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AbstractWhile the complement system is desired for protective immunity, antibody- and complement-mediated neuromuscular junction (NMJ) destruction, a hallmark of myasthenia gravis (MG) or experimental autoimmune MG (EAMG), is a significant concern. Evidence suggests that the binding of complement factors to the pathogenic anti-acetylcholine receptor (AChR) autoantibody induces the formation of membrane attack complexes (MAC), which ultimately lead to NMJ destruction and muscle weakness. Studies corroborating the evidence show that the complement (C3–C6)-deficient or complement inhibitor (anti-C1q, soluble CR1, anti-C6, and C5 inhibiting peptide)-treated animals are highly resistant to EAMG induction, whereas the deficiency of the naturally occurring complement inhibitors, such as the decay-accelerating factor (DAF), increases EAMG susceptibility. Notably, the complement-inhibited animals do not exhibit significant immunosuppression but only a marginal reduction in the production of certain cytokines and immunoglobulin isotypes. A preliminary clinical trial using antibody-based C5 inhibitor eculizumab has been shown to be of potential use for MG treatment. The inhibition of the classic complement pathway (CCP) alone appears to be enough to suppress EAMG, suggesting that the complement inhibitors targeting specifically the classic pathway could effectively treat MG without causing immunosuppressive and other side effects. For instance, a recent non-antibody-based therapeutic approach selectively targeting the CCP component C2 by small interfering RNA (siRNA) has proven useful in EAMG treatment. The treatment strategies developed for MG might also be beneficial for other complement-mediated autoimmune diseases.

We report the first case of an autoimmune thyroid encephalopathy presenting with multifocal motor status epilepticus. A 37-year-old female patient was admitted with multifocal motor seizures intractable to intravenous status epilepticus treatments, asymmetrical quadriparesis, truncal ataxia and continuous semi-rhythmical jerks. Pathological signal alterations were detected in both precentral cortices in MRI examination. Autoimmune thyroiditis was diagnosed after radiological examinations of the thyroid gland and thyroid function tests. Seizures promptly ceased following intravenous steroid treatment. Immunohistochemistry studies showed mild to moderate neuronal staining with the plasma and CSF samples. Remarkably, autoimmune thyroiditis may present with migrating focal motor status epilepticus. We recommend anti-thyroid antibody screening for multifocal motor status epilepticus cases of unspecified cause.

We utilized HLA transgenic mice to identify the dominant epitopes on the human (H)-AChR ? subunit. The cytoplasmic H-AChR peptide ?320-337 was the dominant T cell epitope for DQ8, DR3, and DQ8×DQ6 F1 mice. The H-AChR-immunized HLA-DQ8, DR3, DQ8×DR3 F1 and DQ8×DQ6 F1 mice developed clinical EAMG, whereas HLA-DQ6 mice were less susceptible.

To comprehend the reduced susceptibility of HLA-DQ6 transgenic mice in comparison with HLA-DQ8 mice, to experimental autoimmune myasthenia gravis (EAMG), we immunized them with acetylcholine receptor (AChR) and examined in vitro, the proliferative and cytokine responses to AChR. When immunized with AChR and examined for AChR-specific lymphocyte responses to AChR, EAMG-resistant DQ6 mice exhibited significantly reduced in vitrolymphoproliferative and cytokine responses to AChR, compared to DQ8 mice. The differences in susceptibility were not linked to a difference in peptide recognition by AChR-specific lymphocytes. AChR T cell epitope mapping showed that both DQ6 and DQ8 responded to the same epitopes, although to varying degrees. Resistance of DQ6 transgenic mice to EAMG was linked to a dramatic suppression of AChR-specific IFN-γ, IL-2 and IL-10 productions by AChR-primed lymph node cells.

Myasthenia gravis (MG) is an antibody-mediated, autoimmune neuromuscular disease. Animal models of experimental autoimmune myasthenia gravis (EAMG) can be induced in vertebrates by immunization with Torpedo californicaacetylcholine receptors (AChR) in complete Freund's adjuvant. The MHC class II genes influence the cellular and humoral immune response to AChR and are involved in the development of clinical EAMG in mice. A dominant epitope within the AChR α146–162 region activates MHC class II-restricted CD4 cells and is involved in the production of pathogenic anti-AChR antibodies by B cells. Neonatal or adult tolerance to this T-cell epitope could prevent EAMG. During an immune response to AChR in vivo,multiple TCR genes are used. The CD28–B7 and CD40L–CD40 interaction is required during the primary immune response to AChR. However, CTLA-4 blockade augmented T- and B-cell immune response to AChR and disease. Cytokines IFN-γ and IL-12 upregulate, while IFN-α downregulates, EAMG pathogenesis. However, the Th2 cytokine IL-4 fails to play a significant role in the development of antibody-mediated EAMG. Systemic or mucosal tolerance to AChR or its dominant peptide(s) has prevented EAMG in an antigen-specific manner. Antigen-specific tolerance and downregulation of pathogenic cytokines could achieve effective therapy of EAMG and probably MG.

