We hypothesized that resuscitation with 100% O2 compared with 21% O2 is detrimental to pulmonary tissue. The pulmonary injury was assessed by matrix metalloproteinase (MMP) activity, oxidative stress, IL-8, and histology 2.5 h after resuscitation from a hypoxic state. In pulmonary tissue extracts, MMP activity was analyzed by broad matrix-degrading capacity (total MMP) and zymography. MMP-2 mRNA expression was evaluated by quantitative real-time PCR. Total endogenous antioxidant capacity was measured by the oxygen radical absorbance capacity (ORAC) assay, and IL-8 was analyzed by ELISA technique. In bronchoalveolar lavage (BAL) fluid, MMPs were analyzed by zymography. In pulmonary tissue, pro- and active MMP-2 levels were increased in piglets that were resuscitated with 100% O2 compared with 21% O2. Pro-MMP-9, total MMP activity, and MMP-2 mRNA levels were significantly increased in resuscitated piglets compared with baseline. Net gelatinolytic activity increased in submucosa and blood vessels after 100% O2 and only in the blood vessels after 21% O2. Compared with baseline, ORAC values were considerably lowered in the resuscitated piglets and significantly reduced in the 100% O2 versus 21% O2 group. In BAL fluid, both pro-MMP-9 and pro-MMP-2 increased 2-fold in the 100% O2 group compared with 21% O2. Moreover, IL-8 concentration increased significantly in piglets that were resuscitated with 100% O2 compared with 21% O2, suggesting a marked proinflammatory response in the pulmonary tissue. Altogether, these data strongly suggest that caution must be taken when applying pure O2 to the newborn infant.