Therapeutic gene silencing with siRNA for IL-23 but not for IL-17 suppresses the development of experimental autoimmune encephalomyelitis in rats

Gene silencing with siRNAs is important as a therapeutic tool in autoimmune diseases. In this study, we administered siRNAs specific for cytokines that may be involved in pathogenesis of experimental autoimmune encephalomyelitis (EAE). siRNA specific for IL-23p19 (siRNA-IL-23) suppressed EAE almost completely, whereas siRNA-IL-17A did not modulate the clinical course. Flow cytometric analysis revealed that siRNA-IL-23 significantly reduced the proportion of both IFN-γ⁺IL-17^? Th1 and IFN-γ^?IL-17⁺ Th17 cells in the spinal cord. Consistent with this finding, siRNA-IL-23 treatment downregulated IL-12, IL-17 and IL-23 mRNAs. These findings indicate that IL-23, but not IL-17, play an important role in the development of EAE.     

Leukocyte infiltration into spinal cord of EAE mice is attenuated by removal of endothelial leptin signaling

Leptin, a pleiotropic adipokine, crosses the blood-brain barrier (BBB) and blood–spinal cord barrier (BSCB) from the periphery and facilitates experimental autoimmune encephalomyelitis (EAE). EAE induces dynamic changes of leptin receptors in enriched brain and spinal cord microvessels, leading to further questions about the potential roles of endothelial leptin signaling in EAE progression. In endothelial leptin receptor specific knockout (ELKO) mice, there were lower EAE behavioral scores in the early phase of the disorder, better preserved BSCB function shown by reduced uptake of sodium fluorescein and leukocyte infiltration into the spinal cord. Flow cytometry showed that the ELKO mutation decreased the number of CD3 and CD45 cells in the spinal cord, although immune cell profiles in peripheral organs were unchanged. Not only were CD4⁺ and CD8⁺ T lymphocytes reduced, there were also lower numbers of CD11b⁺Gr1⁺ granulocytes in the spinal cord of ELKO mice. In enriched microvessels from the spinal cord of the ELKO mice, the decreased expression of mRNAs for a few tight junction proteins was less pronounced in ELKO than WT mice, as was the elevation of mRNA for CCL5, CXCL9, IFN-γ, and TNF-α. Altogether, ELKO mice show reduced inflammation at the level of the BSCB, less leukocyte infiltration, and better preserved tight junction protein expression and BBB function than WT mice after EAE. Although leptin concentrations were high in ELKO mice and microvascular leptin receptors show an initial elevation before inhibition during the course of EAE, removal of leptin signaling helped to reduce disease burden. We conclude that endothelial leptin signaling exacerbates BBB dysfunction to worsen EAE.     

Respiratory infection with a bacterial pathogen attenuates CNS autoimmunity through IL-10 induction

Infection with viral or bacterial pathogens has been linked with the development of multiple sclerosis (MS), while infection with helminth parasites has been associated protection against MS and other autoimmune diseases. Here we have used a murine model of MS, experimental autoimmune encephalomyelitis (EAE), to examine the effect of infection with the respiratory pathogen Bordetella pertussis infection on development of CNS inflammation. The data demonstrate that infection of mice with B. pertussis significantly attenuates the clinical course of EAE induced by active immunization or cell transfer. This was reflected in a significant reduction in VLA-4 and LFA-1 expression on T cells and infiltration of IL-17⁺, IFN-γ⁺ and IFN-γ⁺IL-17⁺ CD4 T cells into the CNS. Infection with B. pertussis induced IL-10 production from dendritic cells in vitro and enhanced the frequency of IL-10-producing CD25^-Foxp3^+/− CD4⁺ T cells in vivo. Furthermore, the suppressive effects of B. pertussis infection on EAE were lost in IL-10^−/− mice. Our findings demonstrate that a bacterial infection of the respiratory tract can attenuate EAE by promoting production of the anti-inflammatory cytokine IL-10 that may suppress licensing of autoaggressive T cells in the lungs, thereby preventing their migration into the CNS.     

