MORPHOLOGICAL AND MOLECULAR CHARACTERIZATION OF HUMAN GASTRIC MUCOUS CELLS IN LONG-TERM PRIMARY CULTURE

A primary cell culture of human gastric mucous cells was developed usi ng enzymatic treatment of surgically obtained gastric mucosal specimen s. Preferential attachment of gastric mucous cells during a preincubat ion step resulted in the enrichment of mucous cells [over 90% stained with periodic acid-Schiff (PAS) and mucin-type lectins] in the primary cell culture. Gastric mucous cells could be maintained in culture for 10 days. DNA synthesis peaked during the first 2 days in culture (8+/ -1% bromodeoxyuridine-positive cells). During the entire culture perio d gastric mucous cells released high-molecular-weight glycoproteins in to the medium, as determined by gel chromatography on a Sepharose CL-4 B column and by metabolic labelling with [C-14]-N-acetylglucosamine. G astric mucin was verified by gas chromatographic analysis of the carbo hydrate composition and fractionation of the void-volume fraction by d ensity gradient centrifugation. Determination of the terminal glycosyl ation of the secreted glycoproteins by a lectin-ELISA revealed that th ere was a high quantity of alpha-L-fucose. Prostaglandin E-2 significa ntly stimulated glycoprotein secretion during the entire cultivation p eriod by 29-60%. Analysis of mucin-encoding MUC mRNA expression by rev erse transcriptase polymerase chain reaction revealed that gastric muc ous cells predominantly express MUC1 and MUC5AC, and to a lesser exten t MUC6, which reflects the expression pattern obtained following analy sis of biopsied samples of gastric mucosa. This primary cell culture m odel enables the regulation of mucin secretion and mucin gene expressi on in man to be investigated.