Experimental Autoimmune Encephalomyelitis Disease Severity Research Articles

Gut environment and multiple sclerosis

Multiple sclerosis (MS) is an inflammatory demyelinating disease of the central nervous system (CNS) and T cell-mediated autoimmune processes are assumed to be involved in its pathogenesis. Recently, accumulating evidence has indicated that commensal bacteria interact with the host immune system and that the alteration of commensal bacteria composition, termed dysbiosis, is associated with various autoimmune diseases including CNS autoimmune diseases. The effect of gut microbiota on disease has been initially shown in experimental autoimmune encephalomyelitis (EAE), animal model of MS. Recent analysis of microbiota revealed dysbiosis in patients with MS including the reduction of short chain fatty acid (SCFA). Administration of SCFA ameliorates disease severity of EAE in association with the induction of regulatory T cells. Moreover metabolites of microbiota such as SCFA and tryptophan have been shown to influence glial functions in CNS. In this review, we introduce recent findings regarding the interaction between gut microbiota and MS both in EAE and MS.

Exposure to a mixture of 23 chemicals associated with unconventional oil and gas operations alters immune response to challenge in adult mice

The prevalence of unconventional oil and gas (UOG) operations raises concerns regarding the potential for adverse health outcomes following exposure to water tainted by mixtures of UOG associated chemicals. The potential effects that exposure to complex chemical mixtures has on the immune system have yet to be fully evaluated. In this study, effects on the immune system of adult mice exposed to a mixture of 23 chemicals that have been associated with water near active UOG operations were investigated. Female and male mice were exposed to the mixture via their drinking water for at least 8 weeks. At the end of the exposure, cellularity of primary and secondary immune organs, as well as an immune system function, were assessed using three different models of disease, i.e. house dust mite (HDM)-induced allergic airway disease, influenza A virus infection, and experimental autoimmune encephalomyelitis (EAE). The results indicated exposures resulted in different impacts on T-cell populations in each disease model. Furthermore, the consequences of exposure differed between female and male mice. Notably, exposure to the chemical mixture significantly increased EAE disease severity in females, but not in male, mice. These findings indicated that direct exposure to this mixture leads to multiple alterations in T-cell subsets and that these alterations differ between sexes. This suggested to us that direct exposure to UOG-associated chemicals may alter the adult immune system, leading to dysregulation in immune cellularity and function.

Sex differences in EAE reveal common and distinct cellular and molecular components

Experimental autoimmune encephalomyelitis (EAE) is commonly used as an animal model for evaluating clinical, histological and immunological processes potentially relevant to the human disease multiple sclerosis (MS), for which the mode of disease induction remains largely unknown. An important caveat for interpreting EAE processes in mice is the inflammatory effect of immunization with myelin peptides emulsified in Complete Freund’s Adjuvant (CFA), often followed by additional injections of pertussis toxin (Ptx) in some strains to induce EAE. The current study evaluated clinical, histological, cellular (spleen), and chemokine-driven processes in spinal cords of male vs. female C57BL/6 mice that were immunized with mouse (m)MOG-35-55/CFA/Ptx to induce EAE; immunized with saline/CFA/Ptx only (CFA, no EAE); or were untreated (Naïve, no EAE). Analysis of response curves utilized a rigorous and sophisticated methodology to parse and characterize the effects of EAE and adjuvant alone vs. the Naive baseline responses. The results demonstrated stronger pro-inflammatory responses of immune cells and their associated cytokines, chemokines, and receptors in male vs. female CFA and EAE mice that appeared to be offset partially by increased percentages of male anti-inflammatory, regulatory and checkpoint T cell, B cell, and monocyte/macrophage subsets. These sex differences in peripheral immune responses may explain the reduced cellular infiltration and differing chemokine profiles in the Central Nervous System (CNS) of male vs. female CFA immunized mice and the reduced CNS infiltration and demyelination observed in male vs. female EAE groups of mice that ultimately resulted in the same clinical EAE disease severity in both sexes. Our findings suggest EAE disease severity is governed not only by the degree of CNS infiltration and demyelination, but also by the balance of pro-inflammatory vs. regulatory cell types and their secreted cytokines and chemokines.

