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Ohsawa Ikuroh

    Biological Process of Aging　Vice-director
Last Updated :2025/04/05

Researcher Information

Degree

  • Ph. D

URL

Research funding number

  • 30343586

J-Global ID

Research Interests

  • 酸化ストレス   水素   神経変性疾患   認知症   老化   加齢   Oxidative Stress   Molecular Hydrogen   Dementia   Aging   

Research Areas

  • Life sciences / Cell biology
  • Life sciences / Molecular biology
  • Nanotechnology/Materials / Molecular biochemistry
  • Life sciences / Nutrition and health science
  • Life sciences / Neuroscience - general

Academic & Professional Experience

  • 2010/04 - Today  東京都健康長寿医療センター研究所生体調節機能研究研究副部長
  • 2008  ：日本医科大学老人病研究所教授
  • 2003  ：日本医科大学老人病研究所講師
  • 2001  ：日本医科大学老人病研究所助手
  • 2001  : Researcher Associate, Institute of Gerontology, Nippon Med Sch
  • 1994  ：国立精神・神経センター神経研究所代謝研究部ポスドク
  • 1994  : Researcher, National Institute of Neuroscience, NCNP,
  • 1984  ：日本ゼオン株式会社研究員
  • 1984  : Researcher, Zeon Corporation

Education

  •        -   The University of Tokyo  The Faculty of Engineering  化学工学科
  •        -   Department of Chemical Engineering, Faculty of Engineering, University of Tokyo

Association Memberships

  • 日本ミトコンドリア学会   日本分子生物学会   日本生化学会   日本神経化学会   化学工学会   The Society of Chemical Engineers, Japan   The Japanese Society of Neurochemistry   The Japanese Biochemical Society   日本分子状水素医学生物学会   日本基礎老化学会   

Published Papers

MISC

Industrial Property Rights

Research Grants & Projects

  • Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C)
    Date (from‐to) : 2022/04 -2025/03 
    Author : 松本 浩; 大澤 郁朗; 鈴木 郁郎
  • 日本学術振興会：科学研究費助成事業 基盤研究(C)
    Date (from‐to) : 2022/04 -2025/03 
    Author : 大澤 麻登里; 松本 浩; 大澤 郁朗
  • Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C)
    Date (from‐to) : 2022/04 -2025/03 
    Author : 青景 聡之; 大澤 郁朗; 中尾 篤典
  • Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C)
    Date (from‐to) : 2022/04 -2025/03 
    Author : 野田 義博; 大澤 郁朗
  • 日本学術振興会：科学研究費助成事業 基盤研究(C)
    Date (from‐to) : 2021/04 -2024/03 
    Author : 河田 光弘; 大澤 郁朗
     
    急性大動脈解離発症に伴い、全身性炎症反応症候群が起こり、サイトカインストームに至り、急性肺障害・肺酸素化障害を引き起こす。分子状水素(H2)の投与では、多くの非感染性炎症においてサイトカインストームの抑制効果が報告されている。 本研究では、保存的治療が主となる急性大動脈解離Stanford B型でのH2ガス吸入療法の確立を目指す。大動脈解離の動物モデルを用いて、H2ガスの最適な投与方法、ガス濃度、投与期間を明らかにする。次に急性大動脈解離で重篤な合併症が無い患者を対象に、標準的内科治療にH2ガス吸入療法を併用することにより抗炎症作用で肺酸素化障害を軽減、予防できるか検討する。 令和３年度については、倫理委員会などの諸手続きが完了し、水素ガス吸引療法に必要な混合ガス吸入装置、水素ガス濃度計、安全装置などのセットが完了して、既に数名の急性大動脈解離Stanford B型患者について投与試験を開始し次年度も引き続き研究進行していく。
  • Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (B)
    Date (from‐to) : 2020/04 -2023/03 
    Author : 野々山 恵章; 松本 浩; 大澤 郁朗; 大澤 麻登里; 關中 悠仁
     
    Chediak東症候群(CHS)は神経変性および免疫不全を呈する疾患である。原因遺伝子はLYSTであるが、神経変性の発症機序は不明である。本研究ではChediak東症候群(CHS)患者６症例からiPS細胞を樹立し、神経細胞に分化させた。６症例のLYST遺伝子変異は全て異なっていたが、５症例で神経症状があり１例は幼児のため神経症状は解析不能であった。
    分化誘導した神経細胞で、細胞内小器官であるリソソーム、ミトコンドリア、オートファゴソームの解析を行った。その結果、蛍光顕微鏡および電子顕微鏡による解析で、リソソームでは、巨大化、PAS染色陽性物質の沈着、細胞内部のフィラメント様構造および黒色顆粒の出現などの形態異常を認めた。機能解析ではカルシウム依存性エキソサイトーシス障害を認めた。ミトコンドリアでは、電子顕微鏡の解析で、形態の不整、肥大化、内部の黒色顆粒を認めた。また核周囲への局在化、分布異常も認めた。オートファゴソームについては、DAP Green染色による蛍光顕微鏡の解析で輝度の亢進および閾値以上の細胞の割合の増加を認めた。
    以上の結果から、Chediak東症候群(CHS)では細胞内小器官の異常があることを明らかにした。このことから、Chediak東症候群(CHS)の神経細胞では、リソソーム機能異常により異常な細胞内小器官および老廃物の処理ができず細胞内に蓄積し、これにより神経障害が発症することが示唆された。
  • 日本学術振興会：科学研究費助成事業 基盤研究(B)
    Date (from‐to) : 2020/04 -2023/03 
    Author : 大澤 郁朗; 藤田 泰典; 池谷 真澄; 伊藤 雅史
     
