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1.  Metformin Attenuated the Autoimmune Disease of the Central Nervous System in Animal Models of Multiple Sclerosis1 
Experimental autoimmune encephalomyelitis (EAE) is a T cell-mediated autoimmune disease of the CNS. Metformin is the most widely used drug for diabetes and mediates its action via activating AMP-activated protein kinase (AMPK). We provide evidence that metformin attenuates the induction of EAE by restricting the infiltration of mononuclear cells into the CNS, down-regulating the expression of proinflammatory cytokines (IFN-γ, TNF-α, IL-6, IL-17, and inducible NO synthase (iNOS)), cell adhesion molecules, matrix metalloproteinase 9, and chemokine (RANTES). Furthermore, the AMPK activity and lipids alterations (total phospholipids and in free fatty acids) were restored by metformin treatment in the CNS of treated EAE animals, suggesting the possible involvement of AMPK. Metformin activated AMPK in macrophages and thereby inhibited biosynthesis of phospholipids as well as neutral lipids and also down-regulated the expression of endotoxin (LPS)-induced proinflammatory cytokines and their mediators (iNOS and cyclooxygenase 2). It also attenuated IFN-γ and IL-17-induced iNOS and cyclooxygenase 2 expression in RAW267.4 cells, further supporting its anti-inflammatory property. Metformin inhibited T cell-mediated immune responses including Ag-specific recall responses and production of Th1 or Th17 cytokines, while it induced the generation of IL-10 in spleen cells of treated EAE animals. Altogether these findings reveal that metformin may have a possible therapeutic value for the treatment of multiple sclerosis and other inflammatory diseases.
PMCID: PMC2965405  PMID: 19494326
2.  Calpain inhibitor attenuated optic nerve damage in acute optic neuritis in rats 
Journal of neurochemistry  2012;124(1):133-146.
Optic neuritis (ON), which is an acute inflammatory autoimmune demyelinating disease of the central nervous system (CNS), often occurs in multiple sclerosis (MS). ON is an early diagnostic sign in most MS patients caused by damage to the optic nerve leading to visual dysfunction. Various features of both MS and ON can be studied following induction of experimental autoimmune encephalomyelitis (EAE), an animal model of MS, in Lewis rats. Inflammation and cell death in the optic nerve, with subsequent damage to the retinal ganglion cells in the retina, are thought to correlate with visual dysfunction. Thus, characterizing the pathophysiological changes that lead to visual dysfunction in EAE animals may help develop novel targets for therapeutic intervention. We treated EAE animals with and without the calpain inhibitor calpeptin (CP). Our studies demonstrated that the Ca2+-activated neutral protease calpain was upregulated in the optic nerve following induction of EAE at the onset of clinical signs (OCS) of the disease and these changes were attenuated following treatment with CP. These reductions correlated with decreases in inflammation (cytokines, iNOS, COX-2, NF-κB), and microgliosis (i.e. activated microglia). We observed that calpain inhibition reduced astrogliosis (reactive astroglia) and expression of aquaporin 4 (AQP4). The balance of Th1/Th2 cytokine production and also expression of the Th1-related CCR5 and CXCR3 chemokine receptors influence many pathological processes and play both causative and protective roles in neuron damage. Our data indicated that CP suppressed cytokine imbalances. Also, Bax:Bcl-2 ratio, production of tBid, PARP-1, expression and activities of calpain and caspases, and internucleosomal DNA fragmentation were attenuated after treatment with CP. Our results demonstrated that CP decreased demyelination [loss of myelin basic protein (MBP)] and axonal damage [increase in dephosphorylated neurofilament protein (de-NFP), and also promoted intracellular neuroprotective pathways in optic nerve in EAE rats. Thus, these data suggest that calpain is involved in inflammatory as well as in neurodegenerative aspects of the disease and may be a promising target for treating ON in EAE and MS.
PMCID: PMC3570118  PMID: 23106593
apoptosis; calpain; chemokines; cytokines; EAE; inflammation; optic neuritis
3.  Bowman-Birk Inhibitor Suppresses Autoimmune Inflammation and Neuronal Loss in a Mouse Model of Multiple Sclerosis 
Journal of the neurological sciences  2008;271(1-2):191-202.
The Bowman-Birk inhibitor (BBI) is a soybean-derived serine protease inhibitor. BBI concentrate (BBIC) is an extract enriched with BBI, but predominantly contains other ingredients including several protease inhibitors. We previously found that BBIC administration to Lewis rats with experimental autoimmune encephalomyelitis (EAE) significantly suppresses disease. In the present study we determined whether BBI mediates the suppressive effects of BBIC in EAE, evaluated its potential neuroprotective effects, and investigated mechanisms of BBI action. We tested effects of purified BBI on clinical and histopathological parameters of EAE in two models (relapsing/remitting EAE in SJL/J mice and chronic EAE in C57BL/6 mice). Effects of BBI were compared to BBIC in relapsing/remitting EAE, and effects of BBI on neuronal survival were examined during acute optic neuritis. Treatment with BBI in both EAE models significantly improved EAE disease parameters (onset, severity, weight loss, inflammation and demyelination). BBI significantly reduced the incidence of optic neuritis and prevented loss of retinal ganglion cells. In most experiments proliferation of immune cells derived from BBI-treated mice was significantly lower relative to control groups. Using Boyden’s chamber assay we found that BBI inhibited invasiveness of activated splenocytes through the matrigel barrier. BBI also induced higher production of EAE-suppressive cytokine IL-10 by immune cells. These results demonstrate that BBI is the active component of BBIC that ameliorates clinical EAE. BBI reduces inflammation and attenuates neuronal loss, making it an excellent candidate for oral therapy in MS. BBI likely ameliorates EAE by inhibiting multiple pathways involved in disease pathogenesis.
PMCID: PMC2723951  PMID: 18544456
Multiple sclerosis; EAE; serine proteases; Bowman-Birk inhibitor; neuroprotection; neuroimmunology
4.  Calcium Intervention Ameliorates Experimental Model of Multiple Sclerosis 
Oman Medical Journal  2014;29(3):185-189.
Multiple sclerosis (MS) is the most common inflammatory disease of the CNS. Experimental autoimmune encephalomyelitis (EAE) is a widely used model for MS. In the present research, our aim was to test the therapeutic efficacy of Calcium (Ca) in an experimental model of MS.
