Hepatocyte Differentiation of HNF4α Genome-Edited Pluripotent Stem Cells
(A) Morphology and immunostaining of HNF4α and AFP in the differentiated cells at hepatic progenitor stage. Scale bar, 100 μm for phase contrast and 50 μm for immunostaining images.
(B) Western bloting for HNF4α in differentiated cells at days 9 (hepatic progenitor stage) and 16 (hepatocyte like cell). ACTIN was used as a loading control. See Table S7 for further details.
(C) Real-time PCR quantification of HNF4α, HNF1α, and transthyretin (TTR) mRNA levels in hepatic progenitor stage (day 9) cells. Data were normalized to the housekeeping gene ACTB and expressed relative to the WT cells. The results shown represent three biological replicates, and error bars represent SD. *p < 0.05, ***p < 0.001, ****p < 0.0001; one-way ANOVA with Tukey post hoc test. See Table S8 for further details.
(D) Morphology of differentiated cells at hepatocyte-like cell stage (differentiation day 18). Scale bar, 100 μm.
(E) ELISA quantification of AFP and albumin secretion in differentiated cells at hepatocyte-like cell stage and quantification of cytochrome P450 3A (CYP3A) activity. Data represented three biological replicates, and error bars represented SD. ***p < 0.001, ****p < 0.0001; one-way ANOVA with Tukey post hoc test.