title: The suitability of rat hepatoma cell line H4IIE for evaluating the potentials of compounds to induce CYP3A23 expression.
abstract: To investigate the suitability of H4IIE cells for detecting cytochrome P450 (CYP) induction in vitro, we compared CYP induction by typical CYP inducers in H4IIE cells and rat primary hepatocytes by examining gene expression and enzyme activity, and by immunocytochemistry. The cells were preincubated with 0.1 μM of dexamethasone (DEX) for 24 h, followed by 48 h of exposure to 10 μM of beta-naphthoflavone (bNF), 100 μM of phenobarbital (PB) and 10 μM of DEX. Cyp1a1, Cyp2b1/2 and Cyp3a23/3a1 (Cyp3a23) expressions in H4IIE cells were up-regulated 280-, 1.5- and 65-fold relative to those in vehicle-treated cells, respectively. The fold inductions of those expressions in rat primary hepatocytes were 80-, 33- and 152-fold, respectively. Comprehensive gene expression analysis using DNA microarrays showed that Cyp3a23, Gsta2, Ugt2b12, Udpgt and Sult2a1 expressions were up-regulated in H4IIE cells exposed to 10 μM of DEX. CYP3A activity was not increased, but some H4IIE cells exposed to DEX were stained strongly with anti-CYP3A antibody. We cloned these cells and obtained cloned H4IIE (cH4IIE) cells with expression level of Cyp3a23 higher than those of vehicle-treated cells. It was confirmed that preincubation with 0.1 μM of DEX increased pregnane X receptor (Pxr) expression level and enhanced the Cyp3a23 induction effects of test compounds significantly. Retrospective examination of in vitro CYP induction assay using cH4IIE cells resulted in 80% correlation with the data from in vivo rat toxicity studies. These results suggested that cH4IIE cells are suitable for evaluating the potentials of a compound to induce CYP3A23 expression.
authors.affiliation: Safety Research Laboratory, Mitsubishi Tanabe Pharma Co., 2-50, Kawagishi, 2-Chome, Toda, Saitama 335-8505, Japan. ujimura.hisako@mk.mt-pharma.co.jp
authors.forename: Hisako
authors.initials: H
authors.lastname: Fujimura
authors.forename: Naoko
authors.initials: N
authors.lastname: Murakami
authors.forename: Satoko
authors.initials: S
authors.lastname: Miwa
authors.forename: Chinami
authors.initials: C
authors.lastname: Aruga
authors.forename: Wataru
authors.initials: W
authors.lastname: Toriumi
mesh_terms: D000818:Animals; D001189:Aryl Hydrocarbon Hydroxylases / Q000096:biosynthesis*; D006528:Carcinoma, Hepatocellular / Q000378:metabolism*; D045744:Cell Line, Tumor / Q000187:drug effects*; D051544:Cytochrome P-450 CYP3A; D020869:Gene Expression Profiling; D022781:Hepatocytes / Q000187:drug effects* / Q000378:metabolism; D007150:Immunohistochemistry; D008113:Liver Neoplasms / Q000378:metabolism*; D008297:Male; D018613:Microscopy, Confocal; D020411:Oligonucleotide Array Sequence Analysis; D051381:Rats; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014116:Toxicology / Q000379:methods*
chemical_list: D001189:Aryl Hydrocarbon Hydroxylases; C465138:Cyp3a23-3a1 protein, rat; D051544:Cytochrome P-450 CYP3A