Airway Epithelium and Mucous Secretion




Introduction


When animals migrated from sea to land, gills were exchanged for lungs, thus increasing the surface area for gas exchange, and the respiratory organ (lungs) was relocated deep within the thorax. This new apparatus required a connection from the areas of gas exchange in the alveoli to the external environment, which was provided by the airways. The airway epithelium is exposed to a wide array of environmental “invaders,” such as bacteria, viruses, allergens, and environmental toxins such as cigarette smoke and air pollutants. Particulate invaders first deposit on the luminal airway epithelial surface and move across the surface to enter the body of the host. In healthy individuals the scarcity of secretions, the effectiveness of clearance of invaders from the epithelial surface, and the efficiency of the host systems for defensive cellular responses to invaders allows protection of the host without symptoms or significant pathologic changes. However, in chronic inflammatory airway diseases, pathologic responses are overexuberant, impairing rather than protecting the host. This chapter focuses on mucus and its major constituent, mucins, and the effects of their hypersecretion.




Components of Mucus


Mucous secretion normally plays a protective role in the epithelium. Airway mucus is a complex mixture of proteins and liquids and includes a sol phase composed of water and electrolytes. Mucus consists of water (95%), most of which is bound in a viscoelastic gel containing mucins. The gel-forming mucins in mucus are high-molecular-weight glycoproteins that are key components of mucous cells and are rich in carbohydrates. Mucin oligosaccharides are joined by an initial α 0 -glycoside linkage of N -acetylgalactosamine to the hydroxyl moieties of serine or threonine of the mucin protein backbone. Mucins produced intracellularly are packed tightly within granules. During exocytosis the cells secrete their granule contents in a condensed form, and the secreted mucins undergo rapid hydration to form a gel with unusual viscoelastic properties that allow the mucins to interact with cilia to effect mucociliary clearance. Various mucins may have very different biophysical properties, and future studies are needed to characterize these properties and their potential pathophysiologic implications. Currently approximately 19 mucin (MUC) genes have been cloned. They are divided into two groups: membrane-associated and gel-forming secreted mucins. Of the secreted mucins, two are especially prominent in inflammatory airway diseases: MUC5AC in airway goblet cells and MUC5B in submucosal gland mucous cells.




Normal Airway Mucins


Gel-forming mucins are produced by mucous tubules in submucosal glands in the large conducting airways and by goblet cells located in the surface epithelium in both large and small airways. MUC5AC is the predominant mucin produced by the surface epithelium and by airway epithelial cells in culture. MUC5B is the predominant mucin in the submucosal glands. MUC5AC is more susceptible to proteolytic degradation than is MUC5B. Thus mucins have a complicated structure, which is likely to contribute to their functions. In the normal surface epithelium, goblet cells are sparse but present at all levels, decreasing in number peripherally, presumably reflecting less contact of peripheral airways with invading environmental particulates. Pathogen-free animals have few goblet cells in the airway epithelium. In healthy airways, multiple stimuli induce mucin production, but the amount of mucin production is small.


On the surface of the airways is a periciliary sol phase, above which the mucous gel floats. The contracting cilia propel the mucous blanket mouthward, along with trapped environmental particles that have deposited on the epithelial surface.




Mucous Hypersecretory Diseases and Their Clinical Consequences


Differences in Clinical Presentation among Various Hypersecretory Diseases


Chronic inflammatory airway diseases are associated with mucous hypersecretion, and the stimuli involved may vary. For example, allergic factors are involved in asthma (see Chapter 41 ), genetic abnormalities underlie cystic fibrosis (CF) (see Chapter 47 ), and the inhalation of toxic components of cigarette smoke is the main cause of chronic obstructive pulmonary disease (COPD) (see Chapter 43 ). Regardless of the variety of causes, the similarities that exist in cellular responses among these various disease states (including mucous hypersecretion) suggest that some common mechanisms are likely to exist that could be useful in therapy.


In patients with severe asthma, CF, and COPD, exaggerated airway epithelial mucin production can lead to mucous plugging and sometimes to death. MUC5AC contributes importantly to mucous plugging in these diseases. In patients with hypersecretory diseases, multiple stimuli can exaggerate mucus production and cause clinical exacerbations. Examples of these stimuli are listed in Table 10-1 .



