Vitrification with Dimethyl Sulfoxide Induces Transcriptomic Alteration of Gene and Transposable Element Expression in Immature Human Oocytes

Abstract

Despite substantial advancements in the field of cryobiology, oocyte and embryo cryopreservation still compromise developmental competence. Furthermore, dimethyl sulfoxide (DMSO), one of the most commonly used cryoprotectants, has been found to exert potent effects on the epigenetic landscape of cultured human cells, as well as mouse oocytes and embryos. Little is known about its impact on human oocytes. Additionally, few studies investigate the effects of DMSO on transposable elements (TE), the control of which is essential for the maintenance of genomic instability. The objective of this study was to investigate the impact of vitrification with DMSO-containing cryoprotectant on the transcriptome, including on TEs, of human oocytes. Twenty-four oocytes at the GV stage were donated by four healthy women undergoing elective oocyte cryopreservation. Oocytes were paired such that half from each patient were vitrified with DMSO-containing cryoprotectant (Vitrified Cohort), while the other half were snap frozen in phosphate buffer, unexposed to DMSO (Non-Vitrified Cohort). All oocytes underwent RNA sequencing via a method with high fidelity for single cell analysis, and which allows for the analysis of TE expression through Switching Mechanism at the 5′-end of the RNA Transcript sequencing 2 (SMARTseq2), followed by functional enrichment analysis. Of the 27,837 genes identified by SMARTseq2, 7331 (26.3%) were differentially expressed (p < 0.05). There was a significant dysregulation of genes involved in chromatin and histone modification. Mitochondrial function, as well as the Wnt, insulin, mTOR, HIPPO, and MAPK signaling pathways were also altered. The expression of TEs was positively correlated with the expression of PIWIL2, DNMT3A, and DNMT3B, and negatively correlated with age. These findings suggest that the current standard process of oocyte vitrification, involving DMSO-containing cryoprotectant, induces significant transcriptome changes, including those involving TEs.

New Approach to the Cryopreservation of GV Oocytes and Cumulus Cells through the Lens of Preserving the Intercellular Gap Junctions Based on the Bovine Model

“…We evaluated its mRNA level after exposure to cryopreservation and different IVM systems. The low level of DNMT3A in immCOCs that we obtained is consistent with the results described previously, which demonstrated a general decrease in transcription at the end of the oocytes’ growth phase and during the transition to meiotic competence [ 35 ]. This is evolutionarily possible due to the accumulation of stable mRNA at the beginning of oocytes’ development, which becomes a reservoir for their further use to support the oocytes’ maturation and embryogenesis.…”

“…This is evolutionarily possible due to the accumulation of stable mRNA at the beginning of oocytes’ development, which becomes a reservoir for their further use to support the oocytes’ maturation and embryogenesis. Silencing of transcriptional activity is believed to begin in the GV phase and is not fully reversed until fertilization [ 35 ]. Our work showed that the mRNA level of DNMT3A was significantly higher in DOs regardless of the effect of cryopreservation.…”