Skeletal Muscle Biopsies Research Articles

Multitissue responses to exercise: a MoTrPAC feasibility study.

We assessed the feasibility of the Molecular Transducers of Physical Activity Consortium (MoTrPAC) human adult clinical exercise protocols, while also documenting select cardiovascular, metabolic, and molecular responses to these protocols. After phenotyping and familiarization sessions, 20 subjects (25 ± 2 yr, 12 M, 8 W) completed an endurance exercise bout (n = 8, 40 min cycling at 70% V̇o2max), a resistance exercise bout (n = 6, ∼45 min, 3 sets of ∼10 repetition maximum, 8 exercises), or a resting control period (n = 6, 40 min rest). Blood samples were taken before, during, and after (10 min, 2 h, and 3.5 h) exercise or rest for levels of catecholamines, cortisol, glucagon, insulin, glucose, free fatty acids, and lactate. Heart rate was recorded throughout exercise (or rest). Skeletal muscle (vastus lateralis) and adipose (periumbilical) biopsies were taken before and ∼4 h following exercise or rest for mRNA levels of genes related to energy metabolism, growth, angiogenesis, and circadian processes. Coordination of the timing of procedural components (e.g., local anesthetic delivery, biopsy incisions, tumescent delivery, intravenous line flushes, sample collection and processing, exercise transitions, and team dynamics) was reasonable to orchestrate while considering subject burden and scientific objectives. The cardiovascular and metabolic alterations reflected a dynamic and unique response to endurance and resistance exercise, whereas skeletal muscle was transcriptionally more responsive than adipose 4 h postexercise. In summary, the current report provides the first evidence of protocol execution and feasibility of key components of the MoTrPAC human adult clinical exercise protocols. Scientists should consider designing exercise studies in various populations to interface with the MoTrPAC protocols and DataHub.NEW & NOTEWORTHY This study highlights the feasibility of key aspects of the MoTrPAC adult human clinical protocols. This initial preview of what can be expected from acute exercise trial data from MoTrPAC provides an impetus for scientists to design exercise studies to interlace with the rich phenotypic and -omics data that will populate the MoTrPAC DataHub at the completion of the parent protocol.

An aerobic exercise intervention to improve metabolic health among people living with HIV with at-risk alcohol use: the ALIVE-Ex research study protocol

BackgroundEffective antiretroviral therapy (ART) in people living with HIV (PLWH) has improved life expectancy and increased risk of age-associated cardiometabolic comorbidities. At-risk alcohol use is more frequent among PLWH and increases the risk of health challenges. PLWH with at-risk alcohol use are more likely to meet criteria for prediabetes/diabetes and this is associated with impaired whole-body glucose-insulin dynamics.MethodsThe Alcohol & Metabolic Comorbidities in PLWH: Evidence Driven Interventions Study (ALIVE-Ex Study, NCT03299205) is a longitudinal, prospective, interventional study to determine the effects of an aerobic exercise protocol on improving dysglycemia among PLWH with at-risk alcohol use. The intervention is a moderate intensity aerobic exercise protocol implemented 3 days per week for 10 weeks at the Louisiana State University Health Sciences Center-New Orleans. Participants who have a fasting blood glucose level between 94 and 125 mg/dl will be enrolled in the study. Oral glucose tolerance tests, fitness assessments, and skeletal muscle biopsies will be performed pre- and post-exercise intervention. The primary outcome is to determine whether the exercise protocol improves measures of whole-body glucose-insulin dynamics, cardiorespiratory fitness, and skeletal muscle metabolic and bioenergetic function. Secondary outcomes are to determine whether the exercise intervention improves cognitive function and overall quality of life. Results generated will demonstrate the effect of exercise on glycemic measures in PLWH with subclinical dysglycemia and at-risk alcohol use.ConclusionsThe proposed intervention will also have the potential to be scalable to promote lifestyle changes among PLWH, particularly in underserved communities.

