The corneal epithelium provides a barrier to fluid loss and pathogen entrance. Rapid healing of the epithelium in response to injury is essential for maintenance of the barrier function. The long-term goal of the laboratory is to obtain basic information about the molecular and cell biology of corneal wound-healing. Preliminary data demonstrated that amyloid B/A4 precursor-like protein (APLP2) has increased expression in healing corneal epithelium. APLP2 is very closely related to the amyloid B/AP4 precursor protein (APP) of Alzheimer's disease. The physiological role of both APP and APLP2 is not clear. Suggested functions of these proteins that may apply to epithelial wound healing include: transmembrane APLP2, binding to denuded basement membrane, may act as an adhesion molecule; proteolytically cleaved APLP2, associated with extracellular matrix, may promote cell adhesion; and alternatively spliced APLP2, containing a serine protease inhibitor domain, may regulate the activity of proteases required for effective cell migration during wound healing. Using a corneal debridement-wound as a model, the role of APLP2 will be explored to determine which function(s) of APLP2 are involved in corneal wound- healing. First, the expression of alternatively spliced isoforms in healing and normal corneal epithelium will be examined by cDNA cloning, quantitative PCR, and in situ hybridization. Second, a comparison of APLP2 distribution and proteolytic cleavage products will be made between wounded and unwounded corneas using immunotechniques. The role of APLP2 as serine protease inhibitor in epithelial migration will be investigated. The possibility that APLP2 may contribute to epithelial cell adhesion, especially in the healing cornea where dynamic cell adhesion is required, will be assessed using an in vitro cell adhesion assay. Finally, the role of APLP2 in regulating cell adhesion and migration will be explored by establishing permanent cell lines through gene transfection. Investigation of APLP2 in corneal epithelial wound- healing may add valuable information about this potentially important molecule and about the not-well-understood biological process of epithelial wound healing.
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