The Anti-inflammatory Effect of Gnaphalium affine Through Inhibition of NF-κB and MAPK in Lipopolysaccharide-Stimulated RAW264.7 Cells and Analysis of Its Phytochemical Components

Abstract

Gnaphalium affine is an annual herbaceous plant that is used as a traditional medicine in some Latin American and Asian countries. However, systematic studies on its anti-inflammatory activity and signaling pathways have not yet been reported. In this study, we investigated the anti-inflammatory effect of G. affine methanol extract in lipopolysaccharide (LPS)-stimulated RAW 264.7 murine macrophage cells and fractioned the methanol extract into hexane, chloroform, ethyl acetate (EtOAc), butyl alcohol (BuOH), and distilled water (DW) by measuring the generation of nitric oxide (NO). G. affine inhibited the generation of NO and prostaglandin E2. The chloroform-soluble fraction most effectively inhibited LPS-stimulated NO production. We also examined the cytotoxicity of G. affine in three normal cell lines: RAW264.7, HEK293, and HaCaT. Cell viability assays showed that the methanol extract and chloroform-soluble fraction of G. affine had no cytotoxic effect on normal cell lines. The expression of pro-inflammatory mediators was also investigated. Western blotting and immunofluorescence showed that G. affine reduces the expression of iNOS, COX-2, and MAPKs, as well as activation of NF-κB in LPS-stimulated RAW264.7 cells. RT-PCR showed that G. affine also negatively regulates inflammatory cytokines at the gene expression level. Taken together, G. affine exerts its anti-inflammatory activity through inhibition of NO generation as a result of inhibiting NF-κB and MAPKs-related inflammatory signaling pathways. In addition, the result of GC-MS analysis revealed the presence of nineteen different types of constituents including guaiacol in the chloroform-soluble fraction of G. affine.

Preparation of newly identified polysaccharide from Pleurotus eryngii and its anti‐inflammation activities potential

“…Specifically, the p38 protein has been shown to be an essential factor for the cytokines secretion as well as the main enzymes during the inflammation interaction of the host (Lu et al., 2014) from which we could speculate that the PEPs displayed its key anti‐inflammatory activities through the downregulation effects of the inflammatory cytokines and some immune mediators (Jiang et al., 2017). The expression and function of Erk1/2 has been considered to be related to the activation of LPS‐induced cellular responses including the increased production of TNF‐α, NO, and IL‐6 in the immune cells (Seong, Hwang, & Kim, 2016). We could confirm that the MAPK pathway was one of the functional signal pathways for the alteration effects of WPEP and NPEP on the cell inflammation induced by LPS.…”

Kaempferol‑3‑O‑β‑rutinoside suppresses the inflammatory responses in lipopolysaccharide‑stimulated RAW264.7 cells via the NF‑κB and MAPK pathways

“…The mRNA expression levels of the cytokines TNF-α, IL-6 and IL-1β were analyzed by RT-PcR, according to previous studies (22,23). Total RNA was isolated using the RNeasy ® Plus Mini kit (Qiagen GmbH), and then cdNA was synthesized using AccuPower ® RT premix (Bioneer corporation).…”

Methanol extract of Guettarda speciosa Linn. inhibits the production of inflammatory mediators through the inactivation of Syk and JNK in macrophages

“…Although the promoter region of each proinflammatory cytokine is known to contain binding sites for NF-κB and downstream transcription factors of MAPKs, previous studies by our group demonstrated that extracts of natural plants exert different inhibitory effects on the production of proinflammatory cytokines as a result of differential regulation of NF-κB and MAPKs (41). Of note, the inhibition of TNF-α production was only detected when the extract inhibited NF-κB and all MAPKs, including JNK, ERK and p38 (42). In one instance, TNF-α production was not attenuated when the extract only inhibited one MAPK in addition to NF-κB (43).…”