From mesenchymal stem cell therapy to discovery of drug therapy for systemic sclerosis

Bone marrow mesenchymal stem cells (BMMSCs) are non-hematopoietic multipotent stem cells capable of differentiating into both mesenchymal and non-mesenchymal cell types. In addition to generate bone structure to replace damaged and diseased tissues, preclinical and clinical studies have shown that BMMSCs display profound immunomodulatory functions via inhibiting the proliferation and function of several major immune cells such as T lymphocytes, B lymphocytes, natural killer (NK), and dendritic cells. Thus, systemic infusion of BMMSCs has been used to treat a variety of diseases, including acute graft-versus-host-disease (GVHD), ameliorating HSC engraftment, systemic lupus erythematosus (SLE), intestinal and bowel disease (IBD), and sepsis. However, the detailed mechanism in which BMMSCs regulate immune function is not fully understood.;In the first part of Chapter 2 of this study, we show that systemic infusion of BMMSCs induces a transient T cell apoptosis via the Fas Ligand (FasL)-mediated Fas pathway and ameliorates diseased phenotypes in fibrillin-1 mutated systemic sclerosis (SSc) and dextran sulfate sodium-induced experimental colitis mice. The therapeutic mechanism of BMMSC infusion is associated with phagocytosis of apoptotic T cell debris, leading to a high level of macrophage-mediated transforming growth factor beta (TGF-beta) production and a subsequent immune tolerance. Importantly, we provided clinical evidence to show that MSC infusion in SS patients resulted in a T cell apoptosis and up-regulation of Tregs. Additionally, we revealed that Fas null BMMSCs, with normal FasL function, failed to induce T cell apoptosis and offer therapeutic effect for SS and colitis mice. Mechanistic study showed that Fas governed monocyte chemotactic protein 1 (MCP-1) secretion in BMMSCs, which plays a crucial role in the recruitment of T cells to BMMSCs for FasL-mediated apoptosis. In summary, BMMSCs use Fas to control MCP-1 secretion for the recruitment of T cells and subsequently use FasL to induce activated T cell apoptosis. Macrophages take debris of apoptotic T cells to release a high level of TGF-b, leading to up-regulation of Tregs and, ultimately, immune tolerance for immunotherapies.;In second part of Chapter 2 of this study, we showed for the first time that telomerase activity is required for maintaining the immunomodulatory properties of MSCs. Telomerase deficient MSCs lose their capacity to inhibit T-cells, activate Foxp3-positive regulatory T cells (Tregs), and ameliorate disease phenotype in systemic sclerosis mice, which can be rescued by overexpression of telomerase reverse tran¬scriptase (TERT). Mechanistically, TERT combined with beta-catenin and BRG1 to form a complex to bind to FAS Ligand (FASL) promoter and upregulate FASL expression. Upregulated FASL expression can elevate MSC immunomodulation function, as shown in our recent publication (Akiyama et al., 2012). When MSCs were treated with aspirin, their immunomodulation was significantly improved due to elevated telomerase activity and the number of MSCs required to treat systemic sclerosis mice was markedly reduced.;This study has uncovered the role of telomerase in MSC-based immunotherapies and the mechanism by which TERT binds to the promoter region to upregulate FASL expression. In fact, this is the first study to link telomerase activity to immunomodulatory therapies. Overall, therefore, this study has brought forth experimental evidence for stem cell biology, the molecular mechanism(s) underlying MSC-associated immunotherapies, and pathway-guided drug therapy.;In Chapter 3, we reveal that Fbn1+/- SSc mice show osteopenia phenotype with decreased osteogenic differentiation and increased adipogenic differentiation of bone marrow MSCs by the activation of IL-4 receptor alpha (IL-4Ralpha)/mTOR signaling. We further determine that mTOR signaling blocks osteogenic differentiation via the P70S6K/RUNX2 pathway, while it elevates adipogenic differentiation via P70S6K/PPARgamma2 pathway. Since significantly elevated levels of IL4 and TGF-beta were observed in Fbn1+/- SSc mice, we reveal that upregulation of the IL-4Ralpha in Fbn1+/- MSCs is governed by TGFbeta/SMAD3/SP1 signaling via SP1 biding to the Il4ralpha promoter. Either knockdown of IL-4Ralpha or inhibition of mTOR can rescue Fbn1+/- MSC function by increasing osteogenesis and reducing adipogenesis. Additionally, we showed that conditional knockout of mTOR in MSCs/osteoblasts could ameliorate osteopenia phenotype in Fbn1+/- mice by rescuing impaired osteogenic/adipogenic differentiation.;To translate our findings to potential clinical applications, we used rapamycin treatment to inhibit mTOR signaling, thereby rescuing osteopenia phenotype in Fbn1+/- SSc mice by rescuing osteo/adipo-lineage differentiation in MSCs. This result strongly suggests that rapamycin treatment may provide an anabolic therapy for systemic sclerosis.;In summary, this study establishes the FBN1/TGFbeta/SP1/IL-4Ralpha/mTOR cascade as a key determinant of MSC lineage selection, a finding which may serve as a basis for the development of novel therapies to treat SSc. (Abstract shortened by UMI.).