Fig. 5: LOX-1 was a cell surface protein mediating H. pylori adhesion.

a HFE145 cells expressing GFP-LOX-1 (green) protein were infected with H. pylori (MOI = 100) for 6 h, and cells were further stained to visualize H. pylori (red). The representative visual fields shown are representative of three independent experiments with similar results. Scale bar, 5 μm. b, c HFE145 cells transfected with siRNAs or overexpression plasmid of LOX-1 were infected with H. pylori (MOI = 100) for 6 h, followed by treating with or without 200 μg/ml gentamycin for 2 h. Cells were permeabilized with 1% saponin for 15 min. The diluted samples were then spread on H. pylori-selective blood agar plates and incubated for 5 days to count the colony number. H. pylori adhesion levels were calculated by subtracting the CFU of group with gentamycin treatment from the CFU of the group without (n = 3 replicates for each group). d The lysates of HFE145 cells transfected with negative control or WTAP siRNAs were incubated with soluble or insoluble proteins of H. pylori. A specific antibody was used to pull down LOX-1 and its interacting proteins from the incubated protein mixture. The pulled down proteins were separated in SDS-PAGE followed by silver staining. The red square indicates a protein band that was only observed in the LOX-1-pulled down groups incubated with H. pylori soluble proteins. Mass spectrum analysis revealed the specific protein band as H. pylori catalase. The gel shown is representative of three independent experiments with similar results. e HFE145 cell lysate was incubated with the soluble proteins of two H. pylori strains (TN2GF4 and ATCC 43504), and LOX-1 or catalase antibody was used to perform reciprocal co-immunoprecipitation assay. The predicted molecular size of catalase is 60 kDa, and the molecular size of LOX-1 is 50 kDa (mature form) and 32 kDa (precursor). The blots shown are representative of three independent experiments with similar results. f HFE145 cells were incubated with non-coated, catalase-coated, or BSA-coated fluorescent beads (green) for 6 h (MOI = 100), and cells were stained to visualize nuclei (blue). Ten visual fields of each group were randomly selected to count the attached and internalized fluorescent beads. g, h HFE145 cells transfected with siRNAs or overexpression vector of LOX-1 were incubated with catalase-coated fluorescent beads (MOI 100) for 6 h and were stained to visualize nuclei (blue). Ten visual fields of each group were randomly selected to count the attached and internalized fluorescent beads. Scale bar, 25 μm. i FITC-labeled H. pylori strains (wild-type, ΔkatA and ΔkatA + katA strains) were respectively incubated with human normal stomach tissue array (72 tissue cores from 24 cases (i.e., triplicate sections for each case), US Biomax, BN01011B). Each line represents the binding of the three H. pylori strains in an individual patient as measured by calculating the mean number of adhered H. pylori on triplicate sections, respectively. Quantitative analysis of the bacteria binding to different gastric areas (cardia tissues from 5 cases, gastric body tissues from 11 cases, and gastric antrum tissues from 8 cases) were performed, respectively. Scale bar, 75 μm. All the quantitative data were shown as means ± SD. Statistical analysis of the data was performed using unpaired two-tailed Student’s t test (c, h) or one-way ANOVA (b, f, g, i) followed by Tukey’s multiple comparison tests with adjustments, and the corresponding p-values are included in the figure panels. The statistical significance of the data (b–d, f–h) was calculated from one of three independent experiments with similar results. Source data are provided as a Source Data file.