bims-moremu 2021-09-19 papers

bims-moremu

Biomed News

on Molecular regulators of muscle mass

Issue of 2021‒09‒19
forty papers selected by
Anna Vainshtein
Craft Science Inc.

Filopodia powered by class X myosin promote fusion of mammalian myoblasts.
Sympathetic activity is correlated with satellite cell aging and myogenesis via β2-adrenoceptor.
Mitochondrial-derived peptides and exercise.
A Coupled Mechanobiological Model of Muscle Regeneration In Cerebral Palsy.
Lactate administration does not affect denervation-induced loss of mitochondrial content and muscle mass in mice.
Elucidating exercise-induced skeletal muscle signaling pathways and applying relevant findings to preemptive therapy for lifestyle-related diseases.
Cannabinoid receptor 1 expression is higher in muscle of old vs. young males, and increases upon resistance exercise in older adults.
Sepsis induces muscle atrophy by inhibiting proliferation and promoting apoptosis via PLK1-AKT signalling.
CRISPR/Cas9/AAV9-mediated in vivo editing identifies MYC regulation of 3D genome in skeletal muscle stem cell.
Shared and Distinct Mechanisms of Skeletal Muscle Atrophy: A Narrative Review.
The missing "link"? β-Catenin's role in skeletal muscle glucose uptake during exercise & contraction.
Ablation of USP21 in skeletal muscle promotes oxidative fibre phenotype, inhibiting obesity and type 2 diabetes.
FTO-mediated demethylation of GADD45B promotes myogenesis through the activation of p38 MAPK pathway.
Deciphering the chromatin organization and dynamics for muscle stem cell function.
Effect of dietary L-theanine supplementation on skeletal muscle fiber type transformation in vivo.
Protein Requirements for Master Athletes: Just Older Versions of Their Younger Selves.
Dynamin-related protein 1 regulates substrate oxidation in skeletal muscle by stabilizing cellular and mitochondrial calcium dynamics.
Androgen receptor regulates the proliferation of myoblasts under appropriate or excessive stretch through IGF-1 receptor mediated p38 and ERK1/2 pathways.
Gene Therapy for Duchenne Muscular Dystrophy.
Reduced mitochondrial fission and impaired energy metabolism in human primary skeletal muscle cells of Megaconial Congenital Muscular Dystrophy.
Biofabrication of 3D Human Muscle Model with Vascularization and Endomysium.
Generation and Characterization of a Skeletal Muscle Cell-Based Model Carrying One Single Gne Allele: Implications in Actin Dynamics.
Two Types of Mouse Models for Sarcopenia Research: Senescence Acceleration and Genetic Modification Models.
Simple derivation of skeletal muscle from human pluripotent stem cells using temperature-sensitive Sendai virus vector.
Visualizing Muscle Sialic Acid Expression in the GNED207VTgGne-/- Cmah-/- Model of GNE Myopathy: A Comparison of Dietary and Gene Therapy Approaches.
Fish Oil for Healthy Aging: Potential Application to Master Athletes.
Designer AAV muscle up.
MyoData: An expression knowledgebase at single cell/nucleus level for the discovery of coding-noncoding RNA functional interactions in skeletal muscle.
Myostatin/Activin Receptor Ligands In Muscle And The Development Status Of Attenuating Drugs.
Malat-1-PRC2-EZH1 interaction supports adaptive oxidative stress dependent epigenome remodeling in skeletal myotubes.
Systemic delivery of AAVrh74.tMCK.hCAPN3 rescues the phenotype in a mouse model for LGMD2A/R1.
Effects of microgravity exposure and fructo-oligosaccharide ingestion on the proteome of soleus and extensor digitorum longus muscles in developing mice.
Duchenne muscular dystrophy cell culture models created by CRISPR/Cas9 gene editing and their application in drug screening.
A revised model for mitochondrial dysfunction in Duchenne muscular dystrophy.
Therapies for RYR1-Related Myopathies: Where We Stand and the Perspectives.
Understanding the gut microbiota and sarcopenia: a systematic review.
Cross-Species and Human Inter-Tissue Network Analysis of Genes Implicated in Longevity and Aging Reveal Strong Support for Nutrient Sensing.
Resistance Exercise Trims the Fat and Puts Some Muscle into Cancer Survivorship.
Full-length Dystrophin Restoration via Targeted Genomic Integration by AAV-CRISPR in a Humanized Mouse Model of Duchenne Muscular Dystrophy.
Dietary lysine restriction induces lipid accumulation in skeletal muscle through an increase in serum threonine levels in rats.

Elife. 2021 Sep 14. pii: e72419. [Epub ahead of print]10

Filopodia powered by class X myosin promote fusion of mammalian myoblasts.

David W Hammers, Cora C Hart, Michael K Matheny, Ernest G Heimsath, Young Il Lee, John A Hammer, Richard E Cheney, H Lee Sweeney.

  Skeletal muscle fibers are multinucleated cellular giants formed by the fusion of mononuclear myoblasts. Several molecules involved in myoblast fusion have been discovered, and finger-like projections coincident with myoblast fusion have also been implicated in the fusion process. The role of these cellular projections in muscle cell fusion was investigated herein. We demonstrate that these projections are filopodia generated by class X myosin (Myo10), an unconventional myosin motor protein specialized for filopodia. We further show that Myo10 is highly expressed by differentiating myoblasts, and Myo10 ablation inhibits both filopodia formation and myoblast fusion in vitro. In vivo, Myo10 labels regenerating muscle fibers associated with Duchenne muscular dystrophy and acute muscle injury. In mice, conditional loss of Myo10 from muscle-resident stem cells, known as satellite cells, severely impairs postnatal muscle regeneration. Furthermore, the muscle fusion proteins Myomaker and Myomixer are detected in myoblast filopodia. These data demonstrate that Myo10-driven filopodia facilitate multi-nucleated mammalian muscle formation.

Keywords:  cell biology; human; mouse; regenerative medicine; stem cells

DOI:  https://doi.org/10.7554/eLife.72419
Stem Cell Res Ther. 2021 Sep 16. 12(1): 505

Sympathetic activity is correlated with satellite cell aging and myogenesis via β2-adrenoceptor.

Shiguo Yuan, Sheng Zheng, Kai Zheng, Yanping Gao, Meixiong Chen, Yikai Li, Xiaochun Bai.

  BACKGROUND AND OBJECTIVE: Sympathetic activity plays an important role in the proliferation and differentiation of stem cells, and it changes over time, thereby exerting differential effects on various stem cell types. Aging causes sympathetic hyperactivity in aged tissues and blunts sympathetic nerves regulation, and sympathetic abnormalities play a role in aging-related diseases. Currently, the effect of sympathetic activity on skeletal muscle stem cells, namely satellite cells (SCs), is unclear. This study aimed to investigate the effects of skeletal muscle sympathetic activity on SC aging and skeletal muscle repair.MATERIALS AND METHODS: To evaluate skeletal muscle and fibrotic areas, numbers of SCs and myonuclei per muscle fiber, β2-adrenoceptor (β2-ADR) expression, muscle repair, and sympathetic innervation in skeletal muscle, aged mice, young mice that underwent chemical sympathectomy (CS) were utilized. Mice with a tibialis anterior muscle injury were treated by barium chloride (BaCl2) and clenbuterol (CLB) in vivo. SCs or C2C12 cells were differentiated into myotubes and treated with or without CLB. Immunofluorescence, western blot, sirius red, and hematoxylin-eosin were used to evaluate SCs, myogenic repair and differentiation.
RESULTS: The number of SCs, sympathetic activity, and reparability of muscle injury in aged mice were significantly decreased, compared with those in young mice. The above characteristics of young mice that underwent CS were similar to those of aged mice. While CLB promoted the repair of muscle injury in aged and CS mice and ameliorated the reduction in the SC number and sympathetic activity, the effects of CLB on the SCs and sympathetic nerves in young mice were not significant. CLB inhibited the myogenic differentiation of C2C12 cells in vitro. We further found that NF-κB and ERK1/2 signaling pathways were activated during myogenic differentiation, and this process could be inhibited by CLB.
CONCLUSION: Normal sympathetic activity promoted the stemness of SCs to thereby maintain a steady state. It also could maintain total and self-renewing number of SCs and maintain a quiescent state, which was correlated with skeletal SCs via β2-ADR. Normal sympathetic activity was also beneficial for the myogenic repair of injured skeletal muscle.

Keywords:  Clenbuterol; Skeletal muscle aging; Skeletal satellite cells; Sympathetic nerve; β-Adrenoceptor

DOI:  https://doi.org/10.1186/s13287-021-02571-8
Biochim Biophys Acta Gen Subj. 2021 Sep 11. pii: S0304-4165(21)00170-7. [Epub ahead of print] 130011

Mitochondrial-derived peptides and exercise.

Jonathan S T Woodhead, Troy L Merry.

