The number of myonuclei within a muscle fiber is an important factor in muscle growth, but its regulation during muscle adaptation is not well understood. We aimed to elucidate the time course of ...myonuclear dynamics during endurance training, loaded and concentric resistance training, and nerve silencing-induced disuse atrophy with subsequent recovery. We modified tibialis anterior muscle activity in free-living rats with electrical stimulation from implantable pulse generators, or with implantable osmotic pumps delivering tetrodotoxin (TTX) to silence the motor nerve without transection. We used the updated, automated software MyoVision to measure fiber-type-specific responses in whole tibialis anterior cross sections (∼8,000 fibers each). Seven days of continuous low-frequency stimulation (CLFS) reduced muscle mass (-12%), increased slower myosin isoforms and reduced IIX/IIB fibers (-32%), and substantially increased myonuclei especially in IIX/IIB fibers (55.5%). High-load resistance training (spillover) produced greater hypertrophy (∼16%) in muscle mass and fiber cross-sectional area (CSA) than low-load resistance training (concentric, ∼6%) and was associated with myonuclear addition in all fiber types (35%-46%). TTX-induced nerve silencing resulted in progressive loss in muscle mass, fiber CSA, and myonuclei per fiber cross section (-50.7%, -53.7%, and -40.7%, respectively, at 14 days). Myonuclear loss occurred in a fiber-type-independent manner, but subsequent recovery during voluntary habitual activity suggested that type IIX/IIB fibers contained more new myonuclei during recovery from severe atrophy. This study demonstrates the power and accuracy provided by the updated MyoVision software and introduces new models for studying myonuclear dynamics in training, detraining, retraining, repeated disuse, and recovery.
We introduce new models for studying fiber-type-specific myonuclear dynamics in muscle training, detraining, retraining, disuse, and recovery. We show that the various fiber types do not respond identically and that myonuclear number changes during adaptation. We also critically assess an updated version of MyoVision automated image analysis software to quantify whole muscle immunofluorescent microscopical images in a faster and less computer intensive manner. MyoVision remains open source and freely available with more user-controlled features.
Circadian rhythms that influence mammalian homeostasis and overall health have received increasing interest over the past two decades. The molecular clock, which is present in almost every cell, ...drives circadian rhythms while being a cornerstone of physiological outcomes. The skeletal muscle clock has emerged as a primary contributor to metabolic health, as the coordinated expression of the core clock factors BMAL1 and CLOCK with the muscle-specific transcription factor MYOD1 facilitates the circadian and metabolic programme that supports skeletal muscle physiology. The phase of the skeletal muscle clock is sensitive to the time of exercise, which provides a rationale for exploring the interactions between the skeletal muscle clock, exercise and metabolic health. Here, we review the underlying mechanisms of the skeletal muscle clock that drive muscle physiology, with a particular focus on metabolic health. Additionally, we highlight the interaction between exercise and the skeletal muscle clock as a means of reinforcing metabolic health and discuss the possible implications of the time of exercise as a chronotherapeutic approach.
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GEOZS, IJS, IMTLJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBMB, UL, UM, UPUK, ZAGLJ
Mice are often used in gain or loss of function studies to understand how genes regulate metabolism and adaptation to exercise in skeletal muscle. Once‐daily resistance training with electrical nerve ...stimulation produces hypertrophy of the dorsiflexors in rat, but not in mouse. Using implantable pulse generators, we assessed the acute transcriptional response (1‐h post‐exercise) after 2, 10, and 20 days of training in free‐living mice and rats using identical nerve stimulation paradigms. RNA sequencing revealed strong concordance in the timecourse of many transcriptional responses in the tibialis anterior muscles of both species including responses related to “stress responses/immediate‐early genes, and “collagen homeostasis,” “ribosomal subunits,” “autophagy,” and “focal adhesion.” However, pathways associated with energy metabolism including “carbon metabolism,” “oxidative phosphorylation,” “mitochondrial translation,” “propanoate metabolism,” and “valine, leucine, and isoleucine degradation” were oppositely regulated between species. These pathways were suppressed in the rat but upregulated in the mouse. Our transcriptional analysis suggests that although many pathways associated with growth show remarkable similarities between species, the absence of an actual growth response in the mouse may be because the mouse prioritizes energy metabolism, specifically the replenishment of fuel stores and intermediate metabolites.
Timecourse transcriptional assessments of identical nerve‐stimulated resistance exercise in mice and rats reveal concordant and discordant gene regulation and pathways related to muscle growth.
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
We present the time course of change in the muscle transcriptome 1 h after the last exercise bout of a daily resistance training program lasting 2, 10, 20, or 30 days. Daily exercise in rat tibialis ...anterior muscles (5 sets of 10 repetitions over 20 min) induced progressive muscle growth that approached a new stable state after 30 days. The acute transcriptional response changed along with progressive adaptation of the muscle phenotype. For example, expression of type 2B myosin was silenced. Time courses recently synthesized from human exercise studies do not demonstrate so clearly the interplay between the acute exercise response and the longer‐term consequences of repeated exercise. We highlight classes of transcripts and transcription factors whose expression increases during the growth phase and declines again as the muscle adapts to a new daily pattern of activity and reduces its rate of growth. Myc appears to play a central role.
