Human CD133 (AC133)⁺CD34⁺ stem and progenitor cells derived from fetal liver and from bone marrow and blood incorporate a functional population of circulating endothelial precursor cells. Vascular endothelial growth factor receptor 3 (VEGFR-3) regulates cardiovascular development and physiological and pathological lymphangiogenesis and angiogenesis. However, the origin of VEGFR-3⁺ endothelial cells (ECs) and the mechanisms by which these cells contribute to postnatal physiological processes are not known, and the possible existence of VEGFR-3⁺ lymphatic or vascular EC progenitors has not been studied. Using monoclonal antibodies to the extracellular domain of VEGFR-3, we show that 11% ± 1% of CD34⁺ cells isolated from human fetal liver, 1.9% ± 0.8% CD34⁺ cells from human cord blood, and 0.2% ± 0.1% of CD34⁺ cells from healthy adult blood donors are positive for VEGFR-3. CD34⁺VEGFR-3⁺ cells from fetal liver coexpress the stem/precursor cell marker CD133 (AC133). Because mature ECs do not express CD133, coexpression of VEGFR-3 and CD133 on CD34⁺cells identifies a unique population of stem and progenitor cells. Incubation of isolated CD34⁺VEGFR-3⁺ cells in EC growth medium resulted in a strong proliferation (40-fold in 2 weeks) of nonadherent VEGFR-3⁺ cells. Plating of these cells resulted in the formation of adherent VEGFR-3⁺Ac-LDL⁺ (Ac-LDL = acetylated low-density lipoprotein) EC monolayers expressing various vascular and lymphatic endothelial-specific surface markers, including CD34, VE-cadherin, CD51/61, CD105, LYVE-1, and podoplanin. These data demonstrate that human CD34⁺CD133⁺ cells expressing VEGFR-3 constitute a phenotypically and functionally distinct population of endothelial stem and precursor cells that may play a role in postnatal lymphangiogenesis and/or angiogenesis.