bims-moremu Biomed News
on Molecular regulators of muscle mass
Issue of 2021‒04‒04
forty-nine papers selected by
Anna Vainshtein
Craft Science Inc.

  1. Front Cell Dev Biol. 2021 ;9 606403
      Satellite cells (SCs) are tissue-specific stem cells responsible for adult skeletal muscle regeneration and maintenance. SCs function is critically dependent on two families of transcription factors: the paired box (Pax) involved in specification and maintenance and the Muscle Regulatory Factors (MRFs), which orchestrate myogenic commitment and differentiation. In turn, signaling events triggered by extrinsic and intrinsic stimuli control their function via post-translational modifications, including ubiquitination and phosphorylation. In this context, the Abelson non-receptor tyrosine kinase (c-Abl) mediates the activation of the p38 α/β MAPK pathway, promoting myogenesis. c-Abl also regulates the activity of the transcription factor MyoD during DNA-damage stress response, pausing differentiation. However, it is not clear if c-Abl modulates other key transcription factors controlling SC function. This work aims to determine the role of c-Abl in SCs myogenic capacity via loss of function approaches in vitro and in vivo. Here we show that c-Abl inhibition or deletion results in a down-regulation of Pax7 mRNA and protein levels, accompanied by decreased Pax7 transcriptional activity, without a significant effect on MRF expression. Additionally, we provide data indicating that Pax7 is directly phosphorylated by c-Abl. Finally, SC-specific c-Abl ablation impairs muscle regeneration upon acute injury. Our results indicate that c-Abl regulates myogenic progression in activated SCs by controlling Pax7 function and expression.
    Keywords:  MRFs; c-Abl; muscle differentiation; muscle stem cells; pax7; satellite cells; skeletal muscle regeneration
  2. FASEB J. 2021 May;35(5): e21503
      The muscle regeneration process requires a properly assembled extracellular matrix (ECM). Its homeostasis depends on the activity of different matrix-metalloproteinases (MMPs). The reversion-inducing-cysteine-rich protein with kazal motifs (RECK) is a membrane-anchored protein that negatively regulates the activity of different MMPs. However, the role of RECK in the process of skeletal muscle differentiation, regeneration, and fibrosis has not been elucidated. Here, we show that during skeletal muscle differentiation of C2C12 myoblasts and in satellite cells on isolated muscle fibers, RECK is transiently up regulated. C2C12 myoblasts with reduced RECK levels are more prone to enter the differentiation program, showing an accelerated differentiation process. Notch-1 signaling was reduced, while p38 and AKT signaling were augmented in myoblasts with decreased RECK levels. Overexpression of RECK restores the normal differentiation process but diminished the ability to form myotubes. Transient up-regulation of RECK occurs during skeletal muscle regeneration, which was accelerated in RECK-deficient mice (Reck±). RECK, MMPs and ECM proteins augmented in chronically damaged WT muscle, a model of muscle fibrosis. In this model, RECK ± mice showed diminished fibrosis compared to WT. These results strongly suggest that RECK is acting as a potential myogenic repressor during muscle formation and regeneration, emerging as a new player in these processes, and as a potential target to treat individuals with the muscle-wasting disease.
    Keywords:  MMP; RECK; muscle fibrosis; regeneration; skeletal muscle differentiation
  3. Cells. 2021 Mar 28. pii: 744. [Epub ahead of print]10(4):
      Background: Skeletal muscle is one of the only mammalian tissues capable of rapid and efficient regeneration after trauma or in pathological conditions. Skeletal muscle regeneration is driven by the muscle satellite cells, the stem cell population in interaction with their niche. Upon injury, muscle fibers undergo necrosis and muscle stem cells activate, proliferate and fuse to form new myofibers. In addition to myogenic cell populations, interaction with other cell types such as inflammatory cells, mesenchymal (fibroadipogenic progenitors-FAPs, pericytes) and vascular (endothelial) lineages are important for efficient muscle repair. While the role of the distinct populations involved in skeletal muscle regeneration is well characterized, the quantitative changes in the muscle stem cell and niche during the regeneration process remain poorly characterized. Methods: We have used mass cytometry to follow the main muscle cell types (muscle stem cells, vascular, mesenchymal and immune cell lineages) during early activation and over the course of muscle regeneration at D0, D2, D5 and D7 compared with uninjured muscles. Results: Early activation induces a number of rapid changes in the proteome of multiple cell types. Following the induction of damage, we observe a drastic loss of myogenic, vascular and mesenchymal cell lineages while immune cells invade the damaged tissue to clear debris and promote muscle repair. Immune cells constitute up to 80% of the mononuclear cells 5 days post-injury. We show that muscle stem cells are quickly activated in order to form new myofibers and reconstitute the quiescent muscle stem cell pool. In addition, our study provides a quantitative analysis of the various myogenic populations during muscle repair. Conclusions: We have developed a mass cytometry panel to investigate the dynamic nature of muscle regeneration at a single-cell level. Using our panel, we have identified early changes in the proteome of stressed satellite and niche cells. We have also quantified changes in the major cell types of skeletal muscle during regeneration and analyzed myogenic transcription factor expression in satellite cells throughout this process. Our results highlight the progressive dynamic shifts in cell populations and the distinct states of muscle stem cells adopted during skeletal muscle regeneration. Our findings give a deeper understanding of the cellular and molecular aspects of muscle regeneration.
    Keywords:  CyTOF; muscle niche; muscle stem cells; regeneration; satellite cells; skeletal muscle
  4. J Cachexia Sarcopenia Muscle. 2021 Mar 30.
      Cancer cachexia is a complex multi-organ catabolic syndrome that reduces mobility, increases fatigue, decreases the efficiency of therapeutic strategies, diminishes the quality of life, and increases the mortality of cancer patients. This review provides an exhaustive and comprehensive analysis of cancer cachexia-related phenotypic changes in skeletal muscle at both the cellular and subcellular levels in human cancer patients, as well as in animal models of cancer cachexia. Cancer cachexia is characterized by a major decrease in skeletal muscle mass in human and animals that depends on the severity of the disease/model and the localization of the tumour. It affects both type 1 and type 2 muscle fibres, even if some animal studies suggest that type 2 muscle fibres would be more prone to atrophy. Animal studies indicate an impairment in mitochondrial oxidative metabolism resulting from a decrease in mitochondrial content, an alteration in mitochondria morphology, and a reduction in mitochondrial metabolic fluxes. Immuno-histological analyses in human and animal models also suggest that a faulty mechanism of skeletal muscle repair would contribute to muscle mass loss. An increase in collagen deposit, an accumulation of fat depot outside and inside the muscle fibre, and a disrupted contractile machinery structure are also phenotypic features that have been consistently reported in cachectic skeletal muscle. Muscle function is also profoundly altered during cancer cachexia with a strong reduction in skeletal muscle force. Even though the loss of skeletal muscle mass largely contributes to the loss of muscle function, other factors such as muscle-nerve interaction and calcium handling are probably involved in the decrease in muscle force. Longitudinal analyses of skeletal muscle mass by imaging technics and skeletal muscle force in cancer patients, but also in animal models of cancer cachexia, are necessary to determine the respective kinetics and functional involvements of these factors. Our analysis also emphasizes that measuring skeletal muscle force through standardized tests could provide a simple and robust mean to early diagnose cachexia in cancer patients. That would be of great benefit to cancer patient's quality of life and health care systems.
    Keywords:  Cancer cachexia; Fibre type; Fibrosis; Force; Metabolism; Regeneration; Skeletal muscle
  5. BMC Biol. 2021 Apr 01. 19(1): 44
      BACKGROUND: IK is a splicing factor that promotes spliceosome activation and contributes to pre-mRNA splicing. Although the molecular mechanism of IK has been previously reported in vitro, the physiological role of IK has not been fully understood in any animal model. Here, we generate an ik knock-out (KO) zebrafish using the CRISPR/Cas9 system to investigate the physiological roles of IK in vivo.RESULTS: The ik KO embryos display severe pleiotropic phenotypes, implying an essential role of IK in embryonic development in vertebrates. RNA-seq analysis reveals downregulation of genes involved in skeletal muscle differentiation in ik KO embryos, and there exist genes having improper pre-mRNA splicing among downregulated genes. The ik KO embryos display impaired neuromuscular junction (NMJ) and fast-twitch muscle development. Depletion of ik reduces myod1 expression and upregulates pax7a, preventing normal fast muscle development in a non-cell-autonomous manner. Moreover, when differentiation is induced in IK-depleted C2C12 myoblasts, myoblasts show a reduced ability to form myotubes. However, inhibition of IK does not influence either muscle cell proliferation or apoptosis in zebrafish and C2C12 cells.
    CONCLUSION: This study provides that the splicing factor IK contributes to normal skeletal muscle development in vivo and myogenic differentiation in vitro.
