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Skeletal muscle development is a highly ordered process orchestrated transcriptionally by the myogenic regulatory factors. However, the downstream molecular mechanisms of myogenic regulatory factor functions in myogenesis are not fully understood. Here, we identified the RNA-binding protein Musashi2 (Msi2) as a myogenin target gene and a post-transcriptional regulator of myoblast differentiation. Msi2 knockdown in murine myoblasts blocked differentiation without affecting the expression of MyoD or myogenin. Msi2 overexpression was also sufficient to promote myoblast differentiation and myocyte fusion. Msi2 loss attenuated autophagosome formation via down-regulation of the autophagic protein MAPL1LC3/ATG8 (LC3) at the early phase of myoblast differentiation. Moreover, forced activation of autophagy effectively suppressed the differentiation defects incurred by Msi2 loss. Consistent with its functions in myoblasts in vitro, mice deficient for Msi2 exhibited smaller limb skeletal muscles, poorer exercise performance, and muscle fiber-type switching in vivo. Collectively, our study demonstrates that Msi2 is a novel regulator of mammalian myogenesis and establishes a new functional link between muscular development and autophagy regulation., (© 2024 Wang et al.)

Following traumatic musculoskeletal injuries, prolonged bedrest and loss of physical activity may limit muscle plasticity and drive metabolic dysfunction. One specific injury, volumetric muscle loss (VML), results in frank loss of muscle and is characterized by whole-body and cellular metabolic dysfunction. However, how VML and restricted physical activity limit plasticity of the whole-body, cellular, and metabolomic environment of the remaining uninjured muscle remains unclear. Adult mice were randomized to posterior hindlimb compartment VML or were age-matched injury naïve controls, then randomized to standard or restricted activity cages for 8-wks. Activity restriction in naïve mice resulted in ~5% greater respiratory exchange ratio (RER); combined with VML, carbohydrate oxidation was ~23% greater than VML alone, but lipid oxidation was largely unchanged. Activity restriction combined with VML increased whole-body carbohydrate usage. Together there was a greater pACC:ACC ratio in the muscle remaining, which may contribute to decreased fatty acid synthesis. Further, β-HAD activity normalized to mitochondrial content was decreased following VML, suggesting a diminished capacity to oxidize fatty acids. The muscle metabolome was not altered by the restriction of physical activity. The combination of VML and activity restriction resulted in similar (~91%) up- and down-regulated metabolites and/or ratios, suggesting that VML injury alone is regulating changes in the metabolome. Data supports possible VML-induced alterations in fatty acid metabolism are exacerbated by activity restriction. Collectively, this work adds to the sequala of VML injury, exhausting the ability of the muscle remaining to oxidize fatty acids resulting in a possible accumulation of triglycerides., Competing Interests: Competing interests The authors declare that they have no competing interests.

This study's objective was to investigate how contractile strength loss associated with a volumetric muscle loss (VML) injury affects the adjacent tibial bone structural and functional properties in male C57BL/6J mice. Mice were randomized into one of two experimental groups: VML-injured mice that were injured at age 12 weeks and aged to 20 weeks (8 weeks postinjury, VML) and 20-week-old age-matched uninjured mice (Uninjured-20). Tibial bone strength, mid-diaphysis cortical geometry, intrinsic material properties, and metaphyseal trabecular bone structure were assessed by three-point bending and microcomputed tomography (µCT). The plantar flexor muscle group (gastrocnemius, soleus, plantaris) was analyzed for its functional capacities, that is, peak-isometric torque and peak-isokinetic power. VML-injured limbs had 25% less peak-isometric torque and 31% less peak-isokinetic power compared to those of Uninjured-20 mice (p < 0.001). Ultimate load, but not stiffness, was significantly less (10%) in tibias of VML-injured limbs compared to those from Uninjured-20 (p = 0.014). µCT analyses showed cortical bone thickness was 6% less in tibias of VML-injured limbs compared to Uninjured-20 (p = 0.001). Importantly, tibial bone cross-section moment of inertia, the primary determinant of bone ultimate load, was 16% smaller in bones of VML-injured limbs compared to bones from Uninjured-20 (p = 0.046). Metaphyseal trabecular bone structure was also altered up to 23% in tibias of VML-injured limbs (p < 0.010). These changes in tibial bone structure and function after a VML injury occur during a natural maturation phase between the age of 12 and 20 weeks, as evidenced by Uninjured-20 mice having greater tibial bone size and strength compared to uninjured-aged 12-week mice., (© 2023 The Authors. Journal of Orthopaedic Research® published by Wiley Periodicals LLC on behalf of Orthopaedic Research Society.)

