Skeletal Muscle Fibers Research Articles

Influence of lipid bilayer on the structure of the muscle-type nicotinic acetylcholine receptor

The muscle-type nicotinic acetylcholine receptor is a transmitter-gated ion channel residing in the plasma membrane of electrocytes and striated muscle cells. It is present predominantly at synaptic junctions, where it effects rapid depolarization of the postsynaptic membrane in response to acetylcholine released into the synaptic cleft. Previously, cryo-EM of intact membrane from Torpedo revealed that the lipid bilayer surrounding the junctional receptor has a uniquely asymmetric and ordered structure, due to a high concentration of cholesterol. It is now shown that this special lipid environment influences the transmembrane (TM) folding of the protein. All five submembrane MX helices of the membrane-intact junctional receptor align parallel to the surface of the cholesterol-ordered lipids in the inner leaflet of the bilayer; also, the TM helices in the outer leaflet are splayed apart. However in the structure obtained from the same protein after extraction and incorporation in nanodiscs, the MX helices do not align to a planar surface, and the TM helices arrange compactly in the outer leaflet. Realignment of the MX helices of the nanodisc-solved structure to a planar surface converts their adjoining TM helices into an obligatory splayed configuration, characteristic of the junctional receptor. Thus, the form of the receptor sustained by the special lipid environment of the synaptic junction is the one that mediates fast synaptic transmission; whereas, the nanodisc-embedded protein may be like the extrajunctional form, existing in a disordered lipid environment.

Atypical Intramuscular Lipoma of the Tongue. Case Report.

Although they are frequently benign tumors of mesenchymal origin, lipomas sporadically develop in the oral cavity. Intramuscular lipomas, which originate inside muscles, are a rare subtype. When found in the tongue, they are considerably more uncommon and its etiology remains unclear. Histological examination of the specimen shows mature adipocytes of uniform size with minimal vascularization and sometimes striated muscle fibers are to be seen in the periphery. A yellowish submucosal mass that is slowly expanding, painless, and asymptomatic, is typically seen based on a patient's history and clinical examination. Lipomas that occur in uncommon locations should be mentioned and should be considered in the differential diagnosis of an oral mass. The main treatment option is a complete surgical excision. The purpose of this report is to describe a case of a 43-year-old female patient with an atypical, well-defined intramuscular lipoma in her tongue that began to form around 10 months ago.

Identification of different myofiber types in pigs muscles and construction of regulatory networks

BackgroundSkeletal muscle is composed of muscle fibers with different physiological characteristics, which plays an important role in regulating skeletal muscle metabolism, movement and body homeostasis. The type of skeletal muscle fiber directly affects meat quality. However, the transcriptome and gene interactions between different types of muscle fibers are not well understood.ResultsIn this paper, we selected 180-days-old Large White pigs and found that longissimus dorsi (LD) muscle was dominated by fast-fermenting myofibrils and soleus (SOL) muscle was dominated by slow-oxidizing myofibrils by frozen sections and related mRNA and protein assays. Here, we selected LD muscle and SOL muscle for transcriptomic sequencing, and identified 312 differentially expressed mRNA (DEmRs), 30 differentially expressed miRNA (DEmiRs), 183 differentially expressed lncRNA (DElRs), and 3417 differentially expressed circRNA (DEcRs). The ceRNA network included ssc-miR-378, ssc-miR-378b-3p, ssc-miR-24-3p, XR_308817, XR_308823, SMIM8, MAVS and FOS as multiple core nodes that play important roles in muscle development. Moreover, we found that different members of the miR-10 family expressed differently in oxidized and glycolytic muscle fibers, among which miR-10a-5p was highly expressed in glycolytic muscle fibers (LD) and could target MYBPH gene mRNA. Therefore, we speculate that miR-10a-5p may be involved in the transformation of muscle fiber types by targeting the MYHBP gene. In addition, PPI analysis of differentially expressed mRNA genes showed that ACTC1, ACTG2 and ACTN2 gene had the highest node degree, suggesting that this gene may play a key role in the regulatory network of muscle fiber type determination.ConclusionsWe can conclude that these genes play a key role in regulating muscle fiber type transformation. Our study provides transcriptomic profiles and ceRNA interaction networks for different muscle fiber types in pigs, providing reference for the transformation of pig muscle fiber types and the improvement of meat quality.

Irisin and Its Role in Postmenopausal Osteoporosis and Sarcopenia.

