Responses In Experimental Autoimmune Encephalomyelitis Research Articles

Intrathecal transplantation of neural stem/precursor cells hampers the CNS-confined immune response of experimental autoimmune encephalomyelitis

Intrathecal transplantation of neural stem/precursor cells hampers the CNS-confined immune response of experimental autoimmune encephalomyelitis

Blocking Glutamate Carboxypeptidase II Inhibited Glutamate Excitotoxicity and Regulated Immune Responses in Experimental Autoimmune Encephalomyelitis

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Effects of Vaccination with Altered Peptide Ligand on Chronic Pain in Experimental Autoimmune Encephalomyelitis, an Animal Model of Multiple Sclerosis

Neuropathic pain is a chronic symptom of multiple sclerosis (MS) and affects nearly half of all MS sufferers. A key instigator of this pain is the pro-inflammatory response in MS. We investigated the behavioral effects of immunization with a mutant peptide of myelin basic protein (MBP), termed altered peptide ligand (APL), known to initiate immune deviation from a pro-inflammatory state to an anti-inflammatory response in experimental autoimmune encephalomyelitis (EAE), an animal model of MS. Male and female Lewis rats were injected with vehicle control or with varying doses of 50 or 100 μg guinea pig MBP in combination with or without APL. APL-treated animals established significantly lower disease severity compared to encephalitogenic MBP-treated animals. Animals with EAE developed mechanical, but not thermal pain hypersensitivity. Mechanical pain sensitivities were either improved or normalized during periods of clinical disease in male and female APL-treated animals as compared to the encephalitogenic group. No significant changes to thermal latency were observed upon co-immunization with APL. Together these data indicate that APL ameliorates disease states and selectively mediates an analgesic effect on EAE animals.

The complement system contributes to the pathology of experimental autoimmune encephalomyelitis by triggering demyelination and modifying the antigen-specific T and B cell response

So far, studies of the human autoimmune disease multiple sclerosis (MS) have largely been hampered by the absence of a pathogenic B cell component in its animal model, experimental autoimmune encephalomyelitis (EAE). To overcome this shortcoming, we have previously introduced the myelin basic protein (MBP)-proteolipid protein (PLP) MP4-induced EAE, which is B cell and autoantibody-dependent. Here we show that MP4-immunized wild-type C57BL/6 mice displayed a significantly lower disease incidence when their complement system was transiently depleted by a single injection of cobra venom factor (CVF) prior to immunization. Considering the underlying pathomechanism, our data suggest that the complement system is crucial for MP4-specific antibodies to trigger CNS pathology. Demyelinated lesions in the CNS were colocalized with complement depositions. In addition, B cell deficient JHT mice reconstituted with MP4-reactive serum showed significantly attenuated clinical and histological EAE after depletion of complement by CVF. The complement system was also critically involved in the generation of the MP4-specific T and B cell response: in MP4-immunized wild-type mice treated with CVF the MP4-specific cytokine and antibody response was significantly attenuated compared to untreated wild-type mice. Taken together, we propose two independent mechanisms by which the complement system can contribute to the pathology of autoimmune encephalomyelitis. Our data corroborate the role of complement in triggering antibody-dependent demyelination and antigen-specific T cell immunity and also provide first evidence that the complement system can modify the antigen-specific B cell response in EAE and possibly MS.

Bevacizumab Diminishes Experimental Autoimmune Encephalomyelitis by Inhibiting Spinal Cord Angiogenesis and Reducing Peripheral T-Cell Responses

Angiogenesis in the animal model of multiple sclerosis experimental autoimmune encephalomyelitis (EAE) is regulated by vascular endothelial growth factor (VEGF) and angiopoietin-2. We determined whether VEGF blockade with the anti-VEGF monoclonal antibody bevacizumab could inhibit angiogenesis and affect peripheral pathogenic immune responses in EAE. Mice treated with bevacizumab from the time of onset of clinical signs showed reduced clinical and pathologic scores. Bevacizumab suppressed angiogenesis and reduced angiopoietin-2 expression at Day 21 but had no effect on VEGF upregulation at Day 14. Messenger RNA levels for the angiogenesis-related protein CD105 were increased at Day 14. Bevacizumab reduced vascular permeability in the spinal cord at Day 14 and Day 21. In peripheral lymph nodes, it induced retention of CD4-positive T cells and inhibited T-cell proliferation. It also reduced mononuclear cell infiltration into spinal cord and the relative proportion of T cells. Isolated lymphoid cells showed reduced secretion of the T-helper 17 (Th-17) cell cytokine interleukin 17 and the Th-1 cytokine interferon-γ. When bevacizumab was added to naive T cells or to antigen-stimulated T cells from mice with untreated EAE in vitro, it had no effect on proliferation or the secretion of interleukin 17 or interferon-γ. These data indicate that bevacizumab ameliorates vascular and T-cell responses during EAE, but its effects on T cells may be indirect, possibly by suppressing angiogenesis.

