2). Administration of anti-IL-1R1 to R258W KI mice results in nearly complete reversal of skin inflammation 9. This result parallels findings in humans with CAPS who usually manifest striking

clinical improvement upon treatment with IL-1β Stem Cell Compound Library in vitro blocking agents 31–33, and supports the view that IL-1β is the main, if not the sole, basis of the inflammation. In contrast, administration of an IL-1β blocking agent (mIL-1 Trap) to A350V and L351P KI mice resulted in virtually no improvement in the inflammatory state, although IL-1R1−/− mice bearing these mutations do not show inflammation 10. This outcome could be the result of the intense NLRP3 inflammasome activity in these mice that leads to effects such as cell necrosis that are not easily reversed by an exogenous agent that neutralizes IL-1β 34, 35. Given the Th17-cell bias of the inflammation in R258W KI mice, the effect of administration of anti-IL-17A was also assessed. Interestingly, this agent was also effective in ameliorating inflammation, despite the fact that it does not block the inflammatory effect of IL-17F, an IL-17 isotype also elevated in lesions

9. These studies suggest that if humans with CAPS can also be shown to have inflammations with a Th17-cell bias, it may be possible PLX4032 nmr to control CAPS with anti-IL-17 as well as IL-1β inhibitory agents. It is evident from selleck products the studies described above that mice carrying mutations of the Nlrp3 gene have already yielded valuable new insights into the immunopathology associated with CAPS, including a possible new treatment approach. Further studies in which these mice are used to elucidate the

role of the NLRP3 inflammasome in various types of organ-specific inflammation hold the promise of defining the role of the inflammasome in a host of inflammatory conditions. This work was supported by the Division of Intramural Research, National Institute of Allergy and Infectious Diseases, National Institutes of Health. We apologize to those authors whose work could not be cited due to space limitations. Conflict of interest: The authors declare no financial or commercial conflict of interest. See accompanying Viewpoint: http://dx.doi.org/10.1002/eji.200940225 “
“Department Clinical Genetics, Erasmus MC Rotterdam, The NetherlandsDr. Sabine Middendorp, Pediatric Gastroenterology, Wilhelmina Children’s Hospital, University Medical Center Utrecht, The Netherlands B-cell receptor (BCR)-mediated signals provide the basis for B-cell differentiation in the BM and subsequently into follicular, marginal zone, or B-1 B-cell subsets. We have previously shown that B-cell-specific expression of the constitutive active E41K mutant of the BCR-associated molecule Bruton’s tyrosine kinase (Btk) leads to an almost complete deletion of immature B cells in the BM.

During IFN-α signaling, the activated STAT1 dimer and ISGF3 (STAT1:STAT2:p48) complex each acquires exposed nuclear localization signals. Importin-α/β complex recognizes these signals and induces translocation of the STAT complexes into the nucleus 35, 38, 39. However, no mechanisms governing the nuclear localization Ku-0059436 ic50 of STAT6 have been identified

yet, except that the translocation of STAT6 depends on its phosphorylation on Y641 39. In this regard, we have noted that the cytoplasmic retention complex containing pY-STAT6 did not interact with importin-α (Supporting Information Fig. S3, left panel). Interestingly, with increased association of pY-STAT6 with pY-STAT2 and p48 during IFN-α treatment from 0.5 to 4 h, there is a decreased interaction of pY-STAT1 with pY-STAT2 and p48 (Supporting Luminespib mw Information Fig. S3, right panel). The observation raises a possibility that by the action of IFN-α-induced factors during 4 h treatment, pY-STAT1 gradually dissociates from the ISGF3 complex, which is then replaced with IL-4-activated pY-STAT6. This would result in the sequestration of STAT6 from the translocatable STAT6 homodimer to form the putative pY-STAT6:pY-STAT2:p48 complex incapable of importin binding and nuclear translocation, which is then retained in the cytosol. On the other hand,

it is also possible that pY-STAT6 is accumulated in the cytoplasm upon the inhibition of translocation mediated by IFN-α-induced factors, which then interacts with pY-STAT2 and p48. Several post-translational modifications other than tyrosine phosphorylation may be involved in the formation of the STAT complex retained in the cytosol, since STAT6 and/or STAT2 are thought to undergo serine/threonine

