Comparing in vitro human liver models to in vivo human liver using RNA-Seq
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IN VITRO SYSTEMS
Comparing in vitro human liver models to in vivo human liver using RNA‑Seq Rajinder Gupta1 · Yannick Schrooders1 · Duncan Hauser1 · Marcel van Herwijnen1 · Wiebke Albrecht2 · Bas ter Braak3 · Tim Brecklinghaus2 · Jose V. Castell4 · Leroy Elenschneider5 · Sylvia Escher5 · Patrick Guye6 · Jan G. Hengstler2 · Ahmed Ghallab2,7 · Tanja Hansen5 · Marcel Leist8 · Richard Maclennan9 · Wolfgang Moritz6 · Laia Tolosa10 · Tine Tricot11 · Catherine Verfaillie11 · Paul Walker9 · Bob van de Water3 · Jos Kleinjans1 · Florian Caiment1 Received: 15 June 2020 / Accepted: 12 October 2020 © The Author(s) 2020
Abstract The liver plays an important role in xenobiotic metabolism and represents a primary target for toxic substances. Many different in vitro cell models have been developed in the past decades. In this study, we used RNA-sequencing (RNA-Seq) to analyze the following human in vitro liver cell models in comparison to human liver tissue: cancer-derived cell lines (HepG2, HepaRG 3D), induced pluripotent stem cell-derived hepatocyte-like cells (iPSC-HLCs), cancerous human liver-derived assays (hPCLiS, human precision cut liver slices), non-cancerous human liver-derived assays (PHH, primary human hepatocytes) and 3D liver microtissues. First, using CellNet, we analyzed whether these liver in vitro cell models were indeed classified as liver, based on their baseline expression profile and gene regulatory networks (GRN). More comprehensive analyses using non-differentially expressed genes (non-DEGs) and differential transcript usage (DTU) were applied to assess the coverage for important liver pathways. Through different analyses, we noticed that 3D liver microtissues exhibited a high similarity with in vivo liver, in terms of CellNet (C/T score: 0.98), non-DEGs (10,363) and pathway coverage (highest for 19 out of 20 liver specific pathways shown) at the beginning of the incubation period (0 h) followed by a decrease during long-term incubation for 168 and 336 h. PHH also showed a high degree of similarity with human liver tissue and allowed stable conditions for a short-term cultivation period of 24 h. Using the same metrics, HepG2 cells illustrated the lowest similarity (C/T: 0.51, non-DEGs: 5623, and pathways coverage: least for 7 out of 20) with human liver tissue. The HepG2 are widely used in hepatotoxicity studies, however, due to their lower similarity, they should be used with caution. HepaRG models, iPSC-HLCs, and hPCLiS ranged clearly behind microtissues and PHH but showed higher similarity to human liver tissue than HepG2 cells. In conclusion, this study offers a resource of RNA-Seq data of several biological replicates of human liver cell models in vitro compared to human liver tissue. Keywords In vivo liver · In vitro liver · RNA-seq · CellNet · Non-DEGs · Non-DEGsDTU− · Pathway coverage
Introduction The liver plays a central role in metabolizing exogenous substances. After oral uptake xenobiotics pass through the digestive tract and enter the liver via the portal vein, where Electronic supplement
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