Activation of skeletal muscle is controlled by a dual-filament mechano-sensing mechanism.

Contraction of skeletal muscle is triggered by a transient rise in intracellular calcium concentration leading to a structural change in the actin-containing thin filaments that allows binding of myosin motors from the thick filaments. Most myosin motors are unavailable for actin binding in resting muscle because they are folded back against the thick filament backbone. Release of the folded motors is triggered by thick filament stress, implying a positive feedback loop in the thick filaments. However, it was unclear how thin and thick filament activation mechanisms are coordinated, partly because most previous studies of the thin filament regulation were conducted at low temperatures where the thick filament mechanisms are inhibited. Here, we use probes on both troponin in the thin filaments and myosin in the thick filaments to monitor the activation states of both filaments in near-physiological conditions. We characterize those activation states both in the steady state, using conventional titrations with calcium buffers, and during activation on the physiological timescale, using calcium jumps produced by photolysis of caged calcium. The results reveal three activation states of the thin filament in the intact filament lattice of a muscle cell that are analogous to those proposed previously from studies on isolated proteins. We characterize the rates of the transitions between these states in relation to thick filament mechano-sensing and show how thin- and thick-filament-based mechanisms are coupled by two positive feedback loops that switch on both filaments to achieve rapid cooperative activation of skeletal muscle.

ß3AR-Dependent Brain-Derived Neurotrophic Factor (BDNF) Generation Limits Chronic Postischemic Heart Failure.

BACKGROUND: Loss of brain-derived neurotrophic factor (BDNF)/TrkB (tropomyosin kinase receptor B) signaling accounts for brain and cardiac disorders. In neurons, ß-adrenergic receptor stimulation enhances local BDNF expression. It is unclear if this occurs in a pathophysiological relevant manner in the heart, especially in the ß-adrenergic receptor-desensitized postischemic myocardium. Nor is it fully understood whether and how TrkB agonists counter chronic postischemic left ventricle (LV) decompensation, a significant unmet clinical milestone. METHODS: We conducted in vitro studies using neonatal rat and adult murine cardiomyocytes, SH-SY5Y neuronal cells, and umbilical vein endothelial cells. We assessed myocardial ischemia (MI) impact in wild type, ß3AR knockout, or myocyte-selective BDNF knockout (myoBDNF KO) mice in vivo (via coronary ligation [MI]) or in isolated hearts with global ischemia-reperfusion (I/R). RESULTS: In wild type hearts, BDNF levels rose early after MI (<24 hours), plummeting at 4 weeks when LV dysfunction, adrenergic denervation, and impaired angiogenesis ensued. The TrkB agonist, LM22A-4, countered all these adverse effects. Compared with wild type, isolated myoBDNF KO hearts displayed worse infarct size/LV dysfunction after I/R injury and modest benefits from LM22A-4. In vitro, LM22A-4 promoted neurite outgrowth and neovascularization, boosting myocyte function, effects reproduced by 7,8-dihydroxyflavone, a chemically unrelated TrkB agonist. Superfusing myocytes with the ß3AR-agonist, BRL-37344, increased myocyte BDNF content, while ß3AR signaling underscored BDNF generation/protection in post-MI hearts. Accordingly, the ß1AR blocker, metoprolol, via upregulated ß3ARs, improved chronic post-MI LV dysfunction, enriching the myocardium with BDNF. Last, BRL-37344-imparted benefits were nearly abolished in isolated I/R injured myoBDNF KO hearts. CONCLUSIONS: BDNF loss underscores chronic postischemic heart failure. TrkB agonists can improve ischemic LV dysfunction via replenished myocardial BDNF content. Direct cardiac ß3AR stimulation, or ß-blockers (via upregulated ß3AR), is another BDNF-based means to fend off chronic postischemic heart failure.

Muscle Mechanics and Thick Filament Activation: An Emerging Two-Way Interaction for the Vertebrate Striated Muscle Fine Regulation.

