Article Text
Abstract
BACKGROUND AND AIMS Helicobacter pylori infection induces expression of proinflammatory cytokines such as interleukin (IL)-8 and tumour necrosis factor α (TNF-α) in gastric mucosa, and their genes have AP-1 binding sites in the promoter region. c-Fos is important for transactivation of AP-1 which has SRE in the promoter region. We conducted this study to confirm H pylori induced transactivation of these binding sites.
METHODS Transactivation of SRE and AP-1 was evaluated in human gastric cancer cells TMK1 and MKN45 by luciferase reporter assay in transient transfection. We compared the effects of coculture with four H pylori strains, a cag pathogenicity island (PAI) positive strain TN2, its isogenicvacA negative (TN2-ΔvacA) orcagE negative (TN2-ΔcagE) mutants, and acag PAI negative clinical isolate T68. Phosphorylation of ERK1/2, JNK, and c-Jun was measured by immunoblot, induction of IL-8 secretion by ELISA, and the effects of MEK by inhibitor U0126.
RESULTS Both SRE and AP-1 were transactivated by coculture with TN2. Although TN2-ΔvacA induced comparable transactivation, TN2-ΔcagE and T68 showed decreased transactivation of SRE (65% and 51%) and AP-1 (71% and 54%, respectively, of TN2). Heat killed TN2 or indirect contact using a permeable membrane inhibited transactivation. Levels of phosphorylated ERK1/2, JNK, and c-Jun were increased by coculture with TN2. MEK inhibitor U0126 reduced TN2 induced transactivation of SRE and AP1, as well as secretion of IL-8, by 83%, 87%, and 53%, respectively, of TN2.
CONCLUSIONS Transactivation of SRE and AP-1, through ERK/MAPK and JNK/SAPK cascades, respectively, was found in gastric cancer cells cocultured with H pylori. Direct contact with viable bacteria possessing intactcag PAI is a prerequisite for the onset of intracellular signalling leading to AP-1 transactivation.
- Helicobacter pylori
- SRE
- AP-1
- cag pathogenicity island PAI
- gastric cancer
Abbreviations used in this paper
- IL
- interleukin
- PAI
- pathogenicity island
- NFκB
- nuclear factor κB
- EGF
- epidermal growth factor
- FBS
- fetal bovine serum
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Abbreviations used in this paper
- IL
- interleukin
- PAI
- pathogenicity island
- NFκB
- nuclear factor κB
- EGF
- epidermal growth factor
- FBS
- fetal bovine serum