Characterization of Corneal Epithelial Stem Cells

George Washington University, Washington DC

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Abstract

DESCRIPTION (provided by applicant): The long term goal of this project is to use molecular approaches to define a set of cell specific surface narkers for human as well as rodent limbal stem cells and to use those markers as tools to characterize stem cells for the corneal epithelium in humans, rats, and mice in health and disease. Clinical experience and practice has shown us that limbal stem cells serve as the source for the cells comprising the corneal epithelium. At present, experimental confirmation that cells in the limbus are stem cells relies upon data from studies of vitro cell proliferation capacity and on the identification of a subset of these cells as slow-cycling. We hypothesize that a specific repertoire of cell surface markers exists which can define and select stem cells within the corneal limbus. Identification of such markers will aid in the study of the human ocular surface by providing tools to use to both select for and to target specific drugs or genes to these cells. Further, they will provide tools to allow for the analysis of limbal stem cell properties both in vivo and in vitro. The specific aims of this application are: 1) Identification of corneal stem-like cells from donor human corneas by selection based on cell adhesion properties, integrin expression profiles, and the ability of cells to proliferate in vitro, 2. Using phage displayed peptide libraries, identification of peptides that bind specifically to human, mouse, and rat limbal basal cells and use of those peptides to isolate and characterize stem cell specific surface markers, 3. In the mouse and rat, determine whether the slow-cycling cells observed in the limbus after neonatal BrdU administration possess similar cell surface markers as do human corneal stem cells, and 4. Using the knowledge and tools developed in Aims 1-3, characterize the limbal stem cells in the mouse under conditions where their differentiation and/or proliferation is altered in wild-type and in genetically engineered mice exhibiting symptoms of stem cell disease. Discovering specific molecular markers for the limbal stem cells will aid in the development of treatments for stem cell deficiency diseases of the ocular surface in humans.