Correlation between corneal epithelial cell outgrowth and monoclonal antibody binding to the cell binding domain of adsorbed fibronectin.
The ability of corneal epithelial cells to attach, spread, and migrate on synthetic surfaces is largely determined by the characteristics of the adsorbed protein layer. In previous studies we have described an in vitro model for quantitating epithelial cell outgrowth from explanted corneal buttons onto synthetic materials (Pettit et al., Invest. Ophthalmol. Vis. Sci., 31, 2269 [1990]). We have also described the role of fibronectin (fn) adsorption and binding strength on epithelial cell outgrowth (Pettit et al., J. Biomed. Mater. Res., 26, 1259 [1992]). In the current study we have used a monoclonal antibody against the RGD cell binding domain of fn (mAb 3E3) to further characterize the role of adsorbed fn in promoting epithelial cell outgrowth. Ten materials of diverse chemical and physical properties were adsorbed with fn (0.1 mg/ml) or mixtures of fn and albumin (concentrations totaling 0.1 mg/ml) and tested for antibody recognition of the cell binding domain. The surface density of bound anti-cell binding domain antibody varied from a low of 0.66 +/- 0.11 for fluorinated ethylene propylene copolymer (FEP) to a high of 1.90 +/- 0.26 for tissue culture polystyrene dish substrates (units are OD at 450 nm measured in the ELISA technique normalized to polyethylene). A general increase in cell outgrowth areas was noted, with increases in recognizable cell binding domain. However, several exceptions to this trend were noted as well (e.g., low cell outgrowth but high antibody recognizability for glass). These results suggest that, although the number of cell binding domains exposed on adsorbed fn molecules may influence cell outgrowth, other characteristics of the adsorbed protein, such as the binding strength to the underlying substrate, may be equally important in characterizing epithelial cell-substrate interactions.
Pettit
D K
DK
Center for Bioengineering, University of Washington, Seattle 98195.
Hoffman
A S
AS
Horbett
T A
TA
D000327:Adsorption; D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal; D001665:Binding Sites; D001672:Biocompatible Materials; D002455:Cell Division; D003315:Cornea / Q000166:cytology* / Q000276:immunology; D004847:Epithelial Cells; D005353:Fibronectins / Q000737:chemistry / Q000276:immunology; D066298:In Vitro Techniques; D008422:Materials Testing; D008969:Molecular Sequence Data; D009842:Oligopeptides / Q000737:chemistry / Q000276:immunology; D011817:Rabbits; D013499:Surface Properties
D000911:Antibodies, Monoclonal; D001672:Biocompatible Materials; D005353:Fibronectins; D009842:Oligopeptides; C047981:arginyl-glycyl-aspartic acid