  Acute exercise, and in particular aerobic exercise, increases skeletal muscle energy demand causing mitochondrial stress, and mitochondrial-related adaptations which are a hallmark of exercise training. Given that mitochondria are central players in the exercise response, it is imperative that they have networks that can communicate their status both intra- and inter-cellularly. Peptides encoded by short open-reading frames within mitochondrial DNA, mitochondrial-derived peptides (MDPs), have been suggested to form a newly recognised branch of this retrograde signalling cascade that contribute to coordinating the adaptive response to regular exercise. Here we summarise the recent evidence that acute high intensity exercise in humans can increase concentrations of the MDPs humanin and MOTS-c in skeletal muscle and plasma, and speculate on the mechanisms controlling MDP responses to exercise stress. Evidence that exercise training results in chronic changes in MDP expression within tissues and the circulation is conflicting and may depend on the mode, duration, intensity of training plan and participant characteristics. Further research is required to define the effect of these variables on MDPs and to determine whether MDPs other than MOTS-c have exercise mimetic properties. MOTS-c treatment of young and aged mice improves exercise capacity/performance and leads to adaptions that are similar to that of being physically active (weight loss, increased antioxidant capacity and improved insulin sensitivity), however, studies utilising a MOTS-c inactivating genetic variant or combination of exercise + MOTS-c treatment in mice suggest that there are distinct and overlapping pathways through which exercise and MOTS-c evoke metabolic benefits. Overall, MOTS-c, and potentially other MDPs, may be exercise-sensitive myokines and further work is required to define inter- and intra-tissue targets in an exercise context.

Keywords:  Cellular signalling; Hormesis; Mitochondria; Muscle; Physical activity; ROS

DOI:  https://doi.org/10.1016/j.bbagen.2021.130011
Front Bioeng Biotechnol. 2021 ;9 689714

A Coupled Mechanobiological Model of Muscle Regeneration In Cerebral Palsy.

Stephanie Khuu, Justin W Fernandez, Geoffrey G Handsfield.

  Cerebral palsy is a neuromusculoskeletal disorder associated with muscle weakness, altered muscle architecture, and progressive musculoskeletal symptoms that worsen with age. Pathological changes at the level of the whole muscle have been shown; however, it is unclear why this progression of muscle impairment occurs at the cellular level. The process of muscle regeneration is complex, and the interactions between cells in the muscle milieu should be considered in the context of cerebral palsy. In this work, we built a coupled mechanobiological model of muscle damage and regeneration to explore the process of muscle regeneration in typical and cerebral palsy conditions, and whether a reduced number of satellite cells in the cerebral palsy muscle environment could cause the muscle regeneration cycle to lead to progressive degeneration of muscle. The coupled model consisted of a finite element model of a muscle fiber bundle undergoing eccentric contraction, and an agent-based model of muscle regeneration incorporating satellite cells, inflammatory cells, muscle fibers, extracellular matrix, fibroblasts, and secreted cytokines. Our coupled model simulated damage from eccentric contraction followed by 28 days of regeneration within the muscle. We simulated cyclic damage and regeneration for both cerebral palsy and typically developing muscle milieus. Here we show the nonlinear effects of altered satellite cell numbers on muscle regeneration, where muscle repair is relatively insensitive to satellite cell concentration above a threshold, but relatively sensitive below that threshold. With the coupled model, we show that the fiber bundle geometry undergoes atrophy and fibrosis with too few satellite cells and excess extracellular matrix, representative of the progression of cerebral palsy in muscle. This work uses in silico modeling to demonstrate how muscle degeneration in cerebral palsy may arise from the process of cellular regeneration and a reduced number of satellite cells.

Keywords:  FEM; agent-based modeling; finite element modeling; mechanobiology; satellite cell; skeletal muscle

DOI:  https://doi.org/10.3389/fbioe.2021.689714
FEBS Open Bio. 2021 Sep 12.

Lactate administration does not affect denervation-induced loss of mitochondrial content and muscle mass in mice.

Kenya Takahashi, Yu Kitaoka, Yutaka Matsunaga, Hideo Hatta.

  Lactate is considered to be a signaling molecule that induces mitochondrial adaptation and muscle hypertrophy. The purpose of this study was to examine whether lactate administration attenuates denervation-induced loss of mitochondrial content and muscle mass. Eight-week-old male Institute of Cancer Research (ICR) mice underwent unilateral sciatic nerve transection surgery. The contralateral hindlimb served as a sham-operated control. From the day of surgery, mice were injected intraperitoneally with phosphate-buffered saline (PBS) or sodium lactate (equivalent to 1 g/kg body weight) once daily for 9 days. After 10 days of denervation, gastrocnemius muscle weight decreased to a similar extent in both the PBS- and lactate-injected groups. Denervation significantly decreased mitochondrial enzyme activity, protein content, and MCT4 protein content in the gastrocnemius muscle. However, lactate administration did not have any significant effects. The current observations suggest that daily lactate administration for 9 days does not affect denervation-induced loss of mitochondrial content and muscle mass.

Keywords:  denervation; lactate; mitochondria; skeletal muscle

DOI:  https://doi.org/10.1002/2211-5463.13293
Endocr J. 2021 Sep 11.

Elucidating exercise-induced skeletal muscle signaling pathways and applying relevant findings to preemptive therapy for lifestyle-related diseases.

Masato Iwabu, Miki Okada-Iwabu, Takashi Kadowaki, Toshimasa Yamauchi.

  While it is well recognized that exercise represents a radical preventive and therapeutic measure for lifestyle-related diseases, it is clear that contemporary lifestyles abound with situations where exercise may be found difficult to implement on a continuous basis. Indeed, this has led to global expectations for elucidation of the exercise-activated skeletal muscle signaling pathways as well as for development of exercise mimics that effectively activate such pathways. It is shown that exercise activates the transcriptional coactivator PGC-1α via AMPK/SIRT1 in muscle, thereby not only enhancing mitochondrial function and muscle endurance but upregulating energy metabolism. Further, adipocyte-derived adiponectin is also shown to activate AMPK/SIRT1/PGC-1α via its receptor AdipoR1 in skeletal muscles. Thus, adiponectin/AdipoR1 signaling is thought to constitute exercise-mimicking signaling. Indeed, it has become clear that AMPK, SIRT1 and AdipoR activators act as exercise mimetics. With the crystal structures of AdipoR elucidated and humanized AdipoR mice generated toward optimization of candidate AdipoR-activators for human use, expectations are mounting for the clinical application in the near future of AdipoR activators as exercise mimetics in humans. This review provides an overview of molecules activated by exercise and compounds activating these molecules, with a focus on the therapeutic potential of AdipoR activators as exercise mimetics.

Keywords:  AdipoR; AdipoR activators; Exercise mimetics

DOI:  https://doi.org/10.1507/endocrj.EJ21-0294
Sci Rep. 2021 Sep 15. 11(1): 18349

Cannabinoid receptor 1 expression is higher in muscle of old vs. young males, and increases upon resistance exercise in older adults.

Sebastiaan Dalle, Katrien Koppo.

Aged skeletal muscle undergoes metabolic and structural alterations eventually resulting in a loss of muscle strength and mass, i.e. age-related sarcopenia. Therefore, novel targets for muscle growth purposes in elderly are needed. Here, we explored the role of the cannabinoid system in muscle plasticity through the expression of muscle cannabinoid receptors (CBs) in young and old humans. The CB1 expression was higher (+ 25%; p = 0.04) in muscle of old (≥ 65 years) vs. young adults (20-27 years), whereas CB2 was not differently expressed. Furthermore, resistance exercise tended to increase the CB1 (+ 11%; p = 0.055) and CB2 (+ 37%; p = 0.066) expression in muscle of older adults. Interestingly, increases in the expression of CB2 following resistance exercise positively correlated with changes in key mechanisms of muscle homeostasis, such as catabolism (FOXO3a) and regenerative capacity (Pax7, MyoD). This study for the first time shows that CB1 is differentially expressed with aging and that changes in CB2 expression upon resistance exercise training correlate with changes in mediators that play a central role in muscle plasticity. These data confirm earlier work in cells and mice showing that the cannabinoid system might orchestrate muscle growth, which is an incentive to further explore CB-based strategies that might counteract sarcopenia.

DOI: https://doi.org/10.1038/s41598-021-97859-3
J Cell Mol Med. 2021 Sep 12.

Sepsis induces muscle atrophy by inhibiting proliferation and promoting apoptosis via PLK1-AKT signalling.

Ying-Ya Cao, Zhen Wang, Tao Yu, Yuan Zhang, Zhong-Han Wang, Zi-Meng Lu, Wei-Hua Lu, Jian-Bo Yu.

  Sepsis and sepsis-induced skeletal muscle atrophy are common in patients in intensive care units with high mortality, while the mechanisms are controversial and complicated. In the present study, the atrophy of skeletal muscle was evaluated in sepsis mouse model as well as the apoptosis of muscle fibres. Sepsis induced atrophy of skeletal muscle and apoptosis of myofibres in vivo and in vitro. In cell-based in vitro experiments, lipopolysaccharide (LPS) stimulation also inhibited the proliferation of myoblasts. At the molecular level, the expression of polo-like kinase 1 (PLK1) and phosphorylated protein kinase B (p-AKT) was decreased. Overexpression of PLK1 partly rescued LPS-induced apoptosis, proliferation suppression and atrophy in C2C12 cells. Furthermore, inhibiting the AKT pathway deteriorated LPS-induced atrophy in PLK1-overexpressing C2C12 myotubes. PLK1 was found to participate in regulating apoptosis and E3 ubiquitin ligase activity in C2C12 cells. Taken together, these results indicate that sepsis induces skeletal muscle atrophy by promoting apoptosis of muscle fibres and inhibiting proliferation of myoblasts via regulation of the PLK1-AKT pathway. These findings enhance understanding of the mechanism of sepsis-induced skeletal muscle atrophy.

