College of Pharmacy, Wannan Medical College, Wuhu, 241002, Anhui, China; Institute of Natural Daily Chemistry, Wannan Medical College, Wuhu, 241002, Anhui, China.
College of Pharmacy, Wannan Medical College, Wuhu, 241002, Anhui, China.
J Ethnopharmacol. 2021 May 10;271:113880. doi: 10.1016/j.jep.2021.113880. Epub 2021 Jan 26.
Chloranthus serratus is a traditional Chinese medicine for treating arthritis and bruises.
To investigate the dose-effect relationship and molecular mechanisms of the water extract of C. serratus roots (WECR) in lipopolysaccharide (LPS)-stimulated RAW264.7 cells.
The cell viability was detected by CCK-8 method. One-step method, DCFH-DA fluorescence probe method and immunofluorescence method were used to detect nitric oxide (NO), reactive oxygen species (ROS) and p65 nuclear transcription, respectively. Interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α) and prostaglandin E (PGE) were detected by enzyme linked immunosorbent assay. Inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) mRNA were detected by quantitative real-time PCR. Western blotting was taken to determine the contents of the relevant proteins in the nuclear transcription factor E2 related factor 2/heme oxygenase-1 (Nrf2/HO-1), mitogen-activated protein kinases (MAPK) and nuclear factor-kappa B (NF-κB) pathways.
The concentrations of 3, 30 and 300 μg/mL were optimized as low, medium and high concentrations of the WECR, respectively, and 1 μg/mL was selected as the optimal concentration of LPS to activate macrophages. The dose of the positive drug dexamethasone was 0.13 mg/mL. The WECR could not only inhibit LPS-induced cell differentiation and the overexpression of NO, IL-6, TNF-α, PGE and ROS but also promote the expression of Nrf2 and HO-1, and down-regulate the phosphorylation levels of ERK, JNK, p38 and p65. After the WECR treatment, the expression levels of iNOS and COX-2 mRNA and nuclear translocation of p65 were all inhibited.
The WECR exerts its anti-inflammatory activity by inhibiting the MAPK and NF-κB pathways, activating the Nrf2/HO-1 pathway and down-regulating inflammatory factor levels in a dose-dependent manner.
腺梗豨莶全草是一种传统的中药，用于治疗关节炎和瘀伤。
研究腺梗豨莶根水提物（WECR）在脂多糖（LPS）刺激的 RAW264.7 细胞中的量效关系和分子机制。
用 CCK-8 法检测细胞活力。采用一步法、DCFH-DA 荧光探针法和免疫荧光法分别检测一氧化氮（NO）、活性氧（ROS）和 p65 核转录，酶联免疫吸附法检测白细胞介素-6（IL-6）、肿瘤坏死因子-α（TNF-α）和前列腺素 E（PGE）。用定量实时 PCR 检测诱导型一氧化氮合酶（iNOS）和环氧化酶-2（COX-2）mRNA。采用 Western blotting 法测定核转录因子 E2 相关因子 2/血红素加氧酶-1（Nrf2/HO-1）、丝裂原激活蛋白激酶（MAPK）和核因子-κB（NF-κB）通路中相关蛋白的含量。
将 3、30 和 300μg/mL 的浓度优化为 WECR 的低、中、高浓度，将 1μg/mL 选为激活巨噬细胞的 LPS 最佳浓度。阳性药物地塞米松的剂量为 0.13mg/mL。WECR 不仅能抑制 LPS 诱导的细胞分化和 NO、IL-6、TNF-α、PGE 和 ROS 的过表达，还能促进 Nrf2 和 HO-1 的表达，下调 ERK、JNK、p38 和 p65 的磷酸化水平。经 WECR 处理后，iNOS 和 COX-2mRNA 的表达水平和 p65 的核转位均受到抑制。
WECR 通过抑制 MAPK 和 NF-κB 通路，激活 Nrf2/HO-1 通路，下调炎症因子水平，发挥抗炎活性，呈剂量依赖性。
