Integrin α2β1 plays a critical role in osteoblast response to micron-scale surface structure and surface energy of titanium substrates

Abstract

Efforts to improve bone response to biomaterials have focused on ligands that bind ␣5␤1 integrins. However, antibodies to ␣5␤1 reduce osteoblast proliferation but do not affect differentiation when cells are grown on titanium (Ti). ␤1-silencing blocks the differentiation stimulus of Ti microtopography, suggesting that other ␤1 partners are important. Stably ␣2-silenced MG63 human osteoblast-like cells were used to test whether ␣2␤1 specifically mediates osteoblast response to Ti surface micron-scale structure and energy. WT and ␣2-silenced MG63 cells were cultured on tissue culture polystyrene (TCPS) and Ti disks with different surface microtopographies: machined pretreatment (PT) surfaces [mean peak to valley roughness (R a) < 0.02 m], PT surfaces that were grit-blasted and acid-etched (SLA; R a ‫؍‬ 4 m), and SLA with high surface energy (modSLA). Alkaline phosphatase (ALP), ␣2 and ␤1 mRNA, but not ␣5, ␣v, ␤3, type-I collagen, or osteocalcin, increased on SLA and modSLA at 6 days. ␣2 increased at 8 days on TCPS and PT, but remained unchanged on SLA and modSLA. ␣2-protein was reduced 70% in ␣2-siRNA cells, whereas ␣5-mRNA and protein were unaffected. ␣2-knockdown blocked surface-dependent increases in ␤1 and osteocalcin and decreases in cell number and increases in ALP and local factors typical of MG63 cells grown on SLA and modSLA [e.g., prostaglandin E 2, osteoprotegerin, latent and active TGF-␤1, and stimulatory effects of 1␣,25(OH)2D3 on these parameters]. This finding indicates that ␣2␤1 signaling is required for osteoblastic differentiation caused by Ti microstructure and surface energy, suggesting that conclusions based on cell behavior on TCPS are not predictive of behavior on other substrates or the mechanisms involved.␣-2 integrin siRNA ͉ MG63 human osteoblasts ͉ titanium surface roughness T itanium (Ti) and Ti alloys are commonly used as biomaterials because their surface properties provide a biocompatible interface with peri-implant tissues. Strategies for modifying the nature of this interface frequently involve changes to the surface, thereby affecting protein adsorption, cell-substrate interactions, and tissue development (1). A common modification has been to create micron-scale and submicron scale roughness. Preclinical and clinical studies (2-12) show that these surfaces support greater bone-to-implant contact than smooth surfaces.How surface microstructure promotes an osteogenic response is an important question, because bone-forming osteoblasts preferentially colonize bone surfaces that have been preconditioned by bone-resorbing osteoclasts (13), resulting in complex micron-scale and submicron-scale morphologies (14). In vitro experiments using model surfaces indicate that migration, growth, and colony morphology of rat bone marrow cells (15) and osteoblasts (16-18) are sensitive to microstructure. These observations suggest that structural elements can modulate the spatial organization of cells and their ECM.The topography of osteoclast resorption pits in bone can be modeled by using Ti subs...

Biological Behavior of Osteoblast-like Cells on Titania and Zirconia Films Deposited by Cathodic Arc Deposition

“…Finally, the difference will affect the regulation of cell adhesion, motility, proliferation, and differentiation [36,37].…”

Adhesion and osteogenic differentiation of human mesenchymal stem cells on titanium nanopores

“…As integrins are receptor molecules for extracellular matrix molecules (e.g., the β 1 family), serum components (α v family) and immunoglobulin family adhesion molecules (β 2 family), these molecules are obviously critical in cell behaviour. Previous studies have identifi ed that integrin β 1 is required for osteoblastic differentiation on microscale Ti substrates (Olivares-Navarrete et al, 2008;Chuluyan et al, 1995). β 1 integrin recognises that the VCAM-1 counter receptor presents on endothelial cells and ECM proteins such as fibronectin, collagen and vitronectin (Issekutz, 1995;Meerschaert and Furie 1995;Lai et al, 2000).…”

“…Using an image analysis and PCR array, we show the infl uence of pore diameter on hMSC adhesion. Previous studies have shown that changes in cell shape or integrin expression could regulate commitment of mesenchymal cells to different lineage (Olivares-Navarrete et al, 2008;Lavenus et al, 2011). Hence, the osteogenic differentiation of hMSCs on the nanostructured Ti was compared with Ti planar substrates with or without osteogenic supplements.…”

“…However, these results have not been reproduced on the titanium substrates that are of high relevance for orthopaedic and dental applications. In view of this, Olivares-Navarrete et al (2008) studied integrin α2β1 expression and osteogenic differentiation of osteoblastic cells on Ti microstructures. Here, we have used physical vapour deposition (PVD) to coat nanoporous polycarbonate membranes to produce titanium substrates with controlled size nanostructure (30, 150 and 300 nm in diameter), but random distribution.…”

Osseointegration and Bioscience of Implant Surfaces - Current Concepts at Bone-Implant Interface

“…These results indicate that on rough surfaces osteoblasts exhibit a more differentiated phenotype, even though the proliferation is negatively affected. The mechanism by which topography influences osteoblast differentiation appears to be mediated by integrin signaling (Olivares-Navarrete et al, 2008) and mitogen-activated proteine kinase (MAPK) pathways (Schwartz et al, 2001b). The topography has also an effect on subsequent expression of transcription factors, ECM protein genes and cytokines (Balloni et al, 2009;Marinucci et al, 2006).…”