Reprint of: A three-dimensional in vitro HepG2 cells liver spheroid model for genotoxicity studies.

dc.cclicenceCC-BY-NCen
dc.contributor.authorShah, U.K.
dc.contributor.authorMallia, J.O.
dc.contributor.authorSingh, Neenu
dc.contributor.authorDoak, Shareen H.
dc.contributor.authorJenkins, Gareth J. S.
dc.date.acceptance2018-08-18
dc.date.accessioned2019-05-15T14:40:08Z
dc.date.available2019-05-15T14:40:08Z
dc.date.issued2018-08-18
dc.descriptionThe file attached to this record is the author's final peer reviewed version. The Publisher's final version can be found by following the DOI link.en
dc.description.abstractThe liver's role in metabolism of chemicals makes it an appropriate tissue for toxicity testing. Current testing protocols, such as animal testing and two-dimensional liver cell systems, offer limited resemblance to in vivo liver cell behaviour, in terms of gene expression profiles and metabolic competence; thus, they do not always accurately predict human toxicology. In vitro three-dimensional liver cell models offer an attractive alternative. This study reports on the development of a 3D liver model, using HepG2 cells, by a hanging-drop technique, with a focus on evaluating spheroid growth characteristics and suitability for genotoxicity testing. The cytokinesis-blocked micronucleus assay protocol was adapted to enable micronucleus (MN) detection in the 3D spheroid models. This involved evaluating the difference between hanging vs non-hanging drop positions for dosing of the test agents and comparison of automated Metafer scoring with manual scoring for MN detection in HepG2 spheroids. The initial seeding density, used for all experiments, was 5000 cells/20 μl drop hanging spheroids, harvested on day 4, with >75% cell viability. Albumin secretion (7.8 g/l) and both CYP1A1 and CYP1A2 gene expression were highest in the 3D environment at day 4. Exposure to metabolically activated genotoxicants for 24 h resulted in a 6-fold increase in CYP1A1 enzyme activity (3 μM B[a]P) and a 30-fold increase in CYP1A2 enzyme activity (5 μM PhIP) in 3D hanging spheroids. MN inductions in response to B[a]P or PhIP were 2-fold and 3-fold, respectively, and were greater in 3D hanging spheroids than in 2D format, showing that hanging spheroids are more sensitive to genotoxic agents. HepG2 hanging-drop spheroids are an exciting new alternative system for genotoxicity studies, due to their improved structural and physiological properties, relative to 2D cultures.en
dc.funderNo external funderen
dc.funder.otherSer Cymru- Life Sciences Research Network Walesen
dc.identifier.citationShah, U.K., Mallia, J.O., Singh, N., Chapman, K.E., Doak, S.H., Jenkins, G.J.S. (2018) Reprint of: A three-dimensional in vitro HepG2 cells liver spheroid model for genotoxicity studies. Mutation Research/Genetic Toxicology and Environmental Mutagenesis, 834, pp.35-41.en
dc.identifier.doihttps://doi.org/10.1016/j.mrgentox.2018.06.020
dc.identifier.urihttps://www.dora.dmu.ac.uk/handle/2086/17824
dc.language.isoenen
dc.peerreviewedYesen
dc.publisherElsevieren
dc.researchinstituteLeicester Institute for Pharmaceutical Innovation - From Molecules to Practice (LIPI)en
dc.subjectCYP450 enzymesen
dc.subjectHanging-drops spheroidsen
dc.subjectMN inductionen
dc.subjectgenotoxicantsen
dc.subjectmetabolic activationen
dc.titleReprint of: A three-dimensional in vitro HepG2 cells liver spheroid model for genotoxicity studies.en
dc.typeArticleen

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