Requirement for Down-Regulation of the CCAAT-binding Activity of the NF-Y  Transcription Factor during Skeletal Muscle Differentiation

NF-Y is composed of three subunits, NF-YA, NF-YB, and NF-YC, all required for DNA binding. All subunits are expressed in proliferating skeletal muscle cells, whereas NF-YA alone is undetectable in terminally differentiated cells in vitro. By immunohistochemistry, we show that the NF-YA protein is not expressed in the nuclei of skeletal and cardiac muscle cells in vivo. By chromatin immunoprecipitation experiments, we demonstrate herein that NF-Y does not bind to the CCAAT boxes of target promoters in differentiated muscle cells. Consistent with this, the activity of these promoters is down-regulated in differentiated muscle cells. Finally, forced expression of the NF-YA protein in cells committed to differentiate leads to an impairment in the down-regulation of cyclin A, cyclin B1, and cdk1 expression and is accompanied by a delay in myogenin expression. Thus, our results indicate that the suppression of NF-Y function is of crucial importance for the inhibition of several cell cycle genes and the induction of the early muscle-specific program in postmitotic muscle cells.

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FGF-2–dependent signaling activated in aged human skeletal muscle promotes intramuscular adipogenesis

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.2021013118 ◽

2021 ◽

Vol 118 (37) ◽

pp. e2021013118 ◽

Cited By ~ 1

Author(s):

Sebastian Mathes ◽

Alexandra Fahrner ◽

Umesh Ghoshdastider ◽

Hannes A. Rüdiger ◽

Michael Leunig ◽

...

Keyword(s):

Skeletal Muscle ◽

Transcriptional Activation ◽

Human Skeletal Muscle ◽

Muscle Growth ◽

Muscle Cells ◽

Adeno Associated Virus ◽

Adult Skeletal Muscle

Aged skeletal muscle is markedly affected by fatty muscle infiltration, and strategies to reduce the occurrence of intramuscular adipocytes are urgently needed. Here, we show that fibroblast growth factor-2 (FGF-2) not only stimulates muscle growth but also promotes intramuscular adipogenesis. Using multiple screening assays upstream and downstream of microRNA (miR)-29a signaling, we located the secreted protein and adipogenic inhibitor SPARC to an FGF-2 signaling pathway that is conserved between skeletal muscle cells from mice and humans and that is activated in skeletal muscle of aged mice and humans. FGF-2 induces the miR-29a/SPARC axis through transcriptional activation of FRA-1, which binds and activates an evolutionary conserved AP-1 site element proximal in the miR-29a promoter. Genetic deletions in muscle cells and adeno-associated virus–mediated overexpression of FGF-2 or SPARC in mouse skeletal muscle revealed that this axis regulates differentiation of fibro/adipogenic progenitors in vitro and intramuscular adipose tissue (IMAT) formation in vivo. Skeletal muscle from human donors aged >75 y versus <55 y showed activation of FGF-2–dependent signaling and increased IMAT. Thus, our data highlights a disparate role of FGF-2 in adult skeletal muscle and reveals a pathway to combat fat accumulation in aged human skeletal muscle.

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Expression of steroidogenic enzymes and synthesis of sex steroid hormones from DHEA in skeletal muscle of rats

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.00367.2006 ◽

2007 ◽

Vol 292 (2) ◽

pp. E577-E584 ◽

Cited By ~ 47

Author(s):

Katsuji Aizawa ◽

Motoyuki Iemitsu ◽

Seiji Maeda ◽

Subrina Jesmin ◽

Takeshi Otsuki ◽

...

Keyword(s):

