Anti-CD52 antibody treatment in murine experimental autoimmune encephalomyelitis induces dynamic and differential modulation of innate immune cells in peripheral immune and central nervous systems.

Anti-CD52 antibody (anti-CD52-Ab) leads to a rapid depletion of T and B cells, followed by reconstitution of immune cells with tolerogenic characteristics. However, very little is known about its effect on innate immune cells. In this study, experimental autoimmune encephalomyelitis mice were administered murine anti-CD52-Ab to investigate its effect on dendritic cells and monocytes/macrophages in the periphery lymphoid organs and the central nervous system (CNS). Our data show that blood and splenic innate immune cells exhibited significantly increased expression of MHC-II and costimulatory molecules, which was associated with increased capacity of activating antigen-specific T cells, at first day but not three weeks after five daily treatment with anti-CD52-Ab in comparison with controls. In contrast to the periphery, microglia and infiltrating macrophages in the CNS exhibited reduced expression levels of MHC-II and costimulatory molecules after antibody treatment at both time-points investigated when compared to controls. Furthermore, the transit response of peripheral innate immune cells to anti-CD52-Ab treatment was also observed in the lymphocyte-deficient SCID mice, suggesting the changes are not a direct consequence of the mass depletion of lymphocytes in the periphery. Our study demonstrates a dynamic and tissue-specific modulation of the innate immune cells in their phenotype and function following the antibody treatment. The findings of differential modulation of the microglia and infiltrating macrophages in the CNS in comparison with the innate immune cells in the peripheral organs support the CNS-specific beneficial effect of alemtuzumab treatment on inhibiting neuroinflammation in multiple sclerosis patients.

Interleukin-16 inhibits sodium channel function and GluA1 phosphorylation via CD4- and CD9-independent mechanisms to reduce hippocampal neuronal excitability and synaptic activity.

Interleukin 16 (IL-16) is a cytokine that is primarily associated with CD4+ T cell function, but also exists as a multi-domain PDZ protein expressed within cerebellar and hippocampal neurons. We have previously shown that lymphocyte-derived IL-16 is neuroprotective against excitotoxicity, but evidence of how it affects neuronal function is limited. Here, we have investigated whether IL-16 modulates neuronal excitability and synaptic activity in mouse primary hippocampal cultures. Application of recombinant IL-16 impairs both glutamate-induced increases in intracellular Ca2+ and sEPSC frequency and amplitude in a CD4- and CD9-independent manner. We examined the mechanisms underlying these effects, with rIL-16 reducing GluA1 S831 phosphorylation and inhibiting Na+ channel function. Taken together, these data suggest that IL-16 reduces neuronal excitability and synaptic activity via multiple mechanisms and adds further evidence that alternative receptors may exist for IL-16.

Role of IL-33/ST2 signaling pathway in systemic sclerosis and other fibrotic diseases.

Systemic sclerosis (SSc) is a complex autoimmune disease characterised by fibrosis of the skin and multiple internal organs. Interleukin 33 (IL-33) has recently been investigated as a potential key player in the pathogenesis of SSc and other fibrotic diseases, owing to its effects on tissue fibrosis. Understanding how IL-33 is regulated and how it contributes to the development of fibrosis will be important to elucidate disease pathogenesis and may shed light on new areas for therapeutic development for patients. Here we discuss the recent research progress in our understanding of the role and the underlying mechanisms of IL-33/ST2 signaling pathway in SSc and other fibrotic diseases.

Failure of the Anti-Inflammatory Parasitic Worm Product ES-62 to Provide Protection in Mouse Models of Type I Diabetes, Multiple Sclerosis, and Inflammatory Bowel Disease.

Parasitic helminths and their isolated secreted products show promise as novel treatments for allergic and autoimmune conditions in humans. Foremost amongst the secreted products is ES-62, a glycoprotein derived from Acanthocheilonema viteae, a filarial nematode parasite of gerbils, which is anti-inflammatory by virtue of covalently-attached phosphorylcholine (PC) moieties. ES-62 has been found to protect against disease in mouse models of rheumatoid arthritis, systemic lupus erythematosus, and airway hyper-responsiveness. Furthermore, novel PC-based synthetic small molecule analogues (SMAs) of ES-62 have recently been demonstrated to show similar anti-inflammatory properties to the parent molecule. In spite of these successes, we now show that ES-62 and its SMAs are unable to provide protection in mouse models of certain autoimmune conditions where other helminth species or their secreted products can prevent disease development, namely type I diabetes, multiple sclerosis and inflammatory bowel disease. We speculate on the reasons underlying ES-62's failures in these conditions and how the negative data generated may help us to further understand ES-62's mechanism of action.

