TGF-beta1-mediated control of central nervous system inflammation and autoimmunity through the inhibitory receptor CD26

The T cell marker CD26/dipeptidyl peptidase (DP) IV is associated with an effector phenotype and markedly elevated in the human CNS disorder multiple sclerosis. However, little is known about the in vivo role of CD26/DP IV in health and disease, and the underlying mechanism of its function in CNS inflammation. To directly address the role of CD26/DP IV in vivo, we examined Th1 immune responses and susceptibility to experimental autoimmune encephalomyelitis in CD26(-/-) mice. We show that gene deletion of CD26 in mice leads to deregulation of Th1 immune responses. Although production of IFN-gamma and TNF-alpha by pathogenic T cells in response to myelin Ag was enhanced in CD26(-/-) mice, production of the immunosuppressive cytokine TGF-beta1 was diminished in vivo and in vitro. In contrast to the reduction in TGF-beta1 production, responsiveness to external TGF-beta1 was normal in T cells from CD26(-/-) mice, excluding alterations in TGF-beta1 sensitivity as a mechanism causing the loss of immune regulation. Natural ligands of CD26/DP IV induced TGF-beta1 production in T cells from wild-type mice. However, natural ligands of CD26/DP IV failed to elicit TGF-beta1 production in T cells from CD26(-/-) mice. The striking functional deregulation of Th1 immunity was also seen in vivo. Thus, clinical experimental autoimmune encephalomyelitis scores were significantly increased in CD26(-/-) mice immunized with peptide from myelin oligodendrocyte glycoprotein. These results identify CD26/DP IV as a nonredundant inhibitory receptor controlling T cell activation and Th1-mediated autoimmunity, and may have important therapeutic implications for the treatment of autoimmune CNS disease.     

CD1-dependent regulation of chronic central nervous system inflammation in experimental autoimmune encephalomyelitis

The existence of T cells restricted for the MHC I-like molecule CD1 is well established, but the function of these cells is still obscure; one implication is that CD1-dependent T cells regulate autoimmunity. In this study, we investigate their role in experimental autoimmune encephalomyelitis (EAE), an animal model for multiple sclerosis, using CD1-deficient mice on a C57BL/6 background. We show that CD1-/- mice develop a clinically more severe and chronic EAE compared with CD1+/+ C57BL/6 mice, which was histopathologically confirmed with increased demyelination and CNS infiltration in CD1-/- mice. Autoantigen rechallenge in vitro revealed similar T cell proliferation in CD+/+ and CD1-/- mice but an amplified cytokine response in CD1-/- mice as measured by both the Th1 cytokine IFN-gamma and the Th2 cytokine IL-4. Investigation of cytokine production at the site of inflammation showed a CNS influx of TGF-beta1-producing cells early in the disease in CD1+/+ mice, which was absent in the CD1-/- mice. Passive transfer of EAE using an autoreactive T cell line induced equivalent disease in both groups, which suggested additional requirements for activation of the CD1-dependent regulatory pathway(s). When immunized with CFA before T cell transfer, the CD1-/- mice again developed an augmented EAE compared with CD1+/+ mice. We suggest that CD1 exerts its function during CFA-mediated activation, regulating development of EAE both through enhancing TGF-beta1 production and through limiting autoreactive T cell activation, but not necessarily via effects on the Th1/Th2 balance.     

Th1 and Th2 cytokine immunomodulation by gangliosides in experimental autoimmune encephalomyelitis

In experimental autoimmune encephalomyelitis, a classical model for multiple sclerosis, the cytokines provide the necessary signals to activate specific T cells for self-antigens. Gangliosides have multiple immunomodulatory activities, decreasing the lymphoproliferative responses and modulating cytokine production. Here, we tested the effects of gangliosides on the switching of Th1 to Th2 cytokine expression, in spleen cells obtained from Lewis rats during the acute phase of EAE, and after recovery from the disease. For this purpose, total RNA from spleen cells was isolated and submitted to RT-PCR to investigate Th1 (IL-2, TNF-alpha, and IFN-gamma) and Th2/Th3 (IL-10 and TGF-beta) cytokine gene expression. Results demonstrate that the group treated with gangliosides displays mild disease, with low expression of IFN-gamma mRNA and high TGF-beta mRNA expression. We conclude that the gangliosides may modulate Th1 cells by the synthesis of cytokines shifting the profile to the Th2/Th3 phenotype.     