    Keywords:  CRISPR/Cas9; IK; Myogenesis; Skeletal muscles; Zebrafish
  6. Rev Endocr Metab Disord. 2021 Mar 30.
      Sarcopenia, cachexia, and atrophy due to inactivity and disease states are characterized by a loss of skeletal muscle mass, often accompanied by reduced levels of anabolic hormones (e.g. testosterone). These conditions are associated with an increase in mortality, hospitalization and worsening in quality of life. Both physical exercise (EX) and anabolic-androgenic steroid (AAS) administration can improve the prognosis of patients as they increase physical functionality. However, there is a gap in the literature as to the impact of these therapies on the gains in strength and muscle mass and their implications for patient safety. Accordingly, we performed a random-effects meta-analysis to elucidate the effects of AAS and/or EX interventions on lean body mass (LBM) and muscle strength in conditions involving muscle loss. A systematic search for relevant clinical trials was conducted in MEDLINE, EMBASE, SCOPUS, Web of Science, and SPORTDiscus. Comparisons included AAS vs. Control, EX vs. Control, AAS vs. EX, AAS + EX vs. AAS and AAS + EX vs. EX. A total of 1114 individuals were analyzed. AAS increased LBM (effect size [ES]: 0.46; 95% CI: 0.25, 0.68, P = 0.00) and muscle strength (ES: 0.31; 95% CI: 0.08, 0.53, P = 0.01) when compared to a control group. EX promoted an increase in muscular strength (ES: 0.89; 95% CI: 0.53, 1.25, P = 0.00), with no effect on LBM when compared to the control group (ES: 0.15; 95% CI: -0.07, 0.38, P = 0.17). AAS did not demonstrate statistically significant differences when compared to EX for LBM and muscle strength. The combination of EX + AAS promoted a greater increase in LBM and muscular strength when compared to AAS or EX in isolation. Qualitatively, AAS administration had relatively few side effects. Significant heterogeneity was found in some analyses, which may be explained by the use of different AAS types and EX protocols. Our findings suggest that AAS administration in cachectic and sarcopenic conditions may be a viable interventional strategy to enhance muscle function when exercise is not a possible approach. Moreover, combining AAS with exercise may enhance positive outcomes in this population.
    Keywords:  Anabolic–androgenic steroids; Exercise; Muscle mass; Strength; Testosterone
  7. Int J Mol Sci. 2021 Mar 02. pii: 2488. [Epub ahead of print]22(5):
      The cell-cell/cell-matrix interactions between myoblasts and their extracellular microenvironment have been shown to play a crucial role in the regulation of in vitro myogenic differentiation and in vivo skeletal muscle regeneration. In this study, by harnessing the heparin-mimicking polymer, poly(sodium-4-styrenesulfonate) (PSS), which has a negatively charged surface, we engineered an in vitro cell culture platform for the purpose of recapitulating in vivo muscle atrophy-like phenotypes. Our initial findings showed that heparin-mimicking moieties inhibited the fusion of mononucleated myoblasts into multinucleated myotubes, as indicated by the decreased gene and protein expression levels of myogenic factors, myotube fusion-related markers, and focal adhesion kinase (FAK). We further elucidated the underlying molecular mechanism via transcriptome analyses, observing that the insulin/PI3K/mTOR and Wnt signaling pathways were significantly downregulated by heparin-mimicking moieties through the inhibition of FAK/Cav3. Taken together, the easy-to-adapt heparin-mimicking polymer-based in vitro cell culture platform could be an attractive platform for potential applications in drug screening, providing clear readouts of changes in insulin/PI3K/mTOR and Wnt signaling pathways.
    Keywords:  focal adhesion kinase (FAK); fusion; myoblast; myogenic differentiation; poly(sodium-4-styrenesulfonate); synthetic mimic of heparin
  8. Br J Pharmacol. 2021 Mar 31.
      BACKGROUND AND PURPOSE: Increasing evidence suggests systemic inflammation-caused skeletal muscle atrophy as a major clinical feature of cachexia. Triptolide obtained from Tripterygium wilfordii Hook F possesses potent anti-inflammatory and immunosuppressive effects. The present study aims to evaluate the protective effects and molecular mechanisms of triptolide on inflammation-induced skeletal muscle atrophy.EXPERIMENTAL APPROACH: The effects of triptolide on skeletal muscle atrophy were investigated in lipopolysaccharide (LPS)-treated C2C12 myotubes and C57BL/6 mice. Protein expressions and mRNA levels were analyzed by western blot and qPCR respectively. Skeletal muscle mass, volume and strength were measured by histological analysis, micro-CT and grip strength respectively. Locomotor activity was measured by the open field testing.
    KEY RESULTS: Triptolide (10-100 fM) upregulated protein synthesis signals (IGF-1/p-IGF-1R/IRS-1/p-Akt/p-mTOR) and downregulated protein degradation signal atrogin-1 in C2C12 myotubes. In LPS (100 ng·ml-1 )-treated C2C12 myotubes, triptolide upregulated MyHC, IGF-1, p-IGF-1R, IRS-1 and p-Akt. Triptolide also down-regulated ubiquitin-proteasome molecules (n-FoxO3a/atrogin-1/MuRF1), proteasome activity, autophagy-lysosomal molecules (LC3-II/LC3-I and Bnip3), and inflammatory mediators (NF-κB, Cox-2, NLRP3, IL-1β, and TNF-α). However, AG1024, an IGF-1R inhibitor, suppressed triptolide-mediated effects on MyHC, myotube diameter, MuRF1 and p62 in LPS-treated C2C12 myotubes. In LPS (1 mg·kg-1 , i.p.)-challenged mice, triptolide (5 and 20 μg·kg-1 ·day-1 , i.p.) decreased plasma TNF-α levels, and it increased skeletal muscle volume, cross-sectional area of myofibers, weights of the gastrocnemius and tibialis anterior muscles, forelimb grip strength and locomotion.
    CONCLUSIONS AND IMPLICATIONS: These findings reveal that triptolide prevented LPS-induced inflammation and skeletal muscle atrophy and have implications for the discovery of novel agents for preventing muscle wasting.
    Keywords:  anti-inflammation; autophagy-lysosomal pathway; insulin-like growth factor 1; skeletal muscle atrophy; triptolide; ubiquitin-proteasome system
  9. J Physiol Biochem. 2021 Mar 31.
      Skeletal muscle atrophy (SMA) is a dominant symptom induced by estrogen deficiency which can lead to severe health problems of postmenopausal women. Furthermore, estrogen deficiency has severely compromised the maintenance of muscle stem cells as well as impairs self-renewal and differentiation into muscle fibers. Resistance training is commonly considered as a positive and useful intervention in accelerating the rate of muscle growth. As one of the resistance training, whether the weight-bearing exercise can alleviate SMA induced by estrogen deficiency has not been investigated. The rats were divided into 3 groups randomly: sham group, ovariectomized (OVX) group, and weight-bearing exercise (WBE) therapeutic group. The weight that rats were loaded was 35% of their body weight, and the rats were trained by treadmill training (5° slope, 20 m/min, 30 min/day, 6 days/week) for 8 weeks. After training, the quality and strength of skeletal muscle of the WBE rats were improved; meanwhile, the cross-sectional areas of the skeletal muscle were also increased. Moreover, the WBE activated Akt significantly, upregulated the expression of mTOR, and downregulated the expression of MSTN and its receptor ActRIIB and FoxO1, respectively. The SMA phenomena of rats which induced by estrogen deficiency were prevented effectively via WBE, and the MSTN/Akt/mTOR and FoxO1 signaling pathway may be the predominant way in this improvement.
    Keywords:  Skeletal muscle atrophy,·Estrogen deficiency,·Ovariectomized rats,·Myostatin,·Signaling pathway; Weight-bearing exercise
  10. Stem Cell Reports. 2021 Mar 17. pii: S2213-6711(21)00134-X. [Epub ahead of print]
      The transplantation of muscle progenitor cells (MuPCs) differentiated from human induced pluripotent stem cells (hiPSCs) is a promising approach for treating skeletal muscle diseases such as Duchenne muscular dystrophy (DMD). However, proper purification of the MuPCs before transplantation is essential for clinical application. Here, by using MYF5 hiPSC reporter lines, we identified two markers for myogenic cell purification: CDH13, which purified most of the myogenic cells, and FGFR4, which purified a subset of MuPCs. Cells purified with each of the markers showed high efficiency for regeneration after transplantation and contributed to the restoration of dystrophin expression in DMD-immunodeficient model mice. Moreover, we found that MYF5 regulates CDH13 expression by binding to the promoter regions. These findings suggest that FGFR4 and CDH13 are strong candidates for the purification of hiPSC-derived MuPCs for therapeutical application.
    Keywords:  CDH13; FGFR4; iPSC; muscle stem cell; skeletal muscle; surface marker
  11. Int J Mol Sci. 2021 Mar 08. pii: 2741. [Epub ahead of print]22(5):
      The regulation of skeletal muscle mass and organelle homeostasis is dependent on the capacity of cells to produce proteins and to recycle cytosolic portions. In this investigation, the mechanisms involved in skeletal muscle mass regulation-especially those associated with proteosynthesis and with the production of new organelles-are presented. Thus, the critical roles of mammalian/mechanistic target of rapamycin complex 1 (mTORC1) pathway and its regulators are reviewed. In addition, the importance of ribosome biogenesis, satellite cells involvement, myonuclear accretion, and some major epigenetic modifications related to protein synthesis are discussed. Furthermore, several studies conducted on the topic of exercise training have recognized the central role of both endurance and resistance exercise to reorganize sarcomeric proteins and to improve the capacity of cells to build efficient organelles. The molecular mechanisms underlying these adaptations to exercise training are presented throughout this review and practical recommendations for exercise prescription are provided. A better understanding of the aforementioned cellular pathways is essential for both healthy and sick people to avoid inefficient prescriptions and to improve muscle function with emergent strategies (e.g., hypoxic training). Finally, current limitations in the literature and further perspectives, notably on epigenetic mechanisms, are provided to encourage additional investigations on this topic.
