bims-moremu 2023-03-19 papers

bims-moremu

Biomed News

on Molecular regulators of muscle mass

Issue of 2023–03–19
thirty-two papers selected by
Anna Vainshtein, Craft Science Inc.

Distinct myofibre domains of the human myotendinous junction revealed by single nucleus RNA-seq.
Important Concepts in Protein Nutrition, Aging, and Skeletal Muscle: Honoring Dr Douglas Paddon-Jones (1969-2021) by Highlighting His Research Contributions.
Inhibition of ATP synthase reverse activity restores energy homeostasis in mitochondrial pathologies.
Effect of heme oxygenase-1 on the differentiation of human myoblasts and the regeneration of murine skeletal muscles after acute and chronic injury.
microRNA-501 controls myogenin+/CD74+ myogenic progenitor cells during muscle regeneration.
Kcnma1 is involved in mitochondrial homeostasis in diabetes-related skeletal muscle atrophy.
Fibroblast growth factor 21 is expressed and secreted from skeletal muscle following electrical stimulation via extracellular ATP activation of the PI3K/Akt/mTOR signaling pathway.
The MuSK-BMP pathway maintains myofiber size in slow muscle through regulation of Akt- mTOR signaling.
Identification of C-C motif chemokine ligand 5 as a heat-dependent myokine.
Muscle stem cells get a new look: Dynamic cellular projections as sensors of the stem cell niche.
Single-nucleus profiling unveils a geroprotective role of the FOXO3 in primate skeletal muscle aging.
A protocol for single nucleus RNA-seq from frozen skeletal muscle.
Melatonin improves muscle injury and differentiation by increasing Pax7 expression.
RNF7 Induces Skeletal Muscle Cell Apoptosis and Arrests Cell Autophagy via Upregulation of THBS1 and Inactivation of the PI3K/Akt Signaling Pathway in a Rat Sepsis Model.
Single-cell transcriptome dynamics of the autotaxin-lysophosphatidic acid axis during muscle regeneration reveal proliferative effects in mesenchymal fibro-adipogenic progenitors.
The crucial role of muscle glucocorticoid signaling in accelerating obesity and glucose intolerance via hyperinsulinemia.
Delayed denervation-induced muscle atrophy in Opg knockout mice.
Inter-muscular networks of synchronous muscle fiber activation.
Human and African ape myosin heavy chain content and the evolution of hominin skeletal muscle.
Regulation of adult stem cell quiescence and its functions in the maintenance of tissue integrity.
Myoscaffolds reveal laminin scarring is detrimental for stem cell function while sarcospan induces compensatory fibrosis.
Myonuclear alterations after training: Train your muscles, train your nuclei.
Full-length human dystrophin on human artificial chromosome compensates for mouse dystrophin deficiency in a Duchenne muscular dystrophy mouse model.
Low-intensity pulsed ultrasound promotes skeletal muscle regeneration via modulating the inflammatory immune microenvironment.
Link between insulin resistance and skeletal muscle extracellular matrix remodeling.
Microbiota Mediate Enhanced Exercise Capacity Induced by Exercise Training.
Four anti-aging drugs and calorie-restricted diet produce parallel effects in fat, brain, muscle, macrophages, and plasma of young mice.
Genetic modifiers modulate phenotypic expression of tafazzin deficiency in a mouse model of Barth syndrome.
New advancements in CRISPR based gene therapy of Duchenne Muscular dystrophy.
Using a Network Physiology Approach to Prescribe Exercise for Exercise Oncology.
Virus-free transfection, transient expression, and purification of human cardiac myosin in mammalian muscle cells for biochemical and biophysical assays.
Non-invasive estimation of muscle fibre size from high-density electromyography.

J Cell Sci. 2023 Mar 16. pii: jcs.260913. [Epub ahead of print]

Distinct myofibre domains of the human myotendinous junction revealed by single nucleus RNA-seq.

Anders Karlsen, Ching-Yan Chloé Yeung, Peter Schjerling, Linda Denz, Christian Hoegsbjerg, Jens R Jakobsen, Michael R Krogsgaard, Manuel Koch, Stefano Schiaffino, Michael Kjaer, Abigail L Mackey.

  The myotendinous junction (MTJ) is a specialized domain of the multinucleated myofibre, faced with the challenge of maintaining robust cell-matrix contact with the tendon under high mechanical stress and strain. Here, we profiled 24,161 nuclei in semitendinosus muscle-tendon samples from 3 healthy males by single nucleus RNA-sequencing (snRNA-seq), alongside spatial transcriptomics, to gain insight into the genes characterizing this specialization in humans. We identified a cluster of MTJ myonuclei, represented by 47 enriched transcripts, of which the presence of ABI3BP, ABLIM1, ADAMTSL1, BICD1, CPM, FHOD3, FRAS1 and FREM2 was confirmed at the MTJ at the protein level by immunofluorescence. Four distinct subclusters of MTJ myonuclei were apparent and segregated into two COL22A1-expressing subclusters and two lacking COL22A1 but with a clear fibre type profile expressing MYH7 or MYH1/2. Our findings reveal distinct myonuclei profiles of the human MTJ, a weak link in the musculoskeletal system, which is selectively affected in pathological conditions, from muscle strains to muscular dystrophies.

Keywords:  Myofibre domains; Myotendinous junction; SnRNA-seq; Spatial transcriptomics; Tendon

DOI:  https://doi.org/10.1242/jcs.260913
J Nutr. 2023 Mar;pii: S0022-3166(23)02513-0. [Epub ahead of print]153(3): 615-621

Important Concepts in Protein Nutrition, Aging, and Skeletal Muscle: Honoring Dr Douglas Paddon-Jones (1969-2021) by Highlighting His Research Contributions.

Emily J Arentson-Lantz, Donald K Layman, Heather J Leidy, Wayne W Campbell, Stuart M Phillips.

  This review is a tribute to honor Dr Douglas Paddon-Jones by highlighting his career research contributions. Dr Paddon-Jones was a leader in recognizing the importance of muscle health and the interactions of physical activity and dietary protein for optimizing the health span. Aging is characterized by loss of muscle mass and strength associated with reduced rates of muscle protein synthesis (MPS) and the ability to repair and replace muscle proteins. Research from the team at the University of Texas Medical Branch in Galveston discovered that the age-related decline in MPS could be overcome by increasing the quantity or quality of dietary protein at each meal. Dr Paddon-Jones was instrumental in proposing and testing a "protein threshold" of ∼30 g protein/meal to optimize MPS in older adults. Dr Paddon-Jones demonstrated that physical inactivity greatly accelerates the loss of muscle mass and function in older adults. His work in physical activity led him to propose the "Catabolic Crisis Model" of muscle size and function losses, suggesting that age-related muscle loss is not a linear process, but the result of acute periods of disuse associated with injuries, illnesses, and bed rest. This model creates the opportunity to provide targeted interventions via protein supplementation and/or increased dietary protein through consuming high-quality animal-source foods. He illustrated that nutritional support, particularly enhanced protein quantity, quality, and meal distribution, can help preserve muscle health during periods of inactivity and promote health across the life course.

Keywords:  disuse; meal patterning; muscle protein synthesis; older adults; physical activity

DOI:  https://doi.org/10.1016/j.tjnut.2023.01.011
EMBO J. 2023 Mar 13. e111699

Inhibition of ATP synthase reverse activity restores energy homeostasis in mitochondrial pathologies.

Rebeca Acin-Perez, Cristiane Benincá, Lucia Fernandez Del Rio, Cynthia Shu, Siyouneh Baghdasarian, Vanessa Zanette, Christoph Gerle, Chimari Jiko, Ramzi Khairallah, Shaharyar Khan, David Rincon Fernandez Pacheco, Byourak Shabane, Karel Erion, Ruchi Masand, Sundeep Dugar, Cristina Ghenoiu, George Schreiner, Linsey Stiles, Marc Liesa, Orian S Shirihai.

