Zhang Yi, Guo Hui, Cheng Brian Chi-Yan, Su Tao, Fu Xiu-Qiong, Li Ting, Zhu Pei-Li, Tse Kai-Wing, Pan Si-Yuan, Yu Zhi-Ling
Centre for Cancer and Inflammation Research, School of Chinese Medicine, Hong Kong Baptist University, Kowloon Tong, Hong Kong,
Department of Pharmacology, Beijing University of Chinese Medicine, Beijing, People's Republic of China.
Drug Des Devel Ther. 2018 Sep 4;12:2731-2748. doi: 10.2147/DDDT.S160645. eCollection 2018.
Dingchuan tang (asthma-relieving decoction), a formula of nine herbs, has been used for treating respiratory inflammatory diseases for >400 years in the People's Republic of China. However, the mechanisms underlying the anti-inflammatory action of dingchuan tang is not fully understood. This study aims to investigate the effects of Dingchuan tang essential oil (DCEO) on inflammatory mediators and the underlying mechanism of action.
DCEO was extracted by steam distillation. Lipopolysaccharide (LPS)-stimulated RAW264.7 macrophages were used as the cell model. Production of nitric oxide (NO) was determined by the Griess test. Protein secretion and mRNA levels of inflammatory mediators were measured by the enzyme-linked immunosorbent assay (ELISA) and quantitative real-time polymerase chain reaction (qRT-PCR), respectively. Protein levels were examined by Western blot. Nuclear localization of nuclear factor-kappa B (NF-κB) was detected using immunofluorescence analyses.
DCEO significantly reduced LPS-triggered production of NO and prostaglandin E2 (PGE2) and decreased protein and mRNA levels of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2). LPS induced upregulation of protein and mRNA levels of cytokines (interleukin-1β [IL-1β], interleukin-6 [IL-6], tumor necrosis factor-α [TNF-α]), and chemokines (monocyte chemoattractant protein-1 [MCP-1], chemokine [C-C motif] ligand 5 [CCL-5], and macrophage inflammatory protein [MIP]-1α) were suppressed by DCEO treatment. Phosphorylation and nuclear protein levels of transcription factors (activator protein-1 [AP-1], NF-κB, interferon regulatory factor 3 [IRF3]) were decreased by DCEO. Protein levels of phosphorylated IκB-α, IκB kinase α/β (IKKα/β), phosphatidylinositol 3-kinase (PI3K), protein kinase B (Akt), TGF β-activated kinase 1 (TAK1), TANK-binding kinase 1 (TBK1), extracellular signal-regulated kinase (ERK), p38 mitogen-activated protein kinase (p38), and c-Jun N-terminal kinase (JNK) were lowered by DCEO. Moreover, degradation of interleukin-1 receptor-associated kinase 1 (IRAK1) and IRAK4 induced by LPS was inhibited by DCEO treatment.
Suppression of the interleukin-1 receptor-associated kinase (IRAK)/NF-κB, IRAK/AP-1 and TBK1/IRF3 pathways was associated with the inhibitory effects of DCEO on inflammatory mediators in LPS-stimulated RAW264.7 macrophages. This study provides a pharmacological justification for the use of dingchuan tang in managing inflammatory disorders.
定喘汤是一种由九味草药组成的方剂，在中华人民共和国已用于治疗呼吸道炎性疾病400多年。然而，定喘汤抗炎作用的潜在机制尚未完全明确。本研究旨在探讨定喘汤精油（DCEO）对炎性介质的影响及其潜在作用机制。
采用水蒸气蒸馏法提取DCEO。以脂多糖（LPS）刺激的RAW264.7巨噬细胞作为细胞模型。采用Griess法测定一氧化氮（NO）的产生。分别通过酶联免疫吸附测定（ELISA）和定量实时聚合酶链反应（qRT-PCR）检测炎性介质的蛋白分泌和mRNA水平。通过蛋白质印迹法检测蛋白水平。采用免疫荧光分析检测核因子-κB（NF-κB）的核定位。
DCEO显著降低LPS诱导的NO和前列腺素E2（PGE2）的产生，并降低诱导型一氧化氮合酶（iNOS）和环氧化酶-2（COX-2）的蛋白和mRNA水平。LPS诱导细胞因子（白细胞介素-1β [IL-1β]、白细胞介素-6 [IL-6]、肿瘤坏死因子-α [TNF-α]）和趋化因子（单核细胞趋化蛋白-1 [MCP-1]、趋化因子[C-C基序]配体5 [CCL-5]和巨噬细胞炎性蛋白[MIP]-1α）的蛋白和mRNA水平上调，而DCEO处理可抑制这些上调。DCEO可降低转录因子（激活蛋白-1 [AP-1]、NF-κB、干扰素调节因子3 [IRF3]）的磷酸化和核蛋白水平。DCEO可降低磷酸化IκB-α、IκB激酶α/β（IKKα/β）、磷脂酰肌醇3-激酶（PI3K）、蛋白激酶B（Akt）、转化生长因子β激活激酶1（TAK1）、TANK结合激酶1（TBK1）、细胞外信号调节激酶（ERK）、p38丝裂原活化蛋白激酶（p38）和c-Jun氨基末端激酶（JNK）的蛋白水平。此外，DCEO处理可抑制LPS诱导的白细胞介素-1受体相关激酶1（IRAK1）和IRAK4的降解。
抑制白细胞介素-1受体相关激酶（IRAK）/NF-κB、IRAK/AP-1和TBK1/IRF3信号通路与DCEO对LPS刺激的RAW264.7巨噬细胞中炎性介质的抑制作用有关。本研究为定喘汤用于治疗炎性疾病提供了药理学依据。
