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Clinical Epigenetics Jul 2023Adolescent idiopathic scoliosis (AIS) is characterized by low lean mass without vertebral deformity. The cause-and-effect relationship between scoliosis and paraspinal...
BACKGROUND
Adolescent idiopathic scoliosis (AIS) is characterized by low lean mass without vertebral deformity. The cause-and-effect relationship between scoliosis and paraspinal muscle imbalance has long puzzled researchers. Although FTO has been identified as a susceptibility gene for AIS, its potential role in the asymmetry of paraspinal muscles has not been fully elucidated.
METHODS
We investigated the role of Fto in murine myoblast proliferation, migration, and myogenic differentiation. We examined its precise regulatory influence on murine muscle fiber remodeling in vitro and in vivo. We identified the downstream target gene of Fto by screening key regulators of murine muscle fiber remodeling and identified its mA reader. Deep paraspinal muscle samples were obtained from the concave and convex sides of AIS patients with or without Schroth exercises, and congenital scoliosis served as a control group. We compared the content of type I fibers, expression patterns of fast- and slow-type genes, and levels of FTO expression.
RESULTS
FTO contributed to maintain the formation of murine slow-twitch fibers both in vitro and in vivo. These effects were mediated by the demethylation activity of FTO, which specifically demethylated NFATC1 and prevented YTHDF2 from degrading it. We found a significant reduction in type I fibers, mRNA levels of MYH7 and MYH7B, and expression of FTO on the concave side of AIS. The percentage of type I fibers showed a positive correlation with the expression level of FTO. The asymmetric patterns observed in AIS were consistent with those seen in congenital scoliosis, and the asymmetry of FTO expression and fiber type in AIS was largely restored by Schroth exercises.
CONCLUSIONS
FTO supports the formation of murine slow-twitch fibers in an NFATC1-YTHDF2 dependent manner. The consistent paraspinal muscle features seen in AIS and congenital scoliosis, as well as the reversible pattern of muscle fibers and expression of FTO in AIS suggest that FTO may contribute to the muscle fiber remodeling secondary to scoliosis.
Topics: Adolescent; Humans; Animals; Mice; Scoliosis; DNA Methylation; Muscle Fibers, Skeletal; Transcription Factors; Paraspinal Muscles; NFATC Transcription Factors; Alpha-Ketoglutarate-Dependent Dioxygenase FTO; RNA-Binding Proteins
PubMed: 37408034
DOI: 10.1186/s13148-023-01526-5 -
Nature Communications Dec 2023The cellular prion protein (PrP) is required for skeletal muscle function. Here, we report that a higher level of PrP accumulates in the cytoplasm of the skeletal muscle...
The cellular prion protein (PrP) is required for skeletal muscle function. Here, we report that a higher level of PrP accumulates in the cytoplasm of the skeletal muscle of six myopathy patients compared to controls. PrP inhibits skeletal muscle cell autophagy, and blocks myoblast differentiation. PrP selectively binds to a subset of miRNAs during myoblast differentiation, and the colocalization of PrP and miR-214-3p was observed in the skeletal muscle of six myopathy patients with excessive PrP. We demonstrate that PrP is overexpressed in skeletal muscle cells under pathological conditions, inhibits muscle cell differentiation by physically interacting with a subset of miRNAs, and selectively recruits these miRNAs into its phase-separated condensate in living myoblasts, which in turn enhances liquid-liquid phase separation of PrP, promotes pathological aggregation of PrP, and results in the inhibition of autophagy-related protein 5-dependent autophagy and muscle bundle formation in myopathy patients characterized by incomplete muscle regeneration.
Topics: Humans; Cell Differentiation; Cell Proliferation; MicroRNAs; Muscle Development; Muscle, Skeletal; Muscular Diseases; PrPC Proteins
PubMed: 38065962
DOI: 10.1038/s41467-023-43826-7 -
Pharmaceutical Biology Dec 2023Kazinol B (KB), an isoprenylated flavan derived from Sieb. (Moraceae) root, has long been used in folk medicine.
CONTEXT
Kazinol B (KB), an isoprenylated flavan derived from Sieb. (Moraceae) root, has long been used in folk medicine.
OBJECTIVE
This study examines the protective effects of KB and its underlying mechanisms in hypoxia and reoxygenation (H/R)-induced cardiac injury in H9c2 rat cardiac myoblasts.
