Decreased expression of miR-29 family associated with autoimmune myasthenia gravis.

BACKGROUND: Myasthenia gravis (MG) is a rare autoimmune disease mainly mediated by autoantibodies against the acetylcholine receptor (AChR) at the neuromuscular junction. The thymus is the effector organ, and its removal alleviates the symptoms of the disease. In the early-onset form of MG, the thymus displays functional and morphological abnormalities such as B cell infiltration leading to follicular hyperplasia, and the production of AChR antibodies. Type-I interferon (IFN-I), especially IFN-ß, is the orchestrator of thymic changes observed in MG. As Dicer and miR-29 subtypes play a role in modulating the IFN-I signalization in mouse thymus, we investigated their expression in MG thymus. METHODS: The expression of DICER and miR-29 subtypes were thoroughly investigated by RT-PCR in human control and MG thymuses, and in thymic epithelial cells (TECs). Using miR-29a/b-1-deficient mice, with lower miR-29a/b-1 expression, we investigated their susceptibility to experimental autoimmune MG (EAMG) as compared to wild-type mice. RESULTS: DICER mRNA and all miR-29 subtypes were down-regulated in the thymus of MG patients and DICER expression was correlated with the lower expression of miR-29a-3p. A decreased expression of miR-29 subtypes was similarly observed in MG TECs; a decrease also induced in TECs upon IFN-ß treatment. We demonstrated that miR-29a/b-1-deficient mice were more susceptible to EAMG without higher levels of anti-AChR IgG subtypes. In the thymus, if no B cell infiltration was observed, an increased expression of Ifn-ß associated with Baff expression and the differentiation of Th17 cells associated with increased expression of Il-6, Il-17a and Il-21 and decreased Tgf-ß1 mRNA were demonstrated in miR-29a/b-1-deficient EAMG mice. CONCLUSIONS: It is not clear if the decreased expression of miR-29 subtypes in human MG is a consequence or a causative factor of thymic inflammation. However, our results from the EAMG mouse model indicated that a reduction in miR-29a/b1 may contribute to the pathophysiological process involved in MG by favoring the increased expression of IFN-ß and the emergence of pro-inflammatory Th17 cells.

MicroRNA-653 Inhibits Thymocyte Proliferation and Induces Thymocyte Apoptosis in Mice with Autoimmune Myasthenia Gravis by Downregulating TRIM9.

OBJECTIVES: Myasthenia gravis (MG) is an organ-specific autoimmune neuromuscular disorder that occurs as a result of the impairment in neuromuscular junction and autoantibody attack on the postsynaptic receptors. Increasing evidence suggests that microRNAs (miRs) might be involved in the development of MG. Therefore, the present study aimed to investigate the regulatory function of miR-653 on MG and its relationship with tripartite motif 9 (TRIM9). METHODS: The thymic tissues obtained from MG patients with thymic hyperplasia were prepared for establishing an MG mouse model in BALB/c mice. Afterwards, the miR-653 and TRIM9 expressions were determined in thymic tissues. A dual-luciferase reporter assay was carried out to validate whether miR-653 directly targets TRIM9. Finally, the thymocytes were exposed to mimics or inhibitors of miR-653, or siRNA against TRIM9 with the use of MTT assays and flow cytometry for the verification of the gain or loss function of miR-653 and TRIM9 on viability, cell cycle progression, and apoptosis of thymocytes. RESULTS: There was a decrease in thymocyte miR-653 and an increase in TRIM9 in thymic tissues of MG mice. miR-653 was found to negatively regulate TRIM9. Overexpression of miR-653 or depletion of TRIM9 resulted in the inhibition of cell viability, suppression of cell cycle progression, and induction of apoptosis rate in thymocytes. CONCLUSION: The findings from the present study provided evidence that miR-653 impairs proliferation and promotes apoptosis of thymocytes of MG mice by suppressing TRIM9, indicating that miR-653 could be used as potential therapeutic target in the treatment of autoimmune MG.

