Expression of tumor necrosis factor-α in regenerating muscle fibers in inflammatory and non-inflammatory myopathies

The expression level of tumor necrosis factor (TNF)-alpha is elevated in idiopathic inflammatory myopathies and Duchenne muscular dystrophy (DMD), but the precise role of TNF-alpha is unknown. To elucidate the possible role of TNF-alpha, we investigated the expression of TNF-alpha and its receptor in polymyositis (PM), dermatomyositis (DM), and DMD using in situ hybridization (ISH) and immunohistochemistry. We showed that TNF-alpha mRNA and protein were present in muscle fibers. TNF-alpha-positive fibers were observed in all cases of PM, DM and DMD, but were rare or absent in neurogenic disorders and normal controls. The proportion of TNF-alpha-positive fiber showed a significant positive correlation with the proportion of regenerating fibers that were positive for the developmental form of myosin heavy chain (MHC-d). The number of TNF receptor-positive fibers was small. Some muscle fibers expressed both TNF-alpha and its receptor simultaneously. Our results indicate that TNF-alpha is produced and expressed by muscle fibers and associated with muscle regeneration.     

NOGO is increased and binds to BACE1 in sporadic inclusion-body myositis and in AβPP-overexpressing cultured human muscle fibers

Increased amyloid-β precursor protein (AβPP) and amyloid-β (Aβ) accumulation appear to be upstream steps in the pathogenesis of sporadic inclusion-body myositis (s-IBM). BACE1, participating in Aβ production is also increased in s-IBM muscle fibers. Nogo-B and Nogo-A belong to a family of integral membrane reticulons, and Nogo-B binding to BACE1 blocks BACE1 access to AβPP, decreasing Aβ production. We studied Nogo-B and Nogo-A in s-IBM muscle and in our IBM muscle culture models, based on AβPP-overexpression or ER-stress-induction in cultured human muscle fibers (CHMFs). We report that: (1) in biopsied s-IBM fibers, Nogo-B is increased, accumulates in aggregates, is immuno-co-localized with BACE1, and binds to BACE1; Nogo-A is undetectable. (2) In CHMFs, (a) AβPP overexpression increases Nogo-B, Nogo-A, and BACE1, (b) ER stress increases BACE1 but decreases Nogo-B and Nogo-A, (c) Nogo-B and Nogo-A associate with BACE1. Accordingly, two novel mechanisms, AβPP overexpression and ER stress, are involved in Nogo-B and Nogo-A expression in human muscle. We propose that in s-IBM muscle the Nogo-B increase may represent an attempt by muscle fiber to decrease Aβ production. However, the increase of Nogo-B seems insufficient because Aβ continues to accumulate and the disease progresses. We propose that manipulations, which increase Nogo-B in s-IBM muscle might offer a new therapeutic opportunity. Supported by grants (to VA) from the National Institutes of Health (AG 16768 Merit Award), the Muscular Dystrophy Association, and The Myositis Association (to VA), and the Helen Lewis Research Fund.     

Novel demonstration of amyloid-β oligomers in sporadic inclusion-body myositis muscle fibers

Accumulation of amyloid-β (Aβ) within muscle fibers has been considered an upstream step in the development of the s-IBM pathologic phenotype. Aβ42, which is considered more cytotoxic than Aβ40 and has a higher propensity to oligomerize, is preferentially increased in s-IBM muscle fibers. In Alzheimer disease (AD), low-molecular weight Aβ oligomers and toxic oligomers, also referred to as “Aβ-Derived Diffusible Ligands” (ADDLs), are considered strongly cytotoxic and proposed to play an important pathogenic role. ADDLs have been shown to be increased in AD brain. We now report for the first time that in s-IBM muscle biopsies Aβ-dimer, -trimer, and -tetramer are identifiable by immunoblots. While all the s-IBM samples we studied had Aβ-oligomers, their molecular weights and intensity varied between the patient samples. None of the control muscle biopsies had Aβ oligomers. Dot-immunoblots using highly specific anti-ADDL monoclonal antibodies also showed highly increased ADDLs in all s-IBM biopsies studied, while controls were negative. By immunofluorescence, in some of the abnormal s-IBM muscle fibers ADDLs were accumulated in the form of plaque-like inclusions, and were often increased diffusely in very small fibers. Normal and disease-controls were negative. By gold-immuno-electron microscopy, ADDL-immunoreactivities were in close proximity to 6–10 nm amyloid-like fibrils, and also were immunodecorating amorphous and floccular material. In cultured human muscle fibers, we found that inhibition of autophagy led to the accumulation of Aβ oligomers. This novel demonstration of Aβ42 oligomers in s-IBM muscle biopsy provides additional evidence that intra-muscle fiber accumulation of Aβ42 oligomers in s-IBM may contribute importantly to s-IBM pathogenic cascade.     