Background: This study was designed to develop an understanding of the pathophysiology of traumatic muscle injury in the context of Western diet (WD; high fat and high sugar) and obesity. The objective was to interrogate the combination of WD and injury on skeletal muscle mass and contractile and metabolic function., Methods: Male and female C57BL/6J mice were randomized into four groups based on a two-factor study design: (1) injury (uninjured vs. volumetric muscle loss [VML]) and (2) diet (WD vs. normal chow [NC]). Electrophysiology was used to test muscle strength and metabolic function in cohorts of uninjured + NC, uninjured + WD, VML + NC and VML + WD at 8 weeks of intervention., Results: VML-injured male and female mice both exhibited decrements in muscle mass (-17%, P < 0.001) and muscle strength (-28%, P < 0.001); however, VML + WD females had a 28% greater muscle mass compared to VML + NC females (P = 0.034), a compensatory response not detected in males. VML-injured male and female mice both had lower carbohydrate- and fat-supported muscle mitochondrial respiration (JO 2 ) and less electron conductance through the electron transport system (ETS); however, male VML-WD had 48% lower carbohydrate-supported JO 2 (P = 0.014) and 47% less carbohydrate-supported electron conductance (P = 0.026) compared to male VML + NC, and this diet-injury phenotype was not present in females. ETS electron conductance starts with complex I and complex II dehydrogenase enzymes at the inner mitochondrial membrane, and male VML + WD had 31% less complex I activity (P = 0.004) and 43% less complex II activity (P = 0.005) compared to male VML + NC. This was a diet-injury phenotype not present in females. Pyruvate dehydrogenase (PDH), β-hydroxyacyl-CoA dehydrogenase, citrate synthase, α-ketoglutarate dehydrogenase and malate dehydrogenase metabolic enzyme activities were evaluated as potential drivers of impaired JO 2 in the context of diet and injury. There were notable male and female differential effects in the enzyme activity and post-translational regulation of PDH. PDH enzyme activity was 24% less in VML-injured males, independent of diet (P < 0.001), but PDH enzyme activity was not influenced by injury in females. PDH enzyme activity is inhibited by phosphorylation at serine-293 by PDH kinase 4 (PDK4). In males, there was greater total PDH, phospho-PDH ser293 and phospho-PDH-to-total PDH ratio in WD mice compared to NC, independent of injury (P ≤ 0.041). In females, PDK4 was 51% greater in WD compared to NC, independent of injury (P = 0.025), and was complemented by greater phospho-PDH ser293 (P = 0.001)., Conclusions: Males are more susceptible to muscle metabolic dysfunction in the context of combined WD and traumatic injury compared to females, and this may be due to impaired metabolic enzyme functions., (© 2023 The Authors. Journal of Cachexia, Sarcopenia and Muscle published by Wiley Periodicals LLC.)

Hypophosphatasia (HPP) is a rare metabolic bone disorder characterized by low levels of tissue non-specific alkaline phosphatase (TNAP) that causes under-mineralization of the bone, leading to bone deformity and fractures. In addition, patients often present with chronic muscle pain, reduced muscle strength, and an altered gait. In this work, we explored dynamic muscle function in a homozygous TNAP knockout mouse model of severe juvenile onset HPP. We found a reduction in skeletal muscle size and impairment in a range of isolated muscle contractile properties. Using histological methods, we found that the structure of HPP muscles was similar to healthy muscles in fiber size, actin and myosin structures, as well as the α-tubulin and mitochondria networks. However, HPP mice had significantly fewer embryonic and type I fibers than wild type mice, and fewer metabolically active NADH+ muscle fibers. We then used oxygen respirometry to evaluate mitochondrial function and found that complex I and complex II leak respiration were reduced in HPP mice, but that there was no disruption in efficiency of electron transport in complex I or complex II. In summary, the severe HPP mouse model recapitulates the muscle strength impairment phenotypes observed in human patients. Further exploration of the role of alkaline phosphatase in skeletal muscle could provide insight into mechanisms of muscle weakness in HPP.

The primary objective of this study was to determine if low- or high-resistance voluntary wheel running leads to functional improvements in muscle strength (i.e., isometric and isokinetic torque) and metabolic function (i.e., permeabilized fibre bundle mitochondrial respiration) after a volumetric muscle loss (VML) injury. C57BL/6J mice were randomized into one of four experimental groups at age 12 weeks: uninjured control, VML untreated (VML), low-resistance wheel running (VML-LR) and high-resistance wheel running (VML-HR). All mice, excluding the uninjured, were subject to a unilateral VML injury to the plantar flexor muscles and wheel running began 3 days post-VML. At 8 weeks post-VML, peak isometric torque was greater in uninjured compared to all VML-injured groups, but both VML-LR and VML-HR had greater (∼32%) peak isometric torque compared to VML. All VML-injured groups had less isokinetic torque compared to uninjured, and there was no statistical difference among VML, VML-LR and VML-HR. No differences in cumulative running distance were observed between VML-LR and VML-HR groups. Because adaptations in VML-HR peak isometric torque were attributed to greater gastrocnemius muscle mass, atrophy- and hypertrophy-related protein content and post-translational modifications were explored via immunoblot; however, results were inconclusive. Permeabilized fibre bundle mitochondrial oxygen consumption was 22% greater in uninjured compared to VML, but there was no statistical difference among VML, VML-LR and VML-HR. Furthermore, neither wheel running group demonstrated a change in the relative protein content of the mitochondrial biogenesis transcription factor, peroxisome proliferator-activated receptor γ coactivator 1-α (PGC-1α). These results indicate that resistance wheel running alone only has modest benefits in the VML-injured muscle. NEW FINDINGS: What is the central question of the study? Does initiation of a resistance wheel running regimen following volumetric muscle loss (VML) improve the functional capacity of skeletal muscle? What is the main finding and its importance? Resistance wheel running led to greater muscle mass and strength in mice with a VML injury but did not result in a full recovery. Neither low- nor high-resistance wheel running was associated with a change in permeabilized muscle fibre respiration despite runners having greater whole-body treadmill endurance capacity, suggesting resilience to metabolic adaptations in VML-injured muscle. Resistance wheel running may be a suitable adjuvant rehabilitation strategy, but alone does not fully mitigate VML pathology., (© 2023 The Authors. Experimental Physiology published by John Wiley & Sons Ltd on behalf of The Physiological Society.)

Volumetric muscle loss (VML) is associated with persistent functional impairment due to a lack of de novo muscle regeneration. As mechanisms driving the lack of regeneration continue to be established, adjunctive pharmaceuticals to address the pathophysiology of the remaining muscle may offer partial remediation. Studies were designed to evaluate the tolerance and efficacy of two FDA-approved pharmaceutical modalities to address the pathophysiology of the remaining muscle tissue after VML injury: (1) nintedanib (an anti-fibrotic) and (2) combined formoterol and leucine (myogenic promoters). Tolerance was first established by testing low- and high-dosage effects on uninjured skeletal muscle mass and myofiber cross-sectional area in adult male C57BL/6J mice. Next, tolerated doses of the two pharmaceutical modalities were tested in VML-injured adult male C57BL/6J mice after an 8-week treatment period for their ability to modulate muscle strength and whole-body metabolism. The most salient findings indicate that formoterol plus leucine mitigated the loss in muscle mass, myofiber number, whole-body lipid oxidation, and muscle strength, and resulted in a higher whole-body metabolic rate (p ≤ 0.016); nintedanib did not exacerbate or correct aspects of the muscle pathophysiology after VML. This supports ongoing optimization efforts, including scale-up evaluations of formoterol treatment in large animal models of VML., (© 2023 The Authors. Physiological Reports published by Wiley Periodicals LLC on behalf of The Physiological Society and the American Physiological Society.)

