Int. J. Dev. Biol. 53: 605 - 613 (2009)

https://doi.org/10.1387/ijdb.082607ah

Vol 53, Issue 4

Large-scale production of growing oocytes in vitro from neonatal mouse ovaries

Open Access | Technical Article | Published: 1 May 2009

Arata Honda, Michiko Hirose, Kimiko Inoue, Hitoshi Hiura, Hiromi Miki, Narumi Ogonuki, Michihiko Sugimoto, Kuniya Abe, Mito Kanatsu-Shinohara, Tomohiro Kono, Takashi Shinohara and Atsuo Ogura*

¹Bioresource Center, RIKEN, Tsukuba, Ibaraki, ²Department of BioScience, Tokyo University of Agriculture, Setagaya-ku, Tokyo, ³Department of Molecular Genetics, Graduate School of Medicine, Kyoto University, Kyoto, ⁴Graduate School of Life and Environmental Sciences, University of Tsukuba, Tsukuba, Ibaraki and ⁵The Center for Disease Biology and Integrative Medicine, Faculty of Medicine, University of Tokyo, Bunkyo-ku, Tokyo, Japan

Abstract

Although fetal or neonatal mammalian ovaries contain many non-growing oocytes within primordial follicles, most degenerate and only a few contribute to the oocyte pool in the mature ovary. Here, we report a follicle-free culture system that allows a large number of these arrested oocytes to enter the growth phase in vitro. As many as 800 oocytes from a newborn mouse, corresponding to more than 104 oocytes in large animals, continued to develop, formed a zona pellucida, and were able to fuse with spermatozoa. Some oocytes reached the size of those in normal antral follicles and entered metaphase I, indicating the completion of the growth phase. The key to success was the sequential provision of essential nutrients and growth factors to the oocytes, while preventing the apoptosis that normally occurs in the majority of growing oocytes in vivo. Importantly, maternal genomic imprinting, which is necessary for normal embryonic development, was imposed correctly on their genomes autonomously. Thus, arrested primordial oocytes can be rescued effectively in vitro and can undergo the morphological and genomic modifications necessary for fertilization and subsequent embryonic development. This culture system may provide a significant impetus to the development of new techniques for the efficient production of oocytes from fetal or neonatal ovaries, for research, clinical, and zoological purposes.

Keywords

theca cell, follicle, stem cell factor, genomic imprinting