Osteoclastic resorption of bone-like apatite formed on a plastic disk as an in vitro assay system

We have investigated the applicability of a simple and inexpensive osteoclastic assay system using bone-like apatite-coated polyethyleneterephthalate (PET) disks. A 1 microm thick apatite layer, uniform and homogeneous bone-mineral-like with no organic components, was made on PET disks using a biomimetic process. As substrates for an osteoclastic assay, these coated disks were compared with dentine as well as with bone-like or heat-treated apatite of various thicknesses on apatite- and wollastonite-containing glass ceramic (A-W GC) disks. The unfractionated bone cells, including osteoclasts, of a neonatal rabbit were seeded onto these substrates. By scanning electron microscopic examination, the resorption lacunae of the thick bone-like apatite clearly showed track-like shapes at various depths, similar to those of dentine although the border between the A-W GC and the apatite was unclear. In contrast, those of heat-treated apatite showed small and shallow shapes with irregular margins, quite different from those of dentine. By reducing the thickness of bone-like apatite to 1 microm as well as using PET as its substrate, the margins of the resorption lacunae became quite clear, and with the use of phase-contrast microscopy during culture, osteoclasts and resorption pits could be precisely observed. The resorbed area, easily measured with the aid of bright-field microscopy and an image analyzer, was found to have increased in a time-dependent manner and at the end of 4 days of culture was not statistically different from that of dentine.