Hou Wen, Wei Bao, Liu Hong Sheng
NHC Key Laboratory of Critical Care Medicine, Tianjin First Central Hospital, Tianjin, China.
Department of Surgery, Children's Hospital, Tianjin, China.
Front Pharmacol. 2021 Nov 11;12:756259. doi: 10.3389/fphar.2021.756259. eCollection 2021.
mixture (PNM) has the characteristics of multicomponent, multitarget, and multieffect, which can cope with the multidirectional and multidimensional complex pathological process caused by hepatic ischemia/reperfusion injury (HIRI). Our animal experiments showed that PNM composed of notoginseng, dogwood, and white peony root could significantly reduce the level of aspartate transaminase and alanine aminotransferase in the blood of mice with HIRI, indicating that this preparation had a protective effect on HIRI in mice. Therefore, on this basis, the molecular mechanism of PNM intervention in HIRI was further explored by network pharmacology. First, target genes corresponding to active components and HIRI were obtained through databases such as TCMSP, Pharm Mapper, Swiss Target Prediction, GeneCards, and so on. All target genes were standardized by Uniprot database, and a total of 291 target genes with their intersection were obtained. Then, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways and biological processes (BPs) of 291 target genes were obtained through the online public platform of DAVID. A total of 177 KEGG pathways and 337 BPs were obtained by setting < 0.01 and false discovery rate <0.05. The network mapping map of components and disease targets was drawn by Cytoscape, and the top 10 Hub target genes related to HIRI were obtained. At the same time, the String database was used to obtain the protein-protein interaction dataset, which was imported into Cytoscape, and the first 10 Hub target genes were obtained. The Hub target genes obtained by the above two methods were molecular docking with their corresponding small molecule compounds through DockThor online tool. The results showed that the docking of paeoniflorin with glyceraldehyde 3-phosphate dehydrogenase (GAPDH), paeoniflorin and loganin with SRC, ginsenoside Rb1 with NR3C2, ursolic acid and oleanolic acid with IL-6, paeoniflorin docking VEGFA, and MMP9. Finally, NR3C2, SRC, and GAPDH were identified as target genes in this study by referring to relevant literature reports. After verification by immunohistochemical experiments, compared with the sham group, the above three target genes were highly expressed in the HIRI group ( < 0.01). Compared with the HIRI group, the expression of three target genes in the PNM + HIRI group was significantly decreased ( < 0.01). The results showed that PNM could protect mouse HIRI by decreasing the expression of NR3C2, SRC, and GAPDH.
复方（PNM）具有多成分、多靶点、多效应的特点，能够应对肝脏缺血/再灌注损伤（HIRI）所导致的多向性和多维性复杂病理过程。我们的动物实验表明，由三七、山茱萸和白芍组成的PNM可显著降低HIRI小鼠血液中天冬氨酸转氨酶和丙氨酸转氨酶的水平，表明该制剂对小鼠HIRI具有保护作用。因此，在此基础上，通过网络药理学进一步探究PNM干预HIRI的分子机制。首先，通过中药系统药理学数据库与分析平台（TCMSP）、中药靶点预测（Pharm Mapper）、瑞士靶点预测（Swiss Target Prediction）、基因卡片（GeneCards）等数据库获取活性成分与HIRI对应的靶基因。所有靶基因通过通用蛋白质资源（Uniprot）数据库进行标准化处理，共获得291个有交集的靶基因。然后，通过DAVID在线公共平台获取291个靶基因的京都基因与基因组百科全书（KEGG）通路和生物学过程（BP）。设定P<0.01和错误发现率<0.05，共获得177条KEGG通路和337个BP。利用Cytoscape绘制成分与疾病靶点的网络映射图，得到与HIRI相关的前10个核心靶基因。同时，利用搜索工具检索相互作用基因数据库（String）获取蛋白质-蛋白质相互作用数据集，导入Cytoscape，得到前10个核心靶基因。通过DockThor在线工具将上述两种方法得到的核心靶基因与其相应的小分子化合物进行分子对接。结果显示，芍药苷与甘油醛-3-磷酸脱氢酶（GAPDH）对接，芍药苷和马钱子苷与原癌基因酪氨酸蛋白激酶（SRC）对接，人参皂苷Rb1与核受体亚家族3成员C2（NR3C2）对接，熊果酸和齐墩果酸与白细胞介素-6（IL-6）对接，芍药苷与血管内皮生长因子A（VEGFA）和基质金属蛋白酶9（MMP9）对接。最后，参考相关文献报道，确定本研究中的靶基因为NR3C2、SRC和GAPDH。经免疫组化实验验证，与假手术组相比，上述三个靶基因在HIRI组中高表达（P<0.01）。与HIRI组相比，PNM+HIRI组中三个靶基因的表达显著降低（P<0.01）。结果表明，PNM可通过降低NR3C2、SRC和GAPDH的表达来保护小鼠HIRI。