Infiltration of Th1 and Th17 cells and activation of microglia in the CNS during the course of experimental autoimmune encephalomyelitis

Experimental autoimmune encephalomyelitis (EAE) is a mouse model for multiple sclerosis, where disease is mediated by autoantigen-specific T cells. Although there is evidence linking CD4⁺ T cells that secrete IL-17, termed Th17 cells, and IFN-γ-secreting Th1 cells with the pathogenesis of EAE, the precise contribution of these T cell subtypes or their associated cytokines is still unclear. We have investigated the infiltration of CD4⁺ T cells that secrete IFN-γ, IL-17 or both cytokines into CNS during development of EAE and have examined the role of T cells in microglial activation. Our findings demonstrate that Th17 cells and CD4⁺ T cells that produce both IFN-γ and IL-17, which we have called Th1/Th17 cells, infiltrate the brain prior to the development of clinical symptoms of EAE and that this coincides with activation of CD11b⁺ microglia and local production of IL-1β, TNF-α and IL-6 in the CNS. In contrast, significant infiltration of Th1 cells was only detected after the development of clinical disease. Co-culture experiments, using mixed glia and MOG-specific T cells, revealed that T cells that secreted IFN-γ and IL-17 were potent activators of pro-inflammatory cytokines but T cells that secrete IFN-γ, but not IL-17, were less effective. In contrast both Th1 and Th1/Th17 cells enhanced MHC-class II and co-stimulatory molecule expression on microglia. Our findings suggest that T cells which secrete IL-17 or IL-17 and IFN-γ infiltrate the CNS prior to the onset of clinical symptoms of EAE, where they may mediate CNS inflammation, in part, through microglial activation.     

Male rats develop more severe experimental autoimmune encephalomyelitis than female rats: Sexual dimorphism and diergism at the spinal cord level

Compared with females, male Dark Agouti (DA) rats immunized for experimental autoimmune encephalomyelitis (EAE) with rat spinal cord homogenate in complete Freund’s adjuvant (CFA) exhibited lower incidence of the disease, but the maximal neurological deficit was greater in the animals that developed the disease. Consistently, at the peak of the disease greater number of reactivated CD4+CD134+CD45RC− T lymphocytes was retrieved from male rat spinal cord. Their microglia/macrophages were more activated and produced greater amount of prototypic proinflammatory cytokines in vitro. Additionally, oppositely to the expression of mRNAs for IL-12/p35, IL-10 and IL-27/p28, the expression of mRNA for IL-23/p19 was upregulated in male rat spinal cord mononuclear cells. Consequently, the IL-17+:IFN-γ+ cell ratio within T lymphocytes from their spinal cord was skewed towards IL-17+ cells. Within this subpopulation, the IL-17+IFN-γ+:IL-17+IL-10+ cell ratio was shifted towards IL-17+IFN-γ+ cells, which have prominent tissue damaging capacity. This was associated with an upregulated expression of mRNAs for IL-1β and IL-6, but downregulated TGF-β mRNA expression in male rat spinal cord mononuclear cells. The enhanced GM-CSF mRNA expression in these cells supported the greater pathogenicity of IL-17+ T lymphocytes infiltrating male spinal cord. In the inductive phase of the disease, contrary to the draining lymph node, in the spinal cord the frequency of CD134+ cells among CD4+ T lymphocytes and the frequency of IL-17+ cells among T lymphocytes were greater in male than in female rats. This most likely reflected an enhanced transmigration of mononuclear cells into the spinal cord (judging by the lesser spinal cord CXCL12 mRNA expression), the greater frequency of activated microglia/macrophages and the increased expression of mRNAs for Th17 polarizing cytokines in male rat spinal cord mononuclear cells. Collectively, the results showed cellular and molecular mechanisms underlying the target organ specific sexual dimorphism in the T lymphocyte-dependent immune/inflammatory response, and suggested a substantial role for the target organ in shaping the sexually dimorphic clinical outcome of EAE.     

