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. 2022 Jan 13;119(3):e2115230119. doi: 10.1073/pnas.2115230119

Fig. 4.

Fig. 4.

IL-22 regulates the amount of α-defensin in the intestinal lumen. (A, C, D, and E) α-defensin concentration in the feces of WT (n = 45) and IL-22–deficient mice (n = 20) (A), control mice (Fut2+/+ and Fut2+/− mice) (n = 10) and Fut2-deficient mice (n = 14) (C), WT mice (n = 15) and Fut1-deficient mice (n = 16) (D), and FMT mice that received WT feces (n = 5) or IL-22–deficient feces (n = 5) and WT feces (n = 5) or Fut2-deficient feces (n = 5) (E), as measured by ELISA. Data are presented as mean ± SD **P < 0.01, *P < 0.05, n.s., not significant, Student’s t test. (B) Sections of ileum from WT mice (n = 5) and Fut2-deficient mice (n = 5) were subjected to hematoxylin and eosin staining to detect Paneth cell granules. (Scale bars, 20 µm.) A total of 10 crypts per mouse were examined, and Paneth cell granule area was measured with the ImageJ software. Representative images are shown. Data are presented as mean ± SD **P < 0.01, Student’s t test. (F) α-defensin concentration in the intestinal organoid culture medium was measured by ELISA. Intestinal organoids derived from WT mice were treated with 1 ng/mL recombinant IL-22 for 2 d, and granule secretion was induced by 5 ng/mL recombinant IFN-γ. Four samples per group were analyzed. Data are presented as mean ± SD; ***P < 0.001, Student’s t test. (G) Intestinal organoids derived from WT mice were treated with 1 ng/mL recombinant IL-22 for 2 d. Paneth cell granule secretion was visualized and quantified as previously described (31). Percent granule secretion was calculated using Paneth cell granule area measured before and 10 min after treatment of the organoids with 0.1 µM carbamylcholine, a cholinergic agonist that stimulates secretion of α-defensin by Paneth cells. A total of 10 Paneth cells in five organoids from each subject were analyzed. Data were pooled from four independent experiments and are presented as mean ± SD; **P < 0.01, Student’s t test. (H) Ileal organoids derived from WT or Fut2-deficient mice were cultured in Matrigel. Percent granule secretion was calculated using Paneth cell granule area measured before and 10 min after treatment of the organoids with 0.1 µM carbamylcholine. Six Paneth cells in three organoids from each subject were analyzed. Data were pooled from three independent experiments and are presented as mean ± SD; *P < 0.05, Student’s t test.