Course Of Experimental Autoimmune Neuritis Research Articles

Immune Modifying Effect of Drug Free Biodegradable Nanoparticles on Disease Course of Experimental Autoimmune Neuritis.

Guillain-Barre syndrome (GBS) is an autoimmune disease of demyelination and inflammation of peripheral nerves. Current treatments are limited to plasma exchange and intravenous immunoglobulins. Cargo-free nanoparticles (NPs) have been evaluated here for their therapeutic benefit on the disease course of experimental autoimmune neuritis (EAN), mimicking the human GBS. NPs prepared from poly-lactic co-glycolic acid (PLGA) with variable size and surface charge (i.e., 500 nm vs. 130 nm, polyvinyl alcohol (PVA) vs. sodium cholate), were intravenously administered in before- or early-onset treatment schedules in a rat EAN model. NP treatment mitigated distinctly the clinical severity of EAN as compared to the P2-peptide control group (P2) in all treatments and reduced the trafficking of inflammatory monocytes at inflammatory loci and diverted them towards the spleen. Therapeutic treatment with NPs reduced the expression of proinflammatory markers (CD68 (P2: 34.8 ± 6.6 vs. NP: 11.9 ± 2.3), IL-1β (P2: 18.3 ± 0.8 vs. NP: 5.8 ± 2.2), TNF-α (P2: 23.5 ± 3.7 vs. NP: 8.3 ± 1.7) and elevated the expression levels of anti-inflammatory markers CD163 (P2: 19.7 ± 3.0 vs. NP: 41.1 ± 6.5; all for NP-PVA of 130 nm; relative to healthy control). These results highlight the therapeutic potential of such cargo-free NPs in treating EAN, which would be easily translatable into clinical use due to their well-known low-toxicity profile.

Programmed Death Ligand 1 Plays a Neuroprotective Role in Experimental Autoimmune Neuritis by Controlling Peripheral Nervous System Inflammation of Rats.

Programmed death 1 (PD-1; CD279), a member of the CD28 family, is an inhibitory receptor on T cells and is responsible for T cell dysfunction in infectious diseases and cancers. The ligand for PD-1, programmed death ligand 1 (PD-L1; also known as B7-H1, CD274), is a member of the B7 family. The engagement of PD-1 with programmed death ligand can downregulate autoreactive T cells that participate in multiple autoimmune diseases. Experimental autoimmune neuritis (EAN) is an animal model of Guillain-Barré syndrome, and the pathogenesis of EAN is mediated principally through T cells and macrophages. In this study, we investigated the effects of PD-L1 in EAN rats. For preventative and therapeutic management, we administered PD-L1, which successfully decreased the severity of EAN; it alleviated the neurologic course of EAN, as well as inhibited the infiltration of inflammatory cells and demyelination of sciatic nerves. Our data revealed that PD-L1 treatment inhibited lymphocyte proliferation and altered T cell differentiation by inducing decreases in IFN-γ+CD4+ Th1 cells and IL-17+CD4+ Th17 cells and increases in IL-4+CD4+ Th2 cells and Foxp3+CD4+ regulatory T cells. The expression levels of p-STAT3 and Foxp3 were significantly different in PD-L1-treated groups compared with the control group. Additionally, PD-L1 regulated the expression of Foxp3 and p-STAT3 in EAN, probably by inhibiting PI3K/AKT/mTOR signaling expression. In summary, PD-L1 is a potentially useful agent for the treatment of EAN because of its anti-inflammatory and neuroprotective effects.

Forced Exercise Preconditioning Attenuates Experimental Autoimmune Neuritis by Altering Th1 Lymphocyte Composition and Egress.

