Abstract
Although microRNAs (miRNAs) expressed in cumulus cells (CCs) may be used to select competent oocytes/embryos, only a limited number of such miRNAs has been reported. To identify more miRNAs that regulate cumulus expansion (CE) and CC apoptosis, we first established that mouse cumulus-oocyte complexes (COCs) cultured in expansion-supporting medium supported full CE while undergoing mild apoptosis, whereas mouse oocytectomized COCs (OOXs) cultured in apoptosis-triggering medium underwent severe apoptosis while supporting no CE. RNA- and miRNA-sequencing and bioinformatics using CCs from these cultured COCs/OOXs identified candidate apoptosis- and/or CE-regulating miRNAs. Transfection of COCs/OOXs with miRNA mimic or inhibitor validated that miR-212-5p and 149-5p promoted CE by facilitating Has2 expression; miR-31-5p and 27a-3p promoted CE by increasing both Has2 and Ptx3 expression; and miR-351-5p and 503-5p inhibited CE by suppressing Ptx3 expression. Furthermore, miR-212-5p, 149-5p and Nov798 inhibited CC apoptosis, involving both Bcl2/Bax and Fas signaling. Analysis using in vivo matured COCs further verified the above apoptosis- and/or CE-regulating miRNAs, except for miR-149-5p. In conclusion, this study identified and validated new CE- and apoptosis-regulating miRNAs in CCs, which could be used as biomarkers to select competent oocytes/embryos and for elucidating how the oocyte-derived factors regulate CE and CC apoptosis.
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