In vivo blockade of macrophage migration inhibitory factor ameliorates acute experimental autoimmune encephalomyelitis by impairing the homing of encephalitogenic T cells to the central nervous system

Macrophage migration inhibitory factor (MIF) is a cytokine that plays a critical role in the regulation of macrophage effector functions and T cell activation. However, its role in the pathogenesis of T cell-mediated autoimmune diseases, such as experimental autoimmune encephalomyelitis (EAE), has remained unresolved. In this study, we report that anti-MIF Ab treatment of SJL mice with acute EAE improved the disease severity and accelerated the recovery. Furthermore, the anti-MIF treatment impaired the homing of neuroantigen-reactive pathogenic T cells to the CNS in a VCAM-1-dependent fashion. Interestingly, MIF blockade also decreased the clonal size of the neuroantigen-specific Th1 cells and increased their activation threshold. Taken together, the results demonstrate an important role for MIF in the pathogenesis of EAE/multiple sclerosis and suggest that MIF blockade may be a promising new strategy for the treatment of multiple sclerosis.     

G-CSF therapy of ongoing experimental allergic encephalomyelitis via chemokine- and cytokine-based immune deviation

Converging evidence that G-CSF, the hemopoietic growth factor of the myeloid lineage, also exerts anti-inflammatory and pro-Th2 effects, prompted us to evaluate its direct therapeutic potential in autoimmune diseases. Here we report a novel activity of G-CSF in experimental allergic encephalomyelitis, a murine model for multiple sclerosis, driven by Th1-oriented autoaggressive cells. A short 7-day treatment with G-CSF, initiated at the onset of clinical signs, provided durable protection from experimental autoimmune encephalomyelitis. G-CSF-treated mice displayed limited demyelination, reduced recruitment of T cells to the CNS, and very discrete autoimmune inflammation, as well as barely detectable CNS mRNA levels of cytokines and chemokines. In the periphery, G-CSF treatment triggered an imbalance in the production by macrophages as well as autoreactive splenocytes of macrophage inflammatory protein-1alpha and monocyte chemoattractant protein-1, the prototypical pro-Th1 and pro-Th2 CC chemokines, respectively. This chemokine imbalance was associated with an immune deviation of the autoreactive response, with reduced IFN-gamma and increased IL-4 and TGF-beta1 levels. Moreover, G-CSF limited the production of TNF-alpha, a cytokine also associated with early CNS infiltration and neurological deficit. These findings support the potential application of G-CSF in the treatment of human autoimmune diseases such as multiple sclerosis, taking advantage of the wide clinical favorable experience with this molecule.     

Prevention and treatment of chronic relapsing experimental allergic encephalomyelitis by transforming growth factor-beta 1.

Experimental allergic encephalomyelitis (EAE) is an autoimmune disease characterized by inflammation and demyelination in the central nervous system. The effect of the immunosuppressive molecule transforming growth factor-beta, (TGF-beta 1) on chronic relapsing EAE produced by the transfer of myelin basic protein-specific T cell lines was studied. TGF-beta 1 markedly inhibited the activation and proliferation of myelin-basic protein-specific lymph node cells in vitro. This reduced the capacity of these cells to transfer EAE. In addition, administration of TGF-beta 1 in vivo consistently resulted in an improved clinical course, even when given during ongoing disease. Immunopathologic study demonstrated a marked reduction in central nervous system damage and expression of cell-surface lymphocyte function-associated Ag-1 and class II MHC molecules in TGF-beta 1-treated mice. These findings have identified TGF-beta 1 as a possible therapeutic agent for the human demyelinating disease multiple sclerosis.     

