Autoantibodies in canine masticatory muscle myositis recognize a novel myosin binding protein-C family member

Inflammatory myopathies are a group of autoimmune diseases that affect muscles. In humans, the most common inflammatory myopathies are polymyositis, dermatomyositis, and inclusion body myositis. Autoantibodies may be found in humans with inflammatory myopathies, and these play an important role in diagnosis and disease classification. However, these Abs are typically not muscle specific. Spontaneously occurring canine inflammatory myopathies may be good parallel disorders and provide insights into human myositis. In dogs with inflammatory myopathy, muscle-specific autoantibodies have been found, especially in masticatory muscle myositis. We have identified the major Ag recognized by the autoantibodies in canine masticatory muscle myositis. This Ag is a novel member of the myosin binding protein-C family, which we call masticatory myosin binding protein-C (mMyBP-C). mMyBP-C is localized not only within the masticatory muscle fibers, but also at or near their cell surface, perhaps making it accessible as an immunogen. The gene for mMyBP-C also exists in humans, and mMyBP-C could potentially play a role in certain human inflammatory myopathies. Understanding the role of mMyBP-C in this canine inflammatory myopathy may advance our knowledge of mechanisms of autoimmune inflammatory muscle diseases, not only in dogs, but also in humans.     

Immune mechanisms in the pathogenesis of idiopathic inflammatory myopathies

Idiopathic inflammatory myopathies (IIMs), comprising polymyositis, dermatomyositis, and inclusion-body myositis, are characterized by inflammatory cell infiltrates in skeletal muscle tissue, muscle weakness, and muscle fatigue. The cellular infiltrates often consist of T lymphocytes and macrophages but also, in some cases, B lymphocytes. Emerging data have led to improved phenotypic characterization of the inflammatory cells, including their effector molecules, in skeletal muscle, peripheral blood, and other organs that are frequently involved, such as skin and lungs. In this review we summarize the latest findings concerning the role of T lymphocytes, B lymphocytes, dendritic cells, and other antigen-presenting cells in the pathophysiology of IIMs.     

Daily Supplementation of D-ribose Shows No Therapeutic Benefits in the MHC-I Transgenic Mouse Model of Inflammatory Myositis

Background

Current treatments for idiopathic inflammatory myopathies (collectively called myositis) focus on the suppression of an autoimmune inflammatory response within the skeletal muscle. However, it has been observed that there is a poor correlation between the successful suppression of muscle inflammation and an improvement in muscle function. Some evidence in the literature suggests that metabolic abnormalities in the skeletal muscle underlie the weakness that continues despite successful immunosuppression. We have previously shown that decreased expression of a purine nucleotide cycle enzyme, adenosine monophosphate deaminase (AMPD1), leads to muscle weakness in a mouse model of myositis and may provide a mechanistic basis for muscle weakness. One of the downstream metabolites of this pathway, D-ribose, has been reported to alleviate symptoms of myalgia in patients with a congenital loss of AMPD1. Therefore, we hypothesized that supplementing exogenous D-ribose would improve muscle function in the mouse model of myositis. We treated normal and myositis mice with daily doses of D-ribose (4 mg/kg) over a 6-week time period and assessed its effects using a battery of behavioral, functional, histological and molecular measures.

Results

Treatment with D-ribose was found to have no statistically significant effects on body weight, grip strength, open field behavioral activity, maximal and specific forces of EDL, soleus muscles, or histological features. Histological and gene expression analysis indicated that muscle tissues remained inflamed despite treatment. Gene expression analysis also suggested that low levels of the ribokinase enzyme in the skeletal muscle might prevent skeletal muscle tissue from effectively utilizing D-ribose.

Conclusions

Treatment with daily oral doses of D-ribose showed no significant effect on either disease progression or muscle function in the mouse model of myositis.     

