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Annotating the secretome of mature primary human muscle cells

ABSTRACT: The Skeletal muscle is a metabolic active tissue that secretes various proteins. These so called myokines act auto-, para- and endocrine affecting muscle physiology and exert systemic effects on other tissues and organs. Myokines are also described to play a crucial role in the pathophysiology of metabolic diseases. Combining three different mass spectrometry based non-targeted and one antibody based targeted approach we investigated the secretome of differentiated primary human skeletal muscle cells derived from adult donors. A total of 548 non-redundant proteins were detected by combined proteomic profiling. Expression was confirmed on mRNA level for 501. Stringent consecutive filtering recruiting several database, i.e. SignalP, SecretomeP and ER_retention signals, the computational analysis assigned 306 as secretory proteins including 33 potentially novel myokines. This comprehensive profiling study of the human skeletal muscle secretome expands our knowledge of the composition of the human myokinome and further highlights the pivotal role of myokines in the regulation of multiple biological processes. We performed gene expression microarray analysis of primary human myotubes derived from twelve healthy individuals

ORGANISM(S): Homo sapiens

SUBMITTER: Johannes Beckers

PROVIDER: E-GEOD-45473 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Annotating the secretome of mature primary human muscle cells

Project description:The Skeletal muscle is a metabolic active tissue that secretes various proteins. These so called myokines act auto-, para- and endocrine affecting muscle physiology and exert systemic effects on other tissues and organs. Myokines are also described to play a crucial role in the pathophysiology of metabolic diseases. Combining three different mass spectrometry based non-targeted and one antibody based targeted approach we investigated the secretome of differentiated primary human skeletal muscle cells derived from adult donors. A total of 548 non-redundant proteins were detected by combined proteomic profiling. Expression was confirmed on mRNA level for 501. Stringent consecutive filtering recruiting several database, i.e. SignalP, SecretomeP and ER_retention signals, the computational analysis assigned 306 as secretory proteins including 33 potentially novel myokines. This comprehensive profiling study of the human skeletal muscle secretome expands our knowledge of the composition of the human myokinome and further highlights the pivotal role of myokines in the regulation of multiple biological processes.

2013-12-03 | GSE45473 | GEO

Stem cell secretome-treatment improves whole-body metabolism, reduces adiposity and promotes skeletal muscle function in aged mice

Project description:Aging coincides with the progressive loss of muscle mass and strength, increased adiposity, and diminished physical function. Accordingly, interventions aimed at improving muscle, metabolic, and/or physical health are of interest to mitigate the adverse effects of aging. In this study, we tested a stem-cell secretome product which contains a host of growth, cytoskeletal remodeling, and immunomodulatory factors. We examined the effects of 4 weeks of bi-weekly unilateral intramuscular secretome injections (quadriceps) in ambulatory aged male C57Bl/6 mice (22-23 months) compared to saline injected aged-matched controls. After 4 weeks of treatment, secretome delivery substantially increased whole body lean mass and decreased fat mass corresponding to higher fiber cross sectional area and smaller adipocyte size, respectively. Secretome-treated mice also displayed improved whole-body grip strength and rotarod performance and had higher energy expenditure and physical activity levels compared to control mice. Furthermore, secretome-treated mice had greater skeletal muscle Pax7+ cell abundance, capillary density, and collagen turnover. Finally, the secretome product reduced lipid content and lipid droplet size in adipocytes in vitro. These data suggest intramuscular treatment secretome improves whole-body metabolism, physical function, and remodels skeletal muscle and adipose tissue in aged mice.

2024-07-31 | GSE242211 | GEO

Cytokines dominate the secretome of primary human myotubes in an in vitro exercise model

Project description:Muscle contraction during exercise is the major stimulus for the release of peptides and proteins (myokines) that are supposed to take part in the benefical adaptation to exercise. We hypothesize that application of an in vitro exercise stimulus as electric pulse stimulation (EPS) to human myotubes enables the investigation of the human muscle secretome in a clearly defined model. We applied EPS for 24 h to primary human myotubes and studied the whole genome-wide transcriptional response and as well as the release of candidate myokines. We observed 183 differentially regulated transcripts with fold-changes > 1.3. The transcriptional response resembles several properties of the in vivo situation in the skeletal muscle after endurance exercise, namely significant enrichment of pathways associated with interleukin and chemokine signaling, lipid metabolism, and anti-oxidant defense; notably without increased release of creatin kinase. Multiplex immunoassays verified the translation of the transcriptional response of several myokines into high secretion levels (IL6, IL8, CXCL1, LIF, CSF3, IL1B, TNF) and identified an increased secretion of additional cytokines (IL2, IL4, IL13, IL17A). Inhibitor studies and immunoblotting revealed the participation of ERK1/2 and AMPK dependent pathways in the upregulation of myokines. To conclude our data highlight the importance of skeletal muscle cells per se as endocrine cells. This in vitro exercise model is not only suitable to identify known and novel exercise-regulated myokines but it might be applied to primary human myotubes obtained from different muscle biopsy donors to study molecular mechanisms of the individual response to exercise. We performed gene expression microarray analysis of myotubes in vitro stimulated by EPS and control myotubes

2013-10-09 | E-GEOD-44051 | biostudies-arrayexpress

Widespread multi-targeted therapy resistance via drug-induced secretome fucosylation

