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Kamal KY, Trombetta-Lima M. Mechanotransduction and Skeletal Muscle Atrophy: The Interplay Between Focal Adhesions and Oxidative Stress. Int J Mol Sci 2025; 26:2802. [PMID: 40141444 PMCID: PMC11943188 DOI: 10.3390/ijms26062802] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/30/2024] [Revised: 03/15/2025] [Accepted: 03/18/2025] [Indexed: 03/28/2025] Open
Abstract
Mechanical unloading leads to profound musculoskeletal degeneration, muscle wasting, and weakness. Understanding the specific signaling pathways involved is essential for uncovering effective interventions. This review provides new perspectives on mechanotransduction pathways, focusing on the critical roles of focal adhesions (FAs) and oxidative stress in skeletal muscle atrophy under mechanical unloading. As pivotal mechanosensors, FAs integrate mechanical and biochemical signals to sustain muscle structural integrity. When disrupted, these complexes impair force transmission, activating proteolytic pathways (e.g., ubiquitin-proteasome system) that accelerate atrophy. Oxidative stress, driven by mitochondrial dysfunction and NADPH oxidase-2 (NOX2) hyperactivation, exacerbates muscle degeneration through excessive reactive oxygen species (ROS) production, impaired repair mechanisms, and dysregulated redox signaling. The interplay between FA dysfunction and oxidative stress underscores the complexity of muscle atrophy pathogenesis: FA destabilization heightens oxidative damage, while ROS overproduction further disrupts FA integrity, creating a self-amplifying vicious cycle. Therapeutic strategies, such as NOX2 inhibitors, mitochondrial-targeted antioxidants, and FAK-activating compounds, promise to mitigate muscle atrophy by preserving mechanotransduction signaling and restoring redox balance. By elucidating these pathways, this review advances the understanding of muscle degeneration during unloading and identifies promising synergistic therapeutic targets, emphasizing the need for combinatorial approaches to disrupt the FA-ROS feedback loop.
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Affiliation(s)
- Khaled Y. Kamal
- Department of Kinesiology, Iowa State University, Ames, IA 50011, USA
| | - Marina Trombetta-Lima
- Department of Pharmaceutical Technology and Biopharmacy, Groningen Research Institute of Pharmacy, University of Groningen, 9700 Groningen, The Netherlands;
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Li WZ, Xiong Y, Wang TK, Chen YY, Wan SL, Li LY, Xu M, Tong JJ, Qian Q, Jiang CQ, Liu WC. Quantitative proteomics analysis reveals the pathogenesis of obstructed defecation syndrome caused by abnormal expression of dystrophin. World J Gastroenterol 2024; 30:4817-4835. [PMID: 39649544 PMCID: PMC11606370 DOI: 10.3748/wjg.v30.i45.4817] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/12/2024] [Revised: 09/20/2024] [Accepted: 10/15/2024] [Indexed: 11/13/2024] Open
Abstract
BACKGROUND Obstructed defecation syndrome (ODS) represents the most prevalent form of chronic constipation, affecting a diverse patient population, leading to numerous complications, and imposing a significant burden on healthcare resources. Most ODS patients have insufficient rectal propulsion, but the exact mechanism underlying the pathogenesis of ODS remains unclear. AIM To explore the molecular mechanism underlying the pathogenesis of ODS. METHODS A total of 30 pairs of rectal samples were collected from patients with ODS (ODS group) or grade IV prolapsed hemorrhoids without constipation (control group) for quantitative proteomic and bioinformatic analysis. Subsequently, 50 pairs of paraffin-embedded rectal specimens were selected for immunohistochemistry and immunofluorescence studies to validate the analysis results. Human intestinal smooth cell contractile function experiments and electrophysiological experiments were conducted to verify the physiological functions of target proteins. Cellular ultrastructure was detected using transmission electron microscopy. RESULTS In comparison to the control group, the expression level of dystrophin (DMD) in rectal specimens from ODS patients was markedly reduced. This finding was corroborated using immunohistochemistry and immunofluorescence techniques. The diminished expression of DMD compromised the contractile function of intestinal smooth muscle cells. At the molecular level, nucleoporin protein 153 and L-type voltage-gated calcium channel were found to be overexpressed in intestinal smooth muscle cells exhibiting downregulated DMD expression. Electrophysiological experiments confirmed an excessive influx of calcium ions into these cells. Moreover, vacuolar-like structures which may be associated with excessive calcium influx were observed in the cells by transmission electron microscopy. CONCLUSION Decreased DMD expression in intestinal smooth muscle may upregulate L-type voltage-gated calcium channel expression, leading to excessive calcium influx which may cause a decrease in rectal propulsion, thereby contributing to the pathogenesis of ODS.
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Affiliation(s)
- Wen-Zhe Li
- Department of Colorectal and Anal Surgery (Clinical Center for Pelvic Floor Surgery), Clinical Center of Constipation and Pelvic Floor Disease of Wuhan, Hubei Key Laboratory of Intestinal and Colorectal Diseases, Clinical Center of Intestinal and Colorectal Diseases of Hubei Province, Quality Control Center of Colorectal and Anal Surgery of Health Commission of Hubei Province, Zhongnan Hospital of Wuhan University, Wuhan 430071, Hubei Province, China
| | - Yu Xiong
- Department of Radiation and Medical Oncology for Esophageal Mediastinal and Lymphatic Tumors, Zhongnan Hospital of Wuhan University, Wuhan 430071, Hubei Province, China
| | - Tian-Kun Wang
- Center for Evidence-Based and Translational Medicine, Zhongnan Hospital of Wuhan University, Wuhan 430071, Hubei Province, China
| | - Yan-Yan Chen
- Department of Pathology, Zhongnan Hospital of Wuhan University, Wuhan 430071, Hubei Province, China
| | - Song-Lin Wan
- Department of Colorectal and Anal Surgery (Clinical Center for Pelvic Floor Surgery), Clinical Center of Constipation and Pelvic Floor Disease of Wuhan, Hubei Key Laboratory of Intestinal and Colorectal Diseases, Clinical Center of Intestinal and Colorectal Diseases of Hubei Province, Quality Control Center of Colorectal and Anal Surgery of Health Commission of Hubei Province, Zhongnan Hospital of Wuhan University, Wuhan 430071, Hubei Province, China
| | - Lu-Yao Li
- Center for Evidence-Based and Translational Medicine, Zhongnan Hospital of Wuhan University, Wuhan 430071, Hubei Province, China
| | - Meng Xu
- School of Life Sciences, Central China Normal University, Wuhan 430071, Hubei Province, China
| | - Jing-Jing Tong
- School of Life Sciences, Central China Normal University, Wuhan 430071, Hubei Province, China
| | - Qun Qian
- Department of Colorectal and Anal Surgery (Clinical Center for Pelvic Floor Surgery), Clinical Center of Constipation and Pelvic Floor Disease of Wuhan, Hubei Key Laboratory of Intestinal and Colorectal Diseases, Clinical Center of Intestinal and Colorectal Diseases of Hubei Province, Quality Control Center of Colorectal and Anal Surgery of Health Commission of Hubei Province, Zhongnan Hospital of Wuhan University, Wuhan 430071, Hubei Province, China
| | - Cong-Qing Jiang
- Department of Colorectal and Anal Surgery (Clinical Center for Pelvic Floor Surgery), Clinical Center of Constipation and Pelvic Floor Disease of Wuhan, Hubei Key Laboratory of Intestinal and Colorectal Diseases, Clinical Center of Intestinal and Colorectal Diseases of Hubei Province, Quality Control Center of Colorectal and Anal Surgery of Health Commission of Hubei Province, Zhongnan Hospital of Wuhan University, Wuhan 430071, Hubei Province, China
| | - Wei-Cheng Liu
- Department of Colorectal and Anal Surgery (Clinical Center for Pelvic Floor Surgery), Clinical Center of Constipation and Pelvic Floor Disease of Wuhan, Hubei Key Laboratory of Intestinal and Colorectal Diseases, Clinical Center of Intestinal and Colorectal Diseases of Hubei Province, Quality Control Center of Colorectal and Anal Surgery of Health Commission of Hubei Province, Zhongnan Hospital of Wuhan University, Wuhan 430071, Hubei Province, China
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Vanegas C, Ursitti J, Kallenbach JG, Pinto K, Harriot A, Coleman AK, Shi G, Ward CW. Acute microtubule changes linked to DMD pathology are insufficient to impair contractile function or enhance contraction-induced injury in healthy muscle. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2024:2024.06.19.599775. [PMID: 38948772 PMCID: PMC11212994 DOI: 10.1101/2024.06.19.599775] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 07/02/2024]
Abstract
Duchenne muscular dystrophy (DMD) is marked by the genetic deficiency of the dystrophin protein in striated muscle whose consequence is a cascade of cellular changes that predispose the susceptibility to contraction injury central to DMD pathology. Recent evidence identified the proliferation of microtubules enriched in post-translationally modified tubulin as a consequence of dystrophins absence that increases the passive mechanics of the muscle fiber and the excess mechanotransduction elicited reactive oxygen species and calcium signals that promote contraction injury. Motivated by evidence that acutely normalizing the disease microtubule alterations reduced contraction injury in murine DMD muscle (mdx), here we sought the direct impact of these microtubule alterations independent of dystrophins absence and the multitude of other changes consequent to dystrophic disease. To this end we used acute pharmacologic (epithiolone-D, EpoD; 4 hours) or genetic (vashohibin-2 and small vasohibin binding protein overexpression via AAV9; 2 weeks) strategies to effectively model the proliferation of detyrosination enriched microtubules in the mdx muscle. Quantifying in vivo nerve evoked plantarflexor function we find no alteration in peak torque nor contraction kinetics in WT mice modeling these DMD relevant MT alterations. Quantifying the susceptibility to eccentric contraction injury we show EpoD treatment proffered a small but significant protection from contraction injury while VASH/SVBP had no discernable impact. We conclude that the disease dependent MT alterations act in concert with additional cellular changes to predispose contraction injury in DMD.
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Affiliation(s)
- Camilo Vanegas
- Department of Orthopedics, University of Maryland School of Medicine, Baltimore, MD, USA
| | - Jeanine Ursitti
- Department of Orthopedics, University of Maryland School of Medicine, Baltimore, MD, USA
| | - Jacob G Kallenbach
- Department of Orthopedics, University of Maryland School of Medicine, Baltimore, MD, USA
| | - Kaylie Pinto
- Department of Biochemistry and Molecular Biology, University of Maryland School of Medicine, Baltimore, MD, USA
| | - Anicca Harriot
- Department of Biochemistry and Molecular Biology, University of Maryland School of Medicine, Baltimore, MD, USA
| | - Andrew K Coleman
- Center for Biomedical Engineering and Technology, University of Maryland School of Medicine, Baltimore, MD, USA
| | - Guoli Shi
- Department of Orthopedics, University of Maryland School of Medicine, Baltimore, MD, USA
| | - Christopher W Ward
- Department of Orthopedics, University of Maryland School of Medicine, Baltimore, MD, USA
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Redwan A, Kiriaev L, Kueh S, Morley JW, Houweling P, Perry BD, Head SI. Six weeks of N-acetylcysteine antioxidant in drinking water decreases pathological fiber branching in MDX mouse dystrophic fast-twitch skeletal muscle. Front Physiol 2023; 14:1109587. [PMID: 36866174 PMCID: PMC9971923 DOI: 10.3389/fphys.2023.1109587] [Citation(s) in RCA: 8] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/28/2022] [Accepted: 02/03/2023] [Indexed: 02/16/2023] Open
Abstract
Introduction: It has been proposed that an increased susceptivity to oxidative stress caused by the absence of the protein dystrophin from the inner surface of the sarcolemma is a trigger of skeletal muscle necrosis in the destructive dystrophin deficient muscular dystrophies. Here we use the mdx mouse model of human Duchenne Muscular Dystrophy to test the hypothesis that adding the antioxidant NAC at 2% to drinking water for six weeks will treat the inflammatory phase of the dystrophic process and reduce pathological muscle fiber branching and splitting resulting in a reduction of mass in mdx fast-twitch EDL muscles. Methods: Animal weight and water intake was recorded during the six weeks when 2% NAC was added to the drinking water. Post NAC treatment animals were euthanised and the EDL muscles dissected out and placed in an organ bath where the muscle was attached to a force transducer to measure contractile properties and susceptibility to force loss from eccentric contractions. After the contractile measurements had been made the EDL muscle was blotted and weighed. In order to assess the degree of pathological fiber branching mdx EDL muscles were treated with collagenase to release single fibers. For counting and morphological analysis single EDL mdx skeletal muscle fibers were viewed under high magnification on an inverted microscope. Results: During the six-week treatment phase NAC reduced body weight gain in three- to nine-week-old mdx and littermate control mice without effecting fluid intake. NAC treatment also significantly reduced the mdx EDL muscle mass and abnormal fiber branching and splitting. Discussion: We propose chronic NAC treatment reduces the inflammatory response and degenerative cycles in the mdx dystrophic EDL muscles resulting in a reduction in the number of complexed branched fibers reported to be responsible for the dystrophic EDL muscle hypertrophy.