phosphorylation, acetylation, and sumoylation. Yet, in our experimental system, we have observed no Isotretinoin detectable changes in these modifications on STAT6 or STAT2 by IFN-α and IL-4, which suggests that such post-translational modifications may not play a role in the molecular interaction and cytosolic accumulation of the STAT complex. As shown by the inhibition of the IL-4-induced CD23 expression and the IFN-α-induced IRF7 expression, a novel feature of the IFN-α and IL-4-induced cross-signaling found in the present study is the cytoplasmic co-retention of activated STATs (Figs. 3, and 4). By coimmunoprecipitation experiments, we have verified the molecular interaction among pY-STAT6, pY-STAT2, and p48 induced in cells upon treatment with IL-4 and IFN-α (Fig. 5A), which strongly suggests a possibility that these proteins are present in a molecular complex. To further examine the possibility that the inhibition by IFN-α and IL-4 is mediated via the formation and cytoplasmic retention of the pY-STAT6:pY-STAT2:p48, the effect of STAT over-expression was analyzed.

These PRRs can detect a broad range of molecular patterns that are associated with either infection (pathogen-associated molecular pattern; PAMPs) [2, 3] or cell death and trauma (damage-associated molecular patterns; DAMPs) [2, 4]. Following activation through PRRs, DCs undergo a maturation process that is characterized by upregulation of MHC class II and costimulatory molecules on their cell surface, proinflammatory

cytokine production, and DC migration to draining lymph nodes. In the lymph nodes, mature DCs function as the prototype of professional APCs to prime naïve T cells and control T-cell activation [5]. In addition to detecting pathogens or tissue damage directly through PRRs, DCs can be indirectly activated by factors that signal the presence of pathogens. For example, type I interferons (IFNs), which are produced rapidly in the course of viral and bacterial infections, GSK-3 inhibitor have been reported to enhance the Ag-presentation LY294002 supplier efficiency of DCs, as well as DC migration to lymphoid tissues [6]. Moreover, type I IFN receptor signaling in DCs has been found to be essential for

T-cell priming in response to various PAMPs [7], as well as for the induction of virus-specific [8] and tumor-specific T-cell responses [9]. Notably, the interaction of DCs with CD4+ T cells provides additional important stimuli for DC maturation [10]. For example, ligation of CD40 on DCs by CD154 on T cells promotes DC activation, leading to priming of cytotoxic T lymphocytes (CTLs) [11] and CD4+ T-cell differentiation. Over the past decade, it has become clear that, in addition to their role in priming effector T-cell responses against invading pathogens, DCs have a crucial role in self-tolerance. These opposing DC functions are controlled through the regulation of DC maturation in the steady state, and this checkpoint is crucial for the maintenance

of immune homeostasis. In this article, ID-8 we review the signals that can induce DC maturation in the steady state and discuss the suppressive mechanisms that counterbalance DC-activating signals to preserve peripheral tolerance. The contribution of steady-state DCs to the maintenance of peripheral tolerance was first shown in animal models, in which Ag could be targeted to immature DCs. Immature steady-state DCs had previously been notoriously difficult to study, as their isolation and manipulation rapidly induce DC maturation [12, 13]. To overcome this problem, the group of Ralph Steinman used mAbs against DC surface receptors to target Ags to DCs in vivo. Antigen delivery to steady-state DCs in the absence of inflammatory signals resulted in a transient activation and proliferation of Ag-specific CD4+ and CD8+ T cells, which was followed by deletion of these T cells and the establishment of Ag-specific T-cell tolerance [14, 15].

Coresh et al.20 estimated the population several times, with refinements in assumptions and in the estimating equations used to define estimated glomerular filtration rate (eGFR), most recently with an improved equation21 that corrects for underestimated eGFR more than 60 mL/min per 1.73 m2. The newest estimates place the CKD population at 11% of

the general population, versus 13% based on the older Modification of Diet in Renal Disease (MDRD) estimating equation.20 Of note, the CKD-EPI equation21 reduces bias in underestimating GFR more than 60 mL/min per 1.73 m2 compared with the MDRD estimating equation.20 The CKD-EPI equation should be considered for implementation in screening programs; it will reduce the number of false positives and selleck chemicals llc improve the accuracy of testing for kidney disease. Whether the estimate is 26 million people or the newer 21 million people, the size of this population is substantial. Almost a million C646 people are at stage 4 CKD; they are just one stage from entering the ESRD incident population, but are far more likely to die before developing ESRD. These estimates are consistent around the world, as reports from China,7 Japan,22 Australia10 and the Democratic Republic of the Congo12 give estimates of 10–14% of the population having evidence of

