concanavalin-a and alpha-naphthoflavone

Here, we attempt to reveal how 2-aminofluorene (AF), benzo(alpha)pyrene (BP) and their major metabolites affect T-cell responses to antigenic and mitogenic stimuli. P-450-related metabolism of these parent compounds to metabolites seems to precede the observed immunosuppression; therefore, we investigated the influence of alpha-naphthoflavone (P-450 inhibitor) and beta-naphthoflavone (P-450 inducer) on BP and AF immunosuppression. We used proliferative responses to concanavalin A and the allogeneic mixed lymphocyte response as correlates of immunosuppression. We also attempted to correlate DNA-adduction to the extent of observed immunosuppression for AF and BP metabolites. These data show that the pathway to the strongest immunosuppressive agents for polycyclic aromatic hydrocarbons and arylamines are divergent and related to metabolism by P450 enzymes.

Topics: Acetyl Coenzyme A; Animals; Antigens; Benzo(a)pyrene; Benzoflavones; beta-Naphthoflavone; Carcinogens; Cell Proliferation; Cells, Cultured; Concanavalin A; Cytochrome P-450 Enzyme Inhibitors; Cytochrome P-450 Enzyme System; DNA Adducts; Dose-Response Relationship, Drug; Enzyme Inhibitors; Fluorenes; Immunosuppressive Agents; Interleukin-2; Lymphocyte Culture Test, Mixed; Mice; Mice, Inbred C3H; Mitogens; Spleen; T-Lymphocytes

The metabolism of the polycyclic aromatic hydrocarbon (PAH) 7,12-dimethylbenz[a]anthracene (DMBA) was studied in murine lymphocytes. This carcinogen has previously been shown to be immunosuppressive to lymphocytes regardless of their ability to be induced via the Ah locus and receptor. Experiments were designed to quantify the generation of metabolites of DMBA by lymphocytes incubated with [14C]DMBA and to ascertain whether radioactivity was covalently bound to cellular macromolecules in DMBA-exposed lymphocytes. No significant metabolism of DMBA was detected in culture supernatants, except when cultures were incubated in the presence of Arochlor-induced rat liver 9000 x g supernatants (S9). Covalent binding of 14C to cellular macromolecules was enhanced approximately eightfold in the presence of S9. Inhibition of monooxygenase activity by alpha-naphthoflavone did not modulate the immunosuppressive character of DMBA. Furthermore, addition of S9 did not amplify or ablate DMBA-mediated suppression of lymphocyte proliferation to the mitogen concanavalin A (Con A). Selected metabolites of DMBA were evaluated for immunosuppressive effects in cultures stimulated with mitogens and cellular alloantigens. 7-Hydroxymethyl-12-methylbenz[a]anthracene (OHMe) and 5,6-dihydro-5,6-dihydroxybenz[a]anthracene (Diol) were found to cause only slightly greater suppression of lymphocyte responses than DMBA. Thus, it appears that metabolites of DMBA were not responsible for the immunosuppression observed in lymphocyte cultures and that lymphocytes were not equipped to metabolize any significant amount of DMBA. These data lend support to the hypothesis that parent compound alone is responsible for the immunosuppressive effects observed in murine lymphocyte culture.

Topics: 9,10-Dimethyl-1,2-benzanthracene; Animals; Benzoflavones; Cells, Cultured; Chromatography, High Pressure Liquid; Concanavalin A; Cytochrome P-450 Enzyme Inhibitors; Female; Immunosuppression Therapy; Lymphocyte Activation; Lymphocytes; Mice; Mice, Inbred C3H; Mice, Inbred C57BL; Mice, Inbred DBA; Microsomes; Mixed Function Oxygenases; Spleen