Muscle biology
Abstract
Mutations in the dysferlin gene underlie a group of autosomal recessive muscle-wasting disorders denoted as dysferlinopathies. Dysferlin has been shown to play roles in muscle membrane repair and muscle regeneration, both of which require vesicle-membrane fusion. However, the mechanism by which muscle becomes dystrophic in these disorders remains poorly understood. Although muscle inflammation is widely recognized in dysferlinopathy and dysferlin is expressed in immune cells, the contribution of the immune system to the pathology of dysferlinopathy remains to be fully explored. Here, we show that the complement system plays an important role in muscle pathology in dysferlinopathy. Dysferlin deficiency led to increased expression of complement factors in muscle, while muscle-specific transgenic expression of dysferlin normalized the expression of complement factors and eliminated the dystrophic phenotype present in dysferlin-null mice. Furthermore, genetic disruption of the central component (C3) of the complement system ameliorated muscle pathology in dysferlin-deficient mice but had no significant beneficial effect in a genetically distinct model of muscular dystrophy, mdx mice. These results demonstrate that complement-mediated muscle injury is central to the pathogenesis of dysferlinopathy and suggest that targeting the complement system might serve as a therapeutic approach for this disease.
Authors
Renzhi Han, Ellie M. Frett, Jennifer R. Levy, Erik P. Rader, John D. Lueck, Dimple Bansal, Steven A. Moore, Rainer Ng, Daniel Beltrán-Valero de Bernabé, John A. Faulkner, Kevin P. Campbell
Abstract
The active thyroid hormone 3,5,3′ triiodothyronine (T3) is a major regulator of skeletal muscle function. The deiodinase family of enzymes controls the tissue-specific activation and inactivation of the prohormone thyroxine (T4). Here we show that type 2 deiodinase (D2) is essential for normal mouse myogenesis and muscle regeneration. Indeed, D2-mediated increases in T3 were essential for the enhanced transcription of myogenic differentiation 1 (MyoD) and for execution of the myogenic program. Conversely, the expression of T3-dependent genes was reduced and after injury regeneration markedly delayed in muscles of mice null for the gene encoding D2 (Dio2), despite normal circulating T3 concentrations. Forkhead box O3 (FoxO3) was identified as a key molecule inducing D2 expression and thereby increasing intracellular T3 production. Accordingly, FoxO3-depleted primary myoblasts also had a differentiation deficit that could be rescued by high levels of T3. In conclusion, the FoxO3/D2 pathway selectively enhances intracellular active thyroid hormone concentrations in muscle, providing a striking example of how a circulating hormone can be tissue-specifically activated to influence development locally.
Authors
Monica Dentice, Alessandro Marsili, Raffaele Ambrosio, Ombretta Guardiola, Annarita Sibilio, Ji-Hye Paik, Gabriella Minchiotti, Ronald A. DePinho, Gianfranco Fenzi, P. Reed Larsen, Domenico Salvatore
Abstract
Limb-girdle muscular dystrophy type 2A (LGMD2A) is a genetic disease that is caused by mutations in the calpain 3 gene (CAPN3), which encodes the skeletal muscle–specific calpain, calpain 3 (also known as p94). However, the precise mechanism by which p94 functions in the pathogenesis of this disease remains unclear. Here, using p94 knockin mice (termed herein p94KI mice) in which endogenous p94 was replaced with a proteolytically inactive but structurally intact p94:C129S mutant protein, we have demonstrated that stretch-dependent p94 distribution in sarcomeres plays a crucial role in the pathogenesis of LGMD2A. The p94KI mice developed a progressive muscular dystrophy, which was exacerbated by exercise. The exercise-induced muscle degeneration in p94KI mice was associated with an inefficient redistribution of p94:C129S in stretched sarcomeres. Furthermore, the p94KI mice showed impaired adaptation to physical stress, which was accompanied by compromised upregulation of muscle ankyrin-repeat protein-2 and hsp upon exercise. These findings indicate that the stretch-induced dynamic redistribution of p94 is dependent on its protease activity and essential to protect muscle from degeneration, particularly under conditions of physical stress. Furthermore, our data provide direct evidence that loss of p94 protease activity can result in LGMD2A and molecular insight into how this could occur.
