| Andrographis is a type of traditional Chinese medicine that has many active components with anti-inflammatory properties. Andrographolide, the main active component extracted from Andrographis, exerts anti-inflammatory effects.However,the underlying molecular mechanisms remain clear. Inflammation is a complex process involving the interaction between an organism and pathogens, with the results of these complex interactions to induce macrophage activation. In addition,activated macrophages can eliminate invading infectious microbes and trigger the release of inflammatory cytokines,such as TNF-α,IL-1β,and IL-6, then complete a variety of immune function in response to these cytokines . Existing research shows that LPS can induce inflammation,NF-κB and MAPK signaling pathways are two extremely classical activation pathways. Therefore, inhibiting the inflammatory reaction by targeting signal transduction pathways has become a novel avenue for the treatment of inflammation. The purpose of this experiment was further to explore the anti-inflammatory effect of andrographolide and mechanism of signal transduction on LPS induced inflammatory reaction,and provide the theoretical basis for therapeutical effect of andrographolide.First, it is to explore the anti-inflammatory mechanism of andrographolide in RAW264.7 cells. An in vitro model of inflammation was induced by LPS in mouse RAW264.7 cells before the cells were treated with various concentrations of andrographolide (25、12.5 and 6.25 μg/mL). The concentration and expression levels of proinflammatory cytokines were determined by an enzyme-linked immunosorbent assay (ELISA) and quantitative real-time polymerase chain reaction (qRT-PCR),respectively. The nuclear expression levels of NF-κB were determined by an electrophoretic mobility shift assay (EMSA). The expression levels of NF-κB, p38,ERK, and JNK were determined by Western blot. The results demonstrated that andrographolide dose-dependently inhibited the expression of TNF-a, IL- 6, and IL-1β in LPS-stimulated RAW264.7 cells. The Western blot analysis showed that andrographolide suppressed LPS-induced NF-κB activation and the phosphorylation of I-κB, ERK1/2, JNK, and p38. Thus, andrographolide exerts an anti-inflammatory effect by inhibiting the activation of NF-κB via the MAPK signaling pathway, and the release of proinflammatory cytokines.Then, it is further to explore the anti-inflammatory mechanism of andrographolide and observe its anti-inflammatory effect in mice. Sixty lactating mice at the age of 5 - 7 days were randomly divided into six groups. Mastitis model was established by pouring LPS into mammary gland. Mammary gland of mice with mastitis was collected at 24 h after the mice were treated with various concentrations of Andrographolide (6.25、12.5and 25 μg/mL) and dexamethasone, respectively. The histopathology of mammary gland was observed by histological section. The concentration and expression levels of TNF-a, IL-6, and IL-1β were determined through enzyme-linked immunosorbent assay (ELISA) and quantitative real-time polymerase chain reaction (qRT-PCR), respectively. The expression levels of NF-κB,p38, ERK, and JNK were determined by western blot. In result, compared with LPS group, Andrographolide significantly inhibited the mammary gland pathological damages. The release and mRNA expression of TNF-a, IL-1β, and IL-6 were dose-dependently inhibited in Andrographolide-treated groups. The nuclear expression of p65 protein and phosphorylation of Iκ-B, JNK, ERK1/2, and p38 were markedly decreased in Andrographolide treatment group. These results suggest that Andrographolide exerts an anti-inflammatory effect by inhibiting the activation of NF-κB/MAPK signaling pathway, and the release of proinflammatory cytokines.In conclusion, andrographolide exerts an anti-inflammatory effect by inhibiting the activation of NF-κB/MAPK signaling pathway, the expression and release of proinflammatory cytokines (TNF-α、IL-1β and IL-6). |
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