Effect of PAR-2 Deficiency in Mice on KC Expression after Intratracheal LPS Administration

Protease activated receptors (PAR) have been shown to play a role in inflammation. PAR-2 is expressed by numerous cells in the lung and has either proinflammatory, anti-inflammatory, or no effect depending on the model. Here, we examined the role of PAR-2 in a model of LPS-induced lung inflammation. We found that PAR-2-deficient mice had significantly less KC expression in bronchial lavage fluid compared with wild-type mice but there was no difference in MIP-2 or TNF-α expression. We also found that isolated alveolar and resident peritoneal macrophages lacking PAR-2 showed a similar deficit in KC after LPS stimulation without differences in MIP-2 or TNF-α. Infiltration of neutrophils and macrophages into the lung following LPS administration was not affected by an absence of PAR-2. Our results support the notion that PAR-2 plays a role in LPS activation of TLR4 signaling in macrophages

Nrf2 Modulates Host Defense during Streptococcus pneumoniae Pneumonia in Mice

Nrf2 regulates the transcriptional response to oxidative stress. These studies tested the role of Nrf2 during S. pneumoniae pneumonia and identified Nrf2-dependent genes and pathways in lung tissue and in recruited neutrophils. Nrf2 null and WT mice were studied at 6 and 24 h following instillation of S. pneumoniae or PBS. At 6 h, fewer neutrophils were recruited and the number of bacterial remaining in the lungs tended to be less (p=0.06) in the Nrf2 null compared to WT mice. In uninfected lungs, 53 genes were already differentially expressed in Nrf2 null compared to WT mouse lungs and gene sets involved in phagocytosis, Fc receptor function, complement and immunoglobulin regulation are enhanced in PBS-treated Nrf2 null gene profiles compared to those of WT mice. These results suggest that initial host defense is enhanced in Nrf2 null mice, resulting in less recruitment of neutrophils. At 24 h, neutrophil recruitment was greater. The percentages of early apoptotic and late apoptotic/necrotic neutrophils were similar. At increasing inoculum numbers, mortality strikingly increased from 15% to 31% and 100% in Nrf2 null mice, whereas all WT mice survived, and Nrf2 null mice had a defect in clearance, particularly at the intermediate dose. The mortality was due to enhanced lung injury and greater systemic response. Gene profiling identified differentially regulated genes and pathways in neutrophils and lung tissue, including those involved in redox stress response, metabolism, inflammation, immunoregulatory pathways and tissue repair, providing insight into the mechanisms for the greater tissue damage and increased neutrophil accumulation

A perpetual switching system in pulmonary capillaries

Of the 300 billion capillaries in the human lung, a small fraction meet normal oxygen requirements at rest, with the remainder forming a large reserve. The maximum oxygen demands of the acute stress response require that the reserve capillaries are rapidly recruited. To remain primed for emergencies, the normal cardiac output must be parceled throughout the capillary bed to maintain low opening pressures. The flow-distributing system requires complex switching. Because the pulmonary microcirculation contains contractile machinery, one hypothesis posits an active switching system. The opposing hypothesis is based on passive switching that requires no regulation. Both hypotheses were tested ex vivo in canine lung lobes. The lobes were perfused first with autologous blood, and capillary switching patterns were recorded by videomicroscopy. Next, the vasculature of the lobes was saline flushed, fixed by glutaraldehyde perfusion, flushed again, and then reperfused with the original, unfixed blood. Flow patterns through the same capillaries were recorded again. The 16-min-long videos were divided into 4-s increments. Each capillary segment was recorded as being perfused if at least one red blood cell crossed the entire segment. Otherwise it was recorded as unperfused. These binary measurements were made manually for each segment during every 4 s throughout the 16-min recordings of the fresh and fixed capillaries (>60,000 measurements). Unexpectedly, the switching patterns did not change after fixation. We conclude that the pulmonary capillaries can remain primed for emergencies without requiring regulation: no detectors, no feedback loops, and no effectors-a rare system in biology. NEW & NOTEWORTHY The fluctuating flow patterns of red blood cells within the pulmonary capillary networks have been assumed to be actively controlled within the pulmonary microcirculation. Here we show that the capillary flow switching patterns in the same network are the same whether the lungs are fresh or fixed. This unexpected observation can be successfully explained by a new model of pulmonary capillary flow based on chaos theory and fractal mathematics