  The maintenance of cellular function relies on the close regulation of adenosine triphosphate (ATP) synthesis and hydrolysis. ATP hydrolysis by mitochondrial ATP Synthase (CV) is induced by loss of proton motive force and inhibited by the mitochondrial protein ATPase inhibitor (ATPIF1). The extent of CV hydrolytic activity and its impact on cellular energetics remains unknown due to the lack of selective hydrolysis inhibitors of CV. We find that CV hydrolytic activity takes place in coupled intact mitochondria and is increased by respiratory chain defects. We identified (+)-Epicatechin as a selective inhibitor of ATP hydrolysis that binds CV while preventing the binding of ATPIF1. In cells with Complex-III deficiency, we show that inhibition of CV hydrolytic activity by (+)-Epichatechin is sufficient to restore ATP content without restoring respiratory function. Inhibition of CV-ATP hydrolysis in a mouse model of Duchenne Muscular Dystrophy is sufficient to improve muscle force without any increase in mitochondrial content. We conclude that the impact of compromised mitochondrial respiration can be lessened using hydrolysis-selective inhibitors of CV.

Keywords:  ATP hydrolysis; ATPase Inhibitor (ATPIF1); Complex V; epicatechin; muscular dystrophy

DOI:  https://doi.org/10.15252/embj.2022111699
Pharmacol Rep. 2023 Mar 15.

Effect of heme oxygenase-1 on the differentiation of human myoblasts and the regeneration of murine skeletal muscles after acute and chronic injury.

Urszula Głowniak-Kwitek, Asier Laria Caballero, Iwona Bronisz-Budzyńska, Magdalena Kozakowska, Kalina Andrysiak, Jacek Stępniewski, Agnieszka Łoboda, Józef Dulak.

   BACKGROUND: Impaired muscle regeneration is a hallmark of Duchenne muscular dystrophy (DMD), a neuromuscular disorder caused by mutations in the DMD gene encoding dystrophin. The lack of heme oxygenase-1 (HO-1, Hmox1), a known anti-inflammatory and cytoprotective enzyme, was shown to aggravate DMD pathology.
METHODS: We evaluated the role of HO-1 overexpression in human induced pluripotent stem cell (hiPSC)-derived skeletal muscle cells (hiPSC-SkM) in vitro and in the regeneration process in vivo in wild-type mice. Furthermore, the effect of cobalt protoporphyrin IX (CoPP), a pharmacological inducer of HO-1 expression, on regeneration markers during myogenic hiPSC differentiation and progression of the dystrophic phenotype was analysed in the mdx mouse DMD model.
RESULTS: HO-1 has an impact on hiPSC-SkM generation by decreasing cell fusion capacity and the expression of myogenic regulatory factors and muscle-specific microRNAs (myomiRs). Also, strong induction of HO-1 by CoPP totally abolished hiPSC-SkM differentiation. Injection of HO-1-overexpressing hiPSC-SkM into the cardiotoxin (CTX)-injured muscle of immunodeficient wild-type mice was associated with decreased expression of miR-206 and Myh3 and lower number of regenerating fibers, suggesting some advanced regeneration. However, the very potent induction of HO-1 by CoPP did not exert any protective effect on necrosis, leukocyte infiltration, fibrosis, myofiber regeneration biomarkers, and exercise capacity of mdx mice.
CONCLUSIONS: In summary, HO-1 inhibits the expression of differentiation markers in human iPSC-derived myoblasts. Although moderate overexpression of HO-1 in the injected myoblast was associated with partially advanced muscle regeneration, the high systemic induction of HO-1 did not improve muscle regeneration. The appropriate threshold of HO-1 expression must be established for the therapeutic effect of HO-1 on muscle regeneration.

Keywords:  Duchenne muscular dystrophy; HO-1; Induced pluripotent stem cells; Mdx; miR-206; microRNA; myomiRs

DOI:  https://doi.org/10.1007/s43440-023-00475-3
Mol Metab. 2023 Mar 11. pii: S2212-8778(23)00038-8. [Epub ahead of print] 101704

microRNA-501 controls myogenin+/CD74+ myogenic progenitor cells during muscle regeneration.

Alexandra Fahrner, Edlira Luca, Jan Krützfeldt.

   OBJECTIVE: Skeletal muscle regeneration is markedly impaired during aging. How adult muscle stem cells contribute to this decrease in regenerative capacity is incompletely understood. We investigated mechanisms of age-related changes in myogenic progenitor cells using the tissue-specific microRNA 501.
METHODS: Young and old C57Bl/6 mice were used (3 months or 24 months of age, respectively) with or without global or tissue-specific genetic deletion of miR-501. Muscle regeneration was induced using intramuscular cardiotoxin injection or treadmill exercise and analysed using single cell and bulk RNA sequencing, qRT-PCR and immunofluorescence. Muscle fiber damage was assessed with Evan`s blue dye (EBD). In vitro analysis was performed in primary muscle cells obtained from mice and humans.
RESULTS: Single cell sequencing revealed myogenic progenitor cells in miR-501 knockout mice at day 6 after muscle injury that are characterized by high levels of myogenin and CD74. In control mice these cells were less in number and already downregulated after day 3 of muscle injury. Muscle from knockout mice had reduced myofiber size and reduced myofiber resilience to injury and exercise. miR-501 elicits this effect by regulating sarcomeric gene expression through its target gene estrogen-related receptor gamma (Esrrg). Importantly, in aged skeletal muscle where miR-501 was significantly downregulated and its target Esrrg significantly upregulated, the number of myog+/CD74+ cells during regeneration was upregulated to similar levels as observed in 501 knockout mice. Moreover, myog+/CD74+-aged skeletal muscle exhibited a similar decrease in the size of newly formed myofibers and increased number of necrotic myofibers after injury as observed in mice lacking miR-501.
CONCLUSIONS: miR-501 and Esrrg are regulated in muscle with decreased regenerative capacity and loss of miR-501 is permissive to the appearance of CD74+ myogenic progenitors. Our data uncover a novel link between the metabolic transcription factor Esrrg and sarcomere formation and demonstrate that stem cell heterogeneity in skeletal muscle during aging is under miRNA control. Targeting Esrrg or myog+/CD74+ progenitor cells might improve fiber size and myofiber resilience to exercise in aged skeletal muscle.

Keywords:  CD74; Esrrg; aging; microRNA; regeneration; skeletal muscle; stem cells

DOI:  https://doi.org/10.1016/j.molmet.2023.101704
FASEB J. 2023 Apr;37(4): e22866

Kcnma1 is involved in mitochondrial homeostasis in diabetes-related skeletal muscle atrophy.

Shan-Yan Gao, Yong-Ping Liu, Ri Wen, Xin-Mei Huang, Ping Li, Yu-Hang Yang, Ni Yang, Tie-Ning Zhang.

  Uncontrolled diabetes causes a catabolic state with multi-organic complications, of which impairment on skeletal muscle contributes to the damaged mobility. Kcnma1 gene encodes the pore-forming α-subunit of Ca2+ - and voltage-gated K+ channels of large conductance (BK channels), and loss-of-function mutations in Kcnma1 are in regards to impaired myogenesis. Herein, we observed a time-course reduction of Kcnma1 expression in the tibialis anterior muscles of leptin receptor-deficient (db/db) diabetic mice. To investigate the role of Kcnma1 in diabetic muscle atrophy, muscle-specific knockdown of Kcnma1 was achieved by mice receiving intravenous injection of adeno-associated virus-9 (AAV9)-encoding shRNA against Kcnma1 under the muscle creatine kinase (MCK) promoter. Impairment on muscle mass and myogenesis were observed in m/m mice with AAV9-shKcnma1 intervention, while this impairment was more obvious in diabetic db/db mice. Simultaneously, damaged mitochondrial dynamics and biogenesis showed much severer in db/db mice with AAV9-shKcnma1 intervention. RNA sequencing revealed the large transcriptomic changes resulted by Kcnma1 knockdown, and changes in mitochondrial homeostasis-related genes were validated. Besides, the artificial alteration of Kcnma1 in mouse C2C12 myoblasts was achieved with an adenovirus vector. Consistent results were demonstrated by Kcnma1 knockdown in palmitate-treated cells, whereas opposite results were exhibited by Kcnma1 overexpression. Collectively, we document Kcnma1 as a potential keeper of mitochondrial homeostasis, and the loss of Kcnma1 is a critical event in priming skeletal muscle loss in diabetes.

