内耳幹細胞ホーミング機構を応用した遺伝性難聴への多能性幹細胞治療法の開発

2015 Fiscal Year Annual Research Report

• Project/Area Number: 25462653
• Research Institution: Juntendo University
• Principal Investigator: 神谷 和作 順天堂大学, 医学部, 准教授 (10374159)
• Project Period (FY): 2013-04-01 – 2016-03-31
• Keywords: 遺伝性難聴 / 難聴モデル動物 / コネキシン26 / 幹細胞ホーミング / 内耳細胞治療

Outline of Annual Research Achievements

我々は遺伝性難聴で最も高頻度に発生するコネキシン26遺伝子変異の遺伝子改変モデル動物を開発し、その病態を明らかにするとともに（Kamiya et al., 2014, J Clin Invest）、これらの聴力改善を目指した遺伝性難聴の幹細胞治療法の開発を進めてきた(Kamiya et al., 2007, Am J Pathol)。内耳細胞治療において作製された細胞を蝸牛組織の異常部位へ直接挿入することは蝸牛の構造上困難であり、適切な箇所に幹細胞を導入できる細胞誘導システムが必須であると考えられる。我々は実験的に誘発した蝸牛線維細胞損傷部において、他臓器における幹細胞ホーミング因子とされるMCP1およびSDF1が高発現することを発見した。さらに間葉系幹細胞の培養下でMCP1およびSDF1での前処置を行ったところ、それらの受容体であるCCR2およびCXCR4の発現量が定量的RT-PCRにより有意に上昇することが確認できた。更にフローサイトメトリー分析により間葉系細胞の細胞表面のCCR2およびCXCR4を高発現する細胞数が有意に上昇することも確認できた。同処置により内耳への誘導効率が増強された骨髄間葉系幹細胞を作製することが可能となった。
さらに我々はマウスiPS細胞から分化誘導した内耳前駆細胞においても同様の処置を行ったとことろ、間葉系幹細胞と同様にホーミング因子受容体の発現上昇が確認できた。

