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The effects of hydroxyapatite coated micromachined substrata on osteogenesis Perizzolo, David

Abstract

Osteogenesis was studied using rat bone cells cultured on hydroxyapatite (HA) or titanium (Ti) coated smooth and grooved substrata. Osteoblast cultures were maintained from 24hrs to 6 weeks in culture medium and supplemented with L-ascorbic acid-2-phosphate and β- glycerophosphate to promote mineralization. The HA coatings were characterized using X-ray diffraction, surface roughness and scanning electron microscopy (SEM). The ceramic hydroxyapatite coating, was dense and uniform, containing HA crystals and was ≈ 1µm thick. Time-lapse cinemicrography of osteoblasts locomotion on HA surfaces revealed osteoblasts moved with the direction of the grooves, that is they exhibited contact guidance. Scanning electron microscopic observations revealed osteoblasts were elongated and orientated on both Ti and HA grooved surfaces. Collagen fibers, as assessed by picro-sirius staining and polarized light microscopy, were aligned on both HA and Ti grooved surfaces. Although not quantified, it appeared that nodule formation was greatest under culture conditions that produced aligned collagen. Osteogenesis was measured by counts of tetracycline labelled bone-like nodules and alkaline phosphatase activity. HA coated surfaces produced significantly more mineralized nodules than Ti surfaces. Surfaces with grooves and narrow gaps produced the highest number of nodules. All grooved substrata produced significantly more nodules than smooth surfaces. These results are consistent with the concept that substrata that form a microenvironment by restricting diffusion increase bone-like tissue production. A novel finding in this thesis was that there was a statistically significant interaction between topography and chemistry in the formation of mineralized nodules. An excellent correlation (r= 0.958) between alkaline phosphatase at 2 weeks and nodule counts at 6 weeks was observed, suggesting that Alk-P is a good leading indicator of osteogenesis on microfabricated surfaces. Therefore Alk-P might be used to screen surfaces for their nodule production, thus saving time and expense. The results of this study indicated that topography and chemistry can affect osteogenesis on biomaterials, and that interactions between chemistry and topography can occur.

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