Zhou Kun, Lu Dingyan, You Jingrui, Liu Ting, Sun Jia, Lu Yuan, Pan Jie, Li Yongjun, Liu Chunhua
State Key Laboratory of Functions and Applications of Medicinal Plants, Engineering Research Center for the Development and Application of Ethnic Medicine and TCM (Ministry of Education), Guizhou Medical University, Guiyang, 550004, China; School of Pharmacy, Guizhou Medical University, Guiyang, 550025, China.
State Key Laboratory of Functions and Applications of Medicinal Plants, Engineering Research Center for the Development and Application of Ethnic Medicine and TCM (Ministry of Education), Guizhou Medical University, Guiyang, 550004, China.
J Ethnopharmacol. 2022 Nov 15;298:115624. doi: 10.1016/j.jep.2022.115624. Epub 2022 Aug 12.
Gerberae Piloselloidis Herba (GPH), a commonly used traditional medicine in China, is derived from Gerbera piloselloides (Linn.) Cass. It is featured by its special bioactivities as antitussive, expectorant, anti-asthma, anti-bacterial, anti-tumor, uterine analgesia, and immunity-enhancing. With a long history of medication in ethnic minority areas in China, it is often used as an effective treatment for cough and sore throat as well as allergic asthma. Although our previous investigation also has discovered GPH performed effective treatment on allergic asthma, its underlying mechanism remains unclear.
This research aims to reveal the pharmacological mechanism of GPH in the treatment for allergic asthma through combination of plasma pharmacology and network pharmacology.
Firstly, the components of GPH in blood samples were identified using UHPLC- Q-Orbitrap HRMS. An interaction network of "compound-target-disease" was constructed based on the compounds confirmed in blood and on their corresponding targets of allergic asthma acquired from disease gene databases, predicting the possible biological targets and potential signal pathways of GPH with the network pharmacology analysis. Then, a molecular docking between the blood ingredients and the core targets was carried out using the Autodock Vina software. Subsequently, after establishing a mouse model with allergic asthma induced by ovalbumin (OVA), the effect of GPH on allergic asthma was evaluated by analyzing a series of indicators including behavior, lung pathological changes, inflammatory factors in serum and bronchoalveolar lavage fluid (BALF). Finally, the key pathway and targets predicted by network pharmacology and molecular docking were further verified using Western blot analysis.
Eleven chemical constituents (such as arbutin, neochlorogenic acid, chlorogenic acid, etc.) were identified through the analysis of plasma samples, on which basis a total of 142 genes intersecting GPH and allergic asthma were collected by network pharmacology. After performing enrichment analysis of these genes in gene ontology (GO) and kyoto encyclopedia of genes and genomes (KEGG), it was found that arbutin-related targets mainly focused on phosphatidylinositol 3 kinase (PI3K)/protein kinase B (Akt) signal pathway, while luteolin and marmesin -related targets tended to locate at Interleukin-17 (IL-17) signal pathway. Meanwhile, the findings of molecular docking suggested that such components as arbutin, luteolin and marmesin entering into blood had good binding with the core targets related to PI3K/Akt and IL-17 pathways. In addition, GPH improved the OVA-induced asthma symptoms, the alveolar septa thickening and the infiltration of inflammatory cell around bronchi and bronchioles as well as reduced the levels of IgE, IL-8 and TNF-α in serum or BALF. Furthermore, GPH could inhibit the phosphorylation level of Akt and the expression of PI3K, an efficacy supported by the findings by way of Western blot which suggests that GPH in the treatment of allergic asthma was linked to PI3K/Akt signal pathway.
In this study, a comprehensive strategy to combine the UPLC-Q-Orbitrap HRMS with network pharmacology was employed to clarify the mechanism of GPH against allergic asthma, a finding where GPH may inhibit PI3K/Akt signal pathway to protect mice from OVA-induced allergic asthma. This study provides a deeper understanding of the pharmacological mechanism of GPH in treatment of asthma, offering a scientific reference for further research and clinical application of GPH in terms of allergic asthma.
黄毛紫菀（Gerberae Piloselloidis Herba，GPH）是中国常用的传统药物，来源于毛果一枝香（Gerbera piloselloides (Linn.) Cass.）。它具有止咳、祛痰、抗哮喘、抗菌、抗肿瘤、子宫镇痛和增强免疫力等特殊生物活性。在中国少数民族地区有悠久的用药历史，常用于治疗咳嗽、喉咙痛以及过敏性哮喘。尽管我们之前的研究也发现GPH对过敏性哮喘有有效的治疗作用，但其潜在机制仍不清楚。
本研究旨在通过血浆药理学和网络药理学相结合的方法揭示GPH治疗过敏性哮喘的药理机制。
首先，采用超高效液相色谱-四极杆-轨道阱高分辨质谱（UHPLC-Q-Orbitrap HRMS）鉴定血液样本中GPH的成分。基于血液中确认的化合物及其从疾病基因数据库中获取的过敏性哮喘相应靶点，构建“化合物-靶点-疾病”相互作用网络，通过网络药理学分析预测GPH可能的生物学靶点和潜在信号通路。然后，使用Autodock Vina软件对血液成分与核心靶点进行分子对接。随后，建立卵清蛋白（OVA）诱导的过敏性哮喘小鼠模型，通过分析行为、肺病理变化、血清和支气管肺泡灌洗液（BALF）中的炎症因子等一系列指标，评估GPH对过敏性哮喘的影响。最后，使用蛋白质免疫印迹法（Western blot分析）进一步验证网络药理学和分子对接预测的关键通路和靶点。
通过血浆样本分析鉴定出11种化学成分（如熊果苷、新绿原酸、绿原酸等），在此基础上，通过网络药理学收集到142个GPH与过敏性哮喘相交的基因。对这些基因在基因本体论（GO）和京都基因与基因组百科全书（KEGG）中进行富集分析后发现，熊果苷相关靶点主要集中在磷脂酰肌醇3激酶（PI3K）/蛋白激酶B（Akt）信号通路，而木犀草素和崖椒素相关靶点倾向于定位在白细胞介素-17（IL-17）信号通路。同时，分子对接结果表明，熊果苷、木犀草素和崖椒素等进入血液的成分与PI3K/Akt和IL-17通路相关的核心靶点具有良好的结合。此外，GPH改善了OVA诱导的哮喘症状、肺泡间隔增厚以及支气管和细支气管周围炎症细胞浸润，并降低了血清或BALF中IgE、IL-8和TNF-α的水平。此外，GPH可以抑制Akt的磷酸化水平和PI3K的表达，蛋白质免疫印迹法的结果支持了这一功效，表明GPH治疗过敏性哮喘与PI3K/Akt信号通路有关。
本研究采用超高效液相色谱-四极杆-轨道阱高分辨质谱与网络药理学相结合的综合策略，阐明GPH抗过敏性哮喘的机制，即GPH可能通过抑制PI3K/Akt信号通路保护小鼠免受OVA诱导的过敏性哮喘。本研究为深入了解GPH治疗哮喘的药理机制提供了依据，为GPH在过敏性哮喘方面的进一步研究和临床应用提供了科学参考。