Morphological and molecular comparison of HIV-associated and sporadic inclusion body myositis

ObjectiveThe molecular characteristics of sporadic inclusion body myositis (sIBM) have been intensively studied, and specific patterns on the cellular, protein and RNA level have emerged. However, these characteristics have not been studied in the context of HIV-associated IBM (HIV-IBM). In this study, we compared clinical, histopathological, and transcriptomic patterns of sIBM and HIV-IBM.MethodsIn this cross-sectional study, we compared patients with HIV-IBM and sIBM based on clinical and morphological features as well as gene expression levels of specific T-cell markers in skeletal muscle biopsy samples. Non-disease individuals served as controls (NDC). Cell counts for immunohistochemistry and gene expression profiles for quantitative PCR were used as primary outcomes.Results14 muscle biopsy samples (7 HIV-IBM, 7 sIBM) of patients and 6 biopsy samples from NDC were included. Clinically, HIV-IBM patients showed a significantly lower age of onset and a shorter period between symptom onset and muscle biopsy. Histomorphologically, HIV-IBM patients showed no KLRG1+ or CD57+ cells, while the number of PD1+ cells did not differ significantly between the two groups. All markers were shown to be significantly upregulated at gene expression level with no significant difference between the IBM subgroups.ConclusionDespite HIV-IBM and sIBM sharing important clinical, histopathological, and transcriptomic signatures, the presence of KLRG1+ cells discriminated sIBM from HIV-IBM. This may be explained by longer disease duration and subsequent T-cell stimulation in sIBM. Thus, the presence of TEMRA cells is characteristic for sIBM, but not a prerequisite for the development of IBM in HIV+ patients.

Skeletal muscle fibre type and enzymatic activity in adult offspring following placental and peripheral malaria exposure in foetal life.

Maternal malaria may restrict foetal growth. Impaired utero-placental blood flow due to malaria infection may cause hypoxia-induced altered skeletal muscle fibre type distribution in the offspring, which may contribute to insulin resistance and impaired glucose metabolism. This study assessed muscle fibre distribution 20 years after placental and/or peripheral in-utero malaria exposure compared to no exposure, i.e., PPM+, PM+, and M-, respectively. We traced 101 men and women offspring of mothers who participated in a malaria chemosuppression study in Muheza, Tanzania. Of 76 eligible participants, 50 individuals (29 men and 21 women) had skeletal muscle biopsy taken from m. vastus lateralis in the right leg. As previously reported, fasting and 30 min post-oral glucose challenge plasma glucose values were higher, and insulin secretion disposition index was lower, in the PPM+ group. Aerobic capacity (fitness) was estimated by an indirect VO2max test on a stationary bicycle. Muscle fibre sub-type (myosin heavy chain, MHC) distribution was analysed, as were muscle enzyme activities (citrate synthase (CS), 3-hydroxyacyl-CoA dehydrogenase, myophosphorylase, phosphofructokinase, lactate dehydrogenase, and creatine kinase activities. Between-group analyses were adjusted for MHC-I %. No differences in aerobic capacity were found between groups. Despite subtle elevations of plasma glucose levels in the PPM+ group, there was no difference in MHC sub-types or muscle enzymatic activities between the malaria-exposed and non-exposed groups. The current study did not show differences in MHC towards glycolytic sub-types or enzymatic activity across the sub-groups. The results support the notion of the mild elevations of plasma glucose levels in people exposed to placental malaria in pregnancy being due to compromised pancreatic insulin secretion rather than insulin resistance.

Identifying molecular targets for rational immunosuppressive strategies in patients with immune checkpoint therapy-induced myocarditis and myositis.