Keywords:  Kcnma1; diabetes; mitochondrial homeostasis; skeletal muscle loss

DOI:  https://doi.org/10.1096/fj.202201397RR
Front Endocrinol (Lausanne). 2023 ;14 1059020

Fibroblast growth factor 21 is expressed and secreted from skeletal muscle following electrical stimulation via extracellular ATP activation of the PI3K/Akt/mTOR signaling pathway.

Manuel Arias-Calderón, Mariana Casas, Julián Balanta-Melo, Camilo Morales-Jiménez, Nadia Hernández, Paola Llanos, Enrique Jaimovich, Sonja Buvinic.

  Fibroblast growth factor 21 (FGF21) is a hormone involved in the regulation of lipid, glucose, and energy metabolism. Although it is released mainly from the liver, in recent years it has been shown that it is a "myokine", synthesized in skeletal muscles after exercise and stress conditions through an Akt-dependent pathway and secreted for mediating autocrine and endocrine roles. To date, the molecular mechanism for the pathophysiological regulation of FGF21 production in skeletal muscle is not totally understood. We have previously demonstrated that muscle membrane depolarization controls gene expression through extracellular ATP (eATP) signaling, by a mechanism defined as "Excitation-Transcription coupling". eATP signaling regulates the expression and secretion of interleukin 6, a well-defined myokine, and activates the Akt/mTOR signaling pathway. This work aimed to study the effect of electrical stimulation in the regulation of both production and secretion of skeletal muscle FGF21, through eATP signaling and PI3K/Akt pathway. Our results show that electrical stimulation increases both mRNA and protein (intracellular and secreted) levels of FGF21, dependent on an extracellular ATP signaling mechanism in skeletal muscle. Using pharmacological inhibitors, we demonstrated that FGF21 production and secretion from muscle requires the activation of the P2YR/PI3K/Akt/mTOR signaling pathway. These results confirm skeletal muscle as a source of FGF21 in physiological conditions and unveil a new molecular mechanism for regulating FGF21 production in this tissue. Our results will allow to identify new molecular targets to understand the regulation of FGF21 both in physiological and pathological conditions, such as exercise, aging, insulin resistance, and Duchenne muscular dystrophy, all characterized by an alteration in both FGF21 levels and ATP signaling components. These data reinforce that eATP signaling is a relevant mechanism for myokine expression in skeletal muscle.

Keywords:  Akt signaling; excitation-transcription coupling; extracellular nucleotides; fibroblast growth factor 21; membrane depolarization; muscle activity; myokines

DOI:  https://doi.org/10.3389/fendo.2023.1059020
Res Sq. 2023 Feb 27. pii: rs.3.rs-2613527. [Epub ahead of print]

The MuSK-BMP pathway maintains myofiber size in slow muscle through regulation of Akt- mTOR signaling.

Diego Jaime, Lauren A Fish, Laura A Madigan, Madison D Ewing, Justin R Fallon.

Myofiber size regulation is critical in health, disease, and aging. MuSK (muscle-specific kinase) is a BMP (bone morphogenetic protein) co-receptor that promotes and shapes BMP signaling. MuSK is expressed at all neuromuscular junctions and is also present extrasynaptically in the slow soleus muscle. To investigate the role of the MuSK-BMP pathway in vivo we generated mice lacking the BMP-binding MuSK Ig3 domain. These âˆ†Ig3-MuSKmice are viable and fertile with innervation levels comparable to wild type. In 3-month-old mice myofibers are smaller in the slow soleus, but not in the fast tibialis anterior (TA). Transcriptomic analysis revealed soleus-selective decreases in RNA metabolism and protein synthesis pathways as well as dysregulation of IGF1-Akt-mTOR pathway components. Biochemical analysis showed that Akt-mTOR signaling is reduced in soleus but not TA. We propose that the MuSK-BMP pathway acts extrasynaptically to maintain myofiber size in slow muscle by promoting protein synthetic pathways including IGF1-Akt-mTOR signaling. These results reveal a novel mechanism for regulating myofiber size in slow muscle and introduce the MuSK-BMP pathway as a target for promoting muscle growth and combatting atrophy.

DOI: https://doi.org/10.21203/rs.3.rs-2613527/v1
Endocr J. 2023 Mar 11.

Identification of C-C motif chemokine ligand 5 as a heat-dependent myokine.

Keigo Murata, Yuri Ishiuchi-Sato, Taku Nedachi.

  The skeletal muscle is an endocrine organ that produces proteins and peptides, collectively termed as myokines. The temperature of skeletal muscles varies during exercise and/or with changes in ambient temperature. However, whether myokine secretion is regulated by heat stimulation is unclear. Thus, we aimed to explore the effects of environmental heat stimulation on myokine secretion. We initially investigated the secretome of C2C12 myotubes and identified several novel heat-responsive myokines. The concentration of C-C motif chemokine ligand 5 (CCL5) dramatically decreased by 0.3-fold in response to heat stress. After 3 h heat stimulation of C2C12 cells, the expression of heat shock protein 70 was induced, and the gene expression and secretion of CCL5 was significantly attenuated in C2C12 cells. We then examined the effects of acute heat stress on serum CCL5 levels in mice and Ccl5 gene expression in skeletal muscles. Mice were maintained at 23°C, exposed to 45°C for 30 min, and then returned to the 23°C chamber for recovery. The expression of Ccl5 in the skeletal muscle significantly decreased after 3 h of recovery. Serum CCL5 levels increased by approximately 1.9-fold after 30 min of heat exposure and then significantly decreased by approximately 0.7-fold after 23 h of recovery. This study suggests that heat stimulation decreases CCL5 secretion from the skeletal muscle in vitro and in vivo. Given its fundamental role in inflammation by recruiting several immune cells, CCL5 has a potential role in controlling inflammatory responses in the body after heat stimulation.

Keywords:  C-C motif chemokine ligand 5 (CCL5); Heat; Myokines; Skeletal muscle

DOI:  https://doi.org/10.1507/endocrj.EJ22-0611
Bioessays. 2023 Mar 14. e2200249

Muscle stem cells get a new look: Dynamic cellular projections as sensors of the stem cell niche.

Robert S Krauss, Allison P Kann.

  Cellular mechanisms whereby quiescent stem cells sense tissue injury and transition to an activated state are largely unknown. Quiescent skeletal muscle stem cells (MuSCs, also called satellite cells) have elaborate, heterogeneous projections that rapidly retract in response to muscle injury. They may therefore act as direct sensors of their niche environment. Retraction is driven by a Rac-to-Rho GTPase activity switch that promotes downstream MuSC activation events. These and other observations lead to several hypotheses: (1) projections are morphologically dynamic at quiescence, providing a surveillance function for muscle damage; (2) quiescent projection dynamics are regulated by the relative balance of Rac and Rho activities promoted by niche-derived cues; (3) projections, particularly their associated filopodia, sense tissue damage via changes to the biomechanical properties of the niche and/or detection of signaling cues released by damaged myofibers; and (4) the dynamic nature of projections result in a population of MuSCs with heterogeneous functional properties. These concepts may extend to other types of quiescent stem cells, as well as prove useful in translational research settings.

Keywords:  Rac1; Rho; filopodia; mechanosignaling; muscle stem cell; quiescence; regeneration

DOI:  https://doi.org/10.1002/bies.202200249
Protein Cell. 2022 Nov 22. pii: pwac061. [Epub ahead of print]

Single-nucleus profiling unveils a geroprotective role of the FOXO3 in primate skeletal muscle aging.

