Abstract
Nodakenin, a coumarin isolated from the roots of Angelicae gigas, has been reported to possess neuroprotective, antiaggregatory, antibacterial, and memory-enhancing effects. In the present study, we investigated the anti-inflammatory effects of nodakenin by examining its in vitro inhibitory effects on inducible nitric-oxide synthase (iNOS), cyclooxygenase-2 (COX-2), and proinflammatory cytokines in lipopolysaccharide (LPS)-induced RAW 264.7 macrophages and mouse peritoneal macrophages and its in vivo effects on LPS-induced septic shock in mice. Our results indicate that nodakenin concentration-dependently inhibits iNOS and COX-2 at the protein, mRNA, and promoter binding levels, and these inhibitions cause attendant decreases in the production of nitric oxide (NO) and prostaglandin E2 (PGE2). Furthermore, we found that nodakenin inhibits the production and mRNA expression of tumor necrosis factor-α (TNF-α), interleukin (IL)-6, and IL-1β induced by LPS. Molecular data revealed that nodakenin suppressed the transcriptional activity and translocation of nuclear factor-κB (NF-κB) by inhibiting inhibitory κB-α degradation and IκB kinase-α/β phosphorylation. In addition, nodakenin was found to significantly inhibit the LPS-induced binding of transforming growth factor-β-activated kinase 1 to tumor necrosis factor receptor-associated factor 6 (TRAF6) by reducing TRAF6 ubiquitination. Pretreatment with nodakenin reduced the serum levels of NO, PGE2, and proinflammatory cytokines and increased the survival rate of mice with LPS-induced endotoxemia. Taken together, our data suggest that nodakenin down-regulates the expression of the proinflammatory iNOS, COX-2, TNF-α, IL-6, and IL-1β genes in macrophages by interfering with the activation of TRAF6, thus preventing NF-κB activation.
Footnotes
This research was supported by the Basic Science Research Program through the National Research Foundation of Korea, which is funded by the Korean government [Grants 2011-0023407, 2011-0030724].
Article, publication date, and citation information can be found at http://jpet.aspetjournals.org.
ABBREVIATIONS:
- LPS
- lipopolysaccharide
- AP-1
- activator protein-1
- Con
- control
- COX
- cyclooxygenase
- DMEM
- Dulbecco's modified Eagle's medium
- ELISA
- enzyme-linked immunosorbent assay
- ERK
- extracellular signal-related kinase
- IκB
- inhibitor of κB
- IKK
- IκB kinase
- IL
- interleukin
- IRAK1
- IL-1 receptor-associated kinase 1
- JNK
- c-Jun N-terminal kinase
- l-NIL
- l-N6-(1-iminoethyl) lysine
- MAPK
- mitogen-activated protein kinase
- MTT
- 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide
- NF-κB
- nuclear factor-κB
- NO
- nitric oxide
- NOS
- NO synthase
- iNOS
- inducible NOS
- p-
- phosphorylated
- NS398
- N-(2-cyclohexyloxy-4-nitrophenyl)methanesulfonamide
- PARP
- poly(ADP-ribose) polymerase
- PBS
- phosphate-buffered saline
- PCR
- polymerase chain reaction
- PG
- prostaglandin
- RT
- reverse transcriptase
- TAK1
- transforming growth factor-β-activated kinase 1
- TNF-α
- tumor necrosis factor-α
- TRAF6
- tumor necrosis factor receptor-associated factor 6
- Ub
- ubiquitin
- Ubc
- Ub-conjugating enzyme complex.
- Received March 20, 2012.
- Accepted May 24, 2012.
- Copyright © 2012 by The American Society for Pharmacology and Experimental Therapeutics
JPET articles become freely available 12 months after publication, and remain freely available for 5 years.Non-open access articles that fall outside this five year window are available only to institutional subscribers and current ASPET members, or through the article purchase feature at the bottom of the page.
|