The metabolic activation of the promutagens 2-amino-3,8-dimethylimidazo[4,5-f]quinoline (IQ), 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx) and 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) by rat and mouse lung microsomes was studied using Salmonella mutagenicity (strain TA98). Lungs from uninduced animals were found to activate all three compounds. A 4-6 fold higher mutagenic activity was obtained with IQ compared to MeIQx and the mutagenic response of PhIP was 1-2 orders of magnitude lower than that of IQ. In order to characterize the forms of P450 in the lung responsible for the metabolic activation of these food mutagens Western blots were performed with microsomes and partially purified P450 fractions from the lung. Western blots revealed the presence of cytochrome P450 2A, 2B and 4A forms in untreated rats. In the lung CYP 1A1 was only detectable after BNF treatment of rats. The CYP 4A isozymes, which have not previously been described in the rat lung, were further identified after PCR amplification from lung mRNA as 4A2 and 4A8. Antibody inhibition studies showed that CYP 2A3 catalyzed a major part (70%) of the metabolic activation of IQ by uninduced rat lung microsomes. The metabolic activation of MeIQx was not influenced by this antibody. An antibody against CYP 2B isozymes also partially inhibited the activation of IQ by uninduced rat lung microsomes. However, since induction of CYP 2B isozymes in the liver by phenobarbital treatment did not increase the metabolic activation of the heterocyclic amines over controls it is unlikely that the rat lung CYP 2B1 is participating in the activation of heterocyclic amines. The inhibition of the IQ-dependent mutagenicity by the CYP 2B antibody is probably due to cross-reaction with CYP 2A3. Alfa-napthoflavone (ANF), considered to be a specific inhibitor of CYP 1A isozymes at 10 μM, partly inhibited the activation of IQ (30-40%) and MeIQx (60-80%) by uninduced rat and mouse lung microsomes. Upon pretreatment of rats with BNF, lung microsomes activated MeIQx at a rate that was 2-10-fold higher than control lung microsomes, whereas the increase in EROD activity was approximately 100-fold in the same lung preparations. These results suggest that CYP 1A1 may not be the enzyme responsible for the activation of MeIQx in the control rat despite the inhibition with ANF. It is likely that ANF can inhibit other P450 enzymes in the 'ung, including CYP 2A3. The involvement of CYP 2A3 in the metabolic activation of IQ by uninduced rat lung shows that CYP forms that are not of major importance in the liver may play a significant role in extra-hepatic activation of heterocyclic amines.
ASJC Scopus subject areas
- Cancer Research