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Skeletal Muscle May 2023Skeletal muscle development and regeneration depend on cellular fusion of myogenic progenitors to generate multinucleated myofibers. These progenitors utilize two...
BACKGROUND
Skeletal muscle development and regeneration depend on cellular fusion of myogenic progenitors to generate multinucleated myofibers. These progenitors utilize two muscle-specific fusogens, Myomaker and Myomerger, which function by remodeling cell membranes to fuse to each other or to existing myofibers. Myomaker and Myomerger expression is restricted to differentiating progenitor cells as they are not detected in adult myofibers. However, Myomaker remains expressed in myofibers from mice with muscular dystrophy. Ablation of Myomaker from dystrophic myofibers results in reduced membrane damage, leading to a model where persistent fusogen expression in myofibers, in contrast to myoblasts, is harmful.
METHODS
Dox-inducible transgenic mice were developed to ectopically express Myomaker or Myomerger in the myofiber compartment of skeletal muscle. We quantified indices of myofiber membrane damage, such as serum creatine kinase and IgM myofibers, and assessed general muscle histology, including central nucleation, myofiber size, and fibrosis.
RESULTS
Myomaker or Myomerger expression in myofibers independently caused membrane damage at acute time points. This damage led to muscle pathology, manifesting with centrally nucleated myofibers and muscle atrophy. Dual expression of both Myomaker and Myomerger in myofibers exacerbated several aspects of muscle pathology compared to expression of either fusogen by itself.
CONCLUSIONS
These data reveal that while myofibers can tolerate some level of Myomaker and Myomerger, expression of a single fusogen above a threshold or co-expression of both fusogens is damaging to myofibers. These results explain the paradigm that their expression in myofibers can have deleterious consequences in muscle pathologies and highlight the need for their highly restricted expression during myogenesis and fusion.
Topics: Mice; Animals; Membrane Proteins; Muscle Proteins; Muscle, Skeletal; Myoblasts; Mice, Transgenic; Muscle Development
PubMed: 37127758
DOI: 10.1186/s13395-023-00317-z -
Bioscience Reports Jan 2023Skeletal muscle possesses a high plasticity and a remarkable regenerative capacity that relies mainly on muscle stem cells (MuSCs). Molecular and cellular components of... (Review)
Review
Skeletal muscle possesses a high plasticity and a remarkable regenerative capacity that relies mainly on muscle stem cells (MuSCs). Molecular and cellular components of the MuSC niche, such as immune cells, play key roles to coordinate MuSC function and to orchestrate muscle regeneration. An abnormal infiltration of immune cells and/or imbalance of pro- and anti-inflammatory cytokines could lead to MuSC dysfunctions that could have long lasting effects on muscle function. Different genetic variants were shown to cause muscular dystrophies that intrinsically compromise MuSC function and/or disturb their microenvironment leading to impaired muscle regeneration that contributes to disease progression. Alternatively, many acquired myopathies caused by comorbidities (e.g., cardiopulmonary or kidney diseases), chronic inflammation/infection, or side effects of different drugs can also perturb MuSC function and their microenvironment. The goal of this review is to comprehensively summarize the current knowledge on acquired myopathies and their impact on MuSC function. We further describe potential therapeutic strategies to restore MuSC regenerative capacity.
Topics: Humans; Muscular Diseases; Muscle, Skeletal; Myoblasts; Muscle Development; Inflammation
PubMed: 36538023
DOI: 10.1042/BSR20220284 -
Molecular Therapy : the Journal of the... Sep 2023Rhabdomyosarcoma is the most common pediatric soft tissue tumor, comprising two major subtypes: the PAX3/7-FOXO1 fusion-negative embryonal and the PAX3/7-FOXO1...
