背景:神灵白术(SLBZ)公式,经典中药(TCM)配方,已被广泛用于治疗抗生素相关性腹泻(AAD)。然而,潜在的药理机制尚未得到彻底研究.
目的:探讨SLBZ治疗AAD的缓解机制,我们在体外和体内进行了网络药理学分析和实验验证。
方法:在本研究中,通过超高效液相色谱-质谱(UHPLC-MS)和在线数据库鉴定了SLBZ的主要化合物。通过网络药理学对活性成分的作用靶点和AAD相关靶点进行预测,并获得了SLBZ抗AAD的潜在靶标。然后在蛋白质-蛋白质相互作用(PPI)分析后识别核心靶标。基于这些,进行了基因本体论(GO)和京都基因和基因组百科全书(KEGG)信号通路分析,并对关键途径进行了筛选。随后,使用AutoDockVina进行分子对接,以找到在该途径中起关键作用的关键成分。通过Gromacs软件进行分子动力学模拟以检测结合模式。最后,结果得到了体外和体内实验的证实。
结果:UHPLC-MS检测SLBZ共66种活性成分,通过网络药理学分析筛选出128种活性成分。此外,获得935个药物靶标和1686个AAD相关靶标。选择78个交叉基因作为潜在的治疗靶标,挖掘19个基因作为核心靶标。富集分析显示PI3K-AKT信号通路是SLBZ抗AAD的关键通路。拓扑分析进一步揭示了JAK2、MTOR、TLR4和SYK是SLBZ影响PI3K-AKT通路的关键靶点,SLBZ的52个成分与它们相关。分子对接和动力学模拟揭示了MTOR和皂苷元之间的强结合亲和力。随后,SLBZ治疗后,JAK2、MTOR、在AAD模型大鼠中发现TLR4和SYK显著上调(p<0.05)。细胞实验进一步验证了MTOR与薯不上皂苷元的良好结合能力。
结论:我们证明SLBZ对AAD的治疗作用部分是通过抑制PI3K-AKT途径实现的。
BACKGROUND: Shenlingbaizhu (SLBZ) formula, a classical traditional Chinese medicinal (TCM) formula, has been widely used for treating antibiotic-associated diarrhea (AAD). However, the underlying pharmacological mechanisms have not yet been investigated thoroughly.
OBJECTIVE: To explore the remission mechanism of SLBZ in the treatment of AAD, we conducted network pharmacological analysis and experimental validation in vitro and in vivo.
METHODS: In this study, the main compounds of SLBZ were identified by ultra-high-performance liquid chromatography-mass spectroscopy (UHPLC-MS) and online databases. The targets of the active components and AAD-related targets were predicted by network pharmacology, and the potential targets of SLBZ against AAD were obtained. Then the core targets were recognized after Protein-Protein Interaction (PPI) analysis. Based on these, gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) signaling pathway analyses were conducted, and the key pathway was screened. Subsequently, molecular docking was performed using Auto Dock Vina to find the key components that played a crucial role in that pathway. Molecular dynamics simulation was performed by Gromacs software to detect the binding mode. Finally, the results were confirmed by in vitro and in vivo experiments.
RESULTS: A total of 66 active ingredients of SLBZ were detected by UHPLC-MS, and 128 active ingredients were screened out by network pharmacological analysis. Additionally, 935 drug targets and 1686 AAD-related targets were obtained. Seventy-eight intersected genes were selected as potential therapeutic targets and 19 genes were excavated as core targets. Enrichment analysis revealed PI3K-AKT signaling pathway was the key pathway in SLBZ against AAD. Topological analysis further revealed that JAK2, MTOR, TLR4, and SYK were the key targets affected by SLBZ on the PI3K-AKT pathway, and 52 components of SLBZ were associated with them. Molecular docking and dynamics simulation revealed strong binding affinities between MTOR and diosgenin. Subsequently, after SLBZ treatment, the expression levels of JAK2, MTOR, TLR4, and SYK were found significantly upregulated in the AAD model rats (p < 0.05). The cell experiment further validated the good binding ability between MTOR and diosgenin.
CONCLUSIONS: We demonstrate that the therapeutic effect of SLBZ on AAD was achieved in part by inhibiting the PI3K-AKT pathway.