Ying Jing, Yuesheng Zuo, Yang Yu, Liang Sun, Zhengrong Yu, Shuai Ma, Qian Zhao, Guoqiang Sun, Huifang Hu, Jingyi Li, Daoyuan Huang, Lixiao Liu, Jiaming Li, Zijuan Xin, Haoyan Huang, Juan Carlos Izpisua Belmonte, Weiqi Zhang, Si Wang, Jing Qu, Guang-Hui Liu.

  Age-dependent loss of skeletal muscle mass and function is a feature of sarcopenia, and increases the risk of many aging-related metabolic diseases. Here, we report phenotypic and single-nucleus transcriptomic analyses of non-human primate skeletal muscle aging. A higher transcriptional fluctuation was observed in myonuclei relative to other interstitial cell types, indicating a higher susceptibility of skeletal muscle fiber to aging. We found a downregulation of FOXO3 in aged primate skeletal muscle, and identified FOXO3 as a hub transcription factor maintaining skeletal muscle homeostasis. Through the establishment of a complementary experimental pipeline based on a human pluripotent stem cell-derived myotube model, we revealed that silence of FOXO3 accelerates human myotube senescence, whereas genetic activation of endogenous FOXO3 alleviates human myotube aging. Altogether, based on a combination of monkey skeletal muscle and human myotube aging research models, we unraveled the pivotal role of the FOXO3 in safeguarding primate skeletal muscle from aging, providing a comprehensive resource for the development of clinical diagnosis and targeted therapeutic interventions against human skeletal muscle aging and the onset of sarcopenia along with aging-related disorders.

Keywords:  FOXO3; aging; primate; single-nucleus RNA sequencing; skeletal muscle

DOI:  https://doi.org/10.1093/procel/pwac061
Life Sci Alliance. 2023 May;pii: e202201806. [Epub ahead of print]6(5):

A protocol for single nucleus RNA-seq from frozen skeletal muscle.

Tyler Gb Soule, Carly S Pontifex, Nicole Rosin, Matthew M Joel, Sukyoung Lee, Minh Dang Nguyen, Sameer Chhibber, Gerald Pfeffer.

Single-cell technologies are a method of choice to obtain vast amounts of cell-specific transcriptional information under physiological and diseased states. Myogenic cells are resistant to single-cell RNA sequencing because of their large, multinucleated nature. Here, we report a novel, reliable, and cost-effective method to analyze frozen human skeletal muscle by single-nucleus RNA sequencing. This method yields all expected cell types for human skeletal muscle and works on tissue frozen for long periods of time and with significant pathological changes. Our method is ideal for studying banked samples with the intention of studying human muscle disease.

DOI: https://doi.org/10.26508/lsa.202201806
Int J Biol Sci. 2023 ;19(4): 1049-1062

Melatonin improves muscle injury and differentiation by increasing Pax7 expression.

Chen-Ming Su, Chun-Hao Tsai, Hsien-Te Chen, Yi-Syuan Wu, Jun-Way Chang, Shun-Fa Yang, Chih-Hsin Tang.

  A balance between muscle injury and regeneration is critical for sustaining muscle function during myogenesis. Melatonin is well recognized for its involvement in neuroprotective activities, immune system regulation and suppression of inflammatory responses. This study set out to provide evidence that melatonin improves muscle regeneration during skeletal muscle differentiation. We began with cloning a stable cell line expressing Pax7 knockdown C2C12 cells. We then investigated markers of muscle degradation and regeneration after treating growth medium and differentiated medium with melatonin. Bioinformatics analysis of RNA sequencing results revealed that melatonin regulates muscle differentiation and that Wnt cascades are involved in the mechanism of muscle differentiation. Screening of miRNA online databases revealed that miR-3475-3p is a specific binding site on Pax7 and acts as a negative regulator of Pax7, which is involved in melatonin-induced muscle differentiation. We then investigated the effects of melatonin treatment in the early stage of glycerol-induced skeletal muscle injury in mice. Rotarod performance, micro-computed tomography and immunohistochemistry findings showed that melatonin-induced increases in Pax7 expression rapidly rescue skeletal muscle differentiation and improve muscle fiber morphology in glycerol-induced muscle injury. Our data support the hypothesis that melatonin rapidly rescues skeletal muscle differentiation and the melatonin/Pax7 axis could therefore serve as an important therapeutic target to optimize muscle healing after injury.

Keywords:  Pax7; differentiation; melatonin; miR-3475-3p; muscle injury

DOI:  https://doi.org/10.7150/ijbs.79169
Infect Immun. 2023 Mar 15. e0053522

RNF7 Induces Skeletal Muscle Cell Apoptosis and Arrests Cell Autophagy via Upregulation of THBS1 and Inactivation of the PI3K/Akt Signaling Pathway in a Rat Sepsis Model.

Yu Lei, Xiaoyuan Jin, Mingli Sun, Zhiyong Ji.

  Recently, long noncoding RNAs (lncRNAs) have been highlighted for extensive functionality in sepsis. In this study, we aimed to explore the role of RNF7 in the progression of sepsis. We initially established a rat model of sepsis through cecal ligation and puncture induction, whereupon RNF7 expression was determined by RT-qPCR. Following adenovirus infection, the role of RNF7 in muscle injury, skeletal muscle protein metabolism, oxidative stress, and inflammation in sepsis rats was analyzed. Then, downstream mechanisms of RNF7 were identified and validated. Further, lipopolysaccharide was applied to treat myoblast to further demonstrate the in vitro role of RNF7. Our results showed that RNF7 expression was upregulated during sepsis. Overexpression of RNF7 worsened the sepsis-induced skeletal muscle injury, induced skeletal muscle protein metabolism, oxidative stress, and inflammation in sepsis rats. Meanwhile, overexpression of RNF7 elevated thrombospondin-1 (THBS1) expression. Silencing of RNF7 inhibited THBS1 and activated the PI3K/Akt signaling pathway, arresting the release of inflammatory factors and oxidative stress levels in skeletal muscle cells. Altogether, RNF7 may promote skeletal muscle cell apoptosis while simultaneously inhibiting cell autophagy through the promotion of THBS1 and inactivation of the PI3K/Akt signaling pathway.

Keywords:  PI3K/Akt signaling pathway; THBS1; apoptosis; autophagy; long noncoding RNA RNF7; sepsis; skeletal muscle cells

DOI:  https://doi.org/10.1128/iai.00535-22
Front Cell Dev Biol. 2023 ;11 1017660

Single-cell transcriptome dynamics of the autotaxin-lysophosphatidic acid axis during muscle regeneration reveal proliferative effects in mesenchymal fibro-adipogenic progenitors.

Osvaldo Contreras, Richard P Harvey.

  Lysophosphatidic acid is a growth factor-like bioactive phospholipid recognising LPA receptors and mediating signalling pathways that regulate embryonic development, wound healing, carcinogenesis, and fibrosis, via effects on cell migration, proliferation and differentiation. Extracellular LPA is generated from lysophospholipids by the secreted hydrolase-ectonucleotide pyrophosphatase/phosphodiesterase 2 (ENPP2; also, AUTOTAXIN/ATX) and metabolised by different membrane-bound phospholipid phosphatases (PLPPs). Here, we use public bulk and single-cell RNA sequencing datasets to explore the expression of Lpar 1-6, Enpp2, and Plpp genes under skeletal muscle homeostasis and regeneration conditions. We show that the skeletal muscle system dynamically expresses the Enpp2-Lpar-Plpp gene axis, with Lpar1 being the highest expressed member among LPARs. Lpar1 was expressed by mesenchymal fibro-adipogenic progenitors and tenocytes, whereas FAPs mainly expressed Enpp2. Clustering of FAPs identified populations representing distinct cell states with robust Lpar1 and Enpp2 transcriptome signatures in homeostatic cells expressing higher levels of markers Dpp4 and Hsd11b1. However, tissue injury induced transient repression of Lpar genes and Enpp2. The role of LPA in modulating the fate and differentiation of tissue-resident FAPs has not yet been explored. Ex vivo, LPAR1/3 and ENPP2 inhibition significantly decreased the cell-cycle activity of FAPs and impaired fibro-adipogenic differentiation, implicating LPA signalling in the modulation of the proliferative and differentiative fate of FAPs. Together, our results demonstrate the importance of the ENPP2-LPAR-PLPP axis in different muscle cell types and FAP lineage populations in homeostasis and injury, paving the way for further research on the role of this signalling pathway in skeletal muscle homeostasis and regeneration, and that of other organs and tissues, in vivo.

