title: Mechanisms of arsenite-mediated decreases in benzo[k]fluoranthene-induced human cytochrome P4501A1 levels in HepG2 cells.
abstract: Polycyclic aromatic hydrocarbons (PAHs) and heavy metals are often environmental cocontaminants that could interact to alter PAH carcinogenicity. The heavy metal, arsenite, and the PAH, benzo[k]fluoranthene, were used as prototypes to investigate, in human HepG2 cells, mechanisms whereby the bioactivation of benzo[k]fluoranthene by human CYP1A1 could be diminished by arsenite-mediated decreases in CYP1A1 induction by benzo[k]fluoranthene. To determine whether arsenite down-regulates CYP1A1 transcription, quantitative real-time reverse transcriptase-polymerase chain reaction assays and luciferase reporter gene expression assays were used with HepG2 cells treated with benzo[k]fluoranthene and arsenite, separately and as a mixture. Benzo[k]fluoranthene (0.5 microM) and arsenite (5 microM) markedly decreased benzo[k]fluoranthene-mediated induction of CYP1A1 mRNA by 45%. Plasmids containing the CYP1A1 promoter region (pHu-1A1-FL) were induced 7.4-fold over vehicle by benzo[k]fluoranthene (0.5 microM), whereas arsenite (1, 2.5, or 5 microM) decreased reporter gene expression by 46%, 45%, and 61%, respectively. The plasmid, pHu-1A1-Delta100-FL, lacked xenobiotic response element (XRE) sites at -1061 and -981 and showed greater responsiveness relative to pHu-1A1-FL, by 1.7-fold. Benzo[k]fluoranthene (0.5 microM) and arsenite (1, 2.5, or 5 microM) decreased reporter gene expression by 0%, 27%, and 39%, respectively, relative to expression levels produced by benzo[k]fluoranthene alone. Arsenite is stable for at least 48 h in the HepG2 cell medium with respect to its ability to diminish CYP1A1 benzo[k]fluoranthene induction. Arsenite did not affect benzo[k]fluoranthene induction directly through XRE sites, nor did it affect the stability of CYP1A1 mRNA. Thus, arsenite affects the transcriptional regulation of the benzo[k]fluoranthene-mediated induction of CYP1A1 and could diminish PAH carcinogenicity by decreasing bioactivation by CYP1A1.
authors.affiliation: New York State Department of Health, Wadsworth Center, PO Box 509, Albany, NY 12201-0509, USA.
authors.forename: Erin E
authors.initials: EE
authors.lastname: Bessette
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Fasco
authors.forename: Brian T
authors.initials: BT
authors.lastname: Pentecost
authors.forename: Laurence S
authors.initials: LS
authors.lastname: Kaminsky
mesh_terms: D018053:Arsenites / Q000494:pharmacology*; D002274:Carcinogens, Environmental / Q000633:toxicity; D045744:Cell Line, Tumor; D019363:Cytochrome P-450 CYP1A1 / Q000037:antagonists & inhibitors / Q000096:biosynthesis* / Q000235:genetics; D004785:Environmental Pollutants / Q000037:antagonists & inhibitors* / Q000633:toxicity; D004790:Enzyme Induction; D005449:Fluorenes / Q000037:antagonists & inhibitors*; D005786:Gene Expression Regulation / Q000187:drug effects*; D017930:Genes, Reporter; D006801:Humans; D008156:Luciferases / Q000235:genetics; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction
chemical_list: D018053:Arsenites; D002274:Carcinogens, Environmental; D004785:Environmental Pollutants; D005449:Fluorenes; D012333:RNA, Messenger; D008156:Luciferases; D019363:Cytochrome P-450 CYP1A1; C015001:arsenite; C022921:benzo(k)fluoranthene