Pathogenesis Of Chronic Airway Diseases Research Articles

Impact of particulate air pollution on airway injury and epithelial plasticity; underlying mechanisms.

Air pollution plays an important role in the mortality and morbidity of chronic airway diseases, such as asthma and chronic obstructive pulmonary disease (COPD). Particulate matter (PM) is a significant fraction of air pollutants, and studies have demonstrated that it can cause airway inflammation and injury. The airway epithelium forms the first barrier of defense against inhaled toxicants, such as PM. Airway epithelial cells clear airways from inhaled irritants and orchestrate the inflammatory response of airways to these irritants by secreting various lipid mediators, growth factors, chemokines, and cytokines. Studies suggest that PM plays an important role in the pathogenesis of chronic airway diseases by impairing mucociliary function, deteriorating epithelial barrier integrity, and inducing the production of inflammatory mediators while modulating the proliferation and death of airway epithelial cells. Furthermore, PM can modulate epithelial plasticity and airway remodeling, which play central roles in asthma and COPD. This review focuses on the effects of PM on airway injury and epithelial plasticity, and the underlying mechanisms involving mucociliary activity, epithelial barrier function, airway inflammation, epithelial-mesenchymal transition, mesenchymal-epithelial transition, and airway remodeling.

Vapors produced by electronic cigarettes and e-juices with flavorings induce toxicity, oxidative stress, and inflammatory response in lung epithelial cells and in mouse lung.

Oxidative stress and inflammatory response are the key events in the pathogenesis of chronic airway diseases. The consumption of electronic cigarettes (e-cigs) with a variety of e-liquids/e-juices is alarmingly increasing without the unrealized potential harmful health effects. We hypothesized that electronic nicotine delivery systems (ENDS)/e-cigs pose health concerns due to oxidative toxicity and inflammatory response in lung cells exposed to their aerosols. The aerosols produced by vaporizing ENDS e-liquids exhibit oxidant reactivity suggesting oxidants or reactive oxygen species (OX/ROS) may be inhaled directly into the lung during a “vaping” session. These OX/ROS are generated through activation of the heating element which is affected by heating element status (new versus used), and occurs during the process of e-liquid vaporization. Unvaporized e-liquids were oxidative in a manner dependent on flavor additives, while flavors containing sweet or fruit flavors were stronger oxidizers than tobacco flavors. In light of OX/ROS generated in ENDS e-liquids and aerosols, the effects of ENDS aerosols on tissues and cells of the lung were measured. Exposure of human airway epithelial cells (H292) in an air-liquid interface to ENDS aerosols from a popular device resulted in increased secretion of inflammatory cytokines, such as IL-6 and IL-8. Furthermore, human lung fibroblasts exhibited stress and morphological change in response to treatment with ENDS/e-liquids. These cells also secrete increased IL-8 in response to a cinnamon flavored e-liquid and are susceptible to loss of cell viability by ENDS e-liquids. Finally, exposure of wild type C57BL/6J mice to aerosols produced from a popular e-cig increase pro-inflammatory cytokines and diminished lung glutathione levels which are critical in maintaining cellular redox balance. Thus, exposure to e-cig aerosols/juices incurs measurable oxidative and inflammatory responses in lung cells and tissues that could lead to unrealized health consequences.

The role of airway epithelial cells and innate immune cells in chronic respiratory disease.

An abnormal immune response to environmental agents is generally thought to be responsible for causing chronic respiratory diseases, such as asthma and chronic obstructive pulmonary disease (COPD). Based on studies of experimental models and human subjects, there is increasing evidence that the response of the innate immune system is crucial for the development of this type of airway disease. Airway epithelial cells and innate immune cells represent key components of the pathogenesis of chronic airway disease and are emerging targets for new therapies. In this Review, we summarize the innate immune mechanisms by which airway epithelial cells and innate immune cells regulate the development of chronic respiratory diseases. We also explain how these pathways are being targeted in the clinic to treat patients with these diseases.