CKD using methods similar to methods used by Coresh et al.20 and Levey et al.21 The future number of potential ESRD patients is considerable unless contravening measures limit progression and the competing event of death reduces the number of CKD patients who reach ESRD. Because major public

health programs have been focused on reducing death rates from major diseases, efforts to slow progression of kidney disease will be needed – along with longer-term lifestyle changes – to reduce the at-risk population with diabetes and hypertension. Several reports have shown that hypertension, diabetes and cardiovascular disease increase with decreasing eGFR (Fig. 2). Similar findings were reported in the Taiwanese population studied for evidence of CKD.15 A similar pattern is noted when kidney damage is defined by increasing albumin-to-creatinine ratio (Fig. 3). This level of comorbidity Levetiracetam is associated with increasing cardiovascular event rates and mortality with advancing CKD stage,14,15 providing evidence that the highest rates of complications in the CKD population occur for patients with evidence of diabetes and cardiovascular disease. The observation of low recognition of CKD (12% of the population in Taiwan show evidence of CKD, but only 3% of patients with evidence of CKD were aware of it) demonstrates the challenge of engaging people in proactively seeking care and adhering to medical therapy to reduce the risk of future adverse events, premature death and progression to ESRD. In the study by Go et al.

The purpose of this study was to evaluate the effect of vitamin A supplementation

on expression of Th17 cells-related IL-17 and RORc genes in atherosclerotic patients. Thirty one atherosclerotic patients and 15 healthy controls were studied for 4 months. Atherosclerotic patients were randomly divided into vitamin A or placebo groups. Healthy controls and patients in vitamin A group received 25,000 IU retinyl palmitate per day. Peripheral blood mononuclear cells Ibrutinib cost were isolated, cultured and divided into three groups including fresh cells, phytohemagglutinin (PHA)-activated T cells and ox-LDL-activated T cells. Gene expressions of T cells were studied by real-time PCR. In atherosclerotic patients, vitamin A supplementation resulted in significant decrease in IL-17 gene expression by 0.63-fold in fresh

cell, 0.82-fold in PHA-activated cells and 0.65-fold in ox-LDL-activated cells (P < 0.05 for all). RORc gene expression in fresh cells as well as ox-LDL-activated cells decreased significantly after vitamin A supplementation in atherosclerotic patients (P = 0.0001 for both). In PHA-activated cells, vitamin A supplementation significantly decreased RORc gene NVP-BKM120 manufacturer in both atherosclerotic patients and healthy subjects by 0.87-fold and 0.72, respectively, while in placebo group, the RORc gene expression significantly increased by 1.17-fold (P < 0.05 for all). Findings of this study suggest that vitamin A supplementation may be an effective approach to slow progression of atherosclerosis. "
“Dendritic cells (DCs) are master regulators of T-cell responses. After sensing pathogen-derived molecular patterns (PAMPs), or signals of inflammation Org 27569 and cellular stress, DCs differentiate into potent activators of naïve CD4+ and

CD8+ T cells through a process that is termed DC maturation. By contrast, DCs induce and maintain peripheral T-cell tolerance in the steady state, that is in the absence of overt infection or inflammation. However, the immunological steady state is not devoid of DC-activating stimuli, such as commensal microorganisms, subclinical infections, or basal levels of proinflammatory mediators. In the presence of these activating stimuli, DC maturation must be calibrated to ensure self-tolerance yet allow for adequate T-cell responses to infections. Here, we review the factors that are known to control DC maturation in the steady state and discuss their effect on the tolerogenic function of steady-state DCs. Since their discovery by Steinman and Cohn in the 1970s [1], it has become clear that dendritic cells (DCs) are key inducers and regulators of immune responses.

