Neuromuscular Research Articles

Investigating specimen size and shape effects on compressive mechanical behaviors of recycled aggregate concrete using discrete element mesoscale modeling

In comparison to natural aggregate concrete (NAC), recycled aggregate concrete (RAC) typically exhibits a more diverse composition of materials, heightened variability and greater inherent weaknesses. These characteristics may result in varying effects of specimen size and shape on compressive responses. Within this framework, the present study endeavors to comprehensively investigate these influences on RAC's compressive properties and their underlying mechanisms utilizing discrete element simulation technology. At the mesoscale, RAC is depicted by several components, encompassing both new and old aggregates, mortar and interfacial transition zones. Through integrating contacts for each phase, elucidating mesoscale mechanical responses with a parallel bond model and introducing Weibull random fields to delineate variations in phase properties, this study effectively simulates these mechanical responses. Subsequent extensive calculations scrutinize the repercussions of specimen size and shape on the compressive strength, elastic modulus, peak strain and failure modes of RAC across varying water-to-binder ratios. The study concludes that RAC manifests a size effect, albeit less pronounced than in NAC with an equivalent water-to-cement ratio, yet more significant than in mortar. Moreover, owing to end plate constraints, the reduction in RAC's cubic compressive strength is notably less than its prismatic compressive strength. Based on these findings, it is recommended to establish the strength size reduction coefficient and cube-to-cylinder shape coefficient for RAC at 0.86 and 0.72, respectively. Notably, these values diverge significantly from the respective 0.81 and 0.78 coefficients for NAC.

Un caso di botulismo infantile

A 6-month-old infant had been presenting with weak sucking and reduced spontaneous limb movements for a week. He had constipation problems for 4 days followed by hard stool. Upon admission, blood tests including metabolic screening and instrumental investigations were performed. Upon further inquiry into the family history, it was discovered that the father works as a mason. Due to suspicion of infant botulism, the Poison Control Center Unit of Pavia was contacted, and a faecal sample was sent to the Brescia Zooprophylactic Institute, which tested positive for Clostridium bacteria that produce botulinum toxin subtype B. The infant did not require respiratory or nutritional support. Antitoxin therapy was not initiated, and the infant was discharged upon negative test results. Botulism is a neuroparalytic disease characterized by acute symmetric descending flaccid paralysis caused by neurotoxin. The pathogenetic mechanism involves the botulinum toxin inhibiting the formation of the SNARE complex, crucial for the release of neurotransmitters at the neuromuscular junction.

Sunitinib-mediated inhibition of STAT3 in skeletal muscle and spinal cord does not affect the disease in a mouse model of ALS

Variability in disease onset and progression is a hallmark of amyotrophic lateral sclerosis (ALS), both in sporadic and genetic forms. Recently, we found that SOD1-G93A transgenic mice expressing the same amount of mutant SOD1 but with different genetic backgrounds, C57BL/6JOlaHsd and 129S2/SvHsd, show slow and rapid muscle wasting and disease progression, respectively. Here, we investigated the different molecular mechanisms underlying muscle atrophy. Although both strains showed similar denervation-induced degradation of muscle proteins, only the rapidly progressing mice exhibited early and sustained STAT3 activation that preceded atrophy in gastrocnemius muscle. We therefore investigated the therapeutic potential of sunitinib, a tyrosine kinase inhibitor known to inhibit STAT3 and prevent cancer-induced muscle wasting. Although sunitinib treatment reduced STAT3 activation in the gastrocnemius muscle and lumbar spinal cord, it did not preserve spinal motor neurons, improve neuromuscular impairment, muscle atrophy and disease progression in the rapidly progressing SOD1-G93A mice. Thus, the effect of sunitinib is not equally positive in different diseases associated with muscle wasting. Moreover, given the complex role of STAT3 in the peripheral and central compartments of the neuromuscular system, the present study suggests that its broad inhibition may lead to opposing effects, ultimately preventing a potential positive therapeutic action in ALS.

Effects of membrane cholesterol-targeting chemicals on skeletal muscle contractions evoked by direct and indirect stimulation.

