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. 2019 Dec 23;51:102572. doi: 10.1016/j.ebiom.2019.11.033

Fig. 3.

Fig 3

HH pathway inhibition induced AEC remodelling. A, Representative immunoblots from Shh immunoprecipitations in AEC culture medium (n = 3) with control IgGM (0.1 µg/ml and 1 µg/ml) and AB5E1 (0.1 µg/ml and 1 µg/ml). B, Schema depicting AEC cultures in the ALI model and AB5E1 treatment in culture medium to investigate transcript levels, protein levels and localizations, and TEER during the process of differentiation. C, Examples of micrographs taken from AEC cultures at ALI-14 showing basal cells (p63, green). Nuclei are stained in blue (DAPI). D, Box and whiskers plot (median with IQR) represents the percentage of p63+ cells (n = 5) on total cell population (evaluated on DAPI-stained cells count) at ALI-7 and ALI-14 in normal condition (black) and AB5E1-treated cells (red). *p < 0.05 CTL vs AB5E1. E, Examples of micrographs taken from AEC cultures at ALI-28 showing cilia (Arl13b, red). Nuclei are stained in blue (DAPI). F, Box and whiskers plot (median with IQR) represents the relative mean grey values of cilia-associated fluorescence (n = 8) at ALI-28 and ALI-35 (the baseline is given by the average value obtained from ALI-21 in normal condition) in normal condition (black) and AB5E1-treated cells (red). *p < 0.05 CTL vs AB5E1. G, Examples of micrographs taken from AEC cultures at ALI-21 showing mucins (Muc5b, red; Muc5ac, green). Nuclei are stained in blue (DAPI). H, Box and whiskers plot (median with IQR) represents the relative mean grey values of the mucins associated fluorescence ratios Muc5ac/Muc5b (n = 6) at ALI-21, ALI-28 and ALI-35 (the baseline is given by the average value obtained from ALI-14 in normal condition for each mucin) in normal condition (black) and AB5E1-treated cells (red). *p < 0.05 CTL vs AB5E1. I, Box and whiskers plots (median with IQR) represent the TEER of AEC (n = 5) at ALI-35 in control condition (black) or treated with AB5E1 (red). *p < 0.05 CTL vs AB5E1. J, Schema summarizing Shh inhibition impact on cell differentiation. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)