Screening Based on Structural and Biological Verification of Stachyose as a PPARγ ‐Modulating Ligand for the Treatment of Non‐Alcoholic Fatty Liver Disease

Peroxisome proliferator-activated receptor gamma (PPARγ) is a critical therapeutic target for metabolic disorders like non-alcoholic fatty liver disease (NAFLD). However, PPARγ full agonists such as rosiglitazone (ROSI) exhibit limited efficacy and off-target effects. Intriguingly, transcriptomic analyses revealed dynamic PPARγ expression during NAFLD progression-compensatory upregulation in early stages and downregulation in advanced disease-highlighting the need for novel modulators. This study investigates the therapeutic potential of stachyose (STA), a natural bioactive compound, in NAFLD and its mechanism of action via PPARγ modulation. Using structure-based virtual screening of 4531 natural compounds, STA was identified as a PPARγ-targeted ligand, validated by surface plasmon resonance and molecular docking. Network pharmacology and functional enrichment analyses elucidated STA's multi-target effects. In vitro and in vivo models assessed STA's impacts on lipid metabolism, inflammation, and insulin resistance. Molecular dynamics simulations and post-translational modification studies clarified STA-PPARγ interactions. STA outperformed ROSI in mitigating hepatic lipid accumulation, inflammation, and insulin resistance in both models. STA bound stably to PPARγ via residues GLU259, GLY284, PHE287, ILE341, and LEU270, with reduced PPARγ acetylation mediated by SIRT1 activation. Unlike ROSI, STA preserved PPARγ activity without inhibiting phosphorylation at Ser273, suggesting a distinct mechanism of action. STA emerges as a partial PPARγ agonist with superior efficacy and safety profiles compared to ROSI. Its dual role in enhancing fatty acid oxidation and suppressing lipogenesis, coupled with SIRT1-dependent deacetylation of PPARγ, positions STA as a promising candidate for NAFLD therapy. This study provides a mechanistic foundation for developing PPARγ-targeted interventions with reduced side effects.