Formation of hydroxyapatite on titanium implants in vivo precedes bone-formation during healing

The bone material interface has been an area of intense study over many decades, where studies of the healing process ranging from simple mineral deposition in vitro to actual healing in vivo have given important clues to the importance of calcium minerals in the bone/implant interface. Here, the authors use a combination of in vitro cell culture methods and in vivo implantation to study how the role of the spontaneously formed hydroxyapatite layer on Ti-implants for the in vivo-healing into the bone tissue of rat tibia. Initial experiments were made in reduced systems by incubation of TiO₂ in cell culture medium and analysis by time of flight secondary ion mass spectrometry (ToF-SIMS) and energy-dispersive x-ray spectroscopy followed by subsequent exposure of human embryological stem cells analyzed by von Kossa staining and environmental scanning electron microsopy. In vivo studies of the bone-material interface was analyzed by ToF-SIMS depth profiling using both C₆₀⁺ ions as well as a gas cluster ion source beam, Ar₁₅₀₀⁺ as sputter source. The low ion yield of the Ar₁₅₀₀⁺ for inorganics allowed the inorganic/organic interface of the implant to be studied avoiding the erosion of the inorganic materials caused by the conventional C₆₀⁺ beam.