Isolation, culture and identification of rat bone marrow derived endothelial progenitor cells
LU Yao-liang1, KANG Tao1, LI Xiao-qiang2, HAN Song1
(1.Department of Vascular Surgery, the Affiliated Taicang Hospital of Soochow University, Suzhou Jiangsu 215400; 2. Department of Vascular Surgery, the Second Affiliated Hospital of Soochow University, Suzhou Jiangsu 215004, China)
Abstract：Objective: To explore the culture, differentiation and identification method and biological characteristics of the rat bone marrow derived endothelial progenitor cells (EPCs)．Methods: We separated the rat bone marrow mononuclear cells using density gradient centrifugation combined with differential sidewall separation, cultured endothelial cell with EGM-2, obtained adherent cells within 48 h (early EPCs) and adherent cells after 48 h(late EPCs), respectively. The cell morphological images were observed under inverted microscope; EPCs surface antigen markers, CD34, CD133 and VEGFR-2 were identified by the flow cytometry, the phagocytosis of EPCs were detected by laser scanning confocal microscope, the internal ultrastructure of EPCs were observed by transmission electron microscope. Results: Mononuclear cells were small round after the first incubation, the late EPCs was given priority to spindleshaped, and there was obvious stem cell colony after culture in 7 days; till the 6th generations of EPCs, the cellular morphology was gradually close to the endothelial cells, appeared the tube cavity structure. CD34, CD133, VEGFR2 expressed positive, could absorb Dil-acLDL combined with FITC-UEA-1. There was characteristic organelles Weible-Palade (W-P) small body in the EPCs under inverted microscope. Conclusion: The EPC was successfully isolated from bone marrow, and purified, cultured by density gradient centrifugation combined with differential adherent method; it had strong proliferation ability, abundant cell count and stable biological characteristics. ［Key words］endothelial progenitor cells; bone marrow; stem cells
2013, Vol. 23 
