Myofibrillar Ca2 Research Articles

Effects of intensified training with insufficient recovery on joint level and single muscle fibre mechanical function: the role of myofibrillar Ca2+ sensitivity.

Intense exercise training with insufficient recovery time is associated with reductions in neuromuscular performance. However, it is unclear how single muscle fibre mechanical function and myofibrillar Ca2+ sensitivity contribute to these impairments. We investigated the effects of overload training on joint-level neuromuscular performance and cellular-level mechanical function. Fourteen athletes (4 female and 10 male) underwent a 3-week intensified training protocol consisting of up to 150% of their regular training hours with three additional high-intensity training sessions per week. Neuromuscular performance of the knee extensors was assessed via maximal voluntary contraction (MVC) force, electrically evoked twitch contractions, and a force-frequency relationship. Muscle biopsies were taken from the vastus lateralis to assess single fibre mechanical function. Neither MVC force nor twitch parameters were altered following training (all p>0.05), but a rightward shift in the force-frequency curve was observed with average reduction in force of 6%-27% across frequencies 5-20Hz (all p<0.05). In single fibres, maximal force output was not reduced following training, but there was a rightward shift in the force-pCa curve driven by a 6% reduction in Ca2+ sensitivity (p<0.05). These data indicate intensified training leads to impaired Ca2+ sensitivity at the single fibre level, which in part explains impaired neuromuscular function at the joint level during lower frequencies of activation. This is an important consideration for athletes, as performance is often assessed at maximal levels of activation, and these underlying impairments in force generation may be less obvious.

Piperine enhances contractile force in slow- and fast-twitch muscle.

Piperine has been shown to bind to myosin and shift the distribution of conformational states of myosin molecules from the super-relaxed state to the disordered relaxed state. However, little is known about the implications for muscle force production and potential underlying mechanisms. Muscle contractility experiments were performed using isolated muscles and single fibres from rats and mice. The dose-response effect of piperine on muscle force was assessed at several stimulation frequencies. The potentiation of muscle force was also tested in muscles fatigued by eccentric contractions. Potential mechanisms of force potentiation were assessed by measuring Ca2+ levels during stimulation in enzymatically dissociated muscle fibres, while myofibrillar Ca2+ sensitivity was assessed in chemically skinned muscle fibres. Piperine caused a dose-dependent increase in low-frequency force with no effect on high-frequency force in both slow- and fast-twitch muscle, with similar relative increases in twitch force, rate of force development and relaxation rate. The potentiating effect of piperine on low-frequency force was reversible, and piperine partially recovered low-frequency force in fatigued muscle. Piperine had no effect on myoplasmic free [Ca2+] levels in mouse muscle fibres, whereas piperine substantially augmented the force response to submaximal levels of [Ca2+] in rat MyHCII fibres and MyHCI fibres along with a minor increase in maximum Ca2+-activated force. Piperine enhances low-frequency force production in both fast- and slow-twitch muscle. The effects are reversible and can counteract muscle fatigue. The primary underlying mechanism appears to be an increase in Ca2+ sensitivity. KEY POINTS: Piperine is a plant alkaloid derived from black pepper. It is known to bind to skeletal muscle myosin and enhance resting ATP turnover but its effects on contractility are not well known. We showed for the first time a piperine-induced force potentiation that was pronounced during low-frequency electrical stimulation of isolated muscles. The effect of piperine was observed in both slow and fast muscle types, was reversible, and could counteract the force decrements observed after fatiguing muscle contractions. Piperine treatment caused an increase in myofibrillar Ca2+ sensitivity in chemically skinned muscle fibres, while we observed no effect on intracellular Ca2+ concentrations during electrical stimulation in enzymatically dissociated muscle fibres.

