Isolation,culture and osteogenic induction of rabbit bone marrow mesenchymal stem cells

BACKGROUND:Bone marrow mesenchymal stem cells are important seed cells for tissue engineering because of their multi-directional differentiation potential and possibility to be amplified and cultured in vitro. But there have been no uniformed methods of in vitro culture and oriented differentiation. OBJECTIVE:To investigate the feasiblity of in vitro oriented differentation of rabbit bone marrow mesenchymal stem cells into osteoblasts. METHODS:Rabbit bone marrow mesenchymal stem cells were isolated and purified using density gradient centrifugation (1.073 g/mL Percoll separation solution for centrifugation at 3 000 r/min for 30 minutes, which was different from Ficoll separation solution for centrifugation at 2 000-2 500 r/min for 20-30 minutes as well as whole bone marrow culture method). After in vitro amplification, passage 3 bone marrow mesenchymal stem cells were cultured with common culture medium (control group) and osteoblast induction culture medium (experimental group). RESULTS AND CONCLUSION:A large number of high purity bone marrow mesenchymal stem cells were successfully obtained. After osteogenic induction, the content of osteocalcin in the experimental group was significantly higher than that in the control group (P < 0.05). Alkaline phosphatase and calcium tubercle staining were positive in the experimental group, but they were negative in the control group. These findings suggest that density gradient centrifugation can be used to isolate and culture rabbit bone marrow mesenchymal stem cells, and using this method, bone marrow mesenchymal stem cells can be induce-differentiated into osteoblasts.