DataSheet1_Anti-Inflammatory Effects and Mechanisms of Dandelion in RAW264.7 Macrophages and Zebrafish Larvae.PDF

Dandelions (Taraxacum spp.) play an important role in the treatment of inflammatory diseases. In this study, we investigated the anti-inflammatory effects of Dandelion Extract (DE) in LPS-induced RAW264.7 macrophages and copper sulfate (CuSO4)-induced zebrafish larvae. DE was not toxic to RAW264.7 cells at 75 μg/ml as measured by cell viability, and DE inhibited LPS-induced cell morphological changes as measured by inverted microscopy. In survival experiments, DE at 25 μg/ml had no toxicity to zebrafish larvae. By using an enzymatic standard assay, DE reduced the production of nitric oxide (NO) in LPS-induced RAW264.7 cells. Fluorescence microscopy results show that DE reduced LPS-induced ROS production and apoptosis in RAW264.7 cells. DE also inhibited CuSO4-induced ROS production and neutrophil aggregation in zebrafish larvae. The results of flow cytometry show that DE alleviated the LPS-induced cell cycle arrest. In LPS-induced RAW264.7 cells, RT-PCR revealed that DE decreased the expression of M1 phenotypic genes iNOS, IL-6, and IL-1β while increasing the expression of M2 phenotypic genes IL-10 and CD206. Furthermore, in CuSO4-induced zebrafish larvae, DE reduced the expression of iNOS, TNF-α, IL-6, and IL-10. The findings suggest that DE reduces the LPS-induced inflammatory response in RAW264.7 cells by regulating polarization and apoptosis. DE also reduces the CuSO4-induced inflammatory response in zebrafish larvae.

蒲公英（Taraxacum spp.）在炎症性疾病的治疗中具有重要作用。本研究探讨了蒲公英提取物（DE）在脂多糖（Lipopolysaccharide, LPS）诱导的RAW264.7巨噬细胞及硫酸铜（CuSO₄）诱导的斑马鱼幼鱼中的抗炎效果。通过细胞活力检测可知，75 μg/ml浓度的DE对RAW264.7细胞无毒性；倒置显微镜检测结果显示，DE可抑制LPS诱导的细胞形态学变化。生存实验表明，25 μg/ml浓度的DE对斑马鱼幼鱼无毒性。采用酶学标准检测法，DE可减少LPS诱导的RAW264.7细胞中一氧化氮（nitric oxide, NO）的生成。荧光显微镜结果显示，DE可降低LPS诱导的RAW264.7细胞中活性氧（reactive oxygen species, ROS）的产生及细胞凋亡。DE还可抑制CuSO₄诱导的斑马鱼幼鱼中ROS的生成与中性粒细胞聚集。流式细胞术结果证实，DE可缓解LPS诱导的细胞周期阻滞。在LPS诱导的RAW264.7细胞中，逆转录聚合酶链反应（RT-PCR）检测显示，DE可下调M1表型相关基因诱导型一氧化氮合酶（inducible nitric oxide synthase, iNOS）、白细胞介素-6（interleukin-6, IL-6）及白细胞介素-1β（interleukin-1β, IL-1β）的表达，同时上调M2表型相关基因白细胞介素-10（interleukin-10, IL-10）与CD206的表达。此外，在CuSO₄诱导的斑马鱼幼鱼中，DE可降低iNOS、肿瘤坏死因子-α（tumor necrosis factor-α, TNF-α）、IL-6及IL-10的表达水平。本研究结果提示，DE可通过调控细胞极化状态与细胞凋亡，减轻RAW264.7细胞的LPS诱导炎症反应；同时亦可缓解斑马鱼幼鱼的CuSO₄诱导炎症反应。