1.Jak1/Stat3 Is an Upstream Signaling of NF-kappaB Activation in Helicobacter pylori-Induced IL-8 Production in Gastric Epithelial AGS Cells.
Boram CHA ; Joo Weon LIM ; Hyeyoung KIM
Yonsei Medical Journal 2015;56(3):862-866
Helicobacter pylori (H. pylori) induces the activation of nuclear factor-kB (NF-kappaB) and cytokine expression in gastric epithelial cells. The Janus kinase/signal transducers and activators of transcription (Jak/Stat) cascade is the inflammatory signaling in various cells. The purpose of the present study is to determine whether H. pylori-induced activation of NF-kappaB and the expression of interleukin-8 (IL-8) are mediated by the activation of Jak1/Stat3 in gastric epithelial (AGS) cells. Thus, gastric epithelial AGS cells were infected with H. pylori in Korean isolates (HP99) at bacterium/cell ratio of 300:1, and the level of IL-8 in the medium was determined by enzyme-linked immonosorbent assay. Phospho-specific and total forms of Jak1/Stat3 and IkappaBalpha were assessed by Western blot analysis, and NF-kappaB activation was determined by electrophoretic mobility shift assay. The results showed that H. pylori induced the activation of Jak1/Stat3 and IL-8 production, which was inhibited by a Jak/Stat3 specific inhibitor AG490 in AGS cells in a dose-dependent manner. H. pylori-induced activation of NF-kappaB, determined by phosphorylation of IkappaBalpha and NF-kappaB-DNA binding activity, were inhibited by AG490. In conclusion, Jak1/Stat3 activation may mediate the activation of NF-kappaB and the expression of IL-8 in H. pylori-infected AGS cells. Inhibition of Jak1/Stat3 may be beneficial for the treatment of H. pylori-induced gastric inflammation, since the activation of NF-kappaB is inhibited and inflammatory cytokine expression is suppressed.
Blotting, Western
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DNA, Bacterial/analysis/genetics
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Epithelial Cells/metabolism
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Gastric Mucosa/drug effects/*immunology/microbiology
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Gene Expression Regulation/drug effects/*immunology
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Gene Expression Regulation, Bacterial
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Helicobacter Infections/immunology/*metabolism
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Helicobacter pylori/genetics/pathogenicity/*physiology
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Humans
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Interleukin-8/genetics/*metabolism
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Janus Kinase 1
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NF-kappa B/biosynthesis/*metabolism
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Phosphorylation
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RNA, Messenger/metabolism
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STAT3 Transcription Factor
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Signal Transduction/genetics
2.Jak1/Stat3 Is an Upstream Signaling of NF-kappaB Activation in Helicobacter pylori-Induced IL-8 Production in Gastric Epithelial AGS Cells.
Boram CHA ; Joo Weon LIM ; Hyeyoung KIM
Yonsei Medical Journal 2015;56(3):862-866
Helicobacter pylori (H. pylori) induces the activation of nuclear factor-kB (NF-kappaB) and cytokine expression in gastric epithelial cells. The Janus kinase/signal transducers and activators of transcription (Jak/Stat) cascade is the inflammatory signaling in various cells. The purpose of the present study is to determine whether H. pylori-induced activation of NF-kappaB and the expression of interleukin-8 (IL-8) are mediated by the activation of Jak1/Stat3 in gastric epithelial (AGS) cells. Thus, gastric epithelial AGS cells were infected with H. pylori in Korean isolates (HP99) at bacterium/cell ratio of 300:1, and the level of IL-8 in the medium was determined by enzyme-linked immonosorbent assay. Phospho-specific and total forms of Jak1/Stat3 and IkappaBalpha were assessed by Western blot analysis, and NF-kappaB activation was determined by electrophoretic mobility shift assay. The results showed that H. pylori induced the activation of Jak1/Stat3 and IL-8 production, which was inhibited by a Jak/Stat3 specific inhibitor AG490 in AGS cells in a dose-dependent manner. H. pylori-induced activation of NF-kappaB, determined by phosphorylation of IkappaBalpha and NF-kappaB-DNA binding activity, were inhibited by AG490. In conclusion, Jak1/Stat3 activation may mediate the activation of NF-kappaB and the expression of IL-8 in H. pylori-infected AGS cells. Inhibition of Jak1/Stat3 may be beneficial for the treatment of H. pylori-induced gastric inflammation, since the activation of NF-kappaB is inhibited and inflammatory cytokine expression is suppressed.
