再生肝細胞コロニーに形成された毛細胆管構造のマイクロ・ナノバイオメカニクス

• Project/Area Number: 15086214
• Research Category: Grant-in-Aid for Scientific Research on Priority Areas
• Allocation Type: Single-year Grants
• Review Section: Science and Engineering
• Research Institution: Keio University
• Principal Investigator: 谷下 一夫 Keio University, 理工学部, 教授 (10101776)
• Co-Investigator(Kenkyū-buntansha): 池田 満里子 慶應義塾大学, 名誉教授 (00051368) ; 工藤 奨 芝浦工業大学, 工学部, 講師 (70306926) ; 須藤 亮 慶應義塾大学, 大学院・理工学研究科, 特別研究助手 (20407141)
• Project Period (FY): 2003 – 2006
• Project Status: Completed (Fiscal Year 2006)
• Budget Amount: ¥34,900,000 (Direct Cost: ¥34,900,000); Fiscal Year 2006: ¥8,700,000 (Direct Cost: ¥8,700,000); Fiscal Year 2005: ¥8,700,000 (Direct Cost: ¥8,700,000); Fiscal Year 2004: ¥8,700,000 (Direct Cost: ¥8,700,000); Fiscal Year 2003: ¥8,800,000 (Direct Cost: ¥8,800,000)
• Keywords: 肝細胞 / 毛細胆管 / バイオメカニクス / 再生医療 / バイオ人工肝臓 / 毛細血管ネットワーク / 血管新生 / 内皮細胞 / 胆管上皮細胞 / 管腔形成 / 胆管ネットワーク / コロニー / 細胞間相互作用 / 収縮

Research Abstract

増殖と分化に富む小型肝細胞を肝臓から分離して培養した所、コロニー形成して毛細胆管が形成され、その毛細胆管が一定な周期で収縮し胆汁流動が生じている事が明らかになった。このような胆管の収縮は、小型肝細胞同士のコミュニケーションの結果、肝細胞の収縮がコロニー全体で同期している事実が見出された。本研究では、培養小型肝細胞のコロニーによって発生された毛細胆管構造の微小運動による収縮を生み出す肝細胞収縮の同期機構に関して、マイクロナノバイオメカニクスの観点から明らかにする事を目的とする。そこで、コロニーに形成された毛細胆管であることを立証するため、細胞の極性、発現するタンパクの同定、収縮のダイナミクス、細胞骨格の様子などを詳細に調べた。さらに、3次元小型肝細胞コロニーを機能を持つ集合体にするため、毛細胆管と胆管との連結、さらに毛細血管網の共存を実現すべく検討を行った。胆管形成に関しては、胆管上皮細胞の分離とその管腔構造の再構築を行った。胆管上皮細胞を分離して体外で胆管様構造を形成することに成功した。この胆管様構造は、数10μmの径を持ち、体外で形成された管腔構造としては極めて大きなもので、肝臓再生に一歩近づいたと考えている。さらに、毛細血管ネットワークを形成させるため、血管内皮細胞の共培養の可能性に関して基礎的な実験を試みた。特に興味深い点は、小型肝細胞と内皮細胞の間を伊東細胞が仲介するように遊走する現象で、伊東細胞のような非実質細胞が重要な役割を演じている事が分かった。