Keywords:  apoptosis; muscle atrophy; polo-like kinase 1; proliferation; sepsis

DOI:  https://doi.org/10.1111/jcmm.16921
Stem Cell Reports. 2021 Sep 08. pii: S2213-6711(21)00429-X. [Epub ahead of print]

CRISPR/Cas9/AAV9-mediated in vivo editing identifies MYC regulation of 3D genome in skeletal muscle stem cell.

Liangqiang He, Yingzhe Ding, Yu Zhao, Karl K So, Xianlu L Peng, Yuying Li, Jie Yuan, Zhiming He, Xiaona Chen, Hao Sun, Huating Wang.

  Skeletal muscle satellite cells (SCs) are stem cells responsible for muscle development and regeneration. Although CRISPR/Cas9 has been widely used, its application in endogenous SCs remains elusive. Here, we generate mice expressing Cas9 in SCs and achieve robust editing in juvenile SCs at the postnatal stage through AAV9-mediated short guide RNA (sgRNA) delivery. Additionally, we reveal that quiescent SCs are resistant to CRISPR/Cas9-mediated editing. As a proof of concept, we demonstrate efficient editing of master transcription factor (TF) Myod1 locus using the CRISPR/Cas9/AAV9-sgRNA system in juvenile SCs. Application on two key TFs, MYC and BCL6, unveils distinct functions in SC activation and muscle regeneration. Particularly, we reveal that MYC orchestrates SC activation through regulating 3D genome architecture. Its depletion results in strengthening of the topologically associating domain boundaries thus may affect gene expression. Altogether, our study establishes a platform for editing endogenous SCs that can be harnessed to elucidate the functionality of key regulators governing SC activities.

Keywords:  3D chromatin; CRISPR/Cas9; MYC; muscle stem cell

DOI:  https://doi.org/10.1016/j.stemcr.2021.08.011
Ageing Res Rev. 2021 Sep 14. pii: S1568-1637(21)00210-5. [Epub ahead of print] 101463

Shared and Distinct Mechanisms of Skeletal Muscle Atrophy: A Narrative Review.

Dylan Wilburn, Ahmed Ismaeel, Steven Machek, Emma Fletcher, Panagiotis Koutakis.

  Maintenance of skeletal muscle mass and function is an incredibly nuanced balance of anabolism and catabolism that can become distorted within different pathological conditions. In this paper we intend to discuss the distinct intracellular signaling events that regulate muscle protein atrophy for a given clinical occurrence. Aside from the common outcome of muscle deterioration, several conditions have at least one or more distinct mechanisms that creates unique intracellular environments that facilitate muscle loss. The subtle individuality to each of these given pathologies can provide both researchers and clinicians with specific targets of interest to further identify and increase the efficacy of medical treatments and interventions.

Keywords:  Atrophy; Cachexia; Catabolism; Muscle Loss; Muscle Protein Degradation; Sarcopenia

DOI:  https://doi.org/10.1016/j.arr.2021.101463
J Physiol. 2021 Sep 12.

The missing "link"? β-Catenin's role in skeletal muscle glucose uptake during exercise & contraction.

Evan M Hoecht.

DOI: https://doi.org/10.1113/JP282232
J Cachexia Sarcopenia Muscle. 2021 Sep 15.

Ablation of USP21 in skeletal muscle promotes oxidative fibre phenotype, inhibiting obesity and type 2 diabetes.

Ayoung Kim, Ja Hyun Koo, Xing Jin, Wondong Kim, Shi-Young Park, Sunghyouk Park, Eugene P Rhee, Cheol Soo Choi, Sang Geon Kim.

  BACKGROUND: Skeletal muscle as a metabolic consumer determines systemic energy homeostasis by regulating myofibre type conversion and muscle mass control. Perturbation of the skeletal muscle metabolism elevates the risk of a variety of diseases including metabolic disorders. However, the regulatory pathways and molecules are not completely understood. The discovery of relevant responsible molecules and the associated network could be an attractive strategy to overcome diseases associated with muscle problems.METHODS: An initial screening using quantitative trait locus analysis enabled us to extract a set of genes including ubiquitin-specific proteases21 (USP21) (r = 0.738; P = 0.004) as potential targets associated with fasting blood glucose content. Given tight regulation of the ubiquitination status of proteins in muscle, we focused on USP21 and generated whole-body (KO) and skeletal muscle-specific USP21 knockout (MKO) mice. Transcriptomics, proteomics, and lipidomics assays in combination with various in vivo and in vitro experiments were performed to understand the functions of USP21 and underlying mechanisms. A high-fat diet (60%)-fed mouse model and diabetic patient-derived samples were utilized to assess the effects of USP21 on energy metabolism in skeletal muscle.
RESULTS: USP21 was highly expressed in both human and mouse skeletal muscle, and controlled skeletal muscle oxidative capacity and fuel consumption. USP21-KO or USP21-MKO significantly promoted oxidative fibre type changes (Δ36.6% or Δ47.2%), muscle mass increase (Δ13.8% to Δ22.8%), and energy expenditure through mitochondrial biogenesis, fatty acid oxidation, and UCP2/3 induction (P < 0.05 or P < 0.01). Consistently, cold exposure repressed USP21 expression in mouse skeletal muscle (Δ55.3%), whereas loss of USP21 increased thermogenesis (+1.37°C or +0.84°C; P < 0.01). Mechanistically, USP21 deubiquitinated DNA-PKcs and ACLY, which led to AMPK inhibition. Consequently, USP21 ablation diminished diet-induced obesity (WT vs. USP21-KO, Δ8.02 g, 17.1%, P < 0.01; litter vs. USP21-MKO, Δ3.48 g, 7.7%, P < 0.05) and insulin resistance. These findings were corroborated in a skeletal muscle-specific gene KO mouse model. USP21 was induced in skeletal muscle of a diabetic patient (1.94-fold), which was reciprocally changed to p-AMPK (0.30-fold).
CONCLUSIONS: The outcomes of this research provide novel information as to how USP21 in skeletal muscle contributes to systemic energy homeostasis, demonstrating USP21 as a key molecule in the regulation of myofibre type switch, muscle mass control, mitochondrial function, and heat generation and, thus, implicating the potential of this molecule and its downstream substrates network as targets for the treatment and/or prevention of muscle dysfunction and the associated metabolic diseases.

Keywords:  Diabetes; Muscle mass control; Obesity; Oxidative fibre type; Skeletal muscle; USP21

DOI:  https://doi.org/10.1002/jcsm.12777
Mol Ther Nucleic Acids. 2021 Dec 03. 26 34-48

FTO-mediated demethylation of GADD45B promotes myogenesis through the activation of p38 MAPK pathway.

Kaiping Deng, Yixuan Fan, Yaxu Liang, Yu Cai, Guomin Zhang, Mingtian Deng, Zhibo Wang, Jiawei Lu, Jianfei Shi, Feng Wang, Yanli Zhang.

  N6-methyladenosine (m6A) modification plays a critical role in mammalian development. However, the role of m6A in the skeletal muscle development remains largely unknown. Here, we report a global m6A modification pattern of goat skeletal muscle at two key development stages and identified that the m6A modification regulated the expression of the growth arrest and DNA damage-inducible 45B (GADD45B) gene, which is involved in myogenic differentiation. We showed that GADD45B expression increased during myoblast differentiation, whereas the downregulation of GADD45B inhibits myogenic differentiation and mitochondrial biogenesis. Moreover, the expression of GADD45B regulates the expression of myogenic regulatory factors and peroxisome proliferator-activated receptor gamma coactivator 1 alpha by activating the p38 mitogen-activated protein kinase (MAPK) pathway. Conversely, the inactivation of p38 MAPK abolished the GADD45B-mediated myogenic differentiation. Furthermore, we found that the knockdown of fat mass and obesity-associated protein (FTO) increases GADD45B m6A modification and decreases the stability of GADD45B mRNA, which impairs myogenic differentiation. Our results indicate that the FTO-mediated m6A modification in GADD45B mRNA drives skeletal muscle differentiation by activating the p38 MAPK pathway, which provides a molecular mechanism for the regulation of myogenesis via RNA methylation.

Keywords:  FTO; GADD45B; goat; m6A modification; mitochondrial biogenesis; myogenic differentiation

DOI:  https://doi.org/10.1016/j.omtn.2021.06.013
Curr Opin Cell Biol. 2021 Sep 14. pii: S0955-0674(21)00097-1. [Epub ahead of print]73 124-132

Deciphering the chromatin organization and dynamics for muscle stem cell function.

Anqi Dong, Tom H Cheung.

