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1.
Sci Rep ; 14(1): 20699, 2024 09 05.
Artigo em Inglês | MEDLINE | ID: mdl-39237579

RESUMO

Insulin-like growth factor (IGF)-I mediates long-term activities that determine cell fate, including cell proliferation and differentiation. This study aimed to characterize the mechanisms by which IGF-I determines cell fate from the aspect of IGF-I signaling dynamics. In L6 myoblasts, myogenic differentiation proceeded under low IGF-I levels, whereas proliferation was enhanced under high levels. Mathematical and experimental analyses revealed that IGF-I signaling oscillated at low IGF-I levels but remained constant at high levels, suggesting that differences in IGF-I signaling dynamics determine cell fate. We previously reported that differential insulin receptor substrate (IRS)-1 levels generate a driving force for cell competition. Computational simulations and immunofluorescence analyses revealed that asynchronous IRS-1 protein oscillations were synchronized during myogenic processes through cell competition. Disturbances of cell competition impaired signaling synchronization and cell fusion, indicating that synchronization of IGF-I signaling oscillation is critical for myoblast cell fusion to form multinucleate myotubes.


Assuntos
Diferenciação Celular , Fator de Crescimento Insulin-Like I , Mioblastos , Transdução de Sinais , Fator de Crescimento Insulin-Like I/metabolismo , Mioblastos/metabolismo , Mioblastos/citologia , Animais , Linhagem Celular , Proliferação de Células , Desenvolvimento Muscular , Proteínas Substratos do Receptor de Insulina/metabolismo , Ratos , Fibras Musculares Esqueléticas/metabolismo , Fibras Musculares Esqueléticas/citologia , Fusão Celular
2.
Acta Neuropathol ; 148(1): 43, 2024 Sep 16.
Artigo em Inglês | MEDLINE | ID: mdl-39283487

RESUMO

Amyotrophic Lateral Sclerosis (ALS) is a multisystemic neurodegenerative disorder, with accumulating evidence indicating metabolic disruptions in the skeletal muscle preceding disease symptoms, rather than them manifesting as a secondary consequence of motor neuron (MN) degeneration. Hence, energy homeostasis is deeply implicated in the complex physiopathology of ALS and skeletal muscle has emerged as a key therapeutic target. Here, we describe intrinsic abnormalities in ALS skeletal muscle, both in patient-derived muscle cells and in muscle cell lines with genetic knockdown of genes related to familial ALS, such as TARDBP (TDP-43) and FUS. We found a functional impairment of myogenesis that parallels defects of glucose oxidation in ALS muscle cells. We identified FOXO1 transcription factor as a key mediator of these metabolic and functional features in ALS muscle, via gene expression profiling and biochemical surveys in TDP-43 and FUS-silenced muscle progenitors. Strikingly, inhibition of FOXO1 mitigated the impaired myogenesis in both the genetically modified and the primary ALS myoblasts. In addition, specific in vivo conditional knockdown of TDP-43 or FUS orthologs (TBPH or caz) in Drosophila muscle precursor cells resulted in decreased innervation and profound dysfunction of motor nerve terminals and neuromuscular synapses, accompanied by motor abnormalities and reduced lifespan. Remarkably, these phenotypes were partially corrected by foxo inhibition, bolstering the potential pharmacological management of muscle intrinsic abnormalities associated with ALS. The findings demonstrate an intrinsic muscle dysfunction in ALS, which can be modulated by targeting FOXO factors, paving the way for novel therapeutic approaches that focus on the skeletal muscle as complementary target tissue.


Assuntos
Esclerose Lateral Amiotrófica , Proteína Forkhead Box O1 , Músculo Esquelético , Esclerose Lateral Amiotrófica/metabolismo , Esclerose Lateral Amiotrófica/genética , Esclerose Lateral Amiotrófica/patologia , Humanos , Animais , Músculo Esquelético/metabolismo , Músculo Esquelético/patologia , Proteína Forkhead Box O1/metabolismo , Proteína Forkhead Box O1/genética , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Masculino , Proteína FUS de Ligação a RNA/genética , Proteína FUS de Ligação a RNA/metabolismo , Feminino , Drosophila , Desenvolvimento Muscular/fisiologia , Pessoa de Meia-Idade , Idoso , Neurônios Motores/metabolismo , Neurônios Motores/patologia , Mioblastos/metabolismo
3.
Stem Cell Res Ther ; 15(1): 302, 2024 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-39278936