A short-term exposure to moderately intense physical exercise affords a novel measure of protection against autoimmune-mediated peripheral nerve injury. Here, we investigated the mechanism by which forced exercise attenuates the development and progression of experimental autoimmune neuritis (EAN), an established animal model of Guillain–Barré syndrome. Adult male Lewis rats remained sedentary (control) or were preconditioned with forced exercise (1.2 km/day × 3 weeks) prior to P2-antigen induction of EAN. Sedentary rats developed a monophasic course of EAN beginning on postimmunization day 12.3 ± 0.2 and reaching peak severity on day 17.0 ± 0.3 (N = 12). By comparison, forced-exercise preconditioned rats exhibited a similar monophasic course but with significant (p < .05) reduction of disease severity. Analysis of popliteal lymph nodes revealed a protective effect of exercise preconditioning on leukocyte composition and egress. Compared with sedentary controls, forced exercise preconditioning promoted a sustained twofold retention of P2-antigen responsive leukocytes. The percentage distribution of pro-inflammatory (Th1) lymphocytes retained in the nodes from sedentary EAN rats (5.1 ± 0.9%) was significantly greater than that present in nodes from forced-exercise preconditioned EAN rats (2.9 ± 0.6%) or from adjuvant controls (2.0 ± 0.3%). In contrast, the percentage of anti-inflammatory (Th2) lymphocytes (7–10%) and that of cytotoxic T lymphocytes (∼20%) remained unaltered by forced exercise preconditioning. These data do not support an exercise-inducible shift in Th1:Th2 cell bias. Rather, preconditioning with forced exercise elicits a sustained attenuation of EAN severity, in part, by altering the composition and egress of autoreactive proinflammatory (Th1) lymphocytes from draining lymph nodes.

Administration of SB203580, a p38 MAPK Inhibitor, Reduced the Expression of MMP9, and Relieved Neurologic Severity in the Experimental Autoimmune Neuritis (EAN) in Rats.

Dysfunctional expression of matrix metalloproteinase-9 (MMP-9) has been found to be closely associated with the experimental autoimmune neuritis (EAN). In this study, we explored the role of p38 mitogen-activated protein kinases (p38 MAPK) in the regulation of MMP-9 in sciatic nerves of EAN rats. We analyzed the expression changes of MMP-2, MMP-3, MMP-9, and mitogen-activated protein kinases (MAP kinases) in sciatic nerves of EAN rats and investigated the effect of p38 MAPK inhibitor (SB203580) on MMP-9 expression and pathological changes in EAN. Real-time PCR and western blot analyses showed that the expression of MMP-2 exhibited no significant changes throughout the course of EAN, while MMP-3 and MMP-9 presented the relatively increased expression compared with that in control group. MAP kinases, including p38 MAPK, ERK1/2, and JNK1/2, were activated in the sciatic nerve of EAN rats, and phosphorylated p38 MAPK showed similar patterns of expression to MMP-9. The expression of MMP-9 and phosphorylated p38 MAPK in sciatic nerves were in positive correlation with disease severity. In addition, SB203580 treatment significantly reduced the mRNA and protein level of MMP-9 in sciatic nerves on the peak phase of EAN. Inhibition of p38 MAPK also relieved neurologic severity and ameliorated pathological changes in EAN. In conclusion, SB203580 may ameliorate clinical deficit and pathological changes in EAN by reducing the MMP-9 expression in sciatic nerves, which suggest that p38 MAPK inhibitor such as SB203580 could be considered as a therapeutic candidate in autoimmune neuropathies.

Chrysin attenuates experimental autoimmune neuritis by suppressing immuno-inflammatory responses

Guillain–Barré syndrome (GBS) is an acute, post-infectious, immune-mediated, demyelinating disease of peripheral nerves and nerve roots. Experimental autoimmune neuritis (EAN) is an animal model of GBS. Chrysin, which is a naturally occurring flavonoid, exhibits various biological activities. This study was designed to investigate the anti-inflammatory and neuroprotective properties of preventative and therapeutic chrysin treatment in EAN rats. For preventative treatment, chrysin was administered orally from day 1 to day 16 (50mg/kg once daily) while, for therapeutic treatment, rats received chrysin from day 7 to day 16 at the same dose once daily. Control animals received the same volume of the vehicle (phosphate-buffered saline/2% dimethylsulfoxide). Regardless of the treatment regimen, chrysin attenuated the severity and duration of the clinical course of EAN and reduced inflammatory cell infiltration and demyelination of sciatic nerves. In the sciatic nerves, the expression of inducible nitric oxide synthase, cyclooxygenase-2 and nuclear factor kappa B was reduced. Furthermore, chrysin inhibited the splenic mononuclear cell secretion of interleukin-1β, interleukin-2, interleukin-6, inteleukin-12, interferon γ and tumor necrosis factor α, and elevated the level of inteleukin-4. In summary, our data demonstrate that chrysin is a potentially useful agent for the treatment of EAN with its anti-inflammatory and neuroprotective effects.

