Pathogenesis Of Experimental Autoimmune Neuritis Research Articles

Antibiotics-Induced Intestinal Immunomodulation Attenuates Experimental Autoimmune Neuritis (EAN)

BackgroundThe composition of gut microbiota plays a pivotal role in priming the immune system and thus impacts autoimmune diseases. Data on the effects of gut bacteria eradication via systemic antibiotics on immune neuropathies are currently lacking. This study therefore assessed the effects of antibiotics-induced gut microbiota alterations on the severity of experimental autoimmune neuritis (EAN), a rat model of Guillain-Barré Syndrome (GBS). Myelin P0 peptide 180–199 (P0 180–199)-induced EAN severity was compared between adult Lewis rats (12 weeks old) that received drinking water with or without antibiotics (colistin, metronidazole, vancomycin) and healthy rats, beginning antibiotics treatment immediately after immunization (day 0), and continuing treatment for 14 consecutive days. Neuropathy severity was assessed via a modified clinical score, and then related to gut microbiota alterations observed after fecal 16S rRNA gene sequencing at baseline and after EAN induction. Effectors of gut mucosal and endoneurial immunity were assessed via immunostaining. EAN rats showed increased gut mucosal permeability alongside increased mucosal CD8+ T cells compared to healthy controls. Antibiotics treatment alleviated clinical EAN severity and reduced endoneurial T cell infiltration, decreased gut mucosal CD8+ T cells and increased gut bacteria that may be associated with anti-inflammatory mechanisms, like Lactobacillus or Parasutterella. Our findings point out a relation between gut mucosal immunity and the pathogenesis of EAN, and indicate that antibiotics-induced intestinal immunomodulation might be a therapeutic approach to alleviate autoimmunity in immune neuropathies. Further studies are warranted to evaluate the clinical transferability of these findings to patients with GBS.Graphical

Siponimod ameliorates experimental autoimmune neuritis

BackgroundGuillain–Barré syndrome (GBS) and chronic inflammatory demyelinating polyneuropathy (CIDP) are human autoimmune peripheral neuropathy. Besides humoral immunity, cellular immunity is also believed to contribute to these pathologies, especially CIDP. Sphingosine-1-phosphate receptor 1 (S1PR1) regulates the maturation, migration, and trafficking of lymphocytes. As of date, the therapeutic effect of sphingosine-1-phosphate receptor (S1PR) agonists on patients with GBS or CIDP remains unclear.MethodsTo evaluate the effect of siponimod, an agonist of S1PR1 and S1PR5, on experimental autoimmune neuritis (EAN), an animal model of autoimmune peripheral neuropathy, was used. Lewis rats were immunized with 125 μg of synthetic peptide from bovine P2 protein. Rats in the siponimod group were orally administered 1.0 mg/kg siponimod and those in the EAN group were administrated the vehicle on days 5–27 post-immunization (p.i.) daily. The symptom severity was recorded daily. The changes in the expression of cytokines and transcription factors in the lymph nodes and cauda equina (CE) which correlate with the pathogenesis of EAN and recovery of injured nerve were measured using reverse transcription quantitative PCR. Histological study of CE was also performed.ResultsFlaccid paralysis developed on day 11 p.i. in both groups. Siponimod relieved the symptom severity and decreased the expression of interferon-gamma and IL-10 mRNAs in lymph nodes and CE compared with that in the EAN group. The expression of Jun proto-oncogene (c-Jun) mRNA increased from the peak to the recovery phase and that of Sonic hedgehog signaling molecule (Shh) and Glial cell line-derived neurotrophic factor (Gdnf) increased prior to increase in c-Jun with no difference observed between the two groups. Histologically, siponimod also reduced demyelinating lesions and inflammatory cell invasion in CE.ConclusionsSiponimod has a potential to ameliorate EAN. Shh and Gdnf, as well as C-Jun played a significant role during the recovery of injured nerves.