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Affiliation(s)
- Asma Redwan
- School of Medicine, Western Sydney University, Sydney, NSW, Australia
| | - Leonit Kiriaev
- Murdoch Children’s Research Institute, Melbourne, VIC, Australia
| | - Sindy Kueh
- School of Medicine, Western Sydney University, Sydney, NSW, Australia
| | - John W. Morley
- School of Medicine, Western Sydney University, Sydney, NSW, Australia
| | - Peter Houweling
- Murdoch Children’s Research Institute, Melbourne, VIC, Australia
| | - Ben D. Perry
- School of Science, Western Sydney University, Sydney, NSW, Australia
| | - Stewart I. Head
- School of Medicine, Western Sydney University, Sydney, NSW, Australia,Murdoch Children’s Research Institute, Melbourne, VIC, Australia,*Correspondence: Stewart I. Head,
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Kiriaev L, Houweling PJ, North KN, Head SI. Loss of α-actinin-3 confers protection from eccentric contraction damage in fast-twitch EDL muscles from aged mdx dystrophic mice by reducing pathological fibre branching. Hum Mol Genet 2022; 31:1417-1429. [PMID: 34761268 PMCID: PMC9071495 DOI: 10.1093/hmg/ddab326] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/27/2021] [Revised: 10/27/2021] [Accepted: 11/02/2021] [Indexed: 11/14/2022] Open
Abstract
The common null polymorphism (R577X) in the ACTN3 gene is present in over 1.5 billion people worldwide and results in the absence of the protein α-actinin-3 from the Z-discs of fast-twitch skeletal muscle fibres. We have previously reported that this polymorphism is a modifier of dystrophin-deficient Duchenne Muscular Dystrophy. To investigate the mechanism underlying this, we use a double knockout (dk)Actn3KO/mdx (dKO) mouse model, which lacks both dystrophin and sarcomere α-actinin-3. We used dKO mice and mdx dystrophic mice at 12 months (aged) to investigate the correlation between morphological changes to the fast-twitch dKO EDL and the reduction in force deficit produced by an in vitro eccentric contraction protocol. In the aged dKO mouse, we found a marked reduction in fibre branching complexity that correlated with protection from eccentric contraction induced force deficit. Complex branches in the aged dKO EDL fibres (28%) were substantially reduced compared to aged mdx EDL fibres (68%), and this correlates with a graded force loss over three eccentric contractions for dKO muscles (~36% after first contraction, ~66% overall) compared to an abrupt drop in mdx upon the first eccentric contraction (~75% after first contraction, ~89% after three contractions). In dKO, protection from eccentric contraction damage was linked with a doubling of SERCA1 pump density the EDL. We propose that the increased oxidative metabolism of fast-twitch glycolytic fibres characteristic of the null polymorphism (R577X) and increase in SR Ca2+ pump proteins reduces muscle fibre branching and decreases susceptibility to eccentric injury in the dystrophinopathies.
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Affiliation(s)
- Leonit Kiriaev
- School of Medicine, Western Sydney University, Sydney, NSW 2560, Australia
| | - Peter J Houweling
- Murdoch Children’s Research Institute, Melbourne, Victoria 3052, Australia
- Department of Paediatrics, University of Melbourne, Melbourne, Victoria 3052, Australia
| | - Kathryn N North
- Murdoch Children’s Research Institute, Melbourne, Victoria 3052, Australia
- Department of Paediatrics, University of Melbourne, Melbourne, Victoria 3052, Australia
| | - Stewart I Head
- School of Medicine, Western Sydney University, Sydney, NSW 2560, Australia
- Murdoch Children’s Research Institute, Melbourne, Victoria 3052, Australia
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Swain SM, Romac JMJ, Vigna SR, Liddle RA. Piezo1-mediated stellate cell activation causes pressure-induced pancreatic fibrosis in mice. JCI Insight 2022; 7:158288. [PMID: 35451372 PMCID: PMC9089793 DOI: 10.1172/jci.insight.158288] [Citation(s) in RCA: 40] [Impact Index Per Article: 13.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/10/2022] [Accepted: 03/02/2022] [Indexed: 11/17/2022] Open
Abstract
Pancreatic fibrosis is a complication of chronic pancreatitis and is a prominent feature of pancreatic cancer. Pancreatic fibrosis is commonly observed in patients with prolonged pancreatic duct obstruction, which elevates intrapancreatic pressure. We show here that increased pancreatic duct pressure causes fibrosis and describes the mechanism by which pressure increases deposition of extracellular matrix proteins and fibrosis. We found that pancreatic stellate cells (PSCs), the source of the extracellular matrix proteins in fibrosis, express the mechanically activated ion channel Piezo1. By increasing intracellular calcium, mechanical stress or the Piezo1 agonist Yoda1-activated PSCs manifest by loss of perinuclear fat droplets and increased TGF-β1, fibronectin, and type I collagen expression. These effects were blocked by the Piezo1 inhibitor GsMTx4 and absent in PSCs from mice with conditional genetic deletion of Piezo1 in stellate cells, as was pancreatic duct ligation-induced fibrosis. Although TRPV4 has been proposed to have direct mechanosensing properties, we discovered that PSCs from Trpv4-KO mice were protected against Yoda1-triggered activation. Moreover, mice devoid of TRPV4 were protected from pancreatic duct ligation-induced fibrosis. Thus, high pressure within the pancreas stimulates Piezo1 channel opening, and subsequent activation of TRPV4 leads to stellate cell activation and pressure-induced chronic pancreatitis and fibrosis.
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Affiliation(s)
- Sandip M Swain
- Department of Medicine, Duke University, Durham, North Carolina, USA
| | - Joelle M-J Romac
- Department of Medicine, Duke University, Durham, North Carolina, USA
| | - Steven R Vigna
- Department of Medicine, Duke University, Durham, North Carolina, USA
| | - Rodger A Liddle
- Department of Medicine, Duke University, Durham, North Carolina, USA.,Department of Veterans Affairs Healthcare System, Durham, North Carolina, USA
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7
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Kiriaev L, Kueh S, Morley JW, North KN, Houweling PJ, Head SI. Lifespan Analysis of Dystrophic mdx Fast-Twitch Muscle Morphology and Its Impact on Contractile Function. Front Physiol 2021; 12:771499. [PMID: 34950049 PMCID: PMC8689589 DOI: 10.3389/fphys.2021.771499] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/06/2021] [Accepted: 11/08/2021] [Indexed: 11/13/2022] Open
Abstract
Duchenne muscular dystrophy is caused by the absence of the protein dystrophin from skeletal muscle and is characterized by progressive cycles of necrosis/regeneration. Using the dystrophin deficient mdx mouse model, we studied the morphological and contractile chronology of dystrophic skeletal muscle pathology in fast-twitch Extensor Digitorum Longus muscles from animals 4–22 months of age containing 100% regenerated muscle fibers. Catastrophically, the older age groups lost ∼80% of their maximum force after one eccentric contraction (EC) of 20% strain with the greatest loss of ∼92% recorded in senescent 22-month-old mdx mice. In old age groups, there was minimal force recovery ∼24% after 120 min, correlated with a dramatic increase in the number and complexity of branched fibers. This data supports our two-phase model where a “tipping point” is reached when branched fibers rupture irrevocably on EC. These findings have important implications for pre-clinical drug studies and genetic rescue strategies.
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Affiliation(s)
- Leonit Kiriaev
- Myogenica Laboratory, School of Medicine, Western Sydney University, Sydney, NSW, Australia
- *Correspondence: Leonit Kiriaev,
| | - Sindy Kueh
- Myogenica Laboratory, School of Medicine, Western Sydney University, Sydney, NSW, Australia
| | - John W. Morley
- Myogenica Laboratory, School of Medicine, Western Sydney University, Sydney, NSW, Australia
| | - Kathryn N. North
- Muscle Research Group, Murdoch Children’s Research Institute, Melbourne, VIC, Australia
| | - Peter J. Houweling
- Muscle Research Group, Murdoch Children’s Research Institute, Melbourne, VIC, Australia
| | - Stewart I. Head
- Myogenica Laboratory, School of Medicine, Western Sydney University, Sydney, NSW, Australia
- Muscle Research Group, Murdoch Children’s Research Institute, Melbourne, VIC, Australia
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Bigford GE, Donovan A, Webster MT, Dietrich WD, Nash MS. Selective Myostatin Inhibition Spares Sublesional Muscle Mass and Myopenia-Related Dysfunction after Severe Spinal Cord Contusion in Mice. J Neurotrauma 2021; 38:3440-3455. [PMID: 34714134 DOI: 10.1089/neu.2021.0061] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/25/2022] Open
Abstract
Clinically relevant myopenia accompanies spinal cord injury (SCI), and compromises function, metabolism, body composition, and health. Myostatin, a transforming growth factor (TGF)β family member, is a key negative regulator of skeletal muscle mass. We investigated inhibition of myostatin signaling using systemic delivery of a highly selective monoclonal antibody - muSRK-015P (40 mg/kg) - that blocks release of active growth factor from the latent form of myostatin. Adult female mice (C57BL/6) were subjected to a severe SCI (65 kdyn) at T9 and were then immediately and 1 week later administered test articles: muSRK-015P (40 mg/kg) or control (vehicle or IgG). A sham control group (laminectomy only) was included. At euthanasia, (2 weeks post-SCI) muSRK-015P preserved whole body lean mass and sublesional gastrocnemius and soleus mass. muSRK-015P-treated mice with SCI also had significantly attenuated myofiber atrophy, lipid infiltration, and loss of slow-oxidative phenotype in soleus muscle. These outcomes were accompanied by significantly improved sublesional motor function and muscle force production at 1 and 2 weeks post-SCI. At 2 weeks post-SCI, lean mass was significantly decreased in SCI-IgG mice, but was not different in SCI-muSRK-015P mice than in sham controls. Total energy expenditure (kCal/day) at 2 weeks post-SCI was lower in SCI-immunoglobulin (Ig)G mice, but not different in SCI-muSRK-015P mice than in sham controls. We conclude that in a randomized, blinded, and controlled study in mice, myostatin inhibition using muSRK-015P had broad effects on physical, metabolic, and functional outcomes when compared with IgG control treated SCI animals. These findings may identify a useful, targeted therapeutic strategy for treating post-SCI myopenia and related sequelae in humans.
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Affiliation(s)
- Gregory E Bigford
- Department of Neurological Surgery and the Miami Project to Cure Paralysis, University of Miami Miller School of Medicine, Miami, Florida, USA
| | | | | | - W Dalton Dietrich
- Department of Neurological Surgery and the Miami Project to Cure Paralysis, University of Miami Miller School of Medicine, Miami, Florida, USA
| | - Mark S Nash
- Department of Neurological Surgery and the Miami Project to Cure Paralysis, University of Miami Miller School of Medicine, Miami, Florida, USA.,Department of Physical Medicine and Rehabilitation, University of Miami Miller School of Medicine, Miami, Florida, USA.,Department of Physical Therapy, University of Miami, Miami, Florida, USA
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Boycott HE, Nguyen MN, Vrellaku B, Gehmlich K, Robinson P. Nitric Oxide and Mechano-Electrical Transduction in Cardiomyocytes. Front Physiol 2020; 11:606740. [PMID: 33384614 PMCID: PMC7770138 DOI: 10.3389/fphys.2020.606740] [Citation(s) in RCA: 17] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/15/2020] [Accepted: 11/23/2020] [Indexed: 12/22/2022] Open
Abstract
The ability§ of the heart to adapt to changes in the mechanical environment is critical for normal cardiac physiology. The role of nitric oxide is increasingly recognized as a mediator of mechanical signaling. Produced in the heart by nitric oxide synthases, nitric oxide affects almost all mechano-transduction pathways within the cardiomyocyte, with roles mediating mechano-sensing, mechano-electric feedback (via modulation of ion channel activity), and calcium handling. As more precise experimental techniques for applying mechanical stresses to cells are developed, the role of these forces in cardiomyocyte function can be further understood. Furthermore, specific inhibitors of different nitric oxide synthase isoforms are now available to elucidate the role of these enzymes in mediating mechano-electrical signaling. Understanding of the links between nitric oxide production and mechano-electrical signaling is incomplete, particularly whether mechanically sensitive ion channels are regulated by nitric oxide, and how this affects the cardiac action potential. This is of particular relevance to conditions such as atrial fibrillation and heart failure, in which nitric oxide production is reduced. Dysfunction of the nitric oxide/mechano-electrical signaling pathways are likely to be a feature of cardiac pathology (e.g., atrial fibrillation, cardiomyopathy, and heart failure) and a better understanding of the importance of nitric oxide signaling and its links to mechanical regulation of heart function may advance our understanding of these conditions.