Contraction in striated muscle is classically described as regulated by calcium-mediated structural changes in the actin-containing thin filaments, which release the binding sites for the interaction with myosin motors to produce force. In this view, myosin motors, arranged in the thick filaments, are basically always ready to interact with the thin filaments, which ultimately regulate the contraction. However, a new "dual-filament" activation paradigm is emerging, where both filaments must be activated to generate force. Growing evidence from the literature shows that the thick filament activation has a role on the striated muscle fine regulation, and its impairment is associated with severe pathologies. This review is focused on the proposed mechanical feedback that activates the inactive motors depending on the level of tension generated by the active ones, the so-called mechanosensing mechanism. Since the main muscle function is to generate mechanical work, the implications on muscle mechanics will be highlighted, showing: (i) how non-mechanical modulation of the thick filament activation influences the contraction, (ii) how the contraction influences the activation of the thick filament and (iii) how muscle, through the mechanical modulation of the thick filament activation, can regulate its own mechanics. This description highlights the crucial role of the emerging bi-directional feedback on muscle mechanical performance.

Inducible deletion of raptor and mTOR from adult skeletal muscle impairs muscle contractility and relaxation.

Skeletal muscle weakness has been associated with different pathological conditions, including sarcopenia and muscular dystrophy, and is accompanied by altered mammalian target of rapamycin (mTOR) signalling. We wanted to elucidate the functional role of mTOR in muscle contractility. Most loss-of-function studies for mTOR signalling have used the drug rapamycin to inhibit some of the signalling downstream of mTOR. However, given that rapamycin does not inhibit all mTOR signalling completely, we generated a double knockout for mTOR and for the scaffold protein of mTORC1, raptor, in skeletal muscle. We found that double knockout in mice results in a more severe phenotype compared with deletion of raptor or mTOR alone. Indeed, these animals display muscle weakness, increased fibre denervation and a slower muscle relaxation following tetanic stimulation. This is accompanied by a shift towards slow-twitch fibres and changes in the expression levels of calcium-related genes, such as Serca1 and Casq1. Double knockout mice show a decrease in calcium decay kinetics after tetanus in vivo, suggestive of a reduced calcium reuptake. In addition, RNA sequencing analysis revealed that many downregulated genes, such as Tcap and Fhod3, are linked to sarcomere organization. These results suggest a key role for mTOR signalling in maintaining proper fibre relaxation in skeletal muscle. KEY POINTS: Skeletal muscle wasting and weakness have been associated with different pathological conditions, including sarcopenia and muscular dystrophy, and are accompanied by altered mammalian target of rapamycin (mTOR) signalling. Mammalian target of rapamycin plays a crucial role in the maintenance of muscle mass and functionality. We found that the loss of both mTOR and raptor results in contractile abnormalities, with severe muscle weakness and delayed relaxation following tetanic stimulation. These results are associated with alterations in the expression of genes involved in sarcomere organization and calcium handling and with an impairment in calcium reuptake after contraction. Taken together, these results provide a mechanistic insight into the role of mTOR in muscle contractility.

Neuromuscular junction instability and altered intracellular calcium handling as early determinants of force loss during unloading in humans.

KEY POINTS: Few days of unloading are sufficient to induce a decline of skeletal muscle mass and function; notably, contractile force is lost at a faster rate than muscle mass. The reasons behind this disproportionate loss of muscle force are still poorly understood. We provide strong evidence of two mechanisms only hypothesized until now for the rapid muscle force loss in only 10 days of bed rest. Our results show that an initial neuromuscular junction instability, accompanied by alterations in the innervation status and impairment of single fibre sarcoplasmic reticulum function contribute to the loss of contractile force in front of a preserved myofibrillar function and central activation capacity. Early onset of neuromuscular junction instability and impairment in calcium dynamics involved in excitation-contraction coupling are proposed as eligible determinants to the greater decline in muscle force than in muscle size during unloading. ABSTRACT: Unloading induces rapid skeletal muscle atrophy and functional decline. Importantly, force is lost at a much higher rate than muscle mass. We aimed to investigate the early determinants of the disproportionate loss of force compared to that of muscle mass in response to unloading. Ten young participants underwent 10 days of bed rest (BR). At baseline (BR0) and at 10 days (BR10), quadriceps femoris (QF) volume (VOL) and isometric maximum voluntary contraction (MVC) were assessed. At BR0 and BR10 blood samples and biopsies of vastus lateralis (VL) muscle were collected. Neuromuscular junction (NMJ) stability and myofibre innervation status were assessed, together with single fibre mechanical properties and sarcoplasmic reticulum (SR) calcium handling. From BR0 to BR10, QFVOL and MVC decreased by 5.2% (P = 0.003) and 14.3% (P < 0.001), respectively. Initial and partial denervation was detected from increased neural cell adhesion molecule (NCAM)-positive myofibres at BR10 compared with BR0 (+3.4%, P = 0.016). NMJ instability was further inferred from increased C-terminal agrin fragment concentration in serum (+19.2% at BR10, P = 0.031). Fast fibre cross-sectional area (CSA) showed a trend to decrease by 15% (P = 0.055) at BR10, while single fibre maximal tension (force/CSA) was unchanged. However, at BR10 SR Ca2+ release in response to caffeine decreased by 35.1% (P < 0.002) and 30.2% (P < 0.001) in fast and slow fibres, respectively, pointing to an impaired excitation-contraction coupling. These findings support the view that the early onset of NMJ instability and impairment in SR function are eligible mechanisms contributing to the greater decline in muscle force than in muscle size during unloading.

