Transfer Of Experimental Allergic Encephalomyelitis Research Articles

Transforming growth factor receptor III (Betaglycan) regulates the generation of pathogenic Th17 cells in EAE.

The transforming growth factor receptor III (TβRIII) is commonly recognized as a co-receptor that promotes the binding of TGFβ family ligands to type I and type II receptors. Within the immune system, TβRIII regulates T cell development in the thymus and is differentially expressed through activation; however, its function in mature T cells is unclear. To begin addressing this question, we developed a conditional knock-out mouse with restricted TβRIII deletion in mature T cells, necessary because genomic deletion of TβRIII results in perinatal mortality. We determined that TβRIII null mice developed more severe autoimmune central nervous neuroinflammatory disease after immunization with myelin oligodendrocyte peptide (MOG35-55) than wild-type littermates. The increase in disease severity in TβRIII null mice was associated with expanded numbers of CNS infiltrating IFNγ+ CD4+ T cells and cells that co-express both IFNγ and IL-17 (IFNγ+/IL-17+), but not IL-17 alone expressing CD4 T cells compared to Tgfbr3fl/fl wild-type controls. This led us to speculate that TβRIII may be involved in regulating conversion of encephalitogenic Th17 to Th1. To directly address this, we generated encephalitogenic Th17 and Th1 cells from wild type and TβRIII null mice for passive transfer of EAE into naïve mice. Remarkably, Th17 encephalitogenic T cells from TβRIII null induced EAE of much greater severity and earlier in onset than those from wild-type mice. The severity of EAE induced by encephalitogenic wild-type and Tgfbr3fl/fl.dLcKCre Th1 cells were similar. Moreover, in vitro restimulation of in vivo primed Tgfbr3fl/fl.dLcKCre T cells, under Th17 but not Th1 polarizing conditions, resulted in a significant increase of IFNγ+ T cells. Altogether, our data indicate that TβRIII is a coreceptor that functions as a key checkpoint in controlling the pathogenicity of autoreactive T cells in neuroinflammation probably through regulating plasticity of Th17 T cells into pathogenic Th1 cells. Importantly, this is the first demonstration that TβRIII has an intrinsic role in T cells.

Correction to: Nebulization of RNS60, a Physically-Modified Saline, Attenuates the Adoptive Transfer of Experimental Allergic Encephalomyelitis in Mice: Implications for Multiple Sclerosis Therapy

The original version of this article unfortunately contained a mistake. The Figure3, 4, 5 legends have been misplaced. The updated legends along with the figures are corrected with this erratum.

Nebulization of RNS60, a Physically-Modified Saline, Attenuates the Adoptive Transfer of Experimental Allergic Encephalomyelitis in Mice: Implications for Multiple Sclerosis Therapy.

Developing a new and effective therapeutic approach against multiple sclerosis (MS) is always an important area of research. RNS60 is a bioactive aqueous solution generated by subjecting normal saline to Taylor-Couette-Poiseuille flow under elevated oxygen pressure. Recently we have demonstrated that RNS60, administered through intraperitoneal injection, ameliorated clinical symptoms and disease progression of experimental allergic encephalomyelitis (EAE), an animal model of MS. Since the intravenous route is not preferred for treating a chronic condition, we tested if nebulization of RNS60 could attenuate the disease process of adoptively-transferred EAE in mice. Although we could not directly image RNS60 after nebulization, nebulized Alexa680 reached spleen, spinal cord and different parts of the brain. Nebulization of RNS60 starting from the acute phase attenuated clinical symptoms of relapsing-remitting EAE in female SJL/J mice. RNS60 nebulization also inhibited perivascular cuffing, maintained the integrity of blood-brain and blood-spinal cord barriers, suppressed inflammation, normalized the expression of myelin genes, and blocked demyelination in the CNS of EAE mice. On the immunomodulatory front, nebulization of RNS60 to EAE mice led to the enrichment of anti-autoimmune regulatory T cells (Tregs) and suppression of autoimmune Th17 cells. Together, these results suggest that nebulization of RNS60 may be used to control aberrant immune responses in MS and other autoimmune disorders.