NFAT1 Regulates Systemic Autoimmunity through the Modulation of a Dendritic Cell Property.

The transcription factor NFAT1 plays a pivotal role in the homeostasis of T lymphocytes. However, its functional importance in non-CD4+ T cells, especially in systemic immune disorders, is largely unknown. In this study, we report that NFAT1 regulates dendritic cell (DC) tolerance and suppresses systemic autoimmunity using the experimental autoimmune myasthenia gravis (EAMG) as a model. Myasthenia gravis and EAMG are T cell-dependent, Ab-mediated autoimmune disorders in which the acetylcholine receptor is the major autoantigen. NFAT1-knockout mice showed higher susceptibility to EAMG development with enhanced Th1/Th17 cell responses. NFAT1 deficiency led to a phenotypic alteration of DCs that show hyperactivation of NF-κB-mediated signaling pathways and enhanced binding of NF-κB (p50) to the promoters of IL-6 and IL-12. As a result, NFAT1-knockout DCs produced much higher levels of proinflammatory cytokines such as IL-1ß, IL-6, IL-12, and TNF-α, which preferentially induce Th1/Th17 cell differentiation. Our data suggest that NFAT1 may limit the hyperactivation of the NF-κB-mediated proinflammatory response in DCs and suppress autoimmunity by serving as a key regulator of DC tolerance.

Silencing of Dok-7 in Adult Rat Muscle Increases Susceptibility to Passive Transfer Myasthenia Gravis.

Myasthenia gravis (MG) is an autoimmune disease mediated by autoantibodies that target proteins at the neuromuscular junction, primarily the acetylcholine receptor (AChR) and the muscle-specific kinase. Because downstream of kinase 7 (Dok-7) is essential for the full activation of muscle-specific kinase and consequently for dense clustering of AChRs, we hypothesized that reduced levels of Dok-7 increase the susceptibility to passive transfer MG. To test this hypothesis, Dok-7 expression was reduced by transfecting shRNA-coding plasmids into the tibialis anterior muscle of adult rats by in vivo electroporation. Subclinical MG was subsequently induced with a low dose of anti-AChR monoclonal antibody 35. Neuromuscular transmission was significantly impaired in Dok-7-siRNA-electroporated legs compared with the contralateral control legs, which correlated with a reduction of AChR protein levels at the neuromuscular junction (approximately 25%) in Dok-7-siRNA-electroporated muscles, compared with contralateral control muscles. These results suggest that a reduced expression of Dok-7 may play a role in the susceptibility to passive transfer MG, by rendering AChR clusters less resistant to the autoantibody attack.

DBC1 is a suppressor of B cell activation by negatively regulating alternative NF-κB transcriptional activity.

CD40 and BAFFR signaling play important roles in B cell proliferation and Ig production. In this study, we found that B cells from mice with deletion of Dbc1 gene (Dbc1(-/-)) show elevated proliferation, and IgG1 and IgA production upon in vitro CD40 and BAFF, but not BCR and LPS stimulation, indicating that DBC1 inhibits CD40/BAFF-mediated B cell activation in a cell-intrinsic manner. Microarray analysis and chromatin immunoprecipitation experiments reveal that DBC1 inhibits B cell function by selectively suppressing the transcriptional activity of alternative NF-κB members RelB and p52 upon CD40 stimulation. As a result, when immunized with nitrophenylated-keyhole limpet hemocyanin, Dbc1(-/-) mice produce significantly increased levels of germinal center B cells, plasma cells, and Ag-specific Ig. Finally, loss of DBC1 in mice leads to higher susceptibility to experimental autoimmune myasthenia gravis. Our study identifies DBC1 as a novel regulator of B cell activation by suppressing the alternative NF-κB pathway.

Silencing miR-146a influences B cells and ameliorates experimental autoimmune myasthenia gravis.