P091 Interferon-beta regulates Th17 and dendritic cell responses in EAE

Introduction Multiple sclerosis (MS) is an autoimmune disease that targets the central nervous system. Interferon (IFN)- β was the first approved therapy for relapsing-remitting MS more than 10 years ago, however, its mechanism of action remains ill-defined. Methods Using a murine experimental autoimmune encephalomyelitis (EAE) model for MS, we show that mice null for IFN- β , IFN- β −/− , exhibit an earlier onset and a more rapid progression of disease compared to IFN- β +/+ mice. Results IFN- β −/− mice exhibit increased numbers of Th17 cells in their draining lymph nodes (DLN) as early as 4 days after disease induction. Th17 cells are implicated as critical effectors in different autoimmune diseases, including MS. Increased Th17 cell polarization may be driven by dendritic cells (DC), as DCs derived from IFN- β −/− mice induce greater proliferation of T cells derived from either IFN- β +/+ or IFN- β −/− mice, compared with DCs derived from IFN- β +/+ mice. In addition, adoptive transfer of myelin oligodendrocyte glycoprotein (MOG) peptide-primed IFN- β −/− DC into IFN- β +/+ and IFN- β −/− mice with EAE resulted in their rapid migration into the CNS of recipient mice, visualized by high resolution fluorescence imaging. Cytokine expression analysis revealed that DCs generated from IFN- β −/− mice secrete increased levels of IL-6, yet decreased levels of IL-27, compared with DCs generated from IFN- β +/+ mice. These data suggest that DCs from IFN- β −/− mice have a propensity to secrete cytokines perhaps associated with Th17 rather than Treg polarization. Moreover, the IFN- β −/− DCs produce lower levels of IL-1 and IL-10. The activation profiles of proteins, such as phospho-proteins that drive proliferation and activation signaling cascades, differ in disease states, both from a “normal” profile and among different pathologies. Using our mouse EAE model of MS and human MS patient peripheral blood, we are interrogating the phosphorylation status of signaling effectors, including STATs and MAPKs, by FACS-based multiparametric staining of cell surface and intracellular markers to determine the activation status of circulating discrete immune cell populations. Our staining panels allow us to discern the phosphorylation status of these signaling effectors in T cell populations and DCs. Data will be presented that describe the signature phospho-profiles during active disease/relapse and remission. Conclusion Taken together, the data indicate that IFN- β plays a role in regulating the Th17 and DC responses during EAE, by limiting cell accumulation and trafficking and by regulating cytokine production.

Lipocalin 2 is a novel immune mediator of experimental autoimmune encephalomyelitis pathogenesis and is modulated in multiple sclerosis

Experimental autoimmune encephalomyelitis (EAE) is a widely used animal model of multiple sclerosis (MS), an inflammatory, demyelinating disease of the central nervous system (CNS). EAE pathogenesis involves various cell types, cytokines, chemokines, and adhesion molecules. Given the complexity of the inflammatory response in EAE, it is likely that many immune mediators still remain to be discovered. To identify novel immune mediators of EAE pathogenesis, we performed an Affymetrix gene array screen on the spinal cords of mice at the onset stage of disease. This screening identified the gene encoding lipocalin 2 (Lcn2) as being significantly upregulated. Lcn2 is a multi-functional protein that plays a role in glial activation, matrix metalloproteinase (MMP) stabilization, and cellular iron flux. As many of these processes have been implicated in EAE, we characterized the expression and role of Lcn2 in this disease in C57BL/6 mice. We show that Lcn2 is significantly upregulated in the spinal cord throughout EAE and is expressed predominantly by monocytes and reactive astrocytes. The Lcn2 receptor, 24p3R, is also expressed on monocytes, macrophages/microglia, and astrocytes in EAE. In addition, we show that EAE severity is increased in Lcn2(-/-) mice as compared with wild-type controls. Finally, we demonstrate that elevated levels of Lcn2 are detected in the plasma and cerebrospinal fluid (CSF) in MS and in immune cells in CNS lesions in MS tissue sections. These data indicate that Lcn2 is a modulator of EAE pathogenesis and suggest that it may also play a role in MS.