Correlation analysis of phosphorylation of myofibrillar protein and muscle quality of tilapia during storage in ice

In this study, the correlation between protein phosphorylation and deterioration in the quality of tilapia during storage in ice was examined by assessing changes in texture, water-holding capacity (WHC), and biochemical characteristics of myofibrillar protein throughout 7 days of storage. The hardness significantly decreased from 471.50 to 252.17 g, whereas cooking and drip losses significantly increased from 26.5% to 32.6% and 2.9% to 9.1%, respectively (P < 0.05). Myofibril fragmentation increased, while myofibrillar protein sulfhydryl content and Ca2+-ATPase activity decreased from 119.33 to 89.29 μmol/g prot and 0.85 to 0.46 μmolPi/mg prot/h, respectively (P < 0.05). Correlation analysis revealed that the myofibrillar protein phosphorylation level was positively correlated with hardness and Ca2+-ATPase activity but negatively correlated with WHC. Myofibrillar protein phosphorylation affects muscle contraction by influencing the dissociation of actomyosin, thereby regulating hardness and WHC. This study provides novel insights for the establishment of quality control strategies for tilapia storage based on protein phosphorylation.

Effects of Adjuvant Exercise and Nutrition Therapy on Muscle Fibre Biomechanics in Gastrointestinal Cancer Patients.

Patients with aggressive cancer, e.g., gastrointestinal cancer, are prone (≥50% chance) to developing cancer cachexia (CC). Little is known about the effects of CC on the biomechanical function of muscle. A promising prevention strategy was found in the form of a multi-modal therapy combining mild resistance exercise (e.g., whole-body electro-myostimulation, WB-EMS) and a protein-rich diet. In a previous study of ours, this was effective in counteracting the loss of muscle mass, yet a systematic and comprehensive assessment of active and passive single muscle fibre functions was so far absent. This pilot study investigated the biomechanical function of single muscle fibres (rectus abdominis) from the biopsies of conventionally treated (pre-)cachectic cancer ((pre-)CC) patients (m = 9), those receiving the multi-modal therapy comprising WB-EMS training and protein-rich nutrition (m = 3), and a control group (m = 5). Our findings not only align with previous findings showing the absolute force loss in CC that is accelerated by atrophy but also speak in favour of a different, potentially energy- and Ca2+-homeostasis-related effect that compromises muscle contraction (F ~0.9 mN vs. F ~0.6 mN in control patients). However, myofibrillar Ca2+ sensitivity and the quality of contraction were unaltered (pCa50: 5.6-5.8). Single fibres from the (pre-)CC patients receiving WB-EMS training and protein supplementation were significantly more compliant (p < 0.001 at ≥130% of resting length L0). Those fibres displayed a similar softness to the ones from the control patients (axial compliance ~15 m/N at ≥130% L0), while single fibres from the patients with (developing) cachexia were significantly stiffer (axial compliance ~7 m/N, p < 0.001 at ≥130% L0). Adjuvant multi-modal therapy (WB-EMS training and nutritional support) contributes to maintaining the axial compliance of single fibres and potentially improves the quality of life for patients at risk of developing CC.

Behavior of Hypertrophied Right Ventricle during the Development of Left Ventricular Failure Due to Myocardial Infarction.

In order to determine the behavior of the right ventricle, we have reviewed the existing literature in the area of cardiac remodeling, signal transduction pathways, subcellular mechanisms, β-adrenoreceptor-adenylyl cyclase system and myocardial catecholamine content during the development of left ventricular failure due to myocardial infarction. The right ventricle exhibited adaptive cardiac hypertrophy due to increases in different signal transduction pathways involving the activation of protein kinase C, phospholipase C and protein kinase A systems by elevated levels of vasoactive hormones such as catecholamines and angiotensin II in the circulation at early and moderate stages of heart failure. An increase in the sarcoplasmic reticulum Ca2+ transport without any changes in myofibrillar Ca2+-stimulated ATPase was observed in the right ventricle at early and moderate stages of heart failure. On the other hand, the right ventricle showed maladaptive cardiac hypertrophy at the severe stages of heart failure due to myocardial infarction. The upregulation and downregulation of β-adrenoreceptor-mediated signal transduction pathways were observed in the right ventricle at moderate and late stages of heart failure, respectively. The catalytic activity of adenylate cyclase, as well as the regulation of this enzyme by Gs proteins, were seen to be augmented in the hypertrophied right ventricle at early, moderate and severe stages of heart failure. Furthermore, catecholamine stores and catecholamine uptake in the right ventricle were also affected as a consequence of changes in the sympathetic nervous system at different stages of heart failure. It is suggested that the hypertrophied right ventricle may serve as a compensatory mechanism to the left ventricle during the development of early and moderate stages of heart failure.