IL-33 attenuates the development of experimental autoimmune uveitis.

Interleukin-33 (IL-33) is associated with several important immune-mediated disorders. However, its role in uveitis, an important eye inflammatory disease, is unknown. Here, we investigated the function of IL-33 in the development of experimental autoimmune uveitis (EAU). IL-33 and IL-33 receptor (ST2) were expressed in murine retinal pigment epithelial (RPE) cells in culture, and IL-33 increased the expression of Il33 and Mcp1 mRNA in RPE cells. In situ, IL-33 was highly expressed in the inner nuclear cells of the retina of naïve mice, and its expression was elevated in EAU mice. ST2-deficient mice developed exacerbated EAU compared with WT mice, and administration of IL-33 to WT mice significantly reduced EAU severity. The attenuated EAU in IL-33-treated mice was accompanied by decreased frequency of IFN-γ+ and IL-17(+) CD4+ T cells and reduced IFN-γ and IL-17 production but with increased frequency of IL-5(+) and IL-4(+) CD4 T cells and IL-5 production in the draining lymph node and spleen. Macrophages from the IL-33-treated mice show a significantly higher polarization toward an alternatively activated macrophage phenotype. Our results therefore demonstrate that the endogenous IL-33/ST2 pathway plays an important role in EAU, and suggest that IL-33 represents a potential option for treatment of uveitis.

Emerging role of interleukin-33 in autoimmune diseases.

Interleukin-33 (IL-33) is a member of the IL-1 cytokine family. It predominantly induces type 2 immune responses and thus is protective against atherosclerosis and nematode infections but contributes to allergic airway inflammation. Interleukin-33 also plays a pivotal role in the development of many autoimmune diseases through mechanisms that are still not fully understood. In this review, we focus on the recent advances in understanding of the expression and function of IL-33 in some autoimmune disorders, aiming to provide insight into its potential role in disease development.

IL-33 attenuates EAE by suppressing IL-17 and IFN-γ production and inducing alternatively activated macrophages.

Interleukin (IL)-33, a member of the IL-1 cytokine family, is an important modulator of the immune system associated with several immune-mediated disorders. High levels of IL-33 are expressed by the central nervous system (CNS) suggesting a potential role of IL-33 in autoimmune CNS diseases. We have investigated the expression and function of IL-33 in the development of experimental autoimmune encephalomyelitis (EAE) in mice. We report here that IL-33 and its receptor ST2 (IL-33Rα) are highly expressed in spinal cord tissue, and ST2 expression is markedly increased in the spinal cords of mice with EAE. Furthermore, ST2-deficient (ST2(-/-) ) mice developed exacerbated EAE compared with wild-type (WT) mice while WT, but not ST2(-/-) EAE mice treated with IL-33 developed significantly attenuated disease. IL-33-treated mice had reduced levels of IL-17 and IFN-γ but produced increased amounts of IL-5 and IL-13. Lymph node and splenic macrophages of IL-33-treated mice showed polarization toward an alternatively activated macrophage (M2) phenotype with significantly increased frequency of MR(+) PD-L2(+) cells. Importantly, adoptive transfer of these IL-33-treated macrophages attenuated EAE development. Our data therefore demonstrate that IL-33 plays a therapeutic role in autoimmune CNS disease by switching a predominantly pathogenic Th17/Th1 response to Th2 activity, and by polarization of anti-inflammatory M2 macrophages.

Regulation of autologous immunity to the mouse 5T4 oncofoetal antigen: implications for immunotherapy.

