Xiong Jinrui, Wu Yu, Luo Liuling, Shen Xiaofei, Zeng Yong, Meng Xianli, Zhang Hai
State Key Laboratory of Southwestern Chinese Medicine Resources, Innovative Institute of Chinese Medicine and Pharmacy, Chengdu University of Traditional Chinese Medicine, Chengdu, 611137, China; College of Pharmacy, Chengdu University of Traditional Chinese Medicine, Chengdu, 611137, China.
Hospital of Chengdu University of Traditional Chinese Medicine, Chengdu, 610075, China.
J Ethnopharmacol. 2025 Mar 13;343:119461. doi: 10.1016/j.jep.2025.119461. Epub 2025 Feb 7.
As a natural medicine, Xanthii Fructus (XF) is widely used in traditional Chinese medicine (TCM) and Tibetan medicine. It has been demonstrated to alleviate allergic rhinitis (AR) in modern research. However, the specific molecular mechanism underlying its treatment of AR is still unclear.
To elucidate the effect and mechanism of XF in treating AR through network pharmacology and experimental validation.
In the present study, blood-entry components of XF were analyzed using UPLC-Orbitrap-HRMS. Then, we conducted pharmacodynamic studies in vitro and vivo. In vitro study, Human IL-4 was used to treat HNEpCs cells to establish a vitro model. Subsequently, HNEpCs cells were administrated with XF extracts (0.5, 1, 2 mg/ml). And ovalbumin (OVA) was employed to establish an allergic rhinitis model, and different doses of XF (8, 16, 32 mg/kg) were administered by gavage to BABL/c mice for in vivo experiments. Next, the Swiss Target Prediction database was employed to acquire blood-entry components targets. Meanwhile, from OMIM and GeneCards databases, AR-related targets were obtained. A protein-protein interaction (PPI) network was established through the STRING database, and potential pathways of XF were identified through Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis. In the end, the results of network pharmacology were experimental validation in vivo and vitro experiments.
Fifteen compounds were identified, most of which were phenolic acids. In pharmacodynamic studies, the vitro study revealed that XF-treated gave rise to a significant decline of iNOS and COX2 protein expression in inflammatory conditions, as evidenced by Western blot results, and there was a sharp decline in the mRNA levels of TNF-α, IL-1β and IL-6. Meanwhile, the vivo studies demonstrated that XF exhibited favorable therapeutic efficacy against AR, as evidenced by a decrease in IgE, TNF-α, IL-4, and IL-6 levels in mice serum, an improvement in nasal mucosal injury pathology. Based on these findings, through network pharmacology, we identified 14 core AR-related targets, including HIF-1α, STAT3, TLR4. Using KEGG pathway analysis, it has been revealed that XF can alleviate AR through JAK2/STAT3/HIF-1α signaling pathway. Therefore, further experiments were conducted to verify the molecular mechanism of the anti-AR effect of XF. A decline of the phosphorylation of JAK2, STAT3 and HIF-1α proteins was observed, which resulted in the suppression of JAK2/STAT3/HIF-1α signaling pathway. These findings were corroborated by the same results obtained through IF. The results were verified by RT-qPCR, which demonstrated that XF was capable of downregulating the mRNA levels of TSLP and CCL11. Then, the conclusions were further reinforced with the introduction of WP1066. It could be observed that XF inhibited the STAT3 nuclear translocation. Finally, a restoration of p-JAK2, p-STAT3, HIF-1α expression levels to normal levels in AR mice.
The combined findings led to the conclusion that XF play its therapeutic role in AR by suppressing the JAK2/STAT3/HIF-1α signaling pathway.
作为一种天然药物，苍耳子在中药和藏药中被广泛应用。现代研究已证明其可缓解变应性鼻炎（AR）。然而，其治疗AR的具体分子机制仍不清楚。
通过网络药理学和实验验证阐明苍耳子治疗AR的作用及机制。
在本研究中，采用超高效液相色谱-轨道阱-高分辨质谱（UPLC-Orbitrap-HRMS）分析苍耳子的入血成分。然后，我们进行了体外和体内药效学研究。体外研究中，用人白细胞介素-4（IL-4）处理人鼻上皮细胞（HNEpCs）以建立体外模型。随后，用苍耳子提取物（0.5、1、2mg/ml）处理HNEpCs细胞。并用卵清蛋白（OVA）建立变应性鼻炎模型，不同剂量的苍耳子（8、16、32mg/kg）经灌胃给予BABL/c小鼠用于体内实验。接下来，利用瑞士靶点预测数据库获取入血成分靶点。同时，从在线孟德尔遗传人类疾病数据库（OMIM）和基因卡片数据库（GeneCards）获取AR相关靶点。通过STRING数据库建立蛋白质-蛋白质相互作用（PPI）网络，并通过基因本体论（GO）和京都基因与基因组百科全书（KEGG）富集分析确定苍耳子的潜在作用途径。最后，在体内和体外实验中对网络药理学结果进行实验验证。
鉴定出15种化合物，其中大多数为酚酸类。在药效学研究中，体外研究表明，Western blot结果显示，苍耳子处理可使炎症条件下诱导型一氧化氮合酶（iNOS）和环氧化酶2（COX2）蛋白表达显著下降，肿瘤坏死因子-α（TNF-α）、白细胞介素-1β（IL-1β）和白细胞介素-6（IL-6）的mRNA水平急剧下降。同时，体内研究表明，苍耳子对AR具有良好的治疗效果，小鼠血清中免疫球蛋白E（IgE）、TNF-α、IL-4和IL-6水平降低，鼻黏膜损伤病理改善。基于这些发现，通过网络药理学，我们确定了14个与AR相关的核心靶点，包括缺氧诱导因子-1α（HIF-1α）、信号转导和转录激活因子3（STAT3）、Toll样受体4（TLR4）。通过KEGG通路分析发现，苍耳子可通过Janus激酶2/信号转导和转录激活因子3/HIF-1α信号通路缓解AR。因此，进一步进行实验以验证苍耳子抗AR作用的分子机制。观察到JAK2、STAT3和HIF-1α蛋白磷酸化水平下降，导致JAK2/STAT3/HIF-1α信号通路受到抑制。免疫荧光（IF）获得的相同结果证实了这些发现。实时定量聚合酶链反应（RT-qPCR）验证了结果，其表明苍耳子能够下调胸腺基质淋巴细胞生成素（TSLP）和CC趋化因子配体11（CCL11）的mRNA水平。然后，通过引入WP1066进一步加强了结论。可以观察到苍耳子抑制了STAT3核转位。最后，AR小鼠中磷酸化JAK2、磷酸化STAT3、HIF-1α表达水平恢复至正常水平。
综合研究结果得出结论，苍耳子通过抑制JAK2/STAT3/HIF-1α信号通路在AR中发挥治疗作用。
