Network pharmacology and molecular docking of endogenous active metabolites in diabetic kidney disease

Network pharmacology and molecular docking were used to predict endogenous active metabolites with protective effects in diabetic kidney disease (DKD). We utilized metabolomics to screen differentially expressed metabolites in kidney tissues of mice with type 2 DKD and predicted potential targets using relevant databases. The interaction network between endogenous active metabolites and target proteins was established by integrating differentially expressed metabolites and proteins associated with DKD identified through proteomics. Gene ontology (GO) and signaling pathway enrichment analysis were performed. The biological functions of the active candidate metabolites and their effects on downstream pathways were also verified. Metabolomics revealed 130 differentially expressed metabolites. Through co-expression network analysis coupled with the investigation of differentially expressed proteins in proteomics, 2-hydroxyphenylpropionylglycine (2-HPG) emerged as a key regulator of DKD. 2-HPG was found to modulate the progression of DKD by regulating the conformation and activity of synaptophysin 1 (SYNJ1), with a correlation coefficient of 0.974. <i>In vivo</i> experiments revealed that SYNJ1 expression was significantly downregulated in the Macroalbuminuria Group compared to the Control Group and negatively correlated with proteinuria (<i>r</i> = −0.7137), indicating its important role in DKD progression. Immunofluorescence demonstrated that treatment with 2-HPG restores the expression of the foot process marker protein Wilms tumor-1 (WT-1) in podocytes injured by high glucose levels. Western blot and polymerase chain reaction support the involvement of SYNJ1 in this process. This study demonstrated the significance of the 2-HPG/SYNJ1 signaling axis in safeguarding the foot process of podocytes in DKD.

本研究采用网络药理学与分子对接技术，预测糖尿病肾病（Diabetic Kidney Disease, DKD）中具有保护作用的内源性活性代谢物。我们利用代谢组学（metabolomics）筛选2型DKD小鼠肾组织中的差异表达代谢物，并通过相关数据库预测潜在作用靶点。通过整合蛋白质组学（proteomics）鉴定得到的与DKD相关的差异表达代谢物与蛋白质，构建内源性活性代谢物与靶蛋白的相互作用网络。开展基因本体论（Gene Ontology, GO）与信号通路富集分析，并验证活性候选代谢物的生物学功能及其对下游通路的调控作用。代谢组学分析共筛选得到130种差异表达代谢物。通过共表达网络分析结合蛋白质组学中差异表达蛋白的研究，2-羟苯基丙酰甘氨酸（2-hydroxyphenylpropionylglycine, 2-HPG）被确定为DKD的关键调控因子。研究发现2-HPG通过调节突触膜蛋白1（synaptophysin 1, SYNJ1）的构象与活性，调控DKD的进展，二者相关系数达0.974。体内实验（in vivo）结果显示，与对照组相比，大量白蛋白尿组小鼠的SYNJ1表达显著下调，且与尿蛋白水平呈负相关（r = -0.7137），表明其在DKD进展中发挥重要作用。免疫荧光实验证实，2-HPG处理可恢复高糖损伤足细胞中足突标志物肾母细胞瘤蛋白1（Wilms tumor-1, WT-1）的表达。蛋白质印迹（Western blot）与聚合酶链式反应（polymerase chain reaction, PCR）实验证实，SYNJ1参与了这一调控过程。本研究证实了2-HPG/SYNJ1信号轴在保护DKD小鼠足细胞足突中的重要作用。