Stephen J. Brand

Spasmolytic polypeptide: A trefoil peptide secreted by rat gastric mucous cells

BACKGROUND/AIMS Spasmolytic polypeptide (SP) is a trefoil peptide expressed in the digestive tract. This study aimed to determine the structure and distribution of SP expression in the rat gastrointestinal tract. METHODS The structure of rat SP was determined from the sequence of complementary DNAs isolated from antral RNA. SP gene expression was localized by Northern blotting and in situ hybridization in the adult and fetal rat digestive tract. Expression of the SP peptide was localized by immunocytochemistry and Western blot analysis. RESULTS SP messenger (m)RNA was found predominantly in the stomach with highest expression in the antrum. High levels of SP mRNA were expressed in the fetal stomach before gastrin and somatostatin expression. Surprisingly, SP mRNA and peptide did not colocalize in the gastric mucosa, SP mRNA being superficial to SP peptide immunoreactivity throughout the gastric mucosa. Abundant SP immunoreactivity was seen in the lumen of the gastric glands and the mucus layer adherent to the gastric mucosa, indicating luminal secretion. CONCLUSIONS In the rat, SP is a peptide secreted predominantly from antral mucous cell. The high concentrations of SP in the adherent gastric mucus layer (approximately 10 mumol/L) suggest that SP functions as a structural peptide rather than a regulatory peptide.

Role of the proteasome in rat indomethacin-induced gastropathy☆☆☆

BACKGROUND & AIMS Intercellular adhesion molecule (ICAM)-dependent adhesion of circulating neutrophils to microvascular endothelial cells is thought to be critical in causing indomethacin (nonsteroidal anti-inflammatory drug [NSAID])-induced gastropathy. Indomethacin stimulates tumor necrosis factor (TNF)-alpha expression, which may enhance adhesiveness of gastric capillaries for neutrophils by activating ICAM expression on endothelial cells. Stimulation of ICAM expression is mediated by activation of the transcription factor NF-kappaB. Because activation of NF-kappaB requires proteolytic degradation of IkappaB by the ubiquitin-proteasome pathway of intracellular proteolysis, treatment with proteasome inhibitors was evaluated for efficacy in preventing NSAID gastropathy. METHODS The effect of proteasome inhibitors on gastric injury caused by oral indomethacin was measured, along with their effects on gastric mucosal permeability measured by the blood to lumen EDTA clearance. Gastric ICAM expression was measured in vivo using infusion of a labeled rat ICAM antibody. RESULTS Proteasome inhibitors prevented NSAID gastropathy if administered from 0 to 12 hours before indomethacin. Equivalent efficacy was observed with intravenous and oral administration of proteasome inhibitors. There was a strong correlation between the potency of proteasome inhibitors in preventing NSAID gastropathy and their potency in inhibiting intracellular proteolysis or their anti-inflammatory potency. All three classes of proteasome inhibitors, peptide boronates, aldehydes, and the mechanistically different lactacystin, prevented NSAID gastropathy. Proteasome inhibitor treatment also abolished the increase in gastric mucosal permeability and the increase in gastric endothelial ICAM expression induced by indomethacin. CONCLUSIONS Indomethacin-induced gastric injury and increased ICAM expression are inhibited by inhibition of the proteasome.

Activation of gastrin gene transcription in islet cells by a RAP1‐like cis‐acting promoter element

Gastrin transcription in islet cells is activated by a cis‐regulatory sequence containing a binding site for the yeast transcription factor RAP1. The DNA—protein interactions between RAP1 protein and the gastrin DNA element determined by methylation interference assays are identical to those of RAP1 and yeast genes. Point mutations in the gastrin RAP1 binding site, which abolished RAP1 binding, decreased transcriptional activation by this sequence. Islet cells revealed a DNA binding protein with RAP1 ‐like binding specificity. These findings support the conclusion that gastrin transcription is activated in mammalian cells by a RAP1 ‐like transcription factor.