リアルタイムイメージングを用いた初代大腸上皮培養細胞の増殖・分化機構解析

2011 年度 研究成果報告書

• 研究課題/領域番号: 21590803
• 研究種目: 基盤研究(C)
• 配分区分: 補助金
• 応募区分: 一般
• 研究分野: 消化器内科学
• 研究機関: 東京医科歯科大学
• 研究代表者: 中村 哲也 東京医科歯科大学, 大学院・医歯学総合研究科, 寄附講座教員 (70265809)
• 研究分担者: 渡辺 守 東京医科歯科大学, 医歯学総合研究科, 教授 (10175127)
• 研究期間 (年度): 2009 – 2011
• キーワード: 消化管上皮幹細胞 / ライブセルイメージング / 初代培養技術 / 細胞移植治療

研究概要

新規の培養技術確立により、正常大腸上皮の増殖・分化を可視化するシステムを構築した。この方法で得られた培養大腸上皮細胞は上皮傷害を惹起した大腸の上皮欠損部位に生着したのみならず、機能的大腸上皮を再生した。重要なことに、1個のLgr5+大腸幹細胞から培養し増やした細胞群の移植でも同様の上皮再生を確認できた。本培養技術とイメージング技術は、大腸上皮幹細胞研究の新しいツールとなることが期待される。また、体外培養幹細胞で大腸上皮再生が可能であることを示した本成果は、新しい再生医療の可能性を示すものと考えた。