Cholesterol is one of the major components of plasma membrane, where its distribution is nonhomogeneous and it participates in lipid raft formation. In skeletal muscle cholesterol and lipid rafts seem to be important for excitation-contraction coupling and for neuromuscular transmission, involving cholesterol-rich synaptic vesicles. In the present study, nerve and muscle stimulation-evoked contractions were recorded to assess the role of cholesterol in contractile function of mouse diaphragm. Exposure to cholesterol oxidase (0.2 U/ml) and cholesterol-depleting agent methyl-β-cyclodextrin (1 mM) did not affect markedly contractile responses to both direct and indirect stimulation at low and high frequency. However, methyl-β-cyclodextrin at high concentration (10 mM) strongly decreased the force of both single and tetanus contractions induced by phrenic nerve stimulation. This decline in contractile function was more profoundly expressed when methyl-β-cyclodextrin application was combined with phrenic nerve activation. At the same time, 10 mM methyl-β-cyclodextrin had no effect on contractions upon direct muscle stimulation at low and high frequency. Thus, strong cholesterol depletion suppresses contractile function mainly due to disturbance of the neuromuscular communication, whereas muscle fiber contractility remains resistant to decline.

Spinal Accessory Nerve to Suprascapular Nerve Transfer by Dorsal Approach for Shoulder Reanimation in Cases of Brachial Plexus Injuries: Surgical Technique

AbstractDistal nerve transfers are frequently used for the treatment of patients having brachial plexus injuries. Spinal accessory nerve to suprascapular nerve transfer by dorsal approach is advantageous since it brings the site of neurotization closer to the neuromuscular end plate and minimizes donor muscle weakness. This surgery is found to be challenging owing to the small and deep operative field. In this article, the authors describe the surgical technique for this procedure.

Mis-localization of endogenous TDP-43 leads to ALS-like early-stage metabolic dysfunction and progressive motor deficits

BackgroundThe key pathological signature of ALS/ FTLD is the mis-localization of endogenous TDP-43 from the nucleus to the cytoplasm. However, TDP-43 gain of function in the cytoplasm is still poorly understood since TDP-43 animal models recapitulating mis-localization of endogenous TDP-43 from the nucleus to the cytoplasm are missing.MethodsCRISPR/Cas9 technology was used to generate a zebrafish line (called CytoTDP), that mis-locates endogenous TDP-43 from the nucleus to the cytoplasm. Phenotypic characterization of motor neurons and the neuromuscular junction was performed by immunostaining, microglia were immunohistochemically localized by whole-mount tissue clearing and muscle ultrastructure was analyzed by scanning electron microscopy. Behavior was investigated by video tracking and quantitative analysis of swimming parameters. RNA sequencing was used to identify mis-regulated pathways with validation by molecular analysis.ResultsCytoTDP fish have early larval phenotypes resembling clinical features of ALS such as progressive motor defects, neurodegeneration and muscle atrophy. Taking advantage of zebrafish’s embryonic development that solely relys on yolk usage until 5 days post fertilization, we demonstrated that microglia proliferation and activation in the hypothalamus is independent from food intake. By comparing CytoTDP to a previously generated TDP-43 knockout line, transcriptomic analyses revealed that mis-localization of endogenous TDP-43, rather than TDP-43 nuclear loss of function, leads to early onset metabolic dysfunction.ConclusionsThe new TDP-43 model mimics the ALS/FTLD hallmark of progressive motor dysfunction. Our results suggest that functional deficits of the hypothalamus, the metabolic regulatory center, might be the primary cause of weight loss in ALS patients. Cytoplasmic gain of function of endogenous TDP-43 leads to metabolic dysfunction in vivo that are reminiscent of early ALS clinical non-motor metabolic alterations. Thus, the CytoTDP zebrafish model offers a unique opportunity to identify mis-regulated targets for therapeutic intervention early in disease progression.

