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. 2018 Oct 25;29(1):8–22. doi: 10.1038/s41422-018-0103-x

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© The Author(s) 2018

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 4 — Efficient hepatic differentiation of HepLPCs in vitro. a Schematic of the hepatic-differentiation protocol. TEM/HMM, mixed by 1:1. b QPCR analyses for the expression ALB and CK19 during hepatic-differentiation of HepLPCs (passage 5) from day 2 to day 14. Error bars represent s.d.; n = 3 donors (one-way ANOVA with Tukey correction for multiple comparisons, n.s., non-significant). c Light microscopy images of HepLPCs versus HepLPCs-Hep. Scale bars, 50 μm. d Flow cytometric analysis showing the proportion of ALB and CK19 positive-cells in HepLPCs or HepLPCs-Hep from donor 4 passage 5. Blue, positive-cells; red, negative controls. (e) Euclidean hierarchical clustering of HepG2, HCC, HepLPCs, HepLPCs-Hep and PHCs using differentially expressed genes ( ≥2-fold changes and P < 0.001) in HepLPCs versus HepLPCs-Hep. f Induction of CYP450 expression in HepLPCs-Hep from three donors (passage 5) in response to stimulation with omeprazole, CITCO, and rifampicin for 72 h. 1A2, CYP1A2; 2B6, CYP2B6; 3A4, CYP3A4. Expression normalized to DMSO-treated controls. Error bars represent s.d.; n = 3 technical replicates. g PAS staining with or without diastase, h DiI-LDL uptake and i CDCFDA staining of HepLPCs-Hep. Scale bars, 100 µm. j Albumin production and k ammonia elimination during 24 h measured in supernatant, PHCs were freshly isolated. Error bars represent s.d.; n = 3 donors (one-way ANOVA with Tukey correction for multiple comparisons, **P < 0.01, ***P < 0.001). l CYP metabolic activities of HepLPCs, HepLPCs-Hep (passage 5) and PHCs (freshly isolated). The metabolic products of Acetaminophen, OH-Bupropion, OH-Diclofenac, OH-Testosterone and OH-Coumarin Glu determined by liquid chromatography-tandem mass spectrometry according to standard curves. Error bars represent s.d.; n = 3 donors (one-way ANOVA with Tukey correction for multiple comparisons, n.s., non-significant, *P < 0.05, **P < 0.01). m Representative immunohistochemical staining of hALB of human chimeric mouse liver tissues, PHCs were freshly isolated. Scale bars, 400 µm. n Quantification of the repopulation efficiency estimated by hALB-positive foci. No significant difference is noticed between PHCs and HepLPCs-Hep (passage 5). Error bars represent s.d.; n.s., non-significant