Clinical Severity Of Experimental Autoimmune Encephalomyelitis Research Articles

Acetyl 11-Keto Beta-Boswellic Acid Improves Neurological Functions in a Mouse Model of Multiple Sclerosis.

Acetyl-11-keto-β-boswellic acid is one of the main active components of Boswellia sp. resin with the most potent anti-inflammatory activity. In recent years, herbal therapy has received considerable attention for the treatments of inflammatory and demyelinating diseases such as Multiple sclerosis (MS). Studies have shown that herbal compounds could enhance myelin repair and suppress inflammation. This study was designed to investigate the therapeutic effects of intraperitoneal administration of AKBA in Experimental Autoimmune Encephalomyelitis (EAE), as an animal model of MS. Following EAE induction in female C57BL/6J mice, animals were treated with AKBA and the levels of different serum inflammatory mediators, as well as motor functions, myelination, and inflammatory cell infiltration were assessed. Our results revealed that the application of AKBA alleviated EAE clinical severity, and suppressed inflammation, demyelination, leukocyte infiltration, and gliosis in EAE mice. Our findings suggest that the therapeutic effects of AKBA are likely a consequence of its neuroprotective and anti-inflammatory properties. The beneficial effects of AKBA may therefore provide new insights in various neuroinflammatory diseases such as MS and thereby could serve as a potential treatment candidate.

Ozanimod-mediated remission in experimental autoimmune encephalomyelitis is associated with enhanced activity of CNS CD27low/- NK cell subset.

Ozanimod (RPC1063) is an immunomodulator that has been recently approved by the FDA (2020) for the treatment of relapsing-remitting multiple sclerosis (RRMS). It is a selective agonist of the sphingosine-1-phophate receptors 1 and 5, expressed on naïve and central memory T and B cells, as well as natural killer (NK) cells, and is involved in lymphocyte trafficking. Oral administration of ozanimod was reported to result in rapid and reversible reduction in circulating lymphocytes in multiple sclerosis (MS) patients, however, only minimal effect on NK cells was observed. In this study, we sought to investigate the effect of ozanimod on NK cells and assess whether they play any role in ozanimod-induced remission in experimental autoimmune encephalomyelitis (EAE), the animal model of MS. Active EAE induction was done in C57BL/6 female mice, followed by daily oral treatment with ozanimod (0.6mg/kg) starting at disease onset (score 1). Flow cytometry of blood and CNS was performed 24 hours after the last oral dose of ozanimod treatment in diseased mice. Histological analysis of lumbar spinal cord was performed for evaluating the level of inflammation and demyelination. Depletion of peripheral NK cells was done using anti-NK1.1 mouse antibody (mAb) at day 5 post-EAE induction. Ozanimod was effective in reducing the clinical severity of EAE and reducing the percentage of autoreactive CD4+ and CD8+ T cells along with significant inhibition of lymphocyte infiltration into the spinal cord, accompanied by reversed demyelination. Furthermore, ozanimod treatment resulted in a significant increase in the frequency of total NK cells in the blood and CNS along with upregulation of the activating receptor NKG2D on CD27low/- NK cell subset in the CNS. The effectiveness of ozanimod treatment in inhibiting the progression of the disease was reduced when NK cells were depleted using anti-NK1.1 mAb. The current study demonstrated that ozanimod treatment significantly improved clinical symptoms in EAE mice. Ozanimod and anti-NK1.1 mAb appear to function in opposition to one another. Collectively, our data suggest that ozanimod-mediated remission is associated with an increased percentage of total NK cells and CD27low/- NK cells expressing the activating receptor, NKG2D in the CNS.

The effects of melatonin supplementation on neurobehavioral outcomes and clinical severity in rodent models of multiple sclerosis; a systematic review and meta-analysis.

