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Gastrin, a small polypetide, is secreted by G cells in the gastric antrum. In addition to stimulating acid secretion, gastrin induces a pro-inflammatory response resulting in overexpression of cyclooxygenase-2 (COX-2) and increased production of prostoglandin E2 (PGE2). Gastrin is also known to stimulate growth of malignant cells of colorectal, gastric, and pancreatic origin. However, the mechanism(s), especially in gastric epithelial cells is not completely understood. In colon cancer cells, gastrin signaling through the CCK-2 receptor activates the PI3 Kinase (PI3K) and ERK pathways to induce COX-2 and interleukin-8 (IL-8) expression. To determine the effect of gastrin on gastric epithelial cells, we treated AGS-E cells, a human gastric cancer cell line stably expressing the CCK-B receptor with amidated gastrin-17 peptide in the presence or absence of a cancer therapeutic agent CPT-11, a topoisomerase 1 inhibitor. While gastrin did not affect cell proliferation, it significantly inhibited CPT-11 mediated apoptosis. Western blot analyses demonstrated that gastrin induced AKT and ERK phosphorylation. Real time RT-PCR analyses of total RNA demonstrated a dose- and time-course dependent induction of IL-8 and COX-2 mRNA expression by gastrin. This was further confirmed at the level of transcription by transfection assays using luciferase reporter gene expression under the control of COX-2 and IL-8 promoters. Gastrin-mediated IL-8 and COX-2 gene expression was partially inhibited by LY294002 (PI3K inhibitor) and SB203580 (p38 MAPK inhibitor). In contrast, while MG132, a potent suppressor of IκB degradation, significantly inhibited gastrin-mediated IL-8 expression, it enhanced gastrin-mediated COX-2 expression, suggesting different modes of regulation for the two genes. Both, IL-8 and COX-2 are regulated at the transcriptional level by NF-κB. Electrophoretic mobility shift assays and luciferase reporter assays demonstrated that MG-132 suppressed gastrin-mediated induction of NF-κB activity. IL-8 and COX-2 transcripts are also regulated at the posttranscriptional level of mRNA stability. Gastrin increased the stability of both transcripts, the half-life increasing from ∼3 h to >8 h. Furthermore, there was increase expression of both proteins. However, addition of SB203580, but not LY294002 abrogated gastrin-mediated increased mRNA stability of both COX-2 and IL-8 mRNA. These results suggest that two distinct pathways mediate gastrin-induced up-regulation of COX-2 and IL-8 expression, a PI3K-dependent pathway that induces gene transcription and a PI3K-independent, p38-dependent pathway that induces mRNA stability.

[Proc Amer Assoc Cancer Res, Volume 47, 2006]