Ca2+ and force during dynamic contractions in mouse intact skeletal muscle fibers

Muscle fiber force production is determined by the excitation frequency of motor nerves, which induce transient increases in cytoplasmic free Ca2+ concentration ([Ca2+]i) and the force-generating capacity of the actomyosin cross-bridges. Previous studies suggest that, in addition to altered cross-bridge properties, force changes during dynamic (concentric or eccentric) contraction might be affected by Ca2+-dependent components. Here we investigated this by measuring [Ca2+]i and force in mouse muscle fibers undergoing isometric, concentric, and eccentric contractions. Intact single muscle fibers were dissected from the flexor digitorum brevis muscle of mice. Fibers were electrically activated isometrically at 30–100 Hz and after reaching the isometric force plateau, they were actively shortened or stretched. We calculated the ratio (relative changes) in force and [Ca2+]i attained in submaximal (30 Hz) and near-maximal (100 Hz) contractions under isometric or dynamic conditions. Tetanic [Ca2+]i was similar during isometric, concentric and eccentric phases of contraction at given stimulation frequencies while the forces were clearly different depending on the contraction types. The 30/100 Hz force ratio was significantly lower in the concentric (44.1 ± 20.3%) than in the isometric (50.3 ± 20.4%) condition (p = 0.005), whereas this ratio did not differ between eccentric and isometric conditions (p = 0.186). We conclude that the larger force decrease by decreasing the stimulation frequency during concentric than during isometric contraction is caused by decreased myofibrillar Ca2+ sensitivity, not by the decreased [Ca2+]i.

Effect of insulin insufficiency on ultrastructure and function in skeletal muscle.

Decreased insulin availability and high blood glucose levels, the hallmark features of poorly controlled diabetes, drive disease progression and are associated with decreased skeletal muscle mass. We have shown that mice with β-cell dysfunction and normal insulin sensitivity have decreased skeletal muscle mass. This project asks how insulin deficiency impacts on the structure and function of the remaining skeletal muscle in these animals. Skeletal muscle function was determined by measuring exercise capacity and specific muscle strength prior to and after insulin supplementation for 28days in 12-week-old mice with conditional β-cell deletion of the ATP binding cassette transporters ABCA1 and ABCG1 (β-DKO mice). Abca1 and Abcg1 floxed (fl/fl) mice were used as controls. RNAseq was used to quantify changes in transcripts in soleus and extensor digitorum longus muscles. Skeletal muscle and mitochondrial morphology were assessed by transmission electron microscopy. Myofibrillar Ca2+ sensitivity and maximum isometric single muscle fibre force were assessed using MyoRobot biomechatronics technology. RNA transcripts were significantly altered in β-DKO mice compared with fl/fl controls (32 in extensor digitorum longus and 412 in soleus). Exercise capacity and muscle strength were significantly decreased in β-DKO mice compared with fl/fl controls (P=0.012), and a loss of structural integrity was also observed in skeletal muscle from the β-DKO mice. Supplementation of β-DKO mice with insulin restored muscle integrity, strength and expression of 13 and 16 of the dysregulated transcripts in and extensor digitorum longus and soleus muscles, respectively. Insulin insufficiency due to β-cell dysfunction perturbs the structure and function of skeletal muscle. These adverse effects are rectified by insulin supplementation.

Glutathione Depression Alters Cellular Mechanisms of Skeletal Muscle Fatigue in Early Stage of Recovery and Prolongs Force Depression in Late Stage of Recovery.

