Increased group IV cytosolic phospholipase A2 activity in lungs of sheep after smoke inhalation injury

Increased phospholipase A(2) (PLA(2)) activity was measured in cytosolic fractions of lungs fi om sheep exposed to smoke from burning cotton or to synthetic smoke consisting of carbon and acrolein, a cotton smoke toxin. Three peaks of PLA(2) activity were identified by heparin-Sepharose chromatography. The heparin-nonbinding PLA(2) activity was twofold higher in the extracts from lungs exposed to smoke than in normal lungs. This activity was identified as the group IV 85-kDa cytosolic PLA(2) (cPLA(2)). The activities of the forms of PLA(2) that bound to heparin did not change after smoke exposure. Those activities showed a pH optimum of 9.0, required a millimolar Ca2+ concentration for full activity, and were inhibited by 5 mM dithiothreitol. One activity eluted at an NaCl concentration typical for group Ib and V PLA(2) and had the expected substrate specificity. The other form of lung PLA(2) that bound heparin was a group II PLA(2). Lung myeloperoxidase activity increased progressively with increased exposure to smoke, cPLA(2) was identified in sheep neutrophils. With 30 breaths of smoke exposure, there was an increase in cPLA(2) activity without a difference in immunoreactivity on Western blot, indicating that the increased activity was not due to increased amounts of protein. In conclusion, smoke induces increases in resident lung cell cPLA(2) activity that is likely responsible for eicosanoid production, leading to lung inflammation and bronchoconstriction.