Angiogenesis and Vascular Remodeling in Chronic Airway Diseases

Asthma and chronic obstructive pulmonary disease remain a global health problem, with increasing morbidity and mortality. Despite differences in the causal agents, both diseases exhibit various degrees of inflammatory changes, structural alterations of the airways leading to airflow limitation. The existence of transient disease phenotypes which overlap both diseases and which progressively decline the lung function has complicated the search for an effective therapy. Important characteristics of chronic airway diseases include airway and vascular remodeling, of which the molecular mechanisms are complex and poorly understood. Recently, we and others have shown that airway smooth muscle (ASM) cells are not only structural and contractile components of airways, rather they bear capabilities of producing large number of pro-inflammatory and mitogenic factors. Increase in size and number of blood vessels both inside and outside the smooth muscle layer as well as hyperemia of bronchial vasculature are contributing factors in airway wall remodeling in patients with chronic airway diseases, proposing for the ongoing mechanisms like angiogenesis and vascular dilatation. We believe that vascular changes directly add to the airway narrowing and hyper-responsiveness by exudation and transudation of proinflammatory mediators, cytokines and growth factors; facilitating trafficking of inflammatory cells; causing oedema of the airway wall and promoting ASM accumulation. One of the key regulators of angiogenesis, vascular endothelial growth factor in concerted action with other endothelial mitogens play pivotal role in regulating bronchial angiogenesis. In this review article we address recent advances in pulmonary angiogenesis and remodelling that contribute in the pathogenesis of chronic airway diseases.

Cigarette smoke-induced differential regulation of glutathione metabolism in bronchial epithelial cells is balanced by an antioxidant tetrapeptide UPF1

Airway epithelium is a principal target for inhaled oxidants like cigarette smoke, which induce epithelial injury and thereby provoke pathogenesis of chronic airway diseases. Alterations in airway epithelial glutathione (GSH) metabolism are central in causing a loss of reducing environment, however, data are scarce on epithelial cells from larger bronchi. We showed a transient depletion of intracellular GSH in human bronchial epithelial cells after exposure to cigarette smoke condensate (CSC), which later followed by a prolonged elevation. Of the GSH-regulating enzymes, CSC increased mRNA expression of both catalytic (GCLC) and modifier (GCLM) subunits of glutamate-cysteine ligase. UPF1, a tetrapeptide GSH analogue, 4-methoxy-L-tyrosinyl-γ-L-glutamyl-L-cysteinyl-glycine, known to possess a 50-fold higher hydroxyl radical scavenging efficiency than does GSH, normalized the intracellular GSH level in the human bronchial epithelial cells under oxidative stress caused by CSC. UPF1 restored the GCLM and GSH reductase mRNA levels, which were significantly augmented by CSC treatment, back to the level of untreated control cells, referring to a successful establishment of control by UPF1 upon the over-accumulation of GSH. Moreover, UPF1 showed a significantly more potent antioxidant capacity than did N-acetyl-L-cysteine (NAC) and, compared to NAC, demonstrated a better potential to assure the whole GSH homeostasis in human bronchial epithelial cells. The current study suggests that UPF1 is capable of maintaining intracellular GSH level under CSC-induced oxidative stress in bronchial epithelial cells via balanced control over GSH-regulating enzymes, reflecting an improved perception of cellular redox conditions and thereby warranting improved adjustment of GSH accumulation.

The Rac1/JNK pathway is critical for EGFR-dependent barrier formation in human airway epithelial cells

The airway epithelial barrier provides defenses against inhaled antigens and pathogens, and alterations of epithelial barrier function have been proposed to play a significant role in the pathogenesis of chronic airway diseases. Although the epidermal growth factor receptor (EGFR) plays roles in various physiological and pathological processes on the airway epithelium, the role of EGFR on barrier function in the airway remains largely unknown. In the present study, we assessed the effects of EGFR activation on paracellular permeability in airway epithelial cells (AECs). EGFR activation induced by the addition of EGF increased transepithelial electrical resistance (TER) in AECs. An EGFR-blocking antibody eradicated the development of TER, paracellular influx of dextran, and spatial organization of tight junction. Moreover, the effects of EGFR activation on paracellular permeability were eradicated by knockdown of occludin. To identify the EGFR signaling pathway that regulates permeability barrier development, we investigated the effects of several MAP kinase inhibitors on permeability barrier function. Pretreatment with a JNK-specific inhibitor, but not an ERK- or p38-specific inhibitor, attenuated the development of TER induced by EGFR activation. Rac1 is one of the upstream activators for JNK in EGFR signaling. Rac1 knockdown attenuated the phosphorylation of JNK activation and EGFR-mediated TER development. These results suggest that EGFR positively regulates permeability barrier development through the Rac1/JNK-dependent pathway.

