Induction of human umbilical cord blood-derived CD34~+ cells into endothelial cells in vitro

Objective: To study the method of CD34+ cell isolated from human umbilical cord blood and induced to differentiate into endothelial cells in vitro. Methods: Mononuclear cells were isolated from fresh cord blood by 6 % HES and density gradient centrifugation. CD34+cells selected from mononuclear cells by magnetic activated cell sorting system (MACS) were cultured in M199 medium supplemented with VEGF, bFGF, IGF-1, then observed the growth and shape of the cells. The cells specific surface mark CD34 and endothelial specific surface marker were assessed by fluorescence activated cell sorter ( FACS) analysis. And the cells were also identified by the test of up take of acety-lated low density lipoprotein (Ac-LDL). Results: The purification of obtained CD34+cells were more than 95%. Attached spindle-like cells and a number of cell clusters appeared after 3 days culture and cord-like structure by 14 days. When the cells grow into large number, it displayed cobble -stone morphology. These cells express endothelial specific markers, including VE-cadherin, vWF and CD31.The cells’ function were identified by up take of Ac-LDL. Conclusion: The two methods and the magnetic bead selection are an effective ways to isolate CD34+ cells from cord blood and there are CD34+ cells in cord blood, which can differentiate into endothelial cells.