WanJun Chen

Conversion of Peripheral CD4+CD25− Naive T Cells to CD4+CD25+ Regulatory T Cells by TGF-β Induction of Transcription Factor Foxp3

CD4+CD25+ regulatory T cells (Treg) are instrumental in the maintenance of immunological tolerance. One critical question is whether Treg can only be generated in the thymus or can differentiate from peripheral CD4+CD25− naive T cells. In this paper, we present novel evidence that conversion of naive peripheral CD4+CD25− T cells into anergic/suppressor cells that are CD25+, CD45RB−/low and intracellular CTLA-4+ can be achieved through costimulation with T cell receptors (TCRs) and transforming growth factor β (TGF-β). Although transcription factor Foxp3 has been shown recently to be associated with the development of Treg, the physiological inducers for Foxp3 gene expression remain a mystery. TGF-β induced Foxp3 gene expression in TCR-challenged CD4+CD25− naive T cells, which mediated their transition toward a regulatory T cell phenotype with potent immunosuppressive potential. These converted anergic/suppressor cells are not only unresponsive to TCR stimulation and produce neither T helper cell 1 nor T helper cell 2 cytokines but they also express TGF-β and inhibit normal T cell proliferation in vitro. More importantly, in an ovalbumin peptide TCR transgenic adoptive transfer model, TGF-β–converted transgenic CD4+CD25+ suppressor cells proliferated in response to immunization and inhibited antigen-specific naive CD4+ T cell expansion in vivo. Finally, in a murine asthma model, coadministration of these TGF-β–induced suppressor T cells prevented house dust mite–induced allergic pathogenesis in lungs.

Generation of pathogenic T(H)17 cells in the absence of TGF-β signalling.

CD4+ T-helper cells that selectively produce interleukin (IL)-17 (TH17), are critical for host defence and autoimmunity. Although crucial for TH17 cells in vivo, IL-23 has been thought to be incapable of driving initial differentiation. Rather, IL-6 and transforming growth factor (TGF)-β1 have been proposed to be the factors responsible for initiating specification. Here we show that TH17 differentiation can occur in the absence of TGF-β signalling. Neither IL-6 nor IL-23 alone efficiently generated TH17 cells; however, these cytokines in combination with IL-1β effectively induced IL-17 production in naive precursors, independently of TGF-β. Epigenetic modification of the Il17a, Il17f and Rorc promoters proceeded without TGF-β1, allowing the generation of cells that co-expressed RORγt (encoded by Rorc) and T-bet. T-bet+RORγt+ TH17 cells are generated in vivo during experimental allergic encephalomyelitis, and adoptively transferred TH17 cells generated with IL-23 without TGF-β1 were pathogenic in this disease model. These data indicate an alternative mode for TH17 differentiation. Consistent with genetic data linking IL23R with autoimmunity, our findings re-emphasize the importance of IL-23 and therefore may have therapeutic implications.

Transforming Growth Factor-β Production and Myeloid Cells Are an Effector Mechanism through Which CD1d-restricted T Cells Block Cytotoxic T Lymphocyte–mediated Tumor Immunosurveillance: Abrogation Prevents Tumor Recurrence

Our previous work demonstrated that cytotoxic T lymphocyte (CTL)-mediated tumor immunosurveillance of the 15-12RM tumor could be suppressed by a CD1d-restricted lymphocyte, most likely a natural killer (NK) T cell, which produces interleukin (IL)-13. Here we present evidence for the effector elements in this suppressive pathway. T cell–reconstituted recombination activating gene (RAG)2 knockout (KO) and RAG2/IL-4 receptor α double KO mice showed that inhibition of immunosurveillance requires IL-13 responsiveness by a non–T non–B cell. Such nonlymphoid splenocytes from tumor-bearing mice produced more transforming growth factor (TGF)-β, a potent inhibitor of CTL, ex vivo than such cells from naive mice, and this TGF-β production was dependent on the presence in vivo of both IL-13 and CD1d-restricted T cells. Ex vivo TGF-β production was also abrogated by depleting either CD11b+ or Gr-1+ cells from the nonlymphoid cells of tumor-bearing mice. Further, blocking TGF-β or depleting Gr-1+ cells in vivo prevented the tumor recurrence, implying that TGF-β made by a CD11b+ Gr-1+ myeloid cell, in an IL-13 and CD1d-restricted T cell–dependent mechanism, is necessary for down-regulation of tumor immunosurveillance. Identification of this stepwise regulation of immunosurveillance, involving CD1-restricted T cells, IL-13, myeloid cells, and TGF-β, explains previous observations on myeloid suppressor cells or TGF-β and provides insights for targeted approaches for cancer immunotherapy, including synergistic blockade of TGF-β and IL-13.

