Effect of biomaterial surface properties on fibronectin–α5β1 integrin interaction and cellular attachment (original) (raw)
Related papers
2006
The ability of fibronectin (Fn) to mediate cell adhesion through binding to a 5 b 1 integrins is dependent on the conditions of its adsorption to the surface. Using a model system of alkylsilane SAMs with different functional groups (X ¼ OH, COOH, NH 2 and CH 3 ) and an erythroleukemia cell line expressing a single integrin (a 5 b 1 ), the effect of surface properties on the cellular adhesion with adsorbed Fn layers was investigated. 125 I-labeled Fn, a modified biochemical cross-linking/extraction technique and a spinning disc apparatus were combined to quantify the Fn adsorption, integrin binding and adhesion strength, respectively. This methodology allows for a binding equilibrium analysis that more closely reflects cellular adhesion found in stable tissue constructs in vivo. Differences in detachment strength and integrin binding were explained in terms of changes in the adhesion constant (c, related to affinity) and binding efficiency of the adsorbed Fn for the a 5 b 1 integrins (CH 3 ENH 2 oCOOHEOH) and the resulting average bond strength. Fn interacted more strongly with a 5 b 1 integrins when adsorbed on COOH vs. OH surfaces suggesting that negative charge may be a critical component of inducing efficient cellular adhesion. As evident by the low c values, Fn adsorbed on NH 2 and CH 3 surfaces interacted inefficiently with a 5 b 1 integrins and also possessed significant non-specific components to adhesion. Lastly, comparison of cellular adhesion to Fn adsorbed onto smooth and rough surfaces showed that nano-scale roughness altered cellular adhesion by increasing the surface density of adsorbed Fn. r
Journal of Biomedical Materials Research, 2003
Integrin-mediated cell adhesion to proteins adsorbed onto synthetic surfaces anchors cells and triggers signals that direct cell function. In the case of fibronectin (Fn), adsorption onto substrates of varying properties alters its conformation/structure and its ability to support cell adhesion. In the present study, self-assembled monolayers (SAMs) of alkanethiols on gold were used as model surfaces to investigate the effects of surface chemistry on Fn adsorption, integrin binding, and cell adhesion. SAMs presenting terminal CH 3 , OH, COOH, and NH 2 functionalities modulated adsorbed Fn conformation as determined through differences in the binding affinities of monoclonal antibodies raised against the central cell-binding domain (OH Ͼ COOH ϭ NH 2 Ͼ CH 3 ). Binding of ␣ 5  1 integrin to adsorbed Fn was controlled by SAM surface chemistry in a manner consistent with antibody binding (OH Ͼ COOH ϭ NH 2 Ͼ CH 3 ), whereas ␣ V integrin binding followed the trend:
Cell Adhesion Strength Increases Linearly with Adsorbed Fibronectin Surface Density
Tissue Engineering, 1997
Cell adhesion is involved in numerous physiological processes and is important to biotechnological applications, including tissue engineering and development of artificial organs. The relationship between cell adhesion strength and fibronectin (Fn) surface density was analyzed using a spinning disk device that applied a linear range of forces to attached cells under uniform surface chemical conditions. ROS 17/2.8 cells were seeded onto Fn-coated glass substrates for 15 min. Cells were then subjected to detachment forces for 10 min and adherent cells were counted at different radial distances. The fraction of adherent cells decreased non-linearly with applied force, and the resulting detachment profile was accurately described by a sigmoidal curve. Analysis of detachment profiles for different Fn concentrations and quantitative Fn adsorption measurements revealed that, for short attachment times, cell adhesion strength increased linearly with Fn surface density. This linear dependence of attachment strength on adsorbed Fn was observed for two different glasses, a non-reactive substrate and a surface-active glass, suggesting that this relationship is not substrate specific. The increases in adhesion strength were effectively blocked by a monoclonal antibody directed against the RGD cell binding domain. The linear relationship between attachment strength and ligand density is consistent with theoretical models for initial receptormediated adhesion and suggests the absence of cooperative receptor-ligand binding during the initial phases of cell adhesion.
Targeted cellular adhesion at biomaterial interfaces
Current Opinion in Solid State & Materials Science, 1998
The interface of biomaterials must be carefully designed to elicit and eliminate specific responses when placed in contact with the body. The interaction of cells with the surfaces of biomaterials is a complex phenomenon that depends on a large number of variables. To design novel biomaterials that possess the desired characteristics, materials scientists and engineers rely heavily upon information provided by molecular biologists. Information regarding cell receptor-ligand interactions is used to understand the role of cellular adhesion in the natural environment so that synthetic biomaterials may be developed successfully. The production of new synthetic materials, understanding how native proteins mediate cellular adhesion with these materials, molecularly engineering surfaces with controlled spatial patterns for optimal responses, evaluation of mechanical properties, and analyzing bioadhesion and surface properties of these materials are topics that must be addressed when discussing targeted cellular adhesion at biomaterial surfaces.
Biomaterials, 2005
Integrin binding and focal adhesion assembly are critical to cellular responses to biomaterial surfaces in biomedical and biotechnological applications. While immunostaining techniques to study focal adhesion assembly are well established, a crucial need remains for quantitative methods for analyzing adhesive structures. We present simple yet robust approaches to quantify integrin binding and focal adhesion assembly on biomaterial surfaces. Integrin binding to fibronectin and a RGDcontaining synthetic peptide was quantified by sequentially cross-linking integrin-ligand complexes via a water-soluble homo-bifunctional cross-linker, extracting bulk cellular components in detergent, and detecting bound integrins by ELISA. Focal adhesion components (vinculin, talin, a-actinin) localized to adhesion plaques were isolated from bulk cytoskeletal and cytoplasmic components by mechanical rupture at a plane close to the basal cell surface and quantified by Western blotting. These approaches represent simple and efficient methodologies to analyze structure-function relationships in cell-material interactions.