Research Products

• [Journal Article] Effects of the mechanical properties of collagen gel on the in vitro formation of microvessel networks by endothelial cells (2007)
  • Author(s): N. Yamamura, et al.
  • Journal Title: Tissue Engineering 13 Pages: 1443-1453
  • Peer Reviewed
• [Journal Article] Effects of the mechanical properties of collagen gel on the in vitro formation of microvessel networks by endothelial cells (2007)
  • Author(s): Tanishita et al.
  • Journal Title: Tissue Engineering 13(in press)
• [Journal Article] Bionic design of tube formation in the integrated cellular structure (2007)
  • Author(s): Tanishita, K
  • Journal Title: Proc. TMS 2007 Annual Meeting Pages: 125-125
• [Journal Article] Initial bFGF distribution affects the depth of three-dimensional microvessel networks in vitro (2006)
  • Author(s): A. Ueda, et al.
  • Journal Title: J Biomechanical Science and Engineering 1 Pages: 136-146
  • Peer Reviewed
• [Journal Article] Initial bFGF distribution affects the depth of three-dimensional microvessel networks in vitro (2006)
  • Author(s): Tanishita et al.
  • Journal Title: J Biomechanical Science and Engineering 1 Pages: 136-146
  • NAID: 130000079537
• [Journal Article] Regulation on cellular configuration by cell-cell adhesion in the 3D stacked-up culture of rat primary hepatosytes (2006)
  • Author(s): Tanishita et al.
  • Journal Title: J Biomechanics 39
• [Journal Article] Effect of endothelial progenitor cells on the formation of in vitro three dimensional network (2006)
  • Author(s): Tanishita et al.
  • Journal Title: J Biomechanics 39
• [Journal Article] Evaluation of coculture model composed of sinusoidal endothelial cells and small hepatocytes (2006)
  • Author(s): Tanishita et al.
  • Journal Title: J Biomechanics 39
• [Journal Article] Coordinated movement of bile canalicular networks reconstructed by rat small hepatocytes (2005)
  • Author(s): Sudo, R., et al.
  • Journal Title: Annals of Biomed Eng. 33 Pages: 696-708
  • Peer Reviewed
• [Journal Article] Reconstruction of 3D stacked-up structures by rat small hepatocytes on microporous membrance (2005)
  • Author(s): R. Sudo, et al.
  • Journal Title: The FASEB Journal, 2005 published on line August17 (Published on line)
  • Peer Reviewed
• [Journal Article] 多孔性膜を用いた小型肝細胞の3次元培養と毛細胆管の形成 (2005)
  • Author(s): 谷下一夫 他
  • Journal Title: 第12回肝細胞研究会 12 Pages: 78-78
• [Journal Article] ラット胆管上皮細胞による管構造の形成 (2005)
  • Author(s): 谷下一夫 他
  • Journal Title: 第26回日本炎症・再生医学会 26 Pages: 375-375
• [Journal Article] Reconstruction of 3D liver-like structures by stacking up layers of rat small hepatocytes, (2005)
  • Author(s): Tanishita K et al.
  • Journal Title: Proc The second Japan-Switzerland workshop on biomechanics Pages: 128-128
• [Journal Article] Reconstruction of 3D stacked-up structures by rat small hepatocytes on microporous membrance (2005)
  • Author(s): Tanishita K et al.
  • Journal Title: The FASEB Journal 19 Pages: 1695-1697
• [Journal Article] Coordinated movement of bile canalicular networks reconstructed by rat small hepatocytes (2005)
  • Author(s): Tanishita K et al.
  • Journal Title: Annals of Biomedical Engineering 33 Pages: 696-708
• [Journal Article] Reconstruction of ductular structure by rat biliary epithelial cells (2005)
  • Author(s): Tanishita K et al.
  • Journal Title: Proc.2005 Summer Bioengineering Conference (CD-ROM)
• [Journal Article] Coordinated movement of bile canalicular networks reconstructed by rat small hepatocytes (2005)
  • Author(s): Tanishita, K, Sudo, R et al.
  • Journal Title: Annals of Biomedical Engineering (in press)
• [Journal Article] 多孔性膜の積層による小型肝細胞の三次元培養 (2005)
  • Author(s): 谷下一夫 他
  • Journal Title: 第17回バイオエンジニアリング講演会 講演論文集 04-48 Pages: 83-84
• [Journal Article] Effect of glycocalyx on shear-dependent albumin uptake in endothelial cells (2004)
  • Author(s): Ueda, A., et al.
  • Journal Title: American J Physiology, Heart Circ Physiol 287
  • Peer Reviewed
• [Journal Article] Bile canalicular formation in hepatic organoid reconstructed by rat small hepatocytes and hepatic nonparenchymal cells (2004)
  • Author(s): Sudo, R., et al.
  • Journal Title: J Cellular Physiol. 199 Pages: 252-261
  • Peer Reviewed
• [Journal Article] Effect of shear stress on micro-vessels network formation of endothelial cells with in vitro three-dimensional model (2004)
  • Author(s): Ueda, A., et al.
  • Journal Title: Am J Physiol, Heart Circ Physiol 287
  • Peer Reviewed
• [Journal Article] Relationship between microtubule network structure and intracellular transport incultured endothelial cells affected by shear stress (2004)
  • Author(s): Kudo, S., et al.
  • Journal Title: JSME Int J, Ser. C 47 Pages: 977-984
  • Peer Reviewed
• [Journal Article] Bile canalicular formation in hepatic organoid reconstructed by rat small hepatocytes and nonparenchymal cells (2004)
  • Author(s): Tanishita, K, Sudo, R et al.
  • Journal Title: Journal of Cellular Physiology 199(2) Pages: 252-261
• [Journal Article] 小型肝細胞の積層培養における毛細胆管の形成 (2004)
  • Author(s): 谷下一夫, 他
  • Journal Title: 第11回肝細胞研究会抄録集 Pages: 92-92
• [Journal Article] Bile canalicular formation in pile-up culture of small hepatocytes cultured on porous membranes (2004)
  • Author(s): Tanishita, K, Sudo, R et al.
