Loss Of Muscle Cells Research Articles

Physiological mechanisms of neuromuscular impairment in diabetes-related complications: Can physical exercise help prevent it?

Diabetes mellitus is a chronic disorder that progressively induces complications, compromising daily independence. Among these, diabetic neuropathy is particularly prevalent and contributes to substantial neuromuscular impairments in both types 1 and 2 diabetes. This condition leads to structural damage affecting both the central and peripheral nervous systems, resulting in a significant decline in sensorimotor functions. Alongside neuropathy, diabetic myopathy also contributes to muscle impairment and reduced motor performance, intensifying the neuromuscular decline. Diabetic neuropathy typically implicates neurogenic muscle atrophy, motoneuron loss and clustering of muscle fibres as a result of aberrant denervation-reinervation processes. These complications are associated with compromised neuromuscular junctions, where alterations occur in pre-synaptic vesicles, mitochondrial content and post-synaptic signalling. Neural damage is intensified by chronic hyperglycaemia and oxidative stress, exacerbating vascular dysfunction and reducing oxygen delivery. These complications imply a severe decline in neuromuscular performance, evidenced by reductions in maximal force and power output, rate of force development and muscle endurance. Furthermore, diabetes-related complications are compounded by age-related degenerative changes in long-term patients. Aerobic and resistance training offer promising approaches for managing blood glucose levels and neuromuscular function. Aerobic exercise promotes mitochondrial biogenesis and angiogenesis, supporting metabolic and cardiovascular health. Resistance training primarily enhances neural plasticity, muscle strength and hypertrophy, which are crucial factors for mitigating sarcopenia and preserving functional independence. This topical review examines current evidence on the physiological mechanisms underlying diabetic neuropathy and the potential impact of physical activity in counteracting this decline.

FAPs orchestrate homeostasis of muscle physiology andpathophysiology.

As a common clinical manifestation, muscle weakness is prevalent in people with mobility disorders. Further studies of muscle weakness have found that patients with muscle weakness present with persistent muscle inflammation, loss of muscle fibers, fat infiltration, and interstitial fibrosis. Therefore, we propose the concept of muscle microenvironment homeostasis, which explains the abnormal pathological changes in muscles through the imbalance of muscle microenvironment homeostasis. And we identified an interstitial progenitor cell FAP during the transition from normal muscle microenvironment homeostasis to muscle microenvironment imbalance caused by muscle damage diseases. As a kind of pluripotent stem cell, FAPs do not participate in myogenic differentiation, but can differentiate into fibroblasts, adipocytes, osteoblasts, and chondrocytes. As a kind of mesenchymal progenitor cell, it is involved in the generation of extracellular matrix, regulate muscle regeneration, and maintain neuromuscular junction. However, the muscle microenvironment is disrupted by the causative factors, and the abnormal activities of FAPs eventually contribute to the complex pathological changes in muscles. Targeting the mechanisms of these muscle pathological changes, we have identified appropriate signaling targets for FAPs to improve and even treat muscle damage diseases. In this review, we propose the construction of muscle microenvironmental homeostasis and find the key cells that cause pathological changes in muscle after homeostasis is broken. By studying the mechanism of abnormal differentiation and apoptosis of FAPs, we found a strategy to inhibit the abnormal pathological changes in muscle damage diseases and improve muscle regeneration.

Protein-extending ACTN2 frameshift variants cause variable myopathy phenotypes by protein aggregation.

The objective of the study is to characterize the pathomechanisms underlying actininopathies. Distal myopathies are a group of rare, inherited muscular disorders characterized by progressive loss of muscle fibers that begin in the distal parts of arms and legs. Recently, variants in a new disease gene, ACTN2, have been shown to cause distal myopathy. ACTN2, a gene previously only associated with cardiomyopathies, encodes alpha-actinin-2, a protein expressed in both cardiac and skeletal sarcomeres. The primary function of alpha-actinin-2 is to link actin and titin to the sarcomere Z-disk. New ACTN2 variants are continuously discovered; however, the clinical significance of many variants remains unknown. Thus, lack of clear genotype-phenotype correlations in ACTN2-related diseases, actininopathies, persists. Functional characterization in C2C12 cell model of several ACTN2 variants is conducted, including frameshift and missense variants associated with dominant and recessive actininopathies. We assess the genotype-phenotype correlations of actininopathies using clinical data from several patients carrying these variants. The results show that the missense variants associated with a recessive form of actininopathy do not cause detectable alpha-actinin-2 aggregates in the cell model. Conversely, dominant frameshift variants causing a protein extension do form alpha-actinin-2 aggregates. The results suggest that alpha-actinin-2 aggregation is the disease mechanism underlying some dominant actininopathies, and thus, we recommend that protein-extending frameshift variants in ACTN2 should be classified as pathogenic. However, this mechanism is likely elicited by only a limited number of variants. Alternative functional characterization methods should be explored to further investigate other molecular mechanisms underlying actininopathies.