NF-κB Activation Accounts for the Cytoprotective Effects of PERK Activation on Oligodendrocytes during EAE

Previous studies demonstrate that activation of pancreatic ER kinase (PERK) protects oligodendrocytes against inflammation in the experimental autoimmune encephalomyelitis (EAE) model of multiple sclerosis (MS). Interestingly, data indicate that the cytoprotective effects of PERK activation on oligodendrocytes during EAE are not mediated by activating transcription factor 4 (ATF4) but are accompanied by activation of nuclear factor κB (NF-κB). NF-κB plays a critical role in MS and EAE; however, the effects of NF-κB activation on oligodendrocytes in these diseases remain elusive. Herein, we generated a mouse model that allow for activation of NF-κB specifically in oligodendrocytes and found that enhanced NF-κB activation in oligodendrocytes had a minimal effect on their viability and function under normal conditions (both male and female mice). Interestingly, we found that enhanced NF-κB activation in oligodendrocytes attenuated EAE disease severity and ameliorated EAE-induced oligodendrocyte loss, demyelination, and axon degeneration, without affecting inflammation (female mice). Moreover, we showed that the detrimental effects of PERK inactivation in oligodendrocytes in EAE were accompanied by impaired NF-κB activation in oligodendrocytes, and were completely rescued by enhanced NF-κB activation in oligodendrocytes (female mice). These findings suggest that NF-κB activation accounts for the cytoprotective effects of PERK activation on oligodendrocytes in MS and EAE.SIGNIFICANCE STATEMENT Nuclear factor κB (NF-κB) is activated in oligodendrocytes in multiple sclerosis (MS) and its animal model experimental autoimmune encephalomyelitis (EAE); however, the role of NF-κB activation in oligodendrocytes in MS and EAE remains elusive. Herein, we generated a mouse model that allows for activation of NF-κB selectively in oligodendrocytes and demonstrated that NF-κB activation prevented oligodendrocyte death and myelin damage in the EAE model. We further demonstrated that NF-κB activation contributed to the protective effects of pancreatic ER kinase (PERK) activation on oligodendrocytes in the EAE model. As such, this work will facilitate the development of new treatments that enhance oligodendrocyte survival in MS patients by targeting the PERK-NF-κB pathway.

Deficiency in B Cell Maturation Antigen reveals sex differences in Experimental Autoimmune Encephalomyelitis

Abstract Multiple sclerosis (MS) is immune mediated demyelinating disease of the central nervous system (CNS) leading to neuronal damage. The disease is heterogeneous with respect to onset and clinical course, being more prevalent in women then in men with a ratio of 3:1. Further sex differences have been observed with males having a more severe and debilitating disease course compared to females. The complex interplay of immune cells, sex hormones or genetic differences could be contributing to these clinical phenomena, which remains to be elucidated. In experimental autoimmune encephalomyelitis (EAE), a mouse model of MS, several reports have demonstrated no sex differences in disease onset or severity in C57BL/6 mice. Herein we report that deficiency in B Cell Maturation Antigen (BCMA) in C57BL/6 mice reveals a sex difference in EAE. BCMA plays a prominent role in B cell function, however the role BCMA plays in MS and EAE is unknown. Our initial goal of this project was to determine the impact BCMA deficiency has on EAE. In doing so we confirmed that EAE disease was not different in males and females from BCMA+/+mice. Strikingly in the BCMA−/− mice, there was a significant increase in EAE disease severity in males compared to females. The increased EAE in the BCMA−/− male mice was associated with increased demyelination and infiltration of inflammatory T cells and macrophages into CNS. Also, we observed decreased splenic transitional B cells in male BCMA−/− mice compared to females. Castration of mice significantly reduces EAE in BCMA−/− mice as compared to sham controls. Altogether, our data demonstrate that male hormones exacerbates EAE in BCMA−/− mice by augmenting inflammatory responses compared to females leading to increased neuro-inflammation.

Dimethyl Itaconate, an Itaconate Derivative, Exhibits Immunomodulatory Effects on Neuroinflammation in Experimental Autoimmune Encephalomyelitis

