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IFN‐γ synergizes with LPS to induce nitric oxide biosynthesis through glycogen synthase kinase‐3‐inhibited IL‐10
Author(s) -
Lin ChiouFeng,
Tsai ChengChieh,
Huang WeiChing,
Wang ChiYun,
Tseng HsiangChi,
Wang Yi,
Kai JuiIn,
Wang SzuWen,
Cheng YiLin
Publication year - 2008
Publication title -
journal of cellular biochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.028
H-Index - 165
eISSN - 1097-4644
pISSN - 0730-2312
DOI - 10.1002/jcb.21868
Subject(s) - nitric oxide synthase , gsk 3 , downregulation and upregulation , glycogen synthase , gsk3b , nitric oxide , tyrosine phosphorylation , lipopolysaccharide , phosphorylation , tyrosine kinase , chemistry , microbiology and biotechnology , tumor necrosis factor alpha , biology , signal transduction , biochemistry , endocrinology , gene
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. J. Cell. Biochem. 105: 746–755, 2008. © 2008 Wiley‐Liss, Inc.