Rhabdomyosarcoma is the most common pediatric soft tissue tumor, comprising two major subtypes: the PAX3/7-FOXO1 fusion-negative embryonal and the PAX3/7-FOXO1 fusion-positive alveolar subtype. Here, we demonstrate that the expression levels of the transcriptional repressor TRPS1 are specifically enhanced in the embryonal subtype, resulting in impaired terminal myogenic differentiation and tumor growth. During normal myogenesis, expression levels of TRPS1 have to decrease to allow myogenic progression, as demonstrated by overexpression of TRPS1 in myoblasts impairing myotube formation. Consequentially, myogenic differentiation in embryonal rhabdomyosarcoma in vitro as well as in vivo can be achieved by reducing TRPS1 levels. Furthermore, we show that TRPS1 levels in RD cells, the bona fide model cell line for embryonal rhabdomyosarcoma, are regulated by miR-1 and that TRPS1 and MYOD1 share common genomic binding sites. The myogenin (MYOG) promoter is one of the critical targets of TRPS1 and MYOD1; we demonstrate that TRPS1 restricts MYOG expression and thereby inhibits terminal myogenic differentiation. Therefore, reduction of TRPS1 levels in embryonal rhabdomyosarcoma might be a therapeutic approach to drive embryonal rhabdomyosarcoma cells into myogenic differentiation, thereby generating postmitotic myotubes.
Topics: Humans; Child; Rhabdomyosarcoma, Embryonal; Myogenin; Cell Differentiation; MicroRNAs; Muscle Development; Cell Line, Tumor; Repressor Proteins
PubMed: 37452493
DOI: 10.1016/j.ymthe.2023.07.003 -
Skeletal Muscle Feb 2023Acute injury to skeletal muscle damages myofibers and fragment capillaries, impairing contractile function and local perfusion. Myofibers and microvessels regenerate...
BACKGROUND
Acute injury to skeletal muscle damages myofibers and fragment capillaries, impairing contractile function and local perfusion. Myofibers and microvessels regenerate from satellite cells and from surviving microvessel fragments, respectively, to restore intact muscle. Established models of injury have used myotoxins and physical trauma to demonstrate the concurrence of myogenesis and angiogenesis during regeneration. In these models, efferocytosis removes cellular debris while basal laminae persist to provide guidance during myofiber and microvessel regeneration. It is unknown whether the spatiotemporal coupling between myofiber and microvascular regeneration persists when muscle tissue is completely removed and local guidance cues are lost.
METHODS
To test whether complete removal of skeletal muscle tissue affects the spatiotemporal relationship between myogenesis and angiogenesis during regeneration, subthreshold volumetric muscle loss was created with a biopsy punch (diameter, 2 mm) through the center of the gluteus maximus (GM) in adult mice. Regeneration into the void was evaluated through 21 days post-injury (dpi). Microvascular perfusion was evaluated in vivo by injecting fluorescent dextran into the circulation during intravital imaging. Confocal imaging and histological analyses of whole-mount GM preparations and tissue cross-sections assessed the growth of microvessels and myofibers into the wound.
RESULTS
A provisional matrix filled with PDGFRα and CD45 cells spanned the wound within 1 dpi. Regenerating microvessels advanced from the edges of the wound into the matrix by 7 dpi. Nascent microvascular networks formed by 10 dpi with blood-perfused networks spanning the wound by 14 dpi. In striking contrast, the wound remained devoid of myofibers at 7 and 10 dpi. Myogenesis into the wound was apparent by 14 dpi and traversed the wound by 21 dpi. Regenerated myofibers and microvessels were disorganized compared to the uninjured muscle.
CONCLUSIONS
Following punch biopsy of adult skeletal muscle, regenerating microvessels span the wound and become perfused with blood prior to myofiber regeneration. The loss of residual guidance cues with complete tissue removal disrupts the spatiotemporal correspondence between microvascular and myofiber regeneration. We conclude that angiogenesis precedes myogenesis during regeneration following subthreshold volumetric muscle loss.
