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KEY POINTS Least shrews were studied to explore the relationship between metabolic function, mitochondrial morphology and protein content in different tissues. Liver and kidney mitochondrial content and enzymatic activity approaches the heart indicating similar metabolic demand among tissues that contribute to basal and maximum metabolism. This allows examination of mitochondrial structure and composition in tissues with similar maximum metabolic demands. Mitochondrial networks only occur in striated muscle. In contrast, the liver and kidney maintain individual mitochondria with limited reticulation. Muscle mitochondrial reticulation is the result of dense ATPase activity and cell-spanning myofibrils which require networking for adequate metabolic support. In contrast, liver and kidney ATPase activity is localized to the endoplasmic reticulum and basolateral membrane respectively, generating a locally balanced energy conversion and utilization Mitochondrial morphology is not driven by maximum metabolic demand, but by the cytosolic distribution of energy utilizing systems set by the functions of the tissue. ABSTRACT Mitochondrial adaptations are fundamental to differentiated function and energetic homeostasis in mammalian cells. But the mechanisms that underlie these relationships remain poorly understood. Here, we investigated organ-specific mitochondrial morphology, connectivity and protein composition in a model of extreme mammalian metabolism, the Least shrew (Cryptotis parva). This was achieved through a combination of high-resolution 3D focused-ion-beam EM imaging and tandem-mass-tag MS proteomics. We demonstrate that liver and kidney mitochondrial content are equivalent to the heart permitting assessment of mitochondrial adaptations in different organs with similar metabolic demand. Muscle mitochondrial networks (cardiac and skeletal) are extensive, with a high incidence of nanotunnels - which collectively support the metabolism of large muscle cells. Mitochondrial networks were not detected in the liver and kidney as individual mitochondria are localized with sites of ATP consumption. This configuration is not observed in striated muscle, likely due to a homogenous ATPase distribution and the structural requirements of contraction. These results demonstrate distinct, fundamental mitochondrial structural adaptations for similar metabolic demand that are dependent on the topology of energy utilization process in a mammalian model of extreme metabolism. Abstract figure. This study investigates the role of mitochondrial morphology and protein composition in setting the extreme metabolic rates of one of the smallest extant mammals - the North American least shrew (Cryptotis parva). To do this, mitochondrial characteristics from liver, kidney, skeletal muscle and heart tissues were compared as these tissues are major contributors to basal and maximum metabolic states. Liver and kidney mitochondrial volume density and protein content approach levels observed in the heart - indicating that these former tissues are major contributors to the high basal metabolic rates of small mammals. Despite this high mitochondrial content, the liver and kidney do not exhibit mitochondrial networking - structures that are proposed to conduct mitochondrial proton motive force at the scale of the cell. Shrew skeletal muscle and cardiac mitochondrial network organization is consistent with networks observed in larger mammals while also exhibiting increased connectivity at the nm-scale. Instead of forming networks, kidney and liver mitochondria are directly associated with sites of ATP utilization. These results identify conditions that dictate the formation of mitochondrial networks and processes that drive mammalian allometric scaling of metabolic rates. This article is protected by copyright. All rights reserved.

Fish myofibrillar protein is underutilized due to the formation of insoluble aggregates in low salt media. High pressure homogenization (HPH) at 20, 40, and 60 MPa for four passes was applied on bighead carp myofibrillar protein in order to modify its structure and interfacial properties. Changes in aggregation, conformation, solubility, emulsifying and foaming properties of myofibrillar protein were investigated. The aggregates of myofibrillar protein were obviously disrupted by HPH treatment. The size of myofibrillar protein aggregates became smaller and more uniform as the treating pressure increased, accompanied by notable decreases of cross-sectional height and Rq value in AFM image. Furthermore, the conformation of HPH-treated myofibrillar protein was unfolded into a flexible and open structure. α-helix and β-sheet were converted into β-turn and random coil. Surface hydrophobicity and zeta potential were strengthened, along with the exposure of sulfhydryl groups onto molecule surface. On the other hand, solubility, emulsifying activity index (EAI) and foaming capacity (FC) of HPH-treated myofibrillar protein were markedly enhanced with the increasing pressure. Especially after HPH treatment at 60 MPa, myofibrillar protein was almost dissolved in low salt media (solubility 91.86%) with 4.92 fold for EAI and 3.52 fold for FC. But there was little variation in emulsifying and foaming stabilities. These results suggested that HPH treatment has interesting potential to induce the dissociation and unfolding of myofibrillar protein in low salt media, therefore improving its interfacial properties. PRACTICAL APPLICATION: Carp myofibrillar protein was treated by high pressure homogenization (HPH). Aggregates of myofibrillar protein were disrupted into smaller size form. Conformation of myofibrillar protein was unfolded into open and loose structure. Emulsifying and foaming capacities of myofibrillar protein were improved. HPH treatment modified the structure and interfacial properties of myofibrillar protein.

BACKGROUND The abnormal ultimate pH (pHu ) in postmortem muscles affect the meat quality and results in substantial economic losses. Dark, firm, and dry (DFD) meat linked with the higher postmortem pHu values and exhibited many quality issues such as dark color, tough texture and shorter shelf life. This research aimed to investigate the effect of protein phosphorylation on variations in beef pHu in order to explore the possible mechanisms underlying DFD meat formation. RESULTS Glycogen and lactate contents were higher, while L* and a* were lower in high pHu beef. Shear force was higher in intermediate pHu group. Global phosphorylation of sarcoplasmic proteins was higher in low pHu samples on day 1 and of myofibrillar proteins was higher in intermediate pHu meat on days 1 and 2 postmortem. Sarcoplasmic protein bands with different phosphorylation levels were identified as containing some glycometabolism and stress response proteins and phosphorylated myofibrillar protein bands were identified sarcomeric and metabolic proteins. CONCLUSIONS Phosphorylation of multiple proteins of glycolytic pathway and contractile machinery may play critical roles in development of DFD beef. © 2021 Society of Chemical Industry.