Adoptive immunotherapy of experimental autoimmune encephalomyelitis via T cell delivery of the IL-12 p40 subunit

CD4+ T cells are believed to play a central role in the initiation and perpetuation of autoimmune diseases such as multiple sclerosis. In the murine model for multiple sclerosis, experimental autoimmune encephalomyelitis, pathogenic T cells exhibit a Th1-like phenotype characterized by heightened expression of proinflammatory cytokines. Systemic administration of "regulatory" cytokines, which serve to counter Th1 effects, has been shown to ameliorate autoimmune responses. However, the inherent problems of nonspecific toxicity limit the usefulness of systemic cytokine delivery as a potential therapy. Therefore, we used the site-specific trafficking properties of autoantigen-reactive CD4+ T cells to develop an adoptive immunotherapy protocol that provided local delivery of a Th1 cytokine antagonist, the p40 subunit of IL-12. In vitro analysis demonstrated that IL-12 p40 suppressed IFN-gamma production in developing and effector Th1 populations, indicating its potential to modulate Th1-promoted inflammation. We have previously demonstrated that transduction of myelin basic protein-specific CD4+ T cells with pGC retroviral vectors can result in efficient and stable transgene expression. Therefore, we adoptively transferred myelin basic protein-specific CD4+ T cells transduced to express IL-12 p40 into mice immunized to develop experimental autoimmune encephalomyelitis and demonstrated a significant reduction in clinical disease. In vivo tracking of bioluminescent lymphocytes, transduced to express luciferase, using low-light imaging cameras demonstrated that transduced CD4+ T cells trafficked to the central nervous system, where histological analysis confirmed long-term transgene expression. These studies have demonstrated that retrovirally transduced autoantigen-specific CD4+ T cells inhibited inflammation and promoted immunotherapy of autoimmune disorders.     

Immune modulation in multiple sclerosis patients treated with the pregnancy hormone estriol

The protective effect of pregnancy on putative Th1-mediated autoimmune diseases, such as multiple sclerosis and rheumatoid arthritis, is associated with a Th1 to Th2 immune shift during pregnancy. The hormone estriol increases during pregnancy and has been shown to ameliorate experimental autoimmune encephalomyelitis and collagen-induced arthritis. In addition, estrogens induce cytokine changes consistent with a Th1 to Th2 shift when administered in vitro to human immune cells and in vivo to mice. In a pilot trial, oral estriol treatment of relapsing remitting multiple sclerosis patients caused significant decreases in enhancing lesions on brain magnetic resonance imaging. Here, the immunomodulatory effects of oral estriol therapy were assessed. PBMCs collected longitudinally during the trial were stimulated with mitogens, recall Ags, and glatiramer acetate. Cytokine profiles of stimulated PBMCs were determined by intracellular cytokine staining (IL-5, IL-10, IL-12 p40, TNF-alpha, and IFN-gamma) and cytometric bead array (IL-2, IL-4, IL-5, IL-10, TNF-alpha, and IFN-gamma). Significantly increased levels of IL-5 and IL-10 and decreased TNF-alpha were observed in stimulated PBMC isolated during estriol treatment. These changes in cytokines correlated with reductions of enhancing lesions on magnetic resonance imaging in relapsing remitting multiple sclerosis. The increase in IL-5 was primarily due to an increase in CD4(+) and CD8(+) T cells, the increase in IL-10 was primarily due to an increase in CD64(+) monocytes/macrophages with some effect in T cells, while the decrease in TNF-alpha was primarily due to a decrease in CD8(+) T cells. Further study of oral estriol therapy is warranted in Th1-mediated autoimmune diseases with known improvement during pregnancy.     