Enhanced glycolysis contributes to the pathogenesis of experimental autoimmune neuritis

BackgroundWith the recognition of the key roles of cellular metabolism in immunity, targeting metabolic pathway becomes a new strategy for autoimmune disease treatment. Guillain-Barré syndrome (GBS) is an acute immune-mediated inflammatory demyelinating disease of the peripheral nervous system, characterized by inflammatory cell infiltration. These inflammatory cells, including activated macrophages, Th1 cells, and Th17 cells, generally undergo metabolic reprogramming and rely mainly on glycolysis to exert functions. This study aimed to explore whether enhanced glycolysis contributed to the pathogenesis of experimental autoimmune neuritis (EAN), a classic model of GBS.MethodsPreventive and therapeutic treatments with glycolysis inhibitor, 2-deoxy-d-glucose (2-DG), were applied to EAN rats. The effects of treatments were determined by clinical scoring, weighting, and tissue examination. Flow cytometry and ELISA were used to evaluate T cell differentiation, autoantibody level, and macrophage functions in vivo and in vitro.ResultsGlycolysis inhibition with 2-DG not only inhibited the initiation, but also prevented the progression of EAN, evidenced by the improved clinical scores, weight loss, inflammatory cell infiltration, and demyelination of sciatic nerves. 2-DG inhibited the differentiation of Th1, Th17, and Tfh cells but enhanced Treg cell development, accompanied with reduced autoantibody secretion. Further experiments in vitro proved glycolysis inhibition decreased the nitric oxide production and phagocytosis of macrophages and suppressed the maturation of dendritic cells (DC).ConclusionThe effects of glycolysis inhibition on both innate and adaptive immune responses and the alleviation of animal clinical symptoms indicated that enhanced glycolysis contributed to the pathogenesis of EAN. Glycolysis inhibition may be a new therapy for GBS.

Inhibition of triggering receptor expressed on myeloid cells-1 ameliorates experimental autoimmune neuritis.

Experimental autoimmune neuritis (EAN) is a cluster of differentiation 4+ T helper 1 cell-mediated inflammatory demyelinating disease of the peripheral nervous system and serves as a useful animal model for Guillain-Barré syndrome. Triggering receptor expressed on myeloid cells-1 (TREM-1) is an important receptor involved in sepsis and the innate inflammatory response. Linear plasmid 17 (LP 17) peptide is a competitive antagonist of TREM-1. To investigate the role of TREM-1 in EAN, 64 male Lewis rats were randomly divided into four groups: Normal saline, complete Freund's adjuvant, EAN and LP 17. The present study assessed the mRNA expression levels of TREM-1, tumor necrosis factor-α and interleukin-1β in sciatic nerves and peripheral blood mononuclear cells. The results demonstrated that inhibiting TREM-1 by administering LP 17 ameliorated symptoms and reduced inflammation in EAN rats. The present study concluded that TREM-1 may be involved in the pathogenesis of EAN, and that inhibition of TREM-1 may ameliorate EAN.

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.

Autophagy is involved in the pathogenesis of experimental autoimmune neuritis in rats.

Recent studies have shown that autophagy is involved in peripheral nervous system disease. However, the role of autophagy in the pathogenesis of experimental autoimmune neuritis (EAN) remains unclear. Therefore, EAN was induced by a subcutaneous injection into both hind footpads of synthetic neuritogenic P2(57-81) peptide in male Lewis rats. The clinical evaluation was completed using a 10-point scale method. The histological alteration of sciatic nerves was analyzed by hematoxylin and eosin and luxol fast blue staining. The ultrastructure of sciatic nerves was analyzed by transmission electron microscopy. Expressions of beclin-1 and microtubule-associated protein light chain-3 (LC3) and p62/SQSTM1 were determined by western blot. 3-Methyladenine, the inhibitor of autophagy, was used in this research. Results showed that the clinical scores were significantly increased from day 6 to day 16 after immunization compared with the control group. Compared with the control group, the number of inflammatory cells and the histological score of sciatic nerves were significantly increased, expressions of beclin-1 and LC3-II and the ratio of LC3-II/LC3-I in the sciatic nerve were significantly increased, and the expression of p62 was significantly decreased in the EAN model group. Considerable double-membrane autophagosomes in axons and myelin sheaths of sciatic nerves were observed and the number of autophagosomes in axons and myelin sheaths of sciatic nerves in the EAN model group was obviously increased compared with the control group. 3-Methyladenine ameliorated the neurologic severity of EAN. Our results suggest that autophagy activity in nerve tissue of EAN rats is increased, which may be associated with the pathogenesis of EAN.

