Figure 11.

Nasal polyp exhibited calcium responses to apical PAR-2 stimulation but control turbinate did not. a, representative image of 2-FLI induced calcium response (Calbryte 590) in a thin piece of polyp mucosa mounted in an Ussing chamber holder and imaged with a spinning disk confocal microscope. b and c, representative traces of polyp (b) and turbinate (c) responses to apical 2FLI (50 μm), trypsin (1 μm), Der p 3 (1 μm), or thrombin (1 μm). ATP (100 μm), which activates apical purinergic receptors, was a positive control. d, peak change in calcium in tissue treated apically with PAR-2 agonists. Each data point is an independent experiment using tissue from separate patients (n = 7 per condition). e, peak change in calcium in polyp tissue treated apically with PAR-2 agonist (50 μm 2FLI or 120 μm SLIGRL-NH₂) ± antagonist (80 μm FSLLRY-NH₂), confirming responses are due to PAR-2. Each data point is an independent experiment using tissue from separate patients (n = 4 per condition). f, immunofluorescence showing lack of apical PAR-2 staining in polyp ciliated cells with Na⁺K⁺ ATPase (NKP) as marker for the basolateral membrane and β-tubulin IV (βTubIV) (cilia marker) shown in green. Pearson's correlation and Mander's overlap coefficients for NKP and PAR-2 in ciliated cells were both ≥0.95 in 10 images analyzed. Scale bars are 10 μm. g, peak change in calcium in isolated ciliated cells; n.s., no significant difference was observed between polyp and turbinate. Significance in d and g was determined by one-way ANOVA with Bonferroni post-test and significance in e by one-way ANOVA with Dunnett's post-test comparing all values to HBSS only control; **, p < 0.01.