Figure 4.

Liver function assays of hiHs. (A): hiHs were stained at 37°C for 60 minutes with indocyanine green (ICG) at day 18 after differentiation, and then the cellular uptake of ICG was examined by microscopy after the cells were washed with phosphate-buffered saline (PBS). (B, C): After PBS was replaced by the culture medium and the cells were incubated at 37°C again, the cellular excretion of ICG was examined by microscopy at 1 hour (B) and 3.5 hours (C) after the removal of ICG. (D): hiHs were analyzed by periodic acid-Schiff's staining for glycogen storage at day 20 after differentiation. Scale bars = 100 μm (A–D). (E): The supernatants were collected during the differentiation of human induced pluripotent stem cells, and albumin secretion by hiHs in supernatants was determined by enzyme-linked immunosorbent assay. The total secreted albumin was normalized to the cell numbers at days 26–27. (F): The supernatants were collected at 72 hours after hiHs were treated with the inducers Rifa and Phe at day 22 after differentiation. The increase of CYP3A4 activity was assessed by measurement of luciferase activity with the P450-Glo CYP3A4 assay kit. Abbreviations: ALB, albumin; hiH, human induced pluripotent stem cell-derived hepatocyte; Phe, phenobarbital; Rifa, rifampicin.