哺乳類卵胞の選択的死滅制御機構とその人為支配による潜在的卵巣卵の利用

2003 年度 実績報告書

• 研究課題/領域番号: 13GS0008
• 研究機関: 京都大学
• 研究代表者: 眞鍋 昇 京都大学, 農学研究科, 助教授 (80243070)
• 研究分担者: 杉本 実紀 京都大学, 農学研究科, 助手 (20243074) ; 宮野 隆 神戸大学, 農学部, 教授 (80200195) ; 酒巻 和弘 京都大学, 生命科学研究科, 助教授 (20271017) ; 若山 照彦 理科学研究所神戸研究所, 発生再生科学総合研究センター, チームリーダー (40360672)
• キーワード: 哺乳類 / 卵巣 / 卵胞 / 選択的死滅 / アポトーシス / 細胞死受容体 / カスパーゼ / 異種移植

研究概要

脊椎動物の生殖戦略は多様である。哺乳類は、硬骨魚類のサケのように一生に一度だけ全卵子を排卵して数千から数億の仔を生み、その仔の中で優れたものが生き残る戦略と異なり、性周期毎に1から10個の卵母細胞を成熟・排卵し、極わずかの仔を産み、その仔を大切に育くむ戦略をとって繁栄してきた。哺乳類卵巣には胎児期に減数分裂ディプロテン期で休止した卵母細胞が5〜100万個含まれる。性成熟後は性周期毎に卵母細胞は卵胞ともに発育・成熟する。この時卵巣内では最終的に排卵にいたる卵子の100倍以上の卵胞が発達を開始するが、発育・成熟の過程で99%以上の卵母細胞が選択的に死滅し、1%未満の二次卵母細胞のみが排卵に至る。卵胞の選択的死滅は、より優秀で強靭な子孫を残す戦略として重要であるが、これの分子制御機構は未解明である。本研究は、哺乳類において卵胞の選択的死滅を制御している分子機構を解明し、これをもとに細胞死を制御している遺伝子の発現をin vivo制御することで卵胞の発達を人為的に支配し、卵巣内に潜在する卵母細胞の有効利用を実現することを目指している。今年度は、新規な細胞死受容体を認識するユニークなモノクロナル抗体を用いてデコイ受容体を発見・同定した。卵巣で発現していることが知られている細胞死受容体であるFasの細胞内シグナル伝達系を担うカスパーゼ系の細胞内シグナル伝達を蛍光エネルギー移動(fluorescence resonance transfer:FRET)法にてリアルタイムにin situ解析できる新規な手法を開発した。また多面的機能をもつinterleukin-6が卵胞選抜に重要に関わっていることを示唆する知見を得た。また細胞内アポトーシス阻害因子(cFLIP)を見いだし、遺伝子とアミノ酸配列を決定した。併行して、ノックアウトしたマウス、免疫不全マウスの腎に移植(異種移植)したウシ、ブタなどの大型哺乳類の未成熟な卵胞を用いて、アポトーシスシグナル伝達系の分子制御機構の解析を進めた。