Research Products

• [Journal Article] Deformation of the Outer Hair Cells and the Accumulation of Caveolin-2 in Connexin 26 Deficient Mice. (2015)
  • Author(s): Anzai, T., Fukunaga, I., Hatakeyama, K., Fujimoto, A., Kobayashi, K., Nishikawa, A., Aoki, T., Noda, T., Minowa, O., Ikeda, K., and Kamiya, K
  • Journal Title: PloS one Volume: 10 Pages: e0141258
  • DOI: 10.1371/journal.pone.0141258.
  • Peer Reviewed / Open Access / Acknowledgement Compliant
• [Journal Article] Postnatal Development and Maturation of the Vestibular Organ in Dominant-Negative Connexin 26 Transgenic Mouse. (2015)
  • Author(s): Hiroko Okada, Kazusaku Kamiya., Takashi Iizuka and Katsuhisa Ikeda.
  • Journal Title: J Otol Rhinol Volume: S1 Pages: 37-40
  • DOI: 10.4172/2324-8785.S1-009
  • Peer Reviewed / Open Access
• [Journal Article] Differentiation of iPS Cells to Cochlear Cells are Regulated Depending on the Part of Co-cultured Organs. (2015)
  • Author(s): Kazusaku Kamiya, Keiko Karasawa, Kazuma Kobayashi, Asuka Miwa and Katsuhisa Ikeda.
  • Journal Title: J Otol Rhinol Volume: S1 Pages: 34-36
  • DOI: 10.4172/2324-8785.S1-008
  • Peer Reviewed / Open Access
• [Journal Article] Perinatal Gjb2 gene transfer rescues hearing in a mouse model of hereditary deafness. (2015)
  • Author(s): Iizuka, T., Kamiya, K., Gotoh, S., Sugitani, Y., Suzuki, M., Noda, T., Minowa, O., and Ikeda, K.
  • Journal Title: Human molecular genetics Volume: 24 Pages: 3651-3661
  • DOI: 10.1093/hmg/ddv109.
  • Peer Reviewed / Open Access / Acknowledgement Compliant
• [Journal Article] Connexin26 regulates assembly and maintenance of cochlear gap junction macromolecular complex for normal hearing (2015)
  • Author(s): Kazusaku Kamiya, Ichiro Fukunaga, Kaori Hatakeyama and Katsuhisa Ikeda
  • Journal Title: AIP Conference Proceedings Volume: 1703 Pages: 30018;1-3
  • DOI: 10.1063/1.4939333
  • Acknowledgement Compliant
• [Presentation] GJB2変異遺伝性難聴に対する細胞治療・遺伝子治療法の開発 (2016)
  • Author(s): 神谷 和作
  • Organizer: 第15回日本再生医療学会総会
  • Place of Presentation: 大阪国際会議場(大阪市)
  • Year and Date: 2016-03-17 – 2016-03-19
• [Presentation] iPS細胞からのConnexin26発現細胞への分化誘導と細胞間ギャップ結合プラークの構築 (2016)
  • Author(s): 福永一朗、畠山佳欧里、 青木徹、藤本あゆみ、 西川貴菜、美野輪治、 池田勝久、神谷和作
  • Organizer: 第15回日本再生医療学会総会
  • Place of Presentation: 大阪国際会議場(大阪市)
  • Year and Date: 2016-03-17 – 2016-03-19
• [Presentation] コネキシン26欠損マウスからのiPS細胞の樹立 (2016)
  • Author(s): 青木徹、福永一朗、 畠山佳欧里、藤本あゆみ、西川貴菜、美野輪治、 池田勝久、神谷和作
  • Organizer: 第15回日本再生医療学会総会
  • Place of Presentation: 大阪国際会議場(大阪市)
  • Year and Date: 2016-03-17 – 2016-03-19
• [Presentation] 内耳フィーダー細胞を用いたiPS細胞の分化誘導によるCx26ギャップ結合形成細胞の樹立 (2016)
  • Author(s): 藤本あゆみ、福永一朗、 畠山佳欧里、青木徹、 西川貴菜、池田勝久、 神谷和作
  • Organizer: 第15回日本再生医療学会総会
  • Place of Presentation: 大阪国際会議場(大阪市)
  • Year and Date: 2016-03-17 – 2016-03-19
• [Presentation] Restoration og Cochlear Gap Junction for GJB2 (2016)
  • Author(s): Kazusaku Kamiya, Ichiro Fukunaｇa
  • Organizer: Association for Research in Otolaryngology (ARO), 39th Annual MidWinter Meeting
  • Place of Presentation: Manchester Grand Hyatt, San Diego, CA, USA
  • Year and Date: 2016-02-20 – 2016-02-24
  • Int'l Joint Research
• [Presentation] Deformation of the Outer Hair Cells and the Accumulation of Caveolin-2 in Connexin 26-Deficient Mice (2016)
  • Author(s): Takashi Anzai, Kazusaku Kamiya, Katsuhisa Ikeda
  • Organizer: Association for Research in Otolaryngology (ARO), 39th Annual MidWinter Meeting
  • Place of Presentation: Manchester Grand Hyatt, San Diego, CA, USA
