Skeletal muscle‐derived exosomes regulate endothelial cell functions via reactive oxygen species‐activated nuclear factor‐κB signalling

Abstract

New Findings What is the central question of this study?Capillary rarefaction is found in diabetic and aged muscle, whereas exercise increases skeletal muscle angiogenesis. The association implies a crosstalk between muscle cells and endothelial cells. The underlying mechanisms mediating the crosstalk between these cells remains to be elucidated fully. What is the main finding and its importance?Endothelial cell functions are regulated by skeletal muscle cell‐derived exosomes via a vascular endothelial growth factor‐independent pathway. This study reveals a new mechanism mediating the crosstalk between skeletal muscle cells and endothelial cells. Abstract Loss of skeletal muscle capillarization, known as capillary rarefaction, is found in type 2 diabetes, chronic heart failure and healthy ageing and is associated with impaired delivery of substrates to the muscle. However, the interaction and communication of skeletal muscle with endothelial cells in the regulation of capillaries surrounding the muscle remains elusive. Exosomes are a type of secreted extracellular vesicle containing mRNAs, proteins and, especially, microRNAs that exert paracrine and endocrine effects. In this study, we investigated whether skeletal muscle‐derived exosomes (SkM‐Exo) regulate the endothelial cell functions of angiogenesis. We demonstrated that C2C12 myotube‐derived exosomes improved endothelial cell functions, assessed by the proliferation, migration and tube formation of human umbilical vein endothelial cells (HUVECs), which were increased by 20, 23 and 40%, respectively, after SkM‐Exo exposure. The SkM‐Exo failed to activate HUVEC vascular endothelial growth factor (VEGF) signalling. The SkM‐Exo increased HUVEC reactive oxygen species and activated the nuclear factor‐κB pathway, suggesting that SkM‐Exo‐induced angiogenesis was mediated by a VEGF‐independent pathway. In addition, several angiogenic microRNAs were packaged in SkM‐Exo, with miR‐130a being particularly enriched and successfully transferred from SkM‐Exo to HUVECs. Delivery of miRNAs into endothelial cells might explain the enhancement of reactive oxygen species production and angiogenesis by SkM‐Exo. The potential angiogenic effect of SkM‐Exo could provide an effective therapy for promoting skeletal muscle angiogenesis in diseases characterized by capillary rarefaction or inadequate angiogenesis.

Multivesicular body and exosome pathway responses to acute exercise

“…Our small sample size limits the conclusions we can draw between miRNA processing and MVB biogenesis, but the evidence presented suggests a coordination of these cellular events. Recently, we found relative exosomal miRNA content was different from intracellular miRNA content in myotubes, implying exosomal content packaging is a regulated process (Nie et al., ). A better understanding of exosomal content packaging will likely be necessary to evaluate the impact of exercise and other interventions/conditions on both positive and negative cell‐to‐cell communications.…”

“…Skeletal muscle myotube‐derived EXOs decrease myoblast proliferation and increase myoblast differentiation into myotubes (Forterre et al., ,b). We recently found myotube‐derived exosomes increase endothelial cell proliferation, migration and tube formation, key features of angiogenesis (Nie et al., ). In mice, injected myotube‐derived EXOs can be incorporated into metabolic tissues such as liver and pancreas.…”

Skeletal muscle tissue secretes more extracellular vesicles than white adipose tissue and myofibers are a major source ex vivo but not in vivo

“…Much of the data described in this report was obtained with an ex vivo EV secretion assay using mouse tissue explants. There is precedent for the use of similar approaches in both SkM (1, 25) and WAT (37). For the studies described here, there were multiple advantages to using this ex vivo approach.…”

“…EVs from SkM cells are principally understood based on monocultures of proliferating myocytes and, to a lesser extent, differentiated myotubes (30). Proliferating C2C12 myocytes secrete EVs that improve endothelial function in vitro (25). EV secretion from C2C12 myocytes is increased by the saturated fatty acid palmitate and these palmitate-induced C2C12 myocytederived EVs impair insulin signaling in recipient cells (18).…”

“…Cell monocultures are a relatively simple system for studying SkM myocyte-derived EVs, but they lack the organization and extracellular matrix characteristic of tissues. Ex vivo incubation of SkM tissue explants has been shown to be an alternative approach to study EVs (1,4,18,25). This method yields EVs with a comparable size, morphology and composition to EVs from cultured cells or blood (1).…”