Recombinant (r) human IL-2 was administered in vivo to improve homing and engraftment of rheumatoid arthritis (RA) patients' peripheral blood mononuclear cells (PBMC) into severe combined immunodeficient (SCID) mice. Human rIL-2 treatment resulted in augmented human Ig production and induced IgM rheumatoid factor (RF) of human origin in SCID-RA chimeras. The increment of human serum IgG in SCID-RA chimeras after IL-2 treatment ranged between 15 and 43% and for IgM between 50 and 98% during 2-8 weeks postengraftment. Human IgM-RF was detectable after 1 to 2 weeks after engraftment and persisted over a period of 10-13 weeks. No RF was produced in SCID mice engrafted with PBMC from healthy individuals with or without exogenous rIL-2 administration. Thus, human rIL-2 expanded autoreactive clones involved in the production of RF in the SCIDRA chimeras. The present study provides a novel approach to establish an in vivo SCID-RA model to study the cellular and molecular mechanisms involved in the production of RF and development of a RA-like lesion. Copyright 1995, 1999 Academic Press

To study the involvement of B cells in the immune response to acetylcholine receptor (AChR), B-cell-deficient (μ mutant) and control wild-type C57BL/6 mice were immunized with AChR and assessed for clinical and immunopathological manifestations of experimental autoimmune myasthenia gravis (EAMG). The μ mutant mice failed to generate anti-AChR antibodies and were completely resistant to the induction of EAMG. However, μ mutant mice developed clinical EAMG when antibodies to the AChR main immunogenic region were passively transferred. Further, thein vivoexpansion of lymph node cells after AChR immunization was greatly impaired in μ mutant mice. The μ mutant mice gave an effectivein vitroT cell immune response to the immunodominant pathogenic AChR α chain peptide 146–162 (α146–162) and to the whole AChR protein when tested on day 90 after immunization with AChR, whereas the response to both AChR and its α146–162 peptide was reduced when tested on day 7 after immunization. Thein vitroproduction of IFN-γ and IL-2 by AChR-specific and α146–162 peptide-specific lymphocytes was lower in μ mutant mice. The AChR immune μ mutant T cells proliferated and produced IFN-γ when AChR or α146–162 peptide was presented by wild-type irradiated AChR-primed antigen-presenting cells (APCs). This indicates that B cells are important in the processing and presentation of AChR dominant peptidein vitroduring the initial immune response to AChR. However, APCs of non-B-cell lineage are sufficient to process AChR and prime the T cells to AChR dominant T cell epitope peptides.

To induce autoimmune diseases in animals, the auto-antigen has to be emulsified in adjuvants (e.g., complete Freund's adjuvant) containing microbial products such as Mycobacterium tuberculosis. But these powerful immunoadjuvants are not without undesirable immune response to the microbial proteins and induction of adjuvant arthritis, which could interfere with the antigen specific autoimmune response to be tested. This study was performed to evaluate the requirement of microbial products in the induction of experimental autoimmune diseases, and to identify an adjuvant without unwanted immune responses. C57BL/6 mice were inoculated with Torpedo acetylcholine receptors (T-AChR) emulsified in Titermax (TM), an adjuvant containing nonionic block copolymer and no microbial products, and evaluated for experimental autoimmune myasthenia gravis (EAMG) susceptibility. Mice immunized with T-AChR in TM demonstrated characteristic myasthenic muscle weakness with electrophysiological defect, elevated serum anti-AChR antibodies, and muscle AChR loss. None of the mice that received TM alone had muscle weakness, serum anti-AChR antibodies or muscle AChR loss. The data imply that microbial products are not critical in the induction of autoimmune disease like myasthenia gravis in mice. Further, nonionic block copolymer could be an ideal adjuvant in the induction of autoimmune diseases in animals.

