Bile Acid Reflux Contributes to the Progression from Barrett's Esophagus to Esophageal Adenocarcinoma Via Activation of a Novel Bile Acid Receptor TGR5 and NADPH Oxidase NOX5-S
W Cao, J Hong, J Behar, J Wands, LJ Wang, RA DeLellis, D Lambeth, M Resnick. Rhode Island Hospital and Warren Alpert Medical School of Brown University, Providence, RI; Emory University School of Medicine, Atlanta, GA
Background: Gastroesophageal reflux disease complicated by Barrett's esophagus (BE) is a major risk factor for esophageal adenocarcinoma (EA). However, the mechanisms of the progression from BE to EA are not fully understood. Bile acids may play an important role in this progression. The aim of this study is to determine the expression of a novel bile acid receptor TGR5 in BE and to examine the role of NADPH oxidase NOX5-S and TGR5 in taurodeoxycholic acid (TDCA)-induced increased cell proliferation.
Design: Immunohistochemical and RT-PCR assessment of TGR5 expression were performed on biopsy samples of BE and EA. The human Barrett's EA cell line FLO was transfected with siRNA or plasmids using Lipofectamine 2000 or Amaxa-Nucleofector-System respectively. NOX5 and TGR5 mRNA were measured by real-time PCR. H2O2 was measured by using the Amplex® Red H2O2 assay kit. Cell proliferation was determined by measurement of thymidine incorporation.
Results: Immunohistochemical staining showed that TGR5 was present in BE and EA, and almost undetectable in esophageal squamous epithelial cells. TGR5 mRNA levels were significantly higher in EA tissues than in normal esophageal mucosa or Barrett's mucosa. In FLO cells, NADPH oxidase NOX5-S was present. TDCA significantly increased NOX5-S expression, H2O2 production and thymidine incorporation. This increase in thymidine incorporation was significantly reduced by knockdown of NOX5-S. Knockdown of TGR5 markedly inhibited TDCA-induced increase in NOX5-S expression, H2O2 production and thymidine incorporation. Conversely overexpression of TGR5 significantly enhanced TDCA's effects. TGR5 receptors were coupled with Gαq and Gαi-3 proteins, but only Gαq mediated TDCA-induced increase in NOX5-S expression, H2O2 production and thymidine incorporation.
Conclusions: TDCA-induced increase in cell proliferation depends on upregulation of NOX5-S expression. TDCA-induced NOX5-S expression may be mediated by activation of the TGR5 receptor and Gαq protein in FLO EA cells. Supported by NIH NIDDK R01 DK080703.
Wednesday, March 24, 2010 9:30 AM
Poster Session V # 77, Wednesday Morning