Effects of Ginkgo biloba extract (EGb 761) and quercetin on lipopolysaccharide-induced signaling pathways involved in the release of tumor necrosis factor-alpha.
Administration of bacterial lipopolysaccharide (LPS) to laboratory animals and cultured macrophages induces tumor necrosis factor-alpha (TNF-alpha), a pro-inflammatory cytokine. Pretreatment with Ginkgo biloba extract (EGb 761) inhibited the in vivo production of TNF-alpha (measured by ELISA) after challenge with LPS. To begin to understand the mechanism of this inhibition, we evaluated the in vitro effects of EGb 761 and its flavonoid component, quercetin, on LPS-treated RAW 264.7 macrophages. Pretreatment with EGb 761 or quercetin concentration-dependently inhibited TNF-alpha release, as measured by the L929 fibroblast assay. Northern blotting demonstrated that quercetin inhibited LPS-induced TNF-alpha mRNA, but did not alter its half-life. Activation of mitogen-activated protein kinases (MAPKs) and the redox-sensitive transcription factors, nuclear factor-kappaB (NF-kappaB) and activator protein 1 (AP-1), are key events in the signal transduction pathways mediating TNF-alpha induction. Phosphorylation of extracellular signal-related kinases 1 and 2 (ERK 1/2), p38 MAPK, and Jun N-terminal kinase/stress-activated protein kinase (JNK/SAPK), members of the MAPK family, was analyzed by western blotting. Our results suggest that quercetin is unique in its ability to inhibit TNF-alpha transcription by inhibiting the phosphorylation and activation of JNK/SAPK and, therefore, suppressing AP-1-DNA binding [assessed by electrophoretic mobility shift analysis (EMSA)]. Results from western analysis, EMSA, and transient transfections suggest that EGb 761 diminishes LPS-induced NF-kappaB but has no effect on LPS-induced TNF-alpha transcription. Both EGb 761 and quercetin inhibited ERK1/2 phosphorylation and p38 MAPK activity, which are important in the post-transcriptional regulation of TNF-alpha mRNA.
Wadsworth
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Department of Physiology and Pharmacology, Oregon Health Sciences University, 3181 S.W. Sam Jackson Park Road, Portland, OR 97201-3098, USA.
McDonald
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Koop
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D051696:Activating Transcription Factors; D000818:Animals; D000975:Antioxidants / Q000494:pharmacology*; D001798:Blood Proteins / Q000378:metabolism; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism; D002478:Cells, Cultured; D004247:DNA / Q000187:drug effects / Q000378:metabolism; D004347:Drug Interactions; D005419:Flavonoids / Q000494:pharmacology*; D020441:Ginkgo biloba / Q000737:chemistry; D020672:I-kappa B Proteins / Q000378:metabolism; D048031:JNK Mitogen-Activated Protein Kinases; D008070:Lipopolysaccharides / Q000494:pharmacology; D048670:MAP Kinase Kinase 4; D008264:Macrophages / Q000187:drug effects / Q000378:metabolism; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D019950:Mitogen-Activated Protein Kinase 1 / Q000187:drug effects / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3; D020929:Mitogen-Activated Protein Kinase Kinases / Q000378:metabolism; D020928:Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000187:drug effects / Q000378:metabolism; D016328:NF-kappa B / Q000378:metabolism; D010936:Plant Extracts; D010946:Plants, Medicinal; D011794:Quercetin / Q000494:pharmacology*; D012702:Serotonin Antagonists / Q000494:pharmacology; D015398:Signal Transduction / Q000187:drug effects*; D018808:Transcription Factor AP-1 / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects; D014409:Tumor Necrosis Factor-alpha / Q000235:genetics / Q000378:metabolism*; D048051:p38 Mitogen-Activated Protein Kinases; D000096983:Ginkgo Extract
D051696:Activating Transcription Factors; D000975:Antioxidants; D001798:Blood Proteins; D004247:DNA; D005419:Flavonoids; D020672:I-kappa B Proteins; D048031:JNK Mitogen-Activated Protein Kinases; D008070:Lipopolysaccharides; D048670:MAP Kinase Kinase 4; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020929:Mitogen-Activated Protein Kinase Kinases; D020928:Mitogen-Activated Protein Kinases; D016328:NF-kappa B; D010936:Plant Extracts; D011794:Quercetin; D012702:Serotonin Antagonists; D018808:Transcription Factor AP-1; D014157:Transcription Factors; D014409:Tumor Necrosis Factor-alpha; D048051:p38 Mitogen-Activated Protein Kinases; C063170:Ginkgo biloba extract; D000096983:Ginkgo Extract