bims-moremu Biomed News
on Molecular regulators of muscle mass
Issue of 2023–07–02
28 papers selected by
Anna Vainshtein, Craft Science Inc.



  1. Biochem Biophys Res Commun. 2023 Jun 16. pii: S0006-291X(23)00809-4. [Epub ahead of print]672 177-184
      Extracellular vesicles (EVs) released into the blood during exercise mediate its whole-body health effects. The differentiation of EVs released by skeletal muscle cells in vivo from those released by other cells is challenging, therefore, it is unclear whether exercise increases the number of EVs secreted by skeletal muscle cells. In this study, we investigated whether exercise affects the quantity of EVs released from skeletal muscle cells using in vitro exercise models. C2C12 myotubes were cultured on a gel layer with 1 or 30 Hz electrical pulse stimulation (EPS) to induce contractions as an artificial simulating exercise. We found that tetanic contraction induced by 30 Hz EPS increased the number of secreted EVs. MicroRNA (miRNA)-seq analysis revealed that 30 Hz EPS altered the miRNA in the secreted EVs. Furthermore, expression analysis of genes related to the biogenesis and transport of EVs revealed that the expression of ALG-2 interacting protein X (Alix) was increased in response to 30 Hz EPS, and the peak value of intracellular Ca2+ in myotubes at 30 Hz EPS was higher than that at 1 Hz, indicating that the increase in intracellular Ca2+ concentration may be related to the increased secretion of EVs in response to 30 Hz EPS.
    Keywords:  Electrical pulse stimulation; Extracellular vesicles; Skeletal muscle cells; miRNA
    DOI:  https://doi.org/10.1016/j.bbrc.2023.06.054
  2. Physiol Rev. 2023 Jun 29.
      Mechanisms underlying mechanical overload-induced skeletal muscle hypertrophy have been extensively researched since the landmark report by Morpurgo (1897) of "work-induced hypertrophy" in dogs that were treadmill-trained. Much of the pre-clinical rodent and human resistance training research to date supports that involved mechanisms include enhanced mammalian/mechanistic target of rapamycin complex 1 (mTORC1) signaling, an expansion in translational capacity through ribosome biogenesis, increased satellite cell abundance and myonuclear accretion, and post-exercise elevations in muscle protein synthesis rates. However, several lines of past and emerging evidence suggest additional mechanisms that feed into or are independent of these processes are also involved. This review will first provide a historical account as to how mechanistic research into skeletal muscle hypertrophy has progressed. A comprehensive list of mechanisms associated with skeletal muscle hypertrophy is then outlined and areas of disagreement involving these mechanisms are presented. Finally, future research directions involving many of the discussed mechanisms will be proposed.
    Keywords:  hypertrophy; myofiber; resistance training; skeletal muscle
    DOI:  https://doi.org/10.1152/physrev.00039.2022
  3. Mol Metab. 2023 Jun 26. pii: S2212-8778(23)00095-9. [Epub ahead of print] 101761
       OBJECTIVE: The AMP-activated protein kinase (AMPK) gets activated in response to energetic stress such as contractions and plays a vital role in regulating various metabolic processes such as insulin-independent glucose uptake in skeletal muscle. The main upstream kinase that activates AMPK through phosphorylation of α-AMPK Thr172 in skeletal muscle is LKB1, however some studies have suggested that Ca2+/calmodulin-dependent protein kinase kinase 2 (CaMKK2) acts as an alternative kinase to activate AMPK. We aimed to establish whether CaMKK2 is involved in activation of AMPK and promotion of glucose uptake following contractions in skeletal muscle.
    METHODS: A recently developed CaMKK2 inhibitor (SGC-CAMKK2-1) alongside a structurally related but inactive compound (SGC-CAMKK2-1N), as well as CaMKK2 knock-out (KO) mice were used. In vitro kinase inhibition selectivity and efficacy assays, as well as cellular inhibition efficacy analyses of CaMKK inhibitors (STO-609 and SGC-CAMKK2-1) were performed. Phosphorylation and activity of AMPK following contractions (ex vivo) in mouse skeletal muscles treated with/without CaMKK inhibitors or isolated from wild-type (WT)/CaMKK2 KO mice were assessed. Camkk2 mRNA in mouse tissues was measured by qPCR. CaMKK2 protein expression was assessed by immunoblotting with or without prior enrichment of calmodulin-binding proteins from skeletal muscle extracts, as well as by mass spectrometry-based proteomics of mouse skeletal muscle and C2C12 myotubes.
    RESULTS: STO-609 and SGC-CAMKK2-1 were equally potent and effective in inhibiting CaMKK2 in cell-free and cell-based assays, but SGC-CAMKK2-1 was much more selective. Contraction-stimulated phosphorylation and activation of AMPK were not affected with CaMKK inhibitors or in CaMKK2 null muscles. Contraction-stimulated glucose uptake was comparable between WT and CaMKK2 KO muscle. Both CaMKK inhibitors (STO-609 and SGC-CAMKK2-1) and the inactive compound (SGC-CAMKK2-1N) significantly inhibited contraction-stimulated glucose uptake. SGC-CAMKK2-1 also inhibited glucose uptake induced by a pharmacological AMPK activator or insulin. Relatively low levels of Camkk2 mRNA were detected in mouse skeletal muscle, but neither CaMKK2 protein nor its derived peptides were detectable in mouse skeletal muscle tissue.