Immune monitoring of Trichuris suis egg therapy in multiple sclerosis patients

Initial clinical trials using Trichuris suis eggs (TSO) in autoimmune diseases such as inflammatory bowel disease, revealed a striking suppressive effect on the autoimmune response. Here, we analysed the effect of TSO therapy on the course of multiple sclerosis (MS), as a Th1/Th17-associated autoimmune disease. Different immunological parameters in four patients with secondary progressive MS were surveyed during a 6-month therapy with TSO, focusing on the modulation of T-cell Th1-Th2 balance as well as on the innate immune response. We are able to show a slight downregulation of the Th1-associated cytokine pattern, especially relevant in interleukin (IL)-2 (P < 0.05 after 2 months of therapy), with a temporary increase of Th2-associated cytokines such as IL-4. Furthermore, mild eosinophily and changes in CD4+ and CD8+T cells and natural killer (NK) CD56 bright cell numbers were observed. The findings observed in this group of patients suggest that TSO therapy has a moderate immunomodulatory impact in MS.     

Differential effects of IL-21 during initiation and progression of autoimmunity against neuroantigen

The cytokine IL-21 is closely related to IL-2 and IL-15, a cytokine family that uses the common gamma-chain for signaling. IL-21 is expressed by activated CD4(+) T cells. We examined the role of IL-21 in the autoimmune disease experimental autoimmune encephalomyelitis (EAE), an animal model for human multiple sclerosis. IL-21 administration before induction of EAE with a neuroantigen, myelin oligodendrocyte glycoprotein peptide 35-55, and adjuvant enhanced the inflammatory influx into the CNS, as well as the severity of EAE. Autoreactive T cells purified from IL-21-treated mice transferred more severe EAE than did the control encephalitogenic T cells. No such effects were observed when IL-21 was administered after EAE progressed. Additional studies demonstrated that IL-21 given before the induction of EAE boosted NK cell function, including secretion of IFN-gamma. Depletion of NK cells abrogated the effect of IL-21. Therefore, IL-21, by affecting NK cells, has differential effects during the initiation and progression of autoimmune responses against neuroantigens.     

Cutting edge: CD4+CD25+ regulatory T cells suppress antigen-specific autoreactive immune responses and central nervous system inflammation during active experimental autoimmune encephalomyelitis

Autoreactive CD4(+) T cells exist in normal individuals and retain the capacity to initiate autoimmune disease. The current study investigates the role of CD4(+)CD25(+) T-regulatory (T(R)) cells during autoimmune disease using the CD4(+) T cell-dependent myelin oligodendrocyte glycoprotein (MOG)-specific experimental autoimmune encephalomyelitis model of multiple sclerosis. In vitro, T(R) cells effectively inhibited both the proliferation of and cytokine production by MOG(35-55)-specific Th1 cells. In vivo, adoptive transfer of T(R) cells conferred significant protection from clinical experimental autoimmune encephalomyelitis which was associated with normal activation of autoreactive Th1 cells, but an increased frequency of MOG(35-55)-specific Th2 cells and decreased CNS infiltration. Lastly, transferred T(R) cells displayed an enhanced ability to traffic to the peripheral lymph nodes and expressed increased levels of the adhesion molecules ICAM-1 and P-selectin that may promote functional interactions with target T cells. Collectively, these findings suggest that T(R) cells contribute notably to the endogenous mechanisms that regulate actively induced autoimmune disease.     

