The aim of this project is to investigate the mechanism(s) of tetranitromethane (TNM) pulmonary toxicity and carcinogenicity. TNM is commonly used in protein chemistry because of its specificity for nitrating the ortho position of tyrosine (Tyr) residues in proteins. Because of this highly specific reaction of TNM studies were conducted to evaluate the ability of TNM to nitrate cell membrane Tyr residues and to investigate the subsequent effects of Tyr nitration on cellular functions dependent upon Tyr kinase pathways. One such cellular function is the lipopolysaccharide/interferon (LPS/IFN) stimulated production of nitric oxide (NO) by macrophages. Rat alveolar macrophages (AM) were treated with 0, 0.312, 0.625, or 1.25 M TNM in serum-free media for 2 hours, then stimulated with LPS/IFN for 24 hours, and NO production measured. TNM treatment caused a dose-related decrease in the amount of NO released by stimulated AMs. Cell viability was not affected by these doses of TNM. Direct measurement of Tyr nitration using antibodies to N-tyr revealed an increased staining of AM membranes after exposure to TNM. These data suggest that TNM toxicity may be due in part to inhibition of Tyr kinase pathways caused by nitration of membrane Tyr.
