Adult Muscle Research Articles

Exercise and ageing impact the kynurenine/tryptophan pathway and acylcarnitine metabolite pools in skeletal muscle of older adults.

Exercise-induced perturbation of skeletal muscle metabolites is a probable mediator of long-term health benefits in older adults. Although specific metabolites have been identified to be impacted by age, physical activity and exercise, the depth of coverage of the muscle metabolome is still limited. Here, we investigated resting and exercise-induced metabolite distribution in muscle from well-phenotyped older adults who were active or sedentary, and a group of active young adults. Percutaneous biopsies of the vastus lateralis were obtained before, immediately after and 3h following a bout of endurance cycling. Metabolite profile in muscle biopsies was determined by tandem mass spectrometry. Mitochondrial energetics in permeabilized fibre bundles was assessed by high resolution respirometry and fibre type proportion was assessed by immunohistology. We found that metabolites of the kynurenine/tryptophan pathway were impacted by age and activity. Specifically, kynurenine was elevated in muscle from older adults, whereas downstream metabolites of kynurenine (kynurenic acid and NAD+ ) were elevated in muscle from active adults and associated with cardiorespiratory fitness and muscle oxidative capacity. Acylcarnitines, a potential marker of impaired metabolic health, were elevated in muscle from physically active participants. Surprisingly, despite baseline group difference, acute exercise-induced alterations in whole-body substrate utilization, as well as muscle acylcarnitines and ketone bodies, were remarkably similar between groups. Our data identified novel muscle metabolite signatures that associate with the healthy ageing phenotype provoked by physical activity and reveal that the metabolic responsiveness of muscle to acute endurance exercise is retained [NB]:AUTHOR: Please ensure that the appropriate material has been provide for Table S2, as well as for Figures S1 to S7, as also cited in the text with age regardless of activity levels. KEY POINTS: Kynurenine/tryptophan pathway metabolites were impacted by age and physical activity in human muscle, with kynurenine elevated in older muscle, whereas downstream products kynurenic acid and NAD+ were elevated in exercise-trained muscle regardless of age. Acylcarnitines, a marker of impaired metabolic health when heightened in circulation, were elevated in exercise-trained muscle of young and older adults, suggesting that muscle act as a metabolic sink to reduce the circulating acylcarnitines observed with unhealthy ageing. Despite the phenotypic differences, the exercise-induced response of various muscle metabolite pools, including acylcarnitine and ketone bodies, was similar amongst the groups, suggesting that older adults can achieve the metabolic benefits of exercise seen in young counterparts.

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

ObjectiveSkeletal 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. MethodsYoung 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. ResultsSingle 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. ConclusionsmiR-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.

Goat MyoD1: mRNA expression, InDel and CNV detection and their associations with growth traits

The Myogenic differentiation 1 (MyoD1) gene is a crucial regulator of muscle formation and differentiation. However, there are few studies on the mRNA expression pattern of the goat MyoD1 gene and its effect on goat growth and development. To address this, we investigated the mRNA expression of the MyoD1 gene in several tissues of fetal and adult goats, containing heart, liver, spleen, lung, kidney and skeletal muscle. The results focused on the expression of the MyoD1 gene in skeletal muscle of fetal goats was much higher than adult goats, suggesting its important role in skeletal muscle formation and development. Following, a total of 619 Shaanbei White Cashmere goats (SBWCs) were used to monitor the InDel (Insertion/Deletion) and CNV (Copy Number Variation) variations of the MyoD1 gene. Three InDel loci were identified, and there was no significant correlation with goat growth traits. Furthermore, a CNV locus containing the MyoD1 gene exon with three types (Loss type, Normal type, Gain type) were identified. The association analysis results showed that the CNV locus was significantly associated with body weight, height at hip cross, heart girth and hip width in SBWCs (P < 0.05). Meanwhile, the Gain type of CNV exhibited the best growth traits and good consistency among three types in goats, suggesting its potential as a DNA marker for marker-assisted breeding of goats. Overall, our study provided a scientific basis for breeding goats with better growth and development traits.