In this study the experiment was done on C57BL/6 mice. EAE was induced using 200 μg of the MOG35-55 peptide emulsified in CFA and injected subcutaneously on day 0 over two flank areas. In addition, 250 ng of pertussis toxin was injected on days 0 and 2. In the treatment group, 30 mg/kg Ca was administered intraperitoneally four times at regular 48 hour intervals. The mice were sacrificed 21 days after EAE induction and blood samples were taken from their hearts. The brains of mice were removed for histological analysis and their isolated splenocytes were cultured.
Our results showed that treatment with Ca caused a significant reduction in the severity of the EAE. Histological analysis indicated that there was no plaque in brain sections of Ca treated group of mice whereas 4 ± 1 plaques were detected in brain sections of controls. The density of mononuclear infiltration in the CNS of Ca treated mice was lower than in controls. The serum level of Nitric Oxide in the treatment group was lower than in the control group but was not significant. Moreover, the levels of IFN-γ in cell culture supernatant of splenocytes in treated mice were significantly lower than in the control group.
The data indicates that Ca intervention can effectively attenuate EAE progression.
PMCID: PMC4052390  PMID: 24936267
MS; CNS; EAE; Calcium; Nitric Oxide
5.  Methylprednisolone inhibits IFN-γ and IL-17 expression and production by cells infiltrating central nervous system in experimental autoimmune encephalomyelitis 
Glucocorticoids have been shown to be effective in the treatment of autoimmune diseases of the CNS such as multiple sclerosis and its animal model, experimental autoimmune encephalomyelitis (EAE). However, the mechanisms and the site of glucocorticoids' actions are still not completely defined. The aim of this study was to investigate the in vivo effect of the synthetic glucocorticoid methylprednisolone (MP) on the expression and production of proinflammatory cytokines interferon (IFN)-γ and interleukin (IL)-17 by cells infiltrating CNS tissue.
Experimental autoimmune encephalomyelitis was induced in Dark Agouti (DA) rats by immunization with rat spinal cord homogenate mixed with adjuvants. Commencing on the day when the first EAE signs appeared, DA rats were injected daily for 3 days with MP and/or RU486, an antagonist of glucocorticoid receptor. Cytokine production and gene expression in CNS-infiltrating cells and lymph node cells were measured using ELISA and real time PCR, respectively.
Treatment of rats with MP ameliorated EAE, and the animals recovered without relapses. Further, MP inhibited IFN-γ and IL-17 expression and production in cells isolated from the CNS of DA rats with EAE after the last injection of MP. The observed effect of MP in vivo treatment was not mediated through depletion of CD4+ T cells among CNS infiltrating cells, or through induction of their apoptosis within the CNS. Finally, the glucocorticoid receptor-antagonist RU486 prevented the inhibitory effect of MP on IFN-γ and IL-17 production both in vitro and in vivo, thus indicating that the observed effects of MP were mediated through glucocorticoid receptor-dependent mechanisms.
Taken together, these results demonstrate that amelioration of EAE by exogenous glucocorticoids might be, at least partly, ascribed to the limitation of effector cell functions in the target tissue.
PMCID: PMC2797773  PMID: 20003332
6.  Passive Experimental Autoimmune Encephalomyelitis in C57BL/6 with MOG: Evidence of Involvement of B Cells 
PLoS ONE  2012;7(12):e52361.
Experimental autoimmune encephalomyelitis (EAE) is the most relevant animal model to study demyelinating diseases such as multiple sclerosis. EAE can be induced by active (active EAE) or passive (at-EAE) transfer of activated T cells in several species and strains of rodents. However, histological features of at-EAE model in C57BL/6 are poorly described. The aim of this study was to characterize the neuroinflammatory and neurodegenerative responses of at-EAE in C57BL/6 mice by histological techniques and compare them with that observed in the active EAE model. To develop the at-EAE, splenocytes from active EAE female mice were harvested and cultured in presence of MOG35–55 and IL-12, and then injected intraperitoneally in recipient female C57BL6/J mice. In both models, the development of EAE was similar except for starting before the onset of symptoms and presenting a higher EAE cumulative score in the at-EAE model. Spinal cord histological examination revealed an increased glial activation as well as more extensive demyelinating areas in the at-EAE than in the active EAE model. Although inflammatory infiltrates composed by macrophages and T lymphocytes were found in the spinal cord and brain of both models, B lymphocytes were significantly increased in the at-EAE model. The co-localization of these B cells with IgG and their predominant distribution in areas of demyelination would suggest that IgG-secreting B cells are involved in the neurodegenerative processes associated with at-EAE.
PMCID: PMC3530560  PMID: 23300649
7.  Oral tolerance to myelin basic protein and natural recovery from experimental autoimmune encephalomyelitis are associated with downregulation of inflammatory cytokines and differential upregulation of transforming growth factor beta, interleukin 4, and prostaglandin E expression in the brain 
The Journal of Experimental Medicine  1992;176(5):1355-1364.
Experimental autoimmune encephalomyelitis (EAE) in the Lewis rat is a self-limited inflammatory process localized to the central nervous system that is induced by the injection of myelin basic protein (MBP) in adjuvant. Oral administration of MBP suppresses EAE, and this suppression is mediated by CD8+ T cells that adoptively transfer protection and suppress both in vitro and in vivo by the release of transforming growth factor (TGF) beta after antigen-specific triggering. Furthermore, oral tolerance to MBP is enhanced by the concomitant oral administration of lipopolysaccharide (LPS). The present study was undertaken to determine whether the disease course in EAE and its suppression by oral tolerization to MBP is associated with distinct patterns of cytokine expression in the target organ. Detailed immunohistology of the brain was performed at the peak of clinical disease (day 14 after immunization) and after recovery (day 18) in control (ovalbumin [OVA]-fed), MBP-fed, and MBP plus LPS-fed animals. Brains from OVA-fed animals at the peak of disease showed perivascular infiltration with activated mononuclear cells which secreted the inflammatory cytokines interleukins (IL) 1, 2, 6, 8, TNF-alpha, and interferon gamma. The inhibitory cytokines TGF-beta and IL-4, and prostaglandin E2 (PGE2) were absent. In MBP orally tolerized animals there was a marked reduction of the perivascular infiltrate and downregulation of all inflammatory cytokines. In addition, there was upregulation of the inhibitory cytokine TGF-beta. In MBP plus LPS orally tolerized animals, in addition to upregulation of TGF-beta and reduction of inflammatory cytokines, there was enhanced expression of IL-4 and PGE2, presumably secondary to activation of an additional population of immunoregulatory cells. In OVA-fed animals that had recovered (day 18), staining for inflammatory cytokines diminished, and there was the appearance of TGF-beta and IL-4. These results suggest that suppression of EAE, either induced by oral tolerization or that which occurs during natural recovery is related to the secretion of inhibitory cytokines or factors that actively suppress the inflammatory process in the target organ.
