Excessive Airway Inflammation Research Articles

Vimentin affects inflammation and neutrophil recruitment in airway epithelium during Streptococcus suis serotype 2 infection

Streptococcus suis serotype 2 (SS2) frequently colonizes the swine upper respiratory tract and can cause Streptococcal disease in swine with clinical manifestations of pneumonia, meningitis, and septicemia. Previously, we have shown that vimentin, a kind of intermediate filament protein, is involved in the penetration of SS2 through the tracheal epithelial barrier. The initiation of invasive disease is closely related to SS2-induced excessive local inflammation; however, the role of vimentin in airway epithelial inflammation remains unclear. Here, we show that vimentin deficient mice exhibit attenuated lung injury, diminished production of proinflammatory cytokines interleukin-6 (IL-6), tumor necrosis factor-alpha (TNF-α), and the IL-8 homolog, keratinocyte-derived chemokine (KC), and substantially reduced neutrophils in the lungs following intranasal infection with SS2. We also found that swine tracheal epithelial cells (STEC) without vimentin show decreased transcription of IL-6, TNF-α, and IL-8. SS2 infection caused reassembly of vimentin in STEC, and pharmacological disruption of vimentin filaments prevented the transcription of those proinflammatory cytokines. Furthermore, deficiency of vimentin failed to increase the transcription of nucleotide oligomerization domain protein 2 (NOD2), which is known to interact with vimentin, and the phosphorylation of NF-κB protein p65. This study provides insights into how vimentin promotes excessive airway inflammation, thereby exacerbating airway injury and SS2-induced systemic infection.

Reduced neutrophil elastase inhibitor elafin and elevated transforming growth factor-β1 are linked to inflammatory response in sputum of cystic fibrosis patients with Pseudomonas aeruginosa.

Research questionPulmonary disease progression in patients with cystic fibrosis (CF) is characterised by inflammation and fibrosis and aggravated by Pseudomonas aeruginosa (Pa). We investigated the impact of Pa specifically on: 1) protease/antiprotease balance; 2) inflammation; and 3) the link of both parameters to clinical parameters of CF patients.MethodsTransforming growth factor-β1 (TGF-β1), interleukin (IL)-1β, IL-8, neutrophil elastase (NE) and elastase inhibitor elafin were measured (ELISA assays), and gene expression of the NF-κB pathway was assessed (reverse transcriptase PCR) in the sputum of 60 CF patients with a minimum age of 5 years. Spirometry was assessed according to American Thoracic Society guidelines.ResultsOur results demonstrated the following: 1) NE was markedly increased in Pa-positive sputum, whereas elafin was significantly decreased; 2) increased IL-1β/IL-8 levels were associated with both Pa infection and reduced forced expiratory volume in 1 s, and sputum TGF-β1 was elevated in Pa-infected CF patients and linked to an impaired lung function; and 3) gene expression of NF-κB signalling components was increased in sputum of Pa-infected patients, and these findings were positively correlated with IL-8.ConclusionOur study links Pa infection to an imbalance of NE and NE inhibitor elafin and increased inflammatory mediators. Moreover, our data demonstrate an association between high TGF-β1 sputum levels and a progress in chronic lung inflammation and pulmonary fibrosis in CF. Controlling the excessive airway inflammation by inhibition of NE and TGF-β1 might be promising therapeutic strategies in future CF therapy and a possible complement to cystic fibrosis transmembrane conductance regulator (CFTR) modulators.

Untapped Potential: Therapeutically Targeting Eicosanoids and Endocannabinoids in the Lung.

Inflammation of the airway involves the recruitment of highly active immune cells to combat and clear microbes and toxic factors; however, this inflammatory response can result in unintended damage to lung tissue. Tissue damage resulting from inflammation is often mitigated by resolving factors that limit the scope and duration of the inflammatory response. Both inflammatory and resolving processes require the actions of a vast array of lipid mediators that can be rapidly synthesized through a variety of airway resident and infiltrating immune cells. Eicosanoids and endocannabinoids represent two major classes of lipid mediators that share synthetic enzymes and have diverse and overlapping functions. This review seeks to provide a summary of the major bioactive eicosanoids and endocannabinoids, challenges facing researchers that study them, and their roles in modulating inflammation and resolution. With a special emphasis on cystic fibrosis, a variety of therapeutics are discussed that have been explored for their potential anti-inflammatory or proresolving impact toward alleviating excessive airway inflammation and improving lung function.

