Abstract P070: NEU3 Sialidase Overexpression Activates Cell Survival in Murine Skeletal Myoblasts C2C12 and Protects Them from Hypoxia

2011 ◽  
Vol 109 (suppl_1) ◽  
Author(s):  
Luigi Anastasia ◽  
Raffaella Scaringi ◽  
Nadia Papini ◽  
Andrea Garatti ◽  
Lorenzo Menicanti ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Egf Receptor ◽  
Day Treatment ◽  
Horse Serum ◽  
Critical Role ◽  
Cell Loss ◽  
Myoblast Differentiation ◽  
Signalling Pathways ◽  
Skeletal Myoblasts ◽  
Culturing Conditions

Membrane-bound sialidase NEU3 increase during skeletal muscle differentiation has been shown to protect myoblasts from apoptosis and drive the differentiation process [1]. Thus, the objective of this study was to assess whether up-regulation of NEU3 would enhance the ability of murine skeletal muscle cells to resist to hypoxia, ultimately opposing cell death. We found that C2C12 myoblasts overexpressing NEU3 (L-NEU3) became highly resistant to 1% oxygen or 200 mM deferoxamine induced hypoxia. Moreover, L-NEU3 myoblasts survived a seven-day treatment of combined hypoxia and low serum (2% horse serum used to induce myoblast differentiation), without any significant cell loss. On the contrary, wild type C2C12 could not resist to these culturing conditions and all died within 48h. Real Time PCR showed NEU3 expression increase during all hypoxic treatments both in C2C12 and L-NEU3 cells, suggesting an endogenous NEU3 activation under these conditions. Moreover, we found that NEU3 over-expression activated pro-survival signalling pathways through up-regulation and activation of EGF receptor. Overall, our data support the hypothesis that NEU3 may play a critical role in the response of skeletal myoblasts to hypoxia and the preservation of cell viability by activating pro-survival signalling pathways. [1] Anastasia L. et al. J.Biol.Chem. 2008, 283 (52): 36265–36271.

scholarly journals Zfp422 promotes skeletal muscle differentiation by regulating EphA7 to induce appropriate myoblast apoptosis

2019 ◽  
Vol 27 (5) ◽  
pp. 1644-1659 ◽  
Author(s):  
Yaping Nie ◽  
Shufang Cai ◽  
Renqiang Yuan ◽  
Suying Ding ◽  
Xumeng Zhang ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Zinc Finger ◽  
Muscle Development ◽  
Zinc Finger Protein ◽  
Critical Role ◽  
Muscle Differentiation ◽  
C2c12 Cells ◽  
Myoblast Differentiation ◽  
Skeletal Muscle Differentiation ◽  
Finger Protein

Abstract Zinc finger protein 422 (Zfp422) is a widely expressed zinc finger protein that serves as a transcriptional factor to regulate downstream gene expression, but until now, little is known about its roles in myogenesis. We found here that Zfp422 plays a critical role in skeletal muscle development and regeneration. It highly expresses in mouse skeletal muscle during embryonic development. Specific knockout of Zfp422 in skeletal muscle impaired embryonic muscle formation. Satellite cell-specific Zfp422 deletion severely inhibited muscle regeneration. Myoblast differentiation and myotube formation were suppressed in Zfp422-deleted C2C12 cells, isolated primary myoblasts, and satellite cells. Chromatin Immunoprecipitation Sequencing (ChIP-Seq) revealed that Zfp422 regulated ephrin type-A receptor 7 (EphA7) expression by binding an upstream 169-bp DNA sequence, which was proved to be an enhancer of EphA7. Knocking EphA7 down in C2C12 cells or deleting Zfp422 in myoblasts will inhibit cell apoptosis which is required for myoblast differentiation. These results indicate that Zfp422 is essential for skeletal muscle differentiation and fusion, through regulating EphA7 expression to maintain proper apoptosis.