PMCID: PMC2119419  PMID: 1383385
8.  TREM2-Transduced Myeloid Precursors Mediate Nervous Tissue Debris Clearance and Facilitate Recovery in an Animal Model of Multiple Sclerosis 
PLoS Medicine  2007;4(4):e124.
In multiple sclerosis, inflammation can successfully be prevented, while promoting repair is still a major challenge. Microglial cells, the resident phagocytes of the central nervous system (CNS), are hematopoietic-derived myeloid cells and express the triggering receptor expressed on myeloid cells 2 (TREM2), an innate immune receptor. Myeloid cells are an accessible source for ex vivo gene therapy. We investigated whether myeloid precursor cells genetically modified to express TREM2 affect the disease course of experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis.
Methods and Findings
EAE was induced in mice by immunization with a myelin autoantigen. Intravenous application of TREM2-transduced bone marrow–derived myeloid precursor cells at the EAE peak led to an amelioration of clinical symptoms, reduction in axonal damage, and prevention of further demyelination. TREM2-transduced myeloid cells applied intravenously migrated into the inflammatory spinal cord lesions of EAE-diseased mice, showed increased lysosomal and phagocytic activity, cleared degenerated myelin, and created an anti-inflammatory cytokine milieu within the CNS.
Intravenously applied bone marrow–derived and TREM2-tranduced myeloid precursor cells limit tissue destruction and facilitate repair within the murine CNS by clearance of cellular debris during EAE. TREM2 is a new attractive target for promotion of repair and resolution of inflammation in multiple sclerosis and other neuroinflammatory diseases.
Myeloid precursors genetically modified to express TREM2 supported recovery in animals with experimental autoimmune encephalomyelitis by phagocytosis and removal of apoptotic cellular debris.
Editors' Summary
Multiple sclerosis (MS) is a progressive disease of the central nervous system (CNS; the brain and spinal cord) in which communications within the brain are disrupted. In healthy individuals, the brain controls the body by sending electrical messages along nerve fibers. Like electrical wires, these fibers have to be insulated to allow the efficient transfer of electrical impulses. This insulation is provided by myelin, a fatty tissue that surrounds the nerve fibers. In patients with MS, patchy myelin loss, inflammation, and scarring (sclerosis) disrupt nerve transmissions, causing weakness or paralysis of the limbs, balance and coordination problems, and numerous other symptoms. The damage to the myelin (which increases over time) is caused by the patient's immune system, but what triggers this abnormal behavior is unclear. There is no cure for MS, but drugs that modulate the immune system (for example, interferons and steroids) can slow its progression.
Why Was This Study Done?
Any treatment for MS needs to limit tissue destruction by the immune system and to promote the repair of the damaged myelin and nerve fibers. Although researchers have identified some agents that promote nerve repair, it is difficult to get them into the CNS, which is protected by an impermeable barrier. However, some cell types can migrate into the CNS so it might be possible to adapt them as delivery vehicles for therapeutic agents. In this study, the researchers investigated whether bone marrow–derived myeloid precursor cells (a source of several immune system cell types) might be useful for the treatment of MS, both as delivery vehicles and as a source of cells that can resolve inflammation and clear cellular debris from sites of myelin destruction; this clearance has to occur before nerves and myelin can be repaired. Myeloid precursor cells, the researchers reasoned, might satisfy both these requirements because they naturally migrate into the CNS where some turn into microglia, immune system cells that engulf and digest cellular debris.
What Did the Researchers Do and Find?
The researchers isolated myeloid precursor cells from mouse bone marrow and induced the expression of triggering receptor expressed on myeloid cells-2 (TREM2, a protein made by microglia) in them using a viral vector. They injected these TREM2-expressing cells into the veins of mice with experimental autoimmune encephalomyelitis (EAE, an animal model of MS) and examined their migration into the spinal cord, their effect on EAE symptoms, and their effect on cell debris clearance and inflammatory responses in spinal cord lesions. Neither TREM2-expressing nor control myeloid precursor cells migrated into the spinal cord when injected into healthy mice or into animals just beginning to show the symptoms of EAE. However, both cell types migrated into the spinal cord when injected into animals when EAE symptoms were at their peak. The injection of TREM2-expressing myeloid precursor cells (but not control myeloid precursor cells) at this time reduced EAE symptoms and nerve damage, and halted myelin loss. It also increased the clearance of cell debris and myelin fragments and created an anti-inflammatory environment in the spinal cord lesions.
What Do These Findings Mean?
These findings indicate that the intravenous injection of bone marrow–derived myeloid precursor cells engineered to express TREM2 can reduce clinical symptoms and induce recovery in mice with EAE. Whether the TREM2-expressing myeloid precursor cells act locally in the CNS or indirectly via other immune system cells is not clear yet. In addition, the findings need to be replicated in more animals before this approach is tested in people. Nevertheless, the intravenous injection of cells of this type could provide a two-pronged approach to the treatment of MS (and other neuroinflammatory diseases). First, bone marrow–derived myeloid precursor cells that have been engineered to express TREM2 could help to treat MS by clearing tissue debris and reducing inflammation in spinal cord lesions. Second, the same cells could be used to deliver agents to the lesions that promote nerve repair. Importantly, these results suggest that TREM2-expressing myeloid precursor cells might have the potential to treat MS after the onset of clinical symptoms, an important consideration for a chronic disease that often indicates relapses by new clinical symptoms.
Additional Information.
Please access these Web sites via the online version of this summary at
US National Institute of Neurological Disorders and Stroke patient information on multiple sclerosis (in English and Spanish)
UK National Health Service health encyclopedia information on multiple sclerosis
MedlinePlus encyclopedia pages on multiple sclerosis
The US National MS Society information on multiple sclerosis for patients and professionals (in English and Spanish)
UK MS Society information on multiple sclerosis for patients and professionals
Multiple Sclerosis International Federation information on multiple sclerosis for patients, caregivers, and professionals (in English, Arabic, German, Spanish, French, Italian and Russian)
PMCID: PMC1851623  PMID: 17425404
9.  Deletion of IL-33R (ST2) Abrogates Resistance to EAE in BALB/C Mice by Enhancing Polarization of APC to Inflammatory Phenotype 
PLoS ONE  2012;7(9):e45225.
