Fig. 5. α5SNP reduces Ca²⁺ permeability and enhances basal cell proliferation and TNF-α production.

a Concentration-response curves for acetylcholine (ACh)- or nicotine-evoked changes in intracellular Ca²⁺ measured on csBEC isolated from α5WT, α5SNP or GFP LV mice. Data are expressed as mean ± SEM.The response curves for each nAChR variant are significantly different (n = 3 for ACh, n = 4 for nicotine, per variant). *p < 0.05, **p < 0.01 for α5SNP versus α5WT; ^$p < 0.05, ^$$p < 0.01, ^$$$p < 0.001, ^$$$$p < 0.0001 for GFP vs α5WT; ^#p < 0.05, ^##p < 0.01 for GFP vs α5SNP, two-way ANOVA. b Proliferation of csBECtreated with SQ22536 (non-selective inhibitor of ACs) or GDC0879 (Raf inhibitor) or KT5720 (inhibitor of PKA and PKC). White circles csBEC GFP, gray circles csBEC α5WT, blue circles csBEC α5SNP). Results are expressed as the number of cells determined for each group at D7 (n = 4 per group). *p < 0.05; ***p < 0.001; for inhibitors vs control DMSO group for each genotype, two-way ANOVA. c Relative expression of TNF-α normalized to cyclophilin A (arbitrary units, A.U.) after in vitro scratch assay on csBEC from α5WT (gray circles) and α5SNP mice (blue circles) (n = 3 per group), 15 h after scratch. *p < 0.05 for α5SNP scratch vs control or inhibitors (H89, selective inhibitor of PKA; GF109203X, selective inhibitor of PKC; SQ22536, non-selective inhibitor of adenylyl cyclases); ^#p < 0.05 for WT scratch vs control or inhibitors, Mann–Whitney two-sided test. d Polarized localization of Ca²⁺-sensitive AC isoforms (all in red) in tracheal epithelia from α5WT. Scale bar: 40 μm. Micrographs representative of two independent experiments. e Relative expression of TNF-α normalized to cyclophilin A, on csBEC from α5WT (gray circles) and α5SNP (blue circles) (n = 3 per group), 15 h after scratch. Data are expressed as means ± SEM of three independent experiments (c, e), or mean with 95% CI of four independent experiments (b). Data for c, e were quantified using the 2^−ΔΔCT method⁸³ in log scale. f Scheme summarizing a mechanism of intracellular signaling in basal cells expressing α5SNP nAChR in response to epithelial stress.