Abstract
Oocyte vitrification allows for the storing of endangered breed female gametes. Cryoprotectant (CPA) concentration and exposure time should ensure cell protection with minimal toxicity. In the present study, a high concentration-rapid exposure (HC-RE) and a low concentration-slow exposure (LC-SE) vitrification protocol, using dimethyl sulfoxide (DMSO) and ethylene glycol (EG) as permeating CPAs, were evaluated on meiotic competence and bioenergetic-oxidative status of pre-pubertal lamb immature COCs after in vitro maturation (IVM). For each protocol, COCs vitrified through a traditional protocol and fresh ones were used as controls. Both protocols allowed COC morphology preservation after vitrification-warming (V-W) and cumulus expansion after IVM. The maturation rate (7% and 14%) was comparable to the vitrified control (13% and 21%) but not satisfactory compared to fresh ones (58% and 64%; p < 0.001). The rate of mature oocytes displaying a perinuclear/subcortical (P/S) mitochondrial distribution pattern, an index of cytoplasmic maturity, was comparable between vitrified and fresh oocytes. The LC-SE vitrification protocol did not affect quantitative bioenergetic-oxidative parameters compared to both controls whereas HC-RE protocol significantly reduced intracellular reactive oxygen species (ROS) levels, indicating cell viability loss. In conclusion, to improve pre-pubertal lamb immature COC vitrification, the combination of low CPA concentrations with prolonged exposure time could be more promising to investigate further.