Glyceryl Tribenzoate: A Flavoring Ingredient, Inhibits the Adoptive Transfer of Experimental Allergic Encephalomyelitis via TGF-β: Implications for Multiple Sclerosis Therapy.

Multiple sclerosis (MS) is the most common autoimmune demyelinating disease of the central nervous system (CNS). Here, we have explored a novel use of glyceryl tribenzoate (GTB), a flavoring ingredient, in ameliorating the disease process of experimental allergic encephalomyelitis (EAE), an animal model of MS, via TGF-β. Oral feeding of GTB suppressed clinical symptoms of adoptively-transferred relapsing-remitting (RR) EAE in recipient mice and suppressed the generation of encephalitogenic T cells in donor mice. GTB also attenuated clinical symptoms of RR-EAE in PLP-TCR transgenic mice and chronic EAE in male C57/BL6 mice. Accordingly, GTB also suppressed perivascular cuffing, preserved the integrity of blood-brain barrier and blood-spinal cord barrier, inhibited inflammation, and stopped demyelination in the CNS of EAE mice. Interestingly, GTB treatment upregulated TGF-β and enriched regulatory T cells (Tregs) in splenocytes as well as in vivo in EAE mice. Blocking TGF-β by neutralizing antibodies abrogated GTB-mediated enrichment of Tregs and protection of EAE. These results suggest that oral GTB may be considered as a possible therapy for MS patients.

T Cells derived from Peripheral lymph nodes have different activation requirements than T cells isolated from spleen.

Abstract Primary murine T cells have been used as a model for human immune function. Animal models like EAE have shown disparate activities between splenic T cells and T cells derived from peripheral lymph nodes (PLN T cells). In this study we investigated if PLN T cells had the same activation requirements as splenic T cells in a serum free environment. PLN T cells activated in did not proliferate or secrete cytokines serum free conditions either by the non-specific stimuli of anti-CD3 and anti-CD28, or by PLP139-151 specific antigen re-stimulation. Conversely, splenic T cells cultured in serum free or FBS conditions proliferated and secreted cytokines equally in both activation models. The activation of serum free PLN T cells can be rescued with the addition of serum albumin as proliferation and cytokine production in both activation models was equivalent to those cultured in FBS. The activation requirement was specific to serum albumin, as the addition of other proteins including ovalbumin and serum proteins; gamma globulin, and collagen did not result in PLN T cell activation. Furthermore full antigen specific activity was returned to the PLN T cells with the addition of serum albumin demonstrated by the ability to induce adoptive transfer of EAE. The use of FBS in culture has masked the differential activation requirement of T cells depending on where they were derived and this studied showed serum albumin is requisite for the activation of T cells derived from PLN but not from spleen.

Nebulization of RNS60 Attenuates the Adoptive Transfer of Experimental Allergic Encephalomyelitis in Mice: Implications for Multiple Sclerosis (P5.313)

Nebulization of RNS60 Attenuates the Adoptive Transfer of Experimental Allergic Encephalomyelitis in Mice: Implications for Multiple Sclerosis (P5.313)

Protection of Tregs, Suppression of Th1 and Th17 Cells, and Amelioration of Experimental Allergic Encephalomyelitis by a Physically-Modified Saline