Differential regulation of CD4+ T helper cell responses by curcumin in experimental autoimmune encephalomyelitis

Nutraceuticals and phytochemicals are important regulators of human health and diseases. Curcumin is a polyphenolic phytochemical isolated from the rhizome of the plant Curcuma longa (turmeric) that has been traditionally used for the treatment of inflammation and wound healing for centuries. Systematic analyses have shown that curcumin exerts its beneficial effects through antioxidant, antiproliferative and anti-inflammatory properties. We and others have shown earlier that curcumin ameliorates experimental autoimmune encephalomyelitis (EAE) model for multiple sclerosis. In this study, we show that C57BL/6 mice induced to develop EAE express elevated levels of interferon (IFN) γ and interleukin (IL)-17 in the central nervous system (CNS) and lymphoid organs that decreased significantly following in vivo treatment with curcumin. The EAE mice also showed elevated expression of IL-12 and IL-23 that decreased after treatment with curcumin. Ex vivo and in vitro treatment with curcumin resulted in a dose-dependent decrease in the secretion of IFNγ, IL-17, IL-12 and IL-23 in culture. The inhibition of EAE by curcumin was also associated with an up-regulation of IL-10, peroxisome proliferator activated receptor γ and CD4+CD25+Foxp3+ Treg cells in the CNS and lymphoid organs. These findings highlight that curcumin differentially regulates CD4+ T helper cell responses in EAE.

Interleukin-10 plays a crucial role in suppression of experimental autoimmune encephalomyelitis by Bowman–Birk inhibitor

The Bowman–Birk inhibitor (BBI) is a soybean-derived serine protease inhibitor with anti-inflammatory properties. Experimental autoimmune encephalomyelitis (EAE) serves as an animal model of the central nervous system (CNS) inflammatory disorder multiple sclerosis (MS). EAE is mediated by Th1 and Th17 cells which migrate into the CNS and initiate inflammation directed against myelin components, resulting in CNS pathology and neurological clinical deficit. We have shown previously that oral treatment with BBI delays onset of EAE and reduces its severity. These beneficial effects were associated with an increase in IL-10 secretion by immune cells of BBI-treated mice. It is not known, however, whether this was a causal relationship or simply an epiphenomenon. In the present study we provide evidence that BBI regulates CD4+ T cell immune responses in EAE. BBI administration delayed the onset of EAE and reduced its severity in an IL-10-dependent manner, as BBI-mediated suppression of EAE was abrogated in IL-10 knockout mice. The beneficial effects were accompanied by reduced IFN-γ, IL-17 and increased IL-10 production, as well as increased Foxp3 expression. CD4+ T cells were the major source of IL-10 in the periphery and in the CNS during BBI treatment. Furthermore, BBI-treated mice had reduced numbers of infiltrated cells in the CNS, including Th17 cells, as compared with PBS-treated control animals. In conclusion, our data provide clear evidence for the essential role of IL-10 in BBI-mediated suppression in EAE, and indicate that BBI may be a promising candidate for the development of a novel MS therapy.

Synapsin peptide fused to E. coli heat-labile toxin B subunit induces regulatory T cells and modulates cytokine balance in experimental autoimmune encephalomyelitis

We previously found that the preventive oral administration of a hybrid consisting of the C domain of synapsin and the B subunit of E. coli heat-labile enterotoxin (LTBSC) efficiently suppresses experimental autoimmune encephalomyelitis (EAE) development in rats. We investigated the effect of LTBSC on cytokine expression and on regulatory T (Treg) cells in rats with myelin induced EAE. LTBSC treatment increased the frequency of CD4+FoxP3+ Treg cells in lymph nodes prior to challenge and in the EAE acute stage. LTBSC also up-regulated the expression of anti-inflammatory Th2/Th3 cytokines and diminished myelin basic protein-specific Th1 and Th17 cell responses in lymph nodes. CD4+CD25+ Treg cells from LTBSC treated rats showed stronger suppressive properties than Treg cells from controls in vitro. Our observations indicate that LTBSC is a useful agent for modulating the autoimmune responses in EAE.

