Role of reactive metabolites in suppression of humoral immunity by N-nitrosodimethylamine.

The hypothesis that dimethylnitrosamine (DMN)-induced humoral immune suppression is mediated by reactive intermediates was tested in female B6C3F1 mice using three approaches. First, distribution of reactive species to known target organs (liver and kidney) and to immune organs (peripheral blood components, bone marrow cells, thymus and spleen) was demonstrated by monitoring acid-insoluble radioactivity after single or multiple doses of 6 mg/kg and 2 microCi of [methyl-14C] DMN. Most of the radioactivity resided in acid-insoluble material, which increased with the number of exposures and correlated with suppression of the in vivo antibody response to sheep erythrocytes as determined by enumerating immunoglobulin M antibody-forming cells. Fractionation of spleens from exposed animals into various cell types indicated greater association of acid-insoluble radioactivity with T- and B-lymphocytes as compared to macrophages. Fractionation of spleen cell lysates into protein, DNA and RNA indicated that most radioactivity was associated with the nucleic acid component. Pretreatment of mice with aminoacetonitrile reduced acid-insoluble disintegrations per minute in the spleen. The second approach was to demonstrate the immunosuppressive nature of compounds forming the same toxic intermediate as DMN. The addition of N-nitroso(acetoxymethyl)methylamine to spleen cell cultures resulted in completely suppressed in vitro antibody responses to sheep erythrocytes and dinitrophenylated Ficoll at micromolar concentrations. The third approach involved activating DMN to an immunosuppressive form in vitro. Preincubation of splenocytes with Aroclor-induced mouse hepatocytes resulted in activation of DMN to intermediates capable of markedly suppressing the response of antibody-forming cells to sheep erythrocytes.