  • Year and Date: 2016-02-20 – 2016-02-24
  • Int'l Joint Research
• [Presentation] Generation of induced pluripoten stem cells form connexin26 conditional knock out mouce (2016)
  • Author(s): Toru Aoki, Katsuhisa Ikeda, Kazusaku Kamiya
  • Organizer: Association for Research in Otolaryngology (ARO), 39th Annual MidWinter Meeting
  • Place of Presentation: Manchester Grand Hyatt, SanDiego,CA, USA
  • Year and Date: 2016-02-20 – 2016-02-24
  • Int'l Joint Research
• [Presentation] Differentiation of mouse iPS cell into Cx26-positive cell and formation of inter cellular Cx26-gap junction plaque (2016)
  • Author(s): Ichiro Fukunaga, Kaori Hatakeyama,Toru Aoki, Atena Nishikawa, Ayumi Fujimoto, Katsuhisa Ikeda, Kazusaku Kamiya
  • Organizer: Association for Research in Otolaryngology (ARO), 39th Annual MidWinter Meeting
  • Place of Presentation: Manchester Grand Hyatt, SanDiego,CA, USA
  • Year and Date: 2016-02-20 – 2016-02-24
  • Int'l Joint Research
• [Presentation] Induction of iPS cell differentiation into Connexin26 positive cells with gap junction plaques using cochlear feeder cell (2016)
  • Author(s): Ayumi Fujimoto, Ichiro Fukunaga, Kaori Hatakeyama, Toru Aoki, Atena Nishikawa, Katsuhisa Ikeda, Kazusaku Kamiya
  • Organizer: Association for Research in Otolaryngology (ARO), 39th Annual MidWinter Meeting
  • Place of Presentation: Manchester Grand Hyatt, SanDiego,CA, USA
  • Year and Date: 2016-02-20 – 2016-02-24
  • Int'l Joint Research
• [Presentation] Various cochlear gap junction plaque reproduced in human cell line (2016)
  • Author(s): Kaori Hatakeyama, Katsuhisa Ikeda, Kazusaku Kamiya
  • Organizer: Association for Research in Otolaryngology (ARO), 39th Annual MidWinter Meeting
  • Place of Presentation: Manchester Grand Hyatt, SanDiego,CA, USA
  • Year and Date: 2016-02-20 – 2016-02-24
  • Int'l Joint Research
• [Presentation] Cochlear gap junction plaque, stabilized macromolecular complex composed of specific connexins (2015)
  • Author(s): Kazusaku Kamiya, Ichiro Fukunaga, Kaori Hatakeyama, Toru Aoki, Ayumi Fujimoto, Atena Nishikawa ,Takashi Anzai, Osamu Minowa, Katsuhisa Ikeda
  • Organizer: 52nd Inner Ear Biology Workshop
  • Place of Presentation: Campus of the Catholic University of Rome, Italy
  • Year and Date: 2015-09-12 – 2015-09-15
  • Int'l Joint Research
• [Presentation] Caveolins accumulates at the organ of Corti in GJB2 associated deafness (2015)
  • Author(s): Takashi Anzai, Kazusaku Kamiya, Katsuhisa Ikeda
  • Organizer: 52nd Inner Ear Biology Workshop
  • Place of Presentation: Campus of the Catholic University of Rome, Italy
  • Year and Date: 2015-09-12 – 2015-09-15
  • Int'l Joint Research
• [Presentation] Reproduction of various types of cochlear gap junction plaques in human cell line (2015)
  • Author(s): Kaori Hatakeyama, Katsuhisa Ikeda, Kazusaku Kamiya
  • Organizer: 52nd Inner Ear Biology Workshop
  • Place of Presentation: Campus of the Catholic University of Rome, Italy
  • Year and Date: 2015-09-12 – 2015-09-15
  • Int'l Joint Research
• [Presentation] Differentiation of mouse iPS cell into Cx26-positive cell and formation of inter cellular Cx26-gap junction plaque (2015)
  • Author(s): Ichiro Fukunaga, Kaori Hatakeyama,Toru Aoki, Atena Nishikawa, Ayumi Fujimoto, Osamu Minowa, Katsuhisa Ikeda, Kazusaku Kamiya
  • Organizer: 52nd Inner Ear Biology Workshop
  • Place of Presentation: Campus of the Catholic University of Rome, Italy
  • Year and Date: 2015-09-12 – 2015-09-15
  • Int'l Joint Research