2511 Background: Immune checkpoint therapies (ICTs) can induce durable anti-tumor responses but also life-threatening immune-related adverse events (irAEs). Myocarditis, myositis, and myasthenia gravis (MG) are rare, often concurrent irAEs with mortality up to 40%. Steroids are the standard first-line treatment and have significant adverse events with prolonged use. We and others have identified serum autoantibodies in these irAEs, suggestive of B cell contribution to pathogenesis. Furthermore, although prior studies have shown T cells and myeloid cells in inflamed muscle tissue, their specific subtype and role in disease pathogenesis have yet to be defined. This understanding may contribute to the development of more effective immunosuppressive strategies. Hypothesis: In addition to T cells, plasma B and myeloid cells contribute to the pathogenesis of ICT-induced myocarditis, myositis, and MG. Methods: We enrolled patients with suspected ICT-induced myocarditis, myositis, and MG on an IRB-approved laboratory protocol. Diagnosis required: 1) symptoms requiring hospitalization; 2) elevated serum markers (creatine kinase ≥5x normal, troponin >99th percentile or detection of autoantibodies for MG); and 3) positive biopsy, electromyography, or cardiac MRI. Patients underwent standard diagnostic testing (including endomyocardial and/or skeletal muscle biopsy) and treatment as clinically indicated. Results: 23 patients were enrolled (n=13, confirmed irAE; n=10, negative control). Most patients received combination ICT (10/13, 77%). Anti-striated muscle and/or acetylcholine receptor binding antibodies were detected in 7/13 patients (54%), including 3 without MG (known to have both T and B cell etiologies). Single cell RNA sequencing of patient tissue (confirmed irAEs: cardiac n=10, skeletal n=9; control: cardiac n=4, skeletal n=5) identified specific populations of myeloid and CD8+ T cells conserved between inflamed cardiac and skeletal muscle. Conclusions: Specific populations of T cells, myeloid cells, and plasma B cells are potential targets for rational immunosuppressive strategies in ICT-induced myocarditis and myositis and will be tested in murine models to inform future clinical trial design. [Table: see text]

Skeletal muscle overexpression of sAnk1.5 in transgenic mice does not predispose to type 2 diabetes

Genome-wide association studies (GWAS) and cis-expression quantitative trait locus (cis-eQTL) analyses indicated an association of the rs508419 single nucleotide polymorphism (SNP) with type 2 diabetes (T2D). rs508419 is localized in the muscle-specific internal promoter (P2) of the ANK1 gene, which drives the expression of the sAnk1.5 isoform. Functional studies showed that the rs508419 C/C variant results in increased transcriptional activity of the P2 promoter, leading to higher levels of sAnk1.5 mRNA and protein in skeletal muscle biopsies of individuals carrying the C/C genotype. To investigate whether sAnk1.5 overexpression in skeletal muscle might predispose to T2D development, we generated transgenic mice (TgsAnk1.5/+) in which the sAnk1.5 coding sequence was selectively overexpressed in skeletal muscle tissue. TgsAnk1.5/+ mice expressed up to 50% as much sAnk1.5 protein as wild-type (WT) muscles, mirroring the difference reported between individuals with the C/C or T/T genotype at rs508419. However, fasting glucose levels, glucose tolerance, insulin levels and insulin response in TgsAnk1.5/+ mice did not differ from those of age-matched WT mice monitored over a 12-month period. Even when fed a high-fat diet, TgsAnk1.5/+ mice only presented increased caloric intake, but glucose disposal, insulin tolerance and weight gain were comparable to those of WT mice fed a similar diet. Altogether, these data indicate that sAnk1.5 overexpression in skeletal muscle does not predispose mice to T2D susceptibility.

Dietary Betaine Impacts Metabolic Responses to Moderate Heat Exposure in Sheep.

Dietary betaine supplementation can ameliorate physiological responses to heat exposure (HE) in sheep. This experiment measured metabolic responses to glucose (intravenous glucose tolerance, IVGTT), insulin (insulin tolerance test, ITT), and adrenocorticotropic hormone (ACTH) challenges in Merino ewes (n = 36, 39.7 kg) maintained at thermoneutral (TN, 21 °C) or HE (18-43 °C) and supplemented with either 0, 2, or 4 g/day dietary betaine (n = 6 per group). Sheep had ad libitum access to water and were pair-fed such that the intake of the TN sheep mimicked that of the HE sheep. After 21 days of treatment, sheep were fitted with jugular catheters and subjected to consecutive daily challenges (IVGTT, ITT, and ACTH, d 21-23, respectively), followed by skeletal muscle and subcutaneous adipose tissue biopsy collections for gene expression analysis (d 24). The HE-treated sheep had a greater insulin:glucose ratio (p = 0.033), a greater estimated homeostatic model assessment of insulin resistance (HOMAIR; p = 0.029), and a reduced revised quantitative insulin sensitivity check index (RQUICKI; p = 0.015). Sheep fed betaine (2 + 4 g/day) had a greater basal plasma insulin (p = 0.017) and a reduced basal non-esterified fatty acid (NEFA; p = 0.036) concentration, while the RQUICKI was reduced (p = 0.001) in sheep fed betaine. The results suggested that betaine supplementation alters lipid metabolism by potentially improving insulin signaling, although these responses differ between TN and HE conditions. There was no other impact of temperature or dietary treatments on the tissue gene expressions measured. Our results support the notion that betaine, in part, acts to modify lipid metabolism.

