目的 基于网络药理学和动物实验探究六神凝胶治疗痤疮的作用机制。方法 通过网络药理学筛选六神凝胶的活性成分和作用靶点，采用String数据库对活性成分靶点与痤疮疾病靶点进行蛋白互作网络分析，并通过DAVID数据库对六神凝胶治疗痤疮的靶点进行基因本体（gene ontology，GO）功能及京都基因与基因组百科全书（Kyoto encyclopedia of genes and genomes，KEGG）通路富集分析。大鼠耳部皮内注射痤疮丙酸杆菌联合外部涂抹油酸建立痤疮模型，考察六神凝胶对大鼠耳部外观、病理变化、耳部组织炎性因子水平的影响，采用Western blotting检测大鼠耳部组织Toll样受体2（Toll-like receptor 2，TLR2）/p38/核因子-κB（nuclear factor-κB，NF-κB）通路相关蛋白表达，采用免疫荧光检测大鼠耳部组织NF-κB p65表达。结果 共获得六神凝胶活性成分27个，药物靶点831个，痤疮靶点120个，活性成分与疾病的交集靶点41个。六神凝胶治疗痤疮的主要成分为麝香酮、胆固醇、菜油甾醇、胆酸和吗啡等，核心靶点为肿瘤坏死因子（tumor necrosis factor，TNF）、白细胞介素-1β（interleukin-1β，IL-1β）、瘦素（leptin，LEP）、TLR4和CXC趋化因子配体8（CXC chemokine ligand 8，CXCL8）等，关键信号通路可能包括IL-17信号通路、TLR信号通路、NOD样受体信号通路、C型凝集素受体信号通路、TNF信号通路、NF-κB信号通路等。动物实验结果表明，六神凝胶能够显著缓解大鼠耳部痤疮症状，减轻大鼠耳肿胀程度，显著抑制耳组织中TNF-α、IL-1β、IL-6、IL-8、IL-17、IL-23和基质金属蛋白酶9（matrix metalloprotein 9，MMP9）水平（P＜0.001），并显著降低痤疮组织中TLR2、p38和p-NF-κB p65蛋白表达水平（P＜0.01、0.001）。结论 六神凝胶通过抑制TLR2/p38/NF-κB信号通路的活化，减少炎症因子的分泌，减轻痤疮模型大鼠炎症反应，从而治疗痤疮。

Objective To explore the mechanism of Liushen Gel (六神凝胶) in treatment of acne based on network pharmacology and animal experiments. Methods The active ingredients and targets of Liushen Gel were screened by network pharmacology, and the protein interaction network between the active ingredient targets and acne disease targets was analyzed by String database. Gene ontology (GO) function and Kyoto encyclopedia of genes and genomes (KEGG) pathway enrichment analysis of targets of Liushen Gel for acne treatment were conducted by DAVID database. The rat model of acne was established by intradermal injection of Propionibacterium acnes and external application of oleic acid. The effects of Liushen Gel on the appearance, pathological changes, and levels of inflammatory factors in ear tissue of rats were investigated. Western blotting was used to detect the expressions of Toll-like receptor 2 (TLR2)/p38/nuclear factor-κB (NF-κB) pathway related proteins in ear tissue of rats, and immunofluorescence was used to detect the expression of NF-κB p65 in ear tissue of rats. Results A total of 27 active ingredients from Liushen Gel, 831 drug targets, 120 acne targets and 41 cross targets of active ingredients and diseases were obtained. The main ingredients of Liushen Gel in the treatment of acne were muscone, cholesterol, campesterol, cholalic acid and morphine, the core targets were tumor necrosis factor (TNF), interleukin-1β (IL-1β), leptin (LEP), TLR4 and CXC chemokine ligand 8 (CXCL8), the key signal pathways may included IL-17 signaling pathway, TLR signaling pathway, NOD like receptor signaling pathway, C-type lectin receptor signaling pathway, TNF signaling pathway and NF-κB signaling pathway, etc. The animal experiment results showed that Liushen Gel could significantly alleviate the symptoms of acne in ear of rats, reduce the degree of ear swelling, significantly inhibit the levels of TNF-α, IL-1β, IL-6, IL-8, IL-17, IL-23 and matrix metalloproteinase 9 (MMP9) in ear tissue (P < 0.001), significantly reduce the expression levels of TLR2, p38 and p-NF-κB p65 proteins in acne tissue (P < 0.01, 0.001). Conclusion Liushen Gel could treat acne by inhibiting the activation of TLR2/p38/NF-κB signaling pathway, reducing the secretion of inflammatory factors and alleviating the inflammatory response of acne model rats.

R285.5

雷允上药业集团有限公司技术服务项目（XS-LS-YL-202201）；广东省中医药局项目（20221222）；广州市基础研究计划基础与应用基础研究项目（202201010293）