title: Integrative "-Omics" Analysis in Primary Human Hepatocytes Unravels Persistent Mechanisms of Cyclosporine A-Induced Cholestasis.
abstract: Cyclosporine A (CsA) is an undecapeptide with strong immunosuppressant activities and is used a lot after organ transplantation. Furthermore, it may induce cholestasis in the liver. In general, the drug-induced cholestasis (DIC) pathway includes genes involved in the uptake, synthesis, conjugation, and secretion of bile acids. However, whether CsA-induced changes in the cholestasis pathway in vitro are persistent for repeated dose toxicity has not yet been investigated. To explore this, primary human hepatocytes (PHH) were exposed to a subcytotoxic dose of 30 μM CsA daily for 3 and 5 days. To investigate the persistence of induced changes upon terminating CsA exposure after 5 days, a subset of PHH was subjected to a washout period (WO-period) of 3 days. Multiple -omics analyses, comprising whole genome analysis of DNA methylation, gene expression, and microRNA expression, were performed. The CsA-treatment resulted after 3 and 5 days, respectively, in 476 and 20 differentially methylated genes (DMGs), 1353 and 1481 differentially expressed genes (DEGs), and in 22 and 29 differentially expressed microRNAs (DE-miRs). Cholestasis-related pathways appeared induced during CsA-treatment. Interestingly, 828 persistent DEGs and 6 persistent DE-miRs but no persistent DMGs were found after the WO-period. These persistent DEGs and DE-miRs showed concordance for 22 genes. Furthermore, 29 persistent DEGs changed into the same direction as observed in livers from cholestasis patients. None of those 29 DEGs which among others relate to oxidative stress and lipid metabolism are yet present in the DIC pathway or cholestasis adverse outcome pathway (AOP) thus presenting novel findings. In summary, we have demonstrated for the first time a persistent impact of repeated dose administration of CsA on genes and microRNAs related to DIC in the gold standard human liver in vitro model with PHH.
authors.affiliation: Department of Toxicogenomics, GROW School for Oncology and Developmental Biology, Maastricht University , P.O. Box 616, 6200 MD, Maastricht, The Netherlands.
authors.forename: Jarno E J
authors.identifier: 0000-0002-4949-5230
authors.initials: JE
authors.lastname: Wolters
authors.affiliation: Department of Toxicogenomics, GROW School for Oncology and Developmental Biology, Maastricht University , P.O. Box 616, 6200 MD, Maastricht, The Netherlands.
authors.forename: Marcel H M
authors.initials: MH
authors.lastname: van Herwijnen
authors.affiliation: Department of Toxicogenomics, GROW School for Oncology and Developmental Biology, Maastricht University , P.O. Box 616, 6200 MD, Maastricht, The Netherlands.
authors.forename: Daniel H J
authors.initials: DH
authors.lastname: Theunissen
authors.affiliation: Department of Toxicogenomics, GROW School for Oncology and Developmental Biology, Maastricht University , P.O. Box 616, 6200 MD, Maastricht, The Netherlands.
authors.forename: Danyel G J
authors.initials: DG
authors.lastname: Jennen
authors.affiliation: Department of Toxicogenomics, GROW School for Oncology and Developmental Biology, Maastricht University , P.O. Box 616, 6200 MD, Maastricht, The Netherlands.
authors.forename: Wim F P M
authors.initials: WF
authors.lastname: Van den Hof
authors.affiliation: Department of Toxicogenomics, GROW School for Oncology and Developmental Biology, Maastricht University , P.O. Box 616, 6200 MD, Maastricht, The Netherlands.
authors.forename: Theo M C M
authors.initials: TM
authors.lastname: de Kok
authors.affiliation: Department of Surgery, Maastricht University , 6200 MD, Maastricht, The Netherlands.
authors.forename: Frank G
authors.initials: FG
authors.lastname: Schaap
authors.affiliation: Department of Toxicogenomics, GROW School for Oncology and Developmental Biology, Maastricht University , P.O. Box 616, 6200 MD, Maastricht, The Netherlands.
authors.forename: Simone G J
authors.initials: SG
authors.lastname: van Breda
authors.affiliation: Department of Toxicogenomics, GROW School for Oncology and Developmental Biology, Maastricht University , P.O. Box 616, 6200 MD, Maastricht, The Netherlands.
authors.forename: Jos C S
authors.initials: JC
authors.lastname: Kleinjans
mesh_terms: D002478:Cells, Cultured; D002779:Cholestasis / Q000139:chemically induced*; D016572:Cyclosporine / Q000009:adverse effects*; D019175:DNA Methylation; D023281:Genomics; D022781:Hepatocytes / Q000378:metabolism*; D006801:Humans; D007166:Immunosuppressive Agents / Q000009:adverse effects*; D020411:Oligonucleotide Array Sequence Analysis; D059467:Transcriptome
chemical_list: D007166:Immunosuppressive Agents; D016572:Cyclosporine