analysis of water channel function to decrease of brain edema caused by mild hypothermia with using RNAi technique

2008 Fiscal Year Final Research Report

• Project/Area Number: 18591714
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Single-year Grants
• Section: 一般
• Research Field: Anesthesiology/Resuscitation studies
• Research Institution: Nagoya City University
• Principal Investigator: FUJITA Yoshihito Nagoya City University, 大学院・医学研究科上, 講師 (90238593)
• Co-Investigator(Kenkyū-buntansha): ASAI Kiyofumi 名古屋市立大学, 大学院・医学研究科, 教授 (70212462) ; SOBUE Kazuya 名古屋市立大学, 大学院・医学研究科, 教授 (90264738)
• Project Period (FY): 2006 – 2008
• Keywords: 水チャンネル / 脳浮腫 / RNAi / アクアポリン / 脳低温

Research Abstract

今後の研究の基礎となるアクアポリン(AQP)Knockdown細胞株の確立を重点に行った。Knockdownが確認されているconstructに加えオリジナルのものを作成した。vectorには、invitrogen社製のBlock-IT Pol II miR RNAi expression vector、pcDNA 6.2-GW/miRを使用した。そのプラスミドを大腸菌にtransformationして、大量培養し、Quiagen社のキットを用いて、目的となるconstructを含むプラスミドを大量に、無菌的に精製した。RNAiの効果を確認するが効果が弱く、原因の追求のためGFPをvectorに挿入した。そのGFPつきのvectorの挿入で、視覚的におおざっぱにいって20%ぐらいのtranfection効率があることを確認した。ただRNAi効果をえる効率は、まだ10%程度にとどまった。transfection効率をあげるためレンチウイルスを使用したvectorの作製を進めると同時に、現在作成してあるGFPつきvectorを用い。実際の低温における脳浮腫効果の解明を始めている。また、同時に疾患の重傷度の指標として注目している乳酸値について、乳酸負荷でのアストロサイトにおける水チャンネルの発現の変化についても解析し、論文発表した。

Research Products

• [Journal Article] Before-after study of a restricted fluid infusion strategy for management of donor hepatectomy for living donor liver transplantation (2009)
  • Author(s): Yoshihito Fujita, Akinori Takeuchi, Takeshi Sugiura, Tomonori Hattori, Nobuko Sasano, Yuichiro Mizuochi, Kazuya Sobue
  • Journal Title: J. Anesth (in press)
  • Peer Reviewed
• [Journal Article] Lactic acid increases aquaporin 4 expression on the cell membrane of cultured rat astrocytes (2008)
  • Author(s): Morishima T, Aoyama M, Iida Y, Yamamoto N, Hirate H, Arima H, Fujita Y, Sasano H, Tsuda T, Katsuya H, Asai K, Sobue K.
  • Journal Title: Neurosci Res. 61(1) Pages: 18-26
  • Peer Reviewed
• [Journal Article] Ca2+-dependent activator protein for secretion 1 is critical for constitutive and regulated exocytosis but not for loading of transmitters into dense core vesicles (2007)
  • Author(s): Fujita Yoshihito, Xu A, Xie L, Arunachalam L, Chou TC, Jiang T, Chiew SK, Kourtesis J, Wang L, Gaisano HY, Sugita S.
  • Journal Title: J Biol Chem 282(29) Pages: 21392-403
  • Peer Reviewed
• [Journal Article] Munc18-1 Is Critical for Plasma Membrane Localization of Syntaxinl but Not of SNAP-25 in PC12 Cells (2007)
  • Author(s): Arunachalam L, Han L, Tassew NG, He Y, Wang L, Xie L, Fujita Y, Kwan E, Davletov B, Monnier PP, Gaisano HY, Sugita S.
  • Journal Title: Mol Biol Cell 19(2) Pages: 722-734
  • Peer Reviewed
• [Journal Article] Anesthetic management of a patient with mitochondrial myopathy, encephalopathy, lactic acidosis, and stroke-like episodes(MELAS)during laparotomy (2007)
  • Author(s): Nobuko Sasano, Yoshihito Fujita, MinHye So, Kazuya Sobue, Hiroshi Sasano and Hirotada Katsuya
  • Journal Title: J. Anesth 21 Pages: 72-5
  • Peer Reviewed
• [Journal Article] Inadvertent intrathecal cannulation in an infant demonstrated by three-dimensional computed tomography : a rare complication of internal jugular vein catheterization (2006)
  • Author(s): Fujita Yoshihito, Sobue K, Hattori T, Takeuchi A, Tsuda T, Katsuya H.
  • Journal Title: J. Anesthesia 20 Pages: 122-5
  • Peer Reviewed
• [Journal Article] Intra-operative monitoring of vagal nerve activity with wire electrodes (2006)
  • Author(s): Ito H, Sobue K, So M, Hirate H, Sugiura T, Azami T, Fujita Y, Sasano H, Katsuya H.
  • Journal Title: Acta Anaesthesiol Scand 50(10) Pages: 1304-5
  • Peer Reviewed
• [Journal Article] 硬膜外麻酔併用全身麻酔時に必要な最小限セボフルラン濃度の検討 (2006)
  • Author(s): 藤田義人、森島徹朗、祖父江和哉、勝屋弘忠
  • Journal Title: 神経麻酔・集中治療プロシーディング 2006 Pages: 29-30
• [Journal Article] Failure of the Airway Scope to reach the larynx
  • Author(s): Nobuko Sasano, Hiroki Yamauchi, Yoshihito Fujita
  • Journal Title: Can J Anesth2007 54(9) Pages: 775
  • Peer Reviewed
• [Presentation] Investigation of blood lactate as an indicator for severity of a child undergone cardiac surgery (2008)
  • Author(s): Yoshihito Fujita, Eisuke Kako, Yuichiro Mizuochi, Hironobu Iguchi, Kazuva Sobue
  • Organizer: Annual Meeting of the American Society of Anesthesiologists
  • Place of Presentation: (ASA)in Orland, Florida
  • Year and Date: 20081018-22
• [Presentation] Investigation of blood lactate level as indicator for severity assessment (2008)
  • Author(s): Yoshihito Fujita, Tae Kato, Hirotada Katsuya
  • Organizer: The 6^<th> International Shock Congress in Cologne
  • Place of Presentation: Germany
  • Year and Date: 20080628-0702
• [Presentation] Minimum concentration of sevoflurane for adequate sedation with combined General-epidural anesthesia (2007)
  • Author(s): Yoshihito Fujita, Hiroki Yamauchi, Masato Morita, Rina Kato
  • Organizer: Annual Meeting of the American Society of Anesthesiologists
  • Place of Presentation: (ASA)in San Francisco
  • Year and Date: 20071013-17
• [Presentation] Effect of mild hypothermia on the expression of aquaporin family in cultured rat astrocytes under hypoxic condition (2007)
  • Author(s): Fujita Yoshihito, Sobue Kazuva, Morishima Tetsuro, Asai Kiyofumi
  • Organizer: the 5th International Conference of Aquaporin in Nara
  • Place of Presentation: Japan
  • Year and Date: 20070713-1316
• [Presentation] Restricted-Volume Replacement Reduces Blood Loss during Donor Hepatectomy for Liver Transplantation (2006)
  • Author(s): Fujita Yoshihito, Katsuya Hirotada
  • Organizer: 2006 Annual Meeting of the American Society of Anesthesiologists
  • Place of Presentation: (ASA)in Chicago
  • Year and Date: 20061106-20