Mitigated Tregs and augmented Th17 cells and cytokines are associated with severity of experimental autoimmune neuritis

Experimental autoimmune neuritis (EAN), an animal model of human Guillain–Barré syndrome, has long been considered as a T helper (Th) 1 cell-mediated autoimmune disorder. However, deficiency of IFN-gamma, a signature Th1 cytokine, aggravated EAN, with features of elevated production of IL-17A, despite an alleviated systemic Th1 immune response. We hypothesized that Th17 cells and their cytokines might play a pathogenic role in EAN. To further clarify the roles of these Th and regulatory T cell (Treg) cytokines in the pathogenesis of EAN and their interrelationship, we investigated the expression of Th1/Th2/Th17/Treg cytokines in EAN in the present study.

Augmented Th17 cells and IL-17A are associated with severity of experimental autoimmune neuritis

Experimental autoimmune neuritis (EAN), an animal model of human Guillain–Barré syndrome, has been considered as a T helper (Th) 1 cell-mediated autoimmune disorder. However, deficiency of IFN-γ, a signature Th1 cytokine, aggravated EAN, with features of elevated production of IL-17A, despite an alleviated systemic Th1 immune response. We hypothesized that Th17 cells and their cytokines might play a pathogenic role in EAN. To clarify the roles of these Th and regulatory T cell (Treg) cytokines in the pathogenesis of EAN and their interrelationship, we investigated the expression of Th1/Th2/Th17/Treg cytokines in EAN.

Mitigated Tregs and augmented Th17 cells and cytokines are associated with severity of experimental autoimmune neuritis.

Experimental autoimmune neuritis (EAN), an animal model of human Guillain-Barré syndrome, has long been considered as a T helper (Th) 1 cell-mediated autoimmune disorder. However, deficiency of IFN-γ, a signature Th1 cytokine, aggravated EAN, with features of elevated production of IL-17A, despite an alleviated systemic Th1 immune response. We hypothesized that Th17 cells and their cytokines might play a pathogenic role in EAN. To further clarify the roles of these Th and regulatory T cell (Treg) cytokines in the pathogenesis of EAN and their interrelationship, we investigated the expression of Th1/Th2/Th17/Treg cytokines in EAN in this study. We found that the levels of Th17 cells and IL-17A in cauda equina (CE)-infiltrating cells and splenic mononuclear cells (MNCs) as well as in serum paralleled the disease evolution, which increased progressively during the initiation stage and reached higher value at the peak of EAN. The same pattern was also noticed for the expression of IL-22. The diverse expression profiles of FoxP3, IL-17 receptors A and C were seen in CE-infiltrating cells and splenic MNCs in EAN. These findings indicate a major pro-inflammatory role of Th17 cells and IL-17A in the pathogenesis of EAN. Therapeutic interventions may be focused upon inhibiting Th17 cells and their cytokines in the early phase of EAN, so as to delay and suppress clinical signs of the disease, which has relevance for future studies on pathogenesis and treatment of GBS in humans.

Inactivation of TLR9 by a Suppressive Oligodeoxynucleotides Can Ameliorate the Clinical Signs of EAN

Susceptible-strain animals immunized with P2 peptide could generate the disease of experimental autoimmune neuritis (EAN) with inflammation and demyelination of peripheral nerve. A myriad of transcription factors and inflammatory cytokines have been found to participate in this process; however, the roles of toll-like receptors (TLRs) are poorly understood in EAN. The aim of this study is to explore the role of TLR9 in the pathogenesis of EAN. The EAN was induced in Lewis rat by immunization with P253–78 and complete Freund’s adjuvant. CpG oligodeoxynucleotides (ODN) (cODN), a suppressive ODN (sODN) and a control non-specific ODN (nODN) were respectively administered to explore the role of TLR9 in EAN both in vivo and vitro. Following immunization up to the peak phase of EAN, EAN rats inoculated with sODN had remarkably better clinical score of EAN and expressed a significantly inhibited TLR9 signaling pathway. Our study suggests that TLR9 may be involved in the pathogenesis of EAN.

