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Synthetic colloid and gel hydroxyapatite (HA) nanoparticles (NPs) were spray dried to form microparticles (MPs). These are intended for use as slow release vaccine vectors. The physico-chemical properties of gel and colloid NPs and MPs were compared to those of HA obtained commercially. Their cytotoxicity to human monocytes'-derived macrophages (HMMs) was assessed in vitro using a range of techniques. These included the MTT assay, LDH leakage and a confocal based live-dead cell assay. Cytotoxicity differed significantly between preparations, with the suspended gel preparation being the most toxic (31-500 microg/ml). Other preparations were also toxic but only at higher concentrations (>250 microg/ml). Transmission electron microscopy (TEM) and stereology showed variable cellular uptake and subsequent dissolution of the various forms of HA. We have demonstrated that HA particle toxicity varied considerably and that it was related to their physico-chemical properties. Cell death correlated strongly with particle load. The intracellular dissolution of particles as a function of time in HMM suggests that increased cytoplasmic calcium load is likely to be the cause of cell death. Some HA NPs eluded the phagocytic pathway and a few were even seen to enter the nuclei through nuclear pores.

The incorporation of silicate into hydroxyapatite (HA) has been shown to significantly increase the rate of bone apposition to HA bioceramic implants. However, uncertainty remains about the mechanism by which silicate increases the in vivo bioactivity of HA. In this study, high-resolution transmission electron microscopy was used to observe dissolution from HA, 0.8 wt% Si-HA and 1.5 wt% Si-HA implants after 6 and 12 weeks in vivo. Our observations confirmed that defects, in particular those involving grain boundaries, were the starting point of dissolution in vivo. Dissolution was observed to follow the order 1.5 wt% Si-HA>0.8 wt% Si-HA>pure HA and it was found to be particularly prevalent at grain boundaries and triple-junctions. These observations may help to explain the mechanism by which silicate ions increase the in vivo bioactivity of pure HA, and highlight the enhanced potential of these ceramics for biomedical applications.

The deposition of biological apatite and subsequent formation of bone on hydroxyapatite implants depends on the partial dissolution of the implant surface and the reprecipitation of carbonated apatite from the biological milieu. Previous investigations in vitro have shown that the degree of dissolution and reprecipitation decreases as the coating crystallinity increases. These findings prompted the current study of the effects of coating crystallinity on the mechanism of bone bonding. The process of mineralization of bone associated with a hydroxyapatite coating was compared to the normal process of ossification. Plasma-sprayed hydroxyapatite (PSHA) coated titanium alloy (6% Al-4% V) rods as received and annealed for 0.7 h at 600 degrees C in air to increase the coating crystallinity were implanted in the proximal and distal femora and proximal tibiae of adult mongrel dogs for 3 h, 3 and 10 days. Bony sites containing the implant were prepared for ultramicrotomy and transmission electron microscopy using an anhydrous embedding procedure: fixation in ethylene glycol and embedment in Spurr's resin. The results demonstrated the precipitation of biological apatite crystallites on non-annealed PSHA coatings in vivo within 3 h of implantation. After 3 and 10 days there were differences in the ultrastructure of the mineral phase on the surfaces of non-annealed and annealed surfaces. Observations showed that there was little difference in the mechanism of mineralization of bone associated with HA-coated prostheses and the normal process of ossification.