Inhibition of lipopolysaccharide-inducible nitric oxide synthase, TNF-alpha and COX-2 expression by sauchinone effects on I-kappaBalpha phosphorylation, C/EBP and AP-1 activation.
1. Sauchinone, a lignan isolated from Saururus chinensis (Saururaceae), is a diastereomeric lignan with cytoprotective and antioxidant activities in cultured hepatocytes. The effects of sauchinone on the inducible nitric oxide synthase (iNOS), tumor necrosis factor-alpha (TNF-alpha) and cyclooxygenase 2 (COX-2) gene expression and on the activation of transcription factors, nuclear factor-kappaB (NF-kappaB), CCAAT/enhancer-binding protein (C/EBP), activator protein-1 (AP-1) and cAMP-response element-binding protein (CREB) were determined in Raw264.7 cells as part of the studies on its anti-inflammatory effects. 2. Expression of the iNOS, TNF-alpha and COX-2 genes was assessed by Northern and Western blot analyses. NO production was monitored by chemiluminescence detection using a NO analyzer. To identify the transcriptional factors affected by sauchinone, the extents of NF-kappaB, C/EBP, AP-1 and CREB activation were measured. Activation of the transcription factors was monitored by gel mobility shift assay, whereas p65 and I-kappaBalpha were analyzed by immunocytochemical and immunoblot analyses. 3. Sauchinone inhibited the induction of iNOS, TNF-alpha and COX-2 by lipopolysaccharide (LPS) (IC50</=10 micro M) with suppression of the mRNAs. 4. Sauchinone (1-30 micro M) inhibited LPS-inducible nuclear NF-kappaB activation and nuclear translocation of p65, which was accompanied by inhibition of I-kappaBalpha phosphorylation. 5. LPS-inducible increase in the intensity of C/EBP binding to its consensus sequence was also inhibited by sauchinone. The AP-1, but not CREB, DNA binding activity was weakly inhibited by sauchinone. 6. These results demonstrate that sauchinone inhibits LPS-inducible iNOS, TNF-alpha and COX-2 expression in macrophages through suppression of I-kappaBalpha phosphorylation and p65 nuclear translocation and of C/EBP and/or AP-1 activation, which may constitute anti-inflammatory effects of the lignan.
College of Pharmacy and Research Institute of Pharmaceutical Sciences, Seoul National University, Sillim-dong, Kwanak-gu, Seoul 151-742, South Korea.
Ae Kyung
AK
Lee
Sang Hyun
SH
Sung
Young Choong
YC
Kim
Sang Geon
SG
Kim
D000818:Animals; D001578:Benzopyrans / Q000494:pharmacology*; D015152:Blotting, Northern; D015153:Blotting, Western; D022762:CCAAT-Enhancer-Binding Proteins / Q000378:metabolism*; D002460:Cell Line; D017362:Cyclic AMP Response Element-Binding Protein / Q000378:metabolism; D051546:Cyclooxygenase 2; D004149:Dioxoles / Q000494:pharmacology*; D004790:Enzyme Induction / Q000187:drug effects; D015870:Gene Expression; D020672:I-kappa B Proteins / Q000378:metabolism*; D007150:Immunohistochemistry; D007527:Isoenzymes / Q000096:biosynthesis*; D008070:Lipopolysaccharides / Q000494:pharmacology; D008264:Macrophages / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism; D051379:Mice; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors / Q000096:biosynthesis*; D052247:Nitric Oxide Synthase Type II; D010766:Phosphorylation; D011451:Prostaglandin-Endoperoxide Synthases / Q000096:biosynthesis*; D012333:RNA, Messenger / Q000096:biosynthesis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D018808:Transcription Factor AP-1 / Q000378:metabolism*; D051742:Transcription Factor CHOP; D014157:Transcription Factors / Q000378:metabolism*; D014409:Tumor Necrosis Factor-alpha / Q000037:antagonists & inhibitors / Q000096:biosynthesis*
D001578:Benzopyrans; D022762:CCAAT-Enhancer-Binding Proteins; D017362:Cyclic AMP Response Element-Binding Protein; C497250:Ddit3 protein, mouse; D004149:Dioxoles; D020672:I-kappa B Proteins; D007527:Isoenzymes; D008070:Lipopolysaccharides; D012333:RNA, Messenger; D018808:Transcription Factor AP-1; D014157:Transcription Factors; D014409:Tumor Necrosis Factor-alpha; C410958:sauchinone; D051742:Transcription Factor CHOP; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II; C496317:Nos2 protein, mouse; D051546:Cyclooxygenase 2; D011451:Prostaglandin-Endoperoxide Synthases