Endothelial progenitor cells (EPCs) are present in the mononuclear cells (MNCs) of umbilical cord blood and peripheral blood. To establish the efficiency of angiogenic cell and gene therapies, we transfected the human vascular endothelial growth factor (hVEGF) gene into cord blood MNCs to enhance endothelialization. MNCs from cord blood and peripheral blood were isolated and transfected with pCR3 expressing hVEGF165 or GFP by the Hemagglutinating Virus of Japan (HVJ)-envelope and the cells were cultured in endothelium basal medium-2. The number of attached cells from cord blood was higher than that from peripheral blood. Attached cells expressed Flk-1, VE-cadherin, PECAM-1, CD34, and Tie-2. The increase in the number of attached cells was transient with the transfection of vascular endothelial growth factor (VEGF) gene early in the experimental period. Flt-1 mRNA was not expressed early in the culture period, but was expressed at 2 weeks after separation. VEGF gene transfer into MNCs at 12 days after separation, i.e., when Flt-1 mRNA was expressed continuously, increased the number of attached cells. We evaluated the effects of the transplantation of cord blood MNCs expressing the hVEGF gene on regional blood flow in an ischemic area in a rat model of chronic hindlimb ischemia. Blood flow was significantly improved in nude rats that received transplanted control MNCs. Transplantation of cord blood MNCs transfected with the hVEGF gene yielded greater improvements in blood flow. These results indicate that the hVEGF gene enhances endothelialization of EPCs, and that the transplantation of cord blood MNCs transfected with the VEGF gene may be feasible for the treatment of ischemic diseases as a type of angiogenic cell and gene therapy.