TNF Inhibits Notch-1 in Skeletal Muscle Cells by Ezh2 and DNA Methylation Mediated Repression: Implications in Duchenne Muscular Dystrophy

Caveolae are vesicular invaginations of the plasma membrane. Caveolin-3 is the principal structural component of caveolae in skeletal muscle cells in vivo. We have recently generated caveolin-3 transgenic mice and demonstrated that overexpression of wild-type caveolin-3 in skeletal muscle fibers is sufficient to induce a Duchenne-like muscular dystrophy phenotype. In addition, we have shown that caveolin-3 null mice display mild muscle fiber degeneration and T-tubule system abnormalities. These data are consistent with the mild phenotype observed in Limb-girdle muscular dystrophy-1C (LGMD-1C) in humans, characterized by a ∼95% reduction of caveolin-3 expression. Thus, caveolin-3 transgenic and null mice represent valid mouse models to study Duchenne muscular dystrophy (DMD) and LGMD-1C, respectively, in humans. Here, we derived conditionally immortalized precursor skeletal muscle cells from caveolin-3 transgenic and null mice. We show that overexpression of caveolin-3 inhibits myoblast fusion to multinucleated myotubes and lack of caveolin-3 enhances the fusion process. M-cadherin and microtubules have been proposed to mediate the fusion of myoblasts to myotubes. Interestingly, we show that M-cadherin is downregulated in caveolin-3 transgenic cells and upregulated in caveolin-3 null cells. For the first time, variations of M-cadherin expression have been linked to a muscular dystrophy phenotype. In addition, we demonstrate that microtubules are disorganized in caveolin-3 null myotubes, indicating the importance of the cytoskeleton network in mediating the phenotype observed in these cells. Taken together, these results propose caveolin-3 as a key player in myoblast fusion and suggest that defects of the fusion process may represent additional molecular mechanisms underlying the pathogenesis of DMD and LGMD-1C in humans.

Download Full-text

Peptide-Functionalized Dendrimer Nanocarriers for Targeted Microdystrophin Gene Delivery

Pharmaceutics ◽

10.3390/pharmaceutics13122159 ◽

2021 ◽

Vol 13 (12) ◽

pp. 2159

Author(s):

Jessica Hersh ◽

José Manuel Condor Capcha ◽

Camila Iansen Irion ◽

Guerline Lambert ◽

Mauricio Noguera ◽

...

Keyword(s):

Skeletal Muscle ◽

Duchenne Muscular Dystrophy ◽

Muscular Dystrophy ◽

Gene Delivery ◽

Nuclear Membrane ◽

Targeted Delivery ◽

Intracellular Transport ◽

Muscle Cells ◽

Skeletal Muscle Cells

Gene therapy is a good alternative for determined congenital disorders; however, there are numerous limitations for gene delivery in vivo including targeted cellular uptake, intracellular trafficking, and transport through the nuclear membrane. Here, a modified G5 polyamidoamine (G5 PAMAM) dendrimer–DNA complex was developed, which will allow cell-specific targeting to skeletal muscle cells and transport the DNA through the intracellular machinery and the nuclear membrane. The G5 PAMAM nanocarrier was modified with a skeletal muscle-targeting peptide (SMTP), a DLC8-binding peptide (DBP) for intracellular transport, and a nuclear localization signaling peptide (NLS) for nuclear uptake, and polyplexed with plasmid DNA containing the GFP-tagged microdystrophin (µDys) gene. The delivery of µDys has been considered as a therapeutic modality for patients suffering from a debilitating Duchenne muscular dystrophy (DMD) disorder. The nanocarrier–peptide–DNA polyplexes were prepared with different charge ratios and characterized for stability, size, surface charge, and cytotoxicity. Using the optimized nanocarrier polyplexes, the transfection efficiency in vitro was determined by demonstrating the expression of the GFP and the µDys protein using fluorescence and Western blotting studies, respectively. Protein expression in vivo was determined by injecting an optimal nanocarrier polyplex formulation to Duchenne model mice, mdx4Cv. Ultimately, these nanocarrier polyplexes will allow targeted delivery of the microdystrophin gene to skeletal muscle cells and result in improved muscle function in Duchenne muscular dystrophy patients.

Download Full-text

New aspects of calcium signaling in skeletal muscle cells: implications in Duchenne muscular dystrophy

Biochimica et Biophysica Acta (BBA) - Proteins and Proteomics ◽

10.1016/s1570-9639(02)00442-9 ◽

2002 ◽

Vol 1600 (1-2) ◽

pp. 38-44 ◽

Cited By ~ 93

Author(s):

P Gailly

Keyword(s):

Skeletal Muscle ◽

Duchenne Muscular Dystrophy ◽

Muscular Dystrophy ◽

Calcium Signaling ◽

Muscle Cells ◽

Skeletal Muscle Cells

Download Full-text

Annexins and Membrane Repair Dysfunctions in Muscular Dystrophies

International Journal of Molecular Sciences ◽

10.3390/ijms22105276 ◽

2021 ◽

Vol 22 (10) ◽

pp. 5276

Author(s):