Strong immunoreactivity of α1 co-localizes with β-amyloid protein and ubiquitin in vacuolated muscle fibers of inclusion-body myositis

Summary In 10 of 10 inclusion-body myositis (IBM) patients, including 1 hereditary case, vacuolated muscle fibers contained large or small cytoplasmic inclusions immunoreactive for (₁-ACT). All IBM muscle biopsies had characteristic cytoplasmic tubulo-filaments by electron microscopy. None of 17 control muscle biopsies contained the (₁-ACT) immunoreactive inclusions characteristic of IBM. In vacuolated muscle fibers, (₁-ACT) immunoreactive inclusions colocalized with -amyloid protein and ubiquitin immunoreactivities. Our study provides the first demonstration of (₁-ACT) accumulations in abnormal human muscle, and it suggest that, as in Alzheimer's disease and Down's syndrome, (₁-ACT) may be involved in the pathogenesis of IBM.Supported in part by the Norma Bard Research Fund. M.B. is a Research Postdoctoral Fellow     

Myostatin is increased and complexes with amyloid-β within sporadic inclusion-body myositis muscle fibers

Myostatin is a negative regulator of muscle mass and strength. Sporadic inclusion-body myositis (s-IBM) is the most common degenerative muscle disease of older persons and is characterized by pronounced muscle wasting. s-IBM is of unknown etiology and pathogenesis, and it lacks definitive treatment. We have now demonstrated in samples from 12 s-IBM biopsies that: (1) by light and electron microscopic immunocytochemistry, myostatin/myostatin precursor is accumulated within muscle fibers and co-localized with amyloid- (A); (2) by immunoblots, both myostatin and myostatin precursor are increased; and (3) by immunoprecipitation, myostatin precursor complexes with A. Our study suggests that myostatin/myostatin precursor, either alone, or bound to A, may play a novel role in the pathogenesis of s-IBM.     

Amyloid-β42 is preferentially accumulated in muscle fibers of patients with sporadic inclusion-body myositis

Sporadic inclusion-body myositis (s-IBM) is the only muscle disease in which accumulation of amyloid-β (Aβ) in abnormal muscle fibers appears to play a key pathogenic role. Increased amyloid-β precursor protein (AβPP) and Aβ accumulation have been reported to be upstream steps in the development of the s-IBM pathologic phenotype, based on cellular and animal models. Aβ is released from AβPP as a 40 or 42 aminoacid peptide. Aβ42 is considered more cytotoxic than Aβ40, and it has a higher propensity to aggregate and form amyloid fibrils. Using highly specific antibodies, we evaluated in s-IBM muscle biopsies intra-muscle fiber accumulation of Aβ40 and Aβ42-immunoreactive aggregates by light- and electron-microscopic immunocytochemistry, and quantified their amounts by ELISA. In s-IBM, 80–90% of the vacuolated muscle fibers and 5–20% of the non-vacuolated muscle fibers contained plaque-like Aβ42-immunoreactive inclusions, while only 69% of those fibers also contained Aβ40 deposits. By immuno-electronmicroscopy, Aβ42 was associated with 6–10 nm amyloid-like fibrils, small electron-dense floccular clumps and larger masses of amorphous material. Aβ40 was present only on small patches of floccular clumps and amorphous material; it was not associated with 6–10 nm amyloid fibrils. By ELISA, in s-IBM muscle biopsies Aβ42 was present in values 8.53-44.7 pg/ml, while Aβ40 was not detectable; normal age-matched control biopsies did not have any detectable Aβ42 or Aβ40. Thus, in s-IBM muscle fibers, Aβ42 is accumulated more than Aβ40. We suggest that Aβ42 oligomers and their cytotoxicity may play an important role in the s-IBM pathogenesis. Supported by grants from the National Institutes of Health (Merit Award AG16768), the Muscular Dystrophy Association and The Myositis Association (to VA). FC is supported by the Fondation pour la Recherche Médicale.     

