IFN-γ synergizes with LPS to induce nitric oxide biosynthesis through glycogen synthase kinase-3-inhibited IL-10

Chiou Feng Lin, Cheng Chieh Tsai, Wei Ching Huang, Chi Yun Wang, Hsiang Chi Tseng, Yi Wang, Jui In Kai, Szu Wen Wang, Yi Lin Cheng

Research output: Contribution to journalArticle

33 Citations (Scopus)

Abstract

Interferon-γ (IFN-γ) plays a crucial role in innate immunity and inflammation. It causes the synergistic effect on endotoxin lipopolysaccharide (LPS)-stimulated inducible nitric oxide synthase (iNOS)/NO biosynthesis; however, the mechanism remains unclear. In the present study, we investigated the effects of glycogen synthase kinase-3 (GSK-3)-mediated inhibition of anti-inflammatory interleukin-10 (IL-10). We found, in LPS-stimulated macrophages, that IFN-γ increased iNOS expression and NO production in a time-dependent manner. In addition, ELISA analysis showed the upregulation of tumor necrosis factor-α and regulated on activation, normal T expressed and secreted, and the downregulation of IL-10. RT-PCR further showed changes in the IL-10 mRNA level as well. Treating cells with recombinant IL-10 showed a decrease in IFN-γ/LPS-induced iNOS/NO biosynthesis, whereas anti-IL-10 neutralizing antibodies enhanced this effect, suggesting that IL-10 acts in an anti-inflammatory role. GSK-3-inhibitor treatment blocked IFN-γ/LPS-induced iNOS/NO biosynthesis but upregulated IL-10 production. Inhibiting GSK-3 using short-interference RNA showed similar results. Additionally, treating cells with anti-IL-10 neutralizing antibodies blocked these effects. We further showed that inhibiting GSK-3 increased phosphorylation of transcription factor cyclic AMP response element binding protein. Inhibiting protein tyrosine kinase Pyk2, an upstream regulator of GSK-3β, caused inhibition on IFN-γ/LPS-induced GSK-3β phosphorylation at tyrosine 216 and iNOS/NO biosynthesis. Taken together, these findings reveal the involvement of GSK-3-inhibited IL-10 on the induction of iNOS/NO biosynthesis by IFN-γ synergized with LPS.

Original languageEnglish
Pages (from-to)746-755
Number of pages10
JournalJournal of Cellular Biochemistry
Volume105
Issue number3
DOIs
Publication statusPublished - Oct 15 2008
Externally publishedYes

Fingerprint

Glycogen Synthase Kinase 3
Biosynthesis
Interleukin-10
Interferons
Lipopolysaccharides
Nitric Oxide
Nitric Oxide Synthase Type II
Phosphorylation
Neutralizing Antibodies
Anti-Inflammatory Agents
Cyclic AMP Response Element-Binding Protein
Macrophages
RNA Interference
Innate Immunity
Endotoxins
Protein-Tyrosine Kinases
Tyrosine
Transcription Factors
Up-Regulation
Down-Regulation

Keywords

  • GSK-3
  • IFN-γ
  • IL-10
  • iNOS
  • LPS
  • Macrophage
  • NO

ASJC Scopus subject areas

  • Biochemistry
  • Cell Biology
  • Molecular Biology

Cite this

IFN-γ synergizes with LPS to induce nitric oxide biosynthesis through glycogen synthase kinase-3-inhibited IL-10. / Lin, Chiou Feng; Tsai, Cheng Chieh; Huang, Wei Ching; Wang, Chi Yun; Tseng, Hsiang Chi; Wang, Yi; Kai, Jui In; Wang, Szu Wen; Cheng, Yi Lin.

In: Journal of Cellular Biochemistry, Vol. 105, No. 3, 15.10.2008, p. 746-755.