研究成果

• [雑誌論文] Functional engraftment of colon epithelium expanded in vitro from a single adult Lgr5+stem cell (2012)
  • 著者名/発表者名: Yui S, Nakamura T, Sato T, Nemoto Y, Mizutani T, Zheng X, Ichinose S, Nagaishi T, Okamoto R, Tsuchiya K, Clevers H, and Watanabe M
  • 雑誌名: Nature Medicine
  • DOI: doi:10.1038/nm.2695
• [雑誌論文] Real-time analysis of P-glycoprotein-mediated drug transport across primary intestinal epithelium three-dimensionally cultured in vitro (2012)
  • 著者名/発表者名: Mizutani T, Nakamura T, Morikawa R, Fukuda M, Mochizuki W, Yamauchi Y, Nozaki K, Yui S, Nemoto Y, Nagaishi T, Okamoto R, Tsuchiya K, and Watanabe M
  • 雑誌名: Biochem Biophys Res Commun 巻: 419(2) ページ: 238-243
• [雑誌論文] Gene bookmarking accelerates the kinetics of post-mitotic transcriptional activation (2011)
  • 著者名/発表者名: Zhao R, Nakamura T, Fu Y, Lazar Z, and Spector DL
  • 雑誌名: Nature Cell Biology 巻: 13(11) ページ: 1295-1304
• [雑誌論文] Suppression of Hath1 gene expression directly regulated by Hes1 via Notch signaling is associated with goblet cell depletion in ulcerative colitis (2011)
  • 著者名/発表者名: Zheng X, Tsuchiya K, Okamoto R, Iwasaki M, Kano Y, Sakamoto N, Nakamura T, Watanabe M
  • 雑誌名: Inflamm Bowel Dis 巻: 17(11) ページ: 2251-2260
• [雑誌論文] Longitudinal cell formation in the entire human small intestine is correlated with the localization of Hath1 and Klf4 (2011)
  • 著者名/発表者名: Iwasaki M, Tsuchiya K, Okamoto R, Zheng X, Kano Y, Okamoto E, Okada E, Araki A, Suzuki S, Sakamoto N, Kitagaki K, Akashi T, Eishi Y, Nakamura T, Watanabe M
  • 雑誌名: J Gastroenterol 巻: 46(2) ページ: 191-201
• [雑誌論文] Nuclear organization and dynamics of 7SK RNA in regulating gene expression (2010)
  • 著者名/発表者名: Prasanth KV, Camiolo M, Chan G, Tripathi V, Denis L, Nakamura T, Hubner M, and Spector DL
  • 雑誌名: Mol BiolCell 巻: 21(23) ページ: 4184-4196
• [雑誌論文] A long nuclear retained non-coding RNA regulates synaptogenesis by modulating gene expression (2010)
  • 著者名/発表者名: Bernard D, Prasanth KV, Tripathi V, Colasse S, Nakamura T, Xuan Z, Zhang MQ, Sedel F, Jourdren L, Coulpier F, Triller A, Spector DL, and Bessis A
  • 雑誌名: EMBO J 巻: 29(18) ページ: 3082-3093
• [雑誌論文] Delta-like 1 expression promotes goblet cell differentiation in Notch-inactivated human colonic epithelial cells (2010)
  • 著者名/発表者名: Akiyama J, Okamoto R, Iwasaki M, Zheng X, Yui S, Tsuchiya K, Nakamura T, and Watanabe M
  • 雑誌名: Biochem Biophys Res Commun 巻: 393(4) ページ: 662-667
• [雑誌論文] Signaling pathway via TNF-a/NF-kB in intestinal epithelial cells may be directly involved in colitis-associated carcinogenesis (2009)
  • 著者名/発表者名: Onizawa M, Nagaishi T, Kanai T, Nagano KI, Oshima S, Nemoto Y, Yoshioka A, Totsuka T, Okamoto R, Nakamura T, Sakamoto N, Tsuchiya K, Aoki K, Ohya K, Yagita H, Watanabe M
  • 雑誌名: Am J Physiol Gastrointest Liver Physiol 巻: 296(4) ページ: 850-859
• [雑誌論文] Requirement of Notch activation during regeneration of the intestinal epithelia (2009)
  • 著者名/発表者名: Okamoto R, Tsuchiya K, Nemoto Y, Akiyama J, Nakamura T, Kanai T, Watanabe M
  • 雑誌名: Am J Physiol Gastrointest Liver Physiol 巻: 296(1) ページ: 23-35
• [学会発表] Epithelial regeneration by cultured colonic cells expanded from a single adult LGR5+stem cell (2012)
  • 著者名/発表者名: Tetsuya Nakamura(Invited Speaker)
  • 学会等名: The 6th Korea-JapanIBD Symposium
  • 発表場所: Tokyo, Japan
  • 年月日: 2012-01-28
• [学会発表] Regeneration of Damaged Colonic Tissue by Transplantation of Colonic Epithelial Stem Cells Maintained and Expanded In Vitro (2011)
  • 著者名/発表者名: Shiro Yui, Tetsuya Nakamura, Yasuhiro Nemoto, Tomohiro Mizutani, Xiu Zheng, Shizuko Ichinose, Takashi Nagaishi, Ryuichi Okamoto, Kiichiro Tsuchiya, Toshiro Sato, Hans Clevers, and Mamoru Watanabe
  • 学会等名: Basic Science Plenary Symposium, Digestive DiseaseWeek 2011
  • 発表場所: Chicago, USA
  • 年月日: 20110507-10
• [学会発表] Regeneration of damaged colon epithelium by transplanted colon Lgr5+stem cells maintained and expanded in vitro (2011)
