ULTRASTRUCTURAL CHARACTERIZATION OF THE INTERFACE BETWEEN CYTODEX-3 MICROCARRIER AND BONE FORMED IN-VITRO USING HIGH-RESOLUTION ELECTRON-MICROSCOPY, HISTOCHEMISTRY AND IMMUNOHISTOCHEMISTRY

Enzymatic isolated rat calvaria cells forming a mineralized matrix in vitro were cultured in the presence of Cytodex-3 microcarriers. The in terface between the bone tissue and the microcarrier beads was studied by transmission electron microscopy (TEM), high resolution electron m icroscopy, histochemical and immunohisto-chemical TEM. Ultrastructural observation by routine TEM demonstrated the presence of an electron-d ense collagen-free layer at the Cytodex/tissue interface. High resolut ion microscopy showed that this layer was composed of apatite crystals . The use of cetylpyridinium chloride in glutaraldehyde fixative shows as the presence of polyanions in this interface. This layer was obser ved before bone formation occurred and then an osteoid matrix was synt hesized by active osteoblasts and subsequently mineralized in contact with the first formed electron-dense layer following a classical patte rn of intramembranous osteogenesis. Immunogold labelling was used to e xamine the distribution of bone sialoprotein (BSP). The Cytodex/bone i nterface was strongly immunoreactive for BSP. From the results of this study, it can be concluded that the present in vitro system is a suit able model to study the step-by-step interactions occurring at the sur face of artificial substrates.