Menopause, an extremely delicate phase in a woman's life, is characterized by a drop in estrogen levels. This decrease has been associated with the onset of several diseases, including postmenopausal osteoporosis and sarcopenia, which often coexist in the same person, leading to an increased risk of fractures, morbidity, and mortality. To date, there are no approved pharmacological treatments for sarcopenia, while not all of those approved for postmenopausal osteoporosis are beneficial to muscles. In recent years, research has focused on the field of myokines, cytokines, or peptides secreted by skeletal muscle fibers following exercise. Among these, irisin has attracted great interest as it possesses myogenic properties but at the same time exerts anabolic effects on bone and could therefore represent the link between muscle and bone. Therefore, irisin could represent a new therapeutic strategy for the treatment of osteoporosis and also serve as a new biomarker of sarcopenia, thus facilitating diagnosis and pharmacological intervention. The purpose of this review is to provide an updated summary of what we know about the role of irisin in postmenopausal osteoporosis and sarcopenia.

Zebrafish polg2 knock-out recapitulates human POLG-disorders; implications for drug treatment.

The human mitochondrial DNA polymerase gamma is a holoenzyme, involved in mitochondrial DNA (mtDNA) replication and maintenance, composed of a catalytic subunit (POLG) and a dimeric accessory subunit (POLG2) conferring processivity. Mutations in POLG or POLG2 cause POLG-related diseases in humans, leading to a subset of Mendelian-inherited mitochondrial disorders characterized by mtDNA depletion (MDD) or accumulation of multiple deletions, presenting multi-organ defects and often leading to premature death at a young age. Considering the paucity of POLG2 models, we have generated a stable zebrafish polg2 mutant line (polg2ia304) by CRISPR/Cas9 technology, carrying a 10-nucleotide deletion with frameshift mutation and premature stop codon. Zebrafish polg2 homozygous mutants present slower development and decreased viability compared to wild type siblings, dying before the juvenile stage. Mutants display a set of POLG-related phenotypes comparable to the symptoms of human patients affected by POLG-related diseases, including remarkable MDD, altered mitochondrial network and dynamics, and reduced mitochondrial respiration. Histological analyses detected morphological alterations in high-energy demanding tissues, along with a significant disorganization of skeletal muscle fibres. Consistent with the last finding, locomotor assays highlighted a decreased larval motility. Of note, treatment with the Clofilium tosylate drug, previously shown to be effective in POLG models, could partially rescue MDD in Polg2 mutant animals. Altogether, our results point at zebrafish as an effective model to study the etiopathology of human POLG-related disorders linked to POLG2, and a suitable platform to screen the efficacy of POLG-directed drugs in POLG2-associated forms.

Grape seed proanthocyanidin extract promotes skeletal muscle fiber type transformation through modulation of cecal microbiota and enhanced butyric acid production.

The conversion of fast-twitch fibers into slow-twitch fibers within skeletal muscle plays a crucial role in improving physical stamina and safeguarding against metabolic disorders in individuals. Grape seed proanthocyanidin extract (GSPE) possesses numerous pharmacological and health advantages, effectively inhibiting the onset of chronic illnesses. However, there is a lack of research on the specific mechanisms by which GSPE influences muscle physiology and gut microbiota. This study aims to investigate the role of gut microbiota and their metabolites in GSPE regulation of skeletal muscle fiber type conversion. In this experiment, 54 male BALB/c mice were randomly divided into three groups: basal diet, basal diet supplemented with GSPE, and basal diet supplemented with GSPE and antibiotics. During the feeding period, glucose tolerance and forced swimming tests were performed. After euthanasia, samples of muscle and feces were collected for analysis. The results showed that GSPE increased the muscle mass and anti-fatigue capacity of the mice, as well as the expression of slow-twitch fibers. However, the beneficial effects of GSPE on skeletal muscle fibers disappeared after adding antibiotics to eliminate intestinal microorganisms, suggesting that GSPE may play a role by regulating intestinal microbial structure. In addition, GSPE increased the relative abundance of Blautia, Muribaculaceae, and Enterorhabdus, as well as butyrate production. Importantly, these gut microbes exhibited a significant positive correlation with the expression of slow-twitch muscle fibers. In conclusion, supplementation with GSPE can increase the levels of slow-twitch fibers by modulating the gut microbiota, consequently prolonging the duration of exercise before exhaustion. PRACTICAL APPLICATION: This research suggests that grape seed proanthocyanidin extract (GSPE) has potential applications in improving physical stamina and preventing metabolic disorders. By influencing the gut microbiota and increasing butyric acid production, GSPE contributes to the conversion of fast-twitch muscle fibers into slow-twitch fibers, thereby enhancing anti-fatigue capacity and exercise endurance. While further studies are needed, incorporating GSPE into dietary supplements or functional foods could support individuals seeking to optimize their exercise performance and overall metabolic health.

