Effects of reproductive aging and postovulatory aging on the maintenance of biological competence after oocyte vitrification: insights from the mouse model

Cryopreservation of female reproductive cells allows preservation of fertility and provides materials for research. Although freezing protocols have been optimized, and there is a high survival rate after thawing, the in vitro fertilization (IVF) pregnancy rate is still lower in cycles with cryopreserved oocytes, thus highlighting the importance of identifying intrinsic limiting factors characterizing the cells at time of freezing. The aim of the present study is to investigate in the mouse model the impact of reproductive aging and postovulatory aging on oocyte biological competence after vitrification. Metaphase II oocytes were vitrified soon after retrieval from young and reproductively old mice. Part of the oocytes from young animals was vitrified after 6 h incubation (in vitro aged oocytes). All classes of oocytes showed similar survival rate after vitrification. Moreover, vitrification did not alter chromosomal organization in young cells, whereas in vitro aged and old oocytes presented an increase of slightly aberrant metaphase configurations. Compared to fresh young oocytes, in vitro aged and old oocytes showed increased ROS levels which remained unchanged after vitrification. By contrast, cryopreservation significantly increased ROS production in young oocytes. Both the aging processes negatively impacted oocyte ability to undergo pronucleus formation and first cleavage after vitrification by stimulating cellular fragmentation. These results could be helpful for establishing the correct time table for cryopreservation in the laboratory routine and improving its application in reproductively old females. Moreover, our observations highlight the importance of oxidative stress protection during vitrification procedures.