Mannose Binding Lectin Is Required for Alphavirus-Induced Arthritis/Myositis

Mosquito-borne alphaviruses such as chikungunya virus and Ross River virus (RRV) are emerging pathogens capable of causing large-scale epidemics of virus-induced arthritis and myositis. The pathology of RRV-induced disease in both humans and mice is associated with induction of the host inflammatory response within the muscle and joints, and prior studies have demonstrated that the host complement system contributes to development of disease. In this study, we have used a mouse model of RRV-induced disease to identify and characterize which complement activation pathways mediate disease progression after infection, and we have identified the mannose binding lectin (MBL) pathway, but not the classical or alternative complement activation pathways, as essential for development of RRV-induced disease. MBL deposition was enhanced in RRV infected muscle tissue from wild type mice and RRV infected MBL deficient mice exhibited reduced disease, tissue damage, and complement deposition compared to wild-type mice. In contrast, mice deficient for key components of the classical or alternative complement activation pathways still developed severe RRV-induced disease. Further characterization of MBL deficient mice demonstrated that similar to C3^−/− mice, viral replication and inflammatory cell recruitment were equivalent to wild type animals, suggesting that RRV-mediated induction of complement dependent immune pathology is largely MBL dependent. Consistent with these findings, human patients diagnosed with RRV disease had elevated serum MBL levels compared to healthy controls, and MBL levels in the serum and synovial fluid correlated with severity of disease. These findings demonstrate a role for MBL in promoting RRV-induced disease in both mice and humans and suggest that the MBL pathway of complement activation may be an effective target for therapeutic intervention for humans suffering from RRV-induced arthritis and myositis.     

Viral persistence during the developmental phase of Coxsackievirus B1-induced murine polymyositis.

Mice infected with coxsackievirus B1 (CVB1) develop a chronic hindquarter muscle weakness which resembles human polymyositis. In this study, we used in situ hybridization to screen for persistent viral RNA in hamstring and quadriceps muscles from mice that displayed various degrees of clinical weakness. At 28 to 31 days postinfection, when chronic myositis is well developed but infectious virus can no longer be recovered, persistent CVB1 RNA was found in hindquarter skeletal muscle of all 12 infected animals examined. Persistent CVB1 showed a multifocal distribution within muscle and was associated with three different histopathology patterns (HPPs). These three HPPs (HPP-1, HPP-2, and HPP-3) represent potentially different stages in the mechanism of persistence. They are based on the pattern of grains, the location of hybridization signal within the muscle, and the accompanying histopathology. In HPP-1, virus persisted in nonnecrotic muscle fibers and was not directly associated with foci of inflammatory cells. HPP-2 consisted of virus contained within necrotic myocytes that were surrounded by inflammatory cells. HPP-3 was rare and showed virus inside infiltrating mononuclear cells in a region where muscle tissue had been extensively destroyed. Persistent CVB1 occurred more frequently in severely diseased animals and in tissue sections displaying intense inflammation. Moreover, HPP-2 showed a stronger association with tissue inflammation and hindquarter weakness than did HPP-1. These data demonstrate that CVB1 persists in skeletal muscle for at least 28 to 31 days postinfection and support the concept that this persistence plays a role in the development of murine polymyositis.     

Autoantibodies against a specific nuclear RNP protein in sera of patients with autoimmune rheumatic diseases associated with myositis