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Affiliation(s)
- Hannah E. Boycott
- Division of Cardiovascular Medicine, Radcliffe Department of Medicine, University of Oxford, John Radcliffe Hospital, Oxford, United Kingdom
- Division of Cardiovascular Medicine, Radcliffe Department of Medicine and British Heart Foundation Centre of Research Excellence Oxford, University of Oxford, Oxford, United Kingdom
| | - My-Nhan Nguyen
- Division of Cardiovascular Medicine, Radcliffe Department of Medicine, University of Oxford, John Radcliffe Hospital, Oxford, United Kingdom
- Division of Cardiovascular Medicine, Radcliffe Department of Medicine and British Heart Foundation Centre of Research Excellence Oxford, University of Oxford, Oxford, United Kingdom
| | - Besarte Vrellaku
- Division of Cardiovascular Medicine, Radcliffe Department of Medicine, University of Oxford, John Radcliffe Hospital, Oxford, United Kingdom
- Division of Cardiovascular Medicine, Radcliffe Department of Medicine and British Heart Foundation Centre of Research Excellence Oxford, University of Oxford, Oxford, United Kingdom
| | - Katja Gehmlich
- Division of Cardiovascular Medicine, Radcliffe Department of Medicine, University of Oxford, John Radcliffe Hospital, Oxford, United Kingdom
- Division of Cardiovascular Medicine, Radcliffe Department of Medicine and British Heart Foundation Centre of Research Excellence Oxford, University of Oxford, Oxford, United Kingdom
- Institute of Cardiovascular Sciences, University of Birmingham, Birmingham, United Kingdom
| | - Paul Robinson
- Division of Cardiovascular Medicine, Radcliffe Department of Medicine, University of Oxford, John Radcliffe Hospital, Oxford, United Kingdom
- Division of Cardiovascular Medicine, Radcliffe Department of Medicine and British Heart Foundation Centre of Research Excellence Oxford, University of Oxford, Oxford, United Kingdom
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10
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Nakada S, Yamashita Y, Machida S, Miyagoe-Suzuki Y, Arikawa-Hirasawa E. Perlecan Facilitates Neuronal Nitric Oxide Synthase Delocalization in Denervation-Induced Muscle Atrophy. Cells 2020; 9:cells9112524. [PMID: 33238404 PMCID: PMC7700382 DOI: 10.3390/cells9112524] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/08/2020] [Revised: 11/17/2020] [Accepted: 11/19/2020] [Indexed: 01/01/2023] Open
Abstract
Perlecan is an extracellular matrix molecule anchored to the sarcolemma by a dystrophin–glycoprotein complex. Perlecan-deficient mice are tolerant to muscle atrophy, suggesting that perlecan negatively regulates mechanical stress-dependent skeletal muscle mass. Delocalization of neuronal nitric oxide synthase (nNOS) from the sarcolemma to the cytosol triggers protein degradation, thereby initiating skeletal muscle atrophy. We hypothesized that perlecan regulates nNOS delocalization and activates protein degradation during this process. To determine the role of perlecan in nNOS-mediated mechanotransduction, we used sciatic nerve transection as a denervation model of gastrocnemius muscles. Gastrocnemius muscle atrophy was significantly lower in perinatal lethality-rescued perlecan-knockout (Hspg2−/−-Tg) mice than controls (WT-Tg) on days 4 and 14 following surgery. Immunofluorescence microscopy showed that cell membrane nNOS expression was reduced by denervation in WT-Tg mice, with marginal effects in Hspg2−/−-Tg mice. Moreover, levels of atrophy-related proteins—i.e., FoxO1a, FoxO3a, atrogin-1, and Lys48-polyubiquitinated proteins—increased in the denervated muscles of WT-Tg mice but not in Hspg2−/−-Tg mice. These findings suggest that during denervation, perlecan promotes nNOS delocalization from the membrane and stimulates protein degradation and muscle atrophy by activating FoxO signaling and the ubiquitin–proteasome system.
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Affiliation(s)
- Satoshi Nakada
- Japanese Center for Research on Women in Sport, Juntendo University Graduate School of Health and Sports Science, Chiba 270-1695, Japan; (S.N.); (S.M.)
| | - Yuri Yamashita
- Research Institute for Diseases of Old Age, Juntendo University Graduate School of Medicine, Tokyo 113-8421, Japan;
| | - Shuichi Machida
- Japanese Center for Research on Women in Sport, Juntendo University Graduate School of Health and Sports Science, Chiba 270-1695, Japan; (S.N.); (S.M.)
| | - Yuko Miyagoe-Suzuki
- Department of Molecular Therapy, National Institute of Neuroscience, National Center of Neurology and Psychiatry, Tokyo 187-8502, Japan;
| | - Eri Arikawa-Hirasawa
- Japanese Center for Research on Women in Sport, Juntendo University Graduate School of Health and Sports Science, Chiba 270-1695, Japan; (S.N.); (S.M.)
- Research Institute for Diseases of Old Age, Juntendo University Graduate School of Medicine, Tokyo 113-8421, Japan;
- Correspondence: ; Tel.: +81-3-3813-3111
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Garbincius JF, Merz LE, Cuttitta AJ, Bayne KV, Schrade S, Armstead EA, Converso-Baran KL, Whitesall SE, D'Alecy LG, Michele DE. Enhanced dimethylarginine degradation improves coronary flow reserve and exercise tolerance in Duchenne muscular dystrophy carrier mice. Am J Physiol Heart Circ Physiol 2020; 319:H582-H603. [PMID: 32762558 DOI: 10.1152/ajpheart.00333.2019] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 01/16/2023]
Abstract
Duchenne muscular dystrophy (DMD) is an X-linked disease caused by null mutations in dystrophin and characterized by muscle degeneration. Cardiomyopathy is common and often prevalent at similar frequency in female DMD carriers irrespective of whether they manifest skeletal muscle disease. Impaired muscle nitric oxide (NO) production in DMD disrupts muscle blood flow regulation and exaggerates postexercise fatigue. We show that circulating levels of endogenous methylated arginines including asymmetric dimethylarginine (ADMA), which act as NO synthase inhibitors, are elevated by acute necrotic muscle damage and in chronically necrotic dystrophin-deficient mice. We therefore hypothesized that excessive ADMA impairs muscle NO production and diminishes exercise tolerance in DMD. We used transgenic expression of dimethylarginine dimethylaminohydrolase 1 (DDAH), which degrades methylated arginines, to investigate their contribution to exercise-induced fatigue in DMD. Although infusion of exogenous ADMA was sufficient to impair exercise performance in wild-type mice, transgenic DDAH expression did not rescue exercise-induced fatigue in dystrophin-deficient male mdx mice. Surprisingly, DDAH transgene expression did attenuate exercise-induced fatigue in dystrophin-heterozygous female mdx carrier mice. Improved exercise tolerance was associated with reduced heart weight and improved cardiac β-adrenergic responsiveness in DDAH-transgenic mdx carriers. We conclude that DDAH overexpression increases exercise tolerance in female DMD carriers, possibly by limiting cardiac pathology and preserving the heart's responses to changes in physiological demand. Methylated arginine metabolism may be a new target to improve exercise tolerance and cardiac function in DMD carriers or act as an adjuvant to promote NO signaling alongside therapies that partially restore dystrophin expression in patients with DMD.NEW & NOTEWORTHY Duchenne muscular dystrophy (DMD) carriers are at risk for cardiomyopathy. The nitric oxide synthase inhibitor asymmetric dimethylarginine (ADMA) is released from damaged muscle in DMD and impairs exercise performance. Transgenic expression of dimethylarginine dimethylaminohydrolase to degrade ADMA prevents cardiac hypertrophy, improves cardiac function, and improves exercise tolerance in DMD carrier mice. These findings highlight the relevance of ADMA to muscular dystrophy and have important implications for therapies targeting nitric oxide in patients with DMD and DMD carriers.
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Affiliation(s)
- Joanne F Garbincius
- Department of Molecular and Integrative Physiology, University of Michigan, Ann Arbor, Michigan
| | - Lauren E Merz
- Department of Molecular and Integrative Physiology, University of Michigan, Ann Arbor, Michigan
| | - Ashley J Cuttitta
- Department of Molecular and Integrative Physiology, University of Michigan, Ann Arbor, Michigan
| | - Kaitlynn V Bayne
- Department of Molecular and Integrative Physiology, University of Michigan, Ann Arbor, Michigan
| | - Sara Schrade
- Department of Molecular and Integrative Physiology, University of Michigan, Ann Arbor, Michigan
| | - Emily A Armstead
- Department of Molecular and Integrative Physiology, University of Michigan, Ann Arbor, Michigan
| | | | - Steven E Whitesall
- Department of Molecular and Integrative Physiology, University of Michigan, Ann Arbor, Michigan.,Physiology Phenotyping Core, University of Michigan, Ann Arbor, Michigan
| | - Louis G D'Alecy
- Department of Molecular and Integrative Physiology, University of Michigan, Ann Arbor, Michigan.,Physiology Phenotyping Core, University of Michigan, Ann Arbor, Michigan
| | - Daniel E Michele
- Department of Molecular and Integrative Physiology, University of Michigan, Ann Arbor, Michigan.,Physiology Phenotyping Core, University of Michigan, Ann Arbor, Michigan.,Department of Internal Medicine, University of Michigan, Ann Arbor, Michigan
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12
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Long MH, Zhu XM, Wang Q, Chen Y, Gan XD, Li F, Fu WL, Xing WW, Xu DQ, Xu DG. PM2.5 exposure induces vascular dysfunction via NO generated by iNOS in lung of ApoE-/- mouse. Int J Biol Sci 2020; 16:49-60. [PMID: 31892845 PMCID: PMC6930374 DOI: 10.7150/ijbs.36073] [Citation(s) in RCA: 23] [Impact Index Per Article: 4.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/25/2019] [Accepted: 08/06/2019] [Indexed: 12/20/2022] Open
Abstract
PM2.5 exposure exacerbates cardiovascular diseases via oxidative stress and inflammation, the detailed mechanism of which is unclear. In this study, the effects of oxidative stress and inflammation, as well as vascular structure and function were studied by multiple PM2.5 exposure model of ApoE-/- mice. The results indicated that NO produced by iNOS not cNOS might play important roles in inducing vascular dysfunction after PM2.5 exposure. The occurrence order and causality among NO, other oxidative stress indicators and inflammation is explored by single PM2.5 exposure. The results showed that NO generated by iNOS occurred earlier than that of other oxidative stress indicators, which was followed by the increased inflammation. Inhibition of NOS could effectively block the raise of NO, oxidative stress and inflammation after PM2.5 exposure. All in all, we firstly confirmed that NO was the initiation factor of PM2.5 exposure-induced oxidative stress, which led to inflammation and the following vascular dysfunction.