In multiple sclerosis (MS) and other autoimmune diseases, the autoreactive T cells overcome the resistance provided by the regulatory T cells (Tregs) due to a decrease in the number of Foxp3-expressing Tregs. Therefore, upregulation and/or maintenance of Tregs during an autoimmune insult may have therapeutic efficacy in autoimmune diseases. Although several immunomodulatory drugs and molecules are available, most present significant side effects over long-term use. Here we have undertaken an innovative approach to upregulate Tregs and achieve immunomodulation. RNS60 is a 0.9% saline solution generated by subjecting normal saline to Taylor-Couette-Poiseuille (TCP) flow under elevated oxygen pressure. RNS60, but not NS (normal saline), RNS10.3 (TCP-modified saline without excess oxygen) and PNS60 (saline containing excess oxygen without TCP modification), was found to upregulate Foxp3 and enrich Tregs in MBP-primed T cells. Moreover, RNS60, but not NS, RNS10.3 and PNS60, inhibited the production of nitric oxide (NO) and the expression of iNOS in MBP-primed splenocytes. Incubation of the cells with an NO donor abrogated the RNS60-mediated upregulation of Foxp3. These results suggest that RNS60 boosts Tregs via suppression of NO production. Consistent to the suppressive activity of Tregs towards autoreactive T cells, RNS60, but not NS, RNS10.3, or PNS60, suppressed the differentiation of Th17 and Th1 cells and shifted the balance towards a Th2 response. Finally, RNS60 treatment exhibited immunomodulation and ameliorated adoptive transfer of experimental allergic encephalomyelitis, an animal model of MS, via Tregs. These results describe a novel immunomodulatory property of RNS60 and suggest its exploration for therapeutic intervention in MS and other autoimmune disorders.

Functional Blocking Monoclonal Antibodies against IL-12p40 Homodimer Inhibit Adoptive Transfer of Experimental Allergic Encephalomyelitis

IL-12p70 (p40:p35) and IL-23 (p40:p19) are bioactive cytokines and their role in experimental allergic encephalomyelitis (EAE), an animal model for multiple sclerosis, are becoming clear. On the other hand, the IL-12p40 homodimer (p40(2)) was considered as an inactive or inhibitory molecule and its functions are poorly understood. To facilitate the studies on p40(2), we have recently generated neutralizing mAb against mouse p40(2). The present study demonstrates the effectiveness of p40(2) mAb in treating the disease process of relapsing-remitting EAE in female SJL/J mice. The p40(2) mAb ameliorated clinical symptoms and disease progression of EAE in recipient mice and suppressed the generation of encephalitogenic T cells in donor mice. Histological and blood-brain barrier (BBB) and blood-spinal cord barrier (BSB) permeability studies reveal that p40(2) mAb effectively inhibited the infiltration of mononuclear cells into brain and spinal cord and improved the integrity of BBB and BSB in EAE mice. Consequently, p40(2) mAb also suppressed the expression of proinflammatory molecules, normalized the expression of myelin genes, and blocked demyelination in the CNS of EAE mice. On the other hand, recombinant mouse p40(2) increased the infiltration of mononuclear cells into the CNS, enhanced the permeability through BBB and BSB, stimulated CNS expression of proinflammatory molecules, and aggravated the disease process of EAE. Taken together, our results suggest that p40(2) participates in the pathogenesis of EAE and that neutralization of p40(2) may be beneficial in multiple sclerosis patients.

New Roles for LTA4 Hydrolase, LTB4 and BLT1 in Multiple Sclerosis and Experimental Allergic Encephalomyelitis (48.9)

Abstract 5-lipoxygenase (5-LO), leukotriene A4 hydrolase (LTA4h), leukotriene B4 (LTB4) and its high affinity receptor BLT1 are known drivers of allergy, adaptive immunity and vascular inflammation, but have unknown roles in human multiple sclerosis (MS) and experimental allergic encephalomyelitis (EAE). We show that LTA4h and BLT1 are markers of inflammatory perivascular cuffs and actively demyelinating plaques in relapsing-remitting and progressive human MS, as well as spinal cord inflammatory foci in rodent EAE. Encephalitogenic murine CD3+ T cells experienced with proteolipid protein peptide antigen (PLP139-151) migrate to LTB4 and express functional BLT1. Peripheral blood immune cell LTB4 synthetic capacity is markedly primed prior to/during onset of rat and mouse EAE and mouse spleen 5-LO/LTA4h/BLT1 mRNAs are dynamically regulated during the immune response after T cell adoptive transfer. Potent, selective LTA4h inhibitors decrease clinical score and cumulative disease burden in rat antigen-induced and murine adoptive T cell transfer EAE. LTA4h inhibitors are also synergistic in suppressing rat EAE when combined with agents having an orthogonal mode of immunomodulatory action, such as the PDE-IV inhibitor mesopram. The results define new roles for the 5-LO / LTA4h / LTB4 / BLT1 pathway in autoimmunity and disease progression in EAE. Human MS may be a therapeutic target for inhibitors of LTB4 synthesis.