    本研究はH2による健康長寿促進を目指して、三つの問いを解明するための包括的研究である。第一の研究では、培養細胞系を中心にリピドームとメタボローム解析から、神経芽細胞SH-SY5Yを水素存在下で培養すると、１時間後にはカルジオリピンなどの特定のリン脂質が上昇し、同時にエネルギー代謝経路が広く抑制されることが判明した。この変化は一過的であり、その間にグルタチオンの低下など酸化ストレスの上昇が認められた。これがミトホルミシスなどの細胞防御機構を誘導することがH2の作用機序の一つであることを示しすことができた。さらに細胞死を抑制する機序については、水素ガス吸入による幼弱マウスの麻酔ガス吸入時脳細胞死抑制効果を確認し、その脳内で細胞死シグナル変化を解析している。第二の研究については、デキストラン硫酸塩投与の大腸炎モデルマウスで、水素水を１日１回投与するだけで病態が緩和された。デキストラン硫酸塩投与は小腸パイエル板でFoxp3の発現を減少し、Il6の発現を上昇させる。水素水の飲用はこれを抑制したことから、IL-6の抑制によって制御性T細胞の恒常性が維持されたものと考えられる。H2が免疫恒常性に関与するメカニズムについてさらに解析中である。第三の研究については、倫理委員会などの諸手続きが完了し、水素ガス吸引療法に必要な混合ガス吸入装置、水素ガス濃度計、安全装置などのセットが完了して、既に数名の急性大動脈解離患者について投与試験を完了し引き続き研究進行中である。
  • Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C)
    Date (from‐to) : 2019/04 -2022/03 
    Author : 本田 陽子; 大澤 郁朗
  • Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C)
    Date (from‐to) : 2019/04 -2022/03 
    Author : Toshiyuki Aokage
     
    Acute respiratory distress syndrome causes alveolar fibrosis. We hypothesized that daily repeated inhalation of hydrogen gas could suppress persistent inflammatory and consequently inhibit lung fibrosis. To test this hypothesis, bleomycin lung injury model mouse, that is imitating ARDS, were exposed to hydrogen (3.2% in air) for 6 hours every day for 21 days. Respiratory physiology, tissue pathology, markers of inflammation, and macrophage phenotypes were examined. Mice with bleomycin-induced lung injury that received daily hydrogen therapy for 21 days (BH group) exhibited higher static compliance than mice with bleomycin-induced lung injury exposed only to air. When the mRNA levels of pro-inflammatory cytokines were examined 7 days after bleomycin administration, interleukin (IL)-6, IL-4 and IL-13 were significantly lower in the BH group than in the BA group. There were significantly fewer M2-biased macrophages in the alveolar interstitium of the BH group than in the BA group.
  • Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C)
    Date (from‐to) : 2017/04 -2022/03 
    Author : 野田 義博; 大澤 郁朗
     
    本研究は、近年報告された分子状水素(H2)の医療への応用(水素分子医学)という革新的な研究の一端である。生殖医療におけるH2の応用研究は、本申請時点で申請者の研究グループ以外からの報告はまだない。このため、精子への酸化ストレス障害と、それにより引き起こされる受精能の低下をH2が改善できるか確かめる本研究の独創性は極めて高い。現在までの研究成果より、マウス精子に過酸化水素を添加することで精子の運動機能低下を誘導する酸化ストレス障害モデル精子の実験系を確立した。そして、これにより生じる精子運動機能、体外受精による受精率および胚移植による個体発生率の低下に対するH2の抑制効果を検証した。その結果、H2は酸化ストレス障害を受けたマウス精子の運動機能改善効果を呈した。さらに、精子の機能低下とミトコンドリア活性に着目し、形態学的な観察と蛍光プローブを用いたミトコンドリアの構造解析について検討した。そして、ミトコンドリアの呼吸活性についても更なる検討を進めた。本研究は臨床の観点からも極めて高い意義を持つ。不妊治療では可能な限り体外受精(IVF)が選択される。H2による精子運動性の活性化がIVFによる受精率向上につながれば、安全で効果的な新しい男性不妊治療法となりうる。また、凍結保存精子の出生率改善も期待できる。このように、本研究の遂行により、薬剤治療の困難な 「生殖医療」に安全性の高いH2を応用が、社会的問題となっている高齢出産における不妊を始めとした諸問題の解決への基盤となる知見を得ることができる。
  • Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C)
    Date (from‐to) : 2018/04 -2021/03 
    Author : Iketani Masumi
     