Forced-exercise attenuates experimental autoimmune neuritis

Physical inactivity in combination with a sedentary lifestyle is strongly associated with an increased risk of development of inflammatory-mediated diseases, including autoimmune disorders. Recent studies suggest that anti-inflammatory effects of physical exercise may be of therapeutic value in some affected individuals. In this study, we determined the effects of forced-exercise (treadmill running) on the development and progression of experimental autoimmune neuritis (EAN), an established animal model of Guillain–Barré syndrome. Adult male Lewis rats were subjected to sedentary (control) or forced-exercise (1.2km per day, 5days a week) for three weeks prior to induction of EAN. P2 (53–78)-immunized sedentary control rats developed a monophasic course of EAN beginning on post-injection day 12.33±0.59 (n=18) and reaching peak severity on day 15.83±0.35 (n=18). At near peak of disease, ankle- and sciatic notch-evoked compound muscle action potential (CMAP) amplitudes in sedentary control rats were reduced (∼50%) while motor nerve conduction velocity (MNCV) was slowed (∼30%) compared with pre-induction evoked responses. In marked contrast, rats undergoing forced-exercise exhibited a significantly less severe clinical course of EAN beginning on post-injection day 12.63±0.53 (n=16) and reaching peaking severity on day 14.69±0.73 (n=16). At near peak of disease, ankle- and sciatic-notch-evoked CMAP amplitudes in forced-exercised rats were preserved while EAN-associated slowing of MNCV was modestly attenuated by exercise. Three weeks of forced-exercise reduced by 46% total plasma corticosterone content while elevating the levels of corticosteroid binding globulin. We conclude from this study that forced-exercise administered prior to and during development of EAN affords a novel measure of protection against autoimmune-associated deficits in peripheral nerve evoked responses independent of steroid-induced immune suppression.

APOE ε3 attenuates experimental autoimmune neuritis by modulating T cell, macrophage and Schwann cell functions

Human apolipoprotein E (apoE) is a 34.2 kDa glycosylated protein with three isoforms (apoE2, apoE3 and apoE4). Experimental autoimmune neuritis (EAN), an animal model for human Guillain–Barré syndrome, is an immune-mediated experimental disorder of the peripheral nervous system (PNS). Increased susceptibility to EAN in apoE deficient mice has been previously found. To elucidate the isoform-dependent effects of apoE on EAN, we used human apoE2, E3 and E4 transgenic mice (Tg) immunized with P0 peptide 180–199, as well as T cell proliferation test, macrophage and Schwann cell (SC) cultures to investigate the effects of apoE isoforms on the functions of T cells, macrophages and SCs both under naïve conditions and in EAN. Clinical signs of EAN were most severe in wild type (WT) C57BL/6 mice and apoE4 Tg mice, followed by apoE2 Tg mice and apoE3 Tg mice (WT ≈ E4 > E2 > E3, p < 0.01). At the nadir of EAN, spleen weight and lymphocyte proliferation were in line with the clinical severity of the disease. Proliferation tests of purified T cells from naive mice stimulated with phytohemagglutinin or interleukin-12 showed isoform-specific differences (WT ≈ E4 > E3 ≈ E2, p < 0.01). Macrophages from both naïve and EAN mice produced nitric oxide upon inflammatory stimulation with lipopolysaccharide, interferon-γ, polyinosinic:polycytidylic acid or combinations thereof, in an isoform-dependent manner (WT ≈ E4 > E2 > E3, p < 0.01). Generalized intervention with 1400W, a specific inducible nitric oxide synthase inhibitor, significantly suppressed the clinical course of EAN in apoE2, E3 and E4 Tg mice and in WT mice. During the recovery stage of disease, the highest expression of CD178 (FasL) on SCs was found in apoE3 Tg mice. Our data support an isoform-dependent effect of apoE on EAN. This might be due to the isoform-specific effects of apoE on functions of T cells, macrophages and SCs, which contribute to the distinct clinical courses of EAN. ApoE3 might not only inhibit the onset and suppress the clinical severity of EAN, but also enhance the termination of immune responses in the PNS.

