bims-moremu Biomed News
on Molecular regulators of muscle mass
Issue of 2021‒10‒24
fifty-five papers selected by
Anna Vainshtein
Craft Science Inc.

  1. Front Cell Dev Biol. 2021 ;9 736813
      Adult skeletal muscle has robust regenerative capabilities due to the presence of a resident stem cell population called satellite cells. Muscle injury leads to these normally quiescent cells becoming molecularly and metabolically activated and embarking on a program of proliferation, migration, differentiation, and fusion culminating in the repair of damaged tissue. These processes are highly coordinated by paracrine signaling events that drive cytoskeletal rearrangement and cell-cell communication. Pannexins are a family of transmembrane channel proteins that mediate paracrine signaling by ATP release. It is known that Pannexin1 (Panx1) is expressed in skeletal muscle, however, the role of Panx1 during skeletal muscle development and regeneration remains poorly understood. Here we show that Panx1 is expressed on the surface of myoblasts and its expression is rapidly increased upon induction of differentiation and that Panx1-/- mice exhibit impaired muscle regeneration after injury. Panx1-/- myoblasts activate the myogenic differentiation program normally, but display marked deficits in migration and fusion. Mechanistically, we show that Panx1 activates P2 class purinergic receptors, which in turn mediate a lipid signaling cascade in myoblasts. This signaling induces bleb-driven amoeboid movement that in turn supports myoblast migration and fusion. Finally, we show that Panx1 is involved in the regulation of cell-matrix interaction through the induction of ADAMTS (Disintegrin-like and Metalloprotease domain with Thrombospondin-type 5) proteins that help remodel the extracellular matrix. These studies reveal a novel role for lipid-based signaling pathways activated by Panx1 in the coordination of myoblast activities essential for skeletal muscle regeneration.
    Keywords:  Panx1; blebbing; lipid signaling; myoblast fusion; myoblast migration; pannexins
  2. Front Physiol. 2021 ;12 732308
      Unc51 like autophagy activating kinase 1 (Ulk1), the primary autophagy regulator, has been linked to metabolic adaptation in skeletal muscle to exercise training. Here we compared the roles of Ulk1 and homologous Ulk2 in skeletal muscle insulin action following exercise training to gain more mechanistic insights. Inducible, skeletal muscle-specific Ulk1 knock-out (Ulk1-iMKO) mice and global Ulk2 knock-out (Ulk2-/-) mice were subjected to voluntary wheel running for 6 weeks followed by assessment of exercise capacity, glucose tolerance, and insulin signaling in skeletal muscle after a bolus injection of insulin. Both Ulk1-iMKO and Ulk2-/- mice had improved endurance exercise capacity post-exercise. Ulk1-iMKO did not improve glucose clearance during glucose tolerance test, while Ulk2-/- had only marginal improvement. However, exercise training-induced improvement of insulin action in skeletal muscle, indicated by Akt-S473 phosphorylation, was only impaired in Ulk1-iMKO. These data suggest that Ulk1, but not Ulk2, is required for exercise training-induced improvement of insulin action in skeletal muscle, implicating crosstalk between catabolic and anabolic signaling as integral to metabolic adaptation to energetic stress.
    Keywords:  Unc51 like autophagy activating kinase; autophagy; exercise; insulin signaling; skeletal muscle
  3. PLoS One. 2021 ;16(10): e0258831
      Cancer causes mitochondrial alterations in skeletal muscle, which may progress to muscle wasting and, ultimately, to cancer cachexia. Understanding how exercise adaptations are altered by cancer and cancer treatment is important for the effective design of exercise interventions aimed at improving cancer outcomes. We conducted an exploratory study to investigate how tumor burden and cancer immunotherapy treatment (anti-PD-1) modify the skeletal muscle mitochondrial response to exercise training in mice with transplantable tumors (B16-F10 melanoma and EO771 breast cancer). Mice remained sedentary or were provided with running wheels for ~19 days immediately following tumor implant while receiving no treatment (Untreated), isotype control antibody (IgG2a) or anti-PD-1. Exercise and anti-PD-1 did not alter the growth rate of either tumor type, either alone or in combination therapy. Untreated mice with B16-F10 tumors showed increases in most measured markers of skeletal muscle mitochondrial content following exercise training, as did anti-PD-1-treated mice, suggesting increased mitochondrial content following exercise training in these groups. However, mice with B16-F10 tumors receiving the isotype control antibody did not exhibit increased skeletal muscle mitochondrial content following exercise. In untreated mice with EO771 tumors, only citrate synthase activity and complex IV activity were increased following exercise. In contrast, IgG2a and anti-PD-1-treated groups both showed robust increases in most measured markers following exercise. These results indicate that in mice with B16-F10 tumors, IgG2a administration prevents exercise adaptation of skeletal muscle mitochondria, but adaptation remains intact in mice receiving anti-PD-1. In mice with EO771 tumors, both IgG2a and anti-PD-1-treated mice show robust skeletal muscle mitochondrial exercise responses, while untreated mice do not. Taken together, we postulate that immune modulation may enhance skeletal muscle mitochondrial response to exercise in tumor-bearing mice, and suggest this as an exciting new avenue for future research in exercise oncology.
  4. Cells. 2021 Oct 12. pii: 2725. [Epub ahead of print]10(10):
      Skeletal myogenesis is required to maintain muscle mass and integrity, and impaired myogenesis is causally linked to the etiology of muscle wasting. Recently, it was shown that excessive uptake of saturated fatty acids (SFA) plays a significant role in the pathogenesis of muscle wasting. Although microRNA (miRNA) is implicated in the regulation of myogenesis, the molecular mechanism whereby SFA-induced miRNAs impair myogenic differentiation remains largely unknown. Here, we investigated the regulatory roles of miR-325-3p on CFL2 expression and myogenic differentiation in C2C12 myoblasts. PA impeded myogenic differentiation, concomitantly suppressed CFL2 and induced miR-325-3p. Dual-luciferase analysis revealed that miR-325-3p directly targets the 3'UTR of CFL2, thereby suppressing the expression of CFL2, a crucial factor for actin dynamics. Transfection with miR-325-3p mimic resulted in the accumulation of actin filaments (F-actin) and nuclear Yes-associated protein (YAP) in myoblasts and promoted myoblast proliferation and cell cycle progression. Consequently, miR-325-3p mimic significantly attenuated the expressions of myogenic factors and thereby impaired the myogenic differentiation of myoblasts. The roles of miR-325-3p on CFL2 expression, F-actin modulation, and myogenic differentiation suggest a novel miRNA-mediated regulatory mechanism of myogenesis and PA-inducible miR-325-3p may be a critical mediator between obesity and muscle wasting.
    Keywords:  CFL2; differentiation; miR-325-3p; microRNA; palmitic acid; proliferation
  5. Front Cell Dev Biol. 2021 ;9 721543
      Myogenic differentiation, muscle stem cell functionality, and regeneration of skeletal muscle are cellular processes under tight control of various signaling pathways. Here, we investigated the role of non-canonical NF-κB signaling in myogenic differentiation, muscle stem cell functionality, and regeneration of skeletal muscle. We stimulated non-canonical NF-κB signaling with an agonistically acting antibody of the lymphotoxin beta receptor (LTβR). Interestingly, we found that stimulation of non-canonical NF-κB signaling through the LTβR agonist impairs myogenic differentiation, muscle stem cell function, and regeneration of skeletal muscle. Furthermore, we show that stimulation of non-canonical NF-κB signaling by the LTβR agonist coincides with activation of canonical NF-κB signaling. We suggest a direct crosstalk between canonical and non-canonical NF-κB signaling during myogenic differentiation which is required for proper myogenic differentiation and thereby regeneration of skeletal muscle.
    Keywords:  LTβR; NF-kappa B; NF-κB; lymphotoxin-β-receptor; muscle stem cell; myogenic differentiation; regeneration; satellite cell
  6. J Cachexia Sarcopenia Muscle. 2021 Oct 19.
      BACKGROUND: Chemotherapy is an essential treatment to combat solid tumours and mitigate metastasis. Chemotherapy causes side effects including muscle wasting and weakness. Regulated in Development and DNA Damage Response 1 (REDD1) is a stress-response protein that represses the mechanistic target of rapamycin (mTOR) in complex 1 (mTORC1), and its expression is increased in models of muscle wasting. The aim of this study was to determine if deletion of REDD1 is sufficient to attenuate chemotherapy-induced muscle wasting and weakness in mice.METHODS: C2C12 myotubes were treated with carboplatin, and changes in myotube diameter were measured. Protein synthesis was measured by puromycin incorporation, and REDD1 mRNA and protein expression were analysed in myotubes treated with carboplatin. Markers of mTORC1 signalling were measured by western blot. REDD1 global knockout mice and wild-type mice were treated with a single dose of carboplatin and euthanized 7 days later. Body weight, hindlimb muscle weights, forelimb grip strength, and extensor digitorum longus whole muscle contractility were measured in all groups. Thirty minutes prior to euthanasia, mice were injected with puromycin to measure puromycin incorporation in skeletal muscle.
    RESULTS: C2C12 myotube diameter was decreased at 24 (P = 0.0002) and 48 h (P < 0.0001) after carboplatin treatment. Puromycin incorporation was decreased in myotubes treated with carboplatin for 24 (P = 0.0068) and 48 h (P = 0.0008). REDD1 mRNA and protein expression were increased with carboplatin treatment (P = 0.0267 and P = 0.0015, respectively), and this was accompanied by decreased phosphorylation of Akt T308 (P < 0.0001) and S473 (P = 0.0006), p70S6K T389 (P = 0.0002), and 4E-binding protein 1 S65 (P = 0.0341), all markers of mTORC1 activity. REDD1 mRNA expression was increased in muscles from mice treated with carboplatin (P = 0.0295). Loss of REDD1 reduced carboplatin-induced body weight loss (P = 0.0013) and prevented muscle atrophy in mice. REDD1 deletion prevented carboplatin-induced decrease of protein synthesis (P = 0.7626) and prevented muscle weakness.
    CONCLUSIONS: Carboplatin caused loss of body weight, muscle atrophy, muscle weakness, and inhibition of protein synthesis. Loss of REDD1 attenuates muscle atrophy and weakness in mice treated with carboplatin. Our study illustrates the importance of REDD1 in the regulation of muscle mass with chemotherapy treatment and may be an attractive therapeutic target to combat cachexia.