Polymyositis is an autoimmune, inflammatory disease affecting human skeletal muscle. In the presence of concomitant vasculitis in the skin, the term dermatomyositis is used. In contrast, systemic lupus erythematosus (SLE) is a multisystem disease in which involvement of the skin, kidneys, joints, brain, and other organs may be found. The clinical manifestations vary according to the organ/system involved. It is clinical and therapeutic importance to define which organ/system is involved during the course of the disease. We approached this problem by studying the specificity of autoantibodies that are generated in patients with SLE and polymyositis/dermatomyositis. Among such antibodies are those directed against nuclear components including a variety of ribonucleoprotein (RNP) complexes. We have utilized mammalian nuclear preparations enriched with RNP particles as the antigenic source for immunoblotting studies to identify specific antigenic polypeptides. In the study reported here, sera from five groups of patients were examined: 10 patients with dermatomyositis/polymyositis; six patients with SLE and myositis; 12 lupus patients with cerebral and/or renal disease; eight patients with SLE but no myositis, renal, or cerebral disease; and 5) 11 patients with muscle weakness or muscle disease not due to myositis. In the first two groups of patients with myositis, antibodies against a nuclear RNP protein of 56 KD was identified in 12 of 16 sera. In contrast, such antibodies were found in the serum of only two of 20 patients with SLE but without muscle involvement (groups 3 and 4), and were not found at all in patients with other muscle diseases. This study has identified a new marker, antibodies against a nuclear RNP protein of 56 KD for detecting muscle involvement among the autoimmune rheumatic diseases.     

The inhibition of apoptosis in myositis and in normal muscle cells

The mechanism of injury and death of muscle cells in the inflammatory myopathies (dermatomyositis, polymyositis, and inclusion body myositis) remains obscure. We and others have not detected apoptosis in the muscle biopsies from patients with myositis despite clear evidence of cell damage and loss. We provide evidence in this study that Fas ligand (FasL) as well as Fas is present on muscle cells and inflammatory cells in myositis biopsies: Fas is present on most muscle cells and lymphocytes, and FasL is present on degenerating muscle cells and many infiltrating mononuclear cells. The expression of both Fas and FasL in the inflamed tissue makes the absence of apoptosis more striking. To address the mechanisms of this resistance to classical apoptosis in muscle cells, we have investigated the expression of the antiapoptotic molecule FLICE (Fas-associated death domain-like IL-1-converting enzyme)-inhibitory protein (FLIP) in muscle biopsies of myositis patients and in cultured human skeletal muscle cells. Using laser capture microscopy, we have shown that FLIP is expressed in the muscle fibers and on infiltrating lymphocytes of myositis biopsies. Furthermore, we have shown that FLIP, but not Bcl-2, is expressed in cultured human skeletal muscle cells stimulated with proinflammatory cytokines, and inhibition of FLIP with antisense oligonucleotides promotes significant cleavage of poly(ADP-ribose) polymerase autoantigen, a sensitive indicator of apoptosis. These studies strongly suggest that the resistance of muscle to Fas-mediated apoptosis is due to the expression of FLIP in muscle cells in the inflammatory environment in myositis.     

BLOOD FLOW RESTRICTED RESISTANCE TRAINING ATTENUATES MYOSTATIN GENE EXPRESSION IN A PATIENT WITH INCLUSION BODY MYOSITIS

Inclusion body myositis is a rare idiopathic inflammatory myopathy that produces extreme muscle weakness. Blood flow restricted resistance training has been shown to improve muscle strength and muscle hypertrophy in inclusion body myositis. Objective: The aim of this study was to evaluate the effects of a resistance training programme on the expression of genes related to myostatin (MSTN) signalling in one inclusion body myositis patient. Methods: A 65-year-old man with inclusion body myositis underwent blood flow restricted resistance training for 12 weeks. The gene expression of MSTN, follistatin, follistatin-like 3, activin II B receptor, SMAD-7, MyoD, FOXO-3, and MURF-2 was quantified. Results: After 12 weeks of training, a decrease (25%) in MSTN mRNA level was observed, whereas follistatin and follistatin-like 3 gene expression increased by 40% and 70%, respectively. SMAD-7 mRNA level was augmented (20%). FOXO-3 and MURF-2 gene expression increased by 40% and 20%, respectively. No change was observed in activin II B receptor or MyoD gene expression. Conclusions: Blood flow restricted resistance training attenuated MSTN gene expression and also increased expression of myostatin endogenous inhibitors. Blood flow restricted resistance training evoked changes in the expression of genes related to MSTN signalling pathway that could in part explain the muscle hypertrophy previously observed in a patient with inclusion body myositis.     

