Integrating Network Pharmacology, Molecular Docking, and Experimental Validation: Andrographolide Attenuates Acute Liver Injury via the NLRP3/Caspase-1/GSDMD-Mediated Pyroptosis Pathway.

BACKGROUND/OBJECTIVES: Andrographolide (Andro), a natural diterpenoid lactone, possesses a wide range of pharmacological properties, including notable anti-inflammatory, antioxidant, antitumor, and immunomodulatory activities. Despite its acknowledged therapeutic promise, the exact protective mechanisms underlying its efficacy against acute liver injury (ALI) are still not fully understood. Consequently, determining the molecular mechanisms through which andrographolide alleviates ALI is of substantial scientific and clinical relevance.

METHODS: Andrographolide's potential targets and pharmacological mechanisms against liver injury were initially identified using network pharmacology and molecular docking. An acute liver injury (ALI) rat model was induced by intraperitoneal injection of lipopolysaccharide (LPS). The therapeutic efficacy of andrographolide in ALI was evaluated by examining liver histopathology, measuring liver function and oxidative stress markers, and quantifying pro-inflammatory cytokine levels. Meanwhile, the expression of key constituents along the NLRP3/caspase-1/GSDMD signaling axis was quantified using RT-qPCR and Western blotting. In parallel, the protective effect of andrographolide via the canonical NLRP3/caspase-1/GSDMD pyroptosis pathway was further examined in vitro using LPS-plus-ATP-stimulated rat hepatocyte BRL-3A cells.

RESULTS: Network pharmacology analysis predicted that andrographolide (Andro) protects against liver injury mainly by targeting core regulators of pyroptosis. Molecular docking simulations further indicated stable binding interactions between Andro and key proteins involved in the pyroptotic pathway, such as NLRP3, ASC, GSDMD, and CASP1. These predictions were experimentally confirmed. Andro administration notably mitigated histopathological alterations, restored serum liver function indicators, lowered pro-inflammatory cytokine levels, and alleviated oxidative stress. Importantly, Andro substantially suppressed the expression of critical mediators along the pyroptosis signaling cascade.

CONCLUSIONS: This study demonstrates that andrographolide (Andro) ameliorates acute liver injury (ALI) by specifically inhibiting the NLRP3/Caspase-1/GSDMD-mediated pyroptosis pathway. By elucidating this underlying molecular mechanism, our work highlights Andro's potential as a novel and promising therapeutic candidate for ALI.