Fig. 6: Whole tissue imaging of EpCAM + IHBD development.

a IHBDs develop primarily as an interconnected web of EpCAM+ BECs from E15.5 through P14, originating at the hilum and extending to the periphery by P1, with rare disconnected BECs visible closer to the periphery at E15.5 (yellow arrowhead marks non-continuous BECs; scale bar represents 0.5 mm). b High-magnification images demonstrate discontinuous BECs at E15.5, which are absent by E17.5 (scale bars represent 150 μm; dashed blue lines denote portal veins). c At P1, early duct-ductule hierarchy begins to emerge as some IHBDs become enlarged but remain intertwined with web-like peripheral IHBDs (scale bars represent 150 μm; yellow arrowhead denotes enlarged IHBD; dashed blue lines denote portal veins). d Duct-ductule hierarchy is clearly established at P14, but large hilar ducts continue to be surrounded by web-like peripheral IHBDs (scale bars represent 150 μm). e Co-localization of Ki-67 and EpCAM reveals proliferative BECs throughout IHBD development (scale bar represents 50 μm; white asterisks indicate portal vein). f Quantification of BEC proliferation reveals abundant embryonic proliferation and decreasing, but high proliferation rates relative to adult tissue during postnatal IHBD development (p < 0.0001). g EpCAM+ BECs per portal field trend upwards E15.5-P5 until decreasing P14 and into adulthood (p = 0.0847). f, g significance indicates a p < 0.5, data are presented as mean values ± SEM, Tukey’s test after one-way ANOVA was performed, and letters indicate grouping by significance. Source data are provided as a Source Data file.