Lymphokine mRNA expression in the spinal cords of Lewis rats with experimental autoimmune encephalomyelitis is associated with a host recruited CD45R hi/CD4 population during recovery

To evaluate CD4⁺ T cell subpopulations involved in the induction and recovery from experimental autoimmune encephalomyelitis (EAE), the CD45R phenotype and lymphokine mRNA profile was evaluated for encephalitogenic CD4⁺ T cell lines in vitro and compared to CD4^* T cells islated from the spinal cord of Lewis rats with EAE were > 90% of the myelin basic protein (MBP)-specific T cell lines and clones that adoptively transferred EAE were > 90% CD4⁺ and > 90% CD45R lo. A time course of EAE disease progression was monitored as a function of the percentage of CD45R hi/CD4⁺ T cells isolated from the spinal cords of diseased animals. The majority of CD4⁺ T cells found in the central nervous system during the early phase of passive EAE were CD45R lo (the same as the encephalitogenic lines/clones). A large increase of the CD45R hi/CD4⁺ T cells (up to 45%) was observed during the peak and recovery phases of EAE. Lymphokine mRNA production was analyzed from antigen-stimulated MBP-specific lines, and from spinal cord lymphocytes isolated from rats with EAE. The BP-specific lines produced Th1 lymphokines (IL-2, IFN-γ, and TNF-α), while the spinal cord lymphocytes produced the same Th1 lymphokines as well as IL-4 and IL-10. The CD45R hi/CD4⁺ T cells isolated from the spinal cords were larger and expressed more lymphokine RNA per cell than the CD45R lo/CD4⁺ T cells. The encephalitogenic cells (CD45R hi/CD4⁺ T detected in the spinal cords of rats with a fluorescent dye and by allelic transfers and all of the CD45R hi/CD4⁺ lymphocytes found in the spinal cells were found to be host recruited. Thus it appears that the CD45R hi/CD4⁺ lymphocytes found in the spinal cord represent a host-recruited, activated cellular infiltrate that increased in number in the recovery phase of EAE and synthesized both Th1 and Th2 lymphokines.     

β-Lapachone ameliorization of experimental autoimmune encephalomyelitis

β-Lapachone is a naturally occurring quinine, originally isolated from the bark of the lapacho tree (Tabebuia avellanedae) which is currently being evaluated in clinical trials for the treatment of cancer. In addition, recent investigations suggest its potential application for treatment of inflammatory diseases. Multiple sclerosis (MS) is an autoimmune disorder characterized by CNS inflammation and demyelination. Reactive T cells including IL-17 and IFN-γ-secreting T cells are believed to initiate MS and the associated animal model system experimental autoimmune encephalomyelitis (EAE). IL-12 family cytokines secreted by peripheral dendritic cells (DCs) and CNS microglia are capable of modulating T-cell phenotypes. The present studies demonstrated that β-lapachone selectively inhibited the expression of IL-12 family cytokines including IL-12 and IL-23 by DCs and microglia, and reduced IL-17 production by CD4⁺ T-cells indirectly through suppressing IL-23 expression by microglia. Importantly, our studies also demonstrated that β-lapachone ameliorated the development on EAE. β-Lapachone suppression of EAE was associated with decreased expression of mRNAs encoding IL-12 family cytokines, IL-23R and IL-17RA, and molecules important in Toll-like receptor signaling. Collectively, these studies suggest mechanisms by which β-lapachone suppresses EAE and suggest that β-lapachone may be effective in the treatment of inflammatory diseases such as MS.     

