Skeletal Muscle最新文献

{"title":"Muscle injury-induced hypoxia alters the proliferation and differentiation potentials of muscle resident stromal cells.","authors":"Geneviève Drouin,&nbsp;Vanessa Couture,&nbsp;Marc-Antoine Lauzon,&nbsp;Frédéric Balg,&nbsp;Nathalie Faucheux,&nbsp;Guillaume Grenier","doi":"10.1186/s13395-019-0202-5","DOIUrl":"https://doi.org/10.1186/s13395-019-0202-5","url":null,"abstract":"<p><strong>Background: </strong>Trauma-induced heterotopic ossification (HO) is a complication that develops under three conditions: the presence of an osteogenic progenitor cell, an inducing factor, and a permissive environment. We previously showed that a mouse multipotent Sca1⁺ CD31^- Lin^- muscle resident stromal cell (mrSC) population is involved in the development of HO in the presence of inducing factors, members of the bone morphogenetic protein family. Interestingly, BMP9 unlike BMP2 causes HO only if the muscle is damaged by injection of cardiotoxin. Because acute trauma often results in blood vessel breakdown, we hypothesized that a hypoxic state in damaged muscles may foster mrSCs activation and proliferation and trigger differentiation toward an osteogenic lineage, thus promoting the development of HO.</p><p><strong>Methods: </strong>Three- to - six-month-old male C57Bl/6 mice were used to induce muscle damage by injection of cardiotoxin intramuscularly into the tibialis anterior and gastrocnemius muscles. mrSCs were isolated from damaged (hypoxic state) and contralateral healthy muscles and counted, and their osteoblastic differentiation with or without BMP2 and BMP9 was determined by alkaline phosphatase activity measurement. The proliferation and differentiation of mrSCs isolated from healthy muscles was also studied in normoxic incubator and hypoxic conditions. The effect of hypoxia on BMP synthesis and Smad pathway activation was determined by qPCR and/or Western blot analyses. Differences between normally distributed groups were compared using a Student's paired t test or an unpaired t test.</p><p><strong>Results: </strong>The hypoxic state of a severely damaged muscle increased the proliferation and osteogenic differentiation of mrSCs. mrSCs isolated from damaged muscles also displayed greater sensitivity to osteogenic signals, especially BMP9, than did mrSCs from a healthy muscle. In hypoxic conditions, mrSCs isolated from a control muscle were more proliferative and were more prone to osteogenic differentiation. Interestingly, Smad1/5/8 activation was detected in hypoxic conditions and was still present after 5 days, while Smad1/5/8 phosphorylation could not be detected after 3 h of normoxic incubator condition. BMP9 mRNA transcripts and protein levels were higher in mrSCs cultured in hypoxic conditions. Our results suggest that low-oxygen levels in damaged muscle influence mrSC behavior by facilitating their differentiation into osteoblasts. This effect may be mediated partly through the activation of the Smad pathway and the expression of osteoinductive growth factors such as BMP9 by mrSCs.</p><p><strong>Conclusion: </strong>Hypoxia should be considered a key factor in the microenvironment of damaged muscle that triggers HO.</p>","PeriodicalId":21747,"journal":{"name":"Skeletal Muscle","volume":"9 1","pages":"18"},"PeriodicalIF":4.9,"publicationDate":"2019-06-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1186/s13395-019-0202-5","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"37343635","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Culturing C2C12 myotubes on micromolded gelatin hydrogels accelerates myotube maturation.","authors":"Lance T Denes,&nbsp;Lance A Riley,&nbsp;Joseph R Mijares,&nbsp;Juan D Arboleda,&nbsp;Kendra McKee,&nbsp;Karyn A Esser,&nbsp;Eric T Wang","doi":"10.1186/s13395-019-0203-4","DOIUrl":"https://doi.org/10.1186/s13395-019-0203-4","url":null,"abstract":"<p><strong>Background: </strong>Skeletal muscle contributes to roughly 40% of lean body mass, and its loss contributes to morbidity and mortality in a variety of pathogenic conditions. Significant insights into muscle function have been made using cultured cells, in particular, the C2C12 myoblast line. However, differentiation of these cells in vitro typically yields immature myotubes relative to skeletal muscles in vivo. While many efforts have attempted to improve the maturity of cultured myotubes, including the use of bioengineered substrates, lack of molecular characterization has precluded their widespread implementation. This study characterizes morphological, molecular, and transcriptional features of C2C12 myotubes cultured on crosslinked, micropatterned gelatin substrates fabricated using previously established methods and compares them to myotubes grown on unpatterned gelatin or traditional plasticware.