Airway Defense Research Articles

Lactoperoxidase-mediated degradation of single-walled carbon nanotubes in the presence of pulmonary surfactant

Carbon nanotubes (CNTs) may elicit inflammatory responses following pulmonary exposure. Conversely, enzymatic biodegradation of CNTs by inflammatory cells has also been reported. The aim of this study was to study the degradation of oxidized single-walled CNTs (ox-SWCNTs) by lactoperoxidase (LPO), a secreted peroxidase present in the airways, and whether pulmonary surfactant affects this biodegradation. To this end, ox-SWCNTs were incubated in vitro with recombinant bovine LPO+H2O2+NaSCN in the presence and absence of porcine lung surfactant (Curosurf®) and biodegradation was monitored using UV–Vis–NIR spectroscopy, Raman spectroscopy, and scanning electron microscopy. The interaction of recombinant LPO with bundles of ox-SWCNTs was confirmed by atomic force microscopy. Cell-free biodegradation of ox-SWCNTs was also observed ex vivo in murine bronchoalveolar lavage fluid in the presence of H2O2+NaSCN. Our study provides evidence for biodegradation of ox-SWCNTs with a lung surfactant ‘bio-corona’ and expands the repertoire of mammalian peroxidases capable of biodegradation of ox-SWCNTs. These findings are relevant to inhalation exposure to these materials, as LPO serves as an important component of the airway defense system.

Role of Mucin 5ac in Murine Airways

Mucin is a highly glycosylated protein in epithelial lining of respiratory, gastrointestinal, and reproductive tracts. To determine the role of Muc5ac, the major secreted airway mucin, in respiratory pathogenesis, Muc5ac‐deficient mice (Muc5ac‐/‐), generated by deletion of exons 21‐31 encoding the glycosylation region, and wild type controls (Muc5ac+/+) were examined in models of respiratory syncytial virus (RSV) disease (intranasal infection, 106 plaque forming unit RSV/mouse) and pulmonary fibrosis (intratracheal instillation, 0.5 U bleomycin/Kg body weight). Phenotypes were assessed by histopathologic and bronchoalveolar lavage analyses. Reduced levels of intraepithelial mucus from nasal to bronchial airways, atrophic maxilloturbinates, and enriched nasal glands were manifest in Muc5ac‐/‐, compared to Muc5ac+/+. In Muc5ac‐/‐, RSV caused more severe nasal airway injuries including subepithelial congestion and gland secretion and compensatory increase of mucus secretion than in Muc5ac+/+ at 1‐5 d post‐infection. Bronchoepithelial injury and hyperplasia/hyperproliferation as well as pulmonary perivascular‐peribronchiolar edema and smooth muscle thickening were more evident in Muc5ac‐/‐ relative to Muc5ac+/+ at 1‐5 d post‐infection. Serum IgE level was also significantly higher in Muc5ac‐/‐ than in Muc5ac+/+ at 5 d after infection. Muc5ac‐/‐ exhibited significantly more severe pulmonary inflammation than Muc5ac+/+ at 7‐14 d post‐bleomycin. Overall, lack of epithelial barrier component Muc5ac impaired the airway defense against virus and fibrogen, and caused compensatory regulation of other secretory proteins. Supported by the NIEHS Intramural Program.

Model of mucociliary clearance in cystic fibrosis lungs.

Mucus clearance is a primary innate defense mechanism in the human airways. Cystic fibrosis (CF) is a genetic disease caused by mutations in the gene encoding the cystic fibrosis transmembrane conductance regulator (CFTR) protein. CF is characterized by dehydration of airway surface liquid and impaired mucociliary clearance. As a result, microorganisms are not efficiently removed from the airways, and patients experience chronic pulmonary infections and inflammation. We propose a new physiologically based mathematical model of muco-ciliary transport consisting of the two major components of the mucociliary clearance system: (i) periciliary liquid layer (PCL) and (ii) mucus layer. We study mucus clearance under normal conditions and in CF patients. Restoring impaired clearance of airway secretions in one of the major goals of therapy in patients with CF. We consider the action of the aerosolized and inhaled medication dornase alfa, which reduces the viscosity of cystic fibrosis mucus, by selectively cleaving the long DNA strands it contains. The results of the model simulations stress the potential relevance of the location of the drug deposition in the central or peripheral airways. Mucus clearance was increased in case the drug was primarily deposited peripherally, i.e. in the small airways.