Blotting, Western
;
DNA, Bacterial/analysis/genetics
;
Epithelial Cells/metabolism
;
Gastric Mucosa/drug effects/*immunology/microbiology
;
Gene Expression Regulation/drug effects/*immunology
;
Gene Expression Regulation, Bacterial
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Helicobacter Infections/immunology/*metabolism
;
Helicobacter pylori/genetics/pathogenicity/*physiology
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Humans
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Interleukin-8/genetics/*metabolism
;
Janus Kinase 1
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NF-kappa B/biosynthesis/*metabolism
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Phosphorylation
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RNA, Messenger/metabolism
;
STAT3 Transcription Factor
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Signal Transduction/genetics
3.alpha-Lipoic Acid Inhibits Expression of IL-8 by Suppressing Activation of MAPK, Jak/Stat, and NF-kappaB in H. pylori-Infected Gastric Epithelial AGS Cells.
Ji Hyun CHOI ; Soon Ok CHO ; Hyeyoung KIM
Yonsei Medical Journal 2016;57(1):260-264
The epithelial cytokine response, associated with reactive oxygen species (ROS), is important in Helicobacter pylori (H. pylori)-induced inflammation. H. pylori induces the production of ROS, which may be involved in the activation of mitogen-activated protein kinases (MAPK), janus kinase/signal transducers and activators of transcription (Jak/Stat), and oxidant-sensitive transcription factor, nuclear factor kappa-light-chain-enhancer of activated B cells (NF-kappaB), and thus, expression of interleukin-8 (IL-8) in gastric epithelial cells. alpha-lipoic acid, a naturally occurring thiol compound, is a potential antioxidant. It shows beneficial effects in treatment of oxidant-associated diseases including diabetes. The present study is purposed to investigate whether alpha-lipoic acid inhibits expression of inflammatory cytokine IL-8 by suppressing activation of MAPK, Jak/Stat, and NF-kappaB in H. pylori-infected gastric epithelial cells. Gastric epithelial AGS cells were pretreated with or without alpha-lipoic acid for 2 h and infected with H. pylori in a Korean isolate (HP99) at a ratio of 300:1. IL-8 mRNA expression was analyzed by RT-PCR analysis. IL-8 levels in the medium were determined by enzyme-linked immunosorbent assay. NF-kappaB-DNA binding activity was determined by electrophoretic mobility shift assay. Phospho-specific and total forms of MAPK and Jak/Stat were assessed by Western blot analysis. ROS levels were determined using dichlorofluorescein fluorescence. As a result, H. pylori induced increases in ROS levels, mRNA, and protein levels of IL-8, as well as the activation of MAPK [extracellular signal-regulated kinase 1/2 (ERK1/2), c-Jun NH2-terminal kinase 1/2 (JNK1/2), p38], Jak/Stat (Jak1/2, Stat3), and NF-kappaB in AGS cells, which was inhibited by alpha-lipoic acid. In conclusion, alpha-lipoic acid may be beneficial for prevention and/or treatment of H. pylori infection-associated gastric inflammation.
Enzyme-Linked Immunosorbent Assay
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Epithelial Cells/metabolism
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Gastric Mucosa/*drug effects/metabolism/microbiology
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Gene Expression Regulation, Bacterial
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Helicobacter Infections/immunology/*metabolism
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Helicobacter pylori/drug effects/*pathogenicity
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Humans
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Interleukin-8/genetics/*metabolism
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JNK Mitogen-Activated Protein Kinases
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Janus Kinase 1
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Mitogen-Activated Protein Kinases/*biosynthesis
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NF-kappa B/*metabolism
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RNA, Messenger/isolation & purification/metabolism
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Reactive Oxygen Species/metabolism
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STAT3 Transcription Factor
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Stomach/metabolism/*microbiology
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Thioctic Acid/*pharmacology