Resumen de Study of the effect of cryopreservation on rabbit embryo gene expression
Maria dels Desemparats Sáenz de Juano Ribes
While cryopreservation techniques are becoming a common method in assisted reproduction and embryo storage, more efforts should be made to investigate effects on embryo and foetal development and the non-lethal damage on the embryos that finally develop to term. The general aim of this thesis was to study the effects of embryo freezing and vitrification on rabbit late blastocyst development, implantation and delivery rates and deep in the remaining effects of vitrification after the implantation. The results showed that slow freezing and vitrification procedures induced different distribution of losses through gestation. While slow freezing losses are mostly located during pre-implantatory period, vitrification losses manifested two picks, one before late blastocyst stage and the other in the second part of gestation, after day 14 of development. Concerning to gene expression level, before the onset of implantation, slow freezing and vitrification induce similar alterations in gene expression pattern of late blastocysts. However, transcriptomic changes observed are also influenced by the recovery procedure, and when fresh transferred embryos were used there were any transcriptomic differences between vitrified and control groups. After the implantation, we observed that at day 14 of development the foetus and the maternal placenta weight less in vitrified than in control embryos. In addition, there were alterations in the transcriptome (60 genes) and proteome (89 protein spots) of foetal placentas derived from vitrified embryos. Moreover, we also detected that the protein alterations did not disappear at the last part of gestation, and at day 24 of development, foetal placenta of those vitrified live fetuses showed 32 differentially expressed proteins, some of them previously identified at day 14. Finally another observation was that vitrified newborns weighted more than the control ones. Our findings clarify the development ability and distribution of losses of cryopreserved rabbit embryos and our findings localize the set off of prenatal losses of vitrified embryos in the formation of the placenta and detected alterations at transcriptomic and proteomic level at in the middle of gestation and near the end. Moreover, this thesis arise the question of cryopreservation neutrality, and bring to light an important issue which must be addressed before embryo vitrification becomes common practice in embryo store and assisted reproduction.
