Studies on molecular mechanisms and artificial regulation of maturation and aging of mammalian oocytes

哺乳动物卵母细胞成熟和衰老的分子机制及人工调控研究

• 批准号: 10660267
• 负责人: NAITO Kunihiko
• 金额: $ 2.24万
• 依托单位: The University of Tokyo
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1998
• 资助国家: 日本
• 起止时间: 1998 至 1999
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-10660267/
• 关键词: MPF; oocyte aging; fragmentation; caffeine; vanadate; matured porcine oocyte; 豚卵子; 老化; p34^<cde2>; サイクリンB

Aims of the present study are clarifying the cytoplasmic changes during mammalian oocyte aging at molecular levels and subsequently regulating this process artificially. The present study might contribute to prevent the deterioration of oocyte qualities derived from elongated manipulation period of the in vitro matured mammalian oocytes, used for such as reproductive and developmental technologies.The gradual decrease of maturation promoting factor (MPF) activity during oocyte aging has been reported previously. The present study revealed that the molecular mechanism of the decrease of MPF activity during oocyte aging was completely different from that during oocyte activation Although the decrease of MPF activity at oocyte activation was attributed to the rapid degradation of cyclin B, a regulatory subunit of MPF, the levels of MPF subunits, both p34ィイD1cdc2ィエD1 and cyclin B, were not significant]y changed during oocyte aging but the gradual accumulation of hyperphosphorylated inactiv … More e MPF, so- called pre-MPF, was observed. In order to confirm that this hyperphosphorylation of MPF was the course of the decreased MPF activity in aged oocytes, I treated the fresh and aged oocytes, respectively, with vanadate and caffeine which modulated the phosphorylation states of MPF. These experiments suggested that the change of phosphorylation state of MPF was the main course of the decrease of the activity during oocyte aging and that the MPF activity in aged oocytes could be regulated at lease in part by vanadate and caffeine treatment. These findings might be valuable as the first report showing the possibility of artificial regulation of MPF activity. Furthermore, I revealed that these treatments could change the rates of spontaneous activation and fragmentation of oocytes, both are the parameters of oocyte aging.In summary, the present study showed cytoplasmic changes during mammalian oocyte aging at molecular levels and proposed a simple method for artificial regulation of this process at least partially. Less

本研究旨在从分子水平阐明哺乳动物卵母细胞老化过程中细胞质的变化，并对这一过程进行人工调控。哺乳动物卵母细胞体外成熟过程中促成熟因子（Maturity Promoting Factor，MPF）活性逐渐降低，这一现象已被报道，但由于操作时间过长，导致卵母细胞的质量下降，本研究将有助于防止卵母细胞的老化。本研究揭示了卵母细胞老化过程中MPF活性下降的分子机制与卵母细胞激活过程中MPF活性下降的分子机制完全不同。虽然卵母细胞激活过程中MPF活性下降是由于MPF调节亚基cyclin B的快速降解，但MPF亚基p34 β D1 cdc2 β D1和cyclin B的水平均下降，在卵母细胞老化过程中， ...更多信息 e观察到MPF，即所谓的前MPF。为了证实这种MPF的过度磷酸化是老年卵母细胞MPF活性降低的过程，我分别用钒酸盐和咖啡因处理新鲜和老年卵母细胞，它们调节MPF的磷酸化状态。提示MPF磷酸化状态的改变是卵母细胞老化过程中MPF活性下降的主要原因，钒酸盐和咖啡因对老化卵母细胞MPF活性有一定的调节作用。这些发现可能是有价值的，因为第一份报告显示了人为调节强积金活动的可能性。本研究从分子水平揭示了哺乳动物卵母细胞老化过程中胞质的变化，并提出了一种人工调控的简单方法。少

项目成果

Seiki Haraguchi: "Phosphate exposure during the 1-cell and early 2-cell stages induces a time-specific decrease in cyclin B and cdc25B mRNAs in AKR/N mouse embryos in vitro"Zygote. 7. 87-93 (1999)

Seiki Haraguchi：“1 细胞和早期 2 细胞阶段的磷酸盐暴露会诱导体外 AKR/N 小鼠胚胎中细胞周期蛋白 B 和 cdc25B mRNA 的时间特异性减少”Zygote。

Koji Sugiura: "Analysis of the germinal vesicle requirement for the activation of MPF in maturation of porcine oocytes"J. Mamm. Ova Res.. 16. 130-134 (1999)

Koji Sugiura：“猪卵母细胞成熟过程中 MPF 激活的生发囊泡需求分析”J.

Kazuhiro Kikuchi: "Inactivation of p34^<cdc2> kinase by the accumulation of its phosphorylated forms in porcine oocytes matured and aged in vitro"Zygote. 7. 173-179 (1999)

Kazuhiro Kikuchi：“通过在体外成熟和老化的猪卵母细胞中磷酸化形式的积累，p34^<cdc2>激酶失活”Zygote。

KIKUCHI, K., NAITO, K., NOGUCHI, J., SHIMADA, A., KANEKO, H., YAMASHITA, M., TOJO, H. and TOYODA Y.: "Inactivation of p34ィイD1cdc2ィエD1 kinase by the accumulation of its phosphorylated forms in porcine oocytes matured and aged in vitro."Zygote. 7. 173-179 (

KIKUCHI, K.、NAITO, K.、NOGUCHI, J.、SHIMADA, A.、KANEKO, H.、YAMASHITA, M.、TOJO, H. 和 TOYODA Y.：“p34D1cdc2D1 激酶因磷酸化积累而失活在体外成熟和老化的猪卵母细胞中形成。“Zygote. 7. 173-179 (

Kunihiko Naito: "Establishment of a small-scale western blotting system named as "micro-western blotting" for mammalian ova analysis"J. Mamm. Ova Res.. 16. 154-157 (1999)

Kunihiko Naito：“用于哺乳动物卵子分析的小型蛋白质印迹系统的建立，名为“微型蛋白质印迹””J.