Fig. 2: h-HepOrg gene expression resembles that of in vivo hepatocytes.
From: Human assembloids recapitulate periportal liver tissue in vitro
a–c, h-HepOrgs expanded in EM2 or differentiated in DM or liver tissue was analysed for expression of the indicated markers. Representative images are shown from at least two independent donors from three independent experiments. a, Immunofluorescence staining for Ki-67 (magenta) in h-HepOrgs in EM2 (top) and DM (bottom). DAPI (cyan) stained nuclei. Scale bars, 50 μm. b, Immunofluorescence staining for YAP (magenta), nuclei (DAPI; cyan) and cell borders (F-actin; grey) in liver tissue (top) and h-HepOrgs in EM2 with (middle) and without (bottom) TRULI. Scale bars, 50 μm. White arrowheads indicate cells with nuclear YAP. Yellow arrowheads indicate YAP-negative cells. c, HNF4A (magenta), bile canaliculus (BC) marker CD13 (green) and F-actin (grey) immunofluorescence staining of organoids in EM2 (left) or DM (right). h-HepOrgs in DM exhibit thinner and more interconnected bile canaliculi. Scale bars: 50 μm. d,e, RNA-seq analysis of h-HepOrgs in EM2 or DM, h-CholOrgs and PHHs either freshly isolated (primary PHHs) or cultured for 1 day (1d-PHH monolayer). d, Heat map representing scaled gene expression. BA, bile acid; Cholest., cholesterol; met., metabolism. e, GSEA of h-HepOrgs in DM compared with EM2. Full list in Supplementary Data 2. Results are shown as a dot plot, where dot colour corresponds to the adjusted P value (permutation test implemented in clusterProfiler, adjusted using the Benjamini–Hochberg method). Dot size corresponds to the gene ratio (the number of core enrichment genes divided by the total number of genes in the pathway). KEGG, Kyoto Encyclopaedia of Genes and Genomes; NES, normalized enrichment score. f, Human liver tissue (left) and an h-HepOrg in DM (right) stained for pericentral (GS; magenta, top) and periportal (HAL; yellow, bottom) markers. Nuclei were stained with DAPI (cyan). Fire LUT images (first column) show fluorescence intensities for GS and HAL. CV, central vein; PV, portal vein. Scale bars, 100 μm (tissue) and 50 μm (organoids). Representative images from three independent experiments.
