Identification of a signaling cascade for interleukin-8 production by Helicobacter pylori in human gastric epithelial cells

Biochem Pharmacol. 2002 Jul 1;64(1):21-30. doi: 10.1016/s0006-2952(02)01030-4.


Infecting gastric epithelial cells with Helicobacter pylori (H. pylori) has been shown to induce interleukin-8 (IL-8) production, but the signal transduction mechanism leading to IL-8 production is not defined clearly. In the present study, we investigated the molecular mechanism responsible for H. pylori-induced IL-8 release in human gastric epithelial cells. IL-8 levels in culture supernatants were determined by an enzyme linked-immunosorbent assay. Extracellular signal-regulated kinase (ERK) activity was tested using an in vitro kinase assay, which measured the incorporation of [gamma-33P]ATP into a synthetic peptide that is a specific ERK substrate. ERK phosphorylation and IkappaBalpha degradation by H. pylori infection were assessed by western blotting. In MKN45 cells, H. pylori-induced IL-8 release in a time-dependent manner. This IL-8 release was abolished by treatment with intracellular Ca2+ chelators (BAPTA-AM and TMB-8) but not by EGTA or nifedipine. The Ca2+ ionophore A23187 also induced IL-8 release to an extent similar to that of H. pylori infection. Calmodulin inhibitors (W7 and calmidazolium) and tyrosine kinase inhibitors (genistein and ST638) completely blocked IL-8 release by H. pylori and A23187. PD98059, an ERK pathway inhibitor, completely abolished H. pylori-induced IL-8 release. Moreover, BAPTA-AM, calmidazolium, and genistein, but not nifedipine, suppressed the ERK activation induced by H. pylori infection. PD98059 as well as MG132, an NF-kappaB pathway inhibitor, blocked both IL-8 production and degradation of IkappaBalpha induced by H. pylori infection, whereas only PD98059 inhibited ERK activity in response to H. pylori. There was no significant difference between IL-8 production induced by the cagA positive wild-type strain and the cagA negative isogenic mutant strain of H. pylori; therefore, CagA is not involved in the IL-8 production pathway. H. pylori-induced IL-8 production is dominantly regulated by Ca2+/calmodulin signaling, and ERK plays an important role in signal transmission for the efficient activation of H. pylori-induced NF-kappaB activity, resulting in IL-8 production.

MeSH terms

  • Bacterial Adhesion / drug effects
  • Calcium / metabolism
  • Calmodulin / metabolism
  • Cells, Cultured
  • Egtazic Acid / analogs & derivatives*
  • Egtazic Acid / pharmacology
  • Helicobacter pylori / metabolism*
  • Humans
  • Interleukin-1 / pharmacology
  • Interleukin-8 / biosynthesis*
  • Intestinal Mucosa / microbiology*
  • Mitogen-Activated Protein Kinases / metabolism
  • NF-kappa B / metabolism
  • Protein-Tyrosine Kinases / metabolism
  • Signal Transduction / physiology*
  • Tumor Necrosis Factor-alpha / pharmacology


  • Calmodulin
  • Interleukin-1
  • Interleukin-8
  • NF-kappa B
  • Tumor Necrosis Factor-alpha
  • 1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid acetoxymethyl ester
  • Egtazic Acid
  • Protein-Tyrosine Kinases
  • Mitogen-Activated Protein Kinases
  • Calcium