Volumetric muscle loss (VML) injuries are characterized by non-recoverable loss of tissue resulting in contractile and metabolic dysfunction. The characterization of metabolic dysfunction in volumetric muscle loss-injured muscle has been interpreted from permeabilized myofiber respiration experiments involving saturating ADP levels and non-physiologic ATP:ADP concentration ratios. The extent to which this testing condition obscures the analysis of mitochondrial (dys) function after volumetric muscle loss injury is unclear. An alternative approach is described that leverages the enzymatic reaction of creatine kinase and phosphocreatine to assess mitochondrial respiration and membrane potential at clamped physiologic ATP:ADP ratios, "CK Clamp." The objective of this study was to validate the CK Clamp in volumetric muscle loss-injured muscle and to detect differences that may exist between volumetric muscle loss-injured and uninjured muscles at 1, 3, 5, 7, 10, and 14 days post-injury. Volumetric muscle loss-injured muscle maintains bioenergetic features of the CK Clamp approach, i.e., mitochondrial respiration rate (JO 2 ) titters down and mitochondrial membrane potential is more polarized with increasing ATP:ADP ratios. Pyruvate/malate/succinate-supported JO 2 was significantly less in volumetric muscle loss-injured muscle at all timepoints compared to uninjured controls (-26% to -84%, p < 0.001) and electron conductance was less at day 1 (-60%), 5 (-52%), 7 (-35%), 10 (-59%), and 14 (-41%) ( p < 0.001). Palmitoyl-carnitine/malate-supported JO 2 and electron conductance were less affected following volumetric muscle loss injury. volumetric muscle loss-injury also corresponded with a more polarized mitochondrial membrane potential across the clamped ATP:ADP ratios at day 1 and 10 (pyruvate and palmitoyl-carnitine, respectively) (+5%, p < 0.001). This study supports previous characterizations of metabolic dysfunction and validates the CK Clamp as a tool to investigate bioenergetics in traumatically-injured muscle., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 McFaline-Figueroa, Hunda, Heo, Winders, Greising and Call.)

Introduction: Musculoskeletal injury (MSKI) risk is increased following mild traumatic brain injury (mTBI). Increased MSKI risk is present up to 2 years following post-mTBI return-to-duty/activity relative to both non-mTBI peers and to their pre-mTBI selves across a range of populations, including military service members, and professional, college and high school athletes. Despite the well documented increased post-mTBI MSKI risk, the underlying neuromuscular mechanisms contributing to this increased risk have yet to be definitively determined. A number of potential mechanisms have been suggested (eg, aberrant kinematics, dynamic balance impairments, lower voluntary muscle activation), but none have been confirmed with a comprehensive, prospective study. This study aims to: (1) elucidate the neuromuscular control mechanisms following mTBI that contribute to increased MSKI risk, and (2) prospectively track patient outcomes (up to 12 months; MSKI occurrences and patient-reported outcomes (PRO))., Methods and Analysis: This is a multicentre prospective, case-matched control observational study to identify deficiencies in neuromuscular function following mTBI that may contribute to increased MSKI risk. Participants (aim to recruit 148, complete data collection on 124) will be classified into two cohorts; mTBI and control. All participants will undergo longitudinal (initial, 6 weeks post-initial, 12 weeks post-initial) comprehensive three-dimensional biomechanical (jump-landing; single leg hop; cut; gait), neuromuscular (interpolated twitch technique, muscular ramp contraction) and sensory (joint repositioning; light touch sensation) assessments to elucidate the underlying neuromuscular control mechanisms post-mTBI that may contribute to increased MSKI. Occurrences of MSKI and PROs (National Institutes of Health Patient-Reported Outcome Measurement Information System: Physical Function, Pain Interference, Depression, Anxiety; Brief Resilience Scale; Tampa Scale of Kinesiophobia), will be tracked monthly (up to 1 year) via electronic data capture platforms., Ethics and Dissemination: The study received approval from the Walter Reed National Military Medical Center Institutional Review Board. Results will be made available to the associated funding agency and other researchers via conference proceedings and journal articles., Trial Registration Number: NCT05122728., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2023. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Objective: Balance impairments may suggest somatosensory disruption beyond concussion clinical recovery, but somatosensory subsystems have never been directly assessed. Our objective was to examine somatosensory function between individuals with a concussion and healthy matched-controls at acute (<7 days) and asymptomatic (<72 hours of being symptom-free) time points., Setting: Laboratory., Participants: Participants with a concussion and matched controls ( n = 24; 58% male, age: 19.3 ± 1.1 years, mass: 70.3 ± 16.4 kg, height: 177.3 ± 12.7 cm)., Design: Prospective cohort., Main Measures: Somatosensory assessments on the dominant limb at both time points included: (1) plantar touch sensation threshold via Semmes-Weinstein monofilaments, (2) plantar pressure pain threshold via algometry, and (3) knee absolute passive joint repositioning (PJR) error via Biodex across 3 arcs (105°-75°, 30°-60°, 90°-45° knee-flexion). We used mixed-model analyses of variance, post hoc Tukey honestly significant difference t tests with mean difference, 95% CI, and Hedges' g effect sizes to examine outcomes., Results: Touch sensation had a group effect with the concussion cohort needing 0.95 grams of force (gf) more relative to controls (95% CI: 0.03 to 1.87; P = .043). No touch sensation interaction was present, but medium and large effects were observed for greater gf needed among the concussed cohort at the acute (1.11 gf; 95% CI: 0.17 to 2.05; g = 0.96) and asymptomatic time points (0.79 gf; 95% CI: -0.15 to 1.73; g = 0.73). No plantar pressure pain threshold effects were observed ( P ≥ .311), with negligible pressure difference magnitudes at the acute (0.26 pound force [lbf]/cm 2 ; 95% CI: -1.54 to 2.06; g = 0.13) and medium magnitudes at the asymptomatic time points (0.99 lbf/cm 2 ; 95% CI: -0.81 to 2.80; g = 0.42) for the concussed cohort needing more pressure to detect pain. The 30° to 60° PJR had a time effect, with asymptomatic time point having 3.12° better accuracy (95% CI: 1.23° to 5.02; P = .002). The concussed cohort had small-to-medium magnitude differences relative to controls at the acute time point for PJR during 105° to 75° (0.89°; g = 0.30) and 90° to 45° (0.62°; g = 0.17), but not 30° to 60° (-1.75°; g = -0.40)., Conclusions: Individuals with a concussion exhibited large effects for diminished plantar touch sensation and small to medium effects for inhibited plantar pressure pain sensation compared with controls, which may indicate altered somatosensory function. Negligible PJR differences suggest knee joint position sense is not altered post-concussion. Pre- and postconcussion examination is warranted to understand causal somatosensory mechanisms., Competing Interests: The authors declare no conflicts of interest., (Copyright © 2022 Wolters Kluwer Health, Inc. All rights reserved.)