MicroRNAs have been shown to be important regulators of immune homeostasis as patients with aberrant microRNA expression appeared to be more susceptible to autoimmune diseases. We recently found that miR-146a was up-regulated in activated B cells in response to rat acetylcholine receptor (AChR) α-subunit 97-116 peptide, and this up-regulation was significantly attenuated by AntagomiR-146a. Our data also demonstrated that silencing miR-146a with its inhibitor AntagomiR-146a effectively ameliorated clinical myasthenic symptoms in mice with ongoing experimental autoimmune myasthenia gravis. Furthermore, multiple defects were observed after miR-146a was knocked down in B cells, including decreased anti-R97-116 antibody production and class switching, reduced numbers of plasma cells, memory B cells and B-1 cells, and weakened activation of B cells. Previously, miR-146a has been identified as a nuclear factor-κB-dependent gene and predicted to base pair with the tumour necrosis factor receptor-associated factor 6 (TRAF6) and interleukin-1 receptor-associated kinase 1 (IRAK1) genes to regulate the immune response. However, our study proved that miR-146a inhibition had no effect on the expression of TRAF6 and IRAK1 in B cells. This result suggests that the function of miR-146a in B cells does not involve these two target molecules. We conclude that silencing miR-146a exerts its therapeutic effects by influencing the B-cell functions that contribute to the autoimmune pathogenesis of myasthenia gravis.

Drosophila Nesprin-1 controls glutamate receptor density at neuromuscular junctions.

Nesprin-1 is a core component of a protein complex connecting nuclei to cytoskeleton termed LINC (linker of nucleoskeleton and cytoskeleton). Nesprin-1 is anchored to the nuclear envelope by its C-terminal KASH domain, the disruption of which has been associated with neuronal and neuromuscular pathologies, including autosomal recessive cerebellar ataxia and Emery-Dreifuss muscular dystrophy. Here, we describe a new and unexpected role of Drosophila Nesprin-1, Msp-300, in neuromuscular junction. We show that larvae carrying a deletion of Msp-300 KASH domain (Msp-300 (∆KASH) ) present a locomotion defect suggestive of a myasthenia, and demonstrate the importance of muscle Msp-300 for this phenotype, using tissue-specific RNAi knock-down. We show that Msp-300 (∆KASH) mutants display abnormal neurotransmission at the larval neuromuscular junction, as well as an imbalance in postsynaptic glutamate receptor composition with a decreased percentage of GluRIIA-containing receptors. We could rescue Msp-300 (∆KASH) locomotion phenotypes by GluRIIA overexpression, suggesting that the locomotion impairment associated with the KASH domain deletion is due to a reduction in junctional GluRIIA. In summary, we found that Msp-300 controls GluRIIA density at the neuromuscular junction. Our results suggest that Drosophila is a valuable model for further deciphering how Nesprin-1 and LINC disruption may lead to neuronal and neuromuscular pathologies.

RNA interference targeting Bcl-6 ameliorates experimental autoimmune myasthenia gravis in mice.

Follicular helper T (Tfh) cells are dedicated to providing help to B cells and are strongly associated with antibody-mediated autoimmune disease. B cell lymphoma 6 (Bcl-6) is a key transcription factor of Tfh cells, and IL-21 is known to be a critical cytokine produced by Tfh cells. We silenced Bcl-6 gene expression using RNA interference (RNAi) delivered by a lentiviral vector, to evaluate the therapeutic role of Bcl-6 short hairpin RNAs (shRNAs) in experimental autoimmune myasthenia gravis (EAMG). Our data demonstrate that CD4(+)CXCR5(+)PD-1(+) Tfh cells, Bcl-6 and IL-21 were significantly increased in EAMG mice, compared with controls. In addition, we found that frequencies of Tfh cells were positively correlated with the levels of serum anti-AChR Ab. In-vivo transduction of lenti-siRNA-Bcl6 ameliorates the severity of ongoing EAMG with decreased Tfh cells, Bcl-6 and IL-21 expression, and leads to decreased anti-AChR antibody levels. Furthermore, we found that siRNA knockdown of Bcl-6 expression increases the expression of Th1(IFN-γ, T-bet) and Th2 markers (IL-4 and GATA3), but failed to alter the expression of Th17-related markers (RORγt, IL-17) and Treg markers (FoxP3). Our study suggests that Tfh cells contribute to the antibody production and could be one of the most important T cell subsets responsible for development and progression of EAMG or MG. Bcl-6 provides a promising therapeutic target for immunotherapy not only for MG, but also for other antibody-mediated autoimmune diseases.

