腸管上皮－間質ニッチの包括的理解と自己補完的組織培養技術の確立

2017 Fiscal Year Annual Research Report

• Project/Area Number: 15J00981
• Research Institution: Keio University
• Principal Investigator: 杉本 真也 慶應義塾大学, 医学研究科, 特別研究員(DC1)
• Project Period (FY): 2015-04-24 – 2018-03-31
• Keywords: オルガノイド / 腸管上皮幹細胞 / 炎症性腸疾患 / 粘膜再生治療 / 間質細胞

Outline of Annual Research Achievements

本年度は，前年度までに確立した無血清Wnt3a-Afaminを含む移植細胞培養系と，正常ヒト大腸オルガノイドを免疫不全マウスの大腸へ同所異種移植するマウスモデルを用いて，これまで実験モデルの欠如から困難であったin vivoでのヒト大腸幹細胞の時空間ダイナミクスの検討を行った．また，大動物における内視鏡的なオルガノイド自家移植系の移植手技最適化を行い，安全性を確認した．なお，間質細胞と上皮細胞の共培養系は，がん細胞への間質細胞からのニッチ因子供給を検討する実験にも応用できた（Seino et al. Cell Stem Cell 2018）．
大腸がんの遺伝学的細胞系譜解析において確立したCRISPR-Cas9システムによるLGR5遺伝子へのCreERT2のノックインとCre依存的に発現するRainbowレポーターを組み込んだオルガノイドを作成する手法を（Shimokawa et al. Nature 2017），正常大腸オルガノイドへ応用することでヒト腸管上皮のLGR5陽性細胞から産生される子孫細胞のクローンを異なる色で標識することが可能となった．このオルガノイドをマウス腸管へ同所異種移植することにより，ヒト大腸上皮細胞の遺伝学的細胞系譜解析が可能となり，ヒトLGR5陽性幹細胞の自己複製能および多分化能をin vivoにおいてはじめて実証することに成功した（Sugimoto et al. Cell Stem Cell 2018）．これにより，ヒト腸管上皮オルガノイドの移植細胞としての可能性が示され，in vivoでのヒト大腸幹細胞の機能的挙動の観察が可能となったのみならず，この移植モデルと遺伝子改変オルガノイドを組み合わせることで，腸内におけるヒトの腫瘍形成に関する病態理解を進展させる可能性をもつ有用なリソースになるものと考えられる．

