Figure 3.

The ERK1/2 pathway plays an important role in V. parahaemolyticus infection-triggered AP-1 and NF-κB mediated IL-8 transcription. Activation of IL-8 transcription was estimated by luciferase reporter assay. Caco-2 cells were transfected with an IL-8 promoter (-133 to +44 bp within the 5’ flanking region) construct. A, After a 6- hour infection, gentamicin (100 μg/mL) was added to avoid excess infection. The cells were incubated for 12 hours, and cell extracts were prepared for determination of luciferase activity. Statistical significance: *, P < .05; **, P < .01 compared with untreated (+)VP. B, Mutant (mut) vectors containing AP-1, C/EBP1, or NF-κB binding sites were transfected into Caco-2 cells, which were infected with V. parahaemolyticus (black bar) or left uninfected (white bar) for 6 hours. The next day, the cells were lysed and luciferase activity was measured. The results are presented as the relative increase in activation (n-fold) of control cells (transfected with standard construct). Luciferase activity was normalized against β-galactosidase activity. The results are presented as the relative increase in active control cells (n-%). Data are means ± SD of 3 independent experiments with assays in duplicate. Statistical significance: *, P < .05; **, P < .01 compared with untreated -133 (+)VP.