Isolation,Culture and Identification of Rat Bone Marrow-Derived Endothelial Progenitor Cells

Objective To study the methods of isolation,culture and identification of endothelial progenitor cells(EPCs) from rat bone marrow. Methods Mononuclear cells suspension of rat bone marrow was prepared by density gradient centrifugation on Percoll(1.077 g/ml) . The mononuclear cells were induced by VEGF and bFGF,and then the shape of these cells were observed by light microscopy. EPCs were identified by immunofluorescence staining for the expression of PECAM-1/CD31,VE-cadherin/CD144,fluorescein Ulex Europaeus agglutinin-1(FITC-UEA-1) and uptake of DiI complexed acetylated low-density lipoprotein(DiI-ac-LDL) . Results The clone-like morphology and the cobblestone structure were observed by inverted microscopy after 7 days cultivation. Laser Scanning Confocal Microscopy(LSCM) showed that the adherent cells were positive for CD31 and VE-cadherin. Moreover the cells could incorporate Dil-ac-LDL and bind FITC-UEA-1. Conclusion 1.077 g/ml percoll density centrifugation,with VEGF and bFGF induction may obtain EPCs from rat bone marrow,thus demonstrating the feasibility of the methods used.