Hypertonic Stress Induces VEGF Production in Human Colon Cancer Cell Line Caco-2: Inhibitory Role of Autocrine PGE2

Luciana B Gentile,Bruno Piva,Bruno L Diaz,Ben C B Ko

doi:10.1371/journal.pone.0025193

Abstract

Vascular Endothelial Growth Factor (VEGF) is a major regulator of angiogenesis. VEGF expression is up regulated in response to micro-environmental cues related to poor blood supply such as hypoxia. However, regulation of VEGF expression in cancer cells is not limited to the stress response due to increased volume of the tumor mass. Lipid mediators in particular arachidonic acid-derived prostaglandin (PG)E2 are regulators of VEGF expression and angiogenesis in colon cancer. In addition, increased osmolarity that is generated during colonic water absorption and feces consolidation seems to activate colon cancer cells and promote PGE2 generation. Such physiological stimulation may provide signaling for cancer promotion. Here we investigated the effect of exposure to a hypertonic medium, to emulate colonic environment, on VEGF production by colon cancer cells. The role of concomitant PGE2 generation and MAPK activation was addressed by specific pharmacological inhibition. Human colon cancer cell line Caco-2 exposed to a hypertonic environment responded with marked VEGF and PGE2 production. VEGF production was inhibited by selective inhibitors of ERK 1/2 and p38 MAPK pathways. To address the regulatory role of PGE2 on VEGF production, Caco-2 cells were treated with cPLA2 (ATK) and COX-2 (NS-398) inhibitors, that completely block PGE2 generation. The Caco-2 cells were also treated with a non selective PGE2 receptor antagonist. Each treatment significantly increased the hypertonic stress-induced VEGF production. Moreover, addition of PGE2 or selective EP2 receptor agonist to activated Caco-2 cells inhibited VEGF production. The autocrine inhibitory role for PGE2 appears to be selective to hypertonic environment since VEGF production induced by exposure to CoCl2 was decreased by inhibition of concomitant PGE2 generation. Our results indicated that hypertonicity stimulates VEGF production in colon cancer cell lines. Also PGE2 plays an inhibitory role on VEGF production by Caco-2 cells exposed to hyperosmotic stress through EP2 activation.

Highlights

Formation of new blood vessels from pre-existing vasculature is a central process in the development of most tumors especially solid ones
As PGE2 has been described to play a role in angiogenesis and Vascular Endothelial Growth Factor (VEGF) production we determined whether PGE2 was regulating VEGF production by Caco-2 during the stimulation with hypertonic medium
To verify whether inhibition of VEGF production by Caco-2 stimulated with hypertonic stress was a consequence of PGE2 action and not a consequence of the nonespecific action of pharmacological drugs used in the experiments, we performed the same experiment in HCT116, a colon cancer cell line that does not express COX-2 and produces no detectable levels of PGE2 under hypertonic stress

Summary

Introduction

Formation of new blood vessels from pre-existing vasculature is a central process in the development of most tumors especially solid ones. This process is called angiogenesis and is regulated by the balance of negative and positive biochemical signals. VEGF is the most prominent positive regulator of angiogenesis due to its ability to recruit endothelial cells to hypoxic sites and to stimulate the proliferation of this cellular type, promoting the differentiation of vascular structures [1]. Expression of VEGF correlates with increased metastatic potential [2], while expression of its receptor is a marker of shorter post-operative survival [3]

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Results

Conclusion