Temporal regulation of the Mediator complex during muscle proliferation, differentiation, regeneration, aging, and disease.

Genesis of skeletal muscle relies on the differentiation and fusion of mono-nucleated muscle progenitor cells into the multi-nucleated muscle fiber syncytium. The temporally-controlled cellular and morphogenetic changes underlying this process are initiated by a series of highly coordinated transcription programs. At the core, the myogenic differentiation cascade is driven by muscle-specific transcription factors, i.e., the Myogenic Regulatory Factors (MRFs). Despite extensive knowledge on the function of individual MRFs, very little is known about how they are coordinated. Ultimately, highly specific coordination of these transcription programs is critical for their masterfully timed transitions, which in turn facilitates the intricate generation of skeletal muscle fibers from a naïve pool of progenitor cells. The Mediator complex links basal transcriptional machinery and transcription factors to regulate transcription and could be the integral component that coordinates transcription factor function during muscle differentiation, growth, and maturation. In this study, we systematically deciphered the changes in Mediator complex subunit expression in skeletal muscle development, regeneration, aging, and disease. We incorporated our in vitro and in vivo experimental results with analysis of publicly available RNA-seq and single nuclei RNA-seq datasets and uncovered the regulation of Mediator subunits in different physiological and temporal contexts. Our experimental results revealed that Mediator subunit expression during myogenesis is highly dynamic. We also discovered unique temporal patterns of Mediator expression in muscle stem cells after injury and during the early regeneration period, suggesting that Mediator subunits may have unique contributions to directing muscle stem cell fate. Although we observed few changes in Mediator subunit expression in aging muscles compared to younger muscles, we uncovered extensive heterogeneity of Mediator subunit expression in dystrophic muscle nuclei, characteristic of chronic muscle degeneration and regeneration cycles. Taken together, our study provides a glimpse of the complex regulation of Mediator subunit expression in the skeletal muscle cell lineage and serves as a springboard for mechanistic studies into the function of individual Mediator subunits in skeletal muscle.

RANKL signaling drives skeletal muscle into the oxidative profile.

The bone-muscle unit refers to the reciprocal regulation between bone and muscle by mechanical interaction and tissue communication via soluble factors. The RANKL stimulation induces mitochondrial biogenesis and increases the oxidative capacity in osteoclasts and adipocytes. RANKL may bind to the membrane bound RANK or to osteoprotegerin (OPG), a decoy receptor that inhibits RANK-RANKL activation. RANK is highly expressed in skeletal muscle, but the contribution of RANKL to healthy skeletal muscle fiber remains elusive. Here we show that RANKL stimulation in C2C12-derived myotubes induced activation of mitochondrial biogenesis pathways as detected by RNA-seq and western blot. RANKL expanded the mitochondrial reticulum, as shown by mitochondrial DNA quantification and MitoTracker staining, and boosted the spare respiratory capacity. Using MEK and MAPK inhibitors, we found that RANKL signals via ERK and p38 to induce mitochondrial biogenesis. The soleus from OPG-/- and OPG+/- mice showed higher respiratory rates compared to C57BL6/J WT mice, which correlates with high serum RANKL levels. RANKL infusion using a mini-osmotic pump in WT mice increased the number of mitochondria, boosted the respiratory rate, increased succinate dehydrogenase activity in skeletal muscle, and improved the fatigue resistance of gastrocnemius. Therefore, our findings reveal a new role of RANKL as an osteokine-like protein that impacts muscle fiber metabolism.

