The critical role of Hepes in SIN-1 cytotoxicity, peroxynitrite versus hydrogen peroxide

• Published in: Free radical biology & medicine 1998-03, Vol.24 (4), p.522-528
• Main Authors: Lomonosova, E E, Kirsch, M, Rauen, U, de Groot, H
• Format: Article
• Language: English
• Subjects: Animals; Buffers; Catalase - pharmacology; Cell Death; Cell Line; Fibroblasts; HEPES - administration & dosage; HEPES - pharmacology; Hydrogen Peroxide - metabolism; Hydrogen Peroxide - pharmacology; Kinetics; L-Lactate Dehydrogenase - metabolism; Mice; Molsidomine - analogs & derivatives; Molsidomine - metabolism; Molsidomine - toxicity; Nitrates - metabolism; Nitrates - pharmacology
• ISSN: 0891-5849
• DOI: 10.1016/s0891-5849(97)00295-5

The cytotoxicity of the superoxide anion radical- and nitric oxide-releasing compound SIN-1 to L929 cells was studied in Krebs-Henseleit buffer. pH 7.4, in the presence and absence of Hepes. SIN-1 cytotoxicity was significantly higher in the presence of Hepes than in the absence of Hepes. The available amount of peroxynitrite formed from SIN-1, however, was significantly decreased by Hepes as indicated by decreased oxidation of dihydrorhodamine 123. On the other hand, Hepes largely increased the formation of H2O2 from SIN-1. Catalase protected the L929 cells from SIN-1 cytotoxicity in the buffer with Hepes. In the buffer without Hepes catalase did not have any protective effect. In contrast, tyrosine and tryptophan provided significant protection against SIN-1 cytotoxicity independent of the presence of Hepes. These results demonstrate that the immediate toxic agent formed from SIN-1 decisively depends on the presence of Hepes. In its absence cytotoxicity is most likely mediated by peroxynitrite while in the presence of Hepes, cytotoxicity is conveyed by co-operative action of hydrogen peroxide and reactive nitrogen species.