Fig. 2: Human ovarian cortex contains two distinct populations of follicles.

120 follicles of normal morphology isolated from children and adults were analyzed via polyA-RNA and miRNA. After quality control, 109 and 113 follicles remained in the analysis, respectively. A Principal component analysis (PCA) and B uniform manifold approximation and projection (UMAP) visualization showed two follicle groups based on polyA-RNA expression, with child and adult follicles equally present. Group 1 included all developmental stages, while Group 2 had growing follicles only. C PCA based on miRNA expression did not group the follicles but aligned PC1 with developmental stages from primordial to secondary follicles. D Volcano plot showed a marked upregulation of genes in Group 1 follicles compared to Group 2, Wald test, adjusted using Benjamini–Hochberg (BH) method. E Similar expression profiles were found when specific follicle growth stages were compared; here, primary follicles of Group 1 vs Group 2. F Differentially expressed genes (DEGs) between Group 1 and Group 2 follicles included recognized oocyte (above red line) and granulosa (under red line) cell markers. Group 1 exhibited an upregulation of traditional oocyte markers, whereas Group 2 had upregulation of typical granulosa cell markers. G The top enriched Gene Ontologies (GO), KEGG, and Reactome pathways in Group 1 follicles suggested enrichment in reproductive processes, meiotic cell cycle, and steroidogenesis. Group 2 displayed enrichment in cell communication, developmental processes, and extracellular matrix organization, Fisher’s one-tailed test, adjusted using BH method. H When analyses were restricted to primary follicles only, similar results were obtained. I Immunofluorescence staining of adult and child ovarian cortical tissues revealed follicles with varying DDX4 oocyte marker expression (high expression, green arrows; low expression, blue arrows). J Quantification of the immunofluorescent signal from 343 follicles across two adults and one child presented a bimodal distribution, suggesting the presence of two follicle groups based on their DDX4 expression levels (red line). K Scatter plot of follicle-to-follicle communications in a 2D space. The dot size reflects the number of inferred links associated with each follicle group. Group 2 primary and intermediary follicles were predominant signal senders, while Group 1 primary follicles were the main signal receivers. NS- non-significant. Source data are provided as a Source Data file.