miR-378a influences vascularization in skeletal muscles

AIMS: MicroRNA-378a, highly expressed in skeletal muscles, was demonstrated to affect myoblasts differentiation and to promote tumor angiogenesis. We hypothesized that miR-378a could play a pro-angiogenic role in skeletal muscle and may be involved in regeneration after ischemic injury in mice. METHODS AND RESULTS: Silencing of miR-378a in murine C2C12 myoblasts did not affect differentiation but impaired their secretory angiogenic potential towards endothelial cells. miR-378a knockout (miR-378a-/-) in mice resulted in a decreased number of CD31-positive blood vessels and arterioles in gastrocnemius muscle. In addition, diminished endothelial sprouting from miR-378a-/- aortic rings was shown. Interestingly, although fibroblast growth factor 1 (Fgf1) expression was decreased in miR-378a-/- muscles, this growth factor did not mediate the angiogenic effects exerted by miR-378a. In vivo, miR-378a knockout did not affect the revascularization of the ischemic muscles in both normo- and hyperglycemic mice subjected to femoral artery ligation (FAL). No difference in regenerating muscle fibers was detected between miR-378a-/- and miR-378+/+ mice. miR-378a expression temporarily declined in ischemic skeletal muscles of miR-378+/+ mice already on day 3 after FAL. At the same time, in the plasma, the level of miR-378a-3p was enhanced. Similar elevation of miR-378a-3p was reported in the plasma of patients with intermittent claudication in comparison to healthy donors. Local adeno-associated viral vectors (AAV)-based miR-378a overexpression was enough to improve the revascularization of the ischemic limb of wild type mice on day 7 after FAL, what was not reported after systemic delivery of vectors.In addition, the number of infiltrating CD45+ cells and macrophages (CD45+ CD11b+ F4/80+ Ly6G-) was higher in the ischemic muscles of miR-378a-/- mice, suggesting an anti-inflammatory action of miR-378a. CONCLUSIONS: Data indicates miR-378a role in the pro-angiogenic effect of myoblasts and vascularization of skeletal muscle. After the ischemic insult, the anti-angiogenic effect of miR-378a deficiency might be compensated by enhanced inflammation. A TRANSLATIONAL PERSPECTIVE: The study reports elevated expression of miR-378a-3p in the plasma of mice subjected to femoral artery ligation and in the plasma of patients with intermittent claudication. Thus, miR-378a may be considered as a marker of ischemic muscle injury.