TWIK-related Acid-sensitive K+ Channel 1 (TASK1) and TASK3 Critically Influence T Lymphocyte Effector Functions

Two major K(+) channels are expressed in T cells, (i) the voltage-dependent K(V)1.3 channel and (ii) the Ca(2+)-activated K(+) channel KCa 3.1 (IKCa channel). Both critically influence T cell effector functions in vitro and animal models in vivo. Here we identify and characterize TWIK-related acid-sensitive potassium channel 1 (TASK1) and TASK3 as an important third K(+) conductance on T lymphocytes. T lymphocytes constitutively express TASK1 and -3 protein. Application of semi-selective TASK blockers resulted in a significant reduction of cytokine production and cell proliferation. Interference with TASK channels on CD3(+) T cells revealed a dose-dependent reduction ( approximately 40%) of an outward current in patch clamp recordings indicative of TASK channels, a finding confirmed by computational modeling. In vivo relevance of our findings was addressed in an experimental model of multiple sclerosis, adoptive transfer experimental autoimmune encephalomyelitis. Pretreatment of myelin basic protein-specific encephalitogenic T lymphocytes with TASK modulators was associated with significant amelioration of the disease course in Lewis rats. These data introduce K(2)P channels as novel potassium conductance on T lymphocytes critically influencing T cell effector function and identify a possible molecular target for immunomodulation in T cell-mediated autoimmune disorders.

Myelin Basic Protein-primed T Cells Induce Neurotrophins in Glial Cells via α5β3 Integrin

Increasing the level of neurotrophins within the central nervous system may have therapeutic efficacy in patients with various neurological diseases. Earlier we have demonstrated that myelin basic protein (MBP)-primed T cells induce the expression of various proinflammatory molecules in glial cells via cell-to-cell contact. Here we describe that after Th2 polarization by gemfibrozil or other drugs, MBP-primed T cells induced the expression of neurotrophic molecules such as brain-derived neurotrophic factor (BDNF) and neurotrophin-3 (NT-3), but not proinflammatory molecules in microglia and astroglia via cell-to-cell contact. MBP-primed Th2 cells expressed alpha5 and beta3 integrins and functional blocking antibodies against both alpha5 and beta3 integrins inhibited the ability of MBP-primed Th2 cells to induce glial neurotrophins. On the other hand, glial cells expressed PDGF-Rbeta and neutralization of this glial receptor abrogated the ability of Th2 cells to induce neurotrophins in glia. Activation of glial cAMP response element-binding protein (CREB) by MBP-primed Th2 cell contact and inhibition of contact-mediated expression of neurotrophins by antisense knockdown of glial CREB suggest that MBP-primed Th2 cell-glia contact induces the expression of neurotrophins through glial activation of CREB. Accordingly, blocking of either alpha5beta3 integrins on T cells or PDGF-Rbeta on glial cells impaired the ability of MBP-primed Th2 cells to induce glial activation of CREB. Furthermore, we demonstrate that these MBP-primed Th2 cells entered into the central nervous system and increased the expression of neurotrophins in vivo in the brain. This study illuminates the importance of alpha5beta3 and PDGF-Rbeta in guiding the novel neurotrophic property of neuroantigen-primed T cells via activation of CREB that may be of therapeutic importance in various neurological disorders.

