FIGURE 5.
p40 promotes mucin production in the colonic epithelium of WT, but not Egfrwa5 mice. WT and Egfrwa5 mice were gavaged with 10 μg of p40 in two pectin/zein beads or two control pectin/zein beads without p40 for 5 days. A, colonic epithelial cells were isolated, and cellular soluble proteins were prepared for a PAS assay. The data are presented as μg of mucin per mg of cellular protein. *, p < 0.05 compared with the no-treatment group and control bead-treated group. B and C, Carnoy-fixed colon sections were immunostained using an anti-MUC2 antibody and a FITC-conjugated secondary antibody. The slides were mounted using mounting medium containing DAPI to stain nuclei and observed using fluorescence microscopy. FITC and DAPI images were taken from the same field. The thickness of the attached mucus layer in the proximal colon was determined by measuring the distance between the epithelial surface and the mucus surface in at least 20 fields for each mouse. D, RNA was prepared from colonic epithelial cells for real time PCR analysis of the Muc2 mRNA level. The Muc2 mRNA expression level in the no-treatment group of WT mice was set as 1, and mRNA expression levels in other groups were compared with that in this group. E and F, formalin-fixed colon sections were immunostained using an anti-MUC2 antibody and a HRP-conjugated secondary antibody. The slides were developed using 3, 3′-diaminobenzidine counterstained with hematoxylin, and observed using light microscopy. The number of MUC2-positive cells in the distal colonic tissues was determined by counting the absolute number of positive stained cells in at least 300 crypts for each mouse. *, p < 0.05 compared with the no-treatment group of WT mice (n = 5/group).