Canine IVM With SOF Medium, Insulin-Transferrin-Selenium, and Low O2 Tension Improves Oocyte Meiotic Competence and Decreases Reactive Oxygen Species Levels

Matteo Duque Rodriguez; Camila O Cittadini; Gabriela M Teplitz; Adrian De Stefano; Daniel M Lombardo; Daniel F Salamone

doi:10.3389/fcell.2021.694889

Canine IVM With SOF Medium, Insulin-Transferrin-Selenium, and Low O₂ Tension Improves Oocyte Meiotic Competence and Decreases Reactive Oxygen Species Levels

Front Cell Dev Biol. 2021 Sep 7:9:694889. doi: 10.3389/fcell.2021.694889. eCollection 2021.

Authors

Matteo Duque Rodriguez^{1

2

3}, Camila O Cittadini², Gabriela M Teplitz⁴, Adrian De Stefano², Daniel M Lombardo⁴, Daniel F Salamone^{2

3}

Affiliations

¹ Facultad de Ciencias Agrarias, Politécnico Colombiano Jaime Isaza Cadavid, Medellin, Colombia.
² Laboratorio Biotecnología Animal, Departamento de Producción Animal, Facultad de Agronomía, Universidad de Buenos Aires, Buenos Aires, Argentina.
³ Instituto de Investigaciones en Producción Animal, Universidad de Buenos Aires, Buenos Aires, Argentina.
⁴ Facultad de Ciencias Veterinarias, Instituto de Investigación y Tecnología en Reproducción Animal, Catedra de Histología y Embriología, CONICET-Universidad de Buenos Aires, Buenos Aires, Argentina.

Abstract

Assisted reproductive technologies in canine species are limited due to the low efficiency of in vitro maturation (IVM). Unlike other mammals, bitches ovulate oocytes in the germinal vesicle stage and complete metaphase II (MII) after 48-72 h in the oviductal environment and become fertilizable. For this reason, we compared two different IVM media, synthetic oviductal fluid (SOF) supplemented with 8% bovine serum albumin (BSA) or a mixture of 8% BSA-2.5% fetal bovine serum (FBS) and TCM-199 with 10% FBS. Additionally, we evaluated the effect of supplementation with insulin-transferrin-selenium (ITS) and low O₂ tension in oocyte maturation, reactive oxygen species (ROS) levels, membrane integrity, and embryo development following parthenogenetic activation (PA). After 72 h of culture, SOF + BSA, SOF + BSA + FBS, and TCM-199 + FBS show 5, 7, and 4% of MII, respectively, without a statistical difference. However, SOF + BSA produced significantly higher degeneration rates compared to SOF + BSA + FBS (44 and 23%, respectively). Remarkably, supplementation with 1 μl/ml of ITS under high O₂ tension demonstrated a beneficial effect by improving maturation rates up to 20% compared to the other groups. Low O₂ tension increased maturation rates to 36.5%, although there were no statistical differences compared to high O₂ tension in the presence of ITS. Lower ROS levels and higher integrity of the cytoplasmic membrane were found in the presence of ITS despite no differences in maturation rates under low O₂ tension groups. Additionally, after PA, 1% development until the eight-cell stage was obtained after activation of in vitro-matured oocytes in the presence of ITS. Taken together, these results indicate that SOF supplemented with 8% BSA and 2.5% FBS is suitable for IVM of canine oocytes and ITS supplementation was beneficial for both high and low O₂ tension. Furthermore, the addition of ITS in the cultured system lowers ROS levels and increases membrane integrity in domestic dog oocytes after IVM.

Keywords: SOF medium; canine; in vitro maturation; insulin-transferrin-selenium; oxygen tension.