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Affiliation(s)
- Min-Hui Long
- Institute of Military Cognitive and Brain Sciences, Beijing, 100850, China
| | - Xiao-Ming Zhu
- Institute of Military Cognitive and Brain Sciences, Beijing, 100850, China
| | - Qin Wang
- National Institute of Environmental Health Chinese Center for Disease Control and Prevention, Beijing, 100021, China
| | - Yao Chen
- Institute of Military Cognitive and Brain Sciences, Beijing, 100850, China
| | - Xiang-Dong Gan
- Institute of Military Cognitive and Brain Sciences, Beijing, 100850, China
| | - Fei Li
- Institute of Military Cognitive and Brain Sciences, Beijing, 100850, China
| | - Wen-Liang Fu
- Institute of Military Cognitive and Brain Sciences, Beijing, 100850, China
| | - Wei-Wei Xing
- Institute of Military Cognitive and Brain Sciences, Beijing, 100850, China
| | - Dong-Qun Xu
- National Institute of Environmental Health Chinese Center for Disease Control and Prevention, Beijing, 100021, China
| | - Dong-Gang Xu
- Institute of Military Cognitive and Brain Sciences, Beijing, 100850, China
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13
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Boehler JF, Ricotti V, Gonzalez JP, Soustek-Kramer M, Such L, Brown KJ, Schneider JS, Morris CA. Membrane recruitment of nNOSµ in microdystrophin gene transfer to enhance durability. Neuromuscul Disord 2019; 29:735-741. [PMID: 31521486 DOI: 10.1016/j.nmd.2019.08.009] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/05/2019] [Revised: 07/23/2019] [Accepted: 07/31/2019] [Indexed: 12/25/2022]
Abstract
Several gene transfer clinical trials are currently ongoing with the common aim of delivering a shortened version of dystrophin, termed a microdystrophin, for the treatment of Duchenne muscular dystrophy (DMD). However, one of the main differences between these trials is the microdystrophin protein produced following treatment. Each gene transfer product is based on different selections of dystrophin domain combinations to assemble microdystrophin transgenes that maintain functional dystrophin domains and fit within the packaging limits of an adeno-associated virus (AAV) vector. While domains involved in mechanical function, such as the actin-binding domain and β-dystroglycan binding domain, have been identified for many years and included in microdystrophin constructs, more recently the neuronal nitric oxide synthase (nNOS) domain has also been identified due to its role in enhancing nNOS membrane localization. As nNOS membrane localization has been established as an important requirement for prevention of functional ischemia in skeletal muscle, inclusion of the nNOS domain into a microdystrophin construct represents an important consideration. The aim of this mini review is to highlight what is currently known about the nNOS domain of dystrophin and to describe potential implications of this domain in a microdystrophin gene transfer clinical trial.
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Affiliation(s)
- Jessica F Boehler
- Solid Biosciences, 141 Portland Street, Cambridge, MA 02139, United States
| | - Valeria Ricotti
- Solid Biosciences, 141 Portland Street, Cambridge, MA 02139, United States
| | - J Patrick Gonzalez
- Solid Biosciences, 141 Portland Street, Cambridge, MA 02139, United States
| | | | - Lauren Such
- Solid Biosciences, 141 Portland Street, Cambridge, MA 02139, United States
| | - Kristy J Brown
- Solid Biosciences, 141 Portland Street, Cambridge, MA 02139, United States
| | - Joel S Schneider
- Solid Biosciences, 141 Portland Street, Cambridge, MA 02139, United States
| | - Carl A Morris
- Solid Biosciences, 141 Portland Street, Cambridge, MA 02139, United States.
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14
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Loyer X, Dubroca C, Branchereau M, Griffith G, Garcia L, Heymes C. Neuronal NO synthase mediates plenylephrine induced cardiomyocyte hypertrophy through facilitation of NFAT-dependent transcriptional activity. Biochem Biophys Rep 2019; 18:100620. [PMID: 30899802 PMCID: PMC6412025 DOI: 10.1016/j.bbrep.2019.100620] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/21/2018] [Revised: 01/30/2019] [Accepted: 02/15/2019] [Indexed: 11/19/2022] Open
Abstract
Neuronal nitric oxide synthase (NOS1) has been consistently shown to be the predominant isoform of NOS and/or NOS-derived NO that may be involved in the myocardial remodeling including cardiac hypertrophy. However, the direct functional contribution of NOS1 in this process remains to be elucidated. Therefore, in the present study, we attempted to use silent RNA and adenovirus mediated silencing or overexpression to investigate the role of NOS1 and the associated molecular signaling mechanisms during OKphenylephrine (PE)-induced cardiac hypertrophy growth in neonatal rat ventricular cardiomyocytes (NRVMs). We found that the expression of NOS1 was enhanced in PE-induced hypertrophic cardiomyocytes. Moreover, LVNIO treatment, a selective NOS1 inhibitor, significantly decreased PE-induced NRVMs hypertrophy and [3H]-leucine incorporation. We demonstrated that NOS1 gene silencing attenuated both the increased size and the transcriptional activity of the hypertrophic marker atrial natriuretic factor (ANF) induced by PE stimulation. Further investigation suggested that deficiency of NOS1-induced diminished NRVMS hypertrophy resulted in decreased calcineurin protein expression and activity (assessed by measuring the transcriptional activity of NFAT) and, an increased activity of the anti-hypertrophic pathway, GSK-3β (estimated by its augmented phosphorylated level). In contrast, exposing the NOS1 overexpressed NRVMs to PE-treatment further increased the hypertrophic growth, ANF transcriptional activity and calcineurin activity. Together, the results of the present study suggest that NOS1 is directly involved in controlling the development of cardiomyocyte hypertrophy.
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Affiliation(s)
- Xavier Loyer
- INSERM UMR-S 970, Paris Cardiovascular Research Center, France
- Université Paris Descartes, Sorbonne Paris Cité, France
| | - Caroline Dubroca
- INSERM U1048 -Institut des Maladies Métaboliques et Cardiovacsulaires (I2MC), Toulouse, France
| | - Maxime Branchereau
- INSERM U1048 -Institut des Maladies Métaboliques et Cardiovacsulaires (I2MC), Toulouse, France
- Université Paul Sabatier, Toulouse, France
| | - Graziellia Griffith
- Université de Versailles St-Quentin, INSERM U1179, Montigny-le-Bretonneux, France
| | - Luis Garcia
- Université de Versailles St-Quentin, INSERM U1179, Montigny-le-Bretonneux, France
| | - Christophe Heymes
- INSERM U1048 -Institut des Maladies Métaboliques et Cardiovacsulaires (I2MC), Toulouse, France
- Université Paul Sabatier, Toulouse, France
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15
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Batra A, Vohra RS, Chrzanowski SM, Hammers DW, Lott DJ, Vandenborne K, Walter GA, Forbes SC. Effects of PDE5 inhibition on dystrophic muscle following an acute bout of downhill running and endurance training. J Appl Physiol (1985) 2019; 126:1737-1745. [PMID: 30946638 DOI: 10.1152/japplphysiol.00664.2018] [Citation(s) in RCA: 12] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/22/2022] Open
Abstract
Lack of sarcolemma-localized neuronal nitric oxide synthase mu (nNOSμ) contributes to muscle damage and fatigue in dystrophic muscle. In this study, we examined the effects of compensating for lack of nNOSμ with a phosphodiesterase type 5 (PDE5) inhibitor in mdx mice following downhill running and endurance training. Dystrophic mice (mdx) were treated with sildenafil citrate and compared with untreated mdx and wild-type mice after an acute bout of downhill running and during a progressive low-intensity treadmill running program (5 days/wk, 4 wk). Magnetic resonance imaging (MRI) and spectroscopy (MRS) transverse relaxation time constant (T2) of hindlimb and forelimb muscles were measured as a marker of muscle damage after downhill running and throughout training. The MRI blood oxygenation level dependence (BOLD) response and 31phosphorus MRS (31P-MRS) data were acquired after stimulated muscle contractions. After downhill running, the increase in T2 was attenuated (P < 0.05) in treated mdx and wild-type mice compared with untreated mdx. During training, resting T2 values did not change in wild-type and mdx mice from baseline values; however, the running distance completed during training was greater (P < 0.05) in treated mdx (>90% of target distance) and wild-type (100%) than untreated mdx (60%). The post-contractile BOLD response was greater (P < 0.05) in treated mdx that trained than untreated mdx, with no differences in muscle oxidative capacity, as measured by 31P-MRS. Our findings indicate that PDE5 inhibition reduces muscle damage after a single bout of downhill running and improves performance during endurance training in dystrophic mice, possibly because of enhanced microvascular function. NEW & NOTEWORTHY This study examined the combined effects of PDE5 inhibition and exercise in dystrophic muscle using high-resolution magnetic resonance imaging and spectroscopy. Our findings demonstrated that sildenafil citrate reduces muscle damage after a single bout of downhill running, improves endurance-training performance, and enhances microvascular function in dystrophic muscle. Collectively, the results support the combination of exercise and PDE5 inhibition as a therapeutic approach in muscular dystrophies lacking nNOSμ.
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Affiliation(s)
- Abhinandan Batra
- Department of Physical Therapy, University of Florida , Gainesville, Florida
| | - Ravneet S Vohra
- Department of Physical Therapy, University of Florida , Gainesville, Florida
| | - Steve M Chrzanowski
- Department of Physiology and Therapeutics, University of Florida , Gainesville, Florida
| | - David W Hammers
- Department of Pharmacology and Functional Genomics, University of Florida, Gainesville, Florida
| | - Donovan J Lott
- Department of Physical Therapy, University of Florida , Gainesville, Florida
| | - Krista Vandenborne
- Department of Physical Therapy, University of Florida , Gainesville, Florida
| | - Glenn A Walter
- Department of Physiology and Therapeutics, University of Florida , Gainesville, Florida
| | - Sean C Forbes
- Department of Physical Therapy, University of Florida , Gainesville, Florida
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16
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Balke JE, Zhang L, Percival JM. Neuronal nitric oxide synthase (nNOS) splice variant function: Insights into nitric oxide signaling from skeletal muscle. Nitric Oxide 2018; 82:35-47. [PMID: 30503614 DOI: 10.1016/j.niox.2018.11.004] [Citation(s) in RCA: 25] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/14/2018] [Revised: 11/19/2018] [Accepted: 11/20/2018] [Indexed: 02/07/2023]
Abstract
Defects in neuronal nitric oxide synthase (nNOS) splice variant localization and signaling in skeletal muscle are a firmly established pathogenic characteristic of many neuromuscular diseases, including Duchenne and Becker muscular dystrophy (DMD and BMD, respectively). Therefore, substantial efforts have been made to understand and therapeutically target skeletal muscle nNOS isoform signaling. The purpose of this review is to summarize recent salient advances in understanding of the regulation, targeting, and function of nNOSμ and nNOSβ splice variants in normal and dystrophic skeletal muscle, primarily using findings from mouse models. The first focus of this review is how the differential targeting of nNOS splice variants creates spatially and functionally distinct nitric oxide (NO) signaling compartments at the sarcolemma, Golgi complex, and cytoplasm. Particular attention is given to the functions of sarcolemmal nNOSμ and limitations of current nNOS knockout models. The second major focus is to review current understanding of cGMP-mediated nNOS signaling in skeletal muscle and its emergence as a therapeutic target in DMD and BMD. Accordingly, we address the preclinical and clinical successes and setbacks with the testing of phosphodiesterase 5 inhibitors to redress nNOS signaling defects in DMD and BMD. In summary, this review of nNOS function in normal and dystrophic muscle aims to advance understanding how the messenger NO is harnessed for cellular signaling from a skeletal muscle perspective.
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Affiliation(s)
- Jordan E Balke
- Department of Molecular and Cellular Pharmacology, University of Miami Miller School of Medicine Miami, Florida, 33101, USA
| | - Ling Zhang
- Department of Molecular and Cellular Pharmacology, University of Miami Miller School of Medicine Miami, Florida, 33101, USA
| | - Justin M Percival
- Department of Molecular and Cellular Pharmacology, University of Miami Miller School of Medicine Miami, Florida, 33101, USA.
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17
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Percival JM. Perspective: Spectrin-Like Repeats in Dystrophin Have Unique Binding Preferences for Syntrophin Adaptors That Explain the Mystery of How nNOSμ Localizes to the Sarcolemma. Front Physiol 2018; 9:1369. [PMID: 30349485 PMCID: PMC6186803 DOI: 10.3389/fphys.2018.01369] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/21/2018] [Accepted: 09/10/2018] [Indexed: 11/13/2022] Open
Abstract
Dystrophin is a massive multi-domain protein composed of specialized amino and carboxyl termini that are separated by 24 spectrin-like repeats. Dystrophin performs critical structural and signaling roles that are indispensable for the functional integrity of skeletal muscle. Indeed, the loss of dystrophin protein expression causes the muscle wasting disease, Duchenne muscular dystrophy (DMD). Substantial progress has been made in defining the functions of the domains of dystrophin, which has proven invaluable for the development of miniaturized dystrophin gene and exon skipping therapies for DMD. However, a long-standing mystery regarding dystrophin function is how dystrophin, and its adaptor and neuronal nitric oxide synthase mu (nNOSμ) binding partner α-syntrophin, cooperate to localize nNOSμ to the sarcolemma. Only when localized to the sarcolemma can nNOSμ override sympathetic vasoconstriction and prevent functional ischemia in contracting muscles. Current evidence suggests that spectrin-like repeat 17 of dystrophin and α-syntrophin cooperate to localize nNOSμ to the sarcolemma. However, the exact mechanism remains unclear and controversial because of equivocal evidence for direct binding of dystrophin and nNOSμ. Recently, an important study identified a novel α-syntrophin binding site within spectrin-like repeat 17, leading to a new model whereby α-syntrophin recruits nNOSμ to the sarcolemmal dystrophin complex by binding spectrin-like repeat 17. This model finally appears to solve the mystery of the dual requirement for dystrophin and α-syntrophin for sarcolemmal nNOSμ localization. The aim of the current perspective is to highlight this major advance in understanding of dystrophin’s role in localizing nNOSμ and its implications for current trials.
