Phenogenomics of the gingival fibroblast interaction with implant materials- a standardized system to determine material/surface potential for connective tissue attachment

摘要

The health of the soft-tissue surrounding a dental implant is an integral part of successful dental implant treatment. Human and animal models have shown that the peri-implant connective tissue contains relatively low numbers of fibroblasts, little or no blood vessels, similar to scar-like tissue. Such a tissue forms a poorly functional attachment to the implant surface unlike the gingival attachment to teeth. We report on an in vitro test at the phenotype and genotype level to quickly assess the scar forming potential of implant materials/surfaces. To test our methodology 3 titanium surfaces were used, namely pickled (PT) (Sa =0.3 μm), sandblasted (S) and sand blasted acid etched (SLA) (both Sa =1.2 μm). The surfaces were characterized by scanning electron microscopy (SEM). The initial gingival fibroblast (GF) response was quantified comparing cell adhesion, cell proliferation, immunolabelling and area of cell-implant contact with a focused ion beam/scanning electron microscope (FIB/SEM). Extracellular matrix protein (ECM) synthesis, anglogenesis potential and gene expression on each material surface were also quantified at 1 and 7 days. GFs cultured on all surfaces had distinct morphology after 1 day. On PT they were well spread. On S some cells were well spread and others had a stellate shape. All cells on SLA were stellate. Mature focal adhesions (FA) were visible on PT and S. Stellate shaped GFs cultured on S and SLA had small FA on the edges of surface features. Cross-sections of cell/implant with FIB/SEM confirmed this finding. FA area of adhesion was greatest on SLA which resulted in stronger cell adhesion as assessed by a cell detachment assay. After 1 day cell proliferation on SLA was significantly lower than PT and S but there was no difference after 7 days. Significant differences in expression of wound healing genes were seen relative to tissue culture plastic. Increased roughness caused a decrease in collagen Ⅰ,Ⅲand decorin genes whereas matrix metalloproteinase (MMP)-1 increased after 1d. MMP-3 expression increased only in SLA after 7d. VEGF was highly expressed in SLA but the tube formation assay with HUVEC cells showed no angiogenic effect. ECM synthesis quantification showed significant differences in collagen Ⅰ, decorin and fibronectin-EDA production. Production was highest on PT and the ECM had orientated with the topography in PT and S but not with SLA. This phenogenomic test suggests that surface topography is effective in modifying wound healing behavior at the GF phenotype, proteome and gene level.