OBJECTIVETo investigate the biological properties of CD34+/CD133 +/VEGFR-3 + lymphatic endothelial progenitor cells in peripheral blood and explore the effects of the VEGF-C/VEGFR-3 signaling pathway on differentiation of lymphatic endothelial progenitor cells to lymphatic endothelial cells.

METHODSMononuclear cells were isolated from peripheral blood by density centrifugation with Percoll solution, and VEGFR-3+ cells were sorted from them with flow cytometry. Differentiation of VEGFR-3+ cells was induced with VEGF-C. The morphology and ultrastructures of the cells were observed with scanning and transmission electron microscopes. Expression of surface markers were examined with a confocal laser scanning microscope.

RESULTSVEGFR-3+ cells expressed CD34 and CD133 antigen. The percentage of CD34+/ VEGFR-3+ and VEGFR-3+/CD133+ cells were 0.13% and 0.08% of peripheral blood MNC respectively. The size of CD34+/CD133+/VEGFR-3+ cells was about 15 microm in the diameter. After induction with VEGFC, they were increased. The cells were shuttle-like in shape and extended the lamellipodia and many filopodia. After 1 week induction with VEGF-C, they expressed coagulation factor VII related antigen, and at 2 week induction, they showed caveolae on the surface and Weibel-Palade body inside the cells. The specific lymphatic endothelial marker LYVE-1 was expressed on the cells, and no longer expressed CD133.

CONCLUSIONSCD34+/CD133+/VEGFR-3+ lymphatic endothelial progenitor cells from peripheral blood may differentiate into lymphatic endothelial cells. The VEGF-C/VEGFR-3 signaling pathway has important effects on the differentiation of the lymphatic endothelial progenitor cells.