    Keywords:  DNA methylation; eIF3f; endurance training; epigenetic modifications; hypoxia; mTOR; protein turnover; resistance training; ribosome biogenesis; satellite cells
  12. Int J Mol Sci. 2021 Mar 04. pii: 2567. [Epub ahead of print]22(5):
      Skeletal muscle is an essential organ, responsible for many physiological functions such as breathing, locomotion, postural maintenance, thermoregulation, and metabolism. Interestingly, skeletal muscle is a highly plastic tissue, capable of adapting to anabolic and catabolic stimuli. Skeletal muscle contains a specialized smooth endoplasmic reticulum (ER), known as the sarcoplasmic reticulum, composed of an extensive network of tubules. In addition to the role of folding and trafficking proteins within the cell, this specialized organelle is responsible for the regulated release of calcium ions (Ca2+) into the cytoplasm to trigger a muscle contraction. Under various stimuli, such as exercise, hypoxia, imbalances in calcium levels, ER homeostasis is disturbed and the amount of misfolded and/or unfolded proteins accumulates in the ER. This accumulation of misfolded/unfolded protein causes ER stress and leads to the activation of the unfolded protein response (UPR). Interestingly, the role of the UPR in skeletal muscle has only just begun to be elucidated. Accumulating evidence suggests that ER stress and UPR markers are drastically induced in various catabolic stimuli including cachexia, denervation, nutrient deprivation, aging, and disease. Evidence indicates some of these molecules appear to be aiding the skeletal muscle in regaining homeostasis whereas others demonstrate the ability to drive the atrophy. Continued investigations into the individual molecules of this complex pathway are necessary to fully understand the mechanisms.
    Keywords:  ER stress; UPR; atrophy; muscle wasting; skeletal muscle
  13. FASEB J. 2021 May;35(5): e21480
      Skeletal muscle ischemia is a major consequence of peripheral arterial disease (PAD) or critical limb ischemia (CLI). Although therapeutic options for resolving muscle ischemia in PAD/CLI are limited, the issue is compounded by poor understanding of the mechanisms driving muscle vascularization. We found that nuclear receptor estrogen-related receptor alpha (ERRα) expression is induced in murine skeletal muscle by hindlimb ischemia (HLI), and in cultured myotubes by hypoxia, suggesting a potential role for ERRα in ischemic response. To test this, we generated skeletal muscle-specific ERRα transgenic (TG) mice. In these mice, ERRα drives myofiber type switch from glycolytic type IIB to oxidative type IIA/IIX myofibers, which are typically associated with more vascular supply in muscle. Indeed, RNA sequencing and functional enrichment analysis of TG muscle revealed that "paracrine angiogenesis" is the top-ranked transcriptional program activated by ERRα in the skeletal muscle. Immunohistochemistry and angiography showed that ERRα overexpression increases baseline capillarity, arterioles and non-leaky blood vessel formation in the skeletal muscles. Moreover, ERRα overexpression facilitates ischemic neo-angiogenesis and perfusion recovery in hindlimb musculature of mice subjected to HLI. Therefore, ERRα is a hypoxia inducible nuclear receptor that is involved in skeletal muscle angiogenesis and could be potentially targeted for treating PAD/CLI.
    Keywords:  angiogenesis; gene expression; ischemia; nuclear receptors; skeletal muscle
  14. Cells. 2021 Mar 10. pii: 615. [Epub ahead of print]10(3):
      Muscle-enriched A-type lamin-interacting protein (Mlip) is a recently discovered Amniota gene that encodes proteins of unknown biological function. Here we report Mlip's direct interaction with chromatin, and it may function as a transcriptional co-factor. Chromatin immunoprecipitations with microarray analysis demonstrated a propensity for Mlip to associate with genomic regions in close proximity to genes that control tissue-specific differentiation. Gel mobility shift assays confirmed that Mlip protein complexes with genomic DNA. Blocking Mlip expression in C2C12 myoblasts down-regulates myogenic regulatory factors (MyoD and MyoG) and subsequently significantly inhibits myogenic differentiation and the formation of myotubes. Collectively our data demonstrate that Mlip is required for C2C12 myoblast differentiation into myotubes. Mlip may exert this role as a transcriptional regulator of a myogenic program that is unique to amniotes.
    Keywords:  ChIP; Mlip; skeletal muscle
  15. PLoS Genet. 2021 Mar 29. 17(3): e1009488
      Mitochondria are essential for maintaining skeletal muscle metabolic homeostasis during adaptive response to a myriad of physiologic or pathophysiological stresses. The mechanisms by which mitochondrial function and contractile fiber type are concordantly regulated to ensure muscle function remain poorly understood. Evidence is emerging that the Folliculin interacting protein 1 (Fnip1) is involved in skeletal muscle fiber type specification, function, and disease. In this study, Fnip1 was specifically expressed in skeletal muscle in Fnip1-transgenic (Fnip1Tg) mice. Fnip1Tg mice were crossed with Fnip1-knockout (Fnip1KO) mice to generate Fnip1TgKO mice expressing Fnip1 only in skeletal muscle but not in other tissues. Our results indicate that, in addition to the known role in type I fiber program, FNIP1 exerts control upon muscle mitochondrial oxidative program through AMPK signaling. Indeed, basal levels of FNIP1 are sufficient to inhibit AMPK but not mTORC1 activity in skeletal muscle cells. Gain-of-function and loss-of-function strategies in mice, together with assessment of primary muscle cells, demonstrated that skeletal muscle mitochondrial program is suppressed via the inhibitory actions of FNIP1 on AMPK. Surprisingly, the FNIP1 actions on type I fiber program is independent of AMPK and its downstream PGC-1α. These studies provide a vital framework for understanding the intrinsic role of FNIP1 as a crucial factor in the concerted regulation of mitochondrial function and muscle fiber type that determine muscle fitness.
  16. Skelet Muscle. 2021 Mar 30. 11(1): 9
      BACKGROUND: ALAS2 (delta-aminolevulinate synthase 2) is one of the two isoenzymes catalyzing the synthesis of delta-aminolevulinic acid (ALA), which is the first precursor of heme synthesis. ALAS2-overexpressing transgenic mice (Tg mice) showed syndrome of porphyria, a series of diseases related to the heme anabolism deficiency. Tg mice showed an obvious decrease in muscle size. Muscle atrophy results from a decrease in protein synthesis and an increase in protein degradation, which ultimately leads to a decrease in myofiber size due to loss of contractile proteins, organelles, nuclei, and cytoplasm.METHODS: The forelimb muscle grip strength of age-matched ALAS-2 transgenic mice (Tg mice) and wild-type mice (WT mice) were measured with an automated grip strength meter. The activities of serum LDH and CK-MB were measured by Modular DPP. The histology of skeletal muscle (quadriceps femoris and gastrocnemius) was observed by hematoxylin and eosin (HE) staining, immunohistochemistry, and transmission electron microscope. Real-time PCR was used to detect mtDNA content and UCP3 mRNA expression. Evans blue dye staining was used to detect the membrane damage of the muscle fiber. Single skeletal muscle fiber diameter was measured by single-fiber analyses. Muscle adenosine triphosphate (ATP) levels were detected by a luminometric assay with an ATP assay kit.
    RESULTS: Compared with WT mice, the strength of forelimb muscle and mass of gastrocnemius were decreased in Tg mice. The activities of serum CK-MB and LDH, the number of central nuclei fibers, and Evans blue positive fibers were more than those in WT mice, while the diameter of single fibers was smaller, which were associated with suppressed expression levels of MHC, myoD1, dystrophin, atrogin1, and MuRF1. Re-expression of eMyHC was only showed in the quadriceps of Tg mice, but not in WT mice. Muscle mitochondria in Tg mice showed dysfunction with descented ATP production and mtDNA content, downregulated UCP3 mRNA expression, and swelling of mitochondria.
    CONCLUSION: ALAS2 overexpressing-transgenic mice (Tg mice) showed muscle dystrophy, which was associated with decreased atrogin-1 and MuRF-1, and closely related to mitochondrial dysfunction.
    Keywords:  Delta-aminolevulinate synthase 2; Mitochondrial dysfunction; Muscle atrophy; Transgenic mice
  17. Commun Biol. 2021 Mar 29. 4(1): 427
      Parabiosis experiments suggest that molecular factors related to rejuvenation and aging circulate in the blood. Here, we show that miR-199-3p, which circulates in the blood as a cell-free miRNA, is significantly decreased in the blood of aged mice compared to young mice; and miR-199-3p has the ability to enhance myogenic differentiation and muscle regeneration. Administration of miR-199 mimics, which supply miR-199-3p, to aged mice resulted in muscle fiber hypertrophy and delayed loss of muscle strength. Systemic administration of miR-199 mimics to mdx mice, a well-known animal model of Duchenne muscular dystrophy (DMD), markedly improved the muscle strength of mice. Taken together, cell-free miR-199-3p in the blood may have an anti-aging effect such as a hypertrophic effect in aged muscle fibers and could have potential as a novel RNA therapeutic for DMD as well as age-related diseases. The findings provide us with new insights into blood-circulating miRNAs as age-related molecules.
