Comprehensive comparison on the anti-inflammatory effects of three species of Sigesbeckia plants based on NF-κB and MAPKs signal pathways in vitro

Ethnopharmacological relevance: Sigesbeckiae Herba (SH), a traditional anti-inflammatory Chinese herbal medicine, is originated from the plants of Sigesbeckia pubescens Makino (SP), S. orientalis L. (SO) and S. glabrescens Makino (SG). The current studies reported that the chemical constituents in the three species of plants were different. Aim of the study: The aim of this study is to provide a systemic comparison on the anti-inflammatory effects and the underlying molecular mechanisms among the three plants based on their effects on nuclear factor-kappa B (NF-kappa B) and mitogen-activated protein kinases (MAPKs) signal pathways in vitro. Material and methods: Twenty-four batches of three Sigesbeckia herbs were collected from different regions of China and extracted with 50% ethanol. The distribution of 6 compounds in the 24 batches of SH extracts were characterized by UPLC analysis. The cytotoxicity of all extracts to RAW264.7 cells in the absence or presence of lipopolysaccharide (LPS) were examined by 3- (4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The anti-inflammatory effects of the extracts were investigated using Griess reagent and enzymelinked immunosorbent assay. The underlying mechanisms of the representative samples (SP007, S0005 and SG003) for individual species were examined by western blotting and immunofluorescence staining. Results: The estimated average sub-lethal dose (LD15) of SP, SO and SG on RAW264.7 cells were 181.7 +/- 15.7, 291.5 +/- 33.9 and 317.1 +/- 16.3 mu g/mL, respectively. In LPS-stimulated RAW264.7 cells, the inhibitory effects of SH species were determined to be SP > SO > SG on NO release, while SP similar to SO > SG on secretion of postinflammatory cytokines (TNF-alpha, IL-6 and MCP-1). Moreover, suppression on LPS-induced excessive expressions of cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS), as well as the activation of NF-kappa B and phosphorylation of MAPKs were investigated to be associated to the anti-inflammatory effects for all SH species. Conclusions: We firstly reported a systemic comparison on the anti-inflammatory properties for the three main plant origins of SH. Although SG showed lower toxicity and less anti-inflammatory effects compared with SP and SO in LPS-induced RAW264.7 cells, comparable inhibitory effects on NF-kappa B and MAPKs pathways and the reduction of LPS-induced iNOS and COX-2 were observed in the anti-inflammatory process for all Sigesbeckia plants.