{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,1,13]],"date-time":"2026-01-13T21:14:55Z","timestamp":1768338895789,"version":"3.49.0"},"reference-count":56,"publisher":"MDPI AG","issue":"14","license":[{"start":{"date-parts":[[2023,7,12]],"date-time":"2023-07-12T00:00:00Z","timestamp":1689120000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"project ATLANTIDA\u2014Platform","award":["NORTE-01-0145-FEDER-000040"],"award-info":[{"award-number":["NORTE-01-0145-FEDER-000040"]}]},{"name":"project ATLANTIDA\u2014Platform","award":["NORTE 2020"],"award-info":[{"award-number":["NORTE 2020"]}]},{"name":"project ATLANTIDA\u2014Platform","award":["UIDB\/04423\/2020"],"award-info":[{"award-number":["UIDB\/04423\/2020"]}]},{"name":"project ATLANTIDA\u2014Platform","award":["UIDP\/04423\/2020"],"award-info":[{"award-number":["UIDP\/04423\/2020"]}]},{"name":"European Regional Development Fund (ERDF)","award":["NORTE-01-0145-FEDER-000040"],"award-info":[{"award-number":["NORTE-01-0145-FEDER-000040"]}]},{"name":"European Regional Development Fund (ERDF)","award":["NORTE 2020"],"award-info":[{"award-number":["NORTE 2020"]}]},{"name":"European Regional Development Fund (ERDF)","award":["UIDB\/04423\/2020"],"award-info":[{"award-number":["UIDB\/04423\/2020"]}]},{"name":"European Regional Development Fund (ERDF)","award":["UIDP\/04423\/2020"],"award-info":[{"award-number":["UIDP\/04423\/2020"]}]},{"name":"CIIMAR\/CIMAR","award":["NORTE-01-0145-FEDER-000040"],"award-info":[{"award-number":["NORTE-01-0145-FEDER-000040"]}]},{"name":"CIIMAR\/CIMAR","award":["NORTE 2020"],"award-info":[{"award-number":["NORTE 2020"]}]},{"name":"CIIMAR\/CIMAR","award":["UIDB\/04423\/2020"],"award-info":[{"award-number":["UIDB\/04423\/2020"]}]},{"name":"CIIMAR\/CIMAR","award":["UIDP\/04423\/2020"],"award-info":[{"award-number":["UIDP\/04423\/2020"]}]},{"name":"ICBAS\u2014U.Porto","award":["NORTE-01-0145-FEDER-000040"],"award-info":[{"award-number":["NORTE-01-0145-FEDER-000040"]}]},{"name":"ICBAS\u2014U.Porto","award":["NORTE 2020"],"award-info":[{"award-number":["NORTE 2020"]}]},{"name":"ICBAS\u2014U.Porto","award":["UIDB\/04423\/2020"],"award-info":[{"award-number":["UIDB\/04423\/2020"]}]},{"name":"ICBAS\u2014U.Porto","award":["UIDP\/04423\/2020"],"award-info":[{"award-number":["UIDP\/04423\/2020"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Animals"],"abstract":"<jats:p>Mammal hepatocyte spheroids have been investigated as alternative experimental models in several contexts, since three-dimensional (3D) systems have shown the potential to mimic in vivo scenarios. The description of fish hepatocyte 3D models is still minimal. This study intends to further characterize brown trout primary hepatocyte spheroids at distinct time points up to 25 days in culture. Viability, biometry, histomorphology, and basal expression of a selection of genes (metabolism and detoxification, efflux transport, and estrogenic signalling) were considered. The gene expression of whole liver samples from the same fish donor were evaluated concurrently. After 12 days in culture, the hepatocyte spheroids exhibited biometric and morphological stability. From the 12th to the 20th day in culture, the basal expression levels for most of the selected genes did not vary. The targeted mRNA levels were higher in brown trout liver samples compared to hepatocyte spheroids. Despite that, data supported that this model resembles some in vivo features. As an experimental alternative model, it showed potential to be used in a stable time window that can be exploited for exposure tests to different xenobiotics, namely, estrogenic compounds.