The chromatin landscape represents a critical regulatory layer for precise transcriptional control. Chromosome architecture restrains the physical access to the DNA elements and is one of the determinants that specifies cell identity. Adult stem cells possess the unique ability to differentiate into a specific lineage. One of the underexplored areas in skeletal muscle biology is the molecular mechanism guiding the chromatin organization changes in muscle stem cell specification, myogenic determination, and differentiation. In this review, we focus on the regulatory network guiding the progression of muscle stem cells to differentiated progeny. We summarize recent findings regarding the mechanisms directing myogenic cell fate decision and differentiation, with a particular focus on three-dimensional chromosome architecture and long noncoding RNA-associated chromatin accessibility changes.

DOI: https://doi.org/10.1016/j.ceb.2021.08.001
J Nutr Biochem. 2021 Sep 10. pii: S0955-2863(21)00279-5. [Epub ahead of print] 108859

Effect of dietary L-theanine supplementation on skeletal muscle fiber type transformation in vivo.

Xiaoling Chen, Man Zhang, Yonghong Xue, Dahui Liang, Wenting An, Gang Jia, Hua Zhao, Guangmang Liu, Zhiqing Huang.

  The aim of this study was to investigate the effect of dietary L-theanine supplementation on skeletal muscle fiber type transition in mice. Our data indicated that dietary 0.15% L-theanine supplementation significantly increased the mRNA expression levels of muscle fiber type related genes (MyHC I, MyHC IIa, PGC-1α, Sirt1, Tnnt1, Tnnc1, Tnni1, MEF2C) and the protein expression levels of MyHC IIa, myoglobin, PGC-1α, Sirt1 and Troponin I-SS, but significantly decreased the mRNA and protein expression levels of MyHC IIb. Dietary 0.15% L-theanine supplementation significantly increased the activities of SDH and MDH and decreased the activity of LDH. Furthermore, immunofluorescence demonstrated that dietary 0.15% L-theanine supplementation significantly increased the percentage of type I fibers, and significantly decreased the percentage of type II fibers. In addition, we found that dietary 0.15% L-theanine supplementation increased the fatigue-resistant, antioxidant capacity, mitochondrial biogenesis and function in skeletal muscle of mice. Furthermore, dietary 0.15% L-theanine supplementation significantly increased the mRNA levels of prox1, CaN and NFATc1, the protein levels of prox1, CNA and NFATc1 and the activity of CaN in GAS muscle when compared with the control group. These results indicated that dietary L-theanine supplementation promoted skeletal muscle fiber transition from type II to type I, which might be via activation of CaN/NFATc1 signaling pathway.

Keywords:  CaN/NFATc1 signaling pathway; Fiber type transition; L-theanine; Skeletal muscle

DOI:  https://doi.org/10.1016/j.jnutbio.2021.108859
Sports Med. 2021 Sep 13.

Protein Requirements for Master Athletes: Just Older Versions of Their Younger Selves.

Daniel R Moore.

It is established that protein requirements are elevated in athletes to support their training and post-exercise recovery and adaptation, especially within skeletal muscle. However, research on the requirements for this macronutrient has been performed almost exclusively in younger athletes, which may complicate their translation to the growing population of Master athletes (i.e. > 35 years old). In contrast to older (> 65 years) untrained adults who typically demonstrate anabolic resistance to dietary protein as a primary mediator of the 'normal' age-related loss of muscle mass and strength, Master athletes are generally considered successful models of aging as evidenced by possessing similar body composition, muscle mass, and aerobic fitness as untrained adults more than half their age. The primary physiology changes considered to underpin the anabolic resistance of aging are precipitated or exacerbated by physical inactivity, which has led to higher protein recommendations to stimulate muscle protein synthesis in older untrained compared to younger untrained adults. This review puts forth the argument that Master athletes have similar muscle characteristics, physiological responses to exercise, and protein metabolism as young athletes and, therefore, are unlikely to have protein requirements that are different from their young contemporaries. Recommendations for protein amount, type, and pattern will be discussed for Master athletes to enhance their recovery from and adaptation to resistance and endurance training.

DOI: https://doi.org/10.1007/s40279-021-01510-0
J Biol Chem. 2021 Sep 13. pii: S0021-9258(21)00998-4. [Epub ahead of print] 101196

Dynamin-related protein 1 regulates substrate oxidation in skeletal muscle by stabilizing cellular and mitochondrial calcium dynamics.

William T King, Christopher L Axelrod, Elizabeth R M Zunica, Robert C Noland, Gangarao Davuluri, Hisashi Fujioka, Bernard Tandler, Kathryn Pergola, Gerlinda E Hermann, Richard C Rogers, Sandra López-Domènech, Wagner S Dantas, Krisztian Stadler, Charles L Hoppel, John P Kirwan.

  Mitochondria undergo continuous cycles of fission and fusion to promote inheritance, regulate quality control, and mitigate organelle stress. More recently, this process of mitochondrial dynamics has been demonstrated to be highly sensitive to nutrient supply, ultimately conferring bioenergetic plasticity to the organelle. However, whether regulators of mitochondrial dynamics play a causative role in nutrient regulation remains unclear. In this study, we generated a cellular loss-of-function model for dynamin-related protein 1 (DRP1), the primary regulator of outer membrane mitochondrial fission. Loss of DRP1 (shDRP1) resulted in extensive ultrastructural and functional remodeling of mitochondria, characterized by pleomorphic enlargement, increased electron density of the matrix, and defective NADH and succinate oxidation. Despite increased mitochondrial size and volume, shDRP1 cells exhibited reduced cellular glucose uptake and mitochondrial fatty acid oxidation. Untargeted transcriptomic profiling revealed severe downregulation of genes required for cellular and mitochondrial calcium homeostasis, inhibition of ATP-stimulated calcium flux, and impaired substrate oxidation stimulated by calcium levels. The insights obtained herein suggest that DRP1 regulates fatty acid oxidation by altering whole-cell and mitochondrial calcium dynamics. These findings are relevant to the targetability of mitochondrial fission and have clinical relevance in the identification of treatments for fission-related pathologies such as hereditary neuropathies, inborn errors in metabolism, cancer, and chronic diseases.

Keywords:  calcium signaling; dynamin-related protein 1; mitochondrial dynamics; skeletal muscle; β-oxidation

DOI:  https://doi.org/10.1016/j.jbc.2021.101196
Nutr Metab (Lond). 2021 Sep 15. 18(1): 85

Androgen receptor regulates the proliferation of myoblasts under appropriate or excessive stretch through IGF-1 receptor mediated p38 and ERK1/2 pathways.

Shaoting Fu, Xiaojing Lin, Lijun Yin, Xiaohui Wang.

  BACKGROUND: Androgen receptor (AR) exerts important roles in exercise-induced alterations of muscle mass, in which the proliferation and differentiation of satellite cells or myoblasts are crucial. Our previous study in C2C12 myoblasts demonstrated that 15% (mimic appropriate exercise) and 20% (mimic excessive exercise) stretches promoted and inhibited the proliferation respectively; and AR played a crucial role in 15% stretch-induced pro-proliferation through IGF-1-modulated PI3K/Akt, p38 and ERK1/2 pathways, but AR's role in stretches-modulated proliferation of general myoblasts, especially 20% stretch, remains unclear, and the mechanisms need to be further clarified.METHODS: Firstly, the discrepancy in proliferation and the above indicators between L6 (without AR) and C2C12 (with AR) myoblasts were compared under 15% or 20% stretch. Then the influences of transfection AR or exogenous IGF-1 treatment on proliferation and these indicators were detected in stretched L6 myoblasts.
RESULTS: (1) Under un-stretched state, the proliferation of L6 was slower than C2C12 cells. Furthermore, AR knockdown in C2C12 myoblasts repressed, while AR overexpression in L6 myoblasts promoted the proliferation. (2) 15% stretch-induced increases in the proliferation and activities of p38 and ERK1/2 were lower in L6 than C2C12 cells; AR overexpression enhanced the proliferation of 15% stretched L6 cells accompanied with the increases of p38 and ERK1/2 activities. (3) 20% stretch-induced anti-proliferation and inhibition of p38 activity were severer in L6 than C2C12 myoblasts; AR overexpression reversed the anti-proliferation of 20% stretch and enhanced p38 activity in L6 myoblasts. (4) In stretched L6 myoblasts, AR overexpression increased IGF-1R level despite no detectable IGF-1; and recombinant IGF-1 increased the proliferation, the level of IGF-1R, and the activities of p38 and ERK1/2 in 15% stretched L6 myoblasts.
CONCLUSIONS: The study demonstrated AR's crucial roles in stretches-regulated proliferation of myoblasts, and increased AR fulfilled 15% stretch's pro-proliferation via activating IGF-1R- p38 and ERK1/2 pathways while decreased AR achieved 20% stretch's anti-proliferation via inhibiting IGF-1R- p38 pathway, which is useful to understand in depth the role and mechanisms of AR in appropriate exercise increasing while excessive exercise decreasing muscle mass.

Keywords:  AR; ERK1/2; IGF-1R; Myoblast proliferation; Stretch; p38

DOI:  https://doi.org/10.1186/s12986-021-00610-y
J Neuromuscul Dis. 2021 Sep 07.

Gene Therapy for Duchenne Muscular Dystrophy.

Nertiyan Elangkovan, George Dickson.

