Department of Integrative Biotechnology, Biomedical Institute for Convergence at SKKU, Sungkyunkwan University, Suwon 16419, Korea.
Department of Biocosmetics, Sungkyunkwan University, Suwon 16419, Korea.
Biomolecules. 2020 Apr 15;10(4):611. doi: 10.3390/biom10040611.
Alverine, a smooth muscle relaxant, is used to relieve cramps or spasms of the stomach and intestine. Although the effects of alverine on spontaneous and induced contractile activity are well known, its anti-inflammatory activity has not been fully evaluated. In this study, we investigated the anti-inflammatory effects of alverine in vitro and in vivo. The production of nitric oxide (NO) in RAW264.7 cells activated by lipopolysaccharide (LPS) or polyinosinic:polycytidylic acid (poly (I:C)) was reduced by alverine. The mRNA expression of inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), and tumor necrosis factor-α (TNF-α) was also dose-dependently inhibited by treatment with alverine. In reporter gene assays, alverine clearly decreased luciferase activity, mediated by the transcription factor nuclear factor κB (NF-κB) in TIR-domain-containing adapter-inducing interferon-β (TRIF)- or MyD88-overexpressing HEK293 cells. Additionally, phosphorylation of NF-κB subunits and upstream signaling molecules, including p65, p50, AKT, IκBα, and Src was downregulated by 200 μM of alverine in LPS-treated RAW264.7 cells. Using immunoblotting and cellular thermal shift assays (CETSAs), Src was identified as the target of alverine in its anti-inflammatory response. In addition, HCl/EtOH-stimulated gastric ulcers in mice were ameliorated by alverine at doses of 100 and 200 mg/kg. In conclusion, alverine reduced inflammatory responses by targeting Src in the NF-κB pathway, and these findings provide new insights into the development of anti-inflammatory drugs.
平滑肌松弛剂阿魏酸可用于缓解胃和肠的痉挛或抽搐。尽管阿魏酸对自发性和诱导性收缩活动的作用已被充分了解，但它的抗炎活性尚未得到充分评估。在这项研究中，我们研究了阿魏酸在体外和体内的抗炎作用。阿魏酸可降低脂多糖 (LPS) 或聚肌苷酸：聚胞苷酸 (poly (I:C)) 激活的 RAW264.7 细胞中一氧化氮 (NO) 的产生。阿魏酸还可剂量依赖性地抑制诱导型一氧化氮合酶 (iNOS)、环氧化酶-2 (COX-2) 和肿瘤坏死因子-α (TNF-α) 的 mRNA 表达。在报告基因检测中，阿魏酸明显降低了 TIR 结构域包含衔接诱导干扰素-β (TRIF) 或 MyD88 过表达的 HEK293 细胞中转录因子核因子 κB (NF-κB) 介导的荧光素酶活性。此外，在 LPS 处理的 RAW264.7 细胞中，200 μM 的阿魏酸可下调 NF-κB 亚基和上游信号分子（包括 p65、p50、AKT、IκBα 和 Src）的磷酸化。使用免疫印迹和细胞热转移分析 (CETSA)，鉴定 Src 是阿魏酸在抗炎反应中的作用靶点。此外，阿魏酸在 100 和 200 mg/kg 剂量下可改善 HCl/EtOH 刺激的小鼠胃溃疡。总之，阿魏酸通过靶向 NF-κB 通路中的 Src 来减轻炎症反应，这些发现为开发抗炎药物提供了新的思路。
