Effect of bovine oocyte vitrification with EGTA and post-warming recovery with resveratrol on meiotic spindle, mitochondrial function, reactive oxygen species, and developmental competence

Abstract

The present study aimed to determine the effects of the addition of EGTA to vitrification solutions and a post-warming recovery period supplemented with 1 μM resveratrol on meiotic spindle integrity, mitochondrial activity, ATP content, reactive oxygen species (ROS) levels, and developmental potential of partially denuded, vitrified-warmed bovine oocytes. Results of microtubule distribution and chromosomal arrangement indicated that resveratrol supplementation, irrespective to EGTA addition, reduced the incidence of abnormal meiotic spindles to similar levels of the control group. Mitochondrial membrane potential was similar in all groups, but ATP content was negatively affected by the vitrification-warming procedure and failed to recover after 4 h of post-warming culture. Resveratrol caused the reduction of ROS to lower levels of the control group, and showed the lowest ROS levels when combined with EGTA treatment. Oocytes in all vitrification groups presented lower developmental potential when compared to fresh oocytes. However, oocytes that underwent vitrification supplemented with EGTA and post-warming culture along with resveratrol showed higher developmental competence compared with vitrified-warmed oocytes not supplemented with resveratrol. The results of our study indicate that submitting vitrified-warmed, partially denuded bovine oocytes to a post-warming recovery period supplemented with 1 μM resveratrol improves vitrification outcomes. However, the benefits of EGTA on vitrification and warming of bovine oocytes need to be further investigated.