  Duchenne muscular dystrophy (DMD) is an X-linked, muscle wasting disease that affects 1 in 5000 males. Affected individuals become wheelchair bound by the age of twelve and eventually die in their third decade due to respiratory and cardiac complications. The disease is caused by mutations in the DMD gene that codes for dystrophin. Dystrophin is a structural protein that maintains the integrity of muscle fibres and protects them from contraction-induced damage. The absence of dystrophin compromises the stability and function of the muscle fibres, eventually leading to muscle degeneration. So far, there is no effective treatment for deteriorating muscle function in DMD patients. A promising approach for treating this life-threatening disease is gene transfer to restore dystrophin expression using a safe, non-pathogenic viral vector called adeno-associated viral (AAV) vector. Whilst microdystrophin gene transfer using AAV vectors shows extremely impressive therapeutic success so far in large animal models of DMD, translating this advanced therapy medicinal product from bench to bedside still offers scope for many optimization steps. In this paper, the authors review the current progress of AAV-microdystrophin gene therapy for DMD and other treatment strategies that may apply to a subset of DMD patients depending on the mutations they carry.

Keywords:  Gene therapy; adeno-associated virus; antisense; duchenne; dystrophin; exon skipping; microdystrophin; muscular dystrophy

DOI:  https://doi.org/10.3233/JND-210678
Sci Rep. 2021 Sep 13. 11(1): 18161

Reduced mitochondrial fission and impaired energy metabolism in human primary skeletal muscle cells of Megaconial Congenital Muscular Dystrophy.

Evrim Aksu-Menges, Cemil Can Eylem, Emirhan Nemutlu, Merve Gizer, Petek Korkusuz, Haluk Topaloglu, Beril Talim, Burcu Balci-Hayta.

Megaconial Congenital Muscular Dystrophy (CMD) is a rare autosomal recessive disorder characterized by enlarged mitochondria located mainly at the periphery of muscle fibers and caused by mutations in the Choline Kinase Beta (CHKB) gene. Although the pathogenesis of this disease is not well understood, there is accumulating evidence for the presence of mitochondrial dysfunction. In this study, we aimed to investigate whether imbalanced mitochondrial dynamics affects mitochondrial function and bioenergetic efficiency in skeletal muscle cells of Megaconial CMD. Immunofluorescence, confocal and transmission electron microscopy studies revealed impaired mitochondrial network, morphology, and localization in primary skeletal muscle cells of Megaconial CMD. The organelle disruption was specific only to skeletal muscle cells grown in culture. The expression levels of mitochondrial fission proteins (DRP1, MFF, FIS1) were found to be decreased significantly in both primary skeletal muscle cells and tissue sections of Megaconial CMD by Western blotting and/or immunofluorescence analysis. The metabolomic and fluxomic analysis, which were performed in Megaconial CMD for the first time, revealed decreased levels of phosphonucleotides, Krebs cycle intermediates, ATP, and altered energy metabolism pathways. Our results indicate that reduced mitochondrial fission and altered mitochondrial energy metabolism contribute to mitochondrial dysmorphology and dysfunction in the pathogenesis of Megaconial CMD.

DOI: https://doi.org/10.1038/s41598-021-97294-4
Methods Mol Biol. 2022 ;2373 213-230

Biofabrication of 3D Human Muscle Model with Vascularization and Endomysium.

Simone Bersini, Riccardo Francescato, Matteo Moretti.

  This protocol describes the biofabrication of 3D millimeter-scale human muscle units, embedding non-planar muscle fibers wrapped by fibroblasts-rich endomysium and intertwined with microvascular networks. Suspended muscle fibers are formed through the self-assembly of human myoblasts within cylindrical cavities generated in a sacrificial gelatin template cast in a 3D printed frame. Following myotube differentiation, muscle fibers are embedded in a 3D matrix containing endothelial cells and muscle-derived fibroblasts. The cellular complexity of the environment is instrumental to drive fibroblast migration towards muscle fibers and to induce the organ-specific differentiation of endothelial cells. This advanced 3D muscle model can be applied to analyze the biological mechanisms underlying specific muscle diseases which involve a complex remodeling of the muscle environment (e.g., muscular dystrophies and fibrosis) whereby the pathological interplay among different cell populations drives the onset and progression of the disease.

Keywords:  3D human muscle model; 3D printing; Duchenne muscular dystrophy; Endomysium; Endothelial cell; Fibrosis; Mesoscale; Organ specificity; Vasculature

DOI:  https://doi.org/10.1007/978-1-0716-1693-2_13
Mol Neurobiol. 2021 Sep 12.

Generation and Characterization of a Skeletal Muscle Cell-Based Model Carrying One Single Gne Allele: Implications in Actin Dynamics.

Shamulailatpam Shreedarshanee Devi, Rashmi Yadav, Fluencephila Mashangva, Priyanka Chaudhary, Shweta Sharma, Ranjana Arya.

  UDP-N-Acetyl glucosamine-2 epimerase/N-acetyl mannosamine kinase (GNE) catalyzes key enzymatic reactions in the biosynthesis of sialic acid. Mutation in GNE gene causes GNE myopathy (GNEM) characterized by adult-onset muscle weakness and degeneration. However, recent studies propose alternate roles of GNE in other cellular processes beside sialic acid biosynthesis, particularly interaction of GNE with α-actinin 1 and 2. Lack of appropriate model system limits drug and treatment options for GNEM as GNE knockout was found to be embryonically lethal. In the present study, we have generated L6 rat skeletal muscle myoblast cell-based model system carrying one single Gne allele where GNE gene is knocked out at exon-3 using AAV mediated SEPT homology recombination (SKM-GNEHz). The cell line was heterozygous for GNE gene with one wild type and one truncated allele as confirmed by sequencing. The phenotype showed reduced GNE epimerase activity with little reduction in sialic acid content. In addition, the heterozygous GNE knockout cells revealed altered cytoskeletal organization with disrupted actin filament. Further, we observed increased levels of RhoA leading to reduced cofilin activity and causing reduced F-actin polymerization. The disturbed signaling cascade resulted in reduced migration of SKM-GNEHz cells. Our study indicates possible role of GNE in regulating actin dynamics and cell migration of skeletal muscle cell. The skeletal muscle cell-based system offers great potential in understanding pathomechanism and target identification for GNEM.

Keywords:  Actin dynamics; GNE myopathy; L6 myoblast; SEPT homology recombination; Sialylation; Single gene mutation disorders

DOI:  https://doi.org/10.1007/s12035-021-02549-w
J Bone Metab. 2021 Aug;28(3): 179-191

Two Types of Mouse Models for Sarcopenia Research: Senescence Acceleration and Genetic Modification Models.

Kyung-Wan Baek, Youn-Kwan Jung, Jin Sung Park, Ji-Seok Kim, Young-Sool Hah, So-Jeong Kim, Jun-Il Yoo.

  Sarcopenia leads to loss of skeletal muscle mass, quality, and strength due to aging; it was recently given a disease code (International Classification of Diseases, Tenth Revision, Clinical Modification, M62.84). As a result, in recent years, sarcopenia-related research has increased. In addition, various studies seeking to prevent and treat sarcopenia by identifying the various mechanisms related to the reduction of skeletal muscle properties have been conducted. Previous studies have identified muscle synthesis and breakdown; investigating them has generated evidence for preventing and treating sarcopenia. Mouse models are still the most useful ones for determining mechanisms underlying sarcopenia through correlations and interventions involving specific genes and their phenotypes. Mouse models used to study sarcopenia often induce muscle atrophy by hindlimb unloading, denervation, or immobilization. Though it is less frequently used, the senescence-accelerated mouse can also be useful for sarcopenia research. Herein, we discuss cases where senescence-accelerated and genetically engineered mouse models were used in sarcopenia research and different perspectives to use them.

Keywords:  Aging; Mice; Muscle, skeletal; Sarcopenia

DOI:  https://doi.org/10.11005/jbm.2021.28.3.179
J Cell Mol Med. 2021 Sep 12.

Simple derivation of skeletal muscle from human pluripotent stem cells using temperature-sensitive Sendai virus vector.

Ghee Wan Tan, Takayuki Kondo, Keiko Imamura, Mika Suga, Takako Enami, Ayako Nagahashi, Kayoko Tsukita, Ikuyo Inoue, Jitsutaro Kawaguchi, Tsugumine Shu, Haruhisa Inoue.

  Human pluripotent stem cells have the potential to differentiate into various cell types including skeletal muscles (SkM), and they are applied to regenerative medicine or in vitro modelling for intractable diseases. A simple differentiation method is required for SkM cells to accelerate neuromuscular disease studies. Here, we established a simple method to convert human pluripotent stem cells into SkM cells by using temperature-sensitive Sendai virus (SeV) vector encoding myoblast determination protein 1 (SeV-Myod1), a myogenic master transcription factor. SeV-Myod1 treatment converted human embryonic stem cells (ESCs) into SkM cells, which expressed SkM markers including myosin heavy chain (MHC). We then removed the SeV vector by temporal treatment at a high temperature of 38℃, which also accelerated mesodermal differentiation, and found that SkM cells exhibited fibre-like morphology. Finally, after removal of the residual human ESCs by pluripotent stem cell-targeting delivery of cytotoxic compound, we generated SkM cells with 80% MHC positivity and responsiveness to electrical stimulation. This simple method for myogenic differentiation was applicable to human-induced pluripotent stem cells and will be beneficial for investigations of disease mechanisms and drug discovery in the future.