<\/jats:p>","DOI":"10.3390\/ani13142277","type":"journal-article","created":{"date-parts":[[2023,7,13]],"date-time":"2023-07-13T01:52:25Z","timestamp":1689213145000},"page":"2277","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":4,"title":["A Step Forward in the Characterization of Primary Brown Trout Hepatocytic Spheroids as Experimental Models"],"prefix":"10.3390","volume":"13","author":[{"given":"Rodrigo F.","family":"Alves","sequence":"first","affiliation":[{"name":"Team of Animal Morphology and Toxicology, Interdisciplinary Centre of Marine and Environmental Research (CIIMAR\/CIMAR), University of Porto, Terminal de Cruzeiros do Porto de Leix\u00f5es, Av. General Norton de Matos s\/n, 4450-208 Matosinhos, Portugal"},{"name":"Laboratory of Histology and Embryology, Department of Microscopy, ICBAS\u2014School of Medicine and Biomedical Sciences, University of Porto, Rua Jorge Viterbo Ferreira 228, 4050-313 Porto, Portugal"}]},{"given":"C\u00e9lia","family":"Lopes","sequence":"additional","affiliation":[{"name":"Team of Animal Morphology and Toxicology, Interdisciplinary Centre of Marine and Environmental Research (CIIMAR\/CIMAR), University of Porto, Terminal de Cruzeiros do Porto de Leix\u00f5es, Av. General Norton de Matos s\/n, 4450-208 Matosinhos, Portugal"},{"name":"Laboratory of Histology and Embryology, Department of Microscopy, ICBAS\u2014School of Medicine and Biomedical Sciences, University of Porto, Rua Jorge Viterbo Ferreira 228, 4050-313 Porto, Portugal"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9813-4389","authenticated-orcid":false,"given":"Eduardo","family":"Rocha","sequence":"additional","affiliation":[{"name":"Team of Animal Morphology and Toxicology, Interdisciplinary Centre of Marine and Environmental Research (CIIMAR\/CIMAR), University of Porto, Terminal de Cruzeiros do Porto de Leix\u00f5es, Av. General Norton de Matos s\/n, 4450-208 Matosinhos, Portugal"},{"name":"Laboratory of Histology and Embryology, Department of Microscopy, ICBAS\u2014School of Medicine and Biomedical Sciences, University of Porto, Rua Jorge Viterbo Ferreira 228, 4050-313 Porto, Portugal"}]},{"given":"T\u00e2nia V.","family":"Madureira","sequence":"additional","affiliation":[{"name":"Team of Animal Morphology and Toxicology, Interdisciplinary Centre of Marine and Environmental Research (CIIMAR\/CIMAR), University of Porto, Terminal de Cruzeiros do Porto de Leix\u00f5es, Av. General Norton de Matos s\/n, 4450-208 Matosinhos, Portugal"},{"name":"Laboratory of Histology and Embryology, Department of Microscopy, ICBAS\u2014School of Medicine and Biomedical Sciences, University of Porto, Rua Jorge Viterbo Ferreira 228, 4050-313 Porto, Portugal"}]}],"member":"1968","published-online":{"date-parts":[[2023,7,12]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"25187","DOI":"10.1038\/srep25187","article-title":"Characterization of primary human hepatocyte spheroids as a model system for drug-induced liver injury, liver function and disease","volume":"6","author":"Bell","year":"2016","journal-title":"Sci. Rep."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"422","DOI":"10.1016\/j.xphs.2020.10.043","article-title":"Primary Human Hepatocyte Spheroid Model as a 3D In Vitro Platform for Metabolism Studies","volume":"110","author":"Kanebratt","year":"2020","journal-title":"J. Pharm. Sci."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"2419","DOI":"10.1007\/s10646-012-0965-5","article-title":"Towards a more representative in vitro method for fish ecotoxicology: Morphological and biochemical characterisation of three-dimensional spheroidal hepatocytes","volume":"21","author":"Baron","year":"2012","journal-title":"Ecotoxicology"},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"494","DOI":"10.1039\/C4TX00160E","article-title":"Functional xenobiotic metabolism and efflux transporters in trout hepatocyte spheroid cultures","volume":"4","author":"Uchea","year":"2015","journal-title":"Toxicol. Res."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"1180","DOI":"10.1002\/jcb.21345","article-title":"Functional three-dimensional HepG2 aggregate cultures generated from an ultrasound trap: Comparison with HepG2 spheroids","volume":"102","author":"Liu","year":"2007","journal-title":"J. Cell. Biochem."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"1689","DOI":"10.2174\/1381612824666180404152304","article-title":"Comparative Assay of 2D and 3D Cell Culture Models: Proliferation, Gene Expression and Anticancer Drug Response","volume":"24","author":"Souza","year":"2018","journal-title":"Curr. Pharm. Des."