Hippo pathway downstream effectors Yap and Taz play key roles in cell proliferation and regeneration, regulating gene expression especially via Tead transcription factors. To investigate their role ...in skeletal muscle stem cells, we analyzed Taz in vivo and ex vivo in comparison with Yap. Small interfering RNA knockdown or retroviral‐mediated expression of wild‐type human or constitutively active TAZ mutants in satellite cells showed that TAZ promoted proliferation, a function shared with YAP. However, at later stages of myogenesis, TAZ also enhanced myogenic differentiation of myoblasts, whereas YAP inhibits such differentiation. Functionally, while muscle growth was mildly affected in Taz (gene Wwtr1–/–) knockout mice, there were no overt effects on regeneration. Conversely, conditional knockout of Yap in satellite cells of Pax7Cre‐ERT2/+: Yapfl°x/fl°x:Rosa26Lacz mice produced a regeneration deficit. To identify potential mechanisms, microarray analysis showed many common TAZ/YAP target genes, but TAZ also regulates some genes independently of YAP, including myogenic genes such as Pax7, Myf5, and Myod1 (ArrayExpress–E‐MTAB‐5395). Proteomic analysis revealed many novel binding partners of TAZ/YAP in myogenic cells, but TAZ also interacts with proteins distinct from YAP that are often involved in myogenesis and aspects of cytoskeleton organization (ProteomeXchange–PXD005751). Neither TAZ nor YAP bind members of the Wnt destruction complex but both regulated expression of Wnt and Wnt‐cross talking genes with known roles in myogenesis. Finally, TAZ operates through Tead4 to enhance myogenic differentiation. In summary, Taz and Yap have overlapping functions in promoting myoblast proliferation but Taz then switches to enhance myogenic differentiation. Stem Cells 2017;35:1958–1972
In skeletal muscle, Taz and Yap both enhance satellite cell derived‐myoblast proliferation, but during the later stages of myogenesis, Taz switches to influence satellite cell fate by promoting myogenic differentiation over self‐renewal.
MicroRNAs are evolutionarily conserved small RNAs that post-transcriptionally regulate gene expression and have emerged as critical regulators of skeletal muscle development. Here, we identified ...miR-148a as a novel myogenic microRNA that mediated myogenic differentiation. The expression levels of miR-148a increased during C2C12 myoblast differentiation. Overexpression of miR-148a significantly promoted myogenic differentiation of both C2C12 myoblast and primary muscle cells. Blocking the function of miR-148a with a 2′-O-methylated antisense oligonucleotide inhibitor repressed C2C12 myoblast differentiation. Using a bioinformatics approach, we identified Rho-associated coiled-coil containing protein kinase 1 (ROCK1), a known inhibitor of myogenesis, as a target of miR-148a. A dual-luciferase reporter assay was used to demonstrate that miR-148a directly targeted the 3′-UTR of ROCK1. In addition, the overexpression of miR-148a decreased the protein expression of ROCK1 in C2C12 myoblast and primary muscle cells. Furthermore, ROCK1 inhibition with specific siRNA leaded to accelerated myogenic differentiation progression, underscoring a negative regulatory function of ROCK1 in myogenesis. Therefore, our results revealed a novel mechanism in which miR-148a positively regulates myogenic differentiation via ROCK1 down-regulation.
miRNAs participate in the regulation of skeletal muscle development.
miR-148a promotes myogenic differentiation and down-regulates ROCK1 gene at the translational level.
miR-148a plays a positive role in skeletal muscle development via ROCK1 down-regulation.