IFN-gamma and IL-17 production in experimental autoimmune encephalomyelitis depends on local APC-T cell complement production

IFN-gamma- and IL-17-producing T cells autoreactive across myelin components are central to the pathogenesis of multiple sclerosis. Using direct in vivo, adoptive transfer, and in vitro systems, we show in this study that the generation of these effectors in myelin oligodendrocyte glycoprotein(35-55)-induced experimental autoimmune encephalomyelitis depends on interactions of locally produced C3a/C5a with APC and T cell C3aR/C5aR. In the absence of the cell surface C3/C5 convertase inhibitor decay-accelerating factor (DAF), but not the combined absence of DAF and C5aR and/or C3aR on APC and T cells, a heightened local autoimmune response occurs in which myelin destruction is markedly augmented in concert with markedly more IFN-gamma(+) and IL-17(+) T cell generation. The augmented T cell response is due to increased IL-12 and IL-23 elaboration by APCs together with increased T cell expression of the receptors for each cytokine. The results apply to initial generation of the IL-17 phenotype because naive CD62L(high) Daf1(-/-) T cells produce 3-fold more IL-17 in response to TGF-beta and IL-6, whereas CD62L(high) Daf1(-/-)C5aR(-/-)C3aR(-/-) T cells produce 4-fold less.     

Therapeutic effect of omega-3 fatty acids on T cell-mediated autoimmune diseases

The present study was to demonstrate that the G protein coupled receptors serve as targets for the treatment of autoimmune disease such as rheumatoid arthritis and multiple sclerosis. Rats received pristane at the base of the tail. Affected joints were counted daily. The T cell mediated autoimmune diseases such as pristine-induced arthritis (PIA) and autoimmune encephalomyelitis (EAE) in a rat model were profoundly ameliorated by treatment with the specific G protein couple receptors 120 (GPR120) stimuli omega-3 fatty acids (ω-3 FAs). Our study further revealed that the activation of GPR120 by ω-3 FAs can result in a decrease of phosphorylated transforming growth factor-β activated kinase 1 (TAK1), and further inhibit the downstream IKKβ/I-κB pathway and the terminal NF-κB activation which serves as a mediator of T cell activation. ω-3 Fatty acids exhibited an inhibitory effect on TAK1 by enhancing the association of β-arrestin2 and TAK1 binding protein 1 (TAB1), thus the disassociation of TAB1 from the TAB1/TAK1 complex renders a limited effect on β-arrestin2 signaling as an innate immunity regulation. GPR120 is a functional receptor of ω-3 fatty acids in T cell-mediated autoimmune disease compared with its effect on innate immunity.     

Treatment of passive experimental autoimmune encephalomyelitis in SJL mice with a recombinant TCR ligand induces IL-13 and prevents axonal injury

The major goal of this study was to evaluate the efficacy and mechanism of a rTCR ligand (RTL) construct (I-A(s)/proteolipid protein (PLP)-139-151 peptide = RTL401) for treatment of SJL/J mice developing passive experimental autoimmune encephalomyelitis (EAE) that did not involve coimmunization with the highly inflammatory CFA. Our results demonstrated clearly that RTL401 was highly effective in treating passive EAE, with kinetics of recovery from disease very similar to treatment of actively induced EAE. The potent RTL401 treatment effect was reflected by a partial reduction of infiltrating mononuclear cells into CNS, minimal inflammatory lesions in spinal cord, and preservation of axons injured in vehicle-treated mice during the progression of EAE. Interestingly, in the absence of CFA, RTL401 treatment strongly enhanced production of the Th2 cytokine, IL-13, in spleen, blood, and spinal cord tissue, with variable effects on other Th1 and Th2 cytokines, and no significant effect on the Th3 cytokine, TGF-beta1, or on FoxP3 that is expressed by regulatory T cells. Moreover, pretreatment of PLP-139-151-specific T cells with RTL401 in vitro induced high levels of secreted IL-13, with lesser induction of other pro- and anti-inflammatory cytokines. Given the importance of IL-13 for protection against EAE, these data strongly implicate IL-13 as a dominant regulatory cytokine induced by RTL therapy. Pronounced IL-13 levels coupled with marked reduction in IL-6 levels secreted by PLP-specific T cells from blood after treatment of mice with RTL401 indicate that IL-13 and IL-6 may be useful markers for following effects of RTL therapy in future clinical trials in multiple sclerosis.     