Keywords:  Myod1; Sendai virus; differentiation method; disease modelling; high temperature treatment; human embryonic stem cells; human-induced pluripotent stem cells; skeletal muscle

DOI:  https://doi.org/10.1111/jcmm.16899
J Neuromuscul Dis. 2021 Sep 10.

Visualizing Muscle Sialic Acid Expression in the GNED207VTgGne-/- Cmah-/- Model of GNE Myopathy: A Comparison of Dietary and Gene Therapy Approaches.

Kelly E Crowe, Deborah A Zygmunt, Kristin Heller, Louise Rodino-Klapac, Satoru Noguchi, Ichizo Nishino, Paul T Martin.

  BACKGROUND: GNE myopathy (GNEM) is a rare, adult-onset, inclusion body myopathy that results from partial loss of function mutations in the GNE gene. GNE encodes UDP-GlcNAc epimerase/Mannose-6 kinase, a protein with two enzymatic activities that comprise the committed step in biosynthesis of sialic acid (SA), an essential glycan that appears on the terminal positions of many extracellular oligosaccharide chains. These GNE mutations can cause a reduction of SA in many tissues, although pathology is restricted to skeletal muscles through a poorly understood mechanism.OBJECTIVE: Despite recent advances in the field, it remains unclear which therapeutic avenue is most promising for the restoration of SA level in skeletal muscle affected by GNEM. Our objective was to assess dietary and gene therapy strategies for GNEM in Cmah-deficient GNED207VTgGne-/- mice, a model that allows for the visualization of orally delivered N-glycolylneuraminic acid (Neu5Gc), one of the two predominant SA forms in muscle.
METHODS: Methods included in situ physiology studies of the tibialis anterior muscle, studies of ambulation and limb grip strength, and muscle staining using MAA, SNA, and anti-Neu5Gc antibody, along with qPCR, qRT-PCR, western blot, and HPLC studies to assess virally introduced DNA, GNE gene expression, GNE protein expression, and SA expression.
RESULTS: We found that a diet enriched in Neu5Gc-containing glycoproteins had no impact on Neu5Gc immunostaining in muscles of GNEM model mice. Delivery of a single high dose oral Neu5Gc therapy, however, did increase Neu5Gc immunostaining, though to levels below those found in wild type mice. Delivery of a single dose of GNE gene therapy using a recombinant Adeno Associated Virus (rAAV) vector with a liver-specific or a muscle-specific promoter both caused increased muscle Neu5Gc immunostaining that exceeded that seen with single dose monosaccharide therapy.
CONCLUSIONS: Our findings indicate that dietary loading of Neu5Gc-containing glycoproteins is not effective in increasing muscle Neu5Gc expression, while single dose oral Neu5Gc monosaccharide or GNE gene therapy are. Neu5Gc immunostaining, however, showed greater changes than did lectin staining or HPLC analysis. Taken together, these results suggest that Neu5Gc immunostaining may be more sensitive technique to follow SA expression than other more commonly used methods and that liver expression of GNE may contribute overall muscle SA content.

Keywords:  AAV; Myopathy; gene therapy; hereditary inclusion body myopathy; sialic acid

DOI:  https://doi.org/10.3233/JND-200575
Sports Med. 2021 Sep 13.

Fish Oil for Healthy Aging: Potential Application to Master Athletes.

Caoileann H Murphy, Chris McGlory.

Master athletes perform high volumes of exercise training yet display lower levels of physical functioning and exercise performance when compared with younger athletes. Several reports in the clinical literature show that long chain n-3 polyunsaturated fatty acid (LC n-3 PUFA) ingestion promotes skeletal muscle anabolism and strength in untrained older persons. There is also evidence that LC n-3 PUFA ingestion improves indices of muscle recovery following damaging exercise in younger persons. These findings suggest that LC n-3 PUFA intake could have an ergogenic effect in master athletes. However, the beneficial effect of LC n-3 PUFA intake on skeletal muscle in response to exercise training in both older and younger persons is inconsistent and, in some cases, generated from low-quality studies or those with a high risk of bias. Other factors such as the choice of placebo and health status of participants also confound interpretation of existing reports. As such, when considered on balance, the available evidence does not indicate that ingestion of LC n-3 PUFAs above current population recommendations (250-500 mg/day; 2 portions of oily fish per week) enhances exercise performance or recovery from exercise training in master athletes. Further work is now needed related to how the dose, duration, and co-ingestion of LC n-3 PUFAs with other nutrients such as amino acids impact the adaptive response to exercise training. This work should also consider how LC n-3 PUFA supplementation may differentially alter the lipid profile of cellular membranes of key regulatory sites such as the sarcolemma, mitochondria, and sarcoplasmic reticulum.

DOI: https://doi.org/10.1007/s40279-021-01509-7
Cell. 2021 Sep 16. pii: S0092-8674(21)01014-X. [Epub ahead of print]184(19): 4845-4847

Designer AAV muscle up.

Carsten G Bönnemann.

Directed evolution of AAV capsids has been a successful strategy for generating bespoke serotypes to target gene therapies more specifically to the intended tissue. This has now been achieved for the largest organ, skeletal muscle, by selecting for an RGD containing integrin binding heptamer in a hypervariable region of the capsid of AAV9.

DOI: https://doi.org/10.1016/j.cell.2021.08.031
Comput Struct Biotechnol J. 2021 ;19 4142-4155

MyoData: An expression knowledgebase at single cell/nucleus level for the discovery of coding-noncoding RNA functional interactions in skeletal muscle.

Davide Corso, Francesco Chemello, Enrico Alessio, Ilenia Urso, Giulia Ferrarese, Martina Bazzega, Chiara Romualdi, Gerolamo Lanfranchi, Gabriele Sales, Stefano Cagnin.

  Non-coding RNAs represent the largest part of transcribed mammalian genomes and prevalently exert regulatory functions. Long non-coding RNAs (lncRNAs) and microRNAs (miRNAs) can modulate the activity of each other. Skeletal muscle is the most abundant tissue in mammals. It is composed of different cell types with myofibers that represent the smallest complete contractile system. Considering that lncRNAs and miRNAs are more cell type-specific than coding RNAs, to understand their function it is imperative to evaluate their expression and action within single myofibers. In this database, we collected gene expression data for coding and non-coding genes in single myofibers and used them to produce interaction networks based on expression correlations. Since biological pathways are more informative than networks based on gene expression correlation, to understand how altered genes participate in the studied phenotype, we integrated KEGG pathways with miRNAs and lncRNAs. The database also integrates single nucleus gene expression data on skeletal muscle in different patho-physiological conditions. We demonstrated that these networks can serve as a framework from which to dissect new miRNA and lncRNA functions to experimentally validate. Some interactions included in the database have been previously experimentally validated using high throughput methods. These can be the basis for further functional studies. Using database information, we demonstrate the involvement of miR-149, -214 and let-7e in mitochondria shaping; the ability of the lncRNA Pvt1 to mitigate the action of miR-27a via sponging; and the regulatory activity of miR-214 on Sox6 and Slc16a3. The MyoData is available at https://myodata.bio.unipd.it.

Keywords:  Database; Networks; Non-coding RNAs; Pathways; Single myofiber; Single nucleus

DOI:  https://doi.org/10.1016/j.csbj.2021.07.020
Endocr Rev. 2021 Sep 14. pii: bnab030. [Epub ahead of print]

Myostatin/Activin Receptor Ligands In Muscle And The Development Status Of Attenuating Drugs.

Buel D Rodgers, Christopher W Ward.

  Muscle wasting disease indications are among the most debilitating and often deadly noncommunicable disease states. As a comorbidity, muscle wasting is associated with different neuromuscular diseases and myopathies, cancer, heart failure, chronic pulmonary and renal diseases, peripheral neuropathies, inflammatory disorders and of course, musculoskeletal injuries. Current treatment strategies are relatively ineffective and can at best only limit the rate of muscle degeneration. This includes nutritional supplementation and appetite stimulants as well as immunosuppressants capable of exacerbating muscle loss. Arguably, the most promising treatments in development attempt to disrupt myostatin and activin receptor signaling as these circulating factors are potent inhibitors of muscle growth and regulators of muscle progenitor cell differentiation. Indeed, several studies demonstrated the clinical potential of "inhibiting the inhibitors", increasing muscle cell protein synthesis, decreasing degradation, enhancing mitochondrial biogenesis and preserving muscle function. Such changes can prevent muscle wasting in various disease animal models yet many drugs targeting this pathway failed during clinical trials, some from serious treatment-related adverse events and off-target interactions. More often, however, failures resulted from the inability to improve muscle function despite preserving muscle mass. Drugs still in development include antibodies and gene therapeutics, all with different targets and thus, safety, efficacy and proposed use profiles. Each is unique in design and, if successful, could revolutionize the treatment of both acute and chronic muscle wasting. They could also be used in combination with other developing therapeutics for related muscle pathologies or even metabolic diseases.

Keywords:  ACVR2; ACVR2B; ActRIIa; ActRIIb; GDF11; activin; growth/differentiation factor (GDF)8; muscle atrophy; muscle wasting; myostatin

DOI:  https://doi.org/10.1210/endrev/bnab030
Cell Death Dis. 2021 Sep 16. 12(10): 850

Malat-1-PRC2-EZH1 interaction supports adaptive oxidative stress dependent epigenome remodeling in skeletal myotubes.

