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Abstract Skeletal muscle fibers regulate surrounding endothelial cells (EC) via secretion of numerous angiogenic factors, including extracellular vesicles (SkM-EV). Muscle fibers are broadly classified as oxidative (OXI) or glycolytic (GLY) depending on their metabolic characteristics. OXI fibers secrete more pro-angiogenic factors and have greater capillary densities than GLY fibers. OXI muscle secretes more EV than GLY, however it is unknown whether muscle metabolic characteristics regulate EV contents and signaling potential. EVs were isolated from primarily oxidative or glycolytic muscle tissue from mice. MicroRNA (miR) contents were determined and endothelial cells were treated with OXI- and GLY-EV to investigate angiogenic signaling potential. There were considerable differences in miR contents between OXI- and GLY-EV and pathway analysis identified that OXI-EV miR were predicted to positively regulate multiple endothelial-specific pathways, compared to GLY-EV. OXI-EV improved in vitro angiogenesis, which may have been mediated through nitric oxide synthase (NOS) related pathways, as treatment of endothelial cells with a non-selective NOS inhibitor abolished the angiogenic benefits of OXI-EV. This is the first report to show widespread differences in miR contents between SkM-EV isolated from metabolically different muscle tissue and the first to demonstrate that oxidative muscle tissue secretes EV with greater angiogenic signaling potential than glycolytic muscle tissue.

Abstract Skeletal muscle capillarization is proportional to muscle fibre mitochondrial content and oxidative capacity. Skeletal muscle cells secrete many factors that regulate neighbouring capillary endothelial cells (ECs), including extracellular vesicles (SkM‐EVs). Peroxisome proliferator‐activated receptor γ coactivator 1‐α (PGC‐1α) regulates mitochondrial biogenesis and the oxidative phenotype in skeletal muscle. Skeletal muscle PGC‐1α also regulates secretion of multiple angiogenic factors, but it is unknown whether PGC‐1α regulates SkM‐EV release, contents and angiogenic signalling potential. PGC‐1α was overexpressed via adenovirus in primary human myotubes. EVs were collected from PGC‐1α‐overexpressing myotubes (PGC‐EVs) as well as from green fluorescent protein‐overexpressing myotubes (GFP‐EVs), and from untreated myotubes. EV release and select mRNA contents were measured from EVs. Additionally, ECs were treated with EVs to measure angiogenic potential of EVs in normal conditions and following an oxidative stress challenge. PGC‐1α overexpression did not impact EV release but did elevate EV content of mRNAs for several antioxidant proteins (nuclear factor erythroid 2‐related factor 2, superoxide dismutase 2, glutathione peroxidase). PGC‐EV treatment of cultured human umbilical vein endothelial cells (HUVECs) increased their proliferation (+36.6%), tube formation (length: +28.1%; number: +25.7%) and cellular viability (+52.9%), and reduced reactive oxygen species levels (−41%) compared to GFP‐EVs. Additionally, PGC‐EV treatment protected against tube formation impairments and induction of cellular senescence following acute oxidative stress. Overexpression of PGC‐1α in human myotubes increases the angiogenic potential of SkM‐EVs. These angiogenic benefits coincided with increased anti‐oxidative capacity of recipient HUVECs. High PGC‐1α expression in skeletal muscle may prompt the release of SkM‐EVs that support vascular redox homeostasis and angiogenesis.