Endothelial IL-1R1 is a critical mediator of EAE pathogenesis

Interleukin-1 (IL-1) has been implicated in the disease progression of multiple sclerosis (MS). In the animal model of MS, experimental autoimmune encephalomyelitis (EAE), the induction of disease is significantly attenuated in mice lacking the type I IL-1 receptor (IL-1R1). In this study, we created a transgenic mouse (eIL-1R1 kd) in which IL-1R1 expression is knocked down specifically in endothelial cells. Induction of EAE in eIL-1R1 kd mice results in a decrease in incidence, severity and delayed onset of EAE. In addition, eIL-1R1 kd mice show significant decrease in VCAM-1 expression and diminished CD45⁺ and CD3⁺ infiltrating leukocytes in the spinal cord in animals challenged with EAE. Further, IL-1 and IL-23 stimulate IL-17 production by splenocytes from both wild type and the eIL-1R1 kd animals. Similarly, IL-1 and IL-23 synergistically stimulate splenocytes proliferation in these two strains of animals. After immunization with MOG_79-96, although eIL-1R1 kd mice displayed greatly reduced clinical scores, their splenocytes produced IL-17 and proliferated in response to a second MOG challenge, similar to wild type animals. These findings indicate a critical role for endothelial IL-1R1 in mediating the pathogenesis of EAE, and describe a new model that can be used to study endothelial IL-1R1.     

IL-33 blockade suppresses the development of experimental autoimmune encephalomyelitis in C57BL/6 mice

IL-33 is a recently described member of the IL-1 family that has been reported to have a pathogenic role in several inflammatory diseases. In this study, we evaluated the role of IL-33 in a murine model of multiple sclerosis, experimental autoimmune encephalomyelitis (EAE). We showed that the expression of IL-33 and its receptor, ST2, was markedly elevated in the spinal cord of mice during myelin oligodendrocyte glycoprotein (MOG)(35-55) peptide-induced EAE. Administration of a blocking anti-IL-33 antibody in mice of EAE during the induction phase significantly inhibited the onset and severity of EAE and reduced MOG(35-55)-induced IFN-γ and IL-17 production. In contrast, treatment with recombinant IL-33 worsened the disease course of EAE in association with increased induction of both IFN-γ and IL-17. Furthermore, anti-IL-33 treatment caused a remarkable decrease in expression of IL-17, IFN-γ, T-bet and RORγt, and an upregulation of IL-10 and TGF-β in the spinal cord of EAE mice. These results demonstrate that endogenous IL-33 plays a pivotal role in the pathogenesis of EAE and indicate that blockade of IL-33 has a significant protective effect against EAE.     

Tellurium Compound AS101 Ameliorates Experimental Autoimmune Encephalomyelitis by VLA-4 Inhibition and Suppression of Monocyte and T Cell Infiltration into the CNS

Multiple sclerosis (MS) is an inflammatory autoimmune disease of the central nervous system (CNS) involving demyelinating and neurodegenerative processes. Several of the major pathological CNS alterations and behavioral deficits of MS are recapitulated in the experimental autoimmune encephalitis (EAE) mouse model in which the disease process is induced by administration of myelin peptides. Development of EAE requires infiltration of inflammatory cytokine-generating monocytes and macrophages, and auto-reactive T cells, into the CNS. Very late antigen-4 (VLA-4, α4β1) is an integrin molecule that plays a role in inflammatory responses by facilitating the migration of leukocytes across the blood–brain barrier during inflammatory disease, and antibodies against VLA-4 exhibit therapeutic efficacy in mouse and monkey MS models. Here, we report that the tellurium compound AS101 (ammonium trichloro (dioxoethylene-o,o′) tellurate) ameliorates EAE by inhibiting monocyte and T cell infiltration into the CNS. CD49d is an alpha subunit of the VLA-4 (α4β1) integrin. During the peak stage of EAE, AS101 treatment effectively ameliorated the disease process by reducing the number of CD49d⁺ inflammatory monocyte/macrophage cells in the spinal cord. AS101 treatment markedly reduced the pro-inflammatory cytokine levels, while increasing anti-inflammatory cytokine levels. In contrast, AS101 treatment did not affect the peripheral populations of CD11b⁺ monocytes and macrophages. AS101 treatment reduced the infiltration of CD4⁺ and CD49⁺/VLA4 T cells. In addition, treatment of T cells from MS patients with AS101 resulted in apoptosis, while such treatment did not affect T cells from healthy donors. These results suggest that AS101 reduces accumulation of leukocytes in the CNS by inhibiting the activity of the VLA-4 integrin and provide a rationale for the potential use of Tellurium IV compounds for the treatment of MS.     