Keywords:  ENPP2; autotaxin (ATX); ectonucleotide pyrophosphatase/phosphodiesterase family member 2; fibro-adipogenic progenitors (FAPs); lysophosphatidic acid (LPA); muscle stem cells; myogenesis; regeneration

DOI:  https://doi.org/10.3389/fcell.2023.1017660
JCI Insight. 2023 Mar 14. pii: e162382. [Epub ahead of print]

The crucial role of muscle glucocorticoid signaling in accelerating obesity and glucose intolerance via hyperinsulinemia.

Hiroki Yamazaki, Masaaki Uehara, Noritada Yoshikawa, Akiko Kuribara-Souta, Motohisa Yamamoto, Yasuko Hirakawa, Yasuaki Kabe, Makoto Suematsu, Hirotoshi Tanaka.

  Metabolic crosstalk from skeletal muscle to multiple organs is important for maintaining homeostasis, and its dysregulation can lead to various diseases. Chronic glucocorticoid administration often induces muscle atrophy and metabolic disorders such as diabetes and central obesity; however, the detailed underlying mechanism remains unclear. We previously reported that the deletion of glucocorticoid receptor (GR) in skeletal muscle increases muscle mass and reduces fat mass through muscle-liver-fat communication under physiological conditions. In this study, we show that muscle GR signaling plays a crucial role in accelerating obesity through the induction of hyperinsulinemia. Fat accumulation in liver and adipose tissue, muscle atrophy, hyperglycemia, and hyperinsulinemia induced by chronic corticosterone (CORT) treatment improved in muscle-specific GR knockout (GRmKO) mice. Such CORT-induced fat accumulation was alleviated by suppressing insulin production (streptozotocin injection), indicating that hyperinsulinemia enhanced by muscle GR signaling promotes obesity. Strikingly, glucose intolerance and obesity in ob/ob mice without CORT treatment were also improved in GRmKO mice, indicating that muscle GR signaling contributes to obesity-related metabolic changes, regardless of systemic glucocorticoid levels. Thus, this study provides new insight for the treatment of obesity and diabetes by targeting muscle GR signaling.

Keywords:  Endocrinology; Insulin; Muscle Biology; Obesity

DOI:  https://doi.org/10.1172/jci.insight.162382
Front Physiol. 2023 ;14 1127474

Delayed denervation-induced muscle atrophy in Opg knockout mice.

Mingming Zhang, Ming Chen, Yi Li, Man Rao, Duanyang Wang, Zhongqi Wang, Licheng Zhang, Pengbin Yin, Peifu Tang.

  Recent evidence has shown a crucial role for the osteoprotegerin/receptor activator of nuclear factor κ-B ligand/RANK (OPG/RANKL/RANK) signaling axis not only in bone but also in muscle tissue; however, there is still a lack of understanding of its effects on muscle atrophy. Here, we found that denervated Opg knockout mice displayed better functional recovery and delayed muscle atrophy, especially in a specific type IIB fiber. Moreover, OPG deficiency promoted milder activation of the ubiquitin-proteasome pathway, which further verified the protective role of Opg knockout in denervated muscle damage. Furthermore, transcriptome sequencing indicated that Opg knockout upregulated the expression of Inpp5k, Rbm3, and Tet2 and downregulated that of Deptor in denervated muscle. In vitro experiments revealed that satellite cells derived from Opg knockout mice displayed a better differentiation ability than those acquired from wild-type littermates. Higher expression levels of Tet2 were also observed in satellite cells derived from Opg knockout mice, which provided a possible mechanistic basis for the protective effects of Opg knockout on muscle atrophy. Taken together, our findings uncover the novel role of Opg in muscle atrophy process and extend the current understanding in the OPG/RANKL/RANK signaling axis.

Keywords:  OPG/RANKL/RANK; denervation; muscle atrophy; osteoporosis; transcriptome sequencing

DOI:  https://doi.org/10.3389/fphys.2023.1127474
Front Netw Physiol. 2022 ;2 1059793

Inter-muscular networks of synchronous muscle fiber activation.

Sergi Garcia-Retortillo, Plamen Ch Ivanov.

  Skeletal muscles continuously coordinate to facilitate a wide range of movements. Muscle fiber composition and timing of activation account for distinct muscle functions and dynamics necessary to fine tune muscle coordination and generate movements. Here we address the fundamental question of how distinct muscle fiber types dynamically synchronize and integrate as a network across muscles with different functions. We uncover that physiological states are characterized by unique inter-muscular network of muscle fiber cross-frequency interactions with hierarchical organization of distinct sub-networks and modules, and a stratification profile of links strength specific for each state. We establish how this network reorganizes with transition from rest to exercise and fatigue-a complex process where network modules follow distinct phase-space trajectories reflecting their functional role in movements and adaptation to fatigue. This opens a new area of research, Network Physiology of Exercise, leading to novel network-based biomarkers of health, fitness and clinical conditions.

Keywords:  complex systems; dynamic networks; fatigue; inter-muscular coordination; muscle fibers; network physiology; synchronization

DOI:  https://doi.org/10.3389/fnetp.2022.1059793
Comp Biochem Physiol A Mol Integr Physiol. 2023 Mar 15. pii: S1095-6433(23)00048-X. [Epub ahead of print] 111415

Human and African ape myosin heavy chain content and the evolution of hominin skeletal muscle.

Samantha R Queeno, Peter J Reiser, Caley M Orr, Terence D Capellini, Kirstin N Sterner, Matthew C O'Neill.

  Humans are unique among terrestrial mammals in our manner of walking and running, reflecting 7 to 8 Ma of musculoskeletal evolution since diverging with the genus Pan. One component of this is a shift in our skeletal muscle biology towards a predominance of myosin heavy chain (MyHC) I isoforms (i.e. slow fibers) across our pelvis and lower limbs, which distinguishes us from chimpanzees. Here, new MyHC data from 35 pelvis and hind limb muscles of a Western gorilla (Gorilla gorilla) are presented. These data are combined with a similar chimpanzee dataset to assess the MyHC I content of humans in comparison to African apes (chimpanzees and gorillas) and other terrestrial mammals. The responsiveness of human skeletal muscle to behavioral interventions is also compared to the human-African ape differential. Humans are distinct from African apes and among a small group of terrestrial mammals whose pelvis and hind/lower limb muscle is slow fiber dominant, on average. Behavioral interventions, including immobilization, bed rest, spaceflight and exercise, can induce modest decreases and increases in human MyHC I content (i.e. -9.3% to 2.3%, n = 2033 subjects), but these shifts are much smaller than the mean human-African ape differential (i.e. 31%). Taken together, these results indicate muscle fiber content is likely an evolvable trait under selection in the hominin lineage. As such, we highlight potential targets of selection in the genome (e.g. regions that regulate MyHC content) that may play an important role in hominin skeletal muscle evolution.

Keywords:  Chimpanzee; Enhancers; Fiber type; Gorilla; Hind limb; Lower limb; Myosin heavy chain I

DOI:  https://doi.org/10.1016/j.cbpa.2023.111415
Nat Rev Mol Cell Biol. 2023 Mar 15.