Frailty: Focus on Regenerative Medicine

Frailty, as a specific condition of increased vulnerability and deterioration of general health associated with aging in the elderly, is an emerging problem worldwide with serious implications for clinical practice and public health. Frailty increases the risk of falls, hospitalization, disability, and death. Muscle aging is associated with progressive loss of skeletal muscle mass and function. Loss of muscle mass with age can be related to muscle fiber atrophy and loss of muscle fibers. With increasing human age, remodeling of bone tissue becomes more prone to slow and steady loss of bone mass, which is primarily associated with loss of bone formation by osteoblasts. Recent preclinical and clinical studies have confirmed the safety of mesenchymal stem/stromal cells (MSCs) in the treatment of frailty. MSCs are attracted to sites of damage where they act to reduce inflammation and promote cell repair. Thus, there is an assumption that frailty can be treated with cell therapy

Sarcopenia as a non-motor symptom of Parkinson's disease

Sarcopenia is a progressive generalized skeletal muscle disease that is accompanied by an accelerated loss of muscle mass and function, affecting the quality of life and the ability to perform self-care. The prevalence of sarcopenia in the world today ranges from 10 to 25%, which represents a certain danger as it is a prognostic factor for possible injury and increased disability in the elderly population. Sarcopenia often accompanies a large number of different diseases, including neurodegenerative ones, so it is actively studied in this category of patients, for example, as one of the early symptoms of Parkinson's disease (PD). PD and sarcopenia have overlapping pathophysiological mechanisms of muscle fiber loss: inflammation, muscle autophagy, oxidative stress and apoptosis. Loss of muscle mass due to malnutrition is common in PD. According to some studies, the prevalence of sarcopenia in PD varies from 6 to 55.8%; weakness and sarcopenia are more common in patients with PD than in society as a whole, which is associated with an unfavorable course of the disease. The presence of both diseases simultaneously in one patient can impose certain restrictions on the treatment of the patient, worsen his physical and mental condition, which determines the need for early detection of sarcopenia in patients with PD.

Lifelong dietary protein restriction accelerates skeletal muscle loss and reduces muscle fibre size by impairing proteostasis and mitochondrial homeostasis

The early life environment significantly affects the development of age-related skeletal muscle disorders. However, the long-term effects of lactational protein restriction on skeletal muscle are still poorly defined. Our study revealed that male mice nursed by dams fed a low-protein diet during lactation exhibited skeletal muscle growth restriction. This was associated with a dysregulation in the expression levels of genes related to the ribosome, mitochondria and skeletal muscle development. We reported that lifelong protein restriction accelerated loss of type-IIa muscle fibres and reduced muscle fibre size by impairing mitochondrial homeostasis and proteostasis at 18 months of age. However, feeding a normal-protein diet following lactational protein restriction prevented accelerated fibre loss and fibre size reduction in later life. These findings provide novel insight into the mechanisms by which lactational protein restriction hinders skeletal muscle growth and includes evidence that lifelong dietary protein restriction accelerated skeletal muscle loss in later life.

Redox control of signalling responses to contractile activity and aging in skeletal muscle

During aging loss of skeletal muscle mass and function has a significant effect of an individual’s quality of life and ability to maintain independence. Both loss of muscle fibres and atrophy of the remaining fibres play a role in the muscle decline and this is associated with loss of motor units and a reduction in the number of motor neurons. Increased oxidative damage has long been claimed to be associated with aging and many studies have reported increased amounts of oxidative damage markers are found in tissues from old organisms. Reactive oxygen species (ROS) are recognised to play a major role in cell signalling and in muscle ROS generated during contractile play an important role in signalling adaptations to contractile activity. These ’redox-regulated’ pathways are beneficial adaptations which are attenuated during aging. This review will briefly cover what is currently known about the mechanisms underlying these muscle adaptations to exercise, how they are affected by aging and assess the importance of these pathways in age-related loss of skeletal muscle mass and function.

