ovalbumin and myelin-oligodendrocyte-glycoprotein-(35-55)

T cell development and activation are usually accompanied by expansion and production of numerous proteins that require active translation. The eukaryotic translation initiation factor 4E (eIF4E) binds to the 5' cap structure of mRNA and is critical for cap-dependent translational initiation. It has been hypothesized that MAPK-interacting kinase 1 and 2 (Mnk1/2) promote cap-dependent translation by phosphorylating eIF4E at serine 209 (S209). Pharmacologic studies using inhibitors have suggested that Mnk1/2 have important roles in T cells. However, genetic evidence supporting such conclusions is lacking. Moreover, the signaling pathways that regulate Mnk1/2 in T cells remain unclear. We demonstrate that TCR engagement activates Mnk1/2 in primary T cells. Such activation is dependent on Ras-Erk1/2 signaling and is inhibited by diacylglycerol kinases α and ζ. Mnk1/2 double deficiency in mice abolishes TCR-induced eIF4E S209 phosphorylation, indicating their absolute requirement for eIF4E S209 phosphorylation. However, Mnk1/2 double deficiency does not affect the development of conventional αβ T cells, regulatory T cells, or NKT cells. Furthermore, T cell activation, in vivo primary and memory CD8 T cell responses to microbial infection, and NKT cell cytokine production were not obviously altered by Mnk1/2 deficiency. Although Mnk1/2 deficiency causes decreased IL-17 and IFN-γ production by CD4 T cells following immunization of mice with myelin oligodendrocyte glycoprotein peptide in complete Freund's adjuvant, correlating with milder experimental autoimmune encephalomyelitis scores, it does not affect Th cell differentiation in vitro. Together, these data suggest that Mnk1/2 has a minimal role in T cell development and activation but may regulate non-T cell lineages to control Th1 and Th17 differentiation in vivo.

Topics: Adoptive Transfer; Aniline Compounds; Animals; Cells, Cultured; Encephalomyelitis, Autoimmune, Experimental; Eukaryotic Initiation Factor-4E; Female; Interferon-gamma; Interleukin-17; Listeriosis; Lymphocyte Activation; Lymphocytic Choriomeningitis; Lymphopoiesis; MAP Kinase Signaling System; Mice; Mice, Inbred C57BL; Mice, Knockout; Mice, Transgenic; Myelin-Oligodendrocyte Glycoprotein; Natural Killer T-Cells; Ovalbumin; Peptide Fragments; Phosphorylation; Protein Processing, Post-Translational; Protein Serine-Threonine Kinases; Purines; RNA Caps; T-Lymphocyte Subsets

The ability of activated B cells to protect against various experimental autoimmune or allergic diseases makes them attractive for use in cell-based therapies. We describe an efficient way to generate B cells with strong suppressive functions by incubating naive B cells with a relevant Ag conjugated to cholera toxin B subunit (CTB). This allows most B cells, irrespective of BCR, to take up and present Ag and induces their expression of latency-associated polypeptide (LAP)/TGF-β and after adoptive transfer also their production of IL-10. With OVA as model Ag, when naive T cells were cocultured in vitro with B cells pretreated with OVA conjugated to CTB (OVA/CTB) Ag-specific CD4(+) Foxp3 regulatory T (Treg) cells increased >50-fold. These cells effectively suppressed CD25(-)CD4(+) effector T (Teff) cells in secondary cultures. Adoptive transfer of OVA/CTB-treated B cells to mice subsequently immunized with OVA in CFA induced increase in Foxp3 Treg cells together with suppression and depletion of Teff cells. Likewise, adoptive transfer of B cells pulsed with myelin oligodendrocyte glycoprotein peptide(35-55) (MOGp) conjugated to CTB increased the number of Treg cells, suppressed MOGp-specific T cell proliferation and IL-17 and IFN-γ production, and prevented the development of experimental autoimmune encephalomyelitis. Similar effects were seen when B cells were given "therapeutically" to mice with early-stage experimental autoimmune encephalomyelitis. Our results suggest that B cells pulsed in vitro with relevant Ag/CTB conjugates may be used in cell therapy to induce Ag-specific suppression of autoimmune disease.