scholarly journals Requirement for PINCH in Skeletal Myoblast Differentiation

2021 ◽  
Author(s):  
Siyi Xie ◽  
Chushan Fang ◽  
Yujie Gao ◽  
Jie Yan ◽  
Lina Luo ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Skeletal Muscles ◽  
Molecular Mechanisms ◽  
Myogenic Differentiation ◽  
Critical Role ◽  
The Body ◽  
Myoblast Differentiation ◽  
Skeletal Myogenesis ◽  
Congenital Myopathies

Abstract Background: Skeletal muscle is composed of bundles of myofibers ensheathed by extracellular matrix networks. Malformation of skeletal muscle during embryonic development results in congenital myopathies. Disease mechanisms of congenital myopathies remain unclear. PINCH, an adaptor of focal adhesion complex, plays essential roles in multiple cellular processes and organogenesis. Elucidation of the molecular mechanisms underlying skeletal myogenesis will offer new insights into pathogenesis of myopathies.Methods: We generated muscle-specific PINCH knock-out mice to study the functional role of PINCH in skeletal myogenesis. Histologic and Transmission Electron Microscopy analysis demonstrated that Impaired myogenic differentiation and maturation in mice with PINCH1 being ablated in skeletal muscle progenitors, and Ablation of PINCH1 and PINCH2 resulted in reduced size of muscle fibers and impaired multinucleation; Cell culture and immunostaining showed that defects in myoblast fusion and cytoskeleton assembly in PINCH double mutant mice; Western blotting showed that defects in expression of cytoskeleton proteins and proteins involved in myogenesis in DMUT skeletal muscles.Results: Double ablation of PINCH1 and PINCH2 resulted in early postnatal lethality with reduced size of skeletal muscles and detachment of diaphragm muscles from the body wall. Myofibers of PINCH mutant myofibers failed to undergo multinucleation and exhibited disrupted sarcomere structures. The mutant myoblasts in culture were able to adhere to newly formed myotubes, but impeded in cell fusion and subsequent sarcomere genesis and cytoskeleton organization. Consistent with this, expression of integrin β1 and some cytoskeleton proteins, and phosphorylation of ERK and AKT were significantly reduced in PINCH mutants. Expression of MRF4, the most highly expressed myogenic factor at late stages of myogenesis, was abolished in PINCH mutants, that could contribute to observed phenotypes. In addition, mice with PINCH1 being ablated in myogenic progenitors exhibited only mild centronuclear myopathic changes, suggesting a compensatory role of PINCH2 in myogenic differentiation, indicating a critical role of PINCH proteins in myogenic differentiation.Conclusion: Our results demonstrated an essential role of PINCH in skeletal myogenic differentiation.

scholarly journals Critical Role Played by Cyclin D3 in the MyoD-Mediated Arrest of Cell Cycle during Myoblast Differentiation

1999 ◽  
Vol 19 (7) ◽  
pp. 5203-5217 ◽  
Author(s):  
Carlo Cenciarelli ◽  
Francesca De Santa ◽  
Pier Lorenzo Puri ◽  
Elisabetta Mattei ◽  
Letizia Ricci ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Critical Role ◽  
Cyclin D3 ◽  
Myoblast Differentiation ◽  
Specific Gene ◽  
Gene Expressions ◽  
Adenovirus E1a ◽  
Skeletal Myoblasts ◽  
Myogenic Factors ◽  
Cell Cycle Inhibitors

ABSTRACT During the terminal differentiation of skeletal myoblasts, the activities of myogenic factors regulate not only tissue-specific gene expressions but also the exit from the cell cycle. The induction of cell cycle inhibitors such as p21 and pRb has been shown to play a prominent role in the growth arrest of differentiating myoblasts. Here we report that, at the onset of differentiation, activation by MyoD of the Rb, p21, and cyclin D3 genes occurs in the absence of new protein synthesis and with the requirement of the p300 transcriptional coactivator. In differentiated myocytes, cyclin D3 also becomes stabilized and is found nearly totally complexed with unphosphorylated pRb. The detection of complexes containing cyclin D3, cdk4, p21, and PCNA suggests that cdk4, along with PCNA, may get sequestered into high-order structures held together by pRb and cyclin D3. Cyclin D3 up-regulation and stabilization is inhibited by adenovirus E1A, and this correlates with the ability of E1A to promote pRb phosphorylation; conversely, the overexpression of cyclin D3 in differentiated myotubes counteracts the E1A-mediated reactivation of DNA synthesis. These results indicate that cyclin D3 critically contributes to the irreversible exit of differentiating myoblasts from the cell cycle.

