bims-moremu Biomed News
on Molecular regulators of muscle mass
Issue of 2022‒06‒12
35 papers selected by
Anna Vainshtein
Craft Science Inc.

  1. FEBS J. 2022 Jun 11.
      Muscle regeneration is essential for vertebrate muscle homeostasis and recovery after injury. During regeneration, muscle stem cells differentiate into myocytes, which then fuse with pre-existing muscle fibers. Hence, differentiation, fusion and contraction must be tightly regulated during regeneration to avoid the disastrous consequences of premature fusion of myocytes to actively contracting fibers. Cytosolic calcium (Ca2+ ), which is coupled to both induction of myogenic differentiation and contraction, has more recently been implicated in the regulation of myocyte-to-myotube fusion. In this Viewpoint, we propose that Ca2+ -mediated coordination of differentiation, fusion, and contraction is a feature selected in the amniotes to facilitate muscle regeneration.
    Keywords:  Calcium signaling; Calcium-responsive genes (CRGs); muscle architecture; muscle contraction; muscle regeneration; myoblast fusion
  2. Am J Physiol Cell Physiol. 2022 Jun 08.
      The development of skeletal muscle (myogenesis) is a well-orchestrated process where myoblasts withdraw from the cell cycle and differentiate into myotubes. Signaling by fluxes in intracellular Ca2+ is known to contribute to myogenesis and increased mitochondria biogenesis is required to meet the metabolic demand of mature myotubes. However, gaps remain in the understanding of how intracellular Ca2+ signals can govern myogenesis. Polycystin-2 (PC2 or TRPP1) is a non-selective cation channel permeable to Ca2+. It can interact with intracellular calcium channels to control Ca2+ release, and concurrently modulates mitochondrial function and remodeling. Due to these features, we hypothesized that PC2 is a central protein in mediating both the intracellular Ca2+ responses and mitochondrial changes seen in myogenesis. To test this hypothesis, we created CRISPR/Cas9 knockout C2C12 murine myoblast cell lines. PC2 KO cells were unable to differentiate into myotubes, had impaired spontaneous Ca2+ oscillations and did not develop depolarization-evoked Ca2+ transients. The autophagic-associated pathway beclin-1 was downregulated in PC2 KO cells, and direct activation of the autophagic pathway resulted in decreased mitochondrial remodeling. Re-expression of full length PC2, but not a calcium channel dead pathologic mutant, restored the differentiation phenotype and increased the expression of mitochondrial proteins. Our results establish that PC2 is a novel regulator of in-vitro myogenesis by integrating PC2 dependent Ca2+ signals and metabolic pathways.
    Keywords:  Myogenesis; PC2; TRP channels; calcium signaling; muscle differentiation
  3. Front Cell Dev Biol. 2022 ;10 917771
      Adult skeletal muscle is mainly composed of post-mitotic, multinucleated muscle fibers. Upon injury, it has the unique ability to regenerate thanks to the activation of a subset of quiescent muscle stem cells (MuSCs). Activated MuSCs either differentiate to repair muscle, or self-renew to maintain the pool of MuSC. MuSC fate determination is regulated by an intricate network of intrinsic and extrinsic factors that control the expression of specific subsets of genes. Among these, the myogenic regulatory factors (MRFs) are key for muscle development, cell identity and regeneration. More globally, cell fate determination involves important changes in the epigenetic landscape of the genome. Such epigenetic changes, which include DNA methylation and post-translational modifications of histone proteins, are able to alter chromatin organization by controlling the accessibility of specific gene loci for the transcriptional machinery. Among the numerous epigenetic modifications of chromatin, extensive studies have pointed out the key role of histone methylation in cell fate control. Particularly, since the discovery of the first histone demethylase in 2004, the role of histone demethylation in the regulation of skeletal muscle differentiation and muscle stem cell fate has emerged to be essential. In this review, we highlight the current knowledge regarding the role of histone demethylases in the regulation of muscle stem cell fate choice.
    Keywords:  cell fate and differentiation; epigenetics; histone demethylase; metabolism; muscle stem cell (MuSC)
  4. J Physiol. 2022 Jun 06.
      KEY POINTS: Exercise mediates H3K27me3 at transcriptionally upregulated loci in skeletal muscle, although the role of H3K27me3 in the adaptation of skeletal muscle to exercise training is unclear. Chromatin immunoprecipitation followed by sequencing analysis demonstrated that H3K27me3, as well as H3K4me3 modifications, is the hallmark of sites showing higher responses to acute exercise. GSK343, a selective inhibitor of the enhancer of zeste homologue 2 (EZH2), enhanced the gene responses to a single bout of exercise and accelerated the adaptive changes during exercise training in association with myonuclear H3K27me3 accumulation. Administration of valemetostat, an EZH1/2 dual inhibitor, repressed myonuclear H3K27me3 accumulation during training and caused the failure in adaptive changes. Exercise-induced H3K27me3 may play a key role in inducing exercise-related effects in the skeletal muscle.ABSTRACT: Histone H3 trimethylation at lysine 27 (H3K27me3) is known to act as a transcriptionally repressive histone modification via heterochromatin formation. However, in skeletal muscle, it was also reported that H3K27me3 was enriched at the sites transcriptionally activated by exercise, although the role of H3K27me3 in the adaptation to exercise is unknown. In this study, we first determined the genome-wide enrichment of RNA polymerase II and histone H3 trimethylation at lysine 4 (H3K4me3) and H3K27me3 using chromatin immunoprecipitation followed by sequencing analysis in mouse tibialis anterior muscle. The loci that were transcriptionally upregulated by a single bout of running exercise were marked by both H3K27me3 and H3K4me3, which also correlated with the distribution of RNA polymerase II. The genes that were not responsive to exercise exhibited high H3K4me3 occupancy, similar to the upregulated genes but with fewer H3K27me3. Next, we tested the effects of H3K27 methyltransferase, an enhancer of zeste homologue (EZH) 2-specific inhibitor GSK343. GSK343 administration unexpectedly enhanced the H3K27me3 occupancy at the target loci, leading to the upregulation of gene responses to acute exercise. GSK343 administration also facilitated the phenotypic transformation from IIb to IIa fibres and the upregulation of AMPK phosphorylation and HSP70, PDK4, PGC-1α, and MuRF1 levels. Furthermore, in contrast to the accelerated adaptation to exercise by GSK343, EZH1/2 dual inhibitor valemetostat administration caused the failure in the changes of the aforementioned parameters after exercise training. These results indicate that exercise-induced H3K27me3 plays a key role in inducing exercise-related effects in the skeletal muscle. Abstract figure legend The loci upregulated in response to exercise are characterized by a bivalent modification with histone H3 trimethylation at lysine 27 (H3K27me3) and lysine 4 (H3K4me3) in mouse skeletal muscle. Acute exercise further stimulates both H3K27me3 and H3K4me3 at these loci associated with the upregulation of gene transcription. Lysine methyltransferase EZH2-specific inhibitor GSK343 administration increased H3K27me3 and H3K4me3 occupancies at the target loci after a single bout of exercise. Chronic treatment of GSK343 during exercise training more upregulated H3K27me3 in muscle fibres. In addition, it increased the number of muscle fibres expressing type IIa myosin heavy chain (MyHC) and enhanced the adaptive changes in the related protein levels. In contrast, administration of valemetostat, an EZH1/2 dual inhibitor, decreased H3K27me3 and H3K4me3 occupancies after acute exercise and caused the failure in the exercise-induced effects after training. It was also suggested that EZH1 acted as a modifier of exercise-induced H3K27me3 in skeletal muscle. This article is protected by copyright. All rights reserved.
