PATHOGENEISIS OF CYSTIC FIBROSIS IN THE GI SYSTEM

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The long-term objective is to understand how loss of functional CFTR (the cystic fibrosis transmembrane regulator) affects expression and post- translational processing of sulfated glycoconjugates and the role of these altered glycoproteins in the development of cystic fibrosis (CF). High levels of sulfated glycoconjugates with abnormal post-translational processing are believed to contribute to obstruction in the pancreas and intestines, and digestion and absorption of nutrients is abnormal in CF. The model mucin-like sulfated glycoprotein Muclin will be used to explore these issues. Muclin is over-expressed in the pancreas and small intestine of CFTR (-/-) mice, and immunolocalization of Muclin in these tissues shows that it is associated with protein plugs in the acinar lumen and mucin plugs in the intestinal crypt lumen. The hypothesis is that in the absence of functional CFTR, mucin-like glycoconjugates, such as Muclin, are over-expressed and have altered post-translational processing. They then contribute to the deleterious accumulation of protein aggregates in the lumina of the pancreatic acini and small intestinal crypts. The following specific aims will be addressed. 1. To test the hypothesis that post-translational processing of Muclin is altered in the absence of CFTR in epithelial cells which normally express CFTR, i.e., the intestinal crypt cell, and to determine the mechanism of this change. The carbohydrate structure and protein core size of Muclin in normal and CF intestine will be determined. 2. To test the hypothesis that excess Muclin alters the protein secretory pathway in gastrointestinal tissues of CF mice. Pancreatic secretions will be studied in vivo and in vitro in normal and CF mice, and the effect of pH on the interaction of Muclin with zymogens will be assessed. Also, the luminal pH of secretory compartments in CF and normal cells will be measured to test whether CFTR has a functional role in pH gradients in the cell. 3. To test the hypothesis that abnormal acidity in the intestine mediates CF pathogenesis and the over-expression of Muclin. Pathology and Muclin expression will be compared in CF mice, and CF mice crossed with gastrin deficient mice (lack gastric acid secretion) to normalize intestinal Ph.