BACKGROUND Quite recently, considerable attention has been paid to atmospheric cold plasma (ACP) as an eco-friendly and highly efficient technology to modify the functional properties of foods. This study focuses on the effect of ACP on the myofibril protein and lipid quality of hairtail (Trichiurus lepturus) fish. In achieving this, the samples were treated with ACP at 50 kV for different times (30, 60, 120, 180, 240, 300 s). RESULTS The findings indicated slight changes in peroxide value and thiobarbituric acid reactive substances in the samples treated with ACP. A significant increase (P

NEW FINDINGS What is the main observation in this case? The main observation of this case report is that blood flow-restricted exercise can cause myofibrils to have an aberrant wave-like appearance that is accompanied by irregular pockets of sarcoplasm in the intermyofibrillar space, while traditional forms of damage to the Z-discs and contractile elements are not as apparent. What insights does it reveal? Our findings indicate that blood flow restriction-mediated fluid pooling might cause alterations in skeletal muscle ultrastructure after exercise that might be directly related to myofibre swelling. ABSTRACT The acute effects of blood flow-restricted (BFR) exercise training on skeletal muscle ultrastructure are poorly understood owing to inconsistent findings and the use of largely imprecise systemic markers for indications of muscle damage. The purpose of this study was to compare myofibrillar ultrastructure before and 30 min after normal and BFR resistance exercise using transmission electron microscopy in a single individual to evaluate the feasibility of this more nuanced approach. One apparently healthy male with 13 years of resistance exercise completed six sets of both BFR [30% of one-repetition maximum (1-RM)] and normal non-occluded (70% of 1-RM) unilateral angled leg press on the contralateral leg, as a control, after assessment of 1-RM 72 h before. Vastus lateralis muscle biopsies were collected before and 30 min after each exercise session. The lengths and widths of 250 sarcomeres and the sarcoplasmic area were assessed via 20 individual transmission electron photomicrographs. Analysis revealed that BFR training (1.769 ± 0.12 μm) increased sarcomere length when compared with normal exercise (1.64 ± 0.17 μm; P

Key points The ingestion of protein potentiates the stimulation of myofibrillar protein synthesis rates after an acute bout of resistance exercise. Protein supplementation (eating above the protein Recommended Dietary Allowance) during resistance training has been shown to maximize lean mass and strength gains in healthy young and older adults. Here, we assessed contractile, oxidative, and structural protein synthesis in skeletal muscle in response to a moderate or higher protein diet during the early adaptive phase of resistance training in middle-aged adults. We report that the stimulation of myofibrillar, mitochondrial, or collagen protein synthesis rates during 0-3 weeks of resistance training is not further enhanced by a higher protein diet. These results show that moderate protein diets are sufficient to support the skeletal muscle adaptive response during the early phase of a resistance training program. Abstract Protein ingestion augments muscle protein synthesis (MPS) rates acutely after resistance exercise and can offset age-related loss in muscle mass. Skeletal muscle contains a variety of protein pools, such as myofibrillar (contractile), mitochondrial (substrate oxidation), and collagen (structural support) proteins, and the sensitivity to nutrition and exercise seems to be dependent on the major protein fraction studied. However, it is unknown how free-living conditions of dietary protein density with habitual resistance exercise mediates muscle protein subfraction synthesis. Therefore, we investigated the effect of moderate (MOD: 1.06 ± 0.22 g·kg-1 ·d-1 ) or high (HIGH: 1.55 ± 0.25 g·kg-1 ·d-1 ) protein intake on daily MPS rates within the myofibrillar (MyoPS), mitochondrial (MitoPS), and collagen (CPS) protein fractions in middle-aged men and women (n = 20, 47 ± 1 y, BMI 28 ± 1 kg·m-2 ) during the early phase (0-3 wks) of a dietary counseling-controlled resistance training program. Participants were loaded with deuterated water, followed by daily maintenance doses throughout the intervention. Muscle biopsies were collected at baseline and after weeks 1, 2, and 3. MyoPS in the HIGH condition remained constant (P = 1.000), but MOD decreased over time (P = 0.023). MitoPS decreased after 0-3 wks when compared to 0-1 wks (P = 0.010) with no effects of protein intake (P = 0.827). A similar decline with no difference between groups (P = 0.323) was also observed for CPS (P = 0.007). Our results demonstrated that additional protein intake above moderate amounts does not potentiate the stimulation of longer-term MPS responses during the early stage of resistance training adaptations in middle-aged adults. This article is protected by copyright. All rights reserved.

In the three-step myofibrillogenesis model, mature myofibrils are formed through two intermediate structures: premyofibrils and nascent myofibrils. We have recently reported that several inhibitors of the Ubiquitin Proteosome System, for example, MG-132, and DBeQ, reversibly block progression of nascent myofibrils to mature myofibrils. In this investigation, we studied the effects of MG132 and DBeQ on the expression of various myofibrillar proteins including actin, myosin light and heavy chains, tropomyosin, myomesin, and myosin binding protein-C in cultured embryonic quail myotubes by western blotting using two loading controls-α-tubulin and glyceraldehyde 3-phosphate dehydrogenase (GAPDH). Surprisingly, we found that MG-132 affected the level of expression of GAPDH but DBeQ did not. Reverse transcription polymerase chain reaction (RT-PCR) and quantitative reverse transcription-PCR (qRT-PCR) showed no significant effect of MG-132 on GAPDH transcription. Two-dimensional (2D) western blot analyses with extracts of control and MG-132-treated cells using anti-ubiquitin antibody indicated that MG132-treated myotubes show a stronger emitter-coupled logic signal. However, Spot% and Spot volume calculations for all spots from both western blot film signals and matched Coomassie-stained 2D polyacrylamide gel electrophoresis showed that the intensity of staining in a spot of ~39 kDa protein is 3.5-fold lower in the gel of MG-132-treated extracts. Mass spectrometry analyses identified the ~39 kDa protein as quail GAPDH. Immunohistochemical analysis of fixed MG-132-treated myotubes with anti-GAPDH antibody showed extensive clump formation, which may be analogous to granule formation by stress response factors in MG132-treated cells. This is the first report on in vivo ubiquitination of GAPDH. This may be essential for the moonlighting (Jeffery, 1999) activity of GAPDH for tailoring stress in myotubes.