After 24 h of culture, the CTLL cells were pulsed with [3H]thymidine for an additional 4 h and the net cpm (mean±SD) click here were calculated. HLA-DR2 mice between 8 and 12 wk of age were immunized s.c. at four sites on the flanks with 0.2 mL of an emulsion of 200 μg mouse MOG-35-55 peptide and complete Freund’s adjuvant containing 400 μg

of Mycobacterium tuberculosis H37RA (Difco, Detroit, MI, USA). In addition, mice were given Ptx from List Biological Laboratories (Campbell, CA, USA) on days 0 and 2 post immunization (75 and 200 ng per mouse, respectively). HLA-DR2 mice were treated with vehicle, RTL342m alone, or RTL342m pre-incubated with one of the FAbs beginning on the first day that the combined clinical EAE score for each individual mouse reached

2 or higher. Once-daily treatments were administered to each mouse subcutaneously in the interscapular region for three days. RTL342m and RTL342m+FAb were prepared in 5-Fluoracil in vitro 100 μL of 20 mM Tris-HCl pH 8.0 with 5% w/v D-glucose (Sigma-Aldrich, St. Louis, MO, USA). Vehicle treatments consisted of only Tris-HCl pH 8.0 with 5% w/v D-glucose. Mean EAE scores and SDs for mice grouped according to initiation of RTL or vehicle treatment were calculated for each day. The CDI was determined for each mouse by summing the daily EAE scores. Group CDI scores were calculated by determining the mean±SD of the individual mice in the group. The IACUC Protocol ♯2108, Vandenbark AA PI, was in

place and is currently approved for the animal experiments reported in the manuscript. Detection of RTL-like material in human serum or plasma was determined by ELISA using Fab 1B11. ELISA plates (Falcon) were coated for 2 h with anti-MHC mAb TU39 (10 μg/well). The plates were blocked for 30 min at room temperature with PBS/2% skim milk Tangeritin and subsequently were incubated for 2 h at room temperature with serial dilutions of RTL1000 (for standard curve) and 1:10 serum dilutions. After being washed, the plates were incubated (1 h at room temperature) with 1B11 Fab (10 μg/mL), washed extensively and further incubated (1 h at room temperature) with anti-myc-biotin Ab (9E10 clone, Covance). The plates were washed and incubated for 30 min with HRP-conjugated streptavidin. Further amplification steps were performed using the ELAST ELISA amplification system (PerkinElmer), according to the manufacturer’s protocol. Detection was performed using TMB reagent (Sigma). Detection of RTL1000 in human serum or plasma was determined by ELISA using biotinylated Fab 2E4. ELISA plates (Falcon) were coated overnight with BSA-biotin (1 μg/well). After being washed, the plates were incubated (1 h at room temperature) with streptavidin (10 μg/mL), washed extensively and further incubated (1 h at room temperature) with 5 μg/mL of biotinylated Fab 2E4.

While there is a clear role for MyD88 in the ability of conventional mice to mount neutrophilic inflammation to zymosan, we found that several other innate immune signalling pathways were not required for this response. Although Clarke et al. have reported that commensal bacteria prime neutrophils via NOD1 signalling in ways that enhance their phagocytic potential to various bacteria,[16] we found selleck chemical that RIP2 knockout mice did not show reduced inflammation to zymosan. Since RIP2 is required for NOD1/2 signalling, this finding argued against a role for either NOD1 or NOD2 in mediating a gut flora-induced effect in our system.[32] Therefore, NOD1/2 signalling may be important for phagocytosis

but is not needed for neutrophilic inflammation to this agent. Similarly, we found no contribution of the inflammasome components (NLRP3/ASC/caspase 1) or the RNA-sensing RIG-I like receptors MK-1775 purchase in mediating zymosan-induced inflammation. Hence, we show that intestinal flora affect the ability of the immune system to mount neutrophilic inflammation

via the MyD88 pathway. To examine when the MyD88 pathway was required, we took advantage of the ROSA26-Cre system, in which the MyD88 gene could be temporally deleted by the addition of tamoxifen. We showed that for zymosan-induced peritonitis, the presence of MyD88 was not required at the time of challenge. This eliminates the possibility that zymosan Liothyronine Sodium needs to signal through MyD88 via TLR2 or IL-1R or any other MyD88-dependent receptor. These data therefore, make a strong case for the necessity of priming by intestinal flora-induced MyD88 activation for zymosan-induced neutrophil migration, before the actual zymosan challenge. Hence a significant finding of this study is that although the MyD88 pathway is essential for creating an innate immune system

that is poised to respond to inflammatory agent, this pathway is not needed at the elicitation phase of an inflammatory response (unless of course the pro-inflammatory stimulus was using MyD88-dependent receptors such as TLRs). An implication of our study is that the set point of the naive (i.e. never exposed to microbes) innate immune system may be anti-inflammatory for many stimuli. However, in conventionally reared mice the immune system is perturbed by exposure to microbial flora in ways that alter the cytokines that are made. As part of this process MyD88-dependent pattern recognition receptor signalling by microbial flora appears to alter this set point in ways that promote inflammatory responses. In summary, we postulate that TLR ligands derived from the intestinal flora constitutively enter the blood and tissues. Here, they prime tissue-resident cells via MyD88 signalling, so that they provide appropriate stimulatory signals that condition the innate immune system to be able to respond to future inflammatory insults in ways that promote neutrophil migration into tissue sites.

