Fig. 1: Development and characterization of human ecto-and endo-cervix chips.

a Schematic top (left) and cross-sectional views (right) of the dual channel microfluidic organ chip lined by human cervical epithelium interfaced across an ECM-coated porous membrane with human cervical fibroblasts. b Phase-contrast microscopic view from above of a Cervix Chip lined by cervical epithelium and fibroblasts on the apical and basal sides of the chip porous membrane, respectively, on day 1 (left) as well as after 12 days of culture under continuous (middle) or periodic (right) flow (bar, 200 μm). c Side view, dark field images of living Cervix Chip hours after cell seeding day 0 (top) and after 7 days of differentiation (bottom). White arrow indicates light reflective mucus accumulating above the differentiated epithelium (bar, 1 mm). d Fluorescence microscopic side views of mucus layers in live Cervix Chip cultures stained with fluorescent wheat germ agglutinin (WGA) (green) on day 7 of differentiation under continuous (top) and periodic (bottom) flow regimens (bar, 1 mm). e Immunofluorescence microscopic view from above of the cervical epithelium stained for mucin 5B (MUC5B, green), F-actin (yellow), and nuclei with Hoechst (blue) (bar, 50 μm) stained at day 7 of differentiation. f Immunofluorescence microscopic vertical cross sectional view of the chip showing the cervical epithelium stained for MUC5B (green) overlaying the porous membrane, the underlying layer of fibroblasts stained for vimentin (yellow), and Hoechst-stained nuclei (blue) (bar, 20 μm). g Quantification of the total mucin content produced by Cervix Chips cultured under continuous (black symbols) or periodic (white symbols) flow measured at days 0 and 7 of differentiation. h Cytokine proteins in effluents of Cervix Chips cultured under continuous (gray bars) or periodic flow (white bars) on day 0 versus 7 of differentiation. i RNAseq analysis of genes expressed in cervical epithelial cells from three different donors differentiated in static Transwells or Cervix Chips under periodic or continuous flow. Expression levels of the signature genes associated with the endo- and ecto-cervical epithelial cells⁵⁹ were compared using Z-scores calculated per donor across samples for the three flow conditions, emphasizing common trends among samples of different donors (colors represent gene expression levels; each column represents one of the three donors). Data represent the mean ± s.e.m.; n = 3 (periodic flow) and 5 (continuous flow) (g), 4 (continuous flow) and 5 (periodic flow) (h), and 3 (i) experimental chip replicates. Micrographs in (b–f) are representatives of three separate experiments. Source data and statistical tests are provided as a Source Data file. Left image in (a) was created with BioRender.com.