Figure 3.

Interaction of HopQ with CEACAM is important for activation of the non-canonical NF-κB pathway. NUGC-4 and SNU1 cells were infected with P12 wild type H. pylori strain, or isogenic mutant strains, at an MOI of 10 for 8 h. (a) p100 to p52 processing was detected by Western blot. GAPDH served as a loading control and was used for normalization. (b) IL8 secretion was measured by ELISA. (c) A20 and (d) LTβ mRNA levels were analyzed by qPCR. GAPDH was used as a housekeeping gene. All qPCR values were normalized to uninfected controls and calculated using the 2^-ΔΔct equation. (e) Binding assay. CFSE-labeled bacteria were incubated with gastric cancer cells for 30 min with shaking. After washing and fixation, samples were analyzed for GFP signal on a Beckman Coulter Cytoflex flow cytometer. (f) Representative confocal images of H. pylori P12 wild type, P12ΔhopQ, P12ΔcagA and P12ΔcagE (green) binding to MKN45 and NUGC-4 cells. Nuclei were stained with DAPI (blue), and cell membranes were stained with phalloidin (red). Scale bar: 50 µm. Graphs show mean ± SD of at least three independent experiments. One-way ANOVA with Bonferroni correction was used to test differences between means. * p ≤ 0.05; ** p ≤ 0.01; *** p ≤ 0.001.