Figure 6.

Recognition of Hp by COLEC12 in GSCs and its subsequent effect on DC-derived IL-23 expression. (a) Blockade of IL-23 expression in GSC-DC co-cultures by anti-COLEC12 blocking Abs (30 ug/ml). A trans-well co-culture system was set up as above in the absence or presence of anti-COLEC12 blocking Abs in the upper chamber, and the levels of IL-23 in the lower chamber was measured the same as in Fig. 1. (b) Levels of IL-23 secretion in the co-cultures with Hp wild-type strain (Hp) or a mutant strain of Hp lacking an α3-fucosyltransferase gene (FutB; HP0651). To evaluate the role of COLEC12 in GSCs in regulating PGE₂ expression, GSCs were cultured with (c) anti-COLEC12 blocking Abs (30 ug/ml) for 1 hr or (d) COLEC12 siRNA (50 nM) for 72 hrs before Hp treatment. Twenty-four hrs after Hp stimulation, supernatants were collected and PGE₂ expression was measured by ELISA. (e) As shown in Fig. 6d, GSCs cell pellet were collected and COLEC12 expression was assessed by q-PCR. (f) The glycan structure confers the recognition specificity of COLEC12 on GSCs. Blocking reagents, fucose (25 mM) or galactose (25 mM), were added in GSC cultures for 1 hr before Hp treatment. After 24 hrs, supernatants were collected for PGE₂ measurement by ELISA. The results are expressed as mean ± SE, and the data represent three to five independent experiments from different DC donors. In Fig. 6d and e, data were from analyses of GSCs from three gastric cancer patients (T2, T9 and T21). Mann-Whitney U-test was used and for pair-wise comparisons only. ANOVA followed Scheffe test of multiple post hoc analyses was used for q-PCR experiment. *p < 0.05, **p < 0.01, ***p < 0.001, NS; non significant.