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孙辉,刘翠玲,姚静,何秋婷,谢月,梁奇.葛根素对LPS诱导的RAW264.7细胞炎症反应的作用机制研究[J].湖南中医药大学学报英文版,2023,43(4):627-632.[Click to copy
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| 葛根素对LPS诱导的RAW264.7细胞炎症反应的作用机制研究 |
| 孙辉,刘翠玲,姚静,何秋婷,谢月,梁奇 |
| (广州中医药大学附属宝安区中医院, 广东 深圳 518101) |
| 摘要: |
| 目的 探讨葛根素（puerarin, PUE）在脂多糖（lipopolysaccharide, LPS）诱导的RAW264.7巨噬细胞中的抗炎作用，并揭示其分子机制。方法 采用CCK-8比色法分别设置不同浓度的LPS和PUE，观察对RAW264.7细胞活性的影响，筛选出合适的LPS造模浓度及PUE的给药浓度。将细胞分为正常对照组、模型组（1 μg/mL）、PUE给药组（12.5、25、50 μmol/L）。Griess法检测一氧化氮（nitric oxide, NO）释放量；ELISA法检测肿瘤坏死因子-α（tumor necrosis factor-α, TNF-α）、白细胞介素-1β（interleukin-1β, IL-1β）和白细胞介素-6（interleukin-6, IL-6）的含量；qRT-PCR法检测环氧化酶-2（cyclooxygenase-2, COX-2） mRNA水平；Western blot法检测诱导型一氧化氮合酶（inducible nitric oxide synthase, iNOS）、COX-2、p-IκBα、p-p65蛋白表达水平。结果 与正常对照组比较，不同浓度的LPS诱导24 h，RAW264.7细胞存活率降低，但对细胞增殖无影响（P＞0.05），PUE在浓度100 μmol/L及以下时对RAW264.7细胞增殖也无影响（P＞0.05）；与模型组相比，PUE给药组（12.5、25、50 μmol/L）NO、TNF-α、IL-1β及IL-6的释放量显著降低（P<0.01），COX-2的蛋白及mRNA表达水平显著降低（P<0.05），iNOS、p-IκBα、p-p65的蛋白表达水平呈浓度依赖性降低（P<0.01）。结论 PUE的体外抗炎作用机制与抑制核因子κB（nuclear factor-κB, NF-κB）信号通路相关。 |
| 关键词: 葛根素 脂多糖 RAW264.7细胞 炎症 一氧化氮 一氧化氮合酶 NF-κB信号通路 |
| DOI：10.3969/j.issn.1674-070X.2023.04.008 |
| Received:November 25, 2022 |
| 基金项目:广东省粤深青年联合基金项目（2019A1515111192）。 |
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| Anti-inflammatory effects and mechanisms of Puerarin on LPS-induced RAW264.7 macrophages |
| SUN Hui,LIU Cuiling,YAO Jing,HE Qiuting,XIE Yue,LIANG Qi |
| (Bao'an TCM Hospital of Guangzhou University of Chinese Medicine, Shenzhen, Guangdong 518101, China) |
| Abstract: |
| Objective To explore the anti-inflammatory potential of Puerarin (PUE) in lipopolysaccharide (LPS)-induced RAW264.7 macrophages and reveal its potential molecular mechanisms.Methods The effects of different concentrations of LPS and PUE on the viability of RAW264.7 cells were determined by CCK-8 colorimetric method, and the appropriate LPS modeling concentration and PUE administration concentration were screened. Cells were divided into normal control group, model group (1 μg/mL), and PUE administration group (12.5, 25, 50 μmol/L); Griess assay was used to assess the nitric oxide (NO) concentration; tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β) and interleukin-6 (IL-6) content was measured by ELISA; cyclooxygenase-2 (COX-2) mRNA levels were detected by qRT-PCR; the protein expression levels of inducible nitric oxide synthase (iNOS), COX-2, p-IκBα, and p-p65 were determined by Western blot.Results Compared with the normal control group, the survival rate of RAW264.7 cells decreased after 24 h of LPS induction at different concentrations, and there was no effect on cell proliferation (P>0.05), PUE has no significant cytotoxic effects at or below the concentrations of 100 μmol/L (P>0.05); Compared with the model group, PUE administration group (12.5, 25, 50 μmol/L) displayed favorable inhibitory effects on the release level of NO, TNF-α, IL-1β and IL-6 (P<0.01), and the protein and mRNA expression levels of COX-2 significantly decreased (P<0.05) in LPS-induced RAW264.7 cells, and it is the same for the expression levels of iNOS, p-IκBα, p-p65 (P<0.01).Conclusion The anti-inflammation mechanisms of PUE is related with nuclear factor-κB (NF-κB) signaling pathway. |
| Key words: puerarin lipopolysaccharide RAW264.7 cells inflammation nitric oxide inducible nitric oxide synthase NF-κB signaling pathway |
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