scholarly journals Altered Skeletal Muscle Phenotypes in Calcineurin Aα and Aβ Gene-Targeted Mice

2003 ◽  
Vol 23 (12) ◽  
pp. 4331-4343 ◽  
Author(s):  
Stephanie A. Parsons ◽  
Benjamin J. Wilkins ◽  
Orlando F. Bueno ◽  
Jeffery D. Molkentin
Keyword(s):  
Skeletal Muscle ◽  
Fiber Type ◽  
Critical Role ◽  
Cell Types ◽  
Luciferase Reporter ◽  
Myoblast Differentiation ◽  
Activated T Cells ◽  
Slow Fiber ◽  
Calcineurin Signaling

ABSTRACT Calcineurin is a calcium-regulated serine-threonine protein phosphatase that controls developmental and inducible biological responses in diverse cell types, in part through activation of the transcription factor nuclear factor of activated T cells (NFAT). In skeletal muscle, calcineurin has been implicated in the regulation of myoblast differentiation, hypertrophy of mature myofibers, and fiber type switching in response to alterations in intracellular calcium concentration. However, considerable disagreement persists about the functional role of calcineurin signaling in each of these processes. Here we evaluated the molecular phenotypes of skeletal muscle from both calcineurin Aα and calcineurin Aβ gene-targeted mice. Calcineurin Aα was observed to be the predominant catalytic isoform expressed in nearly all skeletal muscles examined. Neither calcineurin Aα or Aβ null mice showed any gross growth-related alterations in skeletal muscle, nor was fiber size or number altered in glycolytic/fast muscle types. In contrast, both calcineurin Aα and Aβ gene-targeted mice demonstrated an alteration in myofiber number in the soleus, an oxidative/slow-type muscle. More significantly, calcineurin Aα and Aβ gene-targeted mice showed a dramatic down-regulation in the oxidative/slow fiber type program in multiple muscles (both slow and fast). Associated with this observation, NFAT-luciferase reporter transgenic mice showed significantly greater activity in slow fiber-containing muscles than in fast. However, only calcineurin Aα null mice showed a defect in NFAT nuclear occupancy or NFAT-luciferase transgene activity in vivo. Collectively, our results suggest that calcineurin signaling plays a critical role in regulating skeletal muscle fiber type switching but not hypertrophy. Our results also suggest that fiber type switching occurs through an NFAT-independent mechanism.

scholarly journals Decreased myoblast differentiation in chronic binge alcohol-administered simian immunodeficiency virus-infected male macaques: role of decreased miR-206

2017 ◽  
Vol 313 (3) ◽  
pp. R240-R250 ◽  
Author(s):  
L. Simon ◽  
S. M. Ford ◽  
K. Song ◽  
P. Berner ◽  
C. Vande Stouwe ◽  
...  
Keyword(s):  
Gene Expression ◽  
Skeletal Muscle ◽  
Simian Immunodeficiency Virus ◽  
Critical Role ◽  
Myoblast Differentiation ◽  
Differentiation Potential ◽  
Immunodeficiency Virus ◽  
Siv Infection ◽  
Underlying Mechanisms