    CONCLUSIONS: We demonstrate that pharmacological inhibition or genetic loss of CaMKK2 does not affect contraction-stimulated AMPK phosphorylation and activation, as well as glucose uptake in skeletal muscle. Previously observed inhibitory effect of STO-609 on AMPK activity and glucose uptake is likely due to off-target effects. CaMKK2 protein is either absent from adult murine skeletal muscle or below the detection limit of currently available methods.
    Keywords:  AMP-activated protein kinase; Ca(2+)/calmodulin dependent protein kinase kinase 2; SGC-CAMKK2-1; STO-609; glucose uptake
    DOI:  https://doi.org/10.1016/j.molmet.2023.101761
  4. FASEB J. 2023 Aug;37(8): e23050
      Gene expression of the NR4A nuclear orphan receptor NOR-1 is reduced in obesity and in human skeletal muscle during disuse. It has been well established that NOR-1 is highly responsive to both aerobic and resistance exercise and NOR-1 overexpression is coincident with a plethora of metabolic benefits. However, it is unclear whether loss of NOR-1 contributes to inappropriate metabolic signaling in skeletal muscle that could lead to insulin resistance. The purpose of this study was to elucidate the impact of NOR-1 deficiency on C2C12 metabolic signaling. Changes in gene expression after siRNA-mediated NOR-1 knockdown in C2C12 myotubes were determined by qPCR and bioinformatic analysis of RNA-Seq data. Our RNA-Seq data identified several metabolic targets regulated by NOR-1 and implicates NOR-1 as a modulator of mTORC1 signaling via Akt-independent mechanisms. Furthermore, pathway analysis revealed NOR-1 knockdown perturbs the insulin resistance and insulin sensitivity pathways. Taken together, these data suggest skeletal muscle NOR-1 deficiency may contribute to altered metabolic signaling that is consistent with metabolic disease. We postulate that strategies that improve NOR-1 may be important to offset the negative impact that inactivity, obesity, and type 2 diabetes have on mitochondria and muscle metabolism.
    Keywords:  inactivity; metabolism; mitochondria; muscle; myoblast; obesity
    DOI:  https://doi.org/10.1096/fj.202202029R
  5. Development. 2023 Jun 27. pii: dev.201509. [Epub ahead of print]
      The earliest skeletal muscle progenitor cells (SMPCs) derived from human pluripotent stem cells (hPSCs) are often identified by factors expressed by a diverse number of progenitors. An early transcriptional checkpoint that defines myogenic commitment could improve hPSC differentiation to skeletal muscle. Analysis of several myogenic factors in human embryos and early hPSC differentiations found SIX1+PAX3+ co-expression was most indictive of myogenesis. Using dCas9-KRAB hPSCs, we demonstrate early inhibition of SIX1 alone significantly decreased PAX3 expression, reduced PAX7+ SMPCs, and myotubes later in differentiation. Emergence of SIX1+PAX3+ precursors can be improved by manipulating seeding density, monitoring metabolic secretion, and altering the concentration of CHIR99021. These modifications resulted in the co-emergence of hPSC-derived sclerotome, cardiac, and neural crest that we hypothesized enhanced hPSC myogenic differentiation. Inhibition of non-myogenic lineages modulated PAX3 independent of SIX1. To better understand SIX1 expression, we compared directed differentiations to fetal progenitors and adult satellite cells by RNA-seq. While SIX1 continued to be expressed across human development, SIX1 co-factor expression was dependent on developmental timing. We provide a resource to enable efficient derivation of skeletal muscle from hPSCs.
    Keywords:  Development; Directed Differentiation; Myogenesis; Pluripotent Stem Cells; SIX1; Skeletal Muscle
    DOI:  https://doi.org/10.1242/dev.201509
  6. J Sport Health Sci. 2023 Jun 27. pii: S2095-2546(23)00060-1. [Epub ahead of print]
       OBJECTIVE: The aim of this umbrella review was to determine the impact of resistance training (RT) and individual RT prescription variables on muscle mass, strength, and physical function in healthy adults.
    METHODS: Following Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines, we systematically searched and screened eligible systematic reviews reporting the effects of differing RT prescription variables on muscle mass (or its proxies), strength, and/or physical function in healthy adults aged >18 years.
    RESULTS: We identified 44 systematic reviews that met our inclusion criteria. The methodological quality of these reviews was assessed using A Measurement Tool to Assess Systematic Reviews (AMSTAR); standardized effectiveness statements were generated. We found that RT was consistently a potent stimulus for increasing skeletal muscle mass (4/4 reviews provide some or sufficient evidence), strength (4/6 reviews provided some or sufficient evidence), and physical function (1/1 review provided some evidence). RT load (6/8 reviews provided some or sufficient evidence), weekly frequency (2/4 reviews provided some or sufficient evidence), volume (3/7 reviews provided some or sufficient evidence), and exercise order (1/1 review provided some evidence) impacted RT-induced increases in muscular strength. We discovered that 2/3 reviews provided some or sufficient evidence that RT volume and contraction velocity influenced skeletal muscle mass while 4/7 reviews provided insufficient evidence in favor of RT load impacting skeletal muscle mass. There was insufficient evidence to conclude that time of day, periodization, inter-set rest, set configuration, set end point, contraction velocity/time under tension, or exercise order (only pertaining to hypertrophy) influenced skeletal muscle adaptations. A paucity of data limited insights into the impact of RT prescription variables on physical function.
