[In vitro analysis and in vivo tracing of BrdU-labeled rat bone marrow mesenchymal stem cells].

ABSTRACT

OBJECTIVES: To investigate the optimal condition of bromodeoxyuridine (BrdU) labeling for bone marrow mesenchymal stem cells (BMSCs) in vitro and the feasibility of in vivo tracing of BrdU-labeling BMSCs. METHODS: BMSCs were isolated from Wistar rats and were in vitro routinely cultured. The third passage BMSCs was used for identification of special surface antigens by immunohistochemical methods. The purified BMSCs were incubated with BrdU at different concentrations for different incubating time to investigate optimal BrdU concentration and incubating time for cell labeling. The cell labeling index of BrdU was calculated with immunohistochemical analysis. BMSCs labeled BrdU were injected into damaged gastric mucosa of rats by micro injector. The colonization of BMSCs labeled BrdU in gastric mucosa was viewed. RESULTS: After purification and proliferation, the primary cultured BMSCs were uniformly long spindle-shapped form and formed cell colony, which showed the characteristics of stem cell. Immunocytochemistry showed BMSCs were positive for CD44 and CD90, while negative for CD14, CD45. The labeling rate of BrdU increased with the labeling time lasting and reached its height at 48 h. After incubating 48 and 72 hours, the labeling rate of BrdU with a concentration of 10 micromol/L was higher than that of BrdU with a concentration of 5 micromol/L (P < 0.05) and similar with that of BrdU with a concentration of 15 micromol/L (P > 0.05). In addition, the BrdU labeling could be detected after five consecutive passages and the labeling time could keep 21 d. The pathological observation demonstrated that BrdU-labeled BMSCs could colonize the damaged gastric mucosa with normal morphologic characteristics during observation period. CONCLUSION: BrdU labeling might be a feasible method for dynamic observation of the migration, growth and differentiation of migrating BMSCs in colonizing sites.