Denitrosation of 1,3-dimethyl-2-cyano-1-nitrosoguanidine in rat primary hepatocyte cultures.

N-Nitrosoguanidines are potential carcinogens. However, the toxicity of these agents is attenuated significantly in laboratory rodents by processes that remove the nitroso group to generate the relatively innocuous parent guanidinium compound. The denitrosation of 1,3-dimethyl-2-cyano-1-nitrosoguanidine (CyanoDMNG) mediated by rat hepatocytes in primary culture was investigated. At concentrations < or = 200 microM, applied CyanoDMNG was converted efficiently to 1,3-dimethyl-2-cyanoguanidine (CyanoDMG). In trials using 50 microM CyanoDMNG (5 mL dosing solutions), it was demonstrated that hepatocytes are capable of denitrosating a 40 microM concentration of the applied compound with little change in the total or oxidized glutathione levels. The process was inhibited by coincidently applied ethacrynic acid, a glutathione transferase inhibitor. Reduction of hepatocyte glutathione to 20% of control levels by buthionine sulfoximine pretreatment had little effect on CyanoDMG production; total depletion of cytosolic glutathione by diethyl maleate pretreatment arrested CyanoDMNG processing. Hepatocyte-mediated CyanoDMNG denitrosation did not generate nitrite; nitrate yields were 10% relative to the CyanoDMG produced. The mercuric chloride/azo dye response of cultures lysed at times during 50 microM CyanoDMNG processing indicated intact CyanoDMNG as the only dye-sensitive material present. At applied CyanoDMNG > 100 microM, S-nitrosoglutathione (GSNO) yields were detectable; 4 microM GSNO was generated (concentration in 5 mL lysates) and maintained during 60 min at the 200 microM CyanoDMNG treatment level; this yield decayed if CyanoDMNG was withdrawn. Based on these and previous findings, it is hypothesized that CyanoDMNG is converted to CyanoDMG and GSNO by glutathione transferases and that GSNO is catabolized to eventually regenerate reduced glutathione. The fate of most of the NO moiety released remains to be determined.