The effects of reduced glutathione (GSH) on skeletal muscle fatigue were investigated. GSH was depressed by buthionine sulfoximine (BSO) (100 mg/kg body weight/day) treatment for 5 d, which decreased GSH content to ~10%. Male Wistar rats were assigned to the control (N = 18) and BSO groups (N =17). Twelve hours after BSO treatment, the plantar flexor muscles were subjected to fatiguing stimulation (FS). Eight control and seven BSO rats were rested for 0.5 h (early stage of recovery), and the remaining were rested for 6 h (late stage of recovery). Forces were measured before FS and after rest, and physiological functions were estimated using mechanically skinned fibers. The force at 40 Hz decreased to a similar extent in both groups in the early stage of recovery and was restored in the control but not in the BSO group in the late stage of recovery. In the early stage of recovery, sarcoplasmic reticulum (SR) Ca2+ release was decreased in the control greater than in the BSO group, whereas myofibrillar Ca2+ sensitivity was increased in the control but not in the BSO group. In the late stage of recovery, SR Ca2+ release decreased and SR Ca2+ leakage increased in the BSO group but not in the control group. These results indicate that GSH depression alters the cellular mechanism of muscle fatigue in the early stage and delays force recovery in the late stage of recovery, due at least in part, to the prolonged Ca2+ leakage from the SR.

A methylation-dependent checkpoint by SETD7 promotes myocardial ischemic injury in mice and men

Abstract Background Despite appropriate revascularization strategies, a significant number of patients with myocardial infarction (MI) develop ischemic heart failure suggesting that breakthrough therapies are yet to be approved in this setting. Methylation of non-histone proteins is emerging as a central regulatory mechanism in health and disease. The methyltransferase SETD7 has been shown to methylate and alter the function of a variety of proteins in vitro, however, its function in the heart is poorly understood. Purpose To determine the role of SETD7 in myocardial ischemic injury. Methods Neonatal rat ventricular myocytes (NRVM) were exposed to normal glucose levels or glucose deprivation (GD) for 15 h, in the presence of the selective SETD7 inhibitor (R)-PFI-2 or its inactive enantiomer (S)-PFI-2. Western blot and real-time PCR were employed to investigate the effects of energy stress on SETD7 and the Hippo pathway, while apoptosis and oxidative stress were assessed by Caspase-3 activity assay and mitoSOX staining. YAP transcriptional activity was assessed by chromatin immunoprecipitation assay (ChIP) while its localization and methylation were examined by confocal microscopy and immunoblotting, respectively. SETD7 knockout (SETD7−/−) mice and wild-type (WT) littermates underwent myocardial ischemia-reperfusion (I/R) injury (1h coronary ligation /24 h of reperfusion) followed by assessment of cardiac function by echocardiography. Left ventricular (LV) myocardial samples were collected from I/R mice and patients with ischemic cardiomyopathy (ICM), and isolated cardiomyocytes were treated with (R)-PFI-2. Finally, SETD7 expression was also assessed in peripheral blood mononuclear cells (PBMCs) from patients with ST-elevation MI (STEMI). Results SETD7 was activated upon energy deprivation in cultured NRVMs and methylated YAP, leading to its cytosolic retention and impaired transcription of antioxidant genes MnSOD and CAT. Pharmacological inhibition of SETD7 by (R)-PFI-2 restored YAP nuclear localization thus preventing mitochondrial reactive oxygen species (mtROS) and apoptosis. SETD7 deletion in mice attenuated I/R injury, mtROS and LV dysfunction by restoring YAP-dependent transcriptional programs. SETD7/YAP dysregulation was also observed in LV specimens from ICM patients. Moreover, in cardiomyocytes isolated from I/R mice and ICM patients, (R)-PFI-2 restored YAP nuclear localization, prevented mtROS accumulation while improving myofibrillar protein contractility and Ca2+ sensitivity. Finally, SETD7 was upregulated in PBMCs from STEMI patients and negatively correlated with the expression of MnSOD and CAT. Conclusions SETD7-dependent methylation of YAP is an important mechanism underpinning myocardial oxidative stress and apoptosis during ischemia. Pharmacological modulation of SETD7 by (R)-PFI-2 may represent a potential therapeutic approach to prevent myocardial ischemic damage through modulation of the Hippo pathway. Funding Acknowledgement Type of funding sources: Public Institution(s). Main funding source(s): University of Zurich

Sphingomyelinase activity promotes atrophy and attenuates force in human muscle fibres and is elevated in heart failure patients.