Effects of a Myofascial Technique on the Stiffness and Thickness of the Thoracolumbar Fascia and Lumbar Erector Spinae Muscles in Adults with Chronic Low Back Pain: A Randomized before-and-after Experimental Study.

The thoracolumbar fascia (TLF) may be a pain generator, given its rich innervation. Structural and biomechanical changes have also been documented in adults with chronic non-specific low back pain (LBP). Myofascial techniques (MFTs) are commonly used in manual therapy and are hypothesized to reduce tissue stiffness and pain. However, evidence for these effects is limited. The objective of this study was to evaluate the immediate effects of a standardized MFT compared to a simulated MFT on: (1) the stiffness of the TLF and erector spinae muscles (shear-wave sonoelastography), (2) the thickness of the TLF (B-mode ultrasound), and (3) pain intensity (numerical rating scale). Forty-nine participants with chronic non-specific LBP were included in a randomized before-and-after experimental study. Outcome measures were collected before (T0) and immediately after the intervention (T1). Pain intensity was also assessed on day two (T2) and seven (T7). The MFT group showed a significant decrease in left erector spinae muscle stiffness and left TLF thickness compared to the simulated group. In addition, there was a significant reduction in pain intensity in the MFT group compared to the simulated group at T1 and T2. The results of this study suggest that MFT results in immediate tissue changes and transient pain reduction in patients with LBP.

Return of the Tbx5; lineage-tracing reveals ventricular cardiomyocyte-like precursors in the injured adult mammalian heart

The single curative measure for heart failure patients is a heart transplantation, which is limited due to a shortage of donors, the need for immunosuppression and economic costs. Therefore, there is an urgent unmet need for identifying cell populations capable of cardiac regeneration that we will be able to trace and monitor. Injury to the adult mammalian cardiac muscle, often leads to a heart attack through the irreversible loss of a large number of cardiomyocytes, due to an idle regenerative capability. Recent reports in zebrafish indicate that Tbx5a is a vital transcription factor for cardiomyocyte regeneration. Preclinical data underscore the cardioprotective role of Tbx5 upon heart failure. Data from our earlier murine developmental studies have identified a prominent unipotent Tbx5-expressing embryonic cardiac precursor cell population able to form cardiomyocytes, in vivo, in vitro and ex vivo. Using a developmental approach to an adult heart injury model and by employing a lineage-tracing mouse model as well as the use of single-cell RNA-seq technology, we identify a Tbx5-expressing ventricular cardiomyocyte-like precursor population, in the injured adult mammalian heart. The transcriptional profile of that precursor cell population is closer to that of neonatal than embryonic cardiomyocyte precursors. Tbx5, a cardinal cardiac development transcription factor, lies in the center of a ventricular adult precursor cell population, which seems to be affected by neurohormonal spatiotemporal cues. The identification of a Tbx5-specific cardiomyocyte precursor-like cell population, which is capable of dedifferentiating and potentially deploying a cardiomyocyte regenerative program, provides a clear target cell population for translationally-relevant heart interventional studies.

ChIP-seq profiling of H3K4me3 and H3K27me3 in an invasive insect, Bactrocera dorsalis.

Introduction: While it has been suggested that histone modifications can facilitate animal responses to rapidly changing environments, few studies have profiled whole-genome histone modification patterns in invasive species, leaving the regulatory landscape of histone modifications in invasive species unclear. Methods: Here, we screen genome-wide patterns of two important histone modifications, trimethylated Histone H3 Lysine 4 (H3K4me3) and trimethylated Histone H3 Lysine 27 (H3K27me3), in adult thorax muscles of a notorious invasive pest, the Oriental fruit fly Bactrocera dorsalis (Hendel) (Diptera: Tephritidae), using Chromatin Immunoprecipitation with high-throughput sequencing (ChIP-seq). Results: We identified promoters featured by the occupancy of H3K4me3, H3K27me3 or bivalent histone modifications that were respectively annotated with unique genes key to muscle development and structure maintenance. In addition, we found H3K27me3 occupied the entire body of genes, where the average enrichment was almost constant. Transcriptomic analysis indicated that H3K4me3 is associated with active gene transcription, and H3K27me3 is mostly associated with transcriptional repression. Importantly, we identified genes and putative motifs modified by distinct histone modification patterns that may possibly regulate flight activity. Discussion: These findings provide the first evidence of histone modification signature in B. dorsalis, and will be useful for future studies of epigenetic signature in other invasive insect species.

