Nitric Oxide Generation during Cellular Metabolization of the Diabetogenic N-Mefhyl-N-Nitroso-Urea Streptozotozin Contributes to Islet Cell DNA Damage

Abstract

The N-methyl-N-nitroso-urea streptozotocin is an antibiotic with diabetogenic, carcinogenic and antitumor activity thought to act via alkylation of DNA and proteins. Evidence points to a release of bioactive nitric oxide (NO) from streptozotocin as an additional cytotoxic activity of this drug. Here we show by EPR spectroscopy, that NO is not generated during spontaneous decay of streptozotocin but that its metabolization in rat hepatocytes and pancreatic islet cells yields NO. This NO formation is not due to a NO synthase (NOS) activity since NO formation in hepatocytes in the presence of streptozotocin is not blocked by the NOS inhibitor NG-methyl-L-arginine. By iNOS-specific RT-PCR no positive signal for specific mRNA presence was obtained in streptozotocin-treated cells, proving that iNOS activity was not induced during cell isolation procedures and did not account for the NO release. Furthermore, early DNA-strand breaks induced either by SZ or by the NO donor nitroprusside were both significantly reduced in the presence of an intracellular NO scavenger. In contrast, DNA damage found after incubation with the purely alkylating agent methylmethanesulfonate was not inhibited by the NO trap. These results prove that intracellular formation of NO occurs during degradation of SZ within cells. This NO appears to contribute significantly to streptozotocin-induced cytotoxicity.