CRTH2 mediates the activation of human Th2 cells in response to PGD2 released from IgE/anti‐IgE treated nasal polyp tissue

Mast cells release mediators upon stimulation that contribute to the pathogenesis of chronic airway disease, including the recruitment and activation of Th2 lymphocytes. The objective was to determine the involvement of prostaglandin D(2) (PGD(2)) and its receptors in the chemotaxis of Th2 cells, using nasal polyp tissue. Tissue explants from ten patients with nasal polyposis were incubated with RPMI alone or RPMI containing IgE/anti-IgE for 30 min. Some samples were treated with diclofenac to inhibit the production of PGD(2). Supernatants were assayed for PGD(2) content and for their ability to promote human Th2 cell chemotaxis in the presence and absence of a CRTH2 antagonist. Transcript levels of D protanoid receptor type 1 (DP(1)), chemoattractant receptor-homologous receptor expressed on Th2 cells (CRTH2) and PGD(2) synthase were analysed by real time PCR. Increased release of PGD(2) by nasal polyp tissue treated with IgE/anti-IgE was significantly inhibited by preincubation of the tissue with diclofenac. Transcript levels of PGD(2) synthase, DP(1) and CRTH2 receptors increased after stimulation with IgE/anti-IgE. Supernatants from IgE/anti-IgE-stimulated nasal polyp tissue caused significantly increased chemotaxis of Th2 cells. The levels of PGD(2) produced and the degree of Th2 cell chemotaxis were highly correlated. Diclofenac inhibited the production of Th2 cell chemotactic activity, and the chemotactic effect of the supernatant on Th2 cells was inhibited by the CRTH2 antagonist ramatroban. These data suggest that in immunologically activated nasal polyp tissue, PGD(2) produced by mast cells promotes the migration of Th2 cells through a CRTH2 dependent mechanism.

Increased nasal epithelial ciliary beat frequency associated with lifestyle tobacco smoke exposure

The ciliated epithelium of the respiratory airways is one of the first vital systemic surfaces in contact with the ambient air. Ex vivo nasal epithelial ciliary beat frequency (CBF) at room temperature is on the order of 7–8 Hz but may be stimulated by irritant exposure. The upregulation of CBF in response to acute irritant exposure is generally considered to be a transient event with eventual return to baseline. However, studies of CBF dynamics in response to typical lifestyle exposures are limited. This study assessed nasal epithelial CBF among human subjects as a function of quantifiable lifestyle tobacco smoke exposure. Nasal epithelial biopsies were obtained from human subjects with well documented histories of tobacco smoke exposure. CBF was determined using a digital photometric technique and concurrent assays of nasal nitric oxide and urine cotinine and creatinine were performed. Mean CBF among active smokers and non-smokers exposed to environmental tobacco smoke (ETS) was elevated over non-smokers. Although there were dramatic differences in relative levels of tobacco smoke exposure, CBF values among tobacco smoke-exposed groups were comparable. Parallel in vitro studies of cultured nasal epithelium exposed to cigarette smoke condensate further supported these observations. These studies suggest that persistent elevation in nasal epithelial CBF is an early, subtle, physiologic effect associated with lifestyle tobacco smoke exposure. The molecular mechanisms that upregulate CBF may also create a cell molecular milieu capable of provoking the eventual emergence of more overt adverse health effects and the pathogenesis of chronic airway disease.

Regulation of inflammation by airway smooth muscle

Inflammatory mediators play a critical role in the pathogenesis of chronic airway diseases and facilitate the recruitment, activation, and trafficking of inflammatory cells in the airways. Compelling evidence now shows that airway smooth muscle expresses adhesion molecules and secretes inflammatory mediators. Airway myocytes also express a repertoire of immunomodulatory proteins such as Toll-like receptors, chemokines, and cytokines. The underlying mechanisms by which these molecules modulate airway inflammation and the physiological consequences of these molecules are now being elucidated, suggesting that airway smooth muscle plays an important role in orchestrating and perpetuating airway inflammation, remodeling, and fibrosis in chronic airway diseases.

