In Vitro Toxicity of a DEHP and Cadmium Mixture on Sheep Cumulus-Oocyte Complexes

Di-(2-ethylhexyl) phthalate (DEHP) and Cadmium (Cd) are known to affect female reproduction. To date, most of toxicological research focused on the effects of individual contaminants whereas humans and animals are actually exposed to mixtures. This study analyzed the effects of a DEHP/Cd mixture on nuclear and cytoplasmic maturation of sheep cumulus-oocyte complexes (COCs) compared with single compounds. COCs recovered from slaughterhouses-derived sheep ovaries were exposed to 0.5μM DEHP, 0.1μM Cd or DEHP/Cd mixture at same concentrations during 24h in vitro maturation (IVM). Control COCs were cultured in DEHP-vehicle supplemented medium. After IVM, oocyte nuclear chromatin configuration was evaluated, and bioenergetic/oxidative parameters were assessed on expanded cumulus cells (CCs) and matured oocytes. No effects were noticed on oocyte nuclear maturation at any tested condition. However, either the mixture or individual compounds affected oocyte bioenergetics as they significantly reduced the percentages of matured oocytes with healthy perinuclear/subplasmalemmal mitochondrial distribution pattern and decreased mitochondrial membrane potential, intracellular ROS levels and mitochondria/ROS co-localization compared to controls (Chi-square test and one-way ANOVA; p<0.05). Moreover, the mixture increased CC ROS levels. In conclusion, oocytes suffered the same damage when exposed to the mixture or separate compounds whereas CCs displayed oxidative stress only upon mixture exposure.