Abstract
HepaRG cells possess the unique property to differentiate in vitro and to express various functions of mature hepatocytes, including the major cytochromes P450 (P450s). In the present study, we carefully analyzed mRNA expression and activity of the major P450s and their responsiveness to three prototypical inducers, phenobarbital, rifampicin, and omeprazole, in differentiated HepaRG cell cultures over a 4-week period after low and high seeding. Only minor differences were observed in P450 activities when measured by two cocktails of probe substrates, probably related to the choice and/or concentration of substrates. Similar results were obtained from the two cell seeding conditions. Expression and activities of several P450s were dimethyl sulfoxide-dependent. However, basal P450 expression and activities as well as their responsiveness to the prototypical inducers were well maintained over the 4-week period, and a good correlation was observed between transcript levels and corresponding activities. Thus, CYP1A2, CYP2B6, and CYP3A4 were found to accurately respond to their respective prototypical inducers, i.e., omeprazole, phenobarbital, and rifampicin. Likewise, basal expression of several phase II enzymes, transporters, and nuclear receptors, and response to inducers were also well preserved. More genes were found to be induced in HepaRG cells than in primary human hepatocytes, and no marked variation was noticed between the different passages. Taken together, these data support the conclusion that HepaRG cells represent a promising surrogate to primary human hepatocytes for xenobiotic metabolism and toxicity studies.
Footnotes
This work was supported by the European Community [Contracts LIINTOP-STREP-037499 and Predict-IV-202222 (to A.G.)]; and the Ministerio Ciencia e Inovación/Instituto de Salud Carlos III for a Miguel Server [Contract CP08/00125] (to A.L.).
Article, publication date, and citation information can be found at http://dmd.aspetjournals.org.
doi:10.1124/dmd.109.030197.
↵ The online version of this article (available at http://dmd.aspetjournals.org) contains supplemental material.
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- P450
- cytochrome P450
- LD
- low density
- DMSO
- dimethyl sulfoxide
- HD
- high density
- PB
- phenobarbital
- RIF
- rifampicin
- OME
- omeprazole
- FCS
- fetal calf serum
- CDFDA
- carboxydichlorofluorescein di-acetate
- UAM
- Unidad Mixta Fundación Hospital La Fe-Advancell
- NM
- Novamass
- PCR
- polymerase chain reaction
- FIH
- freshly isolated hepatocytes
- HPLC
- high-performance liquid chromatography
- MRP
- multidrug resistance-associated protein
- BSEP
- bile salt export pump
- MDR
- multidrug resistance protein
- RT-qPCR
- reverse transcriptase-quantitative PCR
- GSTA1/A2
- glutathione transferase A1/A2
- UGT1A1
- UDP-glucuronosyl transferase 1A1
- BCRP
- breast cancer resistance protein
- NTCP
- Na+-dependent taurocholic cotransporting polypeptide
- AhR
- aryl hydrocarbon receptor
- CAR
- constitutive androstane receptor
- PXR
- pregnane X receptor.
- Received September 12, 2009.
- Accepted December 17, 2009.
- Copyright © 2010 by The American Society for Pharmacology and Experimental Therapeutics
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