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Background   Panax Notoginseng  flower saponins (PNFS) are the main active component of  Panax notoginseng  (Burk) F. H. Chen flower bud (PNF) and possess significant anti-inflammatory efficacy. This study aims to explore the mechanisms underlying PNFS’ antiflammatory action in RAW264.7 macrophages.    Methods  A cell counting kit-8 assay was used to determine the viability of RAW264.7 macrophages. Anti-inflammation effects of PNFS in lipopolysaccharide (LPS)-stimulated RAW264.7 macrophages were measured based on the detection of nitric oxide (NO) overproduction (Griess method, DAF-FM DA fluorescence assay and NO 2  −  scavenging assay), and interleukin (IL)-6 and tumor necrosis factor (TNF)-alpha gene overexpression (real-time PCR and ELISA). Inducible nitric oxide synthase (iNOS) gene overexpression was determined by real-time PCR and western blotting. iNOS enzyme activity was also assayed. The mechanisms underlying the suppression of iNOS gene overexpression by PNFS were explored using real-time PCR and western blotting to assess mRNA and protein levels of components of the Toll-like receptor 4 mitogen-activated protein kinase (MAPK), phosphatidylinositol 3-kinase (PI3K)/Akt, and nuclear factor-kappa B (NF-kappa B) signaling pathways.    Results  PNFS (50, 100, 200 μg/mL) significantly reduced LPS-induced overproduction of NO ( P <  0.001,  P <  0.001,  P <  0.001) and IL-6 ( P =  0.103,  P <  0.001,  P <  0.001), but did not affect TNF-alpha overproduction. PNFS (50, 100, 200 μg/mL) also markedly decreased LPS-activated iNOS ( P <  0.001,  P <  0.001,  P <  0.001) and TLR4 gene overexpression ( P =  0.858,  P =  0.046,  P =  0.005). Furthermore, treatment with PNFS (200 μg/mL) suppressed the phosphorylation of MAPKs including P38 ( P =  0.001), c-Jun N-terminal kinase (JNK) ( P =  0.036) and extracellular-signal regulated kinase (ERK) 1/2 ( P =  0.021). PNFS (200 μg/mL) inhibited the activation of the NF-kappa B signaling pathway by preventing the phosphorylation of inhibitor of NF-kappa B alpha (I-kappa B alpha) ( P =  0.004) and P65 ( P =  0.023), but PNFS (200 μg/mL) could not activate the LPS-induced PI3K-Akt signaling pathway.    Conclusions  PNFS significantly down-regulated iNOS gene overexpression and thereby decreased NO overproduction via the inhibition of TLR4-mediated MAPK/NF-kappa B signaling pathways, but not the PI3K/Akt signaling pathway.

chinese medicine

Panax Notoginseng flower saponins (PNFS) inhibit LPS-stimulated NO overproduction and iNOS gene overexpression via the suppression of TLR4-mediated MAPK/NF-kappa B signaling pathways in RAW264.7 macrophages