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The effect of novel fluorapatite surfaces on osteoblast-like cell adhesion, growth, and mineralization.

ABSTRACT: There is increasing demand for biomedical implants to correct skeletal defects caused by trauma, disease, or genetic disorder. In this study, the MG-63 cells were grown on metals coated with ordered and disordered fluorapatite (FA) crystal surfaces to study the biocompatibility, initial cellular response, and the underlying mechanisms during this process. The long-term growth and mineralization of the cells were also investigated. After 3 days, the cell numbers on etched metal surface are significantly higher than those on the ordered and disordered FA surfaces, but the initial adherence of a greater number of cells did not lead to earlier mineral formation at the cell-implant interface. Of the 84 cell adhesion and matrix-focused pathway genes, an up- or down-regulation of a total of 15 genes such as integrin molecules, integrin alpha M and integrin alpha 7 and 8 was noted, suggesting a modulating effect on these adhesion molecules by the ordered FA surface compared with the disordered. Osteocalcin expression and the mineral nodule formation are most evident on the FA surfaces after osteogenic induction (OI) for 7 weeks. The binding of the ordered FA surfaces to the metal, with and without OI, was significantly higher than that of the disordered FA surfaces with OI. Most significantly, even without the OI supplement, the MG-63 cells grown on FA crystal surfaces start to differentiate and mineralize, suggesting that the FA crystal could be a simple and bioactive implant coating material.

SUBMITTER: Liu J

PROVIDER: S-EPMC2928044 | biostudies-literature | 2010 Sep

REPOSITORIES: biostudies-literature

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The effect of novel fluorapatite surfaces on osteoblast-like cell adhesion, growth, and mineralization.

Liu Jun J Jin Taocong T Chang Syweren S Czajka-Jakubowska Agata A Zhang Zhaocheng Z Nör Jacques E JE Clarkson Brian H BH

Tissue engineering. Part A 20100901 9

There is increasing demand for biomedical implants to correct skeletal defects caused by trauma, disease, or genetic disorder. In this study, the MG-63 cells were grown on metals coated with ordered and disordered fluorapatite (FA) crystal surfaces to study the biocompatibility, initial cellular response, and the underlying mechanisms during this process. The long-term growth and mineralization of the cells were also investigated. After 3 days, the cell numbers on etched metal surface are signif ...[more]

PMID: 20412028

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Project description:Silicon (Si) is suggested to be an important/essential nutrient for bone and connective tissue health. Silicon-substituted hydroxyapatite (Si-HA) has silicate ions incorporated into its lattice structure and was developed to improve attachment to bone and increase new bone formation. Here we investigated the direct adsorption of silicate species onto an HA coated surface as a cost effective method of incorporating silicon on to HA surfaces for improved implant osseointegration, and determined changes in surface characteristics and osteoblast cell adhesion. Plasma-sprayed HA-coated stainless steel discs were incubated in silica dispersions of different concentrations (0-42 mM Si), at neutral pH for 12 h. Adsorbed Si was confirmed by XPS analysis and quantified by ICP-OES analysis following release from the HA surface. Changes in surface characteristics were determined by AFM and measurement of surface wettability. Osteoblast cell adhesion was determined by vinculin plaque staining. Maximum Si adsorption to the HA coated disc occurred after incubation in the 6 mM silica dispersion and decreased progressively with higher silica concentrations, while no adsorption was observed with dispersions below 6 mM Si. Comparison of the Si dispersions that produced the highest and lowest Si adsorption to the HA surface, by TEM-based analysis, revealed an abundance of small amorphous nanosilica species (NSP) of ~1.5 nm in diameter in the 6 mM Si dispersion, with much fewer and larger NSP in the 42 mM Si dispersions. 29Si-NMR confirmed that the NSPs in the 6 mM silica dispersion were polymeric and similar in composition to the larger NSPs in the 42 mM Si dispersion, suggesting that the latter were aggregates of the former. Amorphous NSP adsorbed from the 6 mM dispersion on to a HA-coated disc surface increased the surface's water contact angle by 53°, whereas that adsorbed from the 42 mM dispersion decreased the contact angle by 18°, indicating increased and decreased hydrophobicity, respectively. AFM showed an increase in surface roughness of the 6 mM Si treated surface, which correlated well with an increase in number of vinculin plaques. These findings suggest that NSP of the right size (relative to charge) adsorb readily to the HA surface, changing the surface characteristics and, thus, improving osteoblast cell adhesion. This treatment provides a simple way to modify plasma-coated HA surfaces that may enable improved osseointegration of bone implants.

Project description:Focal Adhesion Kinase (FAK) is an important regulator of tumor cell proliferation, survival and metastasis. As such it has become a therapeutic target of interest in cancer. Previous studies suggested that use of FAK tyrosine kinase inhibitors (TKIs) blocks osteolysis in in vivo models of bone metastasis. However, from these studies it was not clear whether FAK TKIs blocked bone degradation by osteoclasts or also promoted bone formation by osteoblasts. In this study we evaluated whether use of the FAK TKI PF-562,271 affected the differentiation of pre-osteoblasts, or activity of mature differentiated osteoblasts. MC3T3-E1 pre-osteoblastic cells were treated with various doses of PF-562,271 following 3 or 10 days of differentiation which led to the inhibition of alkaline phosphatase (ALP) expression and reduced viable cell numbers in a dose-dependent manner. MC3T3-E1 cells which had been differentiated for 21 days prior to treatment with PF-562,271 showed a dose dependent decrease in mineralization as assessed by Alizarin Red staining, with concomitant decreased expression of ALP which is known to facilitate the bone mineralization activity of osteoblasts, however mRNA levels of the transcription factors RUNX2 and osterix which are important for osteoblast maturation and mineralization appeared unaffected at this time point. We speculated that this may be due to altered function of RUNX2 protein due to inhibitory phosphorylation by GSK3β. We found treatment with PF-562,271 resulted in increased GSK3β activity as measured by reduced levels of phospho-Ser9-GSK3β which would result in phosphorylation and inhibition of RUNX2. Treatment of 21 day differentiated MC3T3-E1 cells with PF-562,271 in combination with GSK3β inhibitors partially restored mineralization however this was not statistically significant. As we observed that FAK TKI also resulted in suppression of Akt, which is known to alter osterix protein stability downstream of RUNX2, we examined protein levels by western blot and found a dose-dependent decrease in osterix in FAK TKI treated differentiated MC3T3-E1 cells which is likely responsible for the reduced mineralization observed. Taken together our results suggest that use of FAK TKIs as therapeutics in the bone metastatic setting may block new bone formation as an off-target effect and thereby exacerbate the defective bone regulation that is characteristic of the bone metastatic environment.

Dataset Information

The effect of novel fluorapatite surfaces on osteoblast-like cell adhesion, growth, and mineralization.

Publications

The effect of novel fluorapatite surfaces on osteoblast-like cell adhesion, growth, and mineralization.

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