Restrained whole body plethysmography for measure of strain-specific and allergen-induced airway responsiveness in conscious mice

The mouse is the most extensively studied animal species in respiratory research, yet the technologies available to assess airway function in conscious mice are not universally accepted. We hypothesized that whole body plethysmography employing noninvasive restraint (RWBP) could be used to quantify specific airway resistance (sRaw-RWBP) and airway responsiveness in conscious mice. Methacholine responses were compared using sRaw-RWBP vs. airway resistance by the forced oscillation technique (Raw-FOT) in groups of C57, A/J, and BALB/c mice. sRaw-RWBP was also compared with sRaw derived from double chamber plethysmography (sRaw-DCP) in BALB/c. Finally, airway responsiveness following allergen challenge in BALB/c was measured using RWBP. sRaw-RWBP in C57, A/J, and BALB/c mice was 0.51 +/- 0.03, 0.68 +/- 0.03, and 0.63 +/- 0.05 cm/s, respectively. sRaw derived from Raw-FOT and functional residual capacity (Raw*functional residual capacity) was 0.095 cm/s, approximately one-fifth of sRaw-RWBP in C57 mice. The intra- and interanimal coefficients of variations were similar between sRaw-RWBP (6.8 and 20.1%) and Raw-FOT (3.4 and 20.1%, respectively). The order of airway responsiveness employing sRaw-RWBP was AJ > BALBc > C57 and for Raw-FOT was AJ > BALB/c = C57. There was no difference between the airway responsiveness assessed by RWBP vs. DCP; however, baseline sRaw-RWBP was significantly lower than sRaw-DCP. Allergen challenge caused a progressive decrease in the provocative concentration of methacholine that increased sRaw to 175% postsaline values based on sRaw-RWBP. In conclusion, the technique of RWBP was rapid, reproducible, and easy to perform. Airway responsiveness measured using RWBP, DCP, and FOT was equivalent. Allergen responses could be followed longitudinally, which may provide greater insight into the pathogenesis of chronic airway disease.

Commentary on “Restrained whole body plethysmography for measure of strain-specific and allergen-induced airway responsiveness in conscious mice”

The mouse is the most extensively studied animal species in respiratory research, yet the technologies available to assess airway function in conscious mice are not universally accepted. We hypothesized that whole body plethysmography employing noninvasive restraint (RWBP) could be used to quantify specific airway resistance (sRaw-RWBP) and airway responsiveness in conscious mice. Methacholine responses were compared using sRaw-RWBP vs. airway resistance by the forced oscillation technique (Raw-FOT) in groups of C57, A/J, and BALB/c mice. sRaw-RWBP was also compared with sRaw derived from double chamber plethysmography (sRaw-DCP) in BALB/c. Finally, airway responsiveness following allergen challenge in BALB/c was measured using RWBP. sRaw-RWBP in C57, A/J, and BALB/c mice was 0.51 ± 0.03, 0.68 ± 0.03, and 0.63 ± 0.05 cm/s, respectively. sRaw derived from Raw-FOT and functional residual capacity (Raw*functional residual capacity) was 0.095 cm/s, approximately one-fifth of sRaw-RWBP in C57 mice. The intra- and interanimal coefficients of variations were similar between sRaw-RWBP (6.8 and 20.1%) and Raw-FOT (3.4 and 20.1%, respectively). The order of airway responsiveness employing sRaw-RWBP was AJ > BALBc > C57 and for Raw-FOT was AJ > BALB/c = C57. There was no difference between the airway responsiveness assessed by RWBP vs. DCP; however, baseline sRaw-RWBP was significantly lower than sRaw-DCP. Allergen challenge caused a progressive decrease in the provocative concentration of methacholine that increased sRaw to 175% postsaline values based on sRaw-RWBP. In conclusion, the technique of RWBP was rapid, reproducible, and easy to perform. Airway responsiveness measured using RWBP, DCP, and FOT was equivalent. Allergen responses could be followed longitudinally, which may provide greater insight into the pathogenesis of chronic airway disease.