Biomarkers for Duchenne muscular dystrophy progression: impact of age in the mdx tongue spared muscle

BackgroundDuchenne muscular dystrophy (DMD) is a severe form of muscular dystrophy without an effective treatment, caused by mutations in the DMD gene, leading to the absence of dystrophin. DMD results in muscle weakness, loss of ambulation, and death at an early age. Metabolomics studies in mdx mice, the most used model for DMD, reveal changes in metabolites associated with muscle degeneration and aging. In DMD, the tongue muscles exhibit unique behavior, initially showing partial protection against inflammation but later experiencing fibrosis and loss of muscle fibers. Certain metabolites and proteins, like TNF-α and TGF-β, are potential biomarkers for dystrophic muscle characterization.MethodsTo investigate disease progression and aging, we utilized young (1 month old) and old (21–25 months old) mdx and wild-type tongue muscles. Metabolite changes were analyzed using 1H nuclear magnetic resonance, while TNF-α and TGF-β were assessed using Western blotting to examine inflammation and fibrosis. Morphometric analysis was conducted to assess the extent of myofiber damage between groups.ResultsThe histological analysis of the mid-belly tongue showed no differences between groups. No differences were found between the concentrations of metabolites from wild-type or mdx whole tongues of the same age. The metabolites alanine, methionine, and 3-methylhistidine were higher, and taurine and glycerol were lower in young tongues in both wild type and mdx (p < 0.001). The metabolites glycine (p < 0.001) and glutamic acid (p = 0.0018) were different only in the mdx groups, being higher in young mdx mice. Acetic acid, phosphocreatine, isoleucine, succinic acid, creatine, and the proteins TNF-α and TGF-β had no difference in the analysis between groups (p > 0.05).ConclusionsSurprisingly, histological, metabolite, and protein analysis reveal that the tongue of old mdx remains partially spared from the severe myonecrosis observed in other muscles. The metabolites alanine, methionine, 3-methylhistidine, taurine, and glycerol may be effective for specific assessments, although their use for disease progression monitoring should be cautious due to age-related changes in the tongue muscle. Acetic acid, phosphocreatine, isoleucine, succinate, creatine, TNF-α, and TGF-β do not vary with aging and remain constant in spared muscles, suggesting their potential as specific biomarkers for DMD progression independent of aging.

Decoding the transcriptome of Duchenne muscular dystrophy to the single nuclei level reveals clinical-genetic correlations

Duchenne muscular dystrophy is a genetic disease produced by mutations in the dystrophin gene characterized by early onset muscle weakness leading to severe and irreversible disability. The cellular and molecular consequences of the lack of dystrophin in humans are only partially known, which is crucial for the development of new therapies aiming to slow or stop the progression of the disease. Here we have analyzed quadriceps muscle biopsies of seven DMD patients aged 2 to 4 years old and five age and gender matched controls using single nuclei RNA sequencing (snRNAseq) and correlated the results obtained with clinical data. SnRNAseq identified significant differences in the proportion of cell population present in the muscle samples, including an increase in the number of regenerative fibers, satellite cells, and fibro-adipogenic progenitor cells (FAPs) and a decrease in the number of slow fibers and smooth muscle cells. Muscle samples from the younger patients with stable mild weakness were characterized by an increase in regenerative fibers, while older patients with moderate and progressive weakness were characterized by loss of muscle fibers and an increase in FAPs. An analysis of the gene expression profile in muscle fibers identified a strong regenerative signature in DMD samples characterized by the upregulation of genes involved in myogenesis and muscle hypertrophy. In the case of FAPs, we observed upregulation of genes involved in the extracellular matrix regeneration but also several signaling pathways. Indeed, further analysis of the potential intercellular communication profile showed a dysregulation of the communication profile in DMD samples identifying FAPs as a key regulator of cell signaling in DMD muscle samples. In conclusion, our study has identified significant differences at the cellular and molecular levels in the different cell populations present in skeletal muscle samples of patients with DMD compared to controls.

