Person:
Cajas Suárez, Yulia Nathaly

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First Name
Yulia Nathaly
Last Name
Cajas Suárez
URI
https://hdl.handle.net/20.500.14352/87116
Affiliation
Universidad Complutense de Madrid
Faculty / Institute
Veterinaria
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Author: Arias Álvarez, María × Author: Fierro, Natacha ×

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    Improvement of oocyte competence and in vitro oocyte maturation with EGF and IGF-I in Guinea pig model
    (Theriogenology, 2023) Cañón Beltrán, Karina Esperanza; García García, Rosa María; Cajas Suárez, Yulia Nathaly; Fierro, Natacha; Lorenzo González, Pedro Luis; Arias Álvarez, María
    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.
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    Acquisition of fertilization competence in guinea pig spermatozoa under different capacitation protocols
    (Theriogenology, 2023) Cañón Beltrán, Karina Esperanza; Cajas Suárez, Yulia Nathaly; González Martínez, María Encina; Fernández-González, Raul; Fierro, Natacha; Lorenzo González, Pedro Luis; Arias Álvarez, María; García García, Rosa María; Gutiérrez Adán, Alfonso; Rizos, Dimitri
    Guinea pig in vitro fertilization (IVF) are poorly developed due to the limited accessibility to oocytes and the lack of an efficient method of sperm capacitation. Thus, we aimed to evaluate different capacitation protocols that we validated through sperm analysis and using heterologous (He) IVF with zona-intact bovine oocytes. Spermatozoa of guinea pigs were collected and processed separately by 4 different protocols: A) Spermatozoa were obtained by flushing the lumen of one cauda epididymis and incubated in a minimal culture medium (MCM); B) One epididymis was placed in a prewarmed of M2 medium and gently minced with fine scissors. Spermatozoa were incubated in a modified human tubal fluid medium (HTF). In both protocols, the spermatozoa were capacitated at 37 °C under an atmosphere of 5% CO2 for 2 h. In the protocols C and D, the spermatozoa were collected by flushing the lumen of the cauda epididymis and selected by commercial density gradient Bovipure® (Nidacon Laboratories AB, Göthenborg, Sweden), according to the manufacturer's instructions. Then for Protocol C) spermatozoa were incubated in MCM medium supplemented with 10 mg/mL heparin (MCM-Hep); while for Protocol D) spermatozoa were incubated in FERT medium supplemented 10 mg/mL heparin (FERT-Hep). Incubation of C and D protocols were performed at 38.5 °C under an atmosphere of 5% CO2 for 2 h. Capacitation protocols C and D showed a higher percentage of viability, total and hyperactive-like motility, and acrosome reaction compared to protocols A and B. For this reason, protocols C and D were used for further He-IVF analysis. Guinea pig sperm and matured zona-intact bovine oocytes were co-incubated at 5% CO2 and 38.5 °C. Sperm-oocyte interaction was assessed at 2.5 h post-insemination (hpi) and pronuclear formation (PrF) were evaluated at 18, 20, 22, 24 and 26 hpi, while the cleavage rate was evaluated at 48 hpi. In protocol D, PrF was significantly higher than in protocol C (P ≤ 0.05) at every time point evaluated. Also, the cleavage rate at 48 hpi was higher (P ≤ 0.05) in He-IVF protocol D (69.8 ± 1.7%) compared to He-IVF protocol C (49.1 ± 1.1%). In conclusion, we determined the most adequate sperm capacitation conditions for guinea pig that allow zona-intact bovine oocyte penetration and lead to hybrid embryo formation, suggesting that these conditions could be optimal to develop IVF in guinea pigs.