    Keywords:  H3K27me3; exercise epigenetics; histone modification; skeletal muscle
  5. Proc Natl Acad Sci U S A. 2022 Jun 14. 119(24): e2103615119
      Skeletal muscle atrophy is commonly associated with aging, immobilization, muscle unloading, and congenital myopathies. Generation of mature muscle cells from skeletal muscle satellite cells (SCs) is pivotal in repairing muscle tissue. Exercise therapy promotes muscle hypertrophy and strength. Primary cilium is implicated as the mechanical sensor in some mammalian cells, but its role in skeletal muscle cells remains vague. To determine mechanical sensors for exercise-induced muscle hypertrophy, we established three SC-specific cilium dysfunctional mouse models-Myogenic factor 5 (Myf5)-Arf-like Protein 3 (Arl3)-/-, Paired box protein Pax-7 (Pax7)-Intraflagellar transport protein 88 homolog (Ift88)-/-, and Pax7-Arl3-/--by specifically deleting a ciliary protein ARL3 in MYF5-expressing SCs, or IFT88 in PAX7-expressing SCs, or ARL3 in PAX7-expressing SCs, respectively. We show that the Myf5-Arl3-/- mice develop grossly the same as WT mice. Intriguingly, mechanical stimulation-induced muscle hypertrophy or myoblast differentiation is abrogated in Myf5-Arl3-/- and Pax7-Arl3-/- mice or primary isolated Myf5-Arl3-/- and Pax7-Ift88-/- myoblasts, likely due to defective cilia-mediated Hedgehog (Hh) signaling. Collectively, we demonstrate SC cilia serve as mechanical sensors and promote exercise-induced muscle hypertrophy via Hh signaling pathway.
    Keywords:  exercise; mechanical stimulation; muscle hypertrophy; primary cilia
  6. Peptides. 2022 Jun 03. pii: S0196-9781(22)00089-4. [Epub ahead of print]154 170823
      Skeletal muscle is the major effector organ for exercise. It has been proposed that VEGFB is significantly related to apoptosis in various cell types but not yet in skeletal muscle. We hypothesize that the decrease of VEGFB in skeletal muscle participates in the occurrence of skeletal muscle apoptosis and that exercise inhibits apoptosis by elevating the expression of VEGFB in skeletal muscle cells. Based on this hypothesis, we developed in vitro experiments to mimic the effect of exercise through electrical pulse stimulation (EPS) to observe the effect of EPS on apoptosis and the change in VEGFB expression in differentiated myotubes. In addition, we employed RNA interference to explore whether VEGFB is directly involved in the regulation of myotube apoptosis during EPS. Our results showed that exogenous VEGFB167 significantly inhibited C2C12 myotube apoptosis induced by TNF-α treatment and that endogenous VEGFB in differentiated C2C12 myotubes was significantly upregulated by EPS. In addition, EPS significantly changed the expression of the apoptotic indicators Bax and Bcl-2 at the mRNA level and downregulated the protein expression of cleaved caspase-3. The antiapoptotic effect of EPS weakened substantially as VEGFB in C2C12 myotubes was inhibited. Taken together, these results indicate that exercise-like EPS inhibits apoptosis by increasing the expression of C2C12 myotube-derived VEGFB.
    Keywords:  Apoptosis; C2C12 myotubes; EPS; VEGFB
  7. FASEB Bioadv. 2022 Jun;4(6): 402-407
      Physical activity or regular exercise provides many beneficial effects towards human health, helping prevent and ameliorate metabolic diseases. However, certain molecular mechanisms that mediate these health benefits remain poorly understood. Parker et al. provided the first global analysis of exercise-regulated ubiquitin signalling in human skeletal muscle, revealing post-translational modification cross-talk. As a result of their analysis, NEDDylation is thought to promote ubiquitin signalling for the removal of damaged proteins following exercise. The proteomic dataset generated from their study is invaluable for researchers in this field to validate new mechanistic hypotheses. To further reveal molecular mechanisms regulated by exercise, future research could employ more sensitive mass spectrometry-based workflows that increase the detection of both ubiquitylated sites and peptides and subsequently identify more exercise-regulated ubiquitin signalling pathways.
    Keywords:  NEDDylation; mass spectrometry; physical activity; ubiquitylation; ubiquitylome
  8. Cytoskeleton (Hoboken). 2022 Jun 05.
      Skeletal muscle differentiation occurs as muscle precursor cells (myoblasts) elongate and fuse to form multinucleated syncytial myotubes in which the highly-organized actomyosin sarcomeres of muscle fibers assemble. Although less well characterized, the microtubule cytoskeleton also undergoes dramatic rearrangement during myogenesis. The centrosome-nucleated microtubule array found in myoblasts is lost as the nuclear membrane acquires microtubule nucleating activity and microtubules emerge from multiple sites in the cell, eventually rearranging into a grid-like pattern in myotubes. In order to characterize perinuclear microtubule organization using a biochemically tractable system, we isolated nuclei from mouse C2C12 skeletal muscle cells during the course of differentiation and incubated them in cytoplasmic extracts prepared from eggs of the frog Xenopus laevis. Whereas centrosomes associated with myoblast nuclei gave rise to radial microtubule arrays in extracts, myotube nuclei produced a sun-like pattern with microtubules transiently nucleating from the entire nuclear envelope. Perinuclear microtubule growth was suppressed by inhibition of Aurora A kinase or by degradation of RNA, treatments that also inhibited microtubule growth from sperm centrosomes. Myotube nuclei displayed microtubule motor-based movements leading to their separation, as occurs in myotubes. This in vitro assay therefore recapitulates key features of microtubule organization and nuclear movement observed during muscle cell differentiation. This article is protected by copyright. All rights reserved.