RESUMO

BACKGROUND: Cell-based strategies are being explored as a therapeutic option for muscular dystrophies, using a variety of cell types from different origin and with different characteristics. Primary pericytes are multifunctional cells found in the capillary bed that exhibit stem cell-like and myogenic regenerative properties. This unique combination allows them to be applied systemically, presenting a promising opportunity for body-wide muscle regeneration. We previously reported the successful isolation of ALP+ pericytes from skeletal muscle of patients with myotonic dystrophy type 1 (DM1). These pericytes maintained normal growth parameters and myogenic characteristics in vitro despite the presence of nuclear (CUG)n RNA foci, the cellular hallmark of DM1. Here, we examined the behaviour of DM1 pericytes during myogenic differentiation. METHODS: DMPK (CTG)n repeat lengths in patient pericytes were assessed using small pool PCR, to be able to relate variation in myogenic properties and disease hallmarks to repeat expansion. Pericytes from unaffected controls and DM1 patients were cultured under differentiating conditions in vitro. In addition, the pericytes were grown in co-cultures with myoblasts to examine their regenerative capacity by forming hybrid myotubes. Finally, the effect of pericyte fusion on DM1 disease hallmarks was investigated. RESULTS: Small pool PCR analysis revealed the presence of somatic mosaicism in pericyte cell pools. Upon differentiation to myotubes, DMPK expression was upregulated, leading to an increase in nuclear foci sequestering MBNL1 protein. Remarkably, despite the manifestation of these disease biomarkers, patient-derived pericytes demonstrated myogenic potential in co-culture experiments comparable to unaffected pericytes and myoblasts. However, only the unaffected pericytes improved the disease hallmarks in hybrid myotubes. From 20% onwards, the fraction of unaffected nuclei in myotubes positively correlated with a reduction of the number of RNA foci and an increase in the amount of free MBNL1. CONCLUSIONS: Fusion of only a limited number of unaffected myogenic precursors to DM1 myotubes already ameliorates cellular disease hallmarks, offering promise for the development of cell transplantation strategies to lower disease burden.


Assuntos
Diferenciação Celular , Fibras Musculares Esqueléticas , Distrofia Miotônica , Miotonina Proteína Quinase , Pericitos , Humanos , Distrofia Miotônica/metabolismo , Distrofia Miotônica/genética , Distrofia Miotônica/terapia , Distrofia Miotônica/patologia , Fibras Musculares Esqueléticas/metabolismo , Pericitos/metabolismo , Miotonina Proteína Quinase/genética , Miotonina Proteína Quinase/metabolismo , Mioblastos/metabolismo , Mioblastos/citologia , Desenvolvimento Muscular , Células Cultivadas , Masculino , Adulto , Feminino , Técnicas de Cocultura , Pessoa de Meia-Idade , Fusão Celular
4.
Int J Mol Sci ; 25(17)2024 Aug 31.
Artigo em Inglês | MEDLINE | ID: mdl-39273445

RESUMO

Limb muscle is responsible for physical activities and myogenic cell migration during embryogenesis is indispensable for limb muscle formation. Maternal obesity (MO) impairs prenatal skeletal muscle development, but the effects of MO on myogenic cell migration remain to be examined. C57BL/6 mice embryos were collected at E13.5. The GeoMx DSP platform was used to customize five regions along myogenic cell migration routes (myotome, dorsal/ventral limb, limb stroma, limb tip), and data were analyzed by GeomxTools 3.6.0. A total of 2224 genes were down-regulated in the MO group. The GO enrichment analysis showed that MO inhibited migration-related biological processes. The signaling pathways guiding myogenic migration such as hepatocyte growth factor signaling, fibroblast growth factor signaling, Wnt signaling and GTPase signaling were down-regulated in the MO E13.5 limb tip. Correspondingly, the expression levels of genes involved in myogenic cell migration, such as Pax3, Gab1, Pxn, Tln2 and Arpc, were decreased in the MO group, especially in the dorsal and ventral sides of the limb. Additionally, myogenic differentiation-related genes were down-regulated in the MO limb. MO impedes myogenic cell migration and differentiation in the embryonic limb, providing an explanation for the impairment of fetal muscle development and offspring muscle function due to MO.