Skeletal muscle stem cells play a critical role in regeneration of myofibers. We previously demonstrated that chronic binge alcohol (CBA) markedly attenuates myoblast differentiation potential and myogenic gene expression. Muscle-specific microRNAs (miRs) are implicated in regulation of myogenic genes. The aim of this study was to determine whether myoblasts isolated from asymptomatic CBA-administered simian immunodeficiency virus (SIV)-infected macaques treated with antiretroviral therapy (ART) showed similar impairments and, if so, to elucidate potential underlying mechanisms. Myoblasts were isolated from muscle at 11 mo after SIV infection from CBA/SIV macaques and from time-matched sucrose (SUC)-treated SIV-infected (SUC/SIV) animals and age-matched controls. Myoblast differentiation and myogenic gene expression were significantly decreased in myoblasts from SUC/SIV and CBA/SIV animals compared with controls. SIV and CBA decreased muscle-specific miR-206 in plasma and muscle and SIV decreased miR-206 expression in myoblasts, with no statistically significant changes in other muscle-specific miRs. These findings were associated with a significant increase in histone deacetylase 4 (HDAC4) and decrease in myogenic enhancer factor 2C (MEF2C) expression in CBA/SIV muscle. Transfection with miR-206 inhibitor decreased myotube differentiation, increased expression of HDAC4, and decreased MEF2C, suggesting a critical role of miR-206 in myogenesis. Moreover, HDAC4 was confirmed to be a direct miR-206 target. These results support a mechanistic role for decreased miR-206 in suppression of myoblast differentiation resulting from chronic alcohol and SIV infection. The parallel changes in skeletal muscle and circulating levels of miR-206 warrant studies to establish the possible use of plasma miR-206 as an indicator of impaired muscle function.

scholarly journals Smooth muscle myosin light chain kinase expression in cardiac and skeletal muscle

2000 ◽  
Vol 279 (5) ◽  
pp. C1656-C1664 ◽  
Author(s):  
B. Paul Herring ◽  
Shelley Dixon ◽  
Patricia J. Gallagher
Keyword(s):  
Skeletal Muscle ◽  
Smooth Muscle ◽  
Light Chain ◽  
Myosin Light Chain Kinase ◽  
Myosin Light Chain ◽  
Cardiac Myocyte ◽  
Cardiac Tissue ◽  
Specific Protein ◽  
Myoblast Differentiation ◽  
Adult Skeletal Muscle

The purpose of this study was to characterize myosin light chain kinase (MLCK) expression in cardiac and skeletal muscle. The only classic MLCK detected in cardiac tissue, purified cardiac myocytes, and in a cardiac myocyte cell line (AT1) was identical to the 130-kDa smooth muscle MLCK (smMLCK). A complex pattern of MLCK expression was observed during differentiation of skeletal muscle in which the 220-kDa-long or “nonmuscle” form of MLCK is expressed in undifferentiated myoblasts. Subsequently, during myoblast differentiation, expression of the 220-kDa MLCK declines and expression of this form is replaced by the 130-kDa smMLCK and a skeletal muscle-specific isoform, skMLCK in adult skeletal muscle. These results demonstrate that the skMLCK is the only tissue-specific MLCK, being expressed in adult skeletal muscle but not in cardiac, smooth, or nonmuscle tissues. In contrast, the 130-kDa smMLCK is ubiquitous in all adult tissues, including skeletal and cardiac muscle, demonstrating that, although the 130-kDa smMLCK is expressed at highest levels in smooth muscle tissues, it is not a smooth muscle-specific protein.

scholarly journals Adhesive multiplicity in the interaction of embryonic fibroblasts and myoblasts with extracellular matrices.

1984 ◽  
Vol 99 (4) ◽  
pp. 1398-1404 ◽  
Author(s):  
C Decker ◽  
R Greggs ◽  
K Duggan ◽  
J Stubbs ◽  
A Horwitz
Keyword(s):  
Skeletal Muscle ◽  
Cardiac Muscle ◽  
Cardiac Fibroblasts ◽  
Cell Types ◽  
Extracellular Matrices ◽  
Common Denominator ◽  
Cell Matrix ◽  
Skeletal Myoblasts ◽  
A Cell ◽  
Cell Matrix Adhesion

