Fig. 3: Transport of hepatocyte metabolites in HBTO.

a Transport of fluorescein diacetate in HBTOs. Fluorescein derived from chloromethyl fluorescein diacetate (CMFDA) is secreted into BCs and then transported into the biliary network. After 10 min of incubation, fluorescein is strongly detected in the BC network of the hepatocyte cluster (HEP, surrounded by a broken line), as well as in the biliary network (BD). Fluorescein is absent from the hepatocyte clusters and reaches the biliary network after 120 min incubation. HBTOs were incubated in the presence of CMFDA for 10 min. Phase-contrast and fluorescent images were taken after washing with culture medium five times. HBTOs were kept in the culture medium without CMFDA. Dotted lines indicate the boundary between HEP and BD. The experiment on one well was repeated three times, independently. Two fields were examined in each well and the representative images are shown in this figure. Bars represent 100 μm. b Transport of bile acids in HBTO. CLF taken up by hepatocytes is transported into the biliary network. The organoids were incubated in the presence of CLF for 30 min. After five washes, images were taken at 30 min (panels 1 and 2) and at 6 hours (panels 3 and 4). Broken lines indicate the boundary between HEP and BD. The experiment using one well was repeated three times, independently. Three fields were examined in each well and the representative images are shown in this figure. Bars represent 100 μm. c Transport of bilirubin. Bilirubin taken up by hepatocytes is excreted into BCs and then transported into the biliary network. Bilirubin in HBTOs was visualized by turning it into biliverdin. The experiment on one well was repeated three times, independently. Three fields were examined in each well and the representative images are shown in this figure. The box in the left panel is enlarged in the right panel. Bars represent 200 μm.