Cytokine response of primary human myotubes in an in vitro exercise model.
Muscle contraction during exercise is a major stimulus for the release of peptides and proteins (myokines) that are supposed to take part in the beneficial 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 molecular response to exercise in a clearly defined model. We applied EPS for 24 h to primary human myotubes and studied the whole genome-wide transcriptional response 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 antioxidant defense. Multiplex immunoassays verified the translation of the transcriptional response of several cytokines into high-secretion levels (IL-6, IL-8, CXCL1, LIF, CSF3, IL-1B, and TNF) and the increased secretion of further myokines such as angiopoietin-like 4. Notably, EPS did not induce the release of creatine kinase. Inhibitor studies and immunoblotting revealed the participation of ERK1/2-, JNK-, and NF-κB-dependent pathways in the upregulation of myokines. To conclude, our data highlight the importance of skeletal muscle cells as endocrine cells. This in vitro exercise model is not only suitable to identify exercise-regulated myokines, but it might be applied to primary human myotubes obtained from different muscle biopsy donors to study the molecular mechanisms of the individual response to exercise.
Institute of Experimental Genetics, Helmholtz Zentrum Muenchen, German Research Center for Environmental Health, Neuherberg, Germany;
Mika
M
Scheler
Martin
M
Irmler
Stefan
S
Lehr
Sonja
S
Hartwig
Harald
H
Staiger
Hadi
H
Al-Hasani
Johannes
J
Beckers
Martin Hrabé
MH
de Angelis
Hans-Ulrich
HU
Häring
Cora
C
Weigert
D000328:Adult; D000075803:Angiopoietin-Like Protein 4; D042682:Angiopoietins / Q000378:metabolism; D049109:Cell Proliferation; D002478:Cells, Cultured; D054360:Chemokine CXCL1 / Q000378:metabolism; D003115:Colony-Stimulating Factors / Q000378:metabolism; D016207:Cytokines / Q000378:metabolism*; D004558:Electric Stimulation; D015444:Exercise / Q000502:physiology*; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D005260:Female; D006801:Humans; D053583:Interleukin-1beta / Q000378:metabolism; D015850:Interleukin-6 / Q000378:metabolism; D016209:Interleukin-8 / Q000378:metabolism; D048031:JNK Mitogen-Activated Protein Kinases / Q000378:metabolism; D053605:Leukemia Inhibitory Factor / Q000378:metabolism; D008297:Male; D009119:Muscle Contraction / Q000502:physiology*; D018485:Muscle Fibers, Skeletal / Q000378:metabolism*; D018482:Muscle, Skeletal / Q000378:metabolism*; D016328:NF-kappa B / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism
C474772:ANGPTL4 protein, human; D000075803:Angiopoietin-Like Protein 4; D042682:Angiopoietins; D054360:Chemokine CXCL1; D003115:Colony-Stimulating Factors; D016207:Cytokines; D053583:Interleukin-1beta; D015850:Interleukin-6; D016209:Interleukin-8; C507837:LIF protein, human; D053605:Leukemia Inhibitory Factor; D016328:NF-kappa B; D014409:Tumor Necrosis Factor-alpha; D048049:Extracellular Signal-Regulated MAP Kinases; D048031:JNK Mitogen-Activated Protein Kinases
ANGPTL4; ERK; IL-8; electric pulse stimulation; human myotubes; myokine