Fig. 5: Myonectin ameliorates DEX-induced atrophy of myotubes through the AMPK/PGC1α pathway. | Nature Communications

Fig. 5: Myonectin ameliorates DEX-induced atrophy of myotubes through the AMPK/PGC1α pathway.

From: Myonectin protects against skeletal muscle dysfunction in male mice through activation of AMPK/PGC1α pathway

Fig. 5: Myonectin ameliorates DEX-induced atrophy of myotubes through the AMPK/PGC1α pathway.

a, b C2C12 myotubes were pretreated with myonectin protein (5 µg/ml) or vehicle for 1 hour, followed by stimulation with DEX (100 µM) or vehicle for 24 hours. a Upper panels show representative images of myotubes. Scale bars show 200 μm. Lower panel shows quantification of myotube diameter. N = 5 in each group. b Upper panels show representative Western blot analyses of PGC1α, PGC1α4, IGF1 and tubulin. Lower panels show quantification of PGC1α, PGC1α4 and IGF1 signals normalized to tubulin signal. N = 4 in each group. c C2C12 myotubes were treated with myonectin protein (5 µg/ml) or vehicle for the indicated lengths of time. Left panels show representative Western blot analyses of pAMPK, AMPK, pACC, ACC, PGC1α and tubulin. Right graphs show quantitative analyses of the pAMPK/tubulin, pACC/tubulin and PGC1α/tubulin signal ratios. N = 4 in each group. d, e C2C12 myotubes were pretreated with adenoviral vectors expressing dominant-negative mutant form of AMPKα2 tagged by Myc (Ad-dnAMPK) or control Ad-β-gal 24 hours prior to myonectin treatment. d C2C12 myotubes were treated with myonectin protein (5 µg/ml) or vehicle for 1 hour. Left panels show representative Western blots of pACC, ACC, PGC1α, Myc-Tag and tubulin. Right panels show quantitative analyses of pACC/tubulin and PGC1α/tubulin signal ratios. N = 3 in each group. e C2C12 myotubes were pretreated with myonectin protein (5 µg/ml) or vehicle for 1 hour, followed by stimulation with DEX (100 µM) or vehicle for 24 hours. Upper panels show representative images of myotubes. Scale bars show 200 μm. Lower panel shows quantification of myotubes diameter. N = 4 in each group. f C2C12 myotubes were transduced with siRNA targeting PGC1α or unrelated control siRNA at 24 hours before myonectin treatment. Upper panels show representative images of myotubes. Lower panel shows quantification of myotubes diameter. N = 4 in each group. Scale bars show 200 μm. Data are presented as means ± SEM. One-way ANOVA with a post-hoc analysis for a–f was performed. DEX, dexamethasone. Ad-dnAMPK Adenovirus vectors containing the gene for dominant-negative AMPKα2, Ad-β-gal Adenovirus vectors containing the gene for β-galactosidase.

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