RESUMEN
Pulmonary infection with Pseudomonas aeruginosa and neutrophilic lung inflammation significantly contribute to morbidity and mortality in cystic fibrosis (CF). High-mobility group box 1 protein (HMGB1), a ubiquitous DNA binding protein that promotes inflammatory tissue injury, is significantly elevated in CF sputum. However, its mechanistic and potential therapeutic implications in CF were previously unknown. We found that HMGB1 levels were significantly elevated in bronchoalveolar lavage fluids (BALs) of CF patients and cystic fibrosis transmembrane conductance regulator (CFTR )(-/-) mice. Neutralizing anti-HMGB1 monoclonal antibody (mAb) conferred significant protection against P. aeruginosa-induced neutrophil recruitment, lung injury and bacterial infection in both CFTR(-/-) and wild-type mice. Alveolar macrophages isolated from mice treated with anti-HMGB1 mAb had improved phagocytic activity, which was suppressed by direct exposure to HMGB1. In addition, BAL from CF patients significantly impaired macrophage phagocytotic function, and this impairment was attenuated by HMGB1-neutralizing antibodies. The HMGB1-mediated suppression of bacterial phagocytosis was attenuated in macrophages lacking toll-like receptor (TLR)-4, suggesting a critical role for TLR4 in signaling HMGB1-mediated macrophage dysfunction. These studies demonstrate that the elevated levels of HMGB1 in CF airways are critical for neutrophil recruitment and persistent presence of P. aeruginosa in the lung. Thus, HMGB1 may provide a therapeutic target for reducing bacterial infection and lung inflammation in CF.
Asunto(s)
Fibrosis Quística/inmunología , Proteína HMGB1/inmunología , Neumonía Bacteriana/inmunología , Infecciones por Pseudomonas/inmunología , Pseudomonas aeruginosa , Animales , Anticuerpos Monoclonales/farmacología , Anticuerpos Monoclonales/uso terapéutico , Líquido del Lavado Bronquioalveolar/inmunología , Línea Celular , Fibrosis Quística/tratamiento farmacológico , Femenino , Humanos , Macrófagos/inmunología , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos CFTR , Ratones Noqueados , Neutrófilos/inmunología , Fagocitosis/inmunología , Neumonía Bacteriana/tratamiento farmacológico , Infecciones por Pseudomonas/tratamiento farmacológico , Receptor Toll-Like 2/deficiencia , Receptor Toll-Like 2/genética , Receptor Toll-Like 4/deficiencia , Receptor Toll-Like 4/genéticaRESUMEN
Prolonged exposure to hyperoxia results in acute lung injury (ALI), accompanied by a significant elevation in the levels of proinflammatory cytokines and leukocyte infiltration in the lungs. However, the mechanisms underlying hyperoxia-induced proinflammatory ALI remain to be elucidated. In this study, we investigated the role of the proinflammatory cytokine high mobility group box protein 1 (HMGB1) in hyperoxic inflammatory lung injury, using an adult mouse model. The exposure of C57BL/6 mice to ≥99% O2 (hyperoxia) significantly increased the accumulation of HMGB1 in the bronchoalveolar lavage fluids (BALF) prior to the onset of severe inflammatory lung injury. In the airways of hyperoxic mice, HMGB1 was hyperacetylated and existed in various redox forms. Intratracheal administration of recombinant HMGB1 (rHMGB1) caused a significant increase in leukocyte infiltration into the lungs compared to animal treated with a non-specific peptide. Neutralizing anti-HMGB1 antibodies, administrated before hyperoxia significantly attenuated pulmonary edema and inflammatory responses, as indicated by decreased total protein content, wet/dry weight ratio, and numbers of leukocytes in the airways. This protection was also observed when HMGB1 inhibitors were administered after the onset of the hyperoxic exposure. The aliphatic antioxidant, ethyl pyruvate (EP), inhibited HMGB1 secretion from hyperoxic macrophages and attenuated hyperoxic lung injury. Overall, our data suggest that HMGB1 plays a critical role in mediating hyperoxic ALI through the recruitment of leukocytes into the lungs. If these results can be translated to humans, they suggest that HMGB1 inhibitors provide treatment regimens for oxidative inflammatory lung injury in patients receiving hyperoxia through mechanical ventilation.
Asunto(s)
Lesión Pulmonar Aguda/inducido químicamente , Lesión Pulmonar Aguda/patología , Anticuerpos/administración & dosificación , Líquido del Lavado Bronquioalveolar/inmunología , Proteína HMGB1/metabolismo , Piruvatos/administración & dosificación , Acetilación , Lesión Pulmonar Aguda/inmunología , Lesión Pulmonar Aguda/terapia , Animales , Hipoxia de la Célula , Línea Celular , Modelos Animales de Enfermedad , Proteína HMGB1/antagonistas & inhibidores , Inyecciones Espinales , Pulmón/citología , Pulmón/inmunología , Pulmón/patología , Masculino , Ratones , Ratones Endogámicos C57BL , Neutrófilos/metabolismoRESUMEN
Mechanical ventilation with hyperoxia is a necessary treatment for patients with respiratory distress. However, patients on mechanical ventilation have increased susceptibility to infection. Studies including ours have shown that reactive oxygen species (ROS), generated by exposure to prolonged hyperoxia, can cause a decrease in the phagocytic activity of alveolar macrophages. Hydrogen peroxide (H2O2) is a form of ROS generated under hyperoxic conditions. In this study, we examined whether treatment with H2O2 directly affects macrophage phagocytic ability in RAW 264.7 cells that were exposed to either 21% O2 (room air) or 95% O2 (hyperoxia). Moderate concentrations (ranging from 10 to 250 µM) of H2O2 significantly enhanced macrophage phagocytic activity and restored hyperoxia-suppressed phagocytosis through attenuation of hyperoxia-induced disorganization of actin cytoskeleton and actin oxidation. These results indicate that H2O2 at low-moderate concentrations can be beneficial to host immune responses by improving macrophage phagocytic activity.