Aspiration-Related Pulmonary Syndromes

Aspiration of foreign matter into the airways and lungs can cause a wide spectrum of pulmonary disorders with various presentations. The type of syndrome resulting from aspiration depends on the quantity and nature of the aspirated material, the chronicity, and the host responses. Aspiration is most likely to occur in subjects with a decreased level of consciousness, compromised airway defense mechanisms, dysphagia, gastroesophageal reflux, and recurrent vomiting. These aspiration-related syndromes can be categorized into airway disorders, including vocal cord dysfunction, large airway obstruction with a foreign body, bronchiectasis, bronchoconstriction, and diffuse aspiration bronchiolitis, or parenchymal disorders, including aspiration pneumonitis, aspiration pneumonia, and exogenous lipoid pneumonia. In idiopathic pulmonary fibrosis, aspiration has been implicated in disease progression and acute exacerbation. Aspiration may increase the risk of bronchiolitis obliterans syndrome in patients who have undergone a lung transplant. Accumulating evidence suggests that a causative role for aspiration is often unsuspected in patients presenting with aspiration-related pulmonary diseases; thus, many cases go undiagnosed. Herein, we discuss the broadening spectrum of these pulmonary syndromes with a focus on presenting features and diagnostic aspects.

Polymorphisms associated with expression of BPIFA1/BPIFB1 and lung disease severity in cystic fibrosis.

BPI fold containing family A, member 1 (BPIFA1) and BPIFB1 are putative innate immune molecules expressed in the upper airways. Because of their hypothesized roles in airway defense, these molecules may contribute to lung disease severity in cystic fibrosis (CF). We interrogated BPIFA1/BPIFB1 single-nucleotide polymorphisms in data from an association study of CF modifier genes and found an association of the G allele of rs1078761 with increased lung disease severity (P = 2.71 × 10(-4)). We hypothesized that the G allele of rs1078761 is associated with decreased expression of BPIFA1 and/or BPIFB1. Genome-wide lung gene expression and genotyping data from 1,111 individuals with lung disease, including 51 patients with CF, were tested for associations between genotype and BPIFA1 and BPIFB1 gene expression levels. Findings were validated by quantitative PCR in a subset of 77 individuals. Western blotting was used to measure BPIFA1 and BPIFB1 protein levels in 93 lung and 101 saliva samples. The G allele of rs1078761 was significantly associated with decreased mRNA levels of BPIFA1 (P = 4.08 × 10(-15)) and BPIFB1 (P = 0.0314). These findings were confirmed with quantitative PCR and Western blotting. We conclude that the G allele of rs1078761 may be detrimental to lung function in CF owing to decreased levels of BPIFA1 and BPIFB1.

Immunoglobulin replacement therapy in antibody deficiency syndromes: are we really doing enough?

Immunoglobulin replacement therapy in antibody deficiency syndromes: are we really doing enough?