Metabolic cross‐talk between alveolar type 2 cells and resident alveolar macrophages attenuates inflammation in acute lung injury

IntroductionAcute lung injury (ALI) is an inflammatory lung disease, which manifests itself in patients as acute respiratory distress syndrome (ARDS). Our lab previously showed that hypoxia inducible transcription factor (HIF) 1A up‐regulates glycolytic capacity in AT II cells during ALI as a compensatory mechanism to dampen inflammation with the mechanism unknown. In tumor associated macrophages, lactate, the end product of glycolysis can shift macrophages to an anti‐inflammatory Arg‐1 + phenotype. We hypothesized that increased glycolysis in AT2 cells would crosstalk and shift AMs towards a pro‐resolving phenotype and thus attenuate inflammation in ALI.MethodsC57/B6 wild type or AT2 cell‐specific HIF1A knockout (SPC‐ER Hif1a loxp/loxp) animals were utilized. To induce ALI mice received either i.t. LPS or i.t. HCl (acid aspiration). 6 hours after induction of ALI mice received 25 mM Lactate i.t or pH‐controlled PBS in the control group. Resident vs. recruited AMs (CD45+ CD64+) were segregated by expression of Siglec‐F and CD11b respectively by flow cytometry. ALI severity was determined by cytokine expression and histology. Cytokine expression was measured with qPCR. A549 cells (an alveolar epithelial cell line) with stable knockdown of lactate dehydrogenase LDHA (A549 LDHA KD) or controls were co‐cultured with primary human airway macrophages (isolated from deceased donor lungs not suitable for transplantation) and stimulated with LPS.Resultsi.t Lactate significantly attenuated ALI in both models of acute lung injury in C57/B6 mice. The numbers of resident and recruited AMs were not altered by treatment with lactate 6‐hour post‐injury. However, the levels of Arg1, were increased in resident AMs in lactate treated mice. Furthermore, i.t. lactate shifted cytokine expression in murine airway macrophages towards increased IL‐10 but decreased IL‐1b and IL‐6 production. SPC‐ER Hif1a loxp/loxp animals displayed exacerbated injury and inflammation following LPS/Acid ALI. However, i.t. lactate significantly attenuated ALI phenotype in SPC‐ER Hif1a loxp/lox with increased IL‐10 and Arg1 mRNA expression.Human primary macrophages co‐cultured with A549 had significantly reduced inflammatory responses to LPS than macrophages alone. The protective phenotype of the epithelial cells on macrophages was abrogated when LDHA was genetically suppressed in A549 cells (A549 LDHA KD).ConclusionsOur data in 2 murine models of ALI and in an in vitro co‐culture model with primary human airway macrophages show that lactate produced by AT2 cells serves as a mediator to dampen macrophage inflammatory responses by shifting the macrophages towards a more anti‐inflammatory phenotype. Epithelial HIF1A is an activator of glycolysis is sufficient to prevent this shift in resident macrophages as SPC‐ER Hif1a loxp/loxp animals have worse ALI but can be “rescued” with i.t. Lactate. This suggests, that modifying the airway macrophage phenotype via metabolic crosstalk with the alveolar epithelium metabolism acts as an innate protective mechanism to dampen excessive airway inflammation in ALI.Support or Funding InformationNHLBI/NIH, NICHD/NIH and Parker B Francis FoundationFigure 1

Neutrophilic Asthma Is Associated with Increased Airway Bacterial Burden and Disordered Community Composition.

