Project Summary Acute lung injury (ALI) is a life-threatening condition, which affects > 200,000 patients annually in the U.S. It is associated with pneumonia, sepsis, and trauma, leading to pulmonary insufficiency and eventually multisystem organ failure. Since excessive recruitment of activated neutrophils to lung microvessels is a primary cause of ALI, a better understanding of the mechanism mediating neutrophil-endothelial cell interactions will provide insight into developing an effective therapeutic for treating ALI. We previously demonstrated that intravascular protein disulfide isomerase (PDI) enhances the ligand-binding activity of neutrophil and platelet surface receptors and leads to intravascular cell-cell interactions during vascular inflammation. To elucidate how extracellular PDI activity is regulated and whether the regulatory mechanism contributes to the pathology of ALI, we have found that endoplasmic reticulum oxidoreductin 1α (ERO1α, a key oxidase of PDI in the ER) promotes neutrophil recruitment during vascular inflammation. In this proposal, we will test the hypothesis that intravascular ERO1α enhances the ligand-binding function of neutrophil adhesion receptors by modifying disulfide bonds, inducing neutrophil recruitment to sites of acute lung inflammation. In Aim 1, we will determine the mechanism by which ERO1α regulates neutrophil-endothelial cell interactions. In Aim 2, we will utilize lung live imaging techniques to investigate the pathological role of intravascular ERO1α in ALI. In Aim 3, using the blood of patients with acute respiratory distress syndrome (ARDS), we will determine the contribution of ERO1α to the disease progression and severity in patients with ARDS. These studies will employ biochemical, cell biological, genetic, and confocal intravital imaging approaches. Since little is known about extracellular thiol-modifying machinery, the proposed studies will identify an essential, yet unexplored, mechanism that promotes the pathogenesis of ALI/ARDS.

Project Narrative The main question addressed in this proposal is how neutrophils are recruited to sites of acute lung injury (ALI) and whether an intravascular oxidase, endoplasmic reticulum oxidoreductin 1α (ERO1α) contributes to the disease process. Our preliminary data demonstrate that ERO1α enhances the adhesive function of neutrophil surface molecules. The goal of the proposed studies is to determine a previously undefined molecular and cellular mechanism by which intravascular ERO1α promotes neutrophil recruitment and vascular occlusion in ALI and contributes to the disease process. This project will provide valuable evidence that targeting ERO1α and its signaling might be novel therapeutic strategies for treating ALI.