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Received for publication November 22, 2004.
Revised March 14, 2005.
Accepted for publication March 29, 2005.
Fresh human hepatocytes are still considered as the 'gold standard' to screen in vitro for cytochrome P450 (CYP) induction. However, sparse availability of good quality human liver tissue for research purposes, the demand for standardized cell populations together with the need for proper storage of the cells not immediately required, have resulted in the development of cryopreservation techniques that provide adequate viability and plateability of hepatocytes after thawing. This study aimed at validating cryopreserved human hepatocytes as a model to investigate CYP induction. Cryopreserved cells from four different donors were plated and cultured for 48 h followed by incubation in the presence of typical CYP inducers. During the experiments, quality of the cultured cells was monitored both physiologically and morphologically. Concomitantly, the activity of CYPs 1A2, 2B6, 2C9, 2E1 and 3A4 was measured together with their mRNA and protein expression. Determination of CYP1A2, 2B6, 2C9, 2E1 and 3A4 activity in control versus prototypical inducer-treated hepatocytes revealed a maximal significant mean 11.6-, 2.8-, 1.9-, 1.5-, and 9.0-fold induction over their basal expression, respectively. Protein expression analysis of these CYPs confirmed these results. Moreover, a mean 44.9-, 3.5-, 3.2-, and 13.8-fold induction of CYP1A2, 2B6, 2C9, and 3A4 mRNA was observed. Our data demonstrate that cryopreserved human hepatocytes are a valuable tool to study the induction of CYP1A2, 2B6, 2C9, 2E1 and 3A4
Key words:
CYP expression, CYP induction, cytochrome P450, drug-drug interactions, hepatocytes, human CYP enzymes, induction
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