203 Effects of oxygen level on bovine oocyte maturation and embryo development

Oxygen concentrations in the reproductive tract and follicular fluid are between 1.5% and 8.7%; however, the concentration typically used in standard IVM for research is between 20% and 21%. Higher oxygen levels could increase the risk of reactive oxygen species (ROS) formation in oocytes and have a negative impact on embryo development. The use of embryos derived by parthenogenetic activation (PA) of oocytes is a good model for detecting the differences in the oocyte developmental competence without a compounding effect of sperm. Embryos produced by IVF are commonly used as a standard model to evaluate the effect of the treatment on oocyte fertilization and embryonic development. The aim of this study was to evaluate the effect of oxygen levels on bovine oocyte maturation and subsequent embryo development following PA or IVF. Cumulus–oocyte-complexes (COCs) were matured in vitro in either 20% or 5% oxygen for 21–23 h. From each group, half of the COCs were denuded and used to assess the nuclear maturation. The matured metaphase II (MII) oocytes were then activated in 5 µM ionomycin for 5 minutes, followed by 4 h incubation in 6-dimethylaminopurine and 10 µg mL⁻¹ cycloheximide. For IVF, the other half of the COCs from each group were incubated with sperm for 6 h. Then both IVF and PA embryos were cultured in SOF media in 4-well dishes under oil in 5% oxygen. Cleavage and blastocyst rates were assessed on Day 2 and 8, post-activation and IVF, respectively. All the experiments were performed 4 times. Intracellular glutathione and ROS levels were assessed and used as an indicator of oocyte quality (12 oocytes in each group). Data were analysed by one-way analysis of variance and differences with P-values of less than 0.05 were considered statistically significant. The results were represented as means ± standard deviation. In total 296 COCs were placed in maturation and 206 and 266 oocytes were used for PA and IVF, respectively. Our results showed that there was no significant difference in cumulus cell expansion and maturation rates between high and low oxygen levels (73.7 ± 4.0% vs 69.7 ± 3.4%, P > 0.05). Additionally, no difference was observed in cleavage and blastocyst rates between high and low oxygen PA groups (86.6 ± 6.6% vs 79.6 ± 11.6% and 19.8 ± 5.1% vs 13.2 ± 5.1%, respectively; P > 0.05), and between high and low oxygen IVF groups (71.0 ± 12.4% vs 67.5 ± 9.9% and 25.8 ± 16.2% vs 20.3 ± 15.7%, respectively; P > 0.05). Preliminary results showed that slightly higher levels of GSH (P = 0.089) and significantly lower levels of ROS (P < 0.001) were observed in the MII oocytes from the low oxygen group, compared with that in the high oxygen group. In conclusion, the oxygen level during IVM did not affect the maturation and embryo development rates, but potentially improved the quality of the oocytes. We are assessing blastocyst cell numbers in these treatment groups.