This study investigates the influence of 24‐hr marination (with different plant extracts and vinegar) at refrigerated conditions on commonly evaluated physicochemical and textural attributes, including pH, water‐holding capacity (WHC), collagen solubility, moisture, drip loss, and shear force values of beef meat. The results reflected the appropriate correlation between each pair and indicated the efficiency of the household marination procedure to acquire more palatability and tender beef meat. Therefore, to predict beef meat tenderness by applying the Warner–Bratzler shear force (WBSF), a strong positive correlation with the drip loss (p, The results reflected the appropriate correlation between each pair and indicated the efficiency of the household marination procedure to acquire more palatability and tender beef meat. These outcomes of this study were a good tenderness predictor be utilized in retailing and industrial scale.

Passive skeletal muscle mechanical properties of whole muscle are not as well understood as muscle's active mechanical properties. Both the structural basis for passive mechanical properties and the properties themselves are challenging to determine because it is not clear which structures within skeletal muscle actually bear passive loads and there are not established standards by which to make mechanical measurements. Evidence suggests that titin bears the majority of the passive load within the single muscle cell. However, at larger scales, such as fascicles and muscles, there is emerging evidence that the extracellular matrix (ECM) bears the majority of the load. Complicating the ability to quantify and compare across size scales, muscles, and species, definitions of muscle passive properties such as stress, strain, modulus and stiffness can be made relative to many reference parameters. These uncertainties make a full understanding of whole muscle passive mechanical properties and modeling these properties very difficult. Future studies defining the specific load bearing structures and their composition and organization are required to fully understand passive mechanics of the whole muscle and develop therapies to treat disorders in which passive muscle properties are altered such as muscular dystrophy, traumatic laceration, and contracture due to upper motor neuron lesion as seen in spinal cord injury, stroke and cerebral palsy. Abstract figure legend Schematic representation of passive load bearing at various skeletal muscle scales. In the sarcomere (upper left), load is borne by the giant elastic protein titin and, to a lesser extent, desmin. The myofilaments actin and myosin are responsible for active force production. Sarcomeres in series (myofibrils) connect to the fiber surface at specialized focal adhesions called costameres at which desmin and other cytoskeletal proteins converge (upper right). Muscle fibers are embedded in a connective tissue matrix (lower left) composed primarily of basal lamina collagen type IV. At larger scales, passive load is borne in the perimysium (lower right) by perimysial cables that seem to be composed of collagen types 1 and 3 and this is where the majority of passive load is borne. This article is protected by copyright. All rights reserved.

In this study, we analyze radiation images of muscle structure of mice soleus muscles using radiation source-based microtomography and nanotomography. Soleus muscle samples were collected for analysis from 8-week-old male Institute of Cancer Research mice. First, phase-contrast X-ray microtomography was employed in these experiments. Then to obtain images with excellent contrast, imaging was performed using monochromatic light with excellent transmission power. To analyze additional muscle structures in higher magnification images than these images, nanotomography was performed, which facilitated obtaining high-magnification and high-resolution images. Muscle tissue microstructures were confirmed through three-dimensional images obtained from phase-contrast X-ray microtomography. Thus, the muscle tissue's overall shape at microscopic level can be captured. Additionally, a single muscle fiber was examined using hard X-ray nano-imaging, through which we could observe the alignment of countless myofibrils, that is, actin and myosin filaments in the muscle fibers. Thus, the methodology adopted here proved to be advantageous in analyzing the muscle tissue's overall structure with microtomography and in observing the myofibrils in detail using nanotomography.

This study evaluated the effect of air thawing (AT), water thawing (WT), and ultrasound-assisted immersion thawing (UT) at different power levels (200, 300, 400, and 500 W) on the thawing rate, physicochemical properties, and protein structure of chicken breast muscle (pectoralis), and the weight of each sample was approximately 106 ± 3 g. UT shortened the total thawing time and decreased the cutting force with increasing ultrasound power. Additionally, UT at 300 W (UT-300) remarkably reduced the thawing and cooking losses of the samples compared to AT, WT, and other UT powers (p

BACKGROUND: Prolonged critically ill patients frequently develop debilitating muscle weakness that can affect both peripheral nerves and skeletal muscle. In-depth knowledge on the temporal contribution of neural and muscular components to muscle weakness is currently incomplete. METHODS: We used a fluid-resuscitated, antibiotic-treated, parenterally fed murine model of prolonged (5 days) sepsis-induced muscle weakness (caecal ligation and puncture; n = 148). Electromyography (EMG) measurements were performed in two nerve-muscle complexes, combined with histological analysis of neuromuscular junction denervation, axonal degeneration, and demyelination. In situ muscle force measurements distinguished neural from muscular contribution to reduced muscle force generation. In myofibres, imaging and biomechanics were combined to evaluate myofibrillar contractile calcium sensitivity, sarcomere organization, and fibre structural properties. Myosin and actin protein content and titin gene expression were measured on the whole muscle. RESULTS: Five days of sepsis resulted in increased EMG latency (P = 0.006) and decreased EMG amplitude (P

