Figure 3.

Effect of CSP7 on CSE-induced MH and cilia dysfunction. A: Western blot images showing expression of MUC5AC, HDAC6, FOXA2, FOXA3, SPDEF, acetylated tubulin (Ac-Tub), CAV1, p53, PAI-1, and LC3BII in lysates of naïve AECs from human nL or nL AECs exposed to CSE left untreated (None) or treated with CSP7 or CP for 48 h in vitro. Same samples were analyzed for β-actin to assess equal loading. B: total RNA from nL AECs (n = 4) treated as in A tested for MUC5AC, HDAC6, FOXA2, FOXA3, and CAV1 mRNA expression by qPCR. Immunofluorescence staining of AECs from nL treated as Fig. 2A for colocalization of MUC5AC and HDAC6 (scale bar 500 µM; C) or Ac-Tub and LC3B (scale bar 500 µM and 200 µM; D). Representative image from two independent experiments was shown. Bar graph depicting number of ciliated cell (E) and cilia length (F) in AECs (n = 3) from nL treated as in Fig. 2. Each experiment was repeated at least two to three times, and data are presented as means + SD, and *P < 0.05, **P < 0.01, and ***P < 0.001 were obtained by one-way ANOVA with Tukey’s multiple comparison test and log-rank tests, respectively. AECs, airway epithelial cells; CAV1, caveolin‐1; CSE, cigarette smoke extract; CSP7, caveolin-1 scaffolding domain peptide; FOXA2, forkhead box protein A2; HDAC6, histone deacetylase 6; MH, mucus hypersecretion; MUCAC, mucin 5AC; nL, “normal” lung; PAI-1, plasminogen activator inhibitor-1; SPDEF, domain-containing E26 transformation-specific like factor.