Assuntos
Diferenciação Celular , Movimento Celular , Desenvolvimento Muscular , Obesidade Materna , Animais , Movimento Celular/genética , Camundongos , Feminino , Desenvolvimento Muscular/genética , Diferenciação Celular/genética , Gravidez , Obesidade Materna/metabolismo , Obesidade Materna/genética , Camundongos Endogâmicos C57BL , Regulação da Expressão Gênica no Desenvolvimento , Transcriptoma , Desenvolvimento Embrionário/genética , Extremidades/embriologia , Perfilação da Expressão Gênica , Transdução de Sinais , Músculo Esquelético/metabolismo , Músculo Esquelético/embriologia
5.
BMC Biol ; 22(1): 194, 2024 Sep 11.
Artigo em Inglês | MEDLINE | ID: mdl-39256800

RESUMO

BACKGROUND: DNA G-quadruplexes (G4s) represent a distinctive class of non-canonical DNA secondary structures. Despite their recognition as potential therapeutic targets in some cancers, the developmental role of G4 structures remains enigmatic. Mammalian embryonic myogenesis studies are hindered by limitations, prompting the use of chicken embryo-derived myoblasts as a model to explore G4 dynamics. This study aims to reveal the embryonic G4s landscape and elucidate the underlying mechanisms for candidate G4s that influence embryonic myogenesis. RESULTS: This investigation unveils a significant reduction in G4s abundance during myogenesis. G4s stabilizer pyridostatin impedes embryonic myogenesis, emphasizing the regulatory role of G4s in this process. G4 Cut&Tag sequencing and RNA-seq analyses identify potential G4s and DEGs influencing embryonic myogenesis. Integration of G4 and DEG candidates identifies 32 G4s located in promoter regions capable of modulating gene transcription. WGBS elucidates DNA methylation dynamics during embryonic myogenesis. Coordinating transcriptome data with DNA G4s and DNA methylation profiles constructs a G4-DMR-DEG network, revealing nine interaction pairs. Notably, the NFATC2 promoter region sequence is confirmed to form a G4 structure, reducing promoter mCpG content and upregulating NFATC2 transcriptional activity. CONCLUSIONS: This comprehensive study unravels the first embryonic genomic G4s landscape, highlighting the regulatory role of NFATC2 G4 in orchestrating transcriptional activity through promoter DNA methylation during myogenesis.


Assuntos
Quadruplex G , Desenvolvimento Muscular , Desenvolvimento Muscular/genética , Animais , Embrião de Galinha , Mioblastos/metabolismo , Metilação de DNA
6.
Elife ; 132024 Sep 09.
Artigo em Inglês | MEDLINE | ID: mdl-39248331

RESUMO

SRSF2 plays a dual role, functioning both as a transcriptional regulator and a key player in alternative splicing. The absence of Srsf2 in MyoD + progenitors resulted in perinatal mortality in mice, accompanied by severe skeletal muscle defects. SRSF2 deficiency disrupts the directional migration of MyoD progenitors, causing them to disperse into both muscle and non-muscle regions. Single-cell RNA-sequencing analysis revealed significant alterations in Srsf2-deficient myoblasts, including a reduction in extracellular matrix components, diminished expression of genes involved in ameboid-type cell migration and cytoskeleton organization, mitosis irregularities, and premature differentiation. Notably, one of the targets regulated by Srsf2 is the serine/threonine kinase Aurka. Knockdown of Aurka led to reduced cell proliferation, disrupted cytoskeleton, and impaired differentiation, reflecting the effects seen with Srsf2 knockdown. Crucially, the introduction of exogenous Aurka in Srsf2-knockdown cells markedly alleviated the differentiation defects caused by Srsf2 knockdown. Furthermore, our research unveiled the role of Srsf2 in controlling alternative splicing within genes associated with human skeletal muscle diseases, such as BIN1, DMPK, FHL1, and LDB3. Specifically, the precise knockdown of the Bin1 exon17-containing variant, which is excluded following Srsf2 depletion, profoundly disrupted C2C12 cell differentiation. In summary, our study offers valuable insights into the role of SRSF2 in governing MyoD progenitors to specific muscle regions, thereby controlling their differentiation through the regulation of targeted genes and alternative splicing during skeletal muscle development.


Assuntos
Diferenciação Celular , Movimento Celular , Desenvolvimento Muscular , Músculo Esquelético , Proteína MyoD , Fatores de Processamento de Serina-Arginina , Animais , Camundongos , Fatores de Processamento de Serina-Arginina/metabolismo , Fatores de Processamento de Serina-Arginina/genética , Desenvolvimento Muscular/genética , Músculo Esquelético/metabolismo , Músculo Esquelético/crescimento & desenvolvimento , Proteína MyoD/metabolismo , Proteína MyoD/genética , Aurora Quinase A/metabolismo , Aurora Quinase A/genética , Mioblastos/metabolismo , Processamento Alternativo
7.
Biochemistry (Mosc) ; 89(7): 1325-1335, 2024 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-39218028

RESUMO

Effect of succinic acid on the processes of myogenesis was investigated in the study with the cells of C2C12 line. In the concentration range 10-1000 µM, succinic acid stimulated the process of myogenic differentiation, increasing the levels of myogenesis factors MyoD (at all stages of myogenesis) and myogenin (at the stage of terminal differentiation). Presence of the succinate receptors SUCNR1 was revealed in the C2C12 cells using Western blotting, level of which decreased during myogenesis. When succinic acid was added to the cells, the level of intracellular succinate did not change significantly and decreased during myogenic differentiation. Using a specific Gai protein inhibitor, pertussis toxin, it was found that stimulation of myogenesis in the C2C12 cells under the action of succinic acid is realized through SUCNR1-Gai interaction.