Conditioning-induced expression of novel glucose transporters in canine skeletal muscle homogenate.

Athletic conditioning can increase the capacity for insulin-stimulated skeletal muscle glucose uptake through increased sarcolemmal expression of GLUT4 and potentially additional novel glucose transporters. We used a canine model that has previously demonstrated conditioning-induced increases in basal, insulin- and contraction-stimulated glucose uptake to identify whether expression of glucose transporters other than GLUT4 was upregulated by athletic conditioning. Skeletal muscle biopsies were obtained from 12 adult Alaskan Husky racing sled dogs before and after a full season of conditioning and racing, and homogenates from those biopsies were assayed for expression of GLUT1, GLUT3, GLUT4, GLUT6, GLUT8, and GLUT12 using western blots. Athletic conditioning resulted in a 1.31 ± 0.70 fold increase in GLUT1 (p <0.0001), 1.80 ± 1.99 fold increase in GLUT4 (p = 0.005), and 2.46 ± 2.39 fold increase in GLUT12 (p = 0.002). The increased expression of GLUT1 helps explain the previous findings of conditioning-induced increases in basal glucose clearance in this model, and the increase in GLUT12 provides an alternative mechanism for insulin- and contraction-mediated glucose uptake and likely contributes to the substantial conditioning-induced increases in insulin sensitivity in highly trained athletic dogs. Furthermore, these results suggest that athletic dogs can serve as a valuable resource for the study of alternative glucose transport mechanisms in higher mammals.

H19X-encoded non-coding RNAs are associated with loss of muscle and old age in human: A quantitative histological study using the Genotype-Tissue Expression (GTEx) database

Loss of skeletal muscle mass and function is commonly seen in individuals with chronic illness, older age, and altered hormonal and/or endocrine functions. Whole-genome survey of human muscle biopsies has identified several miRNAs which are associated with chronic diseases and thus named “atromiRs.” Using murine models, we have showed that H10X-encoded atromiRs miR-424/miR-503 downregulates muscle mass by directly targeting protein translation initiation factors. We hypothesized that H19X-encoded non-coding RNAs (ncRNAs) are involved in the pathogenesis of muscle atrophy/wasting in humans, and the levels of the ncRNAs are correlated with the loss of skeletal muscle mass in old age. A total of 803 samples from the GTEx project, which contains both skeletal muscle RNA expression information and skeletal muscle biopsy images, were included in the study. The samples were further classified into eight groups based on the levels of miR-503HG (host gene of miR-424/miR-503) reported in the database. Quantification of histopathology in HE-stained muscle sections were manually performed in group 1 (n=89) and 8 (n=100), which had the lowest levels or highest levels of miR-424/miR-503, respectively. The degree of muscle atrophy, fibrosis, and interfascicular/perifascicular fat accumulation were assessed with a grading scale of 0-3. The total muscle histology score was also calculated as a sum of all three scores. Unpaired student t-tests were used to compare histological scores, BMI, and age between the two groups. All tests were two-tailed and p-values less than 0.05 were considered significant. There is no significant difference between the two groups in sex ( p= 0.55). In agreement with our recent study, high levels of miR-503HG (group 8) are associated with old age ( p= 0.02), and a trend of higher BM ( p= 0.07). Notably, high levels of miR-503HG are significantly correlated with muscle atrophy ( p= 0.001) and a total muscle wasting score ( p= 0.03) from histological assessment. These findings suggest the increased levels of H19X-encoded ncRNAs in human skeletal muscle are associated with muscle mass loss and potential sarcopenia. The results of our study may provide insight into the underlying role of H19X-encoded ncRNAs in the pathogenesis of geriatric muscle loss. The research was funded by the University of Houston internal fund to enhance and advance research (to YL). 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.

