Beneficial effects of glutathione supplementation during vitrification of mouse oocytes at the germinal vesicle stage on their preimplantation development following maturation and fertilization in vitro

Oocyte cryopreservation is imperative for assisted reproductive technologies (ART). Although cryopreservation of oocytes at the Metaphase II has been widely used, immature oocytes at the germinal vesicle stage (GV-oocytes) need to be cryopreserved in certain situations such as cancer patients; however, the success rate of embryonic development from the GV-oocytes remains low largely due to the requirement for in vitro maturation (IVM). Our aim was to investigate the effects of glutathione (GSH) supplementation during vitrification and warming of mouse GV-oocytes on the preservation of developmental competence. GV-oocytes within cumulus oocyte complexes (COCs) were collected from C57BL/6J (B6) and (B6.DBA)F1 mouse strains and subjected to vitrification and warming, followed by IVM. The vitrification, warming or IVM medium was supplemented with GSH at 0-4.0 mM. In vitro matured oocytes were then fertilized in vitro and cultured in KSOMaa up to 4 days. The first cleavage and blastocyst development were evaluated morphologically, and their rates were statistically analysed by one-way ANOVA followed by Tukey's multiple comparisons test. The difference was considered significant at P < 0.05. The results showed that GSH supplementation in the IVM medium exhibited no or rather inhibitory effects on the first cleavage or blastocyst development in both mouse strains except that 1.0 mM GSH increased the blastocyst development rate in B6. By contrast, 1 mM GSH supplementation during vitrification and warming increased the blastocyst development rate in both mouse strains, more efficiently in B6 than (B6.DBA)F1. In conclusion, GSH supplementation during vitrification and warming of GV-oocytes protects the oocytes from freezing-inflicted loss of developmental competence.