Even though a lot of reports have suggested the anti-inflammatory activity of kaempferol (KF) in macrophages, little is known about its exact anti-inflammatory mode of action and its immunopharmacological target molecules. In this study, we explored anti-inflammatory activity of KF in LPS-treated macrophages. In particular, molecular targets for KF action were identified by using biochemical and molecular biological analyses. KF suppressed the release of nitric oxide (NO) and prostaglandin E2 (PGE2), downregulated the cellular adhesion of U937 cells to fibronectin (FN), neutralized the generation of radicals, and diminished mRNA expression levels of inflammatory genes encoding inducible NO synthase (iNOS), TNF-α, and cyclooxygenase- (COX-) 2 in lipopolysaccharide- (LPS-) and sodium nitroprusside- (SNP-) treated RAW264.7 cells and peritoneal macrophages. KF reduced NF-κB (p65 and p50) and AP-1 (c-Jun and c-Fos) levels in the nucleus and their transcriptional activity. Interestingly, it was found that Src, Syk, IRAK1, and IRAK4 responsible for NF-κB and AP-1 activation were identified as the direct molecular targets of KF by kinase enzyme assays and by measuring their phosphorylation patterns. KF was revealed to have in vitro and in vivo anti-inflammatory activity by the direct suppression of Src, Syk, IRAK1, and IRAK4, involved in the activation of NF-κB and AP-1.

No abstract

Kaempferol (KF) is the most abundant polyphenol in tea, fruits, vegetables, and beans. However, little is known about its in vivo anti-inflammatory efficacy and mechanisms of action. To study these, several acute mouse inflammatory and nociceptive models, including gastritis, pancreatitis, and abdominal pain were employed. Kaempferol was shown to attenuate the expansion of inflammatory lesions seen in ethanol (EtOH)/HCl- and aspirin-induced gastritis, LPS/caerulein (CA) triggered pancreatitis, and acetic acid-induced writhing.

Korean Red Ginseng (KRG) is an herbal medicine prescribed worldwide that is prepared from Panax ginseng C.A. Meyer (Araliaceae). Out of ginseng's various components, ginsenosides are regarded as the major ingredients, exhibiting anticancer and anti-inflammatory activities. Although recent studies have focused on understanding the anti-inflammatory activities of KRG, compounds that are major anti-inflammatory components, precisely how these can suppress various inflammatory processes has not been fully elucidated yet. In this study, we aimed to identify inhibitory saponins, to evaluate the in vivo efficacy of the saponins, and to understand the inhibitory mechanisms. To do this, we employed in vitro lipopolysaccharide-treated macrophages and in vivo inflammatory mouse conditions, such as collagen (type II)-induced arthritis (CIA), EtOH/HCl-induced gastritis, and lipopolysaccharide (LPS)/D-galactosamine (D-GalN)-triggered hepatitis. Molecular mechanisms were also verified by real-time PCR, immunoblotting analysis, and reporter gene assays. Out of all the ginsenosides, ginsenoside Rc (G-Rc) showed the highest inhibitory activity against the expression of tumor necrosis factor (TNF)-[Formula: see text], interleukin (IL)-1[Formula: see text], and interferons (IFNs). Similarly, this compound attenuated inflammatory symptoms in CIA, EtOH/HCl-mediated gastritis, and LPS/D-galactosamine (D-GalN)-triggered hepatitis without altering toxicological parameters, and without inducing gastric irritation. These anti-inflammatory effects were accompanied by the suppression of TNF-[Formula: see text] and IL-6 production and the induction of anti-inflammatory cytokine IL-10 in mice with CIA. G-Rc also attenuated the increased levels of luciferase activity by IRF-3 and AP-1 but not NF-[Formula: see text]B. In support of this phenomenon, G-Rc reduced TBK1, IRF-3, and ATF2 phosphorylation in the joint and liver tissues of mice with hepatitis. Therefore, our results strongly suggest that G-Rc may be a major component of KRG with useful anti-inflammatory properties due to its suppression of IRF-3 and AP-1 pathways.

Celtis choseniana is the traditional plant used at Korea as a herbal medicine to ameliorate inflammatory responses. Although Celtis choseniana has been traditionally used as a herbal medicine at Korea, no systemic research has been conducted on its anti-inflammatory activity. Therefore, the present study explored an anti-inflammatory effect and its underlying molecular mechanism using Celtis choseniana methanol extract (Cc-ME) in macrophage-mediated inflammatory responses. In vitro anti-inflammatory activity of Cc-ME was evaluated using RAW264.7 cells and peritoneal macrophages stimulated by lipopolysaccharide (LPS), pam3CSK4 (Pam3), or poly(I:C). In vivo anti-inflammatory activity of Cc-ME was investigated using acute inflammatory disease mouse models, such as LPS-induced peritonitis and HCl/EtOH-induced gastritis. The molecular mechanism of Cc-ME-mediated anti-inflammatory activity was examined by Western blot analysis and immunoprecipitation using whole cell and nuclear fraction prepared from the LPS-stimulated RAW264.7 cells and HEK293 cells. Cc-ME inhibited NO production and mRNA expression of inducible nitric oxide synthase (iNOS), cyclooxygenase (COX-2), and tumor necrosis factor-alpha (TNF-α) in the RAW264.7 cells and peritoneal macrophages induced by LPS, pam3, or poly(I:C) without cytotoxicity. High-performance liquid chromatography (HPLC) analysis showed that Cc-ME contained anti-inflammatory flavonoids quercetin, luteolin, and kaempferol. Among those, the content of luteolin, which showed an inhibitory effect on NO production, was highest. Cc-ME suppressed the NF-κB signaling pathway by targeting Src and interrupting molecular interactions between Src and p85, its downstream kinase. Moreover, Cc-ME ameliorated the morphological finding of peritonitis and gastritis in the mouse disease models. Therefore, these results suggest that Cc-ME exerted in vitro and in vivo anti-inflammatory activity in LPS-stimulated macrophages and mouse models of acute inflammatory diseases. This anti-inflammatory activity of Cc-ME was dominantly mediated by targeting Src in NF-κB signaling pathway during macrophage-mediated inflammatory responses.