BackgroundActivation of sphingomyelinase (SMase) as a result of a general inflammatory response has been implicated as a mechanism underlying disease‐related loss of skeletal muscle mass and function in several clinical conditions including heart failure. Here, for the first time, we characterize the effects of SMase activity on human muscle fibre contractile function and assess skeletal muscle SMase activity in heart failure patients.MethodsThe effects of SMase on force production and intracellular Ca2+ handling were investigated in single intact human muscle fibres. Additional mechanistic studies were performed in single mouse toe muscle fibres. RNA sequencing was performed in human muscle bundles exposed to SMase. Intramuscular SMase activity was measured from heart failure patients (n = 61, age 69 ± 0.8 years, NYHA III‐IV, ejection fraction 25 ± 1.0%, peak VO2 14.4 ± 0.6 mL × kg × min) and healthy age‐matched control subjects (n = 10, age 71 ± 2.2 years, ejection fraction 60 ± 1.2%, peak VO2 25.8 ± 1.1 mL × kg × min). SMase activity was related to circulatory factors known to be associated with progression and disease severity in heart failure.ResultsSphingomyelinase reduced muscle fibre force production (−30%, P < 0.05) by impairing sarcoplasmic reticulum (SR) Ca2+ release (P < 0.05) and reducing myofibrillar Ca2+ sensitivity. In human muscle bundles exposed to SMase, RNA sequencing analysis revealed 180 and 291 genes as up‐regulated and down‐regulated, respectively, at a FDR of 1%. Gene‐set enrichment analysis identified ‘proteasome degradation’ as an up‐regulated pathway (average fold‐change 1.1, P = 0.008), while the pathway ‘cytoplasmic ribosomal proteins’ (average fold‐change 0.8, P < 0.0001) and factors involving proliferation of muscle cells (average fold‐change 0.8, P = 0.0002) where identified as down‐regulated. Intramuscular SMase activity was ~20% higher (P < 0.05) in human heart failure patients than in age‐matched healthy controls and was positively correlated with markers of disease severity and progression, and with several circulating inflammatory proteins, including TNF‐receptor 1 and 2. In a longitudinal cohort of heart failure patients (n = 6, mean follow‐up time 2.5 ± 0.2 years), SMase activity was demonstrated to increase by 30% (P < 0.05) with duration of disease.ConclusionsThe present findings implicate activation of skeletal muscle SMase as a mechanism underlying human heart failure‐related loss of muscle mass and function. Moreover, our findings strengthen the idea that SMase activation may underpin disease‐related loss of muscle mass and function in other clinical conditions, acting as a common patophysiological mechanism for the myopathy often reported in diseases associated with a systemic inflammatory response.

Role of Oxidative Stress in Cardiac Dysfunction and Subcellular Defects Due to Ischemia-Reperfusion Injury.

Ischemia-reperfusion (I/R) injury is well-known to be associated with impaired cardiac function, massive arrhythmias, marked alterations in cardiac metabolism and irreversible ultrastructural changes in the heart. Two major mechanisms namely oxidative stress and intracellular Ca2+-overload are considered to explain I/R-induced injury to the heart. However, it is becoming apparent that oxidative stress is the most critical pathogenic factor because it produces myocardial abnormalities directly or indirectly for the occurrence of cardiac damage. Furthermore, I/R injury has been shown to generate oxidative stress by promoting the formation of different reactive oxygen species due to defects in mitochondrial function and depressions in both endogenous antioxidant levels as well as regulatory antioxidative defense systems. It has also been demonstrated to adversely affect a wide variety of metabolic pathways and targets in cardiomyocytes, various resident structures in myocardial interstitium, as well as circulating neutrophils and leukocytes. These I/R-induced alterations in addition to myocardial inflammation may cause cell death, fibrosis, inflammation, Ca2+-handling abnormalities, activation of proteases and phospholipases, as well as subcellular remodeling and depletion of energy stores in the heart. Analysis of results from isolated hearts perfused with or without some antioxidant treatments before subjecting to I/R injury has indicated that cardiac dysfunction is associated with the development of oxidative stress, intracellular Ca2+-overload and protease activation. In addition, changes in the sarcolemma and sarcoplasmic reticulum Ca2+-handling, mitochondrial oxidative phosphorylation as well as myofibrillar Ca2+-ATPase activities in I/R hearts were attenuated by pretreatment with antioxidants. The I/R-induced alterations in cardiac function were simulated upon perfusing the hearts with oxyradical generating system or oxidant. These observations support the view that oxidative stress may be intimately involved in inducing intracellular Ca2+-overload, protease activation, subcellular remodeling, and cardiac dysfunction as a consequence of I/R injury to the heart.

