BioProject

Building on the success of previous pregnancies achieved through blastocyst transfer into D0.5 oviducts or D2.5 uteri. This study aimed to transfer well-developed blastocysts into D1.5 recipient uteri. However, the implantation was unsuccessful. To understand the underlying reasons for this failure. Embryos were transferred into D1.5 (Group A), D2.5 (Group B) and D3.5 (Group C) recipients, and recovered for observation within 3 hours. Subsequently, Multibarcod single-cell RNAseq analysis was conducted on both poor-quality blastocysts after transfer and well-developed blastocysts. Our findings revealed that embryo recovery rate in Group A declined significantly compared to Groups B and C, meanwhile poor-quality embryos rate in Group A reached 68.67% (p<0.01). Transcriptome analysis was conducted on well-developed and poor-quality blastocysts, which revealed the Top 20 significantly up-regulated genes were closely related to cellular and functional regulation. And Top 20 down-regulated genes predominantly pertained to cellular DNA damage repair and structural maintenance. Both GO and KEGG analyses revealed that these differentially expressed genes were predominantly enriched in transcriptional regulation and cell cycle. GSEA analysis uncovered that the basic transcription factor pathway was suppressed, while the apoptosis, lysosome, and ubiquitin-mediated protein degradation pathways were all activated. These findings suggested that when blastocysts were transferred into D1.5 recipient mouse, the embryos' cellular structure and DNA may sustain damage due to the specific uterine environment. This damage not only impaired normal transcriptional activities but also triggered apoptosis by activating the apoptosis pathway, thereby impeding the embryo's further development towards the implantation window and ultimately resulting in implantation failure.