INTESTINAL CELL DIFFERENTIATION IN VIVO AND IN CULTURE

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The objective is the identification of the factors and cellular interactions which induce and modulate maturation and differentiation of the intestinal epithelium. The studies outlined in this proposal will focus on intestinal cell components defined by previously produced monoclonal antibodies which, because of their cell and/or tissue distribution in adult intestine and at different stages of development are potentially involved in those processes. New monoclonal antibodies will be prepared and characterized to further define the biological properties of the intestinal crypt cells and to detect individual cell types early in the process of commitment or differentiation. The composition, structure and biosynthesis of the newly identified antigens will be investigated by a combination of studies conducted in vivo and in culture, using intestinal epithelial cell lines and fetal rat intestinal organ cultures. The antigens recognized by the different monoclonal antibodies will be purified by affinity chromatography and characterized by slab gel electrophoresis, immunoblotting, peptide mapping, amino acid sequence determination, cell-free translation of purified intestinal mRNA, and cDNA cloning. The distribution of the various antigens in the intestinal mucosa at different stages of development and differentiation, and in cultured cells, will be determined by immunofluorescence staining and by immunoelectron-microscopy. Antibodies recognizing distinct crypt cell types will be used for a comparative study of the ultrastructure, relative position in the crypts, proliferative activity and development in fetal and newborn rats of the cells they identify. The studies should provide new insights into the process of intestinal cell differentiation, and also contribute to our knowledge of the intestinal mucosa in disease states such as sprue, inflammatory bowel diseases, interaction with luminal components, and intestinal adaptation.