Fine needle aspiration cytology of focal myositis: a case report

BACKGROUND: Focal myositis is an unusual inflammatwy lesion of the skeletal muscle first described by Heffizer. It is a benign condition and usually involves the muscles of the limbs. CASE: A man presented with a palpable mass in the left leg of 6 months' duration. Nuclear magnetic resonance of the leg showed a mass in the tibial muscle; the presumptive diagnosis was sarcoma of the muscle. Smears showed inflammatory cells, skeletal muscle fibers with degenerative and regenerative changes, and fibrous tissue, suggesting a diagnosis of focal myositis. An incisional muscle biopsy was performed, confirming the diagnosis. CONCLUSION: Focal myositis should always he considered when aspirating muscle masses because it is a clinical mimic of a neoplasm. The prognosis is good, and all cases reported in the literature were self-limiting and gradually resolved.     

Free radical-mediated skeletal muscle dysfunction in inflammatory conditions.

Loss of functional capacity of skeletal muscle is a major cause of morbidity in patients with a number of acute and chronic clinical disorders, including sepsis, chronic obstructive pulmonary disease, heart failure, uremia, and cancer. Weakness in these patients can manifest as either severe limb muscle weakness (even to the point of virtual paralysis), respiratory muscle weakness requiring mechanical ventilatory support, and/or some combination of these phenomena. While factors such as nutritional deficiency and disuse may contribute to the development of muscle weakness in these conditions, systemic inflammation may be the major factor producing skeletal muscle dysfunction in these disorders. Importantly, studies conducted over the past 15 years indicate that free radical species (superoxide, hydroxyl radicals, nitric oxide, peroxynitrite, and the free radical-derived product hydrogen peroxide) play an key role in modulating inflammation and/or infection-induced alterations in skeletal muscle function. Substantial evidence exists indicating that several free radical species can directly alter contractile protein function, and evidence suggests that free radicals also have important effects on sarcoplasmic reticulum function, on mitochondrial function, and on sarcolemmal integrity. Free radicals also modulate activation of several proteolytic pathways, including proteosomally mediated protein degradation and, at least theoretically, may also influence pathways of protein synthesis. As a result, free radicals appear to play an important role in regulating a number of downstream processes that collectively act to impair muscle function and lead to reductions in muscle strength and mass in inflammatory conditions.     

Theiler's murine encephalomyelitis virus-induced cardiac and skeletal muscle disease.

The DA strain of Theiler's murine encephalomyelitis virus, a member of the cardiovirus genus of picornaviruses, induces a restricted and persistent infection associated with a demyelinating process following intracerebral inoculation of mice; both virus infection and the immune response are believed to contribute to the late white matter disease. We now report that intraperitoneal inoculation with DA produces an acute myositis that progresses to a chronic inflammatory muscle disease in CD-1 mice as well as several inbred mouse strains. Some mouse strains also develop central nervous system white matter disease and a focal myocarditis. Infectious virus in skeletal muscle falls to undetectable levels 3 weeks postinoculation (p.i.), although viral genome persists for at least 12 weeks p.i., the longest period of observation. Severe combined immunodeficient animals have evidence of muscle pathology as long as 5 weeks p.i., suggesting that DA virus is capable of inducing chronic muscle disease in the absence of an immune response. The presence in immunocompetent mice, however, of prominent muscle inflammation in the absence of infectious virus suggests that the immune system also contributes to the pathology. T lymphocytes are the predominant cell type infiltrating the skeletal muscle during the chronic disease. This murine model may further our understanding of virus-induced chronic myositis and help to clarify the pathogenesis of human inflammatory myopathies.     