  • Journal Title: 2004 FASEB Summer Research Conferences, Proceedings (CD-ROM)
• [Journal Article] Relationship between microtubule network structure and intracellular transport in cultured endothelial cells affected by shear stress (2004)
  • Author(s): Tanishita, K et al.
  • Journal Title: JSME Int J, Ser.C 47(4) Pages: 977-984
  • NAID: 110004820826
• [Presentation] Evaluation of coculture model composed of sinusoidal endothelial cells and small hepatocytes (2006)
  • Author(s): Kazuo Tanishita
  • Organizer: 5^<th> World Congress of Biomechanics
  • Place of Presentation: Germany
  • Year and Date: 2006-07-31
• [Presentation] Contribution of endothelial progenitor cells on microvessel-like network formation in vitro (2005)
  • Author(s): Yoshiyuki Ozaki
  • Organizer: 2nd Asian pacific conference on biomechanics
  • Place of Presentation: Taipei, Taiwan
  • Year and Date: 2005-10-25
• [Presentation] Effect of sodium orthovanadate on culture of sinusoidal endothelial cells (2005)
  • Author(s): Hiroyuki Asami
  • Organizer: 2nd Asian pacific conference on biomechanics
  • Place of Presentation: Taipei, Taiwan
  • Year and Date: 2005-10-23
• [Presentation] Reconstruction of 3D stacked-up structures by rat small hepatocytes cultured on microporous membranes (2005)
  • Author(s): Ryo Sudo
  • Organizer: 2nd Japan-Switzerland Workshop on Biomechanics
  • Place of Presentation: Kyoto, Japan
  • Year and Date: 2005-09-14
• [Presentation] Bionic design of tube formation in the integrated cellular structure (2005)
  • Author(s): Kazuo Tanishita
  • Organizer: 8th Japanese-polish seminar on new engineering methods supporting human life
  • Place of Presentation: Kanagawa, Japan
  • Year and Date: 2005-09-06
• [Presentation] Bile canalicular formation in 3D stacked-up culture of rat small hepatocytes and nonparenchymal cells (2005)
  • Author(s): Ryo Sudo
  • Organizer: ASME summer bioengineering conference
  • Place of Presentation: Colorado, USA
  • Year and Date: 2005-06-23
• [Presentation] Reconstruction of ductular structure by rat biliary epithelial cells (2005)
  • Author(s): Wataru Hashimoto
  • Organizer: ASME summer bioengineering conference
  • Place of Presentation: Colorado, USA
  • Year and Date: 2005-06-23
• [Presentation] Reconstruction of 3D liver-like structures by stacking cell-layers cultured on microporous membranes (2005)
  • Author(s): Ryo Sudo
  • Organizer: 6th Asian-pacific conference on medical and biological engineering
  • Place of Presentation: Ibaraki, Japan
  • Year and Date: 2005-04-25
• [Presentation] Bile canalicular formation in pile-up culture of rat small hepatocytes cultured on porous membranes (2004)
  • Author(s): Ryo Sudo
  • Organizer: FASEB (federation of American societies for experimental biology) summer research conferences, Mechanisms of liver growth, development and disease
  • Place of Presentation: Colorado, USA
  • Year and Date: 2004-08-08
• [Presentation] In vitro multicellular system reconstructed by small hepatocytes and nonparenchymal cells (2003)
  • Author(s): Ryo Sudo
  • Organizer: International symposium of 21st century COE program, Keio university, Understanding and control of life's function via systems biology
  • Place of Presentation: Kanagawa, Japan
  • Year and Date: 2003-11-20
• [Presentation] Coordinated contraction of bile canaliculi reconstructed in rat small hepatocytes (2003)
  • Author(s): Ryo Sudo
  • Organizer: ASME (American society of mechanical engineers) summer bioengineering conference
  • Place of Presentation: Florida, USA
  • Year and Date: 2003-06-26
• [Book] Biomechanics at Micro- and Nanoscale levels, Vol. IV (2007)
  • Author(s): Hiroshi Wada
  • Publisher: World Scientific Publishing
• [Book] Biomechanics at Micro- and Nanoscale levels, Vol. II (2006)
  • Author(s): Hiroshi Wada
  • Publisher: World Scientific Publishing
• [Book] Biomechanics at Micro- and Nanoscale levels. Vol. I (2005)
  • Author(s): Hiroshi Wada
  • Publisher: World Scientific Publishing
• [Book] Biomechanics at micro-and nanoscale levels (2005)
  • Author(s): Tanishita, K et al.
  • Total Pages: 173
  • Publisher: World Scientific
• [Book] Organ Microcirculation (2004)
  • Author(s): H. Ishi
  • Publisher: World Scientific Publishing
• [Publications] Tanishita, K, Sudo, R, et al.: "Bile canalicular formation in hepatic organoid reconstructed by rat small hepatocytes and nonparenchymal cells"J.Cellular Physiology. (In press). (2004)
• [Publications] Tanishita, K, Sudo, R, Ikeda, M, et al.: "Reconstruction of Ductular Structure by Rat Bile Ductular Epithelial Cells"Proc.First Asian Pacific Conference on Biomechanics. 219-220 (2004)
• [Publications] Tanishita, K, Sudo, R, Ikeda, et al.: "Cellular Biomechanics Applied to Tissue Engineering (Plenary lecture)"Proc.First Asian Pacific Conference on Biomechanics. 12 (2004)
• [Publications] Tanishita, K, Sudo, R, Ikeda, M et al.: "Coordinated Contraction of Bile Canaliculi Reconstructed in Rat Small Hepatocytes."Proc.of the 2003 Summer Bioengineering Conf. (The award was given to this paper.). (CD-ROM). (2003)
• [Publications] 谷下一夫, 池田満里子, 他: "小型肝細胞による毛細胆管の形成とビリルビン排出機能"再生医療. 2. 87 (2003)
• [Publications] 谷下一夫, 池田満里子, 他: "凍結保存小型肝細胞における毛細胆管の形成と運動"生体医工学. 41. 399 (2003)