  18. Int J Mol Sci. 2021 Mar 31. pii: 3627. [Epub ahead of print]22(7):
      Skeletal muscle regeneration is a well-organized process that requires remodeling of the extracellular matrix (ECM). In this study, we revealed the protective role of periostin, a matricellular protein that binds to several ECM proteins during muscle regeneration. In intact muscle, periostin was localized at the neuromuscular junction, muscle spindle, and myotendinous junction, which are connection sites between muscle fibers and nerves or tendons. During muscle regeneration, periostin exhibited robustly increased expression and localization at the interstitial space. Periostin-null mice showed decreased muscle weight due to the loss of muscle fibers during repeated muscle regeneration. Cultured muscle progenitor cells from periostin-null mice showed no deficiencies in their proliferation, differentiation, and the expression of Pax7, MyoD, and myogenin, suggesting that the loss of muscle fibers in periostin-null mice was not due to the impaired function of muscle stem/progenitor cells. Periostin-null mice displayed a decreased number of CD31-positive blood vessels during muscle regeneration, suggesting that the decreased nutritional supply from blood vessels was the cause of muscle fiber loss in periostin-null mice. These results highlight the novel role of periostin in maintaining muscle mass during muscle regeneration.
    Keywords:  extracellular matrix; periostin; skeletal muscle regeneration
  19. Int J Environ Res Public Health. 2021 Mar 01. pii: 2367. [Epub ahead of print]18(5):
      BST204 is a purified ginseng dry extract that has an inhibitory effect on lipopolysaccharide-induced inflammatory responses, but its effect on muscle atrophy is yet to be investigated. In this study, C2C12 myoblasts were induced to differentiate for three days followed by the treatment of dexamethasone (DEX), a corticosteroid drug, with vehicle or BST204 for one day and subjected to immunoblotting, immunocytochemistry, qRT-PCR and biochemical analysis for mitochondrial function. BST204 alleviates the myotube atrophic effect mediated by DEX via the activation of protein kinase B/mammalian target of rapamycin (Akt/mTOR) signaling. Through this pathway, BST204 suppresses the expression of muscle-specific E3 ubiquitin ligases contributing to the enhanced myotube formation and enlarged myotube diameter in DEX-treated myotubes. In addition, BST204 treatment significantly decreases the mitochondrial reactive oxygen species production in DEX-treated myotubes. Furthermore, BST204 improves mitochondrial function by upregulating the expression of peroxisome proliferator-activated receptor-γ coactivator-1α (PGC1α) in DEX-induced myotube atrophy. This study provides a mechanistic insight into the effect of BST204 on DEX-induced myotube atrophy, suggesting that BST204 has protective effects against the toxicity of a corticosteroid drug in muscle and promising potential as a nutraceutical remedy for the treatment of muscle weakness and atrophy.
    Keywords:  Akt/mTOR pathway; BST204; PGC1α; mitochondria; muscle atrophy
  20. Int J Mol Sci. 2021 Mar 18. pii: 3110. [Epub ahead of print]22(6):
      Cachexia is a multifactorial syndrome characterized by muscle loss that cannot be reversed by conventional nutritional support. To uncover the molecular basis underlying the onset of cancer cachectic muscle wasting and establish an effective intervention against muscle loss, we used a cancer cachectic mouse model and examined the effects of aerobic exercise. Aerobic exercise successfully suppressed muscle atrophy and activated adiponectin signaling. Next, a cellular model for cancer cachectic muscle atrophy using C2C12 myotubes was prepared by treating myotubes with a conditioned medium from a culture of colon-26 cancer cells. Treatment of the atrophic myotubes with recombinant adiponectin was protective against the thinning of cells through the increased production of p-mTOR and suppression of LC3-II. Altogether, these findings suggest that the activation of adiponectin signaling could be part of the molecular mechanisms by which aerobic exercise ameliorates cancer cachexia-induced muscle wasting.
    Keywords:  adiponectin; aerobic exercise; cancer cachexia; muscle atrophy
  21. J Physiol. 2021 Mar 26.
      KEY POINTS: Muscle glycogen and intramuscular triglycerides (IMTG, stored in lipid droplets) are important energy substrates during prolonged exercise. Exercise-induced changes in lipid droplet (LD) morphology (i.e., LD size and number) has not yet been studied under nutritional conditions typically adopted by elite endurance athletes, that is, after carbohydrate (CHO) loading and CHO feeding during exercise. We report for the first time that exercise reduces IMTG content in both central and peripheral regions of type I and IIa fibres, reflective of decreased LD number in both fibre types whereas reductions in LD size was exclusive to type I fibres. Additionally, CHO feeding does not alter subcellular IMTG utilisation, LD morphology or muscle glycogen utilisation in type I or IIa/II fibres. In the absence of alterations to muscle fuel selection, CHO feeding does not attenuate cell signalling with regulatory roles in mitochondrial biogenesis.ABSTRACT: We examined the effects of carbohydrate (CHO) feeding on lipid droplet (LD) morphology, muscle glycogen utilisation and exercise-induced skeletal muscle cell signalling. After a 36 h CHO loading protocol and pre-exercise meal (12 and 2 g·kg-1 , respectively), eight trained males ingested 0, 45 or 90 g CHO·h-1 during 180 min cycling at lactate threshold followed by an exercise capacity test (150% lactate threshold). Muscle biopsies were obtained pre- and post-completion of submaximal exercise. Exercise decreased (P<0.01) glycogen concentration to comparable levels (∼700 to 250 mmol·kg-1 dw), though utilisation was greater in type I (∼40%) versus type II fibres (∼10%) (P<0.01). LD content decreased in type I (∼50%) and type IIa fibres (∼30%) (P<0.01) with greater utilisation in type I fibres (P<0.01). CHO feeding did not affect glycogen or IMTG utilisation in type I or II fibres (all P>0.05). Exercise decreased LD number within central and peripheral regions of both type I and IIa fibres, though reduced LD size was exclusive to type I fibres. Exercise induced (all P<0.05) comparable AMPKThr172 (∼4 fold), p53Ser15 (∼2 fold) and CaMKIIThr268 phosphorylation (∼2 fold) with no effects of CHO feeding (all P>0.05). CHO increased exercise capacity where 90 g·h-1 (233 ± 133 s) > 45 g·h-1 (156 ± 66 s; P = 0.06) > 0 g·h-1 (108 ± 54 s; P = 0.03). In conditions of high pre-exercise CHO availability, we conclude CHO feeding does not influence exercise-induced changes in LD morphology, glycogen utilisation or cell signalling pathways with regulatory roles in mitochondrial biogenesis. This article is protected by copyright. All rights reserved.
    Keywords:  glycogen; intramuscular triglyceride; vastus lateralis
  22. Nat Biomed Eng. 2021 Mar 29.
      Because post-mortem human skeletal muscle is not viable, autologous muscle grafts are typically required in tissue reconstruction after muscle loss due to disease or injury. However, the use of autologous tissue often leads to donor-site morbidity. Here, we show that intraspecies and interspecies chimaeric pig embryos lacking native skeletal muscle can be produced by deleting the MYF5, MYOD and MYF6 genes in the embryos via CRISPR, followed by somatic-cell nuclear transfer and the delivery of exogenous cells (porcine blastomeres or human induced pluripotent stem cells) via blastocyst complementation. The generated intraspecies chimaeras were viable and displayed normal histology, morphology and function. Human:pig chimaeras generated with TP53-null human induced pluripotent stem cells led to higher chimaerism efficiency, with embryos collected at embryonic days 20 and 27 containing humanized muscle, as confirmed by immunohistochemical and molecular analyses. Human:pig chimaeras may facilitate the production of exogenic organs for research and xenotransplantation.
  23. Int J Mol Sci. 2021 Mar 23. pii: 3265. [Epub ahead of print]22(6):
      Skeletal muscle regeneration is highly dependent on the inflammatory response. A wide variety of innate and adaptive immune cells orchestrate the complex process of muscle repair. This review provides information about the various types of immune cells and biomolecules that have been shown to mediate muscle regeneration following injury and degenerative diseases. Recently developed cell and drug-based immunomodulatory strategies are highlighted. An improved understanding of the immune response to injured and diseased skeletal muscle will be essential for the development of therapeutic strategies.
    Keywords:  T cell; immunomodulatory therapy; inflammation; macrophage; mesenchymal stem cell; myogenesis
  24. Antioxidants (Basel). 2021 Mar 30. pii: 537. [Epub ahead of print]10(4):
      Physical exercise represents a major challenge to whole-body homeostasis, provoking acute and adaptative responses at the cellular and systemic levels. Different sources of reactive oxygen species (ROS) have been described in skeletal muscle (e.g., NADPH oxidases, xanthine oxidase, and mitochondria) and are closely related to the physiological changes induced by physical exercise through the modulation of several signaling pathways. Many signaling pathways that are regulated by exercise-induced ROS generation, such as adenosine monophosphate-activated protein kinase (AMPK), mitogen activated protein kinase (MAPK), nuclear respiratory factor2 (NRF2), and PGC-1α are involved in skeletal muscle responses to physical exercise, such as increased glucose uptake, mitochondriogenesis, and hypertrophy, among others. Most of these adaptations are blunted by antioxidants, revealing the crucial role played by ROS during and after physical exercise. When ROS generation is either insufficient or exacerbated, ROS-mediated signaling is disrupted, as well as physical exercise adaptations. Thus, an understanding the limit between "ROS that can promote beneficial effects" and "ROS that can promote harmful effects" is a challenging question in exercise biology. The identification of new mediators that cause reductive stress and thereby disrupt exercise-stimulated ROS signaling is a trending on this topic and are covered in this current review.