</p><p><strong>Methods: </strong>We used immunocytochemistry, SDS-PAGE, and RNAseq to characterize C2C12 myotubes grown on micropatterned gelatin hydrogels, unpatterned gelatin hydrogels, and typical cell culture substrates (i.e., plastic or collagen-coated glass) across a differentiation time course. The ability to form aligned sarcomeres and myofilament protein concentration was assessed. Additionally, the transcriptome was analyzed across the differentiation time course.</p><p><strong>Results: </strong>C2C12 myotubes grown on micropatterned gelatin hydrogels display an increased ability to form aligned sarcomeres as well as increased contractile protein content relative to myotubes cultured on unpatterned gelatin and plastic. Additionally, genes related to sarcomere formation and in vivo muscle maturation are upregulated in myotubes grown on micropatterned gelatin hydrogels relative to control myotubes.</p><p><strong>Conclusions: </strong>Our results suggest that growing C2C12 myotubes on micropatterned gelatin hydrogels accelerates sarcomere formation and yields a more fully matured myotube culture. Thus, the use of micropatterned hydrogels is a viable and simple approach to better model skeletal muscle biology in vitro.</p>","PeriodicalId":21747,"journal":{"name":"Skeletal Muscle","volume":"9 1","pages":"17"},"PeriodicalIF":4.9,"publicationDate":"2019-06-07","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1186/s13395-019-0203-4","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"37314450","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Congenital myopathy with hanging big toe due to homozygous myopalladin (MYPN) mutation.","authors":"Luciano Merlini,&nbsp;Patrizia Sabatelli,&nbsp;Manuela Antoniel,&nbsp;Valeria Carinci,&nbsp;Fabio Niro,&nbsp;Giuseppe Monetti,&nbsp;Annalaura Torella,&nbsp;Teresa Giugliano,&nbsp;Cesare Faldini,&nbsp;Vincenzo Nigro","doi":"10.1186/s13395-019-0199-9","DOIUrl":"https://doi.org/10.1186/s13395-019-0199-9","url":null,"abstract":"<p><strong>Background: </strong>Myopalladin (MYPN) is a component of the sarcomere that tethers nebulin in skeletal muscle and nebulette in cardiac muscle to alpha-actinin at the Z lines. Autosomal dominant MYPN mutations cause hypertrophic, dilated, or restrictive cardiomyopathy. Autosomal recessive MYPN mutations have been reported in only six families showing a mildly progressive nemaline or cap myopathy with cardiomyopathy in some patients.</p><p><strong>Case presentation: </strong>A consanguineous family with congenital to adult-onset muscle weakness and hanging big toe was reported. Muscle biopsy showed minimal changes with internal nuclei, type 1 fiber predominance, and ultrastructural defects of Z line. Muscle CT imaging showed marked hypodensity of the sartorius bilaterally and MRI scattered abnormal high-intensity areas in the internal tongue muscle and in the posterior cervical muscles. Cardiac involvement was demonstrated by magnetic resonance imaging and late gadolinium enhancement. Whole exome sequencing analysis identified a homozygous loss of function single nucleotide deletion in the exon 11 of the MYPN gene in two siblings. Full-length MYPN protein was undetectable on immunoblotting, and on immunofluorescence, its localization at the Z line was missed.</p><p><strong>Conclusions: </strong>This report extends the phenotypic spectrum of recessive MYPN-related myopathies showing: (1) the two patients had hanging big toe and the oldest one developed spine and hand contractures, none of these signs observed in the previously reported patients, (2) specific ultrastructural changes consisting in Z line fragmentation, but (3) no nemaline or caps on muscle pathology.