Effect of a Hybrid Micro/Nano-integrated Titanium Surface on Behavior of Rat Osteoblasts

The objective of this study was to investigate the effect of a new combined micro/nanoscale implant surface feature on osteoblasts' behaviors including cell morphology, adhesion, proliferation, differentiation, and mineralization in vitro. A new micro/nano-hybrid topography surface was fabricated on commercial pure titanium (Cp Ti) by a two-step sandblasted acid-etching and subsequent alkali-and heat-treatment (SA-AH). The conventional sandblasted/acid-etching (SA) treatment and alkali and heat (AH) treatment were also carried out on the Cp Ti as controls. Surface microstructures of the Ti disc samples were assessed by scanning electron microscopy (SEM). The neonatal rat calvaria-derived osteoblasts were seeded on these discs and the initial cell morphology was evaluated by SEM and immunofluorescence. Initial adhesion of the cells was then assayed by DAPI staining at 1, 2, and 4 h after seeding. The Cell Counting Kit-8 (CCact K8) assay, gene expression of osteoblastic markers (ALP, Col 1, OCN, BSP, OSX, Cbf alpha 1) and Alizarin Red S staining assays were monitored respectively for cell proliferations, differentiation and mineralization. The results show significant differences in osteoblast's behaviors on the four kinds of Ti surfaces. Compared with Cp Ti surface, the SA and AH treatment can significantly promote cell adhesion, differentiation and mineralization of osteoblasts. In particular, the combined SA and AH treatments exhibit synergistic effects in comparison with the treatment of SA and AH individually, and are more favorable for stimulating a series of osteogenous responses from cell adhesion to mineralization of osteoblasts. In summary, this study provides some new evidence that the integrated micro/nanostructure on the Cp Ti surface may promote bone osseointegration between the Ti implantbone interfaces in vitro.