Neff et al. (1982, J. Cell Biol., 95:654-666) have described a monoclonal antibody, CSAT, directed against a cell surface antigen that participates in the adhesion of skeletal muscle to extracellular matrices. We used the same antibody to compare and parse the determinants of adhesion and morphology on myogenic and fibrogenic cells. We report here that the antigen is present on skeletal and cardiac muscle and on tendon, skeletal, dermal, and cardiac fibroblasts; however, its contribution to their morphology and adhesion is different. The antibody produces large alterations in the morphology and adhesion of skeletal myoblasts and tendon fibroblasts; in contrast, its effects on the cardiac fibroblasts are not readily detected. The effects of CSAT on the other cell types, i.e., dermal and skeletal fibroblasts, cardiac muscle, 5-bromodeoxyuridine-treated skeletal muscle, lie between these extremes. The effects of CSAT on the skeletal myoblasts depends on the calcium concentration in the growth medium and on the culture age. We interpret these differential responses to CSAT as revealing differences in the adhesion of the various cells to extracellular matrices. This interpretation is supported by parallel studies using quantitative assays of cell-matrix adhesion. The likely origin of these adhesive differences is the progressive display of different kinds of adhesion-related molecules and their organizational complexes on increasingly adhesive cells. The antigen to which CSAT is directed is present on all of the above cells and thus appears to be a lowest common denominator of their adhesion to extracellular matrices.

scholarly journals Effects of Cortisol and Dexamethasone on Insulin Signalling Pathways in Skeletal Muscle of the Ovine Fetus during Late Gestation

PLoS ONE ◽  
2012 ◽  
Vol 7 (12) ◽  
pp. e52363 ◽  
Author(s):  
Juanita K. Jellyman ◽  
Malgorzata S. Martin-Gronert ◽  
Roselle L. Cripps ◽  
Dino A. Giussani ◽  
Susan E. Ozanne ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Insulin Signalling ◽  
Late Gestation ◽  
Signalling Pathways ◽  
Ovine Fetus

Anemic Hypoxemia Reduces Myoblast Proliferation and Muscle Growth in Late Gestation Fetal Sheep

2021 ◽  
Author(s):  
Paul J. Rozance ◽  
Stephanie R Wesolowski ◽  
Sonnet S. Jonker ◽  
Laura D Brown
Keyword(s):  
Skeletal Muscle ◽  
Mrna Expression ◽  
Muscle Growth ◽  
Late Gestation ◽  
Whole Body ◽  
Myoblast Differentiation ◽  
Fetal Sheep ◽  
Body Protein ◽  
Skeletal Muscle Growth ◽  
Myoblast Proliferation

Fetal skeletal muscle growth requires myoblast proliferation, differentiation, and fusion into myofibers in addition to protein accretion for fiber hypertrophy. Oxygen is an important regulator of this process. Therefore, we hypothesized that fetal anemic hypoxemia would inhibit skeletal muscle growth. Studies were performed in late gestation fetal sheep that were bled to anemic, and therefore hypoxemic, conditions beginning at ~125 days of gestation (term = 148 days) for 9 ± 0 days (n=19) and compared to control fetuses (n=16). A metabolic study was performed on gestational day ~134 to measure fetal protein kinetic rates. Myoblast proliferation and myofiber area were determined in biceps femoris (BF), tibialis anterior (TA), and flexor digitorum superficialis (FDS) muscles. mRNA expression of muscle regulatory factors was determined in BF. Fetal arterial hematocrit and oxygen content were 28% and 52% lower, respectively, in anemic fetuses. Fetal weight and whole-body protein synthesis, breakdown, and accretion rates were not different between groups. Hindlimb length, however, was 7% shorter in anemic fetuses. TA and FDS muscles weighed less and FDS myofiber area was smaller in anemic fetuses compared to controls. The percentage of Pax7+ myoblasts that expressed Ki67 was lower in BF and tended to be lower in FDS from anemic fetuses indicating reduced myoblast proliferation. There was less MYOD and MYF6 mRNA expression in anemic vs. control BF consistent with reduced myoblast differentiation. These results indicate that fetal anemic hypoxemia reduced muscle growth. We speculate that fetal muscle growth may be improved by strategies that increase oxygen availability.

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