Mucin is the predominant glycoprotein secreted by gastric epithelium, and is the principal component of mucus. Gastric mucus lines the epithelium and protects it from the harmful constituents of the gastric lumen, chief among these being secreted acid. The mucus coat provides an unstirred layer which maintains the pH gradient, and disruption of this layer appears to be associated with mucosal injury. In spite of this important protective role, details about the regulation of gastric mucin synthesis and secretion are limited, primarily because of the complexities of intact gastric mucosa. This proposal describes a novel system in which the gastric mucous cell may be studied in isolated cell culture, so that the regulation of mucin synthesis and its release may be studied at the cellular level. This type of information will permit an integrated interpretation of data derived in the whole animal or organ culture model. Our preliminary data suggest that mucin synthesis may be regulated in a manner parallel to that of acid secretion, and this can be best explored in more depth in an isolated cell culture system. The effects of hormones, neuro- transmitters, and autocoids on the regulation of mucin production will be investigated. The isolated mucous cell preparation permits an investigation of glycoprotein synthesis and secretion in the stomach at a cellular and molecular level. Signal transduction mechanisms will be investigated, as will the molecular processing of gastric mucin. The regulation of oligosaccharide processing (specifically, fucosylation, sulfation, and the synthesis of terminal glycosylation structures), will be studied in the context of the regulation of mucin synthesis. The mucin apoprotein will be explored by harvesting messenger RNA from stimulated cells, and synthesizing a carbohydrate-free mucin in a cell free translation system. In summary, the isolated gastric mucous cell system will be used to answer questions that cannot be addressed in more complex systems involving intact mucosa. Data are required at cellular and molecular level to properly integrate and understand the information that has previously been obtained regarding gastric mucin.