Early Murine T-lymphocyte Activation Is Accompanied by a Switch from N-Glycolyl- to N-Acetyl-neuraminic Acid and Generation of Ligands for Siglec-E

It is well established that murine T-lymphocyte activation is accompanied by major changes in cell-surface sialylation, potentially influencing interactions with sialic acid-binding immunoglobulin-like lectins (siglecs). In the present study, we analyzed early activation of murine CD4+ and CD8+ T-lymphocytes at 24 h. We observed a striking and selective up-regulation in the binding of a recombinant soluble form of siglec-E, an inhibitory siglec, which is expressed on several myeloid cell types including antigen-presenting dendritic cells. In contrast, much lower levels of T cell binding were observed with other siglecs, including sialoadhesin, CD22, and siglec-F and the plant lectins Maackia amurensis leukoagglutinin and Sambucus nigra agglutinin. By mass spectrometry, the sialic acid content of 24-h-activated CD4+ and CD8+ T-lymphocytes exhibited an increased proportion of N-acetyl-neuraminic acid (NeuAc) to N-glycolyl-neuraminic acid (NeuGc) in N-glycans. Reduced levels of NeuGc on the surface of activated T cells were demonstrated using an antibody specific for NeuGc and the expression levels of the gene encoding NeuAc- to NeuGc-converting enzyme, CMP-NeuAc hydroxylase, were also reduced. Siglec-E bound a wide range of sialylated structures in glycan arrays, had a preference for NeuAc versus NeuGc-terminated sequences and could recognize a set of sialoglycoproteins that included CD45, in lysates from activated T-lymphocytes. Collectively, these results show that early in T cell activation, glycan remodelling involves a switch from NeuGc- to NeuAc-terminating oligosaccharides on cell surface glycoproteins. This is associated with a strong up-regulation of siglec-E ligands, which may be important in promoting cellular interactions between early activated T-lymphocytes and myeloid cells expressing this inhibitory receptor.

An acidic polysaccharide of Panax ginseng ameliorates experimental autoimmune encephalomyelitis and induces regulatory T cells

An acidic polysaccharide of Panax ginseng (APG), so called ginsan, is a purified polysaccharide. APG has multiple immunomodulatory effects of stimulating natural killer (NK) and T cells and producing a variety of cytokines that proved to diminish the proinflammatory response, and protect from septic lethality. To determine APG's role in the autoimmune demyelinating disease, we tested whether APG can regulate inflammatory and encephalitogenic response in experimental autoimmune encephalomyelitis (EAE), an animal model of human multiple sclerosis (MS). Here, we demonstrate the therapeutic efficacy of the APG which induces the suppression of an encephalitogenic response during EAE. APG significantly ameliorates the progression of EAE by inhibiting the proliferation of autoreactive T cells and the production of inflammatory cytokines such as IFN-γ, IL-1β and IL-17. More importantly, APG promotes the generation of immunosuppressive regulatory T cells (Tregs) through the activation of transcription factor, Foxp3. Furthermore, the depletion of CD25+ cells from APG-treated EAE mice abrogates the beneficial effects of EAE. The capacity of APG to induce clinically beneficial effects furthers our understanding of the basis for its therapeutic immunosuppression of EAE and, possibly, MS. Thus, our results suggest that APG may serve as an effective therapy for MS and other autoimmune diseases.

IL-4- and IL-10-producing invariant NKT cells inhibit Th1 but not Th17 cells, while IFN-γ-producing invariant NKT cells minimally regulate Th1/Th17 response in experimental autoimmune encephalomyelitis (101.10)

Abstract Experimental autoimmune encephalomyelitis (EAE) is regulated by Th1 and Th17 response. Invariant (i) NKT cells prevent EAE, depending on IL-4, IL-10, and IFN-γ. However, which cytokines produced by iNKT cells regulate Th1 and/or Th17 response in EAE remains unclear. To address this issue, B6 and Jα18-/- mice were immunized with MOG35-55 peptide. Jα18-/- mice showed higher clinical scores, IL-17 and IFN-γ transcript, and intracellular IL-17 and IFN-γ+CD4+ T cells in CNS than B6 mice, which was reduced by adoptive transfer of wild type or IFN-γ-deficient iNKT cells. Adoptive transfer of IL-4- or IL-10-deficient iNKT cells into Jα18-/- mice decreased IL-17 transcript, and intracellular IL-17+CD4+ T cells in CNS but not clinical scores, IFN-γ transcript, and intracellular IFN-γ+CD4+ T cells. Taken together, IL-4- and IL-10-producing iNKT cells inhibit Th1 but not Th17 response in CNS during EAE. Moreover, IFN-γ-producing iNKT cells minimally regulate Th1 and Th17 in EAE.

