目的 通过数据库预测结合体外细胞实验探讨四妙丸调血脂的作用及机制。方法 通过中药系统药理学（TCMSP）数据库与分析平台、GeneCards、Disgenet、OMIM等数据库筛选四妙丸关键成分及高脂血症相关靶点；蛋白质-蛋白质相互作用（PPI）分析四妙丸治疗高脂血症的关键靶点；基因本体（GO）和京都基因与基因组百科全书（KEGG）分析此过程涉及的生物学过程及通路；进一步采用分子对接探讨四妙丸成分与关键靶点间的相互作用。体外培养HepG2细胞，采用CCK-8实验筛选游离脂肪酸造模和四妙丸、辛伐他汀给药的最佳浓度，构建游离脂肪酸诱导的HepG2细胞高脂血症模型，给予四妙丸（25、50、100 mg·L^-1）、辛伐他汀（25 μmol·L^-1）处理24 h，对照组不造模不加药，模型组不加药，通过细胞油红O染色和细胞内总胆固醇（TC）、三酰甘油（TG）含量测定研究细胞内脂质积累情况；实时荧光定量PCR（qRT-PCR）和Westernblotting检测磷脂酰肌醇4,5-二磷酸3-激酶催化亚基α（PIK3CA）、丝氨酸/苏氨酸激酶1（AKT1）、过氧化物酶体增殖物激活受体γ（PPARG）和细胞色素7A1（CYP7A1） mRNA和蛋白表达。结果 共筛选出56个四妙丸核心成分，对应蛋白靶点735个；高脂血症疾病靶点共1 860个；有效成分靶点与疾病靶点的交集靶点为244个。PPI结果显示核心靶点包括AKT1、IL6、EGFR、TNF、PIK3CA、PPARG等。GO和KEGG富集分析揭示四妙丸主要通过PI3K-AKT1信号通路、胰岛素抵抗、炎症反应信号等改善高脂血症。细胞实验结果表明，与模型组比较，四妙丸显著降低游离脂肪酸诱导的HepG2高脂血症细胞模型内脂质沉积（P＜0.05、0.01），显著降低细胞内TC、TG的含量（P＜0.05、0.01）；显著降低细胞内PIK3CA、AKT1mRNA和蛋白表达水平（P＜0.05、0.01），显著升高PPARG、CYP7A1 mRNA和蛋白表达水平（P＜0.05、0.01）。结论 四妙丸改善游离脂肪酸诱导的HepG2细胞高脂血症，作用机制与调节PIK3CA/AKT1/PPARG/CYP7A1信号通路有关。

Objective To explore the lipid-lowering effect and mechanism of Simiao Pill through database prediction and in vitro cell experiments. Methods The key components of Simiao Pill and the targets related to hyperlipidemia were screened through databases such as Traditional Chinese Medicine Systems Pharmacology (TCMSP) Database and Analysis Platform and GeneCards, the key targets of Simiao Pill in treating hyperlipidemia were analyzed by protein-protein interaction (PPI), the biological processes and pathways involved in this process were analyzed by Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG), molecular docking was further used to explore the interaction between the components of Simiao Pill and the key targets. HepG2 cells were cultured in vitro, and the CCK-8 assay was used to screen the optimal concentrations of free fatty acid modeling and Simiao Pill and simvastatin administration. A free fatty acid-induced HepG2 cell hyperlipidemia model was constructed, and the cells were treated with Simiao Pill (25, 50, 100 mg·L^-1) and simvastatin (25 μmol·L^-1) for 24 h. The control group was not modeled and not treated with drugs, and the model group was not treated with drugs. The intracellular lipid accumulation was studied by oil red O staining and determination of total cholesterol (TC) and triglyceride (TG) content in cells. The mRNA and protein expressions of phosphatidylinositol 4,5-bisphosphate 3-kinase catalytic subunit alpha (PIK3CA), serine/threonine kinase 1 (AKT1), peroxisome proliferator-activated receptor gamma (PPARG), and cytochrome P450 7A1 (CYP7A1) were detected by real-time fluorescence quantitative PCR (qRT-PCR) and Western blotting. Results A total of 56 core components of Simiao Pill were screened, corresponding to 735 protein targets; there were 1 860 disease targets for hyperlipidemia; the intersection targets of the effective component targets and disease targets were 244. The PPI results showed that the core targets included AKT1, IL6, EGFR, TNF, PIK3CA, PPARG, et al. GO and KEGG enrichment analysis revealed that Simiao Pill mainly improved hyperlipidemia through the PI3K-AKT signaling pathway, insulin resistance, and inflammatory response signaling. The cell experiment results showed that compared with the model group, Simiao Pill significantly reduced lipid deposition in the free fatty acid-induced HepG2 hyperlipidemia cell model (P <0.05, 0.01), significantly reduced the intracellular TC and TG content (P <0.05, 0.01); significantly reduced the mRNA and protein expression levels of PIK3CA and AKT1 in cells (P <0.05, 0.01), and significantly increased the mRNA and protein expression levels of PPARG and CYP7A1 (P <0.05, 0.01). Conclusion Simiao Pill improves free fatty acid-induced hyperlipidemia in HepG2 cells, and its mechanism of action is related to the regulation of the PIK3CA/AKT1/PPARG/CYP7A1 signaling pathway.

R285.5

广东省中医药研究开发重点实验室开放基金项目（ KFKT02-002）；广东省中医药局开放项目（20233004）