Expression of TLR4 and TLR9 mRNA in Lewis Rats with Experimental Allergic Neuritis

Objective: Experimental autoimmune neuritis (EAN) is a T cell-mediated animal model of the Guillain-Barré syndrome characterized by inflammation and demyelination of the peripheral nervous system (PNS). The key pathogenesis of EAN is the imbalance between Th1- and Th2-type immune response. Toll-like receptor (TLR) is a receptor of the innate immune system. It can recognize the antigen by pathogen-associated molecular patterns, and activate the antigen-presenting cells, producing costimulating molecules to activate T cells. In this study, we observed the expression of mRNA of TLR4 and TLR9 in EAN rats. Methods: Male Lewis rats were immunized with the component of PNS myelin sheath protein P0 180–199 (100 µg) and Freund’s complete adjuvant (CFA). The clinical signs of rats and the pathological changes in the sciatic nerves were observed. The rats in the immunized group and the control group were sacrificed on day 7, 16, 24 and 33 after immunization. To observe the histopathological changes, sciatic nerve was embedded in paraffin, sectioned and stained with HE and Weil’s stain. The mRNA expression of TLR4 and TLR9 in spleen, sciatic nerve, peripheral blood monocytes and lymph nodes was detected by reverse transcriptase PCR dynamically. Results: The EAN group reached the peak of the clinical score at day 16 after the immunization, and the clinical manifestation obviously improved at day 33 after the immunization. The mRNA expression of TLR4 peaked at day 16, then decreased gradually, but at day 33, it was still higher than that of the CFA control group (p < 0.05); there were significant differences among the four time points (p < 0.05). The mRNA expression of TLR9 was upregulated during the entire course of EAN compared to the CFA and the control group (p < 0.05). Conclusions: TLR4 and TLR9 may play a role in the pathogenesis of EAN by taking part in the activation phage and effector phage.

B cells play a cooperative role via CD40L–CD40 interaction in T cell-mediated experimental autoimmune neuritis in Lewis rats

The expression of co-stimulatory molecules CD40 and CD40L was examined over the course of experimental autoimmune neuritis (EAN) induced in Lewis rats by immunization with bovine peripheral nerve myelin. In draining lymph nodes, highest level of CD40L expression was seen on day 7 post immunization (p.i.), i.e. before onset of clinical signs of EAN, while CD40 expression was increased on day 14 p.i., i.e. at peak of clinical disease. In contrast, both CD40 and CD40L expressing cells in sciatic nerves, a target organ of EAN, peaked on day 14 p.i., large numbers of both expressing cells were mainly detected on day 14-21 p.i. After co-culture with EAN rat B cells bearing CD40, P0 peptide 180-199-specific T cell line cells exhibited a rapid down-regulation of CD40L expression. Furthermore, EAN rats had enhanced P0 peptide 180-199-specific antibody responses on day 14 p.i., which might have contributed to their aggravated EAN and further demonstrated the role of antibodies in EAN. The results indicate that CD40L-CD40 interactions are involved in the initiation of the antigen-specific T cell responses associated with the generation and development of EAN, and may mediate autoantibody production in EAN. Evidently, B cells play a cooperative role via CD40L-CD40 interaction in T cell-mediated EAN of Lewis rats.

Immunohistochemical study of caveolin-1 in the sciatic nerves of Lewis rats with experimental autoimmune neuritis

The expression of caveolin-1 and the related molecule endothelial nitric oxide synthase (eNOS) was analyzed in the sciatic nerves of Lewis rats with experimental autoimmune neuritis (EAN). Western blot analysis showed that caveolin-1 significantly increased in the sciatic nerves with EAN upon initiation of cell infiltration during the early and peak stages (days 10 and 14 post-immunization, p.i.) and declined thereafter. The pattern of eNOS expression over the course of EAN largely matched that of caveolin-1. Immunohistochemistry showed that in EAN lesions, intense caveolin-1 immunostaining occurred in ED1-positive macrophages as well as in vessels, while the caveolin-1 immunoreaction was reduced in Schwann cells in the inflammatory lesions. Consequently, we postulated that caveolin-1 expression increased in the sciatic nerves with EAN; this possibly mediated either molecular trafficking or nitric oxide generation partly through the activation of eNOS in vascular endothelial cells, as well as in inflammatory macrophages in EAN and/or cellular apoptosis of inflammatory cells.