Evolution over Time of Leg Length Discrepancy in Patients with Syndromic and Isolated Lateralized Overgrowth.

OBJECTIVE: To provide information on evolution over time of leg length discrepancy in patients with syndromic and isolated lateralized overgrowth. STUDY DESIGN: This retrospective study investigates leg length discrepancy longitudinally in 105 patients with lateralized overgrowth either isolated (n = 37) or associated with Beckwith-Wiedemann spectrum (n = 56) or PIK3CA-related overgrowth spectrum (n = 12). Discrepancy was measured by standard methods and categorized as minor, mild, severe, and critical, based on the thresholds of 1, 2 and 5, respectively. RESULTS: The period of observation from diagnosis was 1.7 ± 2.6 to 9.0 ± 6.0 years. Leg length discrepancy was 11.0 ± 7.2 mm at diagnosis and 17.1 ± 14.4 mm at last visit. Both final leg length discrepancy and change over time were correlated with discrepancy at diagnosis (r2 = 0.45, P < .001 and r2 = 0.05, P = .019, respectively). Among minor leg length discrepancy at diagnosis, 47.5% remained minor, 40.0% become mild, and 12.5% severe. Among patients with discrepancy classified as severe at diagnosis, 84.6% remained severe and 15.4% evolved to critical. The isolated lateralized overgrowth group showed a milder evolution over time compared with Beckwith-Wiedemann spectrum and PIK3CA-related overgrowth spectrum groups. Among patients with Beckwith-Wiedemann, those with paternal chromosome 11 uniparental disomy had more severe leg length discrepancy at diagnosis and evolution over time. CONCLUSIONS: Leg length discrepancy associated with isolated or syndromic lateralized overgrowth tends to worsen with growth and correlates with discrepancy at first observation. Among the genotypic groups, isolated lateralized overgrowth tends to have a milder evolution, whereas Beckwith-Wiedemann spectrum predisposes to a more severe outcome, especially if associated with paternal chromosome 11 uniparental disomy genotype.

The Synergic Role of Actomyosin Architecture and Biased Detachment in Muscle Energetics: Insights in Cross Bridge Mechanism Beyond the Lever-Arm Swing.

Muscle energetics reflects the ability of myosin motors to convert chemical energy into mechanical energy. How this process takes place remains one of the most elusive questions in the field. Here, we combined experimental measurements of in vitro sliding velocity based on DNA-origami built filaments carrying myosins with different lever arm length and Monte Carlo simulations based on a model which accounts for three basic components: (i) the geometrical hindrance, (ii) the mechano-sensing mechanism, and (iii) the biased kinetics for stretched or compressed motors. The model simulations showed that the geometrical hindrance due to acto-myosin spatial mismatching and the preferential detachment of compressed motors are synergic in generating the rapid increase in the ATP-ase rate from isometric to moderate velocities of contraction, thus acting as an energy-conservation strategy in muscle contraction. The velocity measurements on a DNA-origami filament that preserves the motors' distribution showed that geometrical hindrance and biased detachment generate a non-zero sliding velocity even without rotation of the myosin lever-arm, which is widely recognized as the basic event in muscle contraction. Because biased detachment is a mechanism for the rectification of thermal fluctuations, in the Brownian-ratchet framework, we predict that it requires a non-negligible amount of energy to preserve the second law of thermodynamics. Taken together, our theoretical and experimental results elucidate less considered components in the chemo-mechanical energy transduction in muscle.