Neutrophilic asthma (NA) is an important asthma inflammatory phenotype associated with disease severity, airflow limitation, and steroid resistance, and its mechanism is still uncertain. Evidences suggest a potential role for bacteria in its pathogenesis, but, so far, this remains poorly understood. We sought to investigate airway bacterial burden, community composition, and inflammatory response in NA. Fifty-four stable asthmatics without infection were enrolled and separated into either NA group (n = 20) or non-NA group (n = 34). Subject demographics, Asthma Control Test (ACT) scores, medications, and pulmonary functions were documented. Sputum cytology, airway bacterial burden, microbial community composition, and inflammatory cytokines were assessed. The total airway bacterial burden was significantly increased in subjects with NA versus non-NA and was positively correlated with the sputum neutrophil percentage. Airway neutrophilia was associated with less airway bacterial community richness and diversity, along with a distinct community composition. In patients with NA, bacteria in phylum Proteobacteria, especially Haemophilus spp. and Moraxella spp., showed significant increases in both actual loads and relative abundances, while bacteria in phyla Firmicutes, Actinobacteria, and Saccharibacteria showed decreased relative abundances compared with non-NA. Patients with NA demonstrated higher levels of interleukin-1β (IL-1β), IL-6, IL-8, IL-12, IL-17A, and tumor necrosis factor-α (TNF-α) in sputum samples compared with non-NA. Increased bacterial burden and distinct microbiota composition were the key characters of neutrophilic phenotype in asthma, accompanied by excessive airway inflammation. Understanding the relationship between airway microbiota and neutrophilic inflammation may help in treatment and management of asthma, such as targeting airway microbiota.

The Anti-inflammatory Effect of Alpha-1 Antitrypsin in Rhinovirus-infected Human Airway Epithelial Cells.

Excessive airway inflammation is seen in chronic obstructive pulmonary disease (COPD) patients experiencing acute exacerbations, which are often associated with human rhinovirus (HRV) infection. Alpha-1 antitrypsin (A1AT) has anti-inflammatory function in endothelial cells and monocytes, but its anti-inflammatory effect has not been investigated in COPD airway epithelial cells. We determined A1AT's anti-inflammatory function in COPD airway epithelial cells and the underlying mechanisms such as the role of caspase-1. Brushed bronchial epithelial cells from COPD and normal subjects were cultured at air-liquid interface and treated with A1AT or bovine serum albumin (BSA, control) two hours prior to whole cigarette smoke (WCS) or air exposure, followed by HRV-16 infection. After 24 hours of viral infection, cell supernatants were collected for measuring IL-8, and cells were examined for caspase-1. The in vivo anti-inflammatory function of A1AT was determined by infecting mice intranasally with HRV-1B followed by aerosolized A1AT or BSA. A1AT significantly reduced WCS and HRV-16-induced IL-8 production in normal and COPD airway epithelial cells. COPD cells are less sensitive to A1AT's anti-inflammatory effect than normal cells. A1AT exerted the anti-inflammatory function in part via reducing caspase-1 in normal cells, but not in COPD cells. In mice, A1AT significantly reduced HRV-1B induced lung neutrophilic inflammation. A1AT exerts an anti-inflammatory effect in cigarette smoke-exposed and HRV-infected human airway epithelial cells, which may be related to its inhibitory effect on caspase-1 activity.

An observational study on factors influencing antibiotic use in chronic obstructive pulmonary disease at Universitas Academic Hospital, Bloemfontein

Background: Acute exacerbations of chronic obstructive pulmonary disease (AECOPD) are often precipitated by excessive airway inflammation caused by viral or bacterial infections. Current guidelines suggest prescribing antibiotics to patients with AECOPD and purulent sputum production, but this may lead to unnecessary or inappropriate antibiotic use. The aim of this study was to identify clinical and laboratory variables influencing antibiotic prescriptions of clinicians managing patients hospitalised for AECOPD.Methods: An observational study was conducted among patients hospitalised with AECOPD. Antibiotic prescriptions were compared with sputum appearance, white blood cell count, C-reactive protein (CRP) levels and sputum cultures. Treatment outcomes were assessed on days 3 and 5 after admission.Results: Thirteen patients were included in the study, from July to October 2013, at Universitas Academic Hospital, Bloemfontein. Antibiotics were prescribed in seven out of eight patients with an elevated CRP level. None of the patients with reported sputum purulence received antibiotics. White blood cell count and sputum cultures did not seem to influence antibiotic prescription habits.Conclusions: Clinicians managing patients with AECOPD do not follow guidelines that suggest prescribing antibiotics to patients presenting with purulent sputum production. Further studies on whether biomarkers such as CRP may be more acceptable as a deciding factor on which to base antibiotic prescriptions are required.

