Abstract
BackgroundChloranthus serratus (Chloranthaceae) has been used to treat bruises, rheumatoid and bone pain. However, the anti-inflammatory mechanisms of C. serratus in vitro have not been fully elucidated. The present study aimed to explore the anti-inflammatory activity and potential mechanisms of C. serratus’s separated part of water (CSSPW) in lipopolysaccharide (LPS)-induced RAW264.7 cells.MethodsThe concentrations of CSSPW were optimized by CCK-8 method. Nitric oxide (NO) content was detected by one-step method. The levels of inflammatory cytokines were determined by enzyme-linked immunosorbent assay (ELISA). Gene expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) was detected by real-time quantitative PCR (qPCR). Immunofluorescence and DCFH-DA fluorescent probes were used to detect p65 nuclear translocation and reactive oxygen species (ROS) content, respectively. Western blotting was used to assay the protein expression of mitogen-activated protein kinases (MAPK), nuclear factor-kappa B (NF-κB) and nuclear transcription factor E2 related factor 2/haem oxygenase-1 (Nrf2/HO-1) pathways.ResultsThe final concentrations of 15 ng/mL, 1.5 μg/mL and 150 μg/mL were selected as low, medium and high doses of CSSPW, respectively. CSSPW treatment significantly reduced the generation of NO, tumour necrosis factor-α (TNF-α), interleukin-6 (IL-6), prostaglandinE2 (PGE2), iNOS mRNA and COX-2 mRNA in response to LPS stimulation. Furthermore, the protein expression of the MAPK and NF-κB pathways was suppressed by CSSPW treatment, as well as p65 nuclear translocation and ROS production. In contrast, the protein expression of the Nrf2/HO-1 pathway was markedly upregulated.ConclusionsCSSPW exerts its anti-inflammatory effect via downregulating the production of pro-inflammatory mediators, inhibiting the activation of NF-κB and MAPK pathways, as well as activating Nrf2/HO-1 pathway in LPS-induced RAW264.7 cells.
Highlights
Chloranthus serratus (Chloranthaceae) has been used to treat bruises, rheumatoid and bone pain
The occurrence of inflammation is accompanied by the production of inflammatory mediators such as prostaglandin E2 (PGE2), interleukin-6 (IL-6), tumour necrosis factor-α (TNF-α) and nitric oxide (NO)
(p < 0.01), especially those of the 150 μg/mL group (Fig. 4). These results indicated that CSSPW may suppress LPS-induced PGE2 and NO production by downregulating COX-2 and inducible nitric oxide synthase (iNOS) mRNA expression in RAW264.7 cells
Summary
Chloranthus serratus (Chloranthaceae) has been used to treat bruises, rheumatoid and bone pain. The occurrence of inflammation is accompanied by the production of inflammatory mediators such as prostaglandin E2 (PGE2), interleukin-6 (IL-6), tumour necrosis factor-α (TNF-α) and nitric oxide (NO). The overexpression of these inflammatory mediators may cause serious damage to cells and tissues, even resulting in organ failure [3, 5]. Two pro-inflammatory enzymes, inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2), play synergistic roles in inflammatory diseases, and drugs with dual inhibitory effects on iNOS and COX-2 mRNA have great antiinflammatory potential [6]. Inhibiting the overexpression of related inflammatory mediators is an effective method to control inflammation
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