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Background: Acute lung injury (ALI) is associated with high mortality due to the lack of effective therapeutic strategies. Mechanical ventilation itself can cause ventilator-induced lung injury. Pulmonary vascular barrier function, regulated in part by Src kinase-dependent phosphorylation of caveolin-1 and intercellular adhesion molecule-1 (ICAM-1), plays a crucial role in the development of protein-/neutrophil-rich pulmonary edema, the hallmark of ALI. Amide-linked local anesthetics, such as ropivacaine, have anti-inflammatory properties in experimental ALI. We hypothesized ropivacaine may attenuate inflammation in a "double-hit" model of ALI triggered by bacterial endotoxin plus hyperinflation via inhibition of Src-dependent signaling., Methods: C57BL/6 (WT) and ICAM-1 (-/-) mice were exposed to either nebulized normal saline (NS) or lipopolysaccharide (LPS, 10 mg) for 1 hour. An intravenous bolus of 0.33 mg/kg ropivacaine or vehicle was followed by mechanical ventilation with normal (7 ml/kg, NTV) or high tidal volume (28 ml/kg, HTV) for 2 hours. Measures of ALI (excess lung water (ELW), extravascular plasma equivalents, permeability index, myeloperoxidase activity) were assessed and lungs were homogenized for Western blot analysis of phosphorylated and total Src, ICAM-1 and caveolin-1. Additional experiments evaluated effects of ropivacaine on LPS-induced phosphorylation/expression of Src, ICAM-1 and caveolin-1 in human lung microvascular endothelial cells (HLMVEC)., Results: WT mice treated with LPS alone showed a 49% increase in ELW compared to control animals (p = 0.001), which was attenuated by ropivacaine (p = 0.001). HTV ventilation alone increased measures of ALI even more than LPS, an effect which was not altered by ropivacaine. LPS plus hyperinflation ("double-hit") increased all ALI parameters (ELW, EVPE, permeability index, MPO activity) by 3-4 fold compared to control, which were again decreased by ropivacaine. Western blot analyses of lung homogenates as well as HLMVEC treated in culture with LPS alone showed a reduction in Src activation/expression, as well as ICAM-1 expression and caveolin-1 phosphorylation. In ICAM-1 (-/-) mice, neither addition of LPS to HTV ventilation alone nor ropivacaine had an effect on the development of ALI., Conclusions: Ropivacaine may be a promising therapeutic agent for treating the cause of pulmonary edema by blocking inflammatory Src signaling, ICAM-1 expression, leukocyte infiltration, and vascular hyperpermeability.