Myomesin‐1 (encoded by MYOM1 gene) is expressed in almost all cross‐striated muscles, whose family (together with myomesin‐2 and myomesin‐3) helps to cross‐link adjacent myosin to form the M‐line in myofibrils. However, little is known about its biological function, causal relationship and mechanisms underlying the MYOM1‐related myopathies (especially in the heart). Regrettably, there is no MYMO1 knockout model for its study so far. A better and further understanding of MYOM1 biology is urgently needed. Here, we used CRISPR/Cas9 gene‐editing technology to establish an MYOM1 knockout human embryonic stem cell line (MYOM1−/− hESC), which was then differentiated into myomesin‐1 deficient cardiomyocytes (MYOM1−/− hESC‐CMs) in vitro. We found that myomesin‐1 plays an important role in sarcomere assembly, contractility regulation and cardiomyocytes development. Moreover, myomesin‐1‐deficient hESC‐CMs can recapitulate myocardial atrophy phenotype in vitro. Based on this model, not only the biological function of MYOM1, but also the aetiology, pathogenesis, and potential treatments of myocardial atrophy caused by myomesin‐1 deficiency can be studied.

Background Poor recovery from periods of disuse accelerates age‐related muscle loss, predisposing individuals to the development of secondary adverse health outcomes. Exercise prior to disuse (prehabilitation) may prevent muscle deterioration during subsequent unloading. The present study aimed to investigate the effect of short‐term resistance exercise training (RET) prehabilitation on muscle morphology and regulatory mechanisms during 5 days of bed rest in older men. Methods Ten healthy older men aged 65–80 years underwent four bouts of high‐volume unilateral leg RET over 7 days prior to 5 days of inpatient bed rest. Physical activity and step‐count were monitored over the course of RET prehabilitation and bed rest, whilst dietary intake was recorded throughout. Prior to and following bed rest, quadriceps cross‐sectional area (CSA), and hormone/lipid profiles were determined. Serial muscle biopsies and dual‐stable isotope tracers were used to determine integrated myofibrillar protein synthesis (iMyoPS) over RET prehabilitation and bed rest phases, and acute postabsorptive and postprandial myofibrillar protein synthesis (aMyoPS) rates at the end of bed rest. Results During bed rest, daily step‐count and light and moderate physical activity time decreased, whilst sedentary time increased when compared with habitual levels (P

De novo assembly of myofibrils in vertebrate cross-striated muscles progresses in three distinct steps, first from a minisarcomeric alignment of several nonmuscle and muscle proteins in premyofibrils, followed by insertions of additional proteins and increased organization in nascent myofibrils, ending with mature contractile myofibrils. In a search for controls of the process of myofibril assembly, we discovered that the transition from nascent to mature myofibrils could be halted by inhibitors of three distinct functions of the ubiquitin proteasome system (UPS). First, inhibition of pathway to E3 Cullin ligases that ubiquitinate proteins led to an arrest of myofibrillogenesis at the nascent myofibril stage. Second, inhibition of p97 protein extractions of ubiquitinated proteins led to a similar arrest of myofibrillogenesis at the nascent myofibril stage. Third, inhibitors of proteolytic action by proteasomes also blocked nascent myofibrils from transitioning to mature myofibrils. In contrast, inhibitors of autophagy or lysosomes did not affect myofibrillogenesis. To probe for differences in the effects of UPS inhibitors during myofibrillogenesis, we analyzed by fluorescence recovery after photobleaching the exchange rates of two selected sarcomeric proteins (muscle myosin II heavy chains and light chains). In the presence of p97 and proteasomal inhibitors, the dynamics of each of these two myosin proteins decreased in the nascent myofibril stage, but were unaffected in the mature myofibril stage. The increased stability of myofibrils occurring in the transition from nascent to mature myofibril assembly indicates the importance of dynamics and selective destruction in the muscle myosin II proteins for the remodeling of nascent to mature myofibrils.

The coordinated generation of mechanical forces by cardiac myocytes is required for proper heart function. Myofibrils are the functional contractile units of force production within individual cardiac myocytes. At the molecular level, myosin motors form cross-bridges with actin filaments and use ATP to convert chemical energy into mechanical forces. The energetic efficiency of the cross-bridge cycle is influenced by the viscous damping of myofibril contraction. The viscoelastic response of myofibrils is an emergent property of their individual mechanical components. Previous studies have implicated titin-actin interactions, cell-ECM adhesion, and microtubules as regulators of the viscoelastic response of myofibrils. Here we probed the viscoelastic response of myofibrils using laser-assisted dissection. As a proof-of-concept, we found actomyosin contractility was required to endow myofibrils with their viscoelastic response, with blebbistatin treatment resulting in decreased myofibril tension and viscous damping. Focal adhesion kinase (FAK) is a key regulator of cell-ECM adhesion, microtubule stability, and myofibril assembly. We found inhibition of FAK signaling altered the viscoelastic properties of myofibrils. Specifically, inhibition of FAK resulted in increased viscous damping of myofibril retraction following laser ablation. This damping was not associated with acute changes in the electrophysiological properties of cardiac myocytes. These results implicate FAK as a regulator of mechanical properties of myofibrils.

This study examined the effects on conformation and oxidation of myofibrillar protein in largemouth bass by different thawing methods. The conventional thawing, microwave thawing, microwave (MVT) or ultrasound combined with vacuum thawing, microwave or far-infrared thawing (FMT) combined with magnetic nanoparticles were used in this experiment. The physicochemical changes were analyzed by differential scanning calorimetry and dynamic rheology. The protein structure changes were measured by Raman, intrinsic fluorescence, and second-derivative ultraviolet spectrometry. The degree of protein aggregation was evaluated by surface hydrophobicity, particle size, and zeta-potential measurements. Total sulfhydryl content, protein carbonyl content, Ca2+ -ATPase activity, and SDS-PAGE were used to analyze the degree of protein oxidation. Results showed that MVT and FMT samples had better thermal stability, more stable protein conformation, and a lower degree of protein oxidation. Thus, these two methods would be beneficial to sustain the quality of thawed fillets. PRACTICAL APPLICATIONS: In the market circulation, largemouth bass (Micropterus salmoides) need to be frozen. The thawing methods can directly affect the quality of frozen fish, thus causing the changes in the conformation of the myofibrillar protein in fish, and also affecting the degree of protein oxidation. The results showed that the microwave combined with vacuum and the magnetic nanoparticles combined with far-infrared thawing had less effect on myofibrillar protein of fish and were a better thawing method.

