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Lung endothelial dysfunction is a key feature of acute lung injury (ALI) and clinical acute respiratory distress syndrome (ARDS). Previous studies have identified the lipid-generating enzyme, group V phospholipase A2 (gVPLA2), as a mediator of lung endothelial barrier disruption and inflammation. The current study aimed to determine the role of gVPLA2 in mediating lung endothelial responses to methicillin-resistant Staphylococcus aureus (MRSA, USA300 strain), a major cause of ALI/ARDS. In vitro studies assessed the effects of gVPLA2 inhibition on lung endothelial cell (EC) permeability after exposure to heat-killed (HK) MRSA. In vivo studies assessed the effects of intratracheal live or HK-MRSA on multiple indices of ALI in wild-type (WT) and gVPLA2-deficient (KO) mice. In vitro, HK-MRSA increased gVPLA2 expression and permeability in human lung EC. Inhibition of gVPLA2 with either the PLA2 inhibitor, LY311727, or with a specific monoclonal antibody, attenuated the barrier disruption caused by HK-MRSA. LY311727 also reduced HK-MRSA-induced permeability in mouse lung EC isolated from WT but not gVPLA2-KO mice. In vivo, live MRSA caused significantly less ALI in gVPLA2 KO mice compared to WT, findings confirmed by intravital microscopy assessment in HK-MRSA-treated mice. After targeted delivery of gVPLA2 plasmid to lung endothelium using ACE antibody-conjugated liposomes, MRSA-induced ALI was significantly increased in gVPLA2-KO mice, indicating that lung endothelial expression of gVPLA2 is critical in vivo. In summary, these results demonstrate an important role for gVPLA2 in mediating MRSA-induced lung EC permeability and ALI. Thus, gVPLA2 may represent a novel therapeutic target in ALI/ARDS caused by bacterial infection. [ABSTRACT FROM AUTHOR]

R3191 --> Cigarette smoking (CS) is the major cause of Chronic Obstructive Pulmonary Disease (COPD). E‐cigarettes are considered by some as a safer alternative; however, inhalation of nicotine‐containing e‐cigarettes can also cause pathophysiologic changes, and "vaping" of some substances has led to severe lung damage. Our group has previously identified cortactin (CTTN), a central regulator of the actin cytoskeleton, as an important modulator of lung endothelial cell (EC) function. Our present study seeks to characterize the role of CTTN in lung endothelial dysfunction upon CS and e‐cigarette challenge. Human lung EC were exposed to CS (40 µg/ml, 24 h) and cell lysates were subjected to Western blotting for autophagy markers p62 and LC3B‐II. CS exposure increased both p62 and LC3B‐II expression, suggesting autophagic flux inhibition. Further CTTN down‐regulation by siRNA altered CS‐induced p62 and LC3B‐II expression. Excess mitochondrial ROS (mitoROS) production is associated with mitochondrial dysfunction and altered autophagy. Here, the mitoROS production in lung ECs was assessed using the MitoSOX assay. CS (40 µg/ml, 2 h) challenge significantly increased mitoROS production, which was potentiated by cortactin siRNA. Blocking mitoROS with Mito‐TEMPO (10 µM, 3 h), a cell‐permeable antioxidant, decreased mitochondrial superoxide production. We next assessed the role of Mito‐TEMPO on E‐cigarette‐induced endothelial permeability. Lung EC exposure to E‐cigarette extract (50 µg/ml) resulted in disruption of endothelial permeability, assessed by Electric Cell‐substrate Impedance Sensing system (ECIS). Pretreatment with Mito‐TEMPO provided barrier protection upon e‐cigarette challenge. These results suggest a key role of mitoROS in e‐cigarette‐induced endothelial permeability. In conclusion, this study identified new mechanisms by which CS/e‐cigarette exposure induces EC dysfunction and provides evidence for a previously undescribed role for CTTN in these responses. [ABSTRACT FROM AUTHOR]