Role of platelet-activating factor during gram-negative bacteremia-induced acute lung injury

Platelet-activating factor (PAF) is a vasoactive, pro-inflammatory, phospholipid involved in the pathogenesis of multiple inflammatory conditions such as gram-negative sepsis, acute lung injury (ALI) and multisystem organ dysfunction. Blood levels of PAF are elevated in patients with sepsis and exogenous PAF reproduces many of the hemodynamic and respiratory features of gram-negative sepsis induced ALI. Platelets, leukocytes and endothelial cells, among others, produce PAF. PAF exerts biological effects by binding to a membrane specific receptor, which in turn is coupled to a complex signaling pathway. Despite the overwhelming evidence that PAF is one of the primary mediators of gram-negative sepsis-induced ALI, the mechanism by which PAF causes pulmonary edema are not clear. This dissertation studies the hypothesis that during gram-negative sepsis PAF induces pulmonary edema by multiple mechanisms. Those mechanisms include: (1) PAF increases pulmonary microvascular permeability; (2) PAF potentiates gram-negative sepsis induced pulmonary hypertension and injury; (3) PAF facilitates pulmonary neutrophil trapping and neutrophil-induced ALI during gram-negative bacteremia; (4) PAF causes relaxation of pulmonary arteries but contraction of pulmonary veins thus creating a pressure gradient that favors pulmonary edema; (5) PAF induces pulmonary microvascular endothelial cell cytoskeletal changes, cell contraction and intercellular gap formation, which in turn increase permeability; and (6) PAF receptor inhibitors prevent ALI during gram-negative sepsis. To explore these hypotheses experiments were performed: Rat in vivo animal studies (hemodynamic monitoring and fluorescent microscopy pulmonary edema assessment); In vitro vascular ring studies of rat pulmonary arteries and veins; In vitro F-actin fluorescence labeling of human and rat pulmonary microvascular endothelial cells; and ELISAs of nitric oxide metabolites and endothelin from serum samples.; We learned from our experiments that in the rat intravenous PAF causes dose dependent receptor-mediated pulmonary edema, and pulmonary hypotension. We demonstrated that topical PAF applied to the lung surface increases microvascular permeability independently of hemodynarnic changes. We found that PAF potentiates pulmonary hypertension during E. coli bacteremia by upregulating ET-1 (a potent pulimonary vasoconstrictor peptide). PAF facilitates neutropbil trapping and neutropbil-induced lung injury by a receptor-mediator mechanism. We conclude that in the rat PAF favors pulmonary edema by relaxing pulmonary arteries while contracting pulmonary veins. Our results also demonstrate that in contrast to macrovascular pulmonary endothelial cells, in which PAF directly increases permeability by inducing cytoskeletal changes, PAF requires neutrophils or other blood components to increase permeability in pulmonary microvascular endothelial cells. And finally, platelet-activating factor receptor inhibition prevents pulmonary edema and pulmonary hypertension during gram-negative bacteremia and may be beneficial in the treatment of sepsis...