detection of marker groups indicating differentiating and regenerating stages of digestive cells using protein chips

2006 Fiscal Year Final Research Report Summary

• Project/Area Number: 16591340
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Single-year Grants
• Section: 一般
• Research Field: Digestive surgery
• Research Institution: Kyoto Prefectural University of Medicine
• Principal Investigator: HAGIWARA Akeo Kyoto Prefectural University of Medicine, Graduate School of Medicine, Associate Professor, 医学研究科, 准教授 (90198648)
• Co-Investigator(Kenkyū-buntansha): SAKAKURA Chouhei Kyoto Prefectural University of Medicine, Graduate School of Medicine, Lecturer, 医学研究科, 講師 (10285257) ; NAITO Yuji Kyoto Prefectural University of Medicine, School of Medicine, Associate Professor, 医学部, 准教授(寄附講座) (00305575) ; NAKAMURA Tatsuo Kyoto University, Institute of Frontier Medical Sciences, Associate Professor, 再生医科学研究所, 准教授 (70227908) ; YONEMURA Yutaka Shizuoka Cancer Center, Deputy Director, 副院長 (20167042)
• Project Period (FY): 2004 – 2006
• Keywords: mucosa of the digestive tract / regeneration / differentiation / marker / tissue-interaction / endocrine cells of the digestive organs

Research Abstract

Proliferation and differentiation of the small-intestinal epithelium depends in part on intestinal subepithelial myofibroblasts. Although there have been many culture studies of intestinal epithelial-mesenchymal interaction, the observation periods have rarely exceeded ten days. This study investigated the influence of intestinal subepithelial myofibroblasts on proliferation and differentiation of intestinal epithelial cells with long-term observation for three weeks, using a three-dimensional coculture system. Cultured rat intestinal subepithelial myofibroblasts, obtained from the duodenum, were embedded in collagen gel, and cells from the rat intestinal epithelial cell line IEC-6 were seeded onto it. Histologic sections of the cell-embedded gels were made, and histochemical and immunohistochemical examinations were carried out in conjunction with expression analysis of the pdx-1 transcription factor in IEC-6 cells. IEC-6 cells showed increased proliferation and displayed characteristic endocrine features when cocultured with rat intestinal subepithelial myofibroblasts, arranging themselves into multilayer structures and becoming cuboidal, with abundant cytoplasm and oval nuclei. Some IEC-6 cells were immunohistochemically positive for chromogranin A and glicentin. They also expressed the pdx-1 transcription factor at both the mRNA and protein levels. The number and percentage of chromogranin A-positive cells increased with culture time, whereas no increase was observed in cells cultured without rat intestinal subepithelial myofibroblasts. Intestinal subepithelial myofibroblasts influence both proliferation and differentiation, particularly enteroendocrine differentiation, of epithelial cells in the intestinal mucosa. This three-dimensional coculture system provides a useful tool for long-term study of epithelial proliferation and differentiation influenced by intestinal subepithelial myofibroblasts.