Through the antioxidant and anti-inflammation pathways, melatonin is proposed as a safe and effective intervention in neurological diseases. This study aims to evaluate the effects of melatonin supplementation on the neurobehavioral and clinical outcomes in animal models of multiple sclerosis (MS). This study was conducted following the PRISMA statement. Animal studies that reported the effects of melatonin in preclinical MS models, including the experimental autoimmune encephalomyelitis (EAE) and cuprizone model for demyelination are included in this study. A systematic search in PubMed, Web of Science, Embase, and Scopus up was conducted in April 2023. The collaborative Approach to Meta-Analysis and Review of Animal Experimental Studies (CAMARADES) critical appraisal tool was used for the quality assessment of the studies and the quantitative synthetizes were conducted using the comprehensive meta-analysis software. Out of 542 studies, finally 21 studies, including 14 studies in the EAE model and 7 studies of the toxic demyelination method with cuprizone were included. The route of administration was intraperitoneal in 18 studies, oral in 2 studies, and subcutaneous in 1 study. The quantitative synthesis of the EAE clinical severity scale was associated with significant differences (standardized mean difference [SDM]: - 2.52; - 3.61 to - 1.42; p value < 0.01). In subgroup analyses, the difference was statistically significant in the mouse subgroup (SMD: - 2.60; - 3.74 to - 1.46; p value < 0.01). This study encountered that melatonin may be associated with improved behavioral and cognitive outcomes of preclinical models of MS with acceptable safety profiles. The research was supported by the Student Research Committee, Tabriz University of Medical Sciences (grant number: 71005).

Pharmacological targeting of coagulation factor XI attenuates experimental autoimmune encephalomyelitis in mice.

Multiple sclerosis (MS) is the most common causes of non-traumatic disability in young adults worldwide. MS pathophysiologies include the formation of inflammatory lesions, axonal damage and demyelination, and blood brain barrier (BBB) disruption. Coagulation proteins, including factor (F)XII, can serve as important mediators of the adaptive immune response during neuroinflammation. Indeed, plasma FXII levels are increased during relapse in relapsing-remitting MS patients, and previous studies showed that reducing FXII levels was protective in a murine model of MS, experimental autoimmune encephalomyelitis (EAE). Our objective was to determine if pharmacological targeting of FXI, a major substrate of activated FXII (FXIIa), improves neurological function and attenuates CNS damage in the setting of EAE. EAE was induced in male mice using murine myelin oligodendrocyte glycoprotein peptides combined with heat-inactivated Mycobacterium tuberculosis and pertussis toxin. Upon onset of symptoms, mice were treated every other day intravenously with anti-FXI antibody, 14E11, or saline. Disease scores were recorded daily until euthanasia for ex vivo analyses of inflammation. Compared to the vehicle control, 14E11 treatment reduced the clinical severity of EAE and total mononuclear cells, including CD11b+CD45high macrophage/microglia and CD4+ T cell numbers in brain. Following pharmacological targeting of FXI, BBB disruption was reduced, as measured by decreased axonal damage and fibrin(ogen) accumulation in the spinal cord. These data demonstrate that pharmacological inhibition of FXI reduces disease severity, immune cell migration, axonal damage, and BBB disruption in mice with EAE. Thus, therapeutic agents targeting FXI and FXII may provide a useful approach for treating autoimmune and neurologic disorders.

Anemoside B4 ameliorates experimental autoimmune encephalomyelitis in mice by modulating inflammatory responses and the gut microbiota

BackgroundAnemoside B4 (AB4) is a representative component of Pulsatilla decoction that is used in traditional Chinese medicine for treating inflammatory conditions. It is not known whether AB4 has beneficial effects on multiple sclerosis (MS). MethodsIn the present study, we examined the preventative and therapeutic effects of AB4, and the possible mechanism by which it protects female mice against experimental autoimmune encephalomyelitis (EAE). ResultsPreventative treatment with AB4 (given orally at 100 and 200 mg/kg for 18 days) reduced the clinical severity of EAE significantly (from 3.6 ± 1.3 to 1.8 ± 1.5 and 1.6 ± 0.6, respectively), and inhibited demyelination and inflammatory infiltration of the spinal cord. In the therapeutic protocol, oral administration of 200 mg/kg AB4 for 21 days after initiation of EAE significantly alleviated disease severity (from 2.6 ± 1.3 to 0.9 ± 0.6) and was as effective as the clinically used drug fingolimod (0.3 ± 0.6). Furthermore, both doses of AB4 significantly inhibited mRNA expression of TNF-α, IL-6, and IL-17, and STAT3 activation, in the spinal cord; and the ex vivo and iv vitro AB4 treatment markedly inhibited secretion of the three cytokines from lymphocytes of EAE mice upon in vitro restimulation. In addition, AB4 reversed the changes in the composition of the intestinal microbiome observed in EAE mice. ConclusionWe reveal for the first time that AB4 protects against EAE by modulating inflammatory responses and the gut microbiota, demonstrating that AB4 may have potential as a therapeutic agent for treating MS in humans.