    Keywords:  Microtubules; Xenopus laevis; centrosome; microtubule organizing center; myogenesis
  9. J Cachexia Sarcopenia Muscle. 2022 Jun 05.
      Sarcopenia is a progressive loss of muscle mass and strength with a risk of adverse outcomes such as disability, poor quality of life, and death. Increasing evidence indicates that diminished ability of the muscle to activate satellite cell-dependent regeneration is one of the factors that might contribute to its development. Skeletal muscle regeneration following myogenic cell death results from the proliferation and differentiation of myogenic stem cells, called satellite cells, located beneath the basal lamina of the muscle fibres. Satellite cell differentiation is not a satellite cell-autonomous process but depends on signals provided by the surrounding cells. Infiltrating macrophages play a key role in the process partly by clearing the necrotic cell debris, partly by producing cytokines and growth factors that guide myogenesis. At the beginning of the muscle regeneration process, macrophages are pro-inflammatory, and the cytokines produced by them trigger the proliferation and differentiation of satellite cells. Following the uptake of dead cells, however, a transcriptionally regulated phenotypic change (macrophage polarization) is induced in them resulting in their transformation into healing macrophages that guide resolution of inflammation, completion of myoblast differentiation, myoblast fusion and growth, and return to homeostasis. Impaired efferocytosis results in delayed cell death clearance, delayed macrophage polarization, prolonged inflammation, and impaired muscle regeneration. Thus, proper efferocytosis by macrophages is a determining factor during muscle repair. Here we review that both efferocytosis and myogenesis are dependent on the cell surface phosphatidylserine (PS), and surprisingly, these two processes share a number of common PS receptors and signalling pathways. Based on these findings, we propose that stimulating the function of PS receptors for facilitating muscle repair following injury could be a successful approach, as it would enhance efferocytosis and myogenesis simultaneously. Because increasing evidence indicates a pathophysiological role of impaired efferocytosis in the development of chronic inflammatory conditions, as well as in impaired muscle regeneration both contributing to the development of sarcopenia, improving efferocytosis should be considered also in its management. Again applying or combining those treatments that target PS receptors would be expected to be the most effective, because they would also promote myogenesis. A potential PS receptor-triggering candidate molecule is milk fat globule-EGF-factor 8 (MFG-E8), which not only stimulates PS-dependent efferocytosis and myoblast fusion but also promotes extracellular signal-regulated kinase (ERK) and Akt activation-mediated cell proliferation and cell cycle progression in myoblasts.
    Keywords:  Efferocytosis; MFG-E8; Muscle regeneration; Myoblast fusion; Myogenesis; Phosphatidylserine; Phosphatidylserine receptor; Sarcopenia
  10. Bioact Mater. 2023 Feb;20 166-178
      Skeletal muscle disorders have posed great threats to health. Selective delivery of drugs and oligonucleotides to skeletal muscle is challenging. Aptamers can improve targeting efficacy. In this study, for the first time, the human skeletal muscle-specific ssDNA aptamers (HSM01, etc.) were selected and identified with Systematic Evolution of Ligands by Exponential Enrichment (SELEX). The HSM01 ssDNA aptamer preferentially interacted with human skeletal muscle cells in vitro. The in vivo study using tree shrews showed that the HSM01 ssDNA aptamer specifically targeted human skeletal muscle cells. Furthermore, the ability of HSM01 ssDNA aptamer to target skeletal muscle cells was not affected by the formation of a disulfide bond with nanoliposomes in vitro or in vivo, suggesting a potential new approach for targeted drug delivery to skeletal muscles via liposomes. Therefore, this newly identified ssDNA aptamer and nanoliposome modification could be used for the treatment of human skeletal muscle diseases.
    Keywords:  Aptamer; Human skeletal muscle; Nanoliposomes; SELEX; Target delivery
  11. Cell Rep. 2022 Jun 07. pii: S2211-1247(22)00709-4. [Epub ahead of print]39(10): 110927
      Adult muscle stem cells, also known as satellite cells (SCs), play pivotal roles in muscle regeneration, and long non-coding RNA (lncRNA) functions in SCs remain largely unknown. Here, we identify a lncRNA, Lockd, which is induced in activated SCs upon acute muscle injury. We demonstrate that Lockd promotes SC proliferation; deletion of Lockd leads to cell-cycle arrest, and in vivo repression of Lockd in mouse muscles hinders regeneration process. Mechanistically, we show that Lockd directly interacts with RNA helicase DHX36 and the 5'end of Lockd possesses the strongest binding with DHX36. Furthermore, we demonstrate that Lockd stabilizes the interaction between DHX36 and EIF3B proteins; synergistically, this complex unwinds the RNA G-quadruplex (rG4) structure formed at Anp32e mRNA 5' UTR and promotes the translation of ANP32E protein, which is required for myoblast proliferation. Altogether, our findings identify a regulatory Lockd/DHX36/Anp32e axis that promotes myoblast proliferation and acute-injury-induced muscle regeneration.
    Keywords:  ANP32E; CP: Molecular biology; CP: Stem cell research; DHX36; EIF3B; Lockd; RNA G-quadruplexes; muscle regeneration; post-transcriptional regulation; satellite cells; translational regulation
  12. Nat Commun. 2022 Jun 08. 13(1): 3180
      Formation and maintenance of neuromuscular junctions (NMJs) are essential for skeletal muscle function, allowing voluntary movements and maintenance of the muscle tone, thereby preventing atrophy. Generation of NMJs depends on the interaction of motor neurons with skeletal muscle fibers, which initiates a cascade of regulatory events that is essential for patterning of acetylcholine receptor (AChR) clusters at specific sites of the sarcolemma. Here, we show that muscle-specific miRNAs of the miR-1/206/133 family are crucial regulators of a signaling cascade comprising DOK7-CRK-RAC1, which is critical for stabilization and anchoring of postsynaptic AChRs during NMJ development and maintenance. We describe that posttranscriptional repression of CRK by miR-1/206/133 is essential for balanced activation of RAC1. Failure to adjust RAC1 activity severely compromises NMJ function, causing respiratory failure in neonates and neuromuscular symptoms in adult mice. We conclude that miR-1/206/133 serve a specific function for NMJs but are dispensable for skeletal muscle development.
  13. Stem Cell Res Ther. 2022 Jun 03. 13(1): 226
      BACKGROUND: Skeletal muscle mass and function losses in aging individuals are associated with quality of life deterioration and disability. Mesenchymal stromal cells exert immunomodulatory and anti-inflammatory effects and could yield beneficial effects in aging-related degenerative disease.METHODS AND RESULTS: We investigated the efficacy of umbilical cord-derived mesenchymal stromal cells (UC-MSCs) on sarcopenia-related skeletal muscle atrophy and dysfunction in senescence-accelerated mouse prone 10 (SAMP10) mice. We randomly assigned 24-week-old male SAMP10 mice to a UC-MSC treatment group and control group. At 12 weeks post-injection, the UC-MSC treatment had ameliorated sarcopenia-related muscle changes in performance, morphological structures, and mitochondria biogenesis, and it enhanced the amounts of proteins or mRNAs for myosin heavy chain, phospho-AMP-activated protein kinase, phospho-mammalian target of rapamycin, phospho-extracellular signal-regulated kinase1/2, peroxisome proliferator-activated receptor-γ coactivator, GLUT-4, COX-IV, and hepatocyte growth factor in both gastrocnemius and soleus muscles, and it reduced the levels of proteins or mRNAs for cathepsin K, cleaved caspase-3/-8, tumor necrosis factor-α, monocyte chemoattractant protein-1, and gp91phox mRNAs. The UC-MSC treatment retarded mitochondria damage, cell apoptosis, and macrophage infiltrations, and it enhanced desmin/laminin expression and proliferating and CD34+/Integrin α7+ cells in both types of skeletal muscle of the SAMP10 mice. In vitro, we observed increased levels of HGF, PAX-7, and MoyD mRNAs at the 4th passage of UC-MSCs.