Table 10-1

Examples of Stimuli That Induce Mucin Synthesis In Vitro or In Vivo by EGFR Activation In Airways











IN VITRO EXPERIMENTS



  • Bacterial products




    • -Pseudomonas aeruginosa supernatant



    • -Lipopolysaccharide (LPS)



    • -Lipoteichoic acid (LTA)




  • Phorbol 12-myristate 13-acetate (PMA)



  • Cigarette smoke



  • Inflammatory cells




    • -Neutrophils



    • -Eosinophils




  • Serine proteases




    • -Human neutrophil elastase



    • -Human airway trypsin-like protease


IN VIVO EXPERIMENTS



  • Th2 cells




    • -Antigen (ovalbumin)



    • -IL-13




  • Mechanical damage of epithelium



  • Cigarette smoke



  • Leukotrienes



  • Cathelicidin


EGFR, epidermal growth factor receptor; IL, interleukin; Th2, type 2 T helper.

From Burgel PR, Nadel JA: Roles of epidermal growth factor receptor activation in epithelial cell repair and mucin production in airway epithelium. Thorax 59:992, 2004.


Manifestations of Mucous Hypersecretion Depend on Airway Location


Intrathoracic airways branch continuously from the trachea to small airways (terminal bronchioles) before they finally arrive in the alveolar zone. Because of their peripheral location, the small airways remain relatively “silent.”


In the 1950s, clinicians recognized a clinical condition consisting of cough and sputum production (which they called “chronic bronchitis” or “simple bronchitis”). They recognized the symptoms as part of a disease of the conducting airways that was disturbing to patients but did not cause serious clinical deterioration or death. One logical explanation for this clinical condition was as follows: in disease of the conducting airways, overproduction of mucins in the submucosal glands travels to the airway lumen via ducts localized to airway bifurcations. The cough receptors are colocalized at airway bifurcations. Stimulation of these receptors (e.g., by chronic smoking) causes cough and activates mucous secretion ( Fig. 10-1 ).




Figure 10-1


Colocalization of gland duct opening and cough receptors at airway bifurcations.

Tantalum bronchogram outlines airways. Arrow indicates site of submucosal gland duct opening and site of cough receptor concentration at large airway bifurcation.

(From Nadel JA: Mucous hypersecretion and relationship to cough. Pulm Pharmacol Ther 26[5]:510–513, 2013.)


Together, anatomists and physiologists advanced the structural analysis of mucin production in submucosal glands and in epithelial goblet cells. These innovators paid little attention to the importance of the peripheral airways. However, it was pathologists who became intrigued by bronchiolar mucous plugging and suggested that plugging was an early event in the development of emphysema. In the study of mucins, this seminal observation lost out to the interest in mucin studies in the large airways, which led to the discoveries of the mucin genes and the signaling pathways involved in mucin production. However, the importance of the small airways soon resurfaced: in 1963 the anatomist Ewald Weibel analyzed the total airway cross section as a function of generation number. He showed that the first five generations occupy a small cross-sectional area and that the bronchioles occupy an exponentially increased cross-sectional area (see Fig. 4-2 ). Using Weibel’s data, it was predicted that normally most of the airflow resistance resides in the large airways and that the airflow resistance contributed by the bronchioles is normally very small.


Early studies implicated bronchioles in lung disease, but their study has been impeded because (1) bronchoscopy, which is very useful for examination and biopsy of large airways, is unable to visualize bronchioles; (2) clinical radiologic techniques still cannot resolve bronchioles; and (3) for structural reasons, biopsies are not usually useful in bronchiolar pathologic conditions. For these reasons, bronchioles still remain a relatively “silent zone” of the lungs!


In bronchioles, because of striking differences in structure from the large airways, the effects of mucous hypersecretion have different manifestations. Bronchiolar hypersecretion is not associated with cough because bronchioles do not possess cough receptors. Furthermore, the number of bronchioles and therefore the total cross-sectional area is very large, so obstruction of bronchioles will not have a measurable effect on airflow resistance until most of the bronchioles are occluded. In acute asthma the importance of bronchiolar obstruction has often only been discovered postmortem, when microscopic examination of bronchioles has shown extensive bronchiolar plugging. Diffuse bronchiolar plugging is also found in advanced CF at the time of lung transplantation. Remarkably, the potential importance of bronchiolar obstruction was reported as long ago as 1956 by pathologists as an early manifestation of emphysema, but the role of plugging in lung tissue breakdown was not explored.


