Osteogenic responses and bond strength of zirconia with hydroxyapatite coating by aerosol deposition

Abstract

Objectives: The purposes of this study were to evaluate the in vitro osteogenic potential and bonding strength of hydroxyapatite (HA)-coated zirconia by aerosol deposition method for the improved osseointegration.

Materials and Methods: Pure titanium discs (25 mm diameter, 1 mm thickness) were prepared by machining (Ti-machined

Ti-m) or treated by anodizing (Ti-anodizing

Ti-a). Green zirconia discs (15 mm diameter, 1 mm thickness) were prepared by cold isostatic pressing of the powder mixtures at 200 MPa followed by sintering for 5 h at 1650°C in air. After polishing, the (Y, Nb)-TZP and (Y, Ta)-TZP surfaces were roughened by sandblasting with 50 μm Al2O3 at 2 bar and 1 bar pressure, respectively. The HA by aerosol deposition to a thickness of approximately 10 μm was annealed at 700°C for 2 h under an air atmosphere. Specimens were evaluated to measure the average surface roughness (Ra) and topography by confocal laser scanning microscopy, contact angles by automated contact-angle-measuring device, and the crystal structure by X-ray diffraction. Cell attachment was evaluated by confocal laser scanning microscope 24 h after seeding MC3T3-E1 pre-osteoblast cells on the discs. Cellular proliferation on the zirconia and titanium discs was analyzed by Picogreen assay. MC3T3-E1 pre-osteoblast cells were harvested after 2, 6, 10 and 15 days of differentiation and real-time PCR was performed to measure the expression of the bone marker genes such as type I collagen, alkaline phosphatase, and osteocalcin. Bonding strength of HA layer was evaluated by a universal testing machine and microstructure of HA coating layer was examined by scanning electron microscopy. Student t-test was used for the contact angle and Ra value. Statistical analyses for cell proliferation assay, cytotoxicity test and quantitative real time PCR were performed using one-way analysis of variance (ANOVA) with Tukeys post hoc test. Post hoc analysis was used to detect pairs of groups with statistically significance differences. Significance was considered at p < 0.05 (SPSS software, Chicago, IL, USA).

Results: Surface analysis by scanning electron microscopy and X-ray diffraction proved as-deposited thin HA film on the zirconia showing a shallow regular crater like surface. Deposition of dense and uniform HA-films was measured by SEM and contact angle test proved improved wettability of HA-coated surface. Confocal laser scanning microscopy indicated that MC3T3-E1 pre-osteoblasts attachment did not differ notably between the titanium and zirconia surfaces

however, cells on the HA-coated zirconia exhibited a lower proliferation than the uncoated zirconia late in the culture. Nevertheless, ALP, alizarin red S staining and bone marker gene expression analysis indicated good osteogenic responses on HA-coated zirconia. Mean bonding strength of HA coating layer to zirconia was 26.02 MPa ± 3.92.

Conclusions: HA-coating on zirconia by aerosol-deposition improves the quality of surface modification and is favorable to the osteogenesis. Surface configuration, chemical composition of coated surface and bond strength between zirconia substrate and hydroxyapatite demonstrated that aerosol deposition method can be used as a viable option for the HA coating. However, further in vivo study is needed to confirm more about the efficacy of HA-coated zirconia implants with respect to osseointegration.