Coralie Croissant ◽

Romain Carmeille ◽

Charlotte Brévart ◽

Anthony Bouter

Keyword(s):

Skeletal Muscle ◽

Muscular Dystrophy ◽

Genetic Disorders ◽

Muscle Cells ◽

Cell Types ◽

Clinical Severity ◽

Skeletal Muscle Cells ◽

Muscular Dystrophies ◽

Membrane Repair ◽

Human Skeletal Muscle Cells

Muscular dystrophies constitute a group of genetic disorders that cause weakness and progressive loss of skeletal muscle mass. Among them, Miyoshi muscular dystrophy 1 (MMD1), limb girdle muscular dystrophy type R2 (LGMDR2/2B), and LGMDR12 (2L) are characterized by mutation in gene encoding key membrane-repair protein, which leads to severe dysfunctions in sarcolemma repair. Cell membrane disruption is a physiological event induced by mechanical stress, such as muscle contraction and stretching. Like many eukaryotic cells, muscle fibers possess a protein machinery ensuring fast resealing of damaged plasma membrane. Members of the annexins A (ANXA) family belong to this protein machinery. ANXA are small soluble proteins, twelve in number in humans, which share the property of binding to membranes exposing negatively-charged phospholipids in the presence of calcium (Ca2+). Many ANXA have been reported to participate in membrane repair of varied cell types and species, including human skeletal muscle cells in which they may play a collective role in protection and repair of the sarcolemma. Here, we discuss the participation of ANXA in membrane repair of healthy skeletal muscle cells and how dysregulation of ANXA expression may impact the clinical severity of muscular dystrophies.

Download Full-text

O.5 Skeletal muscle-specific calpain, p94/calpain 3, dynamically distributes in skeletal muscle cells to adapt to physical stress, defects of which cause muscular dystrophy

Neuromuscular Disorders ◽

10.1016/j.nmd.2010.07.014 ◽

2010 ◽

Vol 20 (9-10) ◽

pp. 598-599 ◽

Cited By ~ 1

Author(s):

K. Ojima ◽

Y. Ono ◽

N. Doi ◽

F. Kitamura ◽

S. Hata ◽

...

Keyword(s):

Skeletal Muscle ◽

Muscular Dystrophy ◽

Muscle Cells ◽

Physical Stress ◽

Skeletal Muscle Cells ◽

Calpain 3

Download Full-text

THREE-DIMENSIONAL STRUCTURE OF THE T-SYSTEM OF SKELETAL MUSCLE CELLS**This work was supported by grants from the Muscular Dystrophy Association /Henry M. Watts Center/ and the National Institutes of Health /HL-15835, Pennsylvania Muscle Institute/.

Molecular and Cellular Aspects of Muscle Function ◽

10.1016/b978-0-08-026817-0.50047-0 ◽

1981 ◽

pp. 299-311

Author(s):

Lee D. Peachey

Keyword(s):

Skeletal Muscle ◽

Muscular Dystrophy ◽

Three Dimensional ◽

Muscle Cells ◽

National Institutes Of Health ◽

Skeletal Muscle Cells ◽

Dimensional Structure ◽

Three Dimensional Structure ◽

T System

Download Full-text

Lipid exposure elicits differential responses in gene expression and DNA methylation in primary human skeletal muscle cells from severely obese women

Physiological Genomics ◽

10.1152/physiolgenomics.00065.2014 ◽

2015 ◽

Vol 47 (5) ◽

pp. 139-146 ◽

Cited By ~ 17

Author(s):

Jill M. Maples ◽

Jeffrey J. Brault ◽

Brian M. Shewchuk ◽

Carol A. Witczak ◽

Kai Zou ◽

...

Keyword(s):

Skeletal Muscle ◽

Dna Methylation ◽

Severe Obesity ◽

Muscle Cells ◽

Skeletal Muscle Cells ◽

Obese Women ◽

Impaired Ability ◽

Expression Of Genes ◽

Human Skeletal Muscle Cells ◽

Severely Obese

The skeletal muscle of obese individuals exhibits an impaired ability to increase the expression of genes linked with fatty acid oxidation (FAO) upon lipid exposure. The present study determined if this response could be attributed to differential DNA methylation signatures. RNA and DNA were isolated from primary human skeletal muscle cells (HSkMC) from lean and severely obese women following lipid incubation. mRNA expression and DNA methylation were quantified for genes that globally regulate FAO [PPARγ coactivator ( PGC-1α), peroxisome proliferator-activated receptors ( PPARs), nuclear respiratory factors ( NRFs)]. With lipid oversupply, increases in NRF-1, NRF-2, PPARα, and PPARδ expression were dampened in skeletal muscle from severely obese compared with lean women. The expression of genes downstream of the PPARs and NRFs also exhibited a pattern of not increasing as robustly upon lipid exposure with obesity. Increases in CpG methylation near the transcription start site with lipid oversupply were positively related to PPARδ expression; increases in methylation with lipid were depressed in HSkMC from severely obese women. With severe obesity, there is an impaired ability to upregulate global transcriptional regulators of FAO in response to lipid exposure. Transient changes in DNA methylation patterns and differences in the methylation signature with severe obesity may play a role in the transcriptional regulation of PPARδ in response to lipid. The persistence of differential responses to lipid in HSkMC derived from lean and obese subjects supports the possibility of stable epigenetic programming of skeletal muscle cells by the respective environments.

