Figure 3.
ERAdP deficiency leads to abnormal lysozyme secretion. (A) Lysozyme immunohistochemistry of intestines from WT and Cnep1r1ΔIEC mice. Scale bar: 70 μm. (B and C) Whole-mount images of tissues from littermate WT and Cnep1r1ΔIEC mice taken from immediately above the ileal mucosal surface and stained. Red: lysozyme (B) and DEFA5 (C); green: WGA. Scale bar: 100 μm. (D) Schematic diagram of stimulation of small intestinal crypts by CCh and co-incubation of supernatant with S. Typhimurium. (E) Lysozyme in supernatant of stimulated crypts from littermate WT and Cnep1r1ΔIEC mice after treatment of DMSO or CCh was measured by ELISA (n = 6 mice for each group). (F) Bacterial load analysis of S. Typhimurium in secreted supernatants. SI crypts of littermate WT and Cnep1r1ΔIEC mice were isolated and stimulated by DMSO or CCh. The crypt-secreted supernatants were incubated with S. Typhimurium for 30 min, and CFUs were measured (n = 6 mice for each group). (G and H) Intestinal lumen (G) and tissue-associated (H) bacterial load analysis, quantified by qPCR of 16S rRNA gene copy numbers in distal ileums (n = 6 mice for each group). (I–L) Bacterial load analysis of livers (I), spleens (J), ileal contents (K) and PPs (L) of WT and Cnep1r1ΔIEC mice. 3 days after S. Typhimurium infection, livers, spleens, ileal contents, and PPs were collected, and CFUs were calculated (n = 6 mice for each group). (M) Body weight changes of littermate WT and Cnep1r1ΔIEC mice after infection with S. Typhimurium (n = 6 mice for each group). (N) Survival rates of littermate WT and Cnep1r1ΔIEC mice after infection with S. Typhimurium (n = 12 mice for each group). (O) Representative pathological manifestation of intestines from WT and Cnep1r1ΔIEC mice after infected by S. Typhimurium for 7 days. Scale bar: 80 μm. Data are shown as means ± SEM; significance was determined by unpaired two-tailed Student’s t test (E–M) and Mantel–Cox test (N) (*P < 0.05; **P < 0.01; ***P < 0.001; ns, not significant). Data in A–C and E–O are representative of at least three independent experiments.

ERAdP deficiency leads to abnormal lysozyme secretion. (A) Lysozyme immunohistochemistry of intestines from WT and Cnep1r1ΔIEC mice. Scale bar: 70 μm. (B and C) Whole-mount images of tissues from littermate WT and Cnep1r1ΔIEC mice taken from immediately above the ileal mucosal surface and stained. Red: lysozyme (B) and DEFA5 (C); green: WGA. Scale bar: 100 μm. (D) Schematic diagram of stimulation of small intestinal crypts by CCh and co-incubation of supernatant with S. Typhimurium. (E) Lysozyme in supernatant of stimulated crypts from littermate WT and Cnep1r1ΔIEC mice after treatment of DMSO or CCh was measured by ELISA (n = 6 mice for each group). (F) Bacterial load analysis of S. Typhimurium in secreted supernatants. SI crypts of littermate WT and Cnep1r1ΔIEC mice were isolated and stimulated by DMSO or CCh. The crypt-secreted supernatants were incubated with S. Typhimurium for 30 min, and CFUs were measured (n = 6 mice for each group). (G and H) Intestinal lumen (G) and tissue-associated (H) bacterial load analysis, quantified by qPCR of 16S rRNA gene copy numbers in distal ileums (n = 6 mice for each group). (I–L) Bacterial load analysis of livers (I), spleens (J), ileal contents (K) and PPs (L) of WT and Cnep1r1ΔIEC mice. 3 days after S. Typhimurium infection, livers, spleens, ileal contents, and PPs were collected, and CFUs were calculated (n = 6 mice for each group). (M) Body weight changes of littermate WT and Cnep1r1ΔIEC mice after infection with S. Typhimurium (n = 6 mice for each group). (N) Survival rates of littermate WT and Cnep1r1ΔIEC mice after infection with S. Typhimurium (n = 12 mice for each group). (O) Representative pathological manifestation of intestines from WT and Cnep1r1ΔIEC mice after infected by S. Typhimurium for 7 days. Scale bar: 80 μm. Data are shown as means ± SEM; significance was determined by unpaired two-tailed Student’s t test (E–M) and Mantel–Cox test (N) (*P < 0.05; **P < 0.01; ***P < 0.001; ns, not significant). Data in A–C and E–O are representative of at least three independent experiments.

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