Project description:Cancer secretome is a reservoir for aberrant glycosylation. How therapies alter this post59 translational cancer hallmark and the consequences thereof remain elusive. Here we show that an elevated secretome fucosylation is a pan-cancer signature of both response and resistance to multiple targeted therapies. Large-scale pharmacogenomics revealed that fucosylation genes display widespread association with resistance to these therapies. In both cancer cell cultures and patients, targeted kinase inhibitors distinctively induced core fucosylation of secreted proteins less than 60 kDa. Label-free proteomics of N-glycomes revealed that fucosylation of the antioxidant PON1 is a critical component of the therapy66 induced secretome. Core fucosylation in the Golgi impacts PON1 stability and folding prior to secretion, promoting a more degradation-resistant PON1. Non-specific and PON1-specific secretome deglycosylation both limited the expansion of resistant clones in a tumor regression model. Our findings demonstrate that core fucosylation is a common modification indirectly induced by targeted therapies that paradoxically promotes resistance.

2023-04-05 | GSE160205 | GEO

Cytokines dominate the secretome of primary human myotubes in an in vitro exercise model

2013-10-09 | GSE44051 | GEO

RNA sequencing of TA muscle in miR-501KO mice and control littermates at day 6 of CTX-induced muscle regeneration

Project description:Adult muscle stem cells show a high transcriptional and clonal heterogeneity during ageing. Whether microRNAs contribute to this diversity is not known. Here we use global and tissue-specific genetic deletion of microRNA (miR)-501 that is highly enriched in activated myogenic progenitor cells 3 to address this question. Deletion of miR-501 reduced myofiber size and resilience of muscle fibers to injury and exercise in adult skeletal muscle. miR-501 elicits this effect by changing sarcomeric gene expression in newly built myofibers in vivo.

2023-03-16 | GSE199111 | GEO

Transcription profiling by array of the secretome from human myotube and myotube derived vesicles

Project description:Human satellite cells were isolated from muscle biopsies and expanded in vitro. Cells were induced to differentiate into branched multinucleated myotubes by serum depletion and were harvested from three independent cultures after 72 hours. Total RNA was isolated, amplified, labelled and hybridised to Illumina Human WG-6v3 arrays. Data were analysed to identify a "virtual secretome". These data and proteomic data from similar cultures suggested that differentiating muscle cultures produce microvesicles - which were characterised in the ensuing experiment. Human satellite cells were cultured as before (duplicate cultures) and culture supernatants were subjected to a differential centrifugation procedure, resulting in isolation of two distinct vesicle populations: exosomes and microparticles. Total RNA was isolated from these vesicle populations and the cells from which they were derived. RNA was amplified, labelled and hybridised to Illumina Human HT12v3 arrays. Data from myotubes, exosomes and microparticles were analysed to identify transcripts present, differential expression and functional interpretation of transcriptomes. IDAT files are available on the ArrayExpress ftp site in the additional.zip file.

2011-04-30 | E-MTAB-423 | biostudies-arrayexpress

Deciphering the determinants of recombinant protein yield across the human secretome

Project description:Mammalian cells are critical hosts for the production of most therapeutic proteins and many proteins for biomedical research. While cell line engineering and bioprocess optimization have yielded high protein titers of some recombinant proteins, many proteins remain difficult to express. Here we use systems biology methods to deciper the factors influencing yields in Chinese hamster ovary (CHO) cells as they produce proteins from the human secretome.

2024-02-23 | GSE225989 | GEO

A myogenin+/CD74+ adult muscle stem cell population during regeneration of skeletal muscle lacking miR-501

Project description:Adult muscle stem cells show a high transcriptional and clonal heterogeneity during ageing. Whether microRNAs contribute to this diversity is not known. Here we use tissue-specific genetic deletion of microRNA (miR)-501 that is highly enriched in activated myogenic progenitor cells to address this question. Single-cell sequencing revealed a novel subpopulation of committed myogenic progenitor cells in miR-501 knockout mice that express high levels of terminal differentiation markers and inflammatory genes such as CD74. Our data show that stem cell heterogeneity in skeletal muscle during ageing is under miRNA control.

2023-03-21 | GSE200501 | GEO

Mining the secretome of C2C12 muscle cells

Project description:Secretome analysis faces several challenges, including detection of low abundant proteins and the discrimination of bona fide secreted proteins from false-positive identifications stemming from cell leakage or serum. Here, we developed a two-step secretomics approach and applied it for the analysis of secreted proteins of C2C12 skeletal muscle cells as the skeletal muscle has been identified as an important endocrine organ secreting myokines as active signaling molecules. First, we compared culture supernatants and corresponding cell lysates by high-resolution MS-based proteomics and label-free quantification. We found 672 protein groups as candidate secreted proteins, as they showed a higher abundance in the secretome. By Brefeldin A mediated blocking of classical secretory processes, we estimate a sensitivity of > 80% for the detection of classical secreted proteins by our experimental approach. In the second step, peptide level information was integrated with UniProt based protein information by the bioinformatics tool “lysate and secretome peptide feature plotter” (LSPFP) to detect proteolytic protein processing events which might occur during secretion e.g. As a proof on concept, we identified truncations of the cytoplasmic part of the protein Plexin-B2. Our workflow provides an efficient combination of experimental workflow and data analysis to identify putative secreted and proteolytic processed proteins.

2018-01-24 | PXD007527 | Pride

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