Fig. 3: Re-differentiation of dediHeps into functional and transplantable hepatocytes in 3D culture.

a Representative morphology of a dediHep aggregate at the presented hours (h) and days (d) after initiation of 3D culture. Inset, the enlarged micrograph of aggregate. b Co-immunofluorescence staining (E-cad with Alb, Vim, or Ki67) of dediHep aggregates 5 days after initiation of 3D culture. c Volcano plot depicting differences in gene expression between dediHep aggregates and dediHep monolayer cultures. Each dot represents one gene. Blue and red dots represent down- and up-regulated genes, respectively, in dediHep aggregates. Gray dots represent genes that are not significantly differentially expressed. d Top five most significantly enriched pathways associated with down- or up-regulated genes in dediHep aggregates compared to dediHep monolayer cultures, identified by KEGG pathway enrichment analysis. e Immunohistochemical staining of Fah and co-immunofluorescence staining of YFP with Fah on liver sections of Fah^-/- mice 3 months after transplantation of cells dissociated from dediHep aggregates derived from TM-treated Alb-CreER^T2;R26R^YFP/+ mice. f A representative micrograph of Fah⁺ cells on liver sections of Fah^-/- mice 3 months after transplantation of hepatocytes freshly isolated from the liver of adult wild-type mice. Graph at right depicts percentages of Fah⁺ cells in CK8/18⁺ epithelial cells on liver sections of Fah^-/- mice 3 months after transplantation of cells dissociated from dediHep aggregates or hepatocytes. Data represent the mean ± SEM (n = 3 independent experiments). DNA was stained with DAPI. Scale bars, 200 μm (a, e, and f) and 20 μm (b). Statistical difference was determined by quasi-likelihood F-test (c), Fisher’s exact test (d), or two-sided Student’s t test (f). Source data are provided as a Source Data file.