Methods:We attempted to isolate osteoblasts using alveolar bones of 52-, 53- and 66-year-old donors. Those bones sequentially digested and primary human alveolar bone osteoblasts (HAOBs) were isolated. To investigate whether HAOBs have osteogenic activity in vitro, they are treated with rhBmp-2 and investigated by alkaline phosphatase activity and alizarin red staining respectively.To examine bone forming capacity of HAOBs in vivo, HAOBs at 6 PDs were subcutaneously transplanted into the dorsal skin in severe combined immunodeficiency (SCID) mice.
Results:HAOBs from all of the individuals were successfully expanded and grow more than 60 population doublings (PDs). HAOBs exhibited high alkaline phosphatase activity and mineralized nodule formation and expressed osteoblast marker genes such as runx2, osterix, osteocalcin and bone sialoprotein upon treatment with rhBMP-2. Histological examination revealed that the transplanted HAOBs formed bone tissues 4 weeks after the transplantation.
Conclusions:Our results show that the culture technique used here allows us to successfully grow human osteoblasts of aged donors. They also suggest that HAOBs are a useful cell source for future application to cell based bone regeneration therapy for local bone diseases such as periodontal diseas.