Delivery of an miR155 inhibitor by anti-CD20 single-chain antibody into B cells reduces the acetylcholine receptor-specific autoantibodies and ameliorates experimental autoimmune myasthenia gravis.

MicroRNA-155 (miR155) is required for antibody production after vaccination with attenuated Salmonella. miR155-deficient B cells generated reduced germinal centre responses and failed to produce high-affinity immunoglobulin (Ig)G1 antibodies. In this study, we observed up-regulation of miR155 in the peripheral blood mononuclear cells (PBMCs) of patients with myasthenia gravis (MG), and miR155 was also up-regulated in torpedo acetylcholine receptor (T-AChR)-stimulated B cells. We used an inhibitor of miR155 conjugated to anti-CD20 single-chain antibody to treat both the cultured B cells and the experimental autoimmune MG (EAMG) mice. Our results demonstrated that silencing of miR155 by its inhibitor impaired the B cell-activating factor (BAFF)-R-related signalling pathway and reduced the translocation of nuclear factor (NF)-κB into the nucleus. Additionally, AChR-specific autoantibodies were reduced, which may be related to the altered amounts of marginal zone B cells and memory B cells in the spleens of EAMG mice. Our study suggests that miR155 may be a promising target for the clinical therapy of MG.

CD59 Expression in Skeletal Muscles and Its Role in Myasthenia Gravis.

BACKGROUND AND OBJECTIVES: Complement regulatory proteins at the neuromuscular junction (NMJ) could offer protection against complement-mediated damage in myasthenia gravis (MG). However, there is limited information on their expression at the human NMJ. Thus, this study aimed at investigating the expression of the cluster of differentiation 59 (CD59) at the NMJ of human muscle specimens and demonstrating the overexpression of CD59 mRNA and protein in the muscles of patients with MG. METHODS: In this observational study, muscle specimens from 16 patients with MG (9 and 7 patients with and without thymoma, respectively) and 6 nonmyopathy control patients were examined. Immunohistochemical stains, Western blot analysis, and quantitative real-time reverse transcription PCR were used to evaluate the CD59 expression. RESULTS: A strong localized expression of CD59 was observed at the NMJ in both patients with and without MG. Moreover, the CD59/glyceraldehyde-3-phosphate dehydrogenase protein ratio in patients with MG was significantly higher than that in the nonmyopathy controls (MG; n = 16, median 0.16, interquartile range (IQR) 0.08-0.26 and nonmyopathy controls; n = 6, median 0.03, IQR 0.02-0.11, p = 0.01). The proportion of CD59 mRNA expression relative to AChR mRNA expression (ΔCtCD59/AChR) was associated with the quantitative MG score, MG activities of daily living score, and MG of Foundation of America Clinical Classification (r = 0.663, p = 0.01; r = 0.638, p = 0.014; and r = 0.715, p = 0.003, respectively). DISCUSSION: CD59, which acts as a complement regulator, may protect the NMJ from complement attack. Our findings could provide a basis for further research that investigates the underlying pathogenesis in MG and the immunomodulating interactions of the muscle cells.

Ocular and generalized myasthenia gravis induced by human acetylcholine receptor γ subunit immunization.

