Optimization of vitrification methods for equine oocytes.

An important method for preserving equine germplasm is the cryopreservation of equine oocytes. Due to its ease, rapidity and affordability, vitrification freezing has taken over as the primary method of horse oocyte cryopreservation. The vitrification cryoprotectants utilized in this investigation were Ethylene glycol (E), Dimethyl sulfoxide (D), Sucrose (S), and Ficoll (F). According to the oocyte volume alteration, the treatment time was 39 s in equilibrium solution ED10 (10 % EG + 10 % DMSO), 32 s in equilibrium solution ED15 (15 % EG + 15 % DMSO), while 20 s in equilibrium solution ED20 (20 % EG + 20 % DMSO). We prepared three kinds of cryosolutions EDFS30 (E15 %+D15 %+70 %FS), EDFS35 (E17.5 % + D17.5 % + 65 %FS), EDFS40 (E20 % + D20 % + 60 %FS) according to the proportion of protectant components. Among 27 freezing protocols, we selected protocol ED10 (39 s) + EDFS30 + 80 s which has the highest in vitro culture maturation rate of 19.3 % while protocol ED20 (20 s) + EDFS40 + 120 s is the worst. Apoptosis gene analysis revealed that BAX, BID, BOK, and TP53 expression was substantially higher in oocytes from the ED20 (20 s) + EDFS40 + 120 s group than in oocytes from the ED10 (39 s) + EDFS30 + 80 s and control groups (p<0.01). This study investigated several vitrification schemes for equine oocytes.