Sodium Benzoate, a Food Additive and a Metabolite of Cinnamon, Modifies T Cells at Multiple Steps and Inhibits Adoptive Transfer of Experimental Allergic Encephalomyelitis

Experimental allergic encephalomyelitis (EAE) is the animal model for multiple sclerosis. This study explores a novel use of sodium benzoate (NaB), a commonly used food additive and a Food and Drug Administration-approved nontoxic drug for urea cycle disorders, in treating the disease process of relapsing-remitting EAE in female SJL/J mice. NaB, administered through drinking water at physiologically tolerable doses, ameliorated clinical symptoms and disease progression of EAE in recipient mice and suppressed the generation of encephalitogenic T cells in donor mice. Histological studies reveal that NaB effectively inhibited infiltration of mononuclear cells and demyelination in the spinal cord of EAE mice. Consequently, NaB also suppressed the expression of proinflammatory molecules and normalized myelin gene expression in the CNS of EAE mice. Furthermore, we observed that NaB switched the differentiation of myelin basic protein-primed T cells from Th1 to Th2 mode, enriched regulatory T cell population, and down-regulated the expression of various contact molecules in T cells. Taken together, our results suggest that NaB modifies encephalitogenic T cells at multiple steps and that NaB may have therapeutic importance in multiple sclerosis.

Sodium phenylacetate inhibits adoptive transfer of experimental allergic encephalomyelitis in SJL/J mice at multiple steps.

Experimental allergic encephalomyelitis (EAE) is the animal model for multiple sclerosis. The present study underlines the importance of sodium phenylacetate (NaPA), a drug approved for urea cycle disorders, in inhibiting the disease process of adoptively transferred EAE in female SJL/J mice at multiple steps. Myelin basic protein (MBP)-primed T cells alone induced the expression of NO synthase (iNOS) and the activation of NF-kappaB in mouse microglial cells through cell-cell contact. However, pretreatment of MBP-primed T cells with NaPA markedly inhibited its ability to induce microglial expression of iNOS and activation of NF-kappaB. Consistently, adoptive transfer of MBP-primed T cells, but not that of NaPA-pretreated MBP-primed T cells, induced the clinical symptoms of EAE in female SJL/J mice. Furthermore, MBP-primed T cells isolated from NaPA-treated donor mice were also less efficient than MBP-primed T cells isolated from normal donor mice in inducing iNOS in microglial cells and transferring EAE to recipient mice. Interestingly, clinical symptoms of EAE were much less in mice receiving NaPA through drinking water than those without NaPA. Similar to NaPA, sodium phenylbutyrate, a chemically synthesized precursor of NaPA, also inhibited the disease process of EAE. Histological and immunocytochemical analysis showed that NaPA inhibited EAE-induced spinal cord mononuclear cell invasion and normalized iNOS, nitrotyrosine, and p65 (the RelA subunit of NF-kappaB) expression within the spinal cord. Taken together, our results raise the possibility that NaPA or sodium phenylbutyrate taken through drinking water or milk may reduce the observed neuroinflammation and disease process in multiple sclerosis patients.

Beneficial effect of the immunosuppressant LF 15-0195 on passively induced rat experimental autoimmune encephalomyelitis

EXPERIMENTAL allergic encephalomyelitis (EAE), an inflammatory autoimmune disease of the central nervous system (CNS), serves as an experimental model for multiple sclerosis. EAE is a T-cell-dependent paralytic disease that can be induced in experimental animals by injection of myelin-derived antigens emulsified in Complete Freund’s Adjuvant (CFA) or following adoptive transfer of T-cell lines or clones specific for various myelin proteins. The clinical course of EAE varies with the animal model and the protocol. In most models, acute EAE is followed by permanent remission and concomitant resistance to further disease induction or by a transient remission that is followed by subsequent relapses and remissions. A large body of evidence correlates the induction phase of EAE to a Th1-type immune response, whereas the remission phase as well as the subsequent resistance is characterized by an up-regulation of the anti-inflammatory cytokines IL-4, IL-10, and TGF. Deoxyspergualin (DSG) is a synthetic analogue of the anti-tumor antibiotic spergualin, a natural product isolated from Bacillus laterosporus. DSG is claimed to have potent immunosuppressive effects and several advantages compared to available immunosuppressive drugs. However, preclinical and clinical use of DSG is limited by its chemical and metabolic instability. Several analogues of DSG have been recently prepared through organic synthesis, such as LF 15-0195, an analogue that is more potent than DSG due to improved activity and increased metabolic resistance to oxidative metabolism in vivo. This new immunosuppressant is currently in clinical development for autoimmune diseases. The present study was undertaken to analyze the capacity of LF 15-0195 to inhibit the pathogenicity of differentiated effector T cells. We tested (i) the effect of LF 15-0195 administration in vivo on development of EAE induced by adoptive transfer of anti-myelin basic protein (MBP) pathogenic CD4 T cells and (ii) the effect of stimulation of immune lymph node cells in the presence of LF 15-0195 in vitro on their ability to transfer EAE. We showed that LF 15-0195, both in vitro and in vivo, reduced the pathogenicity of autoreactive effector anti-MBP CD4 T cells. This beneficial effect could not be explained by inhibition of the expansion of pathogenic CD4 T cells or by the inhibition of their production of effector cytokines. MATERIALS AND METHODS Animals