    Molecular hydrogen (H2) is known to act as an antioxidant and anti-inflammatory agent in the body, however, the mechanism of action remains unclear. In this study, we found that exposure of cells to H2 for only one hour changed the phospholipids that constitute the cell membrane. The phospholipids, which are known to function in the cellular organelles mitochondria and endosomes, were found to increase. Each of these is a cell organelle that is closely related to disease. We examined for changes in mitochondria-related metabolism, and found that many metabolites decreased within 1 hour of exposure to H2, indicating that transient oxidative stress was induced. We also examined endosomes and found that endosomal transport was delayed.
  • Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (B)
    Date (from‐to) : 2016/04 -2019/03 
    Author : Ikuroh Ohsawa
     
    We found that molecular hydrogen protects against oxidative stress-induced SH-SY5Y neuroblastoma cell death through the process of mitohormesis. Indeed, preadministration of hydrogen-rich water to mice was enough to protect against lipopolysaccharide-induced sepsis and attenuate liver injury. Furthermore, we found that molecular hydrogen attenuates gefitinib-induced exacerbation of acute lung injury in mice through a reduction in oxidative stress and inflammation. Hydrogen-rich water administration also prevents vascular aging of the aorta in LDL receptor-deficient mice.
  • Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Challenging Exploratory Research
    Date (from‐to) : 2015/04 -2018/03 
    Author : Ohsawa Ikuroh; IKETANI Masumi
     
    We applied stimulated emission depletion imaging with subdiffraction resolution to submitochondrial structures. Their shapes depend on both a cell’s type and its physiological state. Staining with a cationic fluorescent dye, TMRM, unveiled intriguing details of lamellar structure, consisting of rapidly changeable, curtain-like formations. The TMRM-positive structure colocalized with neither protein in the matrix nor on the outer membrane, but partially localized with the nucleoid. Suppression of a component in the mitochondrial contact site disrupted the lamellar TMRM-positive structure. Uncoupling of the oxidative phosphorylation system released TMRM from the inner membrane without any alteration in the matrix structure. The approach presented here provides novel insights into the in vivo nature of submitochondrial structures, and can be used for further functional investigations of these complex structures.
  • Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research
    Date (from‐to) : 2014/04 -2017/03 
    Author : Takahashi Hiroshi; Ohsawa Ikuroh
     
    In phacoemulsification, ultrasound induces hydroxyl radical (・OH) formation, damaging corneal endothelium. The effects of H2 in phacoemulsification were evaluated in rabbits, comparing H2-dissolved and control solutions. The whole cornea was excised and subjected to image analysis for corneal edema, real-time semiquantitative PCR (qPCR) and immunohistochemistry for oxidative stress marker. Corneal edema was significantly less and the increases in anti-oxidative mRNA expressions were significantly suppressed in the H2 group. In addition, corneal endothelial cell expressions of oxidative stress markers were significantly lower in the H2 group. In conclusion, H2 dissolved in the ocular irrigating solution protected corneal endothelial cells from phacoemulsification-induced oxidative stress and damage.
  • Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (B)
    Date (from‐to) : 2013/04 -2016/03 
    Author : Tsuyoshi Shimamura; Taketomi Akinobu; Fukai Moto; Kimura Taichi; Nishikawa Yuji; Ohsawa Ikuro
     
    We succeeded to obtain the "Proof of Concept" that combined therapy against the marginal grafts, including steatosis and DCD liver, could reduce ischemia and reperfusion injury. Using novel preservation solution, perfusate for machine perfusion, and hydrogen gas, graft weight gain and intrahepatic resistance during CS , HOPE and/or reperfusion were well controlled. Although bile production was not fully resucitated in DCD graft subjected to extended cold preservation, it would be resolved by perfusion from the artery. We believe that our method would become a choice to utilize the marginal grafts, and thereby resolving, at least to some extent, organshortage.
  • Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C)
    Date (from‐to) : 2012/04 -2015/03 
    Author : OHSAWA Ikuroh
     