Regulation of adult stem cell quiescence and its functions in the maintenance of tissue integrity.

Antoine de Morree, Thomas A Rando.

Adult stem cells are important for mammalian tissues, where they act as a cell reserve that supports normal tissue turnover and can mount a regenerative response following acute injuries. Quiescent stem cells are well established in certain tissues, such as skeletal muscle, brain, and bone marrow. The quiescent state is actively controlled and is essential for long-term maintenance of stem cell pools. In this Review, we discuss the importance of maintaining a functional pool of quiescent adult stem cells, including haematopoietic stem cells, skeletal muscle stem cells, neural stem cells, hair follicle stem cells, and mesenchymal stem cells such as fibro-adipogenic progenitors, to ensure tissue maintenance and repair. We discuss the molecular mechanisms that regulate the entry into, maintenance of, and exit from the quiescent state in mice. Recent studies revealed that quiescent stem cells have a discordance between RNA and protein levels, indicating the importance of post-transcriptional mechanisms, such as alternative polyadenylation, alternative splicing, and translation repression, in the control of stem cell quiescence. Understanding how these mechanisms guide stem cell function during homeostasis and regeneration has important implications for regenerative medicine.

DOI: https://doi.org/10.1038/s41580-022-00568-6
NPJ Regen Med. 2023 Mar 15. 8(1): 16

Myoscaffolds reveal laminin scarring is detrimental for stem cell function while sarcospan induces compensatory fibrosis.

Kristen M Stearns-Reider, Michael R Hicks, Katherine G Hammond, Joseph C Reynolds, Alok Maity, Yerbol Z Kurmangaliyev, Jesse Chin, Adam Z Stieg, Nicholas A Geisse, Sophia Hohlbauch, Stefan Kaemmer, Lauren R Schmitt, Thanh T Pham, Ken Yamauchi, Bennett G Novitch, Roy Wollman, Kirk C Hansen, April D Pyle, Rachelle H Crosbie.

We developed an on-slide decellularization approach to generate acellular extracellular matrix (ECM) myoscaffolds that can be repopulated with various cell types to interrogate cell-ECM interactions. Using this platform, we investigated whether fibrotic ECM scarring affected human skeletal muscle progenitor cell (SMPC) functions that are essential for myoregeneration. SMPCs exhibited robust adhesion, motility, and differentiation on healthy muscle-derived myoscaffolds. All SPMC interactions with fibrotic myoscaffolds from dystrophic muscle were severely blunted including reduced motility rate and migration. Furthermore, SMPCs were unable to remodel laminin dense fibrotic scars within diseased myoscaffolds. Proteomics and structural analysis revealed that excessive collagen deposition alone is not pathological, and can be compensatory, as revealed by overexpression of sarcospan and its associated ECM receptors in dystrophic muscle. Our in vivo data also supported that ECM remodeling is important for SMPC engraftment and that fibrotic scars may represent one barrier to efficient cell therapy.

DOI: https://doi.org/10.1038/s41536-023-00287-2
J Physiol. 2023 Mar 17.

Myonuclear alterations after training: Train your muscles, train your nuclei.

Zakarias Kefil Jensen Ogueboule.



Keywords:  Ageing; Exercise; Mechanotransduction; Nuclear lamina; Nuclear shape; Rho GTPases; Sarcopenia

DOI:  https://doi.org/10.1113/JP284476
Sci Rep. 2023 Mar 16. 13(1): 4360

Full-length human dystrophin on human artificial chromosome compensates for mouse dystrophin deficiency in a Duchenne muscular dystrophy mouse model.

Yosuke Hiramuki, Satoshi Abe, Narumi Uno, Kanako Kazuki, Shuta Takata, Hitomaru Miyamoto, Haruka Takayama, Kayoko Morimoto, Shoko Takehara, Mitsuhiko Osaki, Jun Tanihata, Shin'ichi Takeda, Kazuma Tomizuka, Mitsuo Oshimura, Yasuhiro Kazuki.

Dystrophin maintains membrane integrity as a sarcolemmal protein. Dystrophin mutations lead to Duchenne muscular dystrophy, an X-linked recessive disorder. Since dystrophin is one of the largest genes consisting of 79 exons in the human genome, delivering a full-length dystrophin using virus vectors is challenging for gene therapy. Human artificial chromosome is a vector that can load megabase-sized genome without any interference from the host chromosome. Chimeric mice carrying a 2.4-Mb human dystrophin gene-loaded human artificial chromosome (DYS-HAC) was previously generated, and dystrophin expression from DYS-HAC was confirmed in skeletal muscles. Here we investigated whether human dystrophin expression from DYS-HAC rescues the muscle phenotypes seen in dystrophin-deficient mice. Human dystrophin was normally expressed in the sarcolemma of skeletal muscle and heart at expected molecular weights, and it ameliorated histological and functional alterations in dystrophin-deficient mice. These results indicate that the 2.4-Mb gene is enough for dystrophin to be correctly transcribed and translated, improving muscular dystrophy. Therefore, this technique using HAC gives insight into developing new treatments and novel humanized Duchenne muscular dystrophy mouse models with human dystrophin gene mutations.

DOI: https://doi.org/10.1038/s41598-023-31481-3
Int J Biol Sci. 2023 ;19(4): 1123-1145

Low-intensity pulsed ultrasound promotes skeletal muscle regeneration via modulating the inflammatory immune microenvironment.

Haocheng Qin, Zhiwen Luo, Yaying Sun, Zhong He, Beijie Qi, Yisheng Chen, Junlong Wang, Ce Li, Weiwei Lin, Zhihua Han, Yulian Zhu.

  Background: Low-intensity pulsed ultrasound (LIPUS, a form of mechanical stimulation) can promote skeletal muscle functional repair, but a lack of mechanistic understanding of its relationship and tissue regeneration limits progress in this field. We investigated the hypothesis that specific energy levels of LIPUS mediates skeletal muscle regeneration by modulating the inflammatory microenvironment. Methods: To address these gaps, LIPUS irritation was applied in vivo for 5 min at two different intensities (30mW/cm2 and 60mW/cm2) in next 7 consecutive days, and the treatment begun at 24h after air drop-induced contusion injury. In vitro experiments, LIPUS irritation was applied at three different intensities (30mW/cm2, 45mW/cm2, and 60mW/cm2) for 2 times 24h after introduction of LPS in RAW264.7. Then, we comprehensively assessed the functional and histological parameters of skeletal muscle injury in mice and the phenotype shifting in macrophages through molecular biological methods and immunofluorescence analysis both in vivo and in vitro. Results: We reported that LIPUS therapy at intensity of 60mW/cm2 exhibited the most significant differences in functional recovery of contusion-injured muscle in mice. The comprehensive functional tests and histological analysis in vivo indirectly and directly proved the effectiveness of LIPUS for muscle recovery. Through biological methods and immunofluorescence analysis both in vivo and in vitro, we found that this improvement was attributable in part to the clearance of M1 macrophages populations and the increase in M2 subtypes with the change of macrophage-mediated factors. Depletion of macrophages in vivo eliminated the therapeutic effects of LIPUS, indicating that improvement in muscle function was the result of M2-shifted macrophage polarization. Moreover, the M2-inducing effects of LIPUS were proved partially through the WNT pathway by upregulating FZD5 expression and enhancing β-catenin nuclear translocation in macrophages both in vitro and in vivo. The inhibition and augment of WNT pathway in vitro further verified our results. Conclusion: LIPUS at intensity of 60mW/cm2 could significantly promoted skeletal muscle regeneration through shifting macrophage phenotype from M1 to M2. The ability of LIPUS to direct macrophage polarization may be a beneficial target in the clinical treatment of many injuries and inflammatory diseases.

