scholarly journals Nav1.4 Deregulation in Dystrophic Skeletal Muscle Leads to Na+ Overload and Enhanced Cell Death

2008 ◽  
Vol 132 (2) ◽  
pp. 199-208 ◽  
Author(s):  
Carole Hirn ◽  
George Shapovalov ◽  
Olivier Petermann ◽  
Emmanuelle Roulet ◽  
Urs T. Ruegg
Keyword(s):  
Skeletal Muscle ◽  
Cell Death ◽  
Early Death ◽  
Mdx Mouse ◽  
Mdx Mice ◽  
Strongly Correlated ◽  
Muscle Degeneration ◽  
Voltage Gated ◽  
Voltage Gated Sodium Channels ◽  
Associated Proteins

Duchenne muscular dystrophy (DMD) is a hereditary degenerative disease manifested by the absence of dystrophin, a structural, cytoskeletal protein, leading to muscle degeneration and early death through respiratory and cardiac muscle failure. Whereas the rise of cytosolic Ca2+ concentrations in muscles of mdx mouse, an animal model of DMD, has been extensively documented, little is known about the mechanisms causing alterations in Na+ concentrations. Here we show that the skeletal muscle isoform of the voltage-gated sodium channel, Nav1.4, which represents over 90% of voltage-gated sodium channels in muscle, plays an important role in development of abnormally high Na+ concentrations found in muscle from mdx mice. The absence of dystrophin modifies the expression level and gating properties of Nav1.4, leading to an increased Na+ concentration under the sarcolemma. Moreover, the distribution of Nav1.4 is altered in mdx muscle while maintaining the colocalization with one of the dystrophin-associated proteins, syntrophin α-1, thus suggesting that syntrophin is an important linker between dystrophin and Nav1.4. Additionally, we show that these modifications of Nav1.4 gating properties and increased Na+ concentrations are strongly correlated with increased cell death in mdx fibers and that both cell death and Na+ overload can be reversed by 3 nM tetrodotoxin, a specific Nav1.4 blocker.

scholarly journals Dystrophin-associated proteins are greatly reduced in skeletal muscle from mdx mice.

1991 ◽  
Vol 115 (6) ◽  
pp. 1685-1694 ◽  
Author(s):  
K Ohlendieck ◽  
K P Campbell
Keyword(s):  
Skeletal Muscle ◽  
Immunoblot Analysis ◽  
Protein Product ◽  
Mdx Mouse ◽  
Mdx Mice ◽  
Normal Density ◽  
Mouse Muscle ◽  
Glycoprotein Complex ◽  
Associated Proteins ◽  
Dy Mouse

Dystrophin, the protein product of the human Duchenne muscular dystrophy gene, exists in skeletal muscle as a large oligomeric complex that contains four glycoproteins of 156, 50, 43, and 35 kD and a protein of 59 kD. Here, we investigated the relative abundance of each of the components of the dystrophin-glycoprotein complex in skeletal muscle from normal and mdx mice, which are missing dystrophin. Immunoblot analysis using total muscle membranes from control and mdx mice of ages 1 d to 30 wk found that all of the dystrophin-associated proteins were greatly reduced (80-90%) in mdx mouse skeletal muscle. The specificity of the loss of the dystrophin-associated glycoproteins was demonstrated by the finding that the major glycoprotein composition of skeletal muscle membranes from normal and mdx mice was identical. Furthermore, skeletal muscle membranes from the dystrophic dy/dy mouse exhibited a normal density of dystrophin and dystrophin-associated proteins. Immunofluorescence microscopy confirmed the results from the immunoblot analysis and showed a drastically reduced density of dystrophin-associated proteins in mdx muscle cryosections compared with normal and dy/dy mouse muscle. Therefore, our results demonstrate that all of the dystrophin-associated proteins are significantly reduced in mdx skeletal muscle and suggest that the loss of dystrophin-associated proteins is due to the absence of dystrophin and not due to secondary effects of muscle fiber degradation.