Effects of air pollution on respiratory health

Epidemiological studies have demonstrated that there is an association between increases in air pollution and cardiopulmonary mortality and morbidity. Multi-center studies in North America and Western European countries reported that an increase in the levels of particulate matter (PM), ozone, nitrogen oxides (NOx) and sulphur dioxide (SO2) leads to increases in prevalence, emergency room visits and hospitalization due to chronic respiratory diseases such as asthma and chronic obstructive disease (COPD). Furthermore, it has been shown that increased levels of air pollutants are associated with increased prevalence of respiratory infections, including pneumonia. Although studies of indoor air pollution clearly showed a significant association with tuberculosis (TB), studies investigating the role of outdoor air pollution are lacking. However, it has recently been reported that increased ambient levels of air pollutants such as PM≤2.5 μm and SO2 were associated with increased risk of TB. In vivo human studies demonstrated that exposure to ozone, NO2 and diesel exhaust (DE) leads to irritation in the mouth and nose, dyspnoea, wheezing, chest tightness, decreases in lung function tests, and increased airway hyper-responsiveness. There were also increased levels of neutrophils, lymphocytes and inflammatory mediators in bronchial lavage and bronchial biopsies of subjects exposed to ozone and DE. Furthermore, SO2 and NO2 increased airway response of allergic asthmatic subjects to allergens, such as house dust mite allergen. In vitro studies reported that DE particles (DEP) induce inflammatory mediator expression and synthesis of IgE specific to allergens in human B cells. Studies of human macrophages demonstrated that fine carbon black (CB) induced inflammatory cytokines and impaired phagocytosis of pneumococci and mycobacteria TB with a reduction in the oxidative burst capacity of these cells. This study has demonstrated that DEP, which constitute an important fraction of PM pollution, can decrease ciliary beat frequency (CBF), and induce the release of inflammatory mediators such as interleukin (IL)-8, granulocyte macrophage-colony stimulating factor (GM-CSF), normal T-cell expressed and secreted (RANTES) and soluble intercellular adhesion molecule (sICAM)-1 from primary bronchial epithelial cells (BECs). More recently, this study found that DEP can decrease airway epithelial cell viability and modify the cell cycle progression and apoptosis by inducing oxidative stress-related pathways, such as activator protein (AP)-1 and nuclear factor (NF)-κB, and an increased expression of apoptosis and cell cycle regulating proteins such as p21, p27, p53, cyclin E, c-myc, and cyclin-dependant kinase 2 (CDK2). Interestingly, serum, as can be seen in the inflamed airways of patients with chronic airway diseases such as asthma and COPD because of extravasation, enhanced the detrimental effects of DEP. Furthermore, these studies showed that gaseous air pollutants such as ozone and NO2 induce permeability of BEC cultures with an increase in the release of inflammatory mediators. More importantly, this study demonstrated that BECs from patients with chronic airway diseases such as asthma and COPD are more susceptible to deleterious effects of air pollutants. In conclusion, air pollutants can induce respiratory mortality and morbidity by leading to airway and lung inflammation and impairing the airway defence system against noxious agents and microorganisms such as mycobacteria TB.

Toxicity assessment of aggregated/agglomerated cerium oxide nanoparticles in an in vitro 3D airway model: The influence of mucociliary clearance

We investigated the toxicity of aggregated nanoparticles of cerium oxide (CeO2) using an in vitro 3D human bronchial epithelial model that included a mucociliary apparatus (MucilAir™). CeO2 was dispersed in saline and applied to the apical surface of the model. CeO2 did not induce distinct effects in the model, whereas it did in BEAS-2B and A549 cell cultures. The absence of effects of CeO2 was not because of the model’s insensitivity. Nanoparticles of zinc oxide (ZnO) elicited positive responses in the toxicological assays. Respiratory mucus (0.1% and 1%) added to dispersions increased aggregation/agglomeration to such an extent that most CeO2 sedimented within a few minutes. Also, the mucociliary apparatus of the model removed CeO2 from the central part of the apical surface to the borders. This ‘clearance’ may have prevented the majority of CeO2 from reaching the epithelial cells. Chemical analysis of cerium in the basal tissue culture medium showed only minimal translocation of cerium across the 3D barrier. In conclusion, mucociliary defence appeared to prevent CeO2 reaching the respiratory epithelial cells in this 3D in vitro model. This model and approach can be used to study compounds of specific toxicological concern in airway defence mechanisms in vitro.

Deprived TLR9 expression in apparently healthy nasal mucosa might trigger polyp-growth in chronic rhinosinusitis patients.

BackgroundThe origin of nasal polyps in chronic rhinosinusitis is unknown, but the role of viral infections in polyp growth is clinically well established. Toll-like receptors (TLRs) have recently emerged as key players in our local airway defense against microbes. Among these, TLR9 has gained special interest in viral diseases. Many studies on chronic rhinosinusitis with nasal polyps (CRSwNP) compare polyp tissue with nasal mucosa from polyp-free individuals. Knowledge about changes in the turbinate tissue bordering the polyp tissue is limited.ObjectivesTo analyse the role of TLR9 mediated microbial defense in tissue bordering the polyp.MethodsNasal polyps and turbinate tissue from 11 patients with CRSwNP and turbinate tissue from 11 healthy controls in total were used. Five biopsies from either group were analysed immediately with flow cytometry regarding receptor expression and 6 biopsies were used for in vitro stimulation with a TLR9 agonist, CpG. Cytokine release was analysed using Luminex. Eight patients with CRSwNP in total were intranasally challenged with CpG/placebo 24 hours before surgery and the biopsies were collected and analysed as above.ResultsTLR9 expression was detected on turbinate epithelial cells from healthy controls and polyp epithelial cells from patients, whereas TLR9 was absent in turbinate epithelial cells from patients. CpG stimulation increased the percentage cells expressing TLR9 and decreased percentage cells expressing VEGFR2 in turbinate tissue from patients. After CpG stimulation the elevated levels of IL-6, G-CSF and MIP-1β in the turbinate tissue from patients were reduced towards the levels demonstrated in healthy controls.ConclusionDefects in the TLR9 mediated microbial defense in the mucosa adjacent to the anatomic origin of the polyp might explain virus induced polyp growth. CpG stimulation decreased VEGFR2, suggesting a role for CpG in polyp formation. The focus on turbinate tissue in patients with CRSwNP opens new perspectives in CRSwNP-research.

