Artificial environment impact: O2 concentration changes between IVM and IVF alter embryo production, metabolism, and epigenetic marks

ABSTRACT Creating an optimal in vitro cell culture environment requires careful simulation of all critical components, especially the gaseous atmosphere. Although it is well-documented that embryonic culture under low oxygen tension promotes embryonic development, little is known about the effect of changes in in vitro maturation (IVM) and fertilization (IVF) on the epigenome. This study explores the role of oxygen tension variation in the early stages of in vitro production of bovine embryos and its impact on oxidative stress and epigenetic remodeling. We initially validated our system by scrutinizing the epigenetic effects on bovine fibroblasts. We observed that cell cultures under 20% O2 exhibited reduced H3K9me2 levels in early passages, which stabilized with prolonged cultivation and elevated gene expression of HIF2a and KDM5C. Our results reveal that oocytes maturing in 20% O2 environments have heightened levels of reactive oxygen species (ROS) and glutathione (GSH), whereas blastocyst embryos maturing under reduced oxygen tension exhibit increased oxidative stress markers ( NRF2 , SOD1 , SOD2 ), with upregulated transcripts observed in epigenetic remodelers ( KDM5A , TET1 ). Elevated O2 levels in both IVM and IVF processes showcase improved embryo production. Maintaining consistent O2 levels at either 5% or 20% between IVM and IVF results in heightened GSH, reduced ROS, and increased levels of H3K9me2/3 in embryos. Finally, distinct DNA methylation patterns emerge, indicating higher levels in groups matured under low O2 tension and increased DNA hydroxymethylation in groups fertilized under low O2 tension. In conclusion, our comprehensive investigation underscores the critical role of oxygen concentration in shaping the epigenetic landscape during the early stages of in vitro culture. These findings provide valuable insights for optimizing conditions in assisted reproductive technologies.