33 Smad3 plays an essential

role in TGF-β1-induced EMT.34 Evidence of renal EMT has been obtained by numerous independent studies in different animal models of chronic renal disease and also in human kidney biopsies.35–38 The inverse correlation between increasing numbers of tubular epithelial cells undergoing EMT and decline of excretory renal function suggests a pathological role of EMT in the progression of renal fibrosis.39,40 The observation that reversal of EMT improved renal function and decreased mortality in a mouse model with nephrotoxic serum nephritis further confirmed the importance of EMT in the progression of chronic renal disease.34 Advanced glycation end-product (AGE)-induced EMT has been implicated in the pathogenesis of DN.41 TGF-β1, AGE, high glucose,42 angiotensin II43 and oxidative stress44 are also key EMT inducers, shown to be involved in the development and progression of diabetic renal find more fibrosis. Endothelium is a simple squamous epithelium, a specialized type of epithelial tissue. Y-27632 clinical trial Thus, EndoMT can be considered to be a specific form of EMT. EndoMT is an essential mechanism in cardiac development.45 During heart valve formation, a subset of EC overlying the future valve site delaminate, differentiate into mesenchymal cells and migrate into the cardiac jelly to form cardiac cushions, a process

referred to as endothelial-mesenchymal transition.46 Disruption of Notch signalling results in failure of EndoMT, revealing an essential role for notch in the control of endocardial cushion EndoMT.47,48 Evidence that wnt/β-catenin signalling was restricted to a subset of mesenchymal cells in endocardial cushions in the developing mouse heart49 and that antagonism of wnt/β-catenin signalling in zebrafish embryos inhibited cardiac cushion EndoMT suggested wnt/β-catenin signalling may activate expression of genes crucial for EndoMT.49β-catenin also acts as a structural link between actin and Vascular Endothelial Cadherin

(VE-cadherin) to form the cell–cell adherens junction necessary for polarity of EC.50 Bone morphogenetic proteins 2 and 4 (BMP-2 and 4), TGF-β2 and TGF-β3 are required for initiation click here and completion of EndoMT.46 The role of TGF-β and BMP signalling pathways in endocardial cushion EndoMT has been thoroughly studied.51,52 Recent studies have demonstrated that EndoMT contributes to the development of tissue fibrosis. Zeisberg et al.53 used Tie1Cre; R26RstoplacZ mice to track cells of endothelial origin, and placed aortic bands on the hearts of mice to induce cardiac fibrosis. They showed that EC undergo EndoMT during cardiac fibrosis and contribute to the total pool of cardiac fibroblasts. In addition, they showed that TGF-β1 induced EndoMT, whereas BMP7 abrogated EndoMT, preserved the endothelial phenotype and reversed or prevented TGF-β1-induced EndoMT and cardiac fibrosis.

The tumor cells were periodic acid Schiff positive, diastase resistant, and were positive with S-100 protein, CD68,

inhibin, and neuron-specific enolase immunohistochemistry. The clinical and histologic differential diagnosis includes schwannoma, neurofibroma, meningioma, astrocytoma, melanocytoma, and metastatic tumors. Patients were managed LY2109761 in vitro with excision. One patient had symptomatic and radiographic local recurrence that was subsequently treated with radiation, resulting in stabilization of disease and symptoms. Intradural GCTs of the spine are rare and radiographically indistinguishable from tumors that more commonly arise in this location. Histologic recognition of this rare tumor is important because the subsequent clinical course of the disease differs from other similar lesions. “
“Anaplastic large cell lymphoma (ALCL) is characterized by large anaplastic cells of T-cell or null-cell phenotype expressing CD30 (Ki-1 antigen). In most cases this neoplasm expresses the anaplastic lymphoma kinase (ALK), a chimeric protein resulting from the t(2;5)(p23;q35) translocation. ALK-positive