Potassium ion channels and allergic asthma.

High-conductive calcium-sensitive potassium channels (BK+Ca) and ATP-sensitive potassium (K+ATP) channels play a significant role in the airway smooth muscle cell and goblet cell function, and cytokine production. The present study evaluated the therapeutic potential of BK+Ca and K+ATP openers, NS 1619 and pinacidil, respectively, in an experimental model of allergic inflammation. Airway allergic inflammation was induced with ovalbumine in guinea pigs during 21 days, which was followed by a 14-day treatment with BK+Ca and K+ATP openers. The outcome measures were airway smooth muscle cells reactivity in vivo and in vitro, cilia beating frequency and the level of exhaled NO (ENO), and the level of pro-inflammatory cytokines in the plasma and bronchoalveolar lavage fluid. The openers of both channels decreased airway smooth muscle cells reactivity, cilia beating frequency, and cytokine levels in the serum. Furthermore, NS1619 reduced ENO and inflammatory cells infiltration. The findings confirmed the presence of beneficial effects of BK+Ca and K+ATP openers on airway defence mechanisms. Although both openers dampened pro-inflammatory cytokines and mast cells infiltration, an evident anti-inflammatory effect was provided only by NS1619. Therefore, we conclude that particularly BK+Ca channels represent a promising new drug target in treatment of airway's allergic inflammation.

House Dust Mite Allergy Models—Reliability for Research of Airway Defensive Mechanisms

Extensive research of respiratory physiology and diseases of the airways and lungs provides better understanding of mechanisms of diseases and it is a source of new data with potential clinical application. The majority of the data in the field of airway defensive reflexes were obtained from guinea pig model, as the most convenient one. Neurophysiology and neuropharma-cology of the cough reflex—which is exclusively mediated by the vagus nerve, share close similarities between guinea pigs and humans. The models used to study pathological processes and their influence on airway-defensive reflexes use sensitization with ovalbumin—the protein from chicken egg, which does not mimic allergies within human respiratory system. House dust mites (HDM) (Dermatophagoides pteronyssius and Dermatophagoides farinae) represent frequent human aeroallergens, however the HDM models are used considerably less than ovalbumin models. The primary objective of this review is to focus on already validated models of HDM-induced airway diseases to see, whether some of them are suitable to study mechanisms of peripheral cough plasticity in a condition of HDM-induced pathological processes. The main purpose of future use and validation of HDM model is to produce higher ability to translate the results obtained in animal models to human cough research.

Host Microarray Molecular Signature and Serologic Evaluation of Stages of Pseudomonas aeruginosa Infection in Cystic Fibrosis

Background: Cystic fibrosis (CF) is a genetic disorder caused by mutations in the CF transmembrane regulator gene (CFTR); lung disease is characterized by chronic infection by Pseudomonas aeruginosa, the major cause of morbidity and mortality in patients with CF. However, oropharyngeal cultures for P. aeruginosa in young children result in poor specificity for lower airway infection. We used a novel approach to generate genomewide expression profiles to identify markers for P. aeruginosa infection and CF disease severity. Transcription was induced in healthy unrelated peripheral blood mononuclear cells through coculture with autologous serum (self-baseline control), allogeneic serums (healthy unrelated controls), and serums from patients with CF. Methodology: All CF diagnoses included sweat chloride levels greater than or equal to 60 mEq/L and/or two CFTR mutations. We performed various ELISAs against 192 serum samples (43 individuals sampled over 15 years) and comparative cultures. Concurrently, we completed expression profiling of RNA (HU133 plus 2.0 array; Affymetrix, Santa Clara, CA) from coculture of peripheral blood mononuclear cells with the serum of patients with CF, who possessed increasing degrees of colonization with P. aeruginosa. Differentially expressed probe sets were defined as those possessing a false discovery rate less than 10% and |log2 ratio| greater than 0.5 among comparison groups. Results: Our ELISA data indicated that the host response to P. aeruginosa infection increased over time, suggesting that serology to P. aeruginosa may help determine initial airway pathology. Comparison of array signatures performed before and after seroconversion identified several hundred unique genes; the most significant differences in gene expression included host genes involved with airway defense, antigen presentation, and transcription. Conclusions: Our methodology consistently identified a small number of genes that were unique to patients with CF or to patients with CF with initial P. aeruginosa infection conversion. These genes merit investigation as active participants in the function of the lung microbiome and may serve as therapeutic targets for P. aeruginosa infection in CF.

