The overall goal of this research proposal is to understand the contribution of mucin and non-mucin biliary glycoproteins to the pathogenesis of cholesterol gallstones. Cholesterol gallstones are the predominant type of stone in developed countries and medical or surgical treatment of symptomatic stones is a leading cause for hospitalization. Gallbladder mucin, the major component of the visco-elastic gel that lines the gallbladder epithelium, plays a central role in gallstone formation. Experimental evidence from animals and man indicate that gallbladder mucin is critical for both cholesterol crystal nucleation from cholesterol supersaturated bile, the initial event in gallstone formation, and in the subsequent growth and agglomeration of these crystals into mature gallstones. In addition to mucin, gallbladder bile appears to contain one or more non-mucin glycoproteins which also act as promotors of rapid cholesterol crystal nucleation. These proteins have been incompletely characterized and their contribution to the early events in gallstone formation are unknown.
The specific aims of this proposal are to determine the nucleotide sequence of gall bladder mucin cDNA and identify specific sequences on the protein core which are critical for binding biliary lipids and promoting nucleation. Since these activities have been localized to mucin non-glycosylated domains, these domains will be expressed, and the ability of the expressed polypeptides to bind biliary lipids and promote cholesterol crystal nucleation determined. The pathologic hypersecretion of mucin in the cholesterol-fed prairie dog model of gallstone formation will be used to study mucin gene expression using mucin specific antibodies and cDNA probes. The role of prostaglandins as mediators of mucin hypersecretion will be investigated. Finally, nonmucin glycoproteins which have been shown to be potent cholesterol crystal nucleators will be isolated from lithogenic prairie dog bile and biliary sludge. These proteins will be structurally characterized and regulation of their synthesis in response to cholesterol feeding will be studied in prairie dogs. Comparison of the kinetics of increases of mucin and non-mucin proteins in this model system will provide insights into the roles of these proteins in gallstone pathogenesis. Taken together, the results of these studies may lead to improved therapies for the prevention and treatment of this common disease.
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