Pore Size of Porous Hydroxyapatite as the Cell-Substratum Controls BMP-Induced Osteogenesis

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Abstract
To elucidate the biochemical mechanism of osteogenesis, the effect of matrix geometry upon the osteogenesis induced by bone morphogenetic protein (BMP) was studied. A series of five porous hydroxyapatites with different pore sizes, 106–212, 212–300, 300–400, 400–500, and 500–600 μm, was prepared. A block (approximately 5×5×1 mm, 40.0 mg) of each hydroxyapatite ceramics was combined with 4 μg of recombinant human BMP-2 and implanted subcutaneously into the back skin of rat. Osteoinductive ability of each implant was estimated by quantifying osteocalcin content and alkaline phosphatase activity in the implant up to 4 wk after implantation. In the ceramics of 106–212 μm, the highest alkaline phosphatase activity was found 2 wk after implantation, and the highest osteocalcin content 4 wk after implantation, consistent with the results observed with particulate porous hydroxyapatite [Kuboki, Y. et al. (1995) Connect Tissue Res. 32: 219–226]. Comparison of the alkaline phosphatase activities at 2 wk and the osteocalcin contents at 4 wk after implantation revealed that the highest amount of bone was produced in the ceramics implants with pore size of 300–400 μm. In the ceramics with smaller or larger pore sizes, the amount of bone formation decreased as the pore size deviated from 300–400 μm. The results indicated that the optimal pore size for attachment, differentiation and growth of osteoblasts and vascularization is approximately 300–400 μm. This study using chemically identical but geometrically different cell substrata is the first demonstration that a matrix with a certain geometrical size is most favorable for cell differentiation.

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