BACKGROUND Chloranthus 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. METHODS The 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. RESULTS The 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. CONCLUSIONS CSSPW 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.

中文翻译：

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锯缘青蟹的水分离部分主要通过调节MAPK和Nrf2 / HO-1炎症途径来减轻脂多糖诱导的RAW264.7细胞损伤。

背景技术锯缘青蟹（Chloranthus serratus，Chloranthaceae）已被用于治疗瘀伤，类风湿和骨痛。然而，尚未完全阐明体外的锯齿隐球菌的抗炎机制。本研究旨在探讨脂多糖（LPS）诱导的RAW264.7细胞中锯缘隐球菌的水分离部分（CSSPW）的抗炎活性和潜在机制。方法采用CCK-8法优化CSSPW的浓度。通过一步法检测一氧化氮（NO）含量。通过酶联免疫吸附测定（ELISA）确定炎性细胞因子的水平。通过实时定量PCR（qPCR）检测诱导型一氧化氮合酶（iNOS）和环氧合酶-2（COX-2）的基因表达。免疫荧光和DCFH-DA荧光探针分别用于检测p65核易位和活性氧（ROS）含量。Western印迹法用于检测丝裂原活化蛋白激酶（MAPK），核因子-κB（NF-κB）和核转录因子E2相关因子2 /血红素加氧酶-1（Nrf2 / HO-1）通路的蛋白表达。 。结果最终浓度分别为15 ng / mL，1.5μg/ mL和150μg/ mL，分别作为低，中和高剂量的CSSPW。CSSPW治疗显着减少了对LPS刺激产生的NO，肿瘤坏死因子-α（TNF-α），白细胞介素6（IL-6），前列腺素E2（PGE2），iNOS mRNA和COX-2 mRNA的产生。此外，CSSPW处理可抑制MAPK和NF-κB通路的蛋白表达，以及p65核易位和ROS产生。相反，Nrf2 / HO-1途径的蛋白质表达明显上调。结论CSSPW通过下调促炎性介质的产生，抑制NF-κB和MAPK途径的激活以及激活LPS诱导的RAW264.7细胞中的Nrf2 / HO-1途径发挥其抗炎作用。