Human Mesenchymal Stromal Cell Source and Culture Conditions Influence Extracellular Vesicle Angiogenic and Metabolic Effects on Human Endothelial Cells In Vitro.

BACKGROUND: Mesenchymal stem/stromal cell (MSC) derived extracellular vesicles (EVs) are a possible cell-free alternative to MSCs because they retain the regenerative potential of MSCs, while still mitigating some of their limitations (such as the possible elicitation of host immune responses). The promotion and restoration of angiogenesis, however, is an important component in treating trauma related injuries, and has not been fully explored with EVs. Herein we describe the effects of monolayer adipose-derived EVs (MAd-EVs), spheroid adipose-derived EVs (SAd-EVs), monolayer bone marrow-derived EVs (MBM-EVs), and spheroid bone marrow-derived EVs (SBM-EVs) on human umbilical vein endothelial cell (HUVEC) tube formation and mitochondrial respiration. METHODS: The successful isolation of EVs derived from adipose MSCs or bone marrow MSCs in monolayer or spheroid cultures was confirmed by NanoSight (particle size distribution) and Western blot (surface marker expression). EV angiogenic potential was measured using a 24-hour HUVEC tube formation assay. EV effects on HUVEC mitochondrial function were evaluated using the Seahorse respirometer machine. RESULTS: The number of junctions, branches, and the average length of branches formed at 24 hours of tube formation were significantly affected by cell and culture type; overall adipose-derived EVs outperformed bone marrow-derived EVs, and spheroid-derived EVs outperformed monolayer-derived EVs. Additionally, adipose-derived EVs resulted in significantly increased HUVEC mitochondrial maximal respiration and adenosine triphosphate (ATP) production, while only MBM-EVs negatively impacted HUVEC proton leak. CONCLUSIONS: Adipose-derived EVs promoted HUVEC tube formation significantly more than bone marrow-derived EVs, while also maximizing HUVEC mitochondria function. Results demonstrate that, as with MSC therapies, it is possible to tailor EV culture and production to optimize therapeutic potential. LEVEL OF EVIDENCE: Basic or Foundational Research; Original Article.