研究成果

• [文献書誌] Mizuho Nakayama, Noboru Manabe, Naoko Inoue, Toshikatsu Matsui, Hajime Miyamoto: "Changes in the expression of tumor necrosis factor (TNF)a, TNFa Receptor (TNFR) 2 and TNFR-associated factor 2 in granulosa cells during atresia in pig ovaries."Biology of Reproduction. 68(2). 530-535 (2003)
• [文献書誌] Kohji Komatsu, Noboru Manabe, Minako Kiso, Munetake Shimabe, Hajime Miyamoto: "Changes in localization of immune cells and cytokines in corpora lutea during luteolysis in murine ovaries."Journal of Experimental Zoology. 296(1). 152-159 (2003)
• [文献書誌] Kohji Komatsu, Noboru Manabe, Minako Kiso, Munetake Shimabe, Hajime Miyamoto: "Soluble Fas (FasB) regulates luteal cell apoptosis during luteolysis in murine ovaries."Molecular Reproduction and Development. 65. 345-352 (2003)
• [文献書誌] Toshikatsu Matsui, Noboru Manabe, Yasufumi Goto, Mizuho Nakayama, Naoko Inoue, Susumu Nishihara, Hajime Miyamoto: "Changes in the expression and activity of caspase-9 and Apafl in granulosa cells during follicular atresia in pig ovaries."Reproduction. 126. 113-120 (2003)
• [文献書誌] Istvan Egerszegi, Falk Schneider, Jozsef Ratky, Ferenc Soos, Laszlo Solti, Noboru Manabe, Klaus-Peter Brussow: "Comparison of luteinizing hormone and steroid hormone secretion during the peri-and post-ovulatory periods in mangalica and landrace gilts."Journal of Reproduction and Development. 49(4). 291-296 (2003)
• [文献書誌] Naoko Inoue, Noboru Manabe, Toshikatsu Matsui, Akihisa Maeda, Shynsuke Nakagawa, Satoko Wada, Hajime Miyamoto: "roles of tumor necrosis factor-related apoptosis-inducing ligand signaling pathway in granulosa cell apoptosis during atresia in pig ovaries."Journal of Reproduction and Devevelopment. 49(4). 313-321 (2003)
• [文献書誌] Satoshi Imanishi, Noboru Manabe, Hanako Nishizawa, Maki Morita, Miki Sugimoto, Mineko Iwahori, Hajime Miyamoto: "Effects of oral exposure of bisphenol A on mRNA expression of nuclear receptors in murine placentae assessed by DNA microarray."Journal of Reproduction and Devevelopment. 49(4). 329-336 (2003)
• [文献書誌] Minako Kiso, Noboru Manabe, Kohji Komatsu, Munetake Shimabe, Hajime Miyamoto: "Abnormal structural luteolysis in ovaries of the senescence accelerated mouse (SAM) : expression of Fas Ligand/Fas-mediated apoptosis signaling molecules in luteal cells."Journal of Reproduction and Devevelopment. 49(6). 457-463 (2003)
• [文献書誌] Hanako Nishizawa, Noboru Manabe, Maki Morita, Miki Sugimoto, Satoshi Imanishi, Hajime Miyamoto: "Effects of in utero exposure of bisphenol A on expression of RARα and RXRα mRNAs in murine embryos."Journal of Reproduction and Devevelopment. 49(6). 539-545 (2003)
• [文献書誌] Kozue Uchio-Yamada, Noboru Manabe, Misuzu Yamaguchi-Yamada, Yasufumi Goto, Yoshie Yamamoto, Atsuo Ogura, Hajime Miyamoto: "Changes in the localization of type I, III and IV collagen mRNAs in the kidneys of hereditary nephritic (ICGN) mice with renal fibrosis."The Journal of Veterinary Medical Science. 66(2). 123-128 (2004)
• [文献書誌] Miki Sugimoto, Noboru Manabe, Maki Morita, Kei-Ichi Sezaki, Takako Tanaka, Satoshi Imanishi, Hanako Nishizawa, Mineko Iwahori, Hajime Miyamoto: "NMR spectroscopic assessment of effects of estrogenic compounds on male reproductive organs in mice."Biology of Reproduction. 67. 186 (2003)
• [文献書誌] Ysufumi Goto, Noboru Manabe, Kozue Uchio, Misuzu Yamaguchi-Yamada, Naoko Inoue, Yoshie Yamamoto, Atsuo Ogura, Nobuo Nagano, Hajime Miyamoto: "Myofibroblastic differentiation in tubulointerstitial cells is provoked by augmented cytoplasmic Smad4 protein in hereditary nephrotic ICGN mice with chronic renal fibrosis."Cell and Tissue Research. 315. 209-221 (2004)
• [文献書誌] Miki Sugimoto, Noboru Manabe, Hajime Miyamoto: "Cryopreservation of ovarian tissue."Igakuno Ayumi. 204(13). 921-925 (2003)
• [文献書誌] Noboru Manabe, Naoko Inoue, Chiemi Tajima, Toshikatsu Matsui, Shinsuke Nakagawa, Akihisa Maeda, Hajime Miyamoto: "Monoclonal antibodis recognize novel cell death receptors of granulosa cells in pig ovaries."Biology of Reproduction. 67. 234 (2003)
• [文献書誌] Naoko Inoue, Noboru Manabe, Toshikatsu Matsui, Shinsuke Nakagawa, Akihisa Maeda, Hajime Miyamoto: "Changes in expression of fas-dependent apoptotic signaling molecules in granulosa cells during folliclar atresia in pig ovaries."Biology of Reproduction. 67. 234 (2003)