Because of the small diameters, bronchioles are more likely to become obstructed (“plugged”) than airways with larger diameters. Therapy for airway obstruction usually focuses on smooth muscle contraction (“bronchospasm”). However, in chronic obstructive airway diseases, thickening of the airway wall and mucous hypersecretion are predicted to contribute significantly to the airflow limitation that is present (shown schematically in Fig. 10-2 ). In spite of the fact that chronic progressive mucous plugging is common in chronic inflammatory airway diseases such as COPD, CF, and acute fatal asthma, the clinical detection of peripheral mucous plugging is very difficult. This is due to the fact that these small structures are not visualized radiologically or visually during bronchoscopy and are difficult to sample in biopsies. Novel methods for further examination of bronchiolar structure and function are sorely needed.




Figure 10-2


Schematic representation of the potential impact of goblet cell hyperplasia and mucous plugging on small airway obstruction.

In normal individuals the surface epithelium covering small airways ( blue ) contains few goblet cells and is composed of a thin layer of epithelial cells. A and B, Normal, thin epithelium contributes little to luminal narrowing both when airway smooth muscles are relaxed ( A ) and when smooth muscles shorten ( B ). C and D, In various airway inflammatory diseases (e.g., acute asthma, COPD, cystic fibrosis), epithelial cell hyperplasia is present in small airway epithelium, resulting in epithelial thickening. C, When airway smooth muscles are relaxed, thickened epithelium may contribute little to airflow limitation. D, However, when smooth muscles shorten, the hyperplastic epithelium (composed of epithelium, lamina propria, muscle, and adventitial compartments) becomes an important determinant of airflow limitation. E and F, When hyperplastic goblet cells release mucins in the airway lumen ( pale green circle ), swelling of secreted mucins during hydration may result in mucous obstruction. F, The contribution of mucous secretions to airflow limitation is increased when smooth muscles shorten. Thus both goblet cell hyperplasia and mucous obstruction in small airways are predicted to contribute significantly to the airflow limitation in chronic inflammatory airway diseases.

(Redrawn from Burgel PR, Montani D, Danel C, et al: A morphometric study of mucins and small airway plugging in cystic fibrosis. Thorax 62:153, 2007.)




Importance of Mucous Hypersecretion in Chronic Inflammatory Airway Diseases


In healthy subjects, mucous secretion is involved in innate immune defense of the host (see Chapter 12 ). These host responses normally defend efficiently against inhaled “invaders” (e.g., microbes, inhaled irritants), usually without obvious interference with function. However, in various chronic inflammatory airway diseases, there is evidence of abnormal inflammatory responses, which are involved in the pathophysiologic changes of the diseases. Thus, although secretion of mucus normally plays a protective role in host defense, hypersecretion plays important roles in the pathophysiologic characteristics of various chronic airway diseases. The following section is a brief discussion of some current issues related to mucous hypersecretion in selected obstructive airway diseases. Detailed reviews are provided in the chapters on chronic bronchitis and COPD (see Chapters 43 and 44 ), asthma (see Chapters 41 and 42 ), and cystic fibrosis (see Chapter 47 ).


Brief Review of Cystic Fibrosis, Asthma, and Chronic Bronchitis/COPD


Cystic Fibrosis (see Chapter 47 )


In CF, mucous plugging has long been known to play a major role in the pathophysiologic changes of the disease, but the extent and characteristics of plugging have previously escaped analysis. Various investigators have reported that mucins contribute to the pathophysiologic changes in CF. A biopsy study of conducting airways in CF showed an increase in goblet cell size and an enlarged submucosal gland volume. Kreda and colleagues described the relationship between the cystic fibrosis transmembrane conductance regulator (CFTR), mucins, and mucous obstruction. In a quantitative analysis of lungs removed at the time of lung transplantation, Burgel and associates reported that in CF most small airways contained extensive plugging, whereas control lungs contained only rare plugs ( Fig. 10-3 ). Another example of plugging in small noncartilaginous airways is shown in Figure 10-4 . Pulmonary function studies in these CF patients before transplantation were indicative of severe airway obstruction, and the authors suggested that the extensive peripheral airway plugging contributed to the loss of lung function. In the airways with plugs, there was extensive goblet cell hyperplasia, with mucins streaming into the luminal plugs. These studies indicate that CF patients with late-stage disease and severe airway obstruction have extensive peripheral airway plugging, which contributes to the advanced obstruction present and the need for transplantation.