Localization of sensory nerve terminals containing calcitonin gene-related peptide (CGRP) on striated muscle fibers in the rat esophagus: Evidence for triple innervation via motor endplates

BackgroundMany esophageal striated muscles of mammals are dually innervated by the vagal and enteric nerves. Recently, substance P (SP)-sensory nerve terminals with calcitonin gene-related peptide (CGRP) were found on a few striated muscle fibers in the rat esophagus, implying that these muscle fibers are triply innervated. In this study, we examined the localization and origin of CGRP-nerve endings in striated muscles to consider their possible roles in the esophagus regarding triple innervation. MethodsWholemounts of the rat esophagus were immunolabeled to detect CGRP-nerve endings in striated muscles. Also, retrograde tracing was performed by injecting Fast Blue (FB) into the esophagus, and cryostat sections of the medulla oblongata, nodose ganglion (NG), and the tenth thoracic (T10) dorsal root ganglion (DRG) were immunostained to identify the origin of the CGRP-nerve endings. ResultsCGRP-fine, varicose nerve endings were localized in motor endplates on a few esophageal striated muscle fibers (4 %), most of which received nitric oxide (NO) synthase nerve terminals, and most of the CGRP nerve endings were SP- and transient receptor potential vanilloid member 1 (TRPV1)-positive. Retrograde tracing showed many FB-labeled CGRP-neurons positive for SP and TRPV1 in the NG and T10 DGR. ConclusionsThis study suggests that the CGRP-varicose nerve endings containing SP and TRPV1 in motor endplates are sensory, and a few esophageal striated muscle fibers are triply innervated. The nerve endings may detect acetylcholine-derived acetic acid from the vagal motor nerve endings and NO from esophageal intrinsic nerve terminals in the motor endplates to regulate esophageal motility.

CircUSP13 facilitates the fast-to-slow myofiber shift via the MAPK/ERK signaling pathway in goat skeletal muscles.

Understanding how skeletal muscle fiber proportions are regulated is essential for understanding muscle function and improving the quality of mutton. While circular RNA (circRNA) has a critical function in myofiber type transformation, the specific mechanisms are not yet fully understood. Prior evidence indicates that circular ubiquitin-specific peptidase 13 (circUSP13) can promote myoblast differentiation by acting as a ceRNA, but its potential role in myofiber switching is still unknown. Herein, we found that circUSP13 enhanced slow myosin heavy chain (MyHC-slow) and suppressed MyHC-fast expression in goat primary myoblasts (GPMs). Meanwhile, circUSP13 evidently enhanced the remodeling of the mitochondrial network while inhibiting the autophagy of GPMs. We obtained fast-dominated myofibers, via treatment with rotenone, and further demonstrated the positive role of circUSP13 in the fast-to-slow transition. Mechanistically, activation of the mitogen-activated protein kinase/extracellular signal-regulated kinase (MAPK/ERK) pathway significantly impaired the slow-to-fast shift in fully differentiated myotubes, which was restored by circUSP13 or IGF1 overexpression. In conclusion, circUSP13 promoted the fast-to-slow myofiber type transition through MAPK/ERK signaling in goat skeletal muscle. These findings provide novel insights into the role of circUSP13 in myofiber type transition and contribute to a better understanding of the genetic mechanisms underlying meat quality.

A deep learning-based approach for fully automated segmentation and quantitative analysis of muscle fibers in pig skeletal muscle

Muscle fiber properties exert a significant influence on pork quality, with cross-sectional area (CSA) being a crucial parameter closely associated with various meat quality indicators, such as shear force. Effectively identifying and segmenting muscle fibers in a robust manner constitutes a vital initial step in determining CSA. This step is highly intricate and time-consuming, necessitating an accurate and automated analytical approach. One limitation of existing methods is their tendency to perform well on high signal-to-noise ratio images of intact, healthy muscle fibers but their lack of validation on more complex image datasets featuring significant morphological changes, such as the presence of ice crystals. In this study, we undertake the fully automatic segmentation of muscle fiber microscopic images stained with myosin adenosine triphosphate (mATPase) activity using a deep learning architecture known as SOLOv2. Our objective is to efficiently derive accurate measurements of muscle fiber size and distribution. Tests conducted on actual images demonstrate that our method adeptly handles the intricate task of muscle fiber segmentation, yielding quantitative results amenable to statistical analysis and displaying reliability comparable to manual analysis.

Oscillatory work and the step that generates force in single myofibrils from rabbit psoas.

The elementary molecular step that generates force by cross-bridges (CBs) in active muscles has been under intense investigation in the field of muscle biophysics. It is known that an increase in the phosphate (Pi) concentration diminishes isometric force in active fibers, indicating a tight coupling between the force generation step and the Pi release step. The question asked here is whether the force generation occurs before Pi release or after release. We investigated the effect of Pi on oscillatory work production in single myofibrils and found that Pi-attached state(s) to CBs is essential for its production. Oscillatory work is the mechanism that allows an insect to fly by beating its wings, and it also has been observed in skeletal and cardiac muscle fibers, implying that it is an essential feature of all striated muscle types. With our studies, oscillatory work disappears in the absence of Pi in experiments using myofibrils. This suggests that force is generated during a transition between steps of oscillatory work production, and that the states involved in force production must have Pi attached. With sinusoidal analysis, we obtained the kinetic constants around the Pi release steps, established a CB scheme, and evaluated force generated (and supported) by each CB state. Our results demonstrate that force is generated before Pi is released, and the same force is maintained after Pi is released. Stretch activation and/or delayed tension can also be explained with this CB scheme and forms the basis of force generation and oscillatory work production.

