Acid aspiration-induced lung injury is a common disease in the intensive care unit (ICU) and acute respiratory distress syndrome (ARDS). Hypoxia-inducible factor (HIF)-1α is a major transcription factor responsible for regulating the cellular response to changes in oxygen tension. A clear understanding of the function of HIF-1α in lung inflammatory response is currently lacking. Here, we sought to determine the role of HIF-1α in type 2 alveolar epithelial cells (AEC) in the generation of the acute inflammatory response following gastric aspiration (GA). GA led to profound hypoxia at very early time points following GA. This correlated to a robust increase in HIF-1α, tissue albumin and pro-inflammatory mediators following GA in AECs. The extent of lung injury and the release of pro/anti-inflammatory cytokines were significantly reduced in HIF-1α (−/−) mice. Finally, we report that HIF-1α upregulation of the acute inflammatory response is dependent on NF-κB following GA.

Objectives Lung contusion (LC) is a major risk factor for the development of acute respiratory distress syndrome (ARDS). We set to determine the role of TLR3 and the binding of dsRNA in the pathogenesis of sterile injury following lung contusion. Methods TLR3 expression was analyzed in post-mortem lung samples from patients with LC. Unilateral LC was induced in TLR3 (−/−), TRIF (−/−), and WT mice; subsequently, lung injury and inflammation were evaluated. Apoptotic indices, phagocytic activity and phenotypic characterization of the macrophages were determined. Double stranded (ds) RNA in BAL and serum samples following LC were measured. A TLR3/dsRNA ligand inhibitor was injected into WT mice prior to LC. Measurements and Main Results TLR3 expression was higher in patients and WT mice with LC. The degree of lung injury, inflammation, and macrophage apoptosis was reduced in TLR3 (−/−), TRIF (−/−), and WT mice with TLR3 antibody neutralization. Alveolar macrophages from TLR3 (−/−) mice had a lower early apoptotic index, a predominant M2 phenotype and increased surface translocation of TLR3 from the endosome to the surface. When compared to viral activation pathways, lung injury in LC demonstrated increased p38 MAPK, ERK1/2 phosphorylation with inflammasome activation without a corresponding increase in NF-kB or Type-1 Interferon production. Additionally, pretreatment with TLR3/dsRNA ligand inhibitor led to a reduction in injury, inflammation, and macrophage apoptosis. Conclusions We conclude that the interaction of dsRNA from injured cells with TLR3 drives the acute inflammatory response following LC.

BACKGROUND Lung contusion (LC) is a common injury resulting from blunt thoracic trauma. LC is an important risk factor for the development acute lung injury, adult respiratory distress syndrome, and ventilator-associated pneumonia, all of which increase mortality from trauma. LC produces a nonspecific immune cellular response. Neutrophil recruitment is known to increase the severity of inflammation during LC. However, the exact role of macrophages in modulating the response to LC has not been well described. METHODS We used a cortical contusion impactor to induce unilateral LC in mice. Thoracic micro computed tomographic scans of these animals were obtained to document radiologic changes over time following LC. To understand the role of macrophages during LC, liposomal clodronate was used to deplete macrophage levels before traumatic insult. Acute inflammatory attributes after LC were assessed, by measuring pressure-volume mechanics; quantifying bronchial alveolar lavage levels of leukocytes, albumin, and cytokines; and finally examining lung specimen histopathology at 5, 24, 48, and 72 hours after injury. RESULTS After LC, alveolar macrophage numbers were significantly reduced and exhibited slowed recovery. Simultaneously, there was a significant increase in bronchial alveolar lavage neutrophil counts. The loss of macrophages could be attributed to both cellular apoptosis and necrosis. Pretreatment with clodronate increased the severity of lung inflammation as measured by worsened pulmonary compliance, increased lung permeability, amplification of neutrophil recruitment, and increases in early proinflammatory cytokine levels. CONCLUSION The presence of regulatory alveolar macrophages plays an important role in the pathogenesis of acute inflammation following LC.

Injured patients with lung contusion (LC) are at risk of developing bacterial pneumonia (PNA) followed by sepsis and death. A recent Genome-wide Association Study, showed FER gene expression positively correlating with survival rates among individuals with above conditions. We sought to determine if electroporation-mediated (EP) delivery of FER gene could indeed improve survival, in a lethal model of combined LC and PNA. C57BL/6 mice sustained unilateral LC, which preceded a 500 Klebsiella CFU inoculation by 6 hrs. In-between these insults, human FER plasmid (pFER) was introduced into the lungs followed by eight EP pulses applied externally (10ms at 200V/cm). Control groups included EP of empty vector (pcDNA3) or Na+/K+-ATPase genes (pPump) and no treatment (LC + PNA). We recorded survival, histology, lung mechanics, bronchial alveolar fluid (BAL), FER and inflammatory gene expression and bacteriology. The data shows that 7-day survival was significantly improved by pFER compared to control groups. pFER increased BAL monocytes and activated antibacterial response genes (NOS, Fizz). pFER treatment showed decreased lung and blood Klebsiella counts reaching, in some cases, complete sterilization. In conclusion, FER gene delivery promoted survival in LC+PNA mice via recruitment of activated immune cells, improving efficiency of bacterial clearance within contused lung