Figure 10-3


Mucous glycoconjugates and mucins in airway lumens of controls and patients with cystic fibrosis (CF).

Representative photomicrographs. Sections containing small airways in a typical control subject ( left panels ) and in a patient with CF ( right panels ) were stained with Alcian blue/periodic acid-Schiff (AB/PAS) for mucous glycoconjugates, blue staining ( upper panels ) or with antibodies to mucin MUC5B (brown staining; middle panels ) and MUC5AC (brown staining, lower panels ). In the control subject the airway lumen (L) is empty; no staining for AB/PAS, MUC5B, or MUC5AC is observed in the lumen or in the epithelium. In the patient with CF, the airway lumen is obstructed by a plug containing mucous glycoconjugates (AB/PAS), MUC5B, and MUC5AC only at the periphery of the plug ( arrowheads ), adjacent to the epithelium. Staining for mucins is absent in the center of the plug. Goblet cells in the epithelium are stained positively with AB/PAS and with MUC5AC and MUC5B antibodies ( arrows ). (Original magnification ×100; bar = 100 µm.)

(From Burgel PR, Montani D, Danel C, et al: A morphometric study of mucins and small airway plugging in cystic fibrosis. Thorax 62:153, 2007.)



Figure 10-4


Mucin plug in terminal bronchiole in a patient with cystic fibrosis (CF).

This section was stained with Alcian blue/periodic acid-Schiff (AB/PAS). The plugged bronchiole is surrounded by alveoli.


Although mucins play a role in the production of mucous plugs, other components (e.g., neutrophil products), including DNA and extravasated blood products (such as albumin), also contribute. It should be noted that the observation that mucin production is increased in CF does not necessarily imply that CF epithelial cells have increased responsiveness to mucin production, but only that, in the area of plugs, there are stimuli (e.g., Pseudomonas bacteria) that effectively stimulate production and secretion of mucins.


In CF, Burgel and associates noted that the staining for mucins occupied approximately 20% of the plugged lumen volume, indicating that mucins contributed to the plugs but that other molecules were also present. Neutrophils were also conspicuous in the plugs. Voynow and coworkers reported that elastase, a product secreted by neutrophils, causes the degradation of MUC5AC. Davies and colleagues reported findings that suggested that one airway mucin (MUC5AC) is more vulnerable to proteolytic degradation than the other (MUC5B). The findings reported by Burgel and associates confirmed this observation. The prominent presence of neutrophils in plugged airways suggests the possibility that neutrophil-mediated effects could be involved in proteolytic inactivation of secreted mucins. Many other molecules may also be components of mucous plugs. For example, albumin is a normal component of blood, but, in states of inflammation with increased vascular permeability, albumin is reported to move from the vascular lumen to the airway lumen. Albumin is a plasma protein that can move across the endothelial surface via multiple mechanisms. When albumin is present in renal tubular epithelial cells, it activates epidermal growth factor receptors (EGFRs) and causes the production of inflammatory mediators. If albumin has similar actions in airways, perhaps in diseases with increased vascular permeability, albumin might play a role in activating the EGFR and leading to mucous hypersecretion.