    CONCLUSION: Overall, RT increased muscle mass, strength, and physical function compared to no exercise. RT intensity (load) and weekly frequency impacted RT-induced increases in muscular strength but not muscle hypertrophy. RT volume (number of sets) influenced muscular strength and hypertrophy.
    Keywords:  Resistance training; Resistance training prescription variables hypertrophy; Strength; Umbrella review
    DOI:  https://doi.org/10.1016/j.jshs.2023.06.005
  7. Front Aging. 2023 ;4 1175510
      The voltage-gated potassium channels (Kv) are complex ion channels with distinct roles in neurotransmission, electrical conductivity of the heart, and smooth and striated muscle functions. Previously, we demonstrated that deletion of Kvβ2 in mice results in decreased Pax7 protein levels, hindlimb muscles and body weights, and fiber type switching. In the present study, we tested the hypothesis that Kvβ2 regulates skeletal muscle function in mice. The young and old Kvβ2 knockout (KO) and wildtype (WT) mice were utilized to test the aging phenotype and skeletal muscle function. Consistent with our previous finding, we found a significant reduction in hindlimb skeletal muscles mass and body weight in young Kvβ2 KO mice, which was also significantly reduced in old Kvβ2 KO mice compared with age-matched WT mice. Forelimb grip strength, and the hindleg extensor digitorum longus (EDL) muscles force-frequency relations were significantly decreased in young and old Kvβ2 KO mice compared to age-matched WT mice. Analysis of transmission electron microscopy images of EDL muscles in young mice revealed a significant reduction in the sarcomere length for Kvβ2 KO vs. WT. Hematoxylin and eosin-stained tibialis anterior muscles cryosections displayed a significant decrease in the number of medium (2,000-4,000 µm2) and largest (>4,000 µm2) myofibers area in young Kvβ2 KO vs. WT mice. We also found a significant increase in fibrotic tissue area in young Kvβ2 KO mice compared with age-matched WT mice. Analysis of RNA Seq data of the gastrocnemius muscles (GAS) identified significant increase in genes involved in skeletal muscle development, proliferation and cell fate determination, atrophy, energy metabolism, muscle plasticity, inflammation, and a decrease in circadian core clock genes in young Kvβ2 KO vs. WT mice. Several genes were significantly upregulated (384 genes) and downregulated (40 genes) in young Kvβ2 KO mice compared to age-matched WT mice. Further, RT-qPCR analysis of the GAS muscles displayed a significant increase in pro-inflammatory marker Il6 expression in young Kvβ2 KO mice compared to age-matched WT mice. Overall, the present study shows that deletion of Kvβ2 leads to decreased muscles strength and increased inflammation.
    Keywords:  AKR6; KCNAB2; Kvβ2; inflammation; sarcopenia; skeletal muscle
    DOI:  https://doi.org/10.3389/fragi.2023.1175510
  8. Front Physiol. 2023 ;14 1198390
      Inactivity causes insulin resistance in skeletal muscle and exacerbates various lifestyle-related diseases. We previously found that 24-h hindlimb cast immobilization (HCI) of the predominantly slow-twitch soleus muscle increased intramyocellular diacylglycerol (IMDG) and insulin resistance by activation of lipin1, and HCI after a high-fat diet (HFD) further aggravated insulin resistance. Here, we investigated the effects of HCI on the fast-twitch-predominant plantaris muscle. HCI reduced the insulin sensitivity of plantaris muscle by approximately 30%, and HCI following HFD dramatically reduced insulin sensitivity by approximately 70% without significant changes in the amount of IMDG. Insulin-stimulated phosphorylation levels of insulin receptor (IR), IR substrate-1, and Akt were reduced in parallel with the decrease in insulin sensitivity. Furthermore, tyrosine phosphatase 1B (PTP1B), a protein known to inhibit insulin action by dephosphorylating IR, was activated, and PTP1B inhibition canceled HCI-induced insulin resistance. In conclusion, HCI causes insulin resistance in the fast-twitch-predominant plantaris muscle as well as in the slow-twitch-predominant soleus muscle, and HFD potentiates these effects in both muscle types. However, the mechanism differed between soleus and plantaris muscles, since insulin resistance was mediated by the PTP1B inhibition at IR in plantaris muscle.