TGF-β Released by Apoptotic T Cells Contributes to an Immunosuppressive Milieu

Abstract T cell apoptosis is critical to development and homeostasis of the immune system. The most salient feature of apoptosis is the lack of an attendant inflammatory response or tissue damage. Here, we present evidence that apoptotic T cells release TGF-β, thereby contributing to an immunosuppressive milieu. Apoptotic T cells released not only latent but also bio-active TGF-β. Nonetheless, TGF-β transcription was not upregulated, suggesting release of existing rather than synthesis of new TGF-β. Localized within the intracellular membrane-bound compartment, which includes mitochondria, TGF-β was redistributed into the cytosol following loss of mitochondrial membrane potential. TGF-β secreted from apoptotic T cells inhibited proinflammatory cytokine production by activated macrophages to foster immune suppression. These findings broaden the potential mechanisms whereby induction of immune tolerance or deficiency occurs through T cell deletion.

Mesenchymal stem cell–based tissue regeneration is governed by recipient T lymphocytes via IFN-γ and TNF-α

Stem cell–based regenerative medicine is a promising approach in tissue reconstruction. Here we show that proinflammatory T cells inhibit the ability of exogenously added bone marrow mesenchymal stem cells (BMMSCs) to mediate bone repair. This inhibition is due to interferon γ (IFN-γ)–induced downregulation of the runt-related transcription factor 2 (Runx-2) pathway and enhancement of tumor necrosis factor α (TNF-α) signaling in the stem cells. We also found that, through inhibition of nuclear factor κB (NF-κB), TNF-α converts the signaling of the IFN-γ–activated, nonapoptotic form of TNF receptor superfamily member 6 (Fas) in BMMSCs to a caspase 3– and caspase 8–associated proapoptotic cascade, resulting in the apoptosis of these cells. Conversely, reduction of IFN-γ and TNF-α concentrations by systemic infusion of Foxp3+ regulatory T cells, or by local administration of aspirin, markedly improved BMMSC-based bone regeneration and calvarial defect repair in C57BL/6 mice. These data collectively show a previously unrecognized role of recipient T cells in BMMSC-based tissue engineering.

Mesenchymal-stem-cell-induced immunoregulation involves FAS-ligand-/FAS-mediated T cell apoptosis

Systemic infusion of bone marrow mesenchymal stem cells (BMMSCs) yields therapeutic benefit for a variety of autoimmune diseases, but the underlying mechanisms are poorly understood. Here we show that in mice systemic infusion of BMMSCs induced transient T cell apoptosis via the FAS ligand (FASL)-dependent FAS pathway and could ameliorate disease phenotypes in fibrillin-1 mutated systemic sclerosis (SS) and dextran-sulfate-sodium-induced experimental colitis. FASL(-/-) BMMSCs did not induce T cell apoptosis in recipients, and could not ameliorate SS and colitis. Mechanistic analysis revealed that FAS-regulated monocyte chemotactic protein 1 (MCP-1) secretion by BMMSCs recruited T cells for FASL-mediated apoptosis. The apoptotic T cells subsequently triggered macrophages to produce high levels of TGFβ, which in turn led to the upregulation of CD4(+)CD25(+)Foxp3(+) regulatory T cells and, ultimately, immune tolerance. These data therefore demonstrate a previously unrecognized mechanism underlying BMMSC-based immunotherapy involving coupling via FAS/FASL to induce T cell apoptosis.

