广西医学

目的基于数据挖掘、网络药理学、分子对接及动物实验探讨中医药改善动脉粥样硬化（AS）脂质代谢的作用机制。方法利用数据挖掘方法筛选中医药干预AS脂质代谢的文献，对纳入文献的处方药物的使用频次和高频药物的性、味、归经进行统计分析，并基于Apriori算法分析高频药物的关联规则，确定使用频次最高的药物组合。通过TCMSP、BATMAN⁃TCM数据库收集使用频次最高药物组合的化学成分及作用靶点，通过GeneCard® 和OMIM® 数据库检索AS脂质代谢相关靶点，通过R语言软件获取药物作用靶点与疾病相关靶点的交集，获得核心靶点。将活性成分及核心靶点数据导入Cytoscape软件，构建“中药活性成分⁃核心靶点”网络。通过STRING数据库构建核心靶点的蛋白⁃蛋白相互作用（PPI）网络图。通过R语言软件和Perl程序对核心靶点进行富集分析，使用AutoDock软件进行分子对接。将27只小鼠随机分为空白组、模型组、半夏陈皮组，各9只。空白组小鼠以普通饲料喂养，模型组、半夏陈皮组小鼠以高脂饲料喂养，持续喂养16 周。喂养8周时，半夏陈皮组小鼠给予半夏陈皮药液 200 μL/d 灌胃，空白组与模型组小鼠给予等体积无菌0.9%氯化钠溶液灌胃，连续给药8周。末次给药2 h后，检测小鼠血脂水平变化，通过油红O染色和HE染色观察小鼠心脏主动脉组织斑块情况，采用Western blot检测小鼠主动脉组织中过氧化物酶体增殖激活受体γ（PPARγ）、肝脏X受体α（LXRα）、三磷酸腺苷结合盒转运体A1（ABCA1）蛋白相对表达水平。结果共检索获得125个中医临床处方，处方药物的药性以温性、寒性为主，药味以苦味、甘味、辛味为主，主要归属于肝经、脾经、心经，半夏⁃陈皮药对为使用频次最高的药物组合。半夏⁃陈皮改善AS脂质代谢的核心靶点共96个。“中药活性成分⁃核心靶点”网络拓扑分析度值排名前5的活性成分为豆甾醇、黄芩素、环阿屯醇、顺式⁃11⁃二十碳烯酸、川陈皮素。PPI网络拓扑分析度值排名前10的靶点包括AKT1、TP53、PPARγ、CASP3、VEGFA、ESR1、FOS、MAPK3、HIF1α、CREB1。富集分析结果显示，核心靶点主要涉及对不同营养水平的反应、对类固醇激素的反应、对金属离子的反应等生物过程，膜筏、膜微区和突触膜等细胞结构，DNA⁃结合转录因子结合、核受体活性、配体激活的转录因子活性等分子功能，脂质与动脉粥样硬化、磷脂酰肌醇3激酶/蛋白激酶B信号通路等信号通路。分子对接结果表明，半夏⁃陈皮药对的核心活性成分黄芩素与PPARγ的结合最为稳定。动物实验结果显示，与模型组相比，半夏陈皮组、空白组小鼠的TG、TC、LDL⁃C水平降低，HDL⁃C水平升高，主动脉粥样硬化斑块相对面积减少，主动脉组织中的PPARγ、LXRα和ABCA1蛋白相对表达水平升高（P<0.05）。结论半夏⁃陈皮药对可通过调控PPARγ⁃LXRα⁃ABCA1信号通路，有效改善AS的脂质代谢。

Objective To investigate the mechanism of Traditional Chinese Medicine in improving lipid metabolism in atherosclerosis (AS) based on data mining, network pharmacology, molecular docking, and animal experiments. Methods Data mining was used to screen literature on Traditional Chinese Medicine intervention in AS lipid metabolism. Statistical analysis was performed on the use frequency of prescription drugs, as well as properties, flavors, and meridian entries of high⁃frequency drugs. The Apriori algorithm was applied to analyze association rules among high⁃frequency drugs and identify the most frequently used drug combination. The chemical components and effect targets of the most high⁃frequency used drug combination were collected using the TCMSP and BATMAN⁃TCM databases. Targets related to AS lipid metabolism were retrieved from the GeneCards® and OMIM® databases. Intersection between drug effect targets and disease⁃related targets were obtained using R language software to identify core targets. The data of the active components and core targets were imported into Cytoscape software to construct a “Traditional Chinese Medicine active component⁃core target” network. A protein⁃protein interaction (PPI) network of core targets was constructed using the STRING database. Enrichment analysis of core targets was performed using R language software and Perl programs, and molecular docking was conducted using AutoDock software. A total of 27 mice were randomly divided into three groups: blank group, model group, and Pinellia ternata⁃Citrus reticulata group, with 9 mice in each group. The blank group was fed a normal diet, while the model group and Pinellia ternata⁃Citrus reticulata group were fed a high⁃fat diet for 16 weeks. At week 8 of feeding, the Pinellia ternata⁃Citrus reticulata group received intragastric administration of 200 μL/d Pinellia ternata⁃Citrus reticulata decoction, while the blank group and model group received an equal volume of sterile 0.9% sodium chloride solution for 8 consecutive weeks. Two hours after the last administration, blood lipid levels in mice were measured. The conditions of aortic plaque in the heart tissue was observed using oil red O staining and HE staining. Western blot was performed to detect the relative protein expressions of peroxisome proliferator⁃activated receptor γ (PPARγ), liver X receptor α (LXRα), and ATP⁃binding cassette transporter A1 (ABCA1) in the aorta tissues of mice. Results A total of 125 Traditional Chinese Medicine clinical prescriptions were retrieved. The properties of the prescription drugs were mainly warm and cold, while the flavors were primarily bitter, sweet, and acrid. These drugs were mainly attributed to the liver, spleen, and heart meridians. The Pinellia ternata⁃Citrus reticulata drug pair was the most frequently used drug combination. A total of 96 core targets of Pinellia ternata⁃Citrus reticulata in improving AS lipid metabolism were identified. The top five active components in the “Traditional Chinese Medicine active component⁃core target” network topology analysis were stigmasterol, baicalein, cycloartenol, cis⁃11⁃eicosenoic acid, and nobiletin. The top 10 targets ranked by degree values in the PPI network topology analysis included AKT1, TP53, PPARγ, CASP3, VEGFA, ESR1, FOS, MAPK3, HIF1α, and CREB1. Enrichment analysis results revealed that core targets were mainly involved in biological processes such as responses to different nutrient levels, steroid hormones, and metal ions; cellular structures such as membrane rafts, membrane microdomains, and synaptic membranes; in addition, molecular functions such as DNA⁃binding transcription factor binding, nuclear receptor activity, and ligand⁃activated transcription factor activity. Enriched signaling pathways included lipid and atherosclerosis, phosphatidylinositol 3⁃kinase/protein kinase B signaling pathway, etc. Molecular docking results indicated that baicalein, the core active component of Pinellia ternata⁃Citrus reticulata, exhibited the most stable binding with PPARγ. Animal experiment results expressed that compared with the model group, the Pinellia ternata⁃Citrus reticulata group and blank group exhibited decreased levels of TG, TC, and LDL⁃C, elevated HDL⁃C level, decreased relative area of aortic atherosclerotic plaques, and elevated relative protein expressions of PPARγ, LXRα, and ABCA1 in the aorta tissues (P<0.05). Conclusion The Pinellia ternata⁃Citrus reticulata drug pair effectively ameliorates lipid metabolism in AS by regulating the PPARγ⁃LXRα⁃ABCA1 signaling pathway.