Alterations of Extracellular Matrix Mechanical Properties Contribute to Age-Related Functional Impairment of Human Skeletal Muscles.

Aging of human skeletal muscles is associated with increased passive stiffness, but it is still debated whether muscle fibers or extracellular matrix (ECM) are the determinants of such change. To answer this question, we compared the passive stress generated by elongation of fibers alone and arranged in small bundles in young healthy (Y: 21 years) and elderly (E: 67 years) subjects. The physiological range of sarcomere length (SL) 2.5-3.3 µm was explored. The area of ECM between muscle fibers was determined on transversal sections with picrosirius red, a staining specific for collagen fibers. The passive tension of fiber bundles was significantly higher in E compared to Y at all SL. However, the resistance to elongation of fibers alone was not different between the two groups, while the ECM contribution was significantly increased in E compared to Y. The proportion of muscle area occupied by ECM increased from 3.3% in Y to 8.2% in E. When the contribution of ECM to bundle tension was normalized to the fraction of area occupied by ECM, the difference disappeared. We conclude that, in human skeletal muscles, the age-related reduced compliance is due to an increased stiffness of ECM, mainly caused by collagen accumulation.

Including Thermal Fluctuations in Actomyosin Stable States Increases the Predicted Force per Motor and Macroscopic Efficiency in Muscle Modelling.

Muscle contractions are generated by cyclical interactions of myosin heads with actin filaments to form the actomyosin complex. To simulate actomyosin complex stable states, mathematical models usually define an energy landscape with a corresponding number of wells. The jumps between these wells are defined through rate constants. Almost all previous models assign these wells an infinite sharpness by imposing a relatively simple expression for the detailed balance, i.e., the ratio of the rate constants depends exponentially on the sole myosin elastic energy. Physically, this assumption corresponds to neglecting thermal fluctuations in the actomyosin complex stable states. By comparing three mathematical models, we examine the extent to which this hypothesis affects muscle model predictions at the single cross-bridge, single fiber, and organ levels in a ceteris paribus analysis. We show that including fluctuations in stable states allows the lever arm of the myosin to easily and dynamically explore all possible minima in the energy landscape, generating several backward and forward jumps between states during the lifetime of the actomyosin complex, whereas the infinitely sharp minima case is characterized by fewer jumps between states. Moreover, the analysis predicts that thermal fluctuations enable a more efficient contraction mechanism, in which a higher force is sustained by fewer attached cross-bridges.

Resection of Calcaneonavicular and Talocalcaneal Coalitions With Surgical Correction of the Hindfoot Valgus Deformity in One Step.