Abstract Itaconate has recently emerged as a regulator of immune cell function. Studies show that itaconate is required for the activation of anti-inflammatory transcription factor Nrf2 by LPS in macrophages. Moreover, dimethyl itaconate (DMI), an itaconate derivative, has been shown to inhibit IL-17-induced IκBζ activation in keratinocytes and modulate IL-17-IκBζ pathway-mediated skin inflammation in an animal model of psoriasis. Currently, the effect of itaconate on regulating macrophage functions and peripheral inflammatory immune responses is well established. However, its effect on microglia (MG) and CNS inflammatory immune responses remains unexplored. Thus, in this study we investigated whether itaconate possesses an immunomodulatory effect on regulating MG activation and CNS inflammation in experimental autoimmune encephalomyelitis (EAE). Chronic C57BL/6 EAE and relapsing-remitting SJL/J EAE were induced to assess the therapeutic effect of DMI. Our results show DMI ameliorated disease severity in the chronic C57BL/6 EAE model. Further analysis of the cellular and molecular mechanisms revealed that DMI mitigated blood-brain barrier disruption, inhibited MMP3 and MMP9 production, suppressed microglia activation, and repressed CNS infiltration of Th1 and Th17 cells. Strikingly, DMI also exhibited a therapeutic effect on lessening severity of relapse in the relapsing-remitting SJL/J EAE model. In conclusion, we demonstrate for the first time that DMI suppressed neuroinflammation and ameliorated disease severity in EAE through multiple cellular and molecular mechanisms, and our findings suggest that DMI can be developed as a novel therapeutic agent for the treatment of EAE through its anti-inflammatory properties.

Organic compound farnesol as possible inhibitor of inflammasome complex, in murine macrophages and mouse model of multiple sclerosis

Abstract Many advancements in the understanding of multiple sclerosis (MS) have been made through the use of laboratory models. One commonly used model is experimental autoimmune encephalomyelitis (EAE), a mouse model characterized by central nervous system (CNS) inflammation and demyelination, allowing for symptoms resembling some of the most prominent features of the human disease. Although the exact etiology of MS is still being investigated, experiments with EAE have shown that the NLRP3 inflammasome complex of the innate immune system is critical and necessary for disease development. The inflammasome complex can be assembled in all innate immune cells, including microglia and astrocytes in the CNS. Dysregulation of inflammasome activity can result in uncontrolled inflammation, which underlies many chronic diseases, and metabolic and autoimmune disorders such as MS. Our lab has shown that farnesol a 15-carbon organic sesquiterpene and primary alcohol, reduced EAE disease severity and onset and also decreased T-cell infiltration into the CNS. However, the mechanisms of its action have yet to be fully defined. Therefore, in-vitro work on murine macrophages is being conducted to investigate how farnesol may be potentially affecting the pathway of the inflammasome complex and providing this protection. Furthermore, since farnesol is a quorum-sensing molecule that impacts biofilm formation, other studies of ours are aimed at evaluating how farnesol affects the gut-brain axis and specifically the gut microbiome of EAE mice.

Dimethyl itaconate, an itaconate derivative, exhibits immunomodulatory effects on neuroinflammation in experimental autoimmune encephalomyelitis

BackgroundInflammatory stimuli induce immunoresponsive gene 1 (IRG1) expression that in turn catalyzes the production of itaconate from the tricarboxylic acid cycle. Itaconate has recently emerged as a regulator of immune cell functions, especially in macrophages. Studies show that itaconate is required for the activation of anti-inflammatory transcription factor Nrf2 by LPS in mouse and human macrophages, and LPS-activated IRG1-/- macrophages that lack endogenous itaconate production exhibit augmented inflammatory responses. Moreover, dimethyl itaconate (DMI), an itaconate derivative, inhibits IL-17-induced IκBς activation in keratinocytes and modulates IL-17-IκBς pathway-mediated skin inflammation in an animal model of psoriasis. Currently, the effect of itaconate on regulating macrophage functions and peripheral inflammatory immune responses is well established. However, its effect on microglia (MG) and CNS inflammatory immune responses remains unexplored. Thus, we investigated whether itaconate possesses an immunomodulatory effect on regulating MG activation and CNS inflammation in animal models of multiple sclerosis, experimental autoimmune encephalomyelitis (EAE).MethodsChronic C57BL/6 EAE was induced followed by DMI treatment. The effect of DMI on disease severity, blood-brain barrier (BBB) disruption, MG activation, peripheral Th1/Th17 differentiation, and the CNS infiltration of Th1/Th17 cells in EAE was determined. Primary MG was cultured to study the effect of DMI on MG activation. Relapsing-remitting SJL/J EAE was induced to assess the therapeutic effect of DMI.ResultsOur results show DMI ameliorated disease severity in the chronic C57BL/6 EAE model. Further analysis of the cellular and molecular mechanisms revealed that DMI mitigated BBB disruption, inhibited MMP3/MMP9 production, suppressed microglia activation, inhibited peripheral Th1/Th17 differentiation, and repressed the CNS infiltration of Th1 and Th17 cells. Strikingly, DMI also exhibited a therapeutic effect on alleviating severity of relapse in the relapsing-remitting SJL/J EAE model.ConclusionsWe demonstrate that DMI suppresses neuroinflammation and ameliorates disease severity in EAE through multiple cellular and molecular mechanisms, suggesting that DMI can be developed as a novel therapeutic agent for the treatment of MS/EAE through its immunomodulatory and anti-inflammatory properties.