Nitrite reductase from Pseudomonas aeruginosa induces inflammatory cytokines in cultured respiratory cells.

Persistent infection with Pseudomonas aeruginosa increases interleukin-8 (IL-8) levels and causes dense neutrophil infiltrations in the airway of patients with chronic airway diseases. To investigate the role of P. aeruginosa infection in IL-8 production in the airway of these patients, we examined whether cell lysates of P. aeruginosa could cause IL-8 production from human bronchial epithelial cells. Diluted sonicated supernatants of P. aeruginosa (SSPA) with a mucoid or nonmucoid phenotype stimulated human bronchial epithelial (BET-1A) cells to produce IL-8. In this study, we have purified a 59-kDa heat-stable protein with IL-8-inducing activity from the SSPA by sequential ion-exchange chromatography. The N-terminal sequence of this purified protein completely matched a sequence at the N-terminal part of the mature protein of nitrite reductase from P. aeruginosa. In addition, immunoblotting with a polyclonal immunoglobulin G (IgG) against recombinant Pseudomonas nitrite reductase demonstrated a specific binding to the purified protein. Furthermore, the immunoprecipitates of the SSPA with a polyclonal IgG against recombinant nitrite reductase induced a twofold-higher IL-8 production in the BET-1A cell culture than did the immunoprecipitates of the SSPA with a control IgG. These lines of evidence confirmed that Pseudomonas nitrite reductase was responsible for IL-8 production in the BET-1A cells. The purified nitrite reductase induced maximal expression of IL-8 mRNA in the BET-1A cells at 1 to 3 h after stimulation, and the IL-8 mRNA expression declined by 8 h after stimulation. New protein translation was not required for nitrite reductase-mediated IL-8 mRNA expression in the BET-1A cells. Nitrite reductase stimulated the BET-1A cells, as well as human alveolar macrophages, pulmonary fibroblasts, and neutrophils, to produce IL-8. In contrast, nitrite reductase induced significant levels of tumor necrosis factor alpha and IL-1beta protein only in human alveolar macrophages. These data support the notion that nitrite reductase from P. aeruginosa induces the production of inflammatory cytokines by respiratory cells and may contribute to the pathogenesis of chronic airway diseases and persistent P. aeruginosa infection.

Sulfuric acid-induced changes in the physiology and structure of the tracheobronchial airways.

Sulfuric acid aerosols occur in the ambient particulate mode due to atmospheric conversion from sulfur dioxide (SO2). This paper describes the response of the rabbit tracheobronchial tree to daily exposures to sulfuric acid (H2SO4) aerosol, relating physiological and morphological parameters. Rabbits were exposed to filtered air (sham control) or to submicrometer-sized H2SO4 at 250 micrograms/m3 H2SO4, for 1 hr/day, 5 days/week, with sacrifices after 4, 8, and 12 months of acid (or sham) exposure; some rabbits were allowed a 3-month recovery after all exposures ended. H2SO4 produced a slowing of tracheobronchial mucociliary clearance during the first weeks of exposure; this change became significantly greater with continued exposures and did not improve after exposures ended. Airway hyperresponsiveness was evident by 4 months of acid exposure; the condition worsened by 8 months of exposure and appeared to stabilize after this time. Standard pulmonary mechanics parameters showed no significant trends with repeated acid exposure, except for a decline in dynamic lung compliance in animals exposed to acid for 12 months. Lung tissue samples obtained from exposed animals showed a shift toward a greater frequency of smaller airways compared to control, an increase in epithelial secretory cell density in smaller airways, and a shift from neutral to acidic glycoproteins in the secretory cells. The effect on airway diameter resolved after the exposures ceased, but the secretory cell response did not return to normal within the recovery period. No evidence of inflammatory cell infiltration was found due to H2SO4 exposure. Thus, significant alterations in the physiology of the tracheobronchial tree have been demonstrated due to repeated 1-hr exposures to a concentration of H2SO4 that is one-fourth the current 8-hr threshold limit value for exposure in the work environment. The cumulative dose inhaled by the rabbits is similar to current peak daily doses from ambient exposure in North America. The results obtained in the rabbit model provide insight into early changes in the tracheobronchial tree due to repeated irritant exposure and may be involved in the pathogenesis of chronic airway disease.