The effect of non-specific interactions on cellular adhesion using model surfaces
2005
The contribution of non-specific interactions between cells and model functional surfaces was measured using a spinning disc apparatus. These model functional surfaces were created using self-assembled monolayers (SAM) of alkylsilanes terminated with epoxide, carboxyl (COOH), amine (NH 2 ), and methyl (CH 3 ) groups. These SAMs were characterized using ellipsometry, atomic force microscopy, contact angle goniometry, and X-ray photoelectron spectroscopy to confirm the presence of well-formed monolayers of expected physicochemical characteristics. All substrates also demonstrated excellent stability under prolonged exposure (up to 18 h) to aqueous conditions. The adhesion strength of K100 erythroleukemia cells to the functional substrates followed the trend: CH 3 o COOH E epoxide { NH 2 . The NH 2 SAM surface exhibited nearly an order of magnitude greater adhesion strength than the other SAMs and this non-specific effect exceeded the adhesion measured when RGD tri-peptides were also immobilized on the surface. These findings illustrate the importance of substrate selection in quantitative studies of peptidemediated cellular adhesion. r
ACS Biomaterials Science & Engineering, 2018
Surface modification plays vital role in regulating protein adsorption and subsequently cell adhesion. In the present work, we prepared nanoscaled modified surfaces using silanization and characterized them using Fourier-transform infrared spectroscopy (FTIR), water contact angle (WCA) and Atomic Force microscopy (AFM). Five different (amine, octyl, mixed, hybrid and COOH) surfaces were prepared based on their functionality and varying wettability and their effect on protein adsorption and initial cell adhesion was investigated. AFM analysis revealed nanoscale roughness on all modified surfaces. Fetal bovine serum (FBS) was used for protein adsorption experiment and effect of FBS was analyzed on initial cell adhesion kinetics (upto 6 h) under three different experimental conditions: (a) with FBS in media, (b) with pre-adsorbed FBS on surfaces and (c) incomplete media, i.e., without FBS. Various cell features such as cell morphology/circularity, cell area and nuclei size were also studied for above stated conditions at different time intervals. The cell adhesion rate as well as cell spreaded area were highest in case of surfaces with pre-adsorbed FBS. We observed higher surface coverage rate by adhering cells on hybrid (rate, 0.073 h-1) and amine (0.072 h-1) surfaces followed by COOH (0.062 h-1) and other surfaces under pre-adsorbed FBS condition. Surface treated with cells in incomplete media exhibited least adhesion rate, poor cell spreading and improper morphology. Furthermore, we found that initial cell adhesion rate and ∆ adhered cells (%) linearly increased with the change in α-helix content of adsorbed FBS on surfaces. Amongst all the modified surfaces and under all three experimental conditions, hybrid surface exhibited excellent properties for supporting cell adhesion and growth and hence can be potentially used as surface modifiers in biomedical applications to design biocompatible surfaces.
Biomaterials, 2004
Biomaterial surface properties influence protein adsorption and elicit diverse cellular responses in biomedical and biotechnological applications. However, the molecular mechanisms directing cellular activities remain poorly understood. Using a model system with well-defined chemistries (CH 3 , OH, COOH, NH 2 ) and a fixed density of the single adhesive ligand fibronectin, we investigated the effects of surface chemistry on focal adhesion assembly and signaling. Surface chemistry strongly modulated integrin binding and specificity-a 5 b 1 integrin binding affinity followed the pattern OH>NH 2 QCOOH>CH 3 , while integrin a V b 3 displayed the relationship COOH>NH 2 cOHQCH 3 . Immunostaining and biochemical analyses revealed that surface chemistry modulates the structure and molecular composition of cell-matrix adhesions as well as focal adhesion kinase (FAK) signaling. The neutral hydrophilic OH functionality supported the highest levels of recruitment of talin, a-actinin, paxillin, and tyrosinephosphorylated proteins to adhesive structures. The positively charged NH 2 and negatively charged COOH surfaces exhibited intermediate levels of recruitment of focal adhesion components, while the hydrophobic CH 3 substrate displayed the lowest levels. These patterns in focal adhesion assembly correlated well with integrin a 5 b 1 binding. Phosphorylation of specific tyrosine residues in FAK also showed differential sensitivity to surface chemistry. Finally, surface chemistry-dependent differences in adhesive interactions modulated osteoblastic differentiation. These differences in focal adhesion assembly and signaling provide a potential mechanism for the diverse cellular responses elicited by different material properties.
Effect of Preadsorbed Proteins on Cell Adhesion to Polymer Surfaces
Journal of Colloid and Interface Science, 1993
Adsorption of three different proteins and adhesion of cells onto various substrates in the presence of serum proteins were studied. Both the maximal protein adsorption and the maximal cell adhesion were observed on surfaces with water contact angle around 70°. Preadsorption of serum albumin prevented cell adhesion to all the substrates, whereas preadsorbed fibronectin (FN) enhanced cell adhesion to all the substrates, independent of their water wettability, except for poly(vinyl alcohol) and acrylamide-grafted films. Competitive adsorption of FN from mixed proteins, ranging from 0.03 to 0.07 p.g/cm2 , markedly influenced cell adhesion in the presence of serum. These results suggest that the effect of the water wettability of surfaces on cell adhesion in the presence of serum should occur through protein layers adsorbed directly to the substrate surfaces. ©1993 Academic Press, inc.