Skeletal Muscle ◽

Steroid Hormones ◽

Skeletal Muscles ◽

Muscle Cells ◽

Sex Steroid Hormones ◽

Sex Steroid ◽

Dependent Manner ◽

Steroidogenic Enzymes

The functional importance of sex steroid hormones (testosterone and estrogens), derived from extragonadal tissues, has recently gained significant appreciation. Circulating dehydroepiandrosterone (DHEA) is peripherally taken up and converted to testosterone by 3β-hydroxysteroid dehydrogenase (HSD) and 17β-HSD, and testosterone in turn is irreversibly converted to estrogens by aromatase cytochrome P-450 (P450arom). Although sex steroid hormones have been implicated in skeletal muscle regulation and adaptation, it is unclear whether skeletal muscles have a local steroidogenic enzymatic machinery capable of metabolizing circulating DHEA. Thus, here, we investigate whether the three key steroidogenic enzymes (3β-HSD, 17β-HSD, and P450arom) are present in the skeletal muscle and are capable of generating sex steroid hormones. Consistent with our hypothesis, the present study demonstrates mRNA and protein expression of these enzymes in the skeletal muscle cells of rats both in vivo and in culture (in vitro). Importantly, we also show an intracellular formation of testosterone and estradiol from DHEA or testosterone in cultured muscle cells in a dose-dependent manner. These findings are novel and important in that they provide the first evidence showing that skeletal muscles are capable of locally synthesizing sex steroid hormones from circulating DHEA or testosterone.

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THE FINE STRUCTURE OF EMBRYONIC CHICK SKELETAL MUSCLE CELLS DIFFERENTIATED IN VITRO

The Journal of Cell Biology ◽

10.1083/jcb.35.2.445 ◽

1967 ◽

Vol 35 (2) ◽

pp. 445-453 ◽

Cited By ~ 91

Author(s):

Y. Shimada ◽

D. A. Fischman ◽

A. A. Moscona

Keyword(s):

Electron Microscopy ◽

Skeletal Muscle ◽

Fine Structure ◽

Muscle Cells ◽

Skeletal Muscle Cells ◽

Chick Embryos ◽

Embryonic Chick ◽

Developing Muscle

Dissociated myoblasts from 12-day chick embryos were cultured in monolayer, and the differentiation of skeletal muscle cells was studied by electron microscopy. The results have revealed a striking ultrastructural similarity between the in vivo and the in vitro developing muscle, particularly with respect to the myofibrils and sarcoplasmic reticulum. This study demonstrates that all the characteristic organelles of mature skeletal muscle can develop in vitro in the absence of nerves.

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Monosodium Iodoacetate delays regeneration and inhibits hypertrophy in skeletal muscle cells in vitro

10.1101/577866 ◽

2019 ◽

Author(s):

Rowan P. Rimington ◽

Darren J. Player ◽

Neil R.W. Martin ◽

Mark P. Lewis

Keyword(s):

Skeletal Muscle ◽

Muscle Cells ◽

Rodent Model ◽

Skeletal Muscle Cells ◽

Skeletal Muscle Regeneration ◽

Index Number ◽

Monosodium Iodoacetate ◽

The Impact

AbstractObjectiveOsteoarthritis (OA) is a musculoskeletal disease which contributes to severe morbidity. The monosodium iodoacetate (MIA) rodent model of OA is now well established, however the effect of MIA on surrounding tissues post injection has not been investigated and as such the impact on phenotypic development is unknown. The aim of this investigation was to examine the impact of MIA incubation on skeletal muscle cells in vitro, to provide an indication as to the potential influence of MIA administration of skeletal muscle in vivo.MethodsC2C12 skeletal muscle myotubes were treated with either 4.8μM MIA or 10μM Dexamethasone (DEX, positive atrophic control) up to 72hrs post differentiation and sampled for morphological and mRNA analyses.ResultsSignificant morphological effects (fusion index, number of myotubes and myotube width, p<0.05) were evident, demonstrating a hypertrophic phenotype in control (CON) compared to a hyperplasic phenotype in MIA and DEX. Increases in MAFbx mRNA were also evident between conditions, with post-hoc analysis demonstrating significance between CON and DEX (p<0.001), but not between CON and MIA (p>0.05).ConclusionsThese data indicate a significant impact of both DEX and MIA on regeneration and hypertrophy in vitro and suggest differential activating mechanisms. Future investigations should determine whether skeletal muscle regeneration and hypertrophy is affected in the in vivo rodent model and the potential impact this has on the OA phenotypic outcome.

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Embryonic Stem Cells Differentiated in Vitro as a Novel Source of Cells for Transplantation

Cell Transplantation ◽

10.1177/096368979600500205 ◽

1996 ◽

Vol 5 (2) ◽

pp. 131-143 ◽

Cited By ~ 38

Author(s):

Jonathan Dinsmore ◽

Judson Ratliff ◽

Terry Deacon ◽

Peyman Pakzaba ◽

Douglas Jacoby ◽

...