Intermuscular coherence of plantar and dorsiflexor muscles in older adults with Parkinson's disease and age-matched controls during bipedal and unipedal stance.

Postural instability increases with age and is exacerbated in neurological disorders such as Parkinson's disease (PD). Reducing the base of support from bipedal to unipedal stance increases center of pressure (CoP) parameters and intermuscular coherence in lower-leg muscles of healthy older adults. To further develop an understanding of postural control in an altered state of neurological impairment, we explored intermuscular coherence in lower-leg muscles and CoP displacement in older adults with PD. This study measured surface EMG from the medial (MG) and lateral (LG) gastrocnemii, soleus (SOL), and tibialis anterior (TA), and examined EMG amplitude and intermuscular coherence during bipedal and unipedal stance on a force plate with firm (no foam) and compliant (standing on foam) surface conditions in nine older adults with PD (70±5 years, 6 females) and 8 age-matched non-Parkinsonian older adults (5 females). Intermuscular coherence was analyzed between agonist-agonist and agonist-antagonist muscle pairs in the alpha (8-13 Hz) and beta (15-35 Hz) frequency bands. CoP parameters increased from bipedal to unipedal stance in both groups (p < 0.01), but did not increase from the firm to compliant surface condition (p > 0.05). During unipedal stance, CoP path length was shorter in older adults with PD (2027.9 ± 1074.1 mm) compared to controls (3128.5 ± 1198.7 mm) (p < 0.01). Alpha and beta agonist-agonist and agonist-antagonist coherence increased by 28% from bipedal to unipedal stance (p > 0.05), but did not differ between older adults with PD (0.09 ± 0.07) and controls (0.08 ± 0.05) (p > 0.05). The older adults with PD also had greater normalized EMG amplitude of the LG (63.5 ± 31.7%) and TA (60.6 ± 38.4%) during the balance tasks (p > 0.05) than the non-Parkinsonian counterparts. Older adults with PD had shorter path lengths during unipedal stance and required greater muscle activation than older adults without PD to perform the tasks, but intermuscular coherence did not differ between the groups. This may be attributable to their early disease stage and high motor function.

Effect of exercise modality and weight loss on changes in muscle and bone quality in older adults with obesity

BackgroundLittle is known about the effect of exercise modality during a dietary weight loss program on muscle size and quality, as measured by computed tomography (CT). Even less is known about how CT-derived changes in muscle track with changes in volumetric bone mineral density (vBMD) and bone strength. MethodsOlder adults (66 ± 5 years, 64 % women) were randomized to 18-months of diet-induced weight loss (WL), WL with aerobic training (WL + AT), or WL with resistance training (WL + RT). CT-derived muscle area, radio-attenuation and intermuscular fat percentage at the trunk and mid-thigh were determined at baseline (n = 55) and 18-month follow-up (n = 22–34), and changes were adjusted for sex, baseline value, and weight lost. Lumbar spine and hip vBMD and finite element-derived bone strength were also measured. ResultsAfter adjustment for the weight lost, muscle area losses at the trunk were −7.82 cm2 [−12.30, −3.35] for WL, −7.72 cm2 [−11.36, −4.07] for WL + AT, and −5.14 cm2 [−8.65, −1.63] for WL + RT (p < 0.001 for group differences). At the mid-thigh, decreases were −6.20 cm2 [−10.39, −2.02] for WL, −7.84 cm2 [−11.19, −4.48] for WL + AT, and −0.60 cm2 [−4.14, 2.94] for WL + RT; this difference between WL + AT and WL + RT was significant in post-hoc testing (p = 0.01). Change in trunk muscle radio-attenuation was positively associated with change in lumbar bone strength (r = 0.41, p = 0.04). ConclusionsWL + RT better preserved muscle area and improved muscle quality more consistently than WL + AT or WL alone. More research is needed to characterize the associations between muscle and bone quality in older adults undertaking weight loss interventions.

