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Endothelial progenitor cells (EPCs) have been pursued as a potential cellular therapy for stroke and central nervous system injury. However, their underlying mechanisms remain to be fully defined. Recent experimental studies suggest that mitochondria may be released and transferred between cells. In this proof‐of‐concept study, we asked whether beneficial effects of EPCs may partly involve a mitochondrial phenomenon as well. First, EPC‐derived conditioned medium was collected and divided into supernatant and particle fractions after centrifugation. Electron microscopy, Western blots, and flow cytometry showed that EPCs were able to release mitochondria. ATP and oxygen consumption assays suggested that these extracellular mitochondria may still be functionally viable. Confocal microscopy confirmed that EPC‐derived extracellular mitochondria can be incorporated into normal brain endothelial cells. Adding EPC particles to brain endothelial cells promoted angiogenesis and decreased the permeability of brain endothelial cells. Next, we asked whether EPC‐derived mitochondria may be protective. As expected, oxygen–glucose deprivation (OGD) increased brain endothelial permeability. Adding EPC‐derived mitochondria particles to the damaged brain endothelium increased levels of mitochondrial protein TOM40, mitochondrial DNA copy number, and intracellular ATP. Along with these indirect markers of mitochondrial transfer, endothelial tightness was also restored after OGD. Taken together, these findings suggest that EPCs may support brain endothelial energetics, barrier integrity, and angiogenic function partly through extracellular mitochondrial transfer. S tem  C ells   2018;36:1404–1410

Protective Effects of Endothelial Progenitor Cell‐Derived Extracellular Mitochondria in Brain Endothelium