MATERIALS AND METHODS
H9c2 cells were incubated with various concentrations of KB (0, 0.3, 1, 3, 10 and 30 μM) for 2 h and then subjected to H/R insults. The protective effects of KB and its underlying mechanisms were explored.
RESULTS
KB significantly elevated cell viability (1 μM, 1.21-fold; 3 μM, 1.36-fold, and 10 μM, 1.47-fold) and suppressed LDH release (1 μM, 0.77-fold; 3 μM, 0.68-fold, and 10 μM, 0.59-fold) in H/R-induced H9c2 cells. Further, 10 μM KB blocked apoptotic cascades, as shown by the Annexin-V/PI (0.41-fold), DNA fragmentation (0.51-fold), caspase-3 (0.52-fold), PARP activation (0.27-fold) and Bax/Bcl-2 expression (0.28-fold) assays. KB (10 μM) downregulated reactive oxygen species production (0.51-fold) and lipid peroxidation (0.48-fold); it upregulated the activities of GSH-Px (2.08-fold) and SOD (1.72-fold). KB (10 μM) induced Nrf2 nuclear accumulation (1.94-fold) and increased ARE promoter activity (2.15-fold), HO-1 expression (3.07-fold), AKT (3.07-fold) and AMPK (3.07-fold) phosphorylation. Nrf2 knockdown via using Nrf2 siRNA abrogated KB-mediated protective effects against H/R insults. Moreover, pharmacological inhibitors of AKT and AMPK also abrogated KB-induced Nrf2 activation and its protective function.
DISCUSSION AND CONCLUSIONS
KB prevented H/R-induced cardiomyocyte injury via modulating the AKT and AMPK-mediated Nrf2 induction. KB might be a promising drug candidate for managing ischemic cardiac disorders.
Topics: Rats; Animals; Myocytes, Cardiac; Proto-Oncogene Proteins c-akt; NF-E2-Related Factor 2; AMP-Activated Protein Kinases; Hypoxia; Apoptosis; Oxidative Stress
PubMed: 36740871
DOI: 10.1080/13880209.2023.2173247 -
Acta Neuropathologica Communications Oct 2023Duchenne muscular dystrophy (DMD) is a devastating X-linked muscular disease, caused by mutations in the DMD gene encoding Dystrophin and affecting 1:5000 boys...
Duchenne muscular dystrophy (DMD) is a devastating X-linked muscular disease, caused by mutations in the DMD gene encoding Dystrophin and affecting 1:5000 boys worldwide. Lack of Dystrophin leads to progressive muscle wasting and degeneration resulting in cardiorespiratory failure. Despite the absence of a definitive cure, innovative therapeutic avenues are emerging. Myopathologic studies are important to further understand the biological mechanisms of the disease and to identify histopathologic benchmarks for clinical evaluations. We conducted a myopathologic analysis on twenty-four muscle biopsies from DMD patients, with particular emphasis on regeneration, fibro-adipogenic progenitors and muscle stem cells behavior. We describe an increase in content of fibro-adipogenic progenitors, central orchestrators of fibrotic progression and lipid deposition, concurrently with a decline in muscle regenerative capacity. This regenerative impairment strongly correlates with compromised activation and expansion of muscle stem cells. Furthermore, our study uncovers an early acquisition of a senescence phenotype by DMD-afflicted muscle stem cells. Here we describe the myopathologic trajectory intrinsic to DMD and establish muscle stem cell senescence as a pivotal readout for future therapeutic interventions.
Topics: Humans; Male; Dystrophin; Fibrosis; Muscle, Skeletal; Muscular Dystrophy, Duchenne; Regeneration; Satellite Cells, Skeletal Muscle; Cellular Senescence
PubMed: 37858263
DOI: 10.1186/s40478-023-01657-z -
Advanced Science (Weinheim,... Jul 2023It is well-known that muscle regeneration declines with aging, and aged muscles undergo degenerative atrophy or sarcopenia. While exercise and acute injury are both...
