| Project/Area Number |
13670015
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
General anatomy (including Histology/Embryology)
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| Research Institution | HIROSHIMA UNIVERSITY |
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Principal Investigator |
KATAOKA Katsuko Hiroshima University, Graduate School of Biomedical Sciences, Professor, 大学院・医歯学総合研究科, 教授 (30034002)
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| Co-Investigator(Kenkyū-buntansha) |
SUZAKI Etsuko Hiroshima University, Graduate School of Biomedical Sciences, Instructor, 大学院・医歯薬学総合研究科, 講師 (10274052)
SAITO Akira Hiroshima University, Graduate School of Biomedical Sciences, Research Associate, 大学院・医歯薬学総合研究科, 助手 (80346486)
野村 隆士 広島大学, 医学部, 助手 (20325161)
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| Project Period (FY) |
2001 – 2004
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| Project Status |
Completed (Fiscal Year 2004)
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| Budget Amount *help |
¥2,900,000 (Direct Cost: ¥2,900,000)
Fiscal Year 2004: ¥500,000 (Direct Cost: ¥500,000)
Fiscal Year 2003: ¥900,000 (Direct Cost: ¥900,000)
Fiscal Year 2002: ¥900,000 (Direct Cost: ¥900,000)
Fiscal Year 2001: ¥600,000 (Direct Cost: ¥600,000)
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| Keywords | gastrointestinal mucosa / cytoskeleton / intercellular junction / microtubule / epithelial cell / cell polarity / γ-tubulin / HT-29 cells / MTOC / HT-29 / 免疫組織化学 / カドヘリン / 閉鎖帯 / γ-チュブリン / 組織形成 / 腸上皮 / tubulin / 電子顕微鏡 / 免疫蛍光法 / 腸上皮細胞 / 上皮性培養細胞株 / 結合組織細胞 / α-tubulin / 免疫蛍光染色 |
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| Research Abstract |
Microtubules and Golgi apparatus were histochemically demonstrated in the mouse duodenal mucosa and examined by confocal laser scanning and electron microscopy. In free cells, microtubules radiated from MTOC where γ-tubulin was located. In enterocytes, microtubules ran parallel to the cell axis and especially abundant in the supranuclear area. γ-tubulin-immunoreactivity was associated with Golgi apparatus, which suggests the role of the apparatus on microtubule nucleation. The monoclonal antibody G9, raised against east γ-tubulin, bound along the cytoplasmic aspect of the three-cell-meeting junction. Since protein related to the three-cell-meeting junction has not been reported, we tried proteome analysis of G9-immunoreactive protein. A suggested candidate was Noxo 1(NADPH oxidase organizer 1). However, the relation between Noxo 1 and cell junction has been unknown.
In another experiment, we examined epithelial cell polarity formation in a cultured colon cancer cell line, HT-29. When we
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seeded microtubule-depolymerized HT-29 cells, newly formed microtubules ran perpendicular to the matrix. Then, the cadherin-mediated intercellular junction and stress fibers appeared. γ-tubulin located randomly at first, but gradually relocated near the free surface of the cell. The cells gradually piled up in normal culture medium. Tight junction was not formed, and MUC1(an apical surface glycoprotein) was not expressed. When we added butyrate into the medium, the cell gradually acquired epithelial cell polarity. First, an intracellular cavity lined with MUC1-immunoreactivity and F-actin were formed. Then it fused to cell membrane to bean intercellular cavity sealed with the tight junction. The γ-tubulin spot was seen by the intercellular cavity, from which microtubules were radiated. Finally, the intercellular cavity enlarged to become apical cell surface with brush border and MUC1-immunoreactivity, and the cells arranged to form a simple cuboidal epithelium. This process mimics the villi and crypt formation in prenatal development of the gastrointestinal mucosa. Less
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