Fasciculations in nerve and muscle disorders – A prospective study of muscle ultrasound compared to electromyography

Objectives

We examined the clinical utility of muscle ultrasound (MUS) in detecting fasciculations in patients with nerve and muscle disorders (NMD) and investigated the impact on diagnostic sensitivity when combining electromyography (EMG) and MUS.

Activation of the γ-secretase complex and presence of γ-secretase-activating protein may contribute to Aβ42 production in sporadic inclusion-body myositis muscle fibers

The muscle-fiber phenotype of sporadic inclusion-body myositis (s-IBM), the most common muscle disease associated with aging, shares several pathological abnormalities with Alzheimer disease (AD) brain, including accumulation of amyloid-β 42 (Aβ42) and its cytotoxic oligomers. The exact mechanisms leading to Aβ42 production within s-IBM muscle fibers are not known. Aβ42 and Aβ40 are generated after the amyloid-precursor protein (AβPP) is cleaved by β-secretase and the γ-secretase complex. Aβ42 is considered more cytotoxic than Aβ40, and it has a higher propensity to oligomerize, form amyloid fibrils, and aggregate. Recently, we have demonstrated in cultured human muscle fibers that experimental inhibition of lysosomal enzyme activities leads to Aβ42 oligomerization. In s-IBM muscle, we here demonstrate prominent abnormalities of the γ-secretase complex, as evidenced by: a) increase of γ-secretase components, namely active presenilin 1, presenilin enhancer 2, nicastrin, and presence of its mature, glycosylated form; b) increase of mRNAs of these γ-secretase components; c) increase of γ-secretase activity; d) presence of an active form of a newly-discovered γ-secretase activating protein (GSAP); and e) increase of GSAP mRNA. Furthermore, we demonstrate that experimental inhibition of lysosomal autophagic enzymes in cultured human muscle fibers a) activates γ-secretase, and b) leads to posttranslational modifications of AβPP and increase of Aβ42. Since autophagy is impaired in biopsied s-IBM muscle, the same mechanism might be responsible for its having increased γ-secretase activity and Aβ42 production. Accordingly, improving lysosomal function might be a therapeutic strategy for s-IBM patients.     

Intracytoplasmic vacuoles in αW fibers of dystrophic chicken muscle —Probable early pathologic event initiates massive fiber necrosis

Summary An electron-microscopic study on dystrophic chicken white muscle, posterior latissimus dorsi (PLD), was performed with histochemical identification of three fiber types of R (red), R and W (white) fibers to evaluate the pathophysiology in fiber necrosis. As seen in histochemically stained sections, vacuolar formation in the cytoplasm, an outstanding pathologic feature in chicken dystrophy, was recognized in the W fibers by electron microscopy. The vacuole was membrane-bound and thought to originate from coalescence or dilatation of extensively proliferated sarcotubular system. There was evidence of a delay in fiber type transformation from R to W in dystrophic white muscle, while the initial pathologic event of sarcotubular system proliferation might be expressed only after muscle fibers had attained histochemical characteristics of W fibers. Localized myofibrillar degeneration was encountered in the vicinity of the vacuole with focal membrane defect. An influx of extracellular fluid through the vacuolated sarcotubular system into the sarcoplasm may activate certain proteases, such as calcium-dependent protease because the extracellular fluid contains high concentration of calcium ion. The activated protease then degrades structural protein, especially Z-line protein, followed by fiber necrosis with phagocytosis.Supported in part by grant no. 4, 1980, from the National Center for Nervous, Mental, and Muscular Disorders (NCNMMD) of the Ministry of Health and Welfare, Japan     

Dopamine-depletion and increased α-synuclein load induce degeneration of cortical cholinergic fibers in mice

