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Quantitative analysis of the effect of Helicobacter pylori on the expressions of SOX2, CDX2, MUC2, MUC5AC, MUC6, TFF1, TFF2, and TFF3 mRNAs in human gastric carcinoma cells.

ABSTRACT: OBJECTIVE:To investigate the phenotypic characters of carcinoma cells and the response of gastric epithelial cells to Helicobacter pylori (H. pylori) infection using the gastric carcinoma cell lines. MATERIAL AND METHODS:Real-time reverse transcription-polymerase chain reaction (RT-PCR) was used to assess the effect of H. pylori infection on mRNA levels of transcription factors (SOX2 and CDX2), mucin core proteins (MUC2, MUC5AC, and MUC6), and trefoil factor family peptides (TFF) (TFF1, TFF2, and TFF3) in gastric carcinoma cells (AGS, MKN45, and KATO III cells). H. pylori ATCC 43504 and its isogenic cag pathogenicity island (PAI) deleted mutant were used. RESULTS:These cell lines expressed mixed gastric and intestinal phenotypes. The intestinal phenotype predominated in AGS cells and gastric phenotypes in MKN45 and KATO III cells. In all three cell lines, H. pylori infection inhibited SOX2 mRNA expression, but induced the three TFFs mRNAs. In AGS cells, H. pylori induced cag PAI-dependent mRNA expression of CDX2, MUC2, MUC5AC, and MUC6. mRNA expressions of CDX2, MUC5AC, and MUC6 were inhibited in KATO III cells, whereas MUC2 mRNA expression was unchanged. In MKN45 cells, H. pylori induced the three MUCs mRNAs but inhibited CDX2 mRNA expression. CONCLUSIONS:This study provides a useful platform for selecting appropriate cell lines to model H. pylori-related changes in the gastric epithelium that mirror the changes seen in vivo. The outcome of H. pylori infection may reflect changes in the mucus gel layer caused by altered expression of mucins and TFF peptides.

SUBMITTER: Matsuda K

PROVIDER: S-EPMC3128253 | biostudies-literature | 2008 Jan

REPOSITORIES: biostudies-literature

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Quantitative analysis of the effect of Helicobacter pylori on the expressions of SOX2, CDX2, MUC2, MUC5AC, MUC6, TFF1, TFF2, and TFF3 mRNAs in human gastric carcinoma cells.

Matsuda Kazuyuki K Yamauchi Kazuyoshi K Matsumoto Takehisa T Sano Kenji K Yamaoka Yoshio Y Ota Hiroyoshi H

Scandinavian journal of gastroenterology 20080101 1

<h4>Objective</h4>To investigate the phenotypic characters of carcinoma cells and the response of gastric epithelial cells to Helicobacter pylori (H. pylori) infection using the gastric carcinoma cell lines.<h4>Material and methods</h4>Real-time reverse transcription-polymerase chain reaction (RT-PCR) was used to assess the effect of H. pylori infection on mRNA levels of transcription factors (SOX2 and CDX2), mucin core proteins (MUC2, MUC5AC, and MUC6), and trefoil factor family peptides (TFF) ...[more]

PMID: 18938748

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Project description:Background:Intestinal metaplasia (IM) is a premalignant lesion associated with gastric cancer. Both animal and clinical studies have revealed that bile acid reflux and subsequent chronic inflammation are key causal factors of IM. Previous studies indicated that SOX2, the key transcription factor in gastric differentiation, was downregulated during IM development while CDX2, the pivotal intestine-specific transcription factor was upregulated significantly. However, it remains unclear whether the downregulation of SOX2 promotes gastric IM emergence or is merely a concomitant phenomenon. In addition, the underlying mechanisms of SOX2 downregulation during IM development are unclear. Methods:Gastric cell lines were treated with deoxycholic acid (DCA) in a dose-dependent manner. The expression of CDX2 and miR-21 in gastric tissue microarray were detected by immunohistochemistry and in situ hybridization. Coimmunoprecipitation and immunofluorescence were performed to ascertain the interaction of SOX2 and CDX2. Luciferase reporter assays were used to detect the transcriptional activity of CDX2, and confirm miR-21 binding to SOX2 3'-UTR. The protein level of SOX2, CDX2 and downstream IM-specific genes were investigated using western blotting. mRNA level of miR-21, SOX2, CDX2 and downstream IM-specific genes were detected by qRT-PCR. Results:Bile acid treatment could suppress SOX2 expression and simultaneously induce expression of CDX2 in gastric cell lines. Furthermore, we demonstrated that SOX2 overexpression could significantly inhibit bile acid- and exogenous CDX2-induced IM-specific gene expression, including KLF4, cadherin 17 and HNF4? expression. In contrast, SOX2 knockdown had the opposite effect. A dual-luciferase reporter assay demonstrated that SOX2 overexpression could significantly suppress CDX2 transcriptional activity in HEK293T cells. CDX2 and SOX2 could form protein complexes in the nucleus. In addition, bile acid induced the expression of miR-21. The inhibition of SOX2 in bile acid-treated gastric cell lines was rescued by miR-21 knockdown. Conclusions:These findings suggested that SOX2 can interfere with the transcriptional activity of CDX2 in bile acid-induced IM and that miR-21 might play a key role in this process, which shed new lights in the prevention of gastric cancer.

Dataset Information

Quantitative analysis of the effect of Helicobacter pylori on the expressions of SOX2, CDX2, MUC2, MUC5AC, MUC6, TFF1, TFF2, and TFF3 mRNAs in human gastric carcinoma cells.

Publications

Quantitative analysis of the effect of Helicobacter pylori on the expressions of SOX2, CDX2, MUC2, MUC5AC, MUC6, TFF1, TFF2, and TFF3 mRNAs in human gastric carcinoma cells.

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