International Institute for Translational Chinese Medicine, School of Pharmaceutical Science, Guangzhou University of Chinese Medicine, Guangzhou, Guangdong, China.
School of Pharmacy, Jiangxi University of Traditional Chinese Medicine, Nanchang, China.
J Ethnopharmacol. 2023 Dec 5;317:116852. doi: 10.1016/j.jep.2023.116852. Epub 2023 Jun 28.
Rheumatoid arthritis (RA) is a common difficult disease with a high disability rate. Siegesbeckia orientalis L. (SO), a Chinese medicinal herb that is commonly used for treating RA in clinical practice. While, the anti-RA effect and the mechanisms of action of SO, as well as its active compound(s) have not been elucidated clearly.
We aim to explore the molecular mechanism of SO against RA by using network pharmacology analysis, as well as the in vitro and in vivo experimental validations, and to explore the potential bioactive compound(s) in SO.
Network pharmacology is an advanced technology that provides us an efficient way to study the therapeutic actions of herbs with the underlying mechanisms of action delineated. Here, we used this approach to explore the anti-RA effects of SO, and then the molecular biological approaches were used to verify the prediction. We first established a drug-ingredient-target-disease network and a protein-protein interaction (PPI) network of SO-related RA targets, followed by the Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses. Further, we used lipopolysaccharide (LPS)-stimulated RAW264.7 macrophages and vascular endothelial growth factor-A (VEGFA)-induced human umbilical vein endothelial cell (HUVEC) models, as well as adjuvant-induced arthritis (AIA) rat model to validate the anti-RA effects of SO. The chemical profile of SO was also determined by using the UHPLC-TOF-MS/MS analysis.
Network pharmacology analysis highlighted inflammatory- and angiogenesis-related signaling pathways as promising pathways that mediate the anti-RA effects of SO. Further, in both in vivo and in vitro models, we found that the anti-RA effect of SO is at least partially due to the inhibition of toll like receptor 4 (TLR4) signaling. Molecular docking analysis revealed that luteolin, an active compound in SO, shows the highest degree of connections in compound-target network; moreover, it has a direct binding to the TLR4/MD-2 complex, which is confirmed in cell models. Besides, more than forty compounds including luteolin, darutoside and kaempferol corresponding to their individual peaks were identified tentatively via matching with the empirical molecular formulae and their mass fragments.
We found that SO and its active compound luteolin exhibit anti-RA activities and potently inhibit TLR4 signaling both in vitro and in vivo. These findings not only indicate the advantage of network pharmacology in the discovery of herb-based therapeutics for treating diseases, but also suggest that SO and its active compound(s) could be developed as potential anti-RA therapeutic drugs.
类风湿性关节炎（RA）是一种常见的难治性疾病，致残率高。苍耳草（SO）是一种中药，在临床上常用于治疗 RA。然而，SO 的抗 RA 作用及其作用机制以及其活性化合物尚未得到明确阐明。
本研究旨在通过网络药理学分析，以及体外和体内实验验证，探讨 SO 治疗 RA 的分子机制，并探讨 SO 中的潜在活性化合物。
网络药理学是一种先进的技术，为我们提供了一种有效的方法来研究草药的治疗作用及其作用机制。在这里，我们使用这种方法来探讨 SO 对 RA 的治疗作用，然后使用分子生物学方法来验证预测。我们首先建立了一个 SO 相关 RA 靶点的药物-成分-靶标-疾病网络和一个蛋白质-蛋白质相互作用（PPI）网络，然后进行基因本体（GO）和京都基因与基因组百科全书（KEGG）通路富集分析。此外，我们使用脂多糖（LPS）刺激的 RAW264.7 巨噬细胞和血管内皮生长因子-A（VEGFA）诱导的人脐静脉内皮细胞（HUVEC）模型以及佐剂诱导的关节炎（AIA）大鼠模型来验证 SO 的抗 RA 作用。SO 的化学特征也通过使用 UHPLC-TOF-MS/MS 分析来确定。
网络药理学分析突出了炎症和血管生成相关信号通路作为介导 SO 抗 RA 作用的有前途的途径。此外，在体内和体外模型中，我们发现 SO 的抗 RA 作用至少部分是由于抑制 Toll 样受体 4（TLR4）信号。分子对接分析表明，木犀草素是 SO 中的一种活性化合物，在化合物-靶标网络中显示出最高的连接度；此外，它与 TLR4/MD-2 复合物直接结合，这在细胞模型中得到了证实。此外，通过与经验分子公式及其质量片段匹配，我们初步鉴定出 40 多种化合物，包括木犀草素、达鲁特皂苷和山奈酚，分别对应于各自的峰。
我们发现 SO 和其活性化合物木犀草素在体外和体内均表现出抗 RA 活性，并能有效抑制 TLR4 信号。这些发现不仅表明网络药理学在发现治疗疾病的草药疗法方面具有优势，还表明 SO 和其活性化合物可能被开发为潜在的抗 RA 治疗药物。