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Affiliation(s)
- Justin M Percival
- Department of Molecular and Cellular Pharmacology, University of Miami Miller School of Medicine, Miami, FL, United States
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18
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Todd D, Bonthius DJ, Sabalo LM, Roghair J, Karacay B, Bousquet SL, Bonthius DJ. Regional Patterns of Alcohol-Induced Neuronal Loss Depend on Genetics: Implications for Fetal Alcohol Spectrum Disorder. Alcohol Clin Exp Res 2018; 42:1627-1639. [PMID: 29957842 PMCID: PMC6445660 DOI: 10.1111/acer.13824] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/14/2017] [Revised: 05/11/2018] [Accepted: 06/06/2018] [Indexed: 12/23/2022]
Abstract
BACKGROUND Alcohol exposure during pregnancy can kill developing neurons and lead to fetal alcohol spectrum disorder (FASD). However, affected individuals differ in their regional patterns of alcohol-induced neuropathology. Because neuroprotective genes are expressed in spatially selective ways, their mutation could increase the vulnerability of some brain regions, but not others, to alcohol teratogenicity. The objective of this study was to determine whether a null mutation of neuronal nitric oxide synthase (nNOS) can increase the vulnerability of some brain regions, but not others, to alcohol-induced neuronal losses. METHODS Immunohistochemistry identified brain regions in which nNOS is present or absent throughout postnatal development. Mice genetically deficient for nNOS (nNOS-/- ) and wild-type controls received alcohol (0.0, 2.2, or 4.4 mg/g/d) over postnatal days (PD) 4 to 9. Mice were sacrificed in adulthood (~PD 115), and surviving neurons in the olfactory bulb granular layer and brain stem facial nucleus were quantified stereologically. RESULTS nNOS was expressed throughout postnatal development in olfactory bulb granule cells but was never expressed in the facial nucleus. In wild-type mice, alcohol reduced neuronal survival to similar degrees in both cell populations. However, null mutation of nNOS more than doubled alcohol-induced cell death in the olfactory bulb granule cells, while the mutation had no effect on the facial nucleus neurons. As a result, in nNOS-/- mice, alcohol caused substantially more cell loss in the olfactory bulb than in the facial nucleus. CONCLUSIONS Mutation of the nNOS gene substantially increases vulnerability to alcohol-induced cell loss in a brain region where the gene is expressed (olfactory bulb), but not in a separate brain region, where the gene is not expressed (facial nucleus). Thus, differences in genotype may explain why some individuals are vulnerable to FASD, while others are not, and may determine the specific patterns of neuropathology in children with FASD.
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Affiliation(s)
- Dylan Todd
- Neuroscience Program, University of Iowa, Carver College of Medicine, Iowa City, Iowa 52242
| | | | - Lia Marie Sabalo
- Departments of Pediatrics, University of Iowa, Carver College of Medicine, Iowa City, Iowa 52242
| | - Jasmine Roghair
- Departments of Pediatrics, University of Iowa, Carver College of Medicine, Iowa City, Iowa 52242
| | - Bahri Karacay
- Departments of Pediatrics, University of Iowa, Carver College of Medicine, Iowa City, Iowa 52242
| | | | - Daniel J. Bonthius
- Neuroscience Program, University of Iowa, Carver College of Medicine, Iowa City, Iowa 52242
- Departments of Pediatrics, University of Iowa, Carver College of Medicine, Iowa City, Iowa 52242
- Neurology, University of Iowa, Carver College of Medicine, Iowa City, Iowa 52242
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19
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Kiriaev L, Kueh S, Morley JW, North KN, Houweling PJ, Head SI. Branched fibers from old fast-twitch dystrophic muscles are the sites of terminal damage in muscular dystrophy. Am J Physiol Cell Physiol 2018; 314:C662-C674. [PMID: 29412689 DOI: 10.1152/ajpcell.00161.2017] [Citation(s) in RCA: 20] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/30/2023]
Abstract
A striking pathological feature of dystrophinopathies is the presence of morphologically abnormal branched skeletal muscle fibers. The deterioration of muscle contractile function in Duchenne muscular dystrophy is accompanied by both an increase in number and complexity of these branched fibers. We propose that when number and complexity of branched fibers reaches a critical threshold, or "tipping point," the branches in and of themselves are the site of contraction-induced rupture. In the present study, we use the dystrophic mdx mouse and littermate controls to study the prediseased dystrophic fast-twitch extensor digitorum longus (EDL) muscle at 2-3 wk, the peak myonecrotic phase at 6-9 wk, and finally, "old," at 58-112 wk. Using a combination of isolated muscle function contractile measurements coupled with single-fiber imaging and confocal microscope imaging of cleared whole muscles, we identified a distinct pathophysiology, acute fiber rupture at branch nodes, which occurs in "old" fast-twitch EDL muscle approaching the end stage of the dystrophinopathy muscle disease, where the EDL muscles are entirely composed of complexed branched fibers. This evidence supports our concept of "tipping point" where the number and extent of fiber branching reach a level where the branching itself terminally compromises muscle function, irrespective of the absence of dystrophin.
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Affiliation(s)
- Leonit Kiriaev
- School of Medical Sciences, University of New South Wales , Sidney, New South Wales , Australia
| | - Sindy Kueh
- School of Medical Sciences, University of New South Wales , Sidney, New South Wales , Australia.,School of Medicine, Western Sydney University, Penrith, New South Wales , Australia
| | - John W Morley
- School of Medicine, Western Sydney University, Penrith, New South Wales , Australia
| | - Kathryn N North
- Murdoch Children's Research Institute , Melbourne, Victoria , Australia
| | - Peter J Houweling
- Murdoch Children's Research Institute , Melbourne, Victoria , Australia
| | - Stewart I Head
- School of Medical Sciences, University of New South Wales , Sidney, New South Wales , Australia.,School of Medicine, Western Sydney University, Penrith, New South Wales , Australia
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20
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Victor RG, Sweeney HL, Finkel R, McDonald CM, Byrne B, Eagle M, Goemans N, Vandenborne K, Dubrovsky AL, Topaloglu H, Miceli MC, Furlong P, Landry J, Elashoff R, Cox D. A phase 3 randomized placebo-controlled trial of tadalafil for Duchenne muscular dystrophy. Neurology 2017; 89:1811-1820. [PMID: 28972192 PMCID: PMC5664308 DOI: 10.1212/wnl.0000000000004570] [Citation(s) in RCA: 52] [Impact Index Per Article: 6.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/21/2017] [Accepted: 07/28/2017] [Indexed: 12/14/2022] Open
Abstract
OBJECTIVE To conduct a randomized trial to test the primary hypothesis that once-daily tadalafil, administered orally for 48 weeks, lessens the decline in ambulatory ability in boys with Duchenne muscular dystrophy (DMD). METHODS Three hundred thirty-one participants with DMD 7 to 14 years of age taking glucocorticoids were randomized to tadalafil 0.3 mg·kg-1·d-1, tadalafil 0.6 mg·kg-1·d-1, or placebo. The primary efficacy measure was 6-minute walk distance (6MWD) after 48 weeks. Secondary efficacy measures included North Star Ambulatory Assessment and timed function tests. Performance of Upper Limb (PUL) was a prespecified exploratory outcome. RESULTS Tadalafil had no effect on the primary outcome: 48-week declines in 6MWD were 51.0 ± 9.3 m with placebo, 64.7 ± 9.8 m with low-dose tadalafil (p = 0.307 vs placebo), and 59.1 ± 9.4 m with high-dose tadalafil (p = 0.538 vs placebo). Tadalafil also had no effect on secondary outcomes. In boys >10 years of age, total PUL score and shoulder subscore declined less with low-dose tadalafil than placebo. Adverse events were consistent with the known safety profile of tadalafil and the DMD disease state. CONCLUSIONS Tadalafil did not lessen the decline in ambulatory ability in boys with DMD. Further studies should be considered to confirm the hypothesis-generating upper limb data and to determine whether ambulatory decline can be slowed by initiation of tadalafil before 7 years of age. CLINICALTRIALSGOV IDENTIFIER NCT01865084. CLASSIFICATION OF EVIDENCE This study provides Class I evidence that tadalafil does not slow ambulatory decline in 7- to 14-year-old boys with Duchenne muscular dystrophy.
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Affiliation(s)
- Ronald G Victor
- From the Cedars-Sinai Medical Center (R.G.V.), Los Angeles, CA; University of Florida (H.L.S., B.B., K.V.), Gainesville; Nemours Children's Hospital (R.F.), Orlando, FL; University of California at Davis (C.M.M.), Sacramento; Newcastle University (M.E.), Newcastle Upon Tyne, UK; University Hospitals Leuven (N.G.), Belgium; Instituto de Neurociencias-Fundacion Favaloro (A.L.D.), Buenos Aires, Argentina; Hacettepe University School of Medicine (H.T.), Ankara, Turkey; UCLA (M.C.M., R.E.), Los Angeles, CA; Parent Project Muscular Dystrophy (P.F.), Hackensack, NJ; Eli Lilly Canada, Eli Lilly and Company, Toronto, ON (J.L.); and Eli Lilly and Company (D.C.), Indianapolis, IN.