An observational study on factors influencing antibiotic use in chronic obstructive pulmonary disease at Universitas Academic Hospital, Bloemfontein

Background: Acute exacerbations of chronic obstructive pulmonary disease (AECOPD) are often precipitated by excessive airway inflammation caused by viral or bacterial infections. Current guidelines suggest prescribing antibiotics to patients with AECOPD and purulent sputum production, but this may lead to unnecessary or inappropriate antibiotic use. The aim of this study was to identify clinical and laboratory variables influencing antibiotic prescriptions of clinicians managing patients hospitalised for AECOPD.Methods: An observational study was conducted among patients hospitalised with AECOPD. Antibiotic prescriptions were compared with sputum appearance, white blood cell count, C-reactive protein (CRP) levels and sputum cultures. Treatment outcomes were assessed on days 3 and 5 after admission.Results: Thirteen patients were included in the study, from July to October 2013, at Universitas Academic Hospital, Bloemfontein. Antibiotics were prescribed in seven out of eight patients with an elevated CRP level. None of the patients with reported sputum purulence received antibiotics. White blood cell count and sputum cultures did not seem to influence antibiotic prescription habits.Conclusions: Clinicians managing patients with AECOPD do not follow guidelines that suggest prescribing antibiotics to patients presenting with purulent sputum production. Further studies on whether biomarkers such as CRP may be more acceptable as a deciding factor on which to base antibiotic prescriptions are required.

AMPK Agonists Ameliorate Sodium and Fluid Transport and Inflammation in Cystic Fibrosis Airway Epithelial Cells

The metabolic sensor AMP-activated kinase (AMPK) inhibits both the cystic fibrosis (CF) transmembrane conductance regulator (CFTR) Cl(-) channel and epithelial Na(+) channel (ENaC), and may inhibit secretion of proinflammatory cytokines in epithelia. Here we have tested in primary polarized CF and non-CF human bronchial epithelial (HBE) cells the effects of AMPK activators, metformin and 5-aminoimidazole-4-carboxamide-1-beta-D-riboside (AICAR), on various parameters that contribute to CF lung disease: ENaC-dependent short-circuit currents (I(sc)), airway surface liquid (ASL) height, and proinflammatory cytokine secretion. AMPK activation after overnight treatment with either metformin (2-5 mM) or AICAR (1 mM) substantially inhibited ENaC-dependent I(sc) in both CF and non-CF airway cultures. Live-cell confocal images acquired 60 minutes after apical addition of Texas Red-dextran-containing fluid revealed significantly greater ASL heights after AICAR and metformin treatment relative to controls, suggesting that AMPK-dependent ENaC inhibition slows apical fluid reabsorption. Both metformin and AICAR decreased secretion of various proinflammatory cytokines, both with and without prior LPS stimulation. Finally, prolonged exposure to more physiologically relevant concentrations of metformin (0.03-1 mM) inhibited ENaC currents and decreased proinflammatory cytokine levels in CF HBE cells in a dose-dependent manner. These findings suggest that novel therapies to activate AMPK in the CF airway may be beneficial by blunting excessive sodium and ASL absorption and by reducing excessive airway inflammation, which are major contributors to CF lung disease.

Chemokines and their receptors in respiratory disease: a therapeutic target for respiratory syncytial virus infection

Cell recruitment is a multistep process orchestrated by chemokines and their receptors. The chemokine/receptor system is central to many inflammatory diseases, making it a key target for therapeutic intervention. Despite complexity and redundancy within the system, effective antagonists are in development and undergoing clinical trials, for example, maraviroc, for use in HIV treatment. Respiratory syncytial virus (RSV) is a major cause of lower respiratory tract infection in infants, with global annual infection estimated at 64 million people. Current treatment is purely supportive, with no effective vaccine available. RSV pathology is partly due to excessive airway inflammation. Evidence is growing for a key role for chemokine receptors. Receptor blockade may therefore provide a feasible therapeutic option to inhibit RSV-induced inflammation and thereby reduce disease severity.