    Keywords:  PTP1B; high fat diet; insulin resistance; physical inactivity; skeletal muscle
    DOI:  https://doi.org/10.3389/fphys.2023.1198390
  9. J Mol Histol. 2023 Jun 26.
      Skeletal muscle (SkM) comprises slow and fast-twitch fibers, which differ in molecular composition, function, and systemic energy consumption. In addition, muscular dystrophies (DM), a group of diverse hereditary diseases, present different patterns of muscle involvement, progression, and severity, suggesting that the regeneration-degeneration process may differ depending on the muscle type. Therefore, the study aimed to explore the expression of proteins involved in the repair process in different muscles at an early stage of muscular dystrophy in the δ-sarcoglycan null mice (Sgcd-null), a limb-girdle muscular dystrophy 2 F model. Hematoxylin & Eosin (H&E) Staining showed a high number of central nuclei in soleus (Sol), tibialis (Ta), gastrocnemius (Gas), and extensor digitorum longus (Edl) from four months Sgcd-null mice. However, fibrosis, determined by trichrome of Gomori modified staining, was only observed in Sgcd-null Sol. In addition, the number of Type I and II fibers variated differentially in the Sgcd-null muscles vs. wild-type muscles. Besides, the protein expression level of β-catenin, myomaker, MyoD, and myogenin also presented different expression levels in all the Sgcd-null muscles studied. In summary, our study reveals that muscles with different metabolic characteristics showed distinct expression patterns of proteins involved in the muscle regeneration process. These results could be relevant in designing therapies for genetic and acquired myopathy.
    Keywords:  Delta sarcoglycan; Muscular dystrophy; Myogenic proteins; Skeletal muscle
    DOI:  https://doi.org/10.1007/s10735-023-10136-7
  10. Kobe J Med Sci. 2023 Jun 22. 69(2): E40-E48
      Skeletal muscle is a tissue that contains abundant water. However, by aging a decrease in muscle water content is induced in skeletal muscles, which is one of major age-related alterations in skeletal muscles as common as muscle atrophy. Selective water channel aquaporin 4 (AQP4) is one of major water transport networks in the skeletal muscles. However, the effects of aging on water transport via AQP4 in skeletal muscles remain unclear. Thus, the current study investigated the change of the expression level of AQP4 in the aged skeletal muscles. Eight-week-old (the young group) and 2-year-old (the old group) female Fischer 344 rats were used in this study (n = 6/group). In skeletal muscles of each group, the expression levels of some target proteins were quantified by Western blot analysis. As a result, the relative muscle weight in the old group was significantly decreased, compared with that in the young group (p < 0.05). The decline in the muscle water content was accompanied by the decrease in expression of AQP4 in the aged skeletal muscles (p < 0.05, respectively). Moreover, the expression of transient receptor potential vanilloid 4, which synergistically regulates the osmolality together with AQP4, was significantly reduced in the aged skeletal muscles (p < 0.05). Therefore, the current study suggested that water transport abilities via AQP4 may decrease in the aged skeletal muscles, and thereby may be involved in age-related loss of muscle water content.
    Keywords:  AQP1; AQP4; Aging; Muscle water content; Skeletal muscle
  11. Microsc Res Tech. 2023 Jun 28.
      Down syndrome (DS) is a genetically based disease caused by triplication of chromosome 21. DS is characterized by multi-systemic premature aging associated with deficit in motor coordination, balance, and postural control. Using a morphological, morphometrical, and immunocytochemical ultrastructural approach, this study investigated in vastus lateralis muscle of Ts65Dn mouse, a murine model of DS, the effect of an adapted physical training on the extracellular matrix (ECM) characteristics and whether the forecasted exercise-induced ECM remodeling impacts on sarcomere organization. Morphometry demonstrated thicker basement membrane and larger collagen bundles with larger interfibrillar spacing as well as irregularly arrayed myofibrils and lower telethonin density on Z-lines in trisomic versus euploid sedentary mice. In agreement with the multi-systemic premature aging described in DS, these ECM alterations were similar to those previously observed in skeletal muscle of aged mice. Adapted physical training induced remodeling of ECM in both trisomic and euploid mice, that is, enlargement of the collagen bundles associated with hypertrophy of collagen fibrils and reduction of the interfibrillar spacing. A re-alignment of the myofibrils and a higher telethonin density on Z-line was found in trisomic mice. Altogether, our findings suggest that physical training is an effective tool in limiting/counteracting the trisomy-associated musculoskeletal structural anomalies. The current findings constitute a solid experimental background for further study investigating the possible positive effect of physical training on skeletal muscle performance. RESEARCH HIGHLIGHTS: Vastus lateralis muscle of trisomic mice shows aging-like alterations of extracellular matrix. Training promotes extracellular matrix remodeling. Training may be an effective tool to counteract trisomy-associated alterations of skeletal muscle.
