OBJECTIVETo investigate the optimal dosage and timing for 5-bromodeoxyuridine (BrdU) labeling of endothelial progenitor cells (EPCs) from rat circulating blood.

METHODSThe animal model for rat tooth movement was established. EPCs were obtained by density gradient centrifugation. The expressions of specific antigens on cell surface were analysed by immunocytochemistry and fluorescenceochemistry. EPCs were incubated with BrdU at different concentrations (5, 10, 15 micromol/L) for different incubating time (12, 24, 48, 72, 96 h) to identify the optimal BrdU concentration and incubating time for cell labeling. Immunohistochemistry was performed to calculate the labeling index (LI).

RESULTSThe culture cell positively expressed CD34, CD133 and could be shown to endocytose DiI-ac-LDL, FITC-UEA-1. Incubation of the EPCs with BrdU at 10 micromol/L and for an optimal length of 72 h appeared to achieve the highest LI (66.8+/-2.9)%, which was significantly higher than group of 5 micromol/L (P<0.05), while there was no significant difference between the group of 15 micromol/L and 10 micromol/L (P>0.05).

CONCLUSIONEPCs can be isolated from tooth movement rat circulating blood and cultured. Incubation of the EPCs with BrdU at 10 micromol/L and for an optimal length of 72 h appeared to achieve the optimal LI. This provides a foundation for us to investigate the mechanism of chemiotaxis and differentiation for EPCs.