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Hypoxia promotes proliferation and inhibits myogenesis in broiler satellite cells

ABSTRACT: Breast muscle myopathies in broilers compromise meat quality and continue to plague the poultry industry. Broiler breast muscle myopathies are characterized by impaired satellite cells (SC)-mediated repair, and localized tissue hypoxia and dysregulation of oxygen homeostasis have been implicated as contributing factors. The present study was designed to test the hypothesis that hypoxia disrupts the behavior of SC essential for multinucleated myotube formation in vitro, and to determine the extent to which effects are reversed by restoration of oxygen tension. Primary SC was isolated from pectoralis major of young (5 d) Cobb 700 chicks and maintained in growth conditions or induced to differentiate under normoxic (20% O2) or hypoxic (1% O2) conditions for up to 48 h. Hypoxia enhanced SC proliferation while inhibiting myogenic potential, with decreased fusion index and suppressed myotube formation. Reoxygenation after hypoxia partially reversed effects on both proliferation and myogenesis. Western blotting showed that hypoxia diminished myogenin expression, activated AMPK, upregulated proliferation markers, and increased molecular signaling of cellular stress. Hypoxia also promoted accumulation of lipid droplets in myotubes. Targeted RNAseq identified numerous differentially expressed genes across differentiation under hypoxia, including several genes that have been associated with myopathies in vivo. Altogether, these data demonstrate localized hypoxia may influence SC behavior in ways that disrupt muscle repair and promote the formation of myopathies in broilers.

ORGANISM(S): Gallus gallus

PROVIDER: GSE241619 | GEO | 2023/10/24

REPOSITORIES: GEO

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Project description:Introduction: Skeletal muscle satellite cells (MuSCs, or stem cells) play a crucial role in muscle development, maintenance, and regeneration, serving both hypertrophic support and regenerative myogenesis. Syndecans (SDCs) act as communication bridges within the muscle microenvironment, regulating interactions with extracellular matrix components, and contributing significantly to tissue repair and inflammation. Specifically, syndecan-4 (SDC4) is involved in muscle regeneration at multiple stages. Methods: This study delves into the emerging challenge of wooden breast (WB) myopathy and its connection to SDC4. Our hypothesis proposes that disruptions in MuSC dynamics through SDC4 contribute to the increased incidence of breast myopathies observed in growing broilers. To test our hypothesis, non-affected and affected broilers were systematically selected, and the characteristics of WB myopathy were studied both in vitro and in vivo. SDC4 overexpression in MuSCs and blocking peptides corresponding to the SDC4 ectodomain were used for investigation of the role of SDC4 in muscle development and its shedding levels. Results and Discussion: Examination in vivo of affected muscles revealed smaller fibers and changes in metabolic pathways. In vitro studies unveiled disrupted proliferation of MuSCs in WB myopathy, accompanied by downregulation of several muscle markers. Investigation of a potentially role of SDC4 in the pathogenesis of WB myopathy revealed a decreased tendency in SDC4 gene expression and increased shedding of its ectodomain. Moreover, we showed that SDC4 overexpression is linked to decreased proliferation in MuSCs and affected myogenesis. We detected the impairment proliferation of WB-affected MuSCs, revealing critical insights into the dysfunctional state of these cells in myopathy. Additionally, by treating MuSCs with blocking peptides derived from the SDC4 ectodomain, we identified altered proliferation. Taken together, this work contributes valuable knowledge on the molecular mechanisms underlying WB myopathy and role of SDC4 in this chicken myopathy.

Project description:Background Quiescent (non-dividing) muscle satellite cells (SCs) transition into proliferating progenitor cells that differentiate to repair skeletal muscle after injury. Recently, the AP-1 family member, FOS, was found to be rapidly and transiently induced in SCs in response to muscle damage, and its activity is required for effective SC activation and muscle repair. However, why FOS is rapidly down-regulated before SCs enter the cell cycle as progenitor cells remains unclear. In addition, whether boosting FOS expression in primary cycling muscle progenitor cells can enhance their myogenic properties needs to be evaluated. Methods In this study, we established a lentiviral, doxycycline-inducible system to explore the molecular and functional consequences of continued FOS expression in SC-derived, cycling muscle progenitor cells ex vivo. Specifically, we performed cell growth and EdU incorporation assays to measure cellular proliferation; myotube formation assays to evaluate terminal differentiation potential; and RNA-Sequencing (RNA-Seq) in conjunction with high-throughput chromosome conformation capture (Hi-C) to uncover changes in gene expression and long-range chromatin interactions. Results Persistent FOS activity in cycling muscle progenitor cells modestly enhances progenitor cell proliferation, but severely antagonizes their ability to differentiate and form myotubes. Genome-wide RNA-Seq analysis revealed that ectopic FOS activity in cycling muscle progenitor cells suppresses a global pro-myogenic gene expression program, while activating a stress-induced Mitogen-Activated Protein Kinase (MAPK) transcriptional signature. Importantly, we found various chromosomal re-organization events in A/B compartments, TADs, and long-range genomic loops near FOS-regulated differentially expressed genes in cycling muscle progenitor cells ectopically expressing FOS. Conclusions Altogether, our results suggest that elevated FOS activity in proliferating muscle progenitor cells perturbs cellular differentiation by altering the 3-dimensional (3D) chromosome organization near critical pro-myogenic genes. This work highlights the crucial importance of tightly controlling FOS expression in the muscle lineage, and raises the possibility that in states of chronic stress or disease, persistent FOS activity in muscle stem and/or progenitor cells may actually disrupt the muscle forming process.

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Hypoxia promotes proliferation and inhibits myogenesis in broiler satellite cells

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