Metabolic activation of 2-acetylaminofluorene is required for induction of multidrug resistance gene expression in rat liver cells.
P-Glycoprotein the multidrug resistance (mdr) efflux transporter is encoded by class 1 mdr genes (mdr1) in humans and rodent species. In rat liver and in rat hepatocytes in primary culture, expression of mdr1 genes can be induced with the carcinogenic aromatic amine 2-acetylaminofluorene (2-AAF). As a consequence, increased P-glycoprotein levels led to an accelerated efflux of vinblastine from the hepatocytes and to resistance towards vinblastine-mediated cytotoxicity. N-Hydroxylation, an obligatory initial step in the activation of 2-AAF into electrophilic DNA-binding metabolites is catalyzed predominantly by cytochrome P450 (CYP)1A2, an isozyme present in normal rat liver. In rat hepatocytes in primary culture, mdr1 induction with 2-AAF could be inhibited by addition of the CYP1A-inhibitor alpha-naphthoflavone, indicating the requirement for metabolic conversion of 2-AAF to act as an inducer of mdr1 gene expression. Both N-hydroxy-2-AAF and the mutagenic 2-AAF derivative N-acetoxy-2-AAF (AAAF) were more potent than 2-AAF as mdr1 inducers. mdr1 induction also decreased when deacetylation of AAAF, which strongly accelerates its conversion into a mutagen, was inhibited with paraoxon. Furthermore, rat liver epithelial cells stably transfected with mouse CYP1A2 showed inducibility of mdr1 gene expression with 2-AAF, whereas the parental cell line, which is devoid of CYP1A2 activity, did not. These findings indicate that electrophilic metabolites formed during 2-AAF or AAAF metabolism are responsible for mdr1 induction in rat hepatocytes. The increased mdr1 gene expression may reflect an adaptive cellular response to electrophiles which includes enhanced synthesis of P-glycoprotein aimed to protect the cell from further damage.
Institute of Toxicology, University of Tübingen, Germany.
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D
Schrenk
T W
TW
Gant
A
A
Michalke
A
A
Orzechowski
J A
JA
Silverman
N
N
Battula
S S
SS
Thorgeirsson
D015073:2-Acetylaminofluorene / Q000378:metabolism* / Q000494:pharmacology; D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000032:analysis / Q000235:genetics*; D000818:Animals; D001711:Biotransformation; D002478:Cells, Cultured; D004351:Drug Resistance / Q000235:genetics*; D005786:Gene Expression Regulation / Q000187:drug effects*; D008099:Liver / Q000378:metabolism*; D008297:Male; D051381:Rats; D011916:Rats, Inbred F344
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D015073:2-Acetylaminofluorene