Signal transducer and activator of transcription 6 mediates skeletal muscle cell injury in septic mice by regulating ferroptosis

To explore the effect of signal transducer and activator of transcription 6 (STAT6) on ferroptosis in skeletal muscle cells in sepsis model and its potential mechanism. Twenty-four 8-week-old male specific pathogen free Kunming mice were divided into normal control group, sham group, sepsis model group and STAT6 inhibitor pretreatment group according to random number table method with 6 mice in each group. A mouse sepsis model was reproduced by cecal ligation and perforation (CLP). In the sham group, the skin of mice was sutured after exposing the cecum tissue. In the STAT6 inhibitor pretreatment group, 10 mg/kg AS1517499 was injected intraperitoneally 1 hour before model reproduction. The sham group and the model group were intraperitoneally injected with the same volume of normal saline. Mice in the normal control group did not receive any operation or drug intervention. The mice were sacrificed 24 hours after model reproduction, and the muscle tissue of hind limb was obtained under sterile condition. Hematoxylin-eosin (HE) staining was used to observe the histopathology with optical microscope, and mitochondrial morphological changes were observed by transmission electron microscopy after double staining with uranium acetate lead citrate. The ferroptosis marker proteins expressions of chitinase-3-like protein 1 (CHI3L1), cyclooxygenase-2 (COX-2), acyl-CoA synthetase long-chain family member 4 (ACSL4), ferritin heavy chain 1 (FTH1), and glutathione peroxidase 4 (GPx4) were detected by Western blotting. Under the optical microscope, the morphology and structure of skeletal muscle tissues in the normal control and sham groups were normal. In the model group, the structure of skeletal muscle tissues was loose, the muscle fiber became smaller and atrophic, inflammatory cell infiltration and even muscle fiber loss were found. Compared with the model group, the structure of skeletal muscle tissues was tight and skeletal muscle atrophy was improved in the STAT6 inhibitor pretreatment group. The ultrastructure of skeletal muscle cell in the normal control and sham groups was normal under transmission electron microscope. The ultrastructure characteristics of skeletal muscle in the model group showed that cell membrane was broken and blister, mitochondria became smaller and membrane density increased, the mitochondrial crista decreased or disappeared, the mitochondrial outer membrane was broken, and the nucleus was normal in size but lacked chromatin condensation. Compared with the model group, the STAT6 inhibitor pretreatment group had a significant improvement in the ultrastructure of muscle cells. Compared with the normal control and sham groups, the protein expressions of CHI3L1, COX-2, ACSL4 and FTH1 in the muscle of the model group were significantly increased, while the protein expression of GPx4 was decreased significantly, indicating that the skeletal muscle cells in the mouse sepsis model showed characteristic mitochondrial injury and abnormal expression of ferroptosis markers. Compared with the model group, the protein expressions of CHI3LI, COX-2, ACSL4 and FTH1 in the STAT6 inhibitor pretreatment group were significantly decreased [CHI3L1 protein (CHI3L1/GAPDH): 0.70±0.08 vs. 0.97±0.09, COX-2 protein (COX-2/GAPDH): 0.61±0.03 vs. 0.83±0.03, ACSL4 protein (ACSL4/GAPDH): 0.75±0.04 vs. 1.02±0.16, FTH1 protein (FTH1/GAPDH): 0.49±0.06 vs. 0.76±0.13, all P < 0.05], while the protein expression of GPx4 was significantly increased (GPx4/GAPDH: 1.14±0.29 vs. 0.53±0.03, P < 0.05). Sepsis can induce ferroptosis in skeletal muscle cells of mice. STAT6 may mediate ferroptosis in mouse skeletal muscle cells by regulating the expressions of COX-2, ACSL4, FTH1 and GPx4, thereby inducing skeletal muscle cell injury in sepsis.

Age-related alterations in human motor units properties and in muscle innervation profile