Context: Aberrant movement patterns among individuals with concussion history have been reported during sport-related movement. However, the acute postconcussion kinematic and kinetic biomechanical movement patterns during a rapid acceleration-deceleration task have not been profiled and leaves their progressive trajectory unknown. Our study aimed to examine single-leg hop stabilization kinematics and kinetics between concussed and healthy-matched controls acutely (≤7 d) and when asymptomatic (≤72 h of symptom resolution)., Design: Prospective, cohort laboratory study., Methods: Ten concussed (60% male; 19.2 [0.9] y; 178.7 [14.0] cm; 71.3 [18.0] kg) and 10 matched controls (60% male; 19.5 [1.2] y; 176.1 [12.6] cm; 71.0 [17.0] kg) completed the single-leg hop stabilization task under single and dual task (subtracting by 6's or 7's) at both time points. Participants stood on a 30-cm tall box set 50% of their height behind force plates while in an athletic stance. A synchronized light was illuminated randomly, queuing participants to initiate the movement as rapidly as possible. Participants then jumped forward, landed on their nondominant leg, and were instructed to reach and maintain stabilization as fast as possible upon ground contact. We used 2 (group) × 2 (time) mixed-model analyses of variance to compare single-leg hop stabilization outcomes separately during single and dual task., Results: We observed a significant main group effect for single-task ankle plantarflexion moment, with greater normalized torque (mean difference = 0.03 N·m/body weight; P = .048, g = 1.18) for concussed individuals across time points. A significant interaction effect for single-task reaction time indicated that concussed individuals had slower performance acutely relative to asymptomatic (mean difference = 0.09 s; P = .015, g = 0.64), while control group performance was stable. No other main or interaction effects for single-leg hop stabilization task metrics were present during single and dual task (P ≥ .051)., Conclusions: Greater ankle plantarflexion torque coupled with slower reaction time may indicate stiff, conservative single-leg hop stabilization performance acutely following concussion. Our findings shed preliminary light on the recovery trajectories of biomechanical alterations following concussion and provide specific kinematic and kinetic focal points for future research.

Introduction: Mitochondria are extremely important organelles in the regulation of bone marrow and brain activity. However, live imaging of these subcellular features with high resolution in scattering tissues like brain or bone has proven challenging., Methods: In this study, we developed a two-photon fluorescence microscope with adaptive optics (TPFM-AO) for high-resolution imaging, which uses a home-built Shack-Hartmann wavefront sensor (SHWFS) to correct system aberrations and a sensorless approach for correcting low order tissue aberrations., Results: Using AO increases the fluorescence intensity of the point spread function (PSF) and achieves fast imaging of subcellular organelles with 400 nm resolution through 85 μm of highly scattering tissue. We achieved ~1.55×, ~3.58×, and ~1.77× intensity increases using AO, and a reduction of the PSF width by ~0.83×, ~0.74×, and ~0.9× at the depths of 0, 50 μm and 85 μm in living mouse bone marrow respectively, allowing us to characterize mitochondrial health and the survival of functioning cells with a field of view of 67.5× 67.5 μm. We also investigate the role of initial signal and background levels in sample correction quality by varying the laser power and camera exposure time and develop an intensity-based criteria for sample correction., Discussion: This study demonstrates a promising tool for imaging of mitochondria and other organelles in optically distorting biological environments, which could facilitate the study of a variety of diseases connected to mitochondrial morphology and activity in a range of biological tissues., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Zheng, Liversage, Tehrani, Call, Kner and Mortensen.)