NLRP3 inflammasome as prognostic factor and therapeutic target in primary progressive multiple sclerosis patients.

Primary progressive multiple sclerosis is a poorly understood disease entity with no specific prognostic biomarkers and scarce therapeutic options. We aimed to identify disease activity biomarkers in multiple sclerosis by performing an RNA sequencing approach in peripheral blood mononuclear cells from a discovery cohort of 44 untreated patients with multiple sclerosis belonging to different clinical forms and activity phases of the disease, and 12 healthy control subjects. A validation cohort of 58 patients with multiple sclerosis and 26 healthy control subjects was included in the study to replicate the RNA sequencing findings. The RNA sequencing revealed an interleukin 1 beta (IL1B) signature in patients with primary progressive multiple sclerosis. Subsequent immunophenotyping pointed to blood monocytes as responsible for the IL1B signature observed in this group of patients. Functional experiments at baseline measuring apoptosis-associated speck-like protein containing a CARD (ASC) speck formation showed that the NOD-leucine rich repeat and pyrin containing protein 3 (NLRP3) inflammasome was overactive in monocytes from patients with primary progressive multiple sclerosis, and canonical NLRP3 inflammasome activation with a combination of ATP plus lipopolysaccharide was associated with increased IL1B production in this group of patients. Primary progressive multiple sclerosis patients with high IL1B gene expression levels in peripheral blood mononuclear cells progressed significantly faster compared to patients with low IL1B levels based on the time to reach an EDSS of 6.0 and the Multiple Sclerosis Severity Score. In agreement with peripheral blood findings, both NLRP3 and IL1B expression in brain tissue from patients with primary progressive multiple sclerosis was mainly restricted to cells of myeloid lineage. Treatment of mice with a specific NLRP3 inflammasome inhibitor attenuated established experimental autoimmune encephalomyelitis disease severity and improved CNS histopathology. NLRP3 inflammasome-specific inhibition was also effective in reducing axonal damage in a model of lipopolysaccharide-neuroinflammation using organotypic cerebellar cultures. Altogether, these results point to a role of IL1B and the NLRP3 inflammasome as prognostic biomarker and potential therapeutic target, respectively, in patients with primary progressive multiple sclerosis.

The proportion of myeloid-derived suppressor cells in the spleen is related to the severity of the clinical course and tissue damage extent in a murine model of multiple sclerosis

Multiple Sclerosis (MS) is the second cause of paraplegia among young adults, after all types of CNS traumatic lesions. In its most frequent relapsing-remitting form, the severity of the disease course is very heterogeneous, and its reliable evaluation remains a key issue for clinicians. Myeloid-Derived sSuppressor Cells (MDSCs) are immature myeloid cells that suppress the inflammatory response, a phenomenon related to the resolution or recovery of the clinical symptoms associated with experimental autoimmune encephalomyelitis (EAE), the most common model for MS. Here, we establish the severity index as a new parameter for the clinical assessment in EAE. It is derived from the relationship between the maximal clinical score and the time elapsed since disease onset. Moreover, we relate this new index with several histopathological hallmarks in EAE and with the peripheral content of MDSCs. Based on this new parameter, we show that the splenic MDSC content is related to the evolution of the clinical course of EAE, ranging from mild to severe. Indeed, when the severity index indicates a severe disease course, EAE mice display more intense lymphocyte infiltration, demyelination and axonal damage. A direct correlation was drawn between the MDSC population in the peripheral immune system, and the preservation of myelin and axons, which was also correlated with T cell apoptosis within the CNS (being these cells the main target for MDSC suppression). The data presented clearly indicated that the severity index is a suitable tool to analyze disease severity in EAE. Moreover, our data suggest a clear relationship between circulating MDSC enrichment and disease outcome, opening new perspectives for the future targeting of this population as an indicator of MS severity.