Introduction: Mounting evidence suggests that motoneuron and neuromuscular junction (NMJ) degeneration, muscle fiber denervation and loss of motor units contribute to muscle wasting and weakness in old age. However, these aspects are still poorly investigated in humans. This study aimed at comparing the neuromuscular system integrity and function of young and older adults, with particular focus on NMJ and motor unit potential (MUP) characteristics. We hypothesized that older individuals would present altered electrophysiological properties and innervation profile. Methods: We recruited 55 healthy young individuals (YI) (aged 26.1±4.5 yr; 50% females) and 88 older individuals (OI) (aged 75.9±4.7 yr; 55% females). Muscle force was evaluated by isometric dynamometry, muscle morphology by ultrasound and neuromuscular excitability by compound motor unit action potential (CMAP). MUP properties and motor unit number estimate (iMUNE) were obtained by intramuscular electromyography (iEMG). In a sub-cohort of individuals (n=16 YI and n=34 OI), vastus lateralis (VL) muscle biopsies were obtained. We assessed denervation status by scoring neural cell adhesion molecule (NCAM) positive fibers. Generalized linear mixed models were carried out for iEMG data, while unpaired t-tests were carried out for all the other parameters. Results: OI showed alterations in muscle morphology when compared to YI, evident as smaller whole quadriceps cross-sectional area (p&lt;0.001) and VL fiber diameter (p&lt;0.01), together with smaller VL pennation angles (p&lt;0.001) and fascicle lengths (p&lt;0.05), resulting in an increased ultrasound sarcopenia index (p=0.001). Muscle force (p&lt;0.001) and CMAP (p&lt;0.001) were also largely impaired in OI, suggesting that muscle function and neuromuscular excitability are considerably affected by aging. iEMG analysis revealed changes in MUP turns (p&lt;0.001), near fiber (NF) MUP duration (p&lt;0.001) and NF count (p&lt;0.001) in OI, pointing towards an increased MUP complexity, in absence of sex differences. The iMUNE was reduced (p&lt;0.001) and the NMJ transmission stability was significantly altered in OI, as suggested by elevated NF jiggle (p&lt;0.001) and segmented jitter (p&lt;0.05). These electrophysiological alterations seem likely due to altered muscle innervation profile, as highlighted by an increased percentage of NCAM-positive fiber (p&lt;0.001). Conclusions: This study showed that changes in muscle morphology and function, MUP properties, motor units loss and denervation status occur in older humans; these may be serve as useful markers of human neuromuscular aging. The present work was funded by PRIN project NeuAge (2017CBF8NJ_001) to MVN and the Milky Way Foundation, the Baxter Foundation, and the Li Ka Shing Foundation to HMB. This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Older mice show decreased regeneration of neuromuscular junctions following lengthening contraction-induced injury.

Progressive muscle atrophy and loss of muscle strength associated with old age have been well documented. Although age-associated impairments in skeletal muscle regeneration following injury have been demonstrated, less is known about whether aging impacts the regenerative response of neuromuscular junctions (NMJ) following contraction-induced injury. Reduced ability of NMJs to regenerate could lead to increased numbers of denervated muscle fibers and therefore play a contributing role to age-related sarcopenia. To investigate the relationship between age and NMJ regeneration following injury, extensor digitorum longus (EDL) muscles of middle-aged (18-19 months) and old mice (27-28 months) were subjected to a protocol of lengthening contractions (LC) that resulted in an acute force deficit of ~55% as well as functional and histological evidence of a similar magnitude of injury 3 days post LCs that was not different between age groups. After 28 days, the architecture and innervation of the NMJs were evaluated. The numbers of fragmented endplates increased and of fully innervated NMJs decreased post-injury for the muscle of both middle-aged and old mice and for contralateral uninjured muscles of old compared with uninjured muscles of middle-aged controls. Thus, the diminished ability of the skeletal muscle of old mice to recover following injury may be due in part to an age-related decrease in the ability to regenerate NMJs in injured muscles. The impaired ability to regenerate NMJs may be a triggering factor for degenerative changes at the NMJ contributing to muscle fiber weakness and loss in old age.

Comparison of meat quality and glycolysis potential of two hybrid pigs in three-way hybrid model.

With the improvement of consumers' requirements for pork quality, the method of crossbreeding with excellent local pig breeds to improve meat quality is popular. Saba pig has high reproduction rate, good meat quality and high utilization rate of roughage, but its excellent characteristics have not been fully developed and utilized. To promote the development and utilization of Saba pigs and production of high-quality pork, the meat quality traits and glycolysis potential of Duroc × (Landrace × Yorkshire) (DLY), Berkshire × (Duroc × Saba) (BDS), and Duroc × (Berkshire × Saba) (DBS) three-way crossbred pigs were compared. The results showed that DLY had the highest live weight, carcass weight, lean meat percentage, drip loss, glycolysis potential, muscle diameter, and relative mRNA expression levels of type IIb muscle fibers as well as the lowest ultimate pH (p < 0.05). The lightness value of DBS was the highest (p < 0.05). Among the three crossbred pigs, myristic, arachidic, palmitoleic, and eicosenoic acids were the highest in BDS. These results indicated that the carcass traits of local crossbred pigs were worse than those of DLY pigs, but meat quality was markedly higher, with BDS showing the best meat quality.

