Comparative aflatoxin B(1) activation and cytotoxicity in human bronchial cells expressing cytochromes P450 1A2 and 3A4.
Some epidemiological evidence suggests a link between the inhalation of aflatoxin B(1) (AFB(1))-contaminated grain dusts and increased lung cancer risk. However, the mechanisms of AFB(1) activation and action in human lung are not well understood. We compared AFB(1) action in SV40 immortalized human bronchial epithelial cells (BEAS-2B) with two transfected cell lines that stably express human cytochromes P450 (CYPs) 1A2 (B-CMV1A2) and 3A4 (B3A4), the principal CYPs thought to activate this mycotoxin in human liver. All three cell types retained catalytically active glutathione S-transferase, the key phase II enzyme that detoxifies metabolically activated AFB(1). B-CMV1A2 and B3A4 cells expressed methoxyresorufin-O-demethylase (MROD) and nifedipine oxidase activities, respectively, and were 3000- and 70-fold more susceptible, respectively, to the cytotoxic effects of AFB(1) than the control cell line (BEAS-2B). When cultured with a range of low, environmentally relevant AFB(1) concentrations (0.02-1.5 microM), control cells formed barely detectable AFB(1)-DNA adducts, whereas B-CMV1A2 cells formed significantly more adducts than B3A4 cells. In B-CMV1A2 cells, formation of AFB(1)-DNA adducts was inhibited by the CYP 1A2 inhibitor 7,8-benzoflavone, whereas formation of AFB(1)-DNA adducts in B3A4 cells was inhibited by the CYP 3A4 inhibitor 17alpha-ethynylestradiol. Competitive reverse transcription-PCR analysis showed that only the CYP-transfected cell lines expressed CYP mRNA. When adjusted for CYP mRNA expression, B-CMV1A2 cells were more efficient in the formation of cytotoxic and DNA-alkylating species at low AFB(1) concentrations, whereas B3A4 cells were more efficient at high concentrations. Our results affirm the hypothesis that, as in human liver microsomes, CYP 1A2 in human lung cells appears to have a more important role than CYP 3A4 in the bioactivation of low AFB(1) concentrations associated with many human exposures. Therefore, it is possible that under conditions in which appropriate CYPs are expressed in lung, inhalation of AFB(1) may result in increased risk of lung cancer in exposed persons.
Van Vleet
Terry R
TR
Graduate Program in Toxicology, Department of Veterinary Sciences, Utah State University, Logan, Utah 84322-4620, USA.
Macé
Katherine
K
Coulombe
Roger A
RA
D016604:Aflatoxin B1 / Q000493:pharmacokinetics* / Q000633:toxicity*; D001571:Benzoflavones / Q000494:pharmacology; D001711:Biotransformation; D001980:Bronchi / Q000166:cytology / Q000201:enzymology* / Q000378:metabolism; D002460:Cell Line; D002470:Cell Survival / Q000502:physiology; D019388:Cytochrome P-450 CYP1A2 / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D065609:Cytochrome P-450 CYP1A2 Inhibitors; D051544:Cytochrome P-450 CYP3A; D003577:Cytochrome P-450 Enzyme System / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D006899:Mixed Function Oxygenases / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D010088:Oxidoreductases / Q000378:metabolism; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection
D001571:Benzoflavones; D065609:Cytochrome P-450 CYP1A2 Inhibitors; D004791:Enzyme Inhibitors; D012333:RNA, Messenger; C011512:alpha-naphthoflavone; D003577:Cytochrome P-450 Enzyme System; D016604:Aflatoxin B1; D006899:Mixed Function Oxygenases; D010088:Oxidoreductases; C076307:methoxyresorufin-O-demethylase; C104464:CYP3A protein, human; D019388:Cytochrome P-450 CYP1A2; D051544:Cytochrome P-450 CYP3A