Clinical studies have shown a relationship between markers of oxidative stress and parameters of arterial stiffness. We have previously found that vascular reactive oxygen species (ROS) contribute to aortic hypertrophy in hypertension. In the present study, we hypothesize that vascular oxidative stress induces aortic stiffening by causing oxidative injury to the vessels. To test this hypothesis, we first studied isolated aortas from tgsm/p22phox mice, which have a constitutive increase in vascular ROS production. Six month old tgsm/p22phox mice were normotensive at baseline (118±6 systolic, 89±5 diastolic, mean±S.E.M. mmHg), but had increased aortic stiffness as indicated by the leftward shift on stress-strain curves (p<0.05 vs. wild type, one way ANOVA). While treatment with angiotensin II increased aortic stiffness in wild-type mice, it did not further increase stiffness in the tgsm/p22phox mice. In order to understand the nature of ROS involved in aortic stiffening, we deleted extracellular superoxide dismutase (ecSOD) in vascular smooth muscle using cre-loxp technology. Deletion of ecSOD would increase O2·-, but not H2O2. Surprisingly, we found no difference in aortic stiffness or blood pressure in these mice compared to wild-type mice, either at baseline or in response to angiotensin II. These data indicate that O2·- is unlikely involved in increasing aortic stiffness. It is therefore likely that other ROS, such as H2O2, modulate this response. In this regard, we have performed additional studies in which mice were treated with salicylamine, a scavenger of isoketals, which are oxidized products of arachidonic acid. These are known to cross-link lysines and could promote cross-linking of cellular components leading to vascular stiffening. Amazingly, this drug markedly reduced blood pressure (133±8 vs. 168±9 systolic; 95±6 vs. 125±7 diastolic, salicylamine vs. angiotensin II alone, p<0.01) and aortic stiffening in response to angiotensin II. In conclusion, our data strongly suggest that vascular ROS promotes aortic stiffening by inducing oxidative injury to the vessels.