In skeletal muscle fibers, mitochondria are densely packed adjacent to myofibrils because adenosine triphosphate (ATP) is needed to fuel sarcomere shortening. However, despite this close physical and biochemical relationship, the effects of mitochondrial dynamics on skeletal muscle contractility are poorly understood. In this study, we analyzed the effects of Mitochondrial Division Inhibitor 1 (mdivi-1), an inhibitor of mitochondrial fission, on the structure and function of both mitochondria and myofibrils in skeletal muscle tissues engineered on micromolded gelatin hydrogels. Treatment with mdivi-1 did not alter myotube morphology, but did increase the mitochondrial turbidity and oxidative capacity, consistent with reduced mitochondrial fission. Mdivi-1 also significantly increased basal, twitch, and tetanus stresses, as measured using the Muscular Thin Film (MTF) assay. Finally, mdivi-1 increased sarcomere length, potentially due to mdivi-1-induced changes in mitochondrial volume and compression of myofibrils. Together, these results suggest that mdivi-1 increases contractile stress generation, which may be caused by an increase in maximal respiration and/or sarcomere length due to increased volume of individual mitochondria. These data reinforce that mitochondria have both biochemical and biomechanical roles in skeletal muscle and that mitochondrial dynamics can be manipulated to alter muscle contractility.

Aims: Both left ventricular (LV) and left atrial (LA) dysfunction and remodelling contribute to adverse outcomes in heart failure with reduced ejection fraction (HFrEF). Danicamtiv is a novel, cardiac myosin activator that enhances cardiomyocyte contraction.Methods and results: We studied the effects of danicamtiv on LV and LA function in non-clinical studies (ex vivo: skinned muscle fibres and myofibrils; in vivo: dogs with heart failure) and in a randomized, double-blind, single- and multiple-dose phase 2a trial in patients with stable HFrEF (placebo, n = 10; danicamtiv, n = 30; 50–100 mg twice daily for 7 days). Danicamtiv increased ATPase activity and calcium sensitivity in LV and LA myofibrils/muscle fibres. In dogs with heart failure, danicamtiv improved LV stroke volume (+10.6 mL, P < 0.05) and LA emptying fraction (+10.7%, P < 0.05). In patients with HFrEF (mean age 60 years, 25% women, ischaemic heart disease 48%, mean LV ejection fraction 32%), treatment-emergent adverse events, mostly mild, were reported in 17 patients (57%) receiving danicamtiv and 4 patients (40%) receiving placebo. Danicamtiv (at plasma concentrations ≥2000 ng/mL) increased stroke volume (up to +7.8 mL, P < 0.01), improved global longitudinal (up to −1.0%, P < 0.05) and circumferential strain (up to −3.3%, P < 0.01), decreased LA minimal volume index (up to −2.4 mL/m2, P < 0.01) and increased LA function index (up to 6.1, P < 0.01), when compared with placebo.Conclusions: Danicamtiv was well tolerated and improved LV systolic function in patients with HFrEF. A marked improvement in LA volume and function was also observed in patients with HFrEF, consistent with pre-clinical findings of direct activation of LA contractility.

BACKGROUND Sturgeon is popular for its nutritious value and its taste. However, sturgeon fillets are traditionally heated in 100 °C boiling water, resulting in unfavorable taste and with a negative effect on the quality. This study considered the effect of combinations of vacuum and low-temperature treatments (LTVH groups) on sturgeon fillets compared with the traditional heat treatment (TC groups). RESULTS The results show that the LTVH groups had lower cooking-loss rates. All LTVH fillets were changed to a white color, and appeared 'done', as did the TC fillets. The LTVH and TC methods gave rise to significant differences in texture: the springiness of the LTVH groups decreased with heating time, and decreased rapidly in the TC groups (P

Slice shear force (SSF) and laser diffraction, considered faster methodologies, for measuring beef instrumental tenderness and sarcomere length, were compared with reference methodologies Warner-Bratzler shear force (WBSF) and phase contrast microscopy. Striploin samples (n = 74) were analyzed for pH, sarcomere length, instrumental tenderness, myofibrillar fragmentation index, and sensorial tenderness. Pearson's correlation measured the association of meat evaluation methods with residual analysis of the multivariate analysis of variance model. The n-dimensional profile to evaluated methods was presented by biplot to identify the behavior of the correlation between the methods (variables). There was moderate correlation between SSF and WBSF (r = .63; p

The main mammalian heart pacemakers are spindle‐shaped cells compressed into tangles within protective layers of collagen in the sino‐atrial node (SAN). Two cell types, “dark” and “light,” differ on their high or low content of intermediate filaments, but share scarcity of myofibrils and a high content of glycogen. Sarcoplasmic reticulum (SR) is scarce. The free SR (fSR) occupies 0.04% of the cell volume within ~0.4 µm wide peripheral band. The junctional SR (jSR), constituting peripheral couplings (PCs), occupies 0.03% of the cell volume. Total fSR + jSR volume is 0.07% of cell volume, lower than the SR content of ventricular myocytes. The average distance between PCs is 7.6 µm along the periphery. On the average, 30% of the SAN cells surfaces is in close proximity to others. Identifiable gap junctions are extremely rare, but small sites of close membrane‐to‐membrane contacts are observed. Possibly communication occurs via these very small sites of contact if conducting channels (connexons) are located within them. There is no obvious anatomical detail that might support ephaptic coupling. These observations have implications for understanding of SAN cell physiology, and require incorporation into biophysically detailed models of SAN cell behavior that currently do not include such features.