INTRODUCTION: HLA-DQ8 transgenic mice develop ocular myasthenia gravis (oMG), which then progresses to generalized MG (gMG) when immunized with the human acetylcholine receptor (H-AChR) α subunit. Because the fetal AChR γ subunit is expressed in adult extraocular muscles, we anticipated that γ subunit immunization would generate an immune response to mouse AChR and induce MG in mice. RESULTS: H-AChR γ subunit immunization in HLA-DQ8 mice induced an autoimmune response to mouse AChR and led to the destruction of AChR in the neuromuscular junction (NMJ) by anti-AChR antibody and complement activation, and it triggered upregulation of AChR gene transcription. CONCLUSION: Our findings indicate that oMG may be induced by immunity to the AChR γ subunit.

Metformin inhibits the pathogenic functions of AChR-specific B and Th17 cells by targeting miR-146a.

Myasthenia gravis (MG) is characterized by fatigable skeletal muscle weakness with a fluctuating and unpredictable disease course and is caused by circulating autoantibodies and pathological T helper cells. Regulation of B-cell function and the T-cell network may be a potential therapeutic strategy for MG. MicroRNAs (miRNAs) have emerged as potential biomarkers in immune disorders due to their critical roles in various immune cells and multiple inflammatory diseases. Aberrant miR-146a signal activation has been reported in autoimmune diseases, but a detailed exploration of the relationship between miR-146a and MG is still necessary. Using an experimental autoimmune myasthenia gravis (EAMG) rat model, we observed that miR-146a was highly expressed in the spleen but expressed at low levels in the thymus and lymph nodes in EAMG rats. Additionally, miR-146a expression in T and B cells was also quite different. EAMG-specific Th17 and Treg cells had lower miR-146a levels, while EAMG-specific B cells had higher miR-146a levels, indicating that targeted intervention against miR-146a might have diametrically opposite effects. Metformin, a drug that was recently demonstrated to alleviate EAMG, may rescue the functions of both Th17 cells and B cells by reversing the expression of miR-146a. We also investigated the downstream target genes of miR-146a in both T and B cells using bioinformatics screening and qPCR. Taken together, our study identifies a complex role of miR-146a in the EAMG rat model, suggesting that more caution should be paid in targeting miR-146a for the treatment of MG.

Blocking of IL-6 suppresses experimental autoimmune myasthenia gravis.

Suppressive regulatory T cells (Treg) and pathogenic T helper 17 (Th17) cells are two lymphocyte subsets with opposing activities in autoimmune diseases. The proinflammatory cytokine IL-6 is a potent factor in switching immune responses in vivo from the induction of Treg to pathogenic Th17 cells. We studied the Treg and Th17 cell compartments in experimental autoimmune myasthenia gravis (EAMG) and healthy control rats in order to assess whether the equilibrium between Treg and Th17 cells is perturbed in the disease. We found that Th17 cell-related genes are upregulated and Treg-related genes are downregulated in EAMG. The shift in favor of Th17 cells in EAMG could be reversed by antibodies to IL-6. Administration of anti-IL-6 antibodies to myasthenic rats suppressed EAMG when treatment started at the acute or at the chronic phase of disease. Suppression of EAMG by anti-IL-6 antibodies was accompanied by a decrease in the overall rat anti-AChR antibody titer and by a reduced number of B cells as compared with control treatment. Administration of anti-IL-6 antibodies led to down-regulation of several Th17 related genes including IL-17, IL-17R, IL-23R and IL-21 but did not affect the number of Treg cells in the lymph nodes. These data identify IL-6 as an important target for modulation of autoimmune responses.

IL-4 receptor as a bridge between the immune system and muscle in experimental myasthenia gravis I: up-regulation of muscle IL-15 by IL-4.

The study reported below describes increased expression of IL-4 receptor in cultured rat myocytes following exposure to an antibody reactive with the acetylcholine receptor (AChR). In addition, upon up-regulation of IL-4R, myocytes demonstrated an increased responsiveness to IL-4 by producing increased levels of IL-15. Moreover, following passive transfer of AChR antibody into Lewis rats, both the increased IL-4R expression and IL-15 production were also observed in intact skeletal muscle, co-localizing in particular individual muscle fibers; the same muscle fibers also produced the chemokine MCP-1 to which IL-4-producing T cells were attracted. A model is proposed in which these muscle activities participate in disease progression in experimental myasthenia gravis.

