Increased Primary Cilia Contribute to Airway Epithelium Remodeling in Asthma

The data that support the findings of this study are available from the corresponding author upon reasonable request. Figure S1: Characterization of primary and motile cilia in asthmatic airway epithelium and IL-13-treated HBECs. (A) Immunofluorescent staining of ARL13B and FOXJ1 in bronchial biopsies from asthma patients. (B) Quantification of primary cilia in the epithelium of non-eosinophilic asthma patients (n = 6), eosinophilic asthma patients (n = 24), and control subjects (n = 15). Values indicate the number of primary cilia-positive cells per millimeter. (C) Representative images for immunofluorescent staining of FOXJ1 in normal human bronchial epithelial cells (HBECs) cultured at the air–liquid interface (ALI) under control conditions, IL-13 treatment, and IL-13 plus chloral hydrate (CH) treatment. (D) Quantification of FOXJ1 fluorescence intensity in HBECs. (E) The protein levels of FOXJ1 in control, IL-13, and IL-13 plus CH-treated HBECs cultured at ALI. **p < 0.01, ***p < 0.001. Figure S2: Supplemental analysis of epithelial barrier integrity following IL-13 treatment and intervention. (A) Representative images of PAS staining of NC siRNA plus IL-13, and IFT88 siRNA plus IL-13-treated normal human bronchial epithelial cells (HBECs) cultured at the air–liquid interface (ALI). (B) Quantification of PAS-positive cells in NC siRNA plus IL-13, and IFT88 siRNA plus IL-13-treated HBECs. (C) Representative transmission electron microscopy (TEM) images of HBECs cultured at ALI under control conditions, IL-13 treatment, and IL-13 plus chloral hydrate (CH) treatment. (D) Representative images for immunofluorescent staining of E-cadherin and ZO-1 in control, IL-13, CH, and IL-13 plus CH-treated HBECs at ALI. (E-F) Quantification of ZO-1 and E-cadherin positive area in HBECs. *p < 0.05, **p < 0.01, ***p < 0.001. Figure S3: IL-13 upregulates Hedgehog and Notch signaling, elimination of primary cilia suppresses the IL-13-induced as well as Hedgehog and Notch agonists-augmented IL-13-induced goblet cell metaplasia in vitro. (A) Representative images for immunofluorescent staining of NOTCH3 in normal human bronchial epithelial cells (HBECs) cultured at air–liquid interface (ALI) under control conditions, IL-13 treatment, and IL-13 plus chloral hydrate (CH) treatment. (B) Quantification of NOTCH3 fluorescence intensity in HBECs. (C) Representative images for immunofluorescent staining of SMO of HBECs cultured at ALI under control conditions, IL-13 treatment, and IL-13 plus chloral hydrate (CH) treatment. (D) Quantification of SMO fluorescence intensity in HBECs. (E) The protein levels of NICD3 and SMO in control, IL-13, and IL-13 plus CH-treated HBECs. (F) Representative images of PAS staining and MUC5AC immunostaining of HBECs cultured at ALI following treatment with IL-13, CH, Hedgehog signaling agonist SAG (Smoothened Agonist), and Notch signaling agonist Jagged-1. (G) Quantitative analysis of the percentage of PAS-positive area relative to the total area of HBECs. (H) Quantitative analysis of the percentage of MUC5AC-positive area relative to the total area of HBECs. *p < 0.05, **p < 0.01, ***p < 0.001. Table S1: all70309-sup-0003-Supinfo.docx. Please note: The publisher is not responsible for the content or functionality of any supporting information supplied by the authors. Any queries (other than missing content) should be directed to the corresponding author for the article.