BACKGROUND & AIMS: Gastrin induces the expression of cyclooxygenase (COX)-2 and interleukin (IL)-8; however, the mechanism(s), especially in gastric epithelial cells, is not well understood. Here, we have determined the intracellular mechanisms mediating gastrin-dependent gene expression. METHODS: AGS-E human gastric cancer cell line stably expressing cholecystokinin-2 receptor was treated with amidated gastrin-17. Real-time polymerase chain reaction, Western blot, and enzyme-linked immunosorbent assay were performed to determine COX-2 and IL-8 expression and Akt, Erk, and p38 phosphorylation. Gene promoter activity was determined by luciferase assay. Electrophoretic mobility shift assay analysis was performed for nuclear factor kappaB (NF-kappaB) and activator protein-1 activity. RNA stability was determined after actinomycin D treatment. HuR localization was determined by immunocytochemistry. RESULTS: Gastrin induced COX-2 and IL-8 expression in AGS-E cells, which was inhibited by phosphatidylinositol 3' kinase (PI3K) and p38 inhibitors. Gastrin-mediated Akt activation was observed to be downstream of p38. IL-8 expression was dependent on COX-2-mediated prostaglandin E(2) synthesis. In the presence of an NF-kappaB inhibitor MG132, IL-8 transcription was inhibited, but not that of COX-2. This was confirmed after knockdown of the p65 RelA subunit of NF-kappaB. Further studies showed that COX-2 gene transcription is regulated by activator protein-1. Gastrin increased the stability of both COX-2 and IL-8 messenger RNA (mRNA) in a p38-dependent manner, the half-life increasing from 31 minutes to 8 hours and approximately 4 hours, respectively. Gastrin, through p38 activity, also enhanced HuR expression, nucleocytoplasmic translocation, and enhanced COX-2 mRNA binding. CONCLUSIONS: Gastrin differentially induces COX-2 and IL-8 expression at the transcriptional and posttranscriptional levels by PI3K and p38 mitogen-activated protein kinase pathways, respectively.
BACKGROUND & AIMS:Gastrin induces the expression of cyclooxygenase (COX)-2 and interleukin (IL)-8; however, the mechanism(s), especially in gastric epithelial cells, is not well understood. Here, we have determined the intracellular mechanisms mediating gastrin-dependent gene expression. METHODS: AGS-E humangastric cancer cell line stably expressing cholecystokinin-2 receptor was treated with amidated gastrin-17. Real-time polymerase chain reaction, Western blot, and enzyme-linked immunosorbent assay were performed to determine COX-2 and IL-8 expression and Akt, Erk, and p38 phosphorylation. Gene promoter activity was determined by luciferase assay. Electrophoretic mobility shift assay analysis was performed for nuclear factor kappaB (NF-kappaB) and activator protein-1 activity. RNA stability was determined after actinomycin D treatment. HuR localization was determined by immunocytochemistry. RESULTS:Gastrin induced COX-2 and IL-8 expression in AGS-E cells, which was inhibited by phosphatidylinositol 3' kinase (PI3K) and p38 inhibitors. Gastrin-mediated Akt activation was observed to be downstream of p38. IL-8 expression was dependent on COX-2-mediated prostaglandin E(2) synthesis. In the presence of an NF-kappaB inhibitor MG132, IL-8 transcription was inhibited, but not that of COX-2. This was confirmed after knockdown of the p65RelA subunit of NF-kappaB. Further studies showed that COX-2 gene transcription is regulated by activator protein-1. Gastrin increased the stability of both COX-2 and IL-8 messenger RNA (mRNA) in a p38-dependent manner, the half-life increasing from 31 minutes to 8 hours and approximately 4 hours, respectively. Gastrin, through p38 activity, also enhanced HuR expression, nucleocytoplasmic translocation, and enhanced COX-2 mRNA binding. CONCLUSIONS:Gastrin differentially induces COX-2 and IL-8 expression at the transcriptional and posttranscriptional levels by PI3K and p38 mitogen-activated protein kinase pathways, respectively.
Authors: Asma Al Menhali; Theresa M Keeley; Elise S Demitrack; Linda C Samuelson Journal: Am J Physiol Gastrointest Liver Physiol Date: 2017-04-13 Impact factor: 4.052
Authors: Simon Almeida-Vega; Krista Catlow; Susan Kenny; Rod Dimaline; Andrea Varro Journal: Am J Physiol Gastrointest Liver Physiol Date: 2008-12-12 Impact factor: 4.052
Authors: Judy S Wang; Andrea Varro; Charles J Lightdale; Nantaporn Lertkowit; Kristen N Slack; Michael L Fingerhood; Wei Yann Tsai; Timothy C Wang; Julian A Abrams Journal: Am J Gastroenterol Date: 2009-11-10 Impact factor: 10.864