Cryopreservation of In Vitro Produced Embryos and Immature Oocytes in Pigs

Gamete cryopreservation combined with in vitro embryo production (IVP) and cryopreservation of resultant embryos are important technologies for gene banking in domestic animals. Especially, cryopreservation of embryos and oocytes offers the possibility for the preservation of female genetic lines. Although several vitrification protocols have been developed for the cryopreservation of embryos and oocytes in pigs, their adaptation in IVP systems have resulted in only limited success. In case of IVP embryos, high frequencies of polyspermy and the sensitivity of porcine embryos to culture stresses reduce embryo competence and thus limit the efficacy of their cryopreservation. Several approaches have been introduced to improve efficacy of cryopreservation for IVP embryos such as removal of intracellular lipid, adjustments of the culture systems to improve embryo competence and artificial induction of stress tolerance. Our researches have demonstrated that IVP embryos can be both selected for monospermy and vitrified effectively at the zygote stage and their direct transfer into recipients is an effective approach to overcome the problems of polyspermic development and culture stress. Regarding oocyte cryopreservation, our researches have revealed that using current protocols, vitrification of porcine oocytes at the immature stage is more advantageous compared with vitrification at the matured stage since oocytes preserved at the immature stage show better developmental competence in the IVP system. Optimization of cryoprotectant treatment during vitrification and warming temperature after vitrification specifically for immature oocytes allowed us the production of live piglets from cryopreserved oocytes for the first time. Nevertheless, limited efficacies require further improvement of these technologies.