Hindlimb immobilization applied to 21-day-oldmdx mice prevents the occurrence of muscle degeneration

1999 ◽  
Vol 86 (3) ◽  
pp. 924-931 ◽  
Author(s):  
Asghar Mokhtarian ◽  
Jean Pascal Lefaucheur ◽  
Patrick C. Even ◽  
Alain Sebille
Keyword(s):  
Skeletal Muscle ◽  
Motor Activity ◽  
Skeletal Muscles ◽  
Extensor Digitorum Longus ◽  
Contralateral Side ◽  
Mdx Mouse ◽  
Mdx Mice ◽  
Muscle Degeneration ◽  
Limb Immobilization ◽  
Old Mice

Dystrophin-deficient skeletal muscles of mdx mice undergo their first rounds of degeneration-regeneration at the age of 14–28 days. This feature is thought to result from an increase in motor activity at weaning. In this study, we hypothesize that if the muscle is prevented from contracting, it will avoid the degenerative changes that normally occur. For this purpose, we developed a procedure of mechanical hindlimb immobilization in 3-wk-old mice to restrain soleus (Sol) and extensor digitorum longus (EDL) muscles in the stretched or shortened position. After a 14-day period of immobilization, the striking feature was the low percentage of regenerated (centronucleated) myofibers in Sol and EDL muscles, regardless of the length at which they were fixed, compared with those on the contralateral side (stretched Sol: 8.4 ± 6.5 vs. 46.6 ± 10.3%, P = 0.0008; shortened Sol: 1.2 ± 1.6 vs. 50.4 ± 16.4%, P = 0.0008; stretched EDL: 05 ± 0.5 vs. 32.9 ± 17.5%, P = 0.002; shortened EDL: 3.3 ± 3.1 vs. 34.7 ± 11.1%, P = 0.002). Total numbers of myofibers did not change with immobilization. This study shows that limb immobilization prevents the occurrence of the first round of myofiber necrosis in mdx mice and suggests that muscle contractions play a role in the skeletal muscle degeneration of dystrophin-deficient mdx mouse muscles.

scholarly journals ProNGF/p75NTR Axis Drives Fiber Type Specification by Inducing the Fast-Glycolytic Phenotype in Mouse Skeletal Muscle Cells

Cells ◽  
2020 ◽  
Vol 9 (10) ◽  
pp. 2232
Author(s):  
Valentina Pallottini ◽  
Mayra Colardo ◽  
Claudia Tonini ◽  
Noemi Martella ◽  
Georgios Strimpakos ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Myosin Heavy Chain ◽  
Heavy Chain ◽  
Myogenic Differentiation ◽  
Fiber Type ◽  
Pcr Analysis ◽  
Mdx Mouse ◽  
Mdx Mice ◽  
Muscle Physiology

Despite its undisputable role in the homeostatic regulation of the nervous system, the nerve growth factor (NGF) also governs the relevant cellular processes in other tissues and organs. In this study, we aimed at assessing the expression and the putative involvement of NGF signaling in skeletal muscle physiology. To reach this objective, we employed satellite cell-derived myoblasts as an in vitro culture model. In vivo experiments were performed on Tibialis anterior from wild-type mice and an mdx mouse model of Duchenne muscular dystrophy. Targets of interest were mainly assessed by means of morphological, Western blot and qRT-PCR analysis. The results show that proNGF is involved in myogenic differentiation. Importantly, the proNGF/p75NTR pathway orchestrates a slow-to-fast fiber type transition by counteracting the expression of slow myosin heavy chain and that of oxidative markers. Concurrently, proNGF/p75NTR activation facilitates the induction of fast myosin heavy chain and of fast/glycolytic markers. Furthermore, we also provided evidence that the oxidative metabolism is impaired in mdx mice, and that these alterations are paralleled by a prominent buildup of proNGF and p75NTR. These findings underline that the proNGF/p75NTR pathway may play a crucial role in fiber type determination and suggest its prospective modulation as an innovative therapeutic approach to counteract muscle disorders.

scholarly journals In vitro and in vivo analysis of human fibroblast reprogramming and multipotency

2015 ◽  
Vol 20 (3) ◽  
Author(s):  
Rongqing Pang ◽  
Xiangqing Zhu ◽  
Jia Geng ◽  
Yongyun Zhang ◽  
Qiang Wang ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Human Fibroblasts ◽  
Mdx Mouse ◽  
Mdx Mice ◽  
Therapeutic Effects ◽  
Tail Vein ◽  
Tail Vein Injection ◽  
Activity Measurements ◽  
Human Dystrophin

