Department of Physiology, Veterinary Faculty, Complutense University of Madrid - UCM, 28040, Spain; Departamento de Ciencias Biológicas, Universidad Técnica Particular de Loja - UTPL, Loja, 11-01-608, Ecuador.
Department of Physiology, Veterinary Faculty, Complutense University of Madrid - UCM, 28040, Spain.
Theriogenology. 2024 Jan 15;214:206-214. doi: 10.1016/j.theriogenology.2023.10.029. Epub 2023 Oct 24.
In vitro maturation (IVM) system is an alternative method to superovulation protocols to obtain mature oocytes. Epidermal Growth Factor (EGF) and Insulin-like Growth Factor I (IGF-I) have been widely used in IVM medium in different species. Although the guinea pig is a valuable animal model for reproductive studies, IVM is rarely used. We aimed to establish a suitable in vitro production system using EGF and/or IGF-I during IVM to improve oocyte competence. Firstly, immunolocalization of EGF and IGF-I receptors in the ovary was assessed. An IVM dose-response experiment was performed with cumulus-oocyte complexes (COCs) supplemented with: 1) EGF [0, 10, 50, 100 ng/mL or 10% fetal calf serum (FCS)]; 2) IGF-I [0, 50, 100, 200 ng/mL or 10% FCS]; or 3) the concentrations of EGF and IGF-I which showed the best IVM index in the previous experiments, with or without Fetal Calf Serum (FCS). Cortical granule and mitochondria distribution patterns were determined in in vivo and in vitro-matured oocytes for the first time in this species. Apoptotic rate after IVM and oocyte competence by in vitro embryo development were evaluated. Immunohistochemistry results showed positive immunostaining of EGF and IGF receptors in corpus luteum, oocytes, granulosa and theca cells in follicles in all stages of development. Supplementation of IVM medium with 50 ng/mL EGF or 100 ng/mL IGF-I or their combination with FCS successfully led to oocyte nuclear and cytoplasmic maturation and reduced the apoptotic rate. Both growth factors improved oocyte competence during IVM in this species since early embryos were in vitro developed, showing better results when FCS was used in the IVM medium.
体外成熟(IVM)系统是一种替代超排卵方案以获得成熟卵母细胞的方法。表皮生长因子(EGF)和胰岛素样生长因子 I(IGF-I)已在不同物种的 IVM 培养基中广泛使用。尽管豚鼠是生殖研究的有价值的动物模型,但 IVM 很少使用。我们旨在建立一种使用 EGF 和/或 IGF-I 在 IVM 期间改善卵母细胞能力的合适的体外生产系统。首先,评估了卵巢中 EGF 和 IGF-I 受体的免疫定位。进行了 cumulus-oocyte 复合物(COC)的 IVM 剂量反应实验,其中添加了:1)EGF[0、10、50、100ng/mL 或 10%胎牛血清(FCS)];2)IGF-I[0、50、100、200ng/mL 或 10%FCS];或 3)在先前实验中显示最佳 IVM 指数的 EGF 和 IGF-I 浓度,有或没有胎牛血清(FCS)。首次在该物种中确定了体内和体外成熟卵母细胞中的皮质颗粒和线粒体分布模式。评估了 IVM 后的凋亡率和体外胚胎发育的卵母细胞能力。免疫组织化学结果显示,在所有发育阶段的黄体、卵母细胞、颗粒细胞和卵泡的膜细胞中,EGF 和 IGF 受体均呈阳性免疫染色。在 IVM 培养基中添加 50ng/mL EGF 或 100ng/mL IGF-I 或它们与 FCS 的组合可成功导致卵母细胞核和细胞质成熟,并降低凋亡率。这两种生长因子都通过在该物种的 IVM 期间提高卵母细胞能力,因为早期胚胎在体外发育,当 IVM 培养基中使用 FCS 时,结果更好。