One of the dominant T cell epitopes in the acetylcholine receptor (AChR) α chain lies within the region 146-162 and has been implicated in the pathogenesis of experimental autoimmune myasthenia gravis (EAMG) in C57BL6 (B6) mice. To directly examine the pathogenic potential of α146-162 in EAMG, B6 mice were primed with AChR in complete Freunds adjuvant (CFA) and subsequently boosted twice with either α146-162 or a control peptide in CFA. Seventy percent of the mice boosted with α146-162 developed muscle weakness characteristic of EAMG, while none of the mice boosted with the control peptide showed any clinical signs of the disease. Thus, the data provided evidence for epitope within AChR α146-162 as one of the EAMG-inducing pathogenic epitopes in B6 mice. Copyright 1994, 1999 Academic Press

Mice bearing the H-2bhaplotype are susceptible to the development of experimental autoimmune myasthenia gravis (EAMG), induced by acetylcholine receptor (AChR) autoimmunity. One of the genes influencing EAMG susceptibility has been mapped to the Ablocus of the major histocompatibility complex, and the Aß chain has been implicated in the pathogenesis. Mice of the H-2bhaplotype, including C57BL/10 (B10), have a genomic deletion of the Eagene and therefore fail to express the E molecule on their cell surface. To test the hypothesis that failure to express the cell surface E molecule in B10 mice contributes to EAMG pathogenesis, Einfasupktransgenic B10 mice expressing the T molecule were examined. Expression of the E molecule in Einfasupktransgenic B10 mice partially prevented the development of EAMG.

A gene conversion event between Eb^b and Ab^b in the B6.C-H-2^bm12 (bm12) strain, which alters three amino acids in the C-terminal half of the first domain of Ab^b (Ile-67 → Phe; Arg-70 → Gln; Thr-71 → Lys) resulted in resistance to experimental autoimmune myasthenia gravis (EAMG) pathogenesis. To study the effect of bm12 mutation on the T-cell responses to epitopes of acetylcholine receptor (AChR)-α subunit, C57BL6 (B6) and bm12 mice were primed with Torpedo californica AChR, and the profiles of T-lymphocyte proliferation were determined with 18 synthetic overlapping peptides encompassing the entire extracellular portion of the AChR-α subunit. The proliferative responses of AChR-primed bm12 lymphocytes were markedly reduced to two (α146-162 and α182-198) of the three AChR peptides (α111-126, α146-162, and α182-198) that are immunodominant in B6 mice. Thus, the Ab residues encompassing the region 67-71 determine the immunogenicity of two of the AChR-α subunit T-cell epitopes. To test the involvement of AChR-α chain epitopes within peptide α146-162 in EAMG pathogenesis, B6 mice were neonatally tolerized with soluble peptide α146-162, and subsequently immunized with AChR in complete Freund's adjuvant. Neonatal tolerance to AChR or to peptide α146-162 reduced the incidence of clinical myasthenia gravis and suppressed serum anti-AChR antibodies. This indicates the involvement of T-cell epitopes within AChR-α subunit region α146-162 in EAMG pathogenesis. Neonatal tolerance to peptide α146-162 could have caused specific clonal deletion, and/or clonal anergy, and/or recruited suppressor cells to prevent clinical EAMG. Presumably, epitope(s) with AChR α146-162, in the context of Ab encompassing region 67-71, stimulate specific T helper cells which interact with specific B cells to produce pathogenic antibodies, the primary culprit causing the end plate lesion in patients with myasthenia gravis. Copyright 1993, 1999 Academic Press

Experimental autoimmune myasthenia gravis (EAMG) is an animal model of human myasthenia gravis (MG). In mice, EAMG is induced by immunization with Torpedo californica acetylcholine receptor (AChR) in complete Freund's adjuvant (CFA). However, the role of cytokines in the pathogenesis of EAMG is not clear. Because EAMG is an antibody-mediated disease, it is of the prevailing notion that Th2 but not Th1 cytokines play a role in the pathogenesis of this disease. To test the hypothesis that the Th1 cytokine, interferon (IFN)-γ, plays a role in the development of EAMG, we immunized IFN-γ knockout (IFN-gko) (−/−) mice and wild-type (WT) (+/+) mice of H-2b haplotype with AChR in CFA. We observed that AChR-primed lymph node cells from IFN-gko mice proliferated normally to AChR and to its dominant pathogenic α146–162 sequence when compared with these cells from the WT mice. However, the IFN-gko mice had no signs of muscle weakness and remained resistant to clinical EAMG at a time when the WT mice exhibited severe muscle weakness and some died. The resistance of IFN-gko mice was associated with greatly reduced levels of circulating anti-AChR antibody levels compared with those in the WT mice. Comparatively, immune sera from IFN-gko mice showed a dramatic reduction in mouse AChR-specific IgG1 and IgG2a antibodies. However, keyhole limpet hemocyanin (KLH)–priming of IFN-gko mice readily elicited both T cell and antibody responses, suggesting that IFN-γ regulates the humoral immune response distinctly to self (AChR) versus foreign (KLH) antigens. We conclude that IFN-γ is required for the generation of a pathogenic anti-AChR humoral immune response and for conferring susceptibility of mice to clinical EAMG.