Anlotinib attenuates experimental autoimmune encephalomyelitis mice model of multiple sclerosis via modulating the differentiation of Th17 and Treg cells

Background In multiple sclerosis (MS), the imbalance between T helper (Th)-17 cells and regulatory T (Treg) cells are critical in autoimmune central nervous system (CNS) inflammation and demyelination. Experimental autoimmune encephalomyelitis (EAE) is an established mouse MS model and simulates MS at diverse levels. Objectives This study aims at investigating the impact of anlotinib on the clinical severity of EAE and CD4+ T cell differentiation. Materials and methods EAE-induced mice were treated with water (control) or 6 mg/kg anlotinib by gavage daily. At the peak of EAE, histopathological examination and flow cytometry analysis of CNS-infiltrating CD4+ T cells were performed. In vitro differentiation of CD4+ T cells under different conditions was detected by flow cytometry and quantitative real-time PCR. Finally, the impacts of anlotinib on the phosphorylation of signal transducer and activator of transcription 3 (STAT3) and the transcription levels of key genes involved in Th17 and Treg differentiation were tested. Results Anlotinib attenuated the clinical severity of EAE and changed the frequencies of CNS-infiltrating CD4+ T cell subsets. Anlotinib inhibited the differentiation of Th17 cells in vitro, decreased the phosphorylation of STAT3, and reduced the expression of Rorc. Anlotinib promoted the differentiation of Treg cells and upregulated the expression levels of CD39 and CD73. Discussion and conclusions Anlotinib alleviated the symptoms of EAE via inhibiting the Th17 cell differentiation and promoting Treg cell differentiation. Our study provides new opportunities for the exploitation of anlotinib as a therapeutic agent for the treatment of MS.

Application of a triple‐helical peptide inhibitor of MMP‐2/MMP‐9 to examine T‐cell activation in experimental autoimmune encephalomyelitis

AbstractMultiple sclerosis (MS) is an inflammatory demyelinating disease whereby pathology includes multifocal perivascular mononuclear cell infiltration into the central nervous system, oligodendrocyte loss and demyelination, and variable axonal loss. Little is known about the causative agent in MS, but the matrix metalloproteinase (MMP) family of proteolytic enzymes has been implicated in disease progression and resolution. Recent studies revealed that MMP‐2 and/or MMP‐9 are required for an effective T‐cell‐mediated immune response to antigenic stimulation. In the present studies, we used an MMP‐2/MMP‐9 selective triple‐helical peptide inhibitor (THPI) to examine the role of these enzymes in antigen specific T‐cell responses. We found that treatment of murine CD4+ T‐cells with the THPI, followed by CD3/CD28 stimulation of the T‐cell receptor (TCR), resulted in reduced cell proliferation and production of interleukin 2 (IL‐2), interferon gamma (IFN‐γ), and tumor‐necrosis factor alpha (TNF‐α). Application of the THPI in experimental autoimmune encephalomyelitis (EAE), a widely accepted animal model of MS, reduced clinical severity of the disease and weight loss. Splenocytes isolated from THPI‐treated EAE animals had reduced proliferation compared with untreated animals. In an in vitro recall experiment, splenocytes isolated from EAE mice were treated with the THPI, followed by CD3/CD28 stimulation of the TCR. Treated splenocytes had reduced production of IL‐2, IL‐6, IL‐10, IFN‐γ, and TNF‐α compared with untreated splenocytes isolated from EAE mice. Overall, the MMP‐2/MMP‐9‐targeting THPI modulates T‐cell response to antigenic stimulation, including proliferation and immune cell production of specific cytokines, providing insight as to why the THPI is effective in reducing the clinical severity of EAE.

