Purpose: To investigate the methods of isolation, culture and identification of BMMSCs derived from rabbit mandible.
Methods: BMMSCs were collected from rabbit mandible and isolated by density gradient centrifugation. Cells were adherently cultured in vitro, and P2 or P3 BMMSCs populations were collected and examined. Cell growth was observed by inverted microscopy; the propagation of BMMSCs were tested by MTT and a growth curve was drawn after statistical analysis; colony-forming unit-fibroblast(CFU-F) was detected by examination of colony formation; the potential of multi-directional differentiation into osteoblasts, adipocytes and skeletal muscle cells was estimated by pertinent methods; the surface marks of BMMSCs were detected by flow cytometry, the data was analysed using SPSS16.0 software package.
Results: The majority of adherent cells were long fusiform, and few were small triangle; the growth curve of BMMSCs showed that every passage experienced incubation period, log phase and platform period; the rate of colony formation was 37%. Growth of BMMSCs represented the appearance of CFU-F; BMMSCs after inducted differentiation showed osteogenic and adipogenic potential. The staining of mineralized nodules was positive by alizarin red S and the positive staining of oil red O appeared in lipid drops around cell nucleus. The staining of skeletal muscle cells was positive by desmin immunofluorescence; the cell surface marks assessed with flow cytometry indicated that these BMMSCs expressed CD90 and CD146 in high percentage (about 98.7% and 98.1%, respectively).
Conclusions: The highly uniformed BMMSCs derived from rabbit mandible can be collected. These BMMSCs have the ability of self-replication and propagation, as well as potential of multi-directional differentiation in vitro.