    CONCLUSIONS: Our results suggest that UC-MSCs can improve sarcopenia-related skeletal muscle atrophy and dysfunction via anti-apoptosis, anti-inflammatory, and mitochondrial biogenesis mechanisms that might be mediated by an AMPK-PGC1-α axis, indicating that UC-MSCs may provide a promising treatment for sarcopenia/muscle diseases.
    Keywords:  Apoptosis; Inflammation; Mitochondria; SAMP10; Sarcopenia; Umbilical cord-derived mesenchymal stromal cell
  14. J Physiol. 2022 Jun 10.
    Keywords:  fatigue; magnetic resonance spectroscopy; skeletal muscle
  15. Sports Med. 2022 Jun 08.
      BACKGROUND: Systematic investigation of muscle protein synthesis (MPS) responses with or without protein ingestion has been largely limited to resistance training.OBJECTIVE: This systematic review determined the capacity for aerobic-based exercise or high-intensity interval training (HIIT) to stimulate post-exercise rates of MPS and whether protein ingestion further significantly increases MPS compared with placebo.
    METHODS: Three separate models analysed rates of either mixed, myofibrillar, sarcoplasmic, or mitochondrial protein synthesis (PS) following aerobic-based exercise or HIIT: Model 1 (n = 9 studies), no protein ingestion; Model 2 (n = 7 studies), peri-exercise protein ingestion with no placebo comparison; Model 3 (n = 14 studies), peri-exercise protein ingestion with placebo comparison.
    RESULTS: Eight of nine studies and all seven studies in Models 1 and 2, respectively, demonstrated significant post-exercise increases in either mixed or a specific muscle protein pool. Model 3 observed significantly greater MPS responses with protein compared with placebo in either mixed or a specific muscle fraction in 7 of 14 studies. Seven studies showed no difference in MPS between protein and placebo, while three studies reported no significant increases in mitochondrial PS with protein compared with placebo.
    CONCLUSION: Most studies reporting significant increases in MPS were confined to mixed and myofibrillar PS that may facilitate power generating capacity of working skeletal muscle with aerobic-based exercise and HIIT. Only three of eight studies demonstrated significant increases in mitochondrial PS post-exercise, with no further benefits of protein ingestion. This lack of change may be explained by the acute analysis window in most studies and apparent latency in exercise-induced stimulation of mitochondrial PS.
  16. Cell Stress Chaperones. 2022 Jun 10.
      Indigo is a bis-indolic alkaloid that has antioxidant and anti-inflammatory effects reported in literature and is a promissory compound for treating chronic inflammatory diseases. This fact prompted to investigate the effects of this alkaloid in the experimental model of Duchenne muscular dystrophy. The main aim of this study was to evaluate the potential role of the indigo on oxidative stress and related signaling pathways in primary skeletal muscle cell cultures and in the diaphragm muscle from mdx mice. The MTT and Neutral Red assays showed no indigo dose-dependent toxicities in mdx muscle cells at concentrations analyzed (3.12, 6.25, 12.50, and 25.00 μg/mL). Antioxidant effect of indigo, in mdx muscle cells and diaphragm muscle, was demonstrated by reduction in 4-HNE content, H2O2 levels, DHE reaction, and lipofuscin granules. A significant decrease in the inflammatory process was identified by a reduction on TNF and NF-κB levels, on inflammatory area, and on macrophage infiltration in the dystrophic sample, after indigo treatment. Upregulation of PGC-1α and SIRT1 in dystrophic muscle cells treated with indigo was also observed. These results suggest the potential of indigo as a therapeutic agent for muscular dystrophy, through their action anti-inflammatory, antioxidant, and modulator of SIRT1/PGC-1α pathway.
    Keywords:  Activators mitochondrial biogenesis; Indigo; Inflammatory process; Oxidative stress; mdx mice
  17. Gene. 2022 May 31. pii: S0378-1119(22)00427-9. [Epub ahead of print]834 146608
      Myod and Myf5 are muscle-specific basic helix-loop-helix (bHLH) transcription factors that play essential roles in regulating skeletal muscle development and growth. In order to investigate potential function of myod and myf5 of Megalobrama amblycephala, an economically important freshwater fish species, in the present study, we characterized the sequences and expression profiles of M. amblycephala myod and myf5. The open reading frame (ORF) sequences of myod and myf5 encoded 275 and 240 amino acids, respectively, possessing analogous structure with the highly conserved domains, bHLH and C-terminal helix III domains. Spatio-temporal expression patterns revealed that myod and myf5 were predominant in skeletal muscle with the highest expression in white muscle, and the highest at 10 days post-hatching (dph) and the segmentation period, respectively. Furthermore, we evaluated the effects of lipopolysaccharide (LPS) on the expression of muscle-related genes in white and red muscle, and proliferation and differentiation of satellite cells. The myod, myf5 and pax-7 expression generally increased and then decreased with increase of LPS concentration and treatment time in red muscle, while these genes showed inconsistent expression patterns in white muscle. In addition, LPS administration caused the frequency increase of satellite cells in red and white muscle especially at 3 and 7 days after LPS-injection.
    Keywords:  LPS; Megalobrama amblycephala; Myf5; Myod; Satellite cell; Skeletal muscle
  18. Nat Aging. 2021 Dec;1(12): 1148-1161
      Heterochronic blood exchange (HBE) has demonstrated that circulating factors restore youthful features to aged tissues. However, the systemic mediators of those rejuvenating effects remain poorly defined. We show here that the beneficial effect of young blood on aged muscle regeneration was diminished when serum was depleted of extracellular vesicles (EVs). Whereas EVs from young animals rejuvenate aged cell bioenergetics and skeletal muscle regeneration, aging shifts EV subpopulation heterogeneity and compromises downstream benefits on recipient cells. Machine learning classifiers revealed that aging shifts the nucleic acid, but not protein, fingerprint of circulating EVs. Alterations in sub-population heterogeneity were accompanied by declines in transcript levels of the pro-longevity protein, α-Klotho, and injection of EVs improved muscle regeneration in a Klotho mRNA-dependent manner. These studies demonstrate that EVs play a key role in the rejuvenating effects of HBE and that Klotho transcripts within EVs phenocopy the effects of young serum on aged skeletal muscle.
