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Highly efficient gene transfer into hepatocyte-like HepaRG cells: new means for drug metabolism and toxicity studies

Abstract : HepaRG progenitor cells are capable to differentiate into hepatocyte-like cells expressing a large set of liver specific functions. These cells, however, express little amounts of an important cytochrome P450, the CYP2E1, which limits their use for toxicological studies of drugs metabolized by this pathway. We aimed at establishing efficient protocols of transfection in order to increase CYP2E1 expression in HepaRG cells. Transfection protocols of the GFP reporter gene were evaluated using electroporation and cationic lipids belonging to the lipophosphonates, lipophosphoramidates and lipids derived from glycine betaine. Following optimization of the charge ratios, plasmid DNA amounts and formulations with neutral co-lipids, the lipophosphoramidate compounds KLN47 and BSV10 allowed expression of the GFP in ~50% of adherent progenitor HepaRG cells while electroporation targeted GFP expression in ~85% of both progenitor and differentiated cells in suspension. Then, transient enforced expression of active CYP2E1 was achieved in progenitors and/or differentiated HepaRG cells using the electroporation and the lipophosphoramidate compound BSV10. Importantly, in electroporated cells, CYP2E1 expression level was correlated with a significant increase in CYP2E1-specific enzymatic activity, which opens new perspectives for this CYP-dependent drug metabolism and toxicity studies using HepaRG cells.
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Submitted on : Thursday, January 6, 2011 - 2:55:58 AM
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Véronique Laurent, Aurore Fraix, Tristan Montier, Sandrine Cammas-Marion, Catherine Ribault, et al.. Highly efficient gene transfer into hepatocyte-like HepaRG cells: new means for drug metabolism and toxicity studies. Biotechnology Journal, Wiley-VCH Verlag, 2010, 5 (3), pp.314. ⟨10.1002/biot.200900255⟩. ⟨hal-00552337⟩



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