Research Progress Status

29年度が最終年度であるため、記入しない。

Strategy for Future Research Activity

29年度が最終年度であるため、記入しない。

Research Products

• [Journal Article] Reconstruction of the Human Colon Epithelium In Vivo (2018)
  • Author(s): Shinya Sugimoto, Yuki Ohta, Masayuki Fujii, Mami Matano, Mariko Shimokawa, Kosaku Nanki, Shoichi Date, Shingo Nishikori, Yoshihiro Nakazato, Tetsuya Nakamura, Takanori Kanai, Toshiro Sato
  • Journal Title: Cell Stem Cell Volume: 22 Pages: 171-176.e5
  • DOI: 10.1016/j.stem.2017.11.012
  • Peer Reviewed
• [Journal Article] Human Pancreatic Tumor Organoids Reveal Loss of Stem Cell Niche Factor Dependence during Disease Progression (2018)
  • Author(s): Takashi Seino, Shintaro Kawasaki, Mariko Shimokawa, Hiroki Tamagawa, Kohta Toshimitsu, Masayuki Fujii, Yuki Ohta, Mami Matano, Kosaku Nanki, Kenta Kawasaki, Sirirat Takahashi, Shinya Sugimoto, Eisuke Iwasaki, Junichi Takagi, Takao Itoi, Minoru Kitago, Yuko Kitagawa, Takanori Kanai, Toshiro Sato
  • Journal Title: Cell Stem Cell Volume: 22 Pages: 454-467.e6
  • DOI: 10.1016/j.stem.2017.12.009
  • Peer Reviewed
• [Journal Article] Efficacy of Indigo Naturalis in a Multicenter Randomized Controlled Trial of Patients with Ulcerative Colitis (2018)
  • Author(s): Makoto Naganuma, Shinya Sugimoto, Keiichi Mitsuyama, Taku Kobayashi, Naoki Yoshimura, Hidehisa Ohi, Shinji Tanaka, Akira Andoh, Naoki Ohmiya, Keiichiro Saigusa, Takayuki Yamamoto, Yuichi Morohoshi, Hitoshi Ichikawa, Katsuyoshi Matsuoka, et al.
  • Journal Title: Gastroenterology Volume: 154 Pages: 935-947
  • DOI: 10.1053/j.gastro.2017.11.024
  • Peer Reviewed / Open Access
• [Journal Article] Fecal microbiota transplantation for recurrent Clostridium difficile infection in a patient with ulcerative colitis (2018)
  • Author(s): Kosaku Nanki, Shinta Mizuno, Katsuyoshi Matsuoka, Keiko Ono, Shinya Sugimoto, Hiroki Kiyohara, Mari Arai, Moeko Nakashima, Kozue Takeshita, Keiichiro Saigusa, Mitsutoshi Senoh, Tadashi Fukuda, Makoto Naganuma, Haru Kato, Wataru Suda, Masahira Hattori, Takanori Kanai
  • Journal Title: Intestinal Research Volume: 16 Pages: 142-146
  • DOI: 10.5217/ir.2018.16.1.142
  • Peer Reviewed / Open Access
• [Journal Article] Visualization and targeting of LGR5+ human colon cancer stem cells (2017)
  • Author(s): Mariko Shimokawa, Yuki Ohta, Shingo Nishikori, Mami Matano, Ai Takano, Masayuki Fujii, Shoichi Date, Shinya Sugimoto, Takanori Kanai, Toshiro Sato
  • Journal Title: Nature Volume: 545 Pages: 187-192
  • DOI: 10.1038/nature22081
  • Peer Reviewed
• [Journal Article] Endocytoscopy can be used to assess histological healing in ulcerative colitis patients with clinical and endoscopic remission (2017)
  • Author(s): Yoshihiro Nakazato, Makoto Naganuma, Shinya Sugimoto, Rieko Bessho, Mari Arai, Hiroki Kiyohara, Keiko Ono, Kosaku Nanki, Makoto Mutaguchi, Shinta Mizuno, Taku Kobayashi, Naoki Hosoe, Masayuki Shimoda, Takayuki Abe, Nagamu Inoue, Haruhiko Ogata, Yasushi Iwao, Takanori Kanai
  • Journal Title: Endoscopy Volume: 49 Pages: 560-563
  • DOI: 10.1055/s-0043-106556
  • Peer Reviewed
• [Journal Article] The risk factor of clinical relapse in ulcerative colitis patients with low dose 5-aminosalicylic acid as maintenance therapy: A report from the IBD registry (2017)
  • Author(s): Tomohiro Fukuda, Makoto Naganuma, Shinya Sugimoto, Kosaku Nanki, Shinta Mizuno, Makoto Mutaguchi, Yoshihiro Nakazato, Nagamu Inoue, Haruhiko Ogata, Yasushi Iwao, Takanori Kanai
  • Journal Title: PLoS One Volume: 12 Pages: e0187737
  • DOI: 10.1371/journal.pone.0187737
  • Peer Reviewed / Open Access
• [Presentation] オルガノイドを用いた細胞系譜解析によるヒト正常大腸幹細胞挙動の可視化 (2018)
  • Author(s): 佐藤 俊朗, 杉本 真也, 太田 悠木, 下川 真理子
  • Organizer: 第17回日本再生医療学会総会
  • Invited
• [Presentation] Efficacy of therapeutic intervention for ulcerative colitis patients with the Mayo endoscopic score of 1 (2018)
  • Author(s): Tomohiro Fukuda, Makoto Naganuma, Shinya Sugimoto, Keiko Ono, Kosaku Nanki, Shinta Mizuno, Kayoko Kimura, Makoto Mutaguchi, Kaoru Takabayashi, Nagamu Inoue, Haruhiko Ogata, Yasushi Iwao, Takanori Kanai
  • Organizer: 13th Congress of European Crohn’s and Colitis Organization (ECCO)
  • Int'l Joint Research
• [Presentation] Indigo naturalis induces mucosal healing in patients with ulcerative colitis: A multicentre randomised controlled trial (2018)
  • Author(s): Makoto Naganuma, Shinya Sugimoto, Keiichi Mitsuyama, Taku Kobayashi, Naoki Yoshimura, Hidehisa Ohi, Katsuyoshi Matsuoka, Tadakazu Hisamatsu, Kenji Watanabe, Takayuki Abe, Mamoru Watanabe, Toshifumi Hibi, Yasuo Suzuki, Takanori Kanai, INDIGO study group
  • Organizer: 13th Congress of European Crohn’s and Colitis Organization (ECCO)
  • Int'l Joint Research
• [Presentation] The impact of establishment of organoid-based orthotopic xenograft model on intestinal stem cell research (2017)
  • Author(s): Shinya Sugimoto, Yuki Ohta, Takanori Kanai, Toshiro Sato
  • Organizer: The 54th Annual Meeting of the Japan Society for Mucosal Immunology
  • Invited
• [Presentation] 寛解期潰瘍性大腸炎における腸内・口腔内細菌叢の構成と予後との関連の検討 (2017)
  • Author(s): 大野 恵子, 水野 慎大, 清原 裕貴, 杉本 真也, 南木 康作, 牟田口 真, 中里 圭宏, 筋野 智久, 長沼 誠, 金井 隆典
  • Organizer: 第54回日本消化器免疫学会総会
• [Presentation] 潰瘍性大腸炎症例の散発性大腸腫瘍の臨床像 (2017)
  • Author(s): 牟田口 真, 長沼 誠, 杉本 真也, 南木 康作, 水野 慎大, 緒方 晴彦, 岩男 泰, 金井 隆典
  • Organizer: 第94回消化器内視鏡学会総会（JDDW2017）
• [Presentation] Indigo naturalis is effective for inducing clinical remission and mucosal healing in patients with ulcerative colitis (INDIGO study) (2017)
  • Author(s): Makoto Naganuma, Shinya Sugimoto, Keiichi Mitsuyama, Taku Kobayashi, Naoki Yoshimura, Katsuyoshi Matsuoka, Tadakazu Hisamatsu, Kenji Watanabe, Takayuki Abe, Yasuo Suzuki, Toshifumi Hibi, Takanori Kanai
  • Organizer: UEG Week 2017
  • Int'l Joint Research
• [Presentation] Clinical characteristics in ulcerative colitis patients with colitis associated dysplasia/cancer and sporadic tumor (2017)
  • Author(s): Makoto Mutaguchi, Makoto Naganuma, Yasushi Iwao, Tomohiro Fukuda, Shinya Sugimoto Kosaku Nanki, Shinta Mizuno, Haruhiko Ogata, Takanori Kanai
  • Organizer: UEG Week 2017
  • Int'l Joint Research