In CF, a disease caused by mutations in the CFTR, exaggerated airway epithelial cell interleukin-8 (IL-8) production leads to persistent neutrophilic inflammation, a severe and untreated feature of CF airway disease. Some investigators suggest that inflammatory changes in CF result from infection, and others suggest that the exaggerated effects are intrinsic elements of CF disease. It is known that activation of the EGFR is responsible for the production of IL-8, which is a potent neutrophil chemokine. Kim and associates recently hypothesized that normal CFTR suppresses EGFR-dependent IL-8 production and that loss of CFTR at the epithelial surface exaggerates IL-8 production via activation of a proinflammatory EGFR cascade. Airway epithelial cells known to contain normal CFTR were treated with a CFTR-selective inhibitor, CFTR(inh)-172. The authors found that CFTR(inh)-172 increased IL-8 production via an epithelial surface signaling cascade via the production and release of interleukin-1-α (IL-1α), and binding of IL-1α to its receptor, leading to EGFR activation and hence exag­gerated IL-8 production. (For a description of the cascade leading to exaggerated IL-8 production see Kim. ) Similar exaggerated effects were seen in CF epithelial cells. If this cascade is also activated in CF patients in vivo, one would predict that the increased responsiveness of the airway epithelium accounts for the marked increase in the patient’s responses. In CF this could explain the effects of Pseudomonas infections on the patients’ symptoms and their responses to bacterial therapy. These studies are of interest because they suggest that inflammatory responses such as mucous hypersecretion in CF may be due to exaggeration of EGFR proinflammatory responses when CFTR is absent from the epithelial surface. In addition, in other airway diseases, down-regulation of CFTR may also increase EGFR-mediated proinflammatory responses. A limitation of the study is the fact that it was performed in isolated airway epithelial cells. In addition, CFTR(inh)-172 could conceivably have other side effects. Further studies are indicated to determine the validity of the antagonism of CFTR and EGFR and to further investigate the implications. It is possible that understanding the interactions of the CFTR and the EGFR signaling cascades could provide novel therapies.


Asthma (see Chapters 41 and 42 )


In contrast to normal individuals, for patients with asthma, sputum secretion and cough, symptoms of hypersecretion in large airways, are common. Thus Turner-Warwick and Openshaw reported that 77% of asthma patients had a history of sputum production, and 56% reported maximum sputum production at the peak of asthma attacks. Goblet cell numbers in the conducting airways are increased even in mild asthma.


Chronic asthma has increased as a public health burden over the past 20 years, and acute exacerbations of asthma play important roles in morbidity and cost. Respiratory viruses, mainly rhinoviruses, cause the majority of exacerbations. Asthma attacks cause the acute onset of symptoms and acute deterioration of lung function. Chronic mucus hypersecretion is a common symptom in adults with stable asthma, particularly in smokers and in patients with severe disease. A history of asthma with chronic sputum overproduction is associated with an accelerated decline in maximal expiratory airflow (indicative of airway obstruction). Morgan and colleagues reported that asthmatic subjects show a greater decline in lung function over time than normal subjects.


Mucous hypersecretion in the large conducting airways causes significant symptoms (cough and mucous hypersecretion). Hypersecretion in small airways generally is asymptomatic early but is often ultimately associated with marked airway narrowing (mucous plugging). Further evidence for the involvement of peripheral airways in asthma was provided by Yanai and coworkers, who reported that the majority of the increased airflow resistance in asthmatics resides in small airways.


Widespread mucous hypersecretion was reported in fatal asthma by Cardell and Pearson in 1959 and was confirmed by other studies. Mortality in acute asthma is associated with mucous plugging, which is considered to be a major cause of death in asthma. An example of mucous plugging in fatal asthma is shown in Figure 10-5 .




Figure 10-5


Mucous hypersecretion in fatal asthma.

A, Alcian blue/periodic acid-Schiff (AB/PAS) staining for mucous glycoconjugates in a proximal airway section in a patient with fatal asthma. The airway lumen is plugged (PLUG) with mucus. The epithelium shows marked goblet cell metaplasia. The submucosal gland (GLAND) has intense staining for mucous glycoconjugates and has released its contents into a gland duct (DUCT) that is filled with AB/PAS-stained material. B, Another AB/PAS-stained section of airway epithelium in the same patient at higher magnification. Although goblet cells here remain intact, mucus can be seen streaming from the luminal tips of the goblet cells into the lumen. The lumen is filled with mucus. Both scale bars = 50 µm.


In experimental models of allergic airway disease, mucous hypersecretion has also been reported. For example, allergic sensitization by ovalbumin in rats increased mucin production in the airway epithelium, an effect that was prevented by a selective inhibition of EGFR activation. For details of signaling involved in mucin production, see “ Epithelial Signaling Pathways for Mucin Production ” section.