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"106105","DOI":"10.1016\/j.aquatox.2022.106105","article-title":"Transcriptomic and physiological analysis of endocrine disrupting chemicals Impacts on 3D Zebrafish liver cell culture system","volume":"245","author":"Park","year":"2022","journal-title":"Aquat. Toxicol."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"655","DOI":"10.1093\/toxsci\/kfx289","article-title":"Comparison of Hepatic 2D Sandwich Cultures and 3D Spheroids for Long-term Toxicity Applications: A Multicenter Study","volume":"162","author":"Bell","year":"2018","journal-title":"Toxicol. Sci."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"1021","DOI":"10.1007\/s00204-019-02427-4","article-title":"Integrated in vitro models for hepatic safety and metabolism: Evaluation of a human Liver-Chip and liver spheroid","volume":"93","author":"Foster","year":"2019","journal-title":"Arch. Toxicol."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"eabe4724","DOI":"10.1126\/sciadv.abe4724","article-title":"JAK inhibition reduces SARS-CoV-2 liver infectivity and modulates inflammatory responses to reduce morbidity and mortality","volume":"7","author":"Stebbing","year":"2021","journal-title":"Sci. Adv."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"2696","DOI":"10.1096\/fj.201601375R","article-title":"Endogenous and xenobiotic metabolic stability of primary human hepatocytes in long-term 3D spheroid cultures revealed by a combination of targeted and untargeted metabolomics","volume":"31","author":"Vorrink","year":"2017","journal-title":"FASEB J."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"419","DOI":"10.1124\/dmd.116.074369","article-title":"Transcriptional, Functional, and Mechanistic Comparisons of Stem Cell\u2013Derived Hepatocytes, HepaRG Cells, and Three-Dimensional Human Hepatocyte Spheroids as Predictive In Vitro Systems for Drug-Induced Liver Injury","volume":"45","author":"Bell","year":"2017","journal-title":"Drug Metab. Dispos."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"1227","DOI":"10.1002\/hep.24760","article-title":"Human liver cell spheroids in extended perfusion bioreactor culture for repeated-dose drug testing","volume":"55","author":"Leite","year":"2012","journal-title":"Hepatology"},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"035021","DOI":"10.1088\/1758-5090\/ac7825","article-title":"In vitro long term differentiation and functionality of three-dimensional bioprinted primary human hepatocytes: Application for in vivo engraftment","volume":"14","author":"Cuvellier","year":"2022","journal-title":"Biofabrication"},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"914","DOI":"10.1083\/jcb.101.3.914","article-title":"Spheroidal aggregate culture of rat liver cells: Histotypic reorganization, biomatrix deposition, and maintenance of functional activities","volume":"101","author":"Landry","year":"1985","journal-title":"J. Cell Biol."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"673","DOI":"10.1007\/BF02631045","article-title":"A simplified method for the culturing of primary adult rat and human hepatocytes as multicellular spheroids","volume":"28a","author":"Li","year":"1992","journal-title":"Vitr. Cell. Dev. Biol."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"407","DOI":"10.1242\/jcs.105.2.407","article-title":"Monolayer and aggregate cultures of rainbow trout hepatocytes: Long-term and stable liver-specific expression in aggregates","volume":"105","author":"Flouriot","year":"1993","journal-title":"J. Cell Sci."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"250","DOI":"10.1016\/j.ecoenv.2018.10.009","article-title":"Development of three-dimensional (3D) spheroid cultures of the continuous rainbow trout liver cell line RTL-W1","volume":"167","author":"Lammel","year":"2018","journal-title":"Ecotoxicol. Environ. Saf."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"109470","DOI":"10.1016\/j.cbpc.2022.109470","article-title":"Fish hepatocyte spheroids\u2014A powerful (though underexplored) alternative in vitro model to study hepatotoxicity","volume":"262","author":"Alves","year":"2022","journal-title":"Comp. Biochem. Physiol. Part C Toxicol. Pharmacol."