Blood meal digestion and changes in lipid reserves are associated with the post-ecdysis development of the flight muscle and ovary in young adults of Rhodnius prolixus

Rhodnius prolixus is a hemimetabolous, hematophagous insect, and both nymphs and adults feed exclusively on blood. The blood feeding triggers the molting process and, after five nymphal instar stages, the insect reaches the winged adult form. After the final ecdysis, the young adult still has a lot of blood in the midgut and, thus, we have investigated the changes in protein and lipid contents that are observed in the insect organs as digestion continues after molting. Total midgut protein content decreased during the days after the ecdysis, and digestion was finished fifteen days later. Simultaneously, proteins and triacylglycerols present in the fat body were mobilized, and their contents decreased, whereas they increased in both the ovary and the flight muscle. In order to evaluate the activity of de novo lipogenesis of each organ, the fat body, ovary and flight muscle were incubated in the presence of radiolabeled acetate, and the fat body showed the highest efficiency rate (around 47%) to convert the taken up acetate into lipids. The levels of de novo lipid synthesis in the flight muscle and ovary were very low. When 3H-palmitate was injected into the young females, its incorporation by the flight muscle was higher than by the ovary or the fat body. In the flight muscle, the 3H-palmitate was similarly distributed amongst triacylglycerols, phospholipids, diacylglycerols and free fatty acids, while in the ovary and fat body it was mostly found in triacylglycerols and phospholipids. The flight muscle was not fully developed after the molt, and at day two no lipid droplets were observed. At day five, very small lipid droplets were present, and they increased in size up to day fifteen. The diameter of the muscle fibers also increased from day two to fifteen, as well as the internuclear distance, indicating that muscle hypertrophy occurred along these days. The lipid droplets from the fat body showed a different pattern, and their diameter decreased after day two, but started to increase again at day ten. The data presented herein describes the development of the flight muscle after the final ecdysis, and modifications that occur regarding lipid stores. We show that, after molting, substrates that are present in the midgut and fat body are mobilized and directed to the ovary and flight muscle, for the adults of R. prolixus to be ready to feed and reproduce.

CCN6 influences transcription and controls mitochondrial mass and muscle organization.

Mutations in Cellular Communication Network Factor 6 (CCN6) are linked to the debilitating musculoskeletal disease Progressive Pseudo Rheumatoid Dysplasia (PPRD), which disrupts mobility. Yet, much remains unknown about CCN6 function at the molecular level. In this study, we revealed a new function of CCN6 in transcriptional regulation. We demonstrated that CCN6 localizes to chromatin and associates with RNA Polymerase II in human chondrocyte lines. Using zebrafish as a model organism we validated the nuclear presence of CCN6 and its association with RNA Polymerase II in different developmental stages from 10 hpf embryo to adult fish muscle. In concurrence with these findings, we confirmed the requirement of CCN6 in the transcription of several genes encoding mitochondrial electron transport complex proteins in the zebrafish, both in the embryonic stages and in the adult muscle. Reduction in the expression of these genes upon morpholino-mediated knockdown of CCN6 protein expression led to reduced mitochondrial mass, which correlated with defective myotome organization during zebrafish muscle development. Overall, this study suggests that the developmental musculoskeletal abnormalities linked with PPRD could be contributed at least partly by impaired expression of genes encoding mitochondrial electron transport complexes due to defects in CCN6 associated transcriptional regulation.

Ten-year longitudinal changes in muscle power, force, and velocity in young, middle-aged, and older adults.

