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耐凍性タンパク質遺伝子等のexvivo導入による卵巣の凍結保存

Research Project

Project/Area Number 12876061
Research Category

Grant-in-Aid for Exploratory Research

Allocation TypeSingle-year Grants
Research Field Applied animal science
Research InstitutionKyoto University

Principal Investigator

宮本 元  京都大学, 農学研究科, 教授 (00026618)

Co-Investigator(Kenkyū-buntansha) 眞鍋 昇  京都大学, 農学研究科, 助教授 (80243070)
Project Period (FY) 2000 – 2001
Project Status Completed (Fiscal Year 2001)
Budget Amount *help
¥500,000 (Direct Cost: ¥500,000)
Fiscal Year 2001: ¥500,000 (Direct Cost: ¥500,000)
Keywords卵巣 / 凍結保存 / 耐凍性タンパク質 / ex vivo遺伝子導入 / リポゾーム / 細胞死受容体 / レクチン / センダイウイルスエンベロープタンパク質 / 耐凍タンパク質 / アポトーシス受容体 / Ex vivo遺伝子導入 / リポソーム法 / アポトーシス阻止因子 / NIH-3T3細胞
Research Abstract

これまで生殖器官の凍結保存を実現するため、凍結前に組織を薄切してより急速に冷卸しようとする手法が開発されたが、受精能をもつ卵子をほとんど得られなかった。そこで、器官摘出前に動脈から耐凍剤を灌流した後薄切して凍結する方法が考案されたが、これも満足のいく成績を得られなかった。遊離細胞より格段に巨大な器官の凍結保存を実現するためにはブレークスルー的技術革新が必要である。本研究は、極地に生息する動物の体液氷点を低下させたり氷晶表面に結合して結晶の成長を抑えて細胞破砕を阻止する耐凍タンパク質の遺伝子と申請者らが見いだしたアポトーシス阻止遺伝子を生殖器官にex vivo導入することによる凍害保護や細胞死阻害によって生殖器官の凍結保存を実現しようとするもので、下記の成果をあげた。Ex vivo遺伝子導入のためのリポソーム作製法の検討を進めた。卵巣等に耐凍性タンパク質遺伝子および申請者らが発見した新規アポトーシス阻害因子遺伝子を特異的に発現させるex vivo遺伝子導入法をマウスを用いて確立するために、遺伝子導入に際して遺伝情報の伝達を担う生殖細胞(卵子・精子)に外来遺伝子が不用意に導入されることを未然に防ぐべく、導入効率が高いがウイルス遺伝子を含むウイルスベクター法をとらず、ウイルス遺伝子を全く含まない「リポソーム法」を改良した「センダイウイルスエンベロープタンパク質と卵胞特異的レクチン組込型リポソーム」を作製した。導入遺伝子の安定発現を計るため、極地の節足動物のI型とIII型N型糖鎖耐凍タンパク質ならびに申請者の発見した新規アポトーシス阻害因子をコードするcDNAを合成し、大腸菌プラスミドに組み込んで大量に調製し、生殖器官に導入して、耐凍能の向上をみた。

Report

(2 results)
  • 2001 Annual Research Report
  • 2000 Annual Research Report
  • Research Products

    (22 results)

All Other

All Publications (22 results)

  • [Publications] Satoko Wada, Noboru Manabe, Hajime Miyamoto et al.: "TRAIL-decoy receptor-1 disappears in granulosa cells of atretic follicles in porcine ovaries"Journal of Reproduction and Devevelopment. 48・2. 167-173 (2002)

    • Related Report
      2001 Annual Research Report
  • [Publications] Satoko Wada, Noboru Manabe, Hajime Miyamoto et at.: "TRADD is involved in apoptosis induction in granulosa cells during atresia in pig ovaries"Journal of Reproduction and Devevelopment. 48・2. 175-181 (2002)

    • Related Report
      2001 Annual Research Report
  • [Publications] Hitoshi Okamura, Noboru Manabe et al.: "Protein tyrosine kinase expression in the porcine ovary"Molecular Human Reproduction. 7・4. 723-729 (2001)

    • Related Report
      2001 Annual Research Report
  • [Publications] Eiichi Hondo, Noboru Manabe et al.: "Molecular cloning and expression of suppressor of potassium transport defect 3 (SKD3) in rat testis"Journal of Reproduction and Devevelopment. 47・2. 173-180 (2001)

    • Related Report
      2001 Annual Research Report
  • [Publications] Minako Kiso, Noboru Manabe, Hajime Miyamoto et al.: "Abnormal accumulation of luteal bodies in ovaries of the senescence accelerated mouse (SAM)"Journal of Reproduction and Devevelopment. 47・2. 153-164 (2001)

    • Related Report
      2001 Annual Research Report
  • [Publications] Mizuho Nakayama, Noboru Manabe, Hajime Miyamoto et al.: "Changes in localization of type I, III and IV collagens and their mRNA expression during follicular atresia in bovine and porcine ovaries"Journal of Reproduction and Devevelopment. 47・1. 27-36 (2001)

    • Related Report
      2001 Annual Research Report
  • [Publications] Noboru Manabe, Hajime Miyamoto et al.: "In vivo noninvasive nuclear magnetic resonance (NMR) technique for assessment of teratogenicity induced by chemicals"Chemical Engineering. 47・1. 110-114 (2002)