Aminophylline Improves Ventilator-Induced Diaphragmatic Dysfunction by Targeting IGF-1-FOXO1-MURF1 Pathway.

The usage of life-saving mechanical ventilation (MV) could cause ventilator-induced diaphragmatic dysfunction (VIDD), increasing both mortality and morbidity. Aminophylline (AP) has the potential to enhance the contractility of animal skeletal muscle fibers and improve the activity of human respiratory muscles, and the insulin-like growth factor-1 (IGF-1)- forkhead box protein O1 (FOXO1)-muscle RING finger-1 (MURF1) pathway plays a crucial role in skeletal muscle dysfunction. This study aimed to investigate the impact of AP on VIDD and to elucidate the role of the IGF-1-FOXO1-MURF1 pathway as an underlying mechanism. Rat models of VIDD were established through MV treatment. IGF-1 lentiviral (LV) interference (LV-IGF-1-shRNA; controlled by lentiviral negative control LV-NC) was employed to inhibit IGF-1 expression and thereby block the IGF-1-FOXO1-MURF1 pathway. Protein and mRNA levels of IGF-1, FOXO1, and MURF1 were assessed using western blot and real-time reverse transcriptase-polymerase chain reaction (RT-qPCR), respectively. Diaphragm contractility and morphometry were examined through measurement of compound muscle action potentials (CMAPs) and hematoxylin and eosin (H&E) staining. Oxidative stress was evaluated by levels of hydrogen peroxide (H2O2), superoxide dismutase (SOD), antioxidant glutathione (GSH), and carbonylated protein. Mitochondrial stability was assessed by measuring the mitochondrial membrane potential (MMP), and mitochondrial fission and mitophagy were examined through protein levels of dynamin-related protein 1 (DRP1), mitofusin 2 protein (MFN2), phosphatase and tensin homolog (PTEN)-induced kinase 1 (PINK1), and Parkin (western blot). Apoptosis was evaluated using the terminal deoxynucleotidyl transferase-mediated uridine 5'-triphosphate (UTP) nick-end labeling (TUNEL) assay and levels of Bax, B-cell lymphoma 2 (BCL-2), and Caspase-3. Levels of Atrogin-1, neuronally expressed developmentally downregulated 4 (NEDD4), and muscle ubiquitin ligase of SCF complex in atrophy-1 (MUSA1) mRNA, as well as ubiquitinated protein, were utilized to determine protein degradation. Furthermore, the SUnSET (surface sensing of translation) method was employed to determine rates of protein synthesis. MV treatment upregulated IGF-1 while downregulated FOXO1 and MURF1 (p < 0.05). AP administration reversed IGF-1, FOXO1 and MURF1 (p < 0.05), which was suppressed again by IGF-1 inhibition (p < 0.05), demonstrating the blockage of the IGF-1-FOXO1-MURF1 pathway. MV treatment caused decreased CMAP and cross-sectional areas of diaphragm muscle fibers, and increased time course of CMAP (p < 0.05). Additionally, oxidative stress, cell apoptosis, and protein degradation were increased and mitochondrial stability was decreased by MV treatment (p < 0.05). Conversely, AP administration reversed all these changes induced by MV, but this reversal was disrupted by the blockage of the IGF-1-FOXO1-MURF1 pathway. In this study, MV treatment induced symptoms of VIDD in rats, which were all effectively reversed by AP regulating the IGF-1-FOXO1-MURF1 pathway, demonstrating the potential of AP in ameliorating VIDD.

ADAR2 deficiency ameliorates non-alcoholic fatty liver disease and muscle atrophy through modulating serum amyloid A1.