Up-regulation of AMP-activated Kinase by Dysfunctional Cystic Fibrosis Transmembrane Conductance Regulator in Cystic Fibrosis Airway Epithelial Cells Mitigates Excessive Inflammation

AMP-activated kinase (AMPK) is a ubiquitous metabolic sensor that inhibits the cystic fibrosis (CF) transmembrane conductance regulator (CFTR). To determine whether CFTR reciprocally regulates AMPK function in airway epithelia and whether such regulation is involved in lung inflammation, AMPK localization, expression, and activity and cellular metabolic profiles were compared as a function of CFTR status in CF and non-CF primary human bronchial epithelial (HBE) cells. As compared with non-CF HBE cells, CF cells had greater and more diffuse AMPK staining and had greater AMPK activity than their morphologically matched non-CF counterparts. The cellular [AMP]/[ATP] ratio was higher in undifferentiated than in differentiated non-CF cells, which correlated with AMPK activity under these conditions. However, this nucleotide ratio did not predict AMPK activity in differentiating CF cells. Inhibiting channel activity in non-CF cells did not affect AMPK activity or metabolic status, but expressing functional CFTR in CF cells reduced AMPK activity without affecting cellular [AMP]/[ATP]. Therefore, lack of functional CFTR expression and not loss of channel activity in CF cells appears to up-regulate AMPK activity in CF HBE cells, presumably through non-metabolic effects on upstream regulatory pathways. Compared with wild-type CFTR-expressing immortalized CF bronchial epithelial (CFBE) cells, DeltaF508-CFTR-expressing CFBE cells had greater AMPK activity and greater secretion of tumor necrosis factor-alpha and the interleukins IL-6 and IL-8. Further pharmacologic AMPK activation inhibited inflammatory mediator secretion in both wild type- and DeltaF508-expressing cells, suggesting that AMPK activation in CF airway cells is an adaptive response that reduces inflammation. We propose that therapies to activate AMPK in the CF airway may be beneficial in reducing excessive airway inflammation, a major cause of CF morbidity.

CCL5-CCR5 interaction provides antiapoptotic signals for macrophage survival during viral infection

Host defense against viruses probably depends on targeted death of infected host cells and then clearance of cellular corpses by macrophages. For this process to be effective, the macrophage must presumably avoid its own virus-induced death. Here we identify one such mechanism. We show that mice lacking the chemokine Ccl5 are immune compromised to the point of delayed viral clearance, excessive airway inflammation and respiratory death after mouse parainfluenza or human influenza virus infection. Virus-inducible levels of Ccl5 are required to prevent apoptosis of virus-infected mouse macrophages in vivo and mouse and human macrophages ex vivo. The protective effect of Ccl5 requires activation of the Ccr5 chemokine receptor and consequent bilateral activation of G(alphai)-PI3K-AKT and G(alphai)-MEK-ERK signaling pathways. The antiapoptotic action of chemokine signaling may therefore allow scavengers to finally stop the host cell-to-cell infectious process.

Chemokine CCL5 signals required for survival during viral infection

Rationale Inducible expression of the CCL5 (RANTES) gene is an extremely sensitive indicator of viral infection, but the functional role of CCL5 in anti-viral defense still needs to be defined. We therefore studied newly developed mice with null mutations of the CCL5 gene in a model of viral bronchiolitis/pneumonia. Methods CCL5-null and WT cells (especially T cells and macrophages) isolated from these mice were monitored after inoculation with mouse parainfluenza type I (Sendai) virus. Studies were also extended to chemokine receptor CCR5-null mice and human cells treated with anti-CCR5 antibody and infected with respiratory syncytial virus (RSV). Results CCL5- and CCR5-null mice were immune compromised to the point of delayed viral clearance, excessive airway inflammation, and respiratory death compared to control mice. Inflammation was characterized by accumulation of virus-infected, apoptotic macrophages in the airway mucosa. Isolated CCL5- and CCR5-null macrophages (as well as human macrophages treated with anti-CCR5 antibody) were also more susceptible to virus-induced apoptosis. This susceptibility was rescued by CCL5 add-back in CCL5-null mouse macrophages and was reproduced by inhibition of G-protein/PI3K/Akt or MEK/ERK1/2 signaling in mouse and human macrophages. These same Akt and ERK pathways were resistant to endogenous activation in CCL5-null (but not wild-type) mice after viral infection in vivo. Conclusions Inducible expression of CCL5 is critical for host survival during viral infection, most likely because CCL5 protects macrophages from virus-induced apoptosis and thereby allows for effective clearance of cellular corpses that would otherwise cause excessive airway inflammation and obstruction.