Efgartigimod improves muscle weakness in a mouse model for muscle-specific kinase myasthenia gravis.

Myasthenia gravis is hallmarked by fatigable muscle weakness resulting from neuromuscular synapse dysfunction caused by IgG autoantibodies. The variant with muscle-specific kinase (MuSK) autoantibodies is characterized by prominent cranial and bulbar weakness and a high frequency of respiratory crises. The majority of MuSK MG patients requires long-term immunosuppressive treatment, but the result of these treatments is considered less satisfactory than in MG with acetylcholine receptor antibodies. Emergency treatments are more frequently needed, and many patients develop permanent facial weakness and nasal speech. Therefore, new treatment options would be welcome. The neonatal Fc receptor protects IgG from lysosomal breakdown, thus prolonging IgG serum half-life. Neonatal Fc receptor antagonism lowers serum IgG levels and thus may act therapeutically in autoantibody-mediated disorders. In MuSK MG, IgG4 anti-MuSK titres closely correlate with disease severity. We therefore tested efgartigimod (ARGX-113), a new neonatal Fc receptor blocker, in a mouse model for MuSK myasthenia gravis. This model involves 11 daily injections of purified IgG4 from MuSK myasthenia gravis patients, resulting in overt myasthenic muscle weakness and, consequently, body weight loss. Daily treatment with 0.5 mg efgartigimod, starting at the fifth passive transfer day, reduced the human IgG4 titres about 8-fold, despite continued daily injection. In muscle strength and fatigability tests, efgartigimod-treated myasthenic mice outperformed control myasthenic mice. Electromyography in calf muscles at endpoint demonstrated less myasthenic decrement of compound muscle action potentials in efgartigimod-treated mice. These substantial in vivo improvements of efgartigimod-treated MuSK MG mice following a limited drug exposure period were paralleled by a tendency of recovery at neuromuscular synaptic level (in various muscles), as demonstrated by ex vivo functional studies. These synaptic improvements may well become more explicit upon longer drug exposure. In conclusion, our study shows that efgartigimod has clear therapeutic potential in MuSK myasthenia gravis and forms an exciting candidate drug for many autoantibody-mediated neurological and other disorders.

C5a is not involved in experimental autoimmune myasthenia gravis pathogenesis.

C5 deficient mice are highly resistant to experimental autoimmune myasthenia gravis (EAMG) despite intact immune response to acetylcholine receptor (AChR), validating the pivotal role played by membrane attack complex (MAC, C5b-9) in neuromuscular junction destruction. To distinguish the significance of C5a from that of C5b in EAMG pathogenesis, C5a receptor (C5aR) knockout (KO) and wild-type (WT) mice were immunized with AChR to induce pathogenic anti-AChR antibodies. In contrast with C5 deficient mice, C5aR KO mice were equally susceptible to EAMG as WT mice and exhibited comparable antibody and lymphocyte proliferation response to AChR implicating that C5a is not involved in EAMG development.

Classical complement pathway in experimental autoimmune myasthenia gravis pathogenesis.

Mice deficient for complement factors C3, C4, or C5 are resistant to experimental autoimmune myasthenia gravis (EAMG). Acetylcholine receptor (AChR) immune lymph node cells (LNC) of C3 deficient mice produce less interleukin 6 (IL-6), and EAMG-resistant IL-6 deficient mice have less serum C3. Increased serum C1q-circulating immune complex (CIC) levels correlated with EAMG disease severity in RIIIS/J mice. The CIC promotes EAMG severity by stimulating the production of LNC IL-6, serum C1q, and C3 via FCgammaR interaction. Therefore, EAMG/MG could be treated by blocking the activation of classical complement pathway (CCP) and/or IL-6. Anti-C1q antibody administration before and following AChR immunization suppressed EAMG by reducing LNC IL-6 production and neuromuscular junction deposits of IgG, C3, and C5b-C9 complexes. Treatment with low dose (10 microg) of anti-C1q antibody twice a week for 4 weeks in mice with ongoing clinical EAMG reduced the clinical severity of disease and LNC IL-6 production. Therefore, inhibitors of CCP factors C1q, C2, or C4 could treat MG and would preserve the alternate complement pathway activation. Our goal is to tailor MG therapy using anti-C2/C4 reagents in combination, with or without anti-cytokine (e.g., anti-IL-6) reagents.

