anti-inflammatory signals of DCs in in vitro. Moreover, these data showed that human BM-MSCs highly induced the CD4+CD25+Foxp3+ Tregs in both in vitro and in vivo.ope