Muc5b is required for airway defence

Respiratory surfaces are exposed to billions of particulates and pathogens daily. A protective mucus barrier traps and eliminates them through mucociliary clearance (MCC). However, excessive mucus contributes to transient respiratory infections and to the pathogenesis of numerous respiratory diseases. MUC5AC and MUC5B are evolutionarily conserved genes that encode structurally related mucin glycoproteins, the principal macromolecules in airway mucus. Genetic variants are linked to diverse lung diseases, but specific roles for MUC5AC and MUC5B in MCC, and the lasting effects of their inhibition, are unknown. Here we show that mouse Muc5b (but not Muc5ac) is required for MCC, for controlling infections in the airways and middle ear, and for maintaining immune homeostasis in mouse lungs, whereas Muc5ac is dispensable. Muc5b deficiency caused materials to accumulate in upper and lower airways. This defect led to chronic infection by multiple bacterial species, including Staphylococcus aureus, and to inflammation that failed to resolve normally. Apoptotic macrophages accumulated, phagocytosis was impaired, and interleukin-23 (IL-23) production was reduced in Muc5b(-/-) mice. By contrast, in mice that transgenically overexpress Muc5b, macrophage functions improved. Existing dogma defines mucous phenotypes in asthma and chronic obstructive pulmonary disease (COPD) as driven by increased MUC5AC, with MUC5B levels either unaffected or increased in expectorated sputum. However, in many patients, MUC5B production at airway surfaces decreases by as much as 90%. By distinguishing a specific role for Muc5b in MCC, and by determining its impact on bacterial infections and inflammation in mice, our results provide a refined framework for designing targeted therapies to control mucin secretion and restore MCC.

Aqueous cigarette smoke extract induces a voltage-dependent inhibition of CFTR expressed in Xenopus oocytes

The cystic fibrosis transmembrane conductance regulator (CFTR) chloride channel inhabits the apical membrane of airway epithelia, where its function is essential for mucus hydration, mucociliary clearance, and airway defense. Chronic obstructive pulmonary disease (COPD), most often a consequence of cigarette smoke (CS) exposure, affects 15 million persons in the US. Clinically, COPD is characterized by many of the salient features of cystic fibrosis lung disease, where CFTR is either absent or reduced in function. CS is an acidic aerosol (pH 5.3 to 6.3) reported to contain over 4,000 constituents. Acute CS exposure has been reported to decrease airway transepithelial voltage in vivo and short-circuit current in vitro; however, the mechanistic basis of these effects is uncertain. The goal of the studies described here was to develop a bioassay to characterize the effects of aqueous CS preparations on the channel function of CFTR. We studied aqueous CS extract (CSE) prepared in our laboratory, as well as commercial cigarette smoke condensate (CSC) in Xenopus oocytes expressing human CFTR. Application of CSE at pH 5.3 produced a reversible, voltage-dependent inhibition of CFTR conductance. CSE neutralized to pH 7.3 produced less inhibition of CFTR conductance. Serial dilution of CSE revealed a dose-dependent effect at acidic and neutral pH. In contrast, CSC did not inhibit CFTR conductance in oocytes. We conclude that one or more components of CSE inhibits CFTR in a manner similar to diphenylamine-2-carboxylate, a negatively charged, open-channel blocker.

