Synthesis of ion-substitued nanosized apatites and assessment of dispersibility and cell culture behaviour

The use of hydroxyapatite (HA) based materials is widely spread in the repair of bone tissue and their great acceptance arises from the close similarity of HA to the mineral phase of bone. In the last years HA has been explored in a new area of application where it, in the form of nanoparticles (NPs), has been used as a carrier for the delivery of drugs and genes inside targeted cells. In spite of the promising results, which have been reported up to day, there are fundamental questions that remain unanswered. The goal of this project is to address one specific aspect: if the addition of doping ions in the formulation of hydroxyapatite NPs affects cell behaviour. For this purpose the first objective was to prepare HA NPs with various types and degree of ion substitution: carbonate (CO3)-, magnesium (Mg)- and CO3-Mg-co-substituted HA NPs. They were prepared via a wet precipitation route involving neutralization of calcium hydroxide with phosphoric acid and were characterised by X-ray diffraction (XRD), Fourier-transform infrared spectroscopy (FTIR), inductively coupled plasma (ICP) elemental analysis, transmission electron microscopy (TEM) and nitrogen (N2) absorption. The next objective was to obtain stable aqueous suspensions of the NPs. This was achieved by the simultaneous use of a natural dispersant (sodium citrate) and with the help of a probe sonicator. The dispersibility was evaluated visually and with the help of a particle size analyser. The ultimate objective was to assess the cytotoxicity of the well dispersed and non-dispersed ion-substituted HA NPs in a cell culture. Two types of cells were used for this purpose: osteoblast-like cancer cells (MG63) and rat mesenchymal stem cells (rMSC). The results showed that although the incorporation of the doping ions (e.g. carbonate) changed substantially the morphology and specific surface area of some NPs, such alterations did not seem to have any effect on cell behaviour. Yet, the presence of Mg at percentages of only 1,2 wt% caused no alteration on the nanoparticle morphology, but had a great impact in cell viability. Mg-substituted NPs were cytotoxic for MG63 but did not compromise the viability of rMSC. The results clearly showed that indeed the composition of the NPs is crucial and, though further studies need to be done, one can envisage that fine tuning of the NP composition can be a strategy to improve drug and gene delivery.