In Vitro Culture of Early Secondary Preantral Follicles in Hanging Drop of Ovarian Cell-Conditioned Medium to Obtain MII Oocytes from Outbred Deer Mice
In Vitro Culture of Early Secondary Preantral Follicles in Hanging Drop of Ovarian Cell-Conditioned Medium to Obtain MII Oocytes from Outbred Deer Mice
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DOI：
10.1089/ten.tea.2013.0055
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发表时间：
2013-12-01
影响因子：
4.1
通讯作者：
He, Xiaoming
中科院分区：
文献类型：
--
作者：
Choi, Jung Kyu;Agarwal, Pranay;He, Xiaoming
卵巢卵泡（每个卵泡包含一个卵母细胞）是哺乳动物卵巢的基本功能组织单位。在人类中，长期以来人们一直认为女性出生时卵泡（或卵母细胞）的数量最多，这些卵泡（或卵母细胞）不仅不可再生，而且随着时间的推移而退化，在大约35岁之后卵母细胞质量急剧下降。因此，分离和储存卵巢卵泡进行体外培养，以获得可受精的卵母细胞，对于可能想要推迟生育的职业女性，由于暴露于环境/职业危害或积极的医疗治疗（如放疗和化疗）而可能失去性腺功能的年轻未婚女性，甚至濒危物种和品种都具有重要意义。尽管它们对卵泡科学和生物学的理解做出了重大贡献，但迄今为止关于这一主题的大多数研究都是使用人造的、非自然的近交动物物种完成的。本研究发现，传统的二维微升滴法和三维挂滴法（HD）可有效地体外培养近交系小鼠的腔前卵泡，但不能直接移植到远交系鹿小鼠身上。因此，本研究开发了一种改进的HD方法，以获得更高的（比最佳常规方法高5倍）从远交种小鼠到中央期的早期继发性腔前卵泡发育百分比，为此，使用卵巢细胞条件培养基和每个HD的多个卵泡被认为是至关重要的。进一步发现，体外培养腔前卵泡获得的窦卵泡卵母细胞体外成熟方法与体内激素刺激获得的窦卵泡卵母细胞体外成熟方法有很大不同。因此，本研究对建立有效的体外卵泡培养方案，保护野生动物和人类自然远交的生育能力具有重要的指导意义。
The ovarian follicle (each contains a single oocyte) is the fundamental functional tissue unit of mammalian ovaries. In humans, it has been long held true that females are born with a maximum number of follicles (or oocytes) that are not only nonrenewable, but also undergoing degeneration with time with a sharply decreased oocyte quality after the age of approximate to 35. Therefore, it is of importance to isolate and bank ovarian follicles for in vitro culture to obtain fertilizable oocytes later, to preserve the fertility of professional women who may want to delay childbearing, young and unmarried women who may lose gonadal function because of exposure to environmental/occupational hazards or aggressive medical treatments, such as radiation and chemotherapy, and even endangered species and breeds. Although they contributed significantly to the understanding of follicle science and biology, most studies reported to date on this topic were done using the man-made, unnatural inbred animal species. It was found in this study that the conventional two-dimensional microliter drop and three-dimensional hanging drop (HD) methods, reported to be effective for in vitro culture of preantral follicles from inbred mice, are not directly transferrable to outbred deer mice. Therefore, a modified HD method was developed in this study to achieve a much higher (>5 times compared to the best conventional methods) percentage of developing early secondary preantral follicles from the outbred mice to the antral stage, for which, the use of an ovarian cell-conditioned medium and multiple follicles per HD were identified to be crucial. It was further found that the method for in vitro maturation of oocytes in antral follicles obtained by in vitro culture of preantral follicles could be very different from that for oocytes in antral follicles obtained by hormone stimulation in vivo. Therefore, this study should provide important guidance for establishing effective protocols of in vitro follicle culture to preserve the fertility of wildlife and humans outbred by nature.