基于线粒体功能探讨温脾通络开窍方治疗阿尔茨海默病的分子机制

Exploration of the Molecular Mechanism of Wenpi Tongluo Kaiqiao Prescription in Treating Alzheimer's Disease Based on Mitochondrial Function

目的基于生物信息学、网络药理学、免疫浸润分析和机器学习等方法并结合实验验证探讨温脾通络开窍方调节线粒体功能治疗阿尔茨海默病(Alzheimer’s disease,AD)的分子机制。方法获取差异表达基因(differentially expressed genes,DEGs)进行相关性分析、KEGG分析以及免疫浸润分析。对公用数据集样品聚类分类后分析免疫细胞差异,构建机器学习模型、筛选特征基因并构建风险预测列线图模型。建立AD大鼠模型,进行水迷宫实验、HE染色及RT-qPCR检测,对数据挖掘的结果进行动物实验验证。结果DEGs相互联系、调节免疫系统并调控P53(tumor protein 53,p53)信号通路。结果显示有13个差异表达基因,并和免疫细胞在亚型分布上有差异。最佳机器学习模型是支持向量机模型(support vector machine,SVM),评分前5的特征基因是MAOB、MAOA、CASP9、Bcl-2、ABAT。风险预测列线图模型的准确性高,预测误差的风险小。动物实验结果显示中药组大鼠学习认知功能障碍和神经损伤比模型组明显减轻,特征基因的mRNA相对表达量与数据挖掘的结果具有一致性。结论温脾通络开窍方可能是通过13个差异性分子靶点相互网络调控、P53信号通路和免疫调控作用实现调节线粒体功能以缓解AD症状。

Objective Based on bioinformatics,network pharmacology,immune infiltration analysis,machine learning and other methods,combined with experimental verification,the molecular mechanism of Wenpi Tongluo Kaiqiao prescription regulating mitochondrial function in the treatment of Alzheimer's disease(AD)was discussed.Methods The differentially expressed genes were obtained for correlation analysis,KEGG pathway enrichment analysis and immune infiltration analysis.After clustering and classifying the public data set samples,the differences of immune cells were analyzed,the machine learning model was constructed,the characteristic genes were screened and the risk prediction nomogram model was verified.The AD rat model was established,and water maze test,HE staining and RT-qPCR were performed to further verify the conclusion of data mining.Results DEGs interacted with each other,regulated the immune system and regulated the P53(tumor protein 53,p53)signaling pathway.The results showed that there were 13 differentially expressed genes,which were different from the distribution of immune cells in the subtypes.The best machine learning model was SVM,and the top 5 feature genes were MAOB,MAOA,CASP9,Bcl-2 and ABAT.The nomogram prediction model had high accuracy and low risk of prediction error.In animal experiments,the results showed that the learning and cognitive dysfunction and nerve injury in the Chinese medicine group were significantly reduced compared with the model group.The relative mRNA expressions of the characteristic genes were consistent with the results of data mining.Conclusion The mechanism of Wenpi Tongluo Kaiqiao prescription alleviating AD symptoms by regulating mitochondrial function may be based on the mutual network regulation,P53 signaling pathway and immune regulation of 13 different molecular targets.