    Keywords:  Cachexia; Chemotherapy; Muscle atrophy; Muscle weakness; Protein synthesis; REDD1
  7. J Muscle Res Cell Motil. 2021 Oct 18.
      Skeletal muscles undergo adaptations in response to mechanical stimuli such as stretching. However, there is limited evidence regarding the hypertrophic effects of passive repetitive stretching in vivo. We examined the effect of passive repetitive stretching on skeletal muscle myofiber morphology, satellite cell content, and messenger RNA expression of myogenic regulatory factors and signaling molecules involved in muscle protein synthesis and degradation. The gastrocnemius muscles of mice were stretched 15 times/min by manual ankle dorsiflexion for 15 min, 5 days a week for 2 weeks. We found that passive repetitive stretching significantly increased myofiber cross-sectional area. In stretched gastrocnemius muscles, the messenger RNA expression of p70S6K and myogenin was upregulated, whereas MuRF1, MAFbx, myostatin, and 4E-BP1 were downregulated. The phosphorylation level of p70S6K was significantly increased in stretched muscles. The number of Pax7+ cells was unaffected. Passive repetitive stretching induces muscle hypertrophy by regulating signaling pathways involved in muscle protein turnover. These findings are applicable to clinical muscle strengthening and for the maintenance of skeletal muscle mass and function in patients who are unconscious or paralyzed.
    Keywords:  Gene expression; Mechanical stimulation; Muscle hypertrophy; Passive stretching
  8. J Appl Physiol (1985). 2021 Oct 21.
      Cancer cachexia is a wasting disorder associated with advanced cancer that contributes to mortality. Cachexia is characterized by involuntary loss of body weight and muscle weakness that affects physical function. Regulated in DNA damage and development 1 (REDD1) is a stress-response protein that is transcriptionally upregulated in muscle during wasting conditions and inhibits mechanistic target of rapamycin complex 1 (mTORC1). C2C12 myotubes treated with Lewis lung carcinoma (LLC)-conditioned media increased REDD1 mRNA expression and decreased myotube diameter. To investigate the role of REDD1 in cancer cachexia, we inoculated 12-week old male wild-type or global REDD1 knockout (REDD1 KO) mice with LLC cells and euthanized 28-days later. Wild-type mice had increased skeletal muscle REDD1 expression, and REDD1 deletion prevented loss of body weight and lean tissue mass, but not fat mass. We found that REDD1 deletion attenuated loss of individual muscle weights and loss of myofiber cross sectional area. We measured markers of the Akt/mTORC1 pathway and found that, unlike wild-type mice, phosphorylation of both Akt and 4E-BP1 was maintained in the muscle of REDD1 KO mice after LLC inoculation, suggesting that loss of REDD1 is beneficial in maintaining mTORC1 activity in mice with cancer cachexia. We measured Foxo3a phosphorylation as a marker of the ubiquitin proteasome pathway and autophagy and found that REDD1 deletion prevented dephosphorylation of Foxo3a in muscles from cachectic mice. Our data provides evidence that REDD1 plays an important role in cancer cachexia through the regulation of both protein synthesis and protein degradation pathways.
    Keywords:  REDD1; cachexia; muscle
  9. Life Sci. 2021 Oct 19. pii: S0024-3205(21)01054-7. [Epub ahead of print] 120067
      AIMS: Brain derived neurotrophic factor (BDNF) and the related receptors TrkB and p75NTR are expressed in skeletal muscle, yet their functions remain to be fully understood. Skeletal muscle denervation, which occurs in spinal injury, peripheral neuropathies, and aging, negatively affects muscle mass and function. In this study, we wanted to understand the role of BDNF, TrkB, and p75NTR in denervation-induced adverse effects on skeletal muscle.MAIN METHODS: Mice with unilateral sciatic denervation were used. Protein levels of pro- and mature BDNF, TrkB, p75NTR, activations of their downstream signaling pathways, and inflammation in the control and denervated muscle were measured with Western blot and tissue staining. Treatment with a p75NTR inhibitor and BDNF skeletal muscle specific knockout in mice were used to examine the role of p75NTR and pro-BDNF.
    KEY FINDINGS: In denervated muscle, pro-BDNF and p75NTR were significantly upregulated, and JNK and NF-kB, two major downstream signaling pathways of p75NTR, were activated, along with muscle atrophy and inflammation. Inhibition of p75NTR using LM11A-31 significantly reduced JNK activation and inflammatory cytokines in the denervated muscle. Moreover, skeletal muscle specific knockout of BDNF reduced pro-BDNF level, JNK activation and inflammation in the denervated muscle.
    SIGNIFICANCE: These results reveal for the first time that the upregulation of pro-BDNF and activation of p75NTR pathway are involved in denervation-induced inflammation in skeletal muscle. The results suggest that inhibition of pro-BDNF-p75NTR pathway can be a new target to treat skeletal muscle inflammation.
    Keywords:  BDNF; Denervation; Inflammation; Myokine; p75NTR
  10. Cells. 2021 Oct 12. pii: 2722. [Epub ahead of print]10(10):
      Intracellular Ca2+ ions represent a signaling mediator that plays a critical role in regulating different muscular cellular processes. Ca2+ homeostasis preservation is essential for maintaining skeletal muscle structure and function. Store-operated Ca2+ entry (SOCE), a Ca2+-entry process activated by depletion of intracellular stores contributing to the regulation of various function in many cell types, is pivotal to ensure a proper Ca2+ homeostasis in muscle fibers. It is coordinated by STIM1, the main Ca2+ sensor located in the sarcoplasmic reticulum, and ORAI1 protein, a Ca2+-permeable channel located on transverse tubules. It is commonly accepted that Ca2+ entry via SOCE has the crucial role in short- and long-term muscle function, regulating and adapting many cellular processes including muscle contractility, postnatal development, myofiber phenotype and plasticity. Lack or mutations of STIM1 and/or Orai1 and the consequent SOCE alteration have been associated with serious consequences for muscle function. Importantly, evidence suggests that SOCE alteration can trigger a change of intracellular Ca2+ signaling in skeletal muscle, participating in the pathogenesis of different progressive muscle diseases such as tubular aggregate myopathy, muscular dystrophy, cachexia, and sarcopenia. This review provides a brief overview of the molecular mechanisms underlying STIM1/Orai1-dependent SOCE in skeletal muscle, focusing on how SOCE alteration could contribute to skeletal muscle wasting disorders and on how SOCE components could represent pharmacological targets with high therapeutic potential.
    Keywords:  Orai1; SOCE-related skeletal muscle diseases; STIM1; skeletal muscle; store-operated calcium entry (SOCE)
  11. Cells. 2021 Sep 29. pii: 2586. [Epub ahead of print]10(10):
      Elevated mitochondrial reactive oxygen species (mROS) and an increase in caspase-3 activity are established mechanisms that lead to skeletal muscle atrophy via the upregulation of protein degradation pathways. However, the mechanisms upstream of an increase in mROS and caspase-3 activity in conditions of muscle atrophy have not been identified. Based upon knowledge that an event known as mitochondrial permeability transition (MPT) causes an increase in mROS emission and the activation of caspase-3 via mitochondrial release of cytochrome c, as well as the circumstantial evidence for MPT in some muscle atrophy conditions, we tested MPT as a mechanism of atrophy. Briefly, treating cultured single mouse flexor digitorum brevis (FDB) fibers from adult mice with a chemical inducer of MPT (Bz423) for 24 h caused an increase in mROS and caspase-3 activity that was accompanied by a reduction in muscle fiber diameter that was able to be prevented by inhibitors of MPT, mROS, or caspase-3 (p < 0.05). Similarly, a four-day single fiber culture as a model of disuse caused atrophy that could be prevented by inhibitors of MPT, mROS, or activated caspase-3. As such, our results identify MPT as a novel mechanism of skeletal muscle atrophy that operates through mROS emission and caspase-3 activation.
    Keywords:  ROS; caspase-3; mitochondrial permeability transition pore; skeletal muscle atrophy
  12. J Cell Mol Med. 2021 Oct 22.
      Skeletal muscle plays a pivotal role in the maintenance of physical and metabolic health. Skeletal muscle atrophy usually results in physical disability, inferior quality of life and higher health care costs. The higher incidence of muscle atrophy in obese and ageing groups is due to increased levels of inflammatory factors during obesity and ageing. Dihydromyricetin, as a bioactive polyphenol, has been used for anti-inflammatory, anti-tumour and improving insulin sensitivity. However, there are no published reports demonstrated the dihydromyricetin effect on inflammation-induced skeletal muscle atrophy. In this study, we first confirmed the role of dihydromyricetin in inflammation-induced skeletal muscle atrophy in vivo and in vitro. Then, we demonstrated that dihydromyricetin resisted inflammation-induced skeletal muscle atrophy by activating Ca2+ -CaMKK-AMPK through signal pathway blockers, Ca2+ probes and immunofluorescence. Finally, we clarified that dihydromyricetin activated Ca2+ -CaMKK-AMPK signalling pathway through interaction with the ryanodine receptor, its target protein, by drug affinity responsive target stability (DARTS). Our results not only demonstrated that dihydromyricetin resisted inflammation-induced muscle atrophy via the ryanodine receptor-CaMKK-AMPK signal pathway but also discovered that the target protein of dihydromyricetin is the ryanodine receptor. Our results provided experimental data for the development of dihydromyricetin as a functional food and new therapeutic strategies for treating or preventing skeletal muscle atrophy.
    Keywords:  dihydromyricetin; mice; obesity; skeletal muscle atrophy; target protein
  13. Diagnostics (Basel). 2021 Oct 12. pii: 1879. [Epub ahead of print]11(10):
      BACKGROUND: The potassium channel encoded by the ether-a-gogo-related gene 1A (erg1a) has been detected in the atrophying skeletal muscle of mice experiencing either muscle disuse or cancer cachexia and further evidenced to contribute to muscle deterioration by enhancing ubiquitin proteolysis; however, to our knowledge, ERG1A has not been reported in human skeletal muscle.METHODS AND RESULTS: Here, using immunohistochemistry, we detect ERG1A immunofluorescence in human Rectus abdominis skeletal muscle sarcolemma. Further, using single point brightness data, we report the detection of ERG1A immunofluorescence at low levels in the Rectus abdominis muscle sarcolemma of young adult humans and show that it trends toward greater levels (10.6%) in healthy aged adults. Interestingly, we detect ERG1A immunofluorescence at a statistically greater level (53.6%; p < 0.05) in the skeletal muscle of older cancer patients than in age-matched healthy adults. Importantly, using immunoblot, we reveal that lower mass ERG1A protein is 61.5% (p < 0.05) more abundant in the skeletal muscle of cachectic older adults than in healthy age-matched controls. Additionally, we report that the ERG1A protein is detected in a cultured human rhabdomyosarcoma line that may be a good in vitro model for the study of ERG1A in muscle.
