title: Induction of cytochrome P450 enzymes in primary equine hepatocyte culture.
abstract: In this study, we established cell culture conditions for primary equine hepatocytes allowing cytochrome P450 enzyme (CYP) induction experiments. Hepatocytes were isolated after a modified method of Bakala et al. (2003) and cultivated on collagen I coated plates. Three different media were compared for their influence on morphology, viability and CYP activity of the hepatocytes. CYP activity was evaluated with the fluorescent substrate 7-benzyloxy-4-trifluoromethylcoumarin. Induction experiments were carried out with rifampicin, dexamethasone or phenobarbital. Concentration-response curves for induction with rifampicin were created. Williams' medium E showed the best results on morphology and viability of the hepatocytes and was therefore used for the following induction experiments. Cells cultured in Dulbecco's Modified Eagle Medium were not inducible. Incubation with rifampicin increased the CYP activity in two different hepatocyte preparations in a dose dependent manner (EC₅₀=1.20 μM and 6.06 μM; Emax=4.1- and 3.4-fold induction). No increase in CYP activity was detected after incubation with dexamethasone or phenobarbital. The hepatocyte culture conditions established in this study proved to be valuable for investigation of the induction of equine CYPs. In further studies, other equine drugs can be evaluated for CYP induction with this in vitro system.
authors.affiliation: Vetsuisse Faculty University of Bern, Division of Veterinary Pharmacology and Toxicology, Laenggassstrasse 124, CH-3012 Bern, Switzerland. Electronic address: anja.stefanski@vetsuisse.unibe.ch.
authors.forename: A
authors.initials: A
authors.lastname: Stefanski
authors.forename: M
authors.initials: M
authors.lastname: Mevissen
authors.forename: A-M
authors.initials: AM
authors.lastname: Möller
authors.forename: K
authors.initials: K
authors.lastname: Kuehni-Boghenbor
authors.forename: A
authors.initials: A
authors.lastname: Schmitz
mesh_terms: D000818:Animals; D000893:Anti-Inflammatory Agents / Q000009:adverse effects / Q000494:pharmacology; D000904:Antibiotics, Antitubercular / Q000009:adverse effects / Q000494:pharmacology; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D003374:Coumarins / Q000378:metabolism; D016895:Culture Media, Serum-Free / Q000378:metabolism; D003577:Cytochrome P-450 Enzyme System / Q000096:biosynthesis / Q000187:drug effects* / Q000378:metabolism; D003907:Dexamethasone / Q000009:adverse effects / Q000494:pharmacology; D004353:Drug Evaluation, Preclinical / Q000662:veterinary*; D004790:Enzyme Induction / Q000187:drug effects; D022781:Hepatocytes / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D006736:Horses; D006993:Hypnotics and Sedatives / Q000009:adverse effects / Q000494:pharmacology; D007150:Immunohistochemistry / Q000662:veterinary; D007202:Indicators and Reagents / Q000378:metabolism; D007700:Kinetics; D010634:Phenobarbital / Q000009:adverse effects / Q000494:pharmacology; D012293:Rifampin / Q000009:adverse effects / Q000494:pharmacology; D019155:Veterinary Drugs / Q000009:adverse effects / Q000494:pharmacology*
chemical_list: C419147:7-benzyloxy-4-trifluoromethylcoumarin; D000893:Anti-Inflammatory Agents; D000904:Antibiotics, Antitubercular; D003374:Coumarins; D016895:Culture Media, Serum-Free; D006993:Hypnotics and Sedatives; D007202:Indicators and Reagents; D019155:Veterinary Drugs; D003907:Dexamethasone; D003577:Cytochrome P-450 Enzyme System; D012293:Rifampin; D010634:Phenobarbital
keywords: 7-benzyloxy-4-trifluoromethylcoumarin; 7-ethoxy-4-trifluoromethylcoumarin; 7-hydroxy-4-trifluoromethylcoumarin; BFC; CYPs; Cytochrome P450; DMEM; DMSO; Dulbecco’s Modified Eagle Medium; EFC; EGTA; FBS; HBSS; HFC; HTK; Hanks’ balanced salt solution; Hepatocyte; Horse; Induction; Rifampicin; WEM; Williams’ medium E; cytochrome P450 enzymes; dimethyl-sulphoxide; ethylene glycol tetraacetic acid; fetal bovine serum; histidine-tryptophan-ketoglutarate