Late pregnancy suppresses relapses in experimental autoimmune encephalomyelitis: evidence for a suppressive pregnancy-related serum factor

Women with multiple sclerosis have significantly diminished disease activity during pregnancy. The purpose of our study was to identify the underlying mechanism for the diminished disease activity. We found that during the period of late pregnancy there is protection against paralysis, during both the induction and effector phases of relapsing experimental autoimmune encephalomyelitis, a mouse model of multiple sclerosis. We did not find any changes in the cytokine secretion profiles or the proliferative activity of autoreactive T cells from mice induced during late pregnancy compared with virgin controls. In mice mated after disease onset, the inflammatory histologic lesions did not clear, despite marked clinical improvement during pregnancy. We found evidence for a serum factor present in late pregnancy that suppresses T cell activation. In the presence of sera taken from mice late in pregnancy, the proliferative response and IL-2 production of proteolipid protein p139-151-specific T cells were significantly diminished as compared with stimulation in the presence of normal mouse sera. In conclusion, serum from late pregnancy has the capacity to down-regulate T cell responses and might be associated with the amelioration of disease activity in experimental autoimmune encephalomyelitis.     

TGF-beta as a T cell regulator in colitis and colon cancer

TGF-beta is a pleiotropic cytokine with powerful immunosuppressive functions. Mice deficient for TGF-beta1 show a dramatic phenotype with severe multiorgan inflammation and die shortly after birth. Recent investigations have highlighted the role of TGF-beta in suppression of T cell mediated autoimmune inflammation and anti-tumor immunity. In addition to its direct anti-inflammatory effects on T cells, TGF-beta has been implicated as central regulator of regulatory T cells. TGF-beta not only mediates the suppression of effector T cells by Tregs, recent evidence also reveals a role for TGF-beta along with TCR stimulation in the peripheral induction of regulatory T cells from na?ve CD4+CD25- cells.     

Coronin 1-mediated naive T cell survival is essential for the development of autoimmune encephalomyelitis

Autoimmune encephalomyelitis is a disease of the CNS that can develop when an initial peripheral inflammatory stimulus is followed by infiltration and reactivation of T lymphocytes in the CNS. We report a crucial role for coronin 1, which is essential for maintenance of the naive T cell pool, for the development of murine experimental autoimmune encephalomyelitis (EAE), a model for multiple sclerosis. In the absence of coronin 1, immunization with myelin oligoglycoprotein (MOG(35-55)) peptide largely failed to induce EAE symptoms, despite normal mobilization of leukocyte subsets in the blood, as well as effector cytokine expression comparable with wild-type T cells on polyclonal stimulation. Susceptibility of coronin 1-deficient mice to EAE induction was restored by transfer of wild-type CD4(+) T cells, suggesting that the observed resistance of coronin 1-deficient mice to EAE development is T cell intrinsic. Importantly, although coronin 1-deficient regulatory T cells (Tregs) showed a suppressor activity comparable with wild-type Tregs, Treg depletion failed to restore EAE development in coronin 1-deficient animals. These results suggest a hitherto unrecognized role of naive T cells in the development of autoimmune encephalomyelitis and reveal coronin 1 as a crucial modulator of EAE induction.     

