Xanthatin alleviates LPS-induced inflammatory response in RAW2647 macrophages by inhibiting toll-like receptor 4 pathway

Xanthatin (XT) is a sesquiterpene lactone isolated from the Chinese herb Xanthium, which belongs to the Asteraceae family. Here, we developed an in-flammation model via stimulating RAW 264.7 cells with LPS, which was applied to assess the anti-inflammatory effect and probable mechanisms of xanthatin. Pretreatment with of xanthatin lowered the amount of NO and associated pro-inflammatory factors (TNF-α, IL-1β and IL-6), which were enhanced in the cells after LPS-activated. Interestingly, RAW 264.7 cells were pre-treated with xanthatin, the mRNA expression of iNOS, COX-2, TNF-α, IL-1β and IL-6 was downregulated and repressed as compared with the only LPS-induced group. Furthermore, phosphorylated levels of related proteins (STAT3, ERK1/2, SAPK/JNK, IκBα, p65) were notably increased with the only LPS-activated cells. Interestingly, these phosphorylated proteins could be reverted by pre-treating with xanthatin in a dose-dependent way. Additionally, xanthatin was involved to block NF-κB p65 from translocating into the nucleus and activating inflamma-tory gene transcription. Collectively, these results demonstrated that xanthatin suppresseeds the inflammatory effect caused by LPS through downregulating NF-κB, MAPK and STATs signaling pathways. Taken together, xanthatin pos-sesses the potential to act as a good anti-inflammatory medication candidate.苍耳素（Xanthatin, XT）是从菊科（Asteraceae）药用植物苍耳（Xanthium）中分离得到的倍半萜内酯。本研究通过脂多糖（LPS）刺激RAW 264.7细胞构建炎症模型，用以评估苍耳素的抗炎活性及其潜在作用机制。苍耳素预处理可降低脂多糖激活后细胞中升高的一氧化氮（NO）水平，同时下调相关促炎因子（肿瘤坏死因子-α（TNF-α）、白细胞介素-1β（IL-1β）与白细胞介素-6（IL-6)）的分泌量。值得注意的是，与仅经脂多糖诱导的对照组相比，苍耳素预处理的RAW 264.7细胞中，诱导型一氧化氮合酶（iNOS）、环氧合酶-2（COX-2）、TNF-α、IL-1β及IL-6的mRNA表达均被显著下调抑制。此外，仅经脂多糖激活的细胞内，信号传导与转录激活因子3（STAT3）、细胞外调节蛋白激酶1/2（ERK1/2）、应激活化蛋白激酶/c-Jun氨基末端激酶（SAPK/JNK）、核因子κB抑制蛋白α（IκBα）及p65等相关蛋白的磷酸化水平显著升高。有趣的是，苍耳素预处理可通过剂量依赖性方式逆转上述磷酸化蛋白的异常升高。进一步研究发现，苍耳素可阻断核因子κB p65（NF-κB p65）向细胞核内转位，进而抑制炎症基因的转录激活。综上，本研究结果表明，苍耳素可通过下调核因子κB（NF-κB）、丝裂原活化蛋白激酶（MAPK）及信号传导与转录激活因子（STATs）信号通路，抑制脂多糖诱导的炎症反应。综上所述，苍耳素具备开发为优质抗炎候选药物的潜力。