AbstractMultipotent stem cells have potential therapeutic roles in the treatment of Duchenne muscular dystrophy (DMD). However, the limited access to stem cell sources restricts their clinical application. To address this issue, we established a simple in vitro epigenetic reprogramming technique in which skin fibroblasts are induced to dedifferentiate into multipotent cells. In this study, human fibroblasts were isolated from circumcised adult foreskin and were reprogrammed by co-culture for 72 h with fish oocyte extract (FOE) in serum-free medium. The cells were then observed and analyzed by immunofluorescence staining, flow cytometry and in vitro differentiation assays. Then FOE-treated human fibroblasts were transplanted by tail vein injection into irradiated mdx mice, an animal model of DMD. Two months after injection, the therapeutic effects of FOE-treated fibroblasts on mdx skeletal muscle were evaluated by serum creatine kinase (CK) activity measurements and by immunostaining and RT-PCR of human dystrophin expression. The results indicated that the reprogrammed fibroblasts expressed higher levels of the pluripotent antigen markers SSEA-4, Nanog and Oct-4, and were able to differentiate in vitro into adipogenic cells, osteoblastic cells, and myotube-like cells. Tail vein injection of FOE-treated fibroblasts into irradiated mdx mice slightly reduced serum CK activity and the percentage of centrally nucleated myofibers two months after cell transplantation. Furthermore, we confirmed human dystrophin protein and mRNA expression in mdx mouse skeletal muscle. These data demonstrated that FOE-treated fibroblasts were multipotent and could integrate into mdx mouse myofibers through the vasculature.

scholarly journals Clonal Isolation of Muscle-Derived Cells Capable of Enhancing Muscle Regeneration and Bone Healing

2000 ◽  
Vol 150 (5) ◽  
pp. 1085-1100 ◽  
Author(s):  
Joon Yung Lee ◽  
Zhuqing Qu-Petersen ◽  
Baohong Cao ◽  
Shigemi Kimura ◽  
Ron Jankowski ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Stem Cells ◽  
Stem Cell ◽  
Bone Healing ◽  
Muscle Regeneration ◽  
Mdx Mouse ◽  
Mdx Mice ◽  
Hematopoietic Stem ◽  
Isolation And Characterization ◽  
Osteogenic Lineage

Several recent studies suggest the isolation of stem cells in skeletal muscle, but the functional properties of these muscle-derived stem cells is still unclear. In the present study, we report the purification of muscle-derived stem cells from the mdx mouse, an animal model for Duchenne muscular dystrophy. We show that enrichment of desmin+ cells using the preplate technique from mouse primary muscle cell culture also enriches a cell population expressing CD34 and Bcl-2. The CD34+ cells and Bcl-2+ cells were found to reside within the basal lamina, where satellite cells are normally found. Clonal isolation and characterization from this CD34+Bcl-2+ enriched population yielded a putative muscle-derived stem cell, mc13, that is capable of differentiating into both myogenic and osteogenic lineage in vitro and in vivo. The mc13 cells are c-kit and CD45 negative and express: desmin, c-met and MNF, three markers expressed in early myogenic progenitors; Flk-1, a mouse homologue of KDR recently identified in humans as a key marker in hematopoietic cells with stem cell-like characteristics; and Sca-1, a marker for both skeletal muscle and hematopoietic stem cells. Intramuscular, and more importantly, intravenous injection of mc13 cells result in muscle regeneration and partial restoration of dystrophin in mdx mice. Transplantation of mc13 cells engineered to secrete osteogenic protein differentiate in osteogenic lineage and accelerate healing of a skull defect in SCID mice. Taken together, these results suggest the isolation of a population of muscle-derived stem cells capable of improving both muscle regeneration and bone healing.

scholarly journals Extensive but Coordinated Reorganization of the Membrane Skeleton in Myofibers of Dystrophic (mdx) Mice

1999 ◽  
Vol 144 (6) ◽  
pp. 1259-1270 ◽  
Author(s):  
McRae W. Williams ◽  
Robert J. Bloch
Keyword(s):  
Skeletal Muscle ◽  
Membrane Proteins ◽  
Muscle Fibers ◽  
Membrane Skeleton ◽  
Confocal Imaging ◽  
Mdx Mouse ◽  
Mdx Mice ◽  
Intracellular Membrane ◽  
Dystrophic Muscle ◽  
Dihydropyridine Receptors

We used immunofluorescence techniques and confocal imaging to study the organization of the membrane skeleton of skeletal muscle fibers of mdx mice, which lack dystrophin. β-Spectrin is normally found at the sarcolemma in costameres, a rectilinear array of longitudinal strands and elements overlying Z and M lines. However, in the skeletal muscle of mdx mice, β-spectrin tends to be absent from the sarcolemma over M lines and the longitudinal strands may be disrupted or missing. Other proteins of the membrane and associated cytoskeleton, including syntrophin, β-dystroglycan, vinculin, and Na,K-ATPase are also concentrated in costameres, in control myofibers, and mdx muscle. They also distribute into the same altered sarcolemmal arrays that contain β-spectrin. Utrophin, which is expressed in mdx muscle, also codistributes with β-spectrin at the mutant sarcolemma. By contrast, the distribution of structural and intracellular membrane proteins, including α-actinin, the Ca-ATPase and dihydropyridine receptors, is not affected, even at sites close to the sarcolemma. Our results suggest that in myofibers of the mdx mouse, the membrane- associated cytoskeleton, but not the nearby myoplasm, undergoes widespread coordinated changes in organization. These changes may contribute to the fragility of the sarcolemma of dystrophic muscle.

