Extended Data Fig. 7: STAT signaling in mouse colon organoids and epithelial cell line following cytokine stimulation.
From: The IL-22–oncostatin M axis promotes intestinal inflammation and tumorigenesis
(a) Immunofluorescence staining for pSTAT1 and pSTAT3 in steady-state (n = 7) and inflamed (n = 10) mouse samples with subsequent quantification in colon lamina propria. Data pooled from two experiments. P-values are from Mann-Whitney U tests. (b) Western blot analysis of pSTAT3, total STAT3, and β-actin in mouse colon organoids treated with the indicated cytokines. Data are representative of two independent experiments. (c) Western blot analysis of pSTAT1, total STAT1, and β-actin in mouse colon organoids treated with indicated cytokines. Data are representative of two independent experiments. (d) Western blot analysis of pSTAT1, total STAT1, and β-actin in the human Caco2 cell line (left panel) was performed to confirm functionality of pSTAT1 antibody. Epithelial cells from C57BL/6 J mice with induced inflammation at days 7 and 14 (n = 5) were also analyzed (right panel). pSTAT1, total STAT1, and β-actin in steady state epithelial cells from C57BL/6 J mice are shown in Fig. 4a. (e) Colon epithelial organoids isolated from C57BL/6 mice were treated with JAK/STAT pathway inhibitors (50 μM fludarabine, 25 μM STA-21, 2.5 μM ruxolitinib, or 2.5 μM tofacitinib) for 6 h, and 10 ng/mL of IL-22 was added for another 6 h. 0.1% DMSO was used as inhibitor control. Relative expression of Osmr was analyzed by qPCR. Data are shown for n = 2 biological replicates with n = 3 technical replicates. Graph displays mean ± SEM. P-values are derived from Kruskal-Wallis test.
