Open AccessCharacterization and Functional Analysis of Extracellular Vesicles and Muscle-Abundant miRNAs (miR-1, miR-133a, and miR-206) in C2C12 Myocytes and mdx Mice
Author(s) -
Yasunari Matsuzaka,
Jun Tanihata,
Hirofumi Komaki,
Akihiko Ishiyama,
Yasushi Oya,
Urs T. Rüegg,
Shin’ichi Takeda,
Kazuo Hashido
Publication year - 2016
Publication title -
plos one
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.99
H-Index - 332
ISSN - 1932-6203
DOI - 10.1371/journal.pone.0167811
Subject(s) - c2c12 , myogenesis , myocyte , duchenne muscular dystrophy , skeletal muscle , biology , microbiology and biotechnology , exosome , microvesicles , muscular dystrophy , mdx mouse , microrna , endocrinology , dystrophin , biochemistry , genetics , gene
Duchenne muscular dystrophy (DMD) is a progressive neuromuscular disorder. Here, we show that the CD63 antigen, which is located on the surface of extracellular vesicles (EVs), is associated with increased levels of muscle-abundant miRNAs, namely myomiRs miR-1, miR-133a, and miR-206, in the sera of DMD patients and mdx mice. Furthermore, the release of EVs from the murine myoblast C 2 C 12 cell line was found to be modulated by intracellular ceramide levels in a Ca 2+ -dependent manner. Next, to investigate the effects of EVs on cell survival, C 2 C 12 myoblasts and myotubes were cultured with EVs from the sera of mdx mice or C 2 C 12 cells overexpressing myomiRs in presence of cellular stresses. Both the exposure of C 2 C 12 myoblasts and myotubes to EVs from the serum of mdx mice, and the overexpression of miR-133a in C 2 C 12 cells in presence of cellular stress resulted in a significant decrease in cell death. Finally, to assess whether miRNAs regulate skeletal muscle regeneration in vivo , we intraperitoneally injected GW4869 (an inhibitor of exosome secretion) into mdx mice for 5 and 10 days. Levels of miRNAs and creatine kinase in the serum of GW4869-treated mdx mice were significantly downregulated compared with those of controls. The tibialis anterior muscles of the GW4869-treated mdx mice showed a robust decrease in Evans blue dye uptake. Collectively, these results indicate that EVs and myomiRs might protect the skeletal muscle of mdx mice from degeneration.