Fig. 4: Identification of the cervical cell subpopulation enriched for human cervical stem cells.

a Immunofluorescence staining of ITGB4 and CD24 in human cervical epithelium. Scale bars: 100 μm. b Immunofluorescence staining of ITGB4 and CD24 in human cervical organoids. c Flow cytometry analysis of primary human cervical epithelial cells using ITGB4 and CD24. Expression of basal markers (d), cell cycle genes (e), and suprabasal markers (f) in each subpopulation assessed by qPCR (n = 3). g Cell cycle analysis of each subpopulation. The percentage of cells in S/G2/M phases in each subpopulation was measured (n = 5). Cervical organoids generated from each subpopulation and bulk sorted cells. Representative bright-field images (h) and number (i) of organoids (n = 4). j Representative H&E staining of the human stratified squamous epithelial tissues regenerated from each subpopulation of human ectocervical cells transplanted inside immunocompromised murine tongues. k Frequency of human ectocervical tissue regeneration from each subpopulation of cells. Transplanted murine tongues were harvested at 3–4 weeks after transplantation. l Pseudo-temporal trajectory plot showing the order of cell transitions at the cluster level. The black line starts at basal cluster and ends at suprabasal cluster II. m Density dot plot showing the number of cells in each cluster over time. n Single-cell trajectory reconstructed by Monocle for epithelial cells. o Schematic model of human cervical stem cell differentiation. p Representative flow cytometry analysis of primary HPV-infected human cervical epithelial cells using ITGB4 and CD24. Comparison of the ratios of ITGB4⁺CD24⁻ (q), ITGB4⁻CD24⁻ (r), ITGB4⁻CD24⁺ (s) cells between six HPV-infected and eleven HPV-uninfected normal cervical epithelia. All data are collected from indicated biological replicates and presented as mean ± SEM (*p < 0.05; **p < 0.01; ***p < 0.001). P-values were calculated by one-way ANOVA with Tukey’s multiple comparison test (d–g, i) or two-tailed unpaired Student’s t-test (q–s). Source data and exact p-values are provided as a Source Data file. Scale bars: 100 μm.