Volumetric muscle loss (VML) is the traumatic loss of skeletal muscle, resulting in chronic functional deficits and pathological comorbidities, including altered whole-body metabolic rate and respiratory exchange ratio (RER), despite no change in physical activity in animal models. In other injury models, treatment with β 2 receptor agonists (e.g. formoterol) improves metabolic and skeletal muscle function. We aimed first to examine if restricting physical activity following injury affects metabolic and skeletal muscle function, and second, to enhance the metabolic and contractile function of the muscle remaining following VML injury through treatment with formoterol. Adult male C57Bl/6J mice (n = 32) underwent VML injury to the posterior hindlimb compartment and were randomly assigned to unrestricted or restricted activity and formoterol treatment or no treatment; age-matched injury naïve mice (n = 4) were controls for biochemical analyses. Longitudinal 24 h evaluations of physical activity and whole-body metabolism were conducted following VML. In vivo muscle function was assessed terminally, and muscles were biochemically evaluated for protein expression, mitochondrial enzyme activity and untargeted metabolomics. Restricting activity chronically after VML had the greatest effect on physical activity and RER, reflected in reduced lipid oxidation, although changes were attenuated by formoterol treatment. Formoterol enhanced injured muscle mass, while mitigating functional deficits. These novel findings indicate physical activity restriction may recapitulate following VML clinically, and adjunctive oxidative treatment may create a metabolically beneficial intramuscular environment while enhancing the injured muscle's mass and force-producing capacity. Further investigation is needed to evaluate adjunctive oxidative treatment with rehabilitation, which may augment the muscle's regenerative and functional capacity following VML. KEY POINTS: The natural ability of skeletal muscle to regenerate and recover function is lost following complex traumatic musculoskeletal injury, such as volumetric muscle loss (VML), and physical inactivity following VML may incur additional deleterious consequences for muscle and metabolic health. Modelling VML injury-induced physical activity restriction altered whole-body metabolism, primarily by decreasing lipid oxidation, while preserving local skeletal muscle metabolic activity. The β 2 adrenergic receptor agonist formoterol has shown promise in other severe injury models to improve regeneration, recover function and enhance metabolism. Treatment with formoterol enhanced mass of the injured muscle and whole-body metabolism while mitigating functional deficits resulting from injury. Understanding of chronic effects of the clinically available and FDA-approved pharmaceutical formoterol could be a translational option to support muscle function after VML injury., (© 2022 The Authors. The Journal of Physiology published by John Wiley & Sons Ltd on behalf of The Physiological Society.)

Exercise benefits many organ systems, including having a panacea-like effect on the brain. For example, aerobic exercise improves cognition and attention and reduces the risk of brain-related diseases, such as dementia, stress, and depression. Recent advances suggest that endocrine signaling from peripheral systems, such as skeletal muscle, mediates the effects of exercise on the brain. Consequently, it has been proposed that factors secreted by all organs in response to physical exercise should be more broadly termed the "exerkines." Accumulating findings suggest that exerkines derived from skeletal muscle, liver, and adipose tissues directly impact brain mitochondrial function. Mitochondria play a pivotal role in regulating neuronal energy metabolism, neurotransmission, cell repair, and maintenance in the brain, and therefore exerkines may act via impacting brain mitochondria to improve brain function and disease resistance. Therefore, herein we review studies investigating the impact of muscle-, liver-, and adipose tissue-derived exerkines on brain cognitive and metabolic function via modulating mitochondrial bioenergetics, content, and dynamics under healthy and/or disease conditions.

New Findings: What is the central question of this study? First, how does physical rehabilitation influence recovery from traumatic muscle injury? Second, how does physical activity impact the rehabilitation response for skeletal muscle function and whole-body metabolism? What is the main finding and its importance? The most salient findings were that rehabilitation impaired muscle function and range of motion, while restricting activity mitigated some negative effects but also impacted whole-body metabolism. These data suggest that first, work must continue to explore treatment parameters, including modality, time, type, duration and intensity, to find the best rehabilitation approaches for volumetric muscle loss injuries; and second, restricting activity acutely might enhance rehabilitation response, but whole-body co-morbidities should continue to be considered., Abstract: Volumetric muscle loss (VML) injury occurs when a substantial volume of muscle is lost by surgical removal or trauma, resulting in an irrecoverable deficit in muscle function. Recently, it was suggested that VML impacts whole-body and muscle-specific metabolism, which might contribute to the inability of the muscle to respond to treatments such as physical rehabilitation. The aim of this work was to understand the complex relationship between physical activity and the response to rehabilitation after VML in an animal model, evaluating the rehabilitation response by measurement of muscle function and whole-body metabolism. Adult male mice (n = 24) underwent a multi-muscle, full-thickness VML injury to the gastrocnemius, soleus and plantaris muscles and were randomized into one of three groups: (1) untreated; (2) rehabilitation (i.e., combined electrical stimulation and range of motion, twice per week, beginning 72 h post-injury, for ∼8 weeks); or (3) rehabilitation and restriction of physical activity. There was a lack of positive adaption associated with electrical stimulation and range of motion intervention alone; however, maximal isometric torque of the posterior muscle group was greater in mice receiving treatment with activity restriction (P = 0.008). Physical activity and whole-body metabolism were measured ∼6 weeks post-injury; metabolic rate decreased (P = 0.001) and respiratory exchange ratio increased (P = 0.022) with activity restriction. Therefore, restricting physical activity might enhance an intervention delivered to the injured muscle group but impair whole-body metabolism. It is possible that restricting activity is important initially post-injury to protect the muscle from excess demand. A gradual increase in activity throughout the course of treatment might optimize muscle function and whole-body metabolism., (© 2022 The Authors. Experimental Physiology published by John Wiley & Sons Ltd on behalf of The Physiological Society.)