PI3Kδ drives the pathogenesis of experimental autoimmune encephalomyelitis by inhibiting effector T cell apoptosis and promoting Th17 differentiation

The Class IA phosphoinositide 3-kinase delta (PI3Kδ) has been implicated in multiple signaling pathways involved in leukocyte activation and hence is an attractive target in many human autoimmune diseases, including multiple sclerosis (MS). Here, using mice expressing a catalytically inactive form of the PI3Kδ subunit p110δ, we show that signaling through PI3Kδ is required for full and sustained pathology of experimental autoimmune encephalomyelitis (EAE), a Th17-driven model of MS. In p110δ-inactivated mice, T cell activation and function during EAE was markedly reduced and fewer T cells were observed in the central nervous system (CNS). The decrease in T cell activation is unlikely to be due to defects in dendritic cell (DC) function, as p110δ-inactivated DCs migrate and present antigen normally. However, significant increases in the proportion of T cells undergoing apoptosis at early stages of EAE were evident in the absence of PI3Kδ activity. Furthermore, a profound defect in Th17 cellular responses during EAE was apparent in the absence of PI3Kδ activity while Th1 responses were less affected. A highly selective PI3Kδ inhibitor, IC87114, also had greater inhibitory effects on Th17 cell generation in vitro than it did on Th1 cell generation. Thus, PI3Kδ plays an important role in Th17 responses in EAE, suggesting that small molecule inhibitors of PI3Kδ may be useful therapeutics for treatment of MS and other autoimmune diseases.

The clinical course of EAE is reflected by the dynamics of the neuroantigen-specific T cell compartment in the blood

Due to the limited numbers of PBMCs that can be obtained from the blood of individual mice, the key question whether central disease parameters such as onset, progression and severity correlate with the magnitude and cytokine quality of the T cell response in experimental autoimmune encephalomyelitis (EAE) has remained unanswered. Here we introduce an ELISPOT-based PBMC test system in which as little as 150 μl of murine blood are sufficient, allowing to bleed mice repeatedly while continuing to observe the clinical course of EAE. Using this technique, we demonstrate that longitudinal measurements of antigen-specific IFN-γ and IL-17 production in the blood are a highly suitable approach to predict the disease outcome in remitting–relapsing PLP:139–151- and chronic MOG:35–55-induced EAE of SJL/J and C57BL/6 mice, respectively. Our data propound cytokine monitoring as promising tool in the quest for more efficient diagnostic and prognostic options in human multiple sclerosis and other autoimmune diseases.

Peroxisome proliferator‐activated receptor δ agonists inhibit T helper type 1 (Th1) and Th17 responses in experimental allergic encephalomyelitis

Multiple sclerosis (MS) is a neurological disorder that affects more than a million people world-wide. The aetiology of MS is not known and there is no medical treatment available that can cure MS. Experimental autoimmune encephalomyelitis (EAE) is a T-cell-mediated autoimmune disease model of MS. The pathogenesis of EAE/MS is a complex process involving activation of immune cells, secretion of inflammatory cytokines and destruction of myelin sheath in the central nervous system (CNS). Peroxisome proliferator-activated receptors (PPARs) are nuclear hormone receptor transcription factors that regulate cell growth, differentiation and homeostasis. PPAR agonists have been used in the treatment of obesity, diabetes, cancer and inflammation. We and others have shown that PPARgamma, alpha and delta agonists inhibit CNS inflammation and demyelination in the EAE model of MS. In this study we show that the PPARdelta agonists GW501516 and L165041 ameliorate MOGp35-55-induced EAE in C57BL/6 mice by blocking interferon (IFN)-gamma and interleukin (IL)-17 production by T helper type 1 (Th1) and Th17 cells. The inhibition of EAE by PPARdelta agonists was also associated with a decrease in IL-12 and IL-23 and an increase in IL-4 and IL-10 expression in the CNS and lymphoid organs. These findings indicate that PPARdelta agonists modulate Th1 and Th17 responses in EAE and suggest their use in the treatment of MS and other autoimmune diseases.