It is well-known that muscle regeneration declines with aging, and aged muscles undergo degenerative atrophy or sarcopenia. While exercise and acute injury are both known to induce muscle regeneration, the molecular signals that help trigger muscle regeneration have remained unclear. Here, mass spectrometry imaging (MSI) is used to show that injured muscles induce a specific subset of prostanoids during regeneration, including PGG1, PGD2, and the prostacyclin PGI2. The spike in prostacyclin promotes skeletal muscle regeneration via myoblasts, and declines with aging. Mechanistically, the prostacyclin spike promotes a spike in PPARγ/PGC1a signaling, which induces a spike in fatty acid oxidation (FAO) to control myogenesis. LC-MS/MS and MSI further confirm that an early FAO spike is associated with normal regeneration, but muscle FAO became dysregulated during aging. Functional experiments demonstrate that the prostacyclin-PPARγ/PGC1a-FAO spike is necessary and sufficient to promote both young and aged muscle regeneration, and that prostacyclin can synergize with PPARγ/PGC1a-FAO signaling to restore aged muscles' regeneration and physical function. Given that the post-injury prostacyclin-PPARγ-FAO spike can be modulated pharmacologically and via post-exercise nutrition, this work has implications for how prostacyclin-PPARγ-FAO might be fine-tuned to promote regeneration and treat muscle diseases of aging.
Topics: Muscle, Skeletal; PPAR gamma; Epoprostenol; Chromatography, Liquid; Tandem Mass Spectrometry; Prostaglandins I; Regeneration
PubMed: 37140179
DOI: 10.1002/advs.202301519 -
Journal of Cachexia, Sarcopenia and... Dec 2023Decreased ryanodine receptor type 1 (RyR1) protein levels are a well-described feature of recessive RYR1-related myopathies. The aim of the present study was twofold:...
BACKGROUND
Decreased ryanodine receptor type 1 (RyR1) protein levels are a well-described feature of recessive RYR1-related myopathies. The aim of the present study was twofold: (1) to determine whether RyR1 content is also decreased in other myopathies and (2) to investigate the mechanisms by which decreased RyR1 protein triggers muscular disorders.
METHODS
We used publicly available datasets, muscles from human inflammatory and mitochondrial myopathies, an inducible muscle-specific RYR1 recessive mouse model and RyR1 knockdown in C2C12 muscle cells to measure RyR1 content and endoplasmic reticulum (ER) stress markers. Proteomics, lipidomics, molecular biology and transmission electron microscopy approaches were used to decipher the alterations associated with the reduction of RyR1 protein levels.
RESULTS
RYR1 transcripts were reduced in muscle samples of patients suffering from necrotizing myopathy (P = 0.026), inclusion body myopathy (P = 0.003), polymyositis (P < 0.001) and juvenile dermatomyositis (P < 0.001) and in muscle samples of myotonic dystrophy type 2 (P < 0.001), presymptomatic (P < 0.001) and symptomatic (P < 0.001) Duchenne muscular dystrophy, Becker muscular dystrophy (P = 0.004) and limb-girdle muscular dystrophy type 2A (P = 0.004). RyR1 protein content was also significantly decreased in inflammatory myopathy (-75%, P < 0.001) and mitochondrial myopathy (-71%, P < 0.001) muscles. Proteomics data showed that depletion of RyR1 protein in C2C12 myoblasts leads to myotubes recapitulating the common molecular alterations observed in myopathies. Mechanistically, RyR1 protein depletion reduces ER-mitochondria contact length (-26%, P < 0.001), Ca transfer to mitochondria (-48%, P = 0.002) and the mitophagy gene Parkinson protein 2 transcripts (P = 0.037) and induces mitochondrial accumulation (+99%, P = 0.005) and dysfunction (P < 0.001). This was associated to the accumulation of deleterious sphingolipid species. Our data showed increased levels of the ER stress marker chaperone-binding protein/glucose regulated protein 78, GRP78-Bip, in RyR1 knockdown myotubes (+45%, P = 0.046), in mouse RyR1 recessive muscles (+58%, P = 0.001) and in human inflammatory (+96%, P = 0.006) and mitochondrial (+64%, P = 0.049) myopathy muscles. This was accompanied by increased protein levels of the pro-apoptotic protein CCAAT-enhancer-binding protein homologous protein, CHOP-DDIT3, in RyR1 knockdown myotubes (+27%, P < 0.001), mouse RyR1 recessive muscles (+63%, P = 0.009), human inflammatory (+50%, P = 0.038) and mitochondrial (+51%, P = 0.035) myopathy muscles. In publicly available datasets, the decrease in RYR1 content in myopathies was also associated to increased ER stress markers and RYR1 transcript levels are inversely correlated with ER stress markers in the control population.