    Keywords:  aging; exercise; sarcomere; transmission electron microscopy; trisomy
    DOI:  https://doi.org/10.1002/jemt.24379
  12. Am J Physiol Endocrinol Metab. 2023 Jun 28.
      FOLFOX (5-fluorouracil, leucovorin, oxaliplatin) chemotherapy is used to treat colorectal cancer and can acutely induce metabolic dysfunction. However, the lasting effects on systemic and skeletal muscle metabolism after treatment cessation are poorly understood. Therefore, we investigated the acute and lasting effects of FOLFOX chemotherapy on systemic and skeletal muscle metabolism in mice. Direct effects of FOLFOX in cultured myotubes were also investigated. Male C57BL/6J mice completed four cycles (acute) of FOLFOX or PBS. Subsets were allowed to recover for 4-wks or 10-wks. CLAMS metabolic measurements were performed for 5-days prior to study endpoint. C2C12 myotubes were treated with FOLFOX for 24-hrs. Acute FOLFOX attenuated body mass and body fat accretion independent of food intake or cage activity. Acute FOLFOX decreased blood glucose, oxygen consumption (VO2), carbon dioxide production (VCO2), energy expenditure, and carbohydrate (CHO) oxidation. Deficits in VO2 and energy expenditure remained at 10-wks. CHO oxidation remained disrupted at 4-wks but returned to control levels after 10-wks. Acute FOLFOX reduced muscle COXIV enzyme activity, AMPK(T172), ULK1(S555), and LC3BII protein expression. Muscle LC3BII/I ratio was associated with altered CHO oxidation (r=0.75, p=0.03). In vitro, FOLFOX suppressed myotube AMPK(T172), ULK1(S555), and autophagy flux. Recovery for 4-wks normalized skeletal muscle AMPK and ULK1 phosphorylation. Our results provide evidence that FOLFOX disrupts systemic metabolism, which is not readily recoverable after treatment cessation. FOLFOX effects on skeletal muscle metabolic signaling did recover. Further investigations are warranted to prevent and treat FOLFOX-induced metabolic toxicities that negatively impact cancer patient survival and life quality.
    Keywords:  5-FU; Autophagy; Carbohydrate Oxidation; Colorectal cancer; Energy Expenditure
    DOI:  https://doi.org/10.1152/ajpendo.00096.2023
  13. Metallomics. 2023 Jun 30. pii: mfad042. [Epub ahead of print]
      Cu (Cu) is essential for several biochemical pathways due to its role as a catalytic cofactor or allosteric regulator of enzymes. Its import and distribution are tightly controlled by transporters and metallochaperones and Cu homeostasis is maintained by balancing Cu uptake and export. Genetic diseases are caused by impaired Cu transporters CTR1, ATP7A, or ATP7B but little is known about the regulatory mechanisms by which these proteins meet the fluctuating demands of Cu in specific tissues. Cu is required for differentiation of skeletal myoblasts to myotubes. Here, we demonstrate that ATP7A is needed for myotube formation and that its increased abundance during differentiation is mediated by stabilization of Atp7a mRNA via the 3' untranslated region. Increased ATP7A levels during differentiation resulted in increased Cu delivery to lysyl oxidase, a secreted cuproenzyme that needed for myotube formation. These studies identify a previously unknown role for Cu in regulating muscle differentiation and have broad implications for understanding Cu-dependent differentiation in other tissues.
    DOI:  https://doi.org/10.1093/mtomcs/mfad042
  14. Cytotechnology. 2023 Aug;75(4): 335-348
      The plasticizer di (2-ethylhexyl) phthalate (DEHP) inhibits differentiation, impairs glucose metabolism, and decreases mitochondrial function in murine muscle satellite cells; however, if these effects are translated to human cells is unknown. The goal of this study was to evaluate changes in morphology and proliferation of primary human skeletal muscle cells exposed to DEHP. Rectus abdominis muscle samples were obtained from healthy women undergoing programed cesarean surgery. Skeletal muscle cells were isolated and grown under standard primary culture conditions, generating two independent sample groups of 25 subcultures each. Cells from the first group were exposed to 1 mM DEHP for 13 days and monitored for changes in cell morphology, satellite cell frequency and total cell abundance, while the second group remained untreated (control). Differences between treated and untreated groups were compared using generalized linear mixed models (GLMM). Cell membrane and nuclear envelope boundary alterations, loss of cell volume and presence of stress bodies were observed in DEHP-treated cultures. DEHP-treated cultures also showed a significant reduction in satellite cell frequency compared to controls. Exposure to DEHP reduced human skeletal muscle cell abundance. Statistical differences were found between the GLMM slopes, suggesting that exposure to DEHP reduced growth rate. These results suggest that exposure to DEHP inhibits human skeletal muscle cell proliferation, as evidenced by reduced cell abundance, potentially compromising long-term culture viability. Therefore, DEHP induces human skeletal muscle cell deterioration potentially inducing an inhibitory effect of myogenesis by depleting satellite cells.
    Graphical abstract:
    Keywords:  Cytotoxicity; Emerging pollutants; Phthalates; Primary culture; Striated muscle; Women
    DOI:  https://doi.org/10.1007/s10616-023-00580-4
  15. Life (Basel). 2023 Jun 11. pii: 1367. [Epub ahead of print]13(6):
      Mutations in the DMD gene can cause Duchenne or Becker muscular dystrophy (DMD/BMD) by affecting the giant isoform of dystrophin, a protein encoded by the DMD gene. The role of small dystrophin isoforms is not well investigated yet, and they may play a role in muscle development and molecular pathology. Here, we investigated the nuclear localization of short carboxy-terminal dystrophin isoforms during the in vitro differentiation of human, porcine, and murine myoblast cultures. We could not only confirm the presence of Dp71 in the nucleoplasm and at the nuclear envelope, but we could also identify the Dp40 isoform in muscle nuclei. The localization of both isoforms over the first six days of differentiation was similar between human and porcine myoblasts, but murine myoblasts behaved differently. This highlights the importance of the porcine model in investigating DMD. We could also detect a wave-like pattern of nuclear presence of both Dp71 and Dp40, indicating a direct or indirect involvement in gene expression control during muscle differentiation.