The administration of interleukin 33 and deletion of IL-33 receptor, ST2 molecule, affects the induction of autoimmunity in different experimental models of human autoimmune diseases. The aim of this study was to analyze the effect of ST2 deletion on the induction of experimental autoimmune encephalomyelitis (EAE) in resistant BALB/c mice. Mice were immunized with MOG35–55 peptide or disease was induced by passive transfer of encephalitogenic singenic cells and EAE was clinically and histologically evaluated. Expression of intracellular inflammatory cytokines, markers of activation and chemokine receptors on lymphoid tissue and CNS infiltrating mononuclear cells was analyzed by flow cytometry. We report here that deletion of ST2−/− molecule abrogates resistance of BALB/c mice to EAE induction based on clinical and histopathological findings. Brain and spinal cord infiltrates of ST2−/− mice had significantly higher number of CD4+ T lymphocytes containing inflammatory cytokines compared to BALB/c WT mice. Adoptive transfer of ST2−/− primed lymphocytes induced clinical signs of the disease in ST2−/− as well as in WT mice. MOG35–55 restimulated ST2−/− CD4+ cells as well as ex vivo analyzed lymph node cells had higher expression of T-bet and IL-17, IFN-γ, TNF-α and GM-CSF in comparison with WT CD4+ cells. ST2−/− mice had higher percentages of CD4+ cells expressing chemokine receptors important for migration to CNS in comparison with WT CD4+ cells. Draining lymph nodes of ST2−/− mice contained higher percentage of CD11c+CD11b+CD8− cells containing inflammatory cytokines IL-6 and IL-12 with higher expression of activation markers. Transfer of ST2−/− but not WT dendritic cells induced EAE in MOG35–55 immunized WT mice. Our results indicate that ST2 deficiency attenuates inherent resistance of BALB/c mice to EAE induction by enhancing differentiation of proinflammatory antigen presenting cells and consecutive differentiation of encephalitogenic T cells in the draining lymph node rather than affecting their action in the target tissue.
PMCID: PMC3445483  PMID: 23028861
10.  Lithium Prevents and Ameliorates Experimental Autoimmune Encephalomyelitis1 
Experimental autoimmune encephalomyelitis (EAE) models, in animals, many characteristics of multiple sclerosis, for which there is no adequate therapy. We investigated whether lithium, an inhibitor of glycogen synthase kinase-3 (GSK3), can ameliorate EAE in mice. Pretreatment with lithium markedly suppressed the clinical symptoms of EAE induced in mice by myelin oligodendrocyte glycoprotein peptide (MOG35–55) immunization and greatly reduced demyelination, microglia activation, and leukocyte infiltration in the spinal cord. Lithium administered postimmunization, after disease onset, reduced disease severity and facilitated partial recovery. Conversely, in knock-in mice expressing constitutively active GSK3, EAE developed more rapidly and was more severe. In vivo lithium therapy suppressed MOG35–55-reactive effector T cell differentiation, greatly reducing in vitro MOG35–55-stimulated proliferation of mononuclear cells from draining lymph nodes and spleens, and MOG35–55-induced IFN-γ, IL-6, and IL-17 production by splenocytes isolated from MOG35–55-immunized mice. In relapsing/remitting EAE induced with proteolipid protein peptide139–151, lithium administered after the first clinical episode maintained long-term (90 days after immunization) protection, and after lithium withdrawal the disease rapidly relapsed. These results demonstrate that lithium suppresses EAE and identify GSK3 as a new target for inhibition that may be useful for therapeutic intervention of multiple sclerosis and other autoimmune and inflammatory diseases afflicting the CNS.
PMCID: PMC2704026  PMID: 18566399
11.  Erythropoietin: A Potent Inducer of Peripheral Immuno/Inflammatory Modulation in Autoimmune EAE 
PLoS ONE  2008;3(4):e1924.
Beneficial effects of short-term erythropoietin (EPO) therapy have been demonstrated in several animal models of acute neurologic injury, including experimental autoimmune encephalomyelitis (EAE)-the animal model of multiple sclerosis. We have found that EPO treatment substantially reduces the acute clinical paralysis seen in EAE mice and this improvement is accompanied by a large reduction in the mononuclear cell infiltration and downregulation of glial MHC class II expression within the inflamed CNS. Other reports have recently indicated that peripherally generated anti-inflammatory CD4+Foxp3+ regulatory T cells (Tregs) and the IL17-producing CD4+ T helper cell (Th17) subpopulations play key antagonistic roles in EAE pathogenesis. However, no information regarding the effects of EPO therapy on the behavior of the general mononuclear-lymphocyte population, Tregs or Th17 cells in EAE has emerged.
Methods and Findings
We first determined in vivo that EPO therapy markedly suppressed MOG specific T cell proliferation and sharply reduced the number of reactive dendritic cells (CD11c positive) in EAE lymph nodes during both inductive and later symptomatic phases of MOG35–55 induced EAE. We then determined the effect in vivo of EPO on numbers of peripheral Treg cells and Th17 cells. We found that EPO treatment modulated immune balance in both the periphery and the inflamed spinal cord by promoting a large expansion in Treg cells, inhibiting Th17 polarization and abrogating proliferation of the antigen presenting dendritic cell population. Finally we utilized tissue culture assays to show that exposure to EPO in vitro similarly downregulated MOG-specific T cell proliferation and also greatly suppressed T cell production of pro-inflammatory cytokines.
Taken together, our findings reveal an important new locus whereby EPO induces substantial long-term tissue protection in the host through signaling to several critical subsets of immune cells that reside in the peripheral lymphatic system.
PMCID: PMC2271128  PMID: 18382691
12.  Sinomenine reduces iNOS expression via inhibiting the T-bet IFN-γ pathway in experimental autoimmune encephalomyelitis in rats 
Journal of Biomedical Research  2012;26(6):448-455.
Sinomenine is a bioactive alkaloid isolated from the Chinese medicinal plant Sinomenium acutum. It is widely used as an immunosuppressive drug for treating rheumatic and arthritic diseases. In our previous studies, we found that sinomenine reduced cellular infiltration within the spinal cord and alleviated experimental autoimmune encephalomyelitis (EAE) in rats. In this study, we further investigated the mechanisms of sinomenine treatment in EAE rats. In EAE rats, treatment with sinomenine exerted an anti-inducible NO synthase (anti-iNOS) effect, which is related to the reductions of Th1 cytokine interferon-γ (IFN-γ) and its transcription factor, T-bet, in spinal cords. Moreover, sinomenine treatment of splenocytes stimulated with anti-CD3 antibody and recombinant rat interleukin 12 reduced the expression of T-bet and IFN-γ in vitro and also reduced the capability of supernatants of splenocyte culture to induce iNOS expression by primary astrocytes. However, sinomenine had no direct inhibitory effect on iNOS produced by astrocytes cultured with IFN-γ and tumor necrosis factor α in vitro. In conclusion, the anti-iNOS effect of sinomenine on EAE is mediated via the suppression of T-bet /IFN-γ pathway.
