Fig. 5
From: MicroRNA-9 downregulates the ANO1 chloride channel and contributes to cystic fibrosis lung pathology
miR-9-specific TSB restores deficient parameters in primary CF cells. Primary hAECB and fully differentiated human bronchial air–liquid-interface cultures, isolated from bronchial biopsies from CF (F508del/F508del) patients were transfected with TSB control (control) or ANO1 TSB every day during 3 days. a Confocal microscopic analysis of fluorescein-conjugated TSB transfected into human bronchial cells isolated from CF patients (green). Cells were cultured in ALI and transfected for 24 h. b Isolated cells from ALI cultures transfected with fluorescein-conjugated TSB (green). The nuclei were stained with DAPI (blue), and merged images are shown. Scale bars 10 µm. c ANO1 protein expression was analyzed and quantified by western blotting using anti-ANO1 antibody. β-actin was used for normalization. Histograms represent average values ± SDs (n = 6) and were compared using Student’s t-test. d Representative and original traces of ANO1 channel activity (left) and quantification (right) of hAECB CF cells transfected with ANO1 TSB or a negative control (n = 4). e Migration rates during repair of primary CF cells. Representative photographs were taken during 4 h of wound closure of CF cells (left), and migration rates were quantification (right) (n = 4). Scale bar 20 µm. Histograms represent average values ± SDs and were compared using Student’s t-test. f Effect of TSB control or ANO1 TSB on mucus dynamics after 30 days of transfection. The movements of 100 beads were quantified for each condition, and the average speed (µm/ms) was determined. Scale bar 40 µm. Histograms represent the mean values ± SDs and were compared using Student’s t-test