Matriptase/MT-SP1 is required for postnatal survival, epidermal barrier function, hair follicle development, and thymic homeostasis

Matriptase/MT-SP1 is a novel tumor-associated type II transmembrane serine protease that is highly expressed in the epidermis, thymic stroma, and other epithelia. A null mutation was introduced into the Matriptase/MT-SP1 gene of mice to determine the role of Matriptase/MT-SP1 in epidermal development and neoplasia. Matriptase/MT-SP1-deficient mice developed to term but uniformly died within 48 h of birth. All epidermal surfaces of newborn mice were grossly abnormal with a dry, red, shiny, and wrinkled appearance. Matriptase/MT-SP1-deficiency caused striking malformations of the stratum corneum, characterized by dysmorphic and pleomorphic corneocytes and the absence of vesicular bodies in transitional layer cells. This aberrant skin development seriously compromised both inward and outward epidermal barrier function, leading to the rapid and fatal dehydration of Matriptase/MT-SP1-deficient pups. Loss of Matriptase/MT-SP1 also seriously affected hair follicle development resulting in generalized follicular hypoplasia, absence of erupted vibrissae, lack of vibrissal hair canal formation, ingrown vibrissae, and wholesale abortion of vibrissal follicles. Furthermore, Matriptase/MT-SP1-deficiency resulted in dramatically increased thymocyte apoptosis, and depletion of thymocytes. This study demonstrates that Matriptase/MT-SP1 has pleiotropic functions in the development of the epidermis, hair follicles, and cellular immune system.

TGF-β: the missing link in CD4+CD25+ regulatory T cell-mediated immunosuppression

Abstract A unique population of CD4 + T lymphocytes that constitutively express CD25 has been recognized as anergic/suppressor cells. While the immunosuppressive activity of these CD4 + CD25 + cells has been validated and implicated in tolerance, autoimmunity, transplantation, cancer and infectious diseases, the mechanism(s) by which they function still remains controversial. Although the involvement of TGF-β was initially discounted, emerging evidence now links this cytokine with CD4 + CD25 + T cell-mediated suppression of antigen-activated T cells. In this perspective, we summarize recently published studies, as well as our own data, which shed light on how cell membrane-bound TGF-β can deliver a regulatory signal to target cells via a contact-dependent process. Moreover, suppressor T cell function is a complex process, tightly regulated by multiple factors, including IL-2, cytotoxic T lymphocyte-associated antigen-4 (CTLA-4) and glucocorticoid induced TNF receptor (GITR). Collectively, multiple previously unconnected puzzle pieces are beginning to be linked into a more coherent, albeit incomplete picture of CD4 + CD25 + T cell-mediated suppression.

CD3-specific antibody–induced immune tolerance involves transforming growth factor-β from phagocytes digesting apoptotic T cells

Intact CD3-specific antibody transiently depletes large numbers of T cells and subsequently induces long-term immune tolerance. The underlying mechanisms for the systemic tolerance, however, remain unclear. We show here that treatment of normal mice with intact antibody to CD3 increases systemic transforming growth factor-β (TGF-β) produced by phagocytes exposed to apoptotic T cells. Among the phagocytes, macrophages and immature dendritic cells (iDCs) secrete TGF-β upon ingestion of apoptotic T cells, which induces CD4+Foxp3+ regulatory T cells in culture and contributes to immune tolerance mediated by CD3-specific antibody in vivo. In accordance with these results, depletion of macrophages and iDCs not only abrogates CD3-specific antibody–mediated prevention of myelin oligodendrocyte glycoprotein–induced acute experimental autoimmune encephalomyelitis (EAE), but also reverses the therapeutic effects of antibody to CD3 on established disease in a model of relapsing-remitting EAE. Thus, CD3-specific antibody–induced immune tolerance is associated with TGF-β production in phagocytes involved in clearing apoptotic T cells, which suggests that apoptosis is linked to active suppression in immune tolerance.

TGF-β: the perpetrator of immune suppression by regulatory T cells and suicidal T cells

Innate and adaptive immunity function to eliminate foreign invaders and respond to injury while enabling coexistence with commensal microbes and tolerance against self and innocuous agents. Although most often effective in accomplishing these objectives, immunologic processes are not fail‐safe and may underserve or be excessive in protecting the host. Checks and balances to maintain control of the immune system are in place and are becoming increasingly appreciated as targets for manipulating immunopathologic responses. One of the most recognized mediators of immune regulation is the cytokine transforming growth factor‐β (TGF‐β), a product of immune and nonimmune cells. Emerging data have unveiled a pivotal role for TGF‐β as a perpetrator of suppression by CD4+CD25+ regulatory T (Treg) cells and in apoptotic sequelae. Through its immunosuppressive prowess, TGF‐β effectively orchestrates resolution of inflammation and control of autoaggressive immune reactions by managing T cell anergy, defining unique populations of Treg cells, regulating T cell death, and influencing the host response to infections.