  19. J Gerontol A Biol Sci Med Sci. 2022 Jun 06. pii: glac121. [Epub ahead of print]
      Low skeletal muscle mass is associated with cognitive impairment and dementia in older adults. This review describes the possible underlying pathophysiological mechanisms: systemic inflammation, insulin metabolism, protein metabolism and mitochondrial function. We hypothesize that the central tenet in this pathophysiology is the dysfunctional myokine secretion consequent to minimal physical activity. Myokines, such as fibronectin type III domain containing 5/irisin and cathepsin B are released by physically active muscle and cross the blood-brain-barrier. These myokines upregulate local neurotrophin expression such as brain-derived neurotrophic factor (BDNF) in the brain microenvironment. BDNF exerts anti-inflammatory effects that may be responsible for neuroprotection. Altered myokine secretion due to physical inactivity exacerbates inflammation and impairs muscle glucose metabolism, potentially affecting the transport of insulin across the blood-brain barrier. Our working model also suggests other underlying mechanisms. A negative systemic protein balance, commonly observed in older adults, contributes to low skeletal muscle mass and may also reflect deficient protein metabolism in brain tissues. As a result of age-related loss in skeletal muscle mass, decrease in the abundance of mitochondria and detriments in their function, lead to a decrease in tissue oxidative capacity. Dysfunctional mitochondria in skeletal muscle and brain result in the excessive production of reactive oxygen species, which drives tissue oxidative stress and further perpetuates the dysfunction in mitochondria. Both oxidative stress and accumulation of mitochondrial DNA mutations due to aging drive cellular senescence. A targeted approach in the pathophysiology of low muscle mass and cognition could be to restore myokine balance by physical activity.
    Keywords:  dementia; inflammation; insulin; mitochondria; myokines; protein balance; sarcopenia
  20. J Appl Physiol (1985). 2022 Jun 09.
      This investigation examined the influence of twelve-week ballistic resistance training programs on the IGF-I system in circulation, interstitial fluid and skeletal muscle, at rest and in response to acute exercise. Seventeen college-aged subjects (11 women/6 men; 21.7 ± 3.7 yr) completed an acute ballistic exercise bout before and after the training program. Blood samples were collected pre-, mid- and post-exercise and analyzed for serum total IGF-I, free IGF-I and IGF binding proteins (IGFBPs) 1-4. Dialysate and interstitial free IGF-I were analyzed in vastus lateralis (VL) interstitial fluid collected pre- and post-exercise via microdialysis. Pre- and post-exercise VL muscle biopsies were analyzed for IGF-I protein expression, IGF-I receptor phosphorylation (p-IGF-IR) and AKT phosphorylation (p-AKT). Following training, basal serum IGF-I, free IGF-I, IGFBP-2 and IGFBP-3 decreased while IGFBP-1 and IGFBP-4 increased. Training reduced basal dialysate and interstitial free IGF-I, but had no effect on basal skeletal muscle IGF-I, p-IGF-IR or p-AKT. Acute exercise elicited transient changes in IGF-I system concentrations and downstream anabolic signaling both pre- and post-training; training did not affect this acute exercise response. Post-training, acute exercise induced changes in dialysate/interstitial free IGF-I were strongly correlated with changes in intramuscular IGF-I expression, p-IGF-IR and p-AKT. The divergent influence of resistance training on circulating/interstitial and skeletal muscle IGF-I demonstrates the importance of concurrent, multiple bio-compartment analysis when examining the IGF-I system. As training elicited muscle hypertrophy, these findings indicate that IGF-I's anabolic effects on skeletal muscle are mediated by local, rather than systemic mechanisms.
    Keywords:  Insulin-like growth factor; interstitial fluid; microdialysis; serum; skeletal muscle biopsy
  21. J Sports Sci. 2022 Jun 05. 1-23
      While proximity-to-failure is considered an important resistance training (RT) prescription variable, its influence on physiological adaptations and short-term responses to RT is uncertain. Given the ambiguity in the literature, a scoping review was undertaken to summarise evidence for the influence of proximity-to-failure on muscle hypertrophy, neuromuscular fatigue, muscle damage and perceived discomfort. Literature searching was performed according to PRISMA-ScR guidelines and identified three themes of studies comparing either: i) RT performed to momentary muscular failure versus non-failure, ii) RT performed to set failure (defined as anything other than momentary muscular failure) versus non-failure, and iii) RT performed to different velocity loss thresholds. The findings highlight that no consensus definition for "failure" exists in the literature, and the proximity-to-failure achieved in "non-failure" conditions is often ambiguous and variable across studies. This poses challenges when deriving practical recommendations for manipulating proximity-to-failure in RT to achieve desired outcomes. Based on the limited available evidence, RT to set failure is likely not superior to non-failure RT for inducing muscle hypertrophy, but may exacerbate neuromuscular fatigue, muscle damage, and post-set perceived discomfort versus non-failure RT. Together, these factors may impair post-exercise recovery and subsequent performance, and may also negatively influence long-term adherence to RT.KEY POINTS This scoping review identified three broad themes of studies investigating proximity-to-failure in RT, based on the specific definition of set failure used (and therefore the research question being examined), to improve the validity of study comparisons and interpretations.There is no consensus definition for set failure in RT, and the proximity-to-failure achieved during non-failure RT is often unclear and varies both within and between studies, which together poses challenges when interpreting study findings and deriving practical recommendations regarding the influence of RT proximity-to-failure on muscle hypertrophy and other short-term responses.Based on the limited available evidence, performing RT to set failure is likely not superior to non-failure RT to maximise muscle hypertrophy, but the optimal proximity to failure in RT for muscle hypertrophy is unclear and may be moderated by other RT variables (e.g., load, volume-load). Also, RT performed to set failure likely induces greater neuromuscular fatigue, muscle damage, and perceived discomfort than non-failure RT, which may negatively influence RT performance, post-RT recovery, and long-term adherence.
    Keywords:  Proximity-to-failure; failure; fatigue; hypertrophy; non-failure
  22. Nutrients. 2022 May 27. pii: 2239. [Epub ahead of print]14(11):
      Recently, fasting has been spotlighted from a healthcare perspective. However, the de-tailed biological mechanisms and significance by which the effects of fasting confer health benefits are not yet clear. Due to certain advantages of the zebrafish as a vertebrate model, it is widely utilized in biological studies. However, the biological responses to nutrient metabolism within zebrafish skeletal muscles have not yet been amply reported. Therefore, we aimed to reveal a gene expression profile in zebrafish skeletal muscles in response to fasting-refeeding. Accordingly, mRNA-sequencing and bioinformatics analysis were performed to examine comprehensive gene expression changes in skeletal muscle tissues during fasting-refeeding. Our results produced a novel set of nutrition-related genes under a fasting-refeeding protocol. Moreover, we found that five genes were dramatically upregulated in each fasting (for 24 h) and refeeding (after 3 h), exhibiting a rapid response to the provided conditional changes. The assessment of the gene length revealed that the gene set whose expression was elevated only after 3 h of refeeding had a shorter length, suggesting that nutrition-related gene function is associated with gene length. Taken together, our results from the bioinformatics analyses provide new insights into biological mechanisms induced by fasting-refeeding conditions within zebrafish skeletal muscle.
