Utilizing network pharmacology and experimental validation to investigate the underlying mechanism of Sophora davidii resists aging

Technical Summary: Mechanism of Sophora davidii in Resisting Aging

Problem Statement
Global population aging presents significant public health challenges, with endothelial cell dysfunction serving as a critical factor in the aging process and associated chronic diseases. While Sophora davidii flower (SDF), a traditional Chinese medicine abundant in Guizhou, is known for its anti-aging properties and rich content of flavonoids and alkaloids (such as matrine), the specific molecular mechanisms underlying its anti-aging effects remain unclear. This study aims to elucidate the therapeutic mechanisms of SDF against aging, specifically focusing on human umbilical vein endothelial cells (HUVECs).

Methodology
The research employed a dual approach combining network pharmacology and experimental validation:

1. Network Pharmacology Analysis:

   • Target Acquisition: Potential targets of SDF were predicted using six databases (SwissTargetPrediction, TCSMP, BATMAN-TCM, PharmMapper, SuperPred, and TargetNet). Aging-related targets were retrieved from GeneCards, TTD, PharmGKB, OMIM, CTD, and the NCBI GEO database.
   • Intersection and Network Construction: Venn diagrams identified 81 intersecting targets between SDF and aging. A Protein-Protein Interaction (PPI) network was constructed using the STRING database and visualized via Cytoscape to identify core targets based on degree values.
   • Enrichment and Docking: Gene Ontology (GO) and KEGG pathway analyses were performed using Metascape. Molecular docking (AutoDock Vina) was conducted to evaluate the binding affinity between the top five active ingredients (quercetin, genistein, luteolin, kaempferol, isorhamnetin) and key target proteins.

2. Experimental Validation (In Vitro):

   • Model Establishment: An aging model of HUVECs was induced using 0.2 mmol/L H₂O₂ for 24 hours.
   • Treatment: Cells were treated with varying concentrations of SDF extract (SDFE) at 0.5, 1, and 2 mg/mL.
   • Assays:
     • Senescence Assessment: β-galactosidase staining was used to quantify the percentage of senescent cells.
     • Functional Assessment: A tube formation assay evaluated the angiogenic capability of the cells.
     • Molecular Analysis: Western blotting was employed to detect protein expression levels of TP53, p-AKT/AKT, and p-PI3K/PI3K.

Key Contributions and Results

• Target Identification: The study identified 81 potential SDF anti-aging targets. The PPI network highlighted TP53, AKT1, TNF, ESR1, and IL6 as the top five core targets, with TP53 identified as a pivotal regulator.
• Molecular Docking: Active ingredients, particularly luteolin, isorhamnetin, and genistein, demonstrated strong binding affinities (binding energies ranging from -4.9 to -7.24 kcal/mol) with the TP53 protein, primarily through hydrogen bonds and π-π stacking interactions.
• Anti-Aging Efficacy:
  • Senescence Inhibition: SDFE treatment significantly reduced the positive rate of β-galactosidase staining in H₂O₂-induced HUVECs, indicating a suppression of cellular senescence.
  • Functional Recovery: The tube formation assay revealed that SDFE enhanced the tube-forming capability of HUVECs, suggesting a restoration of endothelial function.
• Mechanistic Pathway: Western blot analysis confirmed that SDFE treatment led to a significant, dose-dependent decrease in the expression levels of p53, p-AKT/Akt, and p-PI3K/PI3K compared to the control group.

Significance and Claims
The paper concludes that Sophora davidii flower extract effectively inhibits HUVEC aging. The proposed mechanism involves the regulation of the TP53-PI3K/AKT signaling pathway. Specifically, the study suggests that SDF modulates this pathway to delay the aging process of vascular endothelial cells.

The authors position this work as providing a scientific basis for the application of SDF in anti-aging therapies. They note that while previous studies have linked components like matrine to delaying age-related diseases, this study offers new perspectives by clarifying the specific network pharmacology and cellular mechanisms of the whole extract. The authors state that further research is required to delve deeper into how SDFE specifically affects HUVECs and the TP53-PI3K/Akt pathway, maintaining a modest scope regarding future implications.