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Reconstructed human intestinal comet assay, a possible alternative in vitro model for genotoxicity assessment

The aim of the present study was to evaluate the compatibility of reconstructed 3D human small intestinal microtissues to perform the in vitro comet assay. The comet assay is a common follow-up genotoxicity test to confirm or supplement other genotoxicity data. Technically, it can be performed utili...

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Autores principales: Hughes, Christopher Owen, Lim, Hui Kheng, Tan, Joseph Choon Wee, Leavesley, David Ian, Smith, Benjamin Paul Chapman
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Oxford University Press 2023
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10281391/
https://www.ncbi.nlm.nih.gov/pubmed/37115513
http://dx.doi.org/10.1093/mutage/gead011
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author Hughes, Christopher Owen
Lim, Hui Kheng
Tan, Joseph Choon Wee
Leavesley, David Ian
Smith, Benjamin Paul Chapman
author_facet Hughes, Christopher Owen
Lim, Hui Kheng
Tan, Joseph Choon Wee
Leavesley, David Ian
Smith, Benjamin Paul Chapman
author_sort Hughes, Christopher Owen
collection PubMed
description The aim of the present study was to evaluate the compatibility of reconstructed 3D human small intestinal microtissues to perform the in vitro comet assay. The comet assay is a common follow-up genotoxicity test to confirm or supplement other genotoxicity data. Technically, it can be performed utilizing a range of in vitro and in vivo assay systems. Here, we have developed a new reconstructed human intestinal comet (RICom) assay protocol for the assessment of orally ingested materials. The human intestine is a major site of food digestion and adsorption, first-pass metabolism as well as an early site of toxicant first contact and thus is a key site for evaluation. Reconstructed intestinal tissues were dosed with eight test chemicals: ethyl methanesulfonate (EMS), ethyl nitrosourea (ENU), phenformin hydrochloride (Phen HCl), benzo[a]pyrene (BaP), 1,2-dimethylhydrazine hydrochloride (DMH), potassium bromate (KBr), glycidamide (GA), and etoposide (Etop) over a span of 48 h. The RICom assay correctly identified the genotoxicity of EMS, ENU, KBr, and GA. Phen HCl, a known non-genotoxin, did not induce DNA damage in the 3D reconstructed intestinal tissues whilst showing high cytotoxicity as assessed by the assay. The 3D reconstructed intestinal tissues possess sufficient metabolic competency for the successful detection of genotoxicity elicited by BaP, without the use of an exogenous metabolic system. In contrast, DMH, a chemical that requires liver metabolism to exert genotoxicity, did not induce detectable DNA damage in the 3D reconstructed intestinal tissue system. The genotoxicity of Etop, which is dependent on cellular proliferation, was also undetectable. These results suggest the RICom assay protocol is a promising tool for further investigation and safety assessment of novel ingested materials. We recommend that further work will broaden the scope of the 3D reconstructed intestinal tissue comet assay and facilitate broader analyses of genotoxic compounds having more varied modes of actions.
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spelling pubmed-102813912023-06-21 Reconstructed human intestinal comet assay, a possible alternative in vitro model for genotoxicity assessment Hughes, Christopher Owen Lim, Hui Kheng Tan, Joseph Choon Wee Leavesley, David Ian Smith, Benjamin Paul Chapman Mutagenesis Original Manuscripts The aim of the present study was to evaluate the compatibility of reconstructed 3D human small intestinal microtissues to perform the in vitro comet assay. The comet assay is a common follow-up genotoxicity test to confirm or supplement other genotoxicity data. Technically, it can be performed utilizing a range of in vitro and in vivo assay systems. Here, we have developed a new reconstructed human intestinal comet (RICom) assay protocol for the assessment of orally ingested materials. The human intestine is a major site of food digestion and adsorption, first-pass metabolism as well as an early site of toxicant first contact and thus is a key site for evaluation. Reconstructed intestinal tissues were dosed with eight test chemicals: ethyl methanesulfonate (EMS), ethyl nitrosourea (ENU), phenformin hydrochloride (Phen HCl), benzo[a]pyrene (BaP), 1,2-dimethylhydrazine hydrochloride (DMH), potassium bromate (KBr), glycidamide (GA), and etoposide (Etop) over a span of 48 h. The RICom assay correctly identified the genotoxicity of EMS, ENU, KBr, and GA. Phen HCl, a known non-genotoxin, did not induce DNA damage in the 3D reconstructed intestinal tissues whilst showing high cytotoxicity as assessed by the assay. The 3D reconstructed intestinal tissues possess sufficient metabolic competency for the successful detection of genotoxicity elicited by BaP, without the use of an exogenous metabolic system. In contrast, DMH, a chemical that requires liver metabolism to exert genotoxicity, did not induce detectable DNA damage in the 3D reconstructed intestinal tissue system. The genotoxicity of Etop, which is dependent on cellular proliferation, was also undetectable. These results suggest the RICom assay protocol is a promising tool for further investigation and safety assessment of novel ingested materials. We recommend that further work will broaden the scope of the 3D reconstructed intestinal tissue comet assay and facilitate broader analyses of genotoxic compounds having more varied modes of actions. Oxford University Press 2023-04-28 /pmc/articles/PMC10281391/ /pubmed/37115513 http://dx.doi.org/10.1093/mutage/gead011 Text en © The Author(s) 2023. Published by Oxford University Press on behalf of the UK Environmental Mutagen Society. https://creativecommons.org/licenses/by/4.0/This is an Open Access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited.
spellingShingle Original Manuscripts
Hughes, Christopher Owen
Lim, Hui Kheng
Tan, Joseph Choon Wee
Leavesley, David Ian
Smith, Benjamin Paul Chapman
Reconstructed human intestinal comet assay, a possible alternative in vitro model for genotoxicity assessment
title Reconstructed human intestinal comet assay, a possible alternative in vitro model for genotoxicity assessment
title_full Reconstructed human intestinal comet assay, a possible alternative in vitro model for genotoxicity assessment
title_fullStr Reconstructed human intestinal comet assay, a possible alternative in vitro model for genotoxicity assessment
title_full_unstemmed Reconstructed human intestinal comet assay, a possible alternative in vitro model for genotoxicity assessment
title_short Reconstructed human intestinal comet assay, a possible alternative in vitro model for genotoxicity assessment
title_sort reconstructed human intestinal comet assay, a possible alternative in vitro model for genotoxicity assessment
topic Original Manuscripts
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10281391/
https://www.ncbi.nlm.nih.gov/pubmed/37115513
http://dx.doi.org/10.1093/mutage/gead011
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