    Keywords:  DMD; Duchenne muscular dystrophy; dystrophin; muscle cell culture; nuclear localization; porcine muscle cells
    DOI:  https://doi.org/10.3390/life13061367
  16. Nat Commun. 2023 Jun 24. 14(1): 3766
      Successful muscle regeneration relies on the interplay of multiple cell populations. However, the signals required for this coordinated intercellular crosstalk remain largely unknown. Here, we describe how the Hedgehog (Hh) signaling pathway controls the fate of fibro/adipogenic progenitors (FAPs), the cellular origin of intramuscular fat (IMAT) and fibrotic scar tissue. Using conditional mutagenesis and pharmacological Hh modulators in vivo and in vitro, we identify DHH as the key ligand that acts as a potent adipogenic brake by preventing the adipogenic differentiation of FAPs. Hh signaling also impacts muscle regeneration, albeit indirectly through induction of myogenic factors in FAPs. Our results also indicate that ectopic and sustained Hh activation forces FAPs to adopt a fibrogenic fate resulting in widespread fibrosis. In this work, we reveal crucial post-developmental functions of Hh signaling in balancing tissue regeneration and fatty fibrosis. Moreover, they provide the exciting possibility that mis-regulation of the Hh pathway with age and disease could be a major driver of pathological IMAT formation.
    DOI:  https://doi.org/10.1038/s41467-023-39506-1
  17. Biomolecules. 2023 05 24. pii: 884. [Epub ahead of print]13(6):
      microRNAs (miRs) have been proposed as a promising new class of biomarkers in the context of training adaptation. Using microarray analysis, we studied skeletal muscle miR patterns in sedentary young healthy females (n = 6) before and after a single submaximal bout of endurance exercise ('reference training'). Subsequently, participants were subjected to a structured training program, consisting of six weeks of moderate-intensity continuous endurance training (MICT) and six weeks of high-intensity interval training (HIIT) in randomized order. In vastus lateralis muscle, we found significant downregulation of myomiRs, specifically miR-1, 133a-3p, and -5p, -133b, and -499a-5p. Similarly, exercise-associated miRs-23a-3p, -378a-5p, -128-3p, -21-5p, -107, -27a-3p, -126-3p, and -152-3p were significantly downregulated, whereas miR-23a-5p was upregulated. Furthermore, in an untargeted approach for differential expression in response to acute exercise, we identified n = 35 miRs that were downregulated and n = 20 miRs that were upregulated by factor 4.5 or more. Remarkably, KEGG pathway analysis indicated central involvement of this set of miRs in fatty acid metabolism. To reproduce these data in a larger cohort of all-female subjects (n = 29), qPCR analysis was carried out on n = 15 miRs selected from the microarray, which confirmed their differential expression. Furthermore, the acute response, i.e., the difference between miR concentrations before and after the reference training, was correlated with changes in maximum oxygen uptake (V̇O2max) in response to the training program. Here, we found that miRs-199a-3p and -19b-3p might be suitable acute-response candidates that correlate with individual degrees of training adaptation in females.
    Keywords:  biomarkers; exercise; females; micro RNAs; skeletal muscle
    DOI:  https://doi.org/10.3390/biom13060884
  18. Int J Mol Sci. 2023 Jun 09. pii: 9954. [Epub ahead of print]24(12):
      Exercise-induced muscle damage (EIMD) is a common occurrence in athletes and can lead to delayed onset muscle soreness, reduced athletic performance, and an increased risk of secondary injury. EIMD is a complex process involving oxidative stress, inflammation, and various cellular signaling pathways. Timely and effective repair of the extracellular matrix (ECM) and plasma membrane (PM) damage is critical for recovery from EIMD. Recent studies have shown that the targeted inhibition of phosphatase and tension homolog (PTEN) in skeletal muscles can enhance the ECM environment and reduce membrane damage in Duchenne muscular dystrophy (DMD) mice. However, the effects of PTEN inhibition on EIMD are unknown. Therefore, the present study aimed to investigate the potential therapeutic effects of VO-OHpic (VO), a PTEN inhibitor, on EIMD symptoms and underlying mechanisms. Our findings indicate that VO treatment effectively enhances skeletal muscle function and reduces strength loss during EIMD by upregulating membrane repair signals related to MG53 and ECM repair signals related to the tissue inhibitor of metalloproteinases (TIMPs) and matrix metalloproteinase (MMPs). These results highlight the potential of pharmacological PTEN inhibition as a promising therapeutic approach for EIMD.
