Abstract
Abstract (a) Introduction: Tumor necrosis factor-α (TNF-α)-inducing protein (Tipα) of H. pylori acts as a carcinogenic factor in the gastric epithelium: it binds to surface nucleolin on human and mouse gastric cancer cell lines, resulting in cancer progression. We studied various phenotypes of epithelial-mesenchymal transition (EMT) to examine the function of surface nucleolin in gastric cancer cell line. (b) Experimental procedures : 1) Induction of cell migration and elongation: After human gastric cancer cell line MKN-1 was transfected with nucleolin-targeted siRNAs (siRNA-n), cell migration and elongation induced by Tipα were determined using Transwell assay and cell morphology. 2) Formation of filopodia in MKN-1 cells was determined by staining filamentous actin with AF488-phalloidin. 3) Phosphorylation of 89 cancer-related proteins was determined using Cancer Signaling Phosphoantibody Array. 4) Expression of EMT markers was examined with Western blotting. 5) Cell stiffness was determined from Young's modulus using atomic force microscope (AFM). (c) A summary of the new data: 1) The transfected cells with siRNA-n showed a reduced amount of surface nucleolin in membrane fraction, but not in cytosol or nuclear fractions. Knockdown of nucleolin inhibited cell migration and elongation induced by Tipα. 2) Tipα induced the formation of filopodia in MKN-1 cells, which suggests the invasive morphological changes. 3) Tipα enhanced the phosphorylation of 11 cancer-related proteins in serine, threonine and tyrosine residues, indicating activation of MEK-ERK signal cascade. 4) Tipα induced the expression of vimentin, a significant marker of EMT, even though down-regulation of E-cadherin was not found in MKN-1 cells. 5) Tipα reduced the Young's modulus of MKN-1 cells as determined by AFM: The results showed that lower cell stiffness and increased cell motility were induced by Tipα. (d) Conclusions: This is the first report that Tipα is a new inducer of EMT in human gastric cancer cells, and that EMT was induced through the binding of Tipα to surface nucleolin. Citation Format: Hirota Fujiki, Tatsuro Watanabe, Atsushi Takahashi, Kaori Suzuki, Miki Kurusu-Kanno, Kensei Yamaguchi, Masami Suganuma. Induction of epithelial-mesenchymal transition (EMT) in human gastric cancer cell lines induced by Tipα of H. pylori mediated through surface nucleolin. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 1143. doi:10.1158/1538-7445.AM2014-1143
Published Version
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