通讯作者: 孙广臣, sungc@glmc.edu.cn

DOI：10.12201/bmr.202504.00003

Investigating the Role of Escin in RAW 264.7 Macrophage Inflammatory Models

• 摘要：目的 研究七叶皂苷（Escin）对巨噬细胞极化因子、炎症因子和炎症相关通路的调控作用，探讨七叶皂苷对类风湿性关节炎（Rheumatoid Arthritis，RA）的治疗机制。 方法 分别使用0、0.5、1.0、2.0、5.0、10.0和20.0的Escin给药治疗RAW 264.7巨噬细胞，噻唑蓝（3-(4,5)-dimethylthiahiazo(-z-y1)-3,5-di-phenytetrazoliumromide，MTT）法测定不同浓度七叶皂苷对细胞活性的影响；通过脂多糖（Lipopolysaccharide, LPS）刺激诱导RAW 264.7巨噬细胞炎症模型，共分为6组：空白组、模型组（500 ng/mL LPS）、Escin治疗组（500 ng/mL LPS+0.5 μM Escin、500 ng/mL LPS+1.0 μM Escin、500 ng/mL LPS+2.0 μM Escin、500 ng/mL LPS+4.0 μM Escin），蛋白印迹实验（Western Blotting, WB）检测七叶皂苷对LPS刺激的RAW 264.7巨噬细胞炎症模型中丝裂原活化蛋白激酶（Mitogen-activated protein kinase，MAPK）信号通路蛋白表达的影响，如细胞外信号调节激酶（Extracellular Signal-Regulated Kinase，ERK）和c-Jun N末端激酶（c-Jun N-terminal Kinase，JNK），对巨噬细胞相关分子表达的影响，如巨噬细胞甘露糖受体（Macrophage Mannose Receptor，CD206）和血红蛋白清道夫受体（Hemoglobin Scavenger Receptor，CD163），对炎症因子肿瘤坏死因子-α（Tumor necrosis factor alpha，TNF-α）表达的影响。 结果 根据MTT法检验，0.5-5 μM Escin对RAW264.7细胞没有毒性。蛋白印迹实验结果显示，4 μM Escin显著下调TNF-α的表达，0.5-4 μM Escin显著上调CD206和CD163的表达；0.5-4 μM Escin显著下调ERK的表达，4 μM Escin显著下调JNK的表达。 结论 七叶皂苷在体外可以一定程度上抑制脂多糖刺激的RAW264.7巨噬细胞M1型极化，促进M2型极化，并通过调控MAPK信号通路，发挥七叶皂苷对类风湿性关节炎的治疗作用。

  关键词： 七叶皂苷; 抗炎; 作用机制; RAW264.7巨噬细胞

   

  Abstract: Objective This study aimed toTo investigate the regulatory effects of escin on macrophage polarization factors, inflammatory cytokines, and inflammation-related pathways, and to explore its therapeutic mechanisms in rheumatoid arthritis (RA). Methods RAW 264.7 macrophages were treated with escin at concentrations of 0 , 0.5, 1.0, 2.0, 5.0, 10.0, and 20.0 μM. Cell viability was assessed using the thiazolyl blue tetrazolium bromide (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, MTT) assay. An inflammatory model was established by stimulating RAW 264.7 macrophages with lipopolysaccharide (LPS, 500 ng/mL). The experimental groups were divided as follows: Control group (untreated), Model group (500 ng/mL LPS), Escin treatment groups(500 ng/mL LPS + 0.5 μM escin, 500 ng/mL LPS + 1.0 μM escin, 500 ng/mL LPS + 2.0 μM escin, 500 ng/mL LPS + 4.0 μM escin). Western blot analysis?was performed to analyze: The expression of?mitogen-activated protein kinase (MAPK) pathway proteins, including?extracellular signal-regulated kinase (ERK)?and?c-Jun N-terminal kinase (JNK), in LPS-stimulated macrophages. The expression of?macrophage polarization markers:?macrophage mannose receptor (CD206)?and?hemoglobin scavenger receptor (CD163). The levels of the?pro-inflammatory cytokine tumor necrosis factor-α (TNF-α). Results MTT assays confirmed that?0.5-5 μM escin?exhibited no cytotoxicity toward RAW 264.7 cells. Western blot results demonstrated that: 4 μM escin?significantly?downregulated TNF-α expression.?0.5–4 μM escin?markedly?upregulated CD206 and CD163 expression, indicating enhanced M2 polarization.?0.5–4 μM escin?significantly?suppressed ERK expression, while?4 μM escin?notably?inhibited JNK expression, suggesting modulation of the MAPK pathway. Conclusion Escin exerts therapeutic effects on RA by?inhibiting LPS-induced M1 polarization?and?promoting M2 polarization?in RAW 264.7 macrophages. These effects are mediated through the?suppression of the MAPK signaling pathway?(specifically ERK and JNK), which correlates with reduced TNF-α levels and enhanced anti-inflammatory activity.

  Key words: Escin; anti-inflammation; mechanism of action; RAW264.7 macrophages

  提交时间：2025-04-02

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• 序号	提交日期	编号	操作
  1	2025-03-18	bmr.202504.00003V1	下载

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