</p>","PeriodicalId":21747,"journal":{"name":"Skeletal Muscle","volume":"9 1","pages":"14"},"PeriodicalIF":4.9,"publicationDate":"2019-05-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1186/s13395-019-0199-9","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"37278740","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A GDF11/myostatin inhibitor, GDF11 propeptide-Fc, increases skeletal muscle mass and improves muscle strength in dystrophic mdx mice.","authors":"Quan Jin,&nbsp;Chunping Qiao,&nbsp;Jianbin Li,&nbsp;Bin Xiao,&nbsp;Juan Li,&nbsp;Xiao Xiao","doi":"10.1186/s13395-019-0197-y","DOIUrl":"https://doi.org/10.1186/s13395-019-0197-y","url":null,"abstract":"<p><strong>Background: </strong>Growth differentiation factor 11 (GDF11) is a member of the transforming growth factor β superfamily. The GDF11 propeptide, which is derived from the GDF11 precursor protein, blocks the activity of GDF11 and its homolog, myostatin, which are both potent inhibitors of muscle growth. Thus, treatment with GDF11 propeptide may be a potential therapeutic strategy for diseases associated with muscle atrophy like sarcopenia and the muscular dystrophies. Here, we evaluate the impact of GDF11 propeptide-Fc (GDF11PRO-Fc) gene delivery on skeletal muscle in normal and dystrophic adult mice.</p><p><strong>Methods: </strong>A pull-down assay was used to obtain physical confirmation of a protein-protein interaction between GDF11PRO-Fc and GDF11 or myostatin. Next, differentiated C2C12 myotubes were treated with AAV6-GDF11PRO-Fc and challenged with GDF11 or myostatin to determine if GDF11PRO-Fc could block GDF11/myostatin-induced myotube atrophy. Localized expression of GDF11PRO-Fc was evaluated via a unilateral intramuscular injection of AAV9-GDF11PRO-Fc into the hindlimb of C57BL/6J mice. In mdx mice, intravenous injection of AAV9-GDF11PRO-Fc was used to achieve systemic expression. The impact of GDF11PRO-Fc on muscle mass, function, and pathological features were assessed.</p><p><strong>Results: </strong>GDF11PRO-Fc was observed to bind both GDF11 and myostatin. In C2C12 myotubes, expression of GDF11PRO-Fc was able to mitigate GDF11/myostatin-induced atrophy. Following intramuscular injection in C57BL/6J mice, increased grip strength and localized muscle hypertrophy were observed in the injected hindlimb after 10 weeks. In mdx mice, systemic expression of GDF11PRO-Fc resulted in skeletal muscle hypertrophy without a significant change in cardiac mass after 12 weeks. In addition, grip strength and rotarod latency time were improved. Intramuscular fibrosis was also reduced in treated mdx mice; however, there was no change seen in central nucleation, membrane permeability to serum IgG or serum creatine kinase levels.</p><p><strong>Conclusions: </strong>GDF11PRO-Fc induces skeletal muscle hypertrophy and improvements in muscle strength via inhibition of GDF11/myostatin signaling. However, GDF11PRO-Fc does not significantly improve the dystrophic pathology in mdx mice.</p>","PeriodicalId":21747,"journal":{"name":"Skeletal Muscle","volume":"9 1","pages":"16"},"PeriodicalIF":4.9,"publicationDate":"2019-05-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1186/s13395-019-0197-y","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"37001830","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Automated image-analysis method for the quantification of fiber morphometry and fiber type population in human skeletal muscle.","authors":"Perla C Reyes-Fernandez,&nbsp;Baptiste Periou,&nbsp;Xavier Decrouy,&nbsp;Fréderic Relaix,&nbsp;François Jérôme Authier","doi":"10.1186/s13395-019-0200-7","DOIUrl":"https://doi.org/10.1186/s13395-019-0200-7","url":null,"abstract":"<p><strong>Background: </strong>The quantitative analysis of muscle histomorphometry has been growing in importance in both research and clinical settings. Accurate and stringent assessment of myofibers' changes in size and number, and alterations in the proportion of oxidative (type I) and glycolytic (type II) fibers is essential for the appropriate study of aging and pathological muscle, as well as for diagnosis and follow-up of muscle diseases. Manual and semi-automated methods to assess muscle morphometry in sections are time-consuming, limited to a small field of analysis, and susceptible to bias, while most automated methods have been only tested in rodent muscle.