IL-15-dependent CD8+ CD122+ T cells ameliorate experimental autoimmune encephalomyelitis by modulating IL-17 production by CD4+ T cells.
ABSTRACT: Interleukin-15 (IL-15) is an inflammatory cytokine whose role in autoimmune diseases has not been fully elucidated. Th17 cells have been shown to play critical roles in experimental autoimmune encephalomyelitis (EAE) models. In this study, we demonstrate that blockade of IL-15 signaling by TM?-1 mAb treatment aggravated EAE severity. The key mechanism was not NK-cell depletion but depletion of CD8+ CD122+ T cells. Adoptive transfer of exogenous CD8+ CD122+ T cells to TM?-1-treated mice rescued animals from severe disease. Moreover, transfer of preactivated CD8+ CD122+ T cells prevented EAE development and significantly reduced IL-17 secretion. Naïve effector CD4+ CD25- T cells cultured with either CD8+ CD122+ T cells from wild-type mice or IL-15 transgenic mice displayed lower frequencies of IL-17A production with lower amounts of IL-17 in the supernatants when compared with production by effector CD4+ CD25- T cells cultured alone. Addition of a neutralizing antibody to IL-10 led to recovery of IL-17A production in Th17 cultures. Furthermore, coculture of CD8+ CD122+ T cells with effector CD4+ T cells inhibited their proliferation significantly, suggesting a regulatory function for IL-15 dependent CD8+ CD122+ T cells. Taken together, these observations suggest that IL-15, acting through CD8+ CD122+ T cells, has a negative regulatory role in reducing IL-17 production and Th17-mediated EAE inflammation.
Project description:IL-15 is an important IL-2-related cytokine whose role in Th17 cell biology has not been fully elucidated. In this study, we show that exogenous IL-15 decreased IL-17A production in Th17 cultures. Neutralization of IL-15 using an Ab led to increases in IL-17A production in Th17 cultures. Both Il15(-/-) and Il15r(-/-) T cell cultures displayed higher frequency of IL-17A producers and higher amounts of IL-17A in the supernatants compared with those of wild-type (WT) cells in vitro. IL-15 down-modulated IL-17A production independently of retinoic acid-related orphan receptor-?t, Foxp3, and IFN-? expression. Both Th17 cells and APCs produced IL-15, which induced binding of STAT5, an apparent repressor to the Il17 locus in CD4 T cells. Also, in a model of myelin oligodendrocyte glycoprotein-induced experimental autoimmune encephalomyelitis (EAE), Il15(-/-) mice displayed exacerbated inflammation-correlating with increased IL-17A production by their CD4(+) T cells-compared with WT controls. Exogenous IL-15 administration and IL-17A neutralization reduced the severity of EAE in Il15(-/-) mice. Taken together, these data indicate that IL-15 has a negative regulatory role in fine-tuning of IL-17A production and Th17-mediated inflammation.
Project description:IL-17-producing CD8+ T (Tc17) cells are detectible in multiple sclerosis (MS) lesions; however, their contribution to the disease is unknown. To identify functions of Tc17 cells, we induced EAE, a murine model of MS, in mice lacking IFN regulatory factor 4 (IRF4). IRF4-deficient mice failed to generate Tc17 and Th17 cells and were resistant to EAE. After adoptive transfer of WT CD8+ T cells and subsequent immunization for EAE induction in these mice, the CD8+ T cells developed a Tc17 phenotype in the periphery but could not infiltrate the CNS. Similarly, transfer of small numbers of WT CD4+ T cells alone did not evoke EAE, but when transferred together with CD8+ T cells, IL-17-producing CD4+ (Th17) T cells accumulated in the CNS and mice developed severe disease. Th17 accumulation and development of EAE required IL-17A production by CD8+ T cells, suggesting that Tc17 cells are required to promote CD4+ T cell-mediated induction of EAE. Accordingly, patients with early-stage MS harbored a greater number of Tc17 cells in the cerebrospinal fluid than in peripheral blood. Our results reveal that Tc17 cells contribute to the initiation of CNS autoimmunity in mice and humans by supporting Th17 cell pathogenicity.