    CONCLUSIONS: The data demonstrate that ERG1A is detected more abundantly in the atrophied skeletal muscle of cancer patients, suggesting it may be related to muscle loss in humans as it has been shown to be in mice experiencing muscle atrophy as a result of malignant tumors.
    Keywords:  cachexia; dystrophin; erg1a; potassium channel; rhabdomyosarcoma; sarcolemma membrane
  14. Nat Rev Mol Cell Biol. 2021 Oct 18.
      Skeletal muscle contains a designated population of adult stem cells, called satellite cells, which are generally quiescent. In homeostasis, satellite cells proliferate only sporadically and usually by asymmetric cell division to replace myofibres damaged by daily activity and maintain the stem cell pool. However, satellite cells can also be robustly activated upon tissue injury, after which they undergo symmetric divisions to generate new stem cells and numerous proliferating myoblasts that later differentiate to muscle cells (myocytes) to rebuild the muscle fibre, thereby supporting skeletal muscle regeneration. Recent discoveries show that satellite cells have a great degree of population heterogeneity, and that their cell fate choices during the regeneration process are dictated by both intrinsic and extrinsic mechanisms. Extrinsic cues come largely from communication with the numerous distinct stromal cell types in their niche, creating a dynamically interactive microenvironment. This Review discusses the role and regulation of satellite cells in skeletal muscle homeostasis and regeneration. In particular, we highlight the cell-intrinsic control of quiescence versus activation, the importance of satellite cell-niche communication, and deregulation of these mechanisms associated with ageing. The increasing understanding of how satellite cells are regulated will help to advance muscle regeneration and rejuvenation therapies.
  15. Front Endocrinol (Lausanne). 2021 ;12 681267
      Sarcopenia, characterized by reduced muscle function as well as muscle mass, has been a public health problem with increasing prevalence. It might result from aging, injury, hormone imbalance and other catabolic conditions. Recently, exosomes were considered to regulate muscle regeneration and protein synthesis. In order to confirm the effect of BMSC-derived exosomes (BMSC-Exos) on muscle, dexamethasone-induced muscle atrophy was built both in vitro and in vivo. In the present research, BMSC-Exos attenuated the decrease of myotube diameter induced by dexamethasone, indicating that BMSC-Exos played a protective role in skeletal muscle atrophy. Further mechanism analysis exhibited that the content of miR-486-5p in C2C12 myotubes was up-regulated after treated with BMSC-Exos. Meanwhile, BMSC-Exos markedly downregulated the nuclear translocation of FoxO1, which plays an important role in muscle differentiation and atrophy. Importantly, the miR-486-5p inhibitor reversed the decreased expression of FoxO1 induced by BMSC-Exos. In animal experiments, BMSC-Exos inhibited dexamethasone-induced muscle atrophy, and miR-486-5p inhibitor reversed the protective effect of BMSC-Exos. These results indicating that BMSC-derived exosomes inhibit dexamethasone-induced muscle atrophy via miR486-5p/Foxo1 Axis.
    Keywords:  FoxO1; bone marrow mesenchymal stem cell; exosomes; miR-486-5p; muscle atrophy
  16. Phytomedicine. 2021 Oct 04. pii: S0944-7113(21)00334-2. [Epub ahead of print]93 153791
      BACKGROUND: Recent advancements in understanding β-escin action provide basis for new therapeutic claims for the drug. β-escin-evoked attenuation of NF-κB-dependent signaling, increase in MMP-14 and decrease in COUP-TFII content and a rise in cholesterol biosynthesis could be beneficial in alleviating muscle-damaging processes.PURPOSE: The aim of this study was to investigate the effect of β-escin on skeletal muscle regeneration.
    METHODS: Rat model of cardiotoxin-induced injury of fast-twich extensor digitorum longus (EDL) and slow-twich soleus (SOL) muscles and C2C12 myoblast cells were used in the study. We evaluated muscles obtained on day 3 and 14 post-injury by histological analyses of muscle fibers, connective tissue, and mononuclear infiltrate, by immunolocalization of macrophages and by qPCR to quantify the expression of muscle regeneration-related genes. Mechanism of drug action was investigated in vitro by assessing cell viability, NF-κB activation, MMP-2 and MMP-9 secretion, and ALDH activity.
    RESULTS: In rat model, β-escin rescues regenerating muscles from atrophy. The drug reduces inflammatory infiltration, increases the number of muscle fibers and decreases fibrosis. β-escin reduces macrophage infiltration into injured muscles and promotes their M2 polarization. It also alters transcription of muscle regeneration-related genes: Myf5, Myh2, Myh3, Myh8, Myod1, Pax3 and Pax7, and Pcna. In C2C12 myoblasts in vitro, β-escin inhibits TNF-α-induced activation of NF-κB, reduces secretion of MMP-9 and increases ALDH activity.
    CONCLUSIONS: The data reveal beneficial role of β-escin in muscle regeneration, particularly in poorly regenerating slow-twitch muscles. The findings provide rationale for further studies on β-escin repositioning into conditions associated with muscle damage such as strenuous exercise, drug-induced myotoxicity or age-related disuse atrophy.
    Keywords:  ALDH; MMP, muscle; NF-κB; macrophage; β-escin
  17. J Biochem. 2021 Oct 22. pii: mvab116. [Epub ahead of print]
      Myostatin (Mstn) and GDF11 are critical factors that are involved in muscle atrophy in the young and sarcopenia in the elderly, respectively. These TGF-β superfamily proteins activate not only Smad signaling but also non-Smad signaling including the Ras-mediated ERK pathway (Raf-MEK-ERK phosphorylation cascade). Although Mstn and GDF11 have been shown to induce muscle atrophy or sarcopenia by Smad2/3-mediated Akt inhibition, participation of the non-Smad Ras-ERK pathway in atrophy and sarcopenia has not been well determined. We show here that both Mstn and GDF11 prevented skeletal myocyte differentiation but that the MEK inhibitor U0126 or trametinib restored differentiation in Mstn- or GDF11-treated myocytes. These MEK inhibitors induced the expression of DA-Raf1 (DA-Raf), which is a dominant-negative antagonist of the Ras-ERK pathway. Exogenous expression of DA-Raf in Mstn- or GDF11-treated myocytes restored differentiation. Furthermore, administration of trametinib to aged mice resulted in an increase in myofiber size, or recovery from muscle atrophy. The trametinib administration downregulated ERK activity in these muscles. These results imply that the Mstn/GDF11-induced Ras-ERK pathway plays critical roles in the inhibition of myocyte differentiation and muscle regeneration, which leads to muscle atrophy. Trametinib and similar approved drugs might be applicable to the treatment of muscle atrophy in sarcopenia or cachexia.
    Keywords:  Ras–ERK pathway; muscle atrophy; muscle differentiation; non-Smad signaling; sarcopenia
  18. Function (Oxf). 2021 ;2(6): zqab045
      Alzheimer's Disease (ad) associates with insulin resistance and low aerobic capacity, suggestive of impaired skeletal muscle mitochondrial function. However, this has not been directly measured in AD. This study ( n  = 50) compared muscle mitochondrial respiratory function and gene expression profiling in cognitively healthy older adults (CH; n = 24) to 26 individuals in the earliest phase of ad-related cognitive decline, mild cognitive impairment (MCI; n  = 11) or MCI taking the ad medication donepezil (MCI + med; n  = 15). Mitochondrial respiratory kinetics were measured in permeabilized muscle fibers from muscle biopsies of the vastus lateralis. Untreated MCI exhibited lower lipid-stimulated skeletal muscle mitochondrial respiration (State 3, ADP-stimulated) than both CH ( P = .043) and MCI + med (P = .007) groups. MCI also exhibited poorer mitochondrial coupling control compared to CH (P = .014). RNA sequencing of skeletal muscle revealed unique differences in mitochondrial function and metabolism genes based on both MCI status (CH vs MCI) and medication treatment (MCI vs MCI + med). MCI + med modified over 600 skeletal muscle genes compared to MCI suggesting donepezil powerfully impacts the transcriptional profile of muscle. Overall, skeletal muscle mitochondrial respiration is altered in untreated MCI but normalized in donepezil-treated MCI participants while leak control is impaired regardless of medication status. These results provide evidence that mitochondrial changes occur in the early stages of AD, but are influenced by a common ad medicine. Further study of mitochondrial bioenergetics and the influence of transcriptional regulation in early ad is warranted.
    Keywords:  Alzheimer's disease; RNA sequencing; donepezil; mild cognitive impairment; mitochondria; respiration; skeletal muscle
  19. Front Cell Dev Biol. 2021 ;9 744171
      N6-methyladenosine (m6A) RNA methylation has emerged as an important factor in various biological processes by regulating gene expression. However, the dynamic profile, function and underlying molecular mechanism of m6A modification during skeletal myogenesis remain elusive. Here, we report that members of the m6A core methyltransferase complex, METTL3 and METTL14, are downregulated during skeletal muscle development. Overexpression of either METTL3 or METTL14 dramatically blocks myotubes formation. Correspondingly, knockdown of METTL3 or METTL14 accelerates the differentiation of skeletal muscle cells. Genome-wide transcriptome analysis suggests ERK/MAPK is the downstream signaling pathway that is regulated to the greatest extent by METTL3/METTL14. Indeed, METTL3/METTL14 expression facilitates ERK/MAPK signaling. Via MeRIP-seq, we found that MNK2, a critical regulator of ERK/MAPK signaling, is m6A modified and is a direct target of METTL3/METTL14. We further revealed that YTHDF1 is a potential reader of m6A on MNK2, regulating MNK2 protein levels without affecting mRNA levels. Furthermore, we discovered that METTL3/14-MNK2 axis was up-regulated notably after acute skeletal muscle injury. Collectively, our studies revealed that the m6A writers METTL3/METTL14 and the m6A reader YTHDF1 orchestrate MNK2 expression posttranscriptionally and thus control ERK signaling, which is required for the maintenance of muscle myogenesis and may contribute to regeneration.
