Multifunctional N6-methyladenosine (m6A) has been revealed to be an important epigenetic component in various physiological and pathological processes, but its role in female ovarian aging remains unclear. Thus, we demonstrated m6A demethylase FTO downregulation and the ensuing increased m6A in granulosa cells (GCs) of human aged ovaries, while FTO-knockdown GCs showed faster aging-related phenotypes mediated. Using the m6A-RNA-sequence technique (m6A-seq), increased m6A was found in the FOS-mRNA-3′UTR, which is suggested to be an erasing target of FTO that slows the degradation of FOS-mRNA to upregulate FOS expression in GCs, eventually resulting in GC-mediated ovarian aging. FTO acts as a senescence-retarding protein via m6A, and FOS knockdown significantly alleviates the aging of FTO-knockdown GCs. Altogether, the abovementioned results indicate that FTO in GCs retards FOS-dependent ovarian aging, which is a potential diagnostic and therapeutic target against ovarian aging and age-related reproductive diseases.

The microbiome of females undergoes extensive remodeling during pregnancy, which is likely to have an impact on the health of both mothers and offspring. Nevertheless, large-scale integrated investigations characterizing microbiome dynamics across key body habitats are lacking. Here, we performed an extensive meta-analysis that compiles and analyzes microbiome profiles from >10,000 samples across the gut, vagina, and oral cavity of pregnant women from diverse geographical regions. We have unveiled unexpected variations in the taxonomic, functional, and ecological characteristics of microbial communities throughout the course of pregnancy. The gut microbiota showed distinct trajectories between Western and non-Western populations. The vagina microbiota exhibited fluctuating transitions at the genus level across gestation, while the oral microbiota remained relatively stable. We also identified distinctive microbial signatures associated with prevalent pregnancy-related disorders, including opposite variations in the oral and gut microbiota of patients with gestational diabetes and disrupted microbial networks in preterm birth. This study establishes a comprehensive atlas of the pregnancy microbiome by integrating multidimensional datasets and offers foundational insights into the intricate interplay between microbes and host factors that underlie reproductive health.

The microbiota in the human body play critical roles in many physiological and pathological processes. However, the diversity and dynamics of the female genital tract (FGT) microbiota have not been fully unveiled. In this study, we characterized the microbiome variations in reproductive-aged Chinese women, and we revealed that the cervicovaginal microbiota were dominated by Lactobacillus. Overall, the composition of microbiota in the uterine cavity was more diverse than that in the vagina and cervix. A positive correlation between Lactobacillus iners and Lactobacillus crispatus was observed in both the vagina and the cervix, suggesting that these two species might have a symbiotic relationship in the cervicovaginal microbiota. Moreover, we, for the first time, stratified the reproductive-aged Chinese women into subgroups, based on their microbiome profiles. Furthermore, we identified the bacteria whose abundance changed in the uterine cavity of infertile patients when compared with healthy controls, such as L. iners and L. crispatus. Functionally, the metabolism-related pathways, neurotrophin signaling pathway, and adipocytokine signaling pathway were predominantly dysregulated in the uterine cavity of infertile patients. In conclusion, we characterized a comprehensive microbial landscape in FGT, as well as their functional roles in female infertility of the Chinese population.

The rise in infertility rates has prompted research into the impact of vaginal microbiota on female fertility and the success of assisted reproductive technology (ART). Our study aimed to compare the vaginal microbiome between fertile and infertile women and explore its influence on ART outcomes. Vaginal secretion samples were collected from 194 infertile women and 100 healthy controls at Shanghai Changzheng Hospital. The V3–V4 region of the 16S rRNA gene was amplified using polymerase chain reaction (PCR). A machine learning model was applied to predict infertility based on genus-level abundance, and the PICRUSt algorithm was employed to predict metabolic pathways related to infertility and ART outcomes. The results showed that infertile women exhibited a significantly different vaginal microbial composition compared to healthy controls, along with increased microbial diversity. Notably, the abundance of Burkholderia, Pseudomonas, and Prevotella was significantly elevated in the vaginal microbiota of the infertility group, while that of Bifidobacterium and Lactobacillus was reduced. Among infertile women, those with recurrent implantation failure (RIF) showed even higher vaginal microbial diversity, with specific genera such as Mobiluncus, Peptoniphilus, Prevotella, and Varibaculum being more abundant. Eleven metabolic pathways were identified to be associated with both RIF and infertility, with Prevotella showing stronger correlations with these pathways. This study elucidates differences in vaginal microbiome between healthy and infertile women, providing novel insights into how vaginal microbiota may impact infertility and ART outcomes. Our findings underscore the importance of specific microbial taxa in women with RIF, suggesting potential avenues for targeted interventions to improve embryo transplantation success rates.

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 bone marrow mesenchymal stromal cells (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 endometrial stromal cells (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 3′-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.

Ovarian aging is a complex process that compromises fertility and elevates the risk of reproductive disorders. To elucidate its spatiotemporal dynamics, we integrated single‐nucleus RNA sequencing and spatial transcriptomics to construct a comprehensive aging atlas of 12 human ovarian tissues spanning ages 12–54 (prepubertal, age 12, n = 1; young, ages 23–29, n = 4; middle‐aged, ages 32–34, n = 2; and older‐aged, ages 42–54, n = 5). Our analysis revealed aging‐related transcriptomic shifts, including impaired mitochondrial oxidative phosphorylation and reproductive structure development in aged human ovaries. We identified a novel endothelial cell (EDC) subtype, CLDN5 ⁺ blood EDCs, which exhibited unique functional specialization as semiprofessional antigen‐presenting cells. In contrast to other cell types that lost cell identity during aging, CLDN5 ⁺ blood EDCs displayed transcriptomic sensitivity to aging, characterized by enhanced antigen‐presenting capabilities, and heightened inflammatory activity. Spatial mapping further uncovered immunoglobulin‐expressing ( IGHG1 ⁺ / IGKC ⁺ ) cell accumulation in the ovarian periphery, correlating with advancing age. Critically, aging disrupted global cellular connectivity while amplifying the DLK1:NOTCH3 axis between theca cells and CLDN5 ⁺ blood EDCs, which may contribute to the dysregulation of ovarian functions. We also detected the upregulation of DLK1 in granulosa cells from patients with primary ovarian insufficiency. This study significantly enhances our comprehension of the underlying mechanisms of human ovarian aging and concurrently pinpoints potential therapeutic avenues for addressing related disorders.