CONCLUSIONS
Decreased RyR1 is commonly observed in myopathies and associated to ER stress in vitro, in mouse muscle and in human myopathy muscles, suggesting a potent role of RyR1 depletion-induced ER stress in the pathogenesis of myopathies.
Topics: Animals; Humans; Mice; Endoplasmic Reticulum Stress; Muscle Fibers, Skeletal; Muscle, Skeletal; Muscular Diseases; Ryanodine Receptor Calcium Release Channel
PubMed: 37964752
DOI: 10.1002/jcsm.13349 -
Stem Cell Reports Oct 2023Skeletal muscle research is transitioning toward 3D tissue engineered in vitro models reproducing muscle's native architecture and supporting measurement of...
Skeletal muscle research is transitioning toward 3D tissue engineered in vitro models reproducing muscle's native architecture and supporting measurement of functionality. Human induced pluripotent stem cells (hiPSCs) offer high yields of cells for differentiation. It has been difficult to differentiate high-quality, pure 3D muscle tissues from hiPSCs that show contractile properties comparable to primary myoblast-derived tissues. Here, we present a transgene-free method for the generation of purified, expandable myogenic progenitors (MPs) from hiPSCs grown under feeder-free conditions. We defined a protocol with optimal hydrogel and medium conditions that allowed production of highly contractile 3D tissue engineered skeletal muscles with forces similar to primary myoblast-derived tissues. Gene expression and proteomic analysis between hiPSC-derived and primary myoblast-derived 3D tissues revealed a similar expression profile of proteins involved in myogenic differentiation and sarcomere function. The protocol should be generally applicable for the study of personalized human skeletal muscle tissue in health and disease.
Topics: Humans; Induced Pluripotent Stem Cells; Proteomics; Cells, Cultured; Muscle, Skeletal; Tissue Engineering; Myoblasts; Cell Differentiation
PubMed: 37774701
DOI: 10.1016/j.stemcr.2023.08.014 -
Journal of Cachexia, Sarcopenia and... Aug 2023Loss of muscle mass is linked with impaired quality of life and an increased risk of morbidity and premature mortality. Iron is essential for cellular processes such as...
BACKGROUND
Loss of muscle mass is linked with impaired quality of life and an increased risk of morbidity and premature mortality. Iron is essential for cellular processes such as energy metabolism, nucleotide synthesis and numerous enzymatic reactions. As the effects of iron deficiency (ID) on muscle mass and function are largely unknown, we aimed to assess the relation between ID and muscle mass in a large population-based cohort, and subsequently studied effects of ID on cultured skeletal myoblasts and differentiated myocytes.
METHODS
In a population-based cohort of 8592 adults, iron status was assessed by plasma ferritin and transferrin saturation, and muscle mass was estimated using 24-h urinary creatinine excretion rate (CER). The relationships of ferritin and transferrin saturation with CER were assessed by multivariable logistic regression. Furthermore, mouse C2C12 skeletal myoblasts and differentiated myocytes were subjected to deferoxamine with or without ferric citrate. Myoblast proliferation was measured with a colorimetric 5-bromo-2'-deoxy-uridine ELISA assay. Myocyte differentiation was assessed using Myh7-stainings. Myocyte energy metabolism, oxygen consumption rate and extracellular acidification rate were assessed using Seahorse mitochondrial flux analysis, and apoptosis rate with fluorescence-activated cell sorting. RNA sequencing (RNAseq) was used to identify ID-related gene and pathway enrichment in myoblasts and myocytes.
RESULTS
Participants in the lowest age- and sex-specific quintile of plasma ferritin (OR vs middle quintile 1.62, 95% CI 1.25-2.10, P < 0.001) or transferrin saturation (OR 1.34, 95% CI 1.03-1.75, P = 0.03) had a significantly higher risk of being in the lowest age- and sex-specific quintile of CER, independent of body mass index, estimated GFR, haemoglobin, hs-CRP, urinary urea excretion, alcohol consumption and smoking status. In C2C12 myoblasts, deferoxamine-induced ID reduced myoblast proliferation rate (P-trend <0.001) but did not affect differentiation. In myocytes, deferoxamine reduced myoglobin protein expression (-52%, P < 0.001) and tended to reduce mitochondrial oxygen consumption capacity (-28%, P = 0.10). Deferoxamine induced gene expression of cellular atrophy markers Trim63 (+20%, P = 0.002) and Fbxo32 (+27%, P = 0.048), which was reversed by ferric citrate (-31%, P = 0.04 and -26%, P = 0.004, respectively). RNAseq indicated that both in myoblasts and myocytes, ID predominantly affected genes involved in glycolytic energy metabolism, cell cycle regulation and apoptosis; co-treatment with ferric citrate reversed these effects.