• [文献書誌] Satoshi Imanishi, Noboru Manabe, Miki Sugimoto, Hajime Miyamoto: "Changes in mRNA expression of GPRC5B/RAIG2, a novel orphan receptor, in mouse placenta during late gestation."Biology of Reproduction. 67. 154 (2003)
• [文献書誌] Noboru Manabe, Naoko Inoue, Toshikatsu Matsui, Shinsuke Nakagawa, Akihisa Maeda, Takashi Miyano, Kazuhiro Sakamaki, Miki Sugimoto, Hajime Miyamoto: "Animal Frontier Science"Hokuto Pub.Co, Kyoto. 363 (2003)
• [文献書誌] Noboru Manabe, Satoko Wada, Naoko Inoue, Toshikatu Matsui, Shinsuke Nakagawa, Akihisa Maeda, Hajime Miyamoto: "Animal Frontier Science"Hokuto Pub.Co, Kyoto. 363 (2003)
• [文献書誌] Mizuho Nakayama, Noboru Manabe, Naoko Inoue, Toshikatsu Matsui, Shinsuke Nakagawa, Akihisa Maeda, Hajime Miyamoto: "Animal Frontier Science"Hokuto Pub.Co, Kyoto. 363 (2003)
• [文献書誌] Naoko Inoue, Noboru Manabe, Toshikatsu Matsui, Shinsuke Nakagawa, Hajime Miyamoto: "Animal Frontier Science"Hokuto Pub.Co, Kyoto. 363 (2003)
• [文献書誌] Miki Sugimoto, Noboru Manabe, Maki Morita, Satoshi Imanishi, Hanako Nishizawa, Mineko Iwahori, Hajime Miyamoto: "Animal Frontier Science"Hokuto Pub.Co, Kyoto. 363 (2003)
• [文献書誌] Noboru Manabe, Yasuyoshi Azuma, Miki Sugimoto, Satoshi Imanishi, Hanako Nishizawa, Maki Morita, Hajime Miyamoto: "Animal Frontier Science"Hokuto Pub.Co, Kyoto. 363 (2003)
• [文献書誌] Noboru Manabe, Naoko Inoue, Takashi Miyano, Kazuhiro Sakamaki, Miki Sugimoto, Hajime Miyamoto: "The Ovary, 2^<nd> edition"Academic Press/Elsevier Science, Amsterdam, The Nederland. 664 (2003)
• [文献書誌] Noboru Manabe: "Veterinary Histology 3^<rd> edition"Gakusoshya Pub.Co, Tokyo, Japan. 315 (2003)
• [文献書誌] Noboru Manabe: "Biology of Reproduction 2^<nd> edition"Asakura Pub.Co, Tokyo, Japan. 315 (2003)
• [文献書誌] Noboru Manabe, Minako Kiso, Munetake Shimabe, Hajime Miyamoto: "The Senescence-Accelerated Mouse (SAM) : An Animal Model of Senescence"Elsevier Science Publishers B.V., Amsterdam, The Nederland. 448 (2004)
• [文献書誌] Toshikatu Matsui, Noboru Manabe, Yasufumi Goto, Naoko Inoue, Shinsuke Nakagawa, Akihisa Maeda, Susumu Nishihara, Hajime Miyamoto: "Animal Frontier Science"Hokuto Pub.Co, Kyoto. 363 (2003)
• [文献書誌] Misuzu Yamaguchi-Yamada, Noboru Manabe, Kozue Uchio-Yamada, Naotsugu Akashi, Yasufumi Gotoh, Masaya Nagao, Yoshie Yamamoto, Atsuo Ogura, Hajime Miyamoto: "Animal Frontier Science"Hokuto Pub.Co, Kyoto. 363 (2003)
• [文献書誌] Minako Kiso, Noboru Manabe, Kohji Komatsu, Munetake Shimabe, Hajime Miyamoto: "Animal Frontier Science"Hokuto Pub.Co, Kyoto. 363 (2003)
• [文献書誌] Noboru Manabe, Kozue Yamada-Uchio, Misuzu Yamada-Yamaguchi, Yasufumi Gotoh, Toshiharu Mori, Chinatsu Sakata, Yoshihiro Furuya, Hajime Miyamoto: "Animal Frontier Science"Hokuto Pub.Co, Kyoto. 363 (2003)
• [文献書誌] Maki Morita, Miki Sugimoto, Takako Tanaka, Satoshi Imanishi, Hanako Nishizawa, Kei-Ichi Sezaki, Mineko Iwahori, Noboru Manabe, Hajime Miyamoto: "Animal Frontier Science"Hokuto Pub.Co, Kyoto. 363 (2003)
• [文献書誌] Munetake Shimabe, Minako Kiso, Kohji Komatsu, Noboru Manabe, Hajime Miyamoto: "The Senescence-Accelerated Mouse (SAM) : An Animal Model of Senescence"Elsevier Science Publishers B.V., Amsterdam, The Nederland. 448 (2004)
• [文献書誌] Minako Kiso, Munetake Shimabe, Kohji Komatsu, Noboru Manabe, Hajime Miyamoto: "The Senescence-Accelerated Mouse (SAM) : An Animal Model of Senescence"Elsevier Science Publishers B.V., Amsterdam, The Nederland. 448 (2004)
• [文献書誌] Morikawa Masahiro, Wakoto Matsuyama, Miki Sugimoto, Maki Morita, Aki Nakayama, Noboru Manabe, Shinsuke Morisawa: "Environmental Toxicity Evaluation and Risk Management"The Environment Risk Management Group of the JSPS-VCC Program on Environmental Science, Engineering and Ethics 2000-2009. Kyoto, Japan. 159 (2003)
• [文献書誌] Noboru Manabe, Minako Kiso, Munetake Shimabe, H.Miyamoto: "Proceedings of the 2^<nd> International Conference on Senescence : The SAM model"The Council for SAM Research, Sapporo, Japan. 120 (2003)
• [文献書誌] Munetake Shimabe, Minako Kiso, Noboru Manabe, Hajime Miyamoto: "Proceedings of the 2^<nd> International Conference on Senescence : The SAM model"The Council for SAM Research, Sapporo, Japan. 120 (2003)
• [文献書誌] Minako Kiso, Munetake Shimabe, Noboru Manabe, Hajime Miyamoto: "Proceedings of the 2^<nd> International Conference on Senescence : The SAM model"The Council for SAM Research, Sapporo, Japan. 120 (2003)