Gynecology World Conference 2026

Abstract

Background: Adenomyosis (AM) is associated with subfertility and poor reproductive outcomes in the context of assisted reproductive technology (ART), yet the mechanisms underlying impaired endometrial receptivity during the window of implantation remain unclear. In particular, the spatially organised cellular crosstalk within the LH+7 eutopic endometrium has not been explored.

Objective: To characterize the single-cell landscape and spatial microenvironmental alterations of the LH+7 receptive endometrium in women with adenomyosis.

Materials and Methods: Eutopic endometrial samples were collected at LH+7 from women with adenomyosis (AM, n=4) and matched controls (CTR, n=4). Single-cell RNA sequencing (scRNA-seq) was performed on three paired samples using the 10x Genomics platform, followed by standard quality control, normalization, and dimensionality reduction. Cell populations were identified based on canonical marker gene expression. To resolve spatial organization, one matched AM–CTR pair was further analyzed using spatial transcriptomics (Stereo-seq), and scRNA-seq data were integrated with spatial profiles to map cell-type distributions and niche-specific gene expression patterns. Downstream analyses included differential gene expression, pathway enrichment, and cell–cell communication inference. Macrophage subpopulations were further characterized through subclustering and functional annotation. Key findings were validated by quantitative real-time PCR (qRT-PCR) in an independent cohort (n=6 per group) and by immunofluorescence staining to confirm spatial localization at the protein level.

Results: Single-cell analysis identified nine major cell populations in the LH+7 eutopic endometrium, with broadly comparable overall cell-type proportions between AM and CTR. Despite similar cellular composition, extensive transcriptional reprogramming was observed across multiple compartments. Differential expression analysis revealed that stromal and pericyte exhibited the highest number of DEGs (Stromal: 5217 upregulated and 3746 downregulated genes, Pericyte: 2955 upregulated and 919 downregulated genes) in AM. Cell–cell communication analysis demonstrated a global increase in interaction number (2633 vs 2232) and interaction strength (55.736 vs 49.046) in AM compared to CTR. Stromal–perivascular interactions emerged as the most prominently altered interaction axis in AM . Within the axis, both BMP8A and ACVR2A were significantly upregulated in AM (P<0.05) and were further validated by qRT-PCR in an independent cohort (n=6 per group). Macrophage heterogeneity was revealed by subclustering, with AM samples showing a shift toward an immunotolerance phenotype. Specifically, macrophages in AM exhibited an immunoregulatory (M2-like) phenotype, characterized by increased expression of CD163 and MRC1, alongside reduced pro-inflammatory M1-associated signatures, including decreased IL1B and CXCL8 expression. Gene set enrichment analysis further revealed altered metabolic signalling pathways (FDR<0.05) in AM macrophages, including reduced oxidative phosphorylation, ATP synthesis, and related mitochondrial pathways. Spatial transcriptomic analysis (Stereo-seq) localized these alterations to stromal and perivascular regions at LH+7, demonstrating spatially restricted disruption of cell-type-specific crosstalk within the receptive endometrium.

Conclusion: Single-cell and spatial transcriptomics reveal an aberrant microenvironment in the mid-secretory (LH+7) endometrium of adenomyosis. Abnormal stromal–perivascular cell-cell interaction, a shift in macrophage polarization and altered metabolic pathways may underlie impaired receptivity in adenomyosis.