TNF-α receptor 1 deficiency reduces antigen-presenting capacity of Schwann cells and ameliorates experimental autoimmune neuritis in mice

Tumor necrosis factor-α (TNF-α) is a pleiotropic pro-inflammatory cytokine with potentially neurodestructive effects and plays a pivotal role in autoimmune demyelinating disease. To address the role of TNF-α in the pathogenesis of experimental autoimmune neuritis (EAN), the current study investigated the antigen-presenting capacity of Schwann cells (SCs) in EAN induced by P0 protein peptide 106–125 in TNF-α recepter 1 deficient (TNFR1 −/−) mice. The antigen-presenting capacity of SCs was assessed by the expression of MHC class II (MHCII), CD40, CD80 and CD86 molecules on activated SCs as well as by induction of T cell proliferation in co-cultures of P0 protein peptide 106–125 specific T cells with activated SCs. In addition, the expression of inducible nitric oxide synthase (iNOS) was measured in activated SCs by flow cytometry. TNFR1 −/− EAN mice developed significantly delayed and reduced clinical signs of EAN compared to wild type EAN mice. In parallel, the expression of MHCII, CD80 and iNOS on SCs were decreased in TNFR1 −/− mice compared to wild type mice. Likewise, proliferation of P0 protein peptide 106–125 specific T cells simulated by activated SCs of TNFR1 −/− EAN mice was lower than that of wild type EAN mice. Our data suggest that TNF-α may exert pro-inflammatory effects in EAN via TNFR1 by up-regulating the antigen-presenting function and iNOS production of SCs.

Aggravation of experimental autoimmune neuritis in TNF-α receptor 1 deficient mice

The role of tumor necrosis factor (TNF)-α and its receptors in the pathogenesis of experimental autoimmune neuritis (EAN) induced by P0 peptide 180–199 in TNFR1 (p55) deficient (TNFR1−/−) mice was investigated. Compared to wild type EAN mice, TNFR1−/− EAN mice developed significantly more severe clinical signs, in parallel with enhanced numbers of inflammatory infiltrating cells in peripheral nerves and splenic P0-reactive T cell proliferation, as well as increased obviously MHC class II and CCR3 expression on the macrophages in the cauda equina. Our data indicated that TNF-α might have anti-inflammatory effect preventing the development of EAN in this mouse model.

IL-18 deficiency inhibits both Th1 and Th2 cytokine production but not the clinical symptoms in experimental autoimmune neuritis

IL-18 deficient (IL-18 −/−) mice were used to investigate the role of IL-18 in the pathogenesis of experimental autoimmune neuritis (EAN) which was induced by immunization of the mice with P0 protein peptide 180–199. The clinical course was not different between IL-18 −/− and wild-type mice. The splenic mononuclear cell (MNC) proliferation was also similar in both animal groups. However, the percentages of IFN-γ, IL-10 and IL-12 positive cells were decreased among infiltrating MNC of cauda equine in IL-18 −/− mice. This indicates that IL-18 deficiency inhibits the production of both Th1 and Th2 cytokines in the target organ of mice with EAN.

The critical role of IL-12p40 in initiating, enhancing, and perpetuating pathogenic events in murine experimental autoimmune neuritis.

