Purpose The receptive endometrium is critical for blastocyst implantation. In mice, after blastocysts enter the uterine cavities on day 4 of pregnancy (day 1 = vaginal plug), blastocyst attachment is completed within 24 h, accompanied by dynamic interactions between the uterine luminal epithelium and the blastocysts. Any failures in this process compromise subsequent pregnancy outcomes. Here, we performed comprehensive analyses of gene expression at the luminal epithelium in the peri‐implantation period. Methods RNA‐seq combined with laser microdissection (LMD) was used to reveal unique gene expression kinetics in the epithelium. Results We found that the prereceptive epithelium on day 3 specifically expresses cell cycle‐related genes. In addition, days 3 and 4 epithelia express glutathione pathway‐related genes, which are protective against oxidative stresses. In contrast, day 5 epithelium expresses genes involved in glycolysis and the regulation of cell proliferation. The genes highly expressed on days 3 and 4 compared to day 5 are related to progesterone receptor signaling, and the genes highly expressed on day 5 compared to days 3 and 4 are associated with the ones regulated by H3K27me3. Conclusions These results suggest that specific gene expression patterns govern uterine functions during early pregnancy, contributing to implantation success.

Adenomyosis is a benign uterine disease and causes dysmenorrhea, heavy menstrual bleeding, and infertility; however, its pathophysiology remains unclear. Since signal transducer and activator of transcription 3 (STAT3) is crucial for endometrial regeneration, we hypothesized that STAT3 participates in adenomyosis pathophysiology. To investigate the influence of STAT3 on adenomyosis development, this study was performed using a novel mouse model of adenomyosis and human specimens of eutopic endometria and adenomyosis lesions. We established a novel mouse model of adenomyosis by puncturing entire mouse uterine layers with a thin needle. Mouse eutopic and ectopic endometria showed a positive immunoreactivity for phosphorylated STAT3 (pSTAT3), the active form of STAT3. Decreased numbers of adenomyotic lesions and reduced expression of Cxcl1, Icam1, and Spp1, which are associated with immune cell chemotaxis and tissue regeneration, were observed in uterine Stat3-deficient mice compared to the controls. In humans, pSTAT3 was intensely expressed at both the eutopic endometrium and the adenomyotic lesions regardless of the menstrual cycle phases. Conversely, it was limitedly expressed in the eutopic endometrium during the menstrual and proliferative phases in women without adenomyosis. Our findings indicate that continuous STAT3 activation promotes adenomyosis development. STAT3 inhibition can be a promising treatment strategy in patients with adenomyosis.

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Recurrent implantation failure is a major problem in assisted reproductive technology (ART). Although ART systems have evolved rapidly over the decades, it is still difficult to diagnose uterine conditions suitable for embryo transfer (ET) without the use of invasive endometrial procedures. Previous studies in mice showed that leukemia inhibitory factor (LIF) is a well-known endometrial biomarker for uterine implantation capacity, also known as uterine receptivity. This study focused on LIF in the mouse and human cervix as a possible biomarker of implantation capacity. We found that high expression of LIF in the cervical epithelium is strongly correlated with that of the uterine epithelium during the peri-implantation period in mice. Likewise, human cervical epithelia also exhibit elevated levels of LIF in the peri-implantation period. In addition, cervical LIF is downregulated in mice with defective implantation caused by pharmacological treatments. These results indicated that cervical LIF is a possible biomarker that detected uterine receptivity without invasive endometrial damage.

Infertility occurs in 15% of couples worldwide. Recurrent implantation failure (RIF) is one of the major problems in in vitro fertilization and embryo transfer (IVF–ET) programs, and how to manage patients with RIF to achieve successful pregnancy outcomes remains unresolved. Here, a uterine polycomb repressive complex 2 (PRC2)-regulated gene network was found to control embryo implantation. Our RNA-seq analyses of the human peri-implantation endometrium obtained from patients with RIF and fertile controls revealed that PRC2 components, including its core enzyme enhancer of zeste homolog 2 (EZH2)-catalyzing H3K27 trimethylation (H3K27me3) and their target genes are dysregulated in the RIF group. Although fertility of uterine epithelium-specific knockout mice of Ezh2 (eKO mice) was normal, Ezh2-deleted mice in the uterine epithelium and stroma (uKO mice) exhibited severe subfertility, suggesting that stromal Ezh2 plays a key role in female fertility. The RNA-seq and ChIP-seq analyses revealed that H3K27me3-related dynamic gene silencing is canceled, and the gene expression of cell-cycle regulators is dysregulated in Ezh2-deleted uteri, causing severe epithelial and stromal differentiation defects and failed embryo invasion. Thus, our findings indicate that the EZH2–PRC2–H3K27me3 axis is critical to preparing the endometrium for the blastocyst invasion into the stroma in mice and humans.