Enhanced expression of netrin-1 protein in the sciatic nerves of Lewis rats with experimental autoimmune neuritis: Possible role of the netrin-1/DCC binding pathway in an autoimmune PNS disorder

Netrin-1 is a chemotropic factor that plays an important role as a survival factor in the adult nervous system. To investigate whether netrin-1 is involved in autoimmune injury of the peripheral nervous system (PNS), the temporal expression of netrin-1 protein was analyzed in the sciatic nerves of Lewis rats with experimental autoimmune neuritis (EAN). Western blot analysis revealed a significant increase in the level of netrin-1 protein in the sciatic nerves of rats on days 11 to 24 post-immunization (p.i.) compared to controls; netrin-1 expression declined by day 30 p.i. Immunohistochemistry revealed that netrin-1 protein was expressed weakly in Schwann cells and vessels in the sciatic nerves of normal and CFA-immunized control rats. In the sciatic nerves of EAN-affected rats, netrin-1 immunoreactivity was increased mainly in the cell membrane and extracellular matrix of OX42-positive macrophages and S100-positive Schwann cells at the peak and recovery phases of EAN. Moreover, the putative netrin-1 receptor, deleted in colorectal cancer (DCC), was expressed mainly in axons, some macrophages, and Schwann cells in EAN-affected sciatic nerves, although the level of protein expression did not change significantly over the course of EAN. We suggest that a significant increase in netrin-1 expression contributes to host cell survival and axon regeneration to counter autoimmune injury and inflammation, which may play a role in recovery from EAN-induced paralysis.

Inhibition of Ras attenuates the course of experimental autoimmune neuritis

EAN induced in Lewis rats by immunization with peripheral bovine myelin was treated by the Ras inhibitor farnesylthiosalicylate (FTS). Treatment from day 0 with FTS (5 mg/kg intraperitoneally twice daily) attenuated peak clinical scores (mean ± S.E., 2.5 ± 0.5 compared to 4.1 ± 0.5 in saline treated controls, p = 0.018, t-test) but not recovery. Treatment from day 10 with FTS attenuated peak disability (2.5 ± 0.6, p = 0.032 compared to saline treated controls) and improved recovery (0.84 ± 0.42, untreated controls 2.4 ± 0.6, p = 0.028 by repeated measures ANOVA). Effects were confirmed by rotarod and nerve conduction studies. An inactive analogue, geranylthiosalicylate, had no clinical effect. Inhibition of Ras is of potential use in the treatment of inflammatory neuropathies.

Activation of p38 mitogen-activated protein kinase in the early and peak phases of autoimmune neuritis in rat sciatic nerves

To examine the involvement of p38 mitogen-activated protein kinase (MAPK) in autoimmune disorders of the peripheral nerve system, we analyzed the phosphorylation of p38 MAPK protein in the sciatic nerves of Lewis rats with experimental autoimmune neuritis (EAN). Western blot analysis showed that phosphorylated p38 (p-p38) MAPK protein was significantly increased in the sciatic nerves of rats in the early and peak phases of EAN, and declined gradually thereafter. Immunohistochemistry showed that p-p38 MAPK levels were increased in the infiltrating inflammatory cells, including T cells and macrophages, as well as in blood vessels and some Schwann cells in EAN-affected sciatic nerves, as compared to the sciatic nerves of controls. Some inflammatory cells and a few Schwann cells were also positive for TUNEL reaction at the peak and recovery phases of EAN. In conclusion, we postulate that the phosphorylation of p38 MAPK is involved in the elimination of infiltrating inflammatory cells during the course of EAN and may possibly modulate recovery in autoimmune disorders of the peripheral nervous system.

0688 The effect of rho-kinase inhibitor fasudil in the clinical course of EAN(experimental autoimmune neuritis)

0688 The effect of rho-kinase inhibitor fasudil in the clinical course of EAN(experimental autoimmune neuritis)

Anti-cytokine autoantibodies in experimental autoimmune neuritis in Lewis rats

Experimental autoimmune neuritis (EAN) is a CD4+ T-cell-mediated, inflammatory demyelinating disease of the peripheral nervous system (PNS) that serves as a model for Guillain–Barré syndrome (GBS) in humans. Cytokine production has been suggested to act a pathogenic role for EAN. To study the potential role of cytokines in context with cytokine autoantibodies (Aabs) in EAN, we used in situ hybridization to detect mRNA expression of interferon (IFN)-γ, tumor necrosis factor (TNF)-α, IL-4, IL-10, and transforming growth factor (TGF)-β in lymph node mononuclear cell (MNC) and in sciatic nerve sections, as well as ELISA for detection of their autoantibodies in sera and cerebrospinal fluid (CSF) over the course of EAN. Increased mRNA expression for IFN-γ and TNF-α was registered correlating with the peak of clinical signs of EAN, and high levels of mRNA expression for IL-4, IL-10, and TGF-β were associated with EAN recovery. The levels of cytokine mRNAs were generally inversely correlated to their autoantibodies in serum and CSF, whereby the CSF levels were equal to or lower than the serum levels. Autoantibodies to IFN-γ dose-dependently inhibited IFN-γ-induced MHC expression by peritoneal macrophages proving a neutralizing biological effect of these autoantibodies. Our data demonstrate the existence of the anti-cytokine autoantibodies in the sera and CSF of rats with EAN; however, the role of anti-cytokine autoantibodies in the disease process of EAN remains to be resolved.