Download Full-text

Reduced mitochondrial fission and impaired energy metabolism in human primary skeletal muscle cells of Megaconial Congenital Muscular Dystrophy

Scientific Reports ◽

10.1038/s41598-021-97294-4 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Evrim Aksu-Menges ◽

Cemil Can Eylem ◽

Emirhan Nemutlu ◽

Merve Gizer ◽

Petek Korkusuz ◽

...

Keyword(s):

Skeletal Muscle ◽

Energy Metabolism ◽

Muscular Dystrophy ◽

Mitochondrial Dynamics ◽

Mitochondrial Fission ◽

Congenital Muscular Dystrophy ◽

Autosomal Recessive Disorder ◽

Muscle Cells ◽

Skeletal Muscle Cells ◽

Mitochondrial Energy Metabolism

AbstractMegaconial Congenital Muscular Dystrophy (CMD) is a rare autosomal recessive disorder characterized by enlarged mitochondria located mainly at the periphery of muscle fibers and caused by mutations in the Choline Kinase Beta (CHKB) gene. Although the pathogenesis of this disease is not well understood, there is accumulating evidence for the presence of mitochondrial dysfunction. In this study, we aimed to investigate whether imbalanced mitochondrial dynamics affects mitochondrial function and bioenergetic efficiency in skeletal muscle cells of Megaconial CMD. Immunofluorescence, confocal and transmission electron microscopy studies revealed impaired mitochondrial network, morphology, and localization in primary skeletal muscle cells of Megaconial CMD. The organelle disruption was specific only to skeletal muscle cells grown in culture. The expression levels of mitochondrial fission proteins (DRP1, MFF, FIS1) were found to be decreased significantly in both primary skeletal muscle cells and tissue sections of Megaconial CMD by Western blotting and/or immunofluorescence analysis. The metabolomic and fluxomic analysis, which were performed in Megaconial CMD for the first time, revealed decreased levels of phosphonucleotides, Krebs cycle intermediates, ATP, and altered energy metabolism pathways. Our results indicate that reduced mitochondrial fission and altered mitochondrial energy metabolism contribute to mitochondrial dysmorphology and dysfunction in the pathogenesis of Megaconial CMD.

Download Full-text

Mutant lamins cause nuclear envelope rupture and DNA damage in skeletal muscle cells

10.1101/364778 ◽

2018 ◽

Cited By ~ 4

Author(s):

Ashley J. Earle ◽

Tyler J. Kirby ◽

Gregory R. Fedorchak ◽

Philipp Isermann ◽

Jineet Patel ◽

...

Keyword(s):

Skeletal Muscle ◽

Dna Damage ◽

Muscular Dystrophy ◽

Nuclear Envelope ◽

Mouse Models ◽

Muscle Cells ◽

Skeletal Muscle Cells ◽

Causative Role ◽

Linc Complex

ABSTRACTMutations in the human LMNA gene, which encodes the nuclear envelope (NE) proteins lamins A and C, cause autosomal dominant Emery-Dreifuss muscular dystrophy, congenital muscular dystrophy, limb-girdle muscular dystrophy, and other diseases collectively known as laminopathies. The molecular mechanisms responsible for these diseases remain incompletely understood, but the muscle-specific defects suggest that mutations may render nuclei more susceptible to mechanical stress. Using three mouse models of muscle laminopathies, we found that Lmna mutations caused extensive NE abnormalities, consisting of chromatin protrusions into the cytoplasm and transient rupture of the NE in skeletal muscle cells. NE damage was associated with DNA damage, activation of DNA damage response pathways, and reduced viability. Intriguingly, NE damage resulted from nuclear migration in maturing skeletal muscle cells, rather than actomyosin contractility. NE damage and DNA damage was reduced by either depletion of kinesin-1 or disruption of the Linker of Nucleoskeleton and Cytoskeleton (LINC) complex. LINC complex disruption rescued myofiber function and viability in Lmna mutant myofibers, indicating that the myofiber dysfunction is the result of mechanically induced NE damage. The extent of NE damage and DNA damage in Lmna mouse models correlated with the disease onset and severity in vivo. Moreover, inducing DNA damage in wild-type muscle cells was sufficient to phenocopy the reduced cell viability of lamin A/C-deficient muscle cells, suggesting a causative role of DNA damage in disease pathogenesis. Corroborating the mouse model data, muscle biopsies from patients with LMNA muscular dystrophy revealed significant DNA damage compared to age-matched controls, particularly in severe cases of the disease. Taken together, these findings point to a new and important role of DNA damage as a pathogenic contributor for LMNA skeletal muscle diseases.

Download Full-text