Non-alcoholic fatty liver disease (NAFLD) is the most common cause of chronic liver disease worldwide. Sarcopenia is a syndrome characterized by progressive and generalized loss of skeletal muscle mass and strength, which is commonly associated with NAFLD. Adenosine-to-inosine editing, catalysed by adenosine deaminase acting on RNA (ADAR), is an important post-transcriptional modification of genome-encoded RNA transcripts. Three ADAR gene family members, including ADAR1, ADAR2 and ADAR3, have been identified. However, the functional role of ADAR2 in obesity-associated NAFLD and sarcopenia remains unclear. ADAR2+/+/GluR-BR/R mice (wild type [WT]) and ADAR2-/-/GluR-BR/R mice (ADAR2 knockout [KO]) were subjected to feeding with standard chow or high-fat diet (HFD) for 20weeks at the age of 5weeks. The metabolic parameters, hepatic lipid droplet, grip strength test, rotarod test, muscle weight, fibre cross-sectional area (CSA), fibre types and protein associated with protein degradation were examined. Systemic and local tissues serum amyloid A1 (SAA1) were measured. The effects of SAA1 on C2C12 myotube atrophy were investigated. ADAR2 KO mice fed with HFD exhibited lower body weight (-7.7%, P<0.05), lower liver tissue weight (-20%, P<0.05), reduced liver lipid droplets in concert with a decrease in hepatic triglyceride content (-24%, P<0.001) and liver injury (P<0.01). ADAR2 KO mice displayed protection against HFD-induced glucose intolerance, insulin resistance and dyslipidaemia. Skeletal muscle mass (P<0.01), muscle strength (P<0.05), muscle endurance (P<0.001) and fibre size (CSA; P<0.0001) were improved in ADAR2 KO mice fed with HFD compared with WT mice fed with HFD. Muscle atrophy-associated transcripts, such as forkhead box protein O1, muscle atrophy F-box/atrogin-1 and muscle RING finger 1/tripartite motif-containing 63, were decreased in ADAR2 KO mice fed with HFD compared with WT mice fed with HFD. ADAR2 deficiency attenuates HFD-induced local liver and skeletal muscle tissue inflammation. ADAR2 deficiency abolished HFD-induced systemic (P<0.01), hepatic (P<0.0001) and muscular (P<0.001) SAA1 levels. C2C12 myotubes treated with recombinant SAA1 displayed a decrease in myotube length (-37%, P<0.001), diameter (-20%, P<0.01), number (-39%, P<0.001) and fusion index (-46%, P<0.01). Myogenic markers (myosin heavy chain and myogenin) were decreased in SAA1-treated myoblast C2C12 cells. These results provide novel evidence that ADAR2 deficiency may be important in obesity-associated sarcopenia and NAFLD. Increased SAA1 might be involved as a regulatory factor in developing sarcopenia in NAFLD.

Skeletal muscle fiber type and TMS-induced muscle relaxation in unfatigued and fatigued knee-extensor muscles.

The force drop after transcranial magnetic stimulation (TMS) delivered to the motor cortex during voluntary muscle contractions could inform about muscle relaxation properties. Because of the physiological relation between skeletal muscle fiber-type distribution and size and muscle relaxation, TMS could be a noninvasive index of muscle relaxation in humans. By combining a noninvasive technique to record muscle relaxation in vivo (TMS) with the gold standard technique for muscle tissue sampling (muscle biopsy), we investigated the relation between TMS-induced muscle relaxation in unfatigued and fatigued states, and muscle fiber-type distribution and size. Sixteen participants (7F/9M) volunteered to participate. Maximal knee-extensor voluntary isometric contractions were performed with TMS before and after a 2-min sustained maximal voluntary isometric contraction. Vastus lateralis muscle tissue was obtained separately from the participants' dominant limb. Fiber type I distribution and relative cross-sectional area of fiber type I correlated with TMS-induced muscle relaxation at baseline (r = 0.67, adjusted P = 0.01; r = 0.74, adjusted P = 0.004, respectively) and normalized TMS-induced muscle relaxation as a percentage of baseline (r = 0.50, adjusted P = 0.049; r = 0.56, adjusted P = 0.031, respectively). The variance in the normalized peak relaxation rate at baseline (59.8%, P < 0.001) and in the fatigue resistance (23.0%, P = 0.035) were explained by the relative cross-sectional area of fiber type I to total fiber area. Fiber type I proportional area influences TMS-induced muscle relaxation, suggesting TMS as an alternative method to noninvasively inform about skeletal muscle relaxation properties.NEW & NOTEWORTHY Transcranial magnetic stimulation (TMS)-induced muscle relaxation reflects intrinsic muscle contractile properties by interrupting the drive from the central nervous system during voluntary muscle contractions. We showed that fiber type I proportional area influences the TMS-induced muscle relaxation, suggesting that TMS could be used for the noninvasive estimation of muscle relaxation in unfatigued and fatigued human muscles when the feasibility of more direct method to study relaxation properties (i.e., muscle biopsy) is restricted.