Sulfuric Acid-Induced Changes in the Physiology and Structure of the Tracheobronchial Airways

Sulfuric acid aerosols occur in the ambient particulate mode due to atmospheric conversion from sulfur dioxide (SO2). This paper describes the response of the rabbit tracheobronchial tree to daily exposures to sulfuric acid (H2SO4) aerosol, relating physiological and morphological parameters. Rabbits were exposed to filtered air (sham control) or to submicrometer-sized H2SO4 at 250 micrograms/m3 H2SO4, for 1 hr/day, 5 days/week, with sacrifices after 4, 8, and 12 months of acid (or sham) exposure; some rabbits were allowed a 3-month recovery after all exposures ended. H2SO4 produced a slowing of tracheobronchial mucociliary clearance during the first weeks of exposure; this change became significantly greater with continued exposures and did not improve after exposures ended. Airway hyperresponsiveness was evident by 4 months of acid exposure; the condition worsened by 8 months of exposure and appeared to stabilize after this time. Standard pulmonary mechanics parameters showed no significant trends with repeated acid exposure, except for a decline in dynamic lung compliance in animals exposed to acid for 12 months. Lung tissue samples obtained from exposed animals showed a shift toward a greater frequency of smaller airways compared to control, an increase in epithelial secretory cell density in smaller airways, and a shift from neutral to acidic glycoproteins in the secretory cells. The effect on airway diameter resolved after the exposures ceased, but the secretory cell response did not return to normal within the recovery period. No evidence of inflammatory cell infiltration was found due to H2SO4 exposure. Thus, significant alterations in the physiology of the tracheobronchial tree have been demonstrated due to repeated 1-hr exposures to a concentration of H2SO4 that is one-fourth the current 8-hr threshold limit value for exposure in the work environment. The cumulative dose inhaled by the rabbits is similar to current peak daily doses from ambient exposure in North America. The results obtained in the rabbit model provide insight into early changes in the tracheobronchial tree due to repeated irritant exposure and may be involved in the pathogenesis of chronic airway disease.

Response of the tracheobronchial mucociliary clearance system to repeated irritant exposure: effect of sulfuric acid mist on function and structure.

This study was designed to determine quantitative and temporal alterations in tracheobronchial mucociliary clearance function and structure due to repeated inhalation exposures to a common irritant, sulfuric acid mist. Rabbits were exposed to 250 micrograms/m3 sulfuric acid (0.3 micron) for 1 h/day, 5 days/week, for up to 1 year, with some animals allowed a 3-month recovery period following the end of the acid exposures. Control animals received temperature- and humidity-conditioned water vapor. At intervals of 2 to 4 weeks, animals inhaled a radioactively tagged tracer aerosol (ferric oxide microspheres, 4.5 micron), and its clearance via mucociliary transport from the thorax was monitored by external serial counting. Clearance became slower during the first month of acid exposure, and this slowing became progressive with time through the end of the 12-month exposure period. After cessation of acid exposure, clearance became extremely slow and did not return to normal by the end of the follow-up period. To assess specific histological changes in the tracheobronchial tree, groups of rabbits were killed after 4, 8, or 12 months of exposure and after the follow-up period. Tissue samples from each lung were embedded in plastic, sectioned at 3 micron, and stained with hematoxylin and eosin or alcian blue/periodic acid-Schiff (AB/PAS). Acid exposure changed the airway diameter distribution compared to the control; except for the follow-up group, all acid-exposed animals had a shift to smaller airways. Acid inhalation also caused an increase in epithelial secretory cell density and a shift from PAS to AB staining of glycoprotein within secretory cells, both of which were unresolved by 3 months after the exposures ceased. No evidence of inflammation was found in any of the animals. Thus, repeated exposures to H2SO4 resulted in a slowing of mucociliary clearance that was associated with alterations in airway morphometry and morphology. Such changes may be involved in the early pathogenesis of chronic airway disease.