Twenty-four hour rhythmicity in mitochondrial network connectivity and mitochondrial respiration; a study in human skeletal muscle biopsies of young lean and older individuals with obesity

ObjectiveMitochondrial network dynamics may play role in metabolic homeostasis. Whether mitochondrial network dynamics are involved in adaptations to day–night fluctuations in energy supply and demand is unclear. Here we visualized and quantified the mitochondrial network morphology in human skeletal muscle of young healthy lean and older individuals with obesity over the course of 24 h MethodsMuscle biopsies taken at 5 timepoints over a 24-hour period obtained from young healthy lean and older metabolically impaired obese males were analyzed for mitochondrial network integrity with confocal laser scanning microscopy. Variation of level of fragmentation over the course of the day were aligned with variation of mitochondrial respiration over the day ResultsYoung healthy lean individuals displayed a day–night rhythmicity in mitochondrial network morphology, which aligned with the day–night rhythmicity of mitochondrial respiratory capacity, with a more fused network coinciding with higher mitochondrial respiratory capacity. In the older individuals with obesity, the mitochondrial network was more fragmented overall compared to young healthy lean individuals and completely lacked 24 h rhythmicity, which was also true for the mitochondrial respiratory capacity ConclusionsOur data shows a paralleled rhythmicity between mitochondrial network morphology and mitochondrial oxidative capacity, which oscillates over the course of a mimicked real-life day in human skeletal muscle of young, healthy lean individuals. In older individuals with obesity, the lack of a 24-hour rhythmicity in mitochondrial network connectivity was also aligned with a lack in respiratory capacity. This suggests that 24-hour rhythmicity in mitochondrial network connectivity is a determinant of rhythmicity in mitochondrial respiratory capacity. Thus, restoring mitochondrial network integrity may promote mitochondrial respiratory capacity and hence contribute to blunting the metabolic aberrations in individuals with a disturbed 24-hour rhythmicity in metabolism, like older individuals with obesity.

Scanning electrochemical microscopy for the stimulation and investigation of human skeletal muscle-derived mesenchymal stem/stromal cells

IntroductionThe human skeletal muscle-derived mesenchymal stem/stromal cells (SM-MSCs) possess myogenic differentiation potential and participate in the muscle regeneration process. However, the question of how to improve human skeletal muscle regeneration remains actual. In this study, the total SM-MSCs population was separated into subpopulations according to the neural cell adhesion molecule (NCAM, CD56), stimulated with an alternating electric field (AC) using scanning electrochemical microscopy (AC-SECM) and the intracellular redox changes, and myogenic differentiation markers were evaluated. MethodsThe total SM-MSC population was isolated from the postoperative human skeletal muscle biopsies by an enzymatic digestion method. The myogenic differentiation of the total SM-MSCs population before and after AC stimulation was evaluated immunohistochemically by the levels of desmin and myogenin. The effect of AC stimulation on the redox capacity of CD56(+) and CD56(-) cell subpopulations as well as on the level of myogenin on indium tin oxide (ITO) surface were also investigated. ResultsThe total SM-MSCs population grown on a glass coverslip weakly responded to AC stimulus, i.e. the level of desmin was slightly increased by the 3rd day of differentiation, while in the not stimulated cells, it increased only by the 7th day. The level of myogenin did not change after AC stimulation. However, the CD56(+) and CD56(-) subpopulations had different redox activities and myogenic differentiation potential: the CD56(+) cells had stronger natural diffusion and were more redox active compared to the CD56(-) cells; the redox activity of CD56(+) cells was more actively stimulated by an alternating electric field than in CD56(-) cells; at control level, the CD56(+) cells had more myogenic differentiation-regulating transcription factor myogenin, which was more intensively stimulated by AC than in CD56(-) cells. Data show that the total population of human SM-MSCs is heterogeneous with different regenerating potential cells that do not equally respond to extracellular stimuli. The SECM can be used in both ways: (i) for extracellular stimulation and (ii) for the investigation of intracellular redox changes of the human SM-MSCs or their subpopulations allowing a deeper understanding of the mechanisms mediating skeletal tissue regeneration both in vitro and in vivo.

Whole-body and forearm muscle protein metabolism in patients with acromegaly before and after treatment.