Project description:Mesenchymal stem cells (MSC) have emerged as a promising candidate for inflammatory suppression and disease amelioration, especially of neuro-inflammatory diseases such as multiple sclerosis (MS). Auto-reactive CD4+ and CD8+ T cells acquire pathogenic IFN?-producing- (Type I) and IL-17A-producing- (Type 17) effector phenotypes in MS and its animal model experimental autoimmune encephalomyelitis (EAE). Although MSC have been extensively demonstrated to suppress pathogenic effector CD4+ T cells and CD4+ T cell-mediated EAE, surprisingly few studies have addressed their modulation of effector CD8+ T cells represented in MS or their impact on CD8+ T cell-mediated EAE. We find that MSC differentially modulate CD8+ T cell development depending on effector T cell subtype. MSC drive activated low-IFN? producers toward an enhanced high-IFN? Tc1-like phenotype but strongly inhibit the production of IL-17A and Tc17 polarization in vitro. These observations are underscored by differential MSC modulation of T cell activation, proliferation, and signature transcription factor up-regulation. In addition, effector CD8+ T cells co-cultured with MSC exhibited increased production of IL-2, a molecule known to enhance IFN?, yet suppress IL-17A, production. Based on these in vitro effects on CD8+ T cells, we next evaluated their impact on the severity of EAE. To better evaluate CD8+ T cells, we immunized mice with MOG37-50 , which is a CD8-targeted epitope. Our results revealed a worsening of disease, consistent with their in vitro stimulation of Tc1 cells. These findings highlight the emerging duality of MSC in immune modulation and provide implications for their future use in immune-related diseases.
Project description:Regulatory CD8(+) T cells are critical for self-tolerance and restricting excessive immune responses. The variety of immune functions they fulfill, the heterogeneity of their phenotype, and the mechanism of action are still poorly understood. Here we describe that regulatory CD8(+) T cells exhibiting immunosuppressive actions in vitro and in vivo are recognized as CD38(high) T cells and present in naive mice. CD38 is a glycosylated membrane protein with ectonucleotidase properties. CD8(+)CD38(high) (CD44(+)CD122(+)CD62L(high)) lymphocytes suppress CD4(+) effector T-cell proliferation in an antigen-non specific manner via IFN-?. While direct cell-to-cell contact is needed for this suppressor activity, it is independent of membrane-bound TGF-? and granzyme B release. IL-15 potentiates the suppressive activity of CD8(+)CD38(high) T cells and controls their survival and expansion. In humans CD8(+)CD38(high) T cells inhibit CD4(+) effector T cell proliferation. In vivo, CD8(+)CD38(high), but not CD8(+)CD38(-) T cells mitigate murine experimental autoimmune encephalomyelitis (EAE) by reducing the clinical score and delaying disease occurrence. EAE suppression is enhanced by pre-treatment of CD8(+)CD38(high) T cells with IL-15. These findings add evidence that the expression of ectoenzyme receptor family members positively correlates with suppressor functions and identifies CD8(+)CD38(high) T cells as potential inhibitors of excessive immune responses.
Project description:NFATc1 is a member of the nuclear factor of activated T cells (NFAT) family of transcription factors. NFAT is activated upon T-cell receptor activation followed by intracytoplasmatic calcium influx where calmodulin, a calcium sensor protein, activates the phosphatase calcineurin that dephosphorylates NFAT proteins and results in NFAT nuclear import. Here, we show the analysis of conditional NFATc1-deficient mice bearing a deletion of NFATc1 in CD4(+) and CD8(+) T cells. NFATc1-deficient CD4(+) T cells polarized under Th17 conditions express reduced levels of the Th17-associated transcription factor RORγT (where ROR is RAR-related orphan receptor) as well as the Th17-associated cytokines IL-17A, IL-17F, IL-21, and IL-10. In the murine model of experimental EAE, we found a strong reduction of the disease outcome in conditional NFATc1-deficient mice, as compared with control littermates. This was accompanied by a diminished inflammation in the brain and spinal cord and reduced IL-17A and IFN-γ expression by antigen-specific spleen, spinal cord, and brain cells. Altogether, these results reveal an important role of NFATc1 in inducing Th17-cell responses and IFN-γ, both being relevant for the EAE development.
