Abstract:

BACKGROUND: Endothelial progenitor cells (EPCs) are involved in angiogenesis, but its isolation, culture and identification methods are not uniform currently.
OBJECTIVE: To explore the methods of separation, culture and identification of rat bone marrow-derived EPCs.
METHODS: EPCs were cultured using density gradient centrifugation and differential velocity adherent methods, growth and morphology changes of cells were observed under inverted microscope. The expression of cell surface antigen CD34 and CD133 were detected by using Dil labeled acetylated low-density lipoprotein (Dil-ac-LDL) and FITC-labeled Ulex europaeus agglutinin 1 (FITC-UEA-1) double dyeing and flow cytometry.  RESULTS AND CONCLUSION: For the first four days, cell proliferation was not obvious, at the 5-10 days, cell proliferation was obvious, and colony-forming unit (CFU) and lineage structure could be seen. At the 7th day, EPCs could swallow the function of DiL-ac-LDL and FITC-UEA-1. At the 10th day, in vitro flow cytometry detection showed that the CD133+ cells were accounted for 19.2%, CD34+ cells were accounted for 28.7%, CD34+/CD133+cells were accounted for 19.1%. Using density gradient centrifugation and differential velocity adherent methods can separate and culture bone marrow derived EPCs.