Assuntos
Diferenciação Celular , Desenvolvimento Muscular , Ácido Succínico , Ácido Succínico/metabolismo , Desenvolvimento Muscular/efeitos dos fármacos , Animais , Camundongos , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular , Receptores Acoplados a Proteínas G/metabolismo , Proteína MyoD/metabolismo , Miogenina/metabolismo
8.
Development ; 151(20)2024 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-39114943

RESUMO

Vertebrates and tunicates are sister groups that share a common fusogenic factor, Myomaker (Mymk), that drives myoblast fusion and muscle multinucleation. Yet they are divergent in when and where they express Mymk. In vertebrates, all developing skeletal muscles express Mymk and are obligately multinucleated. In tunicates, Mymk is expressed only in post-metamorphic multinucleated muscles, but is absent from mononucleated larval muscles. In this study, we demonstrate that cis-regulatory sequence differences in the promoter region of Mymk underlie the different spatiotemporal patterns of its transcriptional activation in tunicates and vertebrates. Although in vertebrates myogenic regulatory factors (MRFs) such as MyoD1 alone are required and sufficient for Mymk transcription in all skeletal muscles, we show that transcription of Mymk in post-metamorphic muscles of the tunicate Ciona requires the combinatorial activity of MRF, MyoD and Early B-cell Factor (Ebf). This macroevolutionary difference appears to be encoded in cis, likely due to the presence of a putative Ebf-binding site adjacent to predicted MRF binding sites in the Ciona Mymk promoter. We further discuss how Mymk and myoblast fusion might have been regulated in the last common ancestor of tunicates and vertebrates, for which we propose two models.


Assuntos
Regiões Promotoras Genéticas , Animais , Regiões Promotoras Genéticas/genética , Proteína MyoD/metabolismo , Proteína MyoD/genética , Regulação da Expressão Gênica no Desenvolvimento , Músculo Esquelético/metabolismo , Fatores de Regulação Miogênica/metabolismo , Fatores de Regulação Miogênica/genética , Urocordados/genética , Urocordados/embriologia , Desenvolvimento Muscular/genética
9.
Int J Mol Sci ; 25(16)2024 Aug 22.
Artigo em Inglês | MEDLINE | ID: mdl-39201775

RESUMO

With the continuous improvement in living standards, people's demand for high-quality meat is increasing. Ningxiang pig has delicious meat of high nutritional value, and is loved by consumers. However, its slow growth and low meat yield seriously restrict its efficient utilization. Gene expression is the internal driving force of life activities, so in order to fundamentally improve its growth rate, it is key to explore the molecular mechanism of skeletal muscle development in Ningxiang pigs. In this paper, Ningxiang boars were selected in four growth stages (30 days: weaning period, 90 days: nursing period, 150 days: early fattening period, and 210 days: late fattening period), and the longissimus dorsi (LD) muscle was taken from three boars in each stage. The fatty acid content, amino acid content, muscle fiber diameter density and type of LD were detected by gas chromatography, acidolysis, hematoxylin eosin (HE) staining and immunofluorescence (IF) staining. After transcription sequencing, weighted gene co-expression network analysis (WGCNA) combined with the phenotype of the LD was used to explore the key genes and signaling pathways affecting muscle development. The results showed that 10 modules were identified by WGCNA, including 5 modules related to muscle development stage, module characteristics of muscle fiber density, 5 modules characteristic of muscle fiber diameter, and a module characteristic of palmitoleic acid (C16:1) and linoleic acid (C18:2n6C). Gene ontology (GO) enrichment analysis found that 52 transcripts relating to muscle development were enriched in these modules, including 44 known genes and 8 novel genes. The Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis showed that these genes were enriched in the auxin, estrogen and cyclic guanosine monophosphate-protein kinase G (cGMP-PKG) pathways. Twelve of these genes were transcription factors, there were interactions among 20 genes, and the interactions among 11 proteins in human, pig and mouse were stable. To sum up, through the integrated analysis of phenotype and transcriptome, this paper analyzed the key genes and possible regulatory networks of skeletal muscle development in Ningxiang pigs at various stages, to provide a reference for the in-depth study of skeletal muscle development.