    Keywords:  ERK signaling; METTL3/14; MNK2; m6A; skeletal muscle differentiation and regeneration
  20. J Cell Physiol. 2021 Oct 19.
      Cold exposure is an unavoidable and severe challenge for people and animals residing in cold regions of the world, and may lead to hypothermia, drastic changes in systemic metabolism, and inhibition of protein synthesis. O-linked-N-acetylglucoseaminylation (O-GlcNAcylation) directly regulates the activity and function of target proteins involved in multiple biological processes by acting as a stress receptor and nutrient sensor. Therefore, our study aimed to examine whether O-GlcNAcylation affected myogenic IL-6 expression, regulation of energy metabolism, and promotion of survival in mouse skeletal muscle under acute cold exposure conditions. Total protein was extracted from C2C12 cells that had been cultured at 32°C for 3, 6, 9, and 12 h. Western blot analysis showed that mild hypothermia enhanced O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA) expression. Furthermore, global OGT-dependent glycosylation and interleukin-6 (IL-6) levels peaked 3 h after induction of mild hypothermia. Enhanced activation of the NF-κB pathway was also observed in response to mild hypothermia. Alloxan and Thiamet G were used to reduce and increase global OGT glycosylation levels in C2C12 cells, respectively. Increased O-GlcNAcylation was associated with significant upregulation of IL-6 expression, as well as enhanced activity and nuclear translocation of p65, while decreased O-GlcNAcylation had the opposite effect. In addition, increased O-GlcNAcylation was associated with significantly increased glucose metabolism, and OGT-mediated O-GlcNAcylation of p65. We generated skeletal muscle-specific OGT knockout mice and exposed them to cold at 4°C for 3 h per day for 1 week. OGT deficiency attenuated the O-GlcNAcylation, activity, and nuclear translocation of p65, resulting in downregulation of IL-6 in mouse skeletal muscle of mice exposed to cold conditions. Taken together, our data suggested that O-GlcNAcylation of p65 enhanced p65 activity and nuclear translocation leading to the upregulation of IL-6, which maintained energy homeostasis and promotes cell survival in mouse skeletal muscle during cold exposure.
    Keywords:  IL-6; O-GlcNAcylation; OGT; glucose metabolism; p65; skeletal muscle
  21. Biomolecules. 2021 Oct 12. pii: 1504. [Epub ahead of print]11(10):
      Duchenne muscular dystrophy (DMD) is an X-linked disease caused by mutations in DMD gene translating in lack of functional dystrophin and resulting in susceptibility of myofibers to rupture during contraction. Inflammation and fibrosis are critical hallmarks of DMD muscles, which undergo progressive degeneration leading to loss of independent ambulation in childhood and death by early adulthood. We reported that intraperitoneal injection of microencapsulated Sertoli cells (SeC) in dystrophic mice translates into recovery of muscle morphology and performance thanks to anti-inflammatory effects and induction of the dystrophin paralogue, utrophin at the muscle level, opening new avenues in the treatment of DMD. The aim of this study is to obtain information about the direct effects of SeC on myoblasts/myotubes, as a necessary step in view of a translational application of SeC-based approaches to DMD. We show that (i) SeC-derived factors stimulate cell proliferation in the early phase of differentiation in C2C12, and human healthy and DMD myoblasts; (ii) SeC delay the expression of differentiation markers in the early phase nevertheless stimulating terminal differentiation in DMD myoblasts; (iii) SeC restrain the fibrogenic potential of fibroblasts, and inhibit myoblast-myofibroblast transdifferentiation; and, (iv) SeC provide functional replacement of dystrophin in preformed DMD myotubes regardless of the mutation by inducing heregulin β1/ErbB2/ERK1/2-dependent utrophin expression. Altogether, these results show that SeC are endowed with promyogenic and antifibrotic effects on dystrophic myoblasts, further supporting their potential use in the treatment of DMD patients. Our data also suggest that SeC-based approaches might be useful in improving the early phase of muscle regeneration, during which myoblasts have to adequately proliferate to replace the damaged muscle mass.
    Keywords:  Sertoli cell; duchenne muscular dystrophy (DMD); fibrosis; heregulin β1; myoblast differentiation; myofibroblast transdifferentiation; utrophin
  22. Cells. 2021 Oct 14. pii: 2746. [Epub ahead of print]10(10):
      Advancements in reprogramming somatic cells into induced pluripotent stem cells (iPSCs) have provided a strong framework for in vitro disease modeling, gene correction and stem cell-based regenerative medicine. In cases of skeletal muscle disorders, iPSCs can be used for the generation of skeletal muscle progenitors to study disease mechanisms, or implementation for the treatment of muscle disorders. We have recently developed an improved directed differentiation method for the derivation of skeletal myogenic progenitors from hiPSCs. This method allows for a short-term (2 weeks) and efficient skeletal myogenic induction (45-65% of the cells) in human pluripotent stem cells (ESCs/iPSCs) using small molecules to induce mesoderm and subsequently myotomal progenitors, without the need for any gene integration or modification. After initial differentiation, skeletal myogenic progenitors can be purified from unwanted cells using surface markers (CD10+CD24-). These myogenic progenitors have been extensively characterized using in vitro gene expression/differentiation profiling as well as in vivo engraftment studies in dystrophic (mdx) and muscle injury (VML) rodent models and have been proven to be able to engraft and form mature myofibers as well as seeding muscle stem cells. The current protocol describes a detailed, step-by-step guide for this method and outlines important experimental details and troubleshooting points for its application in any human pluripotent stem cells.
    Keywords:  differentiation method; human iPSCs; muscle progenitors; skeletal muscle differentiation; stem cells
  23. Int J Mol Sci. 2021 Oct 11. pii: 10972. [Epub ahead of print]22(20):
      MicroRNAs are known to play a critical role in skeletal myogenesis and maintenance, and cofilin-2 (CFL2) is necessary for actin cytoskeleton dynamics and myogenic differentiation. Nonetheless, target molecules and the modes of action of miRNAs, especially those responsible for the inhibitory mechanism on the myogenesis by saturated fatty acids (SFA) or obesity, still remain unclear. Here, we reported the role played by miR-429-3p on CFL2 expression, actin filament dynamics, myoblast proliferation, and myogenic differentiation in C2C12 cells. Palmitic acid (PA), the most abundant SFA in diet, inhibited the myogenic differentiation of myoblasts, accompanied by CFL2 reduction and miR-429-3p induction. Interestingly, miR-429-3p suppressed the expression of CFL2 by targeting the 3'UTR of CFL2 mRNA directly. Transfection of miR-429-3p mimic in myoblasts increased F-actin formation and augmented nuclear YAP level, thereby promoting cell cycle progression and myoblast proliferation. Moreover, miR-429-3p mimic drastically suppressed the expressions of myogenic factors, such as MyoD, MyoG, and MyHC, and impaired myogenic differentiation of C2C12 cells. Therefore, this study unveiled the crucial role of miR-429-3p in myogenic differentiation through the suppression of CFL2 and provided implications of SFA-induced miRNA in the regulation of actin dynamics and skeletal myogenesis.
    Keywords:  CFL2; differentiation; miR-429-3p; myogenesis; proliferation
  24. Biology (Basel). 2021 Oct 18. pii: 1056. [Epub ahead of print]10(10):
      The population of satellite cells (mSCs) is highly diversified. The cells comprising it differ in their ability to regenerate their own population and differentiate, as well as in the properties they exhibit. The heterogeneity of this group of cells is evidenced by multiple differentiating markers that enable their recognition, classification, labeling, and characterization. One of the main tasks of satellite cells is skeletal muscle regeneration. Myofibers are often damaged during vigorous exercise in people who participate in sports activities. The number of satellite cells and the speed of the regeneration processes that depend on them affect the time structure of an athlete's training. This process depends on inflammatory cells. The multitude of reactions and pathways that occur during the regeneration process results in the participation and control of many factors that are activated and secreted during muscle fiber damage and at different stages of its regeneration. However, not all of them are well understood yet. This paper presents the current state of knowledge on satellite cell-dependent skeletal muscle regeneration. Studies describing the effects of various forms of exercise and age on this process were reviewed.
    Keywords:  age; exercise; inflammation; muscle regeneration; myogenic regulatory factors; satellite cells
  25. Cytokine. 2021 Oct 19. pii: S1043-4666(21)00335-5. [Epub ahead of print]149 155746
      Exploring the relationship between exercise inflammation and the peripheral neuroendocrine system is essential for understanding how acute or repetitive bouts of exercise can contribute to skeletal muscle adaption. In severe damage, some evidence demonstrates that peripheral neuroendocrine receptors might contribute to inflammatory resolution, supporting the muscle healing process through myogenesis. In this sense, the current study aimed to evaluate two classic peripheral neuronal receptors along with skeletal muscle inflammation and adaptation parameters in triceps brachii after exercise. We euthanized C57BL (10 to 12 weeks old) male mice before, and one, two, and three days after a downhill running protocol. The positive Ly6C cells, along with interleukin-6 (IL-6), nuclear factor kappa B (NF-κB), glucocorticoid receptor (GR), α7 subunits of the nicotinic acetylcholine receptor (nAChRs), and myonuclei accretion were analyzed. Our main results demonstrated that nAChRs increased with the inflammatory and myonuclei accretion responses regardless of NF-κB and GR protein expression. These results indicate that increased nAChR may contribute to skeletal muscle adaption after downhill running in mice.
    Keywords:  Downhill exercise; Eccentric exercise; Hypertrophy; Macrophage; Skeletal muscle damage
  26. J Muscle Res Cell Motil. 2021 Oct 23.
      To investigate the effects of treadmill running on two different types of skeletal muscle, we established a rat model of collagen-induced arthritis (CIA). The skeletal muscles studied were the extensor digitorum longus (EDL), which is rich in fast-twitch muscle fibers, and the soleus, which is rich in slow-twitch muscle fibers. The histological and transcriptional changes in these muscles at 14 and 44 days after immunosensitization were compared between rats that were forced to exercise (CIA ex group) and free-reared CIA rats (CIA no group). Change in protein expression was examined on day 14 after a single bout of treadmill running. Treadmill running had different effects on the relative muscle weight and total and fiber cross-sectional areas in each muscle type. In the soleus, it prevented muscle atrophy. Transcriptional analysis revealed increased eukaryotic translation initiation factor 4E (Eif4e) expression on day 14 and increased Atrogin-1 and peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α) expression on day 44 in the soleus in the CIA ex group, suggesting an interaction between muscle type and exercise. A single bout of treadmill running increased the level of Eif4e and p70S6K and decreased that of Atrogin-1 in the soleus on day 14. Treadmill running prevented muscle atrophy in the soleus in a rat model of rheumatoid arthritis via activation of mitochondrial function, as evidenced by increased PGC-1α expression.