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
Myonuclei transcriptionally regulate muscle fibers during homeostasis and adaptation to exercise. Their subcellular location and quantity are important when characterizing phenotypes of myopathies, ...the effect of treatments, and understanding the roles of satellite cells in muscle adaptation and muscle "memory." Difficulties arise in identifying myonuclei due to their proximity to the sarcolemma and closely residing interstitial cell neighbors. We aimed to determine to what extent (pericentriolar material-1) PCM1 is a specific marker of myonuclei in vitro and in vivo. Single isolated myofibers and cross sections from mice and humans were studied from several models including wild-type and Lamin A/C mutant mice after functional overload and damage and recovery in humans following forced eccentric contractions. Fibers were immunolabeled for PCM1, Pax7, and DNA. C2C12 myoblasts were also studied to investigate changes in PCM1 localization during myogenesis. PCM1 was detected at not only the nuclear envelope of myonuclei in mature myofibers and in newly formed myotubes but also centrosomes in proliferating myogenic precursors, which may or may not fuse to join the myofiber syncytium. PCM1 was also detected in nonmyogenic nuclei near the sarcolemma, especially in regenerating areas of the
mouse and damaged human muscle. Although PCM1 is not completely specific to myonuclei, the impact that PCM1+ macrophages and interstitial cells have on myonuclei counts would be small in healthy muscle. PCM1 may prove useful as a marker of satellite cell dynamics due to the distinct change in localization during differentiation, revealing satellite cells in their quiescent (PCM1-), proliferating (PCM1+ centrosome), and prefusion states (PCM1+ nuclear envelope).
In this investigation, we addressed the contribution of the core circadian clock factor, BMAL1, in skeletal muscle to both acute transcriptional responses to exercise and transcriptional remodeling ...in response to exercise training. Additionally, we adopted a systems biology approach to investigate how loss of skeletal muscle BMAL1 altered peripheral tissue homeostasis as well as exercise training adaptations in iWAT, liver, heart, and lung of male mice.
Combining inducible skeletal muscle specific BMAL1 knockout mice, physiological testing and standardized exercise protocols, we performed a multi-omic analysis (transcriptomics, chromatin accessibility and metabolomics) to explore loss of muscle BMAL1 on muscle and peripheral tissue responses to exercise.
Muscle-specific BMAL1 knockout mice demonstrated a blunted transcriptional response to acute exercise, characterized by the lack of upregulation of well-established exercise responsive transcription factors including Nr4a3 and Ppargc1a. Six weeks of exercise training in muscle-specific BMAL1 knockout mice induced significantly greater and divergent transcriptomic and metabolomic changes in muscle. Surprisingly, liver, lung, inguinal white adipose and heart showed divergent exercise training transcriptomes with less than 5% of ‘exercise-training’ responsive genes shared for each tissue between genotypes.
Our investigation has uncovered the critical role that BMAL1 plays in skeletal muscle as a key regulator of gene expression programs for both acute exercise and training adaptations. In addition, our work has uncovered the significant impact that altered exercise response in muscle and its likely impact on the system plays in the peripheral tissue adaptations to exercise training. Our work also demonstrates that if the muscle adaptations diverge to a more maladaptive state this is linked to increased gene expression signatures of inflammation across many tissues. Understanding the molecular targets and pathways contributing to health vs. maladaptive exercise adaptations will be critical for the next stage of therapeutic design for exercise mimetics.
•Skeletal muscle BMAL1 is required for the acute exercise induced up-regulation of immediate-early genes including Nr4a3 and Ppargc1a.•Exercise training without muscle BMAL1 results in much larger and more divergent transcriptomic and metabolomic adaptations in muscle.•Skeletal muscle BMAL1 is necessary to support metabolic homeostasis of pyruvate and key TCA cycle intermediates following exercise training.•Muscle training responses impact the system; trained mice without muscle BMAL1 exhibit signatures of inflammation across liver, adipose and lung.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Understanding the role of mechanical loading and exercise in skeletal muscle (SkM) is paramount for delineating the molecular mechanisms that govern changes in muscle mass. However, it is unknown ...whether loading of bioengineered SkM in vitro adequately recapitulates the molecular responses observed after resistance exercise (RE) in vivo. To address this, the transcriptional and epigenetic (DNA methylation) responses were compared after mechanical loading in bioengineered SkM in vitro and after RE in vivo. Specifically, genes known to be upregulated/hypomethylated after RE in humans were analyzed. Ninety‐three percent of these genes demonstrated similar changes in gene expression post‐loading in the bioengineered muscle when compared to acute RE in humans. Furthermore, similar differences in gene expression were observed between loaded bioengineered SkM and after programmed RT in rat SkM tissue. Hypomethylation occurred for only one of the genes analysed (GRIK2) post‐loading in bioengineered SkM. To further validate these findings, DNA methylation and mRNA expression of known hypomethylated and upregulated genes post‐acute RE in humans were also analyzed at 0.5, 3, and 24 h post‐loading in bioengineered muscle. The largest changes in gene expression occurred at 3 h, whereby 82% and 91% of genes responded similarly when compared to human and rodent SkM respectively. DNA methylation of only a small proportion of genes analyzed (TRAF1, MSN, and CTTN) significantly increased post‐loading in bioengineered SkM alone. Overall, mechanical loading of bioengineered SkM in vitro recapitulates the gene expression profile of human and rodent SkM after RE in vivo. Although some genes demonstrated differential DNA methylation post‐loading in bioengineered SkM, such changes across the majority of genes analyzed did not closely mimic the epigenetic response to acute‐RE in humans.
Mechanical loading of mouse bioengineered skeletal muscle in vitro recapitulates the gene expression profile of skeletal muscle after resistance exercise in vivo. However, while some DNA methylation changes were detected following mechanical loading in bioengineered muscle, this did not as closely mimic the DNA methylation response to acute resistance exercise in vivo.
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
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