    Keywords:  fasting; gene length; mRNA-sequencing; refeeding; skeletal muscle; zebrafish
  23. Am J Physiol Regul Integr Comp Physiol. 2022 Jun 07.
      The peroxisome proliferator activated receptor gamma co-activator 1 alpha (PGC-1α) is central to the regulation of cellular and mitochondrial energy homeostasis in mammals, but its role in other vertebrates remains unclear. Indeed, previous work suggests extensive structural and functional divergence of PGC-1α in teleosts but this remains to be directly tested. Here, we describe the initial characterization of heterozygous PGC-1α mutant zebrafish lines created by CRISPR-Cas9 disruptions of an evolutionarily conserved regulatory region of the PGC-1α proximal promoter. Using qPCR, we confirmed the disruption of PGC-1α gene expression in striated muscle, leading to a simultaneous 4-fold increase in mixed skeletal muscle PGC-1α mRNA levels and an opposite 4-fold downregulation in cardiac muscle. In mixed skeletal muscle, most downstream effector genes were largely unaffected yet two mitochondrial lipid transporters, carnitine palmitoyltransferase 1 and 2, were strongly induced. Conversely, PGC-1α depression in cardiac muscle reduced the expression of several transcriptional regulators (estrogen related receptor alpha, nuclear respiratory factor 1 and PGC-1β) without altering metabolic gene expression. Using high resolution respirometry, we determined that white muscle exhibited increased lipid oxidative capacity with little difference in markers of mitochondrial abundance. Finally, using whole animal intermittent respirometry, we show that mutant fish exhibit a 2-fold higher basal metabolism than their wildtype counterparts. Altogether, this new model confirms a central but complex role for PGC-1α in mediating energy utilization in zebrafish and we propose its use as a valuable tool to explore the intricate regulatory pathways of energy homeostasis in a popular biomedical model.
    Keywords:  CRISPR; Metabolism; co-activator; mitochondria; respirometry
  24. Aging Cell. 2022 Jun 03. e13647
      Aging is associated with a loss of metabolic homeostasis, with cofactors such as nicotinamide adenine dinucleotide (NAD+ ) declining over time. The decrease in NAD+ production has been linked to the age-related loss of circulating extracellular nicotinamide phosphoribosyltransferase (eNAMPT), the rate-limiting enzyme in the NAD+ biosynthetic pathway. eNAMPT is found almost exclusively in extracellular vesicles (EVs), providing a mechanism for the distribution of the enzyme in different tissues. Currently, the physiological cause for the release of eNAMPT is unknown, and how it may be affected by age and physical exercise. Here, we show that release of small EVs into the bloodstream is stimulated following moderate intensity exercise in humans. Exercise also increased the eNAMPT content in EVs, most prominently in young individuals with higher aerobic fitness. Both mature fit and young unfit individuals exhibited a limited increase in EV-eNAMPT release following exercise, indicating that this mechanism is related to both the age and physical fitness of a person. Notably, unfit mature individuals were unable to increase the release of eNAMPT in EVs after exercise, suggesting that lower fitness levels and aging attenuate this important signalling mechanism in the body. EVs isolated from exercising humans containing eNAMPT were able to alter the abundance of NAD+ and SIRT1 activity in recipient cells compared to pre-exercise EVs, indicating a pathway for inter-tissue signalling promoted through exercise. Our results suggest a mechanism to limit age-related NAD+ decline, through the systemic delivery of eNAMPT via EVs released during exercise.
    Keywords:  NAD+; NAMPT; SIRT1; aging; exercise; exosome; extracellular vesicles; healthspan
  25. Front Cell Dev Biol. 2022 ;10 788516
      In this study, we examine the cause and progression of mitochondrial diseases linked to the loss of mtRNase P, a three-protein complex responsible for processing and cleaving mitochondrial transfer RNAs (tRNA) from their nascent transcripts. When mtRNase P function is missing, mature mitochondrial tRNA levels are decreased, resulting in mitochondrial dysfunction. mtRNase P is composed of Mitochondrial RNase P Protein (MRPP) 1, 2, and 3. MRPP1 and 2 have their own enzymatic activity separate from MRPP3, which is the endonuclease responsible for cleaving tRNA. Human mutations in all subunits cause mitochondrial disease. The loss of mitochondrial function can cause devastating, often multisystemic failures. When mitochondria do not provide enough energy and metabolites, the result can be skeletal muscle weakness, cardiomyopathy, and heart arrhythmias. These symptoms are complex and often difficult to interpret, making disease models useful for diagnosing disease onset and progression. Previously, we identified Drosophila orthologs of each mtRNase P subunit (Roswell/MRPP1, Scully/MRPP2, Mulder/MRPP3) and found that the loss of each subunit causes lethality and decreased mitochondrial tRNA processing in vivo. Here, we use Drosophila to model mtRNase P mitochondrial diseases by reducing the level of each subunit in skeletal and heart muscle using tissue-specific RNAi knockdown. We find that mtRNase P reduction in skeletal muscle decreases adult eclosion and causes reduced muscle mass and function. Adult flies exhibit significant age-progressive locomotor defects. Cardiac-specific mtRNase P knockdowns reduce fly lifespan for Roswell and Scully, but not Mulder. Using intravital imaging, we find that adult hearts have impaired contractility and exhibit substantial arrhythmia. This occurs for roswell and mulder knockdowns, but with little effect for scully. The phenotypes shown here are similar to those exhibited by patients with mitochondrial disease, including disease caused by mutations in MRPP1 and 2. These findings also suggest that skeletal and cardiac deficiencies induced by mtRNase P loss are differentially affected by the three subunits. These differences could have implications for disease progression in skeletal and heart muscle and shed light on how the enzyme complex functions in different tissues.
    Keywords:  MRPP; arrythmia; cardiomyopathy; drosophila; intravital imaging; mitochondrial RNase P; mitochondrial disease; skeletal muscle
  26. Genes Dis. 2022 Jul;9(4): 1038-1048
      The microRNAs (miRNAs) play an important role in regulating myogenesis by targeting mRNA. However, the understanding of miRNAs in skeletal muscle development and diseases is unclear. In this study, we firstly performed the transcriptome profiling in differentiating C2C12 myoblast cells. Totally, we identified 187 miRNAs and 4260 mRNAs significantly differentially expressed that were involved in myoblast differentiation. We carried out validation of microarray data based on 5 mRNAs and 5 miRNAs differentially expressed and got a consistent result. Then we constructed and validated the significantly up- and down-regulated mRNA-miRNA interaction networks. Four interaction pairs (miR-145a-5p-Fscn1, miR-200c-5p-Tmigd1, miR-27a-5p-Sln and miR-743a-5p-Mob1b) with targeted relationships in differentiated myoblast cells were demonstrated. They are all closely related to myoblast development. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis indicated cell cycle signals important for exploring skeletal muscle development and disease. Functionally, we discovered that miR-743a targeting gene Mps One Binder Kinase Activator-Like 1B (Mob1b) gene in differentiated C2C12. The up-regulated miR-743a can promote the differentiation of C2C12 myoblast. While the down-regulated Mob1b plays a negative role in differentiation. In addition, the expression profile of miR-743a and Mob1b are consistent with skeletal muscle recovery after Cardiotoxin (CTX) injury. Our study revealed that miR-743a-5p regulates myoblast differentiation by targeting Mob1b involved in skeletal muscle development and regeneration. Our findings made a further exploration for mechanisms in myogenesis and might provide potential possible miRNA-based target therapies for skeletal muscle regeneration and disease in the near future.