Authors
Koichi Ojima, Yukiko Kawabata, Harumi Nakao, Kazuki Nakao, Naoko Doi, Fujiko Kitamura, Yasuko Ono, Shoji Hata, Hidenori Suzuki, Hiroyuki Kawahara, Julius Bogomolovas, Christian Witt, Coen Ottenheijm, Siegfried Labeit, Henk Granzier, Noriko Toyama-Sorimachi, Michiko Sorimachi, Koichi Suzuki, Tatsuya Maeda, Keiko Abe, Atsu Aiba, Hiroyuki Sorimachi
Abstract
Duchenne muscular dystrophy (DMD) is a fatal disease of striated muscle deterioration caused by lack of the cytoskeletal protein dystrophin. Dystrophin deficiency causes muscle membrane instability, skeletal muscle wasting, cardiomyopathy, and heart failure. Advances in palliative respiratory care have increased the incidence of heart disease in DMD patients, for which there is no cure or effective therapy. Here we have shown that chronic infusion of membrane-sealing poloxamer to severely affected dystrophic dogs reduced myocardial fibrosis, blocked increased serum cardiac troponin I (cTnI) and brain type natriuretic peptide (BNP), and fully prevented left-ventricular remodeling. Mechanistically, we observed a markedly greater primary defect of reduced cell compliance in dystrophic canine myocytes than in the mildly affected mdx mouse myocytes, and this was associated with a lack of utrophin upregulation in the dystrophic canine cardiac myocytes. Interestingly, after chronic poloxamer treatment, the poor compliance of isolated canine myocytes remained evident, but this could be restored to normal upon direct application of poloxamer. Collectively, these findings indicate that dystrophin and utrophin are critical to membrane stability–dependent cardiac myocyte mechanical compliance and that poloxamer confers a highly effective membrane-stabilizing chemical surrogate in dystrophin/utrophin deficiency. We propose that membrane sealant therapy is a potential treatment modality for DMD heart disease and possibly other disorders with membrane defect etiologies.
Authors
DeWayne Townsend, Immanuel Turner, Soichiro Yasuda, Joshua Martindale, Jennifer Davis, Michael Shillingford, Joe N. Kornegay, Joseph M. Metzger
Abstract
Authors
Ahlke Heydemann, Ermelinda Ceco, Jackie E. Lim, Michele Hadhazy, Pearl Ryder, Jennifer L. Moran, David R. Beier, Abraham A. Palmer, Elizabeth M. McNally
Abstract
Signaling via the neuronal NOS (nNOS) splice variant nNOSμ is essential for skeletal muscle health and is commonly reduced in neuromuscular disease. nNOSμ is thought to be the predominant source of NO in skeletal muscle. Here we demonstrate the existence of what we believe to be a novel signaling pathway, mediated by the nNOS splice variant nNOSβ, localized at the Golgi complex in mouse skeletal muscle cells. In contrast to muscles lacking nNOSμ alone, muscles missing both nNOSμ and nNOSβ were severely myopathic, exhibiting structural defects in the microtubule cytoskeleton, Golgi complex, and mitochondria. Skeletal muscles lacking both nNOSμ and nNOSβ were smaller in mass, intrinsically weak, highly susceptible to fatigue, and exhibited marked postexercise weakness. Our data indicate that nNOSβ is a critical regulator of the structural and functional integrity of skeletal muscle and demonstrate the existence of 2 functionally distinct nNOS microdomains in skeletal muscle, created by the differential targeting of nNOSμ to the sarcolemma and nNOSβ to the Golgi. We have previously shown that sarcolemmal nNOSμ matches the blood supply to the metabolic demands of active muscle. We now demonstrate that nNOSβ simultaneously modulates the ability of skeletal muscle to maintain force production during and after exercise. We conclude therefore that nNOS splice variants are critical regulators of skeletal muscle exercise performance.
Authors
Justin M. Percival, Kendra N.E. Anderson, Paul Huang, Marvin E. Adams, Stanley C. Froehner
Abstract
Muscle wasting is associated with a number of pathophysiologic conditions, including metabolic acidosis, diabetes, sepsis, and high angiotensin II levels. Under these conditions, activation of muscle protein degradation requires endogenous glucocorticoids. As the mechanism(s) underlying this dependence on glucocorticoids have not been identified, we analyzed the effects of glucocorticoids on muscle wasting in a mouse model of acute diabetes. Adrenalectomized, acutely diabetic mice given a physiologic dose of glucocorticoids exhibited decreased IRS-1–associated PI3K activity in muscle and progressive muscle atrophy. These responses were related to increased association of PI3K with the glucocorticoid receptor (GR). In mice with muscle-specific GR deletion (referred to as MGRKO mice), acute diabetes minimally suppressed IRS-1–associated PI3K activity in muscle and did not cause muscle atrophy. However, when a physiologic dose of glucocorticoids was given to mice with muscle-specific IR deletion, muscle protein degradation was accelerated. Fluorescence resonance energy transfer and an in vitro competition assay revealed that activated GRs competed for PI3K, reducing its association with IRS-1. Reexpression of WT GRs or those with a mutation in the nuclear localization signal in the muscle of MGRKO mice indicated that competition for PI3K was a prominent mechanism underlying reduced IRS-1–associated PI3K activity. This nongenomic influence of the GR contributes to activation of muscle protein degradation. We therefore conclude that stimulation of muscle proteolysis requires 2 events, increased glucocorticoid levels and impaired insulin signaling.