The effects of nanoplastics and microcystin-LR coexposure on Aristichthys nobilis at the early developmental stages

Nanoplastics (NPs) and microcystin-LR (MC-LR) are two common and harmful pollutants in water environments, especially at aquafarm where are full of plastic products and algae. It is of great significance to study the toxic effects and mechanisms of the NPs and/or MC-LR on fish at the early stage. In this study, the embryo and larvae of a filtering-feeding fish, Aristichthys nobilis, were used as the research objects. The results showed that the survival and hatching rates of the embryo were not significantly affected by the environmental concentration exposure of these two pollutants. Scanning electron microscopy (SEM) observation displayed that NPs adhered to the surface of the embryo membrane. Transcriptomic and bioinformatic analyses revealed that the NPs exposure activated neuromuscular junction development and skeletal muscle fiber in larvae, and affected C5-Branched dibasic acid metabolism. The metabolic and biosynthetic processes of zeaxanthin, xanthophyll, tetraterpenoid, and carotenoid were suppressed after the MC-LR exposure, which was harmful to the retinol metabolism of fish. Excessive production of superoxide dismutase (SOD) was detected under the MC-LR exposure. The MC-LR and NPs coexposure triggered primary immunodeficiency and adaptive immune response, leading to the possibility of reduced fitness of A.nobilis during the development. Collectively, our results indicate that environmental concentration NPs and MC-LR coexposure could cause toxic damage and enhance sick risk in A.nobilis, providing new insights into the risk of NPs and MC-LR on filtering-feeding fish.

A training goal-oriented categorization model of high-intensity interval training.

There are various categorization models of high-intensity interval training (HIIT) in the literature that need to be more consistent in definition, terminology, and concept completeness. In this review, we present a training goal-oriented categorization model of HIIT, aiming to find the best possible consensus among the various defined types of HIIT. This categorization concludes with six different types of HIIT derived from the literature, based on the interaction of interval duration, interval intensity and interval:recovery ratio. We discuss the science behind the defined types of HIIT and shed light on the possible effects of the various types of HIIT on aerobic, anaerobic, and neuromuscular systems and possible transfer effects into competition performance. We highlight various research gaps, discrepancies in findings and not yet proved know-how based on a lack of randomized controlled training studies, especially in well-trained to elite athlete cohorts. Our HIIT "toolbox" approach is designed to guide goal-oriented training. It is intended to lay the groundwork for future systematic reviews and serves as foundation for meta-analyses.

Myasthenia gravis: the future is here.

Myasthenia gravis (MG) stands as a prototypical antibody-mediated autoimmune disease: it is dependent on T cells and characterized by the presence of autoantibodies targeting proteins located on the postsynaptic surface of skeletal muscle, known as the neuromuscular junction. Patients with MG exhibit a spectrum of weakness, ranging from limited ocular muscle involvement to life-threatening respiratory failure. Recent decades have witnessed substantial progress in understanding the underlying pathophysiology, leading to the delineation of distinct subcategories within MG, including MG linked to AChR or MuSK antibodies as well as age-based distinction, thymoma-associated, and immune checkpoint inhibitor-induced MG. This heightened understanding has paved the way for the development of more precise and targeted therapeutic interventions. Notably, the FDA has recently approved therapeutic inhibitors of complement and the IgG receptor FcRn, a testament to our improved comprehension of autoantibody effector mechanisms in MG. In this Review, we delve into the various subgroups of MG, stratified by age, autoantibody type, and histology of the thymus with neoplasms. Furthermore, we explore both current and potential emerging therapeutic strategies, shedding light on the evolving landscape of MG treatment.

Tensile behavior of novel column-to-column connection for modular steel construction using high-strength bolts

As an important aspect of the city upgrade plan, installing the elevators for existing multi-story residential buildings means a lot to the residents of the higher floors, especially for old people. To minimize the site disruption and overcome the space limitation of installation, the modular elevator shafts have been widely adopted. The external wall decoration panels are normally pre-installed on the elevator shaft frame elements in the workshop, which makes it difficult for the connection between upper and lower segments using the high-strength bolts due to very limited space for applying the pre-tension. A novel column-to-column connection is presented in this study, consisting of an end plate, web opening of column web and strengthening plate at each of the column end. The tensile behavior of the proposed connection is first investigated through an experiment. Subsequently, a detailed finite element model is built and verified using the test results, with which a parametric study is conducted with the main parameters considered. Finally, a simple analytical solution for predicting the design load of the proposed connection is developed employing the yield line theory. Comparisons with the finite element results indicate the high accuracy of the proposed solution. The presented connection is believed to be a good solution for the on-site connection between H-columns of highly assembled modular structures, not limited to the elevator shafts.