PMCID: PMC3597047  PMID: 23554784
sinomenine; experimental autoimmune encephalomyelitis; iNOS; T-bet; interferon-γ (IFN-γ)
13.  Effect of Honey Bee Venom on Lewis Rats with Experimental Allergic Encephalomyelitis, a Model for Multiple Sclerosis 
Multiple sclerosis (MS) is a progressive and autoimmune neurodegenerative disease of the central nervous system (CNS). This disease is recognized through symptoms like inflammation, demyelination and the destruction of neurological actions. Experimental allergic encephalomyelitis (EAE) is a widely accepted animal model for MS. EAE is created in animals by injecting the tissue of myelin basic protein (MBP), CNS, or myelin oligodendrocyte glycoprotein (MOG) along with the adjuvant. EAE and MS are similar diseases. Honey Bee venom (Apis mellifera) contains a variety of low and high molecular weight peptides and proteins, including melittin, apamin, adolapin, mast cell degranulating peptide and phospholipase A2. Bee venom (BV) could exert anti-inflammatory and antinociceptive effects on the inflammatory reactions. The guinea pig spinal cord homogenate (GPSCH) is with the Complete Freund’s Adjuvant (CFA), consisting of 1 mg/mL Mycobacterium tuberculosis. It was used for inducting EAE in Lewis rats for creating the MS model.
The hematoxylin and eosin and luxol fast blue methods were used respectively in analyses of inflammation and detection of demyelination in the central nervous system. Furthermore, the ELISA and the high performance liquid chromatography (HPLC) were used for the assessment of tumor necrosis factor alpha (TNF-α) and nitrate in rats serum. In this study, we indicated that the treatment of EAE with Bee venom decreased the symptoms of clinical disorder, pathological changes, inflammatory cell infiltration, demyelination in the central nervous system, level of serum TNF-α, and the serum nitrates in rat EAE induced through GPSCH.
PMCID: PMC3832151  PMID: 24250492
Bee venom; Multiple sclerosis; Experimental allergic encephalomyelitis; TNF-α; Serum nitrate
14.  Peptide-specific prevention of experimental allergic encephalomyelitis. Neonatal tolerance induced to the dominant T cell determinant of myelin basic protein 
The Journal of Experimental Medicine  1989;169(5):1681-1691.
Experimental allergic encephalomyelitis (EAE) is a model of antigen- specific T cell-mediated autoimmune disease. The alpha-acetylated, NH2- terminal nine amino acids (1-9NAc) of myelin basic protein (MBP) represents the dominant T cell epitope for the induction of EAE in the B10.PL (H-2u) strain. We tolerized neonatal B10.PL mice to 1-9NAc and studied the proliferative responses to this peptide and to whole MBP. Mice exposed to 1-9NAc in the neonatal period were tolerant to subsequent challenge at the proliferative T cell level. Similarly, in the 1-9NAc-tolerant group, both the incidence and severity of 1-9NAc induced EAE were greatly reduced. The fact that we were able to tolerize mice normally responsive to MBP suggests that this self antigen is sequestered (within the central nervous system) and hence tolerance to it is not normally induced. No significant difference in disease incidence was seen in response to rat MBP between control animals and 1-9NAc-tolerized mice (50% in both groups), demonstrating the presence of at least one additional encephalitogenic determinant elsewhere on the molecule. We have successfully prevented disease induction by peptide-induced tolerization. Tolerance induction by peptides provides a new and specific strategy in the prevention of autoimmunity. However, it will be clearly necessary to fully define all epitopes potentially capable of inducing pathogenic T cells to ensure complete and effective therapy of T cell-mediated autoimmune disease.
PMCID: PMC2189310  PMID: 2469764
15.  Dose-Dependent Anti-Inflammatory and Neuroprotective Effects of an ανβ3 Integrin-Binding Peptide 
Mediators of Inflammation  2013;2013:268486.
Previous studies have shown that prevention of leukocyte infiltration by targeting integrins involved in transendothelial migration may suppress the clinical and pathological features of neuroinflammatory disease. This study was designed to investigate the effects of C16, an ανβ3 integrin-binding peptide, in an acute experimental allergic encephalomyelitis (EAE) rat model. Multiple histological and immunohistochemical staining, electron microscopy observation, ELISA assay, Western blot, and magnetic resonance imaging (MRI) were employed to assess the degree of inflammation, axonal loss, neuronal apoptosis, white matter demyelination, and extent of gliosis in the brain and spinal cord of differently treated EAE models. The results showed that C16 treatment could inhibit extensive leukocyte and macrophage accumulation and infiltration and reduce cytokine tumor necrosis factor-α (TNF-α) and interferon-γ (IFN-γ) expression levels. A significantly lower clinical score at the peak time of disease was also demonstrated in the C16 treated group. Moreover, astrogliosis, demyelination, neuronal death, and axonal loss were all alleviated in C16 treated EAE animals, which may be attributed to the improvement of microenvironment. The data suggests that C16 peptide may act as a protective agent by attenuating inflammatory progression and thus affecting the expression of some proinflammatory cytokines during neuroinflammatory disease.
PMCID: PMC3855988  PMID: 24347822
16.  Immunomodulatory activity of polysaccharides isolated from Clerodendrum splendens: Beneficial effects in experimental autoimmune encephalomyelitis 
Extracts of leaves from Clerodendrum have been used for centuries to treat a variety of medicinal problems in tropical Africa. However, little is known about the high-molecular weight active components conferring therapeutic properties to these extracts.
Polysaccharides from the leaves of Clerodendrum splendens were extracted and fractionated by ion exchange and size-exclusion chromatography. Molecular weight determination, sugar analysis, degree of methyl esterification, and other chemical characterization of the fractions were performed. Immunomodulatory activity of the fractions was evaluated by determining their ability to induce monocyte/macrophage nitric oxide (NO), cytokine production, and mitogen-activated protein kinase (MAPK) phosphorylation. Experimental autoimmune encephalomyelitis (EAE) was induced in C57BL/6 mice, and severity of EAE was monitored in mice treated with intraperitoneal (i.p.) injections of the most active polysaccharide fraction. Lymph nodes (LN) and spleen were harvested, and levels of cytokines in supernatants from LN cells and splenocytes challenged with myelin oligodendrocyte glycoprotein peptide were determined.