Therapeutic potential of experimental autoimmune encephalomyelitis by Fasudil,a Rho kinase inhibitor

The migration of aberrant inflammatory cells into the central nervous system plays an important role in the pathogenesis of demyelinating diseases potentially through the Rho/Rho‐kinase (Rock) pathway, but direct evidence from human and animal models remains inadequate. Here we further confirm that Fasudil, a selective Rock inhibitor, has therapeutic potential in a mouse model of myelin oligodendrocyte glycoprotein (MOG)‐induced experimental autoimmune encephalomyelitis (EAE). The results show that Fasudil decreased the development of EAE in C57BL/6 mice. Immunohistochemistry disclosed that expression of Rock‐II in the perivascular spaces and vascular endothelial cells of spleens, spinal cords, and brains was elevated in EAE and was inhibited in the Fasudil‐treated group. T‐cell proliferation specific to MOG_35–55 was markedly reduced, together with a significant down‐regulation of interleukin (IL)‐17, IL‐6, and MCP‐1. In contrast, secretion of IL‐4 was increased, and IL‐10 was slightly elevated. There were no differences in the percentages of CD4⁺CD25⁺, CD8⁺CD28^?, and CD8⁺CD122⁺ in mononuclear cells. Histological staining disclosed a marked decrease of inflammatory cells in spinal cord and brain of Fasudil‐treated mice. These results, together with previous studies showing the inhibitory effect of Fasudil on T‐cell migration, might expand its clinical application as a new therapy for multiple sclerosis by decreasing cell migration and regulating immune balance. © 2010 Wiley‐Liss, Inc.     

Aggravated experimental autoimmune encephalomyelitis in IL-15 knockout mice

IL-15 initially identified as a T proliferating cytokine has several structural and biological similarities with IL-2 and has been associated with a number of autoimmune diseases. Because of the scarcity of information available on the role of IL-15 in MS pathogenesis, we have investigated how the absence of IL-15 affected the development of experimental autoimmune encephalomyelitis, a mouse model of MS. Following immunization of IL-15^?/? and C57BL/6 mice with MOG_35–55, we observed a more severe neurological impairment in the IL-15 knockout mice than in the wild-type group. The enhanced disease severity in IL-15^?/? mice was associated with greater demyelination in the spinal cord, increased immune cell infiltration and inflammation. These events may be related to the higher CD4/CD8 ratio and the almost absent NK cell activity, congenital immune features of IL-15KO mice. Moreover, we found that the fractalkine receptor CX3CR1 was overexpressed in the spinal cord of IL-15^?/? mice, mainly localized on infiltrating CD8⁺ T cells. How these findings are contributing to the aggravated EAE development in IL-15 KO mice remain unclear and need to be further investigated.     

Myelin repair and functional recovery mediated by neural cell transplantation in a mouse model of multiple sclerosis

Cellular therapies are becoming a major focus for the treatment of demyelinating diseases such as multiple sclerosis (MS), therefore it is important to identify the most effective cell types that promote myelin repair. Several components contribute to the relative benefits of specific cell types including the overall efficacy of the cell therapy, the reproducibility of treatment, the mechanisms of action of distinct cell types and the ease of isolation and generation of therapeutic populations. A range of distinct cell populations promote functional recovery in animal models of MS including neural stem cells and mesenchymal stem cells derived from different tissues. Each of these cell populations has advantages and disadvantages and likely works through distinct mechanisms. The relevance of such mechanisms to myelin repair in the adult central nervous system is unclear since the therapeutic cells are generally derived from developing animals. Here we describe the isolation and characterization of a population of neural cells from the adult spinal cord that are characterized by the expression of the cell surface glycoprotein NG2. In functional studies, injection of adult NG2⁺ cells into mice with ongoing MOG_35–55-induced experimental autoimmune encephalomyelitis (EAE) enhanced remyelination in the CNS while the number of CD3⁺ T cells in areas of spinal cord demyelination was reduced approximately three-fold. In vivo studies indicated that in EAE, NG2⁺ cells stimulated endogenous repair while in vitro they responded to signals in areas of induced inflammation by differentiating into oligodendrocytes. These results suggested that adult NG2⁺ cells represent a useful cell population for promoting neural repair in a variety of different conditions including demyelinating diseases such as MS.     