Topics: Animals; Mice; Muscle, Skeletal; Regeneration; Microvessels; Biopsy; Muscle Development
PubMed: 36788624
DOI: 10.1186/s13395-023-00313-3 -
International Journal of Molecular... Jun 2021A decline in the body's motor functions has been linked to decreased muscle mass and function in the oral cavity and throat; however, aging of the junctions of the... (Review)
Review
A decline in the body's motor functions has been linked to decreased muscle mass and function in the oral cavity and throat; however, aging of the junctions of the muscles and bones has also been identified as an associated factor. Basic and clinical studies on the muscles, tendons and bones, each considered independently, have been published. In recent years, however, research has focused on muscle attachment as the muscle-tendon-bone complex from various perspectives, and there is a growing body of knowledge on SRY-box9 (Sox9) and Mohawk(Mkx), which has been identified as a common controlling factor and a key element. Myostatin, a factor that inhibits muscle growth, has been identified as a potential key element in the mechanisms of lifetime structural maintenance of the muscle-tendon-bone complex. Findings in recent studies have also uncovered aspects of the mechanisms of motor organ complex morphostasis in the superaged society of today and will lay the groundwork for treatments to prevent motor function decline in older adults.
Topics: Animals; Biomedical Research; Bone Development; Humans; Morphogenesis; Muscle Development; Muscle Proteins; Tendons
PubMed: 34198655
DOI: 10.3390/ijms22126365 -
Development (Cambridge, England) Jul 2023The earliest skeletal muscle progenitor cells (SMPCs) derived from human pluripotent stem cells (hPSCs) are often identified by factors expressed by a diverse number of...
The earliest skeletal muscle progenitor cells (SMPCs) derived from human pluripotent stem cells (hPSCs) are often identified by factors expressed by a diverse number of progenitors. An early transcriptional checkpoint that defines myogenic commitment could improve hPSC differentiation to skeletal muscle. Analysis of several myogenic factors in human embryos and early hPSC differentiations found SIX1+PAX3+ co-expression was most indictive of myogenesis. Using dCas9-KRAB hPSCs, we demonstrate that early inhibition of SIX1 alone significantly decreased PAX3 expression, reduced PAX7+ SMPCs, and myotubes later in differentiation. Emergence of SIX1+PAX3+ precursors can be improved by manipulating seeding density, monitoring metabolic secretion and altering the concentration of CHIR99021. These modifications resulted in the co-emergence of hPSC-derived sclerotome, cardiac and neural crest that we hypothesized enhanced hPSC myogenic differentiation. Inhibition of non-myogenic lineages modulated PAX3 independent of SIX1. To better understand SIX1 expression, we compared directed differentiations to fetal progenitors and adult satellite cells by RNA-seq. Although SIX1 continued to be expressed across human development, SIX1 co-factor expression was dependent on developmental timing. We provide a resource to enable efficient derivation of skeletal muscle from hPSCs.
Topics: Adult; Humans; PAX3 Transcription Factor; Pluripotent Stem Cells; Cell Differentiation; Muscle, Skeletal; Muscle Development; PAX7 Transcription Factor; Homeodomain Proteins
PubMed: 37366057
DOI: 10.1242/dev.201509 -
American Journal of Physiology. Cell... Oct 2019The αβ-integrin is a transmembrane adhesion protein that connects laminin in the extracellular matrix (ECM) with actin in skeletal muscle fibers. The αβ-integrin is... (Review)
Review
The αβ-integrin is a transmembrane adhesion protein that connects laminin in the extracellular matrix (ECM) with actin in skeletal muscle fibers. The αβ-integrin is highly expressed in skeletal muscle and is concentrated at costameres and myotendious junctions, providing the opportunity to transmit longitudinal and lateral forces across the membrane. Studies have demonstrated that α-integrin subunit mRNA and protein are upregulated following eccentric contractions as a mechanism to reinforce load-bearing structures and resist injury with repeated bouts of exercise. It has been hypothesized for many years that the integrin can also promote protein turnover in a manner that can promote beneficial adaptations with resistance exercise training, including hypertrophy. This review provides basic information about integrin structure and activation and then explores its potential to serve as a critical mechanosensor and activator of muscle protein synthesis and growth. Overall, the hypothesis is proposed that the αβ-integrin can contribute to mechanical-load induced skeletal muscle growth via an mammalian target of rapamycin complex 1-independent mechanism.