    Keywords:  Exercise; Muscle atrophy; Rheumatoid arthritis; Soleus; Treadmill running
  27. Amino Acids. 2021 Oct 21.
      We evaluated whether insulin could stimulate β-alanine uptake by skeletal muscle cells in vitro. Mouse myoblasts (C2C12) (n = 3 wells per condition) were cultured with β-alanine (350 or 700 µmol·L-1), with insulin (100 µU·mL-1) either added to the media or not. Insulin stimulated the β-alanine uptake at the lower (350 µmol·L-1) but not higher (700 µmol·L-1) β-alanine concentration in culture medium, indicating that transporter saturation might blunt the stimulatory effects of insulin.
    Keywords:  Carnosine; Insulin; Taurine transporter; β-alanine
  28. Sci Rep. 2021 Oct 22. 11(1): 20899
      We tested the hypothesis that swim training reverses the impairment of Akt/FOXO3a signaling, ameliorating muscle atrophy in ALS mice. Transgenic male mice B6SJL-Tg (SOD1G93A) 1Gur/J were used as the ALS model (n = 35), with wild-type B6SJL (WT) mice as controls (n = 7). ALS mice were analyzed before ALS onset, at ALS onset, and at terminal ALS. Levels of insulin/Akt signaling pathway proteins were determined, and the body and tibialis anterior muscle mass and plasma creatine kinase. Significantly increased levels of FOXO3a in ALS groups (from about 13 to 21-fold) compared to WT mice were observed. MuRF1 levels in the ONSET untrained group (12.0 ± 1.7 AU) were significantly higher than in WT mice (1.12 ± 0.2 AU) and in the BEFORE ALS group (3.7 ± 0.9 AU). This was associated with body mass and skeletal muscle mass reduction. Swim training significantly ameliorated the reduction of skeletal muscle mass in both TERMINAL groups (p < 0.001) and partially reversed changes in the levels of Akt signaling pathway proteins. These findings shed light on the swimming-induced attenuation of skeletal muscle atrophy in ALS with possible practical implications for anti-cachexia approaches.
  29. J Cell Sci. 2022 Mar 01. pii: jcs258649. [Epub ahead of print]135(5):
      Myoblast fusion is essential for the formation of multinucleated muscle fibers and is promoted by transient changes in the plasma membrane lipid distribution. However, little is known about the lipid transporters regulating these dynamic changes. Here, we show that proliferating myoblasts exhibit an aminophospholipid flippase activity that is downregulated during differentiation. Deletion of the P4-ATPase flippase subunit CDC50A (also known as TMEM30A) results in loss of the aminophospholipid flippase activity and compromises actin remodeling, RAC1 GTPase membrane targeting and cell fusion. In contrast, deletion of the P4-ATPase ATP11A affects aminophospholipid uptake without having a strong impact on cell fusion. Our results demonstrate that myoblast fusion depends on CDC50A and may involve multiple CDC50A-dependent P4-ATPases that help to regulate actin remodeling.
    Keywords:  Aminophospholipid translocase; Myogenesis; P4-ATPase; Phospholipid; Skeletal myoblasts
  30. BMB Rep. 2021 Oct 22. pii: 5463. [Epub ahead of print]
      Cisplatin is widely known as an anti-cancer drug. However, the effects of cisplatin on mitochondrial function and autophagy-related proteins in the skeletal muscle are unclear. The purpose of this study was to investigate the effect of different doses of cisplatin on mito-chondrial function and autophagy-related protein levels in the skeletal muscle of rats. Eight-week-old male Wistar rats (n = 24) were assigned to one of three groups; the first group was administered a saline placebo (CON, n = 10), and the second and third groups were given 0.1 mg/kg body weight (BW) (n = 6), and 0.5 mg/kg BW (n = 8) of cisplatin, respectively. The group that had been administered 0.5 mg cisplatin exhibited a reduced BW, skeletal muscle tissue weight, and mitochondrial function and upregulated levels of autophagy-related proteins, including LC3II, Beclin 1, and BNIP3. Moreover, this group had a high LC3 II/I ratio in the skeletal muscle; i.e., the administration of a high dose of cisplatin decreased the muscle mass and mitochondrial function and increased the levels of autophagy-related proteins. These re-sults, thus, suggest that reducing mitochondrial dysfunction and autophagy pathways may be important for preventing skeletal muscle atrophy following cisplatin administration.
  31. J Cell Physiol. 2021 Nov;236(11): 7612-7624
      Muscle disuse induces atrophy through increased reactive oxygen species (ROS) released from damaged mitochondria. Mitophagy, the autophagic degradation of mitochondria, is associated with increased ROS production. However, the mitophagy activity status during disuse-induced muscle atrophy has been a subject of debate. Here, we developed a new mitophagy reporter mouse line to examine how disuse affected mitophagy activity in skeletal muscles. Mice expressing tandem mCherry-EGFP proteins on mitochondria were then used to monitor the dynamics of mitophagy activity. The reporter mice demonstrated enhanced mitophagy activity and increased ROS production in atrophic soleus muscles following a 14-day hindlimb immobilization. Results also showed an increased expression of multiple mitophagy genes, including Bnip3, Bnip3l, and Park2. Our findings thus conclude that disuse enhances mitophagy activity and ROS production in atrophic skeletal muscles and suggests that mitophagy is a potential therapeutic target for disuse-induced muscle atrophy.
    Keywords:  ROS; disuse-induced muscle atrophy; hindlimb immobilization; mitochondria; mitophagy
  32. Exp Gerontol. 2021 Oct 18. pii: S0531-5565(21)00376-4. [Epub ahead of print] 111594
      With aging comes reductions in the quality and size of skeletal muscle. These changes influence the force-generating capacity of skeletal muscle and contribute to movement deficits that accompany aging. Although declines in strength remain a significant barrier to mobility in older adults, the association between age-related changes in muscle structure and function remain unresolved. In this study, we compared age-related differences in (i) muscle volume and architecture, (ii) the quantity and distribution of intramuscular fat, and (iii) muscle shear modulus (an index of stiffness) in the triceps surae in 21 younger (24.6 ± 4.3 years) and 15 older (70.4 ± 2.4 years) healthy adults. Additionally, we explored the relationship between muscle volume, architecture, intramuscular fat and ankle plantar flexion strength in young and older adults. Magnetic resonance imaging was used to determine muscle volume and intramuscular fat content. B-mode ultrasound was used to quantify muscle architecture, shear-wave elastography was used to measure shear modulus, and ankle strength was measured during maximal isometric plantar flexion contractions. We found that older adults displayed higher levels of intramuscular fat yet similar muscle volumes in the medial (MG) and lateral gastrocnemius (LG) and soleus, compared to younger adults. These age-related higher levels of intramuscular fat were associated with lower muscle shear modulus in the LG and MG. We also found that muscle physiological cross-sectional area (PCSA) that accounted for age-associated differences in intramuscular fat showed a modest increase in its association with ankle strength compared to PCSA that did not account for fat content. This highlights that skeletal muscle fat infiltration plays a role in age-related strength deficits, but does not fully explain the age-related loss in muscle strength, suggesting that other factors play a more significant role.
    Keywords:  B-mode ultrasound; Fascicle length; Magnetic resonance imaging; Muscle stiffness; Shear wave elastography; Skeletal muscle
  33. Free Radic Biol Med. 2021 Oct 13. pii: S0891-5849(21)00751-6. [Epub ahead of print]
      Reactive oxygen species (ROS) are recognized as important signaling molecules in healthy skeletal muscle. Redox sensitive proteins can respond to intracellular changes in ROS by oxidation of reactive thiol groups on cysteine (Cys) residues. Exercise is known to induce the generation of superoxide and nitric oxide, resulting in the activation of several adaptive signaling pathways; however, it has been suggested that aging attenuates these redox-regulated adaptations to acute exercise. In the present study, we used redox proteomics to study the vastus lateralis muscles of Adult (n = 6 male, 6 female; 18-30 yrs) and Old (n = 6 male, 6 female; 64-79 yrs) adults. Participants completed a bout of high intensity cycling exercise consisting of five sets of 2-minute intervals performed at 80% maximal aerobic power output (PPO), with 2 minutes recovery cycling at 40% PPO between sets. Muscle biopsies were collected prior to exercise, and immediately following the first, second, and fifth high intensity interval. Global proteomic analysis indicated differences in abundance of a number of individual proteins between skeletal muscles of Adult and Old subjects at rest with a significant exacerbation of these differences induced by the acute exercise. In particular, we observed an exercise-induced decrease in abundance of mitochondrial proteins in muscles from older subjects only. Redox proteome analysis revealed cysteines from five cytosolic proteins in older subjects with lower oxidation (i.e. greater reduction) than was seen in muscle from the young adults at rest. Redox homeostasis was well maintained in Adult subjects following exercise, but there was significant increase in oxidation of multiple mitochondrial and cytosolic protein cysteines in Old subjects. We also observed that oxidation of peroxiredoxin 3 occurred following exercise in both Adult and Old groups, supporting the possibility that this is a key effector protein for mitochondrial redox signaling. Thus, we show, for the first time that exercise reveals a lack of resilience in muscle of older human participants, that is apparent as a loss of mitochondrial proteins and oxidation of multiple protein cysteines that are not seen in younger subjects. The precise consequences of this redox disruption are unclear, but this likely play a role in the attenuation of multiple adaptations to exercise that are classically seen with aging. Such changes were only seen following the acute stress of exercise., highlighting the need to consider not only basal differences seen during aging but also the difference following physiological challenge.
  34. J Cachexia Sarcopenia Muscle. 2021 Oct 17.
      BACKGROUND: Alzheimer's disease (AD) is an unmet medical need worldwide, and physical inactivity is a risk factor for AD. Performing physical exercise is difficult at old age, and thus, decline in physical movement may be a cause of age-associated lowering of the brain function. This study aimed to elucidate the molecular mechanism and onset of the skeletal muscle atrophy-induced acceleration of AD.METHODS: Pre-symptomatic young 5XFAD or non-transgenic wildtype mice were used. The bilateral hindlimbs were immobilized by placing them in casts for 14 days. Cognitive function was evaluated using the object recognition and spatial memory tests. Further, the hindlimb muscles were isolated for organ culture. Conditioned media (CM) of each muscle was separated by two-dimensional polyacrylamide gel electrophoresis (2D-PAGE). Protein expressions in the CM were analysed by matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry analysis. The expression levels of candidate proteins were quantified using ELISA. After continuous intracerebroventricular (i.c.v.) infusion of recombinant hemopexin, cognitive function was evaluated. Gene microarray analysis of the hippocampus was performed to investigate the molecules involved in the accelerated memory deficit. Real-time reverse transcription polymerase chain reaction and histological analysis confirmed the expression.
