P-204 Transcriptome analysis of cell-free RNAs in embryonic culture medium reveals their potentials on non-invasive testing

Abstract

Abstract Study question Could cell-free RNA (cfRNA) in culture medium be non-invasive strategy to select in vitro fertilized (IVF) embryos? Summary answer CfRNAs in culture medium could be potential markers correlated with embryo qualities. What is known already Infertility is a critical component of reproductive health and couples with infertility may choose assisted reproductive technology (ART) to increase their chances of conception. However, the effectiveness remains low. One way to improve the outcome is to screen the embryo prior to implantation. Noninvasive testing opens new perspectives in analyzing embryo qualities and current pending includes microscopy images coupled with artificial intelligence and omics analysis of the spent blastocyst medium (SBM). Cell-free DNA in SBM could be used to identify the embryo ploidy and metabolomic/proteomic markers could be correlated with embryo potential, while the research of cfRNA is lacking. Study design, size, duration We developed a new library method of polyadenylation ligation-mediated sequencing (PALM-Seq) to capture the low amount of cfRNA and collected twelve culture mediums of in vitro embryos on Day 3 and five samples of embryos on Day 5. Participants/materials, setting, methods We employed PALM-Seq on culture medium of embryos generated by IVF or Intracytoplasmic sperm injection (ICSI). Owing to the new method, we could comprehensively profile the cfRNAs, including miRNA, tRNA, piRNA, lncRNA and mRNA. The morphology and fragmentation of the recruited embryos were also recorded. Main results and the role of chance Generally, thousands of genes could be detected in all culture mediums and the number greatly increased in SBM. We identified several reported miRNA markers in our study, such as miR-21-5p and miR-92a-3p, while highly expressed miRNAs varied across samples and stages. We indeed found piRNA in mediums and several specific piRNAs had large amount of expression. In contrast, detected number and expression of tRNA were stable across samples. Besides of reported mRNA markers, we could profile the expression of all detected mRNAs and no differences were found between embryos with and without fragmentation. We found that the majority of mRNA fragments were shorter than 30 nucleotides. In addition, some granular-specific expressed mRNAs could be detected in medium on Day 3 and they decreased of medium on Day 5. Limitations, reasons for caution We have only conducted preliminary exploration in the culture medium of pre-implantation embryos and studies of larger sample size are needed. Wider implications of the findings Our research proved the feasibility of performing high-throughput sequencing of cfRNAs in SBM. It opens new perspectives on defining non-invasive markers and selection of embryos with the most potential. Our results also contribute a new insight on improving the treatment of infertility. Trial registration number 2018YFC1004900