Free essential amino acid feeding improves endurance during resistance training via DRP1-dependent mitochondrial remodelling.

Loss of muscle strength and endurance with aging or in various conditions negatively affects quality of life. Resistance exercise training (RET) is the most powerful means to improve muscle mass and strength, but it does not generally lead to improvements in endurance capacity. Free essential amino acids (EAAs) act as precursors and stimuli for synthesis of both mitochondrial and myofibrillar proteins that could potentially confer endurance and strength gains. Thus, we hypothesized that daily consumption of a dietary supplement of nine free EAAs with RET improves endurance in addition to the strength gains by RET. Male C57BL6J mice (9weeks old) were assigned to control (CON), EAA, RET (ladder climbing, 3 times a week), or combined treatment of EAA and RET (EAA+RET) groups. Physical functions focusing on strength or endurance were assessed before and after the interventions. Several analyses were performed to gain better insight into the mechanisms by which muscle function was improved. We determined cumulative rates of myofibrillar and mitochondrial protein synthesis using 2H2O labelling and mass spectrometry; assessed ex vivo contractile properties and in vitro mitochondrial function, evaluated neuromuscular junction (NMJ) stability, and assessed implicated molecular singling pathways. Furthermore, whole-body and muscle insulin sensitivity along with glucose metabolism, were evaluated using a hyperinsulinaemic-euglycaemic clamp. EAA+RET increased muscle mass (10%, P<0.05) and strength (6%, P<0.05) more than RET alone, due to an enhanced rate of integrated muscle protein synthesis (19%, P<0.05) with concomitant activation of Akt1/mTORC1 signalling. Muscle quality (muscle strength normalized to mass) was improved by RET (i.e., RET and EAA+RET) compared with sedentary groups (10%, P<0.05), which was associated with increased AchR cluster size and MuSK activation (P<0.05). EAA+RET also increased endurance capacity more than RET alone (26%, P<0.05) by increasing both mitochondrial protein synthesis (53%, P<0.05) and DRP1 activation (P<0.05). Maximal respiratory capacity increased (P<0.05) through activation of the mTORC1-DRP1 signalling axis. These favourable effects were accompanied by an improvement in basal glucose metabolism (i.e., blood glucose concentrations and endogenous glucose production vs. CON, P<0.05). Combined treatment with balanced free EAAs and RET may effectively promote endurance capacity as well as muscle strength through increased muscle protein synthesis, improved NMJ stability, and enhanced mitochondrial dynamics via mTORC1-DRP1 axis activation, ultimately leading to improved basal glucose metabolism.

Re-assessment of the subcellular localization of Bazooka/Par-3 in Drosophila: no evidence for localization to the nucleus and the neuromuscular junction.

Bazooka/Par-3 (Baz) is an evolutionarily conserved scaffold protein that functions as a master regulator for the establishment and maintenance of cell polarity in many different cell types. In the vast majority of published research papers Baz has been reported to localize at the cell cortex and at intercellular junctions. However, there have also been several reports showing localization and function of Baz at additional subcellular sites, in particular the nuclear envelope and the neuromuscular junction. In this study we have re-assessed the localization of Baz to these subcellular sites in a systematic manner. We used antibodies raised in different host animals against different epitopes of Baz for confocal imaging of Drosophila tissues. We tested the specificity of these antisera by mosaic analysis with null mutant baz alleles and tissue-specific RNAi against baz. In addition, we used a GFP-tagged gene trap line for Baz and a bacterial artificial chromosome (BAC) expressing GFP-tagged Baz under control of its endogenous promoter in a baz mutant background to compare the subcellular localization of the GFP-Baz fusion proteins to the staining with anti-Baz antisera. Together, these experiments did not provide evidence for specific localization of Baz to the nucleus or the neuromuscular junction.