Myosin, the most abundant myofibrillar protein in skeletal muscle, functions as a motor protein in muscle contraction. Myosin polymerizes into the thick filaments in the sarcomere where approximately 50% of embryonic myosin (Myh3) are replaced within 3 h (Ojima K, Ichimura E, Yasukawa Y, Wakamatsu J, Nishimura T, Am J Physiol Cell Physiol 309: C669‐C679, 2015). The sarcomere structure including the thick filament is maintained by a balance between protein biosynthesis and degradation. However, the involvement of a protein degradation system in the myosin replacement process remains unclear. Here, we show that the muscle‐specific ubiquitin ligase Ozz regulates replacement rate of Myh3. To examine the direct effect of Ozz on myosin replacement, eGFP‐Myh3 replacement rate was measured in myotubes overexpressing Ozz by fluorescence recovery after photobleaching. Ozz overexpression significantly decreased the replacement rate of eGFP‐Myh3 in the myofibrils, whereas it had no effect on other myosin isoforms. It is likely that ectopic Ozz promoted myosin degradation through increment of ubiquitinated myosin, and decreased myosin supply for replacement, thereby reducing myosin replacement rate. Intriguingly, treatment with a proteasome inhibitor MG132 also decreased myosin replacement rate, although MG132 enhanced the accumulation of ubiquitinated myosin in the cytosol where replaceable myosin is pooled, suggesting that ubiquitinated myosin is not replaced by myosin in the myofibril. Collectively, our findings showed that Myh3 replacement rate was reduced in the presence of overexpressed Ozz probably through enhanced ubiquitination and degradation of Myh3 by Ozz., In our model, ubiquitination of embryonic myosin (Myh3) is enhanced in the cytosol of Ozz‐overexpressed myotubes. Ubiquitinated Myh3 is not incorporated into the thick filaments but degraded through the proteasome system, leading to slow Myh3 replacement rate in the thick filaments.

Key points Changes in intramuscular Ca2+ handling contribute to development of fatigue and disease-related loss of muscle mass and function. To date, no data on human intact living muscle fibres have been described. We manually dissected intact single fibres from human intercostal muscle and simultaneously measured force and myoplasmic free [Ca2+ ] at physiological temperature. Based on their fatigue resistance, two distinct groups of fibres were distinguished: fatigue sensitive and fatigue resistant. Force depression in fatigue and during recovery was due to impaired sarcoplasmic reticulum Ca2+ release in both groups of fibres. Acidification did not affect force production in unfatigued fibres and did not affect fatigue development in fatigue-resistant fibres. The current study provides novel insight into the mechanisms of fatigue in human intercostal muscle. Abstract Changes in intracellular Ca2+ handling of individual skeletal muscle fibres cause a force depression following physical activity and are also implicated in disease-related loss of function. The relation of intracellular Ca2+ handling with muscle force production and fatigue tolerance is best studied in intact living single fibres that allow continuous measurements of force and myoplasmic free [Ca2+ ] during repeated contractions. To this end, manual dissections of human intercostal muscle biopsies were performed to isolate intact single fibres. Based on the ability to maintain tetanic force at >40% of the initial value during 500 fatiguing contractions, fibres were classified as either fatigue sensitive or fatigue resistant. Following fatigue all fibres demonstrated a marked reduction in sarcoplasmic reticulum Ca2+ release, while myofibrillar Ca2+ sensitivity was either unaltered or increased. In unfatigued fibres, acidosis caused a reduction in myofibrillar Ca2+ sensitivity that was offset by increased tetanic myoplasmic free [Ca2+ ] so that force remained unaffected. Acidification did not affect the fatigue tolerance of fatigue-resistant fibres, whereas uncertainties remain whether or not fatigue-sensitive fibres were affected. Following fatigue, a prolonged force depression at preferentially low-frequency stimulation was evident in fatigue-sensitive fibres and this was caused exclusively by an impaired sarcoplasmic reticulum Ca2+ release. We conclude that impaired sarcoplasmic reticulum Ca2+ release is the predominant mechanism of force depression both in the development of, and recovery from, fatigue in human intercostal muscle.

This study aimed to explore the effect exerted by oxidative stress on apoptosis-inducing factors (AIF)-mediated apoptosis and bovine muscle tenderness during postmortem aging. We investigated the reactive oxygen species (ROS) content, mitochondrial membrane permeability, AIF expression level, nucleus apoptosis, shear force, myofibril fragmentation index, pH, and energy level. According to the results, a rise in ROS content was accompanied by the rise in mitochondrial membrane permeability from 6 to 72 hr. In the meantime, the AIF expression in mitochondria was downregulated significantly within 72 hr. However, samples treated with N-acetylcysteine had significantly lower ROS content (6 to 72 hr) and mitochondrial membrane permeability (12 to 72 hr) than the control group. Moreover, during postmortem aging, the variations in AIF levels in mitochondria were closely associated with meat tenderization and nucleus apoptosis. These findings demonstrated that oxidative stress induced by ROS significantly promoted AIF release from mitochondria by enhancing the mitochondrial membrane permeability, and the released AIF mediated nucleus apoptosis that further enhanced bovine muscle tenderness. Besides, results suggest that in the early stage, the environmental factors (ATP content and pH) significantly decreased (0 to 72 hr), whereas ROS-induced oxidative stress had no significant effect on environmental factors. These observations further suggested that during postmortem aging, the decrease of pH and ATP consumption are required by AIF release. We conclude that ROS-induced oxidative stress and internal environment are vital for meat tenderization through the regulation of AIF-mediated apoptosis pathway. PRACTICAL APPLICATION: ROS-induced oxidative stress contributes to bovine muscle tenderization by promoting cell apoptosis. It is likely to lay a theoretical foundation for developing innovative tenderization techniques by altering the internal oxidation environment of postmortem muscles.

