The discovery of endothelial progenitor cells has revolutionized our understanding of postnatal blood vessel formation, with endothelial colony-forming cells (ECFCs) emerging as key players in vasculogenesis. Among various ECFC sources, cord blood-derived ECFCs (CB-ECFCs) are of particular interest due to their superior proliferative and clonogenic potential and their ability to promote vascular network formation. Human embryonic stem cell-derived endothelial cells (hESC-ECs) have also shown potential in regenerative medicine, though their vasculogenic efficacy remains unclear compared to CB- and adult blood-derived ECFCs (AB-ECFCs). This study aimed to directly compare the angiogenic and vasculogenic capabilities of CB-ECFCs, AB-ECFCs, and hESC-ECs in vitro and in vivo. Our results demonstrated that CB-ECFCs had a significantly higher proliferation rate than both AB-ECFCs and hESC-ECs (p <
0.01). In tube formation assays, CB-ECFCs exhibited superior ability to form capillary-like structures compared to hESC-ECs (p <
0.0002) and AB-ECFCs (p <
0.01). In vivo, CB-ECFCs significantly improved blood flow recovery in ischemic tissue (p <
0.01), outperforming both AB-ECFCs and hESC-ECs, with no significant recovery observed in the latter two groups. These findings suggest that CB-ECFCs represent a more effective cell source for therapeutic angiogenesis, while further optimization is needed to enhance the efficacy of hESC-ECs for clinical applications. Future research should explore the molecular mechanisms underlying the superior regenerative potential of CB-ECFCs and focus on improving the stability and functionality of stem cell-derived ECs for therapeutic use.