The droopy eyed toddler – A rare case of juvenile myasthenia gravis

Myasthenia Gravis is a Neuromuscular Junction disorder seen in children. It can have 3 presentations. Transient neonatal form, Immune mediated or Juvenile form and congenital myasthenia variety possibly due to genetic mutation. Amongst these variants Immune mediated form is most common type. It comprises 10% of all Myasthenia Gravis cases. Myasthenia can have myriad of clinical presentations. Our case is of a 18 months old toddler who had drooping of eyes which started insidiously 1 month back.

Behavior of jittering potential before and after impulse blockings: a preliminary study in myasthenia gravis

Neuromuscular junction disorders lead to secession of bioelectrical activity transmission between motor nerve endings and muscle fibers. In diseases that are severe enough, impulse blockings are observed. This study aims to reveal the behavior of neuromuscular junction before and after impulse blockings. Fourteen recordings harboring impulse blockings from nine myasthenia gravis (MG) patients were included. Recordings were made from frontalis muscle by using concentric needle electrode during voluntary contraction. One hundred traces were acquired in each session. In addition to well-known jitter parameters, new parameters were calculated such as number of consecutive impulse blocking groups, number of impulse blockings in each group, ratio of maximum number of consecutive impulse blockings to all number of blockings. Graphics were composed to show location change behavior of jittering potential in all traces. For jittering potential, before or after a single impulse blocking the amount of getting further away from trigger peak was greater than getting closer to trigger peak. However, after consecutive impulse blockings the amount of getting closer to trigger peak was greater than the amount of getting further away. The behavior of neuromuscular junction before and after impulse blockings was demonstrated in MG patients. Moreover, new features were extracted for jitter studies. Building models for different diseases according to their impulse blockings may be possible with the developed algorithm.Graphical abstract

The Neuromuscular Disorder Mediated by Extracellular Vesicles in Amyotrophic Lateral Sclerosis.

Amyotrophic lateral sclerosis (ALS) represents a neurodegenerative disorder characterized by the progressive loss of both upper and lower motor neurons, resulting in muscular atrophy and eventual paralysis. While much research has concentrated on investigating the impact of major mutations associated with ALS on motor neurons and central nervous system (CNS) cells, recent studies have unveiled that ALS pathogenesis extends beyond CNS imbalances, encompassing dysregulation in other tissues such as skeletal muscle. Evidence from animal models and patients supports this broader perspective. Skeletal muscle, once considered solely as an effector organ, is now recognized as possessing significant secretory activity capable of influencing motor neuron survival. However, the precise cellular and molecular mechanisms underlying the detrimental effects observed in muscle and its associated structures in ALS remain poorly understood. Additionally, emerging data suggest that extracellular vesicles (EVs) may play a role in the establishment and function of the neuromuscular junction (NMJ) under both physiological and pathological conditions and in wasting and regeneration of skeletal muscles, particularly in neurodegenerative diseases like ALS. This review aims to explore the key findings about skeletal muscle involvement in ALS, shedding light on the potential underlying mechanisms and contributions of EVs and their possible application for the design of biosensors.

The influence of cortical end plate upon ultrasound transit time spectroscopy estimated bone assessment

Quantitative ultrasound (QUS) is considered a reliable tool for routine assessment of bone. However, none of the QUS parameters has a direct measure of bone quantity or quality. The concept of ultrasound transit time spectroscopy (UTTS) has recently offered the possibility of effective estimates of bone volume fraction (BVF) as an indicator of bone density. This study aimed to investigate the influence of cortical end plate thickness around the cancellous bone on the measurement accuracy of UTTS solid volume fraction (UTTS-SVF). Two categories of cortical discs were designed and three-dimensional printed; category (I) is planer discs of constant different thickness and category (II) is variable-thickness step-wedge discs. In this experimental and simulated study, the discs were placed coaxially with the 3D-printed cylindrical iliac crest cancellous bone sample. A through-transmission 1 MHz ultrasound pulses were recorded, from which the deconvoluted TTS were derived. Then UTTS-SVF was determined and compared with the measured SVF via microcomputed tomography (μCT-SVF) showing a high agreement with an average of 94% ± 3.7% and 95% ± 4% for the planar cortical discs and 95.3% ± 5.5% and 96.5% ± 3.7% for variable-thickness step-wedge cortical discs for experiment and simulation respectively. The variations between the derived UTTS_SVF and the standard μCT-SVF are assumed to be due to the existence of phase interference within the cortical discs. Hence, the cortical end plate creates a negative influence on the derived UTTS-SVF.