Topics: Animals; Extracellular Matrix; Humans; Integrins; Muscle Development; Muscle Fibers, Skeletal; Muscle, Skeletal; Signal Transduction
PubMed: 31314586
DOI: 10.1152/ajpcell.00009.2019 -
Journal of Cachexia, Sarcopenia and... Feb 2022Circular RNAs (circRNAs) represent a novel class of non-coding RNAs formed by a covalently closed loop and play crucial roles in many biological processes. Several...
BACKGROUND
Circular RNAs (circRNAs) represent a novel class of non-coding RNAs formed by a covalently closed loop and play crucial roles in many biological processes. Several circRNAs associated with myogenesis have been reported. However, the dynamic expression, function, and mechanism of circRNAs during myogenesis and skeletal muscle development are largely unknown.
METHODS
Strand-specific RNA-sequencing (RNA-seq) and microarray datasets were used to profile the dynamic circRNAome landscape during skeletal muscle development and myogenic differentiation. Bioinformatics analyses were used to characterize the circRNAome and identify candidate circRNAs associated with myogenesis. Bulk and single-cell RNA-seq were performed to identify the downstream genes and pathways of circFgfr2. The primary myoblast cells, C2C12 cells, and animal model were used to assess the function and mechanism of circFgfr2 in myogenesis and muscle regeneration in vitro or in vivo by RT-qPCR, western blotting, dual-luciferase activity assay, RNA immunoprecipitation, RNA fluorescence in situ hybridization, and chromatin immunoprecipitation.
RESULTS
We profiled the dynamic circRNAome in pig skeletal muscle across 27 developmental stages and detected 52 918 high-confidence circRNAs. A total of 2916 of these circRNAs are conserved across human, mouse, and pig, including four circRNAs (circFgfr2, circQrich1, circMettl9, and circCamta1) that were differentially expressed (|log fold change| > 1 and adjusted P value < 0.05) in various myogenesis systems. We further focused on a conserved circRNA produced from the fibroblast growth factor receptor 2 (Fgfr2) gene, termed circFgfr2, which was found to inhibit myoblast proliferation and promote differentiation and skeletal muscle regeneration. Mechanistically, circFgfr2 acted as a sponge for miR-133 to regulate the mitogen-activated protein kinase kinase kinase 20 (Map3k20) gene and JNK/MAPK pathway. Importantly, transcription factor Kruppel like factor 4 (Klf4), the downstream target of the JNK/MAPK pathway, directly bound to the promoter of circFgfr2 and affected its expression via an miR-133/Map3k20/JNK/Klf4 auto-regulatory feedback loop. RNA binding protein G3BP stress granule assembly factor 1 (G3bp1) inhibited the biogenesis of circFgfr2.
CONCLUSIONS
The present study provides a comprehensive circRNA resource for skeletal muscle study. The functional and mechanistic analysis of circFgfr2 uncovered a circRNA-mediated auto-regulatory feedback loop regulating myogenesis and muscle regeneration, which provides new insight to further understand the regulatory mechanism of circRNAs.
Topics: Animals; DNA Helicases; Feedback; In Situ Hybridization, Fluorescence; Mice; Muscle Development; Muscle, Skeletal; Poly-ADP-Ribose Binding Proteins; RNA Helicases; RNA Recognition Motif Proteins; Regeneration; Swine
PubMed: 34811940
DOI: 10.1002/jcsm.12859 -
Cells Aug 2019Skeletal muscle myogenesis and injury-induced muscle regeneration contribute to muscle formation and maintenance. As myogenic stem cells, skeletal muscle satellite... (Review)
Review
Skeletal muscle myogenesis and injury-induced muscle regeneration contribute to muscle formation and maintenance. As myogenic stem cells, skeletal muscle satellite cells have the ability to proliferate, differentiate and self-renew, and are involved in muscle formation and muscle injury repair. Accumulating evidence suggests that non-coding RNAs (ncRNAs), including microRNAs (miRNAs), long non-coding RNAs (lncRNAs) and circular RNAs (circRNAs), are widely involved in the regulation of gene expression during skeletal muscle myogenesis, and their abnormal expression is associated with a variety of muscle diseases. From the perspective of the molecular mechanism and mode of action of ncRNAs in myogenesis, this review aims to summarize the role of ncRNAs in skeletal muscle satellite cells' myogenic differentiation and in muscle disease, and systematically analyze the mechanism of ncRNAs in skeletal muscle development. This work will systematically summarize the role of ncRNAs in myogenesis and provide reference targets for the treatment of various muscle diseases, such as muscle dystrophy, atrophy and aberrant hypertrophy.
