Paraquat (PQ) is a specific pneumotoxin that causes tissue damage by enhancing oxidative stress within the lung. One likely target for PQ damage is the pulmonary endothelium. Oxidative stress is frequently associated with apoptotic cell death, but the ability of PQ to induce apoptosis in pulmonary endothelium has not been described. Peroxidation and translocation of specific membrane phospholipids such as phosphatidylserine (PS) are hallmarks of apoptosis following oxidative stress. It is hypothesized that oxidation of specific phospholipids can initiate or regulate PS externalization and subsequent intravascular coagulation following PQ-induced apoptosis. The overall objectives of this proposal are two-fold. The first is to define the role of specific phospholipid peroxidation as part of the molecular mechanism leading to PS externalization during PQ-induced apoptosis. The second is to specifically assess the ability of apoptotic pulmonary endothelium to activate coagulation cascades in a PS-dependent manner.
Three specific aims are proposed: (1) To demonstrate the potential for PQ to produce apoptosis and selective oxidation and externalization of PS in human pulmonary endothelium, (2) to determine the effect of PQ and oxidized phospholipid products on specific phospholipid translocating enzymes, and (3) to determine the functional potential of externalized PS on pulmonary endothelium to activate coagulation cascades. To meet these ends the investigators will measure peroxidation of specific phospholipids, movement of phospholipids across membranes, and apoptosis following exposure of pulmonary EC to PQ and specific products of lipid peroxidation. In addition, the ability of EC to activate various clotting factors in vitro in a PS-dependent manner under basal conditions and after PQ-induced apoptosis will be determined. These studies will provide valuable insight for the identification and redox modulation of processes that contribute pulmonary toxicity during environmental oxidative stress.