Prophylactic exposure to oral riluzole reduces the clinical severity and immune-related biomarkers of experimental autoimmune encephalomyelitis

Glutamate-mediated excitotoxicity and immune cell infiltration are hallmarks of multiple sclerosis. The glutamate release inhibitor, riluzole (RIL), has been shown to attenuate the clinical symptoms of experimental autoimmune encephalomyelitis (EAE) in mice, but an association between glutamate excitotoxicity and the progression of MOG35-55-induced EAE has not been well defined. This study investigated the effects of prophylactic and chronic oral RIL on the clinical severity of EAE. Prophylactic+chronic RIL reduced the presence of inflammatory infiltrates, altered GFAP and Foxp3, and attenuated disease severity. These findings indicate a need to delineate the distinct role of glutamate in EAE symptomatology.

Continuous and interval training attenuate encephalomyelitis by separate immunomodulatory mechanisms.

BackgroundStudies have reported beneficial effects of exercise training on autoimmunity, and specifically on multiple sclerosis (MS) and experimental autoimmune encephalomyelitis (EAE). However, it is unknown whether different training paradigms affect disease course via shared or separate mechanisms.ObjectiveTo compare the effects and mechanism of immune modulation of high intensity continuous training (HICT) versus high intensity interval training (HIIT) on systemic autoimmunity in EAE.MethodsWe used the proteolipid protein (PLP)‐induced transfer EAE model to examine training effects on the systemic autoimmune response. Healthy mice performed HICT or HIIT by running on a treadmill. Lymph‐node (LN)‐T cells from PLP‐immunized trained‐ versus sedentary donor mice were transferred to naïve recipients and EAE clinical and pathological severity were assessed. LN cells derived from donor trained and sedentary PLP‐immunized mice were analyzed in vitro for T‐cell activation and proliferation, immune cell profiling, and cytokine mRNA levels and cytokine secretion measurements.ResultsBoth HICT and HIIT attenuated the encephalitogenicity of PLP‐reactive T cells, as indicated by reduced EAE clinical severity and inflammation and tissue pathology in the central nervous system, following their transfer into recipient mice. HICT caused a marked inhibition of PLP‐induced T‐cell proliferation without affecting the T‐cell profile. In contrast, HIIT did not alter T‐cell proliferation, but rather inhibited polarization of T cells into T‐helper 1 and T‐helper 17 autoreactive populations.InterpretationHICT and HIIT attenuate systemic autoimmunity and T cell encephalitogenicity by distinct immunomodulatory mechanisms.

Cnp Promoter-Driven Sustained ERK1/2 Activation Increases B-Cell Activation and Suppresses Experimental Autoimmune Encephalomyelitis.

The ERK1/2 signaling pathway promotes myelin wrapping during development and remyelination, and sustained ERK1/2 activation in the oligodendrocyte (OL) lineage results in hypermyelination of the CNS. We therefore hypothesized that increased ERK1/2 signaling in the OL lineage would 1) protect against immune-mediated demyelination due to increased baseline myelin thickness and/or 2) promote enhanced remyelination and thus functional recovery after experimental autoimmune encephalomyelitis (EAE) induction. Cnp-Cre;Mek1DD-eGFP/+ mice that express a constitutively active form of MEK1 (the upstream activator of ERK1/2) in the OL lineage, exhibited a significant decrease in EAE clinical severity compared to controls. However, experiments using tamoxifen-inducible Plp-CreERT;Mek1DD-eGFP/+ or Pdgfrα-CreERT;Mek1DD-eGFP mice revealed this was not solely due to a protective or reparative effect resulting from MEK1DD expression specifically in the OL lineage. Because EAE is an immune-mediated disease, we examined Cnp-Cre;Mek1DD-eGFP/+ splenic immune cells for recombination. Surprisingly, GFP+ recombined CD19+ B-cells, CD11b+ monocytes, and CD3+ T-cells were noted when Cre expression was driven by the Cnp promoter. While ERK1/2 signaling in monocytes and T-cells is associated with proinflammatory activation, fewer studies have examined ERK1/2 signaling in B-cell populations. After in vitro stimulation, MEK1DD-expressing B-cells exhibited a 3-fold increase in CD138+ plasmablasts and a 5-fold increase in CD5+CD1dhi B-cells compared to controls. Stimulated MEK1DD-expressing B-cells also exhibited an upregulation of IL-10, known to suppress the initiation of EAE when produced by CD5+CD1dhi regulatory B-cells. Taken together, our data support the conclusion that sustained ERK1/2 activation in B-cells suppresses immune-mediated demyelination via increasing activation of regulatory B10 cells.