Background: Calcaneonavicular (CNC) and talocalcaneal (TCC) coalitions are the most common cause of rigid flatfoot in children. After resection, correction of the most frequent valgus-hindfoot deformity usually requires a second-step surgery. We report results of a retrospective study of patients treated with a one-step correction. Methods: Between 2008 and 2019, data were collected on 26 patients (19 male, 7 female) affected by CNC (n = 18) and TCC (n = 13), all with rigid symptomatic flatfeet. Average age at surgery was 12.5 ± 1.1 (SD) years (range, 9.8-15.2). All patients (26/26) underwent resection, 20 of 26 underwent at the same time subtalar extraarticular screw arthroereisis (SESA) for correction of residual hindfoot valgus deformity. Pre- and postoperative talocalcaneal angle according to Costa Bartani and Talar inclination angle in weightbearing were measured. Twenty-five of 26 patients had postoperative American Orthopaedic Foot & Ankle Society (AOFAS) ankle-hindfoot score. Results: Pre- and postoperative talocalcaneal average angle for CNC was respectively 141.5 ± 7.7 degrees and 130.5 ± 5.2 degrees (P < .0001) and 143.7 ± 7.7 degrees and 129.7 ± 7.0 degrees (P < .0001) for TCC. Talar inclination average angle for CNC was 29.2 ± 5.3 degrees and 19.3 ± 1.6 degrees (P < .0001) and 31.2 ± 6.4 degrees and 21.4 ± 3.4 degrees (P < .0001) for TCC. Average follow-up (FU) was 4.7 ± 3.0 years (range, 6 months-11.9 years, median 4.9 years), with a mean age at FU of 17.2 ± 5.8 (SD) years (min 12.1, max 25.3, median 16.8 years). The mean AOFAS ankle-hindfoot score for CNC and for TCC was 96.6 (range 83-100) for resection and valgus correction as one-step procedure with no statistical difference (P = .5) between CNC and TCC. No patients had additional surgery for complications or recurrence. Conclusion: Symptomatic rigid flatfeet affected by CNC and TCC treated with coalition resection and minimally invasive subtalar arthroereisis (SESA) for residual hindfoot valgus correction in one step in adolescent age achieved good to excellent results in all cases. Further surgery to correct malalignment was avoided. Level of Evidence: Level IV, retrospective study.

The role of the extracellular matrix in the reduction of lateral force transmission in muscle bundles: A finite element analysis.

BACKGROUND AND OBJECTIVE: Aging is associated with a reduction in muscle performance, but muscle weakness is characterized by a much greater loss of force loss compared to mass loss. The aim of this work is to assess the contribution of the extracellular matrix (ECM) to the lateral transmission of force in humans and the loss of transmitted force due to age-related modifications. METHODS: Finite element models of muscle bundles are developed for young and elderly human subjects, by considering a few fibers connected through an ECM layer. Bundles of young and elderly subjects are assumed to differ in terms of ECM thickness, as observed experimentally. A three-element-based Hill model is adopted to describe the active behavior of muscle fibers, while the ECM is modeled assuming an isotropic hyperelastic neo-Hookean constitutive formulation. Numerical analyses are carried out by mimicking, at the scale of a bundle, two experimental protocols from the literature. RESULTS: When comparing numerical results obtained for bundles of young and elderly subjects, a greater reduction in the total transmitted force is observed in the latter. The loss of transmitted force is 22 % for the elderly subjects, while it is limited to 7.5 % for the young subjects. The result for the elderly subjects is in line with literature studies on animal models, showing a reduction in the range of 20-34 %. This can be explained by an alteration in the mechanism of lateral force transmission due to the lower shear stiffness of the ECM in elderly subjects, related to its higher thickness. CONCLUSIONS: Computational modeling allows to evaluate at the bundle level how the age-related increase of the ECM amount between fibers affects the lateral transmission of force. The results suggest that the observed increase in ECM thickness in aging alone can explain the reduction of the total transmitted force, due to the impaired lateral transmission of force of each fiber.

Mitochondria can substitute for parvalbumin to lower cytosolic calcium levels in the murine fast skeletal muscle.

AIM: Parvalbumin (PV) is a primary calcium buffer in mouse fast skeletal muscle fibers. Previous work showed that PV ablation has a limited impact on cytosolic Ca2+ ([Ca2+]cyto) transients and contractile response, while it enhances mitochondrial density and mitochondrial matrix-free calcium concentration ([Ca2+]mito). Here, we aimed to quantitatively test the hypothesis that mitochondria act to compensate for PV deficiency. METHODS: We determined the free Ca2+ redistribution during a 2 s 60 Hz tetanic stimulation in the sarcoplasmic reticulum, cytosol, and mitochondria. Via a reaction-diffusion Ca2+ model, we quantitatively evaluated mitochondrial uptake and storage capacity requirements to compensate for PV lack and analyzed possible extracellular export. RESULTS: [Ca2+]mito during tetanic stimulation is greater in knock-out (KO) (1362 ± 392 nM) than in wild-type (WT) (855 ± 392 nM), p < 0.05. Under the assumption of a non-linear intramitochondrial buffering, the model predicts an accumulation of 725 µmoles/L fiber (buffering ratio 1:11 000) in KO, much higher than in WT (137 µmoles/L fiber, ratio 1:4500). The required transport rate via mitochondrial calcium uniporter (MCU) reaches 3 mM/s, compatible with available literature. TEM images of calcium entry units and Mn2+ quenching showed a greater capacity of store-operated calcium entry in KO compared to WT. However, levels of [Ca2+]cyto during tetanic stimulation were not modulated to variations of extracellular calcium. CONCLUSIONS: The model-based analysis of experimentally determined calcium distribution during tetanic stimulation showed that mitochondria can act as a buffer to compensate for the lack of PV. This result contributes to a better understanding of mitochondria's role in modulating [Ca2+]cyto in skeletal muscle fibers.

