Up-regulated microRNAs in blastocoel fluid of human implanted embryos could control circuits of pluripotency and be related to embryo competence

Purpose: The paper aims to investigate the biological role of microRNAs secreted by preimplantation embryo into the blastocoel fluid and to detect a distinctive molecular signature for identifying embryos with the highest implantation potential. Methods: We carried on a multicenter retrospective study involving five European IVF centers. We collected 112 blastocoel fluid samples from embryos on day 5 post-fertilization, cultured individually, along with data on blastocyst grade and embryo transfer outcomes. Using a custom TLDA Array, we compared the expression levels of 89 miRNAs between 33 fluids from high-quality implanted embryos and 30 fluids from high-quality not-implanted embryos. Expression differences were assessed using SAM and t-test. Additionally, correlation and function enrichment analysis and network construction were conducted to identify the biological roles of deregulated microRNAs. Results: We identified six up-regulated microRNAs in the blastocoel fluid from implanted embryos, significantly and positively correlated across all samples (r ≥ 0.7; P ≤ 0.05). They could take part in pluripotency circuits, regulating and being regulated by transcription factors associated with stemness, cell growth, and embryo development. The ROC curve analysis confirmed the potential of these miRNAs as implantation classifiers. Conclusion: The six miRNAs up-regulated in blastocoel fluid from implanted embryos may represent a functional molecular signature for evaluating blastocyst quality and identifying the most competent embryos. Their evaluation associated with non-invasive preimplantation genetic testing, integrating epigenetic and genomic analyses, could enhance implantation grade and allow for identification of the euploid embryo not able to implant.