Fig. 5

HM-induced IL-6 production and secretion through TLR4 signaling and COX-2 generation by the gastric AGS cells. A Representative Western blot showing the optimization of the concentration of the TLR4 signaling pharmacological inhibitor TAK242 blocking TLR4 signaling, based on the loss of phosphorylation of p65 NFκB, its main downstream target. α-Tubulin was used as a loading control. Bar graph indicates the relative expression of phospho-p65 NFκB, calculated as a ratio of the expression of p65 NFκB. B Optimization of the concentration of the COX-2 pharmacological inhibitor for the blockade of COX-2 production, based on the decrease of COX-2 expression. Bar graph indicates the relative expression of COX-2, calculated as a ratio of the expression of α-Tubulin. C Representative Western blot and bar graph showing and summarizing the impact of the blockade of TLR4 signaling (using TAK242) and of COX-2 inhibitor (NS-398) on LPS- and HM-induced COX-2 and IL-6 production in the AGS cells after 72 h incubation, as compared with LPS- and HM-induced COX-2 and IL-6 production in AGS cells-pretreated with DMSO (solvent used for TAK242 and NS-398 reconstitution). D Bar graph showing the impact of the blockade of TLR4 signaling (using TAK242) and of COX-2 inhibitor (NS-398) on the LPS- and HM-induced secreted PGE2 and IL-6 released by AGS cells after 72 h incubation, detected in the conditional media using an ELISA. ^*p < 0.05, ^**p < 0.01, and ^***p < 0.001 compared with the control, from three independent experiments