Nadine Hosny El Said, Francesco Della Valle, Peng Liu, Andreu Paytuví-Gallart, Sabir Adroub, Juliette Gimenez, Valerio Orlando.

PRC2-mediated epigenetic function involves the interaction with long non-coding RNAs (lncRNAs). Although the identity of some of these RNAs has been elucidated in the context of developmental programs, their counterparts in postmitotic adult tissue homeostasis remain uncharacterized. To this aim, we used terminally differentiated postmitotic skeletal muscle cells in which oxidative stress induces the dynamic activation of PRC2-Ezh1 through Embryonic Ectoderm Develpment (EED) shuttling to the nucleus. We identify lncRNA Malat-1 as a necessary partner for PRC2-Ezh1-dependent response to oxidative stress. We show that in this pathway, PRC2-EZH1 dynamic assembly, and in turn stress induced skeletal muscle targeted genes repression, depends specifically on Malat-1. Our study reports about PRC2-RNA interactions in the physiological context of adaptive oxidative stress response and identifies the first lncRNA involved in PRC2-Ezh1 function.

DOI: https://doi.org/10.1038/s41419-021-04082-z
Mol Ther Methods Clin Dev. 2021 Sep 10. 22 401-414

Systemic delivery of AAVrh74.tMCK.hCAPN3 rescues the phenotype in a mouse model for LGMD2A/R1.

Zarife Sahenk, Burcak Ozes, Darren Murrey, Morgan Myers, Kyle Moss, Mehmet E Yalvac, Alicia Ridgley, Lei Chen, Jerry R Mendell.

  Limb girdle muscular dystrophy (LGMD) 2A/R1, caused by mutations in the CAPN3 gene and CAPN3 loss of function, is known to play a role in disease pathogenicity. In this study, AAVrh74.tMCK.CAPN3 was delivered systemically to two different age groups of CAPN3 knockout (KO) mice; each group included two treatment cohorts receiving low (1.17 × 1014 vg/kg) and high (2.35 × 1014 vg/kg) doses of the vector and untreated controls. Treatment efficacy was tested 20 weeks after gene delivery using functional (treadmill), physiological (in vivo muscle contractility assay), and histopathological outcomes. AAV.CAPN3 gene therapy resulted in significant, robust improvements in functional outcomes and muscle physiology at low and high doses in both age groups. Histological analyses of skeletal muscle showed remodeling of muscle, a switch to fatigue-resistant oxidative fibers in females, and fiber size increases in both sexes. Safety studies revealed no organ tissue abnormalities; specifically, there was no histopathological evidence of cardiotoxicity. These results show that CAPN3 gene replacement therapy improved the phenotype in the CAPN3 KO mouse model at both doses independent of age at the time of vector administration. The improvements were supported by an absence of cardiotoxicity, showing the efficacy and safety of the AAV.CAPN3 vector as a potential gene therapy for LGMDR1.

Keywords:  AAV; LGMD2A/R1; calpain3; calpainopathy; functional correlation; gene therapy; muscular dystrophy

DOI:  https://doi.org/10.1016/j.omtm.2021.06.010
NPJ Microgravity. 2021 Sep 17. 7(1): 34

Effects of microgravity exposure and fructo-oligosaccharide ingestion on the proteome of soleus and extensor digitorum longus muscles in developing mice.

Takashi Ohira, Yoko Ino, Yayoi Kimura, Yusuke Nakai, Ayuko Kimura, Yoichi Kurata, Hiroyuki Kagawa, Mitsuo Kimura, Kenji Egashira, Chie Matsuda, Yoshinobu Ohira, Satoshi Furukawa, Hisashi Hirano.

Short-chain fatty acids produced by the gut bacterial fermentation of non-digestible carbohydrates, e.g., fructo-oligosaccharide (FOS), contribute to the maintenance of skeletal muscle mass and oxidative metabolic capacity. We evaluated the effect of FOS ingestion on protein expression of soleus (Sol) and extensor digitorum longus muscles in mice exposed to microgravity (μ-g). Twelve 9-week-old male C57BL/6J mice were raised individually on the International Space Station under μ-g or artificial 1-g and fed a diet with or without FOS (n = 3/group). Regardless of FOS ingestion, the absolute wet weights of both muscles tended to decrease, and the fiber phenotype in Sol muscles shifted toward fast-twitch type following μ-g exposure. However, FOS ingestion tended to mitigate the μ-g-exposure-related decrease in oxidative metabolism and enhance glutathione redox detoxification in Sol muscles. These results indicate that FOS ingestion mildly suppresses metabolic changes and oxidative stress in antigravity Sol muscles during spaceflight.

DOI: https://doi.org/10.1038/s41526-021-00164-6
Sci Rep. 2021 Sep 14. 11(1): 18188

Duchenne muscular dystrophy cell culture models created by CRISPR/Cas9 gene editing and their application in drug screening.

Patricia Soblechero-Martín, Edurne Albiasu-Arteta, Aina Anton-Martinez, Laura de la Puente-Ovejero, Iker Garcia-Jimenez, Gabriela González-Iglesias, Irene Larrañaga-Aiestaran, Andrea López-Martínez, Javier Poyatos-García, Estíbaliz Ruiz-Del-Yerro, Federico Gonzalez, Virginia Arechavala-Gomeza.

Gene editing methods are an attractive therapeutic option for Duchenne muscular dystrophy, and they have an immediate application in the generation of research models. To generate myoblast cultures that could be useful in in vitro drug screening, we have optimised a CRISPR/Cas9 gene edition protocol. We have successfully used it in wild type immortalised myoblasts to delete exon 52 of the dystrophin gene, modelling a common Duchenne muscular dystrophy mutation; and in patient's immortalised cultures we have deleted an inhibitory microRNA target region of the utrophin UTR, leading to utrophin upregulation. We have characterised these cultures by demonstrating, respectively, inhibition of dystrophin expression and overexpression of utrophin, and evaluating the expression of myogenic factors (Myf5 and MyH3) and components of the dystrophin associated glycoprotein complex (α-sarcoglycan and β-dystroglycan). To demonstrate their use in the assessment of DMD treatments, we have performed exon skipping on the DMDΔ52-Model and have used the unedited DMD cultures/ DMD-UTRN-Model combo to assess utrophin overexpression after drug treatment. While the practical use of DMDΔ52-Model is limited to the validation to our gene editing protocol, DMD-UTRN-Model presents a possible therapeutic gene edition target as well as a useful positive control in the screening of utrophin overexpression drugs.

DOI: https://doi.org/10.1038/s41598-021-97730-5
Eur J Transl Myol. 2021 Sep 17.

A revised model for mitochondrial dysfunction in Duchenne muscular dystrophy.

Ai Vu Hong, Mathilde Sanson, Isabelle Richard, David Israeli.

We recetly identified a signaling pathway that links the upregulation of miR-379 with a mitochondrial response in dystrophic muscle. In the present commentary, we explain the significance that this pathway may have in mitochondrial dysfunction in Duchenne muscular dystrophy (DMD). We identified the upregulation of miR-379 in the serum and muscles of DMD animal models and patients. We found that miR-379 is one of very few miRNAs whose expression was normalized in DMD patients treated with glucocorticoid. We identified EIF4G2 as an miR-379 target, which may promote mitochondrial oxidative phosphorylation (OxPhos) in the skeletal muscle. We found enriched EIF4G2 expression in oxidative fibers, and identified the mitochondrial ATP synthase subunit DAPIT as a translational target of EIF4G2. The identified signaling cascade, which comprises miR-379, EIF4G2 and DAPIT, may link the glucocorticoid treatment in DMD to a recovered mitochondrial ATP synthesis rate. We propose an updated model of mitochondrial dysfunction in DMD.

DOI: https://doi.org/10.4081/ejtm.2021.10012
Curr Pharm Des. 2021 Sep 09.

Therapies for RYR1-Related Myopathies: Where We Stand and the Perspectives.

Mathilde Beaufils, Lauriane Travard, John Rendu, Isabelle Marty.

  RyR1-related myopathies are a family of genetic neuromuscular diseases due to mutations in the RYR1 gene. No treatment exists for any of these myopathies today, which could change in the coming years with the growing number of studies dedicated to the pre-clinical assessment of various approaches, from pharmacological to gene therapy strategies, using the numerous models developed up to now. In addition, the first clinical trials for these rare diseases have just been completed or are being launched. We review the most recent results obtained for the treatment of RyR1-related myopathies, and, in view of the progress in therapeutic development for other myopathies, we discuss the possible future therapeutic perspectives for RyR1-related myopathies.

Keywords:  Ryanodine receptor; calcium; excitation-contraction coupling ; gene therapy; myopathy; pharmacological therapy; skeletal muscle

DOI:  https://doi.org/10.2174/1389201022666210910102516
J Cachexia Sarcopenia Muscle. 2021 Sep 14.

Understanding the gut microbiota and sarcopenia: a systematic review.