Enhanced glycolytic metabolism supports transmigration of brain-infiltrating macrophages in multiple sclerosis.

The migration of leukocytes into the CNS drives the neuropathology of multiple sclerosis (MS). This penetration likely utilizes energy resources that remain to be defined. Using the experimental autoimmune encephalomyelitis (EAE) model of MS, we determined that macrophages within the perivascular cuff of post-capillary venules are highly glycolytic as manifested by strong expression of lactate dehydrogenase A (LDHA) that converts pyruvate to lactate. These macrophages expressed prominent levels of monocarboxylate transporter-4 (MCT-4) specialized in secreting lactate from glycolytic cells. The functional relevance of glycolysis was confirmed by siRNA-mediated knockdown of LDHA and MCT-4, which decreased lactate secretion and macrophage transmigration. MCT-4 was in turn regulated by EMMPRIN (CD147) as determined through co-expression/co-immunoprecipitation studies, and siRNA-mediated EMMPRIN silencing. The functional relevance of MCT-4/EMMPRIN interaction was affirmed by lower macrophage transmigration in culture using the MCT-4 inhibitor, α-cyano-4-hydroxy-cinnamic acid (CHCA), a cinnamon derivative. CHCA also reduced leukocyte infiltration and the clinical severity of EAE. Relevance to MS was corroborated by the strong expression of MCT-4, EMMPRIN and LDHA in perivascular macrophages in MS brains. These results detail the metabolism of macrophages for transmigration from perivascular cuffs into the CNS parenchyma and identifies CHCA and diet as potential modulators of neuro-inflammation in MS.

Lactobacillus reuteri reduces the severity of experimental autoimmune encephalomyelitis in mice by modulating gut microbiota

Abstract The gut microbiome plays an important role in immune function and has been implicated in multiple sclerosis (MS). However, how to modulate microbiota to prevent or treat MS remain largely unknown. Probiotic Lactobacillus reuteri DSM 17938 (L. reuteri) has anti-inflammatory effects in mouse models of neonatal necrotizing enterocolitis and Treg-deficiency-induced autoimmune disease. L. reuteri in these models modulated gut microbiota and microbiota-associated metabolites to inhibit TH1 and TH2 differentiation and their-associated cytokines, significantly ameliorating the severity of these diseases. An in vitro study indicated that L. reuteri inhibits the differentiation of naïve CD4+T cells into TH17 and TH1 cells. We determined the effect of L. reuteri on experimental autoimmune encephalomyelitis (EAE) mouse model, a widely used animal model of MS, a model which is primarily mediated by TH17 and TH1 cells. We discovered that L. reuteri treatment reduced TH1/TH17 cells and their associated cytokines IFN-γ/IL-17 in EAE, inhibiting the development of EAE. We also showed that the loss of diversity in gut microbiota induced by EAE was largely restored by L. reuteri treatment. Taxonomy-based analysis of gut microbiota showed that the “beneficial” genera Bifidobacterium, Prevotella, and Lactobacillus were negatively correlated with EAE clinical severity scores, whereas the genera Anaeroplasma, Rikenellaceae, and Clostridium were positively correlated. Notably, L. reuteri treatment altered the relative abundances of these EAE-associated taxa in mice. In conclusion, probiotic L. reuteri changed gut microbiota to modulate immune responses in EAE, making it a novel candidate in future studies to modify the severity of MS.

Continuous cuprizone intoxication allows active experimental autoimmune encephalomyelitis induction in C57BL/6 mice.

Oligodendrocyte degeneration is a hallmark of multiple sclerosis pathology, and protecting oligodendrocytes and myelin is likely to be of clinical relevance. Traditionally, oligodendrocyte and myelin degeneration are viewed as a direct consequence of an inflammatory attack, but metabolic defects might be equally important. Appropriate animal models to study the interplay of inflammation and metabolic injury are, therefore, needed. Here, we describe that in spite of its immunosuppressive effects, a continuous intoxication with cuprizone allows the induction of active experimental autoimmune encephalomyelitis (EAE) by myelin oligodendrocyte glycoprotein (MOG35-55) immunization. Although the clinical severity of EAE is ameliorated in cuprizone-intoxicated mice, the recruitment of granulocytes, and especially, CD3+ lymphocytes into the forebrain is triggered by the cuprizone insult. Such combined lesions are further characterized by oligodendrocyte apoptosis and microglia activation, closely mimicking type III multiple sclerosis lesions. In summary, we provide a protocol that allows to study the direct interplay of immune-mediated and metabolic oligodendrocyte injury and its consequences for the cerebral white and grey matters.