Reduced Nasal Nitric Oxide Production in Cystic Fibrosis Patients with Elevated Systemic Inflammation Markers

BackgroundNitric oxide (NO) is produced within the respiratory tract and can be detected in exhaled bronchial and nasal air. The concentration varies in specific diseases, being elevated in patients with asthma and bronchiectasis, but decreased in primary ciliary dyskinesia. In cystic fibrosis (CF), conflicting data exist on NO levels, which are reported unexplained as either decreased or normal. Functionally, NO production in the paranasal sinuses is considered as a location-specific first-line defence mechanism. The aim of this study was to investigate the correlation between upper and lower airway NO levels and blood inflammatory parameters, CF-pathogen colonisation, and clinical data.Methods and FindingsNasal and bronchial NO concentrations from 57 CF patients were determined using an electrochemical analyser and correlated to pathogen colonisation of the upper and lower airways which were microbiologically assessed from nasal lavage and sputum samples. Statistical analyses were performed with respect to clinical parameters (lung function, BMI), laboratory findings (CRP, leucocytes, total-IgG, fibrinogen), and anti-inflammatory and antibiotic therapy. There were significant correlations between nasal and bronchial NO levels (rho = 0.48, p<0.001), but no correlation between NO levels and specific pathogen colonisation. In patients receiving azithromycin, significantly reduced bronchial NO and a tendency to reduced nasal NO could be found. Interestingly, a significant inverse correlation of nasal NO to CRP (rho = −0.28, p = 0.04) and to leucocytes (rho = −0.41, p = 0.003) was observed. In contrast, bronchial NO levels showed no correlation to clinical or inflammatory parameters.ConclusionGiven that NO in the paranasal sinuses is part of the first-line defence mechanism against pathogens, our finding of reduced nasal NO in CF patients with elevated systemic inflammatory markers indicates impaired upper airway defence. This may facilitate further pathogen acquisition in the sinonasal area, with consequences for lung colonisation and the overall outcome in CF.

Vasoactive intestinal peptide regulates sinonasal mucociliary clearance and synergizes with histamine in stimulating sinonasal fluid secretion

Mucociliary clearance (MCC) is the primary physical airway defense against inhaled pathogens and particulates. MCC depends on both proper fluid/mucus homeostasis and epithelial ciliary beating. Vasoactive intestinal peptide (VIP) is a neurotransmitter expressed in the sinonasal epithelium that is up-regulated in allergy. However, the effects of VIP on human sinonasal physiology are unknown, as are VIP's interactions with histamine, a major regulator of allergic disease. We imaged ciliary beat frequency, mucociliary transport, apical Cl(-) permeability, and airway surface liquid (ASL) height in primary human sinonasal air-liquid-interface cultures to investigate the effects of VIP and histamine. VIP stimulated an increase in ciliary beat frequency (EC50 0.5 μM; maximal increase ∼40% compared with control) and cystic fibrosis transmembrane conductance regulator (CFTR)-dependent and Na(+)K(+)2Cl(-) cotransporter-dependent fluid secretion, all requiring cAMP/PKA signaling. Histamine activated Ca(2+) signaling that increased ASL height but not ciliary beating. Low concentrations of VIP and histamine had synergistic effects on CFTR-dependent fluid secretion, revealed by increased ASL heights. An up-regulation of VIP in histamine-driven allergic rhinitis would likely enhance mucosal fluid secretion and contribute to allergic rhinorrhea. Conversely, a loss of VIP-activated secretion in patients with CF may impair mucociliary transport, contributing to increased incidences of sinonasal infections and rhinosinusitis.

Gene expression of SOAT1, a key enzyme for cholesteryl ester biosynthesis, is regulated by LPS and correlates with the antibacterial activity of human bronchial epithelial cell secretions against Pseudomonas aeruginosa (P1249)