| | - H Lee Sweeney
- From the Cedars-Sinai Medical Center (R.G.V.), Los Angeles, CA; University of Florida (H.L.S., B.B., K.V.), Gainesville; Nemours Children's Hospital (R.F.), Orlando, FL; University of California at Davis (C.M.M.), Sacramento; Newcastle University (M.E.), Newcastle Upon Tyne, UK; University Hospitals Leuven (N.G.), Belgium; Instituto de Neurociencias-Fundacion Favaloro (A.L.D.), Buenos Aires, Argentina; Hacettepe University School of Medicine (H.T.), Ankara, Turkey; UCLA (M.C.M., R.E.), Los Angeles, CA; Parent Project Muscular Dystrophy (P.F.), Hackensack, NJ; Eli Lilly Canada, Eli Lilly and Company, Toronto, ON (J.L.); and Eli Lilly and Company (D.C.), Indianapolis, IN
| | - Richard Finkel
- From the Cedars-Sinai Medical Center (R.G.V.), Los Angeles, CA; University of Florida (H.L.S., B.B., K.V.), Gainesville; Nemours Children's Hospital (R.F.), Orlando, FL; University of California at Davis (C.M.M.), Sacramento; Newcastle University (M.E.), Newcastle Upon Tyne, UK; University Hospitals Leuven (N.G.), Belgium; Instituto de Neurociencias-Fundacion Favaloro (A.L.D.), Buenos Aires, Argentina; Hacettepe University School of Medicine (H.T.), Ankara, Turkey; UCLA (M.C.M., R.E.), Los Angeles, CA; Parent Project Muscular Dystrophy (P.F.), Hackensack, NJ; Eli Lilly Canada, Eli Lilly and Company, Toronto, ON (J.L.); and Eli Lilly and Company (D.C.), Indianapolis, IN
| | - Craig M McDonald
- From the Cedars-Sinai Medical Center (R.G.V.), Los Angeles, CA; University of Florida (H.L.S., B.B., K.V.), Gainesville; Nemours Children's Hospital (R.F.), Orlando, FL; University of California at Davis (C.M.M.), Sacramento; Newcastle University (M.E.), Newcastle Upon Tyne, UK; University Hospitals Leuven (N.G.), Belgium; Instituto de Neurociencias-Fundacion Favaloro (A.L.D.), Buenos Aires, Argentina; Hacettepe University School of Medicine (H.T.), Ankara, Turkey; UCLA (M.C.M., R.E.), Los Angeles, CA; Parent Project Muscular Dystrophy (P.F.), Hackensack, NJ; Eli Lilly Canada, Eli Lilly and Company, Toronto, ON (J.L.); and Eli Lilly and Company (D.C.), Indianapolis, IN
| | - Barry Byrne
- From the Cedars-Sinai Medical Center (R.G.V.), Los Angeles, CA; University of Florida (H.L.S., B.B., K.V.), Gainesville; Nemours Children's Hospital (R.F.), Orlando, FL; University of California at Davis (C.M.M.), Sacramento; Newcastle University (M.E.), Newcastle Upon Tyne, UK; University Hospitals Leuven (N.G.), Belgium; Instituto de Neurociencias-Fundacion Favaloro (A.L.D.), Buenos Aires, Argentina; Hacettepe University School of Medicine (H.T.), Ankara, Turkey; UCLA (M.C.M., R.E.), Los Angeles, CA; Parent Project Muscular Dystrophy (P.F.), Hackensack, NJ; Eli Lilly Canada, Eli Lilly and Company, Toronto, ON (J.L.); and Eli Lilly and Company (D.C.), Indianapolis, IN
| | - Michelle Eagle
- From the Cedars-Sinai Medical Center (R.G.V.), Los Angeles, CA; University of Florida (H.L.S., B.B., K.V.), Gainesville; Nemours Children's Hospital (R.F.), Orlando, FL; University of California at Davis (C.M.M.), Sacramento; Newcastle University (M.E.), Newcastle Upon Tyne, UK; University Hospitals Leuven (N.G.), Belgium; Instituto de Neurociencias-Fundacion Favaloro (A.L.D.), Buenos Aires, Argentina; Hacettepe University School of Medicine (H.T.), Ankara, Turkey; UCLA (M.C.M., R.E.), Los Angeles, CA; Parent Project Muscular Dystrophy (P.F.), Hackensack, NJ; Eli Lilly Canada, Eli Lilly and Company, Toronto, ON (J.L.); and Eli Lilly and Company (D.C.), Indianapolis, IN
| | - Nathalie Goemans
- From the Cedars-Sinai Medical Center (R.G.V.), Los Angeles, CA; University of Florida (H.L.S., B.B., K.V.), Gainesville; Nemours Children's Hospital (R.F.), Orlando, FL; University of California at Davis (C.M.M.), Sacramento; Newcastle University (M.E.), Newcastle Upon Tyne, UK; University Hospitals Leuven (N.G.), Belgium; Instituto de Neurociencias-Fundacion Favaloro (A.L.D.), Buenos Aires, Argentina; Hacettepe University School of Medicine (H.T.), Ankara, Turkey; UCLA (M.C.M., R.E.), Los Angeles, CA; Parent Project Muscular Dystrophy (P.F.), Hackensack, NJ; Eli Lilly Canada, Eli Lilly and Company, Toronto, ON (J.L.); and Eli Lilly and Company (D.C.), Indianapolis, IN
| | - Krista Vandenborne
- From the Cedars-Sinai Medical Center (R.G.V.), Los Angeles, CA; University of Florida (H.L.S., B.B., K.V.), Gainesville; Nemours Children's Hospital (R.F.), Orlando, FL; University of California at Davis (C.M.M.), Sacramento; Newcastle University (M.E.), Newcastle Upon Tyne, UK; University Hospitals Leuven (N.G.), Belgium; Instituto de Neurociencias-Fundacion Favaloro (A.L.D.), Buenos Aires, Argentina; Hacettepe University School of Medicine (H.T.), Ankara, Turkey; UCLA (M.C.M., R.E.), Los Angeles, CA; Parent Project Muscular Dystrophy (P.F.), Hackensack, NJ; Eli Lilly Canada, Eli Lilly and Company, Toronto, ON (J.L.); and Eli Lilly and Company (D.C.), Indianapolis, IN
| | - Alberto L Dubrovsky
- From the Cedars-Sinai Medical Center (R.G.V.), Los Angeles, CA; University of Florida (H.L.S., B.B., K.V.), Gainesville; Nemours Children's Hospital (R.F.), Orlando, FL; University of California at Davis (C.M.M.), Sacramento; Newcastle University (M.E.), Newcastle Upon Tyne, UK; University Hospitals Leuven (N.G.), Belgium; Instituto de Neurociencias-Fundacion Favaloro (A.L.D.), Buenos Aires, Argentina; Hacettepe University School of Medicine (H.T.), Ankara, Turkey; UCLA (M.C.M., R.E.), Los Angeles, CA; Parent Project Muscular Dystrophy (P.F.), Hackensack, NJ; Eli Lilly Canada, Eli Lilly and Company, Toronto, ON (J.L.); and Eli Lilly and Company (D.C.), Indianapolis, IN
| | - Haluk Topaloglu
- From the Cedars-Sinai Medical Center (R.G.V.), Los Angeles, CA; University of Florida (H.L.S., B.B., K.V.), Gainesville; Nemours Children's Hospital (R.F.), Orlando, FL; University of California at Davis (C.M.M.), Sacramento; Newcastle University (M.E.), Newcastle Upon Tyne, UK; University Hospitals Leuven (N.G.), Belgium; Instituto de Neurociencias-Fundacion Favaloro (A.L.D.), Buenos Aires, Argentina; Hacettepe University School of Medicine (H.T.), Ankara, Turkey; UCLA (M.C.M., R.E.), Los Angeles, CA; Parent Project Muscular Dystrophy (P.F.), Hackensack, NJ; Eli Lilly Canada, Eli Lilly and Company, Toronto, ON (J.L.); and Eli Lilly and Company (D.C.), Indianapolis, IN
| | - M Carrie Miceli
- From the Cedars-Sinai Medical Center (R.G.V.), Los Angeles, CA; University of Florida (H.L.S., B.B., K.V.), Gainesville; Nemours Children's Hospital (R.F.), Orlando, FL; University of California at Davis (C.M.M.), Sacramento; Newcastle University (M.E.), Newcastle Upon Tyne, UK; University Hospitals Leuven (N.G.), Belgium; Instituto de Neurociencias-Fundacion Favaloro (A.L.D.), Buenos Aires, Argentina; Hacettepe University School of Medicine (H.T.), Ankara, Turkey; UCLA (M.C.M., R.E.), Los Angeles, CA; Parent Project Muscular Dystrophy (P.F.), Hackensack, NJ; Eli Lilly Canada, Eli Lilly and Company, Toronto, ON (J.L.); and Eli Lilly and Company (D.C.), Indianapolis, IN
| | - Pat Furlong
- From the Cedars-Sinai Medical Center (R.G.V.), Los Angeles, CA; University of Florida (H.L.S., B.B., K.V.), Gainesville; Nemours Children's Hospital (R.F.), Orlando, FL; University of California at Davis (C.M.M.), Sacramento; Newcastle University (M.E.), Newcastle Upon Tyne, UK; University Hospitals Leuven (N.G.), Belgium; Instituto de Neurociencias-Fundacion Favaloro (A.L.D.), Buenos Aires, Argentina; Hacettepe University School of Medicine (H.T.), Ankara, Turkey; UCLA (M.C.M., R.E.), Los Angeles, CA; Parent Project Muscular Dystrophy (P.F.), Hackensack, NJ; Eli Lilly Canada, Eli Lilly and Company, Toronto, ON (J.L.); and Eli Lilly and Company (D.C.), Indianapolis, IN
| | - John Landry
- From the Cedars-Sinai Medical Center (R.G.V.), Los Angeles, CA; University of Florida (H.L.S., B.B., K.V.), Gainesville; Nemours Children's Hospital (R.F.), Orlando, FL; University of California at Davis (C.M.M.), Sacramento; Newcastle University (M.E.), Newcastle Upon Tyne, UK; University Hospitals Leuven (N.G.), Belgium; Instituto de Neurociencias-Fundacion Favaloro (A.L.D.), Buenos Aires, Argentina; Hacettepe University School of Medicine (H.T.), Ankara, Turkey; UCLA (M.C.M., R.E.), Los Angeles, CA; Parent Project Muscular Dystrophy (P.F.), Hackensack, NJ; Eli Lilly Canada, Eli Lilly and Company, Toronto, ON (J.L.); and Eli Lilly and Company (D.C.), Indianapolis, IN
| | - Robert Elashoff
- From the Cedars-Sinai Medical Center (R.G.V.), Los Angeles, CA; University of Florida (H.L.S., B.B., K.V.), Gainesville; Nemours Children's Hospital (R.F.), Orlando, FL; University of California at Davis (C.M.M.), Sacramento; Newcastle University (M.E.), Newcastle Upon Tyne, UK; University Hospitals Leuven (N.G.), Belgium; Instituto de Neurociencias-Fundacion Favaloro (A.L.D.), Buenos Aires, Argentina; Hacettepe University School of Medicine (H.T.), Ankara, Turkey; UCLA (M.C.M., R.E.), Los Angeles, CA; Parent Project Muscular Dystrophy (P.F.), Hackensack, NJ; Eli Lilly Canada, Eli Lilly and Company, Toronto, ON (J.L.); and Eli Lilly and Company (D.C.), Indianapolis, IN
| | - David Cox
- From the Cedars-Sinai Medical Center (R.G.V.), Los Angeles, CA; University of Florida (H.L.S., B.B., K.V.), Gainesville; Nemours Children's Hospital (R.F.), Orlando, FL; University of California at Davis (C.M.M.), Sacramento; Newcastle University (M.E.), Newcastle Upon Tyne, UK; University Hospitals Leuven (N.G.), Belgium; Instituto de Neurociencias-Fundacion Favaloro (A.L.D.), Buenos Aires, Argentina; Hacettepe University School of Medicine (H.T.), Ankara, Turkey; UCLA (M.C.M., R.E.), Los Angeles, CA; Parent Project Muscular Dystrophy (P.F.), Hackensack, NJ; Eli Lilly Canada, Eli Lilly and Company, Toronto, ON (J.L.); and Eli Lilly and Company (D.C.), Indianapolis, IN
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21
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Moon Y, Balke JE, Madorma D, Siegel MP, Knowels G, Brouckaert P, Buys ES, Marcinek DJ, Percival JM. Nitric Oxide Regulates Skeletal Muscle Fatigue, Fiber Type, Microtubule Organization, and Mitochondrial ATP Synthesis Efficiency Through cGMP-Dependent Mechanisms. Antioxid Redox Signal 2017; 26:966-985. [PMID: 27393340 PMCID: PMC5467110 DOI: 10.1089/ars.2016.6630] [Citation(s) in RCA: 35] [Impact Index Per Article: 4.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/22/2022]
Abstract
AIM Skeletal muscle nitric oxide-cyclic guanosine monophosphate (NO-cGMP) pathways are impaired in Duchenne and Becker muscular dystrophy partly because of reduced nNOSμ and soluble guanylate cyclase (GC) activity. However, GC function and the consequences of reduced GC activity in skeletal muscle are unknown. In this study, we explore the functions of GC and NO-cGMP signaling in skeletal muscle. RESULTS GC1, but not GC2, expression was higher in oxidative than glycolytic muscles. GC1 was found in a complex with nNOSμ and targeted to nNOS compartments at the Golgi complex and neuromuscular junction. Baseline GC activity and GC agonist responsiveness was reduced in the absence of nNOS. Structural analyses revealed aberrant microtubule directionality in GC1-/- muscle. Functional analyses of GC1-/- muscles revealed reduced fatigue resistance and postexercise force recovery that were not due to shifts in type IIA-IIX fiber balance. Force deficits in GC1-/- muscles were also not driven by defects in resting mitochondrial adenosine triphosphate (ATP) synthesis. However, increasing muscle cGMP with sildenafil decreased ATP synthesis efficiency and capacity, without impacting mitochondrial content or ultrastructure. INNOVATION GC may represent a new target for alleviating muscle fatigue and that NO-cGMP signaling may play important roles in muscle structure, contractility, and bioenergetics. CONCLUSIONS These findings suggest that GC activity is nNOS dependent and that muscle-specific control of GC expression and differential GC targeting may facilitate NO-cGMP signaling diversity. They suggest that nNOS regulates muscle fiber type, microtubule organization, fatigability, and postexercise force recovery partly through GC1 and suggest that NO-cGMP pathways may modulate mitochondrial ATP synthesis efficiency. Antioxid. Redox Signal. 26, 966-985.
