复方玉液汤通过调控PI3K/Akt信号通路抑制糖尿病大鼠心肌细胞凋亡和炎症反应

Compound Yuye Decoction protects diabetic rats against cardiomyopathy by inhibiting myocardial apoptosis and inflammation via regulating the PI3K/Akt signaling pathway

目的运用网络药理学技术分析复方玉液汤防治糖尿病心肌病(DCM)的作用机制,并通过动物实验进行验证。方法利用TCMSP数据库检索复方玉液汤所含6种中药(黄芪、山药、知母、葛根、五味子、天花粉)中的化学成分,根据口服生物利用度(OB)和药物相似性(DL)筛选出活性化学成分以及与DCM相关的靶点;利用Drugbank、Gene Cards、OMIM和PharmGKb数据库获取与DCM相关靶点;STRING数据库对核心靶点进行PPI网络的构建和分析;Metascape进行核心靶点的GO和KEGG富集分析;利用Cytoscape3.9.1构建“中药-关键成分-核心靶点-关键通路”网络,并对核心靶点对接的关键成分进行分子对接。建立糖尿病心肌病Wistar大鼠动物模型,将大鼠分为正常组、模型组、玉液汤低剂量组(YYT-低组,0.29 g/kg)、玉液汤高剂量组(YYT-高组,1.15 g/kg),15只/组,药物剂量为生药含量。给药8周后观察各组大鼠的心脏组织病理切片,检测各组大鼠心脏电生理变化、血清中的LDH、CK、CK-MB含量变化以及心脏组织中PI3K、P-PI3K、Akt、P-AKT、BAX、IL-6、TNF-α蛋白的表达情况。结果筛选后得到复方玉液汤中61个活性化合物,1057个靶点;3682个DCM相关疾病靶点,共同靶点551个。根据筛选获得的核心靶点富集得到的核心通路发现,凋亡和炎症及相关PI3K/Akt信号通路可能是治疗DCM的关键信号通路;分子对接结果显示,金色酰胺醇酯、山柰酚等活性成分与AKT1和PIK3R1的结合活性较好。利用动物实验对筛选的靶点及通路进行验证,结果显示,与DCM模型组大鼠比较,复方玉液汤能改善模型大鼠心肌组织细胞紊乱及炎性浸润的病理特征,降低模型大鼠血清中LDH、CK、CK-MB水平及模型大鼠心脏组织中BAX、IL-6、TNF-α的蛋白含量并升高P-PI3K和P-AKT的蛋白含量(P<0.05)。结论复方玉液汤防治DCM是多成分、多靶点和多信号通路共同作用的结果。复方中主要活性成分金色酰胺醇酯、山柰酚等通过调控PI3K/Akt信号通路,抑制DCM发病过程中心肌细胞凋亡及炎症反应,发挥对DCM的保护作用。

Objective To explore the therapeutic mechanism of compound Yuye Decoction against diabetic cardiomyopathy(DCM).Methods Drugbank,Gene Cards,OMIM and PharmGKb databases were used to obtain DCM-related targets,and the core targets were identified and functionally annotated by protein-protein interaction network analysis followed by GO and KEGG enrichment analysis.The"Traditional Chinese Medicine-Key Component-Key Target-Key Pathway"network was constructed using Cytoscape 3.9.1,and molecular docking was carried out for the key components and the core targets.In the animal experiment,Wistar rat models of DCM were treated with normal saline or Yuye Decoction by gavage at low(0.29 g/kg)and high(1.15 g/kg)doses for 8 weeks,and the changes in cardiac electrophysiology and histopathology were evaluated.The changes in serum levels of LDH,CK,and CK-MB were examined,and myocardial expressions of PI3K,P-PI3K,Akt,P-AKT,BAX,IL-6,and TNF-α were detected using Western blotting.Results We identified 61 active compounds in Yuye Decoction with 1057 targets,3682 DCM-related disease targets,and 551 common targets between them.Enrichment of the core targets suggested that apoptosis,inflammation and the PI3K/Akt pathways were the key signaling pathways for DCM treatment.Molecular docking studies showed that the active components in Yuye Decoction including gold amidohydroxyethyl ester and kaempferol had strong binding activities with AKT1 and PIK3R1.In DCM rat models,treatment with Yuye Decoction significantly alleviated myocardial pathologies,reduced serum levels of LDH,CK and CK-MB,lowered myocardial expressions of BAX,IL-6 and TNF-α,and increased the expressions of P-PI3K and P-AKT.Conclusion The therapeutic effect of compound Yuye Decoction against DCM is mediated by its multiple active components that act on multiple targets and pathways to inhibit cardiomyocyte apoptosis and inflammatory response by regulating the PI3K/Akt signaling pathway.