Involvement of Oxidative Stress in the Development of Subcellular Defects and Heart Disease.

It is now well known that oxidative stress promotes lipid peroxidation, protein oxidation, activation of proteases, fragmentation of DNA and alteration in gene expression for producing myocardial cell damage, whereas its actions for the induction of fibrosis, necrosis and apoptosis are considered to result in the loss of cardiomyocytes in different types of heart disease. The present article is focused on the discussion concerning the generation and implications of oxidative stress from various sources such as defective mitochondrial electron transport and enzymatic reactions mainly due to the activation of NADPH oxidase, nitric oxide synthase and monoamine oxidase in diseased myocardium. Oxidative stress has been reported to promote excessive entry of Ca2+ due to increased permeability of the sarcolemmal membrane as well as depressions of Na+-K+ ATPase and Na+-Ca2+ exchange systems, which are considered to increase the intracellular of Ca2+. In addition, marked changes in the ryanodine receptors and Ca2+-pump ATPase have been shown to cause Ca2+-release and depress Ca2+ accumulation in the sarcoplasmic reticulum as a consequence of oxidative stress. Such alterations in sarcolemma and sarcoplasmic reticulum are considered to cause Ca2+-handling abnormalities, which are associated with mitochondrial Ca2+-overload and loss of myofibrillar Ca2+-sensitivity due to oxidative stress. Information regarding the direct effects of different oxyradicals and oxidants on subcellular organelles has also been outlined to show the mechanisms by which oxidative stress may induce Ca2+-handling abnormalities. These observations support the view that oxidative stress plays an important role in the genesis of subcellular defects and cardiac dysfunction in heart disease.

Myostatin gene inactivation increases post-mortem calpain-dependent muscle proteolysis in mice

Myostatin deficiency leads to extensive skeletal muscle hypertrophy, but its consequence on post-mortem muscle proteolysis is unknown. Here, we compared muscle myofibrillar protein degradation, and autophagy, ubiquitin-proteasome and Ca2+-dependent proteolysis relative to the energetic and redox status in wild-type (WT) and myostatin knock-out mice (KO) during early post-mortem storage. KO muscles showed higher degradation of myofibrillar proteins in the first 24 h after death, associated with preserved antioxidant status, compared with WT muscles. Analysis of key autophagy and ubiquitin-proteasome system markers indicated that these two pathways were not upregulated in post-mortem muscle (both genotypes), but basal autophagic flux and ATP content were lower in KO muscles. Proteasome and caspase activities were not different between WT and KO mice. Conversely, calpain activity was higher in KO muscles, concomitantly with higher troponin T and desmin degradation. Altogether, these results suggest that calpains but not the autophagy, proteasome and caspase systems, explain the difference in post-mortem muscle protein proteolysis between both genotypes.

Can a chloride channel blocker mitigate muscle fatigue?

Can a chloride channel blocker mitigate muscle fatigue?

Fast skeletal muscle troponin activator CK-2066260 mitigates skeletal muscle weakness independently of the underlying cause.