Curcumin inhibits experimental allergic encephalomyelitis by blocking IL-12 signaling through Janus kinase-STAT pathway in T lymphocytes.

Experimental allergic encephalomyelitis (EAE) is a CD4(+) Th1 cell-mediated inflammatory demyelinating autoimmune disease of the CNS that serves as an animal model for multiple sclerosis (MS). IL-12 is a proinflammatory cytokine that plays a crucial role in the induction of neural Ag-specific Th1 differentiation and pathogenesis of CNS demyelination in EAE and MS. Curcumin (1,7-Bis(4-hydroxy-3-methoxyphenyl)-1,6-heptadiene-3,5-dione) is a naturally occurring polyphenolic phytochemical isolated from the rhizome of the medicinal plant Curcuma longa. It has profound anti-inflammatory activity and been traditionally used to treat inflammatory disorders. In this study we have examined the effect and mechanism of action of curcumin on the pathogenesis of CNS demyelination in EAE. In vivo treatment of SJL/J mice with curcumin significantly reduced the duration and clinical severity of active immunization and adoptive transfer EAE. Curcumin inhibited EAE in association with a decrease in IL-12 production from macrophage/microglial cells and differentiation of neural Ag-specific Th1 cells. In vitro treatment of activated T cells with curcumin inhibited IL-12-induced tyrosine phosphorylation of Janus kinase 2, tyrosine kinase 2, and STAT3 and STAT4 transcription factors. The inhibition of Janus kinase-STAT pathway by curcumin resulted in a decrease in IL-12-induced T cell proliferation and Th1 differentiation. These findings highlight the fact that curcumin inhibits EAE by blocking IL-12 signaling in T cells and suggest its use in the treatment of MS and other Th1 cell-mediated inflammatory diseases.

Poster Sessions BP05: Neuroimmunology. Inhibition of adoptive transfer of EAE by nemo‐binding domain (NBD) peptides

Experimental allergic encephalomyelitis (EAE) is the animal model for Multiple Sclerosis (MS), the chronic autoimmune disease of the central nervous system (CNS). Activation of NF‐κB requires the activity of IkB kinase (IKK) complex containing (IKKa and IKKb) and the regulatory protein NEMO (NF‐κB essential modifier). Recently it has been shown that peptides corresponding to the NEMO‐binding domain (NBD) of IKKa or IKKb specifically inhibit the induction of NF‐κB activation without inhibiting the basal NF‐κB activity. The present study underlines the importance of cell‐permeable NBD peptides in inhibiting the disease process of adoptively‐transferred EAE in female SJL/J mice. Immunocytochemical analysis of spinal cords of EAE mice showed that there was marked induction of NF‐κB activation as evidenced by enhanced p65 (the RelA subunit of NF‐κB) expression compared to that of control mice. Double‐labelling analysis of p65 and cell‐specific markers showed that p65 was mainly expressed by astrocytes, microglia and infiltrating macrophages. Next we examined the effect of NBD peptides on the disease process of EAE. Interestingly, clinical symptoms of EAE were much lower in mice receiving wild type NBD peptides. In contrast, mutated NBD peptides had no effect on the clinical symptoms of EAE. Taken together, our results support the conclusion that activation of NF‐κB participates in the disease process of EAE and that inhibitors of NF‐κB activation may ameliorate the neuroinflammatory disease process in MS patients.Acknowledgements: This study was supported by NIH grants (NS39940 and AG19487.