A prospective clinical study on the mechanisms underlying critical illness myopathy-A time-course approach.

Critical illness myopathy (CIM) is a consequence of modern critical care resulting in general muscle wasting and paralyses of all limb and trunk muscles, resulting in prolonged weaning from the ventilator, intensive care unit (ICU) treatment and rehabilitation. CIM is associated with severe morbidity/mortality and significant negative socioeconomic consequences, which has become increasingly evident during the current COVID-19 pandemic, but underlying mechanisms remain elusive. Ten neuro-ICU patients exposed to long-term controlled mechanical ventilation were followed with repeated muscle biopsies, electrophysiology and plasma collection three times per week for up to 12days. Single muscle fibre contractile recordings were conducted on the first and final biopsy, and a multiomics approach was taken to analyse gene and protein expression in muscle and plasma at all collection time points. (i) A progressive preferential myosin loss, the hallmark of CIM, was observed in all neuro-ICU patients during the observation period (myosin:actin ratio decreased from 2.0 in the first to 0.9 in the final biopsy, P<0.001). The myosin loss was coupled to a general transcriptional downregulation of myofibrillar proteins (P<0.05; absolute fold change >2) and activation of protein degradation pathways (false discovery rate [FDR] <0.1), resulting in significant muscle fibre atrophy and loss in force generation capacity, which declined >65% during the 12day observation period (muscle fibre cross-sectional area [CSA] and maximum single muscle fibre force normalized to CSA [specific force] declined 30% [P<0.007] and 50% [P<0.0001], respectively). (ii) Membrane excitability was not affected as indicated by the maintained compound muscle action potential amplitude upon supramaximal stimulation of upper and lower extremity motor nerves. (iii) Analyses of plasma revealed early activation of inflammatory and proinflammatory pathways (FDR<0.1), as well as a redistribution of zinc ions from plasma. The mechanical ventilation-induced lung injury with release of cytokines/chemokines and the complete mechanical silencing uniquely observed in immobilized ICU patients affecting skeletal muscle gene/protein expression are forwarded as the dominant factors triggering CIM.

P.61 Imaging Mass Cytometry reveals new clues to understand the pathogenesis of Becker muscular dystrophy

Muscular dystrophies are genetic diseases characterised by muscle fiber loss and its replacement by fibrotic and adipose tissue. Several cell types, including macrophages and fibroadipogenic precursor cells (FAPS) interact with degenerative muscle fibers locally in what is known as the degenerative niche. Recent technological advances known as spatial biology allow dissecting cell-to-cell interactions in vivo using samples of tissue, although this has not been applied to human muscle tissue yet. We analysed muscle biopsies of controls and patients with Becker muscular dystrophy (BMD) at early, moderate and advanced stages of degeneration using Hyperion Imaging Mass Cytometry (IMC). We labelled single sections of the biopsies with 40 markers labelling different components of the muscle structure. We developed a deep learning algorithm to analyse the images and studied changes in the composition and spatial correlations of the different markers across disease progression. We included 2 control and 8 muscle biopsies of BMD. We observed a progressive increase in the amount of collagen-I and III, that significantly correlated with the amount of CD68+ macrophages (R=0.93 and R=0.94), most of them positive for the M2 marker CD206. Fat and collagen I surrounded vessels during the first stages of disease progression, substituting muscle fibres in later stages. We observed a negative correlation between regenerative muscle fibers and collagen-I (R=-0.83) and with CD68 cells (R:-0.90). We also observed a modification in the number of satellite cells in the biopsies across disease progression. IMC enables the study of changes in muscle structure along disease progression in patients with muscular dystrophies establishing spatio-temporal correlations opening the door to confirm and understand new potential pathogenic pathways in vivo in human samples.

Antisense Morpholino-Based In Vitro Correction of a Pseudoexon-Generating Variant in the SGCB Gene.

