DNA strand breakage, activation of poly(ADP-ribose) synthetase, and cellular energy depletion are involved in the cytotoxicity in macrophages and smooth muscle cells exposed to peroxynitrite

Csaba Szabó, Basilia Zingarelli, Michael O'Connor, Andrew L. Salzman

    Research output: Contribution to journalArticle

    597 Scopus citations

    Abstract

    The free radicals nitric oxide and superoxide anion react to form peroxynitrite (ONOO-), a highly toxic oxidant species. In vivo formation of ONOO+ has been demonstrated in shock and inflammation. Herein we provide evidence that cytotoxicity in cells exposed to ONOO- is mediated by DNA strand breakage and the subsequent activation of the DNA repair enzyme poly(ADP ribose) synthetase (PARS). Exposure to ONOO- (100 μM to 1 mM) inhibited mitochondrial respiration in cultured J774 macrophages and in rat aortic smooth muscle cells. The loss of cellular respiration was rapid, peaking 1-3 h after ONOO- exposure, and reversible, with recovery after a period of 6-24 h. The inhibition of mitochondrial respiration was paralleled by a dose-dependent increase in DNA strand breakage, reaching its maximum at 20-30 min after exposure to ONOO-. We observed a dose-dependent increase in the activity of PARS in cells exposed to ONOO+. Inhibitors of PARS such as 3-aminobenzamide (1 mM) prevented the inhibition of cellular respiration in cells exposed to ONOO-. Activation of PARS by ONOO--mediated DNA strand breakage resulted in a significant decrease in intracellular energy stores, as reflected by a decline of intracellular NAD+ and ATP content. 3- Aminobenzamide prevented the loss of NAD+ and ATP in cells exposed to ONOO- . In contrast, impairment of cellular respiration by the addition of the nitric oxide donors S-nitroso-N-acetyl-DL-penicillamine or diethyltriamine nitric oxide complex, was not associated with the development of DNA strand breaks, in concentrations up to 1 mM, and was largely refractory to PARS inhibition. Our results suggest that DNA damage and activation of PARS, an energy-consuming futile repair cycle, play a central role in ONOO-mediated cellular injury.

    Original languageEnglish (US)
    Pages (from-to)1753-1758
    Number of pages6
    JournalProceedings of the National Academy of Sciences of the United States of America
    Volume93
    Issue number5
    DOIs
    StatePublished - Mar 5 1996

    Keywords

    • endotoxin shock
    • inflammation
    • nitric oxide
    • nitric oxide synthase
    • superoxide

    ASJC Scopus subject areas

    • General

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