目的 基于网络药理学、分子对接和动物实验探索丹皮酚治疗酒精性肝病（alcoholic liver disease，ALD）的作用及作用机制。方法 利用PharmMapper数据平台检索丹皮酚相关靶点，在DisGeNET、GeneCards和OMIM疾病数据库筛选ALD相关基因，通过检索GEO数据库获得ALD芯片并筛选差异基因，基于WGCNA筛选与疾病相关的基因模块，在疾病数据库、差异基因及WGCNA分析基因取交集，获得ALD相关基因，与丹皮酚靶点取交集，获得丹皮酚治疗ALD的相关靶点，进行相关性分析及基因本体（gene ontology，GO）功能和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，利用最小绝对收缩和选取算子（least absolute shrinkage and selection operator，LASSO）回归筛选靶点，然后进行分子对接验证。采用Lieber-DeCarli酒精液体饲料自由饮食的方法建立ALD小鼠模型，设置对照组、模型组、水飞蓟宾（36.8 mg/kg）组和丹皮酚高、低剂量（480、120 mg/kg）组，每组8只，给予药物进行干预。通过检测血脂、肝功能、炎症因子水平评估药效作用，采用苏木素-伊红（HE）与油红O染色观察肝脏病理学变化，采用Western blotting和qRT-PCR检测肝组织中脂肪酸结合蛋白4（fatty acid binding protein 4，FABP4）、过氧化物酶体增殖物激活受体α（peroxisome proliferator-activated receptor alpha，PPARα）、肉碱棕榈酰转移酶1A（carnitine palmitoyltransferase 1A，CTP1A）表达水平。结果 筛选药物靶点88个、疾病靶点516个，交集靶点13个。GO和KEGG富集分析共得到835条生物功能和70条通路。通过LASSO回归算法筛选得到7个靶点。分子对接结果显示丹皮酚与FABP4、PPARα结合能均小于−5 kcal/mol。动物实验结果显示，与对照组比较，模型组小鼠肝功能、血脂及炎症因子水平显著升高（P＜0.01、0.001），肝脏组织出现炎症浸润和脂肪空泡；与模型组比较，丹皮酚改善了小鼠的肝功能、血脂及炎症因子水平（P＜0.05、0.01、0.001），恢复了肝细胞的正常结构，减少了肝脏脂肪空泡。Western blotting和qRT-PCR结果显示，与对照组比较，模型组小鼠肝脏组织中FABP4的蛋白和mRNA表达水平显著升高（P＜0.01、0.001），PPARα和CPT1A的蛋白和mRNA表达水平显著降低（P＜0.001）；与模型组比较，丹皮酚显著降低FABP4的蛋白和mRNA表达水平（P＜0.05、0.001），上调PPARα和CPT1A的蛋白和mRNA表达水平（P＜0.05、0.01）。结论 丹皮酚通过下调FABP4并上调PPARα和CPT1A表达以改善脂质代谢。

Objective To explore the effect and mechanism of paeonol in treatment of alcoholic liver disease (ALD) based on network pharmacology, molecular docking and animal experiments. Methods PharmMapper data platform was utilized to retrieve targets related to paeonol. ALD related genes were screened from DisGeNET, GeneCards and OMIM disease databases, ALD chips was obtained by searching GEO databases and screening for differentially expressed genes, disease-related gene modules was screened based on WGCNA, ALD related genes was obtained by analyzing the intersection of genes from disease databases, differentially expressed genes and WGCNA derived genes, and then intersecting with paeonol targets to obtain relevant targets for paeonol in treatment of ALD, correlation analysis and gene ontology (GO) function and Kyoto encyclopedia of genes and genomes (KEGG) pathway enrichment analysis were performed, least absolute shrinkage and selection operator (LASSO) regression was used to screen targets, followed by molecular docking validation. The Lieber DeCarli alcohol liquid feed free diet method was used to establish an ALD mouse model. Control group, model group, silibinin (36.8 mg/kg) group, and paeonol high-, low-dose (480, 120 mg/kg) groups were set up, with eight mice in each group, and drugs were administered for intervention. The pharmacological effect was evaluated by detecting blood lipid levels, liver function indexes and levels of inflammatory factors. Hematoxylin-eosin (HE) and oil red O staining were used to observe pathological changes in liver. Western blotting and qRT-PCR were used to detect the expression levels of fatty acid binding protein 4 (FABP4), peroxisome proliferator activated receptor alpha (PPARα) and carnitine palmitoyltransferase 1A (CPT1A) in liver tissue. Results A total of 88 drug targets, 516 disease targets and 13 intersecting targets were screened. A total of 835 biological functions and 70 pathways were obtained by GO and KEGG enrichment analysis. Seven targets were screened by LASSO regression algorithm. Molecular docking results showed that the binding energies of paeonol to FABP4 and PPARα were less than −5 kcal/mol. The results of animal experiments showed that compared with control group, levels of liver function indexes, blood lipid levels and inflammatory factors in model group mice were significantly increased (P < 0.01, 0.001), inflammation infiltration and fat vacuoles were appeared in liver tissue. Compared with model group, paeonol improved the levels of liver function indexes, blood lipid levels and inflammatory factors in mice (P < 0.05, 0.01, 0.001), restored the normal structure of liver cells, and reduced hepatic fat vacuoles. Western blotting and qRT-PCR results showed that compared with control group, the protein and mRNA expression levels of FABP4 in liver tissue of model group mice were significantly increased (P < 0.01, 0.001), while the protein and mRNA expression levels of PPARα and CPT1A were significantly decreased (P < 0.001). Compared with model group, paeonol significantly reduced the protein and mRNA expression levels of FABP4 (P < 0.05, 0.001), and up-regulated the protein and mRNA expression levels of PPARα and CPT1A (P < 0.05, 0.01). Conclusion Paeonol improves lipid metabolism by down-regulating FABP4 and up-regulating PPARα and CPT1A expressions.

R285.5

安徽省自然科学基金资助项目（2022AH050521）