Introductions: Previous studies in isolated perfused rat lungs have revealed that endothelial barrier disruption after intra-tracheal administration of pseudomonas aeruginosa (strain 103; PA103) only occurs after accumulation of extracellular mitochondrial DNA (mtDNA) Damage Associated Molecular Patterns (DAMPs) in the perfusate and is suppressed by addition of DNase to the perfusion medium. Herein, we tested the hypothesis that intra-tracheal DNase - a route of administration readily translatable to patient with ventilator-associated pneumonia (VAP) - also enhances degradation of mtDNA and prevents bacteria-induced lung injury. Methods: Intra-tracheal DNase was administered to isolated rat lungs either before or after intra-tracheal challenge with PA103 to determine if bacteria-induced, mtDNA DAMP-dependent lung injury could be prevented or reversed by enhanced mtDNA degradation. To explore whether this concept is translatable to patients with VAP, consecutive patients suspected of VAP were prospectively enrolled. All patients suspected of VAP received a bronchoalvelolar lavage (BAL) with quantitative culture for the diagnosis of VAP. Mitochondrial and nuclear DNA were measured from the BAL. MtDNA DAMPs (i.e. ND6) were measured from serum at time of suspected diagnosis and at 24-48 hours afterward. Results: Intra-tracheal PA103 caused significantly increased the vascular filtration coefficient (Kf) and perfusate mtDNA DAMPs. In contrast, lungs pre- or post-treated with intra-tracheal DNase were protected from increases in Kf and mtDNA DAMPs. Patients with the diagnosis of VAP had significantly higher mtDNA DAMPs in the BAL (248.70+/-109.7 vs 43.91+/-16.61, p
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