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. 2020 Mar 13;11:1393. doi: 10.1038/s41467-020-15058-6

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© The Author(s) 2020

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. 6 — a Relative expression of CYP3A4 in HepG2 differentiated with AAGLY, in combination with rapamycin addition (RAPA), or using HepG2 wherein TSC1, GCN2, or LKB1 were knocked out. N = 3 independent differentiations. Significance was calculated by comparing against HepG2 CTL. Significance was also calculated between Hepg2 AAGLY D30 and HepG2 AAGLY RAPA, and between HepG2 AAGLY D30 and TSC1 KO AAGLY D30 when indicated. Significance was calculated by unpaired two-tailed Student’s t-test. b Relative gene expression analysis for CYP3A4 in HLC and HC3X differentiated with AAGLY until D20, with or without rapamycin (RAPA). Expression of HLCs and HC3X cells was related to differentiations in LDM. N = 3 independent differentiations. Significance was calculated by comparing against the LDM and between LDM AA3GLY and LDM AA3GLY RAPA by two-tailed unpaired Student’s t-test. c Western blotting analysis comparing the phosphorylation of S6 and the expression of ATF4 and CYP3A4 in HepG2 differentiated in control media or AAGLY media, with or without KO of TSC1. Representative blot of two repeats. d LISA analysis identifying the top TFs responsible for the genome wide difference between HepG2 and HLC grown in control or AAGLY media. e BFC metabolization comparing HC6X LDM D40 and HC6X AAGLY D40/50, as well as HC6X AAGLY D40 where doxycycline was removed from d30 onwards (HC6X AAGLY D40-DOX from D30). N = 3 independent differentiations. N = 3 donors for PHHs. Significance was calculated as compared with PHH 0 h by unpaired two-tailed Student’s t-test. Significance was also calculated between HC6X AAGLY D40 and HC6X AAGLY –DOX D30-D40. f PCA plot showing the distribution of HLCs, HC3X, and HC6X differentiated in different media and compared with PHHs. Data in all panels represent mean ± SEM with P-values indicated when significant. Source data are provided as a Source Data file.