Lactobacillus reuteri Reduces the Severity of Experimental Autoimmune Encephalomyelitis in Mice by Modulating Gut Microbiota

The gut microbiome plays an important role in immune function and has been implicated in multiple sclerosis (MS). However, how and if the modulation of microbiota can prevent or treat MS remain largely unknown. In this study, we showed that probiotic Lactobacillus reuteri DSM 17938 (L. reuteri) ameliorated the development of murine experimental autoimmune encephalomyelitis (EAE), a widely used animal model of MS, a model which is primarily mediated by TH17 and TH1 cells. We discovered that L. reuteri treatment reduced TH1/TH17 cells and their associated cytokines IFN-γ/IL-17 in EAE mice. We also showed that the loss of diversity of gut microbiota induced by EAE was largely restored by L. reuteri treatment. Taxonomy-based analysis of gut microbiota showed that three “beneficial” genera Bifidobacterium, Prevotella, and Lactobacillus were negatively correlated with EAE clinical severity, whereas the genera Anaeroplasma, Rikenellaceae, and Clostridium were positively correlated with disease severity. Notably, L. reuteri treatment coordinately altered the relative abundance of these EAE-associated taxa. In conclusion, probiotic L. reuteri changed gut microbiota to modulate immune responses in EAE, making it a novel candidate in future studies to modify the severity of MS.

Nasal delivery of Fasudil-modified immune cells exhibits therapeutic potential in experimental autoimmune encephalomyelitis.

SummaryAimMultiple sclerosis (MS) is a relapsing‐remitting inflammatory demyelinating disease that requires long‐term treatment. Although Rho kinase inhibitor Fasudil shows good therapeutic effect in experimental autoimmune encephalomyelitis (EAE), an animal model of MS, certain side effects may limit its clinical use. This study aimed at observing the therapeutic potential of Fasudil‐modified encephalitogenic mononuclear cells (MNCs) via nasal delivery in EAE and exploring possible mechanisms of action.MethodsExperimental autoimmune encephalomyelitis was induced with myelin oligodendrocyte glycoprotein 35‐55 in C57BL/6 mice, and encephalitogenic MNCs were treated with Fasudil in vitro. Mice received 3 × 106 cells/10 μL per nasal cavity on day 3 and 11 postimmunization, respectively.ResultsFasudil‐modified MNCs reduced clinical severity of EAE, improved demyelination, and decreased inflammatory cells in spinal cords. Immunohistochemical results indicated that CD4+ T cells and CD68+ macrophages were barely detected in Fasudil‐MNCs group. Fasudil‐modified MNCs decreased CD4+IFN‐γ+ and CD4+IL‐17+ T cells, increased CD4+IL‐10+ T cells, restrained M1 markers CD16/32, CCR7, IL‐12, CD8a, enhanced M2 markers CD206, CD200, CD14 in spleen. Fasudil‐modified MNCs inhibited the activation of inflammatory signaling p‐NF‐kB/P38, accompanied by the decrease of COX‐2 and the increase of Arg‐1 in spinal cord, as well as the reduction of IL‐17, TNF‐α, IL‐6 and the elevation of IL‐10 in cultured supernatant of splenocytes. Fasudil‐modified MNCs enhanced the levels of neurotrophic factors BDNF and NT‐3 in spinal cord.ConclusionOur results indicate that intranasal delivery of Fasudil‐modified MNCs have therapeutic potential in EAE, providing a safe and effective cell therapeutic strategy to MS and/or other related disorders.

