<Originals>Application of periosteal derived osteoblasts onto synthetic biodegradable polymer to induce bone formation in vivo : effect of cell concentration

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We isolated osteoblastic cells derived from bovine periosteum in an explant culture system. The periosteum was cultured in M 199 medium supplemented with 10% fetal calf serum and 5% ascorbic acid. The total number of isolated cell per dish was counted after digestive treatment with 0.25% trypsin and 0.3% collagenase. These cells have positively reacted with alkaline phosphatase staining and demonstrated the ability to mineralize. Western blot analysis showed a maximum level of osteopontin expression at 3 weeks after cell culture. After periosteal derived osteoblasts of 10×10^6/ml (Group I) and 100×10^6/ml (Group II) were seeded onto a 1×1cm polyglycolic polymer sheet, cell polymer constructs were implanted subcutaneously in the athymic mice and harvested after 3,8,and 12 weeks. The results showed the development of bone-like tissue through endochondral ossification, in which minerals were initially localized around the periphery of the construct with remnants of biodegraded polymer. Mineralization was enhanced significantly and bone sialoprotein production was stimulated in Group II over time. These observations indicated that cultured cells from bovine periosteum are osteogenic in nature that have the capacity to differentiate into osteoblasts and to deposit minerals whose level was in accordance with the cell number seeded onto the biodegradable polymer.
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