    Keywords:  NADPH oxidase; ROS; exercise; oxidative stress; redox signaling; reductive stress; skeletal muscle
  25. Int J Mol Sci. 2021 Mar 13. pii: 2928. [Epub ahead of print]22(6):
      Pre-mRNA splicing plays an important role in muscle function and diseases. The RNA binding motif 20 (RBM20) is a splicing factor that is predominantly expressed in muscle tissues and primarily regulates pre-mRNA splicing of Ttn, encoding a giant muscle protein titin that is responsible for muscle function and diseases. RBM20-mediated Ttn splicing has been mostly studied in heart muscle, but not in skeletal muscle. In this study, we investigated splicing specificity in different muscle types in Rbm20 knockout rats and hormonal effects on RBM20-mediated splicing both in cellulo and in vivo studies. The results revealed that RBM20 is differentially expressed across muscles and RBM20-mediated splicing is muscle-type specific. In the presence of RBM20, Ttn splicing responds to hormones in a muscle-type dependent manner, while in the absence of RBM20, Ttn splicing is not affected by hormones. In differentiated and undifferentiated C2C12 cells, RBM20-mediated splicing in response to hormonal effects is mainly through genomic signaling pathway. The knowledge gained from this study may help further understand muscle-specific gene splicing in response to hormone stimuli in different muscle types.
    Keywords:  RBM20; hormones; pre-mRNA splicing; skeletal muscle; titin
  26. Int J Mol Sci. 2021 Mar 04. pii: 2558. [Epub ahead of print]22(5):
      The loss of skeletal muscle mass (muscle atrophy or wasting) caused by aging, diseases, and injury decreases quality of life, survival rates, and healthy life expectancy in humans. Although long non-coding RNAs (lncRNAs) have been implicated in skeletal muscle formation and differentiation, their precise roles in muscle atrophy remain unclear. In this study, we used RNA-sequencing (RNA-Seq) to examine changes in the expression of lncRNAs in four muscle atrophy conditions (denervation, casting, fasting, and cancer cachexia) in mice. We successfully identified 33 annotated lncRNAs and 18 novel lncRNAs with common expression changes in all four muscle atrophy conditions. Furthermore, an analysis of lncRNA-mRNA correlations revealed that several lncRNAs affected small molecule biosynthetic processes during muscle atrophy. These results provide novel insights into the lncRNA-mediated regulatory mechanism underlying muscle atrophy and may be useful for the identification of promising therapeutic targets.
    Keywords:  RNA-sequencing; long non-coding RNA; mRNA; skeletal muscle atrophy
  27. Antioxidants (Basel). 2021 Mar 27. pii: 524. [Epub ahead of print]10(4):
      Muscular aging is a complex process and underlying physiological mechanisms are not fully clear. In recent years, the participation of the NF-kB pathway and the NLRP3 inflammasome in the chronic inflammation process that accompanies the skeletal muscle's aging has been confirmed. microRNAs (miRs) form part of a gene regulatory machinery, and they control numerous biological processes including inflammatory pathways. In this work, we studied the expression of four miRs; three of them are considered as inflammatory-related miRs (miR-21, miR-146a, and miR-223), and miR-483, which is related to the regulation of melatonin synthesis, among other targets. To investigate the changes of miRs expression in muscle along aging, the impact of inflammation, and the role of melatonin in aged skeletal muscle, we used the gastrocnemius muscle of wild type (WT) and NLRP3-knockout (NLRP3-) mice of 3, 12, and 24 months-old, with and without melatonin supplementation. The expression of miRs and pro-caspase-1, caspase-3, pro-IL-1β, bax, bcl-2, and p53, was investigated by qRT-PCR analysis. Histological examination of the gastrocnemius muscle was also done. The results showed that age increased the expression of miR-21 (p < 0.01), miR-146a, and miR-223 (p < 0.05, for both miRs) in WT mice, whereas the 24-months-old mutant mice revealed decline of miR-21 and miR-223 (p < 0.05), compared to WT age. The lack of NLRP3 inflammasome also improved the skeletal muscle fibers arrangement and reduced the collagen deposits compared with WT muscle during aging. For the first time, we showed that melatonin significantly reduced the expression of miR-21, miR-146a, and miR-223 (p < 0.05 for all ones, and p < 0.01 for miR-21 at 24 months old) in aged WT mice, increased miR-223 in NLRP3- mice (p < 0.05), and induced miR-483 expression in both mice strains, this increase being significant at 24 months of age.
    Keywords:  NF-kB; NLRP3 inflammasome; aging; melatonin; microRNAs
  28. ACS Omega. 2021 Mar 23. 6(11): 7422-7433
      Aging is associated with loss of muscle mass and strength that leads to a condition termed sarcopenia. Impaired conditions, morbidity, and malnutrition are the factors of devaluation of muscle fibers in aged animals. Satellite cells play an important role in maintaining muscle homeostasis during tissue regeneration and repair. Proteomic profiling on the skeletal muscle tissues of different age group rats helps to determine the differentially expressed (DE) proteins, which may eventually lead to the development of biomarkers in treating the conditions of sarcopenia. In this study, nanoscale liquid chromatography coupled to tandem mass spectrometry (nano-LC-MS/MS) analysis was implemented in the calf tissues of young and old groups of rats. The mass spectrometry (MS) analysis revealed the presence of 335 differentially expressed proteins between the two different age conditions, among which those based on log-fold change 25 proteins were upregulated and 77 were downregulated. The protein-protein interaction network analysis revealed 18 upregulated proteins with three distinct interconnected networks and 57 downregulated proteins with two networks. Further, gene ontology (GO) enrichment analysis showed the biological process, cellular component, and molecular function of the differential proteins. Pathway enrichment analysis of the DE proteins identified nine significantly enriched pathways with a list of eight significant genes (Cryab, Hspb2, Acat1, Ak1, Adssl1, Anxa5, Gys1, Ogdh, Gc, and Adssl1). Quantification of significant genes by quantitative real-time polymerase chain reaction (qRT-PCR) confirmed the downregulation at the mRNA level. Western blot analysis of their protein expression showed concordant results on two candidate proteins (Ogdh and annexin 5) confirming their differential regulation between the two age groups of rats. Thus, these proteomic approaches on young and aged rats provide insights into the development of protein targets in the treatment of sarcopenia (muscle loss).
  29. Int J Mol Sci. 2021 Mar 09. pii: 2755. [Epub ahead of print]22(5):
      The SCN5A gene encodes the α-subunit of the voltage-gated cardiac sodium channel (NaV1.5), a key player in cardiac action potential depolarization. Genetic variants in protein-coding regions of the human SCN5A have been largely associated with inherited cardiac arrhythmias. Increasing evidence also suggests that aberrant expression of the SCN5A gene could increase susceptibility to arrhythmogenic diseases, but the mechanisms governing SCN5A expression are not yet well understood. To gain insights into the molecular basis of SCN5A gene regulation, we used rat gastrocnemius muscle four days following denervation, a process well known to stimulate Scn5a expression. Our results show that denervation of rat skeletal muscle induces the expression of the adult cardiac Scn5a isoform. RNA-seq experiments reveal that denervation leads to significant changes in the transcriptome, with Scn5a amongst the fifty top upregulated genes. Consistent with this increase in expression, ChIP-qPCR assays show enrichment of H3K27ac and H3K4me3 and binding of the transcription factor Gata4 near the Scn5a promoter region. Also, Gata4 mRNA levels are significantly induced upon denervation. Genome-wide analysis of H3K27ac by ChIP-seq suggest that a super enhancer recently described to regulate Scn5a in cardiac tissue is activated in response to denervation. Altogether, our experiments reveal that similar mechanisms regulate the expression of Scn5a in denervated muscle and cardiac tissue, suggesting a conserved pathway for SCN5A expression among striated muscles.
    Keywords:  H3K27 acetylation; Scn5a; cardiac arrhythmias; epigenetic mechanisms; histone modifications; skeletal muscle denervation; transcriptional regulation
  30. Cells. 2021 Mar 27. pii: 742. [Epub ahead of print]10(4):
      Diabetes promotes an angiostatic phenotype in the microvascular endothelium of skeletal muscle and skin. Angiogenesis-related microRNAs (angiomiRs) regulate angiogenesis through the translational repression of pro- and anti-angiogenic genes. The maturation of micro-RNA (miRs), including angiomiRs, requires the action of DROSHA and DICER proteins. While hyperglycemia modifies the expression of angiomiRs, it is unknown whether high glucose conditions alter the maturation process of angiomiRs in dermal and skeletal muscle microvascular endothelial cells (MECs). Compared to 5 mM of glucose, high glucose condition (30 mM, 6-24 h) decreased DROSHA protein expression, without changing DROSHA mRNA, DICER mRNA, or DICER protein in primary dermal MECs. Despite DROSHA decreasing, high glucose enhanced the maturation and expression of one angiomiR, miR-15a, and downregulated an miR-15a target: Vascular Endothelial Growth Factor-A (VEGF-A). The high glucose condition increased Murine Double Minute-2 (MDM2) expression and MDM2-binding to DROSHA. Inhibition of MDM2 prevented the effects evoked by high glucose on DROSHA protein and miR-15a maturation in dermal MECs. In db/db mice, blood glucose was negatively correlated with the expression of skeletal muscle DROSHA protein, and high glucose decreased DROSHA protein in skeletal muscle MECs. Altogether, our results suggest that high glucose reduces DROSHA protein and enhances the maturation of the angiostatic miR-15a through a mechanism that requires MDM2 activity.