Authors
Zhaoyong Hu, Huiling Wang, In Hee Lee, Jie Du, William E. Mitch
Abstract
Duchenne muscular dystrophy (DMD) is an X-linked, degenerative muscle disease that is exacerbated by secondary inflammation. Here, we characterized the immunological milieu of dystrophic muscle in mdx mice, a model of DMD, to identify potential therapeutic targets. We identified a specific subpopulation of cells expressing the Vβ8.1/8.2 TCR that is predominant among TCR-β+ T cells. These cells expressed high levels of osteopontin (OPN), a cytokine that promotes immune cell migration and survival. Elevated OPN levels correlated with the dystrophic process, since OPN was substantially elevated in the serum of mdx mice and muscle biopsies after disease onset. Muscle biopsies from individuals with DMD also had elevated OPN levels. To test the role of OPN in mdx muscle, mice lacking both OPN and dystrophin were generated and termed double-mutant mice (DMM mice). Reduced infiltration of NKT-like cells and neutrophils was observed in the muscle of DMM mice, supporting an immunomodulatory role for OPN in mdx muscle. Concomitantly, an increase in CD4+ and FoxP3+ Tregs was also observed in DMM muscle, which also showed reduced levels of TGF-β, a known fibrosis mediator. These inflammatory changes correlated with increased strength and reduced diaphragm and cardiac fibrosis. These studies suggest that OPN may be a promising therapeutic target for reducing inflammation and fibrosis in individuals with DMD.
Authors
Sylvia A. Vetrone, Encarnacion Montecino-Rodriguez, Elena Kudryashova, Irina Kramerova, Eric P. Hoffman, Scot D. Liu, M. Carrie Miceli, Melissa J. Spencer
Abstract
Congenital myasthenias (CMs) arise from defects in neuromuscular junction–associated proteins. Deciphering the molecular bases of the CMs is required for therapy and illuminates structure-function relationships in these proteins. Here, we analyze the effects of a mutation in 1 of 4 homologous subunits in the AChR from a CM patient, a Leu to Pro mutation at position 42 of the δ subunit. The mutation is located in a region of contact between subunits required for rapid opening of the AChR channel and impedes the rate of channel opening. Substitutions of Gly, Lys, or Asp for δL42, or substitutions of Pro along the local protein chain, also slowed channel opening. Substitution of Pro for Leu in the ε subunit slowed opening, whereas this substitution had no effect in the β subunit and actually sped opening in the α subunit. Analyses of energetic coupling between residues at the subunit interface showed that δL42 is functionally linked to αT127, a key residue in the adjacent α subunit required for rapid channel opening. Thus, δL42 is part of an intersubunit network that enables ACh binding to rapidly open the AChR channel, which may be compromised in patients with CM.
Authors
Xin-Ming Shen, Taku Fukuda, Kinji Ohno, Steven M. Sine, Andrew G. Engel
Abstract
Numerous studies have suggested that muscle atrophy is accompanied by apoptotic loss of myonuclei and therefore recovery would require replenishment by muscle stem cells. We used in vivo time-lapse microscopy to observe the loss and replenishment of myonuclei in murine muscle fibers following induced muscle atrophy. To our surprise, imaging of single fibers for up to 28 days did not support the concept of nuclear loss during atrophy. Muscles were inactivated by denervation, nerve impulse block, or mechanical unloading. Nuclei were stained in vivo either acutely by intracellular injection of fluorescent oligonucleotides or in time-lapse studies after transfection with a plasmid encoding GFP with a nuclear localization signal. We observed no loss of myonuclei in fast- or slow-twitch muscle fibers despite a greater than 50% reduction in fiber cross-sectional area. TUNEL labeling of fragmented DNA on histological sections revealed high levels of apoptotic nuclei in inactive muscles. However, when costained for laminin and dystrophin, virtually none of the TUNEL-positive nuclei could be classified as myonuclei; apoptosis was confined to stromal and satellite cells. We conclude that disuse atrophy is not a degenerative process, but is rather a change in the balance between protein synthesis and proteolysis in a permanent cell syncytium.
Authors
Jo C. Bruusgaard, Kristian Gundersen
Copyright © 2018 American Society for Clinical Investigation
ISSN: 0021-9738 (print), 1558-8238 (online)