Topics: Adoptive Transfer; Animals; Antigens; B-Lymphocytes; CD4-Positive T-Lymphocytes; Cell Proliferation; Cholera Toxin; Encephalomyelitis, Autoimmune, Experimental; Female; Forkhead Transcription Factors; Glycoproteins; Immune Tolerance; Interferon-gamma; Interleukin-17; Lymphocyte Activation; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Myelin-Oligodendrocyte Glycoprotein; Ovalbumin; Peptide Fragments; Recombinant Fusion Proteins; T-Lymphocytes, Regulatory

DC are professional APC that instruct T cells during the inflammatory course of EAE. We have previously shown that MAPK3 (Erk1) is important for the induction of T-cell anergy. Our goal was to determine the influence of MAPK3 on the capacity of DC to arm T-cell responses in autoimmunity. We report that DC from Mapk3(-/-) mice have a significantly higher membrane expression of CD86 and MHC-II and--when loaded with the myelin oligodendrocyte glycoprotein--show a superior capacity to prime naïve T cells towards an inflammatory phenotype than Mapk3(+/+) DC. Nonetheless and as previously described, Mapk3(-/-) mice were only slightly but not significantly more susceptible to myelin oligodendrocyte glycoprotein-induced EAE than WT littermate mice. However, Mapk3(+/+) mice engrafted with Mapk3(-/-) BM (KO-->WT) developed a severe form of EAE, in direct contrast to WT-->KO mice, which were even less sick than control WT-->WT mice. An infiltration of DC and accumulation of Th17 cells was also observed in the CNS of KO-->WT mice. Therefore, triggering of MAPK3 in the periphery might be a therapeutic option for the treatment of neuroinflammation since absence of this kinase in the immune system leads to severe EAE.

Topics: Animals; Autoimmunity; B7-2 Antigen; Cytokines; Dendritic Cells; Encephalomyelitis, Autoimmune, Experimental; Glycoproteins; Histocompatibility Antigens Class II; MAP Kinase Signaling System; Mice; Mice, Inbred C57BL; Mice, Transgenic; Mitogen-Activated Protein Kinase 3; Myelin-Oligodendrocyte Glycoprotein; Ovalbumin; Peptide Fragments; Radiation Chimera; Specific Pathogen-Free Organisms; T-Cell Antigen Receptor Specificity; T-Lymphocyte Subsets

Although macrophages (MPhi) are known as essential players in wound healing, their contribution to recovery from spinal cord injury (SCI) is a subject of debate. The difficulties in distinguishing between different MPhi subpopulations at the lesion site have further contributed to the controversy and led to the common view of MPhi as functionally homogenous. Given the massive accumulation in the injured spinal cord of activated resident microglia, which are the native immune occupants of the central nervous system (CNS), the recruitment of additional infiltrating monocytes from the peripheral blood seems puzzling. A key question that remains is whether the infiltrating monocyte-derived MPhi contribute to repair, or represent an unavoidable detrimental response. The hypothesis of the current study is that a specific population of infiltrating monocyte-derived MPhi is functionally distinct from the inflammatory resident microglia and is essential for recovery from SCI.. We inflicted SCI in adult mice, and tested the effect of infiltrating monocyte-derived MPhi on the recovery process. Adoptive transfer experiments and bone marrow chimeras were used to functionally distinguish between the resident microglia and the infiltrating monocyte-derived MPhi. We followed the infiltration of the monocyte-derived MPhi to the injured site and characterized their spatial distribution and phenotype. Increasing the naïve monocyte pool by either adoptive transfer or CNS-specific vaccination resulted in a higher number of spontaneously recruited cells and improved recovery. Selective ablation of infiltrating monocyte-derived MPhi following SCI while sparing the resident microglia, using either antibody-mediated depletion or conditional ablation by diphtheria toxin, impaired recovery. Reconstitution of the peripheral blood with monocytes resistant to ablation restored the lost motor functions. Importantly, the infiltrating monocyte-derived MPhi displayed a local anti-inflammatory beneficial role, which was critically dependent upon their expression of interleukin 10.. The results of this study attribute a novel anti-inflammatory role to a unique subset of infiltrating monocyte-derived MPhi in SCI recovery, which cannot be provided by the activated resident microglia. According to our results, limited recovery following SCI can be attributed in part to the inadequate, untimely, spontaneous recruitment of monocytes. This process is amenable to boosting either by active vaccination with a myelin-derived altered peptide ligand, which indicates involvement of adaptive immunity in monocyte recruitment, or by augmenting the naïve monocyte pool in the peripheral blood. Thus, our study sheds new light on the long-held debate regarding the contribution of MPhi to recovery from CNS injuries, and has potentially far-reaching therapeutic implications.

