An in vitro system for studying osteointegration of dental implants utilizing cells grown on dense hydroxyapatite disks.

Abstract

A proposed in vitro system is described where chick osteoblasts are cultured on the flat surfaces of dense, nonporous HA disks to facilitate the study of bone formation at the cell-HA interface. During early bone formation cell-coated HA disks were retrieved, fixed with buffered 2% glutaraldehyde, and embedded in epon/araldite. The underlying HA disks were demineralized in diluted acid, and the intact cell-HA interfaces were re-embedded and thin sectioned for routine transmission electron microscopy. Morphologic studies indicated that osteoblasts proliferated and formed nodules of cells on the surfaces of HA disks. With increasing time in culture, they deposited orthogonally packed collagen fibrils between the cell layers that were enveloped by electron-dense mineralized globules. Eventually, small spicules of mineralized HA formed along collagen fibrils. An electron-dense layer about 50 nm thick was observed on the surface of the HA disks. Biochemical studies indicated that cell proliferation, as judged by 3H-thymidine uptake, increased rapidly during the first 3 days, reached a maximum around 6 days, and then declined by 12 days in culture. AP activity and collagen synthesis, as determined by 3H-hydroxyproline formation, increased as cellular proliferation declined. Mineralization, as judged by 45Ca uptake and spicule formation, occurred, as expected, following the increase in AP activity and deposition of densely packed collagen fibrils. Thus, all morphological and biochemical parameters studied indicate that the proposed in vitro system is reproducible and can facilitate the study of the osteointegration of HA-coated implants.