Keyword(s):

Skeletal Muscle ◽

Es Cells ◽

Embryonic Stem ◽

Muscle Cells ◽

Cell Types ◽

Cell Type ◽

Skeletal Myoblasts ◽

Muscle Cell Type

The controlled differentiation of mouse embryonic stem (ES) cells into near homogeneous populations of both neurons and skeletal muscle cells that can survive and function in vivo after transplantation is reported. We show that treatment of pluripotent ES cells with retinoic acid (RA) and dimethylsulfoxide (DMSO) induce differentiation of these cells into highly enriched populations of γ-aminobutyric acid (GABA) expressing neurons and skeletal myoblasts, respectively. For neuronal differentiation, RA alone is sufficient to induce ES cells to differentiate into neuronal cells that show properties of postmitotic neurons both in vitro and in vivo. In vivo function of RA-induced neuronal cells was demonstrated by transplantation into the quinolinic acid lesioned striatum of rats (a rat model for Huntington's disease), where cells integrated and survived for up to 6 wk. The response of embryonic stem cells to DMSO to form muscle was less dramatic than that observed for RA. DMSO-induced ES cells formed mixed populations of muscle cells composed of cardiac, smooth, and skeletal muscle instead of homogeneous populations of a single muscle cell type. To determine whether the response of ES cells to DMSO induction could be further controlled, ES cells were stably transfected with a gene coding for the muscle-specific regulatory factor, MyoD. When induced with DMSO, ES cells constitutively expressing high levels of MyoD differentiated exclusively into skeletal myoblasts (no cardiac or smooth muscle cells) that fused to form myotubes capable of spontaneous contraction. Thus, the specific muscle cell type formed was controlled by the expression of MyoD. These results provided evidence that the specific cell type formed (whether it be muscle, neuronal, or other cell types) can be controlled in vitro. Further, these results demonstrated that ES cells can provide a source of multiple differentiated cell types that can be used for transplantation.

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Differentiation of cardiac myocytes after mitogen withdrawal exhibits three sequential states of the ventricular growth response.

The Journal of Cell Biology ◽

10.1083/jcb.107.5.1911 ◽

1988 ◽

Vol 107 (5) ◽

pp. 1911-1918 ◽

Cited By ~ 51

Author(s):

H Ueno ◽

M B Perryman ◽

R Roberts ◽

M D Schneider

Keyword(s):

Dna Replication ◽

Dna Synthesis ◽

Muscle Cells ◽

Mitotic Division ◽

Neonatal Rat ◽

Growth Responses ◽

Down Regulation ◽

Cardiac Muscle Cells ◽

Serum Factors

During cardiac myogenesis, ventricular muscle cells lose the capacity to proliferate soon after birth. It is unknown whether this developmental block to mitotic division and DNA replication might involve irreversible repression of the cellular oncogene c-myc. Ventricular myocytes from 2 d-old rats continued to differentiate in vitro during 15 d of mitogen withdrawal, as shown by the formation of cross-striations, increased proportion of the muscle isoenzyme of creatine kinase, stable expression of alpha-cardiac actin and myosin heavy chain mRNAs, and appropriate down-regulation of alpha-skeletal actin mRNA. After mitogen withdrawal for 2 d, serum evoked both DNA synthesis and mitotic division; after 7 d, DNA replication was uncoupled from cell division; after 15 d, DNA synthesis itself was markedly attentuated. These three distinct phenotypic states resemble the sequential properties of growth found in the neonatal rat heart in vivo. Despite failure to induce DNA replication or division after 15 d of mitogen withdrawal, serum elicited both c-myc and alpha-skeletal actin as found during hypertrophy of the intact heart. The results agree with previous evidence that one or more functional pathways that transduce the effects of serum factors may persist in older cardiac muscle cells, and indicate that irreversible down-regulation of c-myc cannot be the basis for the loss of growth responses.

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Amelioration of High-Insulin-Induced Skeletal Muscle Cell Insulin Resistance by Resveratrol Is Linked to Activation of AMPK and Restoration of GLUT4 Translocation

Nutrients ◽

10.3390/nu12040914 ◽

2020 ◽

Vol 12 (4) ◽

pp. 914 ◽

Cited By ~ 3

Author(s):

Filip Vlavcheski ◽

Danja J. Den Hartogh ◽

Adria Giacca ◽

Evangelia Tsiani

Keyword(s):