    Molecular hydrogen (H2) selectively reduces toxic reactive oxygen species including hydroxyl radical and ameliorates oxidative stress-induced injuries in vivo. However, precise mechanisms underlying the remarkable effect with a small amount of H2 remain unclear. In the current study, we investigated the effect of H2 on mitochondria in cultured neuroblastoma SH-SY5Y and found that H2 enhanced mitochondrial membrane potential and cellular ATP accompanying a decrease in reduced glutathione and an increase in superoxide. Pretreatment of cells with H2 suppressed the H2O2-induced cell death, whereas posttreatment did not. An increase in antioxidative enzymes of the pretreated cells indicated the possibility that a mild stress with H2 induce an increased resistance to exacerbated oxidative stress. We propose here that H2 functions both as a radical scavenger and “a mitohormetic effector” against oxidative stress.
  • Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C)
    Date (from‐to) : 2010 -2012 
    Author : TERASAKI Yasuhiro; OHSAWA Ikuro; FUKUDA Yuh; TERASAKI Mika
     
    Various lung injuries such as irradiation or anticancer drug induced injuries are reportedly initiated and sustained by oxidative stress (reactive oxygen species:ROS), especially hydroxyl radicals (-OH), which are the primary cause of the damage. As yet,no ideal ROS scavenge therapy has been established. Because H2 was recently reported as an efficient antioxidant that diffuses rapidly across cell membranes, selectively reduces -OH, and suppresses oxidative stress-induced injury with no known toxicity, we studied the possibility that H2 could protect against irradiation or anticancer drug induced lung damage. We show here that H2 scavenged -OH and protected against apoptotic damage related to oxidative stress induced by irradiation and anticancer drug in lung epithelial cells and in lungs of mice. H2 treatment will thus be valuable for protection against oxidative stress induced various lung diseases.
  • Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C)
    Date (from‐to) : 2009 -2011 
    Author : OHARAZAWA Hideaki
     
    This study was conducted to explore the neuroprotective effect of H_2-loaded eye drops on retinal ischemia-reperfusion(I/R) injury in rats. H_2-loaded eye drops reduced the number of retinal apoptotic and oxidative stress marker. positive cells and prevented retinal thinning with an accompanying activation of Muller glia, astrocytes, and microglia. The results suggest that H_2-loaded eye drops are a highly useful neuroprotective and antioxidative therapeutic treatment for acute retinal I/R injury.
  • Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (B)
    Date (from‐to) : 2008 -2010 
    Author : OHTA Shigeo; OHSAWA Ikuroh; KAMIMURA Naomi
     
    Persistent oxidative stress is one of the major causes of most lifestyle-related diseases, cancer and the aging process. We have proposed that molecular hydrogen (H2) has potential as a "novel" antioxidant in preventive and therapeutic applications [Ohsawa et al., Nat Med. 2007 : 13 ; 688-94]. H2 has a number of advantages as a potential antioxidant : H2 rapidly diffuses into tissues and cells, and it is mild enough neither to disturb metabolic redox reactions nor to affect reactive oxygen species (ROS) that function in cell signaling, thereby, there should be little adverse effects of consuming H2. There are several methods to ingest or consume H2, including drinking H2-dissolved water (hydrogen water). In this study, we have shown that model mice for dementia, diabetes, and arteriosclerosis were improved by drinking hydrogen
  • Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C)
    Date (from‐to) : 2008 -2010 
    Author : OHSAWA Ikuroh; OHTA Shigeo; MIKAMI Toshio
     
    We investigated the effect of molecular hydrogen on a stress-induced brain dysfunction including cognitive and memory impairment, depression, and anxiety. We have recently reported that molecular hydrogen reduces oxidative stress and can penetrate the blood-brain barrier to protect neurons. Oxidative stress is widely accepted as a contributor to neuronal vulnerability and prevents hippocampal neurogenesis, which may be involved in cognitive function. The main results in this study have been published in Neuropsycopharmacology. Immobilization stress was given to mice 10 hours per day for 6 weeks. Water containing high concentration of hydrogen molecule (hydrogen water) was available ad libitum. We examined learning and memory ability using the passive avoidance test, novel recognition test and Morris water maze, and found that hydrogen water improved a stress-induced decline of cognitive function. Hydrogen water also restored the neurogenesis declined by restraint stress and reduced oxidative stress in the brain. Furthermore, we made depression-model mice with a combination of mild stress and did not find any effect of hydrogen water on this model, whereas hydrogen water restored a decline of neurogenesis. Precise molecular mechanisms concerning the effect of molecular hydrogen on neuronal cells is under investigation.
  • Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C)
    Date (from‐to) : 2008 -2010 
    Author : KAMIMURA Naomi; OHSAWA Ikuroh; OHTA Shigeo
     
    Oxidative stress is one of the causes of diabetes. While some ROS (reactive oxygen species) such as NO play physiological roles. Thus, cytotoxic radicals such as ?OH must be neutralized without compromising the essential biological activities of physiologically beneficial ROS. In this study we showed that consumption of molecular hydrogen markedly reduces hepatic oxidative stress levels and improves hyperglycemia and obesity in diabetes model mice.
  • 日本学術振興会：科学研究費助成事業 萌芽研究
    Date (from‐to) : 2007 -2008 
    Author : 太田 成男; 大澤 郁朗
     