</p><p><strong>Methods: </strong>We developed a new macro script for Fiji-ImageJ to automatically assess human fiber morphometry in digital images of the entire muscle. We tested the functionality of our method in deltoid muscle biopsies from a heterogeneous population of subjects with histologically normal muscle (male, female, old, young, lean, obese) and patients with dermatomyositis, necrotizing autoimmune myopathy, and anti-synthetase syndrome myopathy.</p><p><strong>Results: </strong>Our macro is fully automated, requires no user intervention, and demonstrated improved fiber segmentation by running a series of image pre-processing steps before the analysis. Likewise, our tool showed high accuracy, as compared with manual methods, for identifying the total number of fibers (r = 0.97, p < 0.001), fiber I and fiber II proportion (r = 0.92, p < 0.001), and minor diameter (r = 0.86, p < 0.001) while conducting analysis in ~ 5 min/sample. The performance of the macro analysis was maintained in pectoral and deltoid samples from subjects of different age, gender, body weight, and muscle status. The output of the analyses includes excel files with the quantification of fibers' morphometry and color-coded maps based on the fiber's size, which proved to be an advantageous feature for the fast and easy visual identification of location-specific atrophy and a potential tool for medical diagnosis.</p><p><strong>Conclusion: </strong>Our macro is reliable and suitable for the study of human skeletal muscle for research and for diagnosis in clinical settings providing reproducible and consistent analysis when the time is of the utmost importance.</p>","PeriodicalId":21747,"journal":{"name":"Skeletal Muscle","volume":"9 1","pages":"15"},"PeriodicalIF":4.9,"publicationDate":"2019-05-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1186/s13395-019-0200-7","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"37015630","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Targeted ablation of the cellular inhibitor of apoptosis 1 (cIAP1) attenuates denervation-induced skeletal muscle atrophy.","authors":"Neena Lala-Tabbert,&nbsp;Rim Lejmi-Mrad,&nbsp;Kristen Timusk,&nbsp;Marina Fukano,&nbsp;Janelle Holbrook,&nbsp;Martine St-Jean,&nbsp;Eric C LaCasse,&nbsp;Robert G Korneluk","doi":"10.1186/s13395-019-0201-6","DOIUrl":"https://doi.org/10.1186/s13395-019-0201-6","url":null,"abstract":"<p><strong>Background: </strong>Skeletal muscle atrophy is a pathological condition that contributes to morbidity in a variety of conditions including denervation, cachexia, and aging. Muscle atrophy is characterized as decreased muscle fiber cross-sectional area and protein content due, in part, to the proteolytic activities of two muscle-specific E3 ubiquitin ligases: muscle RING-finger 1 (MuRF1) and muscle atrophy F-box (MAFbx or Atrogin-1). The nuclear factor-kappa B (NF-κB) pathway has emerged as a critical signaling network in skeletal muscle atrophy and has become a prime therapeutic target for the treatment of muscle diseases. Unfortunately, none of the NF-κB targeting drugs are currently being used to treat these diseases, likely because of our limited knowledge and specificity, for muscle biology and disease. The cellular inhibitor of apoptosis 1 (cIAP1) protein is a positive regulator of tumor necrosis factor alpha (TNFα)-mediated classical NF-κB signaling, and cIAP1 loss has been shown to enhance muscle regeneration during acute and chronic injury.</p><p><strong>Methods: </strong>Sciatic nerve transection in wild-type, cIAP1-null and Smac mimetic compound (SMC)-treated mice was performed to investigate the role of cIAP1 in denervation-induced atrophy. Genetic in vitro models of C2C12 myoblasts and primary myoblasts were also used to examine the role of classical NF-κB activity in cIAP1-induced myotube atrophy.</p><p><strong>Results: </strong>We found that cIAP1 expression was upregulated in denervated muscles compared to non-denervated controls 14 days after denervation. Genetic and pharmacological loss of cIAP1 attenuated denervation-induced muscle atrophy and overexpression of cIAP1 in myotubes was sufficient to induce atrophy. The induction of myotube atrophy by cIAP1 was attenuated when the classical NF-κB signaling pathway was inhibited.</p><p><strong>Conclusions: </strong>These results demonstrate the cIAP1 is an important mediator of NF-κB/MuRF1 signaling in skeletal muscle atrophy and is a promising therapeutic target for muscle wasting diseases.