    Keywords:  Differentiation; Mob1b; Myoblast; Skeletal muscle; Transcriptome; miR-743a-5p
  27. Cells. 2022 May 30. pii: 1789. [Epub ahead of print]11(11):
      O-GlcNAcylation is a highly dynamic, reversible and atypical glycosylation that regulates the activity, biological function, stability, sublocation and interaction of target proteins. O-GlcNAcylation receives and coordinates different signal inputs as an intracellular integrator similar to the nutrient sensor and stress receptor, which target multiple substrates with spatio-temporal analysis specifically to maintain cellular homeostasis and normal physiological functions. Our review gives a brief description of O-GlcNAcylation and its only two processing enzymes and HBP flux, which will help to better understand its physiological characteristics of sensing nutrition and environmental cues. This nutritional and stress-sensitive properties of O-GlcNAcylation allow it to participate in the precise regulation of skeletal muscle metabolism. This review discusses the mechanism of O-GlcNAcylation to alleviate metabolic disorders and the controversy about the insulin resistance of skeletal muscle. The level of global O-GlcNAcylation is precisely controlled and maintained in the "optimal zone", and its abnormal changes is a potential factor in the pathogenesis of cancer, neurodegeneration, diabetes and diabetic complications. Although the essential role of O-GlcNAcylation in skeletal muscle physiology has been widely studied and recognized, it still is underestimated and overlooked. This review highlights the latest progress and potential mechanisms of O-GlcNAcylation in the regulation of skeletal muscle contraction and structural properties.
    Keywords:  O-GlcNAc; insulin resistance; metabolism; sarcomere contraction and structure; skeletal muscle
  28. Int J Mol Sci. 2022 May 27. pii: 6026. [Epub ahead of print]23(11):
      Skeletal muscle consists of long plurinucleate and contractile structures, able to regenerate and repair tissue damage by their resident stem cells: satellite cells (SCs). Reduced skeletal muscle regeneration and progressive atrophy are typical features of sarcopenia, which has important health care implications for humans. Sarcopenia treatment is usually based on physical exercise and nutritional plans, possibly associated with rehabilitation programs, such as vibratory stimulation. Vibrations stimulate muscles and can increase postural stability, balance, and walking in aged and sarcopenic patients. However, the possible direct effect of vibration on SCs is still unclear. Here, we show the effects of focused vibrations administered at increasing time intervals on SCs, isolated from young and aged subjects and cultured in vitro. After stimulations, we found in both young and aged subjects a reduced percentage of apoptotic cells, increased cell size and percentage of aligned cells, mitotic events, and activated cells. We also found an increased number of cells only in young samples. Our results highlight for the first time the presence of direct effects of mechanical vibrations on human SCs. These effects seem to be age-dependent, consisting of a proliferative response of cells derived from young subjects vs. a differentiative response of cells from aged subjects.
    Keywords:  aging; atrophy; focused mechanoacoustic vibration; human satellite cells; rehabilitation; sarcopenia
  29. J Biol Chem. 2022 Jun 06. pii: S0021-9258(22)00539-7. [Epub ahead of print] 102098
      Sarcopenia is an aging-associated attenuation of muscular volume and strength and is the major cause of frailty and falls in elderly individuals. The number of individuals with sarcopenia is rapidly increasing worldwide; however, little is known about the underlying mechanisms of the disease. Sarcopenia often copresents with obesity, and some patients with sarcopenia exhibit accumulation of peri- or intra-organ adipose tissue as ectopic fat deposition, including atrophied skeletal muscle. In this study, we showed that transplantation of the peri-muscular adipose tissue (PMAT) to the hindlimb thigh muscles of young mice decreased the number of Integrin α7/CD29-double positive muscular stem/progenitor cells and that the reaction was mediated by PMAT-derived exosomes. We also found that the inhibition of cell proliferation was induced by Let-7d-3p miRNA that targets HMGA2, which is an important transcription factor for stem cell self-renewal, in muscular stem/progenitor cells and the composite molecular reaction in aged adipocytes. Reduction of Let-7 miRNA repressor Lin28A/B and activation of NFκB signaling can lead to the accumulation of Let-7d-3p in the exosomes of aged PMAT. These findings suggest a novel crosstalk between adipose tissue and skeletal muscle in the development of aging-associated muscular atrophy and indicate that adipose tissue-derived miRNAs may play a key role in sarcopenia.
    Keywords:  HMGA2; Let-7; Peri-Muscular Adipose Tissue; inter-organ communication; muscular satellite cells; stem cell aging
  30. Data Brief. 2022 Aug;43 108321
      This data article reports the level of expression of messenger RNA (mRNA) obtained from a set of 18 skeletal muscle samples using Affymetrix Genechips Exon arrays. Data were obtained from Gastrocnemius muscle of C57BL6 male mice at 3 distinct age groups, 2, 11 and 25 months old representing young, mature adult and aged groups. The data submitted to GEO constitute a large dataset of 15,300 mRNA levels. The data include eighteen .CEL files obtained after scanning mouse exon arrays and one .xls file obtained after processing with Genespring GX 14.9. Three distinct files containing affymetrix data processed using Genespring and analyzed for differences between stages 2 per 2 are provided as supplementary data.
    Keywords:  ageing; exon array; mature adult; mouse; muscle; transcriptomic; young
  31. Int J Mol Sci. 2022 May 26. pii: 6002. [Epub ahead of print]23(11):
      Myogenesis is a central step in prenatal myofiber formation, postnatal myofiber hypertrophy, and muscle damage repair in adulthood. RNA-Seq technology has greatly helped reveal the molecular mechanism of myogenesis, but batch effects in different experiments inevitably lead to misinterpretation of differentially expressed genes (DEGs). We previously applied the robust rank aggregation (RRA) method to effectively circumvent batch effects across multiple RNA-Seq datasets from 3T3-L1 cells. Here, we also used the RRA method to integrate nine RNA-Seq datasets from C2C12 cells and obtained 3140 robust DEGs between myoblasts and myotubes, which were then validated with array expression profiles and H3K27ac signals. The upregulated robust DEGs were highly enriched in gene ontology (GO) terms related to muscle cell differentiation and development. Considering that the cooperative binding of transcription factors (TFs) to enhancers to regulate downstream gene expression is a classical epigenetic mechanism, differentially expressed TFs (DETFs) were screened, and potential novel myogenic factors (MAF, BCL6, and ESR1) with high connection degree in protein-protein interaction (PPI) network were presented. Moreover, KLF5 cooperatively binds with the three key myogenic factors (MYOD, MYOG, and MEF2D) in C2C12 cells. Motif analysis speculates that the binding of MYOD and MYOG is KLF5-independent, while MEF2D is KLF5-dependent. It was revealed that KLF5-binding sites could be exploited to filter redundant MYOD-, MYOG-, and MEF2D-binding sites to focus on key enhancers for myogenesis. Further functional annotation of KLF5-binding sites suggested that KLF5 may regulate myogenesis through the PI3K-AKt signaling pathway, Rap1 signaling pathway, and the Hippo signaling pathway. In general, our study provides a wealth of untapped candidate targets for myogenesis and contributes new insights into the core regulatory mechanisms of myogenesis relying on KLF5-binding signal.