Dexmedetomidine, an Alpha 2a Adrenergic Receptor Agonist, Mitigates Experimental Autoimmune Encephalomyelitis by Desensitization of CXCR7 in Microglia

The autoimmune disease multiple sclerosis (MS) and its animal model, experimental autoimmune encephalomyelitis (EAE), is characterized by an ascending paralysis that is characterized by extensive infiltration of the central nervous system by inflammatory cells. Although several studies to some extent uncover the cellular mechanisms of microglia that govern EAE pathogenesis, the molecular mechanisms that orchestrate the movement of microglia remain unknown, and potential novel therapeutic strategies are still required. In this study, we report that dexmedetomidine, an alpha 2a adrenergic receptor agonist, attenuates the clinical severity of EAE with less infiltration of microglia. During EAE, dexmedetomidine inhibits SDF-1- and I-TAC-induced chemotaxis of microglia mediated by CXCR7 but not CXCR4 or CXCR3. Most importantly, the alpha 2a adrenergic receptor is essential in dexmedetomidine-induced CXCR7 desensitization in microglia. Further experiments confirmed that CXCR7 desensitization required atypical protein kinase C ζ activation, while conventional and novel protein kinase C isoforms were not involved. Altogether, our data elucidate the mechanism of dexmedetomidine-induced CXCR7 desensitization in microglia and amelioration in EAE, which might lead to a better understanding of the therapeutic effects of dexmedetomidine as well as its implications for CXCR7 desensitization in autoimmune disease.

Isoprenol-Induced Neuroprotection in Experimental Multiple Sclerosis

Abstract Farnesol is a 15-carbon organic acyclic sesquiterpene alcohol (isoprenol) produced by plants and animals. It is a potent blocker of neuronal voltage-gated Ca2+ found in the human brain. It has potent anti-oxidant and antiinflammatory effects in vitro, and neuroprotection based on the regulation of free radicals production by glial cells was demonstrated in a murine model of neurotoxicity. Because inflammation, oxidative stress and intraneuronal Ca2+ overload are associated with central nervous system (CNS) demyelinating diseases, we sought to determine if farnesol treatment would result in protection against experimental autoimmune encephalomyelitis (EAE), a model of multiple sclerosis (MS). We compared the progression of EAE in C57BL/6 mice treated orally with farnesol (100 mg/kg/daily by gavage in olive oil) versus EAE progression in untreated mice (control) (n = 10/group). Treatment started at EAE induction and continued for 25 days. EAE clinical severity was monitored daily. Factorial repeated measures ANOVA of the scores showed: 1) a significant Day effect (disease onset 9 days after induction), 2) a significant Group effect (approx. 80% reduction in scores in the Farnesol group), and 3) a significant Day * Group interaction with scores being significantly lower in the Farnesol group every day starting one day after disease onset. At day 25, 50% of control mice had been euthanized due to scores ≥ 3.5 vs. none in the Farnesol group. Our results indicate that Farnesol significantly reduces disease onset in EAE. Studies to elucidate the mechanism of action of farnesol, screening of structural analogues of farnesol for anti-EAE activity, and assessment of farnesol therapeutic efficacy in other animal models of MS are being conducted.

Isoprenol‐Induced Neuroprotection in Experimental Multiple Sclerosis

Study ObjectivesFarnesol is a 15‐carbon organic acyclic sesquiterpene alcohol (isoprenol) produced by plants and animals. It is a potent blocker of neuronal voltage‐gated Ca2+ channels (L‐ and N‐type) and it is found in the human brain (Roullet, J. Biol. Chem., 1999). Other studies have shown farnesol has potent anti‐oxidant and anti‐inflammatory effects in vitro. Neuroprotection by farnesol was demonstrated in a murine model of neurotoxicity in a mechanism based on the regulation of the production of free radicals by glial cells, and the production of proinflammatory cytokines within the central nervous system. Because inflammation, oxidative stress and intraneuronal Ca2+ overload are associated with CNS demyelinating diseases, we sought to determine if farnesol treatment would result in protection against experimental autoimmune encephalomyelitis (EAE), a well‐established model of multiple sclerosis (MS).MethodsWe compared the progression of EAE in myelin‐oligodendrocyte‐glycoprotein (MOG35–55) immunized C57BL/6 mice (male, approx. 20 g body weight) treated orally with farnesol (100 mg/kg administered daily by gavage in olive oil) versus EAE progression in untreated mice (control) (n = 10/group). Farnesol treatment was started at the time of EAE induction and continued for 25 days at which time animals were euthanized. EAE clinical severity (CS) was monitored daily using a 0 to 5 neurological (paralysis) scoring system (0, normal, 5 most severe or death).ResultsFactorial repeated measures ANOVA of the CS scores showed: 1) a significant (p&lt;0.001) Day effect (disease onset 9 days after induction), 2) a significant (p&lt;0.002) Group effect (approx. 80% reduction in CS scores in the Farnesol group on average), and 3) a significant (p&lt;0.001) [Day * Group] interaction with CS scores being significantly lower in the Farnesol group at all time points every day starting one day after disease onset until day 25). At day 25, 5 out of 10 mice had died or had been euthanized due to scores ≥ 3.5 in controls vs. none in the Farnesol group. Mice treated with farnesol maintained a constant body weight (BW, in g) throughout the study (BW = 19.1, 19.2, 18.6 and 19.6 on average at days 0, 7, 14 and 21 respectively), and displayed normal locomotor activity (video not shown). In contrast, control mice lost weight: 18.1, 18.8, 16.1 and 17.5 on average at days 0, 7, 14 and 21 (ANOVA: p&lt;0.012 [Group], p &lt; 0.001 [Day], and p&lt;0.012 [Group * Day] interaction).Discussion/ConclusionFarnesol significantly reduces disease onset in experimental MS, suggesting that farnesol may have therapeutic efficacy in MS patients. The molecular and cellular mechanisms underlying disease protection by farnesol remain to be determined but could involve protection against either intracellular Ca2+ overload, oxidative stress, or inflammation. Studies to elucidate the mechanism of action of farnesol, screening of structural analogues of farnesol for anti‐MS activity, and assessment of farnesol therapeutic efficacy in other animal models of MS are being conducted.

