Cytochrome P450 expression in human hepatocytes and hepatoma cell lines: molecular mechanisms that determine lower expression in cultured cells
Abstract
1. Cultured hepatic cells have reduced cytochrome P450 (CYP) activities in comparison with human liver, but the mechanism(s) that underlies this circumstance is not clear. We investigated the causes of this low CYP activity by analysing the activity, protein, mRNA and heterologous nuclear RNA contents of the most important CYPs involved in drug metabolism (1A1, 1A2, 2A6, 2B6, 2C9, 2C19, 2D6, 2E1, 3A4, 3A5) in cultured human hepatocytes, and in HepG2 and Mz-Hep-1 hepatoma cell lines. 2. After 24 h of culture, hepatocytes retained most of their CYP activities and protein contents, but the mRNA decreased 20-fold. However, the mRNA content of most CYPs in 24-h hepatocytes was still 400-fold higher than in hepatoma cells. When we examined the transcriptional activity of the CYP genes, this decreased during culture time in hepatocytes and it was poor in hepatoma cell lines. 3. We investigated the abundance of key hepatic transcription factors that govern CYP transcription (C/EBP-beta: LAP and LIP, HNF-3alpha, HNF-4alpha, RXR-alpha) and observed that the expression of some factors was altered in the hepatoma cells. 4. In conclusion, the loss of biotransformation activity in cultured hepatic cells is caused by a decrease in CYP transcription, which correlates with an alteration in the expression of key transcription factors.
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Few of the models contain the full complement and functionality of metabolic enzymes and transporters present in human hepatocytes in vivo, which is also true with plated primary human hepatocytes (PHH) that rapidly loose liver phenotype and CYP450 activity in traditional monolayer cultures.
“However, few of the models contain the full complement and functionality of metabolic enzymes and transporters present in human hepatocytes in vivo (Gustafsson et al. 2014; Wilkening and Bader 2003), which is also true with plated primary human hepatocytes (PHH) that rapidly loose liver phenotype and CYP450 activity in traditional monolayer cultures (Rodriguez-Antona et al. 2002).”
From Utility of spherical human liver microtissues for prediction of clinical drug-induced liver injury (2017) · cites this paper as Rodriguez-Antona et al. 2002
Hepatocytes maintained in conventional static cultures rapidly reduce transcription of relevant liver-specific genes.
“Because hepatocytes maintained in conventional static cultures rapidly reduce transcription of relevant liver-specific genes28”
From Performance assessment and economic analysis of a human Liver-Chip for predictive toxicology (2022) · cites this paper as [28]
HepG2 has sharply reduced expression of Phase I CYP450 enzymes compared to primary human hepatocytes, with mRNA levels of key isoforms hundreds of times lower.
“HepG2 is characterized by a sharply reduced expression of Phase I biotransformation enzymes— CYP450 —compared to primary human hepatocytes [102,103,104,105].”
From The HepG2 Cell Line as a Model for Studying Metabolic Dysfunction-Associated Steatotic Liver Disease (2026) · cites this paper as [102]
Reduced CYP450 expression in HepG2 is due to reduced transcriptional activity of CYP450 genes, correlated with altered expression of hepatic transcription factors HNF-4α, C/EBP, and HNF-3.
“This is not due to post-transcriptional events but to reduced transcriptional activity of CYP450 genes, which in turn correlates with an altered expression profile of hepatic transcription factors such as HNF-4α, C/EBP, and HNF-3 [102].”
From The HepG2 Cell Line as a Model for Studying Metabolic Dysfunction-Associated Steatotic Liver Disease (2026) · cites this paper as [102]
Cytochrome P450s have been strongly associated with a loss of expression during culture.
“Cytochrome P450s has been strongly associated with a loss of expression during culture (Rodriguez-Antona et al. 2002)”
From Mechanistic evaluation of primary human hepatocyte culture using global proteomic analysis reveals a selective dedifferentiation profile (2017) · cites this paper as Rodriguez-Antona et al. 2002
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- Authors
- Rodríguez-Antona, C., Donato, M. T., Boobis, A., Edwards, R. J., Watts, P. S., Castell, J. V., & Gómez-Lechón, M.-J.
- Affiliations
- Hospital Universitari i Politècnic La Fe (ES)
- Imperial College London (GB)
- Universitat de València (ES)
- Venue
- Xenobiotica
- Publisher
- Informa UK Limited
- Published
- 2002-01
- Volume
- 32
- Issue
- 6
- Pages
- 505-520
- ISSN
- 0049-8254 (ISSN-L)
- 1366-5928
- OpenAlex
- W2095853592
- Cited by
- 420
- References
- 60
- Citations by year
- 2026: 8 · 2025: 35 · 2024: 7 · 2023: 11 · 2022: 12 · 2021: 11 · 2020: 11 · 2019: 19 · 2018: 24 · 2017: 21 · 2016: 17 · 2015: 21 · 2014: 17 · 2013: 23 · 2012: 28
- FWCI
- 8.14
- Type
- journal article
- Language
- en
- Open access
- closed
- Locations
- Xenobiotica (publishedVersion)
- PubMed (publishedVersion)
- Funders
- Generalitat Valenciana
- Topics
- Life Sciences › Pharmacology, Toxicology and Pharmaceutics › Pharmacology › Pharmacogenetics and Drug Metabolism
- Health Sciences › Medicine › Oncology › Drug Transport and Resistance Mechanisms
- Life Sciences › Pharmacology, Toxicology and Pharmaceutics › Pharmacology › Drug-Induced Hepatotoxicity and Protection
- Keywords
- cytochrome P450, hepatocytes, hepatoma cell lines, HepG2 cells, CYP gene expression, transcriptional regulation, C/EBPβ, HNF4α, RXRα, drug metabolism, cultured hepatocytes
- MeSH terms
- Cytochrome P-450 Enzyme System, Pharmaceutical Preparations, HeLa Cells, Carcinoma, Hepatocellular, Humans, Indicators and Reagents, Introns, Liver Neoplasms, RNA, Messenger, Testosterone, Transcription Factors, Tumor Cells, Cultured, Blotting, Western, Gene Expression Regulation, Enzymologic, Reverse Transcriptase Polymerase Chain Reaction, Hepatocytes
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