    • Related Report
      2001 Annual Research Report
  • [Publications] Noboru Manabe, Hajime Miyamoto et al.: "Cell death receptor dependent regulation of granulosa cell apoptosis in porcine ovarian follicle atresia. In : Miyamoto H, Manabe N (eds), Reproductive Biotechnology : Reproductive biotechnology update and its related physiology"Hokuto Publishing Co., (Kyoto, Japan). 424 (2001)

    • Related Report
      2001 Annual Research Report
  • [Publications] Noboru Manabe,Miki Sugimoto,Maki Morita,Hajime Miyamoto et al.: "A novel in vivo 31P-nuclear magnetic resonance technique for assessment of teratogenicity induced by environmental endocrine disrupters in mice."Japanese Journal of Clinical Medicine. 58・2. 108-111 (2000)

    • Related Report
      2000 Annual Research Report
  • [Publications] Noboru Manabe,Miki Sugimoto,Maki Morita,Hajime Miyamoto et al.: "In vivo noninvasive nuclear magnetic resonance techinque for assessment of teratogenicity induced by environmental pollutants."Environment Measurement and Chemical Analysis. 27・1. 60-66 (2000)

    • Related Report
      2000 Annual Research Report
  • [Publications] Noboru Manabe,Yasuyoshi Azuma,Hajime Miyamoto et al.: "Effects of the mycelial extract of cultured Cordy ceps sinensis on in vivo hepatic energy metabolism and blood flow in dietary hypoferric anemic mice."British Journal of Nutrition. 83・2. 197-204 (2000)

    • Related Report
      2000 Annual Research Report
  • [Publications] Noboru Manabe,Akira Myoumoto,Chiemi Tajima,Hajime Miyamoto et al.: "Immunochemical characteristics of a novel cell death receptor and a decoy receptor on granulosa cells of porcine ovarian follicles."Cytotechnology. 33・2. 189-201 (2000)

    • Related Report
      2000 Annual Research Report
  • [Publications] Mizuho Nakayama,Noboru Manabe,Hajime Miyamoto et al.: "Species specific differences in apoptotic cell localization in granulosa and theca interna cells during follicular atresia in porcine and bovine ovaries."Journal of Reproduction and Development. 46・2. 147-156 (2000)

    • Related Report
      2000 Annual Research Report
  • [Publications] Susumu Nishihara,Noboru Manabe,Hajime Miyamoto et al.: "Changes in cell adhesion molecules during follicular atresia in porcine ovaries."Journal of Reproduction and Development. 46・3. 325-334 (2000)

    • Related Report
      2000 Annual Research Report
  • [Publications] Mizuho Nakayama,Noboru Manabe,Hajime Miyamoto et al.: "Changes in localization of type I, III and IV collagens and their mRNA expression during follicular atresia in bovine and porcine ovaries."Journal of Reproduction and Development. 47・1. 27-36 (2001)

    • Related Report
      2000 Annual Research Report
  • [Publications] Kozue Uchio,Noboru Manabe,Kotaro Tamura,Makoto Miyamoto et al.: "Decreased matrix metalloproteinase activity in the kidneys of hereditary nephrotic mice (ICGN strain)."Nephron. 86・1. 145-141 (2000)

    • Related Report
      2000 Annual Research Report
  • [Publications] Kotaro Tamura,Noboru Manabe,Kozue Uchio,Makoto Miyamoto et al.: "Characteristic changes in carbohydrate profile in the kidneys of hereditary nephrotic mice (ICGN strain)."The Journal of Veterinary Medical Science. 52・2. 379-390 (2000)

    • Related Report
      2000 Annual Research Report
  • [Publications] Noboru Manabe,Akiko Kinoshita,Kozue Yamada-Uchio et al.: "Changes in quantitative profile of extracellular matrix components in the kidneys of rats with adriamycin-induced nephropathy."The Journal of Veterinary Medical Science. 53・1. 125-133 (2001)

    • Related Report
      2000 Annual Research Report
  • [Publications] Miki Sugimoto,Noboru Manabe,Hajime Miyamoto.: "Future functional-morphological studies in embryology."Journal of Reproduction and Development. 46・1. 71-77 (2000)

    • Related Report
      2000 Annual Research Report
  • [Publications] Noboru Manabe,Hajime Miyamoto et al.: "Immunohistochemical quantification of fast-myosin in upper hind limb muscles of the pig."Animal Science Journal. 72・2. 154-163 (2000)

    • Related Report
      2000 Annual Research Report
  • [Publications] Miki Sugimoto,Noboru Manabe,Hajime Miyamoto.: "Development of infantile rat ovaries autotransplanted after cryopreservation by vitrification."Theriogenology. 53・11. 1093-1103 (2000)

    • Related Report
      2000 Annual Research Report
  • [Publications] Noboru Manabe,Akira Myoumoto,Chiemi Tajima,Hajime Miyamoto et al.: "A novel cell death receptor and a decoy receptor on granulosa cells of porcine ovarian follicles."Tohoku University Press, Sendai, Japan.. 357 (2000)

    • Related Report
      2000 Annual Research Report

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Published: 2001-04-01   Modified: 2016-04-21  

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