Active acromegaly is characterized by increased lean body mass but the mechanisms underlying the protein anabolic effect are unclear. To study if active acromegaly induces reversible changes in whole-body and skeletal muscle protein kinetics. Eighteen patients with acromegaly were investigated before and 47 ± 10 weeks after disease control by surgery (n = 8) and/or medical treatment (n = 10). Labelled phenylalanine and tyrosine tracers were employed to assess whole-body and regional forearm muscle protein kinetics. Intramyocellular protein signaling was assessed in skeletal muscle biopsies, and whole-body DXA scan and indirect calorimetry assessed lean body mass (LBM) and resting energy expenditure, respectively. Disease control induced a 7% decrease in lean body mass (p < 0.000) and a 14% decrease in LBM-adjusted energy expenditure. Whole-body phenylalanine breakdown decreased after disease control (p = 0.005) accompanied by a decrease in the degradation of phenylalanine to tyrosine (p= 0.005) and a decrease in whole-body phenylalanine synthesis (p = 0.030). Skeletal muscle protein synthesis tended to decrease after disease control (p = 0.122), whereas the muscle protein breakdown (p = 0.437) and muscle protein loss were unaltered (p = 0.371). ULK1 phosphorylation, an activator of protein breakdown, increased after disease control (p= 0.042). Active acromegaly represents a reversible high flux state in which both whole-body protein breakdown and synthesis are increased, whereas forearm muscle protein kinetics are unaltered. Future studies are needed to decipher the link between protein kinetics and the structure and function of the associated GH-induced increase in lean body mass.

Skeletal muscle transcriptomics dissects the pathogenesis of Friedreich's ataxia.

In Friedreich's ataxia (FRDA), the most affected tissues are not accessible to sampling and available transcriptomic findings originate from blood-derived cells and animal models. Herein, we aimed at dissecting for the first time the pathophysiology of FRDA by means of RNA-sequencing in an affected tissue sampled in vivo. Skeletal muscle biopsies were collected from seven FRDA patients before and after treatment with recombinant human Erythropoietin (rhuEPO) within a clinical trial. Total RNA extraction, 3'-mRNA library preparation and sequencing were performed according to standard procedures. We tested for differential gene expression with DESeq2 and performed gene set enrichment analysis with respect to control subjects. FRDA transcriptomes showed 1873 genes differentially expressed from controls. Two main signatures emerged: (1) a global downregulation of the mitochondrial transcriptome as well as of ribosome/translational machinery and (2) an upregulation of genes related to transcription and chromatin regulation, especially of repressor terms. Downregulation of the mitochondrial transcriptome was more profound than previously shown in other cellular systems. Furthermore, we observed in FRDA patients a marked upregulation of leptin, the master regulator of energy homeostasis. RhuEPO treatment further enhanced leptin expression. Our findings reflect a double hit in the pathophysiology of FRDA: a transcriptional/translational issue and a profound mitochondrial failure downstream. Leptin upregulation in the skeletal muscle in FRDA may represent a compensatory mechanism of mitochondrial dysfunction, which is amenable to pharmacological boosting. Skeletal muscle transcriptomics is a valuable biomarker to monitor therapeutic interventions in FRDA.

Microscopic and Biochemical Hallmarks of BICD2-Associated Muscle Pathology toward the Evaluation of Novel Variants

BICD2 variants have been linked to neurodegenerative disorders like spinal muscular atrophy with lower extremity predominance (SMALED2) or hereditary spastic paraplegia (HSP). Recently, mutations in BICD2 were implicated in myopathies. Here, we present one patient with a known and six patients with novel BICD2 missense variants, further characterizing the molecular landscape of this heterogenous neurological disorder. A total of seven patients were genotyped and phenotyped. Skeletal muscle biopsies were analyzed by histology, electron microscopy, and protein profiling to define pathological hallmarks and pathogenicity markers with consecutive validation using fluorescence microscopy. Clinical and MRI-features revealed a typical pattern of distal paresis of the lower extremities as characteristic features of a BICD2-associated disorder. Histological evaluation showed myopathic features of varying severity including fiber size variation, lipofibromatosis, and fiber splittings. Proteomic analysis with subsequent fluorescence analysis revealed an altered abundance and localization of thrombospondin-4 and biglycan. Our combined clinical, histopathological, and proteomic approaches provide new insights into the pathophysiology of BICD2-associated disorders, confirming a primary muscle cell vulnerability. In this context, biglycan and thrombospondin-4 have been identified, may serve as tissue pathogenicity markers, and might be linked to perturbed protein secretion based on an impaired vesicular transportation.