Assuntos
Redes Reguladoras de Genes , Desenvolvimento Muscular , Músculo Esquelético , Animais , Suínos , Músculo Esquelético/metabolismo , Músculo Esquelético/crescimento & desenvolvimento , Desenvolvimento Muscular/genética , Perfilação da Expressão Gênica , Ácidos Graxos/metabolismo , Transcriptoma , Transdução de Sinais , Regulação da Expressão Gênica no Desenvolvimento , Masculino
11.
Life Sci ; 354: 122941, 2024 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-39098595

RESUMO

AIMS: Study of the role of mitochondria-generated reactive oxygen species (mtROS) and mitochondrial polarization in mitochondrial fragmentation at the initial stages of myogenesis. MAIN METHODS: Mitochondrial morphology, Drp1 protein phosphorylation, mitochondrial electron transport chain components content, mtROS and mitochondrial lipid peroxidation levels, and mitochondrial polarization were evaluated on days 1 and 2 of human MB135 myoblasts differentiation. A mitochondria-targeted antioxidant SkQ1 was used to elucidate the effect of mtROS on mitochondria. KEY FINDINGS: In immortalized human MB135 myoblasts, mitochondrial fragmentation began on day 1 of differentiation before the myoblast fusion. This fragmentation was preceded by dephosphorylation of p-Drp1 (Ser-637). On day 2, an increase in the content of some mitochondrial proteins was observed, indicating mitochondrial biogenesis stimulation. Furthermore, we found that myogenic differentiation, even on day 1, was accompanied both by an increased production of mtROS, and lipid peroxidation of the inner mitochondrial membrane. SkQ1 blocked these effects and partially reduced the level of mitochondrial fragmentation, but did not affect the dephosphorylation of p-Drp1 (Ser-637). Importantly, mitochondrial fragmentation at early stages of MB135 differentiation was not accompanied by depolarization, as an important stimulus for mitochondrial fragmentation. SIGNIFICANCE: Mitochondrial fragmentation during early myogenic differentiation depends on mtROS production rather than mitochondrial depolarization. SkQ1 only partially inhibited mitochondrial fragmentation, without significant effects on mitophagy or early myogenic differentiation.


Assuntos
Diferenciação Celular , Peroxidação de Lipídeos , Mitocôndrias , Mioblastos , Espécies Reativas de Oxigênio , Humanos , Espécies Reativas de Oxigênio/metabolismo , Diferenciação Celular/efeitos dos fármacos , Mioblastos/metabolismo , Mioblastos/citologia , Mioblastos/efeitos dos fármacos , Mitocôndrias/metabolismo , Mitocôndrias/efeitos dos fármacos , Peroxidação de Lipídeos/efeitos dos fármacos , Desenvolvimento Muscular/fisiologia , Desenvolvimento Muscular/efeitos dos fármacos , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Dinaminas/metabolismo , Fosforilação , Linhagem Celular
12.
Int J Mol Sci ; 25(15)2024 Jul 30.
Artigo em Inglês | MEDLINE | ID: mdl-39125909

RESUMO

Skeletal muscle, which is predominantly constituted by multinucleated muscle fibers, plays a pivotal role in sustaining bodily movements and energy metabolism. Myoblasts, which serve as precursor cells for differentiation and fusion into muscle fibers, are of critical importance in the exploration of the functional genes associated with embryonic muscle development. However, the in vitro proliferation of primary myoblasts is inherently constrained. In this study, we achieved a significant breakthrough by successfully establishing a chicken myoblast cell line through the introduction of the exogenous chicken telomerase reverse transcriptase (chTERT) gene, followed by rigorous G418-mediated pressure screening. This newly developed cell line, which was designated as chTERT-myoblasts, closely resembled primary myoblasts in terms of morphology and exhibited remarkable stability in culture for at least 20 generations of population doublings without undergoing malignant transformation. In addition, we conducted an exhaustive analysis that encompassed cellular proliferation, differentiation, and transfection characteristics. Our findings revealed that the chTERT-myoblasts had the ability to proliferate, differentiate, and transfect after multiple rounds of population doublings. This achievement not only furnished a valuable source of homogeneous avian cell material for investigating embryonic muscle development, but also provided valuable insights and methodologies for establishing primary cell lines.