Insulin Resistance ◽

Skeletal Muscle ◽

Glucose Transporter ◽

Muscle Cells ◽

Skeletal Muscle Cells ◽

Muscle Insulin Resistance ◽

Skeletal Muscle Insulin Resistance ◽

High Insulin

Insulin resistance, the hallmark of type 2 diabetes mellitus (T2DM), is linked to hyperinsulinemia, which develops to counterbalance initial peripheral hormone resistance. Studies indicate that chronically elevated levels of insulin lead to skeletal muscle insulin resistance by deregulating steps within the insulin signaling cascade. The polyphenol resveratrol (RSV) has been shown to have antidiabetic properties in vitro and in vivo. In the present study, we examined the effect of RSV on high insulin (HI)-induced insulin resistance in skeletal muscle cells in vitro and investigated the mechanisms involved. Parental and GLUT4myc-overexpressing L6 rat skeletal muscle cells were used. [3H]2-deoxyglucose (2DG) uptake was measured, and total and phosphorylated levels of specific proteins were examined by immunoblotting. Exposure of L6 cells to HI levels (100 nM) for 24 h decreased the acute-insulin-stimulated 2DG uptake, indicating insulin resistance. HI increased ser307 and ser636/639 phosphorylation of IRS-1 (to 184% ± 12% and 225% ± 28.9% of control, with p < 0.001 and p < 0.01, respectively) and increased the phosphorylation levels of mTOR (174% ± 6.7% of control, p < 0.01) and p70 S6K (228% ± 33.5% of control, p < 0.01). Treatment with RSV abolished these HI-induced responses. Furthermore, RSV increased the activation of AMPK and restored the insulin-mediated increase in plasma membrane GLUT4 glucose transporter levels. These data suggest that RSV has a potential to counteract the HI-induced muscle insulin resistance.

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Factors Controlling Movement of Skeletal Muscles

Leonardo ◽

10.1162/leon_a_01028 ◽

2015 ◽

Vol 48 (3) ◽

pp. 270-271

Author(s):

Miranda D. Grounds

Keyword(s):

Skeletal Muscle ◽

Extracellular Matrix ◽

Cell Membrane ◽

Skeletal Muscles ◽

Muscle Cells ◽

The Body ◽

Skeletal Muscle Cells ◽

3 Dimensional

The contraction of specialized skeletal muscle cells results in classic movements of bones and other parts of the body that are vital for life. There is exquisite control over the movement of diverse types of muscles. This paper indicates the way in which skeletal muscles (myofibres) are formed; then factors that contribute to generating the movement are outlined: these include the nerve, sarcomeres, cytoskeleton, cell membrane and the extracellular matrix. The factors controlling the movement of mature myofibres in 3-dimensional tissues in vivo are vastly more complex than for tissue cultured immature muscle cells in an artificial in vitro environment.

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Prion Infection of Muscle Cells In Vitro

Journal of Virology ◽

10.1128/jvi.02628-06 ◽

2007 ◽

Vol 81 (9) ◽

pp. 4615-4624 ◽

Cited By ~ 21

Author(s):

Wendy M. Dlakic ◽

Eric Grigg ◽

Richard A. Bessen

Keyword(s):

Skeletal Muscle ◽

Cell Line ◽

Muscle Cells ◽

C2c12 Cells ◽

C2c12 Myotubes ◽

C2c12 Myoblasts ◽

Peripheral Cell ◽

Prion Infection

ABSTRACT The prion agent has been detected in skeletal muscle of humans and animals with prion diseases. Here we report scrapie infection of murine C2C12 myoblasts and myotubes in vitro following coculture with a scrapie-infected murine neuroblastoma (N2A) cell line but not following incubation with a scrapie-infected nonneuronal cell line or a scrapie brain homogenate. Terminal differentiation of scrapie-infected C2C12 myoblasts into myotubes resulted in an increase in the expression of the disease-specific prion protein, PrPSc. The amount of scrapie infectivity or PrPSc in C2C12 myotubes was comparable to the levels found in scrapie-infected N2A cells, indicating that a high level of infection was established in muscle cells. Subclones of scrapie-infected C2C12 cells produced high levels of PrPSc in myotubes, and the C-terminal C2 polypeptide fragment of PrPSc was found based on deglycosylation and PrPSc-specific immunoprecipitation of cell lysates. This is the first report of a stable prion infection in muscle cells in vitro and of a long-term prion infection in a nondividing, differentiated peripheral cell type in culture. These in vitro studies also suggest that in vivo prion infection of skeletal muscle requires contact with prion-infected neurons or, possibly, nerve terminals.

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