Mesenchymal stromal cells inhibit Th17 but not regulatory T-cell differentiation
Introduction
Mesenchymal stromal cells (MSC), also called mesenchymal stem cells (1,2), have been shown to have immunomodulatory functions (1., 2., 3., 4., 5.). Although cell–cell contact may be required for full immunomodulation (6,7), we and many other groups have suggested that soluble factors are important for this function (7., 8., 9.). Such soluble factors include transforming growth factor (TGF-β, hepatocyto growth factor (HGF), indoleamine-2,3-dioxygenase (IDO) and prostaglandin E2 (PGE2) (8,10,11). MSC have been reported to be the source of IDO and PGE2 (10,11). We have previously reported that nitric oxide (NO), which is produced by MSC and suppresses T-cell proliferation, is a novel regulator of immunomodulation by MSC (7). We have also demonstrated that a combination of interferon (IFN)-γ and lipopolysaccharide (LPS) or tumor necrosis factor (TNF)-α enables MSC to produce NO (12). Moreover, we have demonstrated that IFN-γ is crucial for NO production (12).
Cell therapy with MSC has been proposed as a salvage therapy for steroid-refractory severe, acute graft-versus-host disease (GvHD) (13), which is a major complication following hematopoietic stem cell transplantation (14,15), sometimes with a fatal outcome. Indeed, MSC therapy induced a high response rate in a phase II study conducted by the European Group for Blood and Marrow Transplantation (EBMT) (16). However, the molecular mechanisms underlying the MSC effect are largely unknown (3,17). GvHD may involve T helper 17 cells (Th17) and regulatory T cells (Treg), which are subtypes of CD4 T cells that have been identified fairly recently (18., 19., 20., 21., 22., 23., 24.) and which can be induced in the presence of TGF-β and interleukin (IL)-6 (25., 26., 27.) or TGF-β and IL-2 (28., 29., 30.), respectively. Treg have been reported to ameliorate mouse GvHD (31), and the induction of Treg following MSC treatment has been reported (11,32., 33., 34.). A role for IL-17 (also referred to as IL-17A) in GvHD has been controversial (35., 36., 37.). Yi et al. (35) reported that the lack of IL-17 promotes GvHD, suggesting a suppressive effect of IL-17. However, Kappel et al. (36) reported that IL-17−/− CD4 T cells ameliorate very early GvHD, suggesting a promoting effect of IL-17. Carlson et al. (37) reported that ex vivo differentiated Th17 cells induce organ-specific, skin and lung GvHD. As a reciprocal relationship is known to exist between Th17 and Treg differentiation (27), it is of great interest, in terms of future GvHD treatment, to determine the effect of MSC on Th17 and Treg differentiation and to understand the molecular mechanisms of MSC immunomodulation.
Section snippets
Preparation of MSC and CD4 T cells
C57BL/6 mice were purchased from Clea Japan (Tokyo, Japan). All mice were housed in our mouse facility (Jichi Medical University, Tochigi, Japan), which is regulated by an intramural small animal committee, and were treated in accordance with the Jichi Medical University guidelines. MSC were harvested from mouse bone marrow as described previously (7). Briefly, femurs and tibias were excised and bone marrow cells were flushed out with a small amount of phosphate-buffered saline (PBS) using a
MSC inhibit Th17 but not Treg differentiation
We first assayed the effect of MSC on the differentiation of Th17 and Treg cells. MSC were prepared from mouse bone marrow, cultured in vitro, and characterized as described previously (7). In the absence of MSC, and in the presence of TGF-β and IL-6, CD4 T cells purified from mouse spleen (Figure 1A, upper left panel) as well as naive CD4+ CD25− CD62Lhigh CD44low T cells (Figure 1A, middle left panel) differentiated into Th17 cells, which are CD4 cells that produce IL-17 (also referred to as
Discussion
In this report we provide evidence indicating that MSC inhibit Th17, but not Treg, differentiation. We also show that, although NO is an important mediator of MSC effects under neutral and Th1 conditions (7,12), NO is not produced by MSC under Th17 conditions. Our results demonstrated that PGE2 and IDO, but not NO, may play a role in the inhibition of Th17 differentiation by MSC.
It is difficult to dissociate the effect of MSC on differentiation from its effects on proliferation, because T-cell
Acknowledgments
This work was supported in part by grants from the Ministry of Health, Labor and Welfare of Japan, and by Grants-in-Aid for Scientific Research from the Ministry of Education, Culture, Sports, Science and Technology of Japan.
Disclosure of potential conflicts of interest: The authors indicate no potential conflicts of interest.
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