Assuntos
Diferenciação Celular , Proliferação de Células , Galinhas , Mioblastos , Telomerase , Animais , Mioblastos/citologia , Mioblastos/metabolismo , Linhagem Celular , Telomerase/metabolismo , Telomerase/genética , Desenvolvimento Muscular/genética , Técnicas de Cultura de Células/métodos , Transfecção , Embrião de Galinha
13.
Mol Cell Biol ; 44(9): 391-409, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-39133076

RESUMO

Myogenesis is a highly orchestrated process whereby muscle precursor cells, myoblasts, develop into muscle fibers to form skeletal muscle during embryogenesis and regenerate adult muscle. Here, we studied the RNA-binding protein FUS (fused in sarcoma), which has been implicated in muscular and neuromuscular pathologies but is poorly characterized in myogenesis. Given that FUS levels declined in human and mouse models of skeletal myogenesis, and that silencing FUS enhanced myogenesis, we hypothesized that FUS might be a repressor of myogenic differentiation. Interestingly, overexpression of FUS delayed myogenesis, accompanied by slower production of muscle differentiation markers. To identify the mechanisms through which FUS inhibits myogenesis, we uncovered RNA targets of FUS by ribonucleoprotein immunoprecipitation (RIP) followed by RNA-sequencing (RNA-seq) analysis. Stringent selection of the bound transcripts uncovered Tnnt1 mRNA, encoding troponin T1 (TNNT1), as a major effector of FUS influence on myogenesis. We found that in myoblasts, FUS retained Tnnt1 mRNA in the nucleus, preventing TNNT1 expression; however, reduction of FUS during myogenesis or by silencing FUS released Tnnt1 mRNA for export to the cytoplasm, enabling TNNT1 translation and promoting myogenesis. We propose that FUS inhibits myogenesis by suppressing TNNT1 expression through a mechanism of nuclear Tnnt1 mRNA retention.


Assuntos
Diferenciação Celular , Desenvolvimento Muscular , Mioblastos , Proteína FUS de Ligação a RNA , Troponina T , Desenvolvimento Muscular/genética , Proteína FUS de Ligação a RNA/metabolismo , Proteína FUS de Ligação a RNA/genética , Animais , Camundongos , Humanos , Troponina T/metabolismo , Troponina T/genética , Mioblastos/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Músculo Esquelético/metabolismo , Linhagem Celular
14.
Artigo em Inglês | MEDLINE | ID: mdl-39179021

RESUMO

Fish skeletal muscle is a component of the human diet, and understanding the mechanisms that control muscle growth can contribute to improving production in this sector and benefits the human health. In this sense, fish such as tambacu can represent a valuable source for exploring muscle growth regulators due to the indeterminate muscle growth pattern. In this context, the genes responsible for the indeterminate and determinate muscle growth pattern of fish are little explored, with piwi genes being possible candidates involved with these growth patterns. Piwi genes are associated with the proliferation and self-renewal of germ cells, and there are descriptions of these same functions in somatic cells from different tissues. However, little is known about the function of these genes in fish somatic cells. Considering this, our objective was to analyze the expression pattern of piwi 1 and 2 genes in cardiac muscle, skeletal muscle, liver, and gonad of zebrafish (species with determinate growth) and tambacu (species with indeterminate growth). We observed a distinct expression of piwi1 and piwi2 between tambacu and zebrafish, with both genes more expressed in tambacu in all tissues evaluated. Piwi genes can represent potential candidates involved with indeterminate muscle growth control.


Assuntos
Proteínas Argonautas , Músculo Esquelético , Peixe-Zebra , Animais , Peixe-Zebra/genética , Peixe-Zebra/crescimento & desenvolvimento , Peixe-Zebra/metabolismo , Proteínas Argonautas/genética , Proteínas Argonautas/metabolismo , Músculo Esquelético/metabolismo , Músculo Esquelético/crescimento & desenvolvimento , Desenvolvimento Muscular/genética , Regulação da Expressão Gênica no Desenvolvimento , Proteínas de Peixes/genética , Proteínas de Peixes/metabolismo , Fígado/metabolismo , Fígado/crescimento & desenvolvimento , Masculino
15.
Dev Biol ; 516: 82-95, 2024 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-39111615

RESUMO

The Myocyte enhancer factor-2 (MEF2) transcription factor plays a vital role in orchestrating muscle differentiation. While MEF2 cannot effectively induce myogenesis in naïve cells, it can potently accelerate myogenesis in mesodermal cells. This includes in Drosophila melanogaster imaginal disc myoblasts, where triggering premature muscle gene expression in these adult muscle progenitors has become a paradigm for understanding the regulation of the myogenic program. Here, we investigated the global consequences of MEF2 overexpression in the imaginal wing disc myoblasts, by combining RNA-sequencing with RT-qPCR and immunofluorescence. We observed the formation of sarcomere-like structures that contained both muscle and cytoplasmic myosin, and significant upregulation of muscle gene expression, especially genes essential for myofibril formation and function. These transcripts were functional since numerous myofibrillar proteins were detected in discs using immunofluorescence. Interestingly, muscle genes whose expression is restricted to the adult stages were not activated in these adult myoblasts. These studies confirm a broad activation of the myogenic program in response to MEF2 expression and suggest that additional regulatory factors are required for promoting the adult muscle-specific program. Our findings contribute to understanding the regulatory mechanisms governing muscle development and highlight the multifaceted role of MEF2 in orchestrating this intricate process.


