Effects of Different Vitrification Temperatures and Cryoprotective Agent Concentrations on Ultrastructure and Apoptosis Related Proteins in Bovine GV Stage Oocytes

Abstract

The effects of different vitrification temperatures (VTs) and cryoprotective agent concentrations (CPAs) on the viability, ultrastructure, and apoptosis related proteins Bcl-2, Bax, and TP53 of bovine oocytes at the GV (germinal vesicle) stage were analyzed in this study. Cumulus oocyte complexes (COCs) were randomized into five groups: fresh (control), LHe 5.6 M (liquid helium, -269\dg, cryoprotectant concentration of 5.6 M), LHe 6.6 M (liquid helium, –269°C, cryoprotectant concentration of 6.6 M), LN 5.6 M (liquid nitrogen, –196°C, cryoprotectant concentration of 5.6 M), LN 6.6 M (liquid nitrogen, –196°C, cryoprotectant concentration of 6.6 M). Statistically, in the vitrification groups, LN 5.6 M group showed the lowest rate of morphologically normal, mature, and cleavage (58.1%; 32.9%; 12.9%). LHe 5.6 M showed the highest cleavage rate (39.6%), as well as effectively ameliorated ultrastructural damage, significantly reduced Bax protein expression, effectively increased Bcl-2 protein expression, reduced TP53 protein expression, in general, it increased the anti-apoptotic ability of the oocytes after vitrification. It was concluded that lower vitrification temperature and decreased cryoprotectant concentrations upregulate the expression of Bcl-2 protein and downregulate the expression of Bax and TP53 protein in oocytes, and enhance the anti-apoptotic capacity of oocytes, ameliorated the damage of the ultrastructure of the oocytes, perhaps improved the developmental capacity of bovine GV stage oocytes after vitrification can be partially attributed to the above changes.