IL-17 plays an important role in the development of experimental autoimmune encephalomyelitis

IL-17 is a proinflammatory cytokine that activates T cells and other immune cells to produce a variety of cytokines, chemokines, and cell adhesion molecules. This cytokine is augmented in the sera and/or tissues of patients with contact dermatitis, asthma, and rheumatoid arthritis. We previously demonstrated that IL-17 is involved in the development of autoimmune arthritis and contact, delayed, and airway hypersensitivity in mice. As the expression of IL-17 is also augmented in multiple sclerosis, we examined the involvement of this cytokine in these diseases using IL-17(-/-) murine disease models. We found that the development of experimental autoimmune encephalomyelitis (EAE), the rodent model of multiple sclerosis, was significantly suppressed in IL-17(-/-) mice; these animals exhibited delayed onset, reduced maximum severity scores, ameliorated histological changes, and early recovery. T cell sensitization against myelin oligodendrocyte glycoprotein was reduced in IL-17(-/-) mice upon sensitization. The major producer of IL-17 upon treatment with myelin digodendrocyte glycopritein was CD4+ T cells rather than CD8+ T cells, and adoptive transfer of IL-17(-/-) CD4+ T cells inefficiently induced EAE in recipient mice. Notably, IL-17-producing T cells were increased in IFN-gamma(-/-) cells, while IFN-gamma-producing cells were increased in IL-17(-/-) cells, suggesting that IL-17 and IFN-gamma mutually regulate IFN-gamma and IL-17 production. These observations indicate that IL-17 rather than IFN-gamma plays a crucial role in the development of EAE.     

TGF-beta1: immunosuppressant and viability factor for T lymphocytes

Transforming growth factor-beta (TGF-beta) is a multifunctional cytokine with multiple roles in the immune system. To date, it has been difficult to develop a comprehensive picture of the effect of TGF-beta on T lymphocytes, because TGF-beta not only acts directly on T lymphocytes, but also acts indirectly by regulating the function of antigen-presenting cells. In early studies, it was mostly the inhibitory function of TGF-beta that was demonstrated; recently, however TGF-beta was recognized as an antiapoptotic survival factor for T lymphocytes. The outcome of the TGF-beta effect on T lymphocytes was shown to strongly depend on their stage of differentiation and on the cytokine milieu. TGF-beta cannot be classified as a classical Th1 or Th2 cytokine. However, recently the existence of the TGF-beta-producing Th3 subset was described which might play an important regulatory role during an immune response. A better understanding of the molecular mechanism of how TGF-beta inhibits or stimulates T lymphocytes will help to predict the complex functions of this cytokine.     

Critical requirement of CD11b (Mac-1) on T cells and accessory cells for development of experimental autoimmune encephalomyelitis

Mac-1 (CD18/CD11b) is a member of the beta2-integrin family of adhesion molecules and is implicated in the development of many inflammatory diseases. The role of Mac-1 in the development of CNS demyelinating diseases, including multiple sclerosis, is not understood, and Ab inhibition studies in experimental allergic encephalomyelitis (EAE), the animal model for multiple sclerosis, have produced conflicting findings. To clarify these results and to determine Mac-1-mediated mechanisms in EAE, we performed EAE using Mac-1-deficient mice. Mac-1 homozygous-deficient, but not Mac-1 heterozygous-deficient mice, had significantly delayed onset and attenuated EAE. Leukocyte infiltration was similar in both groups of mice in early disease but significantly reduced in spinal cords of receptor-deficient mice in late disease. Adoptive transfer of Ag-restimulated T cells from wild-type to Mac-1-deficient mice produced significantly attenuated EAE, whereas transfer of Mac-1-deficient Ag-restimulated T cells to control mice failed to induce EAE. T cells from myelin oligodendrocyte glycoprotein (MOG)35-55 peptide-primed Mac-1-deficient mice displayed an altered cytokine phenotype with elevated levels of TGF-beta and IL-10, but reduced levels of IL-2, IFN-gamma, TNF-alpha, IL-12, and IL-4 compared with control mice. Mac-1-deficient T cells from primed mice proliferated comparably to that of control T cells on MOG35-55 restimulation in vitro. However, the draining lymph nodes of MAC-1-deficient mice on day 10 after MOG35-55 immunization contained lower frequency of blast T cells than in control mice, suggesting poor priming. Our results indicate that Mac-1 expression is critical on both phagocytic cells and T cells for the development of demyelinating disease.