Project description:Nuclear factor of activated T cells (NFATs) is an important transcription factor for T cell activation and proliferation. Recent studies have highlighted the role of NFATs in regulating the differentiation of effector CD4 T helper (Th) subsets including Th1 and Th17 cells. Because controlling the effector T cell function is important for the treatment of autoimmune diseases, regulation of NFAT functions in T cells would be an important strategy to control the pathogenesis of autoimmune diseases. Here, we demonstrated that an NFAT inhibitory peptide, VIVIT conjugated to dNP2 (dNP2-VIVIT), a blood-brain barrier-permeable peptide, ameliorated experimental autoimmune encephalomyelitis (EAE) by inhibiting Th1 and Th17 cells, but not regulatory T (Treg) cells. dNP2-VIVIT negatively regulated spinal cord-infiltrating interleukin-17A (IL-17A) and interferon (IFN)-?-producing CD4+ T cells without affecting the number of Foxp3+ CD4+ Treg cells, whereas dNP2-VEET or 11R-VIVIT could not significantly inhibit EAE. In comparison with cyclosporin A (CsA), dNP2-VIVIT selectively inhibited Th1 and Th17 differentiation, whereas CsA inhibited the differentiation of all T cell subsets including that of Th2 and Treg cells. Collectively, this study demonstrated the role of dNP2-VIVIT as a novel agent for the treatment of autoimmune diseases such as multiple sclerosis by regulating the functions of Th1 and Th17 cells.
Project description:To assess potential roles of effector cells and immunologic markers in demyelinating CNS lesion formation, and their modulation by interferon ?-1a (IFN-?-1a).Twenty-three patients with relapsing-remitting multiple sclerosis (RRMS) received IFN-?-1a for 6 months. Immunologic marker results were correlated with brain MRI lesion volumes, and volumes of normal-appearing brain tissue (NABT) with decreasing or increasing voxel-wise magnetization transfer ratio (VW-MTR), suggestive of demyelination and remyelination, respectively.Baseline expression of Th22 cell transcription factor aryl hydrocarbon receptor (AHR) and interleukin (IL)-17F, and percentages of IL-22-expressing CD4(+) and CD8(+) cells, were significantly higher in patients vs 15 healthy controls; IL-4 in CD4(+) cells was lower. Baseline percentage of IL-22-producing CD8(+) cells positively correlated with T2 lesion volumes, while percentage of IL-17A-producing CD8(+) cells positively correlated with T2 and T1 lesion volumes. IFN-?-1a induced reductions in transcription factor AHR, T-bet, and retinoic acid-related orphan nuclear hormone receptor C (RORc) gene expression, while it increased GATA3's expression in CD4(+) cells. Percentages of IL-22-, IL-17A-, and IL-17F-expressing T cells significantly decreased following treatment. Increased percentages of IL-10-expressing CD4(+) and CD8(+) cells correlated with greater NABT volume with increasing VW-MTR, while decreased percentage of IL-17F-expressing CD4(+) cells positively correlated with decreased NABT volume with decreasing VW-MTR.Findings indicate that IFN-?-1a suppresses Th22 and Th17 cell responses, which were associated with decreased MRI-detectable demyelination.This pilot study provides Class III evidence that reduced Th22 and Th17 responses are associated with decreased demyelination following IFN-?-1a treatment in patients with RRMS.
