Decoding Cholesterol Metabolic Regulation by Naotai Formula in Stroke Management: An Integrated Network Pharmacology...

Naotai formula (NTF) is clinically used for stroke treatment, yet its molecular mechanisms involving vascular and metabolic regulation remain unclear. This study combines network pharmacology (NP) and Mendelian randomization to explore NTF's therapeutic targets and pathways in stroke. Stroke-related genes were sourced from public databases, and NTF's active compounds were screened using SwissADME. Summary-data-based Mendelian randomization (SMR) analysis, combined with colocalization, integrated stroke genome-wide association study data with blood expression quantitative trait loci and protein quantitative trait loci datasets to identify genes/proteins causally linked to stroke risk. Protein-protein interaction (PPI) network and drug-compound-target networks were constructed using Cytoscape and R. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses identified functional roles. Molecular docking assessed interactions between key compounds and prioritized targets. A total of 579 overlapping genes linked NTF and stroke. SMR identified 44 stroke-associated genes, with vascular endothelial growth factor A, angiotensinogen (AGT), and lipoprotein(a) replicated. Validation in Brain-eMeta supported eight of these targets, supporting tissue relevance. Enrichment analyses highlighted pathways including PPAR signaling, cholesterol metabolism, and vascular function. Core targets (Adiponectin, C1Q and collagen domain containing (ADIPOQ), Scavenger Receptor Class B Member 1 (SCARB1), and AGT) emerged from PPI networks. Molecular docking confirmed strong binding between NTF's Calycosin and AGT, a key renin-angiotensin system protein. NTF likely mitigates stroke by modulating genes involved in cholesterol metabolism and vascular regulation. The predicted Calycosin-AGT interaction provides a genetically informed hypothesis for a possible role in renin-angiotensin modulation. This integrative approach provides genetic and mechanistic insights into NTF's therapeutic efficacy.