Effects And Mechanism Of In Vitro Fertilization On Embryo Implantation And Placental Development In Mice

Assisted reproductive technology(ART)including in vitro fertilization(IVF)and embryo transfer has been widely applied in the treatment of human infertility.The safety of assisted reproductive technology has received increasing attention.ART,including IVF,has been associated with increased miscarriage rates,placental defects,and metabolic diseases in offspring.Changes in epigenetic modification may be an important cause of abnormal embryonic development in IVF,but the mechanism is not clear.In this study,we compared the development of IVF embryos and normal embryos in mouse peri-and post-implantation.We found multiple developmental defects in the embryonic and extraembryonic tissues of IVF embryos,especially in the extraembryonic tissue before and after implantation,including reduced cell numbers,placental overgrowth,reduced placental efficiency and structural disorders.By analyzing the transcriptomes of IVF embryos and normal embryos,we found large number of developmental genes differentially expressed in both embryonic and extraembryonic tissues at embryonic day 7.5(E7.5)stage of IVF embryos.Further analysis revealed that two groups of up-regulated genes in IVF extraembryonic ectoderm(ExE)were mainly enriched in pathways related to embryonic development rather than extraembryonic development.Meanwhile,IVF manipulation notably disrupts tissue-specific gene expression in extraembryonic tissue,and 334 epiblast(Epi)-specific genes and 24 Epi-specific transcription factors were abnormally expressed in ExE of IVF embryos at E7.5.Data on histone H3K4me3 and H3K27me3 modifications before and after implantation in IVF and normal embryos were collected using the laboratory-improved ULI-NCh IP(Ultra-low-input micrococcal nuclease-based native chromatin immunoprecipitation)technique.We found that among the major differential histone modifications regions,the ExE of IVF embryos showed great similarity to Epi of NM embryos,and this similarity was due to ectopic increase of H3K4me3 modification.In addition,abnormal H3K4me3 modification located in the transcription start sites(TSS)and promoters induced the ectopic genes expression in E7.5 ExE of IVF embryos.Analysis the histone modification of 124 Epi-active promoters and 85 ExE-active promoters found that ectopic H3K4me3 modifications located in Epi-active promoters of IVF ExE results in abnormal gene expression.At the same time,abnormal widening of H3K4me3 modification in IVF ExE induced abnormal lineage identity.In order to further verify whether abnormally increased H3K4me3 modification causes abnormal embryonic development and gene expression,we interfered with the H3K4me3 demethylase Kdm5 b in normal embryos.We found that ectopic increase of H3K4me3 modification in normal embryos can lead to abnormal development of extraembryonic tissues,especially placenta,resulting in a series of developmental abnormalities similar to IVF embryos,such as reduced placental efficiency and disordered placental structure.Importantly,we demonstrate that knockdown of the H3K4me3-recruited regulator Kmt2 e,which is highly expressed in IVF embryos,greatly improves the development of IVF embryos and reduces abnormal gene expression in ExE.Knockdown the expression of Kmt2 e at the early stage can improve placental development in IVF embryos,including placental efficiency and placental tissue structure.Our study identifies that abnormal H3K4me3 modification in extraembryonic tissue is a major cause of implantation failure and abnormal placental development of IVF embryos.Meanwhile,our study provides theoretical basis and practical guidance for improving ART in clinical treatment.