武藤 弘行

Background SOX9 is a marker for stem cells in the intestine and overexpression of SOX9 is found in some types of cancer. However, the expression of SOX9 in normal stomach, precancerous intestinal metaplasia, and gastric carcinoma has not yet been clarified. This study aimed to investigate SOX9 expression in the corpus and pyloric regions of the normal human stomach, premalignant intestinal metaplasia, and gastric carcinoma by using immunohistochemistry. Methods We evaluated SOX9 expression in 46 clinical samples (early gastric well-differentiated adenocarcinoma including surrounding intestinal metaplasia) resected under esophagogastroduodenoscopy. Results A small amount of SOX9 was expressed in the neck/isthmus of the corpus region and SOX9 expression was predominantly restricted to the neck/isthmus of the pyloric region in normal human stomach. In the intestinal metaplastic mucosa, SOX9- and PCNA-positive cells were located at the base of the intestinal metaplastic mucosa. Almost all of the gastric carcinoma cells expressed SOX9. Conclusion SOX9 is expressed in intestinal metaplasia and gastric carcinoma in humans.

Gene expression in the early stage of the transition to intestinal metaplasia in human gastric mucosa has not been determined. In this study, we investigated the temporal relationship between cell lineage changes and intestine-specific gene expression in the process leading to intestinal metaplasia, using Cdx2-transgenic mice. Cellular phenotypes were analyzed by immunohistochemistry and were compared with the gene expression profiles of cell lineage markers by real-time polymerase chain reaction. Up to postnatal day (PD) 20, the gastric mucosae of Cdx2-transgenic mice were histologically similar to those of their normal littermates. However, at approximately PD 20, we observed the sporadic appearance of glands in which all the epithelial cells expressed Cdx2 (Cdx2-diffuse positive glands). In the Cdx2-diffuse positive glands, parietal cells had disappeared, the proliferating zone had moved from the isthmus to the base, and absorptive cells and goblet cells were recognized. In contrast, the surrounding mucosa retained the phenotype of the gastric gland in which only some of the epithelial cells expressed Cdx2. During PDs 30 and 40, the entire fundic mucosa changed to transdifferentiated mucosa that was a composite of intestinal metaplasia and spasmolytic polypeptide-expressing metaplasia. An increase in the expression of intestine-specific genes, with a reciprocal decrease in gastric-specific gene expression, began much earlier than the emergence of Cdx2-diffuse positive glands. A dramatic increase in intestine-specific gene expression precedes the morphological appearance of intestinal metaplasia and spasmolytic polypeptide-expressing metaplasia.

Sox2 is closely related to the gastric phenotype. Sox2 plays a pivotal role in gastric epithelial differentiation in the adult. Sox2 expression is reduced in Helicobacter pylori-associated intestinal metaplastic change of the gastric epithelium. The gastric mucosa is replaced by intestinal metaplastic mucosa in the stomach of caudal type homeobox 2 (Cdx2)-transgenic mice. The aim of this study was to use Cdx2-transgenic mice to investigate: (i) Sox2 expression in the intestinal metaplastic mucosa of the Cdx2-transgenic mouse stomach; and (ii) the relationship between Sox2 and Cdx2. Quantitative real-time PCR was performed to determine Sox2, Cdx2, Muc5Ac, and alkaline phosphatase mRNA expression levels and single- or double-label immunohistochemistry was used to evaluate the localization of Sox2, Cdx2, gastric mucin and alkaline phosphatase activity. We determined that Sox2 mRNA in the intestinal metaplastic mucosa of the Cdx2-transgenic mouse stomach was expressed 3.5-fold compared to the normal mouse stomach. Immunohistochemical analysis showed that the same cells in the intestinal metaplastic mucosa expressed both Cdx2 and Sox2. Gastric mucin was not expressed while alkaline phosphatase activity was recognized in the intestinal metaplastic mucosa in spite of the Sox2 expression. Cdx2 increased the transcriptional activity of the Sox2 gene, and Sox2 increased the transcriptional activity of the Muc5Ac gene, which was reduced by cotransfecion of Cdx2 together with Sox2 in the human gastric carcinoma cell line AGS. In conclusion, Sox2 expression is maintained while gastric phenotype is completely lost in the intestinal metaplastic mucosa of Cdx2-transgenic mouse stomach. (C) 2010 International Society of Differentiation. Published by Elsevier Ltd. All rights reserved.

Objective. Cdx2 is expressed in human intestinal metaplastic mucosa and induces intestinal metaplastic mucosa in Cdx2-transgenic mouse stomach. Claudin-2 is a structural component of tight junctions in the intestine and Cdx2 activates the Claudin-2 promoter in the human intestinal epithelial cell line Caco-2. Our aim is to evaluate the expression of Claudin-2 in intestinal metaplastic mucosa of Cdx2-transgenic mouse stomach. Material and methods. The Claudin-2 expression in the normal gastric mucosa and normal intestinal mucosa of wild type mice and the intestinal metaplastic mucosa of Cdx2-transgenic mice was analyzed by immunohistochemistry, Western blotting and quantitative real-time PCR (qRT-PCR). Results. Claudin-2 was expressed in the base of the glands in intestine and intestinal metaplasia while it was not expressed in the body of stomach. Claudin-2 expression was found in the antrum of stomach, while it was weaker than that in the intestine and the intestinal metaplasia. Claudin-2 was also detected in intestinal metaplasia, colon and ileum by both Western blotting and qRT-PCR while it was not detected in gastric body. Conclusion. These results suggest that Cdx2 plays an important role in the expression of Claudin-2 in vivo.

Helicobacter pylori (H. pylori) stimulates secretion of monocyte chemoattractant protein 1 (MCP-1) from gastric mucosa. Monocyte chemoattractant protein-1 (MCP-1) expression and macrophage infiltration are recognized in human gastric carcinoma. We have previously generated Cdx2-transgenic mice as model mice for intestinal metaplasia. Both chronic H. pylori-associated gastritis and Cdx2-transgenic mouse stomach develop intestinal metaplasia and finally gastric carcinoma. In this study we have directed our attention to MCP-1 expression in the intestinal metaplastic mucosa and the gastric carcinoma of Cdx2-transgenic mouse stomach. Quantitative real-time PCR was performed to determine MCP-1 and transforming growth factor-beta 1 (TGF-beta 1) mRNA expression levels and single- or double-label immunohistochemistry was used to evaluate the localization of MCP-1, TGF-beta type I receptor, and alpha-smooth muscle actin (alpha SMA). We determined that MCP-1 mRNA dramatically increased in the intestinal metaplastic mucosa and the gastric carcinoma of Cdx2-transgenic mouse stomach, compared with normal mouse stomach. Both MCP-1 and TGF-beta type I receptor were co-expressed in the alpha SMA-positive myofibroblasts of intestinal metaplastic mucosa and gastric carcinoma. Exogenous application of TGF-beta 1 increased MCP-1 mRNA expression levels in the intestinal metaplastic tissue. Furthermore, TGF-beta 1 was overexpressed and macrophage was strongly infiltrated in the gastric carcinoma. In conclusion, MCP-1 expression, which was stimulated by TGF-beta 1, was recognized in the TGF-beta type I receptor-expressing myofibroblasts of the intestinal metaplastic mucosa and the gastric carcinoma of Cdx2-transgenic mouse stomach. The present results suggest that intestinal metaplasia and gastric carcinoma themselves induce MCP-1 expression independently of H. pylori infection. (Cancer Sci 2010).