Volumetric muscle loss (VML) injuries represent a majority of military service member casualties and are common in civilian populations following blunt and/or penetrating traumas. Characterized as a skeletal muscle injury with permanent functional impairments, there is currently no standard for rehabilitation, leading to lifelong disability. Toward developing rehabilitative strategies, previous research demonstrates that the remaining muscle after a VML injury lacks similar levels of plasticity or adaptability as healthy, uninjured skeletal muscle. This may be due, in part, to impaired innervation and vascularization of the remaining muscle, as well as disrupted molecular signaling cascades commonly associated with muscle adaptation. The primary objective of this study was to assess the ability of four pharmacological agents with a strong record of modulating muscle contractile and metabolic function to improve functional deficits in a murine model of VML injury. Male C57BL/6 mice underwent a 15% multimuscle VML injury of the posterior hindlimb and were randomized into drug treatment groups (formoterol [FOR], 5-aminoimidazole-4-carboxamide riboside [AICAR], pioglitazone [PIO], or sildenafil [SIL]) or untreated VML group. At the end of 60 days, the injury model was first validated by comparison to age-matched injury-naive mice. Untreated VML mice had 22% less gastrocnemius muscle mass, 36% less peak-isometric torque, and 27% less maximal mitochondrial oxygen consumption rate compared to uninjured mice ( p  < 0.01). Experimental drug groups were, then, compared to VML untreated, and there was minimal evidence of efficacy for AICAR, PIO, or SIL in improving contractile and metabolic functional outcomes. However, FOR-treated VML mice had 18% greater peak isometric torque ( p  < 0.01) and permeabilized muscle fibers had 36% greater State III mitochondrial oxygen consumption rate ( p  < 0.01) compared to VML untreated mice, suggesting an overall improvement in muscle condition. There was minimal evidence that these benefits came from greater mitochondrial biogenesis and/or mitochondrial complex protein content, but could be due to greater enzyme activity levels for complex I and complex II. These findings suggest that FOR treatment is candidate to pair with a rehabilitative approach to maximize functional improvements in VML-injured muscle. Impact statement Volumetric muscle loss (VML) injuries result in deficiencies in strength and mobility, which have a severe impact on patient quality of life. Despite breakthroughs in tissue engineering, there are currently no treatments available that can restore function to the affected limb. Our data show that treatment of VML injuries with clinically available and FDA-approved formoterol (FOR), a beta-agonist, significantly improves strength and metabolism of VML-injured muscle. FOR is therefore a promising candidate for combined therapeutic approaches (i.e., regenerative rehabilitation) such as pairing FOR with structured rehabilitation or cell-seeded biomaterials as it may provide greater functional improvements than either strategy alone.

Context: Neuromuscular function is altered acutely following concussion and theoretically linked to the subsequent postconcussion musculoskeletal injury risk. Existing research has only examined voluntary muscle activation, limiting mechanistic understanding. Therefore, our study aimed to examine voluntary and involuntary muscle activation between college-aged, concussed individuals when symptom-free and healthy matched controls., Design: Prospective, cross-sectional cohort laboratory study., Methods: Concussed and healthy participants (n = 24; 58% male, age: 19.3 [1.1] y, mass: 70.3 [16.4] kg, height: 177.3 [12.7] cm) completed the superimposed burst (SB) neuromuscular assessment on their dominant limb within 72 hours after self-reporting asymptomatic (22.4 [20.2] d postinjury). Unnormalized and bodyweight-normalized quadriceps maximal voluntary isometric contraction torque (in newton meters), unnormalized and bodyweight-normalized electrically stimulated SB torque, pain (numeric 1-10) during SB, and the central activation ratio (in percentage) were assessed via the SB. Parametric and nonparametric analyses, 95% confidence intervals (95% CIs), and Hedges g (parametric) and Spearman ρ (nonparametric) effect sizes were used to examine group differences (α = .05)., Results: The maximal voluntary isometric contraction torque (concussed: 635.60 N·m [300.93] vs control: 556.27 N·m [182.46]; 95% CI, -131.36 to 290.02; P = .443; d = 0.33), SB torque (concussed: 203.22 N·m [97.17], control: 262.85 N·m [159.07]; 95% CI, -171.22 to 51.97; P = .280; d = -0.47), and central activation ratio (concussed: 72.16% [17.16], control: 70.09% [12.63]; 95% CI, -10.68 to 14.83; P = .740; d = 0.14) did not differ between the concussed and control groups regardless of bodyweight normalization (P ≥ .344). Pain during the SB was significantly higher with a medium effect for participants with a concussion versus healthy controls (concussed: median = 7, control: median = 5; P = .046; ρ = -0.42)., Discussion: These findings suggest concussed participants do not have statistically altered voluntary or involuntary quadricep neuromuscular function once asymptomatic compared with controls. Therefore, the elevated postconcussion musculoskeletal injury risk may not be attributed to lower-extremity muscle activation. Concussed participants displayed greater pain perception during the SB, which suggests somatosensory or perception changes requiring further examination.

A Western diet (WD), high in sugars and saturated fats, impairs learning and memory function and contributes to weight gain. Mitochondria in the brain provide energy for neurocognitive function and may play a role in body weight regulation. We sought to determine whether a WD alters behavior and metabolic outcomes in male and female rodents through impacting hippocampal and hypothalamic mitochondrial bioenergetics. Results revealed a sexually dimorphic macronutrient preference, where males on the WD consumed a greater percentage of calories from fat/protein and females consumed a greater percentage of calories from a sugar-sweetened beverage. Both males and females on a WD gained body fat and showed impaired glucose tolerance when compared to same-sex controls. Males on a WD demonstrated impaired hippocampal functioning and an elevated tendency toward a high membrane potential in hippocampal mitochondria. Comprehensive bioenergetics analysis of WD effects in the hypothalamus revealed a tissue-specific adaption, where males on the WD oxidized more fat, and females oxidized more fat and carbohydrates at peak energy demand compared to same-sex controls. These results suggest that adult male rats show a susceptibility toward hippocampal dysfunction on a WD, and that hypothalamic mitochondrial bioenergetics are altered by WD in a sex-specific manner.

Reliable assessment of skeletal muscle strength is arguably the most important outcome measure in neuromuscular and musculoskeletal disease and injury studies, particularly when evaluating regenerative therapies' efficacy. Additionally, a critical aspect of translating many regenerative therapies is the demonstration of scalability and effectiveness in a large animal model. Various physiological preparations have been established to evaluate intrinsic muscle function properties in basic science studies, primarily in small animal models. The practices may be categorized as: in vitro (isolated fibers, fiber bundles, or whole muscle), in situ (muscle with intact vascularization and innervation but distal tendon attached to a force transducer), and in vivo (structures of the muscle or muscle unit remain intact). There are strengths and weaknesses to each of these preparations; however, a clear advantage of in vivo strength testing is the ability to perform repeated measurements in the same animal. Herein, the materials and methods to reliably assess isometric torque produced by the hindlimb dorsiflexor muscles in vivo in response to standard peroneal electrical stimulation in anesthetized pigs are presented.

