SummaryIn vitro activation of primordial follicles could serve as a safe method to preserve fertility in patients with cancer subjected to ovarian tissue cryopreservation during oncotherapy, however the culture medium for this purpose requires to be optimized. Granulosa cell conditioned medium (GCCM) has been recognized to enhance primordial follicle activation and the present study was conducted to understand whether addition of pyruvate, a combination of insulin, transferrin and selenium (ITS) or testosterone to GCCM could improve its efficiency in this regard. To this end, 1-day-old mouse ovaries were cultured in four different media including CON (control; containing GGCM only), PYR (containing GCCM plus pyruvate), ITS (containing GCCM plus ITS) or TES (containing GCCM plus testosterone) for 11 days. Furthermore, follicular dynamics and gene expression of factors involved in follicular development were assessed using histological examination and RT-PCR, respectively, on days 5 and 11 of culture. Pyruvate decreased follicular activation, but it enhanced the progression of follicles to the primary stage. Moreover, it upregulated Bmp15 and Cx37 (P < 0.05). In the ITS group, activation of follicles was not affected and total number of follicles was reduced by day 11 of culture. Additionally, ITS downregulated Pi3k, Gdf9, Bmp15 and Cx37 (P < 0.05). Although testosterone did not affect primordial follicle activation, it enhanced the development of follicles up to the preantral stage (P < 0.05). Furthermore, testosterone inhibited the expression of Pten but stimulated the expression of Gdf9 and Cx37 (P < 0.05). In conclusion, the present study revealed that inclusion of pyruvate and testosterone into GCCM could enhance the early development of follicles in cultured 1-day-old mouse ovaries.

中文翻译：

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用丙酮酸和睾酮补充颗粒细胞条件培养基可以改善培养的 1 日龄小鼠卵巢的早期卵泡发育

概括体外激活原始卵泡可以作为一种安全的方法来保护在肿瘤治疗期间接受卵巢组织冷冻保存的癌症患者的生育能力，但是需要优化用于此目的的培养基。颗粒细胞条件培养基 (GCCM) 已被认为可以增强原始卵泡的活化，本研究旨在了解在 GCCM 中添加丙酮酸、胰岛素、转铁蛋白和硒 (ITS) 或睾酮的组合是否可以提高其在这方面的效率. 为此，将 1 日龄小鼠卵巢培养在四种不同的培养基中，包括 CON（对照；仅含 GGCM）、PYR（含 GCCM 加丙酮酸盐）、ITS（含 GCCM 加 ITS）或 TES（含 GCCM 加睾酮） 11天。此外，在培养的第 5 天和第 11 天，分别使用组织学检查和 RT-PCR 评估了参与卵泡发育的因素的卵泡动力学和基因表达。丙酮酸降低了卵泡的活化，但它促进了卵泡向初级阶段的进展。此外，它上调了Bmp15和CX37(磷< 0.05)。在 ITS 组中，卵泡的激活不受影响，并且在培养第 11 天时卵泡总数减少。此外，ITS 下调Pi3k,Gdf9,Bmp15和CX37(磷< 0.05)。虽然睾酮不影响原始卵泡的激活，但它促进卵泡发育到前腔期。磷< 0.05)。此外，睾酮抑制普腾但刺激了表达Gdf9和CX37(磷< 0.05)。总之，本研究表明，在 GCCM 中加入丙酮酸和睾酮可以促进培养的 1 日龄小鼠卵巢中卵泡的早期发育。