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Endothelial glucose transporter 1 (GLUT1) is a definitive and diagnostic marker for infantile hemangioma (IH), a vascular tumor of infancy. To date, GLUT1‐positive endothelial cells in IH have not been quantified nor directly isolated and studied. We isolated GLUT1‐positive and GLUT1‐negative endothelial cells from IH specimens and characterized their proliferation, differentiation, and response to propranolol, a first‐line therapy for IH, and to rapamycin, an mTOR pathway inhibitor used to treat an increasingly wide array of proliferative disorders. Although freshly isolated GLUT1‐positive cells, selected using anti‐GLUT1 magnetic beads, expressed endothelial markers CD31, VE‐Cadherin, and vascular endothelial growth factor receptor 2, they converted to a mesenchymal phenotype after 3 weeks in culture. In contrast, GLUT1‐negative endothelial cells exhibited a stable endothelial phenotype in vitro. GLUT1‐selected cells were clonogenic when plated as single cells and could be induced to redifferentiate into endothelial cells, or into pericytes/smooth muscle cells or into adipocytes, indicating a stem cell‐like phenotype. These data demonstrate that, although they appear and function in the tumor as bona fide endothelial cells, the GLUT1‐positive endothelial cells display properties of facultative stem cells. Pretreatment with rapamycin for 4 days significantly slowed proliferation of GLUT1‐selected cells, whereas propranolol pretreatment had no effect. These results reveal for the first time the facultative nature of GLUT1‐positive endothelial cells in IH. S tem  C ells   2015;33:133–145

Glucose Transporter 1‐Positive Endothelial Cells in Infantile Hemangioma Exhibit Features of Facultative Stem Cells