Syk and Src are major pharmacological targets of a Cerbera manghas methanol extract with kaempferol-based anti-inflammatory activity

Cerbera manghas L. (Apocynaceae), a semi-mangrove medicinal plant distributed throughout tropical and subtropical countries, is traditionally known to possess analgesic, anti-inflammatory, anti-convulsant, cardiotonic, and hypotensive activity. In vitro and in vivo anti-inflammatory activities of a methanol extract of the leaves of Cerbera manghas and the underlying molecular mechanisms were investigated to validate the ethnopharmacological use of this plant.The effect of Cerbera manghas methanol extract (Cm-ME) on the production of inflammatory mediators and the induction of HCl/EtOH-treated gastritis was explored using macrophages, HEK293 cells, and ICR mice. The molecular targets of this extract and potential active components in Cm-ME were also investigated.Cm-ME inhibited the production of nitric oxide (NO) in lipopolysaccharide (LPS)-treated RAW264.7 cells and peritoneal macrophages in a dose-dependent manner. This extract also suppressed the expression of NO synthase (iNOS) and cyclooxygenase (COX)-2. NF-κB-mediated enhancement of luciferase activity, nuclear translocation of p50 and p65, and phosphorylation of IκBα were markedly reduced by Cm-ME treatment. Direct enzyme assays, reporter gene assays, and immunoprecipitation analysis of kinases revealed Syk and Src as immunopharmacological targets of Cm-ME. Moreover, this extract strongly ameliorated the gastric symptoms induced by HCl/EtOH treatment of mice. Finally, HPLC analysis and pharmacological tests identified kaempferol as an active component of the extract with Src/Syk inhibitory activities.Inhibition of Syk/Src and the NF-κB pathway by kaempferol could play a key role in the anti-inflammatory pharmacological action of Cerbera manghas.