Assuntos
Proteínas de Drosophila , Drosophila melanogaster , Regulação da Expressão Gênica no Desenvolvimento , Discos Imaginais , Fatores de Transcrição MEF2 , Desenvolvimento Muscular , Mioblastos , Animais , Drosophila melanogaster/metabolismo , Drosophila melanogaster/genética , Desenvolvimento Muscular/genética , Proteínas de Drosophila/metabolismo , Proteínas de Drosophila/genética , Mioblastos/metabolismo , Discos Imaginais/metabolismo , Fatores de Transcrição MEF2/metabolismo , Fatores de Transcrição MEF2/genética , Asas de Animais/metabolismo , Asas de Animais/crescimento & desenvolvimento , Diferenciação Celular , Fatores de Regulação Miogênica
16.
Differentiation ; 139: 100805, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-39214748

RESUMO

Fibroblast Growth Factor 6 (FGF6), also referred to as HST2 or HBGF6, is a member of the Fibroblast Growth Factor (FGF), the Heparin Binding Growth Factor (HBGF) and the Heparin Binding Secretory Transforming Gene (HST) families. The genomic and protein structure of FGF6 is highly conserved among varied species, as is its expression in muscle and muscle progenitor cells. Like other members of the FGF family, FGF6 regulates cell proliferation, differentiation, and migration. Specifically, it plays key roles in myogenesis and muscular regeneration, angiogenesis, along with iron transport and lipid metabolism. Similar to others from the FGF family, FGF6 also possesses oncogenic transforming activity, and as such is implicated in a variety of cancers.


Assuntos
Diferenciação Celular , Fator 6 de Crescimento de Fibroblastos , Humanos , Animais , Fator 6 de Crescimento de Fibroblastos/genética , Fator 6 de Crescimento de Fibroblastos/metabolismo , Desenvolvimento Muscular/genética , Proliferação de Células , Movimento Celular , Neoplasias/metabolismo , Neoplasias/genética , Neoplasias/patologia
17.
Acta Biomater ; 186: 95-107, 2024 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-39117115

RESUMO

A goal of regenerative engineering is the rational design of materials to restore the structure-function relationships that drive reparative programs in damaged tissues. Despite the widespread use of extracellular matrices for engineering tissues, their application has been limited by a narrow range of tunable features. The primary objective of this study is to develop a versatile platform for evaluating tissue-specific cellular interactions using Type I collagen scaffolds with highly tunable biophysical properties. The kinetics of collagen fibrillogenesis were modulated through a combination of varied shear rate and pH during neutralization, to achieve a broad range of fibril anisotropy, porosity, diameter, and storage modulus. The role that each of these properties play in guiding muscle, bone, and vascular cell types was comprehensively identified, and informed the in vitro generation of three distinct musculoskeletal engineered constructs. Myogenesis was highly regulated by smaller fibrils and larger storage moduli, endothelial inflammatory phenotype was predominantly guided by fibril anisotropy, and osteogenesis was enhanced by highly porous collagen with larger fibrils. This study introduces a novel approach for dynamically modulating Type I collagen materials and provides a robust platform for investigating cell-material interactions, offering insights for the future rational design of tissue-specific regenerative biomaterials. STATEMENT OF SIGNIFICANCE: The biophysical properties of regenerative materials facilitate key cell-substrate interactions that can guide the morphology, phenotype, and biological response of cells. In this study, we describe the fabrication of an engineered collagen hydrogel that can be modified to exhibit control over a wide range of biophysical features, including fibril organization and size, nanoscale porosity, and mechanics. We identified the unique combination of collagen features that optimally promote regenerative muscle, bone, and vascular cell types while also delineating the properties that hinder these same cellular responses. This study presents a highly accessible method to control the biophysical properties of collagen hydrogels that can be adapted for a broad range of tissue engineering and regenerative applications.


