Molecular characterization of Helicobacter pylori VacA induction of IL-8 in U937 cells reveals a prominent role for p38MAPK in activating transcription factor-2, cAMP response element binding protein, and NF-kappaB activation.
Helicobacter pylori VacA induces multiple effects on susceptible cells, including vacuolation, mitochondrial damage, inhibition of cell growth, and enhanced cyclooxygenase-2 expression. To assess the ability of H. pylori to modulate the production of inflammatory mediators, we examined the mechanisms by which VacA enhanced IL-8 production by promonocytic U937 cells, which demonstrated the greatest VacA-induced IL-8 release of the cells tested. Inhibitors of p38 MAPK (SB203580), ERK1/2 (PD98059), IkappaBalpha ((E)-3-(4-methylphenylsulfonyl)-2-propenenitrile), Ca(2+) entry (SKF96365), and intracellular Ca(2+) channels (dantrolene) blocked VacA-induced IL-8 production. Furthermore, an intracellular Ca(2+) chelator (BAPTA-AM), which inhibited VacA-activated p38 MAPK, caused a dose-dependent reduction in VacA-induced IL-8 secretion by U937 cells, implying a role for intracellular Ca(2+) in mediating activation of MAPK and the canonical NF-kappaB pathway. VacA stimulated translocation of NF-kappaBp65 to the nucleus, consistent with enhancement of IL-8 expression by activation of the NF-kappaB pathway. In addition, small interfering RNA of activating transcription factor (ATF)-2 or CREB, which is a p38MAPK substrate and binds to the AP-1 site of the IL-8 promoter, inhibited VacA-induced IL-8 production. VacA activated an IL-8 promoter containing an NF-IL-6 site, but not a mutated AP-1 or NF-kappaB site, suggesting direct involvement of the ATF-2/CREB binding region or NF-kappaB-binding regions in VacA-induced IL-8 promoter activation. Thus, in U937 cells, VacA directly increases IL-8 production by activation of the p38 MAPK via intracellular Ca(2+) release, leading to activation of the transcription factors, ATF-2, CREB, and NF-kappaB.
Hisatsune
Junzo
J
Department of Bacteriology, Institute of Tropical Medicine, Nagasaki University, Nagasaki, Japan.
Nakayama
Masaaki
M
Isomoto
Hajime
H
Kurazono
Hisao
H
Mukaida
Naofumi
N
Mukhopadhyay
Asish K
AK
Azuma
Takeshi
T
Yamaoka
Yoshio
Y
Sap
Jan
J
Yamasaki
Eiki
E
Yahiro
Kinnosuke
K
Moss
Joel
J
Hirayama
Toshiya
T
D051698:Activating Transcription Factor 2 / Q000378:metabolism*; D021581:Active Transport, Cell Nucleus; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D002083:Butylated Hydroxyanisole / Q000494:pharmacology; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism; D017362:Cyclic AMP Response Element-Binding Protein / Q000378:metabolism*; D003620:Dantrolene / Q000494:pharmacology; D004533:Egtazic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D004789:Enzyme Activation / Q000187:drug effects; D016480:Helicobacter pylori / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology; D016209:Interleukin-8 / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism; D016328:NF-kappa B / Q000378:metabolism*; D009570:Nitriles / Q000494:pharmacology; D010766:Phosphorylation / Q000187:drug effects; D011401:Promoter Regions, Genetic / Q000235:genetics; D013450:Sulfones / Q000494:pharmacology; D019284:Thapsigargin / Q000494:pharmacology; D020298:U937 Cells; D015854:Up-Regulation; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism*
C434003:3-(4-methylphenylsulfonyl)-2-propenenitrile; C497105:ATF2 protein, human; D051698:Activating Transcription Factor 2; D001426:Bacterial Proteins; D017362:Cyclic AMP Response Element-Binding Protein; D007093:Imidazoles; D016209:Interleukin-8; D016328:NF-kappa B; D009570:Nitriles; D013450:Sulfones; C074689:VacA protein, Helicobacter pylori; C070379:1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid acetoxymethyl ester; D002083:Butylated Hydroxyanisole; D004533:Egtazic Acid; D019284:Thapsigargin; D048051:p38 Mitogen-Activated Protein Kinases; D003620:Dantrolene; C063159:1-(2-(3-(4-methoxyphenyl)propoxy)-4-methoxyphenylethyl)-1H-imidazole