Fig 2: Schematic illustration of the role of uterine epithelial and stromal STAT3 in uterine receptivity and embryo attachment. Epithelial STAT3 regulates the formation of slit-like structure in uterine lumen, and stromal STAT3 suppresses proliferative activity and estrogenic response in the epithelium, thus controlling uterine receptivity and embryo attachment. PGR, progesterone receptor; LIF, leukemia inhibitory factor; LIFR, LIF receptor.

Fig 3: Single-cell RNA-seq analysis of the NETB patient-derived organoids. (A) The t-SNE plot showing 17 clusters. The bar chart showing the proportion of each cluster. Each cluster was shown in a different color. (B) The t-SNE plot showing 3 cell types. The bar chart showing the proportion of each type. Each type was shown in a different color. (C) The heatmap separately displayed the expression proportions and relative expression levels of marker genes in three clusters. (D) The images of the ESR1, ESR2, PGR and ERBB2 expression. (E) The pseudotime trajectory analysis generated a single-branch trajectory that depicted the tumor’s development. (F) Different cell types in the trajectory were marked with different colors. (G) Heatmap showing all cells along the pseudo-time, which was clustered into three profiles. Color key differentially coding from blue to red indicated the relative expression levels from low to high. (H) The GO enrichment dot plot showing enriched pathways on the vertical axis and the ratio of differentially expressed genes annotated to GO terms to the total number of differentially expressed genes on the horizontal axis. The size of the dots represents the number of genes, and the color gradient from red to blue indicates the level of enrichment significance, with red representing higher significance and blue lower significance. (I) The KEGG enrichment dot plot displayed enriched pathways on the vertical axis and the ratio of differentially expressed genes annotated to KEGG pathway IDs to the total number of differentially expressed genes on the horizontal axis. The size of the dots indicates the number of genes, while the color gradient from red to blue represents the enrichment significance, with red indicating higher significance and blue indicating lower significance. (J) The network structure diagram corresponding to the protein-protein interaction analysis

Fig 4: A close interactive dialogue between endometrial stromal cells and epithelial cells. (A) Potential interactions between four FIB (FIBp, FIB1, FIB2, and FIB3) and four EC (Ciliated-EC, EC1, EC2 and EC3) subpopulations based on receptor-ligand pairs. The width of the line represented the number of receptor-ligand pairs. (B) Protein-protein interaction (PPI) network of IGF, endometrial receptivity-related genes and autophagy-related genes. Different edges color presented different interactions. (C) Protein-protein interaction network of IGF and endometrial receptivity-related genes and exosome-related gene. Different edges color presented different interactions. (D) Proportion of the four sub-clusters of EC in the Ctrl and RIF groups. (E) Expressions of CD63 and PGR in endometrial stroma from controls and RIF patients at the WOI time were analyzed by immunofluorescence staining (n = 6 for each group). Data were presented as mean ± SEM and analyzed by t test (***, p < 0.001). Scale bar, 20 µm (times of endometrial biopsy were showed in Table S5). (F) Volcano plot showing differentially expressed genes of ECs between the Ctrl and RIF groups. Red color indicates genes that are upregulated and blue color indicates downregulated genes in the RIF groups when compared with the Ctrl groups.

Fig 5: Single-cell RNA sequencing analysis of the PMBC organoids. Single-cell RNA sequencing was performed on the 7th generation organoids. a The t-SNE visualization of all cells in PMBC organoid sample, with clusters color-coded. The number in brackets is the cell number of the cluster. b Cells are annotated into three main types, with different colors representing different cell types. The bar chart shows the proportion of each cell type in the total cell count. Basal, Basal cells; Lumina epi, Luminal epithelial cells; Pro basal, Proliferating basal cells. c Dot plot of marker genes display. The FindAllMarkers module of Seurat software was used to identify marker genes for all clusters. The dot plot is horizontally represented by cluster ID and vertically by gene ID. Larger circles indicate a higher proportion of gene expression in the cluster, and redder colors indicate higher average expression levels of the gene. d The expression values of ESR1, PGR, and ERBB2 genes are mapped onto the t-SNE plot to show their expression levels in each cell. Darker red indicates higher expression