Meat color is important for consumer acceptability, with excessively dark meat often associated with consumer rejection. It is determined chromatically by pigment content (measured by hue and chroma) and achromatically by scattering of light by the microstructure (measured by lightness), the latter of which has received minimal research focus. This review discusses the individual components of the meat microstructure that cause differences in achromatic contributions to color. Differences in achromatic light scattering between light and dark extremes of meat color are most likely explained by structural attributes within the muscle cell. These differences are proposed to arise from variations in (a) transverse shrinkage of the structural lattice of the myofilaments, myofibrils, and muscles fibers, (b) longitudinal shrinkage of the sarcomere, and (c) different protein composition of the surrounding medium (sarcoplasm and extracellular space). These are discussed at a mechanistic level, in relation to six parameters of the muscle cell: (a) protein surface charge altering the myofilament spacing, (b) protein solubility, (c) sarcoplasmic protein binding to myofilaments and myofibrils, (d) integrity of the cytoskeleton and cell adhesion proteins, (e) sarcomere integrity and myofibrillar proteins, and (f) myosin denaturation and rigor bond modification. New data are presented to support the proposed role of structural elements in muscle causing achromatic light scattering and their contribution to the surface color of meat. In addition, the relationships between lightness and water holding capacity and pH are explored and the economic impact of dark meat for the meat industry is discussed.

BACKGROUND: Composite gels were individually prepared from 20 g kg⁻¹ myofibrillar protein (MP) imbedded with typical native starch (potato, tapioca, rice or corn starch) in 0.6 mol L⁻¹ NaCl at pH 6.2. The gel strength, water holding capacity, rheological properties and microstructure of the obtained myofibrillar protein–starch composite gels were evaluated. RESULTS: Tapioca starch improved (P

BACKGROUND: T‐2 toxin (T‐2) is a potent mycotoxin and a common contaminant of aquatic animal feed, posing a serious risk to health and aquatic animals. We investigated the effect of T‐2 on shrimp muscle proteins using proteomics and conventional biochemical methods. Shrimp were fed a diet containing T‐2 at 0–12.2 mg kg⁻¹ for 20 days, and changes to the muscle protein composition, ATPase activities, and the sulfhydryl (SH) content and hydrophobicity of actomyosin (AM) were determined. A proteomics study of the proteins was conducted with sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS‐PAGE), two‐dimensional (2D) electrophoresis, and matrix‐assisted laser desorption/ionization – time of flight mass spectrometry (MALDI‐TOF/TOF MS). RESULTS: Exposure to T‐2 markedly affected the muscle protein composition of shrimp in a concentration‐responsive manner that displayed a diphasic effect. At a low T‐2 concentration (1.2 mg kg⁻¹), the levels of three major muscle proteins (myofibrillar, sarcoplasmic, and stroma) increased but at higher concentrations they declined progressively. T‐2 exposure also led to a breakdown of muscle proteins as evidenced by increases in alkali‐soluble protein and the surface hydrophobicity (SoANS) of AM. Thirty differentially expressed proteins were detected, 12 of which showed a concentration‐response relationship with T‐2 exposure. Among them, 11 homologous proteins were identified by mass spectrometry (MS), with several being key enzymes in energy metabolism. CONCLUSION: This study demonstrated that T‐2 exposure at medium to high concentrations could significantly affect the protein composition and quality of shrimp muscle, and potentially some of its key metabolisms. One of the arginine kinases (spot 27) was particularly responsive to T‐2 and could potentially be used as a biomarker protein for T‐2 intoxication by shrimp. © 2019 Society of Chemical Industry

The water holding capacity (WHC) of sodium‐reduced (0.3 m sodium chloride, corresponding to the salt percentage (w/v) of 1.755%) myofibrillar protein (MP) gel in response to combined calcium chloride (CaCl₂, 20, 60, 100 mm) and high‐pressure processing (HPP, 200 MPa, 10 min) was investigated. The results showed that 200 MPa + 20 mm CaCl₂ synergistically increased the WHC of MP gel via reducing particle size of MP solutions, strengthening hydrogen‐bonding and disulphide‐bonding, promoting formation of β‐sheet and uncoiling of α‐helix, exposing tryptophan residues, enhancing hydrophobic interactions of aliphatic residues and forming a compact and continuous networked gel structure. However, high concentrations (≥60 mm) of CaCl₂ could attenuate the enhancing effects of HPP on the WHC by inducing decreased hydrogen bonds, fewer tryptophan residues exposed and coarser and aggregated gel structures with large cavities. Therefore, a combined moderate HPP and low concentration of CaCl₂ is a potential alternative for developing sodium‐reduced meat products.

This study was conducted to evaluate the comparative efficacy of actinidin from green and gold kiwifruit extract on in vitro simulated gastrointestinal protein digestion of cooked beef Semitendinosus and its myofibrillar protein fraction. Samples collected during gastrointestinal digestion were analysed for protein profile, protein digestibility (%), soluble protein (%) and free amino acid analysis. The addition of kiwifruit extracts significantly (P

