Erythropoetin (EPO), a stimulator of erythropoiesis, was previously shown to stimulate angiogenesis and proliferation of endothelial cells. Here, we investigated and compared the influence of EPO on cell number, proliferation, apoptosis, migration and differentiation of endothelial cells in intact mouse embryoid bodies (EB), isolated endothelial cells from EB (EBEC) and adult human endothelial progenitor cells (hEPC). EB were treated with EPO (0.5 U/ ml) immediately after plating (day 5+0) or 3 days later. EPO treatment was continued until days 5+3 or 5+6. Cultured endothelial cells from EB (EBEC) were treated 3 days after passage 3 and primary human EPC from young healthy adults 5 days after plating with EPO for 48 h. Immunohistochemistry was performed with anti-PECAM (CD31), anti-Ki67, anti-CD34, anti-CD133, anti-EphB4 and anti-ephrinB2 antibodies. In all, mouse EB, EBEC and human EPC, EPO-treatment resulted in increased number of endothelial cells, increased proliferation, decreased apoptosis and enhanced migration. In EB, this EPO-effect was most pronounced when treatment was begun early (day 5+0) and was accompanied by an enhanced endothelial tube formation. In EBEC and hEPC, EPO shifted the phenotypic differentiation towards an increased ratio of EphB4 positive cells, i.e. towards a venous phenotype. These results are consistent with an important role of EPO for the number, proliferation, apoptosis, function and phenotypical development of immature endothelial cells which persists from early development through adulthood. They provide additional and further evidence for a strong interrelation between haematopoiesis and vasculogenesis/ angiogenesis (sharing the same pathways) which may be important in many physiological and pathophysiological conditions.