Chronic Bronchitis and COPD (see Chapters 43 and 44 )


In the past, mucous hypersecretion was often considered to be an annoying but otherwise benign aspect of airway diseases associated with smoking. Gradually researchers have realized that the clinical implications of the effects of cigarette smoke and other chronic irritants depend on the location and composition of the inhaled irritant. Originally, clinical researchers described a group of patients with symptoms of cough and sputum that were only weakly associated with decline in lung function. The disease was called “chronic bronchitis.” Unlike those in normal healthy persons, submucosal glands located in the large conducting airways of these individuals became enlarged and produced copious secretions, which were cleared in the sputum, assisted by cough. It was reasoned that, if disease in smokers is limited to the large airways and airway obstruction is not prominent, mucous hypersecretion may be disturbing but deterioration of lung function may not become conspicuous. A correlation was found between the presence of bronchitis and the presence of submucosal gland hypertrophy in the central airways of smokers. However, it was subsequently shown that chronic mucous hypersecretion is often associated with excessive decline in pulmonary function and increased risk for hospitalization. Mucin production is increased, especially with disease exacerbation and is associated with a decline in forced expiratory volume in 1 second.


Chronic smoking can also lead to progressive obstruction in the small airways, but the changes have been difficult to visualize because of their peripheral location. In addition, the airways are large in number, so the majority of small airways must be obstructed before the airflow resistance is increased and causes symptoms. For these reasons, these peripheral airways remain a relatively silent zone until late in the disease. However, morphologic specimens of small airways in COPD, such as in the study of surgical specimens, showed increased expression of mucins in bronchioles in COPD and an increased number of goblet cells in peripheral airways. From these results it was concluded that exaggerated mucin production takes place in the bronchioles in COPD.


To examine the role of peripheral airway narrowing, in 1968 Hogg and colleagues showed that the major sites of airway obstruction in COPD are the smaller bronchi and bronchioles less than 2 mm in diameter. In 2004 Hogg and associates performed a quantitative assessment of small airways in surgically resected lung tissue in patients with COPD with different degrees of airway obstruction. The progression of COPD was strongly associated with an increase in inflammatory mucous exudates in the lumens of small airways. In addition, they reported an increase in the volume of tissue in the airway wall, which potentiates the effects of luminal plugging (see Fig. 10-2 ). These results emphasize the roles of repair processes in the airway wall that decrease the luminal area and thus play important roles in airway obstruction in COPD.


In a review in 2010 it was concluded that mucus accumulation in conducting airways accounts for the symptoms of patients with COPD. Thus COPD affects central and peripheral airways; effects of the large airways leads to the majority of symptoms, but the peripheral airways appear to be the major sites of airway obstruction. The nature of small airway obstruction in COPD is reviewed by Hogg and colleagues. For a more recent review of the roles of small airways in COPD, see Burgel and associates.




Epithelial Signaling Pathways for Mucin Production


Early Studies of Mucins


Chronic airway diseases have long been associated with mucous hypersecretion. The gel-forming mucins, which are large glycoproteins, became recognized as being responsible for the major characteristics of mucus. Cloning of mucin genes (reviewed by Rose and coworkers ) provided tools for studying mucin regulation. In the 1990s, intense research on mucins began, and many stimuli were shown to produce mucins in airway epithelial cells. Based on the newly available cell and molecular tools, rapid advances began to clarify the pathways involved in mucin production. In 1997 Li and associates reported that Pseudomonas bacteria stimulate epithelial mucin production via a Src-dependent Ras/Raf/MAPK pathway.


Epidermal Growth Factor Receptor Activation


Many of the stimuli, such as microbes and cigarette smoke, that induce mucin production in airways are derived from the environment and are inhaled, depositing on the airway epithelial surface. Takeyama and colleagues hypothesized that receptors on the surface of the airway epithelium could be candidates for epithelial signaling, resulting in mucin production in response to the deposition of the inhaled foreign particulates. EGF, an EGFR ligand, was discovered by Cohen, and subsequently his group expanded the understanding of EGF and its receptor EGFR, focusing on epithelial cell proliferation and cancer. EGFR is a 170-kd membrane glycoprotein that is activated by multiple ligands.


Takeyama and coworkers hypothesized that epithelial responses to inhaled microbes such as Pseudomonas are initiated on the epithelial luminal surface, where the microbes are deposited, by activation of EGFR. The authors discovered that activation of EGFR by its ligands (such as EGF, transforming growth factor [TGF]-α) cause mucin production in cultured human airway epithelial cells, an effect that is prevented by EGFR inhibition. They confirmed these results in rats in vivo and also found that ovalbumin sensitization increases MUC5AC mucin.