Cytosolic Ca2+ gradients and mitochondrial Ca2+ uptake in resting muscle fibers: A model analysis.

Calcium ions (Ca2+) enter mitochondria via the mitochondrial Ca2+ uniporter, driven by electrical and concentration gradients. In this regard, transgenic mouse models, such as calsequestrin knockout (CSQ-KO) mice, with higher mitochondrial Ca2+ concentrations ([Ca2+]mito), should display higher cytosolic Ca2+ concentrations ([Ca2+]cyto). However, repeated measurements of [Ca2+]cyto in quiescent CSQ-KO fibers never showed a difference between WT and CSQ-KO. Starting from the consideration that fluorescent Ca2+ probes (Fura-2 and Indo-1) measure averaged global cytosolic concentrations, in this report we explored the role of local Ca2+ concentrations (i.e., Ca2+ microdomains) in regulating mitochondrial Ca2+ in resting cells, using a multicompartmental diffusional Ca2+ model. Progressively including the inward and outward fluxes of sarcoplasmic reticulum (SR), extracellular space, and mitochondria, we explored their contribution to the local Ca2+ distribution within the cell. The model predicts Ca2+ concentration gradients with hot spots or microdomains even at rest, minor but similar to those of evoked Ca2+ release. Due to their specific localization close to Ca2+ release units (CRU), mitochondria could take up Ca2+ directly from high-concentration microdomains, thus sensibly raising [Ca2+]mito, despite minor, possibly undetectable, modifications of the average [Ca2+]cyto.

Non-pegylated liposomal doxorubicin in older adjuvant early breast cancer patients: cardiac safety analysis and final results of the COLTONE study.

AIMS: To explore the cardiac safety of adjuvant Non-Pegylated Liposomal Doxorubicin (NPL-DOX) plus Cyclophosphamide (CTX) followed by weekly Paclitaxel, in elderly women (≥ 65 years) with high-risk breast cancer. Previously, we described no symptomatic cardiac events within the first 12 months from starting treatment. We now reported the updated results after a median follow-up 76 months. METHODS: The cardiac activity was evaluated with left ventricular ejection fraction (LVEF) echocardiograms assessments, before starting chemotherapy and every 6 months, until 30 months from baseline, then yearly for at least 5 years. RESULTS: Forty-seven women were recruited by two Units of Medical Oncology (Ethics Committee authorization CESM-AOUP, 3203/2011; EudraCT identification number: 2010-024067-41, for Pisa and Pontedera Hospitals). An episode of grade 3 CHF (NCI-CTCAE, version 3.0) occurred after 18 months the beginning of chemotherapy. The echocardiograms assessments were performed comparing the LVEF values of each patient evaluated at fixed period of time, compared to baseline. We observed a slight changed in terms of mean values at 48, 60, 72 and 84 months. At these time points, a statistically significant reduction of - 3.2%, - 4.6%, - 6.4% and - 7.1%, respectively, was observed. However, LVEF remained above 50% without translation in any relevant clinical signs. No other cardiac significant episodes were reported. To this analysis, in 13 patients (28%) occurred disease relapse and,  of them, 11 (23%) died due to metastatic disease. Eight patients died of cancer-unrelated causes. CONCLUSIONS: The combination including NPL-DOX in elderly patients revealed low rate of cardiac toxic effects. Comparative trials are encouraged.

Defective excitation-contraction coupling and mitochondrial respiration precede mitochondrial Ca2+ accumulation in spinobulbar muscular atrophy skeletal muscle.