anaplastic large cell lymphoma is most frequent in the first three decades of life and shows a male predominance, involving both nodal and extranodal sites, but rarely the CNS. We report a 21-year-old patient with a previous history of nodal ALK-positive ALCL, lymphohistiocytic subtype, who was admitted for recent occurrence of left-sided anesthesia with pain and progressive motor weakness of both legs. An MRI of the spine documented an intradural extramedullary buy CP-868596 mass dislocating the thoracic cord, suggesting a meningioma and the patient underwent

surgical decompression. Histological examination revealed a lymphoproliferative neoplasm with morphology and immunophenotype of ALK-positive anaplastic large cell lymphoma. After surgery, all preoperative selleckchem symptoms disappeared. To our knowledge, no cases of ALCL presenting as secondary localization with an intradural extramedullary spinal mass have been reported in the literature. “
“M. Jansen, G. Mohapatra, R. A. Betensky, C. Keohane and D. N. Louis (2012) Neuropathology and Applied Neurobiology38, 213–219 Gain of chromosome arm 1q in atypical meningioma correlates with shorter progression-free survival Aims: Atypical (World Health Organization grade II) meningiomas have moderately high recurrence rates; even for completely resected tumours, approximately one-third will recur. Post-operative radiotherapy may aid local control and improve survival, but carries the risk of side effects. More accurate prediction of recurrence risk is therefore needed for patients with atypical meningioma. Previously, we used high-resolution array comparative genomic hybridization to identify genetic variations in 47 primary atypical meningiomas and found that approximately 60% of tumours show gain of 1q at 1q25.1 and 1q25.3 to 1q32.

We also reported that Tim-4 could bind to Tim-1 and regulate T-cell responses

12. Interestingly, treatment with Tim-4-hFc fusion proteins did not change DCs function in terms of the expression of CD80, CD86, and MHC class II molecules (Supporting Information Fig. 7). However, Tim-4 also binds to PS 35, 36 and potentially another unknown receptor 38. Thus, without knowing whether DCs express other Tim-4-binding protein(s) Sunitinib manufacturer in addition to Tim-1, it is difficult to understand whether the effect of Tim-4-hFc on DCs is through Tim-1 and/or other pathway(s). These issues will only be clearly addressed using Tim-1 deficient mice, which just became available most recently 15. In summary, we show that Tim-1 plays different roles in the innate and adaptive Raf inhibitor immune responses. Since Tim-1 is constitutively expressed on DCs in the steady state, Tim-1 is readily available for crosslinking on DCs before it is even expressed on adaptive immune cells. The present study highlights the role of Tim-1 expressed on DCs in regulating the balance between effector and regulatory T cells and thus regulating immune responses. A better

understanding of the mechanism by which Tim-1 regulates DC and T cell responses will provide a target by which DC/T cell functions can be regulated so as to treat inflammatory diseases including autoimmune diseases, and to improve vaccination and tumor immunotherapy. SJL mice were purchased from The Jackson Laboratory. B10.S mice and 5B6 SJL mice transgenic for the PLP139–151-specific TCR 5B6 have been described previously 20. Foxp3/GFP ‘knock-in’ mice originally generated on the C57BL/6 background 26 were back-crossed for >10 generations onto the B10.S background. The mice were maintained, and all animal experiments were performed according to the animal protocol guidelines of Harvard Medical

School. PLP139–151 and OVA323–339 peptides were synthesized by Quality Controlled Biochemicals. Anti-Tim-1 antibodies 3B3 and RMT1-10 have been described previously 11, 16. Cytokines and antibodies Interleukin-3 receptor for FACS and ELISA were obtained from eBioscience, BD Biosciences, and R&D Systems. Different populations of immune cells were purified with MACS beads (Miltenyi Biotec). Naïve CD4+ T cells (CD4+CD62LhiCD25–) and DCs (CD11c+CD3−CD19−) were purified using a FACSAria cell sorter following MACS bead-isolation of CD4+ and CD11c+ cells, respectively. CNS-infiltrating mononuclear cells were isolated from mice with EAE as previously described 26, 27. Naïve CD4+ cells (1×106/well) were activated with either plate-bound anti-CD3/CD28 (1 μg/mL for both) or with PLP139–151 (25 μg/mL) plus syngeneic DCs (2×105/well) in the presence or absence of anti-Tim-1 (10 μg/mL).