Transcriptomic difference in bovine blastocysts following vitrification and slow freezing at morula stage
Citation
Gupta, A., Singh, J., Dufort, I., Robert, C., Dias, F.C.F., Anzar, M. (2017). Transcriptomic difference in bovine blastocysts following vitrification and slow freezing at morula stage. PLoS ONE, [online] 12(11), http://dx.doi.org/10.1371/journal.pone.0187268
Plain language summary
Embryo cryopreservation is very common animal breeding technology to conserve the animal genetic resources and exploitation of genetic production traits. Slow freezing and vitrification are two important techniques commonly used for cryopreservation of mammalian embryos. In this study, the bovine compact morulae were either vitrified or slow-frozen, and post-warm morulae were cultured to the expanded blastocyst (embryo) stage. The embryos developed from the vitrified and slow-frozen morulae were subjected to microarray analysis and compared with blastocysts developed from the unfrozen control morulae for differential gene expression. Morula to blastocyst conversion rate was almost similar between control and vitrified groups, and very low in control group. The vitrified embryos exhibited significant changes in gene expression mainly involving embryo implantation, lipid peroxidation and reactive oxygen species generation and cell differentiation. The slow-frozen embryos, however, showed changes in the expression of genes related to cell signaling, cell structure and differentiation, and lipid metabolism.
Abstract
Cryopreservation is known for its marked deleterious effects on embryonic health. Bovine compact morulae were vitrified or slow-frozen, and post-warm morulae were cultured to the expanded blastocyst stage. Blastocysts developed from vitrified and slow-frozen morulae were subjected to microarray analysis and compared with blastocysts developed from unfrozen control morulae for differential gene expression. Morula to blastocyst conversion rate was higher (P < 0.05) in control (72%) and vitrified (77%) than in slow-frozen (34%) morulae. Total 20 genes were upregulated and 44 genes were downregulated in blastocysts developed from vitrified morulae (fold change ≥ ± 2, P < 0.05) in comparison with blastocysts developed from control morulae. In blastocysts developed from slow-frozen morulae, 102 genes were upregulated and 63 genes were downregulated (fold change ≥ ± 1.5, P < 0.05). Blastocysts developed from vitrified morulae exhibited significant changes in gene expression mainly involving embryo implantation (PTGS2, CALB1), lipid peroxidation and reactive oxygen species generation (HSD3B1, AKR1B1, APOA1) and cell differentiation (KRT19, CLDN23). However, blastocysts developed from slow-frozen morulae showed changes in the expression of genes related to cell signaling (SPP1), cell structure and differentiation (DCLK2, JAM2 and VIM), and lipid metabolism (PLA2R1 and SMPD3). In silico comparison between blastocysts developed form vitrified and slow-frozen morulae revealed similar changes in gene expression as between blastocysts developed from vitrified and control morulae. In conclusion, blastocysts developed form vitrified morulae demonstrated better post-warming survival than blastocysts developed from slow-frozen morulae but their gene expression related to lipid metabolism, steroidogenesis, cell differentiation and placentation changed significantly (≥ 2 fold). Slow freezing method killed more morulae than vitrification but those which survived up to blastocyst stage did not express ≥ 2 fold change in their gene expression as compared with blastocysts from control morulae.