Transcriptomic regulatory analysis of skeletal muscle development in landrace pigs

The development of pig skeletal muscle is a complex dynamic regulation process, which mainly includes the formation of primary and secondary muscle fibers, the remodeling of muscle fibers, and the maturation of skeletal muscle; However, the regulatory mechanism of the entire developmental process remains unclear. This study analyzed the whole-transcriptome data of skeletal muscles at 27 developmental nodes (E33-D180) in Landrace pigs, and their key regulatory factors in the development process were identified using the bioinformatics method. Firstly, we constructed a transcriptome expression map of skeletal muscle development from embryo to adulthood in Landrace pig. Subsequently, due to drastic change in gene expression, the perinatal periods including E105, D0 and D9, were focused, and the genes related to the process of muscle fiber remodeling and volume expansion were revealed. Then, though conjoint analysis with miRNA and lncRNA transcripts, a ceRNA network were identified, which consist of 11 key regulatory genes (such as CHAC1, RTN4IP1 and SESN1), 7 miRNAs and 43 lncRNAs, and they potentially play an important role in the process of muscle fiber differentiation, muscle fiber remodeling and volume expansion, intramuscular fat deposition, and other skeletal muscle developmental events. In summary, we reveal candidate genes and underlying molecular regulatory networks associated with perinatal skeletal muscle fiber type remodeling and expansion. These data provide new insights into the molecular regulation of mammalian skeletal muscle development and diversity.

Trilobatin contributes to the improvement of myopathy in a mouse model of Duchenne muscular dystrophy.

Duchenne muscular dystrophy (DMD) occurs due to genetic mutations that lead to a deficiency in dystrophin production and consequent progressive degeneration of skeletal muscle fibres, through oxidative stress and an exacerbated inflammatory process. The flavonoid trilobatin (TLB) demonstrates antioxidant and anti-inflammatory potential. Its high safety profile and effective action make it a potent therapy for the process of dystrophic muscle myonecrosis. Thus, we sought to investigate the action of TLB on damage in a DMD model, the mdx mouse. Eight-week-old male animals were treated with 160 mg/kg/day of trilobatin for 8 weeks. Control animals were treated with saline. Following treatment, muscle strength, serum creatine kinase (CK) levels, histopathology (necrotic myofibres, regenerated fibres/central nuclei, Feret's diameter and inflammatory area) and the levels of catalase and NF-κB (western blotting) of the quadriceps (QUA), diaphragm (DIA) and tibialis anterior (TA) muscles were measured. TLB was able to significantly increase muscle strength and reduce serum CK levels in dystrophic animals. The QUA of mdx mice showed a reduction in catalase and the number of fibres with a centralized nucleus after treatment with TLB. In the DIA of dystrophic animals, TLB reduced the necrotic myofibres, inflammatory area and NF-κB and increased the number of regenerated fibres and the total fibre diameter. In TA, TLB increased the number of regenerated fibres and reduced catalase levels in these animals. It is concluded that in the mdx experimental model, treatment with TLB was beneficial in the treatment of DMD.

Quantal Properties of Voltage-Dependent Ca2+ Release in Frog Skeletal Muscle Persist After Reduction of [Ca2+] in the Sarcoplasmic Reticulum.

In skeletal muscle, the Ca2+ release flux elicited by a voltage clamp pulse rises to an early peak that inactivates rapidly to a much lower steady level. Using a double pulse protocol the fast inactivation follows an arithmetic rule: if the conditioning depolarization is less than or equal to the test depolarization, then decay (peak minus steady level) in the conditioning release is approximately equal to suppression (unconditioned minus conditioned peak) of the test release. This is due to quantal activation by voltage, analogous to the quantal activation of IP3 receptor channels. Two mechanisms are possible. One is the existence of subsets of channels with different sensitivities to voltage. The other is that the clusters of Ca2+-gated Ryanodine Receptor (RyR) β in the parajunctional terminal cisternae might constitute the quantal units. These Ca2+-gated channels are activated by the release of Ca2+ through the voltage-gated RyR α channels. If the RyR β were at the basis of quantal release, it should be modified by strong inhibition of the primary voltage-gated release. This was attained in two ways, by sarcoplasmic reticulum (SR) Ca2+ depletion and by voltage-dependent inactivation. Both procedures reduced global Ca2+ release flux, but SR Ca2+ depletion reduced the single RyR current as well. The effect of both interventions on the quantal properties of Ca2+ release in frog skeletal muscle fibers were studied under voltage clamp. The quantal properties of release were preserved regardless of the inhibitory maneuver applied. These findings put a limit on the role of the Ca2+-activated component of release in generating quantal activation.