  • 著者名/発表者名: Shiro Yui, Tetsuya Nakamura, Toshiro Sato, Yasuhiro Nemoto, Tomohiro Mizutani, Xiu Zheng, Shizuko Ichinose, Takashi Nagaishi, Ryuichi Okamoto, Kiichiro Tsuchiya, Hans Clevers, and Mamoru Watanabe
  • 学会等名: The5^<th> Japan & US Collaboration Conference in Gastroenterology
  • 発表場所: Tokyo, Japan
  • 年月日: 2011-11-10
• [学会発表] Flagellin via TLR5 on basolateral membrane of primary intestinal epithelial cells(IEC) shows the role of IEC in the response to bacteria. (2011)
  • 著者名/発表者名: Tsuchiya K, Zheng X, Kano Y, Okamoto R, Nakamura T, Watanabe M
  • 学会等名: United European Gastroenterology Week 2011
  • 発表場所: Stockholm, Sweden
  • 年月日: 2011-10-26
• [学会発表] Regeneration of damaged colonic tissue by transplanted colonic epithelial stem cells maintained and expanded in vitro (2011)
  • 著者名/発表者名: Yui S, Nakamura T, Nemoto Y, Mizutani T, Zheng X, Nagaishi T, Tsuchiya K, Watanabe M, Okamoto R, Ichinose S, Sato T, Clevers H
  • 学会等名: GI Research Academy 2011
  • 発表場所: Kyoto, Japan
  • 年月日: 2011-06-17
• [学会発表] A long-term, mesenchyme-free in vitro culture system of colonic epithelial cells retaining stem cell compartment with proliferation/differentiation properties (2010)
  • 著者名/発表者名: Tetsuya Nakamura, Shiro Yui, Tomohiro Mizutani, Yasuhiro Nemoto, Ryuichi Okamoto, Kiichiro Tsuchiya, Shizuko Ichinose, and Mamoru Watanabe
  • 学会等名: Digestive Disease Week 2010
  • 発表場所: New Orleans, USA
  • 年月日: 20100501-05
• [学会発表] A well-defined culture system for colonic epithelial cells thet allows efficient enrichment of LGR5+stem cells. (2010)
  • 著者名/発表者名: Yui S, Nakamura T, Nemoto Y, Mizutani T, Zheng X, Ichinose S, Nagaishi T, Okamoto R, Tsuchiya K, Watanabe M
  • 学会等名: The 1st JSGE International Topic Conference-Stem Cells in Digestive Organs-
  • 発表場所: Kamakura, Japan
  • 年月日: 2010-09-26
• [学会発表] Up-reguration of DELTA1 expression is required for goblet cell differentiation in human intestinal epithelial cells (2010)
  • 著者名/発表者名: Akiyama J, Okamoto R, Tsuchiya K, Nakamura T, Watanabe M
  • 学会等名: The 1st JSGE International Topic Conference-Stem Cells in Digestive Organs-
  • 発表場所: Kamakura, Japan
  • 年月日: 2010-09-26
• [学会発表] HES1 via notch signaling directly suppresses ATOH1/HATH1 gene transcription, resulting in the undifferentiated state of intestinal epithelial cells. (2010)
  • 著者名/発表者名: Zheng X, Tsuchiya K, Okamoto R, Okamoto E, Iwasaki M, Kano Yoshihito, Nakamura T, Watanabe M
  • 学会等名: The 1st JSGE International Topic Conference-Stem Cells in Digestive Organs-
  • 発表場所: Kamakura, Japan
  • 年月日: 2010-09-26
• [学会発表] TNF-αand notch signaling synergistically up-regulate OLFM4 expression in human intestinal epithelial cells (2010)
  • 著者名/発表者名: Murano T, Okamoto R, Shimizu H, Akiyama J, Tsuchiya K, Nakamura T, Watanabe M
  • 学会等名: The 1st JSGE International Topic Conference-Stem Cells in Digestive Organs-
  • 発表場所: Kamakura, Japan
  • 年月日: 2010-09-26
• [学会発表] A long-term, fully-deffined culture system for colonic epithelial cells that alows efficient expansion of stem cell compartment (2010)
  • 著者名/発表者名: Tetsuya Nakamura(Invited speaker), Watanabe M
  • 学会等名: The 1st JSGE International Topic Conference-Stem Cells in Digestive Organs-
  • 発表場所: Kamakura, Japan
  • 年月日: 2010-09-25
• [学会発表] Hes1 via Notch signaling directly suppresses Atoh1/Hath1 gene transcription, resulting in the goblet cell depletion of Ulcerative Colitis (2010)
  • 著者名/発表者名: Zheng X, Tsuchiya K, Okamoto R, Iwasaki M, Kano Y, Nakamura T, Watanabe M
  • 学会等名: Digestive Disease Week 2010
  • 発表場所: New Orleans, USA
  • 年月日: 2010-05-03
• [学会発表] GSK3 inhibitor induces the intestinal differen tiation by the proteinstabilization of Atoh1 (2009)
  • 著者名/発表者名: Tsuchiya K, Okamoto R, Nakamura T, Watanabe M
  • 学会等名: Digestive Disease Week 2009
  • 発表場所: Chicago, USA
  • 年月日: 2009-06-02
• [産業財産権] 大腸上皮幹細胞の単離・培養技術とこれを用いた大腸上皮移植技術 (2011)
  • 発明者名: 渡辺守・中村哲也
  • 権利者名: 国立大学法人東京医科歯科大学
  • 産業財産権番号: 特願、2011-236469
  • 出願年月日: 2011-10-27