Transcriptomic evaluation of canine suspension-shipped and pre-plated hepatocytes: comparison to liver

Abstract

Introduction: In vitro assays using rat and human hepatocytes are used for hepatotoxicity studies; however, in vitro methods are less established for canine hepatocytes. In particular, little is known about the effects of plating and culture on canine hepatocytes. The goal of this study was to conduct a descriptive analysis of an in vitro canine hepatocyte system to evaluate its utility and limitations. The study objectives were to determine if canine hepatocytes shipped in suspension or pre-plated were transcriptomically different from one another and their liver of origin, and to understand temporal transcriptomic changes.

Materials and methods: Frozen canine liver samples were delivered on dry ice; hepatocytes from these livers were delivered in a cell/media suspension (S) or pre-plated (P). Hepatocytes were harvested at arrival and after up to 120 hr of culture in naïve media, or after 48 hr treatment with prototypical enzyme inducing xenobiotics (phenobarbital or rifampin).

Results: A global transcriptomic comparison between liver and hepatocyte preparations indicated that the transcriptome was affected post-plating; transporters and genes involved in xenobiotic metabolism were generally down-regulated. Basal mRNA levels of CYP3A12 and CYP2B11 decreased temporally; after 120 hr CYP3A12 levels decreased by 1000-fold. CYP3A12 and CYP2B11 induction after phenobarbital or rifampin treatment was robust in both cell types but stronger in S cells.

Conclusions: These results indicate that S and P hepatocytes cultured under the current conditions are appropriate for specific in vitro tests. Further characterization of endpoints should be conducted for a thorough understanding of the model’s limitations.

• Cytochrome P450
• gene expression
• methods
• toxicology

Notes

¹Food and Drug Administration Center for Drug Evaluation and Research (2010).