Berberine Protects Brain Endothelial Barrier via β-Catenin: A Network Pharmacology in Pigs and In Vitro Mechanistic Analysis

Glaesserella parasuis (G. parasuis) is an important swine pathogen that causes Glässer’s disease, of which meningitis is among the most severe clinical symptoms. However, the molecular mechanisms underlying G. parasuis-induced meningitis remain poorly understood. In this study, we employed a network pharmacology-based strategy combined with in vitro experimental validation to investigate the protective effects and mechanisms of berberine against G. parasuis meningitis. Network pharmacology analysis identified CTNNB1 (encodes the β-catenin protein) as a core target among multiple candidate targets. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses revealed that the predicted targets of berberine are significantly associated with inflammatory signaling (mitogen-activated protein kinase MAPK/nuclear factor kappa B NF-κB) and stress-response pathways (forkhead box O FoxO/p53), suggesting a multifaceted protective mechanism against G. parasuis-induced blood–brain barrier (BBB) disruption. Molecular docking results combined with drug affinity responsive target stability (DARTS) assays demonstrated that berberine binds directly to β-catenin with high affinity (docking score = −7.52 kcal/mol), as evidenced by enhanced proteolytic stability of β-catenin upon berberine (2.5, 10, and 20 μg/mL) treatment. To further validate the functional relevance, we introduced zonula occludens-1 (ZO-1), a representative tight junction protein, as a functional verification indicator. Our results demonstrated that G. parasuis infection significantly downregulated the expression of both ZO-1 and β-catenin in human brain microvascular endothelial cells (hBMECs), while berberine (2.5, 10, and 20 μg/mL) treatment effectively reversed this downregulation in a concentration-dependent manner. These findings reveal that berberine exerts its protective effects against G. parasuis meningitis by targeting CTNNB1 to restore intercellular junctional integrity—including tight junctions (ZO-1) and adherens junctions (β-catenin)—thereby preserving BBB function. This study provides a mechanistic basis for the development of berberine as a promising alternative therapeutic agent against G. parasuis meningitis.