α4β1 Integrin Mediates the Recruitment of Immature Dendritic Cells across the Blood-Brain Barrier during Experimental Autoimmune Encephalomyelitis

Dendritic cells (DCs) within the CNS are recognized to play an important role in the effector phase and propagation of the immune response in experimental autoimmune encephalomyelitis (EAE), a mouse model for multiple sclerosis. However, the mechanisms regulating DC trafficking into the CNS still need to be characterized. In this study, we show by performing intravital fluorescence videomicroscopy of the inflamed spinal cord white-matter microvasculature in SJL mice with EAE that immature, and to a lesser extent, LPS-matured, bone marrow-derived DCs efficiently interact with the CNS endothelium by rolling, capturing, and firm adhesion. Immature but not LPS-matured DCs efficiently migrated across the wall of inflamed parenchymal microvessels into the CNS. Blocking alpha4 integrins interfered with the adhesion but not the rolling or capturing of immature and LPS-matured DCs to the CNS microvascular endothelium, inhibiting their migration across the vascular wall. Functional absence of beta1 integrins but not of beta7 integrins or alpha4beta7 integrin similarly reduced the adhesion of immature DCs to the CNS microvascular endothelium, demonstrating that alpha4beta1 but not alpha4beta7 integrin mediates this step of immature DCs interaction with the inflamed blood-brain barrier during EAE. Our study shows that during EAE, especially immature DCs migrate into the CNS, where they may be crucial for the perpetuation of the CNS-targeted autoimmune response. Thus therapeutic targeting of alpha4 integrins affects DC trafficking into the CNS and may therefore lead to the resolution of the CNS autoimmune inflammation by reducing the number of CNS professional APCs.

Toll‐like Receptors in Multiple Sclerosis Mouse Experimental Models

Certain pathogen molecules trigger innate immune responses and drive subsequent adaptive immune responses toward an antigen presented simultaneously. Such bacterial or viral molecules constitute pathogen-associated molecular patterns (PAMPs) that bind to pattern-recognition receptors such as toll-like receptors (TLRs). Recently, endogenous molecules were identified that ligate the same receptors. The role of these receptors' response to complete Freund's adjuvant during initiation of CD4 T cell responses in EAE, the animal model for multiple sclerosis, is here discussed. Myeloid differentiation primary response gene 88 (MyD88) is necessary for the induction of experimental autoimmune encephalomyelitis (EAE), and it is required for the activation of myeloid dendritic cells and differentiation of T helper 17 cells. The role of individual TLR, in particular TLR3, TLR4, and TLR9, signaling in modulation of EAE inflammation varies with the experimental model employed and the immune cells that drive pathology. The TLR-dependent production of proinflammatory cytokines is regulated by mechanisms that dampen the pathway and prevent excess damage. Development of TLR antagonists to treat autoimmune diseases must acknowledge the possibility of interference with regulatory mechanisms.

Augmenting DAF levels in vivo ameliorates experimental autoimmune encephalomyelitis

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 OVA323–339 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 MOG35–55 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 MOG35–55 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.

Signal Transduction Inhibition of APCs Diminishes Th17 and Th1 Responses in Experimental Autoimmune Encephalomyelitis

IL-17- and IFN-gamma-secreting T cells play an important role in autoimmune responses in multiple sclerosis and the model system experimental autoimmune encephalomyelitis (EAE). Dendritic cells (DCs) in the periphery and microglia in the CNS are responsible for cytokine polarization and expansion of this T cell subset. Our results indicate that in vivo administration of a signal transduction inhibitor that targets DCs to mice with EAE led to a decrease in CNS infiltration of pathogenic Ag-specific T cells. Since this approach does not target T cells directly, we assessed the effects on the APCs that are involved in generating the T cell responses. Since in EAE and multiple sclerosis, both microglia and peripheral DCs are likely to contribute to disease, we utilized a bone marrow chimera system to distinguish between these two populations. These studies show that peripheral DCs are the primary target but that microglia are also modestly affected by CEP-701, as numbers and activation states of the cells in the CNS are decreased after therapy. Our results also showed a decrease in secretion of TNF-alpha, IL-6, and IL-23 by DCs as well as a decrease in expression of costimulatory molecules. We further determined that levels of phospho-Stat1, Stat3, Stat5, and NF-kappaB, which are signaling molecules that have been implicated in these pathways, were decreased. Thus, use of this class of signal transduction inhibitors may represent a novel method to treat autoimmunity by dampening the autoreactive polarizing condition driven by DCs.