Effective vaccination against tumour-associated antigens (TAA) such as the 5T4 oncofoetal glycoprotein may be limited by the nature of the T cell repertoire and the influence of immunomodulatory factors in particular T regulatory cells (Treg). Here, we identified mouse 5T4-specific T cell epitopes using a 5T4 knock out (5T4KO) mouse and evaluated corresponding wild-type (WT) responses as a model to refine and improve immunogenicity. We have shown that 5T4KO mice vaccinated by replication defective adenovirus encoding mouse 5T4 (Adm5T4) generate potent 5T4-specific IFN-γ CD8 and CD4 T cell responses which mediate significant protection against 5T4 positive tumour challenge. 5T4KO CD8 but not CD4 primed T cells also produced IL-17. By contrast, Adm5T4-immunized WT mice showed no tumour protection consistent with only low avidity CD8 IFN-γ, no IL-17 T cell responses and no detectable CD4 T cell effectors producing IFN-γ or IL-17. Treatment with anti-folate receptor 4 (FR4) antibody significantly reduced the frequency of Tregs in WT mice and enhanced 5T4-specific IFN-γ but reduced IL-10 T cell responses but did not reveal IL-17-producing effectors. This altered balance of effectors by treatment with FR4 antibody after Adm5T4 vaccination provided modest protection against autologous B16m5T4 melanoma challenge. The efficacy of 5T4 and some other TAA vaccines may be limited by the combination of TAA-specific T regs, the deletion and/or alternative differentiation of CD4 T cells as well as the absence of distinct subsets of CD8 T cells.

IL-33 exacerbates autoantibody-induced arthritis.

Rheumatoid arthritis pathogenesis comprises dysregulation in both innate and adaptive immunity. There is therefore intense interest in the factors that integrate these immunologic pathways in rheumatoid arthritis. In this paper, we report that IL-33, a novel member of the IL-1 family, can exacerbate anti-glucose-6-phosphate isomerase autoantibody-induced arthritis (AIA). Mice lacking ST2 (ST2(-/-)), the IL-33 receptor alpha-chain, developed attenuated AIA and reduced expression of articular proinflammatory cytokines. Conversely, treatment of wild-type mice with rIL-33 significantly exacerbated AIA and markedly enhanced proinflammatory cytokine production. However, IL-33 failed to increase the severity of the disease in mast cell-deficient or ST2(-/-) mice. Furthermore, mast cells from wild-type, but not ST2(-/-), mice restored the ability of ST2(-/-) recipients to mount an IL-33-mediated exacerbation of AIA. IL-33 also enhanced autoantibody-mediated mast cell degranulation in vitro and in synovial tissue in vivo. Together these results demonstrate that IL-33 can enhance autoantibody-mediated articular inflammation via promoting mast cell degranulation and proinflammatory cytokine production. Because IL-33 is derived predominantly from synovial fibroblasts, this finding provides a novel mechanism whereby a host tissue-derived cytokine can regulate effector adaptive immune response via enhancing innate cellular activation in inflammatory arthritis.

Protease-activated receptor 2 activation induces behavioural changes associated with depression-like behaviour through microglial-independent modulation of inflammatory cytokines.

RATIONALE: Major depressive disorder (MDD) is a leading cause of disability worldwide but currently prescribed treatments do not adequately ameliorate the disorder in a significant portion of patients. Hence, a better appreciation of its aetiology may lead to the development of novel therapies. OBJECTIVES: In the present study, we have built on our previous findings indicating a role for protease-activated receptor-2 (PAR2) in sickness behaviour to determine whether the PAR2 activator, AC264613, induces behavioural changes similar to those observed in depression-like behaviour. METHODS: AC264613-induced behavioural changes were examined using the open field test (OFT), sucrose preference test (SPT), elevated plus maze (EPM), and novel object recognition test (NOR). Whole-cell patch clamping was used to investigate the effects of PAR2 activation in the lateral habenula with peripheral and central cytokine levels determined using ELISA and quantitative PCR. RESULTS: Using a blood-brain barrier (BBB) permeable PAR2 activator, we reveal that AC-264613 (AC) injection leads to reduced locomotor activity and sucrose preference in mice but is without effect in anxiety and memory-related tasks. In addition, we show that AC injection leads to elevated blood sera IL-6 levels and altered cytokine mRNA expression within the brain. However, neither microglia nor peripheral lymphocytes are the source of these altered cytokine profiles. CONCLUSIONS: These data reveal that PAR2 activation results in behavioural changes often associated with depression-like behaviour and an inflammatory profile that resembles that seen in patients with MDD and therefore PAR2 may be a target for novel antidepressant therapies.

Galectin-3 deficiency reduces the severity of experimental autoimmune encephalomyelitis.

