No abstract

Mammalian airways normally regulate the volume of a thin liquid layer, the periciliary liquid (PCL), to facilitate the mucus clearance component of lung defense. Studies under standard (static) culture conditions revealed that normal airway epithelia possess an adenosine-regulated pathway that blends Na ؉ absorption and Cl ؊ secretion to optimize PCL volume. In cystic fibrosis (CF), the absence of CF transmembrane conductance regulator results in a failure of adenosine regulation of PCL volume, which is predicted to initiate mucus stasis and infection. However, under conditions that mimic the phasic motion of the lung in vivo, ATP release into PCL was increased, CF ion transport was rebalanced, and PCL volume was restored to levels adequate for lung defense. This ATP signaling system was vulnerable, however, to insults that trigger CF bacterial infections, such as viral (respiratory syncitial virus) infections, which up-regulated extracellular ATPase activity and abolished motion-dependent ATP regulation of CF PCL height. These studies demonstrate (i) how the normal coordination of opposing ion transport pathways to maintain PCL volume is disrupted in CF, (ii) the hitherto unknown role of phasic motion in regulating key aspects of normal and CF innate airways defense, and (iii) that maneuvers directed at increasing motion-induced nucleotide release may be therapeutic in CF patients.The lung must continually defend itself against bacteria that deposit on airway surfaces during normal tidal breathing. It appears that mechanical clearance of bacteria mediated by mucus transport is the principal innate defense mechanism of mammalian airways (1-4). Recent data have shown that a critical component of this defense system is the thin (ϳ7) m liquid layer lining airway surfaces, the periciliary liquid (PCL), 2 that provides a low viscosity solution for ciliary beating and acts a lubricant layer for mucus transport (5, 6). In cystic fibrosis (CF) lung disease, it appears that the primary pathophysiologic defect is the depletion of PCL volume, resulting in a failure of mucus clearance of bacteria and persistent airways infection (7,8).However, questions have been raised as to the relevance of PCL depletion to CF pathogenesis in vivo (9). For example, whereas in vitro data from standard (static) culture systems describe rapid depletion of PCL height and a complete failure of mucus transport (7), young CF patients exhibit reduced but measurable rates of mucus clearance in vivo (10). This inconsistency suggests that mechanisms for PCL height regulation operating in vivo are absent from standard static culture systems. In addition, clinical observations suggest that CF lung disease exacerbates intermittently and is heterogeneous. Often, viral infections trigger these disease exacerbations (11, 12), but no links between viral infection and PCL regulation have been reported.To investigate these questions, we used a well differentiated airway epithelial culture system that exhibits PCL volume regulation and mucus transport (7). Ba...

Inflammation of human bronchial epithelia (HBE) activates the endoplasmic reticulum (ER) stress transducer inositolrequiring enzyme 1 (IRE1)α, resulting in IRE1α-mediated cytokine production. Previous studies demonstrated ubiquitous expression of IRE1α and gut-restricted expression of IRE1β.We found that IRE1β is also expressed in HBE, is absent in human alveolar cells, and is upregulated in cystic fibrosis and asthmatic HBE. Studies with Ire1β−/− mice and Calu-3 airway epithelia exhibiting IRE1β knockdown or overexpression revealed that IRE1β is expressed in airway mucous cells, is functionally required for airway mucin production, and this function is specific for IRE1β vs. IRE1α. IRE1β-dependent mucin production is mediated, at least in part, by activation of the transcription factor X-box binding protein-1 (XBP-1) and the resulting XBP-1-dependent transcription of anterior gradient homolog 2, a gene implicated in airway and intestinal epithelial mucin production. These novel findings suggest that IRE1β is a potential mucous cell-specific therapeutic target for airway diseases characterized by mucin overproduction.

Inflamed cystic fibrosis (CF) human bronchial epithelia (HBE), or normal HBE exposed to supernatant from mucopurulent material (SMM) from CF airways, exhibit endoplasmic reticulum (ER)/Ca 2؉ store expansion and amplified Ca 2؉ -mediated inflammation. HBE inflammation triggers an unfolded protein response (UPR) coupled to mRNA splicing of X-box binding protein-1 (XBP-1). Because spliced XBP-1 (XBP-1s) promotes ER expansion in other cellular models, we hypothesized that XBP-1s is responsible for the ER/Ca 2؉ store expansion in inflamed HBE. XBP-1s was increased in freshly isolated infected/inflamed CF in comparison with normal HBE. The link between airway epithelial inflammation, XBP-1s, and ER/Ca

Airway epithelia are confronted with distinct signals emanating from the luminal and/or serosal environments. This study tested whether airway epithelia exhibit polarized intracellular free calcium (Ca2+ i) and anion secretory responses to 5′ triphosphate nucleotides (ATP/UTP), which may be released across both barriers of these epithelia. In both normal and cystic fibrosis (CF) airway epithelia, mucosal exposure to ATP/UTP increased Ca2+ i and anion secretion, but both responses were greater in magnitude for CF epithelia. In CF epithelia, the mucosal nucleotide–induced response was mediated exclusively via Ca2+ i interacting with a Ca2+-activated Cl− channel (CaCC). In normal airway epithelia (but not CF), nucleotides stimulated a component of anion secretion via a chelerythrine-sensitive, Ca2+-independent PKC activation of cystic fibrosis transmembrane conductance regulator. In normal and CF airway epithelia, serosally applied ATP or UTP were equally effective in mobilizing Ca2+ i. However, serosally applied nucleotides failed to induce anion transport in CF epithelia, whereas a PKC-regulated anion secretory response was detected in normal airway epithelia. We conclude that (1) in normal nasal epithelium, apical/basolateral purinergic receptor activation by ATP/UTP regulates separate Ca2+-sensitive and Ca2+-insensitive (PKC-mediated) anion conductances; (2) in CF airway epithelia, the mucosal ATP/UTP-dependent anion secretory response is mediated exclusively via Ca2+ i; and (3) Ca2+ i regulation of the Ca2+-sensitive anion conductance (via CaCC) is compartmentalized in both CF and normal airway epithelia, with basolaterally released Ca2+ i failing to activate CaCC in both epithelia.