Maturation and Release of Interleukin-1β by Lipopolysaccharide-primed Mouse Schwann Cells Require the Stimulation of P2X7 Receptors

The P2X7 receptor, mainly expressed by immune cells, is a ionotropic receptor activated by high concentration of extracellular ATP. It is involved in several processes relevant to immunomodulation and inflammation. Among these processes, the production of extracellular interleukin-1beta (IL-1beta), a pro-inflammatory cytokine, plays a major role in the activation of the cytokine network. We have investigated the role of P2X7 receptor and of an associated calcium-activated potassium conductance (BK channels) in IL-1beta maturation and releasing processes by Schwann cells. Lipopolysaccharide-primed Schwann cells synthesized large amounts of pro-IL-1beta but did not release detectable amounts of pro or mature IL-1beta. ATP on its own had no effect on the synthesis of pro-IL-1beta, but a co-treatment with lipopolysaccharide and ATP led to the maturation and the release of IL-1beta by Schwann cells. Both mechanisms were blocked by oxidized ATP. IL-1beta-converting enzyme (ICE), the caspase responsible for the maturation of pro-IL-1beta in IL-1beta, was activated by P2X7 receptor stimulation. The specific inhibition of ICE by the caspase inhibitor Ac-Tyr-Val-Ala-Asp-aldehyde blocked the maturation of IL-1beta. In searching for a link between the P2X7 receptor and the activation of ICE, we found that enhancing potassium efflux from Schwann cells upregulated the production of IL-1beta, while strongly reducing potassium efflux led to opposite effects. Blocking BK channels actually modulated IL-1beta release. Taken together, these results show that P2X7 receptor stimulation and associated BK channels, through the activation of ICE, leads to the maturation and the release of IL-1beta by immune-challenged Schwann cells.

Efficacy of leukaemia inhibitory factor in experimental autoimmune neuritis

Objective: To test the efficacy of leukaemia inhibitory factor (LIF) in experimental autoimmune neuritis (EAN), an animal model of Guillain–Barre Syndrome. Background: Growth factors might be promising treatment in human inflammatory neuropathy. LIF is a cytokine, which exerts neurotrophic and myotrophic properties. Materials and methods: EAN was induced in 24 Lewis rats by the injection of peripheral bovine myelin. At the onset of the disease they were injected in pairs on a random basis either with placebo or murine LIF 10 µg/kg. The injections continued until day 25 when they were killed. The clinical course of the disease and histological pattern of the cauda equina were examined. Results: The severity of the disease was slightly less in the LIF treated group and these animals also had a more rapid recovery. The main pathological features in the cauda equina of both groups were demyelination and remyelination with varying degrees of axonal degeneration, but there were no significant differences in the pathological changes between the groups. Conclusion: The administration of LIF showed a slight, but not significant, improvement in the clinical course of EAN, but no significant evidence on nerve histology. Further investigation of the therapeutic role of LIF would be worthwhile in view of its favourable effects in nerve regeneration.

P0 glycoprotein peptides 56–71 and 180–199 dose-dependently induce acute and chronic experimental autoimmune neuritis in Lewis rats associated with epitope spreading

Two synthetic peripheral nerve myelin P0 protein peptides, an immunodominant (amino acids 180–199) and a cryptic (amino acids 56–71) one, induced an acute or chronic course of experimental autoimmune neuritis (EAN) in Lewis rats, when given at low dose (50–100 μg/rat) or high dose (250 μg/rat), respectively. Corresponding to the different clinical course, pathological changes and immune responses were found: (1) Onset of clinical signs of P0 peptide 56–71 (P0 56–71) induced EAN was 1–3 days later than in P0 peptide 180–199 (P0 180–199) induced EAN at all immunizing doses, whereas the peak of the disease occurred at a similar time point post immunization (p.i.), i.e. at days 14–16 p.i. in P0 56–71 induced EAN and at day 16 p.i. in P0 180–199 induced EAN. (2) Intramolecular epitope spreading as assessed by delayed type hypersensitivity response occurred in P0 56–71 induced EAN at both low and high antigen doses and in P0 180–199 induced EAN at high antigen dose (250 μg/rat) only. (3) P0 180–199 stimulated higher levels of interferon-γ production in P0 180–199 induced EAN than in P0 56–71 induced EAN and vice versa. (4) Histopathologic evaluation revealed a similar grade of mononuclear cell infiltration in the sciatic nerves of both types of EAN, but more severe demyelination was found in P0 180–199 induced EAN compared to P0 56–71 induced EAN. The results support the hypothesis that high dose autoantigen immunization induces extensive determinant spreading and chronic course of autoimmune diseases.