Maximum muscle power (Pmax ) is a biomarker of physical performance in all ages. No longitudinal studies have assessed the effects of aging on Pmax obtained from the torque-velocity (T-V) relationship, which should be considered the 'gold standard'. This study evaluated the longitudinal changes in the T-V relationship and Pmax of the knee-extensor muscles in young, middle-aged, and older adults after 10years of follow-up. Four hundred eighty-nine subjects (311 men and 178 women; aged 19-68years) were tested at baseline and after a 10-year follow-up. Anthropometric data, daily protein intake, physical activity level (PAL), and knee-extension muscle function (isometric, isokinetic, and isotonic) were evaluated. A novel hybrid equation combining a linear and a hyperbolic (Hill-type) region was used to obtain the T-V relationship and Pmax of the participants, who were grouped by sex and age (young: 20-40years; middle-aged: 40-60years; and old: ≥60years). Linear mixed-effect models were used to assess effects of time, sex, and age on T-V parameters, Pmax , and body mass index (BMI). Additional analyses were performed to adjust for changes in daily protein intake and PAL. Pmax decreased in young men (-0.6% per year; P<0.001), middle-aged men and women (-1.1% to -1.4% per year; P<0.001), and older men and women (-2.2% to -2.4% per year; P≤0.053). These changes were mainly related to decrements in torque at Pmax at early age and to decrements in both torque and velocity at Pmax at older age. BMI increased among young and middle-aged adults (0.2% to 0.5% per year; P<0.001), which led to greater declines in relative Pmax in those groups. S/T0 , that is, the linear slope of the T-V relationship relative to maximal torque, exhibited a significant decline over time (-0.10%T0 ·rad·s-1 per year; P<0.001), which was significant among middle-aged men and old men and women (all P<0.05). Annual changes in PAL index were significantly associated to annual changes in Pmax (P=0.017), so the overall decline in Pmax was slightly attenuated in the adjusted model (-5.26 vs. -5.05W per year; both P<0.001). Pmax decreased in young, middle-aged, and older adults after a 10-year follow-up. The early declines in Pmax seemed to coincide with declines in force, whereas the progressive decline at later age was associated with declines in both force and velocity. A progressively blunted ability to produce force, especially at moderate to high movement velocities, should be considered a specific hallmark of aging.

Angiogenesis precedes myogenesis during regeneration following biopsy injury of skeletal muscle

BackgroundAcute injury to skeletal muscle damages myofibers and fragment capillaries, impairing contractile function and local perfusion. Myofibers and microvessels regenerate from satellite cells and from surviving microvessel fragments, respectively, to restore intact muscle. Established models of injury have used myotoxins and physical trauma to demonstrate the concurrence of myogenesis and angiogenesis during regeneration. In these models, efferocytosis removes cellular debris while basal laminae persist to provide guidance during myofiber and microvessel regeneration. It is unknown whether the spatiotemporal coupling between myofiber and microvascular regeneration persists when muscle tissue is completely removed and local guidance cues are lost.MethodsTo test whether complete removal of skeletal muscle tissue affects the spatiotemporal relationship between myogenesis and angiogenesis during regeneration, subthreshold volumetric muscle loss was created with a biopsy punch (diameter, 2 mm) through the center of the gluteus maximus (GM) in adult mice. Regeneration into the void was evaluated through 21 days post-injury (dpi). Microvascular perfusion was evaluated in vivo by injecting fluorescent dextran into the circulation during intravital imaging. Confocal imaging and histological analyses of whole-mount GM preparations and tissue cross-sections assessed the growth of microvessels and myofibers into the wound.ResultsA provisional matrix filled with PDGFRα+ and CD45+ cells spanned the wound within 1 dpi. Regenerating microvessels advanced from the edges of the wound into the matrix by 7 dpi. Nascent microvascular networks formed by 10 dpi with blood-perfused networks spanning the wound by 14 dpi. In striking contrast, the wound remained devoid of myofibers at 7 and 10 dpi. Myogenesis into the wound was apparent by 14 dpi and traversed the wound by 21 dpi. Regenerated myofibers and microvessels were disorganized compared to the uninjured muscle.ConclusionsFollowing punch biopsy of adult skeletal muscle, regenerating microvessels span the wound and become perfused with blood prior to myofiber regeneration. The loss of residual guidance cues with complete tissue removal disrupts the spatiotemporal correspondence between microvascular and myofiber regeneration. We conclude that angiogenesis precedes myogenesis during regeneration following subthreshold volumetric muscle loss.

