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Updated: Sep 5, 2025
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Advanced 3D Liver Models for In vitro Genotoxicity Testing Following Long-Term Nanomaterial Exposure
Published on: June 5, 2020
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Evaluation of an in vitro three-dimensional HepaRG spheroid model for genotoxicity testing using the high-throughput
Ji-Eun Seo1, Xiaobo He2, Levan Muskhelishvili3
1Division of Genetic and Molecular Toxicology, National Center for Toxicological Research, U.S. Food and Drug Administration, Jefferson, AR, USA.
ALTEX
|July 6, 2022
Summary
Three-dimensional (3D) HepaRG spheroids enhance in vitro genotoxicity testing by improving sensitivity and predicting in vivo responses. This optimized 3D culture system offers a reliable approach for hazard identification of genotoxic carcinogens.
Area of Science:
- Toxicology
- Cell Biology
- Biotechnology
Background:
- Three-dimensional (3D) cell cultures offer more physiologically relevant models than traditional 2D cultures for genotoxicity studies.
- HepaRG cells are metabolically competent liver cells suitable for in vitro toxicity assessments.
- Existing 2D models have limitations in replicating complex cell-to-cell interactions and tissue structures.
Purpose of the Study:
- To optimize a 3D HepaRG spheroid culture system for in vitro genotoxicity testing.
- To evaluate the sensitivity and predictive capacity of 3D cultures compared to 2D cultures.
- To assess the utility of 3D HepaRG spheroids for high-throughput genotoxicity screening.
Main Methods:
- Formation of 3D HepaRG spheroids in ultra-low attachment plates.
- Exposure of 3D and 2D cultures to 34 diverse test chemicals.
- Assessment of DNA damage using the CometChip assay.
- Concurrent cytotoxicity evaluation via ATP assay.
Main Results:
- 3D HepaRG spheroids maintained stability for up to 30 days with enhanced liver-specific functions (albumin secretion, CYP450 expression).
- 3D cultures demonstrated higher sensitivity in detecting genotoxicants/carcinogens compared to 2D cultures.
- Benchmark dose values from 3D spheroids showed better agreement with primary human hepatocytes.
Conclusions:
- Optimized 3D HepaRG spheroids are a reliable and sensitive in vitro model for genotoxicity testing.
- The 3D CometChip assay system facilitates high-throughput hazard identification.
- This approach improves the prediction of in vivo genotoxicity and carcinogenicity for risk assessment.