},{"key":"ref_20","doi-asserted-by":"crossref","unstructured":"Baron, M.G., Mintram, K.S., Owen, S.F., Hetheridge, M.J., Moody, A.J., Purcell, W.M., Jackson, S.K., and Jha, A.N. (2017). Pharmaceutical Metabolism in Fish: Using a 3-D Hepatic In Vitro Model to Assess Clearance. PLoS ONE, 12.","DOI":"10.1371\/journal.pone.0168837"},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"1738","DOI":"10.1002\/etc.4476","article-title":"Performance of Three-Dimensional Rainbow Trout (Oncorhynchus mykiss) Hepatocyte Spheroids for Evaluating Biotransformation of Pyrene","volume":"38","author":"Hultman","year":"2019","journal-title":"Environ. Toxicol. Chem."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"106331","DOI":"10.1016\/j.aquatox.2022.106331","article-title":"Establishing brown trout primary hepatocyte spheroids as a new alternative experimental model\u2014Testing the effects of 5\u03b1-dihydrotestosterone on lipid pathways","volume":"253","author":"Pereira","year":"2022","journal-title":"Aquat. Toxicol."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"133","DOI":"10.1016\/j.aquatox.2015.10.010","article-title":"Estrogenic and anti-estrogenic influences in cultured brown trout hepatocytes: Focus on the expression of some estrogen and peroxisomal related genes and linked phenotypic anchors","volume":"169","author":"Madureira","year":"2015","journal-title":"Aquat. Toxicol."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1016\/j.cbpc.2018.06.001","article-title":"Sex-steroids and hypolipidemic chemicals impacts on brown trout lipid and peroxisome signaling\u2014Molecular, biochemical and morphological insights","volume":"212","author":"Madureira","year":"2018","journal-title":"Comp. Biochem. Physiol. Part C Toxicol. Pharmacol."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"105586","DOI":"10.1016\/j.aquatox.2020.105586","article-title":"Disruption of classical estrogenic targets in brown trout primary hepatocytes by the model androgens testosterone and dihydrotestosterone","volume":"227","author":"Lopes","year":"2020","journal-title":"Aquat. Toxicol."},{"key":"ref_26","first-page":"221","article-title":"Trout hepatocyte culture: Isolation and primary culture","volume":"21","author":"Klaunig","year":"1985","journal-title":"In Vitr. Cell. Dev. Biol. Plant"},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"237","DOI":"10.1007\/BF02624453","article-title":"Isolated trout liver cells: Establishing short-term primary cultures exhibiting cell-to-cell interactions","volume":"26","author":"Blair","year":"1990","journal-title":"Vitr. Cell. Dev. Biol."},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"43","DOI":"10.1016\/j.cmpb.2015.02.006","article-title":"AnaSP: A software suite for automatic image analysis of multicellular spheroids","volume":"119","author":"Piccinini","year":"2015","journal-title":"Comput. Methods Programs Biomed."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"574","DOI":"10.1002\/jcb.22741","article-title":"Cellular stress induced by resazurin leads to autophagy and cell death via production of reactive oxygen species and mitochondrial impairment","volume":"111","author":"Erikstein","year":"2010","journal-title":"J. Cell. Biochem."},{"key":"ref_30","unstructured":"Koetsier, G., and Cantor, E. (2019). A practical guide to analyzing nucleic acid concentration and purity with microvolume spectrophotometers. N. Engl. Biolabs, 1\u20138. Available online: https:\/\/prd-sccd01-international.neb.com\/-\/media\/nebus\/files\/application-notes\/technote_mvs_analysis_of_nucleic_acid_concentration_and_purity.pdf?rev=c24cea043416420d84fb6bf7b554dbbb."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"e45","DOI":"10.1093\/nar\/29.9.e45","article-title":"A new mathematical model for relative quantification in real-time RT-PCR","volume":"29","author":"Pfaffl","year":"2001","journal-title":"Nucleic Acids Res."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"5245","DOI":"10.1158\/0008-5472.CAN-04-0496","article-title":"Normalization of Real-Time Quantitative Reverse Transcription-PCR Data: A Model-Based Variance Estimation Approach to Identify Genes Suited for Normalization, Applied to Bladder and Colon Cancer Data Sets","volume":"64","author":"Andersen","year":"2004","journal-title":"Cancer Res"},{"key":"ref_33","unstructured":"Batista-Pinto, C. (2007). Peroxisomes in Brown Trout (Salmo trutta f. fario): Regulation by Estrogens, Institute of Biomedical Sciences Abel Salazar."