Association of gross motor function and activities of daily living with muscle mass of the trunk and lower extremity muscles, range of motion, and spasticity in children and adults with cerebral palsy

ABSTRACT Purpose We examined the association of gross motor function and activities of daily living (ADL) with muscle mass of the trunk and lower extremity muscles in children and adults with cerebral palsy (CP). Methods The subjects were 32 children and adults with CP. Muscle thickness of the trunk and lower extremity muscles was measured using an ultrasound imaging device. Results Stepwise regression analysis revealed that the thoracic erector spinae muscle thickness was a significant and independent factor of gross motor function. Stepwise regression analysis also showed that the thickness of the rectus abdominis and vastus lateralis muscles were significant and independent factors of ADL. Conclusions Our findings suggest that declined gross motor function is associated with decreased thoracic erector spinae muscle mass in children and adults with CP. The results also indicate that declined ADL is associated with decreased muscle mass of the rectus abdominis and vastus lateralis muscles.

Characterizing the functional impact of embryonic myosin mutations in stem cell-derived skeletal muscle.

Divergent contractile properties of different myosins suggest that each isoform performs distinct functions in embryonic and adult skeletal muscles. To better understand the role of embryonic myosin heavy chain 3 (MYH3) in regulating skeletal muscle pattern formation, growth and regeneration, our group investigated the functional defects caused by multiple mutations. Our team generated human induced pluripotent stem cell (iPSC) lines bearing T178I or R672C mutations in MYH3. Human iPSCs bearing these mutations were differentiated into skeletal muscle and evaluated for differences in the structure, maturation and functional performance of the sarcomere. Isolated myofibril mechanics showed T178I myotubes generated significantly greater specific force compared to control myofibrils. Diseased myofibrils also expressed significantly slower rates of relaxation and prolonged thin filament deactivation compared to controls. Myofibril preparations were also used to compare ATP binding rates and showed homozygous R672C myofibrils have faster ATP binding rates compared to heterozygous and control preparations. When incorporated into 3D engineered tissues, these changes in MYH3 mechanics resulted in slower relaxation kinetics and an incomplete relaxation of the tissue in response to repetitive tetanic stimulation, which may explain the emergence of developmental contractures in patients possessing MYH3 mutations. Molecular dynamics simulations of R672C and T178I mutations in the post rigor state showed greater separation between the β-sheet and SH-Helix than in controls. Mutations also disrupted local salt bridges and hydrogen bond formation in some residues that help stabilize the nucleotide binding pocket and converter-connected helix. Interrupted structural communication between the nucleotide binding pocket and surrounding functional regions may underly the impaired relaxation phenotype. Ongoing studies will further characterize the effect of mutation on actin-myosin binding, crossbridge cycling kinetics, the maturation of skeletal myotubes over time, and myosin isoform switching dynamics.

Glu-rich N-terminal variable region of troponin T functions as a myofilament Ca2+-reservoir to reduce SR Ca2+-cycling.

The long Glu-rich N-terminal segment of avian pectoral muscle troponin T (TnT) and C-terminal extension of insect TnT are multi-valent Ca2+-binding sites with physiologically relevant affinities. Genetic deletion of the Glu-rich C-terminal extension of insect TnT decreases muscle and heart functions but increases the tolerance of cardiomyocytes to Ca2+ overloading, indicating decreased [Ca2+]i. In contrast, cardiomyocytes from gene-edited mice expressing modified cardiac TnT with N-terminal variable region replaced by a C-terminal segment of insect TnT (51E-cTnT) show higher shortening amplitude and contraction and relaxation velocities than that of wild type (WT) control. Isolated 51E-cTnT cardiomyocytes exhibited shorter resting sarcomeres, which can be corrected by inhibiting myosin ATPase, indicating increased resting [Ca2+]i. A similar gain-of-function model,fast skeletal muscle TnTgene-edited mice with the N-terminal variable region replaced by the N-terminal segment of adult turkey pectoral muscle TnT (Tx9-fsTnT), shows normal baseline muscle functions but slower and less rising of contractile force during caffeine-induced Ca2+ release from sarcoplasmic reticulum (SR) than that of WT control, indicating a decreased SR Ca2+ store. Consistently, 51E-cTnT mouse hearts respond to beta-adrenergic stimulation or Ca2+ agonist with higher than WT ventricular peak pressure and diastolic velocity. The data support a hypothesis that the Glu-rich segments of insect and avian flight muscle TnT function as myofilament Ca2+ reservoirs to tune muscle kinetics and reduce the amount of Ca2+ cycled during contraction and relaxation. The functional effects are prominent in heterozygotes of the genetically modified flies and mice whereas the loss-of-function in Drosophila TnT and gain-of-function in 51E-cTnT and Tx9-fsTnT mice both produced homozygote lethality, indicating sensitive dose effects of the myofilament Ca2+ reservoir for use as a potent mechanism to improve energetic efficiency of cardiac and skeletal muscles.

