DNA methylation and gene expression changes in mouse pre- and post-implantation embryos generated by intracytoplasmic sperm injection with artificial oocyte activation

Abstract

Background The application of artificial oocyte activation (AOA) after intracytoplasmic sperm injection (ICSI) is successful in mitigating fertilization failure problems in assisted reproductive technology (ART). Nevertheless, there is no relevant study to investigate whether AOA procedures increase developmental risk by disturbing subsequent gene expression at different embryonic development stages. Methods We used a mouse model to explore the influence of AOA treatment on pre- and post-implantation events. Firstly, the developmental potential of embryos with or without AOA treatment were assessed by the rates of fertilization and blastocyst formation. Secondly, transcriptome high-throughput sequencing was performed among the three groups (ICSI, ICSI-AOA and dICSI-AOA groups). The hierarchical clustering and Principal Component Analysis (PCA) analysis were used. Subsequently, Igf2r/Airn methylation analysis were detected using methylation-specific PCR sequencing following bisulfite treatment. Finally, birth rate and birth weight were examined following mouse embryo transfer. Results The rates of fertilization and blastocyst formation were significantly lower in oocyte activation-deficient sperm injection group (dICSI group) when compared with the ICSI group (30.8 % vs. 84.4 %, 10.0 % vs. 41.5 %). There were 133 differentially expressed genes (DEGs) between the ICSI-AOA group and ICSI group, and 266 DEGs between the dICSI-AOA group and ICSI group. In addition, the imprinted gene, Igf2r is up regulated in AOA treatment group compared to control group. The Igf2r/Airn imprinted expression model demonstrates that AOA treatment stimulates maternal allele-specific mehtylation spreads at differentially methylated region 2, followed by the initiation of paternal imprinted Airn long non-coding (lnc) RNA, resulting in the up regulated expression of Igf2r. Furthermore, the birth weight of newborn mice originating from AOA group was significantly lower compared to that of ICSI group. The pups born following AOA treatment did not show any other abnormalities during early development. All offspring mated successfully with fertile controls. Conclusions AOA treatment affects imprinted gene Igf2r expression and mehtylation states in mouse pre- and post-implantation embryo, which is regulated by the imprinted Airn. Nevertheless, no significant differences were found in post-natal growth of the pups in the present study. It is hoped that this study could provide valuable insights of AOA technology in assisted reproduction biology.

Good practice recommendations on add-ons in reproductive medicine

“…Recently, changes in DNA methylation and gene expression have been observed using ionomycin in a mouse model ( Yin etal. , 2021 ).…”

Long-run real-time PCR analysis of repetitive nuclear elements as a novel tool for DNA damage quantification in single cells: an approach validated on mouse oocytes and fibroblasts

“…Finally, development and optimization of biotechnically efficient L1-LORD-Q-mediated approaches to identify, in a highly detectable manner, the diminished molecular quality of single fibroblasts and/or oocytes might provide an excellent tool largely applicable to the negative selection of nuclear donor somatic cells and/or nuclear recipient germ cells that seem to be genetically undesirable/unsuitable for a wide variety of modern assisted reproductive technologies (ARTs) in mice and other mammalian species. This variety of ARTs encompasses such innovative strategies of in vitro embryo production (IVP) as somatic cell nuclear transfer (SCNT)-based cloning combined or not combined with transgenesis (Song et al 2020 ; Skrzyszowska and Samiec 2021 ; Wiater et al 2021 ; Moradi-Hajidavaloo et al 2023 ) and in vitro fertilization (IVF) by either gamete coincubation (Treleaven et al 2021 ; Gorczyca et al 2022 ; Montgomery et al 2023 ) or intracytoplasmic sperm injection (ICSI) (Yin et al 2021 ; Campos et al 2023 ; Ramírez-Agámez et al 2023 ).…”

Artificial oocyte activation with Ca2+ ionophore improves reproductive outcomes in patients with fertilization failure and poor embryo development in the previous ICSI cycles

“…Problems with oocyte-derived and de ciencies in downstream signaling pathways may be the main cause of infertility in women of advanced age and POI [28]. Furthermore, agerelated defects may lead to mitochondrial DNA alignment, accompanied by mitochondrial dysfunction and changes in gene expression pro le [29]. However, these studies suggested that AOA may not improve embryo development with advanced age and POI patients, but may improve fertilization rates and thus potentially provide more transferable embryos [17].…”