Galectin-3 (Gal-3) is a member of the beta-galactoside-binding lectin family and plays an important role in inflammation. However, the precise role of Gal-3 in autoimmune diseases remains obscure. We have investigated the functional role of Gal-3 in experimental autoimmune encephalomyelitis (EAE) following immunization with myelin oligodendrocyte glycoprotein (MOG)35-55 peptide. Gal-3 deficient (Gal-3-/-) mice developed significantly milder EAE and markedly reduced leukocyte infiltration in the CNS compared with similarly treated wild-type (WT) mice. Gal-3-/- mice also contained fewer monocytes and macrophages but more apoptotic cells in the CNS than did WT mice. Following Ag stimulation in vitro, lymph node cells from the immunized Gal-3-/- mice produced less IL-17 and IFN-gamma than did those of the WT mice. In contrast, Gal-3-/- mice produced more serum IL-10, IL-5, and IL-13 and contained higher frequency of Foxp3+ regulatory T cells in the CNS than did the WT mice. Furthermore, bone marrow-derived dendritic cells from Gal-3-/- mice produced more IL-10 in response to LPS or bacterial lipoprotein than did WT marrow-derived dendritic cells. Moreover, Gal-3-/- dendritic cells induced Ag-specific T cells to produce more IL-10, IL-5, and IL-12, but less IL-17, than did WT dendritic cells. Taken together, our data demonstrate that Gal-3 plays an important disease-exacerbating role in EAE through its multifunctional roles in preventing cell apoptosis and increasing IL-17 and IFN-gamma synthesis, but decreasing IL-10 production.

IL-33 exacerbates antigen-induced arthritis by activating mast cells.

IL-33, a cytokine of the IL-1 family, is closely associated with type II T cell responses. Here, we report an unexpected proinflammatory role of IL-33 in inflammatory arthritis. IL-33 was expressed in synovial fibroblasts from patients with rheumatoid arthritis (RA). Expression was markedly elevated in vitro by inflammatory cytokines. Mice lacking ST2, the IL-33 receptor alpha-chain, developed attenuated collagen-induced arthritis (CIA) and reduced ex vivo collagen-specific induction of proinflammatory cytokines (IL-17, TNFalpha, and IFNgamma), and antibody production. Conversely, treatment of wild-type (WT) but not ST2(-/-) mice with IL-33 exacerbated CIA and elevated production of both proinflammatory cytokines and anti-collagen antibodies. Mast cells expressed high levels of ST2 and responded directly to IL-33 to produce a spectrum of inflammatory cytokines and chemokines in vitro. In vivo, IL-33 treatment exacerbated CIA in ST2(-/-) mice engrafted with mast cells from WT but not from ST2(-/-) mice. Disease exacerbation was accompanied by elevated expression levels of proinflammatory cytokines. Our results demonstrate that IL-33 is a critical proinflammatory cytokine for inflammatory joint disease that integrates fibroblast activation with downstream immune activation mainly via an IL-33-driven, mast-cell-dependent pathway. Thus, this IL-1 superfamily member represents a therapeutic target for RA.

Increased Levels of IL-16 in the Central Nervous System during Neuroinflammation Are Associated with Infiltrating Immune Cells and Resident Glial Cells.

Interleukin (IL)-16, a CD4+ immune cell specific chemoattractant cytokine, has been shown to be involved in the development of multiple sclerosis, an inflammatory demyelinating disease of the central nervous system (CNS). While immune cells such as T cells and macrophages are reported to be the producers of IL-16, the cellular source of IL-16 in the CNS is less clear. This study investigates the correlation of IL-16 expression levels in the CNS with the severity of neuroinflammation and determines the phenotype of cells which produce IL-16 in the CNS of experimental autoimmune encephalomyelitis (EAE) mice. Our data show that IL-16 expression is significantly increased in the brain and spinal cord tissues of EAE mice compared to phosphate buffered saline (PBS) immunised controls. Dual immunofluorescence staining reveals that the significantly increased IL-16+ cells in the CNS lesions of EAE mice are likely to be the CD45+ infiltrating immune cells such as CD4+ or F4/80+ cells and the CNS resident CD11b+ microglia and GFAP+ astrocytes, but not NeuN+ neurons. Our data suggest cytokine IL-16 is closely involved in EAE pathology as evidenced by its increased expression in the glial and infiltrating immune cells, which impacts the recruitment and activation of CD4+ immune cells in the neuroinflammation.

Quantification of the influence of drugs on zebrafish larvae swimming kinematics and energetics.