Topics: Animals; Humans; Muscle Development; Muscular Diseases; RNA, Untranslated; Satellite Cells, Skeletal Muscle
PubMed: 31461973
DOI: 10.3390/cells8090988 -
Journal of Cachexia, Sarcopenia and... Apr 2022Most patients with pancreatic cancer develop cachexia, which is characterized by progressive muscle loss. The mechanisms underlying muscle loss in cancer cachexia remain...
BACKGROUND
Most patients with pancreatic cancer develop cachexia, which is characterized by progressive muscle loss. The mechanisms underlying muscle loss in cancer cachexia remain elusive. Pancreatic tumour organoids are 3D cell culture models that retain key characteristics of the parent tumour. We aimed to investigate the effect of pancreatic tumour organoid-derived factors on processes that determine skeletal muscle mass, including the regulation of muscle protein turnover and myogenesis.
METHODS
Conditioned medium (CM) was collected from human pancreatic cancer cell lines (PK-45H, PANC-1, PK-1, and KLM-1), pancreatic tumour organoid cultures from a severely cachectic (PANCO-9a) and a non-cachectic patient (PANCO-12a), and a normal pancreas organoid culture. Differentiating C2C12 myoblasts and mature C2C12 myotubes were exposed to CM for 24 h or maintained in control medium. In myotubes, NF-kB activation was monitored using a NF-κB luciferase reporter construct, and mRNA expression of E3-ubiquitin ligases and REDD1 was analysed by RT-qPCR. C2C12 myoblast proliferation and differentiation were monitored by live cell imaging and myogenic markers and myosin heavy chain (MyHC) isoforms were assessed by RT-qPCR.
RESULTS
Whereas CM from PK-1 and KLM-1 cells significantly induced NF-κB activation in C2C12 myotubes (PK-1: 3.1-fold, P < 0.001; KLM-1: 2.1-fold, P = 0.01), Atrogin-1/MAFbx and MuRF1 mRNA were only minimally and inconsistently upregulated by the CM of pancreatic cancer cell lines. Similarly, E3-ubiquitin ligases and REDD1 mRNA expression in myotubes were not altered by exposure to pancreatic tumour organoid CM. Compared with the control condition, CM from both PANCO-9a and PANCO-12a tumour organoids increased proliferation of myoblasts, which was accompanied by significant downregulation of the satellite cell marker paired-box 7 (PAX7) (PANCO-9a: -2.1-fold, P < 0.001; PANCO-12a: -2.0-fold, P < 0.001) and myogenic factor 5 (MYF5) (PANCO-9a: -2.1-fold, P < 0.001; PANCO-12a: -1.8-fold, P < 0.001) after 48 h of differentiation. Live cell imaging revealed accelerated alignment and fusion of myoblasts exposed to CM from PANCO-9a and PANCO-12a, which was in line with significantly increased Myomaker mRNA expression levels (PANCO-9a: 2.4-fold, P = 0.001; PANCO-12a: 2.2-fold, P = 0.004). These morphological and transcriptional alterations were accompanied by increased expression of muscle differentiation markers such as MyHC-IIB (PANCO-9a: 2.5-fold, P = 0.04; PANCO-12a: 3.1-fold, P = 0.006). Although the impact of organoid CM on myogenesis was not associated with the cachexia phenotype of the donor patients, it was specific for tumour organoids, as CM of control pancreas organoids did not modulate myogenic fusion.
CONCLUSIONS
These data show that pancreatic tumour organoid-derived factors alter the kinetics of myogenesis, which may eventually contribute to impaired muscle mass maintenance in cancer cachexia.
Topics: Cachexia; Humans; Muscle Development; Myoblasts; Organoids; Pancreatic Neoplasms
PubMed: 35146962
DOI: 10.1002/jcsm.12917