MicroRNAs as disease progression biomarkers and therapeutic targets in experimental autoimmune encephalomyelitis model of multiple sclerosis.

Multiple sclerosis is an autoimmune neurodegenerative disease of the central nervous system characterized by pronounced inflammatory infiltrates entering the brain, spinal cord and optic nerve leading to demyelination. Focal demyelination is associated with relapsing-remitting multiple sclerosis, while progressive forms of the disease show axonal degeneration and neuronal loss. The tests currently used in the clinical diagnosis and management of multiple sclerosis have limitations due to specificity and sensitivity. MicroRNAs (miRNAs) are dysregulated in many diseases and disorders including demyelinating and neuroinflammatory diseases. A review of recent studies with the experimental autoimmune encephalomyelitis animal model (mostly female mice 6–12 weeks of age) has confirmed miRNAs as biomarkers of experimental autoimmune encephalomyelitis disease and importantly at the pre-onset (asymptomatic) stage when assessed in blood plasma and urine exosomes, and spinal cord tissue. The expression of certain miRNAs was also dysregulated at the onset and peak of disease in blood plasma and urine exosomes, brain and spinal cord tissue, and at the post-peak (chronic) stage of experimental autoimmune encephalomyelitis disease in spinal cord tissue. Therapies using miRNA mimics or inhibitors were found to delay the induction and alleviate the severity of experimental autoimmune encephalomyelitis disease. Interestingly, experimental autoimmune encephalomyelitis disease severity was reduced by overexpression of miR-146a, miR-23b, miR-497, miR-26a, and miR-20b, or by suppression of miR-182, miR-181c, miR-223, miR-155, and miR-873. Further studies are warranted on determining more fully miRNA profiles in blood plasma and urine exosomes of experimental autoimmune encephalomyelitis animals since they could serve as biomarkers of asymptomatic multiple sclerosis and disease course. Additionally, studies should be performed with male mice of a similar age, and with aged male and female mice.

CNS delivery of anti-CD52 antibodies modestly reduces disease severity in an animal model for multiple sclerosis

Background and aims:Alemtuzumab is a humanized monoclonal antibody that depletes CD52-bearing B and T lymphocytes. Clinical trials defined that systemic administration of alemtuzumab reduces disease severity in the relapsing–remitting phase of multiple sclerosis (MS). However, its efficacy in progressive MS patients is limited, which may reflect the inability of alemtuzumab to cross the reconstituted BBB in these patients. Objective: to study whether central nervous system (CNS) delivery of anti-CD52 antibodies reduces disease severity and the neuroinflammatory burden in the experimental autoimmune encephalomyelitis (EAE) model.Methods:Anti-CD52 antibodies were administered intrathecally during the acute and chronic phases of EAE. Flow cytometry and immunohistochemistry were utilized to define immunological and pathological parameters.Results:We show that subcutaneously administrated anti-CD52 antibodies completely abolish EAE disease severity. CNS delivery of anti-CD52 antibodies during both the acute and chronic phases of EAE moderately reduces disease severity and the neuroinflammatory burden. Our findings further suggest that CNS delivery of anti-CD52 antibodies impacts both the peripheral and CNS immune cell compartments in the EAE model but not in healthy mice.Conclusion:Collectively, our findings highlight the therapeutic potential of CNS delivery of alemtuzumab for the treatment of progressive as well as early MS.

Scoring disease in an animal model of multiple sclerosis using a novel infrared-based automated activity-monitoring system

Multiple sclerosis (MS) is a chronic demyelinating disorder of the central nervous system (CNS). Its corresponding animal model, experimental autoimmune encephalomyelitis (EAE), is widely used to understand disease pathogenesis and test novel therapeutic agents. However, existing methods to score EAE disease severity are subjective and often vary between individual researchers, making it difficult to translate findings across different studies. An enhanced automated method of disease scoring would eliminate subjectivity and reduce operator-dependent errors. Here, we used an Infra-Red Activity Monitoring System (IRAMS) to measure murine locomotor activity as a surrogate measure of disease severity and compared it to standard EAE scoring methods. In mice immunized with CNS-specific myelin antigens, we observed an inverse correlation between disease severity and mouse activity, with the IRAMS showing enhanced disease scoring compared to standard EAE scoring methods. Relative to standard EAE scoring methods, IRAMS showed comparable measurement of disease relapses and remissions in the SJL/J-relapsing-remitting model of EAE, and could comparably assess the therapeutic efficiency of the MS drug, Copaxone (Glatiramer acetate-GA). Thus, the IRAMS is a method to measure disease severity in EAE without subjective bias and is a tool to consistently assess the efficacy of novel therapeutic agents for MS.

