[PubMed] [CrossRef] [Google Scholar] 27. to induces airway mucus hypersecretion by modulating the STAT/EGFR-FOXA2 signaling pathways. Intro Airway mucus forms a protecting covering that entraps foreign particles and microbes, facilitating their clearance by mucociliary transport. Mucus is composed primarily of mucin glycoproteins, water, ions, and cellular debris. Mucins are the major macromolecular component of the mucus gel responsible for its viscoelastic, rheological, and clearance properties. MUC5AC and MUC5B are the major mucins of human being airways (1,C3). Although a deficient mucous barrier intuitively leaves the lungs vulnerable to injury, aberrant mucin secretion and build up contribute significantly to the pathogenesis of airway diseases such as asthma, chronic obstructive pulmonary disease (COPD), and cystic JNJ-47117096 hydrochloride fibrosis (CF) (1,C4). Mucus plugging in asthmatic and COPD lungs is definitely a major cause of airway narrowing and death (5, 6). Furthermore, hypersecretion of MUC5AC is definitely detrimental during acute lung injury (7). The ability of microbial pathogens to induce mucus secretion suggests that it is one of the mechanisms of infection-induced exacerbation in airway diseases (8,C11). is the most common cause of community-acquired pneumonia. Moreover, has long been recognized as a result in of both chronic illness and acute exacerbation in multiple chronic airway diseases, including asthma (12, 13). Several virulence mechanisms of are known, including cytoadherence through a polar attachment organelle (14), generation of reactive oxygen varieties (ROS) (15), and secretion of the community-acquired respiratory stress syndrome (CARDS) toxin (16). Although and CARDS toxin induce mucin manifestation (9, 10, 17), the transmission pathways within the airway epithelium that regulate the response to remain unknown. Previously, we have demonstrated that induces goblet cell hyperplasia and metaplasia (GCHM) and mucus hypersecretion by secreting the redox-active toxin pyocyanin, which induces STAT6 and epidermal growth element receptor (EGFR) signaling to inactivate FOXA2, a major transcriptional repressor of GCHM and mucin biosynthesis (18, 19). Pyocyanin also redox inactivates FOXA2 through posttranslational modifications, resulting in the overexpression of mucins (20). Importantly, the manifestation of FOXA2 is definitely depleted in the Rabbit Polyclonal to LAT airways of bronchiectatic and asthmatic individuals, as well as with the ovalbumin-induced mouse model of asthma (19, 21). In this study, we examined the airway signaling mechanisms modulated by that resulted in FOXA2 depletion and overexpression of mucins. MATERIALS AND METHODS preparation. strain M129 was cultured in SP-4 broth medium at 37C until the color of the medium changed to peach yellow. cells were dislodged having a dish scraper and suspended in sterile saline. The combination was JNJ-47117096 hydrochloride approved through a 25-gauge needle 10 instances. The stock was stored at ?80C. A portion of the stock was serially diluted and plated onto pleuropneumonia-like organism (PPLO) blood agar to determine the CFU count as previously explained (22). M129 grew slowly, yielding extremely small colonies on PPLO blood agar after 7 days at 37C. Plates were then overlaid with blood agar, and 2 days later, colonies were visible as hemolytic plaques. The original stock was diluted to 1 1 108 cells/50-l aliquot and stored at ?80C. All subsequent experiments were performed with aliquots of same frozen stock preparation. Mouse handling and exposure to M129 (1 108 CFU in 50 JNJ-47117096 hydrochloride l) on day time 0. Control mice were exposed to 50 l of phosphate-buffered saline (PBS). On day time 3, mouse lungs were collected for analyses. These animal studies were carried out in strict accordance with the Guidebook for the Care and Use of Laboratory Animals of the National Institutes of Health. The protocol was authorized by the Institutional Animal Care and Use Committee JNJ-47117096 hydrochloride in the University or college of Illinois at Urbana-Champaign. Cytokine analyses. The levels of interleukin-4 (IL-4), IL-6, and IL-13 in the bronchoalveolar lavage (BAL) fluid of mouse lungs were determined with specific enzyme-linked immunosorbent assay (ELISA) packages according to the manufacturer’s protocols (R&D Systems) and by immunohistochemistry (IHC) analysis. Histopathology and IHC analyses. Mouse lungs were processed as explained previously (18, 20). Briefly, a cannula was put into the trachea and the lungs were instilled with 10% neutral buffered formalin at a constant pressure.
