High Mobility Group Box–1 Mediates Hyperoxia-Induced Impairment ofPseudomonas aeruginosaClearance and Inflammatory Lung Injury in Mice

Abstract

Mechanical ventilation with supraphysiological concentrations of oxygen (hyperoxia) is routinely used to treat patients with respiratory distress. However, a significant number of patients on ventilators exhibit enhanced susceptibility to infections and develop ventilatorassociated pneumonia (VAP). Pseudomonas aeruginosa (PA) is one of the most common species of bacteria found in these patients. Previously, we demonstrated that prolonged exposure to hyperoxia can compromise the ability of alveolar macrophages (AMs), an essential part of the innate immunity, to phagocytose PA. This study sought to investigate the potential molecular mechanisms underlying hyperoxiacompromised innate immunity against bacterial infection in a murine model of PA pneumonia. Here, we show that exposure to hyperoxia (> 99% O 2 ) led to a significant elevation in concentrations of airway high mobility group box-1 (HMGB1) and increased mortality in C57BL/6 mice infected with PA. Treatment of these mice with a neutralizing anti-HMGB1 monoclonal antibody (mAb) resulted in a reduction in bacterial counts, injury, and numbers of neutrophils in the lungs, and an increase in leukocyte phagocytic activity compared with mice receiving control mAb. This improved phagocytic function was associated with reduced concentrations of airway HMGB1. The correlation between phagocytic activity and concentrations of extracellular HMGB1 was also observed in cultured macrophages. These results indicate a pathogenic role for HMGB1 in hyperoxia-induced impairment with regard to a host's ability to clear bacteria and inflammatory lung injury. Thus, HMGB1 may provide a novel molecular target for improving hyperoxia-compromised innate immunity in patients with VAP.

Partially-desulfated heparin improves survival inPseudomonaspneumonia by enhancing bacterial clearance and ameliorating lung injury

“…For the studies involving assessment of the impact of ODSH on host resistance to Pseudomonas aeruginosa (PA), mice were anesthetized with isoflurane or sodium pentobarbital (60 mg/kg) as described previously (Patel et al, 2013) and then intranasally inoculated with PA (as indicated in figure legends) that had been prepared as outlined below. ODSH was then administered by subcutaneous injection at various timepoints post-infection.…”

“…BAL samples were prepared as described previously (Entezari et al 2012;Patel et al 2013). Briefly, after mice were euthanized, the trachea was exposed and dissected, and a 20-Gauge (Â1¼ in) catheter was inserted.…”

“…Viable bacterial counts in the airways and lungs were determined as described previously (Entezari et al 2012;Patel et al 2013). Briefly, viable bacterial counts in the airways and lungs were determined using by plating serial dilutions of the BAL and lung homogenates, respectively, onto Pseudomonas Isolation Agar (PIA, Difco; Sparks, MD) and culturing at 37 C. Each dilution was plated in duplicates.…”

“…BAL HMGB1 levels were assessed via immunoblot analysis using anti-HMGB1 antibodies as noted previously in Patel et al (2013). In brief, samples were separated on SDS-PAGE.…”

“…It has been shown that extracellular HMGB1 mediates neutrophilic lung injury (Abraham et al, 2000). Our recent studies showed that extracellular HMGB1 in airways can directly compromise host ability to clear PA infections, exacerbating PA-induced lung injury in animal models of cystic fibrosis and ventilator-associated pneumonia (Entezari et al, 2012;Patel et al, 2013).…”