The culture of human osteoblasts upon bone graft substitutes

In order to assess the ability of six potential bone graft substitutes to support the growth of human osteoblasts, these cells were grown in culture and then plated onto fragments of the six materials and cultured for a further period of 15 days. Tests to confirm the osteoblastic phenotype of the cells included spectrophotometric alkaline phosphatase assay; Western blotting of secreted osteocalcin, osteonectin, bone sialoprotein, and collagen type I; and mineralization within the cultures provided with a supplemented medium. Cells were seeded onto the materials in 24-well plates (Nunc, NaperviUe, IL) at density levels 12 500 cells/cm 2 and 25 000 cells/cm 2. Specimens were examined after the 15-day culture period by scanning electron microscopy. At a seeding density of 12 500 cells/cm 2 results showed that the human osteoblasts had greatest affinities for demineralized rat bone and demineralized Surgibone ®, whilst few osteoblasts were found attached to Pyrost, Surgibone ®, or coral. The collagen matrix of Callopat ® hydrated in the culture media exposing the hydroxyapatite crystals within it, and these became the foci for cell attachment and growth. At a seeding density of 25 000 cells/cm 2 the osteoblasts had attached to and proliferated upon the surfaces of all the materials, forming multilayers, with the exception of Surgibone ®. These experiments demonstrated that all the materials, with the exception of nondemineralized Surgibone R, were biocompatable for human osteoblasts. The final population of osteoblasts on the bone graft substitutes appeared to be influenced by cell plating density, surface topography, and antigenicity of the materials.