Expression of mRNA, before and after freezing, in bovine blastocysts cultured under different conditions

Abstract

Production methods and culture systems have been shown to affect blastocyst mRNA expression and cryopreservability, which may serve as sensitive indicators of embryo quality and developmental competence. In the present study, the impact of four established culture conditions for producing bovine blastocysts ( in vitro production, IVP; gamete intra-fallopian transfer, GIFT; transfer of cleaved stages into the oviduct, CLVT; multiple ovulation embryo transfer, MOET) was assessed, in terms of both cryosurvival and levels of mRNA expression of several selected genes (occludin, desmocollin 2, solute carrier family 2 member 3, BAX, BCL-XL, heat shock protein 1A, aquaporin 3, DNA methyltransferase 1a) detected with RT-qPCR. At 24 hours post-thawing, blastocysts derived from in vitro production showed a significantly higher re-expansion rate compared to the other groups. At later times, this difference was no longer significant. Before freezing, embryos of the MOET group showed significantly more desmocollin 2 mRNA compared to embryos produced using other culture methods. After freezing, significant upregulation was found in transcripts of heat shock protein 1A in embryos of all groups; of solute carrier family 2 member 3, only in IVP derived embryos; of BAX, BCL-XL, occludin, desmocollin 2, only in the MOET and IVP groups. Aquaporin 3 and DNA methyltransferase 1a were neither up- nor downregulated in blastocysts of any group. In conclusion, these findings suggest that, after freezing, embryos seem to have switched on mRNA synthesis, an active metabolism, operational cell connections, and are prepared for hatching and beyond.