</p>","PeriodicalId":21747,"journal":{"name":"Skeletal Muscle","volume":"9 1","pages":"13"},"PeriodicalIF":4.9,"publicationDate":"2019-05-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1186/s13395-019-0201-6","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"36995343","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Preclinical rationale for entinostat in embryonal rhabdomyosarcoma.","authors":"Narendra Bharathy, Noah E Berlow, Eric Wang, Jinu Abraham, Teagan P Settelmeyer, Jody E Hooper, Matthew N Svalina, Zia Bajwa, Martin W Goros, Brian S Hernandez, Johannes E Wolff, Ranadip Pal, Angela M Davies, Arya Ashok, Darnell Bushby, Maria Mancini, Christopher Noakes, Neal C Goodwin, Peter Ordentlich, James Keck, Douglas S Hawkins, Erin R Rudzinski, Atiya Mansoor, Theodore J Perkins, Christopher R Vakoc, Joel E Michalek, Charles Keller","doi":"10.1186/s13395-019-0198-x","DOIUrl":"10.1186/s13395-019-0198-x","url":null,"abstract":"<p><strong>Background: </strong>Rhabdomyosarcoma (RMS) is the most common soft tissue sarcoma in the pediatric cancer population. Survival among metastatic RMS patients has remained dismal yet unimproved for years. We previously identified the class I-specific histone deacetylase inhibitor, entinostat (ENT), as a pharmacological agent that transcriptionally suppresses the PAX3:FOXO1 tumor-initiating fusion gene found in alveolar rhabdomyosarcoma (aRMS), and we further investigated the mechanism by which ENT suppresses PAX3:FOXO1 oncogene and demonstrated the preclinical efficacy of ENT in RMS orthotopic allograft and patient-derived xenograft (PDX) models. In this study, we investigated whether ENT also has antitumor activity in fusion-negative eRMS orthotopic allografts and PDX models either as a single agent or in combination with vincristine (VCR).</p><p><strong>Methods: </strong>We tested the efficacy of ENT and VCR as single agents and in combination in orthotopic allograft and PDX mouse models of eRMS. We then performed CRISPR screening to identify which HDAC among the class I HDACs is responsible for tumor growth inhibition in eRMS. To analyze whether ENT treatment as a single agent or in combination with VCR induces myogenic differentiation, we performed hematoxylin and eosin (H&E) staining in tumors.</p><p><strong>Results: </strong>ENT in combination with the chemotherapy VCR has synergistic antitumor activity in a subset of fusion-negative eRMS in orthotopic \"allografts,\" although PDX mouse models were too hypersensitive to the VCR dose used to detect synergy. Mechanistic studies involving CRISPR suggest that HDAC3 inhibition is the primary mechanism of cell-autonomous cytoreduction in eRMS. Following cytoreduction in vivo, residual tumor cells in the allograft models treated with chemotherapy undergo a dramatic, entinostat-induced (70-100%) conversion to non-proliferative rhabdomyoblasts.</p><p><strong>Conclusion: </strong>Our results suggest that the targeting class I HDACs may provide a therapeutic benefit for selected patients with eRMS. ENT's preclinical in vivo efficacy makes ENT a rational drug candidate in a phase II clinical trial for eRMS.</p>","PeriodicalId":21747,"journal":{"name":"Skeletal Muscle","volume":"9 1","pages":"12"},"PeriodicalIF":5.3,"publicationDate":"2019-05-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6528217/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"37262066","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Exogenous expression of the glycosyltransferase LARGE1 restores α-dystroglycan matriglycan and laminin binding in rhabdomyosarcoma.","authors":"Daniel Beltrán,&nbsp;Mary E Anderson,&nbsp;Narendra Bharathy,&nbsp;Teagan P Settelmeyer,&nbsp;Matthew N Svalina,&nbsp;Zia Bajwa,&nbsp;John F Shern,&nbsp;Sakir H Gultekin,&nbsp;Marco A Cuellar,&nbsp;Takahiro Yonekawa,&nbsp;Charles Keller,&nbsp;Kevin P Campbell","doi":"10.1186/s13395-019-0195-0","DOIUrl":"https://doi.org/10.1186/s13395-019-0195-0","url":null,"abstract":"<p><strong>Background: </strong>α-Dystroglycan is the highly glycosylated component of the dystrophin-glycoprotein complex (DGC) that binds with high-affinity to extracellular matrix (ECM) proteins containing laminin-G-like (LG) domains via a unique heteropolysaccharide [-GlcA-beta1,3-Xyl-alpha1,3-]_n called matriglycan. Changes in expression of components of the DGC or in the O-glycosylation of α-dystroglycan result in muscular dystrophy but are also observed in certain cancers. In mice, the loss of either of two DGC proteins, dystrophin or α-sarcoglycan, is associated with a high incidence of rhabdomyosarcoma (RMS). In addition, glycosylation of α-dystroglycan is aberrant in a small cohort of human patients with RMS. Since both the glycosylation of α-dystroglycan and its function as an ECM receptor require over 18 post-translational processing enzymes, we hypothesized that understanding its role in the pathogenesis of RMS requires a complete analysis of the expression of dystroglycan-modifying enzymes and the characterization of α-dystroglycan glycosylation in the context of RMS.