Normal conjunctival epithelium contains goblet cells, which are a primary source of mucin. These cells and their secretory product are crucial for ocular surface integrity. When conjunctival epithelium covers corneal wounds, goblet cells are initially present after stratification; then the epithelium gradually loses goblet cells and morphologically transforms into a cornea-like appearance. This process is termed conjunctival transdifferentiation. Our working hypothesis is that vitamin A or retinoids are the responsible factor(s) in modulating conjunctival transdifferentiation and are necessary for goblet cell differentiation. Our preliminary studies suggest that several retinoids can modulate conjunctival transdifferentiation in vivo. The major objective of this proposal is to test the hypothesis in vitro, so that the mechanisms by which conjunctival goblet cells are lost or maintained can be better understood. We have the following specific aims: (1) Establish an organ culture system for conjunctival transdifferentiation. Experiments involve manipulating various culture media conditions to determine the best chemically defined medium for optimal growth of normal rabbit conjunctival epithelium as well as of transdifferentiating epithelium. To assess goblet cell differentiation, our recently developed topographical method for counting goblet cells can be used in conjunction with routine histology and ultrastructural studies. (2) Concurrently, develop monoclonal antibodies against conjunctival mucin so that the degree of goblet cell differentiation can be rapidly examined in vitro by immunoassay. With monoclonal antibodies to conjunctival mucins, the qualitative change(s) of mucin during the loss of goblet cells can also be explored. (3) After the successful completion of the studies in Aims 1 and 2, various retinoid analogues will be added to the media at different concentrations to examine the capability of maintaining goblet cell differentiation. (4) Using the same in vitro system, the possible modulating effects of other soluble factors on goblet cell differentiation will be investigated.
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