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Affiliation(s)
- Younghye Moon
- 1 Department of Molecular and Cellular Pharmacology, University of Miami Miller School of Medicine , Miami, Florida
| | - Jordan E Balke
- 1 Department of Molecular and Cellular Pharmacology, University of Miami Miller School of Medicine , Miami, Florida
| | - Derik Madorma
- 1 Department of Molecular and Cellular Pharmacology, University of Miami Miller School of Medicine , Miami, Florida
| | - Michael P Siegel
- 2 Department of Bioengineering, University of Washington , Seattle, Washington
| | - Gary Knowels
- 2 Department of Bioengineering, University of Washington , Seattle, Washington
| | - Peter Brouckaert
- 3 Department for Molecular Biomedical Research and Biomedical Molecular Biology, Ghent University , Ghent, Belgium
| | - Emmanuel S Buys
- 4 Department of Anesthesia, Critical Care and Pain Medicine, Anesthesia Center for Critical Care Research , Massachusetts General Hospital, Boston, Massachusetts
| | - David J Marcinek
- 2 Department of Bioengineering, University of Washington , Seattle, Washington.,5 Department of Radiology, University of Washington , Seattle, Washington
| | - Justin M Percival
- 1 Department of Molecular and Cellular Pharmacology, University of Miami Miller School of Medicine , Miami, Florida
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22
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A. Glemser P, Jaeger H, M. Nagel A, E. Ziegler A, Simons D, Schlemmer HP, Lehmann-Horn F, Jurkat-Rott K, Weber MA. 23Na MRI and myometry to compare eplerenone vs. glucocorticoid treatment in Duchenne dystrophy. ACTA MYOLOGICA : MYOPATHIES AND CARDIOMYOPATHIES : OFFICIAL JOURNAL OF THE MEDITERRANEAN SOCIETY OF MYOLOGY 2017; 36:2-13. [PMID: 28690388 PMCID: PMC5479105] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Indexed: 11/09/2022]
Abstract
In this pilot study we tested whether a low dose application of a mild diuretic substance such as eplerenone is beneficial in early stages of Duchenne muscular dystrophy using 23Na und 1H imaging, myometry, and clinical testing versus the glucocorticoid gold standard. Two 7-years old patients with DMD were examined on a 3T MRI system. 1H MRI and 23Na density-adapted 3-dimensional radial MRI sequences were performed both before and 1, 3 and 6 months after therapy with eplerenone respectively cortisone. We quantified fatty infiltration on T1-weighted images using subcutaneous fat as reference and fat fraction with a two-point DIXON sequence. Muscle oedema was quantified on STIR images and DIXON water maps with background noise as reference. We quantified Na+ by a muscular tissue concentration sequence with a 51.3mM Na+ with 5% agarose reference tube. A Na+ IR-sequence was used for determination of mainly myoplasmic Na+. Correspondingly myometry of muscles and tendons were assessed. Clinical tests (i.e. 4-steps-test) and blood counts (i.e. K+) were done by a pediatrician. Under eplerenone therapy we detected a reduction of muscular oedema, intracellular-weighted sodium IR signal and muscular sodium concentration. The oedema reduction in the DMD patient receiving eplerenone was more pronounced to the patient with cortisone. Myometric-measured tissue parameters such as muscle stiffness had a more pronounced effect in the child treated with eplerenone after a first increase in muscle stiffness both after eplerenone and cortisone treatment. Clinical abilities during both therapies were mostly constant. Eplerenone might be a possible new therapy option in DMD patients.
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Affiliation(s)
- Philip A. Glemser
- Department of Radiology, German Cancer Research Center (DKFZ), Heidelberg, Germany;, Institute of Forensic Medicine, University of Heidelberg, Heidelberg, Germany;,Address for correspondence: Dr. Philip Alexander Glemser, German Cancer Research Center (DKFZ), Department of Radiology, Im Neuenheimer Feld 280, 69120 Heidelberg, Germany. Tel. +49 6221 42 24 08. Fax +49 6221 42 25 57. E-mail:
| | - Heike Jaeger
- Division of Neurophysiology, Ulm University, Ulm, Germany
| | - Armin M. Nagel
- Division of Medical Physics in Radiology, German Cancer Research Center (DKFZ), Heidelberg, Germany;, Institute of Radiology, University Hospital Erlangen, Erlangen, Germany
| | - Andreas E. Ziegler
- Center for Child and Adolescent Medicine, University Hospital Heidelberg, Heidelberg, Germany
| | - David Simons
- Department of Radiology, German Cancer Research Center (DKFZ), Heidelberg, Germany
| | | | | | | | - Marc-André Weber
- Diagnostic and Interventional Radiology, University Hospital Heidelberg, Heidelberg, Germany
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23
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Moon Y, Cao Y, Zhu J, Xu Y, Balkan W, Buys ES, Diaz F, Kerrick WG, Hare JM, Percival JM. GSNOR Deficiency Enhances In Situ Skeletal Muscle Strength, Fatigue Resistance, and RyR1 S-Nitrosylation Without Impacting Mitochondrial Content and Activity. Antioxid Redox Signal 2017; 26:165-181. [PMID: 27412893 PMCID: PMC5278832 DOI: 10.1089/ars.2015.6548] [Citation(s) in RCA: 15] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/13/2022]
Abstract
AIM Nitric oxide (NO) plays important, but incompletely defined roles in skeletal muscle. NO exerts its regulatory effects partly though S-nitrosylation, which is balanced by denitrosylation by enzymes such as S-nitrosoglutathione reductase (GSNOR), whose functions in skeletal muscle remain to be fully deciphered. RESULTS GSNOR null (GSNOR-/-) tibialis anterior (TA) muscles showed normal growth and were stronger and more fatigue resistant than controls in situ. However, GSNOR-/- lumbrical muscles showed normal contractility and Ca2+ handling in vitro, suggesting important differences in GSNOR function between muscles or between in vitro and in situ environments. GSNOR-/- TA muscles exhibited normal mitochondrial content, and capillary densities, but reduced type IIA fiber content. GSNOR inhibition did not impact mitochondrial respiratory complex I, III, or IV activities. These findings argue that enhanced GSNOR-/- TA contractility is not driven by changes in mitochondrial content or activity, fiber type, or blood vessel density. However, loss of GSNOR led to RyR1 hypernitrosylation, which is believed to increase muscle force output under physiological conditions. cGMP synthesis by soluble guanylate cyclase (sGC) was decreased in resting GSNOR-/- muscle and was more responsive to agonist (DETANO, BAY 41, and BAY 58) stimulation, suggesting that GSNOR modulates cGMP production in skeletal muscle. INNOVATION GSNOR may act as a "brake" on skeletal muscle contractile performance under physiological conditions by modulating nitrosylation/denitrosylation balance. CONCLUSIONS GSNOR may play important roles in skeletal muscle contractility, RyR1 S-nitrosylation, fiber type specification, and sGC activity. Antioxid. Redox Signal. 26, 165-181.
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Affiliation(s)
- Younghye Moon
- 1 Department of Molecular and Cellular Pharmacology, University of Miami Miller School of Medicine , Miami, Florida
| | - Yenong Cao
- 1 Department of Molecular and Cellular Pharmacology, University of Miami Miller School of Medicine , Miami, Florida.,2 The Interdisciplinary Stem Cell Institute, University of Miami Miller School of Medicine , Miami, Florida
| | - Jingjing Zhu
- 1 Department of Molecular and Cellular Pharmacology, University of Miami Miller School of Medicine , Miami, Florida
| | - Yuanyuan Xu
- 3 Department of Physiology and Biophysics, University of Miami Miller School of Medicine , Miami, Florida
| | - Wayne Balkan
- 2 The Interdisciplinary Stem Cell Institute, University of Miami Miller School of Medicine , Miami, Florida.,4 Department of Medicine, University of Miami Miller School of Medicine , Miami, Florida
| | - Emmanuel S Buys
- 5 Department of Anesthesia, Critical Care and Pain Medicine, Anesthesia Center for Critical Care Research , Harvard Medical School, Massachusetts General Hospital Boston, Boston, Massachusetts
| | - Francisca Diaz
- 6 Department of Neurology, University of Miami Miller School of Medicine , Miami, Florida
| | - W Glenn Kerrick
- 3 Department of Physiology and Biophysics, University of Miami Miller School of Medicine , Miami, Florida
| | - Joshua M Hare
- 1 Department of Molecular and Cellular Pharmacology, University of Miami Miller School of Medicine , Miami, Florida.,2 The Interdisciplinary Stem Cell Institute, University of Miami Miller School of Medicine , Miami, Florida.,4 Department of Medicine, University of Miami Miller School of Medicine , Miami, Florida
| | - Justin M Percival
- 1 Department of Molecular and Cellular Pharmacology, University of Miami Miller School of Medicine , Miami, Florida
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24
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Latroche C, Gitiaux C, Chrétien F, Desguerre I, Mounier R, Chazaud B. Skeletal Muscle Microvasculature: A Highly Dynamic Lifeline. Physiology (Bethesda) 2016; 30:417-27. [PMID: 26525341 DOI: 10.1152/physiol.00026.2015] [Citation(s) in RCA: 68] [Impact Index Per Article: 7.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/25/2022] Open
Abstract
Skeletal muscle is highly irrigated by blood vessels. Beyond oxygen and nutrient supply, new vessel functions have been identified. This review presents vessel microanatomy and functions at tissue, cellular, and molecular levels. Mechanisms of vessel plasticity are described during skeletal muscle development and acute regeneration, and in physiological and pathological contexts.
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Affiliation(s)
- Claire Latroche
- Institut Cochin, INSERM U1016, Paris, France; CNRS 8104, Paris, France; Université Paris Descartes, Paris, France; Institut Pasteur, Paris, France
| | - Cyril Gitiaux
- Institut Cochin, INSERM U1016, Paris, France; CNRS 8104, Paris, France; Université Paris Descartes, Paris, France; Institut Pasteur, Paris, France
| | | | - Isabelle Desguerre
- Institut Cochin, INSERM U1016, Paris, France; CNRS 8104, Paris, France; Université Paris Descartes, Paris, France
| | - Rémi Mounier
- CGPhyMC, CNRS UMR5534, Villeurbanne, France; and Université Claude Bernard Lyon1, Villeurbanne, France
| | - Bénédicte Chazaud
- Institut Cochin, INSERM U1016, Paris, France; CNRS 8104, Paris, France; Université Paris Descartes, Paris, France; CGPhyMC, CNRS UMR5534, Villeurbanne, France; and Université Claude Bernard Lyon1, Villeurbanne, France
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25
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Gentil C, Le Guiner C, Falcone S, Hogrel JY, Peccate C, Lorain S, Benkhelifa-Ziyyat S, Guigand L, Montus M, Servais L, Voit T, Piétri-Rouxel F. Dystrophin Threshold Level Necessary for Normalization of Neuronal Nitric Oxide Synthase, Inducible Nitric Oxide Synthase, and Ryanodine Receptor-Calcium Release Channel Type 1 Nitrosylation in Golden Retriever Muscular Dystrophy Dystrophinopathy. Hum Gene Ther 2016; 27:712-26. [PMID: 27279388 DOI: 10.1089/hum.2016.041] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/16/2023] Open
Abstract
At present, the clinically most advanced strategy to treat Duchenne muscular dystrophy (DMD) is the exon-skipping strategy. Whereas antisense oligonucleotide-based clinical trials are underway for DMD, it is essential to determine the dystrophin restoration threshold needed to ensure improvement of muscle physiology at the molecular level. A preclinical trial has been conducted in golden retriever muscular dystrophy (GRMD) dogs treated in a forelimb by locoregional delivery of rAAV8-U7snRNA to promote exon skipping on the canine dystrophin messenger. Here, we exploited rAAV8-U7snRNA-transduced GRMD muscle samples, well characterized for their percentage of dystrophin-positive fibers, with the aim of defining the threshold of dystrophin rescue necessary for normalization of the status of neuronal nitric oxide synthase mu (nNOSμ), inducible nitric oxide synthase (iNOS), and ryanodine receptor-calcium release channel type 1 (RyR1), crucial actors for efficient contractile function. Results showed that restoration of dystrophin in 40% of muscle fibers is needed to decrease abnormal cytosolic nNOSμ expression and to reduce overexpression of iNOS, these two parameters leading to a reduction in the NO level in the muscle fibers. Furthermore, the same percentage of dystrophin-positive fibers of 40% was associated with the normalization of RyR1 nitrosylation status and with stabilization of the RyR1-calstabin1 complex that is required to facilitate coupled gating. We concluded that a minimal threshold of 40% of dystrophin-positive fibers is necessary for the reinstatement of central proteins needed for proper muscle contractile function, and thus identified a rate of dystrophin expression significantly improving, at the molecular level, the dystrophic muscle physiology.