Abstract Airway epithelial cells contribute to the first line of mucosal defense by secretion of antimicrobial peptides and antimicrobial lipids, including the cholesteryl ester (CE) cholesteryl arachidonate. Sterol O-acyltransferase 1 (SOAT1) is a key enzyme for CE biosynthesis. However, little is known about the regulation of the lipid-mediated arm of innate host defense. Polarized human bronchial epithelial cells (HBE) were incubated for 6 h with 100 ng/mL LPS, 10 μg/mL peptidoglycan (PG), 25 μg/mL PamCys3, and solvent control. SOAT1 mRNA was quantified by qRT-PCR with duplexing and RPLP0α as housekeeping gene. Lipid content of apical HBE secretions was analyzed by rpHPLC. SOAT1 expression was blocked with siRNA/lipofection. Antibacterial activity of HBE secretions was assessed by a 3 h colony forming unit assay with Pseudomonas aeruginosa (PA). SOAT1 gene expression was significantly increased by LPS (14.6 ±11.4, means ± SD, n=4, p=0.037). Stimulation with PG and PamCys3 increased SOAT1 expression by ~ 4- and 2-fold, respectively. Secretions from LPS-stimulated HBE showed an increase of total lipids including CA by ~30-50%. Selective inhibition of SOAT1 mRNA expression by 80% reduced the antibacterial activity of HBE whereby PA showed enhanced bacterial growth reaching &amp;gt;170% of the controls. These data suggest that SOAT1 is regulated by conserved innate immunity pathways and further support the role of antimicrobial lipids as effector molecules in mucosal airway defense.

Interactions of mechanically induced coughing and sneezing

Mutual interactions of cough and sneeze were studied on 11 spontaneously breathing pentobarbitone anesthetized cats (3.5±0.2 kg). Reflexes were induced mechanically in the nose and tracheobronchial airways, respectively. The amplitude of styloglossus muscle electromyogram (EMG) moving average during the expulsive phase of sneeze was about 16‐fold higher than that during cough (p&lt;0.01; 9 cats). Higher inspiratory efforts occurred during coughing (p&lt;0.01) vs. those in sneeze. Mechanical nasal stimulation (when no sneeze was evoked) had no significant effect on cough (7 cats). In the combined trials (both cough and sneeze were executed; 7 cats), the responses were classified as either sneeze or cough (no hybrid responses occurred) and they were potentiated comparing to control reflexes. Higher expiratory maxima (esophageal pressure and the amplitudes of abdominal muscles EMG) compared to control coughs and sneezes were found (p&lt;0.05). In sneeze during combined trials also inspiratory efforts and the number of sneezes were increased (p&lt;0.05). We conclude that mechanosensitive nasal afferents have limited effect on the tracheobronchial cough response. However, co‐expression of coughing and sneezing results in a variety of enhanced cough and sneeze responses supposedly representing improved airway defense.“This work was supported by the Slovak Research and Development Agency under the contract No. APVV‐0189–11“ The project is co‐financed from EU sources“ – ERDF – European Regional Developmental Fund, and by VEGA 1/0126/12

THE ROLE OF LIPOXIN A4 IN CYSTIC FIBROSIS LUNG DISEASE

In Cystic Fibrosis (CF), mutations of the CFTR gene result in defective Cl− secretion and Na+ hyperabsorption by epithelia which leads to airway lumen dehydration and mucus plugging and favours chronic bacterial colonization, persistent inflammation and progressive lung destruction. Beyond this general description, the pathogenesis of CF lung disease remains obscure due to an incomplete understanding of normal innate airway defense. This mini-review aims to highlight the role of the pro-resolution lipid mediator, Lipoxin A4, which is inadequately produced in CF, on several aspects of innate immunity that are altered in CF airway disease.

Ultrastructural Comparison of the Nasal Epithelia of Healthy and Naturally Affected Rabbits with Pasteurella multocida A.

An ultrastructural comparison between the nasal cavities of healthy rabbits and those suffering from two forms of spontaneous infection with Pasteurella multocida was undertaken. Twelve commercially produced rabbits of different ages and respiratory health status were divided into four groups: healthy from 0 to 21 days (G1, n = 2); healthy from 23 to 49 days (G2, n = 2); healthy from 51 to 69 days (G3, n = 2); diseased rabbits with septicemia and the rhinitic form of P. multocida infection (G4, n = 3). The main ultrastructural changes observed were a widening of the interepithelial spaces, increased activity and number of goblet cells, the formation of two types of vacuoles in epithelial cells, the degranulation and migration of heterophils between the epithelial cells, and the association of this migration with some of the other changes. No bacteria were observed adhering to the epithelium, and very few were observed free in the mucus. Scant inter-epithelial spaces were found in healthy rabbits, but they were not as large and numerous as those found in diseased animals. We discuss the origin and meaning of these changes but, we focus on the significance of the inter-epithelial spaces and goblet cells for the defense of the upper respiratory airways against the bacterium and its lipopolysaccharide.