</p><p><strong>Methods: </strong>A series of cell lines and biopsy samples from human and mouse RMS were analyzed for the glycosylation status of α-dystroglycan and for expression of the genes encoding the responsible enzymes, in particular those required for the addition of matriglycan. Furthermore, the glycosyltransferase LARGE1 was ectopically expressed in RMS cells to determine its effects on matriglycan modifications and the ability of α-dystroglycan to function as a laminin receptor.</p><p><strong>Results: </strong>Immunohistochemistry and immunoblotting of a collection of primary RMS tumors show that although α-dystroglycan is consistently expressed and glycosylated in these tumors, α-dystroglycan lacks matriglycan and the ability to bind laminin. Similarly, in a series of cell lines derived from human and mouse RMS, α-dystroglycan lacks matriglycan modification and the ability to bind laminin. RNAseq data from RMS cell lines was analyzed for expression of the genes known to be involved in α-dystroglycan glycosylation, which revealed that, for most cell lines, the lack of matriglycan can be attributed to the downregulation of the dystroglycan-modifying enzyme LARGE1. Ectopic expression of LARGE1 in these cell cultures restored matriglycan to levels comparable to those in muscle and restored high-affinity laminin binding to α-dystroglycan.</p><p><strong>Conclusions: </strong>Collectively, our findings demonstrate that a lack of matriglycan on α-dystroglycan is a common feature in RMS due to the downregulation of LARGE1, and that ectopic expression of LARGE1 can restore matriglycan modifications and the ability of α-dystroglycan to function as an ECM receptor.</p>","PeriodicalId":21747,"journal":{"name":"Skeletal Muscle","volume":"9 1","pages":"11"},"PeriodicalIF":4.9,"publicationDate":"2019-05-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1186/s13395-019-0195-0","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"37211067","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Muscle Gene Sets: a versatile methodological aid to functional genomics in the neuromuscular field.","authors":"Apostolos Malatras,&nbsp;Stephanie Duguez,&nbsp;William Duddy","doi":"10.1186/s13395-019-0196-z","DOIUrl":"10.1186/s13395-019-0196-z","url":null,"abstract":"<p><strong>Background: </strong>The approach of building large collections of gene sets and then systematically testing hypotheses across these collections is a powerful tool in functional genomics, both in the pathway analysis of omics data and to uncover the polygenic effects associated with complex diseases in genome-wide association study. The Molecular Signatures Database includes collections of oncogenic and immunologic signatures enabling researchers to compare transcriptional datasets across hundreds of previous studies and leading to important insights in these fields, but such a resource does not currently exist for neuromuscular research. In previous work, we have shown the utility of gene set approaches to understand muscle cell physiology and pathology.</p><p><strong>Methods: </strong>Following a systematic survey of public muscle data, we passed gene expression profiles from 4305 samples through a robust pre-processing and standardized data analysis pipeline. Two hundred eighty-two samples were discarded based on a battery of rigorous global quality controls. From among the remaining studies, 578 comparisons of interest were identified by a combination of text mining and manual curation of the study meta-data. For each comparison, significantly dysregulated genes (FDR adjusted p < 0.05) were identified.