Chen Shuaishuai, Xue Weina, Wu Zhongxiu, Lu Dingyan, Zheng Lin, Zhou Meng, Li Yongjun, Wang Yonglin, Liu Ting
State Key Laboratory of Discovery and Utilization of Functional Components in Traditional Chinese Medicine, Engineering Research Center for the Development and Application of Ethnic Medicine and TCM (Ministry of Education), Guizhou Provincial Engineering Research Center for the Development and Application of Ethnic Medicine and TCM, Guizhou Medical University, Guian New Area, 561113, People's Republic of China.
Guizhou Institute of Precision Medicine, Affiliated Hospital of Guizhou Medical University, Guiyang, 550009, People's Republic of China.
J Inflamm Res. 2025 Feb 26;18:2879-2898. doi: 10.2147/JIR.S497166. eCollection 2025.
The total flavonoids of Schltr. (TFPF) exhibit notable anti-rheumatoid arthritis (RA) properties, highlighted by their inhibitory action on TNF-α. This study investigates the specific compounds of TFPF responsible for TNF-α inhibition and delineates their mechanism of action.
L929 cells and luciferase-based assays were used to assess anti-TNF-α activity. Additionally, MH7A cells and Wistar rats were employed to establish in vitro and in vivo models of RA. Chemical chromatography, thermal proteome profiling (TPP), molecular docking, surface plasmon resonance (SPR), cellular thermal shift assays (CETSA), drug affinity responsive target stability tests (DARTS), and transcriptomic analysis were used to study the potent molecules of TFPF and their protein targets and pathways. H&E staining, immunofluorescence staining, ELISA, and Western blot were employed for pharmacodynamic and mechanism studies of these potent molecules.
TFPF mitigated cellular injury in L929 cells and inhibited luciferase expression, both of which were induced by TNF-α. Among the eight compounds identified, quercetin exhibited the most potent anti-TNF-α activity. Quercetin improved cellular injury and reduced the cell apoptosis rate in L929 cells treated with TNF-α. TPP revealed the interaction between quercetin and TNF-α, which was verified by molecular docking, SPR, CETSA, and DARTS assays. Transcriptomic analysis highlighted the TNF signaling pathway, suggesting that quercetin may target TNF-α to inhibit the activation of the NF-κB signaling pathway. Quercetin inhibited luciferase expression, suppressed the phosphorylation of IκBα and p65, blocked p65 nuclear translocation, and reduced the mRNA expression of , and . Moreover, quercetin decreased inflammation and joint damage in RA.
Quercetin, a compound of TFPF, interacts with and inhibits the activity of TNF-α, thereby suppressing the TNF-α/NF-κB signaling pathway and alleviating the symptoms of RA. These findings position quercetin as a promising TNF-α inhibitor for the treatment of RA.
Schltr. 的总黄酮（TFPF）具有显著的抗类风湿性关节炎（RA）特性，对肿瘤坏死因子-α（TNF-α）的抑制作用尤为突出。本研究旨在探究 TFPF 中负责抑制 TNF-α 的具体化合物，并阐明其作用机制。
采用 L929 细胞和基于荧光素酶的检测方法评估抗 TNF-α 活性。此外，利用 MH7A 细胞和 Wistar 大鼠建立 RA 的体外和体内模型。通过化学色谱法、热蛋白质组分析（TPP）、分子对接、表面等离子体共振（SPR）、细胞热位移分析（CETSA）、药物亲和力响应靶点稳定性测试（DARTS）和转录组分析，研究 TFPF 的有效分子及其蛋白质靶点和信号通路。采用苏木精-伊红（H&E）染色、免疫荧光染色、酶联免疫吸附测定（ELISA）和蛋白质免疫印迹法对这些有效分子进行药效学和作用机制研究。
TFPF 减轻了 L929 细胞的细胞损伤并抑制了由 TNF-α 诱导的荧光素酶表达。在所鉴定的 8 种化合物中，槲皮素表现出最有效的抗 TNF-α 活性。槲皮素改善了 TNF-α 处理的 L929 细胞的细胞损伤并降低了细胞凋亡率。TPP 揭示了槲皮素与 TNF-α 之间的相互作用，这通过分子对接、SPR、CETSA 和 DARTS 检测得到了验证。转录组分析突出了 TNF 信号通路，表明槲皮素可能靶向 TNF-α 以抑制核因子-κB（NF-κB）信号通路的激活。槲皮素抑制了荧光素酶表达，抑制了 IκBα 和 p65 的磷酸化，阻止了 p65 的核转位，并降低了 、 和 的 mRNA 表达。此外，槲皮素减轻了 RA 中的炎症和关节损伤。
槲皮素作为 TFPF 的一种化合物，与 TNF-α 相互作用并抑制其活性，从而抑制 TNF-α/NF-κB 信号通路并减轻 RA 的症状。这些发现使槲皮素成为一种有前景的用于治疗 RA 的 TNF-α 抑制剂。