Corticosteroid treatment of experimental autoimmune encephalomyelitis in the Lewis rat results in loss of Vβ8.2 and myelin basic protein-reactive cells from the spinal cord, with increased total T-cell apoptosis but reduced apoptosis of Vβ8.2 cells

We have studied the effects of corticosteroid treatment on the numbers of lymphocytes obtained from the spinal cords of Lewis rats with acute experimental autoimmune encephalomyelitis (EAE) induced by inoculation with myelin basic protein (MBP) and adjuvants. Flow cytometric studies showed that treatment with dexamethasone (4 mg/kg) 8–12 h prior to study on day 14 after inoculation resulted in a reduction in the numbers of CD5⁺, TCRαβ⁺ and Vβ8.2⁺ cells in the spinal cord. Limiting dilution analysis indicated that dexamethasone treatment 12 h prior to study on day 12 after inoculation reduced the frequencies of MBP-reactive and interleukin-2-responsive lymphocytes in the spinal cord to low levels, but reduced the frequency of concanavalin-A-responsive lymphocytes to a lesser extent. Using propidium iodide staining of nuclear chromatin we also studied lymphocyte apoptosis. Greater numbers of apoptotic cells were found in the cells extracted from the spinal cords of rats, examined on day 14, that had been treated 1–12 h previously with dexamethasone, than in saline-treated controls. This increased level of apoptosis was observed in the CD5⁺ and TCRαβ⁺ cell populations. At 1–4 h after dexamethasone treatment there was a reduction in the selective apoptosis of Vβ8.2⁺ cells that normally occurs during spontaneous recovery from EAE. Therefore apoptosis of Vβ38.2⁺ cells cannot explain the reduction in the numbers of Vβ8.2⁺ cells and MBP-reactive cells in the CNS after dexamethasone treatment. By 8–12 h after dexamethasone treatment the selectivity of the apoptotic process was restored. These studies suggest that a reduction in the number of T-lymphocytes in the central nervous system contributes to the beneficial effects of corticosteroids in EAE.     

Inflammatory cells, microglia and MHC class II antigen-positive cells in the spinal cord of Lewis rats with acute and chronic relapsing experimental autoimmune encephalomyelitis

Chronic relapsing experimental autoimmune encephalomyelitis (CR-EAE) was induced in Lewis rats by inoculation with guinea pig spinal cord and adjuvants and treatment with low dose cyclosporin A (CsA). Acute EAE was induced by the same method without CsA treatment. Immunocytochemistry and flow cytometry were used to assess inflammatory cells and MHC class II (Ia) antigen expression in the central nervous system of these rats. The inflammatory infiltrate was composed mainly of CD4⁺ T cells and macrophages, and αß T cells constituted about 65% of the CD2⁺ T cells. After recovery from acute EAE and during the first remission of CR-EAE, the number of T cells was significantly less than in the preceding episodes. The number of T cells was higher in the second episode of CR-EAE than in the first remission. Throughout the course of CR-EAE, the majority of the CD2⁺ T cells were CD45RC⁻. The ratio of IL-2R⁺ cells to CD2⁺ cells ranged from 10.5 to 24.0%. The ratio of CD4⁺ T cells to B cells was lower in the later episodes of CR-EAE than in the first episode. Ia antigen was expressed on filtrating round cells at all stages of CR-EAE and on microglial cells (identified by dendritic morphology) with increasing intensity throughout the course of CR-EAE. With flow cytometry, the number of Ia⁺ cells obtained from the spinal cord rose throughout the course of CR-EAE. The number of FSC^lowOX1^low cells, which we consider represent microglia, also increased during the course of CR-EAE.     

