miR-340-3p-modified bone marrow mesenchymal stem cell-derived exosomes inhibit ferroptosis through METTL3-mediated m6A modification of HMOX1 to promote recovery of injured rat uterus

Abstract Background Ferroptosis is associated with the pathological progression of hemorrhagic injury and ischemia-reperfusion injury. According to our previous study, exosomes formed through bone marrow mesenchymal stem cells modified with miR-340-3p (MB-exos) can restore damaged endometrium. However, the involvement of ferroptosis in endometrial injury and the effect of MB-exos on ferroptosis remain elusive. Methods The endometrial injury rat model was developed. Exosomes were obtained from the supernatants of BMSCs and miR-340/BMSCs through differential centrifugation. We conducted RNA-seq analysis on endometrial tissues obtained from the PBS and MB-exos groups. Ferroptosis was induced in ESCs by treating them with erastin or RSL3, followed by treatment with B-exos or MB-exos. We assessed the endometrial total m⁶A modification level after injury and subsequent treatment with B-exos or MB-exos by methylation quantification assay. We performed meRIP-qPCR to analyze m⁶A modification-regulated endogenous mRNAs. Results We reveal that MB-exos facilitate the injured endometrium to recover by suppressing ferroptosis in endometrial stromal cells. The injured endometrium showed significantly upregulated N⁶-methyladenosine (m⁶A) modification levels; these levels were attenuated by MB-exos through downregulation of the methylase METTL3. Intriguingly, METTL3 downregulation appears to repress ferroptosis by stabilizing HMOX1 mRNA, thereby potentially elucidating the mechanism through which MB-exos inhibit ferroptosis in ESCs. We identified YTHDF2 as a critical m⁶A reader protein that contributes to HMOX1 mRNA degradation. YTHDF2 facilitates HMOX1 mRNA degradation by identifying the m⁶A binding site in the 3ʹ-untranslated regions of HMOX1. In a rat model, treatment with MB-exos ameliorated endometrial injury-induced fibrosis by inhibiting ferroptosis in ESCs. Moreover, METTL3 short hairpin RNA-mediated inhibition of m⁶A modification enhanced the inhibitory effect of MB-exos on ferroptosis in endometrial injury. Conclusions Thus, these observations provide new insights regarding the molecular mechanisms responsible for endometrial recovery promotion by MB-exos and highlight m⁶A modification-dependent ferroptosis inhibition as a prospective therapeutic target to attenuate endometrial injury.