Interleukin 12 (IL-12) is a proinflammatory cytokine with important immunoregulatory activities and is critical in determining the differentiation and generation of Th1 cells. For the present study, we investigated the role of endogenous IL-12 in the pathogenesis of experimental autoimmune neuritis (EAN), which is a CD4+ T-cell mediated autoimmune inflammatory disease of the peripheral nervous system. EAN is used as an animal model for Guillain-Barré syndrome of humans. Here, EAN was established in IL-12 p40 deficient mutant (IL-12-/-) C57BL/6 mice by immunization with P0 peptide 180-199, a purified component of peripheral nerve myelin, and Freund's complete adjuvant. In these IL-12-/- mice the onset of clinical disease was delayed, and the incidence and severity of EAN were significantly reduced compared to that in wild-type mice.The former group's clinical manifestations were associated with less P0-peptide 180-199 induced secretion of interferon-gamma (IFN-gamma) by splenocytes in vitro and low production of anti-P0-peptide 180-199 IgG2b antibodies in serum. Fewer IFN-gamma and TNF-alpha producing cells, but more cells secreting IL-4, were found in sciatic nerve sections from IL-12-/- mice, consistent with impaired Th1 functions and response. However, the IL-12 deficiency appeared not to affect P0 peptide 180-199-specific T-cell proliferation. These results indicate that IL-12 has a major role in the initiation, enhancement and perpetuation of pathogenic events in EAN by promoting a Th1 cell-mediated immune response and suppressing the Th2 response. This information augments consideration of IL-12 as a therapeutic target in Guillain-Barré syndrome and other T-cell-mediated autoimmune diseases.

INCREASED EXPRESSION OF PHOSPHOLIPASE D1 IN THE SCIATIC NERVE OF RATS WITH EXPERIMENTAL AUTOIMMUNE NEURITIS

Phospholipase D1 (PLD1) expression in the sciatic nerve was studied in induced experimental autoimmune neuritis (EAN) in Lewis rats. PLD1 immunoreactivity was seen in some Schwann cells in the sciatic nerves of normal rats. In parallel with the progression of EAN, PLD1-positive Schwann cells significantly increased in number and showed intense immunoreactivity. PLD1 was also detected in some ED1+ macrophages in EAN lesions. These results suggest that PLD1 in macrophages and Schwann cells plays an important role in the activation of these cells in the pathogenesis of EAN, an animal model of human peripheral demyelinating disease.

Dominance of autoreactive T cell-mediated delayed-type hypersensitivity or antibody-mediated demyelination results in distinct forms of experimental autoimmune neuritis in the Lewis rat.

The role of anti-myelin antibodies in the pathogenesis of experimental autoimmune neuritis (EAN) induced in the Lewis rat by immunization with peripheral nerve myelin has been assessed. Passive transfer with lymph node cells (LNC) or purified serum immunoglobulin from rats with EAN was employed to directly measure the contribution of B cells and anti-myelin antibodies to demyelination and disease. Lewis rats with EAN transferred by LNC or purified serum immunoglobulin from EAN donors in conjunction with a low dose of P2-specific CD4+ T cells demonstrated profound histopathological and neurophysiological evidence of demyelination during disease. In contrast, the classical adoptive transfer model of EAN in the Lewis rat induced by the injection of P2-specific CD4+ T cells was characterized by histopathological and neurophysiological evidence of axonal dysfunction and degeneration with limited demyelination. These findings demonstrate that the synergistic action of T cells and anti-myelin antibodies mediating demyelination or purely T cell mediated axonal dysfunction and degeneration are distinct pathways by which a specific autoimmune response in the peripheral nervous system can cause neurological disease.

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.

The role of the very late antigen-4 and its counterligand vascular cell adhesion molecule-1 in the pathogenesis of experimental autoimmune neuritis of the Lewis rat.

