Abstract
Although hydroxyapatite (HAp) has been doped with dozens of different ions, the quest for an ion imparting a combination of properties conducive to bone healing is still ongoing. Because of its protean potency and the similarity in size and shape to the phosphate tetrahedron, selenite ion presents a natural ionic substitute in HAp. The incorporation of selenite into synthetic HAp using two different methods - co-precipitation and ion-exchange sorption - was studied for its effect on crystal properties and on a triad of biological responses: antibacterial, anticancer and osteoinductive. Co-precipitation yielded HAp with higher selenite contents than sorption and the stoichiometry of HAp richest in selenite was represented as Ca(9.75)(PO(4))(5.75)(SeO(3))(0.25)(OH)(1.75). Crystallinity of HAp decreased in direct proportion with the amount of selenite incorporated. Because of their lower selenite content, HAp powders prepared by ion-exchange exhibited a consistently higher crystallinity compared to the co-precipitated ones. Annealing partially recovered the crystallinity, yet the difference in crystallinity between powders prepared by co-precipitation and by ion-exchange remained, suggesting that the amorphization is mainly due to structural incorporation of selenite, not its effect on the crystal growth kinetics. The addition of selenite changed the morphology of HAp nanoparticles from acicular to rounded and affected the crystal lattice parameters in different ways depending on whether the powders were annealed or not. As for the annealed powders, the incorporation of selenite contracted the lattice in both a and c crystallographic directions. In the agar diffusion assay, the effectiveness of HAp was more dependent on the presence or absence of selenite in it than on its concentration and was highest against E. coli and S. aureus, moderately high against S. enteritidis and ineffective against P. aeruginosa. In liquid inoculation tests, on the other hand, the antibacterial activity of HAp was directly proportional to the amount of selenite contained in it. The viability of K7M2 osteosarcoma cells decreased in direct proportion with the amount of selenite in HAp and was significantly different from the untreated control and from pure HAp at contents equal to or higher than 1.9 wt.%. In contrast, no reduction was observed in the viability of primary fibroblasts treated with HAp incorporating different amounts of selenite ions, suggesting their potentially selective anticancer activity: lethal for the cancer cells and harmless for the healthy cells. Finally, mRNA expression of bone gamma-carboxyglutamate protein (BGLAP3) was higher in differentiated MC3T3-E1 osteoblastic cells treated with selenite-incorporated HAp particles than in cells treated with pure HAp. The osteoinductive effect was due to an overall higher metabolic activity of cells treated with the particles and not due to increased proliferation. In such a way, a triad of antibacterial, osteoinductive and anticancer activities was attributed to selenite-incorporated HAp.