Cardiovascular risk factors cause premature rarefaction of the collateral circulation and greater ischemic tissue injury

Collaterals lessen tissue injury in occlusive disease. However, aging causes progressive decline in their number and smaller diameters in those that remain (collateral rarefaction), beginning at 16 months-age in mice (ie, middle age), and worse ischemic injury—effects that are accelerated in even 3 months-old eNOS−/− mice. These findings have found indirect support in recent human studies

Neonatal Pulmonary Macrophage Depletion Coupled to Defective Mucus Clearance Increases Susceptibility to Pneumonia and Alters Pulmonary Immune Responses

Resident immune cells (e.g., macrophages [MΦs]) and airway mucus clearance both contribute to a healthy lung environment. To investigate interactions between pulmonary MΦ function and defective mucus clearance, a genetic model of lysozyme M (LysM) promoter–mediated MΦ depletion was generated, characterized, and crossed with the sodium channel β subunit transgenic (Scnn1b-Tg) mouse model of defective mucus clearance. Diphtheria toxin A–mediated depletion of LysM+ pulmonary MΦs in wild-type mice with normal mucus clearance resulted in lethal pneumonia in 24% of neonates. The pneumonias were dominated by Pasteurella pneumotropica and accompanied by emaciation, neutrophilic inflammation, and elevated Th1 cytokines. The incidence of emaciation and pneumonia reached 51% when LysM+ MΦ depletion was superimposed on the airway mucus clearance defect of Scnn1b-Tg mice. In LysM+ MΦ-depleted Scnn1b-Tg mice, pneumonias were associated with a broader spectrum of bacterial species and a significant reduction in airway mucus plugging. Bacterial burden (CFUs) was comparable between Scnn1b-Tg and nonpneumonic LysM+ MΦ-depleted Scnn1b-Tg mice. However, the nonpneumonic LysM+ MΦ-depleted Scnn1b-Tg mice exhibited increased airway inflammation, the presence of neutrophilic infiltration, and increased levels of inflammatory cytokines in bronchoalveolar lavage fluid compared with Scnn1b-Tg mice. Collectively, these data identify key MΦ–mucus clearance interactions with respect to both infectious and inflammatory components of muco-obstructive lung disease

Mechanisms of Interferon-γ Production by Neutrophils and Its Function during Streptococcus pneumoniae Pneumonia

Bacterial pneumonia is a common public health problem associated with significant mortality, morbidity, and cost. Neutrophils are usually the earliest leukocytes to respond to bacteria in the lungs. Neutrophils rapidly sequester in the pulmonary microvasculature and migrate into the lung parenchyma and alveolar spaces, where they perform numerous effector functions for host defense. Previous studies showed that migrated neutrophils produce IFN-γ early during pneumonia induced by Streptococcus pneumoniae and that early production of IFN-γ regulates bacterial clearance. IFN-γ production by neutrophils requires Rac2, Hck/Lyn/Fgr Src family tyrosine kinases, and NADPH oxidase. Our current studies examined the mechanisms that regulate IFN-γ production by lung neutrophils during acute S. pneumoniae pneumonia in mice and its function. We demonstrate that IFN-γ production by neutrophils is a tightly regulated process that does not require IL-12. The adaptor molecule MyD88 is critical for IFN-γ production by neutrophils. The guanine nucleotide exchange factor CalDAG-GEFI modulates IFN-γ production. The CD11/CD18 complex, CD44, Toll-like receptors 2 and 4, TRIF, and Nrf2 are not required for IFN-γ production by neutrophils. The recently described neutrophil–dendritic cell hybrid cell, identified by its expression of Ly6G and CD11c, is present at low numbers in pneumonic lungs and is not a source of IFN-γ. IFN-γ produced by neutrophils early during acute S. pneumoniae pneumonia induces transcription of target genes in the lungs, which are critical for host defense. These studies underline the complexity of the neutrophil responses during pneumonia in the acute inflammatory response and in subsequent resolution or initiation of immune responses