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"188","DOI":"10.1016\/j.aquatox.2008.08.012","article-title":"Water temperature and concomitant waterborne ethinylestradiol exposure affects the vitellogenin expression in juvenile brown trout (Salmo trutta)","volume":"90","author":"Kohno","year":"2008","journal-title":"Aquat. Toxicol."},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"104920","DOI":"10.1016\/j.pestbp.2021.104920","article-title":"The insecticide chlorpyrifos modifies the expression of genes involved in the PXR and AhR pathways in the rainbow trout, Oncorhynchus mykiss","volume":"178","author":"Darraz","year":"2021","journal-title":"Pestic. Biochem. Physiol."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"420","DOI":"10.1152\/physiolgenomics.00123.2014","article-title":"Identification of conserved hepatic transcriptomic responses to 17\u03b2-estradiol using high-throughput sequencing in brown trout","volume":"47","author":"Webster","year":"2015","journal-title":"Physiol. Genom."},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"94","DOI":"10.1016\/j.cbpb.2017.04.005","article-title":"Genome specific PPAR\u03b1B duplicates in salmonids and insights into estrogenic regulation in brown trout","volume":"208\u2013209","author":"Madureira","year":"2017","journal-title":"Comp. Biochem. Physiol. Part B Biochem. Mol. Biol."},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"153","DOI":"10.1016\/j.aquatox.2017.04.001","article-title":"Cross-interference of two model peroxisome proliferators in peroxisomal and estrogenic pathways in brown trout hepatocytes","volume":"187","author":"Madureira","year":"2017","journal-title":"Aquat. Toxicol."},{"key":"ref_39","first-page":"9","article-title":"PAST: Paleontological statistics software package for education and data analysis","volume":"4","author":"Hammer","year":"2001","journal-title":"Palaeontol. Electron."},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"656","DOI":"10.1290\/1071-2690(2001)037<0656:EOMCOT>2.0.CO;2","article-title":"Effects of medium composition on the morphology and function of rat hepatocytes cultured as spheroids and monolayers","volume":"37","author":"Hamilton","year":"2001","journal-title":"Vitr. Cell. Dev. Biol.-Anim."},{"key":"ref_41","doi-asserted-by":"crossref","unstructured":"Langan, M., Dodd, N.J., Owen, S.F., Purcell, W.M., Jackson, S.K., and Jha, A.N. (2016). Direct measurements of oxygen gradients in spheroid culture system using electron parametric resonance oximetry. PLoS ONE, 11.","DOI":"10.1371\/journal.pone.0160795"},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"srep19103","DOI":"10.1038\/srep19103","article-title":"3D tumor spheroid models for in vitro therapeutic screening: A systematic approach to enhance the biological relevance of data obtained","volume":"6","author":"Zanoni","year":"2016","journal-title":"Sci. Rep."},{"key":"ref_43","doi-asserted-by":"crossref","unstructured":"Malh\u00e3o, F., Macedo, A.C., Ramos, A.A., and Rocha, E. (2022). Morphometrical, Morphological, and Immunocytochemical Characterization of a Tool for Cytotoxicity Research: 3D Cultures of Breast Cell Lines Grown in Ultra-Low Attachment Plates. Toxics, 10.","DOI":"10.3390\/toxics10080415"},{"key":"ref_44","doi-asserted-by":"crossref","first-page":"8430","DOI":"10.1016\/j.biomaterials.2012.08.040","article-title":"An oxygen-permeable spheroid culture system for the prevention of central hypoxia and necrosis of spheroids","volume":"33","author":"Anada","year":"2012","journal-title":"Biomaterials"},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"281","DOI":"10.1159\/000272316","article-title":"Comparative Analysis of Gene Expression in Rat Liver Tissue and Monolayer- and Spheroid-Cultured Hepatocytes","volume":"191","author":"Sakai","year":"2009","journal-title":"Cells Tissues Organs"},{"key":"ref_46","first-page":"2623","article-title":"Dependence of liver-specific transcription on tissue organization","volume":"5","author":"Clayton","year":"1985","journal-title":"Mol. Cell. Biol."