Fractions containing type II arabinogalactan had potent immunomodulatory activity. Specifically, the high-molecular weight sub-fraction CSP-AU1 (average of 38.5 kDa) induced NO and cytokine [interleukin (IL)-1α, -1β, -6, -10, tumor necrosis factor (TNF; designated previously as TNF-α), and granulocyte macrophage-colony stimulating factor (GM-CSF)] production by human peripheral blood mononuclear cells (PBMCs) and monocyte/macrophages. CSP-AU1-induced secretion of TNF was prevented by Toll-like receptor 4 (TLR4) antagonist LPS-RS, indicating a role for TLR4 signaling. Treatment with CSP-AU1 also induced phosphorylation of a number of MAPKs in human PBMC and activated AP-1/NF-κB. In vivo treatment of mice with CSP-AU1 and CSP-NU1 resulted in increased serum IL-6, IL-10, TNF, monocyte chemoattractant protein-1 (MCP-1), macrophage inflammatory protein (MIP)-1α/CCL3, and MIP-1β/CCL4. CSP-AU1 treatment of mice with EAE (50 mg/kg, i.p., daily, 13 days) resulted in significantly reduced disease severity in this experimental model of multiple sclerosis. Levels of IL-13, TNF, interferon (IFN)-γ, IL-17, and GM-CSF were also significantly decreased, whereas transforming growth factor (TGF)-β was increased in LN cells from CSP-AU1-treated EAE mice.
Polysaccharide CSP-AU1 is a potent natural innate immunomodulator with a broad spectrum of agonist activity in vitro and immunosupressive properties after chronic administration in vivo.
PMCID: PMC3717075  PMID: 23806004
Clerodendrum splendens; Polysaccharide; Macrophage; Cytokine; Immunomodulation
17.  Increase in Th17 and T-reg Lymphocytes and Decrease of IL22 Correlate with the Recovery Phase of Acute EAE IN Rat 
PLoS ONE  2011;6(11):e27473.
Experimental autoimmune encephalomyelitis (EAE), a well-established model of multiple sclerosis, is characterised by microglial activation and lymphocyte infiltration. Induction of EAE in Lewis rats produces an acute monophasic disease characterised by a single peak of disability followed by a spontaneous and complete recovery and a subsequent tolerance to further immunizations. In the current study we have performed a detailed analysis of the dynamics of different lymphocyte populations and cytokine profile along the induction, peak, recovery and post-recovery phases in this paradigm. MBP-injected rats were sacrificed attending exclusively to their clinical score, and the different populations of T-lymphocytes as well as the dynamics of different pro- and anti-inflammatory cytokines were analysed in the spinal cord by flow cytometry, immunohistochemistry and ELISA. Our results revealed that, during the induction and peak phases, in parallel to an increase in symptomatology, the number of CD3+ and CD4+ cells increased progressively, showing a Th1 phenotype, but unexpectedly during recovery, although clinical signs progressively decreased, the number and proportion of CD3+ and CD4+ populations remained unaltered. Interestingly, during this recovery phase, we observed a marked decrease of Th1 and an important increase in Th17 and T-reg cells. Moreover, our results indicate a specific cytokine expression profile along the EAE course characterized by no changes of IL10 and IL17 levels, decrease of IL21 on the peak, and high IL22 levels during the induction and peak phases that markedly decrease during recovery. In summary, these results revealed the existence of a specific pattern of lymphocyte infiltration and cytokine secretion along the different phases of the acute EAE model in Lewis rat that differs from those already described in chronic or relapsing-remitting mouse models, where Th17-cells were found mostly during the peak, suggesting a specific role of these lymphocytes and cytokines in the evolution of this acute EAE model.
PMCID: PMC3217052  PMID: 22110656
18.  Oral administration of the KATP channel opener diazoxide ameliorates disease progression in a murine model of multiple sclerosis 
Multiple Sclerosis (MS) is an acquired inflammatory demyelinating disorder of the central nervous system (CNS) and is the leading cause of nontraumatic disability among young adults. Activated microglial cells are important effectors of demyelination and neurodegeneration, by secreting cytokines and others neurotoxic agents. Previous studies have demonstrated that microglia expresses ATP-sensitive potassium (KATP) channels and its pharmacological activation can provide neuroprotective and anti-inflammatory effects. In this study, we have examined the effect of oral administration of KATP channel opener diazoxide on induced experimental autoimmune encephalomyelitis (EAE), a mouse model of MS.
Anti-inflammatory effects of diazoxide were studied on lipopolysaccharide (LPS) and interferon gamma (IFNγ)-activated microglial cells. EAE was induced in C57BL/6J mice by immunization with myelin oligodendrocyte glycoprotein peptide (MOG35-55). Mice were orally treated daily with diazoxide or vehicle for 15 days from the day of EAE symptom onset. Treatment starting at the same time as immunization was also assayed. Clinical signs of EAE were monitored and histological studies were performed to analyze tissue damage, demyelination, glial reactivity, axonal loss, neuronal preservation and lymphocyte infiltration.
Diazoxide inhibited in vitro nitric oxide (NO), tumor necrosis factor alpha (TNF-α) and interleukin-6 (IL-6) production and inducible nitric oxide synthase (iNOS) expression by activated microglia without affecting cyclooxygenase-2 (COX-2) expression and phagocytosis. Oral treatment of mice with diazoxide ameliorated EAE clinical signs but did not prevent disease. Histological analysis demonstrated that diazoxide elicited a significant reduction in myelin and axonal loss accompanied by a decrease in glial activation and neuronal damage. Diazoxide did not affect the number of infiltrating lymphocytes positive for CD3 and CD20 in the spinal cord.
Taken together, these results demonstrate novel actions of diazoxide as an anti-inflammatory agent, which might contribute to its beneficial effects on EAE through neuroprotection. Treatment with this widely used and well-tolerated drug may be a useful therapeutic intervention in ameliorating MS disease.