    組織が虚血再灌流(I/R)に曝露されると、再灌流の早期段階でROSが大規模に生成され、肝、脳、心臓および腎など様々な臓器の組織に深刻な障害を引き起こす。これまで酸化ストレスによる1/R傷害は基礎研究および臨床研究の重要な焦点とされてきた。 I/R誘発性臓器障害の考えられる基礎的な機序は、多くの因子が関与し、相互依存的であり、低酸素症、炎症反応およびフリーラジカル障害に関与している。I/R傷害の病因は未だ解明されていないが、酸素フリーラジカルが重要な役割を担っていることは明らかである。それゆえI/R傷害への臨床的対応としては、フリーラジカル・スカベンジャーが実用的であると考えられている。実際に、これまでにもnicaraven, MCL-186,MESNA,およびαトコフェロールとGdCl_3などの多くの薬剤が、I/R傷害を予防するためのスカベンジャーとして試みられてきた。 私たちは、2007年に水素分子がラットの中脳動脈閉塞モデルを用いた研究で、水素分子が治療的抗酸化活性を呈することを報告した。また、昨年は、肝臓の虚血再灌流障害が、水素ガス吸引によって軽減されること、ヘリウムでは効果がなかったことを示した。虚血再灌流障害で、最も患者が多く、適用可能性が高いのは心筋梗塞であると予測されるので、心筋の虚血再灌流障害に対する水素ガスの吸引効果をラットモデルを用いて調べた。 結果は、水素ガスを吸引させると虚血状態でも心臓内に水素が浸透しることが確認できた。また、心筋梗塞モデルラットで水素ガスを吸引させることで、梗塞層が小さくなり、心機能の低下も抑制された。分離した心臓を用いても虚血再灌流障害を水素は抑制することが明らかとなった。
  • 日本学術振興会：科学研究費助成事業 特定領域研究
    Date (from‐to) : 2006 -2007 
    Author : 太田 成男; 大澤 郁朗; 上村 尚美
     
    大部分のがん細胞のミトコンドリアDNA(mtDNA)には、体細胞変異が蓄積している。発がん初期段階でmtDNAに変異が生じると変異mtDNAと正常mtDNAが混在した状態から、細胞内で変異mtDNAだけの状態に移行し、ついで変異mtDNAだけをもつ細胞のみの集団に移行する.本研究の目的は、(1)発がんにおける過程におけるmtDNA体細胞変異の蓄積の分子機構を明らかにする。(2)発がん時における変異mtDNAが原因のアポトーシス抑制機構を明らかにする。(3)ミトコンドリア機能が低下するにもかかわらず、細胞増殖が促進される機構を明らかにする。 患者のがん組織のミトコンドリアDNAの全塩基配列を決定し、がん細胞と正常細胞のミトコンドリアDNAの配列の違いを比較し、抗癌剤耐性との相関関係を調べ、正の相関関係があることを明らかにした。また、抗癌剤耐性の細胞株のミトコンドリアDNA塩基配列を決定し、そのミトコンドリアDNAを別細胞に移植し、抗癌剤耐性が消失することから抗癌剤耐性はミトコンドリアDNAに存在することを明確にした。 さらに抗癌剤耐性細胞を作り出し、その60%にはミトコンドリアDNAの変異が生じており、ミトコンドリアを移植する方法によって抗癌剤耐性が消失することから、これもミトコンドリアに抗癌剤耐性の原因があることを明確にした。 抗癌剤はアポトーシスを誘導するので、ミトコンドリアDNAの変異によってアポトーシスが阻害され、がん細胞にミトコンドリアDNAの変異が蓄積することを明確にした。さらに、ミトコンドリア機能が低下することと相関していることを明らかにした。乳酸が上昇しているので、ミトコンドリア機能を代償するために解糖系が亢進していることを明らかにした。
  • Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C)
    Date (from‐to) : 2005 -2007 
    Author : OHSAWA Ikuroh; KAMIMURA Naomi; OHTA Shigeo
     