Limb-girdle muscular dystrophies (LGMD) are clinically and genetically heterogenous presentations displaying predominantly proximal muscle weakness due to the loss of skeletal muscle fibers. Beta-sarcoglycanopathy (LGMDR4) results from biallelic molecular defects in SGCB and features pediatric onset with limb-girdle involvement, often complicated by respiratory and heart dysfunction. Here we describe a patient who presented at the age of 12 years reporting high creatine kinase levels and onset of cramps after strenuous exercise. Instrumental investigations, including a muscle biopsy, pointed towards a diagnosis of beta-sarcoglycanopathy. NGS panel sequencing identified two variants in the SGCB gene, one of which (c.243+1548T>C) was found to promote the inclusion of a pseudoexon between exons 2 and 3 in the SGCB transcript. Interestingly, we detected the same genotype in a previously reported LGMDR4 patient, deceased more than twenty years ago, who had escaped molecular diagnosis so far. After the delivery of morpholino oligomers targeting the pseudoexon in patient-specific induced pluripotent stem cells, we observed the correction of the physiological splicing and partial restoration of protein levels. Our findings prompt the analysis of the c.243+1548T>C variant in suspected LGMDR4 patients, especially those harbouring monoallelic SGCB variants, and provide a further example of the efficacy of antisense technology for the correction of molecular defects resulting in splicing abnormalities.

Neuromuscular junction pathology is correlated with differential motor unit vulnerability in spinal and bulbar muscular atrophy

Spinal and bulbar muscular atrophy (SBMA) is an X-linked, neuromuscular neurodegenerative disease for which there is no cure. The disease is characterized by a selective decrease in fast-muscle power (e.g., tongue pressure, grip strength) accompanied by a selective loss of fast-twitch muscle fibers. However, the relationship between neuromuscular junction (NMJ) pathology and fast-twitch motor unit vulnerability has yet to be explored. In this study, we used a cross-model comparison of two mouse models of SBMA to evaluate neuromuscular junction pathology, glycolytic-to-oxidative fiber-type switching, and cytoskeletal alterations in pre- and postsynaptic termini of tibialis anterior (TA), gastrocnemius, and soleus hindlimb muscles. We observed significantly increased NMJ and myofiber pathology in fast-twitch, glycolytic motor units of the TA and gastrocnemius compared to slow-twitch, oxidative motor units of the soleus, as seen by decreased pre- and post-synaptic membrane area, decreased pre- and post-synaptic membrane colocalization, increased acetylcholine receptor compactness, a decrease in endplate area and complexity, and deficits in neurofilament heavy chain. Our data also show evidence for metabolic dysregulation and myofiber atrophy that correlate with severity of NMJ pathology. We propose a model in which the dynamic communicative relationship between the motor neuron and muscle, along with the developmental subtype of the muscle, promotes motor unit subtype specific vulnerability, metabolic alterations, and NMJ pathology.

Impaired Mitochondrial Folate Metabolism Reduces Mitochondrial Respiration in Mouse Muscle Progenitor Cells

ObjectivesMitochondrial dysfunction is a central contributor to aging. Mitochondrial DNA mutations increase with aging and are associated with low serum folate. Furthermore, both increased age and low serum folate are suggested to increase skeletal muscle fiber loss and impair muscle function. However, no research has focused on the metabolic pathway linking the causal relationship. Data from our lab indicate that folate-mediated one-carbon metabolism may play a role, as loss of the mitochondrial specific isozyme (serine hydroxymethyltransferase 2, SHMT2) and folate depletion both result in mitochondrial DNA instability and impaired mitochondrial function. Furthermore, serum folate and SHMT2 gene expression levels decrease with aging, potentially providing a causal mechanism for age-associated mitochondrial DNA mutations and skeletal muscle impairments. To date, little is known about the relationship between loss of SHMT2 enzyme expression and folate status in skeletal muscles.MethodsTibialis anterior muscles were harvested from 10-week-old wild-type (Shmt2+/+) and heterozygous (Shmt2+/−) mice that were weaned onto either a folate-sufficient diet or a modified diet lacking folic acid. Skeletal muscles used to isolate primary muscle progenitor cells (MPCs) were isolated from 20-week-old Shmt2+/+ and Shmt2+/− mouse hindlimbs. MPCs were cultured to 80–90% confluency and then differentiated in low-serum medium for 7–10 days. Skeletal muscles were examined for mitochondrial DNA content and MPCs were examined for mitochondrial function.ResultsHeterozygous loss of Shmt2 resulted in an 80% reduction in SHMT2 protein levels in mouse tibialis anterior muscle. Mice fed a folate-depleted diet for 7 weeks exhibited a 25% reduction in mitochondrial DNA content compared to the folate-sufficient group. In MPCs, reduced Shmt2 expression impaired basal and maximal respiration. Furthermore, protein levels of mitochondrial NADH dehydrogenase 3 were reduced and protein levels of PGC1α and mitofusin 2 were elevated with Shmt2 heterozygosity in MPCs.ConclusionsOur results demonstrate that folate depletion reduced mitochondrial DNA content in tibialis anterior skeletal muscle and loss of SHMT2 impairs MPCs mitochondrial respiration and may influence mitochondrial biogenesis and dynamics.Funding SourcesPresident's Council of Cornell Women Award.

