Contrasting effects of NO and peroxynitrites on HSP70 expression and apoptosis in human monocytes

Abstract

The free radicals nitric oxide (·NO) and superoxide (O2 −·) react to form peroxynitrite (ONOO−), a highly toxic oxidant species. In this study we investigated the respective effects of NO and ONOO− in monocytes from healthy human donors. Purified monocytes were incubated for 6 or 16 h with a pure NO donor ( S-nitroso- N-acetyl-dl-penicillamine, 0–2 mM), an ·NO/ONOO− donor (3-morpholinosydnonimine chlorhydrate, 0–2 mM) with and without superoxide dismutase (200 IU/ml), or pure ONOO−. We provide evidence that 3-morpholinosydnonimine chlorhydrate alone represents a strong stress to human monocytes leading to a dose-dependent increase in heat shock protein-70 (HSP70) expression, mitochondrial membrane depolarization, and cell death by apoptosis and necrosis. These phenomena were abolished by superoxide dismutase, suggesting that ONOO−, but not ·NO, was responsible for the observed effects. This observation was further strengthened by the absence of a stress response in cells exposed to S-nitroso- N-acetyl-dl-penicillamine. Conversely, exposure of cells to ONOO− alone also induced mitochondrial membrane depolarization and cell death by apoptosis and necrosis. Thus ONOO− formation may well explain the toxic effect generally attributed to ·NO.