It is possible to calculate agonist efficiency from receptor structure.

Agonists turn on nicotinic acetylcholine receptors (AChRs) because they bind weakly to resting-C and strongly to active-O conformations of their target sites. Agonist efficiency (η) is the percent of chemical binding energy converted into the mechanical work of C-O gating that depends on the weak/strong binding energy ratio, η=1-ΔΔGC/ΔΔGO. κ has been measured in adult muscle nicotinic AChRs for many agonists and binding site mutations by using electrophysiology. We have compared these experimental values to those calculated from binding free energy estimates to C versus O states at the α-δ binding site. MD simulations were done with an agonist bound to the neurotransmitter site of a muscle AChR (6UVW.pdb). After removing the bound Btx, ligands were docked into the site of the extracellular domain dimer. A five-step equilibration (pre-production) was followed by 200 ns of un-restrained MD simulation using AMBER 2019. The three energy minimas was identified in sequence (m1, m2 and m3) using Principle component analysis (PCA), furthermore their free energies were calculated via MM-PBSA. In general the calculated binding free energy didn’t corroborate well with experimental ones, the calculated binding free energy ratios (η) values for epibatidine, epiboxidine, carbamylcholine and ACh were similar to those estimated by using electophysiology. The results confirm the correctness of our identification of closed-C (m1) versus open-O (m3) binding pocket structures, and suggest that it might be possible in general to calculate agonist efficiency using crystal and MD studies. The structural correlates of agonist efficiency are under investigation.

Adult skeletal muscle peroxisome proliferator-activated receptor γ -related coactivator 1 is involved in maintaining mitochondrial content.

The peroxisome proliferator-activated receptor γ coactivator-1 (PGC-1) family of transcriptional coactivators are regulators of mitochondrial oxidative capacity and content in skeletal muscle. Many of these conclusions are based primarily on gain-of-function studies using muscle-specific overexpression of PGC1s. We have previously reported that genetic deletion of both PGC-1α and PGC-1β in adult skeletal muscle resulted in a significant reduction in oxidative capacity with no effect on mitochondrial content. However, the contribution of PGC-1-related coactivator (PRC), the third PGC-1 family member, in regulating skeletal muscle mitochondria is unknown. Therefore, we generated an inducible skeletal muscle-specific PRC knockout mouse (iMS-PRC-KO) to assess the contribution of PRC in skeletal muscle mitochondrial function. We measured mRNA expression of electron transport chain (ETC) subunits as well as markers of mitochondrial content in the iMS-PRC-KO animals and observed an increase in ETC gene expression and mitochondrial content. Furthermore, the increase in ETC gene expression and mitochondrial content was associated with increased expression of PGC-1α and PGC-1β. We therefore generated an adult-inducible PGC-1 knockout mouse in which all PGC-1 family members are deleted (iMS-PGC-1TKO). The iMS-PGC-1TKO animals exhibited a reduction in ETC mRNA expression and mitochondrial content. These data suggest that in the absence of PRC alone, compensation occurs by increasing PGC-1α and PGC-1β to maintain mitochondrial content. Moreover, the removal of all three PGC-1s in skeletal muscle results in a reduction in both ETC mRNA expression and mitochondrial content. Taken together, these results suggest that PRC plays a role in maintaining baseline mitochondrial content in skeletal muscle.

The myonuclear domain in adult skeletal muscle fibres: past, present and future.