Metabolic dysfunction in obesity is related to impaired suppression of fatty acid release from adipose tissue by insulin.

The aims of this study were: 1) to assess relationships among insulin-mediated glucose uptake with standard clinical outcomes and deep-phenotyping measures (including fatty acid [FA] rate of appearance [FA Ra] into the systemic circulation); and 2) to examine the contribution of adipocyte size, fibrosis, and proteomic profile to FA Ra regulation. A total of 66 adults with obesity (BMI=34 [SD 3] kg/m2 ) were assessed for insulin sensitivity (hyperinsulinemic-euglycemic clamp), and stable isotope dilution methods quantified glucose, FA, and glycerol kinetics in vivo. Abdominal subcutaneous adipose tissue (aSAT) and skeletal muscle biopsies were collected, and magnetic resonance imaging quantified liver and visceral fat content. Insulin-mediated FA Ra suppression associated with insulin-mediated glucose uptake (r=0.51; p < 0.01) and negatively correlated with liver (r=-0.36; p < 0.01) and visceral fat (r=-0.42; p < 0.01). aSAT proteomics from subcohorts of participants with low FA Ra suppression (n=8) versus high FA Ra suppression (n=8) demonstrated greater extracellular matrix collagen protein in low versus high FA Ra suppression. Skeletal muscle lipidomics (n=18) revealed inverse correlations of FA Ra suppression with acyl-chain length of acylcarnitine (r=-0.42; p=0.02) and triacylglycerol (r=-0.51; p < 0.01), in addition to insulin-mediated glucose uptake (acylcarnitine: r=-0.49; p < 0.01, triacylglycerol: r=-0.40; p < 0.01). Insulin's ability to suppress FA release from aSAT in obesity is related to enhanced insulin-mediated glucose uptake and metabolic health in peripheral tissues.

The Landscape of Accessible Chromatin and Developmental Transcriptome Maps Reveal a Genetic Mechanism of Skeletal Muscle Development in Pigs

The epigenetic regulation mechanism of porcine skeletal muscle development relies on the openness of chromatin and is also precisely regulated by transcriptional machinery. However, fewer studies have exploited the temporal changes in gene expression and the landscape of accessible chromatin to reveal the underlying molecular mechanisms controlling muscle development. To address this, skeletal muscle biopsy samples were taken from Landrace pigs at days 0 (D0), 60 (D60), 120 (D120), and 180 (D180) after birth and were then analyzed using RNA-seq and ATAC-seq. The RNA-seq analysis identified 8554 effective differential genes, among which ACBD7, TMEM220, and ATP1A2 were identified as key genes related to the development of porcine skeletal muscle. Some potential cis-regulatory elements identified by ATAC-seq analysis contain binding sites for many transcription factors, including SP1 and EGR1, which are also the predicted transcription factors regulating the expression of ACBD7 genes. Moreover, the omics analyses revealed regulatory regions that become ectopically active after birth during porcine skeletal muscle development after birth and identified 151,245, 53,435, 30,494, and 40,911 peaks. The enriched functional elements are related to the cell cycle, muscle development, and lipid metabolism. In summary, comprehensive high-resolution gene expression maps were developed for the transcriptome and accessible chromatin during postnatal skeletal muscle development in pigs.

The prevalence and associated factors of myocardial involvement in Duchenne muscular dystrophy patients in the first decade of life.

The prevalence and associated factors of myocardial involvement in Duchenne muscular dystrophy patients in the first decade of life.

Identification of two novel RRM2B variants associated with autosomal recessive progressive external ophthalmoplegia in a family with pseudodominant inheritance pattern.