A Kv1.3 channel-specific blocker alleviates neurological impairment through inhibiting T-cell activation in experimental autoimmune encephalomyelitis.

Multiple sclerosis (MS) is a neurological autoimmune disorder characterized by mistaken attacks of inflammatory cells against the central nervous system (CNS), resulting in demyelination and axonal damage. Kv1.3 channel blockers can inhibit T-cell activation and have been designed for MS therapy. However, little is known about the effects of Kv1.3 blockers on protecting myelin sheaths/axons in MS. This study aimed at investigating the neuroprotection efficacy of a selective Kv1.3 channel blocker ImKTx88 (ImK) in MS animal model. Experimental autoimmune encephalomyelitis (EAE) rat model was established. The neuroprotective effect of ImK was assessed by immunohistochemistry and transmission electron microscopy (TEM). In addition, the antiinflammatory effect of ImK by suppressing T-cell activation was assessed by flow cytometry and ELISA in vitro. Our results demonstrated that ImK administration ameliorated EAE clinical severity. Moreover, ImK increased oligodendrocytes survival, preserved axons, and myelin integrity and reduced the infiltration of activated T cells into the CNS. This protective effect of the peptide may be related to its suppression of autoantigen-specific T-cell activation via calcium influx inhibition. ImK prevents neurological damage by suppressing T-cell activation, suggesting the applicability of this peptide in MS therapy.

ATG-dependent phagocytosis in dendritic cells drives myelin-specific CD4+ T cell pathogenicity during CNS inflammation

Although reactivation and accumulation of autoreactive CD4+ T cells within the CNS are considered to play a key role in the pathogenesis of multiple sclerosis (MS) and its animal model, experimental autoimmune encephalomyelitis (EAE), the mechanisms of how these cells recognize their target organ and induce sustained inflammation are incompletely understood. Here, we report that mice with conditional deletion of the essential autophagy protein ATG5 in classical dendritic cells (DCs), which are present at low frequencies in the nondiseased CNS, are completely resistant to EAE development following adoptive transfer of myelin-specific T cells and show substantially reduced in situ CD4+ T cell accumulation during the effector phase of the disease. Endogenous myelin peptide presentation to CD4+ T cells following phagocytosis of injured, phosphatidylserine-exposing oligodendroglial cells is abrogated in the absence of ATG5. Pharmacological inhibition of ATG-dependent phagocytosis by the cardiac glycoside neriifolin, an inhibitor of the Na+, K+-ATPase, delays the onset and reduces the clinical severity of EAE induced by myelin-specific CD4+ T cells. These findings link phagocytosis of injured oligodendrocytes, a pathological hallmark of MS lesions and during EAE, with myelin antigen processing and T cell pathogenicity, and identify ATG-dependent phagocytosis in DCs as a key regulator in driving autoimmune CD4+ T cell-mediated CNS damage.