Characterization of Ross River virus tropism and virus-induced inflammation in a mouse model of viral arthritis and myositis

Mosquito-borne alphaviruses are a significant cause of both encephalitic and arthritic disease in humans worldwide. In contrast to the encephalitic alphaviruses, the pathogenesis of alphavirus-induced arthritic disease is not well understood. Utilizing a mouse model of Ross River virus (RRV) disease, we found that the primary targets of RRV infection are bone, joint, and skeletal muscle tissues of the hind limbs in both outbred CD-1 mice and adult C57BL/6J mice. Moreover, histological analyses demonstrated that RRV infection resulted in severe inflammation of these tissues. Characterization of the inflammatory infiltrate within the skeletal muscle tissue identified inflammatory macrophages, NK cells, and CD4+ and CD8+ T lymphocytes. To determine the contribution of the adaptive immune system, the outcome of RRV-induced disease was examined in C57BL/6J RAG-1(-/-) mice, which lack functional T and B lymphocytes. RAG-1(-/-) and wild-type mice developed similar disease signs, infiltration of inflammatory macrophages and NK cells, and muscle pathology, suggesting that the adaptive immune response does not play a critical role in the development of disease. These results establish the mouse model of RRV disease as a useful system for the identification of viral and host factors that contribute to alphavirus-induced arthritis and myositis.     

Defective endoplasmic reticulum-mitochondria contacts and bioenergetics in SEPN1-related myopathy

SEPN1-related myopathy (SEPN1-RM) is a muscle disorder due to mutations of the SEPN1 gene, which is characterized by muscle weakness and fatigue leading to scoliosis and life-threatening respiratory failure. Core lesions, focal areas of mitochondria depletion in skeletal muscle fibers, are the most common histopathological lesion. SEPN1-RM underlying mechanisms and the precise role of SEPN1 in muscle remained incompletely understood, hindering the development of biomarkers and therapies for this untreatable disease. To investigate the pathophysiological pathways in SEPN1-RM, we performed metabolic studies, calcium and ATP measurements, super-resolution and electron microscopy on in vivo and in vitro models of SEPN1 deficiency as well as muscle biopsies from SEPN1-RM patients. Mouse models of SEPN1 deficiency showed marked alterations in mitochondrial physiology and energy metabolism, suggesting that SEPN1 controls mitochondrial bioenergetics. Moreover, we found that SEPN1 was enriched at the mitochondria-associated membranes (MAM), and was needed for calcium transients between ER and mitochondria, as well as for the integrity of ER-mitochondria contacts. Consistently, loss of SEPN1 in patients was associated with alterations in body composition which correlated with the severity of muscle weakness, and with impaired ER-mitochondria contacts and low ATP levels. Our results indicate a role of SEPN1 as a novel MAM protein involved in mitochondrial bioenergetics. They also identify a systemic bioenergetic component in SEPN1-RM and establish mitochondria as a novel therapeutic target. This role of SEPN1 contributes to explain the fatigue and core lesions in skeletal muscle as well as the body composition abnormalities identified as part of the SEPN1-RM phenotype. Finally, these results point out to an unrecognized interplay between mitochondrial bioenergetics and ER homeostasis in skeletal muscle. They could therefore pave the way to the identification of biomarkers and therapeutic drugs for SEPN1-RM and for other disorders in which muscle ER-mitochondria cross-talk are impaired.Subject terms: Chaperones, Respiratory tract diseases     

Attenuation of experimental autoimmune myositis by blocking ICOS-ICOS ligand interaction