    Keywords:  PTEN3; VO-OHpic2; exercise-induced muscle damage1; muscle extracellular matrix4; muscle membranes5
    DOI:  https://doi.org/10.3390/ijms24129954
  19. Proc Natl Acad Sci U S A. 2023 07 04. 120(27): e2211041120
      The molecular events governing skeletal muscle glucose uptake have pharmacological potential for managing insulin resistance in conditions such as obesity, diabetes, and cancer. With no current pharmacological treatments to target skeletal muscle insulin sensitivity, there is an unmet need to identify the molecular mechanisms that control insulin sensitivity in skeletal muscle. Here, the Rho guanine dissociation inhibitor α (RhoGDIα) is identified as a point of control in the regulation of insulin sensitivity. In skeletal muscle cells, RhoGDIα interacted with, and thereby inhibited, the Rho GTPase Rac1. In response to insulin, RhoGDIα was phosphorylated at S101 and Rac1 dissociated from RhoGDIα to facilitate skeletal muscle GLUT4 translocation. Accordingly, siRNA-mediated RhoGDIα depletion increased Rac1 activity and elevated GLUT4 translocation. Consistent with RhoGDIα's inhibitory effect, rAAV-mediated RhoGDIα overexpression in mouse muscle decreased insulin-stimulated glucose uptake and was detrimental to whole-body glucose tolerance. Aligning with RhoGDIα's negative role in insulin sensitivity, RhoGDIα protein content was elevated in skeletal muscle from insulin-resistant patients with type 2 diabetes. These data identify RhoGDIα as a clinically relevant controller of skeletal muscle insulin sensitivity and whole-body glucose homeostasis, mechanistically by modulating Rac1 activity.
    Keywords:  GLUT4 translocation; glucose uptake; insulin sensitivity; skeletal muscle; type 2 diabetes
    DOI:  https://doi.org/10.1073/pnas.2211041120
  20. Int J Mol Sci. 2023 Jun 06. pii: 9798. [Epub ahead of print]24(12):
      Autophagy dysregulation is commonplace in the pathogenesis of several invalidating diseases, such as musculoskeletal diseases. Polyamines, as spermidine and spermine, are small aliphatic cations essential for cell growth and differentiation, with multiple antioxidant, anti-inflammatory, and anti-apoptotic effects. Remarkably, they are emerging as natural autophagy regulators with strong anti-aging effects. Polyamine levels were significantly altered in the skeletal muscles of aged animals. Therefore, supplementation of spermine and spermidine may be important to prevent or treat muscle atrophy. Recent in vitro and in vivo experimental studies indicate that spermidine reverses dysfunctional autophagy and stimulates mitophagy in muscles and heart, preventing senescence. Physical exercise, as polyamines, regulates skeletal muscle mass inducing proper autophagy and mitophagy. This narrative review focuses on the latest evidence regarding the efficacy of polyamines and exercise as autophagy inducers, alone or coupled, in alleviating sarcopenia and aging-dependent musculoskeletal diseases. A comprehensive description of overall autophagic steps in muscle, polyamine metabolic pathways, and effects of the role of autophagy inducers played by both polyamines and exercise has been presented. Although literature shows few data in regard to this controversial topic, interesting effects on muscle atrophy in murine models have emerged when the two "autophagy-inducers" were combined. We hope these findings, with caution, can encourage researchers to continue investigating in this direction. In particular, if these novel insights could be confirmed in further in vivo and clinical studies, and the two synergic treatments could be optimized in terms of dose and duration, then polyamine supplementation and physical exercise might have a clinical potential in sarcopenia, and more importantly, implications for a healthy lifestyle in the elderly population.
    Keywords:  aging; autophagy; exercise; mitophagy; physical activity; polyamines; sarcopenia; spermidine; spermine
    DOI:  https://doi.org/10.3390/ijms24129798
  21. J Cell Biol. 2023 Aug 07. pii: e202211073. [Epub ahead of print]222(8):
      A balance between self-renewal and differentiation is critical for the regenerative capacity of tissue-resident stem cells. In skeletal muscle, successful regeneration requires the orchestrated activation, proliferation, and differentiation of muscle satellite cells (MuSCs) that are normally quiescent. A subset of MuSCs undergoes self-renewal to replenish the stem cell pool, but the features that identify and define self-renewing MuSCs remain to be elucidated. Here, through single-cell chromatin accessibility analysis, we reveal the self-renewal versus differentiation trajectories of MuSCs over the course of regeneration in vivo. We identify Betaglycan as a unique marker of self-renewing MuSCs that can be purified and efficiently contributes to regeneration after transplantation. We also show that SMAD4 and downstream genes are genetically required for self-renewal in vivo by restricting differentiation. Our study unveils the identity and mechanisms of self-renewing MuSCs, while providing a key resource for comprehensive analysis of muscle regeneration.
    DOI:  https://doi.org/10.1083/jcb.202211073
  22. Int J Mol Sci. 2023 Jun 16. pii: 10227. [Epub ahead of print]24(12):
      Skeletal muscle is continuously exposed during its activities to mechanical/oxidative damage [...].
