Peroxynitrite induces haem oxygenase-1 in vascular endothelial cells: a link to apoptosis

Peroxynitrite (ONOO-) is a potent oxidizing agent generated by the interact ion of nitric oxide (NO) and the superoxide anion. In physiological solutio n, ONOO- rapidly decomposes to a hydroxyl radical, one of the most reactive free radicals, and nitrogen dioxide, another species able to cause oxidati ve damage. In the present study we investigated the effect of ONOO- on the expression of haem oxygenase-l (HO-1), an inducible protein that is highly up-regulated by oxidative stress. Exposure of bovine aortic endothelial cel ls to ONOO- (250-1000 mu M) produced a concentration-dependent increase in haem oxygenase activity and HO-1 protein expression. This effect was comple tely abolished by the ONOO- scavengers uric acid and N-acetyl-cysteine, and partly attenuated by 1,3-dimethyl-2-thiourea, a scavenger of hydroxyl radi cals. ONOO- also produced a concentration-dependent increase in apoptosis a nd cytotoxicity, which were considerably decreased by uric acid and N-acety lcysteine. A 70 % decrease in apoptosis was observed when cells were expose d to ONOO- in the presence of 10 mu M tin protoporphyrin IX (SnPPIX), an in hibitor of haem oxygenase activity. When SnPPIX was added 5 min after ONOO- , apoptosis decreased by only 40%,, which suggests that an interaction betw een ONOO- and the protoporphyrin occurs in our system. Increased haem oxyge nase activity by pretreatment of cells with haemin resulted in elevated bil irubin production and was associated with a substantial decrease (35%) in O NOO--mediated apoptosis. These results indicate the ability of ONOO- to mod ulate the expression of the stress protein HO-1 and suggest that the haem o xygenase pathway contributes to protection against the cytotoxic action of ONOO-.