Anti-inflammatory effect of erythropoietin therapy on experimental autoimmune encephalomyelitis

Previous studies report that erythropoietin (EPO) has a neuroprotective role in some neurodegenerative diseases, but the mechanisms are not completely elucidated. The aim of this study was to investigate whether EPO exerts neuroprotective role in experimental autoimmune encephalomyelitis (EAE) via the routes of anti-inflammation. We established an EAE mice model treated intraperitoneally with EPO at the dose of 5,000 IU/kg on schedule, and recorded the clinical score and weight fluctuation. The infiltration of inflammatory cells in the spinal cord of EAE mice was observed with hemotoxylin and eosin (HE) staining, and the levels of IL-10, IFN-γ, IL-17, and MHC-II in central nervous system (CNS)-infiltrating cells and peripheral mononuclear cells were detected by flow cytometry or ELISA. EPO therapy ameliorates clinical signs of EAE mice, inhibits the body weight loss, and decreases the infiltration of inflammatory cells in spinal cords. IL-17 and IFN-γ are reduced, while IL-10 is not increased significantly, in both CNS-infiltrating cells and peripheral mononuclear cells of EPO-treated EAE mice, as compared with EAE control group. EPO also reduces the expression of MHC-II on peripheral antigen presentation cells. Our results indicate that EPO exerts a beneficial role in EAE by inhibiting the levels of IL-17 and IFN-γ in peripheral splenic cells and CNS-infiltrating cells.     

Antibodies to the RNA binding protein heterogeneous nuclear ribonucleoprotein A1 contribute to neuronal cell loss in an animal model of multiple sclerosis

Neurodegeneration, including loss of neurons and axons, is a feature of progressive forms of multiple sclerosis (MS). The mechanisms underlying neurodegeneration are mostly unknown. Research implicates autoimmunity to nonmyelin self-antigens as important contributors to disease pathogenesis. Data from our lab implicate autoimmunity to the RNA binding protein (RBP) heterogeneous nuclear ribonucleoprotein A1 (hnRNP A1) as a possible mechanism of neurodegeneration in MS. MS patients make antibodies to hnRNP A1, which have been shown to lead to neuronal dysfunction in vitro. Using an animal model of MS, experimental autoimmune encephalomyelitis (EAE), we show here that injection of anti-hnRNP A1 antibodies, in contrast to control antibodies, resulted in worsened disease and increased neurodegeneration. We found a reduction of NeuN⁺ neuronal cell bodies in areas of the ventral gray matter of the spinal cord where anti-hnRNP A1 antibodies localized. Neurons displayed increased levels of hnRNP A1 nucleocytoplasmic mislocalization and stress granule formation, both markers of neuronal injury. Anti-hnRNP A1 antibodies were found to surround neuronal cell bodies and interact with CD68⁺ immune cells via Fc receptors. Additionally, anti-hnRNP A1 antibodies were found within neuronal cell bodies including those of the ventral spinocerebellar tract (VSCT), a tract previously shown to undergo neurodegeneration in anti-hnRNP A1 antibody injected EAE mice. Finally, both immune cells and neurons showed increased levels of inducible nitric oxide synthase, another indicator of cell damage. These findings suggest that autoimmunity to RBPs, such as hnRNP A1, play a role in neurodegeneration in EAE with important implications for the pathogenesis of MS.     

The proximal peripheral nervous system is a major site of demyelination in experimental autoimmune encephalomyelitis induced in the Lewis rat by a myelin basic protein-specific T cell clone

Experimental autoimmune encephalomyelitis (EAE) was induced in the Lewis rat by the passive transfer of a cytotoxic CD4⁺ T cell clone specific for the 72–89 peptide of guinea-pig myelin basic protein (MBP). Histological studies on rats with neurological signs showed that inflammation was present in the proximal peripheral nervous system (PNS), namely the spinal roots, as well as in the central nervous system (CNS). The main sites of demyelination were the spinal roots in the PNS, and the spinal cord root entry and exit zones in the CNS. The major involvement of the proximal PNS in autoimmune disease directed at MBP is in marked contrast to EAE induced by immunisation with myelin proteolipid protein, where the inflammation and demyelination are restricted to the CNS. These findings may have implications for the human inflammatory demyelinating diseases including multiple sclerosis, in which MBP is a putative target antigen.