Our findings reveal a novel mechanism in which miR-148a controls myogenesis through the RhoA/ROCK pathyway.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Biophysical/biochemical cues from the environment contribute to regulation of the regenerative capacity of resident skeletal muscle stem cells called satellites cells. This can be observed in vitro, ...where muscle cell behaviour is influenced by the particular culture substrates and whether culture is performed in a 2D or 3D environment, with changes including morphology, nuclear shape and cytoskeletal organization. To create a 3D skeletal muscle model we compared collagen I, Fibrin or PEG-Fibrinogen with different sources of murine and human myogenic cells. To generate tension in the 3D scaffold, biomaterials were polymerised between two flexible silicone posts to mimic tendons. This 3D culture system has multiple advantages including being simple, fast to set up and inexpensive, so providing an accessible tool to investigate myogenesis in a 3D environment. Immortalised human and murine myoblast lines, and primary murine satellite cells showed varying degrees of myogenic differentiation when cultured in these biomaterials, with C2 myoblasts in particular forming large multinucleated myotubes in collagen I or Fibrin. However, murine satellite cells retained in their niche on a muscle fibre and embedded in 3D collagen I or Fibrin gels generated aligned, multinucleated and contractile myotubes.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Myogenic progenitors (MPs) generate myocytes that fuse to form myofibers during skeletal muscle development while maintaining the progenitor pool, which is crucial for generating sufficient muscle. ...Notch signaling has been known to reserve a population of embryonic MPs during primary myogenesis by promoting cell cycle exit and suppressing premature differentiation. However, the roles of individual Notch receptors (Notch1-4) during embryonic/fetal myogenesis are still elusive. In this study, we found that Notch1 and Notch2, which exhibit the highest structural similarity among Notch receptors, maintain the MP population by distinct mechanisms: Notch1 induces cell cycle exit and Notch2 suppresses premature differentiation. Moreover, genetic and cell culture studies showed that Notch1 and Notch2 signaling in MPs are distinctively activated by interacting with Notch ligand-expressing myofibers and MP-lineage cells, respectively. These results suggest that through different activation modes, Notch1 and Notch2 distinctively and cooperatively maintain MP population during fetal myogenesis for proper muscle development.
Neuromuscular junction degeneration is a prominent aspect of sarcopenia, the age-associated loss of skeletal muscle integrity. Previously, we showed that muscle stem cells activate and contribute to ...mouse neuromuscular junction regeneration in response to denervation (Liu et al., 2015). Here, we examined gene expression profiles and neuromuscular junction integrity in aged mouse muscles, and unexpectedly found limited denervation despite a high level of degenerated neuromuscular junctions. Instead, degenerated neuromuscular junctions were associated with reduced contribution from muscle stem cells. Indeed, muscle stem cell depletion was sufficient to induce neuromuscular junction degeneration at a younger age. Conversely, prevention of muscle stem cell and derived myonuclei loss was associated with attenuation of age-related neuromuscular junction degeneration, muscle atrophy, and the promotion of aged muscle force generation. Our observations demonstrate that deficiencies in muscle stem cell fate and post-synaptic myogenesis provide a cellular basis for age-related neuromuscular junction degeneration and associated skeletal muscle decline.
Myocardial infarction (MI)-induced heart failure (HF) is commonly accompanied with profound effects on skeletal muscle. With the process of MI-induced HF, perturbations in skeletal muscle contribute ...to muscle atrophy. Exercise is viewed as a feasible strategy to prevent muscle atrophy. The aims of this study were to investigate whether exercise could alleviate MI-induced skeletal muscle atrophy via insulin-like growth factor 1 (IGF-1) pathway in mice. Male C57/BL6 mice were used to establish the MI model and were divided into three groups: sedentary MI group (MI), MI with aerobic exercise group, and MI with resistance exercise group; sham-operated group was used as control. Exercise-trained animals were subjected to 4 wk of aerobic exercise (AE) or resistance exercise (RE). Cardiac function, muscle weight, myofiber size, levels of IGF-1 signaling and proteins related to myogenesis, protein synthesis, and degradation and apoptosis in gastrocnemius muscle were detected. H
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-treated C2C12 cells were intervened with recombinant human IGF-1, IGF-1 receptor (IGF-1R) inhibitor NVP-AEW541, and PI3K inhibitor LY294002 to explore the mechanism. Exercises upregulated the IGF-1/IGF-1R-phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt) signaling; increased the expressions of Pax7, myogenic regulatory factors (MRFs), and protein synthesis; and reduced protein degradation and cell apoptosis in MI mice. In vitro, IGF-1 upregulated the levels of Pax7, MRFs, mTOR, and P70S6K; reduced MuRF1 and MAFbx; and inhibited cell apoptosis via IGF-1R-PI3K/Akt pathway. AE and RE, safely and effectively, alleviate skeletal muscle atrophy by regulating the levels of myogenesis, protein degradation, and cell apoptosis in mice with MI via activating IGF-1/IGF-1R-PI3K/Akt signaling pathway.
