EPITHELIALIZATION OF TISSUE ENGINEERED CORNEAS

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DESCRIPTION (Adapted from applicant's abstract): Attempts to augment and replace the cornea with synthetic materials have met with limited success primarily because these devices have been unable to support and maintain a normal stratified epithelium. The fundamental problem is that the host epithelial cells recognize the polymer surface as other than stroma or "self" and, therefore, a degree of foreign body response is present throughout the life of the implant. Although many surfaces have been tested that allow some epithelial growth, the resulting epithelium is generally irregular, lacking the normal tri-layer appearance. Attempts to adsorb growth factors (GFs) and extracellular matrix proteins (ECMPs) onto the surfaces of these materials have not improved the epithelial cell response because 1) adsorption of these biopolymers to synthetic materials is non-specific and therefore they lay down on the surface in many different conformations, 2) some of the biopolymers bind irreversibly to the artificial surface, and 3) the biopolymer- coated surface, when placed in the eye, is itself coated by non-specific protein from the tear fluid. The applicants propose to investigate whether these problems can be avoided by tethering Extracellular matrix proteins (ECMPs) and growth factors (GFs; laminin, fibronectin, substance P, and insulin-like growth factor-1 [IGF-1]) onto synthetic hydrogel (polymer) surfaces. They predict that this tethering should significantly enhance the epithelial cell response for the following reasons. The tethers (or chains that fasten the ECMPs and GFs to the hydrogel surface) will hold the biological substances away from the polymer surface thus preserving their native conformational state. Additionally, tethers will hold the ECMPs and GFs away from the hydrogel surface; they will not be obscured by non-specific protein binding when placed in the tear environment. Specific experiments are designed to verify 1) that tether-modified hydrogel surfaces can be developed that retain the cell-adhesion activity of the tethered molecules, and 2) that tether-modified surfaces enhance the epithelial cell adhesion and migration across the hydrogel surfaces. Methodology includes Attenuated Total Reflectance-Fourier Transformed Infrared Spectroscopy (ATR-FTIR) to characterize the hydrogel surfaces, immunochemistry for tyrosine phosphorylation and F-actin, jet impingement to determine cellular adhesion, and immunohistochemistry for laminin, bullous pemphigoid antigen, and collagen VII. The results of these studies will provide insight into the processes involved in the success or failure of corneal augmentation and replacement by artificial implants.