Immunohistochemistry and Morphometric Analysis of Drosophila Larval Body Wall Neuromuscular Junction Preparations.

The Drosophila neuromuscular junction (NMJ) is an excellent model for studying vertebrate glutamatergic synapses. Researchers have uncovered fundamental mechanisms at the fly NMJ that are conserved in higher-order organisms. To gain molecular and structural insight into these and other structures, immunolabeling is invaluable. In this protocol, we describe how to use immunolabeling to visualize embryonic/larval presynaptic and postsynaptic structures at the NMJ. We also include details about amplification of weak immunohistochemistry signals and how to use these signals to quantify synaptic growth via bouton counting. Boutons are bead-like structures at motor axon terminals that house synapses, and the number of boutons reflects the size of the NMJ. We also describe how to identify the different bouton types.

The Drosophila Larval Neuromuscular Junction: Developmental Overview.

For decades, the Drosophila larval neuromuscular junction (NMJ) has been a go-to model for synaptic development. This simple, accessible system is composed of a repeating pattern of 33 distinct neurons that stereotypically innervate 30 muscles. Fundamental mechanisms that underlie diverse aspects of axon pathfinding, synaptic form, and function have been uncovered at the NMJ, and new pathways continue to be uncovered. These discoveries are fueled by the ease of dissections and an extensive array of techniques. Chief among these techniques are various microscopy approaches, including super-resolution and electron microscopy. Functionally, the Drosophila NMJ is glutamatergic, similar to the vertebrate central synapses, making it a great model to study normal development and neurological diseases. Here we provide a brief overview of the larval neuromuscular system, highlighting the connectivity patterns, development, and some of the mechanisms underlying these processes.

Using the Proximity Ligation Assay to Visualize Colocalization of Proteins at the Drosophila Larval Neuromuscular Junction.

In the nearly 50 years since the neuromuscular junction (NMJ) was first established as a model synapse, its molecular composition has been extensively characterized. Early work relied on fluorescent signals to determine whether proteins localized to the pre- and postsynaptic regions. As more synaptic molecules were identified, determining the localization of these proteins relative to each other became important. Conventional microscopy lacks the resolving power to assess whether two proteins are within an appropriate distance to bind directly or be part of a larger complex. Super-resolution and immunoelectron microscopies can improve spatial resolution, but these techniques can be difficult to execute and troubleshoot, and access to these instruments is limiting. However, another approach, proximity labeling, overcomes many of these limitations by using a DNA secondary label that can only be amplified if the two proteins of interest are within 40 nm of each other, which is ∼5× greater than the resolving power of conventional microscopy. In this protocol, we describe the use of the proximity ligation assay, which combines immunohistochemistry with DNA amplification, to reveal protein colocalization in the Drosophila NMJ.

A case of SBMA presenting with myasthenic syndrome limited to a dropped head

AbstractA 76‐year‐old man with spinal and bulbar muscular atrophy (SBMA) developed an acute course of the dropped head with diurnal fluctuations that worsened in the evening. He was diagnosed with myasthenic syndrome based on a decreased response to repeated stimulation, as well as increased jitter and blocking on single‐fiber electromyography, indicating disturbed neuromuscular transmission. Dropped head symptom was successfully alleviated and maintained by immunotherapy. Although limited reports are available on SBMA cases accompanied by myasthenic syndrome and on the clinical impact of impaired neuromuscular transmission on SBMA, myasthenic syndrome may be a common condition and even a part of the phenotypic spectrum of SBMA. To our knowledge, this is the first case of SBMA comorbid with myasthenic syndrome limited to the dropped head.