    RESULTS: Casting for 2 weeks reduced skeletal muscle weight. Object recognition memory in the cast-attached 5XFAD mice (n = 7, training vs. test, P = 0.3390) was impaired than that in age-matched wildtype (n = 7, training vs. test, P = 0.0523) and non-cast 5XFAD mice (n = 7, training vs. test, P = 0.0473). On 2D-PAGE, 88 spots were differentially expressed in muscle CM. The most increased spot in the cast-attached 5XFAD CM was hemopexin. Hemopexin levels in the skeletal muscle (n = 3, P = 0.0064), plasma (n = 3, P = 0.0386), and hippocampus (n = 3, P = 0.0164) were increased in cast-attached 5XFAD mice than those in non-cast 5XFAD mice. Continuous i.c.v. infusion of hemopexin for 2 weeks induced memory deficits in young 5XFAD mice (n = 4, training vs. test, P = 0.6764). Lipocalin-2 (Lcn2) messenger RNA (mRNA), neuroinflammation-associated factor, was increased in the hippocampus in hemopexin-infused 5XFAD mice than in control mice. LCN2 protein in the hippocampus was localized in the neurons, but not glial cells. Lcn2 mRNA levels in the hippocampus were also increased by cast-immobilization of the hindlimbs (n = 6, P = 0.0043).
    CONCLUSIONS: These findings provide new evidence indicating that skeletal muscle atrophy has an unbeneficial impact on the occurrence of memory impairment in young 5XFAD mice, which is mediated by the muscle secreted hemopexin.
    Keywords:  Alzheimer's disease; Dementia; Hemopexin; Memory deficit; Muscle atrophy; Neuroinflammation
  35. Biomedicines. 2021 Sep 26. pii: 1321. [Epub ahead of print]9(10):
      Currently, an increasing number of people are suffering from fatigue due to the state of their lifestyles, such as sedentary work in a relatively small space, irregular sleep patterns, or the lack of movement and exercise. The present study was designed to simulate the occurrence of fatigue in the above populations through a chronic restraint stress (CRS) model, and to reveal its dynamic processes and potential underlying molecular mechanisms. ICR mice were subjected to 8 h of restraint stress each day for 5, 10, or 15 days. It was found that the weight-loaded swimming performance, grip strength, and locomotor activity of the mice all decreased under CRS treatment, and that up to 15 days of CRS induced notable fatigue. Gastrocnemius muscle atrophy and some abnormal biochemical parameters related to fatigue under CRS were observed. Furthermore, transcriptome data showed that the changes in muscle cell metabolism and mitochondrial dysfunction were associated with the AMPK signaling pathway in CRS-treated mice. Western blotting analysis of the AMPK/PGC-1α signaling pathway revealed that CRS could decrease mitochondrial biogenesis and reduce the numbers of type I skeletal muscle fibers in the gastrocnemius of mice. CRS could also block the protective mitophagic flux to inhibit the abnormal clearance of damaged mitochondria. Our study suggests a critical link between muscle atrophy and CRS-induced fatigue in mice, suggesting that the pharmacological promotion of muscle and mitochondrial function can be used as a treatment for stress-induced fatigue.
    Keywords:  AMPK; chronic restraint stress; fatigue; mitochondrial dysfunction; mitophagy; muscle atrophy
  36. Front Pharmacol. 2021 ;12 751095
      The voltage-gated sodium channel Nav1.4 is a major actor in the excitability of skeletal myofibers, driving the muscle force in response to nerve stimulation. Supporting further this key role, mutations in SCN4A, the gene encoding the pore-forming α subunit of Nav1.4, are responsible for a clinical spectrum of human diseases ranging from muscle stiffness (sodium channel myotonia, SCM) to muscle weakness. For years, only dominantly-inherited diseases resulting from Nav1.4 gain of function (GoF) were known, i.e., non-dystrophic myotonia (delayed muscle relaxation due to myofiber hyperexcitability), paramyotonia congenita and hyperkalemic or hypokalemic periodic paralyses (episodic flaccid muscle weakness due to transient myofiber hypoexcitability). These last 5 years, SCN4A mutations inducing Nav1.4 loss of function (LoF) were identified as the cause of dominantly and recessively-inherited disorders with muscle weakness: periodic paralyses with hypokalemic attacks, congenital myasthenic syndromes and congenital myopathies. We propose to name this clinical spectrum sodium channel weakness (SCW) as the mirror of SCM. Nav1.4 LoF as a cause of permanent muscle weakness was quite unexpected as the Na+ current density in the sarcolemma is large, securing the ability to generate and propagate muscle action potentials. The properties of SCN4A LoF mutations are well documented at the channel level in cellular electrophysiological studies However, much less is known about the functional consequences of Nav1.4 LoF in skeletal myofibers with no available pertinent cell or animal models. Regarding the therapeutic issues for Nav1.4 channelopathies, former efforts were aimed at developing subtype-selective Nav channel antagonists to block myofiber hyperexcitability. Non-selective, Nav channel blockers are clinically efficient in SCM and paramyotonia congenita, whereas patient education and carbonic anhydrase inhibitors are helpful to prevent attacks in periodic paralyses. Developing therapeutic tools able to counteract Nav1.4 LoF in skeletal muscles is then a new challenge in the field of Nav channelopathies. Here, we review the current knowledge regarding Nav1.4 LoF and discuss the possible therapeutic strategies to be developed in order to improve muscle force in SCW.
    Keywords:  congenital myasthenic syndrome (CMS); congenital myopathy (CM); loss of function; skeletal muscle; sodium channel; therapeutics
  37. Medicina (Kaunas). 2021 Sep 25. pii: 1015. [Epub ahead of print]57(10):
      Muscle is one of the main targets for the biological effects of vitamin D. This hormone modulates several functions of skeletal muscles, from development to tissue repair after injury, through genomic and non-genomic mechanisms. Vitamin D deficiency and supplementation seem to significantly affect muscle strength in different populations, including athletes, although optimal serum 25(OH)D3 level for sport performance has not been defined so far. Additionally, vitamin D deficiency results in myopathy characterized by fast-twitch fiber atrophy, fatty infiltration, and fibrosis. However, less is known about regenerative effects of vitamin D supplementation after sport-related muscle injuries. Vitamin D receptor (VDR) is particularly expressed in the embryonic mesoderm during intrauterine life and in satellite cells at all stages of life for recovery of the skeletal muscle after injury. Vitamin D supplementation enhances muscle differentiation, growth, and regeneration by increasing the expression of myogenic factors in satellite cells. The objective of this narrative review is to describe the role of vitamin D in sport-related muscle injury and tissue regeneration.
    Keywords:  athletes; muscle fibers; return to sport; satellite cells; skeletal; skeletal muscle; sports; vitamin D
  38. J Orthop Res. 2021 Oct 17.
      Microenergy acoustic pulses (MAP) is a modified low-intensity extracorporeal shock wave therapy that currently used for treating musculoskeletal disorders. However, its function on muscle regeneration after ischemia-reperfusion injury (IRI) remains unknown. This study aimed to explore the effect of MAP on muscle injury after IRI and its underlying mechanisms. Ten-week-old C57BL/6J mice underwent unilateral hindlimb IRI followed with or without MAP treatment. Wet weight of tibialis anterior muscles at both injury and contralateral sides were measured followed with histology analysis at 3 weeks after IRI. In in vitro study, the myoblasts, endothelial cells and fibro-adipogenic progenitors (FAP) were treated with MAP. Cell proliferation and differentiation were assessed, and related gene expressions were measured by real-time PCR. Our results showed that MAP significantly increased the muscle weight and centrally nucleated regenerating muscle fiber size along with a trend in activating satellite cells. In vitro data indicated that MAP promoted myoblast proliferation and differentiation and endothelial cells migration. MAP also induced FAP brown/beige adipogenesis, a promyogenic phenotype of FAPs. Our findings demonstrate the beneficial function of MAP in promoting muscle regeneration after IR injury by inducing muscle stem cells proliferation and differentiation.
    Keywords:  endothelial cells; fibro-adipogenic progenitors; ischemia-reperfusion; microenergy acoustic pulses; muscle regeneration; myoblast
  39. Cells. 2021 Oct 14. pii: 2753. [Epub ahead of print]10(10):
      Terminal differentiation is an ill-defined, insufficiently characterized, nonproliferation state. Although it has been classically deemed irreversible, it is now clear that at least several terminally differentiated (TD) cell types can be brought back into the cell cycle. We are striving to uncover the molecular bases of terminal differentiation, whose fundamental understanding is a goal in itself. In addition, the field has sought to acquire the ability to make TD cells proliferate. Attaining this end would probe the very molecular mechanisms we are trying to understand. Equally important, it would be invaluable in regenerative medicine, for tissues depending on TD cells and devoid of significant self-repair capabilities. The skeletal muscle has long been used as a model system to investigate the molecular foundations of terminal differentiation. Here, we summarize more than 50 years of studies in this field.
    Keywords:  cell cycle; postmitotic state; regenerative medicine; skeletal muscle; terminal differentiation
  40. FASEB J. 2021 Nov;35(11): e21988
      Interval training (IT) results in improved fatigue resistance in skeletal muscle mainly due to an increased aerobic capacity, which involves increased muscle mitochondrial content and/or improved mitochondrial function. We hypothesized that IT with high-intensity contractions is more effective in increasing mitochondrial function, and hence fatigue resistance, than low-intensity contractions. To study this hypothesis without interference from differences in muscle fiber recruitment obliged to occur during voluntary contractions, IT was performed with in situ supramaximal electrical stimulation where all muscle fibers are recruited. We compared the effect of IT with repeated low-intensity (20 Hz stimulation, IT20) and high-intensity (100 Hz stimulation, IT100) contractions on fatigue resistance and mitochondrial content and function in mouse plantar flexor muscles. Muscles were stimulated every other day for 4 weeks. The averaged peak torque during IT bouts was 4.2-fold higher with IT100 than with IT20. Both stimulation protocols markedly improved in situ fatigue resistance, although the improvement was larger with IT100. The citrate synthase activity, a biomarker of mitochondrial content, was similarly increased with IT20 and IT100. Conversely, increased expression of mitochondrial respiratory chain (MRC) complexes I, III, and IV was only observed with IT100 and this was accompanied by increases in MRC supercomplex formation and pyruvate-malate-driven state 3 respiration in isolated mitochondria. In conclusion, the IT-induced increase in fatigue resistance is larger with high-intensity than with low-intensity contractions and this is linked to improved mitochondrial function due to increased expression of MRC complexes and assembly of MRC supercomplexes.
