Effect of Gamete Cryopreservation on miRNA Expression and Embryonic Development

Background: Fertility preservation is a meaningful option to retain fertility due to the medical or social needs. Sperm or oocytes preservation is the most extensive way for personal fertility preservation. Although the cryopreservation of sperm is well-used while the oocyte cryopreservation technology is improving, the effects and embryonic development potential of gamete cryopreservation need further investigation. Methods: The miRNAs expression of cryopreserved gamete was detected using miRNA sequencing in human and mouse, differentially expressed miRNAs (DEmiRs) were identified by comparing their expression in frozen-thawed and fresh gamete. Function of DEmiRs were detected by GO enrichment analysis, and common DEmiRs were validated by qRT-PCR. The embryonic developmental potential of cryopreserved sperm or oocytes were estimated in in vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI) cases by retrospective cohort study. Results: 21 and 106 DEmiRs were detected in human and mouse frozen-thawed sperm compared with fresh group, respectively. miR-148b-3p and miR-30b-5p were validated that downregulation in both human and mouse frozen-thawed sperm, while miR-140-5p was upregulated, these 3 common DEmiRs were enriched in positive regulation of mitotic cell cycle, in utero embryonic development and others. Meanwhile, 285 DEmiRs were identified in mouse frozen-thawed oocytes, the number of DEmiRs was 2.61-fold than that in mouse sperm. The downregulation of miR-148b-3p and miR-30b-5p were also validated in mouse frozen-thawed oocytes. Clinic data of embryonic development which used cryopreserved sperm indicated the significant decrease of fertilization rete in both IVF and ICSI cases, as well as the decrease of blastocyst formation rate in ICSI cases, while the significant decrease of high qualified embryo rate was observed when used the cryopreserved oocytes compared with paired fresh oocytes which retrieved from the same female in ICSI cases, suggesting the potential negative effects of gamete cryopreservation on embryonic development. Conclusion: Cryopreservation could change the expression of miRNAs in both sperm and oocyte, and the altered miRNAs expression pattern further affected the fertilization and development of embryo. Funding Statement: This research was funded by National Key R&D Program of China (2017YFC1001300). The National Natural Science Foundation of China (81771651, 81873832). The Research Projects of Shanghai Science and Technology Commission (18411964300). Declaration of Interests: The authors declare there are no competing interests. Ethics Approval Statement: The animal experimental procedures were approved by the Institutional Animal Care and Use Committee of Tongji University and the Ethics Committee of the Institute of Animal Science, Tongji University (TJAA06420101). Human sperm samples were donated for research from patients who had provided informed consent at Tongji hospital, with the approval of the Institutional Ethical Committee for Scientific Research (2019-062).