PMCID: PMC3215935  PMID: 22047130
Diazoxide; experimental autoimmune encephalomyelitis; KATP channel; microglia; multiple sclerosis; neuroprotection
19.  Mycobacterium bovis Bacille Calmette-Guérin Infection in the CNS Suppresses Experimental Autoimmune Encephalomyelitis and Th17 Responses in an IFN-gamma-independent Manner1 
Multiple sclerosis (MS) and an animal model resembling MS, experimental autoimmune encephalomyelitis (EAE), are inflammatory demyelinating diseases of the central nervous system (CNS) that are suppressed by systemic mycobacterial infection in mice and BCG vaccination in humans. Host defense responses against Mycobacterium in mice are influenced by T lymphocytes and their cytokine products, particularly IFN-γ, which plays a protective regulatory role in EAE. To analyze the counter-regulatory role of mycobacterial infection-induced IFN-γ in the CNS on the function of the pathological Th17 cells and the clinical outcome of EAE, we induced EAE in mice that were intracerebrally infected with Mycobacterium bovis bacille Calmette-Guerin (BCG). Here we demonstrate that intracerebral (i.c.) BCG infection prevented inflammatory cell recruitment to the spinal cord and suppressed the development of EAE. Concomitantly, there was a significant decrease in the frequency of MOG-specific IFN-γ-producing CD4+ T cells in the CNS. IL-17+CD4+ T cell responses were significantly suppressed in i.c. BCG-infected mice following EAE induction regardless of T cell specificity. The frequency of Foxp3+CD4+ T cells in these mice was equivalent to that of control mice. The i.c. BCG infection-induced protection of EAE and suppression of MOG-specific IL-17+CD4+ T cell responses were similar in both wild type (WT) and IFN-γ deficient mice. These data show that live BCG infection in the brain suppresses CNS autoimmunity. These findings also reveal that the regulation of Th17-mediated autoimmunity in the CNS can be independent of IFN-γ-mediated mechanisms.
PMCID: PMC2735452  PMID: 18941210
Autoimmune disease; T lymphocytes; mycobacterial infection; BCG; IL-17; IFN-γ; immunoregulation; CNS
20.  Astragaloside IV Attenuates Experimental Autoimmune Encephalomyelitis of Mice by Counteracting Oxidative Stress at Multiple Levels 
PLoS ONE  2013;8(10):e76495.
Multiple sclerosis (MS) is a chronic autoimmune neuroinflammatory disease found mostly in young adults in the western world. Oxidative stress induced neuronal apoptosis plays an important role in the pathogenesis of MS. In current study, astragaloside IV (ASI), a natural saponin molecule isolated from Astragalus membranceus, given at 20 mg/kg daily attenuated the severity of experimental autoimmune encephalomyelitis (EAE) in mice significantly. Further studies disclosed that ASI treatment inhibited the increase of ROS and pro-inflammatory cytokine levels, down-regulation of SOD and GSH-Px activities, and elevation of iNOS, p53 and phosphorylated tau in central nervous system (CNS) as well as the leakage of BBB of EAE mice. Meanwhile, the decreased ratio of Bcl-2/Bax was reversed by ASI. Moreover, ASI regulated T-cell differentiation and infiltration into CNS. In neuroblast SH-SY5Y cells, ASI dose-dependently reduced cellular ROS level and phosphorylation of tau in response to hydrogen peroxide challenge by modulation of Bcl-2/Bax ratio. ASI also inhibited activation of microglia both in vivo and in vitro. iNOS up-regulation induced by IFNγ stimulation was abolished by ASI dose-dependently in BV-2 cells. In summary, ASI prevented the severity of EAE progression possibly by counterbalancing oxidative stress and its effects via reduction of cellular ROS level, enhancement of antioxidant defense system, increase of anti-apoptotic and anti-inflammatory pathways, as well as modulation of T-cell differentiation and infiltration into CNS. The study suggested ASI may be effective for clinical therapy/prevention of MS.
PMCID: PMC3790693  PMID: 24124567
21.  Extracellular adenosine signaling induces CX3CL1 expression in the brain to promote experimental autoimmune encephalomyelitis 
Multiple sclerosis and its animal model experimental autoimmune encephalomyelitis (EAE) are debilitating neuroinflammatory diseases mediated by lymphocyte entry into the central nervous system (CNS). While it is not known what triggers lymphocyte entry into the CNS during neuroinflammation, blockade of lymphocyte migration has been shown to be effective in controlling neuroinflammatory diseases. Since we have previously shown that extracellular adenosine is a key mediator of lymphocyte migration into the CNS during EAE progression, we wanted to determine which factors are regulated by adenosine to modulate EAE development.
We performed a genetic analysis of wild type and CD73−/− (that are unable to produce extracellular adenosine and are protected from EAE development) to identify factors that are both important for EAE development and controlled by extracellular adenosine signaling.
We show that extracellular adenosine triggered lymphocyte migration into the CNS by inducing the expression of the specialized chemokine/adhesion molecule CX3CL1 at the choroid plexus. In wild type mice, CX3CL1 is upregulated in the brain on Day 10 post EAE induction, which corresponds with initial CNS lymphocyte infiltration and the acute stage of EAE. Conversely, mice that cannot synthesize extracellular adenosine (CD73−/− mice) do not upregulate CX3CL1 in the brain following EAE induction and are protected from EAE development and its associated lymphocyte infiltration. Additionally, blockade of the A2A adenosine receptor following EAE induction prevents disease development and the induction of brain CX3CL1 expression. The CX3CL1 induced during EAE is found on the choroid plexus, which is the barrier between the blood and cerebral spinal fluid in the brain and is a prime entry point into the CNS for immune cells. Furthermore, CX3CL1 expression can be induced in the brains of mice and in choroid plexus cell line following A2A adenosine receptor agonist administration. Most importantly, we show that CX3CL1 blockade protects against EAE development and inhibits lymphocyte entry into the CNS.
We conclude that extracellular adenosine is an endogenous modulator of neuroinflammation during EAE that induces CX3CL1 at the choroid plexus to trigger lymphocyte entry into the brain.
PMCID: PMC3458968  PMID: 22883932
Extracellular adenosine; CD73; A2A adenosine receptor; CX3CL1 (fractalkine); Experimental autoimmune encephalomyelitis; Multiple sclerosis; Neuroinflammation; Choroid plexus
22.  High-mobility group box 1 protein (HMGB1) neutralization ameliorates experimental autoimmune encephalomyelitis* 
Journal of autoimmunity  2013;43:32-43.
