The aim was to study whether ozone affects vascular endothelium by causing inducible nitric oxide synthase (iNOS) expression and tyrosine nitration. We also studied biomarkers of endothelial function. Male Wistar rats were exposed to ozone (0.25 ppm, 4 h/day) or filtered air (control, ozone <0.05 ppm). After ozone exposure, blood samples were taken to measure 6-keto prostaglandin F1α (6-keto PGF1α), dehydro-thromboxane B2 (DH-TxB 2), endothelin-1 and NO2-/NO3 - (NOx-). iNOS and nitrotyrosine were detected in aorta by immunohistochemistry. Nitrotyrosine was also detected by immunoelectromicroscopy. Control aortae failed to show either iNOS or nitrotyrosine. Time-dependent positive iNOS and nitrotyrosine cells were observed in exposed animals. Except for NOx-, endothelial markers decreased after 14 days of ozone exposure (P<0.05). After 28 days of ozone, 6-keto PGF1α remained low (P<0.05) while DH-TxB2 increased (P<0.05). It is concluded that ozone causes endothelial dysfunction manifested early with peroxynitrite formation and lately with changes in endothelial markers.