Deficiency in B Cell Maturation Antigen reveals gender differences in Experimental Autoimmune Encephalomyelitis

Abstract Multiple sclerosis (MS) is immune mediated demyelinating disease of the central nervous system (CNS) leading to neuronal damage. The disease is heterogeneous with respect to onset and clinical course, being more prevalent in women than in men with a ratio of 3:1. Further sex differences have been observed with males having a more severe and debilitating disease course compared to female. The complex interplay of immune cells, sex hormones or genetic differences could be contributing to these clinical phenomena, which remains to be elucidated. In experimental autoimmune encephalomyelitis (EAE), a mouse model of MS, several reports have demonstrated no sex differences in disease onset or severity in C57BL/6 mice. Herein we report that deficiency in B Cell Maturation Antigen (BCMA) in C57BL/6 mice reveals a sex difference in EAE. BCMA plays a prominent role in B cell function, however, the role BCMA plays in MS and EAE is unknown. Our initial goal of this project was to determine the impact BCMA deficiency has on EAE. In doing so we confirmed that EAE disease was not different in males and females from BCMA+/+mice. Strikingly in the BCMA−/− mice, there was a significant increase in EAE disease severity in the males compared to females. The increased EAE in the BCMA−/− male mice was associated with increased demyelination and infiltration of inflammatory T cells and macrophages into CNS. Also, we observed decreased splenic transitional B cells in male BCMA−/− mice compared to females. Altogether, our data demonstrate that male BCMA−/− have elevated inflammatory responses compared to females leading to increased neuro-inflammation. Further experiments are underway to define the specific mechanisms behind the sex differences in BCMA deficient mice.

Oligodendrocyte-specific ATF4 inactivation does not influence the development of EAE

BackgroundMultiple sclerosis (MS) and its animal model, experimental autoimmune encephalomyelitis (EAE), are inflammatory demyelinating and neurodegenerative diseases of the CNS. Although recent studies suggest the neuroprotective effects of oligodendrocytes in neurodegenerative diseases, it remains unknown whether oligodendrocyte death induced by inflammatory attacks contributes to neurodegeneration in MS and EAE. Upon endoplasmic reticulum (ER) stress, activation of pancreatic ER kinase (PERK) promotes cell survival through induction of activating transcription factor 4 (ATF4) by phosphorylating eukaryotic translation initiation factor 2α (eIF2α). We have generated a mouse model that allows for temporally controlled activation of PERK specifically in oligodendrocytes. Our previous study has demonstrated that PERK activation specifically in oligodendrocytes attenuates EAE disease severity and ameliorates EAE-induced oligodendrocyte apoptosis, demyelination, and axon degeneration, without altering inflammation.MethodsWe determined whether oligodendrocyte-specific PERK activation reduced neuron loss in the CNS of EAE mice using the mouse model that allows for temporally controlled activation of PERK specifically in oligodendrocytes. We further generated a mouse model that allows for inactivation of ATF4 specifically in oligodendrocytes, and determined the effects of ATF4 inactivation in oligodendrocytes on mice undergoing EAE.ResultsWe showed that protection of oligodendrocytes resulting from PERK activation led to attenuation of neuron loss in the CNS gray matter of EAE mice. Surprisingly, we found that ATF4 inactivation specifically in oligodendrocytes did not alter EAE disease severity and had no effect on oligodendrocyte loss, demyelination, axon degeneration, neuron loss, and inflammation in EAE mice.ConclusionsThese findings suggest the neuroprotective effects of PERK activation in oligodendrocytes in EAE, and rule out the involvement of ATF4 in oligodendrocytes in the development of EAE. These results imply that the protective effects of PERK activation in oligodendrocytes in MS and EAE are not mediated by ATF4.

Systemic exposure to a single dose of ferucarbotran aggravates neuroinflammation in a murine model of experimental autoimmune encephalomyelitis.