    Keywords:  DROSHA; MDM2; VEGF-A; angiogenesis; angiostatic; dermal; hyperglycemia; micro-RNA (miRs); microvascular endothelial cells (MECs); skeletal muscle
  31. Int J Mol Sci. 2021 Mar 23. pii: 3254. [Epub ahead of print]22(6):
      Among multiple mechanisms, low-grade inflammation is critical for the development of insulin resistance as a feature of type 2 diabetes. The nucleotide-binding oligomerization domain-like receptor family (NOD-like) pyrin domain containing 3 (NLRP3) inflammasome has been linked to the development of insulin resistance in various tissues; however, its role in the development of insulin resistance in the skeletal muscle has not been explored in depth. Currently, there is limited evidence that supports the pathological role of NLRP3 inflammasome activation in glucose handling in the skeletal muscle of obese individuals. Here, we have centered our focus on insulin signaling in skeletal muscle, which is the main site of postprandial glucose disposal in humans. We discuss the current evidence showing that the NLRP3 inflammasome disturbs glucose homeostasis. We also review how NLRP3-associated interleukin and its gasdermin D-mediated efflux could affect insulin-dependent intracellular pathways. Finally, we address pharmacological NLRP3 inhibitors that may have a therapeutical use in obesity-related metabolic alterations.
    Keywords:  NALP3; chronic inflammation; glucose transport; muscle
  32. Int J Mol Sci. 2021 Mar 07. pii: 2692. [Epub ahead of print]22(5):
      microRNAs (miRNAs) regulate messenger RNA (mRNA) abundance and translation during key developmental processes including muscle differentiation. Assessment of miRNA targets can provide insight into muscle biology and gene expression profiles altered by disease. mRNA and miRNA libraries were generated from C2C12 myoblasts during differentiation, and predicted miRNA targets were identified based on presence of miRNA binding sites and reciprocal expression. Seventeen miRNAs were differentially expressed at all time intervals (comparing days 0, 2, and 5) of differentiation. mRNA targets of differentially expressed miRNAs were enriched for functions related to calcium signaling and sarcomere formation. To evaluate this relationship in a disease state, we evaluated the miRNAs differentially expressed in human congenital myotonic dystrophy (CMD) myoblasts and compared with normal control. Seventy-four miRNAs were differentially expressed during healthy human myocyte maturation, of which only 12 were also up- or downregulated in CMD patient cells. The 62 miRNAs that were only differentially expressed in healthy cells were compared with differentiating C2C12 cells. Eighteen of the 62 were conserved in mouse and up- or down-regulated during mouse myoblast differentiation, and their C2C12 targets were enriched for functions related to muscle differentiation and contraction.
    Keywords:  congenital myotonic dystrophy; microRNA; muscle differentiation
  33. Exp Gerontol. 2021 Mar 24. pii: S0531-5565(21)00095-4. [Epub ahead of print] 111320
      BACKGROUND: Deterioration of neuromuscular function is a major mechanism of age-related strength loss. Resistance training (RT) improves muscle strength and mass. However, the effects of RT on neuromuscular adaptations in middle-aged and older adults are unclear.METHODS: Randomised controlled RT interventions (≥2 weeks) involving adults aged ≥50 years were identified. Primary outcome measures were voluntary activation (VA), electromyographic (EMG) activity during maximal voluntary contraction (MVC), and antagonist coactivation. Data were pooled using a weighted random-effect model. Sub-analyses were conducted by muscle or muscle group and health status of participants. Sensitivity analysis was based on study quality. P < 0.05 indicated statistical significance.
    RESULTS: Twenty-seven studies were included. An effect was found for VA (standardised mean difference [SMD] 0.54, 0.01 to 1.07, P = 0.04), This result remained significant following sensitivity analysis involving only studies that were low risk of bias. Subgroup analyses showed an effect for plantar flexor VA (SMD 1.13, 0.20 to 2.06, P = 0.02) and VA in healthy participants (SMD 1.04, 0.32 to 1.76, P = 0.004). There was no effect for EMG activity or antagonist coactivation of any muscle group (P > 0.05).
    DISCUSSION: Resistance training did not alter EMG activity or antagonist coactivation in older adults. Sensitivity analysis resulted in the effect for VA remaining significant, indicating that this finding was not dependent on study quality. Studies predominantly involved healthy older adults (78%), limiting the generalisability of these findings to clinical cohorts. Future research should determine the effects of RT on neuromuscular function in people with sarcopenia and age-related syndromes.
    Keywords:  Ageing; Muscle activation; Neuromuscular; Resistance training
  34. Int J Mol Sci. 2021 Mar 11. pii: 2850. [Epub ahead of print]22(6):
      Mutations in the acidic alpha-glucosidase (GAA) coding gene cause Pompe disease. Late-onset Pompe disease (LOPD) is characterized by progressive proximal and axial muscle weakness and atrophy, causing respiratory failure. Enzyme replacement therapy (ERT), based on recombinant human GAA infusions, is the only available treatment; however, the efficacy of ERT is variable. Here we address the question whether proteins at variance in LOPD muscle of patients before and after 1 year of ERT, compared withhealthy age-matched subjects (CTR), reveal a specific signature. Proteins extracted from skeletal muscle of LOPD patients and CTR were analyzed by combining gel based (two-dimensional difference gel electrophoresis) and label-free (liquid chromatography-mass spectrometry) proteomic approaches, and ingenuity pathway analysis. Upstream regulators targeting autophagy and lysosomal tethering were assessed by immunoblotting. 178 proteins were changed in abundance in LOPD patients, 47 of them recovered normal level after ERT. Defects in oxidative metabolism, muscle contractile protein regulation, cytoskeletal rearrangement, and membrane reorganization persisted. Metabolic changes, ER stress and UPR (unfolded protein response) contribute to muscle proteostasis dysregulation with active membrane remodeling (high levels of LC3BII/LC3BI) and accumulation of p62, suggesting imbalance in the autophagic process. Active lysosome biogenesis characterizes both LOPD PRE and POST, unparalleled by molecules involved in lysosome tethering (VAMP8, SNAP29, STX17, and GORASP2) and BNIP3. In conclusion this study reveals a specific signature that suggests ERT prolongation and molecular targets to ameliorate patient's outcome.
    Keywords:  autophagy; mass spectrometry; pompe disease; proteomics; rare disease; sarcopenia
  35. Int J Mol Sci. 2021 Mar 09. pii: 2748. [Epub ahead of print]22(5):
      Palmitic acid (PA), a saturated fatty acid enriched in high-fat diet, has been implicated in the development of sarcopenic obesity. Herein, we chose two non-cytotoxic concentrations to better understand how excess PA could impact myotube formation or diameter without inducing cell death. Forty-eight hours of 100 µM PA induced a reduction of myotube diameter and increased the number of type I fibers, which was associated with increased miR-206 expression. Next, C2C12 myotube growth in the presence of PA was evaluated. Compared to control cells, 150 µM PA reduces myoblast proliferation and the expression of MyoD and miR-206 and miR-133a expression, leading to a reduced number and diameter of myotubes. PA (100 µM), despite not affecting proliferation, impairs myotube formation by reducing the expression of Myf5 and miR-206 and decreasing protein synthesis. Interestingly, 100 and 150 µM PA-treated myotubes had a higher number of type II fibers than control cells. In conclusion, PA affects negatively myotube diameter, fusion, and metabolism, which may be related to myomiRs. By providing new insights into the mechanisms by which PA affects negatively skeletal muscle, our data may help in the discovery of new targets to treat sarcopenic obesity.
    Keywords:  atrophy; fatty acids; microRNAs; obesity; skeletal muscle
  36. FEBS J. 2021 Apr 02.
      During development, resident stem cell populations contribute to the growth and maturation of tissue and organs. In skeletal muscle, muscle stem cells, or satellite cells, are responsible for the maturation of postnatal myofibers. However, the role satellite cells play in later stages of postnatal growth, and thus, when they enter a mature quiescent state is controversial. Here we discuss the current literature regarding the role satellite cells play in all stages of postnatal growth, from birth to puberty onset to young adulthood. We additionally highlight the implications of satellite cell loss or dysfunction during developmental stages, both in the context of experimental paradigms and disease settings.
    Keywords:  aging; cancer; development; pediatric; regeneration; satellite cells; skeletal muscle
  37. Int J Mol Sci. 2021 Mar 31. pii: 3645. [Epub ahead of print]22(7):
      Bone marrow-mesenchymal stem/stromal cells (MSCs) may offer promise for skeletal muscle repair/regeneration. Growing evidence suggests that the mechanisms underpinning the beneficial effects of such cells in muscle tissue reside in their ability to secrete bioactive molecules (secretome) with multiple actions. Hence, we examined the effects of MSC secretome as conditioned medium (MSC-CM) on ex vivo murine extensor digitorum longus muscle injured by forced eccentric contraction (EC). By combining morphological (light and confocal laser scanning microscopies) and electrophysiological analyses we demonstrated the capability of MSC-CM to attenuate EC-induced tissue structural damages and sarcolemnic functional properties' modifications. MSC-CM was effective in protecting myofibers from apoptosis, as suggested by a reduced expression of pro-apoptotic markers, cytochrome c and activated caspase-3, along with an increase in the expression of pro-survival AKT factor. Notably, MSC-CM also reduced the EC-induced tissue redistribution and extension of telocytes/CD34+ stromal cells, distinctive cells proposed to play a "nursing" role for the muscle resident myogenic satellite cells (SCs), regarded as the main players of regeneration. Moreover, it affected SC functionality likely contributing to replenishment of the SC reservoir. This study provides the necessary groundwork for further investigation of the effects of MSC secretome in the setting of skeletal muscle injury and regenerative medicine.