Project description:Multiple sclerosis (MS) is an inflammatory, demyelinating disease of the CNS mediated by self-reactive, myelin-specific T cells. Both CD4(+) and CD8(+) T cells play important roles in the pathogenesis of MS. MS is studied using experimental autoimmune encephalomyelitis (EAE), an animal model mediated by myelin-specific T cells. T cell Ig mucin-3 (Tim-3) is a cell surface receptor expressed on CD4(+) IFN-?-secreting Th1 cells, and triggering Tim-3 signaling ameliorated EAE by inducing death in pathogenic Th1 cells in vivo. This suggested that enhancing Tim-3 signaling might be beneficial in patients with MS. However, Tim-3 is also expressed on activated CD8(+) T cells, microglia, and dendritic cells, and the combined effect of manipulating Tim-3 signaling on these cell types during CNS autoimmunity is unknown. Furthermore, CD4(+) IL-17-secreting Th17 cells also play a role in MS, but do not express high levels of Tim-3. We investigated Tim-3 signaling in EAE models that include myelin-specific Th17, Th1, and CD8(+) T cells. We found that preventing Tim-3 signaling in CD4(+) T cells altered the inflammatory pattern in the CNS due to differential effects on Th1 versus Th17 cells. In contrast, preventing Tim-3 signaling during CD8(+) T cell-mediated EAE exacerbated disease. We also analyzed the importance of Tim-3 signaling in EAE in innate immune cells. Tim-3 signaling in dendritic cells and microglia did not affect the manifestation of EAE in these models. These results indicate that the therapeutic efficacy of targeting Tim-3 in EAE is dependent on the nature of the effector T cells contributing to the disease.
Project description:The transcription factor Tbet is critical for the differentiation of Th1 CD4 T cells and is associated with the induction of multiple autoimmune diseases, including experimental autoimmune encephalomyelitis (EAE). Herein, we demonstrate that Tbet suppresses IL-17A and Th17 differentiation both in vitro and in vivo in a cell-intrinsic manner, and that in fact, Tbet is not necessary for EAE induction. Moreover, we find that IFN? inhibits the production of IL-17A and IL-17F in a STAT1-dependent, Tbet-independent manner. These findings illustrate multiple mechanisms utilized by developing Th1 cells to silence the Th17 program.
Project description:Myeloid-derived suppressor cells (MDSCs) have been a focus of recent study on tumor-mediated immune suppression. However, its role in Th17 cell differentiation and the pathogenesis of autoimmune diseases (e.g., multiple sclerosis) has not been determined. We show in this study that development of experimental autoimmune encephalomyelitis (EAE) in mice is associated with a profound expansion of CD11b(+)Gr-1(+) MDSCs, which display efficient T cell inhibitory functions in vitro. Unexpectedly, these MDSCs enhance the differentiation of naive CD4(+) T cell precursors into Th17 cells in a highly efficient manner under Th17-polarizing conditions, as indicated by significantly increased number of Th17 cells, elevation of IL-17A production, and upregulation of the orphan nuclear receptor RORA and RORC. Mechanistic studies show that IL-1? represents a major mediator of MDSC-facilitated Th17 differentiation, which depends on the IL-1 receptor on CD4(+) T cells but not MDSCs. Selective depletion of MDSCs using gemcitabine results in a marked reduction in the severity of EAE (e.g., decreased clinical scores and myelin injury), which correlates with reduced Th17 cells and inflammatory cytokines (IL-17A and IL-1?) in the lymphoid tissues and spinal cord. Adoptive transfer of MDSCs after gemcitabine treatment restores EAE disease progression. Together, we demonstrate for the first time, to our knowledge, that excessive and prolonged presence of MDSCs can drive a Th17 response and consequently contributes to the pathogenesis of EAE. These new findings provide unique insights into the pleiotropic functions of MDSCs and may help explain the failure of immunosuppressive MDSCs to control Th17/IL-17-dependent autoimmune disorders.