The use of zebrafish larvae has aroused wide interest in the medical field for its potential role in the development of new therapies. The larvae grow extremely quickly and the embryos are nearly transparent which allows easy examination of its internal structures using fluorescent imaging techniques. Medical treatment of zebrafish larvae can directly influence its swimming behaviours. These behaviour changes are related to functional changes of central nervous system and transformations of the zebrafish body such as muscle mechanical power and force variation, which cannot be measured directly by pure experiment observation. To quantify the influence of drugs on zebrafish larvae swimming behaviours and energetics, we have developed a novel methodology to exploit intravital changes based on observed zebrafish locomotion. Specifically, by using an in-house MATLAB code to process the recorded live zebrafish swimming video, the kinematic locomotion equation of a 3D zebrafish larvae was obtained, and a customised Computational Fluid Dynamics tool was used to solve the fluid flow around the fish model which was geometrically the same as experimentally tested zebrafish. The developed methodology was firstly verified against experiment, and further applied to quantify the fish internal body force, torque and power consumption associated with a group of normal zebrafish larvae vs. those immersed in acetic acid and two neuroactive drugs. As indicated by our results, zebrafish larvae immersed in 0.01% acetic acid display approximately 30% higher hydrodynamic power and 10% higher cost of transport than control group. In addition, 500 µM diphenylhydantoin significantly decreases the locomotion activity for approximately 50% lower hydrodynamic power, whereas 100 mg/L yohimbine has not caused any significant influences on 5 dpf zebrafish larvae locomotion. The approach has potential to evaluate the influence of drugs on the aquatic animal's behaviour changes and thus support the development of new analgesic and neuroactive drugs.

Expression and Function of IL-33/ST2 Axis in the Central Nervous System Under Normal and Diseased Conditions.

Interleukin-33 (IL-33) is a well-recognized immunomodulatory cytokine which plays critical roles in tissue function and immune-mediated diseases. The abundant expression of IL-33 in brain and spinal cord prompted many scientists to explore its unique role in the central nervous system (CNS) under physiological and pathological conditions. Indeed emerging evidence from over a decade's research suggests that IL-33 acts as one of the key molecular signaling cues coordinating the network between the immune and CNS systems, particularly during the development of neurological diseases. Here, we highlight the recent advances in our knowledge regarding the distribution and cellular localization of IL-33 and its receptor ST2 in specific CNS regions, and more importantly the key roles IL-33/ST2 signaling pathway play in CNS function under normal and diseased conditions.

The therapeutic effect of anti-CD52 treatment in murine experimental autoimmune encephalomyelitis is associated with altered IL-33 and ST2 expression levels.

Experimental autoimmune encephalomyelitis (EAE) mice were administered with murine anti-CD52 antibody to investigate its therapeutic effect and whether the treatment modulates IL-33 and ST2 expression. EAE severity and central nervous system (CNS) inflammation were reduced following the treatment, which was accompanied by peripheral T and B lymphocyte depletion and reduced production of various cytokines including IL-33, while sST2 was increased. In spinal cords of EAE mice, while the number of IL-33+ cells remained unchanged, the extracellular level of IL-33 protein was significantly reduced in anti-CD52 antibody treated mice compared with controls. Furthermore the number of ST2+ cells in the spinal cord of treated EAE mice was downregulated due to decreased inflammation and immune cell infiltration in the CNS. These results suggest that treatment with anti-CD52 antibody differentially alters expression of IL-33 and ST2, both systemically and within the CNS, which may indicate IL-33/ST2 axis is involved in the action of the antibody in inhibiting EAE.

Effect of sphingosine kinase modulators on interleukin-1ß release, sphingosine 1-phosphate receptor 1 expression and experimental autoimmune encephalomyelitis.