Perrais and associates confirmed that EGFR ligands activate EGFR to produce mucins and reported that downstream pathways include a Ras/Raf/MAPK cascade. Subsequent reports identified EGFR as a key player in mucin synthesis/goblet cell metaplasia in response to multiple stimuli in vitro and in vivo (see Table 10-1 ).


Metalloproteases Cleave Membrane-Bound EGFR Ligands to Produce Mucins


Airway epithelial cells themselves synthesize EGFR ligands in the airway epithelium as transmembrane precursors that can be cleaved by metalloproteases, releasing soluble active growth factors. In 1999 Dong and colleagues reported that general metalloprotease inhibitors prevent the effects of EGFR proliferation in proportion to the release of the membrane-bound EGFR ligand TGF-α, and they concluded that the soluble ligand was responsible for the effects of EGFR activation. Prenzel and coworkers similarly reported that a general metalloprotease inhibitor prevented EGFR ligand release and subsequent EGFR activation. These papers were the first to implicate cleavage of EGFR ligands by metalloprotease(s) in EGFR surface signaling. Subsequently Kohri and colleagues linked Pseudomonas bacterial supernatant to mucin production. EGFR activation was required, but the mode of EGFR signaling in the airway epithelium was unclear. One clue came from the study of tumor necrosis factor- α– converting enzyme (TACE), a member of the ADAM family of proteases, which is bound to the luminal surface of the airway epithelium. Shao and Nadel recognized that TACE was capable of cleaving membrane-bound EGFR proligands (e.g., tumor necrosis factor alpha). Shao and associates showed that Pseudomonas and their product lipopolysaccharide increase mucin production in human airway epithelial cells, an effect that was reported to be EGFR dependent and associated with the cleavage and release of the membrane-bound EGFR ligand, TGF-α. Selective knockdown of TACE prevented the sequence leading to mucin production, establishing TACE as important in autocrine signaling involved in mucin production. These results established the role of TACE in the autocrine activation of EGFR and in mucin production in airways. Subsequently a variety of stimuli have been shown to stimulate mucin production via autocrine EGFR signaling in the airway epithelium.


Roles of Reactive Oxygen Species in Airway Epithelial Mucin Production


Excess production of reactive oxygen species (ROS) is known to cause cell damage in chronic inflammatory diseases. For example, ROS scavengers inhibit IL-8 production. Recent studies have contributed to the understanding of the role of ROS in epithelial signaling and mucin production. Thus ROS stimulation (e.g., with H 2 O 2 ) activates EGFR and results in mucin production.


Dual oxidase-1 (DUOX1) was found in human airway epithelial cells and was reported to generate ROS. Shao and associates suggested that DUOX1 could release ROS and thereby activate TACE. They used human neutrophil elastase as the stimulus, because human neutrophil elastase had been previously reported to produce mucin via EGFR ligand-dependent activation. Shao and associates reported that human neutrophil elastase induced TACE activation, TGF-α release, and mucin expression, effects that were inhibited by ROS scavengers, implicating ROS in the response. Knockdown of DUOX1 expression with small interfering RNA prevented the responses. Furthermore, the protein kinase C (PKC)-δ/PKC-θ inhibitor rottlerin prevented the effects of human neutrophil elastase, suggesting that DUOX1 plays a critical role in mucin production via a PKC-δ/PKC-θ-DUOX1-ROS–TACE-proligand-EGFR cascade. The scheme is shown diagrammatically in Figure 10-6 . It is of interest that DUOX1 may release ROS in a limited cell compartment adjacent to TACE, thus producing a signal in defense of the host without harmful effects on epithelial cell viability. In fact, DUOX1 is now recognized as playing a role in multiple epithelial defensive responses such as wound healing. Activation of DUOX1 stimulates the production of the chemokine IL-8. It is hoped that future studies in various epithelia will evaluate the importance of the proinflammatory cascade in a variety of epithelial-based diseases.


Jul 21, 2019 | Posted by in CARDIOLOGY | Comments Off on Airway Epithelium and Mucous Secretion

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