BackgroundMuscle weakness is a common symptom in numerous diseases and a regularly occurring problem associated with ageing. Prolonged low‐frequency force depression (PLFFD) is a form of exercise‐induced skeletal muscle weakness observed after exercise. Three different intramuscular mechanisms underlying PLFFD have been identified: decreased sarcoplasmic reticulum Ca2+ release, decreased myofibrillar Ca2+ sensitivity, and myofibrillar dysfunction. We here used these three forms of PLFFD as models to study the effectiveness of a fast skeletal muscle troponin activator, CK‐2066260, to mitigate muscle weakness.MethodsExperiments were performed on intact single muscle fibres or fibre bundles from mouse flexor digitorum brevis, which were stimulated with electrical current pulses, while force and the free cytosolic [Ca2+] ([Ca2+]i) were measured. PLFFD was induced by three different stimulation protocols: (i) repeated isometric contractions at low intensity (350 ms tetani given every 5 s for 100 contractions); (ii) repeated isometric contractions at high intensity (250 ms tetani given every 0.5 s for 300 contractions); and (iii) repeated eccentric contractions (350 ms tetani with 20% length increase given every 20 s for 10 contractions). The extent and cause of PLFFD were assessed by comparing the force–[Ca2+]i relationship at low (30 Hz) and high (120 Hz) stimulation frequencies before (control) and 30 min after induction of PLFFD, and after an additional 5 min of rest in the presence of CK‐2066260 (10 μM).ResultsProlonged low‐frequency force depression following low‐intensity and high‐intensity fatiguing contractions was predominantly due to decreased sarcoplasmic reticulum Ca2+ release and decreased myofibrillar Ca2+ sensitivity, respectively. CK‐2066260 exposure resulted in marked increases in 30 Hz force from 52 ± 16% to 151 ± 13% and from 6 ± 4% to 98 ± 40% of controls with low‐intensity and high‐intensity contractions, respectively. Following repeated eccentric contractions, PLFFD was mainly due to myofibrillar dysfunction, and it was not fully reversed by CK‐2066260 with 30 Hz force increasing from 48 ± 8% to 76 ± 6% of the control.ConclusionsThe fast skeletal muscle troponin activator CK‐2066260 effectively mitigates muscle weakness, especially when it is caused by impaired activation of the myofibrillar contractile machinery due to either decreased sarcoplasmic reticulum Ca2+ release or reduced myofibrillar Ca2+ sensitivity.

The Popeye mythology may be true: eating spinach can make you fatigue resistant!

The Popeye mythology may be true: eating spinach can make you fatigue resistant!

Cryoprotective characteristics of different sugar alcohols on peeled Pacific white shrimp (Litopenaeus vannamei) during frozen storage and their possible mechanisms of action

ABSTRACT Sugar alcohols, also known as polyols, have been widely applied in food products for their unique properties. In this work, the effect of soaking treatments composed of the sugar alcohols xylitol, sorbitol, erythritol, lactitol, mannitol, maltitol, and isomaltitol on the quality properties of peeled Pacific white shrimp during frozen storage were evaluated. Samples treated with water and sodium pyrophosphate (Na4P2O7) were designated as the negative and positive controls, respectively. The results indicated that significant improvements were found in thawing, pressing, and cooking loss, myofibrillar protein content, myofibrillar Ca2+-ATPase activity, and textural variables in xylitol- and mannitol-treated shrimp as compared to that of control samples. Water mobility analyses showed that xylitol and mannitol effectively increased the relaxation times T 21 and T 22 and the percentage of A 21 (trapped water) but decreased the percentage of A 22 (free water) in muscle tissue. Additionally, the microstructural results indicated that soaking in xylitol and mannitol significantly slowed the damage caused to muscle tissues by large ice crystals. Here, the incorporated xylitol and mannitol molecules presumably replaced water molecules by forming large hydrogen bonds with the muscle proteins, thereby stabilizing their structures in the absence of water while in the frozen state and suppressing the destruction of tissue structure. To note, the lactitol, isomaltitol, and maltitol used in this study did not seem to significantly contribute to cryoprotective effects on shrimp muscle during the experimental period.

