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Updated: Jul 20, 2026
Generation of a Humanized Mouse Liver Using Human Hepatic Stem Cells
Published on: August 29, 2016
Characterization of humanized liver from chimeric mice using coumarin as a human CYP2A6 and mouse CYP2A5 probe
Kimiko Aoki1, Yasuharu Kashiwagura, Toru Horie
1Faculty of Pharmaceutical Sciences, Showa University, Tokyo, Japan. kaoki@pharm.showa-u.ac.jp
Abstract:
Coumarin 7-hydroxylation (COH), which is catalyzed almost solely by human CYP2A6 and mouse CYP2A5, shows large differences in activity (humans>>mice) and inhibitor specificity between mice and humans. To differentiate human and mouse liver functions of chimeric mice (CM1, CM2 and CM3) prepared with hepatocytes from 3 donors, the microsomal COH activities were measured with and without benzaldehyde and undecanoic gamma-lactone as a specific inhibitor of human CYP2A6 and mice CYP2A5, respectively. The replacement % to human hepatocytes designated as replacement index (RI) was calculated from human specific cytokeratin 8/18 expression in the liver section. The COH activities correlated well with RIs in CM2 (R(2)=0.98) and CM3 (R(2)=0.94), except CM1 whose genotype of donor is CYP2A6*4/*4. However, the COH activities expressed as % of donor activities were not always coincident with RIs, and the inhibition pattern of CM2 and CM3 was human-type after RI exceeded approximately 50%. Subsequently, our attempts to use % of COH activities or inhibition patterns as an accurate functional replacement index were unsuccessful. Since the detection of human CYP2A6 protein in the liver and the steep increase of human albumin (hAlb) levels in the blood were begun from almost RI=50% similarly to the changes of inhibition pattern, RI=50% is the turning point for chimeric mice to have humanized liver function.
Insights
Chimeric mice liver function was assessed using coumarin 7-hydroxylation (COH) activity. A replacement index (RI) of approximately 50% indicates a turning point for humanized liver function in these mice.
Area of Science:
- Pharmacology
- Hepatology
- Biochemistry
Background:
- Coumarin 7-hydroxylation (COH) activity differs significantly between humans and mice, primarily due to CYP2A6 and CYP2A5 enzymes.
- Chimeric mice with human hepatocytes offer a model to study human liver function.
- Assessing the functional engraftment of human hepatocytes in chimeric mice is crucial for drug metabolism studies.
Purpose of the Study:
- To differentiate human and mouse liver functions in chimeric mice using COH activity.
- To establish a reliable index for assessing the degree of human hepatocyte engraftment and functional replacement.
- To identify the threshold for achieving humanized liver function in chimeric mice.
Main Methods:
- Microsomal COH activities were measured in chimeric mice (CM1, CM2, CM3) with and without specific inhibitors for human CYP2A6 and mouse CYP2A5.
- The replacement index (RI) was calculated based on human-specific cytokeratin 8/18 expression.
- Correlation analysis was performed between COH activity, inhibition patterns, and RI.
Main Results:
- COH activities correlated well with RI in CM2 and CM3, but not in CM1 (CYP2A6*4/*4 genotype).
- COH activity as a percentage of donor activity did not consistently align with RI.
- A human-type inhibition pattern emerged when RI exceeded approximately 50%, coinciding with human CYP2A6 protein detection and increased human albumin levels.
Conclusions:
- A replacement index (RI) of approximately 50% serves as a critical turning point for chimeric mice to exhibit humanized liver function.
- Simple COH activity or inhibition patterns alone are insufficient as accurate functional replacement indices.
- This study provides a benchmark for evaluating humanized liver function in chimeric mouse models.
