Evaluation of differentially expressed microRNAs in vitrified oocytes by next generation sequencing

MicroRNAs (miRNAs) play crucial roles in gametogenesis and embryo development. The present study was undertaken to identify differentially expressed miRNAs and explore their functions in oocyte vitrification. Small RNA sequencing data revealed that 22 miRNAs were differentially expressed in vitrified oocytes compared with that of the fresh counterparts. The potential target genes for the differentially expressed miRNAs were enriched in anatomical development in biological process, cell in cellular components, and protein binding in molecular functions by gene ontology annotation analysis. In addition, endometrial cancer and metabolic pathway were the two pathways enriched in KEGG with the lowest p-value and highest count number genes, respectively. RT-qPCR data showed that miR-134-5p, miR-210-5p, and miR-21-3p were significantly up-regulated (P < 0.01), whereas miR-465c-5p was dramatically down-regulated (P < 0.01) in vitrified oocytes, which were consistent with that of the sequencing result. Moreover, the expression of potential target PTEN was significantly reduced both at transcriptional (P < 0.01) and post-transcriptional level (P < 0.01) in vitrified oocytes. The expression pattern of PTEN was negatively correlated with that of miR-21-3p. Dual-luciferase reporter assay further demonstrated that miR-21-3p could down-regulate PTEN by targeting its 3'UTR. In conclusion, our results demonstrated that specific miRNAs were differentially expressed in warmed oocytes, and decreased expression of PTEN was involved in response to vitrification stress.