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Bitter taste receptors (T2Rs) are G protein-coupled receptors (GPCRs) expressed in various cell types including ciliated airway epithelial cells and macrophages. T2Rs in these two innate immune cell types are activated by bitter products, including those secreted by Pseudomonas aeruginosa, leading to Ca2+-dependent activation of endothelial nitric oxide (NO) synthase (eNOS). NO enhances mucociliary clearance and has direct antibacterial effects in ciliated epithelial cells. NO also increases phagocytosis by macrophages. Using biochemistry and live-cell imaging, we explored the role of heat shock protein 90 (HSP90) in regulating T2R-dependent NO pathways in primary sinonasal epithelial cells, primary monocyte-derived macrophages, and a human bronchiolar cell line (H441). Immunofluorescence showed that H441 cells express eNOS and T2Rs and that the bitter agonist denatonium benzoate activates NO production in a Ca2+- and HSP90-dependent manner in cells grown either as submerged cultures or at the air–liquid interface. In primary sinonasal epithelial cells, we determined that HSP90 inhibition reduces T2R-stimulated NO production and ciliary beating, which likely limits pathogen clearance. In primary monocyte-derived macrophages, we found that HSP-90 is integral to T2R-stimulated NO production and phagocytosis of FITC-labeled Escherichia coli and pHrodo-Staphylococcus aureus. Our study demonstrates that HSP90 serves as an innate immune modulator by regulating NO production downstream of T2R signaling by augmenting eNOS activation without impairing upstream Ca2+ signaling. These findings suggest that HSP90 plays an important role in airway antibacterial innate immunity and may be an important target in airway diseases such as chronic rhinosinusitis, asthma, or cystic fibrosis. [ABSTRACT FROM AUTHOR]

T2R bitter taste receptors in airway motile cilia increase ciliary beat frequency (CBF) and nitric oxide (NO) production. Polymorphisms in some T2Rs are linked to disease outcomes in chronic rhinosinusitis (CRS) and cystic fibrosis (CF). We examined the expression of cilia T2Rs during the differentiation of human nasal epithelial cells grown at air–liquid interface (ALI). The T2R expression increased with differentiation but did not vary between CF and non-CF cultures. Treatment with Pseudomonas aeruginosa flagellin decreased the expression of diphenhydramine-responsive T2R14 and 40, among others. Diphenhydramine increased both NO production, measured by fluorescent dye DAF-FM, and CBF, measured via high-speed imaging. Increases in CBF were disrupted after flagellin treatment. Diphenhydramine impaired the growth of lab and clinical strains of P. aeruginosa, a major pathogen in CF and CF-related CRS. Diphenhydramine impaired biofilm formation of P. aeruginosa, measured via crystal violet staining, as well as the surface attachment of P. aeruginosa to CF airway epithelial cells, measured using colony-forming unit counting. Because the T2R agonist diphenhydramine increases NO production and CBF while also decreasing bacterial growth and biofilm production, diphenhydramine-derived compounds may have potential clinical usefulness in CF-related CRS as a topical therapy. However, utilizing T2R agonists as therapeutics within the context of P. aeruginosa infection may require co-treatment with anti-inflammatories to enhance T2R expression. [ABSTRACT FROM AUTHOR]

Background: Epithelial solitary chemosensory cell (tuft cell) bitter taste signal transduction occurs through G protein coupled receptors and calcium-dependent signaling pathways. Type II taste cells, which utilize the same bitter taste signal transduction pathways, may also utilize cyclic adenosine monophosphate (cAMP) as an independent signaling messenger in addition to calcium.Methods: In this work we utilized specific pharmacologic inhibitors to interrogate the short circuit current (Isc) of polarized nasal epithelial cells mounted in Ussing chambers to assess the electrophysiologic changes associated with bitter agonist (denatonium) treatment. We also assessed release of human β-defensin-2 from polarized nasal epithelial cultures following treatment with denatonium benzoate and/or potassium channel inhibitors.Results: We demonstrate that the bitter taste receptor agonist, denatonium, decreases human respiratory epithelial two-pore potassium (K2P) current in polarized nasal epithelial cells mounted in Ussing chambers. Our data further suggest that this occurs via a cAMP-dependent signaling pathway. We also demonstrate that this decrease in potassium current lowers the threshold for denatonium to stimulate human β-defensin-2 release.Conclusions: These data thus demonstrate that, in addition to taste transducing calcium-dependent signaling, bitter taste receptor agonists can also activate cAMP-dependent respiratory epithelial signaling pathways to modulate K2P currents. Bitter-agonist regulation of potassium currents may therefore serve as a means of rapid regional epithelial signaling, and further study of these pathways may provide new insights into regulation of mucosal ionic composition and innate mechanisms of epithelial defense. [ABSTRACT FROM AUTHOR]

Taste receptors in the oral cavity guide our preferences for foods, preventing toxic ingestions and encouraging proper nutrient consumption. More recently, expression of taste receptors has been demonstrated in other locations throughout the body, including the airway, gastrointestinal tract, pancreas, and brain. The extent and specific roles of extraoral taste receptors are largely unknown, but a growing body of evidence suggests that taste receptors in the airway serve a critical role in sensing bacteria and regulating innate immunity. This review will focus on the function of bitter and sweet taste receptors in the human airway, with particular emphasis on T2R38, a bitter taste receptor found in sinonasal ciliated cells, and the bitter and sweet receptors found on specialized sinonasal solitary chemosensory cells. The importance of these novel taste receptor-immune circuits in the human airway and their clinical relevance in airway disease will also be reviewed. [ABSTRACT FROM AUTHOR]

Innate and adaptive defense mechanisms protect the respiratory system from attack by microbes. Here, we present evidence that the bitter taste receptor T2R38 regulates the mucosal innate defense of the human upper airway. Utilizing immunofluorescent and live cell imaging techniques in polarized primary human sinona-sal cells, we demonstrate that T2R38 is expressed in human upper respiratory epithelium and is activated in response to acyl-homoserine lactone quorum-sensing molecules secreted by Pseudomonas aeruginosa and other gram-negative bacteria. Receptor activation regulates calcium-dependent NO production, resulting in stimu-lation of mucociliary clearance and direct antibacterial effects. Moreover, common polymorphisms of the TAS2R38 gene were linked to significant differences in the ability of upper respiratory cells to clear and kill bacteria. Lastly, TAS2R38 genotype correlated with human sinonasal gram-negative bacterial infection. These data suggest that T2R38 is an upper airway sentinel in innate defense and that genetic variation contributes to individual differences in susceptibility to respiratory infection. [ABSTRACT FROM AUTHOR]