BackgroundMedicinal preparations of iron oxide nanoparticles (IONPs) have been used as MRI contrast agents for the diagnosis of hepatic tumors and the assessment of neuroinflammation and blood–brain barrier integrity. However, it remains mostly unclear whether exposure to IONPs affects neuroinflammation under disease conditions. The present study aims to investigate the impact of IONPs on autoimmune-mediated neuroinflammation using a murine model of experimental autoimmune encephalomyelitis (EAE) that mimics human multiple sclerosis.MethodsMice were either left untreated or immunized with myelin oligodendrocyte glyco-protein on day 0 followed by two injections of pertussis toxin for EAE induction. The EAE mice were intravenously administered with a single dose of the carboxydextran-coated IONPs, ferucarbotran (20 mg Fe/kg) and/or saline (as vehicle) on day 18. Symptoms of EAE were daily monitored until the mice were killed on day 30. Tissue sections of the brain and spinal cord were prepared for histopathological examinations. Iron deposition, neuron demyelination and inflammatory cell infiltration were examined using histochemical staining. The infiltration of microglial and T cells, and cytokine expression were examined by immunohistochemical staining and/or reverse transcription polymerase chain reaction (RT-PCR).ResultsIron deposition was detected in both the brain and spinal cord of EAE mice 3 days post-ferucarbotran treatment. The clinical and pathological scores of EAE, percentage of myelin loss and infiltration of inflammatory cells into the spinal cord were significantly deteriorated in EAE mice treated with ferucarbotran. Furthermore, ferucarbotran treatment increased the number of CD3+, Iba-1+, IL-6+, Iba-1+TNF-α+ and CD3+IFN-γ+ cells in the spinal cord of EAE mice.ConclusionA single exposure to ferucarbotran exacerbated neuroinflammation and disease severity of EAE, which might be attributed to the enhanced activation of microglia and T cells. These results demonstrated that the pro-inflammatory effect of ferucarbotran on the central nervous system is closely associated with the deterioration of autoimmunity.

Leonurine suppresses neuroinflammation through promoting oligodendrocyte maturation.

Focal inflammation and remyelination failure are major hallmarks of multiple sclerosis and its animal model, experimental autoimmune encephalomyelitis (EAE). In this study, we found that leonurine, a bioactive alkaloid, alleviated EAE disease severity along with reduced central nervous system inflammation and myelin damage. During the pathogenesis of EAE, leonurine dramatically suppressed the recruitment of encephalitogenic T cells into the central nervous system, whereas did not impair periphery immune responses and microglia activation. Mechanistically, leonurine protected mice against demyelination along with enhanced remyelination through promoting the maturation of oligodendrocytes in both EAE and cuprizone‐induced demyelination mouse models. Moreover, we identified that the expression of demethylase jumonji domain‐containing protein D3 was significantly enhanced upon treatment of leonurine, which suppressed the trimethylation of histone H3 lysine‐27 and enhanced oligodendrocyte maturation accordingly. Collectively, our study identified the therapeutic effect of leonurine on EAE model, which potentially represents a promising therapeutic strategy for multiple sclerosis, even other demyelination disorders.

Ambient urban dust particulate matter reduces pathologic T cells in the CNS and severity of EAE