Impaired sarcoplasmic reticulum Ca2+ release is the major cause of fatigue-induced force loss in intact single fibres from human intercostal muscle.

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. 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.

Ultrasound treatment of frozen crayfish with chitosan Nano-composite water-retaining agent: Influence on cryopreservation and storage qualities.

The synergistic impact of ultrasound and Nano-water retaining agent on the cryoprotective effect on crayfish during frozen storage was investigated. The samples soaked in water-retaining agent (WRA), ultrasonic 60 W treatment combined with water-retaining agent (WRA-US60W) and ultrasonic 80 W treatment combined with water-retaining agent (WRA-US80W) were frozen storage at -18 °C. The indices of frozen storage in 0, 7, 14, 21, 28, 35 and 42 days were measured. The results showed that there was a significant difference in the soaking weight gain, thawing loss, water content and water activity between control group and WRA groups (P < .05). The ultrasound combined with WRA treatment showed better water retention effect. The water holding capacity of heat induced gel decreased continuously during the frozen storage period, and the WRA-US60W group exhibited significantly higher values than that of other treatment groups (P < .05). The texture characteristics of hardness, elasticity and chewiness had a significant change (P < .05). The development of total volatile base nitrogen content, myofibrillar protein content and Ca2+-ATPase activity of muscle protein were significantly delayed by WRA-US60W treatment, maintaining the integrity of tissue structure. Therefore, WRA-US60W treatment was found effectively improving the quality of crayfish during frozen storage.

Fast skeletal muscle troponin activator CK-2066260 increases fatigue resistance by reducing the energetic cost of muscle contraction.

Key points Skeletal muscle fatigue limits performance in various physical activities, with exercise intolerance being a key symptom in a broad spectrum of diseases.We investigated whether a small molecule fast skeletal troponin activator (FSTA), CK‐2066260, can mitigate muscle fatigue by reducing the cytosolic free [Ca2+] required to produce a given submaximal force and hence decreasing the energy requirement.Isolated intact single mouse muscle fibres and rat muscles in‐situ treated with CK‐2066260 showed improved muscle endurance., which was accompanied by decreased ATP demand and reduced glycogen usage.CK‐2066260 treatment improved in‐vivo exercise capacity in healthy rats and in a rat model of peripheral artery insufficiency.In conclusion, we show that the FSTA CK‐2066260 effectively counteracts muscle fatigue in rodent skeletal muscle in vitro, in situ, and in vivo. This may translate to humans and provide a promising pharmacological treatment to patients suffering from severe muscle weakness and exercise intolerance. Skeletal muscle fatigue limits performance during physical exercise and exacerbated muscle fatigue is a prominent symptom among a broad spectrum of diseases. The present study investigated whether skeletal muscle fatigue is affected by the fast skeletal muscle troponin activator (FSTA) CK‐2066260, which increases myofibrillar Ca2+ sensitivity and amplifies the submaximal force response. Because more force is produced for a given Ca2+, we hypothesized that CK‐2066260 could mitigate muscle fatigue by reducing the energetic cost of muscle activation. Isolated single mouse muscle fibres were fatigued by 100 repeated 350 ms contractions while measuring force and the cytosolic free [Ca2+] or [Mg2+] ([Mg2+]i). When starting fatiguing stimulation at matching forces (i.e. lower stimulation frequency with CK‐2066260): force was decreased by ∼50% with and by ∼75% without CK‐2066260; [Mg2+]i was increased by ∼10% with and ∼32% without CK‐2066260, reflecting a larger decrease in [ATP] in the latter. The glycogen content in in situ stimulated rat muscles fatigued by repeated contractions at matching forces was about two times higher with than without CK‐2066260. Voluntary exercise capacity, assessed by rats performing rotarod exercise and treadmill running, was improved in the presence of CK‐2066260. CK‐2066260 treatment also increased skeletal muscle fatigue resistance and exercise performance in a rat model of peripheral artery insufficiency. In conclusion, we demonstrate that the FSTA CK‐2066260 mitigates skeletal muscle fatigue by reducing the metabolic cost of force generation.