    Keywords:  KLF5; differentially expressed genes; enhancer; myogenesis; robust rank aggregation
  32. Skelet Muscle. 2022 Jun 10. 12(1): 12
      BACKGROUND: The analysis of in vitro cultures of human adult muscle stem cells obtained from biopsies delineates the potential of skeletal muscles and may help to understand altered muscle morphology in patients. In these analyses, the fusion index is a commonly used quantitative metric to assess the myogenic potency of the muscle stem cells. Since the fusion index only partly describes myogenic potency, we developed the Myotube Analyzer tool, which combines the definition of the fusion index with extra features of myonuclei and myotubes obtained from satellite cell cultures.RESULTS: The software contains image adjustment and mask editing functions for preprocessing and semi-automatic segmentation, while other functions can be used to determine the features of nuclei and myotubes. The fusion index and a set of five novel parameters were tested for reliability and validity in a comparison between satellite cell cultures from children with cerebral palsy and typically developing children. These novel parameters quantified extra nucleus and myotube properties and can be used to describe nucleus clustering and myotube shape. Two analyzers who were trained in cell culture defined all parameters using the Myotube Analyzer app. Out of the six parameters, five had good reliability reflected by good intra-class correlation coefficients (> 0.75). Children with cerebral palsy were significantly different from the typically developing children (p < 0.05) for five parameters, and for three of the six parameters, these differences exceeded the minimal detectable differences.
    CONCLUSIONS: The Myotube Analyzer can be used for the analysis of fixed differentiated myoblast cultures with nuclear and MyHC staining. The app can calculate the fusion index, an already existing parameter, but also provides multiple new parameters to comprehensively describe myogenic potential in its output. The raw data used to determine these parameters are also available in the output. The parameters calculated by the tool can be used to detect differences between cultures from children with cerebral palsy and typically developing children. Since the program is open source, users can customize it to fit their own analysis requirements.
    Keywords:  Cerebral palsy; Fusion index; Image analysis; Satellite cell cultures
  33. Semin Cell Dev Biol. 2022 Jun 06. pii: S1084-9521(22)00173-2. [Epub ahead of print]
      The purpose of this review is to explore and discuss the impacts of augmented training volume, intensity, and duration on the phosphorylation/activation of key signaling protein - AMPK, CaMKII and PGC-1α - involved in the initiation of mitochondrial biogenesis. Specifically, we explore the impacts of augmented exercise protocols on AMP/ADP and Ca2+ signaling and changes in post exercise PGC - 1α gene expression. Although AMP/ADP concentrations appear to increase with increasing intensity and during extended durations of higher intensity exercise AMPK activation results are varied with some results supporting and intensity/duration effect and others not. Similarly, CaMKII activation and signaling results following exercise of different intensities and durations are inconsistent. The PGC-1α literature is equally inconsistent with only some studies demonstrating an effect of intensity on post exercise mRNA expression. We present a novel meta-analysis that suggests that the inconsistency in the PGC-1α literature may be due to sample size and statistical power limitations owing to the effect of intensity on PGC-1α expression being small. There is little data available regarding the impact of exercise duration on PGC-1α expression. We highlight the need for future well designed, adequately statistically powered, studies to clarify our understanding of the effects of volume, intensity, and duration on the induction of mitochondrial biogenesis by exercise.
    Keywords:  AMPK; CaMK; Duration; Exercise; Intensity; Mitochondrial biogenesis; PGC-1a; Volume
  34. Eur J Neurol. 2022 Jun 06.
      BACKGROUND: Among post-COVID-19 symptoms, fatigue is reported as one of the most common, even after mild acute infection, and as the cause of fatigue, myopathy diagnosed by electromyography has been proposed in previous reports. This study aimed to explore the histopathological changes in patients with post-COVID-19 fatigue.METHODS: Sixteen patients (mean age:46 years) with post-COVID-19 complaints of fatigue, myalgia or weakness persisting for up to 14 months were included. In all patients, quantitative electromyography and muscle biopsies analysed with light and electron microscopy were taken.
    RESULTS: Muscle weakness was present in 50%, myopathic electromyography in 75% while in all patients, there were histological changes. Muscle fiber atrophy was found in 38%, and 56% showed indications of fiber regeneration. Mitochondrial changes, comprising loss of COX activity, subsarcollemmal accumulation and/or abnormal cristae, were present in 62%. Inflammation was found in 62%, seen as T-lymphocytes and/or muscle fiber HLA-ABC expression. In 75%, capillaries were affected involving basal lamina and cells. In two patients, uncommon amounts of basal lamina were found, not only surrounding muscle fibers but also around nerves and capillaries.
    CONCLUSIONS: The wide variety of histological changes in this study suggest that skeletal muscles may be a major target of SARS-CoV-2 causing muscular post-COVID-19 symptoms. The mitochondrial changes, inflammation and capillary injury in muscle biopsies can cause fatigue in part due to reduced energy supply. Since most patients had mild-moderate acute affection, the new variants that might cause less severe acute disease could still have the ability to cause long-term myopathy.
    Keywords:  Electromyography; Fatigue; Long-term COVID; Muscle biopsy; Myopathy; Post COVID Syndrome
  35. Cell. 2022 Jun 09. pii: S0092-8674(22)00529-3. [Epub ahead of print]185(12): 2057-2070.e15
      Spinal muscular atrophy (SMA) is a motor-neuron disease caused by mutations of the SMN1 gene. The human paralog SMN2, whose exon 7 (E7) is predominantly skipped, cannot compensate for the lack of SMN1. Nusinersen is an antisense oligonucleotide (ASO) that upregulates E7 inclusion and SMN protein levels by displacing the splicing repressors hnRNPA1/A2 from their target site in intron 7. We show that by promoting transcriptional elongation, the histone deacetylase inhibitor VPA cooperates with a nusinersen-like ASO to promote E7 inclusion. Surprisingly, the ASO promotes the deployment of the silencing histone mark H3K9me2 on the SMN2 gene, creating a roadblock to RNA polymerase II elongation that inhibits E7 inclusion. By removing the roadblock, VPA counteracts the chromatin effects of the ASO, resulting in higher E7 inclusion without large pleiotropic effects. Combined administration of the nusinersen-like ASO and VPA in SMA mice strongly synergizes SMN expression, growth, survival, and neuromuscular function.
    Keywords:  alternative splicing; antisense oligonucleotide therapy; chromatin; histone acetylation; spinal muscular atrophy; transcriptional elongation