Background: This study sought to determine the impact of an acute prior bout of high-intensity interval aerobic exercise on attenuating the vascular dysfunction associated with a prolonged sedentary bout. Methods: Ten young (24 ± 1 y) healthy males completed two 3-hour sessions of prolonged sitting with (SIT-EX) and without (SIT) a high-intensity interval aerobic exercise session performed immediately prior. Prior to and 3 hours into the sitting bout, leg vascular function was assessed with the passive leg movement technique, and blood samples were obtained from the lower limb to evaluate changes in oxidative stress (malondialdehyde and superoxide dismutase) and inflammation (interleukin-6). Results: No presitting differences in leg vascular function (assessed via passive leg movement technique-induced hyperemia) were revealed between conditions. After 3 hours of prolonged sitting, leg vascular function was significantly reduced in the SIT condition, but unchanged in the SIT-EX. Lower limb blood samples revealed no alterations in oxidative stress, antioxidant capacity, or inflammation in either condition. Conclusions: This study revealed that lower limb vascular dysfunction was significantly attenuated by an acute presitting bout of high-intensity interval aerobic exercise. Further analysis of lower limb blood samples revealed no changes in circulating oxidative stress or inflammation in either condition.