BackgroundAutoimmune diseases have increased in incidence and prevalence worldwide. While genetic predispositions play a role, environmental factors are a major contributor. Atmospheric particulate matter (PM) is a complex mixture composed of metals, nitrates, sulfates and diverse adsorbed organic compounds like polycyclic aromatic hydrocarbons (PAHs) and dioxins. Exposure to atmospheric PM aggravates autoimmune diseases such as type 1 diabetes, rheumatoid arthritis, multiple sclerosis, and systemic lupus erythematosus, among others. PAHs and dioxins are known aryl hydrocarbon receptor (AHR) ligands. The AHR modulates T cell differentiation and directs the balance between effector and regulatory T cells in vitro and in experimental autoimmune encephalomyelitis (EAE), a murine model of autoimmune disease. This study aims to identify pathways that contribute to autoimmune disease and their potential use as therapeutic targets to alleviate symptoms and the need for global immunosuppression. This study tests the hypothesis that atmospheric PM enhances effector T cell differentiation and aggravates autoimmune disease. ResultsAn atmospheric ambient urban dust PM sample, standard reference material (SRM)1649b, was tested for its effects on autoimmunity. SRM1649b PM enhanced Th17 differentiation in an AHR-dependent manner in vitro, however intranasal treatment of SRM1649b PM delayed onset of EAE and reduced cumulative and peak clinical scores. Chronic and acute intranasal exposure of SRM1649b PM delayed onset of EAE. Chronic intranasal exposure did not reduce severity of EAE while acute intranasal exposure significantly reduced severity of disease. Acute intranasal treatment of low dose SRM1649b PM had no effect on clinical score or day of onset in EAE. Delayed onset of EAE by intranasal SRM1649b PM was AHR-dependent in vivo. Oral gavage of SRM1649b PM, in the absence of AHR ligands in the diet, had no effect on day of disease onset or severity of EAE. Day 10 analysis of T cells in the CNS after intranasal treatment of SRM1649b PM showed a reduction of pathologic T cell subsets in vivo. Moreover, MOG-specific splenocytes require AHR to generate or maintain IL-10 producing cells and reduce IFNγ producing cells in vitro. ConclusionsThese results identify the AHR pathway as a potential target for driving targeted immunosuppression in the CNS in the context of atmospheric PM-mediated autoimmune disease. The effects of SRM1649b PM on EAE are dependent on route of exposure, with intranasal treatment reducing severity of EAE and delaying disease onset while oral gavage has no effect. Intranasal SRM1649b PM reduces pathologic T cells in the CNS, specifically Th1 cells and Th1Th17 double positive cells, leading to reduced severity of EAE and AHR-dependent delayed disease onset. Additionally, SRM1649b PM treatment of antigen-specific T cells leads to AHR-dependent increase in percent IL-10 positive cells in vitro. These findings may shed light on the known increase of infection after exposure to atmospheric PM and serve as the first step in identifying components of the AHR pathway responsible for Th1-mediated immunosuppression in response to atmospheric PM exposure.

Silencing miR-150 Ameliorates Experimental Autoimmune Encephalomyelitis.

MiR-150 regulates maturation and differentiation of T cells but how it functions in multiple sclerosis (MS) is unclear. In miR-150 knockout (KO) mice, we examined the effect of miR-150 deletion on disease severity of experimental autoimmune encephalomyelitis (EAE), an animal model of MS. After deleting miR-150, EAE disease severity was reduced according to clinical score. Histological staining and MBP immunofluorescence staining revealed that miR-150 deletion limited the extent of inflammatory demyelination and axonal damage in the spinal cord. Flow cytometry showed that CD3+, CD4+, and CD8+ T cells were increased in WT-EAE mice, but miR-150 deletion significantly reversed EAE-mediated up-regulation of CD3+, CD4+, and CD8+ T cells and down-regulation of CD19+ B cells. In addition, miR-150 deletion reduced the mRNA expression of IL-1β, IL-6, IL-17, and TNF-α in spleen and spinal cord after EAE induction. Thus, miR-150 deletion reduces EAE severity and demyelination, probably through inhibiting the activated immune response and the inflammation in the central nervous system.

Arginine methylation drives inflammatory T cell responses and autoimmunity

Abstract Multiple sclerosis (MS) is a chronic inflammatory disease of the central nervous system that is the leading cause of disability in 2 million young adults aged 20–40 worldwide. Heightened pro-inflammatory T helper (Th) 1 and Th17 cell responses and deficient Th2 and regulatory T (Treg) cell responses drive myelin sheath damage and disability in MS. Thus, therapeutic strategies that modulate inflammatory T cell responses would be beneficial to the field. Epigenetic modifications are well known to provide control over cellular phenotype, and pan-methyltransferase inhibitors have been shown to suppress Experimental Autoimmune Encephalomyelitis (EAE), the mouse model of MS. However, the methylation enzyme responsible for this therapeutic effect remains unknown. Protein arginine methyltransferase 5 (PRMT5) is the only enzyme known to catalyze symmetric dimethylation (SDM) on arginine residues of histones, mediating transcriptional repression. Although PRMT5 is a known cancer driver, until recently, there was no known role for PRMT5 in T cells. We found that PRMT5 protein was up-regulated during re-activation of memory human Th1 cells, and that PRMT5-selective inhibitors suppressed inflammatory T cell proliferation. Further, we developed a conditional PRMT5 knockout mouse model in which PRMT5 knockout can be induced in CD4+ T cells. With this model, we genetically validated our findings that PRMT5 drives CD4+ T cell proliferation. Importantly, we translated our findings in vivo by showing that PRMT5 inhibitors suppressed established EAE disease severity. Overall, these data support a role for PRMT5 in pathogenic memory T cell responses and suggest its promise as a potential therapeutic target for autoimmune diseases such as MS.