Multiple sclerosis (MS) is an autoimmune, demyelinating disease and as such, the gold standard of treatment is to selectively suppress the pathogenic autoimmune response without compromising the entire arm of the adaptive immune response. One target of this strategy lying upstream of the pathologic adaptive immune response is the local, innate immune signaling that initiates and drives autoimmunity and sterile injury. High-mobility group box 1 protein (HMGB1) is a ubiquitous nuclear protein that when released from necrotic cells, such as damaged oligodendrocytes in MS lesions, drives pro-inflammatory responses. Here we demonstrate that HMGB1 drives neuroinflammatory responses in experimental autoimmune encephalomyelitis (EAE), a murine model for MS, and that inhibition of HMGB1 signaling ameliorates disease. Specifically i.v. injection of an HMGB1 neutralizing antibody in the C57BL/6 model of chronic EAE or SJL/J model of relapsing-remitting EAE ameliorated clinical disease prophylactically or during ongoing disease, blocked T cell infiltration of the central nervous system, and inhibited systemic CD4+ T cell responses to myelin epitopes. Additionally, lymphocytes from EAE mice restimulated in vitro in the presence of recombinant HMGB1 exhibited increased proliferation and pro-inflammatory cytokine production, an effect that was blocked by anti-HMGB1 antibody. Similarly recombinant HMGB1 promoted proliferation and pro-inflammatory cytokine production of human PBMCs stimulated in vitro, and anti-HMGB1 antibody blocked this effect. These findings indicate that HMGB1 contributes to neuroinflammatory responses that drive EAE pathogenesis and that HMGB1 blockade may be a novel means to selectively disrupt the pro-inflammatory loop that drives MS autoimmunity.
PMCID: PMC3672339  PMID: 23514872
HMGB1; innate immunity; neuroinflammation; autoimmune disease; EAE; tolerance
23.  A2A adenosine receptor signaling in lymphocytes and the central nervous system regulates inflammation during experimental autoimmune encephalomyelitis‡ 
Extracellular adenosine has an important role in regulating the severity of inflammation during an immune response. While there are four adenosine receptor (AR) subtypes, the A2AAR is both highly expressed on lymphocytes and known as a prime mediator of adenosine’s anti-inflammatory effects. To define the importance of A2AAR signaling during neuroinflammatory disease progression, we utilized the experimental autoimmune encephalomyelitis (EAE) animal model for multiple sclerosis. In EAE induction experiments, A2AAR antagonist treatment protected mice from disease development and its associated CNS lymphocyte infiltration. However, A2AAR−/− mice developed a more severe acute EAE phenotype characterized by more proinflammatory lymphocytes and activated microglial/macrophages. Interestingly, very high levels of A2AAR were expressed on the choroid plexus, a well-established CNS lymphocyte entry point. To determine the contribution of A2AAR signaling in lymphocytes and the CNS during EAE, we utilized bone marrow chimeric mice. Remarkably, A2AAR−/− donor hematopoietic cells potentiated severe EAE, while lack of A2AAR expression on non-hematopoietic cells protected against disease development. While no defect in the suppressive ability of A2AAR−/− regulatory T cells was observed, A2AAR−/− lymphocytes where shown to proliferate more and produced more IFNγ following stimulation. Despite this more proinflammatory phenotype, A2AAR antagonist treatment still protected against EAE when A2AAR−/− lymphocytes were adoptively transferred to T cell deficient A2AAR+/+ mice. These results indicate that A2AAR expression on non-immune cells (likely in the CNS) is required for efficient EAE development, while A2AAR lymphocyte expression is essential for limiting the severity of the inflammatory response.
PMCID: PMC3358473  PMID: 22529293
24.  Protective effect of an elastase inhibitor in a neuromyelitis optica-like disease driven by a peptide of myelin oligodendroglial glycoprotein 
The pathology of neuromyelitis optica (NMO), in contrast to multiple sclerosis, comprises granulocyte infiltrates along extensive lengths of spinal cord, as well as optic nerve. Furthermore, IFN-β treatment worsens NMO. We recently found that experimental autoimmune encephalomyelitis (EAE) induced with Th17 cells is exacerbated by IFN-β, in contrast to disease induced with Th1 where treatment attenuated symptoms.
This study demonstrates the similarities between NMO and Th17 EAE and how neutrophils mediate pathology in Th17 disease.
Levels of blood biomarkers in NMO were assessed by Luminex and ELISA. Effects of IFN-β on neutrophils were assessed by culture assays and immunofluorescence. EAE was induced by transfer of myelin-specific Th1 orTh17 cells and treated with Sivelestat sodium hydrate, a neutrophil elastase inhibitor.
We show Th17 cytokines, granulocyte chemokines, type I interferon and neutrophil elastase are elevated in patients with definitive NMO. In culture, we find that IFN-β stimulates neutrophils to release neutrophil elastase. In Th17 EAE, we demonstrate neutrophilic infiltration in the optic nerve and spinal cord which was not present in Th1 EAE. Blockade of neutrophil elastase with Sivelestat had efficacy in Th17 EAE but not Th1 EAE.
The similarities between Th17 EAE and NMO indicate that this model represents several aspects of NMO. Neutrophils are critical in the pathologies of both Th17-EAE and NMO, and therefore blockade of neutrophil elastase is a promising target in treating NMO.
PMCID: PMC3319834  PMID: 22343184
multiple sclerosis; experimental autoimmune encephalomyelitis; Th17 cells; neutrophil elastase; neutrophil elastase inhibitor
25.  Increased osteopontin expression in dendritic cells amplifies IL-17 production by CD4+ T cells in experimental autoimmune encephalomyelitis and in multiple sclerosis 
Osteopontin (Opn) is a broadly expressed pleiotropic cytokine and has been shown to play an important role in various autoimmune diseases including multiple sclerosis (MS) and its animal model experimental autoimmune encephalomyelitis (EAE). It is reported that Opn exacerbates EAE by skewing T cell differentiation towards IFN-γ producing Th1 cells. Opn expression in dendritic cells (DCs) and its role in IL-17 induction from T cells during EAE or MS is unknown. We found that during EAE Opn expression is elevated in DCs both in the periphery and in the central nervous system. There was increased expression of Opn receptor on T cells and Opn induced IL-17 production by CD4+ T cells via the β3 integrin receptor and Opn inhibited IL-10 production via the CD44 receptor. Furthermore, anti-Opn treatment reduced clinical severity of EAE by reducing IL-17 production. Anti-Opn was also effective in reducing clinical severity of EAE when given after the appearance of clinical symptoms. Analogous to EAE, in subjects with MS we found increased expression of Opn in DCs and increased expression of the Opn receptors CD44, β3 and αv on T cells. Furthermore, Opn stimulated CD4+ T cells from MS patients produced significantly higher amounts of IL-17. Our results demonstrate a role for DC produced Opn both in EAE and MS that is linked to the production of IL-17.
PMCID: PMC2653058  PMID: 19017937
Dendritic cells; T cells; EAE/MS and Osteopontin

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