Department of Gastroenterology, Affiliated Hospital of Nantong University, Nantong University, NO.20, Xisi Road, Nantong 226001, Jiangsu, China.
Department of Gastroenterology, Affiliated Hospital of Nantong University, Nantong University, NO.20, Xisi Road, Nantong 226001, Jiangsu, China.
Phytomedicine. 2024 Jul;129:155564. doi: 10.1016/j.phymed.2024.155564. Epub 2024 Mar 22.
The incidence of ulcerative colitis (UC) is on the rise globally and the development of drugs targeting UC is urgent. Finding the target of action of natural products is important for drug discovery, elucidation of drug action mechanism, and disease mechanism. San-Ye-Qing (SYQ), is an ancient herbal medicine, but whether the powder of its rhizome has pharmacological effects against UC and its mechanism of action are not clear.
To evaluate the therapeutic effectiveness of rhizome powder of SYQ in treating UC, and conduct an isolation and characterization of the chemical constituents of the powder. Further, screen the most potent compounds among them and determine the potential mechanism for treating UC.
In vivo, the therapeutic effect of SYQ's rhizome powder on UC was assessed by mice's body weight, DAI score, colon length, tissue MPO activity, serum inflammatory markers, etc. Additionally, HPLC was used to isolate and identify the specific chemical components of SYQ's rhizome powder. Then, the most effective compounds and their therapeutic targets were analysed and screened in SYQ rhizome powder using network pharmacology, combined with CCK-8 assay, NO release assay and molecular docking assay, in conjunction with CETSA, DARTS, SPR and enzyme activity assay. Finally, the biological effects of the key compound on the targets were validated using Western blot and ELISA.
In vivo, SYQ rhizome powder effectively restored mice's body weight, lowered DAI and pathological score, downregulated the expression of inflammatory biomarkers, and restored colon length, as well as the colonic epithelial and mucus barriers. Afterward, 9 compounds were isolated and identified from the powder of the rhizomes of SYQ by HPLC. Nicotiflorin is the primary compound in SYQ with the highest concentration. According to both CCK-8 and NO release tests, Nicotiflorin is also the most efficacious compound. Combined with network pharmacological prediction, molecular docking analysis, CETSA, DARTS, SPR and enzyme activity assay, Nicotiflorin may ultimately suppress inflammation by targeting p65 and inhibiting the NF-κB pathway, thereby attenuating the activation of NLRP3 inflammasome. To verify this conclusion, Western blot and ELISA experiments were conducted.
Our results suggest that the extract from SYQ rhizomes has therapeutic properties for UC. Its active ingredient Nicotiflorin exerted potent anti-UC effects by binding to p65 and inhibiting the activation of NF-κB and NLRP3 inflammasomes.
溃疡性结肠炎（UC）的发病率在全球呈上升趋势，针对 UC 的药物研发迫在眉睫。寻找天然产物的作用靶点对于药物发现、药物作用机制阐明和疾病机制研究具有重要意义。三叶青（SYQ）是一种古老的草药，但它的根茎粉是否对 UC 具有药理作用及其作用机制尚不清楚。
评价 SYQ 根茎粉治疗 UC 的疗效，并对其化学成分进行分离和表征。进一步筛选其中最有效的化合物，并确定其治疗 UC 的潜在机制。
体内实验，通过小鼠体重、DAI 评分、结肠长度、组织 MPO 活性、血清炎症标志物等评估 SYQ 根茎粉对 UC 的治疗效果。此外，采用 HPLC 法分离鉴定 SYQ 根茎粉的特定化学成分。然后，采用网络药理学结合 CCK-8 检测、NO 释放检测和分子对接检测，联合 CETSA、DARTS、SPR 和酶活性检测，分析筛选 SYQ 根茎粉中最有效的化合物及其治疗靶点。最后，采用 Western blot 和 ELISA 验证关键化合物对靶标的生物学效应。
体内实验结果表明，SYQ 根茎粉能有效恢复小鼠体重，降低 DAI 和病理评分，下调炎症标志物的表达，恢复结肠长度，以及结肠上皮和黏液屏障。随后，通过 HPLC 从 SYQ 根茎粉中分离鉴定出 9 种化合物。其中，野黄芩苷含量最高，为主要成分。根据 CCK-8 和 NO 释放试验，野黄芩苷也是最有效的化合物。结合网络药理学预测、分子对接分析、CETSA、DARTS、SPR 和酶活性检测，野黄芩苷可能通过靶向 p65 并抑制 NF-κB 通路，从而抑制 NLRP3 炎症小体的激活，发挥抑制炎症的作用。为验证这一结论，进行了 Western blot 和 ELISA 实验。
本研究结果表明，SYQ 根茎提取物对 UC 具有治疗作用。其活性成分野黄芩苷通过与 p65 结合并抑制 NF-κB 和 NLRP3 炎症小体的激活，发挥强大的抗 UC 作用。
