This study was undertaken to determine if fibronectin containing an extra type III domain (ED1) is released during inflammatory injury of pulmonary tissue in vivo. ED1 fibronectin, measured by quantitative immunoassay, was markedly increased in bronchoalveolar lavage (BAL) fluid (2 and 4 h posttreatment) from rabbits with lung injury resulting from intrabronchial treatment with glucose oxidase and glucose to generate H2O2. This protein comprised a greater portion of total fibronectin in BAL fluid than in plasma, suggesting local release. Leukocyte-mediated lung injury after intrabronchial or intravenous treatment with phorbol myristate acetate (PMA) also affected ED1 fibronectin, triggering specific accumulation of this fibronectin variant in both BAL fluid and plasma. ED1 fibronectin in tissue fluids was largely intact and dimeric during injury, as demonstrated by Western blot analysis. Compartmental release of soluble ED1 fibronectin reflects acute pulmonary injury induced by oxidants or leukocytes in vivo.