Aspects of signal transduction in the gastrointestinal tract
This study was undertaken to increase knowledge of the mechanisms of inter- and intracellular signalling in the gastrointestinal tract. Specific aims were: to use cell lines to elucidate factors affecting growth of gastric cells, to investigate the distribution and aspects of function of isoforms of protein kinase C in a gastric cell line and in the rat gastrointestinal tract and to determine the presence and regulation of nitric oxide synthase in gastrointestinal tissues from the rat and in cell lines. The gastric cancer cell line HGT-1 was used to investigate control of growth. Increases in cell number were found to be dependent on the seeding density of the cells. In cells plated at low density insulin, epidermal growth factor and gastrin all increased cell number. Gastrin produced a bell-shaped dose response curve with a maximum activity at 5nM. No effect of gastrin was apparent in cells plated at high density. and isoforms of protein kinase C were found, by immunoblotting procedures, to be widespread in the gastrointestinal tract of the rat, but protein kinase C was confined to the gastric mucosa and gastrointestinal smooth muscle. HGT-1 cells contained protein kinase C and but or were not detected. Preincubation of HGT-1 cells for 24h with 1M phorbol-12,13-dibutyrate down-regulated protein kinase C but not . The inhibition by the activator of protein kinase C, 12-O-tetradecanoylphorbol 13-acetate (TPA) of the histamine-stimulated increase in cAMP in HGT-1 cells was down regulated by phorbol-12,13-dibutyrate. Inhibition of histamine-stimulation of adenylate cyclase by TPA was Ca2+-dependent and inhibited by the addition of an antibody to protein kinase C . A role for protein kinase C in modulating the effect of histamine on adenylate cyclase in HGT-1 cells is suggested.