},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1089\/aivt.2017.0022","article-title":"Transcriptomic, Proteomic, and Functional Long-Term Characterization of Multicellular Three-Dimensional Human Liver Microtissues","volume":"4","author":"Messner","year":"2018","journal-title":"Appl. Vitr. Toxicol."},{"key":"ref_48","doi-asserted-by":"crossref","first-page":"1251","DOI":"10.1007\/s00204-020-02682-w","article-title":"Functionality of primary hepatic non-parenchymal cells in a 3D spheroid model and contribution to acetaminophen hepatotoxicity","volume":"94","author":"Bell","year":"2020","journal-title":"Arch. Toxicol."},{"key":"ref_49","doi-asserted-by":"crossref","first-page":"359","DOI":"10.1124\/dmd.32.3.359","article-title":"Kupffer cell-mediated il-2 suppression of cyp3a activity in human hepatocytes","volume":"32","author":"Sunman","year":"2004","journal-title":"Drug Metab. Dispos."},{"key":"ref_50","doi-asserted-by":"crossref","first-page":"621","DOI":"10.1042\/bj2890621","article-title":"Altered expression of cytochrome P-450 mRNAs, and potentially of other transcripts encoding key hepatic functions, are triggered during the isolation of rat hepatocytes","volume":"289","author":"Padgham","year":"1993","journal-title":"Biochem. J."},{"key":"ref_51","doi-asserted-by":"crossref","first-page":"438","DOI":"10.1016\/j.aquatox.2010.11.010","article-title":"Constitutive mRNA expression and protein activity levels of nine ABC efflux transporters in seven permanent cell lines derived from different tissues of rainbow trout (Oncorhynchus mykiss)","volume":"101","author":"Fischer","year":"2011","journal-title":"Aquat. Toxicol."},{"key":"ref_52","doi-asserted-by":"crossref","first-page":"322","DOI":"10.1016\/j.aquatox.2008.09.012","article-title":"Cloning and molecular characterization of apical efflux transporters (ABCB1, ABCB11 and ABCC2) in rainbow trout (Oncorhynchus mykiss) hepatocytes","volume":"90","author":"Zaja","year":"2008","journal-title":"Aquat. Toxicol."},{"key":"ref_53","doi-asserted-by":"crossref","first-page":"209","DOI":"10.1016\/j.cbpc.2009.10.009","article-title":"Gene expression analysis of the ABC efflux transporters in rainbow trout (Oncorhynchus mykiss)","volume":"151","author":"Zaja","year":"2010","journal-title":"Comp. Biochem. Physiol. Part C Toxicol. Pharmacol."},{"key":"ref_54","doi-asserted-by":"crossref","first-page":"536","DOI":"10.1016\/j.clinre.2012.06.006","article-title":"The bile salt export pump (BSEP) in health and disease","volume":"36","author":"Kubitz","year":"2012","journal-title":"Clin. Res. Hepatol. Gastroenterol."},{"key":"ref_55","doi-asserted-by":"crossref","first-page":"216","DOI":"10.1016\/j.taap.2004.10.012","article-title":"Multidrug resistance proteins: Role of P-glycoprotein, MRP1, MRP2, and BCRP (ABCG2) in tissue defense","volume":"204","author":"Leslie","year":"2005","journal-title":"Toxicol. Appl. Pharmacol."},{"key":"ref_56","doi-asserted-by":"crossref","first-page":"2485","DOI":"10.1021\/acs.chemrestox.1c00227","article-title":"Evaluation of a Three-Dimensional Primary Human Hepatocyte Spheroid Model: Adoption and Industrialization for the Enhanced Detection of Drug-Induced Liver Injury","volume":"34","author":"Schofield","year":"2021","journal-title":"Chem. Res. Toxicol."}],"container-title":["Animals"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2076-2615\/13\/14\/2277\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,10]],"date-time":"2025-10-10T20:11:13Z","timestamp":1760127073000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2076-2615\/13\/14\/2277"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2023,7,12]]},"references-count":56,"journal-issue":{"issue":"14","published-online":{"date-parts":[[2023,7]]}},"alternative-id":["ani13142277"],"URL":"https:\/\/doi.org\/10.3390\/ani13142277","relation":{},"ISSN":["2076-2615"],"issn-type":[{"value":"2076-2615","type":"electronic"}],"subject":[],"published":{"date-parts":[[2023,7,12]]}}}