Background: Acute and chronic alcohol use can cause skeletal muscle myopathy in concert with impairments in skeletal muscle strength, function and fatigue resistance. However, the fundamental contractile deficits induced in the presence of alcohol versus those observed in the recovery period following the clearance of alcohol have not yet been characterized nor is it known whether sex influences these outcomes., Methods: Male and female mice received an intraperitoneal injection of either saline (Control) or ethanol (EtOH; 5g/kg body weight). Muscle force, fatigue, fatigue recovery and twitch characteristics of the posterior crural muscle complex were measured in situ 1 hour and 24 hours post alcohol., Results: In the presence of alcohol (1-hour post treatment) absolute and normalized force generated at 80-150 Hertz was decreased in male and female mice with concurrent reductions in the rate of force development and increases in ½ relaxation time. When expressed as a percentage of maximum force, both males and females also displayed an alcohol-induced leftward shift in the force frequency curve indicative of a type I contractile phenotype. Alcohol enhanced fatigue in both males and females but had no effect on force recovery. Following clearance of alcohol (24-hour post treatment), contractile function was completely restored in females while alcohol treated males experienced sustained reductions in absolute force and had enhanced fatigue compared with male controls., Conclusions: In the presence of alcohol, both males and females exhibited significant declines in muscle force production and enhanced fatigue; however, following complete clearance of the alcohol, females recovered all functional parameters, while males did not., Competing Interests: The authors have declared that no competing interests exist.

Skeletal muscle mitochondria are highly adaptable, highly dynamic organelles that maintain the functional integrity of the muscle fiber by providing ATP for contraction and cellular homeostasis (e.g., Na + /K + ATPase). Emerging as early modulators of inflammation, mitochondria sense and respond to cellular stress. Mitochondria communicate with the environment, in part, by release of physical signals called mitochondrial-derived damage-associated molecular patterns (mito-DAMPs) and deviation from routine function (e.g., reduced ATP production, Ca 2+ overload). When skeletal muscle is compromised, mitochondria contribute to an acute inflammatory response necessary for myofibril regeneration; however, exhaustive signaling associated with altered or reduced mitochondrial function can be detrimental to muscle outcomes. Here, we describe changes in mitochondrial content, structure, and function following skeletal muscle injury and disuse and highlight the influence of mitochondria-cytokine crosstalk on muscle regeneration and recovery. Although the appropriate therapeutic modulation following muscle stressors remains unknown, retrospective gene expression analysis reveals that interleukin-6 (IL-6), interleukin-1β (IL-1β), chemokine C-X-C motif ligand 1 (CXCL1), and monocyte chemoattractant protein 1 (MCP-1) are significantly upregulated following three unique muscle injuries. These cytokines modulate mitochondrial function and execute bona fide pleiotropic roles that can aid functional recovery of muscle, however, when aberrant, chronically disrupt healing partly by exacerbating mitochondrial dysfunction. Multidisciplinary efforts to delineate the opposing regulatory roles of inflammatory cytokines in the muscle mitochondrial environment are required to modulate regenerative behavior following skeletal muscle injury or disuse. Future therapeutic directions to consider include quenching or limited release of mito-DAMPs and cytokines present in cytosol or circulation.

The remarkable regeneration capability of skeletal muscle depends on the coordinated proliferation and differentiation of satellite cells (SCs). The self-renewal of SCs is critical for long-term maintenance of muscle regeneration potential. Hypoxia profoundly affects the proliferation, differentiation, and self-renewal of cultured myoblasts. However, the physiological relevance of hypoxia and hypoxia signaling in SCs in vivo remains largely unknown. Here, we demonstrate that SCs are in an intrinsic hypoxic state in vivo and express hypoxia-inducible factor 2A (HIF2A). HIF2A promotes the stemness and long-term homeostatic maintenance of SCs by maintaining their quiescence, increasing their self-renewal, and blocking their myogenic differentiation. HIF2A stabilization in SCs cultured under normoxia augments their engraftment potential in regenerative muscle. Conversely, HIF2A ablation leads to the depletion of SCs and their consequent regenerative failure in the long-term. In contrast, transient pharmacological inhibition of HIF2A accelerates muscle regeneration by increasing SC proliferation and differentiation. Mechanistically, HIF2A induces the quiescence and self-renewal of SCs by binding the promoter of the Spry1 gene and activating Spry1 expression. These findings suggest that HIF2A is a pivotal mediator of hypoxia signaling in SCs and may be therapeutically targeted to improve muscle regeneration.