    DOI:  https://doi.org/10.3390/ijms241210227
  23. Metabolites. 2023 May 26. pii: 694. [Epub ahead of print]13(6):
      Exercise has many benefits for physical and mental well-being. Metabolomics research has allowed scientists to study the impact of exercise on the body by analyzing metabolites released by tissues such as skeletal muscle, bone, and the liver. Endurance training increases mitochondrial content and oxidative enzymes, while resistance training increases muscle fiber and glycolytic enzymes. Acute endurance exercise affects amino acid metabolism, fat metabolism, cellular energy metabolism, and cofactor and vitamin metabolism. Subacute endurance exercise alters amino acid metabolism, lipid metabolism, and nucleotide metabolism. Chronic endurance exercise improves lipid metabolism and changes amino acid metabolism. Acute resistance exercise changes several metabolic pathways, including anaerobic processes and muscular strength. Chronic resistance exercise affects metabolic pathways, resulting in skeletal muscle adaptations. Combined endurance-resistance exercise alters lipid metabolism, carbohydrate metabolism, and amino acid metabolism, increasing anaerobic metabolic capacity and fatigue resistance. Studying exercise-induced metabolites is a growing field, and further research can uncover the underlying metabolic mechanisms and help tailor exercise programs for optimal health and performance.
    Keywords:  combined endurance–resistance exercise; endurance exercise; exercise; metabolites; metabolomics; resistance exercise
    DOI:  https://doi.org/10.3390/metabo13060694
  24. Aging (Albany NY). 2023 Jun 24. 15
      
    Keywords:  aging; mitochondria; mitochondrial dynamics; mitophagy; sarcopenia
    DOI:  https://doi.org/10.18632/aging.204857
  25. Nat Metab. 2023 Jun 26.
      Physical endurance and energy conservation are essential for survival in the wild. However, it remains unknown whether and how meal timing regulates physical endurance and muscle diurnal rhythms. Here, we show that day/sleep time-restricted feeding (DRF) enhances running endurance by 100% throughout the circadian cycle in both male and female mice, compared to either ad libitum feeding or night/wake time-restricted feeding. Ablation of the circadian clock in the whole body or the muscle abolished the exercise regulatory effect of DRF. Multi-omics analysis revealed that DRF robustly entrains diurnal rhythms of a mitochondrial oxidative metabolism-centric network, compared to night/wake time-restricted feeding. Remarkably, muscle-specific knockdown of the myocyte lipid droplet protein perilipin-5 completely mimics DRF in enhancing endurance, enhancing oxidative bioenergetics and outputting rhythmicity to circulating energy substrates, including acylcarnitine. Together, our work identifies a potent dietary regimen to enhance running endurance without prior exercise, as well as providing a multi-omics atlas of muscle circadian biology regulated by meal timing.
    DOI:  https://doi.org/10.1038/s42255-023-00826-7
  26. Nature. 2023 Jun 28.
      Caloric restriction that promotes weight loss is an effective strategy for treating non-alcoholic fatty liver disease and improving insulin sensitivity in people with type 2 diabetes1. Despite its effectiveness, in most individuals, weight loss is usually not maintained partly due to physiological adaptations that suppress energy expenditure, a process known as adaptive thermogenesis, the mechanistic underpinnings of which are unclear2,3. Treatment of rodents fed a high-fat diet with recombinant growth differentiating factor 15 (GDF15) reduces obesity and improves glycaemic control through glial-cell-derived neurotrophic factor family receptor α-like (GFRAL)-dependent suppression of food intake4-7. Here we find that, in addition to suppressing appetite, GDF15 counteracts compensatory reductions in energy expenditure, eliciting greater weight loss and reductions in non-alcoholic fatty liver disease (NAFLD) compared to caloric restriction alone. This effect of GDF15 to maintain energy expenditure during calorie restriction requires a GFRAL-β-adrenergic-dependent signalling axis that increases fatty acid oxidation and calcium futile cycling in the skeletal muscle of mice. These data indicate that therapeutic targeting of the GDF15-GFRAL pathway may be useful for maintaining energy expenditure in skeletal muscle during caloric restriction.
    DOI:  https://doi.org/10.1038/s41586-023-06249-4
  27. iScience. 2023 Jul 21. 26(7): 107047
      We examined the effects of ∼30 days of spaceflight on glycogen synthase kinase 3 (GSK3) content and inhibitory serine phosphorylation in murine muscle and bone samples from four separate missions (BION-M1, rodent research [RR]1, RR9, and RR18). Spaceflight reduced GSK3β content across all missions, whereas its serine phosphorylation was elevated with RR18 and BION-M1. The reduction in GSK3β was linked to the reduction in type IIA fibers commonly observed with spaceflight as these fibers are particularly enriched with GSK3. We then tested the effects of inhibiting GSK3 before this fiber type shift, and we demonstrate that muscle-specific Gsk3 knockdown increased muscle mass, preserved muscle strength, and promoted the oxidative fiber type with Earth-based hindlimb unloading. In bone, GSK3 activation was enhanced after spaceflight; and strikingly, muscle-specific Gsk3 deletion increased bone mineral density in response to hindlimb unloading. Thus, future studies should test the effects of GSK3 inhibition during spaceflight.
    Keywords:  Musculoskeletal medicine; Space medicine
    DOI:  https://doi.org/10.1016/j.isci.2023.107047