RRM2B encodes the p53-inducible small subunit (p53R2) of ribonucleotide reductase, a key protein for mitochondrial DNA (mtDNA) synthesis. Pathogenic variants in this gene result in familial mitochondrial disease in adults and children, secondary to a maintenance disorder of mtDNA. This study describes two patients, mother and son, with early-onset chronic progressive external ophthalmoplegia (PEO). Skeletal muscle biopsy from the latter was examined: cytochrome c oxidase (COX)-negative fibres were shown, and molecular studies revealed multiple mtDNA deletions. A next-generation sequencing gene panel for nuclear-encoded mitochondrial maintenance genes identified two unreported heterozygous missense variants (c.514 G > A and c.682 G > A) in the clinically affected son. The clinically affected mother harboured the first variant in homozygous state, and the clinically unaffected father harboured the remaining variant in heterozygous state. In silico analyses predicted both variants as deleterious. Cell culture studies revealed that patients' skin fibroblasts, but not fibroblasts from healthy controls, responded to nucleoside supplementation with enhanced mtDNA repopulation, thus suggesting an in vitro functional difference in patients' cells. Our results support the pathogenicity of two novel RRM2B variants found in two patients with autosomal recessive PEO with multiple mtDNA deletions inherited with a pseudodominant pattern.

Muscle Lipid Oxidation Is Not Affected by Obstructive Sleep Apnea in Diabetes and Healthy Subjects

The molecular mechanisms linking obstructive sleep apnea (OSA) with type 2 diabetes mellitus (T2DM) remain unclear. This study investigated the effect of OSA on skeletal muscle lipid oxidation in nondiabetic controls and in type 2 diabetes (T2DM) patients. Forty-four participants matched for age and adiposity were enrolled: nondiabetic controls (control, n = 14), nondiabetic patients with severe OSA (OSA, n = 9), T2DM patients with no OSA (T2DM, n = 10), and T2DM patients with severe OSA (T2DM + OSA, n = 11). A skeletal muscle biopsy was performed; gene and protein expressions were determined and lipid oxidation was analyzed. An intravenous glucose tolerance test was performed to investigate glucose homeostasis. No differences in lipid oxidation (178.2 ± 57.1, 161.7 ± 22.4, 169.3 ± 50.9, and 140.0 ± 24.1 pmol/min/mg for control, OSA, T2DM, and T2DM+OSA, respectively; p > 0.05) or gene and protein expressions were observed between the groups. The disposition index, acute insulin response to glucose, insulin resistance, plasma insulin, glucose, and HBA1C progressively worsened in the following order: control, OSA, T2DM, and T2DM + OSA (p for trend <0.05). No association was observed between the muscle lipid oxidation and the glucose metabolism variables. We conclude that severe OSA is not associated with reduced muscle lipid oxidation and that metabolic derangements in OSA are not mediated through impaired muscle lipid oxidation.

Outside the fiber: Endomysial stromal and capillary pathology in skeletal muscle may impede infusion therapy in infantile-onset Pompe disease.

The survival of infantile-onset Pompe disease (IOPD) patients has improved dramatically since the introduction of enzyme replacement therapy (ERT) with a1glucosidase alfa. However, long-term IOPD survivors on ERT demonstrate motor deficits indicating that current therapy cannot completely prevent disease progression in skeletal muscle. We hypothesized that in IOPD, skeletal muscle endomysial stroma and capillaries would show consistent changes that could impede the movement of infused ERT from blood to muscle fibers. We retrospectively examined 9 skeletal muscle biopsies from 6 treated IOPD patients using light and electron microscopy. We found consistent ultrastructural endomysial stromal and capillary changes. The endomysial interstitium was expanded by lysosomal material, glycosomes/glycogen, cellular debris, and organelles, some exocytosed by viable muscle fibers and some released on fiber lysis. Endomysial scavenger cells phagocytosed this material. Mature fibrillary collagen was seen in the endomysium, and both muscle fibers and endomysial capillaries showed basal laminar reduplication and/or expansion. Capillary endothelial cells showed hypertrophy and degeneration, with narrowing of the vascular lumen. Ultrastructurally defined stromal and vascular changes likely constitute obstacles to movement of infused ERT from capillary lumen to muscle fiber sarcolemma, contributing to the incomplete efficacy of infused ERT in skeletal muscle. Our observations can inform approaches to overcoming these barriers to therapy.