Most cells in the body are mononuclear whereas skeletal muscle fibres are uniquely multinuclear. The nuclei of muscle fibres (myonuclei) are usually situated peripherally which complicates the equitable distribution of gene products. Myonuclear abundance can also change under conditions such as hypertrophy and atrophy. Specialised zones in muscle fibres have different functions and thus distinct synthetic demands from myonuclei. The complex structure and regulatory requirements of multinuclear muscle cells understandably led to the hypothesis that myonuclei govern defined 'domains' to maintain homeostasis and facilitate adaptation. The purpose of this review is to provide historical context for the myonuclear domain and evaluate its veracity with respect to mRNA and protein distribution resulting from myonuclear transcription. We synthesise insights from past and current in vitro and in vivo genetically modified models for studying the myonuclear domain under dynamic conditions. We also cover the most contemporary knowledge on mRNA and protein transport in muscle cells. Insights from emerging technologies such as single myonuclear RNA-sequencing further inform our discussion of the myonuclear domain. We broadly conclude: (1) the myonuclear domain can be flexible during muscle fibre growth and atrophy, (2) the mechanisms and role of myonuclear loss and motility deserve further consideration, (3) mRNA in muscle is actively transported via microtubules and locally restricted, but proteins may travel far from a myonucleus of origin and (4) myonuclear transcriptional specialisation extends beyond the classic neuromuscular and myotendinous populations. A deeper understanding of the myonuclear domain in muscle may promote effective therapies for ageing and disease.

Comparison of shear wave elastography with gray-scale USG and CT for quantitative evaluation of rectus femoris muscle.

The purpose of this study was to compare the measurement of shear wave elastography (SWE) and gray scale ultrasonography (GSU) and CT attenuation of mid-rectus femoris (RF) muscle in healthy adults. This prospective study included 70 participants with a healthy body mass index (<25 kg/m2 ) between June 2019 and January 2020. Echo intensity (EI) grading of RF on GSU was performed. SWE was performed for the three levels of the RF. Measurements were repeated 10min after the first measurement. The mid-RF attenuation on CT was also measured. Interobserver agreement of EI grade among three readers was assessed using weighted-kappa statistics. The reliability of SWE was assessed using intraclass correlation coefficient. The correlations between the SWE and CT/GSU measurements were analyzed. Interobserver agreement of EI grade on GSU by the three radiologists was moderate to substantial (k=0.562-0.767). The inter-session agreements for SWE were almost perfect for mid RF (k=0.822-0.829) and substantial for proximal and distal RF (k=0.767-0.795). There were significant correlations between SWE-EI and SWE-CT attenuation (p < 0.001, respectively) at the mid-RF. SWE measurements on mid-RF demonstrated the highest reliability. SWE parameters showed a strong correlation with EI on GSU and attenuation on CT.

Activation of PDGF Signaling in the Adult Muscle Stem Cell Niche in Patients With Type 2 Diabetes Mellitus.

Type 2 diabetes mellitus (T2D) negatively affects muscle mass and function throughout life. Whether adult muscle stem cells contribute to the decrease in muscle health is not clear and insights into the stem cell niche are difficult to obtain. To establish the upstream signaling pathway of microRNA (miR)-501, a marker of activated myogenic progenitor cells, and interrogate this pathway in muscle biopsies from patients with T2D. Analysis of primary muscle cell cultures from mice and 4 normoglycemic humans and muscle biopsies from 7 patients with T2D and 7 normoglycemic controls using gene expression, information on histone methylation, peptide screening, and promoter assays. miR-501 shares the promoter of its host gene, isoform 2 of chloride voltage-gated channel 5 (CLCN5-2), and miR-501 expression increases during muscle cell differentiation. We identify platelet-derived growth factor (PDGF) as an upstream regulator of CLCN5-2 and miR-501 via Janus kinase/signal transducer and activator of transcription. Skeletal muscle biopsies from patients with T2D revealed upregulation of PDGF (1.62-fold, P = .002), CLCN5-2 (2.85-fold, P = .03), and miR-501 (1.73-fold, P = .02) compared with normoglycemic controls. In addition, we observed a positive correlation of PDGF and miR-501 in human skeletal muscle (r = 0.542, P = .045, n = 14). We conclude that paracrine signaling in the adult muscle stem cells niche is activated in T2D. Expression analysis of the PDGF-miR-501 signaling pathway could represent a powerful tool to classify patients in clinical trials that aim to improve muscle health and glucose homeostasis in patients with diabetes.