Characterization of rat or human hepatocytes cultured in microphysiological systems (MPS) to identify hepatotoxicity
Abstract
The liver is the main site for drug and xenobiotics metabolism, including inactivation or bioactivation. In order to improve the predictability of drug safety and efficacy in clinical development, and to facilitate the evaluation of the potential human health effects from exposure to environmental contaminants, there is a critical need to accurately model human organ systems such as the liver in vitro. We are developing a microphysiological system (MPS) based on a new commercial microfluidic platform (Nortis, Inc.) that can utilize primary liver cells from multiple species (e.g., rat and human). Compared to conventional monolayer cell culture, which typically survives for 5-7days or less, primary rat or human hepatocytes in an MPS exhibited higher viability and improved hepatic functions, such as albumin production, expression of hepatocyte marker HNF4α and canaliculi structure, for up to 14days. Additionally, induction of Cytochrome P450 (CYP) 1A and 3A4 in cryopreserved human hepatocytes was observed in the MPS. The acute cytotoxicity of the potent hepatotoxic and hepatocarcinogen, aflatoxin B 1 , was evaluated in human hepatocytes cultured in an MPS, demonstrating the utility of this model for acute hepatotoxicity assessment. These results indicate that MPS-cultured hepatocytes provide a promising approach for evaluating chemical toxicity in vitro.
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Conventional 2D cell culture systems are insufficient for sustained preservation of primary hepatocyte differentiation, polarity, and metabolic function.
“Conventional 2D cell culture systems have been acknowledged as insufficient for the sustained preservation of PHs differentiation, polarity, and metabolic function [24,25].”
From The Hepatocyte Expansion Paradox: A Review of In Vitro Challenges and Advances (2026) · cites this paper as [25]
Hepatocytes cultured in 2D systems swiftly impair liver-specific gene expression, CYP450 activity, and synthetic capability, leading to reduced viability and inadequate predictive accuracy for drug testing.
“Across multiple studies, hepatocytes cultured in 2D systems swiftly impair liver-specific gene expression, CYP450 activity, and synthetic capability, ultimately leading to reduced viability and inadequate predictive accuracy for drug testing [24,25].”
From The Hepatocyte Expansion Paradox: A Review of In Vitro Challenges and Advances (2026) · cites this paper as [25]
Hepatocyte spheroids and organoid-like aggregates are notably robust and reproducible models.
“Within 3D methodologies, hepatocyte spheroids and organoid-like aggregates emerged as notably robust and reproducible models [19,20,25,26].”
From The Hepatocyte Expansion Paradox: A Review of In Vitro Challenges and Advances (2026) · cites this paper as [25]
Self-assembled microtissues facilitate improved cell-cell interactions and have greater physiological relevance in cell polarity than 2D systems.
“These self-assembled microtissues facilitate improved cell–cell interactions and have greater physiological relevance in cell polarity than 2D systems [19,20,25,26].”
From The Hepatocyte Expansion Paradox: A Review of In Vitro Challenges and Advances (2026) · cites this paper as [25]
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Gut-on-a-Chip microenvironment induces human intestinal cells to undergo villus differentiation
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Microfluidic titer plate for stratified 3D cell culture
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Long-Term Culture and Coculture of Primary Rat and Human Hepatocytes
Shulman, M., & Nahmias, Y. (2012) · Methods in Molecular Biology · doi:10.1007/978-1-62703-125-7_17 · Cited by 117 · cited 1× · see citing works
“Primary rat hepatocytes were isolated by two-step collagenase digestion following isolation of the liver from male Sprague Dawley® rats (Charles River Laboratories, Seattle, WA) (Shulman and Nahmias, 2013).”
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Reproducing human and cross-species drug toxicities using a liver-chip
Jang, K.-J., Otieno, M. A., Ronxhi, J., Lim, H.-K., Ewart, L., Kodella, K. R., Petropolis, D. B., Kulkarni, G., Rubins, J. E., Conegliano, D., et al. (2019) · Science Translational Medicine · doi:10.1126/scitranslmed.aax5516 · Cited by 511 · see citing works
Recent advances in microfluidic technologies for cell-to-cell interaction studies
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A high-throughput microphysiological system to quantify key events leading to liver fibrosis
Schmidt, S., & Suter-Dick, L. (2025) · Toxicology · doi:10.1016/j.tox.2025.154248 · Cited by 3 · see citing works
The Hepatocyte Expansion Paradox: A Review of In Vitro Challenges and Advances
Kolahdouzmohammadi, M., Tjandra, N., Wu, K., Nikoumaram, R., & Oldani, G. (2026) · Cells · doi:10.3390/cells15161465
“…decade that utilize pharmacological changes to develop viable, expandable hepatocytes. https://doi.org/10.3390/cells15161465 Conventional 2D cell culture systems have been acknowledged as insufficient for the sustained preservation of PHs differentiation, polarity, and metabolic function [24,25].”
Influential
- Authors
- Chang, S.-Y., Voellinger, J. L., Van Ness, K. P., Chapron, B., Shaffer, R. M., Neumann, T., White, C. C., Kavanagh, T. J., Kelly, E. J., & Eaton, D. L.
- Affiliations
- University of Washington (US)
- Nortis (United States) (US)
- Corresponding author
- Eaton, David L.
- Venue
- Toxicology in Vitro
- Publisher
- Elsevier BV
- Published
- 2017-04
- Volume
- 40
- Pages
- 170-183
- ISSN
- 0887-2333 (ISSN-L)
- OpenAlex
- W2572957358
- Cited by
- 49
- References
- 6
- Citations by year
- 2026: 6 · 2025: 4 · 2024: 5 · 2023: 7 · 2022: 3 · 2021: 9 · 2020: 4 · 2019: 4 · 2018: 4 · 2017: 3
- FWCI
- 4.36
- Type
- journal article
- Language
- en
- Open access
- closed
- Locations
- Toxicology in Vitro (publishedVersion)
- PubMed (publishedVersion)
- Funders
- University of Washington (NIH/NIEHS P30ES007033)
- NIH (N01-DK-7-0004 / HHSN267200700004C)
- National Center for Advancing Translational Sciences (UH3 5UH3TR000504-04)
- U.S. Environmental Protection Agency (83573801)
- Topics
- Life Sciences › Pharmacology, Toxicology and Pharmaceutics › Pharmacology › Drug-Induced Hepatotoxicity and Protection
- Physical Sciences › Engineering › Biomedical Engineering › 3D Printing in Biomedical Research
- Health Sciences › Veterinary › Small Animals › Animal testing and alternatives
- Keywords
- microphysiological systems, hepatocytes, hepatotoxicity assessment, primary human hepatocytes, rat hepatocytes, hepatocyte viability, albumin synthesis, HNF4α, CYP1A induction, CYP3A4 induction, aflatoxin B1, acute cytotoxicity, hepatocarcinogens, xenobiotic metabolism
- MeSH terms
- Adult, Animals, Cell Survival, Cells, Cultured, Child, Drug Evaluation, Preclinical, Female, Humans, Male, Middle Aged, Aflatoxin B1, Toxicity Tests, Cell Culture Techniques, Cytochrome P-450 CYP1A1, Cytochrome P-450 CYP1A2, Hepatocytes, Rats, Cytochrome P-450 CYP3A, Hepatocyte Nuclear Factor 4, Young Adult, Chemical and Drug Induced Liver Injury
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