Assuntos
Nanofibras , Osteogênese , Osteogênese/efeitos dos fármacos , Humanos , Nanofibras/química , Animais , Engenharia Tecidual/métodos , Desenvolvimento Muscular , Alicerces Teciduais/química , Colágeno Tipo I/química , Células Endoteliais/citologia , Células Endoteliais/metabolismo , Colágeno/química
18.
EMBO Rep ; 25(9): 3812-3841, 2024 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-39143258

RESUMO

Satellite cells are skeletal muscle stem cells that contribute to postnatal muscle growth, and they endow skeletal muscle with the ability to regenerate after a severe injury. Here we discover that this myogenic potential of satellite cells requires a protein called tripartite motif-containing 28 (TRIM28). Interestingly, different from the role reported in a previous study based on C2C12 myoblasts, multiple lines of both in vitro and in vivo evidence reveal that the myogenic function of TRIM28 is not dependent on changes in the phosphorylation of its serine 473 residue. Moreover, the functions of TRIM28 are not mediated through the regulation of satellite cell proliferation or differentiation. Instead, our findings indicate that TRIM28 regulates the ability of satellite cells to progress through the process of fusion. Specifically, we discover that TRIM28 controls the expression of a fusogenic protein called myomixer and concomitant fusion pore formation. Collectively, the outcomes of this study expose the framework of a novel regulatory pathway that is essential for myogenesis.


Assuntos
Diferenciação Celular , Desenvolvimento Muscular , Células Satélites de Músculo Esquelético , Proteína 28 com Motivo Tripartido , Desenvolvimento Muscular/genética , Animais , Proteína 28 com Motivo Tripartido/metabolismo , Proteína 28 com Motivo Tripartido/genética , Células Satélites de Músculo Esquelético/metabolismo , Camundongos , Linhagem Celular , Fosforilação , Proliferação de Células , Fusão Celular , Músculo Esquelético/metabolismo
19.
Int J Mol Sci ; 25(16)2024 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-39201586

RESUMO

Skeletal muscle satellite cells (SMSCs), a type of myogenic stem cell, play a pivotal role in postnatal muscle regeneration and repair in animals. Circular RNAs (circRNAs) are a distinct class of non-coding RNA molecules capable of regulating muscle development by modulating gene expression, acting as microRNAs, or serving as protein decoys. In this study, we identified circ_14820, an exonic transcript derived from adenosine triphosphatase family protein 2 (ATAD2), through initial RNA-Seq analysis. Importantly, overexpression of circ_14820 markedly enhanced the proliferation of goat SMSCs while concomitantly suppressing their differentiation. Moreover, circ_14820 exhibited predominant localization in the cytoplasm of SMSCs. Subsequent small RNA and mRNA sequencing of circ_14820-overexpressing SMSCs systematically elucidated the molecular regulatory mechanisms associated with circ_14820. Our preliminary findings suggest that the circ_14820-miR-206-CCND2 regulatory axis may govern the development of goat SMSCs. These discoveries contribute to a deeper understanding of circRNA-mediated mechanisms in regulating skeletal muscle development, thereby advancing our knowledge of muscle biology.


Assuntos
Diferenciação Celular , Proliferação de Células , Cabras , RNA Circular , Células Satélites de Músculo Esquelético , Animais , Cabras/genética , Células Satélites de Músculo Esquelético/metabolismo , Células Satélites de Músculo Esquelético/citologia , RNA Circular/genética , RNA Circular/metabolismo , Diferenciação Celular/genética , Proliferação de Células/genética , MicroRNAs/genética , MicroRNAs/metabolismo , Desenvolvimento Muscular/genética , Células Cultivadas , Ciclina D2/genética , Ciclina D2/metabolismo , Músculo Esquelético/metabolismo , Músculo Esquelético/citologia
20.
Protein Sci ; 33(9): e5156, 2024 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-39180494

RESUMO

Executioner caspases, such as caspase-3, are known to induce apoptosis, but in other contexts, they can control very different fates, including cell differentiation and neuronal plasticity. While hundreds of caspase substrates are known to be specifically targeted during cell death, we know very little about how caspase activity brings about non-apoptotic fates. Here, we report the first proteome identification of cleavage events in C2C12 cells undergoing myogenic differentiation and its comparison to undifferentiated or dying C2C12 cells. These data have identified new caspase substrates, including caspase substrates specifically associated with differentiation, and show that caspases are regulating proteins involved in myogenesis in myotubes, several days after caspase-3 initiated differentiation. Cytoskeletal proteins emerged as a major group of non-apoptotic caspase substrates. We also identified proteins with well-established roles in muscle differentiation as substrates cleaved in differentiating cells.


Assuntos
Diferenciação Celular , Desenvolvimento Muscular , Animais , Camundongos , Linhagem Celular , Caspase 3/metabolismo , Caspases/metabolismo , Proteoma/metabolismo , Proteoma/análise
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