Angiogenic Potential of Small Extracellular Vesicles Produced by Stimulated Mesenchymal Stromal Cells Under Hypoxic Conditions

Regenerative vascular medicine research has positioned mesenchymal stromal cells (MSCs) as a leading candidate to treat ischemic diseases. Recent studies have highlighted the emerging role of small extracellular vesicles (sEVs) produced by MSCs in their own potential. This study explores a strategy to improve the angiogenic potential of MSCs through the acquisition of endothelial features. Umbilical cord Wharton's jelly MSCs were cultured in fetal bovine serum-free endothelial growth medium under hypoxic conditions (SH-MSCs). sEVs were characterised by a multimodal approach: visualisation, count and particle size distribution, sEVs surface antigen, proangiogenic potential and ability to internalise into recipient cells. Compared with MSCs, SH-MSCs exhibited significant morphological and phenotypical change characterised by the up-regulation of CD31 and CD144 mRNA as well as a marked increase in sEVs secretion. MSC- and SH-MSC-derived sEVs had the capacity to internalise into endothelial cells, myoblasts and macrophages; exhibited a strong proangiogenic effect in vitro, particularly in promoting endothelial cell proliferation and pseudotube formation, likely due to an enriched cargo of angiogenic factors. These results highlight the dual benefit of hypoxia conditioning and endothelial differentiation of MSCs to optimise the angiogenic potential of their secreted sEVs, thus paving the way for innovative regenerative therapies in ischemic diseases.