Histological evidence of skeletal muscle myopathy has been reported in ≈40–70% (Martin et al., 1985; Shenkman et al., 2019; Urbano-Marquez et al., 1995) of drinkers with a sustained (e.g., at least 3–4 years) history of alcohol consumption that approximates what has been defined as excessive alcohol consumption (i.e., more than 8 drinks per week for women and 15 drinks per week for men) by the 2015–2020 Dietary Guidelines for Americans (U.S. Department of Health and Human Services and U.S. Department of Agriculture 2015). This alcohol-induced myopathy manifests prominently in fast-twitch type muscles and results, at least in part, from a reduction in protein synthesis [as reviewed in (Kimball and Lang, 2018)]. As a result, both men and women who exhibit excessive drinking behavior as defined above have a reduction of muscle mass and the cross-section area (CSA) of such muscles (Shenkman et al., 2019; Thapaliya et al., 2014; Urbano-Marquez et al., 1995), with proximal muscle weakness being reported in ≈40% of these individuals (Estruch et al., 1998; Urbano-Marquez et al., 1995). This alcohol-induced myopathy in humans appears proportional to the total lifetime dose of alcohol consumed and may not be fully reversed in those who abstain from drinking alcohol (Estruch et al., 1998; Urbano-Marquez et al., 1995). Importantly, in other catabolic disease states, the loss of muscle and the accompanying muscle fatigue is directly proportional to survival (VanderVeen et al., 2018). Preclinical studies have also demonstrated similar changes in muscle mass and CSA in rodent models where alcohol consumption is prolonged (i.e., at least 6 weeks duration), and that this atrophy resulted, at least in part, in a reduction in mTOR (mechanistic target of rapamycin) complex 1 kinase activity that antagonizes basal and nutrient-stimulated synthesis of both myofibrillar and sarcoplasmic proteins (Lang, 2018; Lang et al., 1999; Thapaliya et al., 2014). While there has been considerable research into the mechanisms underlying the alcohol-induced loss of muscle mass (Kimball and Lang, 2018), there is a paucity of information pertaining to the effect of alcohol on skeletal muscle function. Many of the early studies in this regard revealed no or relatively isolated changes in specific parameters of contractility when skeletal muscles from chronic alcohol-fed rats and mice were stimulated via the sciatic nerve (Berk et al., 1975; Edmonds et al., 1987; Martyn and Munsat, 1980). However, in vitro incubation of muscle with ethanol decreased contractile function; although the concentrations used (1.8–2.5 g/dL) were certainly pharmacological and are far in excess of those typically reported in the blood of alcohol-consuming individuals (Martyn and Munsat, 1980; Taylor et al., 1992). Moreover, there are no studies that directly compare the effect of acute alcohol intoxication and chronic consumption on muscle function using the same experimental protocol. Hence, the purpose of the present study was to determine whether acute alcohol intoxication or chronic alcohol consumption decrease the intrinsic contractile function of fast-twitch skeletal muscle, independent of a change in nerve conduction, by assessing isometric and tetanic twitch force under basal conditions and after a fatiguing stimuli. To assess possible mechanisms for the observed alcohol-induced defects in contractile function, we also assessed the content of high-energy phosphates as well as the relative abundance of proteins in oxidative phosphorylation (OXPHOS) complexes 1–4 and selected contractile proteins under basal non-stimulated conditions.

BACKGROUND: Emulsification is important for food quality and processing functionality. Most emulsification occurs under high‐fat conditions that eventually cause health concerns. Protein emulsifiers also have drawbacks such as lower dispersity. This study considered the effects of different high‐speed shear homogenization (HSH) speeds on the emulsifying and structural properties of myofibrillar proteins (MPs) under low‐fat conditions. RESULTS: High‐speed shear homogenization significantly increased the emulsifying activity and emulsifying stability of MPs at lower speeds (8000 to 14 500 rpm). The primary structure of MP was not altered significantly by HSH, whereas its secondary, tertiary, and quaternary structures were changed. Particle size decreased first and then increased significantly, and reached a minimum when the HSH speed was 14 500 rpm. The absolute zeta potential values increased significantly and the dendritic fibrous structure of sample was destroyed when the speed exceeded 14 500 rpm. High‐speed shear homogenization (14 500 rpm) decreased the particle size and unfolded the protein, which improved the emulsifying properties of MPs. Excessive HSH speeds (20 500 rpm or higher) caused an aggregation of MP molecules, which was not conducive to improving their emulsifying properties. CONCLUSION: Optimal HSH speed was achieved at 14 500 rpm to modify MPs' emulsifying and structural properties under low‐fatconditions. © 2019 Society of Chemical Industry

Background Heat treatment induces both structural and digestive change of meat protein. However, little has been revealed regarding the associations between structural changes and digested peptides of myofibrillar proteins. This work investigated the effects of heat treatment on the structures and in vitro digestibility of actomyosin, and the peptidomics of the digests were analyzed using liquid chromatography tandem mass spectrometry (LC-MS/MS). Results Heat treatment resulted in unfolding and aggregation behavior of actomyosin according to the results of surface hydrophobicity and particle size. Formation of disulfide bonds and increase in carbonyl groups that occurred during heat treatment of actomyosin indicated the oxidation of specific residues. Unfolding behavior could elevate digestibility of actomyosin by exposing residues, based on the identification of peptides in digests of actomyosin using LC-MS/MS. However, the disulfide bond proved to reduce the action of digestive proteases, since the peptides number (increased from 56 to 86 in sample heated at 70 °C for 30 min) and peptides intensity in digests largely increased after the addition of dithiothreitol (DTT). Heating at higher temperature (100 °C) induced severer aggregation and oxidation, which resulted in lower digestibility of actomyosin than that heated at 70 °C by burying or damaging partial cleavage sites for digestive proteases. Conclusions This work highlights the huge influence of heat treatment on the multi-scale structures of myofibrillar proteins, which largely changed the peptides composition in protein digests. © 2019 Society of Chemical Industry.

This study aimed to investigate the effect of cold atmospheric plasma (CAP) on the physicochemical and functional properties of myofibrillar protein isolated from Alaska pollock (Theragra chalcogramma) muscle. Free sulfhydryl content, surface hydrophobicity, turbidity, differential scanning calorimetry, SDS‐PAGE, heat‐induced myofibrillar protein gel properties and water holding capacity were determined to evaluate the effect of CAP treatment on protein properties. CAP treatment from 10 to 60 kV for 10 min significantly (P