Fig. 5.
Colitis-induced mouse model treated with GlcNAc showed increased branched N-glycosylation associated with suppression of T cell function. (A) L-PHA histochemistry and CD3 immunohistochemistry. L-PHA lectin reactivity showed an increased expression of β1,6-branched structures in the intestinal inflammatory infiltrate (positive to CD3) as well as an increase in mucus lining in mice treated with GlcNAc enemas (arrowheads). (Magnification: 63×.) (B) Immunoprecipitation (IP) of TCR followed by β1,6-GlcNAc branched N-glycan recognition in mouse colon, DSS (DSS-induced colitis) vs. DSS + GlcNAc treatment (Tx). WB, Western blot. (B1) Scatter plot: ratio of densities of L-PHA reactivity normalized to that of TCR depicted as the mean ± SEM comparing DSS (n = 2) mice with DSS + GlcNAc Tx (n = 3) mice. Student’s t test: *P ≤ 0.05. (C) TCR signaling by Western blot analysis of the phosphorylation levels of ZAP70 and LAT in LPLs. Values of pZAP70 densities normalized to tubulin are indicated. (D) Immunofluorescence of T-bet in colonic sections of DSS vs. DSS + GlcNAc Tx. (Insets) T-bet–expressing cells at intestinal inflammatory infiltrate are highlighted. (Magnification: 20×.) (E) Concentration of IFN-γ and IL-17A in the supernatants of 24-h colonic explant cultures from DSS and DSS + GlcNAc Tx MGAT5wt (n = 5) mice by ELISA. Plots depict the mean ± SEM of two to three animals per group. Student’s t test: **P ≤ 0.01. NS, not significant.