Endomysial fibrosis in duchenne muscular dystrophy: A marker of poor outcome associated with macrophage alternative activation
There is considerable interindividual variability in motor function among patients with Duchenne muscular dystrophy (DMD); moreover, pathogenetic mechanisms of motor dysfunction in DMD are not understood. Using multiparametric analysis, we correlated initial histologic alterations in quadriceps muscle biopsies from 25 steroid therapy-free patients with DMD with 13 relevant clinical features assessed by a single clinical team during a long-term period (mean, >10 years). There was no residual muscle dystrophin by immunohistochemistry and Western blot analysis in the biopsies. Myofiber size, hypercontracted fibers, necrotic/basophilic fibers, endomysial and perimysial fibrosis, and fatty degeneration were assessed by morphometry. Endomysial fibrosis was the only myopathologic parameter that significantly correlated with poor motor outcome as assessed by quadriceps muscle strength, manual muscle testing of upper and lower limbs at 10 years, and age at ambulation loss (all p<0.002). Motor outcome and fibrosis did not correlate with genotype. Myofibers exhibited oxidative stress-induced protein alterations and became separated from capillaries by fibrosis that was associated with both increase of CD206+ alternatively activated macrophages and a relative decrease of CD56+ satellite cells (both p<0.0001). This study provides a strong rationale for antifibrotic therapeutic strategies in DMD and supports the view that alternatively activated macrophages that are known to inhibit myogenesis while promoting cellular collagen production play a key role in myofibrosis.
Available from: Ashling Holland
- "Abnormal calcium handling and a concomitant increase in calpain activity were shown to play a critical secondary role in the molecular pathogenesis of DMD   . Importantly, extensive accumulation of collagen and the substitution of contractile fibers with nonfunctional fibrotic tissue are a critical myopathological parameter in dystrophinopathy    and endomysial fibrosis significantly correlates with poor motor outcome in X-linked muscular dystrophy   . "
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ABSTRACT: Proteomic profiling plays a decisive role in the identification of novel biomarkers of muscular dystrophy and the elucidation of new pathobiochemical mechanisms that underlie progressive muscle wasting. Building on the findings of recent comparative analyses of tissue samples and body fluids from dystrophic animals and patients afflicted with Duchenne muscular dystrophy, we have used here label-free MS to study the severely dystrophic diaphragm from the not extensively characterized mdx-4cv mouse. This animal model of progressive muscle wasting exhibits less dystrophin-positive revertant fibres than the conventional mdx mouse, making it ideal for the future monitoring of experimental therapies. The pathoproteomic signature of the mdx-4cv diaphragm included a significant increase in the fibrosis marker collagen and related extracellular matrix proteins (asporin, decorin, dermatopontin, prolargin) and cytoskeletal proteins (desmin, filamin, obscurin, plectin, spectrin, tubulin, vimentin, vinculin), as well as decreases in proteins of ion homeostasis (parvalbumin) and the contractile apparatus (myosin binding protein). Importantly, one of the most substantially increased proteins was identified as periostin, a matricellular component and apparent marker of fibrosis and tissue damage. Immunoblotting confirmed a considerable increase of periostin in the dystrophin-deficient diaphragm from both mdx and mdx-4cv mice, suggesting an involvement of this matricellular protein in dystrophinopathy-related fibrosis. This article is protected by copyright. All rights reserved.
This article is protected by copyright. All rights reserved.
Available from: Joe N Kornegay
- "Fibrosis is readily seen histologically in DMD (Pearce and Walton 1962) and the mdx (Stedman et al. 1991) and GRMD (Figure 4D) (Kornegay et al. 1988; Valentine et al. 1989) models. Increased endomysial connective tissue occurs early in DMD and correlates with clinical disease progression (Desguerre et al. 2009). Mediators of fibrosis, therefore, are logical therapeutic targets (Klingler et al. 2012). "
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ABSTRACT: Duchenne muscular dystrophy (DMD) is an X-linked human disorder in which absence of the protein dystrophin causes degeneration of skeletal and cardiac muscle. For the sake of treatment development, over and above definitive genetic and cell-based therapies, there is considerable interest in drugs that target downstream disease mechanisms. Drug candidates have typically been chosen based on the nature of pathologic lesions and presumed underlying mechanisms and then tested in animal models. Mammalian dystrophinopathies have been characterized in mice (mdx mouse) and dogs (golden retriever muscular dystrophy [GRMD]). Despite promising results in the mdx mouse, some therapies have not shown efficacy in DMD. Although the GRMD model offers a higher hurdle for translation, dogs have primarily been used to test genetic and cellular therapies where there is greater risk. Failed translation of animal studies to DMD raises questions about the propriety of methods and models used to identify drug targets and test efficacy of pharmacologic intervention. The mdx mouse and GRMD dog are genetically homologous to DMD but not necessarily analogous. Subcellular species differences are undoubtedly magnified at the whole-body level in clinical trials. This problem is compounded by disparate cultures in clinical trials and preclinical studies, pointing to a need for greater rigor and transparency in animal experiments. Molecular assays such as mRNA arrays and genome-wide association studies allow identification of genetic drug targets more closely tied to disease pathogenesis. Genes in which polymorphisms have been directly linked to DMD disease progression, as with osteopontin, are particularly attractive targets.
Available from: Juan L Hancke
- "Pathologic features of DMD include myofiber atrophy, fatty degeneration, necrosis and fibrosis, but only fibrosis has been shown through clinical studies to correlate with poor motor outcome, evaluated by muscle strength and age at loss of ambulation . This finding supports the notion that fibrosis directly contributes to progressive muscle dysfunction and the lethal phenotype of DMD . "
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ABSTRACT: Duchenne muscular dystrophy (DMD) is characterized by the absence of the cytoskeletal protein dystrophin, muscle wasting, increased transforming growth factor type beta (TGF-beta) signaling, and fibrosis. At the present time, the only clinically validated treatments for DMD are glucocorticoids. These drugs prolong muscle strength and ambulation of patients for a short term only and have severe adverse effects. Andrographolide, a bicyclic diterpenoid lactone, has traditionally been used for the treatment of colds, fever, laryngitis, and other infections with no or minimal side effects. We determined whether andrographolide treatment of mdx mice, an animal model for DMD, affects muscle damage, physiology, fibrosis, and efficiency of cell therapy.
mdx mice were treated with andrographolide for three months and skeletal muscle histology, creatine kinase activity, and permeability of muscle fibers were evaluated. Fibrosis and TGF-beta signaling were evaluated by indirect immunofluorescence and Western blot analyses. Muscle strength was determined in isolated skeletal muscles and by a running test. Efficiency of cell therapy was determined by grafting isolated skeletal muscle satellite cells onto the tibialis anterior of mdx mice.
mdx mice treated with andrographolide exhibited less severe muscular dystrophy than untreated dystrophic mice. They performed better in an exercise endurance test and had improved muscle strength in isolated muscles, reduced skeletal muscle impairment, diminished fibrosis and a significant reduction in TGF-beta signaling. Moreover, andrographolide treatment of mdx mice improved grafting efficiency upon intramuscular injection of dystrophin-positive satellite cells.
These results suggest that andrographolide could be used to improve quality of life in individuals with DMD.
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