Cognitive dysfunction can be common among Parkinson's disease (PD) patients, and multiplication of the gene α-synuclein (αsyn) increases the risk of dementia. Here, we studied the role of dopamine-depletion and increased αsyn load and aggregation on cholinergic structures in vivo. Wild-type (WT) and mice with A30P αsyn overexpression were treated subacutely with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), and the number of cholinergic cells in their nucleus basalis magnocellularis-substantia innominata (NBM-SI), their cortical fiber density and their expression of different genes 1day or 90 days after the last MPTP-injection were measured. Long-term dopamine depletion decreased the expression of choline acetyl transferase (ChAT) in the NBM-SI of WT mice, but no neuron loss was observed. In contrast, cortical cholinergic fiber density was decreased three months after MPTP-injection. Increased brain-derived neurotrophic factor expression could maintain cholinergic functions under these conditions. Expression of A30P αsyn in six-months-old transgenic mice resulted in decreased tyrosine receptor kinase B expression, and lower cortical cholinergic fiber density. Dopamine-depletion by MPTP induced cholinergic cell loss in the NBM-SI and increased cortical fiber loss. Our findings may explain why cholinergic cells are more vulnerable in PD, leading to an increased probability of dementia.     

Does non-quantum secretion of acetylcholine from motor neuron endings participate in the neurotrophic control of the membrane potential of muscle fibers in the rat?

The resting membrane potential of synaptic zone of muscle fibres within 2-3 mm (near) and 9-11 mm (far) of the nerve section of the rat diaphragm muscle was measured after 3 hours or on the 5th day after the motor nerve section. The membrane potential of "near" fibres was lower than that of "far" fibres. The presence of carbamylcholine or cGMP in the culture medium maintained the membrane potential of "near" fibres close to that of "far" fibres. Ouabain prevented these effects. On the 5th day after the nerve section the presence of carbamylcholine or cGMP had no effect on the membrane potential. In the in vitro experiments the nonquantal acetycholine release was equal in "near" and "far" fibres and just the same as in the control preparations. A conclusion is made that synaptic acetylcholine from motor nerve endings in nonquantal form does not play a significant role in neurotrophic control of the resting membrane potential of muscle fibres membrane in mammals.     

Functional recovery and muscle atrophy in pre-clinical models of peripheral nerve transection and gap-grafting in mice:effects of 4-aminopyridine

We recently demonstrated a repurposing beneficial effect of 4-aminopyridine(4-AP),a potassium channel blocker,on functional recove ry and muscle atrophy after sciatic nerve crush injury in rodents.However,this effect of 4-AP is unknown in nerve transection,gap,and grafting models.To evaluate and compare the functional recovery,nerve morphology,and muscle atrophy,we used a novel stepwise nerve transection with gluing(STG),as well as 7-mm irreparable nerve gap(G-7/0) and 7-mm isografting in 5-mm g...     

Low-symptomatic skeletal muscle disease in patients with a cardiac disease – Diagnostic approach in skeletal muscle laminopathies

Mild skeletal muscle symptoms might be accompanied with severe cardiac disease, sometimes indicating a serious inherited disorder. Very often it is a cardiologist who refers a patient with cardiomyopathy and/or cardiac arrhythmia and discrete muscle disease for neurological consultation, which helps to establish a proper diagnosis. Here we present three families in which a diagnosis of skeletal muscle laminopathy was made after careful examination of the members, who presented with cardiac arrhythmia and/or heart failure and a mild skeletal muscle disease, which together with positive family history allowed to direct the molecular diagnostics and then provide appropriate treatment and counseling.     

Could muscle deformity in children with spastic cerebral palsy be related to an impairment of muscle growth and altered adaptation?

Skeletal muscle deformity is common in children with spastic cerebral palsy (CP), but the underlying mechanisms are unclear. This review explores some possible factors which may influence the development of muscle deformity in CP. Normal muscle function and growth appear to depend on the interaction of neuronal, endocrinal, nutritional, and mechanical factors, and also on the development of an appropriate balance between muscle protein synthesis and degradation, and between the development of contractile and non-contractile components. In this context, the changes seen in muscle in children with CP are reviewed and discussed. It is suggested that the development of muscle deformity in children with CP may be related to a multifactorial impairment of muscle growth, on which adaptation of the extracellular matrix due to altered loading may be imposed.     

Evidence for inflammation in Fabry’s disease? Headache and muscle involvement responding to corticosteroid and methotrexate treatment

Journal of Neurology -