Table8_Integrated Chemical Interpretation and Network Pharmacology Analysis to Reveal the Anti-Liver Fibrosis Effect of Penthorum chinense.xls

Liver fibrosis is a disease with complex pathological mechanisms. Penthorum chinense Pursh (P. chinense) is a traditional Chinese medicine (TCM) for liver injury treatment. However, the pharmacological mechanisms of P. chinense on liver fibrosis have not been investigated and clarified clearly. This study was designed to investigate the chemicals in P. chinense and explore its effect on liver fibrosis. First, we developed a highly efficient method, called DDA-assisted DIA, which can both broaden mass spectrometry (MS) coverage and MS2 quality. In DDA-assisted DIA, data-dependent acquisition (DDA) and data-independent acquisition (DIA) were merged to construct a molecular network, in which 1,094 mass features were retained in Penthorum chinense Pursh (P. chinense). Out of these, 169 compounds were identified based on both MS1 and MS2 analysis. After that, based on a network pharmacology study, 94 bioactive compounds and 440 targets of P. chinense associated with liver fibrosis were obtained, forming a tight compound–target network. Meanwhile, the network pharmacology experimental results showed that multiple pathways interacted with the HIF-1 pathway, which was first identified involved in P. chinense. It could be observed that some proteins, such as TNF-α, Timp1, and HO-1, were involved in the HIF-1 pathway. Furthermore, the pharmacological effects of P. chinense on these proteins were verified by CCl4-induced rat liver fibrosis, and P. chinense was found to improve liver functions through regulating TNF-α, Timp1, and HO-1 expressions. In summary, DDA-assisted DIA could provide more detailed compound information, which will help us to annotate the ingredients of TCM, and combination with computerized network pharmacology provided a theoretical basis for revealing the mechanism of P. chinense.

肝纤维化（Liver fibrosis）是一种病理机制复杂的疾病。赶黄草（Penthorum chinense Pursh，P. chinense）是用于治疗肝损伤的传统中药（traditional Chinese medicine, TCM），但其抗肝纤维化的药理作用机制尚未得到充分研究与阐明。本研究旨在分析赶黄草中的化学成分，并探究其对肝纤维化的干预效果。首先，我们开发了一种高效的分析方法——DDA辅助DIA（DDA-assisted DIA），该方法可同时拓宽质谱（mass spectrometry, MS）的检测覆盖范围并提升MS2级图谱质量。在DDA辅助DIA策略中，研究融合了数据依赖型采集（data-dependent acquisition, DDA）与数据非依赖型采集（data-independent acquisition, DIA）技术以构建分子网络，最终从赶黄草中鉴定得到1094个质谱特征。基于MS1与MS2级质谱分析，其中169种化合物被成功鉴定。随后，依托网络药理学（network pharmacology）分析，我们筛选得到94种活性化合物及440个与肝纤维化相关的赶黄草作用靶点，构建了紧密的化合物-靶点调控网络。同时，网络药理学实验结果显示，多条通路与低氧诱导因子-1通路（HIF-1 pathway）存在相互作用，本研究首次证实该通路参与赶黄草的药理调控过程。研究观察到，肿瘤坏死因子-α（TNF-α）、金属蛋白酶组织抑制剂-1（Timp1）及血红素氧合酶-1（HO-1）等多种蛋白均参与了HIF-1通路。进一步地，我们通过四氯化碳（CCl4）诱导的大鼠肝纤维化模型验证了赶黄草对上述蛋白的药理作用，发现赶黄草可通过调节TNF-α、Timp1及HO-1的表达改善肝脏功能。综上，DDA辅助DIA技术可提供更为详尽的化学成分信息，有助于传统中药的组分注释；结合计算机辅助网络药理学分析，本研究为阐明赶黄草抗肝纤维化的作用机制提供了理论依据。