Abstract
Primary human and rat hepatocyte cultures are well established in vitro systems used in toxicological studies. However, whereas transgenic mouse models provide an opportunity for studying mechanisms of toxicity, mouse primary hepatocyte cultures are less well described. The potential usefulness of a mouse hepatocyte-based in vitro model was assessed in this study by investigating time-dependent competence for xenobiotic metabolism and gene expression profiles. Primary mouse hepatocytes, isolated using two-step collagenase perfusion, were cultured in a collagen sandwich configuration. Gene expression profiles and the activities of various cytochrome P450 (P450) enzymes were determined after 0, 42, and 90 h in culture. Principal component analysis of gene expression profiles shows that replicates per time point are similar. Gene expression levels of most phase I biotransformation enzymes decrease to approximately 69 and 57% of the original levels at 42 and 90 h, respectively, whereas enzyme activities for most of the studied P450s decrease to 59 and 34%. The decrease for phase II gene expression is only to 96 and 92% of the original levels at 42 and 90 h, respectively. Pathway analysis reveals initial effects at the level of proteins, external signaling pathways, and energy production. Later effects are observed for transcription, translation, membranes, and cell cycle-related gene sets. These results indicate that the sandwich-cultured primary mouse hepatocyte system is robust and seems to maintain its metabolic competence better than that of the rat hepatocyte system.
Footnotes
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This work was supported in part by The Netherlands Genomics Initiative; The Netherlands Organisation for Scientific Research; and the carcinoGENOMICS FP6 project sponsored by the European Union [Grant PL037712].
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Article, publication date, and citation information can be found at http://dmd.aspetjournals.org.
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doi:10.1124/dmd.108.025775.
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ABBREVIATIONS: P450, cytochrome P450; DMSO, dimethylsulfoxide; SAM, significance analysis of microarrays; PCA, principal component analysis.
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↵ The online version of this article (available at http://dmd.aspetjournals.org) contains supplemental material.
- Received November 20, 2008.
- Accepted February 26, 2009.
- The American Society for Pharmacology and Experimental Therapeutics
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