</p><p><strong>Results: </strong>Lists of dysregulated genes were divided between upregulated and downregulated to give 1156 Muscle Gene Sets (MGS). This resource is available for download ( www.sys-myo.com/muscle_gene_sets ) and is accessible through three commonly used functional genomics platforms (GSEA, EnrichR, and WebGestalt). Basic guidance and recommendations are provided for the use of MGS through these platforms. In addition, consensus muscle gene sets were created to capture the overlap between the results of similar studies, and analysis of these highlighted the potential for novel disease-relevant findings.</p><p><strong>Conclusions: </strong>The MGS resource can be used to investigate the behaviour of any list of genes across previous comparisons of muscle conditions, to compare previous studies to one another, and to explore the functional relationship of muscle dysregulation to the Gene Ontology. Its major intended use is in enrichment testing for functional genomics analysis.</p>","PeriodicalId":21747,"journal":{"name":"Skeletal Muscle","volume":"9 1","pages":"10"},"PeriodicalIF":4.9,"publicationDate":"2019-05-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1186/s13395-019-0196-z","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"37209501","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"RNA-sequencing reveals altered skeletal muscle contraction, E3 ligases, autophagy, apoptosis, and chaperone expression in patients with critical illness myopathy.","authors":"Monica Llano-Diez,&nbsp;Wen Fury,&nbsp;Haruka Okamoto,&nbsp;Yu Bai,&nbsp;Jesper Gromada,&nbsp;Lars Larsson","doi":"10.1186/s13395-019-0194-1","DOIUrl":"https://doi.org/10.1186/s13395-019-0194-1","url":null,"abstract":"<p><strong>Background: </strong>Critical illness myopathy (CIM) is associated with severe skeletal muscle wasting and impaired function in intensive care unit (ICU) patients. The mechanisms underlying CIM remain incompletely understood. To elucidate the biological activities occurring at the transcriptional level in the skeletal muscle of ICU patients with CIM, the gene expression profiles, potential upstream regulators, and enrichment pathways were characterized using RNA sequencing (RNA-seq). We also compared the skeletal muscle gene signatures in ICU patients with CIM and genes perturbed by mechanical loading in one leg of the ICU patients, with an aim of reducing the loss of muscle function.</p><p><strong>Methods: </strong>RNA-seq was used to assess gene expression changes in tibialis anterior skeletal muscle samples from seven critically ill, immobilized, and mechanically ventilated ICU patients with CIM and matched control subjects. We also examined skeletal muscle gene expression for both legs of six ICU patients with CIM, where one leg was mechanically loaded for 10 h/day for an average of 9 days.</p><p><strong>Results: </strong>In total, 6257 of 17,221 detected genes were differentially expressed (84% upregulated; p < 0.05 and fold change ≥ 1.5) in skeletal muscle from ICU patients with CIM when compared to control subjects. The differentially expressed genes were highly associated with gene changes identified in patients with myopathy, sepsis, long-term inactivity, polymyositis, tumor, and repeat exercise resistance. Upstream regulator analysis revealed that the CIM signature could be a result of the activation of MYOD1, p38 MAPK, or treatment with dexamethasone. Passive mechanical loading only reversed expression of 0.74% of the affected genes (46 of 6257 genes).</p><p><strong>Conclusions: </strong>RNA-seq analysis revealed that the marked muscle atrophy and weakness observed in ICU patients with CIM were associated with the altered expression of genes involved in muscle contraction, newly identified E3 ligases, autophagy and calpain systems, apoptosis, and chaperone expression. In addition, MYOD1, p38 MAPK, and dexamethasone were identified as potential upstream regulators of skeletal muscle gene expression in ICU patients with CIM. Mechanical loading only marginally affected the skeletal muscle transcriptome profiling of ICU patients diagnosed with CIM.</p>","PeriodicalId":21747,"journal":{"name":"Skeletal Muscle","volume":"9 1","pages":"9"},"PeriodicalIF":4.9,"publicationDate":"2019-04-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1186/s13395-019-0194-1","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"37334695","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}