Umbilical cord mesenchymal stem cell-derived extracellular vesicles improve excessive autophagy of granulosa cells through <i>METTL3</i>

Polycystic ovary syndrome (PCOS) is a prevalent endocrine disorder impacting women's fertility. We assessed the effect of umbilical cord mesenchymal stem cell-derived extracellular vesicles (UC-MSC-EVs) on PTEN-induced kinase 1 (<i>PINK1</i>)/<i>Parkin</i>-mediated excessive autophagy of ovarian granulosa cells (GCs) through methyltransferase-like 3 (<i>METTL3</i>). Human ovarian GC line KGN was cultured and treated with dehydroepiandrosterone (DHEA) and UC-MSC-EVs. Cell apoptosis and viability, autophagy-related protein levels, adenosine triphosphate (ATP) and mitochondrial membrane potential (MMP) level, and microtubule-associated protein 1 light chain 3 β (LC3B) and translocase of outer mitochondrial membrane 20 (TOMM20) colocalization were assessed by flow cytometry, CCK-8, Western blot, kit, and immunofluorescence. <i>PINK1</i> N6-methyladenosine (m6A) modification, <i>METTL3</i> levels, and <i>PINK1</i> mRNA stability were determined by methylated RNA immunoprecipitation, reverse transcription quantitative polymerase chain reaction, and Western blot. The PCOS mouse model was established and treated with UC-MSC-EVs. Serum hormone and ovarian tissue autophagy-related protein levels were determined by enzyme-linked immunosorbent assay. DHEA decreased KGN cell viability and p62 level, increased <i>PINK1</i>, <i>Parkin</i>, LC3BII/I, and Beclin-1 protein levels, ATP content, MMP level, TOMM20⁺LC3B⁺ cell number, and apoptosis, which were partly abrogated by UC-MSC-EV treatment. PINK1 had m6A modification sites. METTL3 was a <i>PINK1</i> m6A-modified writer protein. After DHEA treatment, KGN cells showed elevated <i>METTL3</i> and <i>PINK1</i> m6A modification levels and mRNA stability, whereas UC-MSC-EV treatment caused the opposite results. METTL3 overexpression partly averted UC-MSC-EVs-improved <i>PINK1</i>/<i>Parkin</i>-mediated mitophagy. UC-MSC-EVs curbed PINK1/Parkin-mediated excessive autophagy through <i>METTL3</i> and improved ovarian function in PCOS mice. In conclusion, UC-MSC-EVs suppressed PINK1/Parkin-mediated mitophagy of ovarian GCs through METTL3, thereby improving PCOS.<b>NEW & NOTEWORTHY</b> Polycystic ovary syndrome (PCOS) is a prevalent endocrine disorder impacting women's fertility. The authors in this study using DHEA-induced granulosa cells (GCs) demonstrated that umbilical cord mesenchymal stem cell-derived extracellular vesicles (UC-MSC-EVs) suppressed <i>PINK1</i>/<i>Parkin</i>-mediated mitophagy of ovarian GCs through METTL3, thereby improving PCOS.