endothelial progenitor cells.pdf

Cardiovascular Research 58 (2003) 478–486 / locate /cardiores E ndothelial progenitor cell culture and differentiation in vitro: a methodological comparison using human umbilical cord blood a a ,1 b a Juliane Eggermann , Stefanie Kliche , Gergely Jarmy , Karin Hoffmann , Ulrike Mayr- a b a , b *Beyrle , Klaus-Michael Debatin , Johannes Waltenberger , Christian Beltinger a Department of Internal Medicine II (Cardiology), Ulm University Medical Centre, Robert-Koch-Str. 8, D-89081 Ulm, Germany b University Children’s Hospital, Ulm University Medical Centre, 89081 Ulm, Germany Received 6 September 2002; accepted 22 January 2003 Abstract Objective: Endothelial progenitor cells (EPC) can contribute to vascular repair and targeted tumour therapy. Little is known about generating EPC from human umbilical cord blood. We therefore compared methods for purification of EPC from human umbilical cord blood. Methods: Mononuclear cells were isolated from human umbilical cord blood by density gradient centrifugation and used either unselected or after CD34 preselection. Unselected mononuclear cells were cultured for 9 days. Culture-dish-adherent (CDAC) and non-adherent (CDNAC) CD341 cells were cultured separately for 4 weeks. Surface markers were assessed by immunofluorescence staining and FACS analysis. Results: In unselected mononuclear cells,VEGF-R2 and VE-cadherin expression increased up to day 6. They stained positive with UEA-1 and took up acetylated LDL. Expression of CD45 and CD14 decreased over time, but remained strong. CD133 and CD34 were not expressed. CD341-CDNAC acquired an endothelial phenotype over time with an increase of VEGFR-2 and von Willebrand factor (vWF). CD45 and CD14 decreased, while CD34 and the progenitor-cell marker CD133 remained strongly 1 expressed. CD34 -CDAC showed a strong increase in VEGFR-2, CD133, CD34 and vWF, while CD14 decreased, and CD45 did not change. Conclusion: Putative EPC can be obtained from human umbilical cord blood. When selected f