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Affiliation(s)
- Christel Gentil
- 1 Sorbonne Universités , UPMC Univ Paris 06/INSERM/CNRS/Institut de Myologie/Centre de Recherche en Myologie (CRM), GH Pitié Salpêtrière, Paris, France
| | - Caroline Le Guiner
- 2 Atlantic Gene Therapies/INSERM UMR 1089 Université de Nantes , CHU de Nantes, IRT1, Nantes, France.,3 Généthon , Evry, France
| | - Sestina Falcone
- 1 Sorbonne Universités , UPMC Univ Paris 06/INSERM/CNRS/Institut de Myologie/Centre de Recherche en Myologie (CRM), GH Pitié Salpêtrière, Paris, France
| | | | - Cécile Peccate
- 1 Sorbonne Universités , UPMC Univ Paris 06/INSERM/CNRS/Institut de Myologie/Centre de Recherche en Myologie (CRM), GH Pitié Salpêtrière, Paris, France
| | - Stéphanie Lorain
- 1 Sorbonne Universités , UPMC Univ Paris 06/INSERM/CNRS/Institut de Myologie/Centre de Recherche en Myologie (CRM), GH Pitié Salpêtrière, Paris, France
| | - Sofia Benkhelifa-Ziyyat
- 1 Sorbonne Universités , UPMC Univ Paris 06/INSERM/CNRS/Institut de Myologie/Centre de Recherche en Myologie (CRM), GH Pitié Salpêtrière, Paris, France
| | - Lydie Guigand
- 5 Atlantic Gene Therapies /INRA UMR 703, ONIRIS, Nantes-Atlantic National College of Veterinary Medicine , Food Science, and Engineering, Nantes, France
| | | | - Laurent Servais
- 4 Institut de Myologie , GH Pitié-Salpêtrière, Paris, France
| | - Thomas Voit
- 1 Sorbonne Universités , UPMC Univ Paris 06/INSERM/CNRS/Institut de Myologie/Centre de Recherche en Myologie (CRM), GH Pitié Salpêtrière, Paris, France
| | - France Piétri-Rouxel
- 1 Sorbonne Universités , UPMC Univ Paris 06/INSERM/CNRS/Institut de Myologie/Centre de Recherche en Myologie (CRM), GH Pitié Salpêtrière, Paris, France
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26
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Allen DG, Whitehead NP, Froehner SC. Absence of Dystrophin Disrupts Skeletal Muscle Signaling: Roles of Ca2+, Reactive Oxygen Species, and Nitric Oxide in the Development of Muscular Dystrophy. Physiol Rev 2016; 96:253-305. [PMID: 26676145 DOI: 10.1152/physrev.00007.2015] [Citation(s) in RCA: 309] [Impact Index Per Article: 34.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/26/2022] Open
Abstract
Dystrophin is a long rod-shaped protein that connects the subsarcolemmal cytoskeleton to a complex of proteins in the surface membrane (dystrophin protein complex, DPC), with further connections via laminin to other extracellular matrix proteins. Initially considered a structural complex that protected the sarcolemma from mechanical damage, the DPC is now known to serve as a scaffold for numerous signaling proteins. Absence or reduced expression of dystrophin or many of the DPC components cause the muscular dystrophies, a group of inherited diseases in which repeated bouts of muscle damage lead to atrophy and fibrosis, and eventually muscle degeneration. The normal function of dystrophin is poorly defined. In its absence a complex series of changes occur with multiple muscle proteins showing reduced or increased expression or being modified in various ways. In this review, we will consider the various proteins whose expression and function is changed in muscular dystrophies, focusing on Ca(2+)-permeable channels, nitric oxide synthase, NADPH oxidase, and caveolins. Excessive Ca(2+) entry, increased membrane permeability, disordered caveolar function, and increased levels of reactive oxygen species are early changes in the disease, and the hypotheses for these phenomena will be critically considered. The aim of the review is to define the early damage pathways in muscular dystrophy which might be appropriate targets for therapy designed to minimize the muscle degeneration and slow the progression of the disease.
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Affiliation(s)
- David G Allen
- Sydney Medical School & Bosch Institute, University of Sydney, New South Wales, Australia; and Department of Physiology & Biophysics, University of Washington, Seattle, Washington
| | - Nicholas P Whitehead
- Sydney Medical School & Bosch Institute, University of Sydney, New South Wales, Australia; and Department of Physiology & Biophysics, University of Washington, Seattle, Washington
| | - Stanley C Froehner
- Sydney Medical School & Bosch Institute, University of Sydney, New South Wales, Australia; and Department of Physiology & Biophysics, University of Washington, Seattle, Washington
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Rebolledo DL, Kim MJ, Whitehead NP, Adams ME, Froehner SC. Sarcolemmal targeting of nNOSμ improves contractile function of mdx muscle. Hum Mol Genet 2015; 25:158-66. [PMID: 26604149 DOI: 10.1093/hmg/ddv466] [Citation(s) in RCA: 26] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/29/2015] [Accepted: 11/09/2015] [Indexed: 11/13/2022] Open
Abstract
Nitric oxide (NO) is a key regulator of skeletal muscle function and metabolism, including vasoregulation, mitochondrial function, glucose uptake, fatigue and excitation-contraction coupling. The main generator of NO in skeletal muscle is the muscle-specific form of neuronal nitric oxide synthase (nNOSμ) produced by the NOS1 gene. Skeletal muscle nNOSμ is predominantly localized at the sarcolemma by interaction with the dystrophin protein complex (DPC). In Duchenne muscular dystrophy (DMD), loss of dystrophin leads to the mislocalization of nNOSμ from the sarcolemma to the cytosol. This perturbation has been shown to impair contractile function and cause muscle fatigue in dystrophic (mdx) mice. Here, we investigated the effect of restoring sarcolemmal nNOSμ on muscle contractile function in mdx mice. To achieve this, we designed a modified form of nNOSμ (NOS-M) that is targeted to the sarcolemma by palmitoylation, even in the absence of the DPC. When expressed specifically in mdx skeletal muscle, NOS-M significantly attenuates force loss owing to damaging eccentric contractions and repetitive isometric contractions (fatigue), while also improving force recovery after fatigue. Expression of unmodified nNOSμ at similar levels does not lead to sarcolemmal association and fails to improve muscle function. Aside from the benefits of sarcolemmal-localized NO production, NOS-M also increased the surface membrane levels of utrophin and other DPC proteins, including β-dystroglycan, α-syntrophin and α-dystrobrevin in mdx muscle. These results suggest that the expression of NOS-M in skeletal muscle may be therapeutically beneficial in DMD and other muscle diseases characterized by the loss of nNOSμ from the sarcolemma.
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Affiliation(s)
- Daniela L Rebolledo
- Department of Physiology and Biophysics, University of Washington, Seattle, WA 98195-7290, USA and Departamento de Biología Celular y Molecular, Pontificia Universidad Católica de Chile, Santiago 8331150, Chile
| | - Min Jeong Kim
- Department of Physiology and Biophysics, University of Washington, Seattle, WA 98195-7290, USA and
| | - Nicholas P Whitehead
- Department of Physiology and Biophysics, University of Washington, Seattle, WA 98195-7290, USA and
| | - Marvin E Adams
- Department of Physiology and Biophysics, University of Washington, Seattle, WA 98195-7290, USA and
| | - Stanley C Froehner
- Department of Physiology and Biophysics, University of Washington, Seattle, WA 98195-7290, USA and
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De Palma C, Morisi F, Pambianco S, Assi E, Touvier T, Russo S, Perrotta C, Romanello V, Carnio S, Cappello V, Pellegrino P, Moscheni C, Bassi MT, Sandri M, Cervia D, Clementi E. Deficient nitric oxide signalling impairs skeletal muscle growth and performance: involvement of mitochondrial dysregulation. Skelet Muscle 2014; 4:22. [PMID: 25530838 PMCID: PMC4272808 DOI: 10.1186/s13395-014-0022-6] [Citation(s) in RCA: 49] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/26/2014] [Accepted: 11/18/2014] [Indexed: 12/18/2022] Open
Abstract
BACKGROUND Nitric oxide (NO), generated in skeletal muscle mostly by the neuronal NO synthases (nNOSμ), has profound effects on both mitochondrial bioenergetics and muscle development and function. The importance of NO for muscle repair emerges from the observation that nNOS signalling is defective in many genetically diverse skeletal muscle diseases in which muscle repair is dysregulated. How the effects of NO/nNOSμ on mitochondria impact on muscle function, however, has not been investigated yet. METHODS In this study we have examined the relationship between the NO system, mitochondrial structure/activity and skeletal muscle phenotype/growth/functions using a mouse model in which nNOSμ is absent. Also, NO-induced effects and the NO pathway were dissected in myogenic precursor cells. RESULTS We show that nNOSμ deficiency in mouse skeletal muscle leads to altered mitochondrial bioenergetics and network remodelling, and increased mitochondrial unfolded protein response (UPR(mt)) and autophagy. The absence of nNOSμ is also accompanied by an altered mitochondrial homeostasis in myogenic precursor cells with a decrease in the number of myonuclei per fibre and impaired muscle development at early stages of perinatal growth. No alterations were observed, however, in the overall resting muscle structure, apart from a reduced specific muscle mass and cross sectional areas of the myofibres. Investigating the molecular mechanisms we found that nNOSμ deficiency was associated with an inhibition of the Akt-mammalian target of rapamycin pathway. Concomitantly, the Akt-FoxO3-mitochondrial E3 ubiquitin protein ligase 1 (Mul-1) axis was also dysregulated. In particular, inhibition of nNOS/NO/cyclic guanosine monophosphate (cGMP)/cGMP-dependent-protein kinases induced the transcriptional activity of FoxO3 and increased Mul-1 expression. nNOSμ deficiency was also accompanied by functional changes in muscle with reduced muscle force, decreased resistance to fatigue and increased degeneration/damage post-exercise. CONCLUSIONS Our results indicate that nNOSμ/NO is required to regulate key homeostatic mechanisms in skeletal muscle, namely mitochondrial bioenergetics and network remodelling, UPR(mt) and autophagy. These events are likely associated with nNOSμ-dependent impairments of muscle fibre growth resulting in a deficit of muscle performance.
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Affiliation(s)
- Clara De Palma
- Unit of Clinical Pharmacology, National Research Council-Institute of Neuroscience, Department of Biomedical and Clinical Sciences "Luigi Sacco", University Hospital "Luigi Sacco", Università di Milano, Milano, Italy
| | - Federica Morisi
- Unit of Clinical Pharmacology, National Research Council-Institute of Neuroscience, Department of Biomedical and Clinical Sciences "Luigi Sacco", University Hospital "Luigi Sacco", Università di Milano, Milano, Italy
| | - Sarah Pambianco
- Unit of Clinical Pharmacology, National Research Council-Institute of Neuroscience, Department of Biomedical and Clinical Sciences "Luigi Sacco", University Hospital "Luigi Sacco", Università di Milano, Milano, Italy
| | - Emma Assi
- Scientific Institute IRCCS Eugenio Medea, Bosisio Parini, Italy
| | - Thierry Touvier
- Unit of Clinical Pharmacology, National Research Council-Institute of Neuroscience, Department of Biomedical and Clinical Sciences "Luigi Sacco", University Hospital "Luigi Sacco", Università di Milano, Milano, Italy
| | - Stefania Russo
- Scientific Institute IRCCS Eugenio Medea, Bosisio Parini, Italy
| | - Cristiana Perrotta
- Unit of Clinical Pharmacology, National Research Council-Institute of Neuroscience, Department of Biomedical and Clinical Sciences "Luigi Sacco", University Hospital "Luigi Sacco", Università di Milano, Milano, Italy
| | - Vanina Romanello
- Dulbecco Telethon Institute at Venetian Institute of Molecular Medicine, Padova, Italy
| | - Silvia Carnio
- Dulbecco Telethon Institute at Venetian Institute of Molecular Medicine, Padova, Italy
| | - Valentina Cappello
- National Research Council-Institute of Neuroscience, Department of Medical Biotechnology and Translational Medicine, Università di Milano, Milano, Italy ; CNI@NEST, Italian Institute of Technology, Pisa, Italy
| | - Paolo Pellegrino
- Unit of Clinical Pharmacology, National Research Council-Institute of Neuroscience, Department of Biomedical and Clinical Sciences "Luigi Sacco", University Hospital "Luigi Sacco", Università di Milano, Milano, Italy
| | - Claudia Moscheni
- Unit of Morphology, Department of Biomedical and Clinical Sciences "Luigi Sacco", Università di Milano, Milano, Italy
| | | | - Marco Sandri
- Dulbecco Telethon Institute at Venetian Institute of Molecular Medicine, Padova, Italy ; Department of Biomedical Science, Università di Padova, Padova, Italy
| | - Davide Cervia
- Unit of Clinical Pharmacology, National Research Council-Institute of Neuroscience, Department of Biomedical and Clinical Sciences "Luigi Sacco", University Hospital "Luigi Sacco", Università di Milano, Milano, Italy ; Department for Innovation in Biological, Agro-food and Forest Systems, Università della Tuscia, Viterbo, Italy
| | - Emilio Clementi
- Unit of Clinical Pharmacology, National Research Council-Institute of Neuroscience, Department of Biomedical and Clinical Sciences "Luigi Sacco", University Hospital "Luigi Sacco", Università di Milano, Milano, Italy ; Scientific Institute IRCCS Eugenio Medea, Bosisio Parini, Italy
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