Research output: Contribution to journalArticle

Lin, CF, Tsai, CC, Huang, WC, Wang, CY, Tseng, HC, Wang, Y, Kai, JI, Wang, SW & Cheng, YL 2008, 'IFN-γ synergizes with LPS to induce nitric oxide biosynthesis through glycogen synthase kinase-3-inhibited IL-10', Journal of Cellular Biochemistry, vol. 105, no. 3, pp. 746-755. https://doi.org/10.1002/jcb.21868
Lin, Chiou Feng ; Tsai, Cheng Chieh ; Huang, Wei Ching ; Wang, Chi Yun ; Tseng, Hsiang Chi ; Wang, Yi ; Kai, Jui In ; Wang, Szu Wen ; Cheng, Yi Lin. / IFN-γ synergizes with LPS to induce nitric oxide biosynthesis through glycogen synthase kinase-3-inhibited IL-10. In: Journal of Cellular Biochemistry. 2008 ; Vol. 105, No. 3. pp. 746-755.
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AU - Lin, Chiou Feng

AU - Tsai, Cheng Chieh

AU - Huang, Wei Ching

AU - Wang, Chi Yun

AU - Tseng, Hsiang Chi

AU - Wang, Yi

AU - Kai, Jui In

AU - Wang, Szu Wen

AU - Cheng, Yi Lin

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N2 - Interferon-γ (IFN-γ) plays a crucial role in innate immunity and inflammation. It causes the synergistic effect on endotoxin lipopolysaccharide (LPS)-stimulated inducible nitric oxide synthase (iNOS)/NO biosynthesis; however, the mechanism remains unclear. In the present study, we investigated the effects of glycogen synthase kinase-3 (GSK-3)-mediated inhibition of anti-inflammatory interleukin-10 (IL-10). We found, in LPS-stimulated macrophages, that IFN-γ increased iNOS expression and NO production in a time-dependent manner. In addition, ELISA analysis showed the upregulation of tumor necrosis factor-α and regulated on activation, normal T expressed and secreted, and the downregulation of IL-10. RT-PCR further showed changes in the IL-10 mRNA level as well. Treating cells with recombinant IL-10 showed a decrease in IFN-γ/LPS-induced iNOS/NO biosynthesis, whereas anti-IL-10 neutralizing antibodies enhanced this effect, suggesting that IL-10 acts in an anti-inflammatory role. GSK-3-inhibitor treatment blocked IFN-γ/LPS-induced iNOS/NO biosynthesis but upregulated IL-10 production. Inhibiting GSK-3 using short-interference RNA showed similar results. Additionally, treating cells with anti-IL-10 neutralizing antibodies blocked these effects. We further showed that inhibiting GSK-3 increased phosphorylation of transcription factor cyclic AMP response element binding protein. Inhibiting protein tyrosine kinase Pyk2, an upstream regulator of GSK-3β, caused inhibition on IFN-γ/LPS-induced GSK-3β phosphorylation at tyrosine 216 and iNOS/NO biosynthesis. Taken together, these findings reveal the involvement of GSK-3-inhibited IL-10 on the induction of iNOS/NO biosynthesis by IFN-γ synergized with LPS.

AB - Interferon-γ (IFN-γ) plays a crucial role in innate immunity and inflammation. It causes the synergistic effect on endotoxin lipopolysaccharide (LPS)-stimulated inducible nitric oxide synthase (iNOS)/NO biosynthesis; however, the mechanism remains unclear. In the present study, we investigated the effects of glycogen synthase kinase-3 (GSK-3)-mediated inhibition of anti-inflammatory interleukin-10 (IL-10). We found, in LPS-stimulated macrophages, that IFN-γ increased iNOS expression and NO production in a time-dependent manner. In addition, ELISA analysis showed the upregulation of tumor necrosis factor-α and regulated on activation, normal T expressed and secreted, and the downregulation of IL-10. RT-PCR further showed changes in the IL-10 mRNA level as well. Treating cells with recombinant IL-10 showed a decrease in IFN-γ/LPS-induced iNOS/NO biosynthesis, whereas anti-IL-10 neutralizing antibodies enhanced this effect, suggesting that IL-10 acts in an anti-inflammatory role. GSK-3-inhibitor treatment blocked IFN-γ/LPS-induced iNOS/NO biosynthesis but upregulated IL-10 production. Inhibiting GSK-3 using short-interference RNA showed similar results. Additionally, treating cells with anti-IL-10 neutralizing antibodies blocked these effects. We further showed that inhibiting GSK-3 increased phosphorylation of transcription factor cyclic AMP response element binding protein. Inhibiting protein tyrosine kinase Pyk2, an upstream regulator of GSK-3β, caused inhibition on IFN-γ/LPS-induced GSK-3β phosphorylation at tyrosine 216 and iNOS/NO biosynthesis. Taken together, these findings reveal the involvement of GSK-3-inhibited IL-10 on the induction of iNOS/NO biosynthesis by IFN-γ synergized with LPS.

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