Single-cell transcriptomic analysis reveals subtle fluctuations of oocyte cryopreservation on mouse early embryo development.

BACKGROUND: Oocyte cryopreservation is widely used in assisted reproductive technology, but its effects on embryonic development remain a concern. This study aimed to determine whether oocyte cryopreservation induces transcriptional alterations at the early cleavage and blastocyst stages. METHODS: We performed single-cell RNA sequencing on mouse Metaphase II oocyte and embryos at early/late 1 cell, early/late 2 cell, morula and blastocyst stages derived from cryopreserved (cryo) and fresh (fresh) oocytes. Transcriptomic profiles were compared between the two groups. RESULTS: Global transcriptomes were highly similar between cryo and fresh blastocysts. Oocyte cryopreservation did not alter the sex ratio or the developmental progression through preimplantation development. In blastocysts, cryopreservation affected only a few individual genes. In both male and female blastocysts, although oocyte cryopreservation increased the proportion of low Xist-expressing cells, it did not disrupt X-linked gene dosage. In the inner cell mass (ICM) of blastocysts, a negative correlation between X-linked gene expression and cell cycle progression was identified, and this negative correlation was more pronounced in trophectoderm (TE) cells. Overall, this stage-dependent coordination remained unperturbed by cryopreservation. CellChat analysis revealed that the signalling interaction strength between ICM and TE also showed no significant changes in the cryo group. CONCLUSIONS: Oocyte cryopreservation does not severely disrupt overall transcriptional integrity, developmental potency and X-chromosome dosage compensation of embryos, but it may cause subtle molecular changes. These findings highlight the need for continued refinement of cryopreservation methods and further investigation into the sub-acute safety of assisted reproductive technology procedures.