Morphological changes in striated muscle fibres caused by components of the Thiel embalming method.

Thiel-fixed body donors are highly valued for surgical training courses. The pronounced flexibility of Thiel-fixed tissue has been postulated to be caused by histologically visible fragmentation of striated muscle. The aim of this study was to analyse whether a specific ingredient, pH, decay, or autolysis could cause this fragmentation in order to modulate the Thiel solution to adapt specimen flexibility specifically to the needs of different courses. Striated muscle of the mouse was fixed for different time periods in formalin, Thiel solution, and its individual ingredients, and analysed by light microscopy. Further, pH-values of Thiel solution and its ingredients were measured. In addition, unfixed muscle tissue was histologically analysed including Gram staining to investigate a relationship between autolysis, decomposition, and fragmentation. Muscle fixed with Thiel solution for 3 months was slightly more fragmentated than muscle fixed for 1 day. Fragmentation was more pronounced after 1 year of immersion. Three individual salt ingredients showed slight fragmentation. Decay and autolysis had no effect on fragmentation, which occurred regardless of the pH of all solutions. Fragmentation of Thiel-fixed muscle is dependent on fixation time and most likely occurs due to salts present in the Thiel solution. Adjustment of the salt composition in the Thiel solution with verification of the influence on the fixation effect, fragmentation and flexibility of the cadavers could be performed in further studies.

Mustn1 is a smooth muscle cell-secreted microprotein that modulates skeletal muscle extracellular matrix composition

ObjectiveSkeletal muscle plasticity and remodeling are critical for adapting tissue function to use, disuse, and regeneration. The aim of this study was to identify genes and molecular pathways that regulate the transition from atrophy to compensatory hypertrophy or recovery from injury. Here, we have used a mouse model of hindlimb unloading and reloading, which causes skeletal muscle atrophy, and compensatory regeneration and hypertrophy, respectively. MethodsWe analyzed mouse skeletal muscle at the transition from hindlimb unloading to reloading for changes in transcriptome and extracellular fluid proteome. We then used qRT-PCR, immunohistochemistry, and bulk and single-cell RNA sequencing data to determine Mustn1 gene and protein expression, including changes in gene expression in mouse and human skeletal muscle with different challenges such as exercise and muscle injury. We generated Mustn1-deficient genetic mouse models and characterized them in vivo and ex vivo with regard to muscle function and whole-body metabolism. We isolated smooth muscle cells and functionally characterized them, and performed transcriptomics and proteomics analysis of skeletal muscle and aorta of Mustn1-deficient mice. ResultsWe show that Mustn1 (Musculoskeletal embryonic nuclear protein 1, also known as Mustang) is highly expressed in skeletal muscle during the early stages of hindlimb reloading. Mustn1 expression is transiently elevated in mouse and human skeletal muscle in response to intense exercise, resistance exercise, or injury. We find that Mustn1 expression is highest in smooth muscle-rich tissues, followed by skeletal muscle fibers. Muscle from heterozygous Mustn1-deficient mice exhibit differences in gene expression related to extracellular matrix and cell adhesion, compared to wild-type littermates. Mustn1-deficient mice have normal muscle and aorta function and whole-body glucose metabolism. We show that Mustn1 is secreted from smooth muscle cells, and that it is present in arterioles of the muscle microvasculature and in muscle extracellular fluid, particularly during the hindlimb reloading phase. Proteomics analysis of muscle from Mustn1-deficient mice confirms differences in extracellular matrix composition, and female mice display higher collagen content after chemically induced muscle injury compared to wild-type littermates. ConclusionsWe show that, in addition to its previously reported intracellular localization, Mustn1 is a microprotein secreted from smooth muscle cells into the muscle extracellular space. We explore its role in muscle ECM deposition and remodeling in homeostasis and upon muscle injury. The role of Mustn1 in fibrosis and immune infiltration upon muscle injury and dystrophies remains to be investigated, as does its potential for therapeutic interventions.