CONCLUSIONS
In population-dwelling individuals, ID is related to lower muscle mass, independent of haemoglobin levels and potential confounders. ID impaired myoblast proliferation and aerobic glycolytic capacity, and induced markers of myocyte atrophy and apoptosis. These findings suggest that ID contributes to loss of muscle mass.
Topics: Animals; Female; Male; Mice; Atrophy; Cell Proliferation; Deferoxamine; Ferritins; Independent Living; Iron; Iron Deficiencies; Muscles; Myoblasts, Skeletal; Quality of Life; Transferrins; Humans; Adult
PubMed: 37386912
DOI: 10.1002/jcsm.13277 -
Nutrients Jan 2024Epicatechin is a polyphenol compound that promotes skeletal muscle differentiation and counteracts the pathways that participate in the degradation of proteins. Several... (Review)
Review
Epicatechin is a polyphenol compound that promotes skeletal muscle differentiation and counteracts the pathways that participate in the degradation of proteins. Several studies present contradictory results of treatment protocols and therapeutic effects. Therefore, the objective of this systematic review was to investigate the current literature showing the molecular mechanism and clinical protocol of epicatechin in muscle atrophy in humans, animals, and myoblast cell-line. The search was conducted in Embase, PubMed/MEDLINE, Cochrane Library, and Web of Science. The qualitative analysis demonstrated that there is a commonness of epicatechin inhibitory action in myostatin expression and atrogenes MAFbx, FOXO, and MuRF1. Epicatechin showed positive effects on follistatin and on the stimulation of factors related to the myogenic actions (MyoD, Myf5, and myogenin). Furthermore, the literature also showed that epicatechin can interfere with mitochondrias' biosynthesis in muscle fibers, stimulation of the signaling pathways of AKT/mTOR protein production, and amelioration of skeletal musculature performance, particularly when combined with physical exercise. Epicatechin can, for these reasons, exhibit clinical applicability due to the beneficial results under conditions that negatively affect the skeletal musculature. However, there is no protocol standardization or enough clinical evidence to draw more specific conclusions on its therapeutic implementation.
Topics: Animals; Humans; Catechin; Muscle Fibers, Skeletal; Muscle, Skeletal; Muscular Atrophy; MyoD Protein; TOR Serine-Threonine Kinases
PubMed: 38276564
DOI: 10.3390/nu16020326 -
Molecular & Cellular Proteomics : MCP Aug 2023Proteomic studies in facioscapulohumeral muscular dystrophy (FSHD) could offer new insight into disease mechanisms underpinned by post-transcriptional processes. We used...
Proteomic studies in facioscapulohumeral muscular dystrophy (FSHD) could offer new insight into disease mechanisms underpinned by post-transcriptional processes. We used stable isotope (deuterium oxide; DO) labeling and peptide mass spectrometry to investigate the abundance and turnover rates of proteins in cultured muscle cells from two individuals affected by FSHD and their unaffected siblings (UASb). We measured the abundance of 4420 proteins and the turnover rate of 2324 proteins in each (n = 4) myoblast sample. FSHD myoblasts exhibited a greater abundance but slower turnover rate of subunits of mitochondrial respiratory complexes and mitochondrial ribosomal proteins, which may indicate an accumulation of "older" less viable mitochondrial proteins in myoblasts from individuals affected by FSHD. Treatment with a 2'-O-methoxyethyl modified antisense oligonucleotide targeting exon 3 of the double homeobox 4 (DUX4) transcript tended to reverse mitochondrial protein dysregulation in FSHD myoblasts, indicating the effect on mitochondrial proteins may be a DUX4-dependent mechanism. Our results highlight the importance of post-transcriptional processes and protein turnover in FSHD pathology and provide a resource for the FSHD research community to explore this burgeoning aspect of FSHD.
Topics: Humans; Muscular Dystrophy, Facioscapulohumeral; Proteome; Proteomics; Homeodomain Proteins; Myoblasts; Muscle, Skeletal
PubMed: 37353005
DOI: 10.1016/j.mcpro.2023.100605