    Keywords:  contraction intensity; fatigue resistance; interval training; mitochondria; skeletal muscle; supercomplex
  41. Front Endocrinol (Lausanne). 2021 ;12 751488
      Hyperthyroidism triggers a glycolytic shift in skeletal muscle (SKM) by altering the expression of metabolic proteins, which is often accompanied by peripheral insulin resistance. Our previous results show that smoothelin-like protein 1 (SMTNL1), a transcriptional co-regulator, promotes insulin sensitivity in SKM. Our aim was to elucidate the role of SMTNL1 in SKM under physiological and pathological 3,3',5-Triiodo-L-thyronine (T3) concentrations. Human hyper- and euthyroid SKM biopsies were used for microarray analysis and proteome profiler arrays. Expression of genes related to energy production, nucleic acid- and lipid metabolism was changed significantly in hyperthyroid samples. The phosphorylation levels and activity of AMPKα2 and JNK were increased by 15% and 23%, respectively, in the hyperthyroid samples compared to control. Moreover, SMTNL1 expression showed a 6-fold decrease in the hyperthyroid samples and in T3-treated C2C12 cells. Physiological and supraphysiological concentrations of T3 were applied on differentiated C2C12 cells upon SMTNL1 overexpression to assess the activity and expression level of the elements of thyroid hormone signaling, insulin signaling and glucose metabolism. Our results demonstrate that SMTNL1 selectively regulated TRα expression. Overexpression of SMTNL1 induced insulin sensitivity through the inhibition of JNK activity by 40% and hampered the non-genomic effects of T3 by decreasing the activity of ERK1/2 through PKCδ. SMTNL1 overexpression reduced IRS1 Ser307 and Ser612 phosphorylation by 52% and 53%, respectively, in hyperthyroid model to restore the normal responsiveness of glucose transport to insulin. SMTNL1 regulated glucose phosphorylation and balances glycolysis and glycogen synthesis via the downregulation of hexokinase II by 1.3-fold. Additionally, mitochondrial respiration and glycolysis were measured by SeaHorse analysis to determine cellular metabolic function/phenotype of our model system in real-time. T3 overload strongly increased the rate of acidification and a shift to glycolysis, while SMTNL1 overexpression antagonizes the T3 effects. These lines of evidence suggest that SMTNL1 potentially prevents hyperthyroidism-induced changes in SKM, and it holds great promise as a novel therapeutic target in insulin resistance.
    Keywords:  glucose metabolism; hyperthyroidism; insulin receptor substrate 1; insulin sensitivity; insulin signaling; phosphorylation; skeletal muscle
  42. Eur J Pharmacol. 2021 Oct 14. pii: S0014-2999(21)00724-X. [Epub ahead of print] 174568
      Muscular dystrophy is a well-known genetically heterogeneous group of rare muscle disorders. This progressive disease causes the breakdown of skeletal muscles over time and leads to grave weakness. This breakdown is caused by a diverse pattern of mutations in dystrophin and dystrophin associated protein complex. These mutations lead to the production of altered proteins in response to which, the body stimulates production of various cytokines and immune cells, particularly reactive oxygen species and NFκB. Immune cells display/exhibit a dual role by inducing muscle damage and muscle repair. Various anti-oxidants, anti-inflammatory and glucocorticoid drugs serve as potent therapeutics for muscular dystrophy. Along with the above mentioned therapeutics, induced pluripotent stem cells also serve as a novel approach paving a way for personalized treatment. These pluripotent stem cells allow regeneration of large numbers of regenerative myogenic progenitors that can be administered in muscular dystrophy patients which assist in the recovery of lost muscle fibers. In this review, we have summarized gene-editing, immunological and induced pluripotent stem cell based therapeutics for muscular dystrophy treatment.
    Keywords:  Muscular dystrophy; Mutation; Myogenic progenitor; NFκB; iPSC
  43. Int J Mol Sci. 2021 Oct 14. pii: 11080. [Epub ahead of print]22(20):
      Mitochondrial DNA deletions affect energy metabolism at tissue-specific and cell-specific threshold levels, but the pathophysiological mechanisms determining cell fate remain poorly understood. Chronic progressive external ophthalmoplegia (CPEO) is caused by mtDNA deletions and characterized by a mosaic distribution of muscle fibers with defective cytochrome oxidase (COX) activity, interspersed among fibers with retained functional respiratory chain. We used diagnostic histochemistry to distinguish COX-negative from COX-positive fibers in nine muscle biopsies from CPEO patients and performed laser capture microdissection (LCM) coupled to genome-wide gene expression analysis. To gain molecular insight into the pathogenesis, we applied network and pathway analysis to highlight molecular differences of the COX-positive and COX-negative fiber transcriptome. We then integrated our results with proteomics data that we previously obtained comparing COX-positive and COX-negative fiber sections from three other patients. By virtue of the combination of LCM and a multi-omics approach, we here provide a comprehensive resource to tackle the pathogenic changes leading to progressive respiratory chain deficiency and disease in mitochondrial deletion syndromes. Our data show that COX-negative fibers upregulate transcripts involved in translational elongation and protein synthesis. Furthermore, based on functional annotation analysis, we find that mitochondrial transcripts are the most enriched among those with significantly different expression between COX-positive and COX-negative fibers, indicating that our unbiased large-scale approach resolves the core of the pathogenic changes. Further enrichments include transcripts encoding LIM domain proteins, ubiquitin ligases, proteins involved in RNA turnover, and, interestingly, cell cycle arrest and cell death. These pathways may thus have a functional association to the molecular pathogenesis of the disease. Overall, the transcriptome and proteome show a low degree of correlation in CPEO patients, suggesting a relevant contribution of post-transcriptional mechanisms in shaping this disease phenotype.
    Keywords:  disease models; mtDNA deletions; myopathy; proteomics; skeletal muscle; transcriptomics
  44. Biomolecules. 2021 Oct 15. pii: 1519. [Epub ahead of print]11(10):
      Duchenne muscular dystrophy (DMD), caused by the loss of dystrophin, remains incurable. Reduction in muscle regeneration with DMD is associated with the accumulation of fibroadipogenic progenitors (FAPs) differentiating into myofibroblasts and leading to a buildup of the collagenous tissue aggravating DMD pathogenesis. Mesenchymal stromal cells (MSCs) expressing platelet-derived growth factor receptors (PDGFRs) are activated in muscle during DMD progression and give rise to FAPs promoting DMD progression. Here, we hypothesized that muscle dysfunction in DMD could be delayed via genetic or pharmacologic depletion of MSC-derived FAPs. In this paper, we test this hypothesis in dystrophin-deficient mdx mice. To reduce fibro/adipose infiltration and potentiate muscle progenitor cells (MPCs), we used a model for inducible genetic ablation of proliferating MSCs via a suicide transgene, viral thymidine kinase (TK), expressed under the Pdgfrb promoter. We also tested if MSCs from fat tissue, the adipose stromal cells (ASCs), contribute to FAPs and could be targeted in DMD. Pharmacological ablation was performed with a hunter-killer peptide D-CAN targeting ASCs. MSC depletion with these approaches resulted in increased endurance, measured based on treadmill running, as well as grip strength, without significantly affecting fibrosis. Although more research is needed, our results suggest that depletion of pathogenic MSCs mitigates muscle damage and delays the loss of muscle function in mouse models of DMD.
    Keywords:  adipose mesenchymal stromal cell; duchenne muscular dystrophy; platelet-derived growth factor receptor
  45. Mol Nutr Food Res. 2021 Oct 22. e2100526
      SCOPE: This study investigated the combined effect of the angiotensin II (AT-II) receptor blocker losartan and branched-chain amino acids (BCAAs) on skeletal muscle atrophy in rats with cirrhosis and steatohepatitis.METHOD AND RESULTS: Fischer 344 rats were fed a choline-deficient l-amino acid-defined (CDAA) diet for 12 weeks and treated with oral losartan (30 mg/kg/day) and/or BCAAs (Aminoleban® EN, 2500 mg/kg/day). Treatment with losartan and BCAAs attenuated hepatic inflammation and fibrosis and improved skeletal muscle atrophy and strength in CDAA-fed rats. Both agents reduced intramuscular myostatin and pro-inflammatory cytokine levels, resulting in inhibition of the ubiquitin-proteasome system (UPS) through interference with the SMAD and nuclear factor-kappa B pathways, respectively. Losartan also augmented the BCAA-mediated increase of skeletal muscle mass by promoting insulin growth factor-I production and mitochondrial biogenesis. Moreover, losartan decreased the intramuscular expression of transcription factor EB (TFEB), a transcriptional inducer of E3 ubiquitin ligase regulated by AT-II. In vitro assays illustrated that losartan promoted mitochondrial biogenesis and reduced TFEB expression in AT-II-stimulated rat myocytes, thereby potentiating the inhibitory effects of BCAAs on the UPS and caspase-3 cleavage.
    CONCLUSION: These results indicate that this regimen could serve as a novel treatment for patients with sarcopenia and liver cirrhosis. This article is protected by copyright. All rights reserved.
    Keywords:  Angiotensin-II; BCAA; Liver cirrhosis; Muscle atrophy; Myokine
  46. Cells. 2021 Sep 27. pii: 2556. [Epub ahead of print]10(10):
      Duchenne muscular dystrophy (DMD) is a genetic disorder that results from deficiency of the dystrophin protein. In recent years, DMD pathological models have been created using induced pluripotent stem (iPS) cells derived from DMD patients. In addition, gene therapy using CRISPR-Cas9 technology to repair the dystrophin gene has been proposed as a new treatment method for DMD. However, it is not known whether the contractile function of myotubes derived from gene-repaired iPS cells can be restored. We therefore investigated the maturation of myotubes in electrical pulse stimulation culture and examined the effect of gene repair by observing the contractile behaviour of myotubes. The contraction activity of myotubes derived from dystrophin-gene repaired iPS cells was improved by electrical pulse stimulation culture. The iPS cell method used in this study for evaluating muscle contractile activity is a useful technique for analysing the mechanism of hereditary muscular disease pathogenesis and for evaluating the efficacy of new drugs and gene therapy.