BACKGROUND AND PURPOSE: The sphingosine analogue, FTY720 (GilenyaR ), alleviates clinical disease progression in multiple sclerosis. Here, we variously assessed the effects of an azide analogue of (S)-FTY720 vinylphosphonate (compound 5; a sphingosine kinase 1 activator), (R)-FTY720 methyl ether (ROMe, a sphingosine kinase 2 inhibitor) and RB-020 (a sphingosine kinase 1 inhibitor and sphingosine kinase 2 substrate) on IL-1ß formation, sphingosine 1-phosphate levels and expression of S1P1 receptors. We also assessed the effect of compound 5 and ROMe in an experimental autoimmune encephalomyelitis (EAE) model in mice. EXPERIMENTAL APPROACH: We measured IL-1ß formation by macrophages, sphingosine 1-phosphate levels and expression levels of S1P1 receptors in vitro and clinical score in mice with EAE and the extent of inflammatory cell infiltration into the spinal cord in vivo. KEY RESULTS: Treatment of differentiated U937 macrophages with compound 5, RB-020 or sphingosine (but not ROMe) enhanced IL-1ß release. These data suggest that these compounds might be pro-inflammatory in vitro. However, compound 5 or ROMe reduced disease progression and infiltration of inflammatory cells into the spinal cord in EAE, and ROMe induced a reduction in CD4+ and CD8+ T-cell levels in the blood (lymphopenia). Indeed, ROMe induced a marked decrease in expression of cell surface S1P1 receptors in vitro. CONCLUSION AND IMPLICATIONS: This is the first demonstration that an activator of sphingosine kinase 1 (compound 5) and an inhibitor of sphingosine kinase 2 (ROMe, which also reduces cell surface S1P1 receptor expression) have an anti-inflammatory action in EAE.

A review on continuous-flow microfluidic PCR in droplets: Advances, challenges and future.

Significant advances have been made in developing microfluidic polymerase chain reaction (PCR) devices in the last two decades. More recently, microfluidic microdroplet technology has been exploited to perform PCR in droplets because of its unique features. For example, it can prevent crossover contamination and PCR inhibition, is suitable for single-cell and single-molecule analyses, and has the potential for system integration and automation. This review will therefore focus on recent developments on droplet-based continuous-flow microfluidic PCR, and the major research challenges. This paper will also discuss a new way of on-chip flow control and a rational design simulation tool, which are required to underpin fully integrated and automated droplet-based microfluidic systems. We will conclude with a scientific speculation of future autonomous scientific discoveries enabled by microfluidic microdroplet technologies.

MAP kinase phosphatase 2 deficient mice develop attenuated experimental autoimmune encephalomyelitis through regulating dendritic cells and T cells.

Mitogen-activated protein kinase phosphatases (MKPs) play key roles in inflammation and immune mediated diseases. Here we investigated the mechanisms by which MKP-2 modulates central nervous system (CNS) inflammation in experimental autoimmune encephalomyelitis (EAE). Our results show that MKP-2 mRNA levels in the spinal cord and lymphoid organs of EAE mice were increased compared with naive controls, indicating an important role for MKP-2 in EAE development. Indeed, MKP-2-/- mice developed reduced EAE severity, associated with diminished CNS immune cell infiltration, decreased proinflammatory cytokine production and reduced frequency of CD4+ and CD8+ T cells in spleens and lymph nodes. In addition, MKP-2-/- CD11c+ dendritic cells (DCs) had reduced expression of MHC-II and CD40 compared with MKP-2+/+ mice. Subsequent experiments revealed that CD4+ T cells from naïve MKP-2-/- mice had decreased cell proliferation and IL-2 and IL-17 production relative to wild type controls. Furthermore, co-culture experiments showed that bone marrow derived DCs of MKP-2-/- mice had impaired capability in antigen presentation and T cell activation. While MKP-2 also modulates macrophage activation, our study suggests that MKP-2 is essential to the pathogenic response of EAE, and it acts mainly via regulating the important antigen presenting DC function and T cell activation.

Role of IL-33 and ST2 signalling pathway in multiple sclerosis: expression by oligodendrocytes and inhibition of myelination in central nervous system.

Recent research findings have provided convincing evidence indicating a role for Interleukin-33 (IL-33) signalling pathway in a number of central nervous system (CNS) diseases including multiple sclerosis (MS) and Alzheimer's disease. However, the exact function of IL-33 molecule within the CNS under normal and pathological conditions is currently unknown. In this study, we have mapped cellular expression of IL-33 and its receptor ST2 by immunohistochemistry in the brain tissues of MS patients and appropriate controls; and investigated the functional significance of these findings in vitro using a myelinating culture system. Our results demonstrate that IL-33 is expressed by neurons, astrocytes and microglia as well as oligodendrocytes, while ST2 is expressed in the lesions by oligodendrocytes and within and around axons. Furthermore, the expression levels and patterns of IL-33 and ST2 in the lesions of acute and chronic MS patient brain samples are enhanced compared with the healthy brain tissues. Finally, our data using rat myelinating co-cultures suggest that IL-33 may play an important role in MS development by inhibiting CNS myelination.