    Only a few human neuroblastoma cell-lines maintain the potential for neuronal differentiation. We noticed by chance that differentiable human neuroblastomas including SH-SY5Y have a specific mutation in the dihydrolipoamide-succinyltransferase (DLST) gene, whose product is a component of the α-ketoglutarate-dehydrogenase complex involved in mitochondrial energy metabolism. Here we demonstrate a requirement of energy-reduction for neuronal differentiation by restoring SH-SY5Y with the wild-type DLSTgene. Up-regulation of DLST activity increased mitochondrial membrane potential and impaired neuronal differentiation. The impairment was partially rescued by treatment with inhibitors of energy metabolism. Additionally, down-regulation of DLST expression with small interfering RNA enhanced neuronal differentiation in rat pheochromocytoma PC12 and primary cultured neocortical cells. We further found that mitochondrial membrane potential among SH-SY5Y cells was heterogeneous and lower in neuroblastic type cells. It is thus concluded that the casual mutation conferred the potential for differentiation on the neuroblastomas, suggesting that such mutants had been preferentially selected as a model cell-line for neuronal differentiation. The energy-reduction seemed to contribute to the differentiation by preventing cell-death. These results provide a new insight into neuronal differentiation.
  • Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C)
    Date (from‐to) : 2005 -2007 
    Author : AOSH Sadamitsu; OHSAWA Ikuroh; MORI Takashi
     
    The aim of this project is to ameliorate liver injury induced by drug or ischemia/reperfusion using the super anti-cell death protein FNK, which was generated from an anti-apoptotic protein Bcl-xL by substitution of three amino acid residues for another one within the same group, respectively. To introduce FNK protein in cells, FNK was genetically fused with the Protein Transduction Domain (PTD) of the HIV/Tat protein. The resultant fusion protein is named PTD-FNK. PTD-FNK was delivered into neuronal cells in the brain across the blood brain barrier, when intraperitoneally injected, to inhibit delayed neuronal cell death in hippocampus induced by global ischemia (Asoh, S., et. al., Proc. Natl. Acad. Sci. USA. 2002). In this project, we have shown that systemic injection of PTD-FNK inhibits zonal necrosis of the liver induced by carbon tetrachloride, liver degeneration caused by ethanol and dexamethasone, and an early stage of liver injury (apoptosis) induced by ischemia/reperfusion. PTD-FNK suppressed an elevation of serum ALT and AST levels induced by carbon tetrachloride. It was shown that PTD-FNK maintains mitochondrial membrane potential and confer upon HepG2 cells resistance against carbon tetrachloride and TNFα. These results encouraged us to apply PTD-FNK for other experimental disease models. PTD-FNK successfully reduced brain and heart injuries caused by ischemia/reperfusion. Furthermore, PTD-FNK was delivered into inner ear cross the blood-inner ear barrier to ameliorate aminoglycoside-induced hearing loss. It was also shown that PTD-FNK enhances transplantation efficiency of bone marrow mononucleocytes and ameliorates neuronal damage in amyotrophic lateral sclerosis model, acute lung injury caused by lipopolysaccharide, and alopecia caused by anti-cancer drugs. The studies on protein therapy using PTD-FNK have been published in many international journals.
  • Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research
    Date (from‐to) : 2005 -2006 
    Author : TAKAMATSU Ken; HAMANOUE Makoto; OSAWA Ikrou
     
    p38 Mitogen-activated protein kinase (p38 MAPK) is expressed in the oligodendrocyte lineage, and its activity has been implicated in the proliferation and transition of early progenitors into late progenitors. Although p38 MAPK expression has been found in the myelin sheath, however, its role in mature oligodendrocytes remains unknown. In the present study, in order to address the role of p38 MAPK in mature oligodendrocytes, we analyzed 1. the expression of p38 MAPK in the mature oligodendrocytes, 2. the effects of p38 MAPK inhibition on the survival of mature oligodendrocytes, and 3. the effect of introduction of HIV-TAT-fused p38 MAPK protein on the survival of mature oligodendrocytes. 1. Immunocytochemical and Western blot analysis revealed that mature oligodendrocytes express p38 MAPK, and a part of p38 MAPK is an active form. 2. The inhibition of p38 MAPK with specific inhibitors decreased the number of cultured mature oligodendrocyte by the induction of apoptosis, but did not alter the number of oligodendrocyte progenitor cells. These results indicate that p38 MAPK is essential for the mature oligodendrocyte survival. 3. The fusion proteins containing a cell-permeable TAT-peptide of human immunodeficiency virus could be efficiently introduced into mature oligodendrocyte. TAT-fusion protein with a dominant negative form of p38 MAPK caused the decrease in the number of mature oligodendrocyte, indicating the survival of mature oligodendrocyte can be directly regulated by protein transduction of TAT-p38 MAPK fusion proteins. We are investigating the mechanism of the survival of mature oligodendrocyte in vivo and in vitro using the cell permeable-TAT-p38 MAPK system.
  • 日本学術振興会：科学研究費助成事業 特定領域研究
    Date (from‐to) : 2005 -2005 
    Author : 太田 成男; 宮戸 靖幸; 大澤 郁朗; 上村 尚美
     