Chaoran Liu, Wing-Hoi Cheung, Jie Li, Simon Kwoon-Ho Chow, Jun Yu, Sunny Hei Wong, Margaret Ip, Joseph Jao Yiu Sung, Ronald Man Yeung Wong.

  BACKGROUND: Gut microbiota dysbiosis and sarcopenia commonly occur in the elderly. Although the concept of the gut-muscle axis has been raised, the casual relationship is still unclear. This systematic review analyses the current evidence of gut microbiota effects on muscle/sarcopenia.METHODS: A systematic review was performed in PubMed, Embase, Web of Science, and The Cochrane Library databases using the keywords (microbiota* OR microbiome*) AND (sarcopen* OR muscle). Studies reporting the alterations of gut microbiota and muscle/physical performance were analysed.
RESULTS: A total of 26 pre-clinical and 10 clinical studies were included. For animal studies, three revealed age-related changes and relationships between gut microbiota and muscle. Three studies focused on muscle characteristics of germ-free mice. Seventy-five per cent of eight faecal microbiota transplantation studies showed that the recipient mice successfully replicated the muscle phenotype of donors. There were positive effects on muscle from seven probiotics, two prebiotics, and short-chain fatty acids (SCFAs). Ten studies investigated on other dietary supplements, antibiotics, exercise, and food withdrawal that affected both muscle and gut microbiota. Twelve studies explored the potential mechanisms of the gut-muscle axis. For clinical studies, 6 studies recruited 676 elderly people (72.8 ± 5.6 years, 57.8% female), while 4 studies focused on 244 young adults (29.7 ± 7.8 years, 55.4% female). The associations of gut microbiota and muscle had been shown in four observational studies. Probiotics, prebiotics, synbiotics, fermented milk, caloric restriction, and exercise in six studies displayed inconsistent effects on muscle mass, function, and gut microbiota.
CONCLUSIONS: Altering the gut microbiota through bacteria depletion, faecal transplantation, and various supplements was shown to directly affect muscle phenotypes. Probiotics, prebiotics, SCFAs, and bacterial products are potential novel therapies to enhance muscle mass and physical performance. Lactobacillus and Bifidobacterium strains restored age-related muscle loss. Potential mechanisms of microbiome modulating muscle mainly include protein, energy, lipid, and glucose metabolism, inflammation level, neuromuscular junction, and mitochondrial function. The role of the gut microbiota in the development of muscle loss during aging is a crucial area that requires further studies for translation to patients.

Keywords:  Aging; Function; Gut microbiota; Gut-muscle axis; Muscle; Sarcopenia

DOI:  https://doi.org/10.1002/jcsm.12784
Front Genet. 2021 ;12 719713

Cross-Species and Human Inter-Tissue Network Analysis of Genes Implicated in Longevity and Aging Reveal Strong Support for Nutrient Sensing.

Avijit Podder, Anish Raju, Nicholas J Schork.

  Intensive research efforts have been undertaken to slow human aging and therefore potentially delay the onset of age-related diseases. These efforts have generated an enormous amount of high-throughput data covering different levels in the physiologic hierarchy, e.g., genetic, epigenetic, transcriptomic, proteomic, and metabolomic, etc. We gathered 15 independent sources of information about genes potentially involved in human longevity and lifespan (N = 5836) and subjected them to various integrated analyses. Many of these genes were initially identified in non-human species, and we investigated their orthologs in three non-human species [i.e., mice (N = 967), fruit fly (N = 449), and worm (N = 411)] for further analysis. We characterized experimentally determined protein-protein interaction networks (PPIN) involving each species' genes from 9 known protein databases and studied the enriched biological pathways among the individually constructed PPINs. We observed three important signaling pathways: FoxO signaling, mTOR signaling, and autophagy to be common and highly enriched in all four species (p-value ≤ 0.001). Our study implies that the interaction of proteins involved in the mechanistic target of rapamycin (mTOR) signaling pathway is somewhat limited to each species or that a "rewiring" of specific networks has taken place over time. To corroborate our findings, we repeated our analysis in 43 different human tissues. We investigated conserved modules in various tissue-specific PPINs of the longevity-associated genes based upon their protein expression. This analysis also revealed mTOR signaling as shared biological processes across four different human tissue-specific PPINs for liver, heart, skeletal muscle, and adipose tissue. Further, we explored our results' translational potential by assessing the protein interactions with all the reported drugs and compounds that have been experimentally verified to promote longevity in the three-comparator species. We observed that the target proteins of the FDA-approved drug rapamycin (a known inhibitor of mTOR) were conserved across all four species. Drugs like melatonin and metformin exhibited shared targets with rapamycin in the human PPIN. The detailed information about the curated gene list, cross-species orthologs, PPIN, and pathways was assembled in an interactive data visualization portal using RStudio's Shiny framework (https://agingnetwork.shinyapps.io/frontiers/).

Keywords:  cross-species orthologs; drug targets; human aging; nutrient sensing; pathway enrichments; protein-protein interaction network

DOI:  https://doi.org/10.3389/fgene.2021.719713
Sports Med. 2021 Sep 13.

Resistance Exercise Trims the Fat and Puts Some Muscle into Cancer Survivorship.

Justin C Brown.

DOI: https://doi.org/10.1007/s40279-021-01557-z
Mol Ther. 2021 Sep 09. pii: S1525-0016(21)00458-5. [Epub ahead of print]

Full-length Dystrophin Restoration via Targeted Genomic Integration by AAV-CRISPR in a Humanized Mouse Model of Duchenne Muscular Dystrophy.

Adrian Pickar-Oliver, Veronica Gough, Joel D Bohning, Siyan Liu, Jacqueline N Robinson-Hamm, Heather Daniels, William H Majoros, Garth Devlin, Aravind Asokan, Charles A Gersbach.

Targeted gene editing strategies have emerged as promising therapeutic approaches for the permanent treatment of inherited genetic diseases. However, precise gene correction and insertion approaches using homology-directed repair are still limited by low efficiencies. Consequently, many gene editing strategies have focused on removal or disruption, rather than repair, of genomic DNA. In contrast, homology-independent targeted integration (HITI) has been reported to effectively insert DNA sequences at targeted genomic loci. This approach could be particularly useful for restoring full-length sequences of genes affected by a spectrum of mutations that are also too large to deliver by conventional adeno-associated virus (AAV) vectors. Here, we utilize an AAV-based HITI-mediated approach for correction of full-length dystrophin expression in a humanized mouse model of Duchenne muscular dystrophy (DMD). We co-deliver CRISPR-Cas9 and a donor DNA sequence to insert the missing human exon 52 into its corresponding position within the DMD gene and achieve full-length dystrophin correction in skeletal and cardiac muscle. Additionally, as a proof-of-concept strategy to correct genetic mutations characterized by diverse patient mutations, we deliver a superexon donor encoding the last 28 exons of the DMD gene as a therapeutic strategy to restore full-length dystrophin in >20% of the DMD patient population. This work highlights the potential of HITI-mediated gene correction for diverse DMD mutations and advances genome editing towards realizing the promise of full-length gene restoration to treat genetic disease.

DOI: https://doi.org/10.1016/j.ymthe.2021.09.003
J Biol Chem. 2021 Sep 08. pii: S0021-9258(21)00981-9. [Epub ahead of print] 101179

Dietary lysine restriction induces lipid accumulation in skeletal muscle through an increase in serum threonine levels in rats.

Yuki Goda, Daisuke Yamanaka, Hiroki Nishi, Masato Masuda, Hiroyasu Kamei, Mikako Kumano, Koichi Ito, Masaya Katsumata, Keitaro Yamanouchi, Naoyuki Kataoka, Fumihiko Hakuno, Shin-Ichiro Takahashi.

  We previously reported that dietary amino acid restriction induces the accumulation of triglycerides (TAG) in the liver of growing rats. However, differences in TAG accumulation in individual cell types or other tissues were not examined. In this study, we show that TAG also accumulates in the muscle and adipose tissues of rats fed a low amino acid (low-AA) diet. In addition, dietary lysine restriction (low-Lys) induces lipid accumulation in muscle and adipose tissues. In adjusting the nitrogen content to that of the control (CN) diet, we found that glutamic acid supplementation to the low-AA diet blocked lipid accumulation, but supplementation with the low-Lys diet did not, suggesting that a shortage of nitrogen caused lipids to accumulate in the skeletal muscle in the rats fed a low-AA diet. Serum amino acid measurement revealed that, in rats fed a low-Lys diet, serum lysine levels were decreased, while serum threonine levels were significantly increased compared to the control rats. When the threonine content was restricted in the low-Lys diet, TAG accumulation induced by the low-Lys diet was completely abolished in skeletal muscle. Moreover, in L6 myotubes cultured in medium containing high threonine and low lysine, fatty acid uptake was enhanced compared to that in cells cultured in control medium. These findings suggest that the increased serum threonine in rats fed a low-Lys diet resulted in lipid incorporation into skeletal muscle, leading to the formation of fatty muscle tissue. Collectively, we propose conceptual hypothesis that "amino-acid signal" based on lysine and threonine regulates lipid metabolism.

Keywords:  amino acid; amino acid restriction; fatty acid; fiber twitch; lipid accumulation; lipid metabolism; lipid transport; lysine; muscle; threonine

DOI:  https://doi.org/10.1016/j.jbc.2021.101179