Topics: Adoptive Transfer; Animals; Glycoproteins; Inflammation; Interleukin-10; Macrophages; Male; Mice; Mice, Inbred C57BL; Mice, Transgenic; Monocytes; Myelin-Oligodendrocyte Glycoprotein; Ovalbumin; Peptide Fragments; Spinal Cord; Spinal Cord Injuries

Recent studies in experimental autoimmune encephalomyelitis (EAE) have found that CNS injury in Daf1(-/-) mice is much greater than in wild types (WTs), suggesting that upregulating DAF levels in vivo might ameliorate disease. To test this, we generated a Daf1 transgenic (Tg) mouse which had elevated DAF levels on its cell surfaces. In by-stand C3b uptake assays, Daf1 Tg mouse erythrocytes took up less C3b on their surfaces than WT erythrocytes. When co-cultured with OT-II CD4(+) T cells together with OVA(323-339) peptide, Daf1 Tg mouse bone marrow derived dendritic cells (BM-DCs) produced less C5a and C3a than WT BM-DCs and stimulated a lesser T cell response. In MOG(35-55) immunization induced EAE model, Daf1 Tg mice exhibited delayed disease onset and decreased clinical scores compared to WTs. Histological analyses showed that there were less inflammation and demyelination in spinal cords in Daf1 Tg mice than those in WTs. In accordance with these results, Daf1 Tg mice had decreased MOG(35-55) specific Th1 and Th17 responses. These data provide further evidence that DAF suppresses autoreactive T cell responses in EAE, and indicate that augmenting its expression levels could be effective therapeutically in treating multiple sclerosis as well as other T cell mediated diseases.

Topics: Animals; CD55 Antigens; Complement C3a; Complement C5a; Dendritic Cells; Disease Models, Animal; Encephalomyelitis, Autoimmune, Experimental; Erythrocytes; Glycoproteins; Interferon-gamma; Interleukin-17; Lymphocyte Activation; Mice; Mice, Inbred C57BL; Mice, Knockout; Mice, Transgenic; Myelin-Oligodendrocyte Glycoprotein; Ovalbumin; Peptide Fragments; Spinal Cord; T-Lymphocyte Subsets; T-Lymphocytes, Helper-Inducer

Although many self-reactive T cells are eliminated by negative selection in the thymus, some of these cells escape into the periphery, where they must be controlled by additional mechanisms. However, the molecular mechanisms underlying peripheral T cell tolerance and its maintenance remain largely undefined. In this study, we report that sirtuin 1 (Sirt1), a type III histone deacetylase, negatively regulates T cell activation and plays a major role in clonal T cell anergy in mice. In vivo, we found that loss of Sirt1 function resulted in abnormally increased T cell activation and a breakdown of CD4+ T cell tolerance. Conversely, upregulation of Sirt1 expression led to T cell anergy, in which the activity of the transcription factor AP-1 was substantially diminished.Furthermore, Sirt1 interacted with and deacetylated c-Jun, yielding an inactive AP-1 factor. In addition, Sirt1-deficient mice were unable to maintain T cell tolerance and developed severe experimental allergic encephalomyelitis as well as spontaneous autoimmunity. These findings provide insight into the molecular mechanisms of T cell activation and anergy, and we suggest that activators of Sirt1 may be useful as therapeutic agents for the treatment and/or prevention of autoimmune diseases.

Topics: Adoptive Transfer; Animals; CD4-Positive T-Lymphocytes; Cell Line; Clonal Anergy; Encephalomyelitis, Autoimmune, Experimental; Glycoproteins; Humans; Immune Tolerance; Lymphocyte Activation; Mice; Mice, Inbred C57BL; Mice, Knockout; Mice, Transgenic; Myelin-Oligodendrocyte Glycoprotein; Ovalbumin; Peptide Fragments; Proto-Oncogene Proteins c-jun; Sirtuin 1

The characteristics, importance, and molecular requirements for interactions between mast cells (MCs) and CD8(+) T cells have not been elucidated. Here, we demonstrated that MCs induced antigen-specific CD8(+) T cell activation and proliferation. This process required direct cell contact and MHC class I-dependent antigen cross-presentation by MCs and induced the secretion of interleukin-2, interferon-gamma, and macrophage inflammatory protein-1alpha by CD8(+) T cells. MCs regulated antigen-specific CD8(+) T cell cytotoxicity by increasing granzyme B expression and by promoting CD8(+) T cell degranulation. Because MCs also upregulated their expression of costimulatory molecules (4-1BB) and released osteopontin upon direct T cell contact, MC-T cell interactions probably are bidirectional. In vivo, adoptive transfer of antigen-pulsed MCs induced MHC class I-dependent, antigen-specific CD8(+) T cell proliferation, and MCs regulated CD8(+) T cell-specific priming in experimental autoimmune encephalomyelitis. Thus, MCs are important players in antigen-specific regulation of CD8(+) T cells.