    (1)癌細胞におけるミトコンドリアDNAの体細胞変異の蓄積 多くの癌組織、癌細胞のmtDNAに体細胞変異が蓄積していることが明らかにされている。しかし、この体細胞変異の蓄積が、癌の形成過程の二次的結果として生じているのか、癌形成の原因のひとつとして、癌の促進に関与したために変異mtDNAをもつ癌細胞が多くなったのか、明らかではなかった。 (2)サイブリドを用いたmtDNAの役割の研究 細胞には核ゲノムとミトコンドリアゲノムが共存しているので、mtDNAの役割を明らかにするためには、核が共通でmtDNAだけが異なる細胞を比較しなければならない。そこで、mtDNAを消失しているHeLa細胞と脱核した細胞質を融合し、核が共通で正常mtDNAをもつ細胞と変異mtDNAをもつ細胞を作製した。この融合細胞を細胞質の融合であるのでサイブリド(cybrid)と呼ぶ。このサイブリドを用いて、mtDNAの癌の増殖速度へ対する効果とアポトーシ耐性効果、抗癌剤への耐性効果を調べた。 (3)癌の増殖に対する変異mtDNAの効果 変異mtDNAを持つサイブリド細胞と正常mtDNAもつサイブリド細胞をヌードマウスに移植すると、移植直後からの腫瘍体積の増加が正常mtDNAをもつサイブリド細胞より速く、mtDNA変異が癌増殖促進に寄与していることが示された。変異mtDNAをもつサイブリドでは、自発的におきるアポトーシス頻度が低下しており、アポトーシスの頻度の低下が見かけ上の増殖促進に寄与していることが示唆された。 (4)抗癌剤へ対する耐性獲得とmtDNA変異 mtDNA変異によってアポトーシスが抑制されたので、抗癌剤への耐性へのmtDNAの役割をサイブリドを用いて解析した。変異mtDNAをもつサイブリドは正常mtDNAをもつサイブリドよりも抗癌剤耐性であった。
  • Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C)
    Date (from‐to) : 2002 -2004 
    Author : ASOH Sadamitsu; OHTA Shigeo; KATSURA Ken-ichiro; OHSAWA Ikuroh
     
    Preventing massive cell death is an important therapeutic strategy for various injuries and disorders. Few practical therapies have been explored for clinical applications to ischemic cerebral infarction. Protein therapeutics has the advantage of delivering proteins in a short period of time. We have engineered the anti-apoptotic bcl-x gene to generate the super anti-apoptotic factor, FNK, with a more powerful cytoprotective activity. In this project, we fused the protein transduction domain(PTD) of the HIV/Tat protein to FNK, and used the construct in an animal model of ischemic brain injury. When added into culture media of human neuroblastoma cells and rat neocortical neurons, PTD-FNK rapidly transduced into cells to protect the neuroblastomas and neurons from staurosporine-induced apoptosis and glutamate-induced excitotoxicity, respectively. PTD-FNK affected the cytosolic movement of calcium ions, which may relate to its neuroprotective action. When injected intraperitoneally into gerbils, PTD-FNK prevented delayed neuronal death in the hippocampus caused by transient global ischemia. When a focal ischemic model rat was intravenously post-treated with the PTD-FNK protein, ischemic injury was improved in terms of infarction volume (total and cortical) and neurological symptoms. Interestingly, combined injection of PTD-FNK and an immunosuppressant FK506 showed the stronger protective effect to reduce the infarction volume, exhibit a continuous effect for 1 week after the treatment and extend the therapeutic window by 3 hr. These results suggest that PTD-FNK has a potential for clinical utility as a novel protein therapeutic strategy to prevent cell death in the brain. During performing the project, we, in addition, revealed that PTD-FNK penetrates the dense matrix of cartilage to reach chondrocytes in slice culture of cartilage and protect the chondrocytes from death induced by death stimuli, prevents necrosis and hepatic injury with zonal death induced by carbon tetrachloride, and protect cells from death induced by freezing and thawing.
  • Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C)
    Date (from‐to) : 2002 -2004 
    Author : OHSAWA Ikuroh; OHTA Shigeo; ASOH Sadamitsu
     
