Fig. 3
Loss of JAM-A on intestinal epithelial cells increases intestinal permeability and results in decreased recruitment of PMN in models of acute peritonitis as well as diminished production of CXCL1 by peritoneal macrophages that is dependent on the presence of gut microbiota. a JAM-A expression (green) in the ileum of Jam-afl/fl versus Villin-Cre+;Jam-afl/fl mice. Shown are representative images of immunostained tissue sections. Nuclei are stained with Dapi (blue). Asterisks represent crypt lumen. Arrows show vessels and immune cells in the sub-epithelial space (lamina propria). Scale bars are 100 μm. b Expression of JAM-A mRNA in intestinal epithelial cells and lamina propria-enriched fractions from ileal tissue of Jam-a fl/fl versus Villin-Cre+;Jam-afl/fl mice. Symbols represent individual mice from two independent experiments. Data are means ± SEM. ns; not significant. ***P < 0.001 by two-way ANOVA with a Bonferroni multiple comparison post hoc test. c Permeability to FITC-dextran (4 kDa) using an ileal loop in vivo model in Jam-afl/fl versus Villin-Cre+;Jam-afl/fl mice. Symbols represent individual mice and data are pooled from four independent experiments. Data are means ± SEM. **P < 0.01 by Student’s t-test. d Number of PMN in the peritoneal cavity of Villin-Cre+;Jam-afl/fl and control mice 2 h post-injection with zymosan or LPS by flow cytometry. Each data point represents individual mice from two independent experiments. Data represent means ± SEM. **P < 0.01, *P < 0.05 by Student’s t-test. Expression of CXCL1 by ELISA (e) or mRNA level (f) in peritoneal macrophages from Jam-afl/fl versus Villin-Cre+;Jam-afl/fl mice either treated or untreated ex vivo with zymosan for 1 h. Symbols represent individual mice and data are pooled from three independent experiments (e) or two independent experiments (f). Data are means ± SEM. ns; not significant. ***P < 0.001 by two-way ANOVA with a Bonferroni multiple comparison post hoc test. g Number of PMN in the peritoneal cavity of germ-free (GF) Villin-Cre+;Jam-afl/fl and control mice 2 h post-injection with zymosan or LPS by flow cytometry. Each data point represents an individual mouse from two independent experiments. Data are means ± SEM. ns; not significant by Student’s t-test. Expression of CXCL1 by ELISA (h) or mRNA level (i) in peritoneal macrophages from germ-free (GF) Jam-afl/fl versus Villin-Cre+;Jam-afl/fl mice, either treated or untreated ex vivo with zymosan for 1 h. Each data point represents an individual mouse from two independent experiments. Data are means ± SEM. ***P < 0.001 by two-way ANOVA with a Bonferroni multiple comparison post hoc test. ns; not significant. ANOVA analysis of variance, LPS lipopolysaccharide