Ciglitazone is a member of the thiazolidinedione family, and specifically binds to peroxisome proliferator-activated receptor-γ (PPARγ), thereby promoting adipocyte differentiation. We hypothesized ...that ciglitazone as a PPARγ ligand in the absence of an adipocyte differentiation cocktail would increase adiponectin and adipogenic gene expression in bovine satellite cells (BSC). Muscle-derived BSCs were isolated from six, 18-month-old Yanbian Yellow Cattle. The BSC were cultured for 96 h in differentiation medium containing 5 µM ciglitazone (CL), 10 µM ciglitazone (CM), or 20 µM ciglitazone (CH). Control (CON) BSC were cultured only in a differentiation medium (containing 2% horse serum). The presence of myogenin, desmin, and paired box 7 (Pax7) proteins was confirmed in the BSC by immunofluorescence staining. The CL, CM, and CH treatments produced higher concentrations of triacylglycerol and lipid droplet accumulation in myotubes than those of the CON treatment. Ciglitazone treatments significantly increased the relative expression of PPARγ, CCAAT/enhancer-binding protein alpha (C/EBPα), C/EBPβ, fatty acid synthase, stearoyl-CoA desaturase, and perilipin 2. Ciglitazone treatments increased gene expression of Pax3 and Pax7 and decreased expression of myogenic differentiation-1, myogenin, myogenic regulatory factor-5, and myogenin-4 (p < 0.01). Adiponectin concentration caused by ciglitazone treatments was significantly greater than CON (p < 0.01). RNA sequencing showed that 281 differentially expressed genes (DEGs) were found in the treatments of ciglitazone. DEGs gene ontology (GO) analysis showed that the top 10 GO enrichment significantly changed the biological processes such as protein trimerization, negative regulation of cell proliferation, adipocytes differentiation, and cellular response to external stimulus. Kyoto Encyclopedia of Genes and Genomes pathway analysis showed that DEGs were involved in the p53 signaling pathway, PPAR signaling pathway, biosynthesis of amino acids, tumor necrosis factor signaling pathway, non-alcoholic fatty liver disease, PI3K-Akt signaling pathway, and Wnt signaling pathway. These results indicate that ciglitazone acts as PPARγ agonist, effectively increases the adiponectin concentration and adipogenic gene expression, and stimulates the conversion of BSC to adipocyte-like cells in the absence of adipocyte differentiation cocktail.
The development of three dimensional (3D) scaffolds for promoting and stimulating cell growth is one of the greatest concerns in biomedical and tissue engineering. In the present study, novel ...biomimetic 3D scaffolds composed of polyurethane (PU) foam and graphene oxide (GO) nanosheets were designed, and their potential as 3D scaffolds for skeletal tissue regeneration was explored. The GO-coated PU foams (GO-PU foams) were characterized by scanning electron microscopy and Raman spectroscopy. It was revealed that the 3D GO-PU foams consisted of an interconnected foam-like network structure with an approximate 300 μm pore size, and the GO was uniformly distributed in the PU foams. On the other hand, the myogenic stimulatory effects of GO on skeletal myoblasts were also investigated. Moreover, the cellular behaviors of the skeletal myoblasts within the 3D GO-PU foams were evaluated by immunofluorescence analysis. Our findings showed that GO can significantly promote spontaneous myogenic differentiation without any myogenic factors, and the 3D GO-PU foams can provide a suitable 3D microenvironment for cell growth. Furthermore, the 3D GO-PU foams stimulated spontaneous myogenic differentiation via the myogenic stimulatory effects of GO. Therefore, this study suggests that the 3D GO-PU foams are beneficial to myogenesis, and can be used as biomimetic 3D scaffolds for skeletal tissue engineering.
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BFBNIB, DOBA, GIS, IJS, IZUM, KILJ, KISLJ, NUK, PILJ, PNG, SAZU, UILJ, UKNU, UL, UM, UPUK
RNA is likely to be the most rediscovered macromolecule in biology. Periodically, new non-canonical functions have been ascribed to RNA, such as the ability to act as a catalytic molecule or to work ...independently from its coding capacity. Recent annotations show that more than half of the transcriptome encodes for RNA molecules lacking coding activity. Here we illustrate how these transcripts affect skeletal muscle differentiation and related disorders. We discuss the most recent scientific discoveries that have led to the identification of the molecular circuitries that are controlled by RNA during the differentiation process and that, when deregulated, lead to pathogenic events. These findings will provide insights that can aid in the development of new therapeutic interventions for muscle diseases.
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