Bioactivation of aflatoxin B1 by lipoxygenases, prostaglandin H synthase and cytochrome P450 monooxygenase in guinea-pig tissues.
In the present investigation, we have examined the role of lipoxygenases in the bioactivation of aflatoxin B1 (AFB1) in hepatic and extrahepatic tissues. The enzyme activities were evaluated by determining [3H]AFB1-DNA adduct formation. The results demonstrated that both purified soybean lipoxygenase and guinea-pig tissue cytosolic lipoxygenases were able to activate AFB1 to form [3H]AFB1-DNA adduct(s). The reaction was completely inhibited by nordihydroguaiaretic acid (NDGA, 0.1 mM), a lipoxygenase inhibitor and an antioxidant, but not by indomethacin (0.1 mM), an inhibitor of prostaglandin H synthase (PHS), indicating that this reaction is associated with lipoxygenase activity, and/or is involved in a peroxyl radical process. While purified lipoxygenase showed arachidonic acid (AA)-dependent properties, the omission of AA did not diminish guinea-pig tissue cytosolic [3H]AFB1-DNA adduct formation, possibly because AA was released from lipid particles by AFB1. Within the range of hemoglobin (Hb) concentrations found in lung, kidney and liver cytosols (1.4-11.1 microM) and microsomes (0-0.5 microM), neither pure Hb, nor Hb of cytosols or microsomes from whole blood caused detectable AA-dependent AFB1-DNA binding. This indicates that Hb, as a contaminant with quasi-lipoxygenase activity, did not contribute to AFB1 activation attributed to guinea-pig tissue lipoxygenases. [3H]AFB1 concentrations at half-maximal DNA binding rate of pulmonary cytochrome P450 monooxygenases (P450) and lipoxygenases were similar, though P450 had a much higher maximum DNA binding rate. Pulmonary microsomal PHS activity for AFB1 activation was too low for its half-maximal binding concentrations of [3H]AFB1 and maximum rate to be accurately determined. In kidney, maximum rates for lipoxygenase, PHS and P450 were similar, whereas half-maximal binding concentrations for reactions by lipoxygenase and P450 were lower compared to that of PHS. The half-maximal binding concentration of hepatic lipoxygenase was significantly lower than those for PHS and P450. Hepatic half-maximal binding concentrations for PHS and P450 were similar, though P450 had a much higher maximum rate than PHS and lipoxygenases. These data suggest that lipoxygenase-catalyzed AFB1 activation can occur at low AFB1 concentrations. This may be important in view of human exposure to low AFB1 concentrations and predominant lipoxygenase activity in human airway epithelial cells. When expressed per gram of tissue, renal and hepatic PHS activities and renal lipoxygenase activities for AFB1 activation were similar, and higher than the activity of pulmonary PHS, while pulmonary PHS activity for the oxidation of N,N,N',N'-tetramethyl-p-phenylenediamine (TMPD) was similar to that in liver and lower than that in kidney.(ABSTRACT TRUNCATED AT 400 WORDS)
Liu
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Department of Pharmacology and Toxicology, Queen's University, Kingston, Ontario, Canada.
Massey
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D016604:Aflatoxin B1 / Q000378:metabolism*; D000818:Animals; D001711:Biotransformation; D003577:Cytochrome P-450 Enzyme System / Q000502:physiology*; D004247:DNA / Q000378:metabolism; D005609:Free Radicals; D006168:Guinea Pigs; D066298:In Vitro Techniques; D008084:Lipoxygenase / Q000502:physiology*; D008099:Liver / Q000378:metabolism; D008168:Lung / Q000378:metabolism; D008297:Male; D010105:Oxygenases / Q000502:physiology*; D011451:Prostaglandin-Endoperoxide Synthases / Q000502:physiology*; D014810:Vitamin E / Q000494:pharmacology
D005609:Free Radicals; D014810:Vitamin E; D004247:DNA; D003577:Cytochrome P-450 Enzyme System; D016604:Aflatoxin B1; D010105:Oxygenases; D008084:Lipoxygenase; D011451:Prostaglandin-Endoperoxide Synthases