Keywords:  Inflammation; Low-intensity pulsed ultrasound; Macrophage polarization; Muscle injury; WNT signaling

DOI:  https://doi.org/10.7150/ijbs.79685
Endocr Connect. 2023 Mar 01. pii: EC-23-0023. [Epub ahead of print]

Link between insulin resistance and skeletal muscle extracellular matrix remodeling.

Róża Aleksandrowicz, Marek Straczkowski.

Skeletal muscle is the main metabolic tissue responsible for glucose homeostasis in the body. It is surrounded by the extracellular matrix (ECM) consisting of three layers: epimysium, perimysium, and endomysium. ECM plays an important role in the muscle, as it provides integrity and scaffolding cells. The observed disturbances in this structure are related to the abnormal remodeling of the ECM (through an increase in the concentration of its components). ECM rearrangement may impair insulin action by increasing the physical barrier to insulin transport and reducing insulin transport into muscle cells as well as by directly inhibiting insulin action through integrin signaling. Thus, improper ECM remodeling may contribute to the development of insulin resistance (IR) and related comorbidities. In turn, insulin resistance-associated conditions may further aggravate disturbances of ECM in skeletal muscle. This review describes the major components of the ECM that are necessary for it is proper function. Particular attention was also paid to receptors (integrins) involved in the signaling of metabolic pathways. Finally, changes in ECM components in the context of clinical and animal studies are discussed. This article will help the reader to systematize knowledge related to the ECM and to better understand the relationship between ECM remodeling and IR, and its role in the pathogenesis of T2DM. The information in this article presents the concept of the role of ECM and its remodeling in the pathogenesis of IR, which may contribute to developing new therapeutic solutions.

DOI: https://doi.org/10.1530/EC-23-0023
Med Sci Sports Exerc. 2023 Mar 14.

Microbiota Mediate Enhanced Exercise Capacity Induced by Exercise Training.

Robert A Dowden, Paul J Wisniewski, Candace R Longoria, Marko Oydanich, Tara McNulty, Esther Rodriguez, Jie Zhang, Mark Cavallo, John J Guers, Dorothy E Vatner, Stephen F Vatner, Sara C Campbell.

PURPOSE: We investigated the effects of gut microbes, and the mechanisms mediating the enhanced exercise performance induced by exercise training, i.e., skeletal muscle blood flow, and mitochondrial biogenesis and oxidative function in male mice.
METHODS: All mice received a graded exercise test prior to (PRE) and following exercise training via forced treadmill running at 60-70% of maximal running capacity 5 d/wk for 5 weeks (POST). To examine the role of the gut microbes, the graded exercise was repeated after 7 days of access to antibiotic treated water (ABX), used to eliminate gut microbes. Peripheral blood flow, mitochondrial oxidative capacity, and markers of mitochondrial biogenesis were collected at each time point.
RESULTS: Exercise training led to increases of 60 ± 13% in maximal running distance and 63 ± 11% work to exhaustion (p < 0.001). These increases were abolished following ABX (p < 0.001). Exercise training increased hindlimb blood flow and markers of mitochondrial biogenesis and oxidative function, including AMPK, sirtuin-1, PGC-1α citrate synthase, complex IV, and nitric oxide; all of which were also abolished by ABX treatment.
CONCLUSIONS: Our results support the concept that gut microbiota mediate enhanced exercise capacity following exercise training and the mechanisms responsible, i.e., hindlimb blood flow, mitochondrial biogenesis, and metabolic profile. Finally, results of this study emphasize the need to fully examine the impact of prescribing ABX to athletes during their training regimens and how this may affect their performance.

DOI: https://doi.org/10.1249/MSS.0000000000003170
Geroscience. 2023 Mar 15.

Four anti-aging drugs and calorie-restricted diet produce parallel effects in fat, brain, muscle, macrophages, and plasma of young mice.

Xinna Li, Madaline McPherson, Mary Hager, Michael Lee, Peter Chang, Richard A Miller.

  Average and maximal lifespan can be increased in mice, in one or both sexes, by four drugs: rapamycin, acarbose, 17a-estradiol, and canagliflozin. We show here that these four drugs, as well as a calorie-restricted diet, can induce a common set of changes in fat, macrophages, plasma, muscle, and brain when evaluated in young adults at 12 months of age. These shared traits include an increase in uncoupling protein UCP1 in brown fat and in subcutaneous and intra-abdominal white fat, a decline in proinflammatory M1 macrophages and corresponding increase in anti-inflammatory M2 macrophages, an increase in muscle fibronectin type III domain containing 5 (FNDC5) and its cleavage product irisin, and higher levels of doublecortin (DCX) and brain-derived neurotrophic factor (BDNF) in brain. Each of these proteins is thought to play a role in one or more age-related diseases, including metabolic, inflammatory, and neurodegenerative diseases. We have previously shown that the same suite of changes is seen in each of four varieties of slow-aging single-gene mutant mice. We propose that these changes may be a part of a shared common pathway that is seen in slow-aging mice whether the delayed aging is due to a mutation, a low-calorie diet, or a drug.

Keywords:  Brain-derived neurotrophic factor (BDNF); Doublecortin (DCX); Fibronectin type III domain containing 5 (FNDC5); Inflammatory; Neurodegenerative; Slow-aging mice; Uncoupling protein UCP1

DOI:  https://doi.org/10.1007/s11357-023-00770-0
Hum Mol Genet. 2023 Mar 14. pii: ddad041. [Epub ahead of print]

Genetic modifiers modulate phenotypic expression of tafazzin deficiency in a mouse model of Barth syndrome.

Suya Wang, Erika Yazawa, Erin Keating, Neil Mazumdar, Alexander Hauschild, Qing Ma, Haiyan Wu, Yang Xu, Xu Shi, Douglas Strathdee, Robert E Gerszten, Michael Schlame, William T Pu.

Barth syndrome is an X-linked disorder caused by loss-of-function mutations in Tafazzin (TAZ), an acyltransferase that catalyzes remodeling of cardiolipin, a signature phospholipid of the inner mitochondrial membrane. Patients develop cardiac and skeletal muscle weakness, growth delay, and neutropenia, although phenotypic expression varies considerably between patients. Taz knockout mice recapitulate many of the hallmark features of the disease. We used mouse genetics to test the hypothesis that genetic modifiers alter the phenotypic manifestations of Taz inactivation. We crossed TazKO/X females in the C57BL6/J inbred strain to males from 8 inbred strains and evaluated the phenotypes of first generation (F1) TazKO/Y progeny, compared to TazWT/Y littermates. We observed that genetic background strongly impacted phenotypic expression. C57BL6/J and CAST/EiJ[F1] TazKO/Y mice developed severe cardiomyopathy, whereas A/J[F1] TazKO/Y mice had normal heart function. C57BL6/J and WSB/EiJ[F1] TazKO/Y mice had severely reduced treadmill endurance, whereas endurance was normal in A/J[F1] and CAST/EiJ[F1] TazKO/Y mice. In all genetic backgrounds, cardiolipin showed similar abnormalities in knockout mice, and transcriptomic and metabolomic investigations identified signatures of mitochondrial uncoupling and activation of the integrated stress response. TazKO/Y cardiac mitochondria were small, clustered, and had reduced cristae density in knockouts in severely affected genetic backgrounds but were relatively preserved in the permissive A/J[F1] strain. Gene expression and mitophagy measurements were consistent with reduced mitophagy in knockout mice in genetic backgrounds intolerant of Taz mutation. Our data demonstrate that genetic modifiers powerfully modulate phenotypic expression of Taz loss-of-function and act downstream of cardiolipin, possibly by altering mitochondrial quality control.

DOI: https://doi.org/10.1093/hmg/ddad041
Gene. 2023 Mar 11. pii: S0378-1119(23)00199-3. [Epub ahead of print] 147358

New advancements in CRISPR based gene therapy of Duchenne Muscular dystrophy.

Atieh Eslahi, Farzaneh Alizadeh, Amir Avan, Gordon A Ferns, Meysam Moghbeli, Mohammad Reza Abbaszadegan, Majid Mojarrad.

