Department of Animal Science, Tokyo University of Agriculture, Funako 1737, Atsugi, Kanagawa 243-0034, Japan.
Reproduction. 2013 Jan 8;145(1):85-96. doi: 10.1530/REP-12-0319. Print 2013 Jan.
Antrum formation and estradiol (E(2)) secretion are specific features of oocyte and granulosa cell complexes (OGCs). This study investigates the effect of E(2) on the in vitro development of bovine OGCs derived from early antral follicles as well as on the expression of genes in granulosa cells (GCs). The supplementation of culture medium with either E(2) or androstenedione (A(4)) improved the in vitro development of OGCs and the nuclear maturation of enclosed oocytes. When OGCs were cultured in medium containing A(4), developmentally competent OGCs secreted more E(2) than OGCs that were not competent. In addition, fulvestrant inhibited the effect of both E(2) and A(4) on OGCs development. Comprehensive gene expression analysis using next-generation sequence technology was conducted for the following three types of GCs: i) GCs of OGCs cultured for 4 days with E(2) (1 μg/ml; E(2)(+)), ii) GCs of OGCs cultured for 4 days without E(2) (E(2)(-)) or iii) OGCs that formed clear antrum after 8 days of in vitro culture in medium containing E(2) (1 μg/ml; AF group). GCs of the E(2)(+) group had a similar gene expression profile to the profile reported previously for the in vivo development of large follicles. This genetic profile included factors implicated in the up-regulation of E(2) biosynthesis and down-regulation of cytoskeleton and extracellular matrices. In addition, a novel gene expression profile was found in the AF group. In conclusion, E(2) impacts the gene expression profile of GCs to support the in vitro development of OGCs.
腔的形成和雌二醇（E(2)）的分泌是卵母细胞和颗粒细胞复合体（OGCs）的特有特征。本研究调查了 E(2)对来源于早期腔前卵泡的牛 OGCs 的体外发育以及颗粒细胞（GCs）中基因表达的影响。用 E(2)或雄烯二酮（A(4)）补充培养基可改善 OGCs 的体外发育和包被卵母细胞核成熟。当 OGCs 在含有 A(4)的培养基中培养时，具有发育能力的 OGCs 分泌的 E(2)多于不具有发育能力的 OGCs。此外，氟维司群抑制了 E(2)和 A(4)对 OGCs 发育的影响。使用下一代测序技术对以下三种类型的 GCs 进行了综合基因表达分析：i）用 E(2)（1μg/ml；E(2)(+)）培养 4 天的 OGCs 的 GCs，ii）未用 E(2)培养 4 天的 OGCs 的 GCs（E(2)(-)）或 iii）在含有 E(2)（1μg/ml）的培养基中体外培养 8 天后形成明显腔的 OGCs（AF 组）。E(2)(+)组的 GCs 与先前报道的大卵泡体内发育的基因表达谱相似。这种遗传谱包括上调 E(2)生物合成和下调细胞骨架和细胞外基质的因素。此外，还发现了 AF 组中的一个新的基因表达谱。总之，E(2)影响 GCs 的基因表达谱以支持 OGCs 的体外发育。