Polyglutamine expansion in the androgen receptor (AR) causes spinobulbar muscular atrophy (SBMA). Skeletal muscle is a primary site of toxicity; however, the current understanding of the early pathological processes that occur and how they unfold during disease progression remains limited. Using transgenic and knock-in mice and patient-derived muscle biopsies, we show that SBMA mice in the presymptomatic stage develop a respiratory defect matching defective expression of genes involved in excitation-contraction coupling (ECC), altered contraction dynamics, and increased fatigue. These processes are followed by stimulus-dependent accumulation of calcium into mitochondria and structural disorganization of the muscle triads. Deregulation of expression of ECC genes is concomitant with sexual maturity and androgen raise in the serum. Consistent with the androgen-dependent nature of these alterations, surgical castration and AR silencing alleviate the early and late pathological processes. These observations show that ECC deregulation and defective mitochondrial respiration are early but reversible events followed by altered muscle force, calcium dyshomeostasis, and dismantling of triad structure.

A controversial issue: Can mitochondria modulate cytosolic calcium and contraction of skeletal muscle fibers?

Mitochondria are characterized by a high capacity to accumulate calcium thanks to the electrochemical gradient created by the extrusion of protons in the respiratory chain. Thereby calcium can enter crossing the inner mitochondrial membrane via MCU complex, a high-capacity, low-affinity transport mechanism. Calcium uptake serves numerous purposes, among them the regulation of three dehydrogenases of the citric cycle, apoptosis via permeability transition, and, in some cell types, modulation of cytosolic calcium transients. This Review is focused on mitochondrial calcium uptake in skeletal muscle fibers and aims to reanalyze its functional impact. In particular, we ask whether mitochondrial calcium uptake is relevant for the control of cytosolic calcium transients and therefore of contractile performance. Recent data suggest that this may be the case, at least in particular conditions, as modified expression of MCU complex subunits or of proteins involved in mitochondrial dynamics and ablation of the main cytosolic calcium buffer, parvalbumin.

Surgical Treatment of Clubfoot in Children with Moebius Syndrome.

Moebius syndrome (MS) is a rare disease, with paralysis of the VI and VII cranial nerves, frequently associated with clubfoot (CF). The aim of this study was to evaluate surgical treatment of CF in MS, providing its peculiarities. Between 1990 and 2019, we collected data of 11 MS patients with unilateral (n = 5) or bilateral (n = 6) CF, for a total of 17 feet (9R,8L). Six patients (3M,3F) for a total of 10 feet (6R,4L) were treated elsewhere, performing first surgery at an average age of nine months, and in our hospital for relapse surgery at an average age of 4.5 years (Group 1). Five patients (3M, 2F), for a total of seven feet (3R,4L), were primarily treated in our hospital with a peritalar release according to McKay at an average age of 9.4 months (Group 2). Diméglio score was used to assess CF severity. Three questionnaires were submitted for evaluation of subjective and functional results: American Orthopedics Foot and Ankle Society for Hindfoot (AOFAS), Foot and Ankle Outcome Score (FAOS), and Foot and Ankle Ability Measure (FAAM). Average AOFAS/FAOS/FAMM scores were 82.8, 84.8, and 82.3 for Group 1, and 93.2, 94.7, and 95.1 for Group 2 at an average follow-up of 16.9 and 13.3 years, respectively. The average Diméglio score improved from 15.5 to 4.8 in the long-term follow-up in Group 1 and from 14.6 to 3.8 in Group 2. The comparison between the groups showed better results for AOFAS, FAOS, and FAAM scores for Group 2, particularly for pain, function, and foot alignment and for the post-surgical Diméglio score. CF in MS is more severe and presented a higher relapse rate (58.8%) than idiopathic CF. Peritalar release showed no relapse and better subjective and functional results in the long-term follow-up compared to other surgical techniques.

Double Diapering Ineffectiveness in Avoiding Adduction and Extension in Newborns Hips.