    Keywords:  biophysical properties; eccentric contraction; mesenchymal stem/stromal cell (MSC); repair/regeneration; satellite cells; secretome; skeletal muscle; stem cell; telocytes/CD34+ stromal cells
  38. Ann Med. 2021 Dec;53(1): 508-522
      Background: Recent evidence indicates that host-gut microbiota crosstalk has nonnegligible effects on host skeletal muscle, yet gut microbiota-regulating mechanisms remain obscure.Methods: C57BL/6 mice were treated with a cocktail of antibiotics (Abx) to depress gut microbiota for 4 weeks. The profiles of gut microbiota and microbial bile acids were measured by 16S rRNA sequencing and ultra-performance liquid chromatography (UPLC), respectively. We performed qPCR, western blot and ELISA assays in different tissue samples to evaluate FXR-FGF15/19 signaling.Results: Abx treatment induced skeletal muscle atrophy in mice. These effects were associated with microbial dysbiosis and aberrant bile acid (BA) metabolism in intestine. Ileal farnesoid X receptor (FXR)-fibroblast growth factor 15 (FGF15) signaling was inhibited in response to microbial BA disturbance. Mechanistically, circulating FGF15 was decreased, which downregulated skeletal muscle protein synthesis through the extracellular-signal-regulated protein kinase 1/2 (ERK1/2) signaling pathway. Treating Abx mice with FGF19 (human FGF15 ortholog) partly reversed skeletal muscle loss.Conclusions: These findings indicate that the BA-FXR-FGF15/19 axis acts as a regulator of gut microbiota to mediate host skeletal muscle.
    Keywords:  FGF15/19; FXR; Gut microbiota; bile acid; skeletal muscle
  39. Diabetes Metab Syndr Obes. 2021 ;14 1233-1241
      Purpose: Skeletal muscle has a major influence on whole-body metabolic homeostasis. In the present study, we aimed to determine the metabolic effects of the β3 adrenergic receptor agonist CL316243 (CL) in the skeletal muscle of high-fat diet-fed rats.Methods: Sprague-Dawley rats were randomly allocated to three groups, which were fed a control diet (C) or a high-fat diet (HF), and half of the latter were administered 1 mg/kg CL by gavage once weekly (HF+CL), for 12 weeks. At the end of this period, the serum lipid profile and glucose tolerance of the rats were evaluated. In addition, the phosphorylation and protein and mRNA expression of AMP-activated protein kinase (AMPK), peroxisome proliferator-activated receptor γ coactivator (PGC)-1α, and carnitine palmitoyl transferase (CPT)-1b in skeletal muscle were measured by Western blot analysis and qPCR. The direct effects of CL on the phosphorylation (p-) and expression of AMPK, PGC-1α, and CPT-1b were also evaluated by Western blotting and immunofluorescence in L6 myotubes.
    Results: CL administration ameliorated the abnormal lipid profile and glucose tolerance of the high-fat diet-fed rats. In addition, the expression of p-AMPK, PGC-1α, and CPT-1b in the soleus muscle was significantly increased by CL. CL (1 µM) also increased the protein expression of p-AMPK, PGC-1α, and CPT-1b in L6 myotubes. However, the effect of CL on PGC-1α protein expression was blocked by the AMPK antagonist compound C, which suggests that CL increases PGC-1α protein expression via AMPK.
    Conclusion: Activation of the β3 adrenergic receptor in skeletal muscle ameliorates the metabolic abnormalities of high-fat diet-fed rats, at least in part via activation of the AMPK/PGC-1α pathway.
    Keywords:  AMPK; CL316243; L6 myotubes; PGC-1α; carnitine palmitoyl transferase
  40. Regen Eng Transl Med. 2020 Dec;6(4): 365-372
      Volumetric muscle loss (VML) is traumatic, degenerative, or surgical loss of skeletal muscle that exceeds the regenerative capacity of the remaining muscle, thus resulting in impaired muscle function. In humans, the loss of 30% or more mass of any one muscle will result in permanent structural and functional loss. Current VML repair treatments are limited by donor site morbidity and graft tissue availability, necessitating alternative muscle graft sources. To address this need, our lab has fabricated tissue-engineered skeletal muscle units (SMUs) for implantation into a 30 % VML model in the tibialis anterior (TA) muscle of rat. Previous results showed that after 28 days in vivo, muscle with a 30% VML repaired with our SMUs produced significantly more force than muscle with acute VML. But repair with our SMU did not fully restore muscle force production to that of native muscle. Thus, we hypothesized that more time for in vivo tissue regeneration would allow for greater force recovery. Therefore, the purpose of this study was to examine the long-term (3-month) effects of our SMUs on a 30% VML repair. We also assessed the effects of reinnervation by redirecting a branch of the peroneal nerve to the repair site. Thirty-nine, 2-month old female F344 rats were separated into a nonsurgical control group (n=5) and four surgical experimental groups (VML Only, n=5; VML+Nerve Redirect, n=6; VML+SMU, n=5; VML+SMU+ Nerve Redirect, n=8). Experimental rats were allowed a 3-month recovery period post-surgery before undergoing in situ force testing of the surgical (left) TA. The left TA of the control animals also underwent in situ force testing. Finally, the surgical (left) and contralateral (right) TAs of the experimental animals, as well as the left TA of the control animals, were explanted for histological analysis. Results for specific force showed that while all groups recovered specific forces similar to that of native muscle, the two SMU groups had significantly higher specific forces, on average, compared to the uninjured control group. Histological staining showed small muscle fibers in the repair site in animals that received an SMU. The average cross-sectional area of the native fibers just outside the area of repair (or the equivalent area in control animals) was not significantly different between groups, indicating that hypertrophy of remaining fibers did not contribute to the recovery of force following the VML. Our results suggest that following a 30% VML of the TA muscle, all surgical groups were able to recover TA mass, maximum tetanic and specific force production. Thus, creating a 30% VML in the TA in a rat model is not enough a sufficient VML to produce the sustained VML seen in humans following similar 30% loss of muscle volume.
    Keywords:  nerve redirect; satellite cell; scaffold-free; skeletal muscle; tissue engineering; volumetric muscle loss
  41. Cells. 2021 Mar 02. pii: 525. [Epub ahead of print]10(3):
      Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disorder characterized by the selective degeneration of upper and lower motor neurons and by the progressive weakness and paralysis of voluntary muscles. Despite intense research efforts and numerous clinical trials, it is still an incurable disease. ALS had long been considered a pure motor neuron disease; however, recent studies have shown that motor neuron protection is not sufficient to prevent the course of the disease since the dismantlement of neuromuscular junctions occurs before motor neuron degeneration. Skeletal muscle alterations have been described in the early stages of the disease, and they seem to be mainly involved in the "dying back" phenomenon of motor neurons and metabolic dysfunctions. In recent years, skeletal muscles have been considered crucial not only for the etiology of ALS but also for its treatment. Here, we review clinical and preclinical studies that targeted skeletal muscles and discuss the different approaches, including pharmacological interventions, supplements or diets, genetic modifications, and training programs.
    Keywords:  amyotrophic lateral sclerosis; genetic intervention; pharmacological approaches; physical activity; skeletal muscle
  42. Front Genet. 2021 ;12 640807
      microRNAs are a kind of endogenous, non-coding, single-strand small RNA. They have been reported as an important regulatory factor in skeletal myogenesis. In this study, miR-452 was selected from RNA high-throughput sequencing data to explore its regulatory role in myogenesis. Functionally, miR-452 overexpression could promote C2C12 myoblast proliferation while inhibiting myogenic differentiation. On the contrary, inhibition of miR-452 could suppress C2C12 myoblast proliferation but accelerate myogenic differentiation. Bioinformatics analysis and dual luciferase report assays showed that Angiopoietin 1 (ANGPT1), RB1, and CACNB4 were the potential target genes of miR-452. To further confirm the target relationship between ANGPT1, RB1, and CACNB4 with miR-452, the mRNA level and protein level of these genes were detected by using RT-qPCR and Western blot, respectively. Result analysis indicated that ANGPT1 was a target gene of miR-452. In addition, knockdown of ANGPT1 could obviously promote C2C12 myoblast proliferation but block their differentiation. In summary, these results demonstrated that miR-452 promoted C2C12 myoblast proliferation and inhibited their differentiation via targeting ANGPT1.
    Keywords:  ANGPT1; C2C12 myoblasts; differentiation; miR-452; proliferation
  43. Cell Biosci. 2021 Mar 31. 11(1): 65
      Extracellular matrix (ECM) is a kind of connective tissue in the cell microenvironment, which is of great significance to tissue development. ECM in muscle fiber niche consists of three layers: the epimysium, the perimysium, and the endomysium (basal lamina). These three layers of connective tissue structure can not only maintain the morphology of skeletal muscle, but also play an important role in the physiological functions of muscle cells, such as the transmission of mechanical force, the regeneration of muscle fiber, and the formation of neuromuscular junction. In this paper, detailed discussions are made for the structure and key components of ECM in skeletal muscle tissue, the role of ECM in skeletal muscle development, and the application of ECM in biomedical engineering. This review will provide the reader with a comprehensive overview of ECM, as well as a comprehensive understanding of the structure, physiological function, and application of ECM in skeletal muscle tissue.
