Regulation Of Myokine Expression In Exosome-like Vesicles By Electric Pulse Stimulation Of C2C12 Myotubes

Abstract

Physical exercise (PE) is a well-known non-pharmacological intervention to overcome chronic low-graded inflammation-induced sarcopenia through humoral factors. However, it is not fully elucidated whether PE-induced maintenance of muscle homeostasis against inflammation is associated with muscle cell-derived myokines and extracellular vesicles. PURPOSE: To determine the effects of inflammation of muscle cell on the myokine expression in exosome-like vesicles (ELVs), and the effects of electric pulse stimulation (EPS), as an exercise mimetic on the myokine expression using C2C12 myotubes. METHODS: Inflammation of C2C12 was induced by treatment of a cytokine mixture (CM, TNF-α+INF-γ), and insulin resistance was induced by palmitate (0.75 mM) for 24 hrs. ELVs were enriched from conditioned media by differential ultracentrifugation. EPS was set as 11.5V, 2m/s, 2Hz for 24 hrs. We considered P < 0.05 as significant, using GraphPad Prism ver 2.0 program. RESULTS: Treatment of C2C12 by CM significantly inhibited the expression of myogenic regulators (myogenic transcription factors, myogenic myokine, and signaling proteins), while induced the expression of atrophic factors (atrogin-1, myostatin and signaling proteins). In addition, the inflamed C2C12 myotubes released anti-myogenic ELVs which contain abundant myostatin and scanty level of decorin, comparing with control ELVs. When we stimulated C2C12 myotubes by EPS system, levels of myogenic regulators (MyoD and myogenin), myogenic myokines (FDNC5, decorin, FGF21 and cathepsin B), and metabolic function of myotubes were significantly increased, however the levels of myostatin and atrogin-1 were down-regulated. Furthermore, EPS increased the mitochondrial activity and activated mitochondrial biogenesis pathways. CONCLUSIONS: Inflammation, expression of anti-myogenic regulators and mitochondrial dysfunction are major contributors in metabolic diseases- or aging-induced sarcopenia. Therefore, our results suggested that activation of anti-myogenic activity in muscle cells by contraction (i.e., EPS in vitro and skeletal muscle contraction during PE in vivo) through myokine-containing ELVs may be a mechanism of beneficial effects of PE against sarcopenic factors.