Hip flexion and abduction is fundamental for developmental dysplasia of the hip (DDH) treatment. At present, double diaper treatment has been inappropriately adopted when DDH is suspected. The aim of this study was to verify whether double diapers influence a newborn's hip position. Here, we studied 50 children (23 female; 27 male; average age 62.33 ± 20.50 days; average birth weight 3230 ± 447 g) with type I hips according to Graf. At the same time of the ultrasound (US) examination, the following hip positions were measured using a manual protractor: (1) spontaneous position, supine on the outpatient bed without a diaper; (2) spontaneous position, with a double diaper; and (3) squatting position on the caretakers' side. Statistical analysis was performed with a t-test to compare between (1) the spontaneous position without a diaper and with double diapers; (2) the spontaneous position with double diapers as well as the squatting position on the caretakers' side with a diaper. The comparison between the hip position without diaper and with double diapers was statistically not significant for all measurements, i.e., right hip flexion (p < 0.33), left hip flexion (p < 0.34), and right and left hip abduction (p < 0.87). The comparison between the hip position with double diapers and on the caretakers' side was statistically significant for all measurements, i.e., right hip flexion (p < 0.001), left hip flexion (p < 0.001) and right and left hip abduction (p < 0.001). We found that the use of double diapers did not affect hip position, while the position formed on the caretaker's side shows favorable influence.

Guided Growth in Leg Length Discrepancy in Beckwith-Wiedemann Syndrome: A Consecutive Case Series.

Beckwith-Wiedemann Syndrome (BWS) is a rare genetic disorder characterized by overgrowth, macroglossia, abdominal wall defects, neonatal hypoglycemia, predisposition to embryonal tumor, lateralized overgrowth, and leg length discrepancy (LLD), which can affect normal posture and gait. Aim of this study was to evaluate the effects of guided growth (temporary epiphysiodesis technique) as LLD management in BWS patients. Between 2007 and 2021, 22 BWS patients (15 F, 7 M) with a mean age of 7.9 years (2.9-14.4) and a mean LLD at first surgery of 3.65 cm (2-10), underwent temporary proximal tibial (PTE) and distal femur epiphysiodesis (DFE). In 18 patients the first surgical procedure was PTE, in one, DFE, and in 3 cases, PTE and DFE at the same time, respectively. Eleven patients reached equality of leg length after a mean follow-up of 7.7 years (3.7-13.0) and mean age of 13.3 years (12.7-27.5); 10 patients underwent 3 surgical procedures, one 7 procedures. Fifteen patients had no complications. No severe complications, infection, articular stiffness, or neuro-vascular lesions occurred in remaining patients; complications included secondary varus or valgus axial deviation in a total of 6 patients, and two screw breakages in two patients. Guided growth as a minimally invasive procedure seems efficient for LLD treatment with low complication rate in BWS patients.

Are muscle fibres of body builders intrinsically weaker? A comparison with single fibres of aged-matched controls.

AIM: Skeletal muscles of Body Builders (BB) represent an interesting model to study muscle mass gains in response to high volume resistance training. It is debated whether muscle contractile performance improves in proportion to mass. Here, we aim to assess whether muscle hypertrophy does not occur at the expense of performance. METHODS: Six BB and Six untrained controls (CTRL) were recruited. Cross-sectional area (CSA) and maximum voluntary contraction (MVC) of quadriceps femoris muscle (QF) and CSA and architecture of vastus lateralis (VL) were determined. Moreover, a biopsy was taken from VL mid-portion and single fibres were analysed. RESULTS: QF CSA and MVC were 32% (n.s., P = .052) and 58% (P = .009) higher in BB than in CTRL, respectively. VL CSA was 37% higher in BB (P = .030). Fast 2A fibres CSA was 24% (P = .048) greater in BB than in CTRL, when determined in immunostained sections of biopsy samples. Single permeabilized fast fibres CSA was 37% (n.s., P = .052) higher in BB than in CTRL, and their force was slightly higher in BB (n.s.), while specific tension (P0 ) was 19% (P = .024) lower. The lower P0 was not explained either by lower myosin content or by impaired calcium diffusion. Conversely, the swelling caused by skinning-induced permeabilization was different and, when used to correct P0 , differences between populations disappeared. CONCLUSIONS: The results show that high degree of muscle hypertrophy is not detrimental for force generation capacity, as increases in fibre size and force are strictly proportional once the differential swelling response is accounted for.