We have investigated the functional role of very late antigen-4 [VLA-4 (alpha4/beta1) integrin] and vascular cell adhesion molecule-1 (VCAM-1) in experimental autoimmune neuritis (EAN), an animal model of the Guillain-Barré syndrome, using neutralizing monoclonal antibodies (mAbs) as probes. Disease was induced by intravenous adoptive transfer of P2 specific T cells (AT-EAN), or by immunization with bovine myelin (active EAN). Preventive treatment with 500 microg anti-VLA-4 mAb (TA-2) or with an isotype control antibody was given in AT-EAN 4 h before cell transfer and at day 3. Intravenous injection of 500 microg anti-VCAM-1 mAb (5F10) or a corresponding isotype control was given in AT-EAN 4 h before disease induction, and at days 2, 4 and 6. Preventive treatment of active EAN with mAb to VLA-4 or VCAM-1 was performed at days 5, 9 and 13. On immunohistochemical examination, VCAM-1 in sciatic nerve was found to be upregulated at early stages of EAN (days 3-5 after T-cell transfer), whilst no expression was noted in healthy controls. In both EAN models, blockade of VLA-4 markedly attenuated disease severity. Blockade of VCAM-1 also significantly ameliorated the disease course and diminished T-cell infiltration in sciatic nerve, but to a lesser degree. These experiments demonstrate the critical role of VLA-4 in the pathogenesis of EAN and show that upregulation of VCAM-1 expression contributes, at least in part, to the progression of the disease in the early stages. Future studies are needed to assess the potential contribution of other VLA-4 ligands.

Cytokine production and the pathogenesis of experimental autoimmune neuritis and Guillain–Barré syndrome

Acute inflammatory demyelinating polyradiculoneuropathy (Guillain–Barré syndrome, GBS) and its animal model experimental autoimmune neuritis (EAN) are prototypes of T cell-mediated autoimmune diseases affecting the peripheral nervous system (PNS). Perivascular accumulation of macrophages and T lymphocytes in the PNS, and high levels systemically of PNS myelin antigen–reactive T cells are characteristic features of both diseases, thereby suggesting a pathogenic role for immunoregulatory cytokines. Here we summarise recent studies that have clearly documented that Th1/Th2/Th3 cytokines are differently upregulated during various clinical phases of EAN and GBS. The observations indicate that the role of cytokines in immune regulation and autoimmune disease is more complex than a simple Th1–Th2 dichotomy would suggest. New treatments may be searched for that counteract this complex cytokine imbalance. Treatments with antibodies that selectively target certain pro-inflammatory cytokines, as well as with immunomodulatory preparations that promote cytokines that beneficially influence the disease course should be in focus of future therapeutic trials.

Matrix metalloproteinase expression during experimental autoimmune neuritis.

Experimental autoimmune neuritis (EAN) is an animal model of Guillain-Barré syndrome. We have shown recently that BB-1101, a broad-spectrum matrix metalloproteinase (MMP) inhibitor, prevents development of EAN when given from the day of immunization and, more important clinically, reduces disease severity when given from symptom onset. This suggests the involvement of MMP activity in the pathogenesis of EAN. However, the exact function and expression patterns of MMPs in acute inflammation of the PNS have not been investigated. MMP-like enzymes are also involved in the processing of tumour necrosis factor-alpha (TNF-alpha), which has been implicated previously in the pathology associated with EAN. In the present study we investigated the profile of MMP and TNF-alpha expression and their localization in sciatic nerve tissue during EAN, using a semiquantitative competitive reverse transcriptase-coupled polymerase chain reaction and immunohistochemistry. In the normal rat PNS, four of the 10 MMPs studied were constitutively expressed and four MMPs were differentially regulated during EAN. Expression of TNF-alpha was elevated at peak disease severity and localized to Schwann cells, macrophages and endoneurial blood vessels. Expression levels of 92 kDa gelatinase and stromelysin-1 were significantly increased early in the development of EAN and continued to rise, peaking at day 15 coincident with maximum disease severity. Schwann cells and endothelial cells were the main cellular source of these enzymes. Prominent infiltration of inflammatory cells into the sciatic nerve was concordant with a significant increase in the expression levels of matrilysin and macrophage metalloelastase. Both matrilysin and macrophage metalloelastase were detected in invading macrophages, T lymphocytes and resident Schwann cells. The selective upregulation of specific MMPs during EAN and their varied cellular localization suggests that MMPs play a multifactorial role in the aetiology of EAN. Activity of MMPs could participate in the disruption of the blood-nerve barrier, breakdown of the myelin sheath, the release of TNF-alpha, and facilitate leukocyte invasion into the PNS. These observations highlight MMPs as potential targets for therapeutic intervention in acute peripheral neuropathies, such as Guillain-Barré syndrome.