Augmented expression of daintain/allograft inflammatory factor-1 is associated with clinical disease: dynamics of daintain/allograft inflammatory factor-1 expression in spleen, peripheral nerves and sera during experimental autoimmune neuritis.

Experimental autoimmune neuritis (EAN) is an animal model of Guillain-Barré syndrome, characterized by inflammation and demyelination of the peripheral nervous system (PNS). Daintain/allograft inflammatory factor-1 (daintain/AIF-1) is a novel interferon-gamma-inducible protein expressed by macrophages during organ specific autoimmune diseases. To study the involvement of daintain/AIF-1 in EAN we induced EAN in Lewis rats by immunizing with bovine PNS myelin (BPM) and complete Freund's adjuvant (CFA). The expression of daintain/AIF-1 was examined in the spleen, peripheral nerves and sera during the course of EAN by immunohistochemistry and radioimunoassay (RIA). The expression of daintain/AIF-1 in the spleen and in the sciatic nerves peaked at the preclinical stage (day 7 post immunization (p.i.)) and at the height (day 15 p.i.) of clinical EAN, consistent with a disease promoting role for daintain/AIF-1. Daintain/AIF-1 expressing cells represented a subset of ED1+ or CD11b/c+ mononuclear cells. A significant increase of daintain/AIF-1-like immunoreactivity in sera occurred at the preclinical stage of EAN. Taken together, these data indicate that daintain/AIF-1 may play a proinflammatory role in the pathogenesis of EAN.

Dynamics of production of MIP-1α, MCP-1 and MIP-2 and potential role of neutralization of these chemokines in the regulation of immune responses during experimental autoimmune neuritis in Lewis rats

Experimental autoimmune neuritis (EAN) is an inflammatory autoimmune demyelinating disease of the peripheral nervous system (PNS) and represents an animal model of Guillain–Barré syndrome (GBS), which is a major inflammatory demyelinating disease of the PNS in humans. In the present study, the dynamics of the expression of the chemokines macrophage inflammatory protein-1α (MIP-1α), MIP-2 and monocyte chemotactic protein-1 (MCP-1) were determined in the sciatic nerves of EAN rats. Additionally, the effect of neutralizing antibodies against MIP-1α, MIP-2 and MCP-1 on the clinical course of EAN and the chemokine expression was investigated. The maximum of MIP-1α positive cells in the sciatic nerves was seen on day 14 post immunization (p.i.) correlating with the development of severe clinical signs. Administration of an anti-MIP-1α antibody suppressed the clinical signs of EAN and inhibited inflammation and demyelination in the sciatic nerve. Peak numbers of MCP-1 positive cells in the sciatic nerves were detected on day 7 p.i. Administration of an anti-MCP-1 antibody caused a delay of onset of EAN. However, 4 of the 6 EAN rats receiving the anti-MCP-1 antibody showed the same degree of inflammatory cell infiltration and demyelination in the sciatic nerves as sham-treated EAN rats, whereas only 2 EAN rats had less inflammation and demyelination. The numbers of MIP-2 positive cells reached a maximum on day 21 p.i. Anti-MIP-2 antibody failed to suppress the clinical signs of EAN and the inflammation and demyelination in the sciatic nerves. Only administration of the anti-MIP-1α antibody resulted in a significant reduction in the number of chemokine (MIP-1α)-positive cells and ED1-positive macrophages in the sciatic nerves. The present results demonstrate that MIP-1α and MCP-1 may play a role in the immunopathogenesis of EAN, and that MIP-1α induced trafficking of inflammatory cells can be inhibited by immunoneutralization. Further elucidation of the regulation and coordination of MIP-1α and MCP-1 production may lead to new therapeutic approaches to GBS in humans.