Stem cell therapy may transform the treatment of acute and chronic heart disease, with an anticipated impact rivalling the results of revascularization and device therapies. Human embryonic stem cells and their cardiac derivatives are currently being developed as sources of tissue-specific cells for organ regeneration. In this preclinical study, we tested the feasibility and efficacy of human embryonic stem cell-derived endothelial cells (hESC-EC) engraftment. To favour endothelial formation, undifferentiated H7 embryonic stem cells were treated with serum and endothelial growth factors and CD31 + cell population were sorted subsequently by FACS from differentiating cultures. A highly expandable population of hESC-EC was obtained thereby which express endothelial cell markers and form vascular networks in vitro. Using extracellular matrix Matrigel as scaffold, hESC-EC were implanted subcutaneously or intramyocardially into 3-months-old athymic nude rats. For studying angiogenesis, a small animal imaging system, a high-resolution NanoSPECT/CT was used to acquire whole-body SPECT/CT images. By radiolabelled albumin, a local significant increase in perfusion was detected at the grafted sites after 2 weeks, suggesting the functional incorporation of hESC-EC into the microvasculature. Post-mortem histology further showed that hESC-EC survive, engraft successfully and form capillary-like structures. As assessed by quantitative PCR, their expression of angiogenic factors such as angiopoietin-2 and apelin are induced after 2 weeks as compared with preimplanted cells, suggesting that hESC-EC may undergo an in vivo maturation. Cardiac cell replacement therapy by using hESC-EC may be a promising future approach to repair of ischemic tissues and induce the formation of blood vessel networks.