The green tea polyphenol EGCG blocks H. pylori-induced scattering of gastric carcinoma cells, a process dependent on the activation of PI3K, p38 MAPK, and Rac1 pathways

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Helicobacter pylori has been implicated in gastric cancer, although the signaling pathways altered by this bacterium remain to be determined. Virulent strains (Type I) inject the cytotoxin-associated gene (Cag) A protein into gastric epithelial cells, causing them to become motile and scatter, a change important for invasive tumor cells. In order to determine which signaling components were involved in H. pylori-induced scattering, phospho-specific antibodies were used to probe western blots of lysates of AGS cells exposed to Type I H. pylori. As early as 30 minutes following exposure to bacteria, the levels of phosphorylated Erk, JNK, Akt, and p38 significantly increased. Additionally, induced levels of Erk-P, JNK-P, and Akt-P were sustained for 12 hours. Next, a pharmacological approach was used to determine which pathways were necessary for H. pylori-induced scattering. AGS cells pretreated with the inhibitors LY294002 or SB203580 prior to the addition of Type I H. pylori did not scatter, indicating that PI3-kinase and p38 MAP kinase pathways are necessary for this event to occur. Inhibition of the MAPK pathway using U0126 did not prevent bacterially-induced scattering.To determine if Rho family GTPases regulated scattering, AGS cells were transiently transfected with dominant negative (DN) or constitutively active (CA) forms of RhoA, Rac1, and Cdc42. Only expression of DN-Rac1 prevented H. pylori-induced scattering, demonstrating that this GTPase was necessary for scattering, although expression of CA-Rac1 was not sufficient to induce scattering in the absence of bacteria. The Met receptor has previously been shown to play an important regulatory role in tumor progression and metastasis. Exposure of AGS cells to Type I H. pylori induced Met phosphorylation by 30 minutes after the addition of bacteria, and high levels of phosphorylated Met were observed for at least 12 hours. In addition, the level of Met protein increased, beginning 6 hours after the addition of bacteria. Met phosphorylation is associated with scattering and invasion of other cancer cells, including DU145 prostate cancer cells. Interestingly, DU145 cells were induced to scatter by H. pylori in a protein synthesis-independent manner, while the Met ligand, HGF, induced scattering dependent on protein synthesis; HGF does not induce scattering in AGS cells. These results suggest that H. pylori- and HGF-induced scattering occur through distinct pathways. Finally, a large number of animal and epithelial studies support a role for the green tea polyphenol epigallocatechin 3-gallate (EGCG) in the prevention of cancer progression. Pretreatment of AGS cells with physiologically relevant levels of EGCG blocked H. pylori-induced scattering. Current studies are designed to study the nature of the bacterially induced pathways contributing to scattering inhibited by EGCG.