  Duchenne muscular dystrophy (DMD) is caused by the dystrophin gene mutations and is one of the most common and lethal human hereditary disorders. A novel therapeutic approach using CRISPR technology has gained attention in the treatment of DMD. Gene replacement strategies are being proposed as a promising therapeutic option to compensate the loss of function mutations. Although, the large size of the dystrophin gene and the limitations of the existing gene replacement approach, could mean the gene delivery of shortened versions of dystrophin such as midystrophin and microdystrophins. There are also other approaches: including Targeted removal of dystrophin exons to restore the reading-frame; Dual sgRNA-directed DMD exon deletion, CRISPR-SKIP strategy; reframing of dystrophin using Prime Editing technology; exon removal using twin prime technology; TransCRISTI technology to targeted exon integration into dystrophin gene. Here we provide an overview of recent progresses in dystrophin gene editing using updated versions of CRISPR to introduce novel opportunities in DMD gene therapy. Overall, the novel CRISPR based technologies are improving and expanding to allow the application of more precise gene editing for the treatment of DMD.

Keywords:  CRISPR technology; CRISPR-SKIP strategy; Duchenne muscular dystrophy (DMD); Prime Editing; TransCRISTI; gene therapy

DOI:  https://doi.org/10.1016/j.gene.2023.147358
Front Netw Physiol. 2022 ;2 877676

Using a Network Physiology Approach to Prescribe Exercise for Exercise Oncology.

Gwendolyn A Thomas.

  Current American College of Sports Medicine (ACSM) exercise guidelines for exercise oncology survivors are generic one-size fits all recommendations, which assume ideal or prototypic health and fitness state in order to prescribe. Individualization is based on the objective evaluation of the patient's baseline physiological status based on a linear dose response relationship of endpoints. This is only a partial snapshot of both the acute and chronic responses exercise can provide. Each acute exercise session represents a unique challenge to whole-body homeostasis and complex acute and adaptive responses occur at the cellular and systemic levels. Additionally, external factors must be considered when prescribing exercise. Network physiology views the human organism in terms of physiological and organ systems, each with structural organization and functional complexity. This organizational approach leads to complex, transient, fluctuating and nonlinear output dynamics which should be utilized in exercise prescription across health states. Targeting health outcomes requires a multi-system approach as change doesn't happen in only one system at a time or in one direction Utilizing a multi-system or person-centered approach, allows for targeting and personalization and understands and targets non-linear dynamics of change. Therefore, the aims of this review are to propose a paradigm shift towards a Network Physiology approach for exercise prescription for cancer survivors. Cancer treatment affects multiple systems that interact to create symptoms and disruptions across these and therefore, prescribing exercise utilizing both external daily factors and internal physiological networks is of the highest order.

Keywords:  exercise oncology; exercise prescription; network physiology; non-linear; person centered

DOI:  https://doi.org/10.3389/fnetp.2022.877676
Sci Rep. 2023 Mar 12. 13(1): 4101

Virus-free transfection, transient expression, and purification of human cardiac myosin in mammalian muscle cells for biochemical and biophysical assays.

Lok Priya Velayuthan, Luisa Moretto, Sven Tågerud, Marko Ušaj, Alf Månsson.

Myosin expression and purification is important for mechanistic insights into normal function and mutation induced changes. The latter is particularly important for striated muscle myosin II where mutations cause several debilitating diseases. However, the heavy chain of this myosin is challenging to express and the standard protocol, using C2C12 cells, relies on viral infection. This is time and work intensive and associated with infrastructural demands and biological hazards, limiting widespread use and hampering fast generation of a wide range of mutations. We here develop a virus-free method to overcome these challenges. We use this system to transfect C2C12 cells with the motor domain of the human cardiac myosin heavy chain. After optimizing cell transfection, cultivation and harvesting conditions, we functionally characterized the expressed protein, co-purified with murine essential and regulatory light chains. The gliding velocity (1.5-1.7 µm/s; 25 °C) in the in vitro motility assay as well as maximum actin activated catalytic activity (kcat; 8-9 s-1) and actin concentration for half maximal activity (KATPase; 70-80 µM) were similar to those found previously using virus based infection. The results should allow new types of studies, e.g., screening of a wide range of mutations to be selected for further characterization.

DOI: https://doi.org/10.1038/s41598-023-30576-1
J Physiol. 2023 Mar 16.

Non-invasive estimation of muscle fibre size from high-density electromyography.

Andrea Casolo, Sumiaki Maeo, Thomas G Balshaw, Marcel B Lanza, Neil R W Martin, Stefano Nuccio, Tatiana Moro, Antonio Paoli, Francesco Felici, Nicola Maffulli, Bjoern Eskofier, Thomas M Kinfe, Jonathan P Folland, Dario Farina, Alessandro Del Vecchio.

  Because of the biophysical relation between muscle fibre diameter and the propagation velocity of action potentials along the muscle fibres, motor unit conduction velocity could be a non-invasive index of muscle fibre size in humans. However, the relation between motor unit conduction velocity and fibre size has been only assessed indirectly in animal models and in human patients with invasive intramuscular EMG recordings, or it has been mathematically derived from computer simulations. By combining advanced non-invasive techniques to record motor unit activity in vivo, i.e., high-density surface EMG, with the gold standard technique for muscle tissue sampling, i.e., muscle biopsy, here we investigated the relation between the conduction velocity of populations of motor units identified from the biceps brachii muscle, and muscle fibre diameter. We demonstrate the possibility to predict muscle fibre diameter (R2 = 0.66) and cross-sectional area (R2 = 0.65) from conduction velocity estimates with low systematic bias (∼2% and ∼4% respectively) and a relatively low margin of individual error (∼8% and ∼16%, respectively). The proposed neuromuscular interface opens new perspectives in the use of high-density EMG as a non-invasive tool to estimate muscle fibre size without the need of surgical biopsy sampling. The non-invasive nature of high-density surface EMG for the assessment of muscle fibre size may be useful in studies monitoring child development, aging, space and exercise physiology, although the applicability and validity of the proposed methodology needs to be more directly assessed in these specific populations by future studies. KEY POINTS: Because of the biophysical relation between muscle fibre size and the propagation velocity of action potentials along the sarcolemma, motor unit conduction velocity could represent a potential non-invasive candidate to estimate muscle fibre size in vivo. This relation has been previously assessed in animal models and humans with invasive techniques, or it has been mathematically-derived from simulations. By combining high-density surface EMG with muscle biopsy, here we explored the relation between the conduction velocity of populations of motor units and muscle fibre size in healthy individuals. Our results confirmed that motor unit conduction velocity can be considered as a novel biomarker of fibre size, which can be adopted to predict muscle fibre diameter and cross-sectional area with low systematic bias and margin of individual error. The proposed neuromuscular interface opens new perspectives in the use of high-density EMG as a non-invasive tool to estimate muscle fibre size without the need of surgical biopsy sampling Abstract figure legend In this study, we investigated the relation between the conduction velocity of populations of motor units identified from biceps brachii muscle and muscle fibre size. We adopted high-density surface EMG to decode the activity of voluntarily activated motor units and estimated their conduction velocity. Similarly, we adopted muscle biopsy to measure muscle fibre size. We revealed the possibility to accurately transform motor unit conduction velocity values into estimated measures of muscle fibre size, which in turn showed a good degree of association with the muscle fibre size measured directly by muscle biopsy. Furthermore, we demonstrated that the proposed neuromuscular interface allows to predict the mean measured fibre diameter and cross-sectional area from an EMG-derived parameter with a relatively low bias and error, thus opening new perspectives in the use of high-density EMG as a non-invasive tool to estimate muscle fibre size without the need of surgical biopsy sampling This article is protected by copyright. All rights reserved.

Keywords:  Motor unit; conduction velocity; high-density surface electromyography; muscle fibre size

DOI:  https://doi.org/10.1113/JP284170