Polymyositis (PM) is an acquired, systemic, connective tissue disease characterized by the proximal muscle weakness and infiltration of mononuclear cells into the affected muscles. To understand its etiology and immunopathogenesis, appropriate animal model is required. It has been demonstrated that immunization with native human skeletal C protein induces severe and reproducible experimental autoimmune myositis (EAM) in Lewis rats, and that the muscle inflammatory lesions in the EAM mimic those of human PM. In the present study, we prepared recombinant skeletal C protein fragment and succeeded in inducing as severe EAM as that by native C protein. We found ICOS expression on muscle fiber-infiltrating T cells in the EAM rats, but not in normal rats. Treatment with anti-ICOS mAb reduced incidence and severity of myositis; decreased the number of muscle-infiltrating CD11b/c+, TCR+, and CD8a+ cells; and inhibited the expression of IL-1alpha and CCL2 in the hamstring muscles of the EAM rats. However, the treatment neither inhibited serum anti-C protein IgG level, C protein-induced proliferation of lymph node (LN) cells, or LN T cells, nor production of IFN-gamma by C protein-stimulated LN cells in EAM rats. These data indicate that analysis of C protein-induced EAM provides not only insights into pathogenesis of PM, but also useful information regarding development of effective immunotherapy against the disease. ICOS-ICOS ligand interaction would be a novel therapeutic target for PM.     

The TWEAK-Fn14 system: breaking the silence of cytokine-induced skeletal muscle wasting

The occurrence of skeletal muscle atrophy, a devastating complication of a large number of disease states and inactivity/disuse conditions, provides a never ending quest to identify novel targets for its therapy. Proinflammatory cytokines are considered the mediators of muscle wasting in chronic diseases; however, their role in disuse atrophy has just begun to be elucidated. An inflammatory cytokine, tumor necrosis factor (TNF)- like weak inducer of apoptosis (TWEAK), has recently been identified as a potent inducer of skeletal muscle wasting. TWEAK activates various proteolytic pathways and stimulates the degradation of myofibril protein both in vitro and in vivo. Moreover, TWEAK mediates the loss of skeletal muscle mass and function in response to denervation, a model of disuse atrophy. Adult skeletal muscle express very low to minimal levels of TWEAK receptor, Fn14. Specific catabolic conditions such as denervation, immobilization, or unloading rapidly increase the expression of Fn14 in skeletal muscle which in turn stimulates the TWEAK activation of various catabolic pathways leading to muscle atrophy. In this article, we have discussed the emerging roles and the mechanisms of action of TWEAK-Fn14 system in skeletal muscle with particular reference to different models of muscle atrophy and injury and its potential to be used as a therapeutic target for prevention of muscle loss.     

Novel regulatory mechanisms in inflammatory arthritis: a role for microRNA

Elucidating pathways that regulate cytokine production in the context of autoimmune disease will likely lead to the development of novel therapeutics. Herein, we review data suggesting that microRNAs (miRs) represent one such level of regulatory activity, with particular emphasis on the pathogenesis of rheumatoid arthritis (RA). A series of miRs have been identified to be dysregulated in cell subsets within the articular compartment of patients with RA. These have a critical role in regulating cartilage-invading phenotype of RA synovial fibroblasts. More recently, several studies suggest that miRs also regulate leukocyte activation and cytokine production that in turn contribute to the immunologic component of effector synovial pathology. Together, these observations open an exciting new vista of understanding and therapeutic opportunity for this difficult and common disease.     

Novel autoantibodies and clinical phenotypes in adult and juvenile myositis

Autoantibodies targeting intracellular proteins involved in key processes are detected in patients with idiopathic inflammatory myopathies. These myositisspecific autoantibodies have been increasingly demonstrated to correlate with distinct clinical phenotypes within the myositis spectrum. This review highlights the clinical associations of the myositisspecific autoantibodies, with particular attention to the recently identified and characterized novel myositis autoantibodies: p155/140, p140 (MJ), CADM-140 (MDA5), SAE, and 200/100.     

Diaphragmatic weakness and myositis associated with systemic juvenile rheumatoid arthritis.

An unusual presentation of systemic juvenile rheumatoid arthritis in a young adult is reported. Among the major manifestations were severe muscle weakness and dyspnea, which were found to be due to myositis and diaphragmatic weakness. The evolution of the disease and its response to therapy are described.