从凋亡网络的调控研究地黄饮子治疗阿尔兹海默病的分子机制

Researches on molecular mechanism of Dihuang Yinzi Decoction for application to Alzheimer’s disease from regulation of apoptotic network

目的利用中药网络药理学和分子生物学从凋亡网络的调控研究地黄饮子(DHYZ)治疗阿尔兹海默病(AD)的分子机制。方法利用网络药理学方法筛选和预测DHYZ的活性成分治疗AD的潜在作用靶点,构建药物-靶标-疾病网络和蛋白与蛋白相互作用网络,运用生物信息学分析DHYZ调控凋亡网络的主要靶点和信号途径;Aβ1-42处理构建AD细胞模型,使用不同浓度含药血清处理,运用MTT法检测细胞活率,流式细胞术检测细胞凋亡情况,qPCR和Western blotting检测细胞凋亡相关基因和蛋白的表达。结果筛选出DHYZ中108种活性成分有222个AD的潜在治疗靶点,其中有202个靶点之间存在的3737种蛋白蛋白相互作用关系,DHYZ对凋亡网络的调控是其防治AD的重要生物学途径,预测其核心调控靶点有TNF、AKT1、MAPK3、NFKB1、TP53、CASP3等,qPCR和Western blotting验证了预测结果。细胞实验证明不同浓度DYHZ含药血清呈浓度依赖性抑制Aβ1-42处理的SH-SY5Y细胞凋亡,且与调控细胞凋亡相关基因和蛋白有关。结论DHYZ活性化合物对细胞凋亡相关的多个靶点和多条信号通路的调控是其防治AD的重要作用机制。

Objective To confirm evidence regarding the potential therapeutic effect of Dihuang Yinzi Decoction(DHYZ)on Alzheimer’s disease(AD)from the regulation of apoptotic network using network pharmacology and molecular biology.Methods The active ingredients of DHYZ were screened and the potential therapeutic targets on AD were predicted,and the drug-target-disease network and protein-protein interactions(PPI)network were built by using network pharmacology.The main targets and signaling pathways of apoptotic networks of DHYZ were performed by bioinformatics analysis.AD cell model was bulit by Aβ1-42 treating serum containing different concentrations.The cell viability was detected by MTT assay.The cell apoptosis was detected by flow cytometry.The cell apoptosis related gene and protein express were detected by qPCR and Western blotting.Results A total of 108 active ingredients of DHYZ were screened out,and 222 potential therapeutic targets for AD were predicted by network pharmacology.Among the 222 potential therapeutic targets,202 of them had 3737 kinds of PPI network,which were closely related to the apoptotic network signaling pathways and biological processes regulation,involving TNF,AKT1,MAPK3,NFKB1,TP53,CASP3,etc.The predicted results were verified by qPCR and Western blotting.Meanwhile,cell experiments showed that DHYZ contained serum of different concentrations inhibited apoptosis o f Aβ1-42-induced SH-SY5Y cells in a concentration-dependent,which was related to the regulation of apoptosis related genes and proteins.Conclusion The results indicated that the active compounds in DHYZ interact with apoptosis-related multiple targets and multiple pathways,which is an important mechanism of DHYZ for application to AD.