    Keywords:  Application; Cell; ECM; Function; Skeletal muscle; Structure
  44. Nat Commun. 2021 Apr 01. 12(1): 2014
      Age-associated changes in gene expression in skeletal muscle of healthy individuals reflect accumulation of damage and compensatory adaptations to preserve tissue integrity. To characterize these changes, RNA was extracted and sequenced from muscle biopsies collected from 53 healthy individuals (22-83 years old) of the GESTALT study of the National Institute on Aging-NIH. Expression levels of 57,205 protein-coding and non-coding RNAs were studied as a function of aging by linear and negative binomial regression models. From both models, 1134 RNAs changed significantly with age. The most differentially abundant mRNAs encoded proteins implicated in several age-related processes, including cellular senescence, insulin signaling, and myogenesis. Specific mRNA isoforms that changed significantly with age in skeletal muscle were enriched for proteins involved in oxidative phosphorylation and adipogenesis. Our study establishes a detailed framework of the global transcriptome and mRNA isoforms that govern muscle damage and homeostasis with age.
  45. Appl Physiol Nutr Metab. 2021 Mar 27.
      We investigated the effects of the acute and chronic exercise, prescribed in different intensity zones, but with total load-matched on mitochondrial markers (COX-IV, Tfam, and citrate synthase (CS) activity in skeletal muscles, heart, and liver), glycogen stores, aerobic capacity and anaerobic index in swimming rats. For this, two experimental designs were performed (acute and chronic efforts). Load-matched exercises were prescribed below and above and on the anaerobic threshold (AnT), determined by the Lactate Minimum test. In chronic programs, two training prescription strategies were assessed (monotonous and linear periodized model). Results show changes in glycogen stores but no modification in the COX-IV and Tfam contents after acute exercises. In the chronic protocols, COX-IV and Tfam proteins and CS adaptations were intensity and tissue dependents. Monotonous training promoted better adaptations than the periodized model. Training at 80% of the AnT improved both performance variables, emphasizing the anaerobic index, concomitant to CS and COX-IV improvement (soleus muscle). The aerobic capacity and CS activity (gastrocnemius) were increased after 120% AnT training. In conclusion, acute exercise protocol did not promote responses in mitochondrial target proteins. An intensity and tissue dependence are reported in the chronic protocols, highlighting training at 80 and 120% of the AnT. Novelty: • Load-matched acute exercise did not enhance COX-IV and Tfam contents in skeletal muscles, heart, and liver. • In chronic exercise, COX-IV, Tfam, and citrate synthase activity adaptations were intensity and tissue dependents. •Monotonous training was more efficient than the periodized linear model in adaptations of target proteins and enzymatic activity.
  46. Int J Mol Sci. 2021 Mar 23. pii: 3252. [Epub ahead of print]22(6):
      Insufficient stress response and elevated oxidative stress can contribute to skeletal muscle atrophy during mechanical unloading (e.g., spaceflight and bedrest). Perturbations in heat shock proteins (e.g., HSP70), antioxidant enzymes, and sarcolemmal neuronal nitric oxidase synthase (nNOS) have been linked to unloading-induced atrophy. We recently discovered that the sarcolemmal NADPH oxidase-2 complex (Nox2) is elevated during unloading, downstream of angiotensin II receptor 1, and concomitant with atrophy. Here, we hypothesized that peptidyl inhibition of Nox2 would attenuate disruption of HSP70, MnSOD, and sarcolemmal nNOS during unloading, and thus muscle fiber atrophy. F344 rats were divided into control (CON), hindlimb unloaded (HU), and hindlimb unloaded +7.5 mg/kg/day gp91ds-tat (HUG) groups. Unloading-induced elevation of the Nox2 subunit p67phox-positive staining was mitigated by gp91ds-tat. HSP70 protein abundance was significantly lower in HU muscles, but not HUG. MnSOD decreased with unloading; however, MnSOD was not rescued by gp91ds-tat. In contrast, Nox2 inhibition protected against unloading suppression of the antioxidant transcription factor Nrf2. nNOS bioactivity was reduced by HU, an effect abrogated by Nox2 inhibition. Unloading-induced soleus fiber atrophy was significantly attenuated by gp91ds-tat. These data establish a causal role for Nox2 in unloading-induced muscle atrophy, linked to preservation of HSP70, Nrf2, and sarcolemmal nNOS.
    Keywords:  HSP70; MnSOD; NADPH oxidase; Nrf2; atrophy; nNOS; oxidative stress; skeletal muscle; unloading
  47. Cell Metab. 2021 Mar 23. pii: S1550-4131(21)00112-1. [Epub ahead of print]
      Neurodegeneration in the central nervous system (CNS) is a defining feature of organismal aging that is influenced by peripheral tissues. Clinical observations indicate that skeletal muscle influences CNS aging, but the underlying muscle-to-brain signaling remains unexplored. In Drosophila, we find that moderate perturbation of the proteasome in skeletal muscle induces compensatory preservation of CNS proteostasis during aging. Such long-range stress signaling depends on muscle-secreted Amyrel amylase. Mimicking stress-induced Amyrel upregulation in muscle reduces age-related accumulation of poly-ubiquitinated proteins in the brain and retina via chaperones. Preservation of proteostasis stems from the disaccharide maltose, which is produced via Amyrel amylase activity. Correspondingly, RNAi for SLC45 maltose transporters reduces expression of Amyrel-induced chaperones and worsens brain proteostasis during aging. Moreover, maltose preserves proteostasis and neuronal activity in human brain organoids challenged by thermal stress. Thus, proteasome stress in skeletal muscle hinders retinal and brain aging by mounting an adaptive response via amylase/maltose.
    Keywords:  aging; amylase; brain organoids; maltose; muscle-to-brain signaling; muscle-to-retina signaling; myokine; proteasome; proteostasis; stress response
  48. Front Physiol. 2021 ;12 640973
      Background: Intensive care patients commonly develop muscle wasting and functional impairment. However, the role of severe COVID-19 in the magnitude of muscle wasting and functionality in the acute critical disease is unknown. Objective: To perform a prospective characterization to evaluate the skeletal muscle mass and functional performance in intensive care patients with severe COVID-19. Methods: Thirty-two critically ill patients (93.8% male; age: 64.1 ± 12.6 years) with the diagnosis of the severe COVID-19 were prospectively recruited within 24 to 72 h following intensive care unit (ICU) admission, from April 2020 to October 2020, at Hospital Sírio-Libanês in Brazil. Patients were recruited if older than 18 years old, diagnosis of severe COVID-19 confirmed by RT-PCR, ICU stay and absence of limb amputation. Muscle wasting was determined through an ultrasound measurement of the rectus femoris cross-sectional area, the thickness of the anterior compartment of the quadriceps muscle (rectus femoris and vastus intermedius), and echogenicity. The peripheral muscle strength was assessed with a handgrip test. The functionality parameter was determined through the ICU mobility scale (IMS) and the International Classification of Functioning, Disability and Health (ICF). All evaluations were performed on days 1 and 10. Results: There were significant reductions in the rectus femoris cross-section area (-30.1% [95% IC, -26.0% to -34.1%]; P < 0.05), thickness of the anterior compartment of the quadriceps muscle (-18.6% [95% IC, -14.6% to 22.5%]; P < 0.05) and handgrip strength (-22.3% [95% IC, 4.7% to 39.9%]; P < 0.05) from days 1 to 10. Patients showed increased mobility (0 [0-5] vs 4.5 [0-8]; P < 0.05), improvement in respiratory function (3 [3-3] vs 2 [1-3]; P < 0.05) and structure respiratory system (3 [3-3] vs 2 [1-3]; P < 0.05), but none of the patients returned to normal levels. Conclusion: In intensive care patients with severe COVID-19, muscle wasting and decreased muscle strength occurred early and rapidly during 10 days of ICU stay with improved mobility and respiratory functions, although they remained below normal levels. These findings may provide insights into skeletal muscle wasting and function in patients with severe COVID-19.
    Keywords:  COVID-19; SARS-CoV-2 (2019-nCoV); function; functionality; muscle; ultrasound; wasting
  49. PLoS One. 2021 ;16(4): e0249601
      Skeletal muscle tissue has a highly complex and heterogeneous structure comprising several physical length scales. In the simplest model of muscle tissue, it can be represented as a one dimensional nonlinear spring in the direction of muscle fibres. However, at the finest level, muscle tissue includes a complex network of collagen fibres, actin and myosin proteins, and other cellular materials. This study shall derive an intermediate physical model which encapsulates the major contributions of the muscle components to the elastic response apart from activation-related along-fibre responses. The micro-mechanical factors in skeletal muscle tissue (eg. connective tissue, fluid, and fibres) can be homogenized into one material aggregate that will capture the behaviour of the combination of material components. In order to do this, the corresponding volume fractions for each type of material need to be determined by comparing the stress-strain relationship for a volume containing each material. This results in a model that accounts for the micro-mechanical features found in muscle and can therefore be used to analyze effects of neuro-muscular diseases such as cerebral palsy or muscular dystrophies. The purpose of this study is to construct a model of muscle tissue that, through choosing the correct material parameters based on experimental data, will accurately capture the mechanical behaviour of whole muscle. This model is then used to look at the impacts of the bulk modulus and material parameters on muscle deformation and strain energy-density distributions.