Topics: Adoptive Transfer; Animals; Antigen Presentation; CD8-Positive T-Lymphocytes; Cell Degranulation; Chemokine CCL3; Coculture Techniques; Cross-Priming; Encephalomyelitis, Autoimmune, Experimental; Glycoproteins; Granzymes; Histocompatibility Antigens Class I; Interferon-gamma; Interleukin-2; Listeria monocytogenes; Listeriosis; Mast Cells; Mice; Mice, Inbred C57BL; Mice, Transgenic; Myelin-Oligodendrocyte Glycoprotein; Osteopontin; Ovalbumin; Peptide Fragments; Tumor Necrosis Factor Receptor Superfamily, Member 9

Upon stimulation by microbial products through TLR, dendritic cells (DC) acquire the capacity to prime naive T cells and to initiate a proinflammatory immune response. Recently, we have shown that APC within the CNS of multiple sclerosis (MS) patients contain peptidoglycan (PGN), a major cell wall component of Gram-positive bacteria, which signals through TLR and NOD. In this study, we report that Staphylococcus aureus PGN as a single component can support the induction of experimental autoimmune encephalomyelitis (EAE) in mice, an animal model for MS. Mice immunized with an encephalitogenic myelin oligodendrocyte glycoprotein peptide in IFA did not develop EAE. In contrast, addition of PGN to the emulsion was sufficient for priming of autoreactive Th1 cells and development of EAE. In vitro studies demonstrate that PGN stimulates DC-mediated processes, reflected by increased Ag uptake, DC maturation, Th1 cell expansion, activation, and proinflammatory cytokine production. These data indicate that PGN-mediated interactions result in proinflammatory stimulation of Ag-specific effector functions, which are important in the development of EAE. These PGN-mediated processes may occur both within the peripheral lymph nodes as well as in the CNS and likely involve recognition by TLR on DC. Thus, PGN may provide a physiological trigger of DC maturation, and in this way disrupt the normal tolerance to self Ag. As such, PGN signaling pathways may serve as novel targets for the treatment of MS.

Topics: Adjuvants, Immunologic; Amino Acid Sequence; Animals; Cell Differentiation; Dendritic Cells; Encephalomyelitis, Autoimmune, Experimental; Epitopes, T-Lymphocyte; Female; Glycoproteins; Inflammation Mediators; Lymph Nodes; Lymphocyte Activation; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Transgenic; Molecular Sequence Data; Myelin-Oligodendrocyte Glycoprotein; Organ Specificity; Ovalbumin; Peptide Fragments; Peptidoglycan; Protein Transport; T-Lymphocyte Subsets; Th1 Cells

Previous studies suggested that depending on their maturation state, dendritic cells (DC) could either induce T cell tolerance (immature and semimature DC) or T cell activation (mature DC). Pretreatment of C57BL/6 mice with encephalitogenic myelin oligodendrocyte glycoprotein (MOG)(35-55) peptide-loaded semimature DC protected from MOG-induced autoimmune encephalomyelitis. This protection was mediated by IL-10-producing CD4 T cells specific for the self Ag. Here we show that semimature DC loaded with the MHC class II-restricted nonself peptide Ag (OVA) induce an identical regulatory T cell cytokine pattern. However, semimature DC loaded simultaneously with MHC class II- and MHC class I-restricted peptides, could efficiently initiate CD8 T cell responses leading to autoimmune diabetes in a TCR-transgenic adoptive transfer model. Double-peptide-loaded semimature DC also induced simultaneously in the same animal partially activated CD8 T cells with cytolytic function as well as protection from MOG-induced autoimmune encephalomyelitis. Our study suggests that the decision between tolerance and immunity not only depends on the DC, but also on the type and activation requirements of the responding T cell.

Topics: Animals; Antigen Presentation; CD4-Positive T-Lymphocytes; CD8-Positive T-Lymphocytes; Cytotoxicity, Immunologic; Dendritic Cells; Diabetes Mellitus, Experimental; Encephalomyelitis, Autoimmune, Experimental; Glycoproteins; Immune Tolerance; Immunity, Cellular; Lymphocyte Activation; Mice; Mice, Inbred C57BL; Myelin-Oligodendrocyte Glycoprotein; Ovalbumin; Peptide Fragments