Toll-like receptor 9 antagonist suppresses humoral immunity in experimental autoimmune myasthenia gravis.

Recent studies have demonstrated the important role of toll-like receptor 9 (TLR9) signalling in autoimmune diseases, but its role in myasthenia gravis (MG) has not been fully established. We show herein that blocking TLR9 signalling via the suppressive oligodeoxynucleotide (ODN) H154 alleviated the symptoms of experimental autoimmune myasthenia gravis (EAMG). With the downregulation of dendritic cells (DCs), TLR9 interruption reduced follicular helper T cells (Tfh) and germinal centre (GC) B cells, leading to decreased antibody production. In addition, TLR9+ B cells as well as total B cells in the spleen were inhibited by H154. These findings highlight the critical role of TLR9 in EAMG and suggest that the inhibition of the TLR9 pathway might be a potential pharmacological strategy for the treatment of myasthenia gravis.

A new mouse model of autoimmune ocular myasthenia gravis.

PURPOSE: To establish a novel model of autoimmune ocular myasthenia gravis (oMG) in mice and study the pathogenic mechanisms of oMG. METHODS: oMG was induced in HLA-DQ8 transgenic, HLA-DR3 transgenic, major histocompatibility complex (MHC) class II-deficient, C57BL/6, and C57BL/10 mice by immunization with an Escherichia coli plasmid expressing the recombinant human acetylcholine receptor (AChR) alpha subunit. RESULTS: All strains of immunized mice developed ocular myasthenia gravis with varying disease incidence and severity. HLA-DQ8 transgenic mice were highly susceptible to oMG. Mice with oMG had serum autoantibodies to the mouse extraocular AChR, pathologic deposits of IgG, C3, and C5b-C9 in their extraocular and limb neuromuscular junctions, and droopiness of eyelids. HLA-DR3 transgenic and MHC class II-deficient mice were relatively resistant to oMG induced by AChR alpha subunit immunization and had minimal ocular abnormalities. CONCLUSIONS: These findings suggest that oMG pathogenesis could be triggered by immunity to the human AChR alpha subunit and that MHC class II molecule is required for human AChR alpha subunit presentation and CD4 cell-mediated anti-AChR antibody class switching. Differential oMG susceptibility observed in DQ8 and DR3 transgenic mice correlated with the intensity of lymphocytes to respond to the human AChR alpha subunit. This new model of oMG will be a valuable tool for studying the mechanism of oMG and gMG pathogenesis in humans and for preclinical therapeutic analysis.

Complement regulator CD59 deficiency fails to augment susceptibility to actively induced experimental autoimmune myasthenia gravis.

Complement deficient mice are resistant to experimental autoimmune myasthenia gravis (EAMG), suggesting a pivotal role for the membrane attack complex (MAC) in EAMG pathogenesis. To test the significance of MAC regulation in EAMG pathogenesis, CD59 KO and wild type mice were immunized with acetylcholine receptor (AChR). Interestingly, deletion of CD59, the regulator of MAC assembly, failed to augment EAMG susceptibility. The CD59 KO mice had reduced serum anti-AChR IgG1, IgG2b and complement levels. Their lymph node cell IL-2 production and lymphocyte proliferation response to AChR were reduced. The data challenge the current paradigm that CD59 is solely involved in MAC regulation and suggest a role for this molecule in antigen-driven T cell and B cell activation.