Recent studies suggested that cancer stem cells (CSCs) are capable of differentiating into endothelial cells and tumor endothelium may be derived from CSCs. But the mechanism remains unclear. We showed that vascular endothelial growth factor (VEGF) induced the expression of endothelial markers in breast cancer stem like cells (BCSLCs). In addition, the VEGF-treated BCSLCs formed capillary structure in matrigel and released vWF upon histamine treatment. The miR-27a expression was significantly increased in VEGF-treated BCSLCs. Antagonizing miR-27a by miR-27a anti-sense oligos (ASOs) in VEGF-treated BCSLCs led to decreased endothelial markers and function, while increasing miR-27a in BCSLCs resulted in enhanced endothelial properties. VEGF enhanced the transcription of miR-27a by increasing RUNX1 binding to miR-27a promoter. Increased miR-27a paralleled the reduced expression of ZBTB10, a known miR-27a target. Both expression of miR-27a and knockdown of ZBTB10 in BCSLCs promoted in vivo angiogenesis and tumor metastasis. Further, we demonstrated that VEGF-treated BCSLCs secreted more endogenous VEGF compared with undifferentiated BCSLCs. Thus, miR-27a promotes angiogenesis by mediating endothelial differentiation of BCSLCs and it may be a new target for anti-angiogenesis cancer therapy.