    Keywords:  CRISPR/Cas9; Duchenne muscular dystrophy; contractile activity; human induced pluripotent stem cell; myotube
  47. Cell Metab. 2021 Oct 19. pii: S1550-4131(21)00479-4. [Epub ahead of print]
      Type 2 diabetes mellitus (T2DM) is associated with impaired skeletal muscle function and degeneration of the skeletal muscles. However, the mechanisms underlying the degeneration are not well described in human skeletal muscle. Here we show that skeletal muscle of T2DM patients exhibit degenerative remodeling of the extracellular matrix that is associated with a selective increase of a subpopulation of fibro-adipogenic progenitors (FAPs) marked by expression of THY1 (CD90)-the FAPCD90+. We identify platelet-derived growth factor (PDGF) as a key FAP regulator, as it promotes proliferation and collagen production at the expense of adipogenesis. FAPsCD90+ display a PDGF-mimetic phenotype, with high proliferative activity, clonogenicity, and production of extracellular matrix. FAPCD90+ proliferation was reduced by in vitro treatment with metformin. Furthermore, metformin treatment reduced FAP content in T2DM patients. These data identify a PDGF-driven conversion of a subpopulation of FAPs as a key event in the fibrosis development in T2DM muscle.
    Keywords:  adipocytes; extracellular matrix; fatty degeneration; fibro-adipogenic progenitors; fibroblast; fibrosis; mesenchymal stem cells; skeletal muscle; type 2 diabetes
  48. J Gen Physiol. 2021 11 01. pii: e202112896. [Epub ahead of print]153(11):
      The myosin motors in resting skeletal muscle are folded back against their tails in the thick filament in a conformation that makes them unavailable for binding to actin. When muscles are activated, calcium binding to troponin leads to a rapid change in the structure of the actin-containing thin filaments that uncovers the myosin binding sites on actin. Almost as quickly, myosin motors leave the folded state and move away from the surface of the thick filament. To test whether motor unfolding is triggered by the availability of nearby actin binding sites, we measured changes in the x-ray reflections that report motor conformation when muscles are activated at longer sarcomere length, so that part of the thick filaments no longer overlaps with thin filaments. We found that the intensity of the M3 reflection from the axial repeat of the motors along the thick filaments declines almost linearly with increasing sarcomere length up to 2.8 µm, as expected if motors in the nonoverlap zone had left the folded state and become relatively disordered. In a recent article in JGP, Squire and Knupp challenged this interpretation of the data. We show here that their analysis is based on an incorrect assumption about how the interference subpeaks of the M3 reflection were reported in our previous paper. We extend previous models of mass distribution along the filaments to show that the sarcomere length dependence of the M3 reflection is consistent with <10% of no-overlap motors remaining in the folded conformation during active contraction, confirming our previous conclusion that unfolding of myosin motors on muscle activation is not due to the availability of local actin binding sites.
  49. Sci Rep. 2021 Oct 22. 11(1): 20880
      Cell adhesion molecule-related/downregulated by oncogenes (Cdon) is a cell-surface receptor that mediates cell-cell interactions and positively regulates myogenesis. The cytoplasmic region of Cdon interacts with other proteins to form a Cdon/JLP/Bnip-2/CDC42 complex that activates p38 mitogen-activated protein kinase (MAPK) and induces myogenesis. However, Cdon complex may include other proteins during myogenesis. In this study, we found that Cullin 2-interacting protein zinc finger SWIM type containing 8 (ZSWIM8) ubiquitin ligase is induced during C2C12 differentiation and is included in the Cdon complex. We knocked-down Zswim8 in C2C12 cells to determine the effect of ZSWIM8 on differentiation. However, we detected neither ZSWIM8-dependent ubiquitination nor the degradation of Bnip2, Cdon, or JLP. In contrast, ZSWIM8 knockdown accelerated C2C12 differentiation. These results suggest that ZSWIM8 is a Cdon complex-included myogenic protein that prevents C2C12 differentiation without affecting the stability of Bnip2, Cdon, and JLP.
  50. Antioxidants (Basel). 2021 Sep 25. pii: 1522. [Epub ahead of print]10(10):
      Respiratory failure from progressive respiratory muscle weakness is the most common cause of death in amyotrophic lateral sclerosis (ALS). Defects in neuromuscular junctions (NMJs) and progressive NMJ loss occur at early stages, thus stabilizing and preserving NMJs represents a potential therapeutic strategy to slow ALS disease progression. Here we demonstrate that NMJ damage is repaired by MG53, an intrinsic muscle protein involved in plasma membrane repair. Compromised diaphragm muscle membrane repair and NMJ integrity are early pathological events in ALS. Diaphragm muscles from ALS mouse models show increased susceptibility to injury and intracellular MG53 aggregation, which is also a hallmark of human muscle samples from ALS patients. We show that systemic administration of recombinant human MG53 protein in ALS mice protects against injury to diaphragm muscle, preserves NMJ integrity, and slows ALS disease progression. As MG53 is present in circulation in rodents and humans under physiological conditions, our findings provide proof-of-concept data supporting MG53 as a potentially safe and effective therapy to mitigate ALS progression.
    Keywords:  MG53; amyotrophic lateral sclerosis; diaphragm; neuromuscular junction; sarcolemma damage
  51. Wiley Interdiscip Rev RNA. 2021 Oct 19. e1700
      Naturally occurring post-transcriptional chemical modifications serve critical roles in impacting RNA structure and function. More directly, modifications may affect RNA stability, intracellular transport, translational efficiency, and fidelity. The combination of effects caused by modifications are ultimately linked to gene expression regulation at a genome-wide scale. The latter is especially true in systems that undergo rapid metabolic and or translational remodeling in response to external stimuli, such as the presence of stressors, but beyond that, modifications may also affect cell homeostasis. Although examples of the importance of RNA modifications in translation are accumulating rapidly, still what these contribute to the function of complex physiological systems such as muscle is only recently emerging. In the present review, we will introduce key information on various modifications and highlight connections between those and cellular malfunctions. In passing, we will describe well-documented roles for modifications in the nervous system and use this information as a stepping stone to emphasize a glaring paucity of knowledge on the role of RNA modifications in heart and skeletal muscle, with particular emphasis on mitochondrial function in those systems. This article is categorized under: RNA in Disease and Development > RNA in Disease RNA Processing > RNA Editing and Modification.
    Keywords:  mRNA; mitochondria; modifications; muscle; rRNA; tRNA; translation
  52. Front Pharmacol. 2021 ;12 698714
      Background: p38 regulated/activated protein kinase (PRAK) plays a crucial role in modulating cell death and survival. However, the role of PRAK in the regulation of metabolic stress remains unclear. We examined the effects of PRAK on cell survival and mitochondrial function in C2C12 myoblasts in response to high glucose stresses. Methods: PRAK of C2C12 myoblasts was knocked out by using CRISPR/Cas-9 genome editing technology. Both wild type and PRAK-/- C2C12 cells were exposed to high glucose at the concentration of 30 mmol/L to induce metabolic stress. The effect of irisin, an adipomyokine, on both wild type and PRAK-/- cells was determined to explore its relationship with RPAK. Cell viability, ATP product, glucose uptake, mitochondrial damage, and insulin signaling were assessed. Results: PRAK knockout decreased C2C12 viability in response to high glucose stress as evident by MTT assay in association with the reduction of ATP and glucose uptake. PRAK knockout enhanced apoptosis of C2C12 myoblasts in response to high glucose, consistent with an impairment in mitochondrial function, by decreasing mitochondrial membrane potential. PRAK knockout induced impairment of mitochondrial and cell damage were rescued by irisin. PRAK knockout caused decrease in phosphorylated PI3 kinase at Tyr 485, IRS-1 and AMPKα and but did not affect non-phosphorylated PI3 kinase, IRS-1 and AMPKα signaling. High glucose caused the further reduction of phosphorylated PI3 kinase, IRS-1 and AMPKα. Irisin treatment preserved phosphorylated PI3 kinase, IRS-1by rescuing PRAK in high glucose treatment. Conclusion: Our finding indicates a pivotal role of PRAK in preserving cellular survival, mitochondrial function, and high glucose stress.
    Keywords:  C2C12 myoblasts; high glucose; insulin signaling; metabolic stress; mitochondria
  53. Cells. 2021 Oct 02. pii: 2639. [Epub ahead of print]10(10):
      Exercise itself is fundamental for good health, and when practiced regularly confers a myriad of metabolic benefits in a range of tissues. These benefits are mediated by a range of adaptive responses in a coordinated, multi-organ manner. The continued understanding of the molecular mechanisms of action which confer beneficial effects of exercise on the body will identify more specific pathways which can be manipulated by therapeutic intervention in order to prevent or treat various metabolism-associated diseases. This is particularly important as exercise is not an available option to all and so novel methods must be identified to confer the beneficial effects of exercise in a therapeutic manner. This review will focus on key emerging molecular mechanisms of mitochondrial biogenesis, autophagy and mitophagy in selected, highly metabolic tissues, describing their regulation and contribution to beneficial adaptations to exercise.
    Keywords:  autophagy; exercise; mitophagy and mitochondrial biogenesis; molecular signaling
  54. Cells. 2021 Sep 23. pii: 2512. [Epub ahead of print]10(10):
      The ability of the ribonucleic acid (RNA) to self-replicate, combined with a unique cocktail of chemical properties, suggested the existence of an RNA world at the origin of life. Nowadays, this hypothesis is supported by innovative high-throughput and biochemical approaches, which definitively revealed the essential contribution of RNA-mediated mechanisms to the regulation of fundamental processes of life. With the recent development of SARS-CoV-2 mRNA-based vaccines, the potential of RNA as a therapeutic tool has received public attention. Due to its intrinsic single-stranded nature and the ease with which it is synthesized in vitro, RNA indeed represents the most suitable tool for the development of drugs encompassing every type of human pathology. The maximum effectiveness and biochemical versatility is achieved in the guise of non-coding RNAs (ncRNAs), which are emerging as multifaceted regulators of tissue specification and homeostasis. Here, we report examples of coding and ncRNAs involved in muscle regeneration and discuss their potential as therapeutic tools. Small ncRNAs, such as miRNA and siRNA, have been successfully applied in the treatment of several diseases. The use of longer molecules, such as lncRNA and circRNA, is less advanced. However, based on the peculiar properties discussed below, they represent an innovative pool of RNA biomarkers and possible targets of clinical value.
    Keywords:  RNA therapeutics; cardiac regeneration; noncoding RNAs (ncRNAs); skeletal muscle regeneration