Multiple organ dysfunction syndrome (MODS) is a detrimental clinical complication in critically ill patients with high mortality. Emerging evidence suggests that oxidative stress and endothelial activation (induced expression of adhesion molecules) of vital organ vasculatures are key, early steps in the pathogenesis. We aimed to ascertain the role and mechanism(s) of enhanced extracellular superoxide dismutase (EcSOD) expression in skeletal muscle in protection against MODS induced by endotoxemia. We showed that EcSOD overexpressed in skeletal muscle-specific transgenic mice (TG) redistributes to other peripheral organs through the circulation and enriches at the endothelium of the vasculatures. TG mice are resistant to endotoxemia (induced by lipopolysaccharide [LPS] injection) in developing MODS with significantly reduced mortality and organ damages compared with the wild type littermates (WT). Heterogenic parabiosis between TG and WT mice conferred a significant protection to WT mice, whereas mice with R213G knock-in mutation, a human single nucleotide polymorphism leading to reduced binding EcSOD in peripheral organs, exacerbated the organ damages. Mechanistically, EcSOD inhibits vascular cell adhesion molecule 1 expression and inflammatory leukocyte adhesion to the vascular wall of vital organs, blocking an early step of the pathology in organ damage under endotoxemia. Therefore, enhanced expression of EcSOD in skeletal muscle profoundly protects against MODS by inhibiting endothelial activation and inflammatory cell adhesion, which could be a promising therapy for MODS., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Autophagy is a conserved cellular process for degrading aggregate proteins and dysfunctional organelle. It is still debatable if autophagy and mitophagy (a specific process of autophagy of mitochondria) play important roles in myogenic differentiation and functional regeneration of skeletal muscle. We tested the hypothesis that autophagy is critical for functional regeneration of skeletal muscle. We first observed time-dependent increases (3- to 6-fold) of autophagy-related proteins (Atgs), including Ulk1, Beclin1, and LC3, along with reduced p62 expression during C2C12 differentiation, suggesting increased autophagy capacity and flux during myogenic differentiation. We then used cardiotoxin (Ctx) or ischemia-reperfusion (I/R) to induce muscle injury and regeneration and observed increases in Atgs between days 2 and 7 in adult skeletal muscle followed by increased autophagy flux after day 7 Since Ulk1 has been shown to be essential for mitophagy, we asked if Ulk1 is critical for functional regeneration in skeletal muscle. We subjected skeletal muscle-specific Ulk1 knockout mice (MKO) to Ctx or I/R. MKO mice had significantly impaired recovery of muscle strength and mitochondrial protein content post-Ctx or I/R. Imaging analysis showed that MKO mice have significantly attenuated recovery of mitochondrial network at 7 and 14 days post-Ctx. These findings suggest that increased autophagy protein and flux occur during muscle regeneration and Ulk1-mediated mitophagy is critical for recovery for the mitochondrial network and hence functional regeneration., (Copyright © 2017 the American Physiological Society.)

Background: An immediate loss of strength follows virtually all types of muscle injury but there is debate whether the initial strength loss is maximal or if a secondary loss of strength occurs during the first 3 days post-injury., Objective: The objective of this analysis was to conduct a systematic review and meta-analysis of the research literature to determine if a secondary loss of strength occurs after an injurious initiating event., Methods: Literature searches were performed using eight electronic databases (e.g., PubMed, Cochrane Library). Search terms included skeletal muscle AND (injur* OR damage*) AND (strength OR force OR torque). The extracted strength data were converted to a standard format by calculating the standardized mean difference, which is reported as the effect size (ES) along with its 95 % confidence interval (CI). The calculation of ES was designed so that a negative ES that was statistically less than zero would be interpreted as indicating a secondary loss of strength., Results: A total of 223 studies with over 4000 human and animal subjects yielded data on 262 independent groups and a total of 936 separate ESs. Our overall meta-analysis yielded a small-to-medium, positive overall ES that was statistically greater than zero (overall ES = +0.34, 95 % CI 0.27-0.40; P < 0.00000001). Considerable variation in ES was observed among studies (I 2  = 86 %), which could be partially explained by the research group conducting the study, sex of the subject, day of post-injury strength assessment, whether fatigue was present immediately post-injury, and the muscle group injured. From the subgroup meta-analyses probing these variables, 36 subgroup ESs were calculated and none were statistically less than zero., Conclusion: Overall, our findings do not support the presence of a secondary loss of strength following an acute muscle injury, and strongly suggest that strength, on average, recovers steadily over the first 3 days post-injury., Competing Interests: Compliance with Ethical Standards Funding No sources of funding were used to assist in the preparation of this article. Conflicts of interest Gordon Warren, Jarrod Call, Amy Farthing, and Bemene Baadom-Piaro declare that they have no conflicts of interest relevant to the content of this review.

In order to investigate the molecular and cellular mechanisms of muscle regeneration an experimental injury model is required. Advantages of eccentric contraction-induced injury are that it is a controllable, reproducible, and physiologically relevant model to cause muscle injury, with injury being defined as a loss of force generating capacity. While eccentric contractions can be incorporated into conscious animal study designs such as downhill treadmill running, electrophysiological approaches to elicit eccentric contractions and examine muscle contractility, for example before and after the injurious eccentric contractions, allows researchers to circumvent common issues in determining muscle function in a conscious animal (e.g., unwillingness to participate). Herein, we describe in vitro and in vivo methods that are reliable, repeatable, and truly maximal because the muscle contractions are evoked in a controlled, quantifiable manner independent of subject motivation. Both methods can be used to initiate eccentric contraction-induced injury and are suitable for monitoring functional muscle regeneration hours to days to weeks post-injury.

Background & Aims: Duchenne muscular dystrophy is a debilitating genetic disorder characterized by severe muscle wasting and early death in afflicted boys. The primary cause of this disease is mutations in the dystrophin gene resulting in massive muscle degeneration and inflammation. The purpose of this study was to determine if dystrophic muscle pathology and inflammation were decreased by pre-natal and early dietary intervention with green tea extract., Methods: Mdx breeder mice and pups were fed diets containing 0.25% or 0.5% green tea extract and compared to untreated mdx and C57BL/6J mice. Serum creatine kinase was assessed as a systemic indicator of muscle damage. Quantitative histopathological and immunohistochemical techniques were used to determine muscle pathology, macrophage infiltration, and NF-kappaB localization., Results: Early treatment of mdx mice with green tea extract significantly decreased serum creatine kinase by approximately 85% at age 42 days (P< or =0.05). In these mice, the area of normal fiber morphology was increased by as much as approximately 32% (P< or =0.05). The primary histopathological change was a approximately 21% decrease in the area of regenerating fibers (P< or =0.05). NF-kappaB staining in regenerating muscle fibers was also significantly decreased in green tea extract-treated mdx mice when compared to untreated mdx mice (P< or =0.05)., Conclusion: Early treatment with green tea extract decreases dystrophic muscle pathology potentially by regulating NF-kappaB activity in regenerating muscle fibers., (Copyright 2009 Elsevier Ltd and European Society for Clinical Nutrition and Metabolism. All rights reserved.)