Extracellular polysaccharides purified (Polycan) from Aureobasidium pullulans SM‑2001 improves pathophysiology of dystrophin-deficient mdx mice

BackgroundDuchenne muscular dystrophy is a hereditary muscular disease involving degeneration (i.e. atrophy and loss of muscle fibres) of skeletal muscles, including the diaphragm, and progressively severe functional decline. A previous study shows Polycan, a type of β-glucan derived from the black yeast Aureobasidium pullulans (SM-2001), promotes osteogenicity and bone loss, and possesses anti-inflammatory activity to induce inflammatory cytokines in human immune and cancer cells.ObjectiveIn this study, we evaluated changes in exercise load behaviour measurements and changes in muscle-related physiological indicators following oral administration of Polycan in mdx mice, an experimental animal model of Duchenne muscular dystrophy.ResultIn mdx mice, Polycan prevented weight loss and thickness of skeletal muscle. In addition, by monitoring increases in running time of mice on treadmills and performing a grip strength test, we confirmed reduced muscle function was recovered to some extent after administering Polycan to mdx mice. In addition, we confirmed that Polycan significantly altered mRNA expression in a concentration-dependent manner, whereby myogenic transcription factors (MyoD, Myf5 and Myogenin) increased and FoxO3α, MuRF1 and Atrogin-1 decreased. We aimed to investigate the mechanism of action in Polycan on energy metabolism of p-AMPK, SIRT1 and PGC1α with apoptosis expression levels as factors related to signalling pathways. Expression ratios of cleaved-caspase-3/caspase-3 and Bax/Bcl-2 in the Polycan extract-administered group increased compared with the control group.ConclusionThese results demonstrate that Polycan can improve and protect muscle atrophy by preventing apoptosis via pathway regulation related to myogenic transcription factors and energy metabolism in mdx mice.

Breast Augmentation in Athletic Women: A Retrospective Survey Assessing Pectoral Muscle Function and Implant Aesthetics Post-Augmentation.

Breast augmentation is the most common cosmetic, surgical procedure (1). Implant insertion planes include subglandular or submuscular. Submuscular augmentation is often preferred in women with less soft tissue coverage; requiring pectoralis major dissection (2). However, loss of functional muscle fibres is undesirable in athletic women who actively train the pectoral region. 1. Assessment of pectoralis function and strength after breast augmentation in athletic women. 2. Augmentation aesthetical satisfaction at rest and whilst training. A retrospective survey was sent to female, fitness competitors with breast implants via social media over a six-month period. The survey assessed baseline demographics, pectoral strength (bench press, push-ups, pectoral fly), aesthetical satisfaction and issues during training including pain, rippling, firmness and implant movement. Eighty-one participants were surveyed. The mean age of augmentation was 29.7 (±8.41), and mean age of survey completion was 37.6 (±7.22) years. Submuscular augmentation was performed in 72.8% of participants. Recovery post-operative and time spent training was synonymous. The majority of participants' pectoral strength was unaffected or positively affected by breast augmentation. Assessment of post-operative performance and training issues of pain, rippling and firmness yielded no statistically significant difference between groups. Implant movement during pectoral exercises was 2.5 times more likely with submuscular augmentations, p= 0.038. Overall, the majority of the population were very satisfied or satisfied with their breast aesthetic at rest (92.6%) and during training (79.0%). Breast augmentation in athletic women has a high satisfaction rate with the majority maintaining or improving the strength of the pectoral region regardless of augmentation plane. This journal requires that authors assign a level of evidence to each article. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the online Instructions to Authors https://www.springer.com/journal/00266 .