Profiling the Epigenetic State of Cell-Free DNA From IVF Embryos [ID 1044]

INTRODUCTION: In vitro fertilization (IVF) is a common form of assisted reproductive technology. While current embryo selection focuses on morphology and genetic aneuploidy, examining nucleic acids in blastocoel fluid may provide an additional molecular marker for selection. The project aims to assess the epigenetic state of cell-free DNA contained in the blastocoel fluid-conditioned media from day 5 human embryos to profile the post-translational modifications on histones associated with apoptosis. METHODS: Blastocoel fluid-conditioned media was collected from day 5 IVF blastocysts that underwent preimplantation genetic testing for aneuploidies. Media from euploid embryos (grouped by successful and unsuccessful implantation outcomes) and aneuploid embryos were pooled. Pooled samples were acid extracted to isolate histones present in the media (histone acid extraction kit, Active Motif). Histone samples were run on SDS-PAGE gels and Western blotted for histone modification-specific antibodies: anti-histone H3K4me3, anti-histone H3K9me3, anti-histone H3K27me3, and anti-histone H3K6ac (Active Motif). RESULTS: Western blot analysis detected the histone H3K27 trimethylation modification in the histones extracted from pooled euploid blastocoel fluid-conditioned media samples. CONCLUSIONS/IMPLICATIONS: This study shows that histone modifications, specifically H3K27 trimethylation, can be detected in pooled blastocoel fluid-conditioned media from euploid IVF embryos. H3K27 trimethylation is known to be associated with apoptosis and is crucial for early embryo development. Our finding that this apoptotic histone modification is present supports previous research from our laboratory on apoptotic gene expression in similar media. Future studies will investigate whether H3K27 trimethylation levels can serve as a marker for embryo viability based on implantation outcomes.