INGESTED INTERFERON ALPHA INDUCES Mx mRNA

We have demonstrated that ingested murine interferon alpha (IFN-α) suppressed clinical relapse in chronic relapsing experimental autoimmune encephalomyelitis (CR-EAE), decreased inflammation and suppressed the adoptive transfer of EAE, and is a biological response modifier in patients with multiple sclerosis. We examined the relative levels of the Mx mRNA signal using semiquantitative reverse transcription-polymerase chain reaction analysis on splenocytes from mice and peripheral blood mononuclear cells from man after IFN-α ingestion. Both mice and man demonstrated inducible levels of Mx mRNA after ingesting IFN-α. Murine spleen T cells and CD8+T cells also demonstrated upregulation of Mx mRNA. Murine whole splenocytes demonstrated upregulation of Mx mRNA after IFN-α ingestion of 10 and 100U, but not after 0, 1000, 5000U. Ingested IFN-α acts via established pathways of type 1 IFN signalling.

Immune Invasion of the Central Nervous System Parenchyma and Experimental Allergic Encephalomyelitis, But Not Leukocyte Extravasation from Blood, Are Prevented in Macrophage-Depleted Mice

Organ-specific autoimmune diseases are characterized by infiltrates, including T lymphocytes and activated macrophages. Macrophages and secondarily activated tissue resident counterparts can both present Ag to and contribute to cytokine secretion by T lymphocytes. We have previously shown a crucial role of peripheral macrophages in experimental allergic encephalomyelitis (EAE), a Th1-mediated demyelinating disease that serves as a an animal model for multiple sclerosis (MS), by their depletion using mannosylated liposome-encapsulated dichloromethylene diphosphonate (Cl2MDP). Here we describe studies to investigate the mechanisms by which macrophages contribute to the lesion formation in EAE, by studying the effect of Cl2MDP-containing mannosylated liposomes (Cl2MDP-mnL) on adoptively transferred EAE in SJL/J mice. Adoptive transfer of EAE with myelin basic protein-reactive CD4+ T cells to SJL/J mice was abrogated by Cl2MDP-mnL treatment. CD4+ T cell and MHC II+ B220+ B cell extravasation from blood vessels and Th1 cytokine production were not inhibited. However, invasion of the central nervous system intraparenchymal tissues by lymphocytes, F4/80+, Mac-1+, and MOMA-1+ macrophages was almost completely blocked after treatment with Cl2MDP-mnL. Furthermore, in Cl2MDP-mnL-treated mice, the myelin sheaths appeared completely normal, whereas, in the control groups, marked demyelination occurred. Production of TNF-alpha and inducible nitric oxide synthase, both associated with macrophage/microglial activation, was inhibited. This intervention reveals a role for macrophages in regulating the invasion of autoreactive T cells and secondary glial recruitment that ordinarily lead to demyelinating pathology in EAE and multiple sclerosis.

Role of Th2 cytokines in adoptive transfer of EAE

Role of Th2 cytokines in adoptive transfer of EAE

Adoptively Transferred EAE in Mice Bearing the lprMutation

We have recently developed approaches for the generation of encephalitogenic T cell clones from mouse strains considered resistant to experimental allergic encephalomyelitis (EAE). By allowing for the direct use of knockout and mutant strains of mice, such clones allow for the efficient characterization of the relevance of specific gene products in the effector phase of EAE. Recent studies have suggested that Fas/FasL-mediated cell death may play a role in the pathogenesis of MS. To assess the role of Fas/FasL in EAE, we have tested the ability of wild-type C57BL/6-derived, encephalitogenic T cell clones to mediate adoptively transferred EAE in Fas-deficient C57BL/6- lprmice. We now report that mice with the lprmutation are fully susceptible to the adoptive transfer of EAE. Our results suggest that Fas/FasL-mediated cell death in the central nervous system does not play an integral role in the effector phase of acute EAE.