基于网络药理学对藏药四味辣根菜汤散抗慢性缺氧性气管炎症的药效物质基础及作用机制研究

Hypoxia play an important role in inflammatory microenvironment. Chronic hypoxia in inflammatory tissue will be casued by inflammation. Another, transcription factor hypoxia-inducible factor (HIF-1α) induced by chronic hypoxia has been demonastrated to further induce and aggravate inflammation by activation of NF-κ-B，Toll-like receptor and other signaling way. And inflammation would be further aggravated by hypoxia in plateau, even more, inhabitant with lung disease living in plateau has shorter lifetime than those living in low altitude. Tibet medicine Siwei-Lagencai-Tang-San has been used to treat and have good effect on chronic bronchitis in the Qinghai-Tibet plateau for centuries. We propose a hypothesis that Tibet medicine Siwei-Lagencai-Tang-San should treat chronic bronchitis by synergism of anti-inflammation and anti-hypoxia. The aim of this study is to clarify that how the compatibility of medicines influence the solubility and absorption of ingredients of Siwei-Lagencai-Tang-San, and to clarify the effect meterial and mechanism based on human bronchial epithelial cell (BEAS-2B) and rat models of chronic bronchitis in hypoxia using serum medicinal chemistry and network pharmacology methods. At last, a reliable and effective method would be established for screening the potentially bioactive components of Tibet medicine Siwei-Lagencai-Tang-San. And all of those will provide a novel approach of "drug-discovery of chronic hypoxic diseases and second development of Tibet medicines.

低氧在炎症的发展过程中扮演着重要的角色,炎症通过释放炎症因子导致炎症部位的慢性缺氧,并通过转录因子NF-k-B激活HIF-1α的转录,而由于缺氧诱导HIF-1α的表达可激活转录因子NF-k-B及其他炎症因子在炎症部位的表达，加剧炎症的发展。而高原缺氧，更能促使炎症的发展过程，使高原人因肺部疾病而死亡的平均年龄比平原提前。藏药四味辣根菜汤散在高原低氧环境下广泛用于慢性气管炎症的治疗、疗效确切，推测该方从抗炎和调节低氧信号通路两方面发挥协同药效作用。本课题从该方中诸药配伍对化学成分溶出和吸收的影响、入血化学成分、网络药理学，并进一步通过低氧培养人支气管细胞模型（BEAS-2B）和高海拔低氧环境下大鼠慢性气管炎模型来研究其药效，以期阐明四味辣根菜汤散治疗慢性缺氧性气管炎症药效物质基础和作用机理，为高原缺氧性疾病治疗用药和藏药二次开发提供新思路。

本项目利用UPLC-Q-Exactive 四级杆-静电场轨道阱高分辨质谱对宽果丛菔、岩白菜、唐古特红景天、甘草、紫草茸的主要化学成分进行了分析。对索罗西汤水提取物的主要化学成分分析及各成分的药材归属。进一步确定了索洛西汤中能直接吸收入血的化学成分25个。并通过反相分子对接，构建了 “入血化学成分-靶点-通路”作用网络图。并建立了索洛西汤水提取物中主要化学成分熊果苷、没食子酸、岩白菜素、甘草苷、异甘草素、甘草酸的含量测定方法。.通过烟熏的方法成功建立慢性支气管炎大鼠模型。经索罗西汤药物干预后，显著降低大鼠血液中WBC、中性粒细胞和单核细胞计数，降低支气管肺泡灌洗液中炎症因子TNF-α、IL-8和IL-13的浓度，此外还能明显减轻肺组织中炎性细胞浸润和淋巴聚集，抑制支气管上皮杯状细胞数量的增减及黏液的分泌，表现出显著的抗炎活性。.并对其肺组织进行了蛋白质组学研究。发现药物干预所涉及的信号通路主要有NF-κB Signaling Pathway、Cytosolic DNA-sensing Pathway、B Cell Receptor Signaling Pathway、Fc epsilon RI Signaling Pathway、Calcium Signaling Pathway。在机制研究方面，发现索洛西汤主要通过抑制IL13-STAT6信号通路实现抑制气管黏液的分泌，通过NF-κB信号通路实现其抗炎作用。.此外，发现索洛西汤对低氧诱导的肺动脉高压也具有明显的干预作用，亦能干预肺小血管的重构作用。而且发现唐古特红景天对低氧诱导的肺动脉高压具有明显的干预作用，且对肺小血管重构具有抑制作用，其主要机制为下调CDK4、 CyclinD1蛋白的表达、上调P27Kip1蛋白，从而阻断肺动脉平滑肌细胞的增殖。.阐明四味辣根菜汤散治疗慢性缺氧性气管炎症作用机理，为高原缺氧性疾病治疗用药和藏药二次开发提供新思路。