FIG 3.

Compromised intestinal barrier and transport functions in villin-TLR4 mice promote bacterial dissemination. (a) Intestinal mucosal permeability was measured by 4-kDa FITC-dextran levels in the serum of villin-TLR4 and WT mice. Results represent the mean ± the standard error of the mean of fold changes in FITC-dextran levels relative to the WT (seven to nine mice per group). (b) Relative mRNA expression levels of epithelial barrier genes in the colonic ECs of villin-TLR4 and WT mice. Results represent the mean ± the standard error of the mean of relative mRNA expression normalized to GAPDH and β-actin (five mice per group). (c) Correlation between relative JAM-A mRNA expression levels and mucosal bacterial richness as determined by bacterial 16S rRNA gene sequencing (r = −0.77; P < 0.05). (d) Correlation between relative Cldn15 mRNA expression levels and bacterial invasion as determined by CFU counts in MLN cultures of villin-TLR4 and WT littermates (r = −0.75; P < 0.05). (e) Pearson correlation between epithelial barrier genes and markers of bacterial translocation (bacterial richness in mucosa and bacterial CFU counts in MLN and spleen tissues, respectively) showed a mostly negative correlation with cell-cell adhesion genes and a positive correlation with junctional paracellular transporters. The Pearson correlation coefficients (r values) of these variables were mapped by a one-matrix heat map with CIMminer. The scale bar, from left to right, represents negative correlations in red to positive correlations in green. Epithelial barrier genes were clustered by Euclidean distance. *, P < 0.05; **, P < 0.01; ***, P < 0.001 by two-tailed Student t test. (f, g) Western blot analysis of claudin-3 and occludin protein levels in colonic and small IECs of WT and villin-TLR4 littermate mice. β-Actin was used as a protein loading control for each Western blot gel.