    Precise mechanism for survival of neurons in central nervous system remains to be elucidated. It includes many anti-apoptotic proteins. Survivin is a member of the inhibitor-of-apoptosis protein. Its expression is determined in the developing embryo and in rapidly dividing cells including many human cancers. Previous studies demonstrated that survivin is expressed at G2/M in a cell-cycle dependent manner and associated with kinetochores of metaphase chromosomes and the central spindle midzone at anaphase, suggesting that survivin participates as a chromosomal passenger protein in cleavage furrow formation. Forced expression of survivin counteracted cell death induced by various apoptotic stimuli, whereas interference with the expression or function of survivin by a dominant negative form or antisense of survivin caused spontaneous apoptosis and multiple cell division defects, suggesting that survivin acts both as a mitotic regulator and as a cytoprotective factor at cell division. Then, we prepared polyclonal anti-survivin serum to investigate the expression of survivin in central nervous system. At first, we stained primary cultured cells derived from neocortex in rat embryo and found that survivin was expressed in MAP2 positive differentiated neurons. The expression was also observed in a part of astrocytes. Immunological staining of mouse brain section confirmed the expression of survivin in neurons. These results suggested that survivin might be included in anti-apoptotic machineries of differentiated neurons. On the other hand, we examined whether survivin expression in human gliomas would be a correlative of prognosis, and found that higher expression of survivin was correlated with the survival of gliomas. Thus, immunostaining of glioma with anti-survivin serum would be a powerful tool for a clinical prognosis. We also investigated whether urine surviving is useful for diagnosing bladder cancer, and found that ELISA system for survivin might be a useful tumor marker.
  • Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (B)
    Date (from‐to) : 2000 -2002 
    Author : OHTA Shigeo; OHOSAWA Ikurho; KANAMORI Takashi; ASHO Sadamitsu
     
    Mitochondrial aldehyde dehydrogenase 2 (ALDH2) is involved in ethanol metabolism by playing a major role in acetaldehyde detoxification : A polymorphism of the ALDH2 gene is specific to north Asians. Sensitivity to ethanol is highly associated with this polymorphism (ALDH2^*2 allele), which is responsible for a deficiency of ALDH2 activity. We at first show that this deficiency influences the risk for late-onset Alzheimer's disease (LOAD) by a case-control study in a Japanese population. In a comparison of 447 patients with sex, age and region-matched non-demented controls, the genotype frequency for carrying the ALDH2^*2 allele was significantly higher in the patients than in the controls (p=0.001). Next, we examined the combined effect of the ALDH2^*2 and apolipoprotein E 4 allele (APOE-e4), which has been confirmed be a risk factor for LOAD. The ALDH2^*2 allele more significantly affected frequency and onset-age in patients with APOE-e4 than without. These results indicate that the ALDH2 deficiency is a risk factor for LOAD, acting synergistically with the APOE-e allele. Next, to elucidate the molecular mechanism involved, we obtained ALDH2-deficient cell lines by introducing mouse mutant Aldh2 cDNA into PC12 cells. We speculate that ALDH2 may function to oxidize toxic aldehyde derivatives. Then, we found that the ALDH2-deficient transfectants were highly vulnerable to exogenous 4-hydroxy-2-nonenal, an aldehyde derivative generated from peroxidized fatty acids. In addition, the ALDH2-deficient transfectants were sensitive to oxidative insult induced by antimycin A, accompanied by an accumulation of proteins modified with 4-hydroxy-2-nonenal. Mitochondrial ALDH2 functions as a protector against oxidative stress.
  • 日本学術振興会：科学研究費助成事業 特定領域研究(C)
    Date (from‐to) : 2001 -2001 
    Author : 太田 成男; 大澤 郁朗; 金森 崇; 麻生 定光
     
    ミトコンドリアゲノムは核ゲノムとは空間的に独立して存在しており、その挙動は多くの疾患において重要な役割を果たしていることが明らかになりつつある。糖尿病全体の1%におよぶ患者にはmtDNAの変異が蓄積していることがすでに報告されている。しかし、この変異が糖尿病にどのように影響を及ぼすのかは不明である。糖尿病患者と健常人において、ヘテロプラズミー変異の比率の相関関係を定量的に検討し、糖尿病とmtDNAのヘテロプラズミー変異の因果関係を明らかにし、糖尿病の危険因子となりうることを証明するのが本研究の目的である。 (1)糖尿病の原因のミトコンドリアtRNAの点変異によって、アンチコドンの塩基修飾が変化して、mRNAへの親和力が低下し、蛋白合成が正常に行われなくなることを明らかにした。この結果は、わずかな量の変異mtDNAでも不完全が蛋白を合成することによって細胞に影響を及ぼす可能性を示した。 (2)mtDNAのヘテロプラズミー変異の検出法を確立した。この方法の確立によって、ヘテロプラズミー変異を定量的に検討することが可能になった。 糖尿病患者の血液細胞において、体細胞変異が健常人よりも4倍多いことを明らかにした。さらに、多変量解析により、糖尿病の羅患期間に依存してmtDNAの塩基番号3243変異が増加することを明らかにした。 ミトコンドリアtRNA遺伝子変異によって生じるミトコンドリア機能低下の分子機構は多くの研究室で研究されているが、当研究室がはじめて分子機構を明らかにした。mtDNAのヘテロプラズミー変異と糖尿病の関連については多数の報告があるが、微量なヘテロプラズミー変異の定量についての研究はなく、定量的解析は当研究がリードしている。
  • 水素分子の生体における機能解析、医学への応用．加齢に伴う神経変性疾患における酸化ストレスの影響．
  • Hydrogen as a therapeutic antioxidant, its molecular and cellular mechanisms

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