Ganoderic acid L alleviates lung injury through PI3K/MAPKs signaling revealed by network pharmacology, molecular dynamics and transcriptomics

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Abstract

Abstract Background/Objectives: Ganoderma lucidum is widely used for inflammatory disorders, but its key active compounds and direct targets in acute lung injury (ALI) remain unclear. This study aimed to identify the major active triterpenoid from G. lucidum and clarify its mechanism against lipopolysaccharide (LPS)-induced ALI. Methods: Untargeted LC-MS was used to characterize the chemical profile of G. lucidum extract. Network pharmacology, GO/KEGG enrichment, and molecular docking were applied to predict active compounds and targets. Ganoderic acid L was evaluated in an LPS-induced ALI model by histopathology, BALF protein, inflammatory cytokines, oxidative stress assays, transcriptomics, RT-qPCR, and western blotting. Molecular dynamics, SPR, and CETSA were used to validate target binding. Results: A total of 660 compounds were identified, with terpenoids as the predominant class. Network pharmacology revealed 480 overlapping targets and highlighted PI3K-related signaling as a core regulatory node. Ganoderic acid L showed the strongest docking affinity toward PIK3CA among representative compounds. In vivo, ganoderic acid L alleviated lung histopathological injury, collagen deposition, pulmonary edema, and BALF protein leakage, reduced TNF-α, IL-6, IL-1β, MPO, and MDA, and restored SOD and GSH-Px activities. Transcriptomic and RT-qPCR analyses showed suppression of inflammatory and stress-responsive genes. Western blotting confirmed inhibition of Akt/NF-κB/MAPK signaling. SPR demonstrated direct binding to PI3K with a KD of 4.77 μM, and CETSA confirmed target engagement. Conclusions: Ganoderic acid L is a key active triterpenoid from G. lucidum that protects against LPS-induced ALI by directly targeting PI3K and suppressing downstream Akt/NF-κB/MAPK signaling.