117 Dysregulated cross talk between alveolar epithelial cells and stromal cells in IPF reduces epithelial regenerative capacity to isolated EpCAM+ cells (Supplementary Figure 2), suggesting the presence of a supportive cell population. Interestingly, when unfractionated organoid cultures were compared between the groups, we observed that significantly less organoids were formed from IPF lung-derived cells compared to non-IPF lung-derived cells (Figure 2A). In addition to the number of organoids formed, the size distribution of the organoids was compared between the groups. Notably, IPF-derived cells formed significantly larger organoids compared to the non-IPF group, for both unfractionated (non-IPF organoids mean size 78.45 ± 8.00 µm, IPF organoids mean size 92.70 ± 26.83 µm) and EpCAM+ (non-IPF organoids mean size 64.45 ± 1.47 µm, IPF organoids mean size 82.51 ± 14.42 µm) cultures . (Figure 2B). Figure 2: Abnormalities in organoid formation of unfractionated lung cell suspensions from IPF patients. A) Quantification of organoid numbers at day 7 comparing non-IPF (n=3) and IPF (n=4) unfractionated and EpCAM+ cultures. Organoid counts were normalized to the number of EpCAM+ cells to correct for differences in epithelial cell input during the organoid assay. Means ± SD are indicated. One-way ANOVA with Šídák’s multiple comparisons test was used to assess for statistical differences after testing the data for normality with Q-Q plots and Shapiro-Wilk test. B) Quantification of organoid size distribution at day 7 comparing non-IPF (n=3) and IPF (n=4) unfractionated and EpCAM+ cultures. The Kruskal–Wallis test was used to assess for statistical differences. 5
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