Elevated MMP2 abundance and activity in mdx mice are alleviated by prenatal taurine supplementation

2020 ◽  
Vol 318 (6) ◽  
pp. C1083-C1091
Author(s):  
Xiaoyu Ren ◽  
Hongyang Xu ◽  
Robert G. Barker ◽  
Graham D. Lamb ◽  
Robyn M. Murphy
Keyword(s):  
Muscle Fibers ◽  
Muscle Growth ◽  
Early Death ◽  
Active Role ◽  
Matrix Metalloproteinase 2 ◽  
Mdx Mouse ◽  
Mdx Mice ◽  
Taurine Supplementation ◽  
Naturally Occurring ◽  
Age Related

Duchenne muscular dystrophy (DMD) is a severe, progressive muscle-wasting disorder that leads to early death. The mdx mouse is a naturally occurring mutant model for DMD. It lacks dystrophin and displays peak muscle cell necrosis at ~28 days (D28), but in contrast to DMD, mdx mice experience muscle regeneration by D70. We hypothesized that matrix metalloproteinase-2 (MMP2) and/or MMP9 play key roles in the degeneration/regeneration phases in mdx mice. MMP2 abundance in muscle homogenates, measured by calibrated Western blotting, and activity, measured by zymogram, were lower at D70 compared with D28 in both mdx and wild-type (WT) mice. Importantly, MMP2 abundance was higher in both D28 and D70 mdx mice than in age-matched WT mice. The higher MMP2 abundance was not due to infiltrating macrophages, because MMP2 content was still higher in isolated muscle fibers where most macrophages had been removed. Prenatal supplementation with the amino acid taurine, which improved muscle strength in D28 mdx mice, produced approximately twofold lower MMP2 activity, indicating that increased MMP2 abundance is not required when muscle damage is attenuated. There was no difference in MMP9 abundance between age-matched WT and mdx mice ( P > 0.05). WT mice displayed decreased MMP9 abundance as they aged. While MMP9 may have a role during age-related skeletal muscle growth, it does not appear essential for degeneration/regeneration cycles in the mdx mouse. Our findings indicate that MMP2 plays a more active role than MMP9 in the degenerative phases of muscle fibers in D28 mdx mice.

Long-term administration of antisense oligonucleotides into the paraspinal muscles of mdx mice reduces kyphosis

2008 ◽  
Vol 105 (2) ◽  
pp. 662-668 ◽  
Author(s):  
Nicola Laws ◽  
Renée A. Cornford-Nairn ◽  
Nicole Irwin ◽  
Russell Johnsen ◽  
Susan Fletcher ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Rna Splicing ◽  
Antisense Oligonucleotides ◽  
Latissimus Dorsi ◽  
Stop Codon ◽  
Premature Stop Codon ◽  
Mdx Mouse ◽  
Mdx Mice ◽  
Paraspinal Muscles ◽  
Reading Frame

The mdx mouse model of muscular dystrophy has a premature stop codon preventing production of dystrophin. This results in a progressive phenotype causing centronucleation of skeletal muscle fibers, muscle weakness, and fibrosis and kyphosis. Antisense oligonucleotides alter RNA splicing to exclude the nonsense mutation, while still maintaining the open reading frame to produce a shorter, but partially functional dystrophin protein that should ameliorate the extent of pathology. The present study investigated the benefits of chronic treatment of mdx mice by once-monthly deep intramuscular injections of antisense oligonucleotides into paraspinal muscles. After 8 mo of treatment, mdx mice had reduced development of kyphosis relative to untreated mdx mice, a benefit that was retained until completion of the study at 18 mo of age (16 mo of treatment). This was accompanied by reduced centronucleation in the latissimus dorsi and intercostals muscles and reduced fibrosis in the diaphragm and latissimus dorsi. These benefits were accompanied by a significant increase in dystrophin production. In conclusion, chronic antisense oligonucleotide treatment provides clear and ongoing benefits to paralumbar skeletal muscle, with associated marked reduction in kyphosis.

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