Study on the Mechanism of Action of Qi Zhu Formula in the Treatment of Metabolic-associated Fatty Liver Disease based on Network Pharmacologyand Experimental Validation

Introduction: The aim of the study was to investigate the mechanism of Qi Zhu Formula (QZF) against Metabolic-Associated Fatty Liver Disease (MAFLD) via network pharmacology and experimental validation. Methods: Network pharmacology identified QZF components, targets, and pathways for MAFLD. Key predicted AMPK pathway targets (SREBP1C, FASN, ACC1) were validated. MAFLD was induced in rats with a 16-week high-fat/high-sugar diet. Low/medium/high QZF doses and positive control (YSF) were administered for 8 weeks. Serum parameters (liver function, lipids, glucose, cytokines, oxidative stress markers), liver histopathology (HE, Oil Red O), and hepatic mRNA/protein levels (SREBP1C, FASN, ACC1, p-AMPK) were assessed. In vitro, lipid accumulation and protein expression (p-AMPK, SREBP1C, FASN, ACC1) were measured in fatty AML12 cells treated with control/model/normal serum/QZF serum/AMPK inhibitor/ QZF serum + inhibitor. Results: Network pharmacology identified 36 QZF components, 236 targets, and 138 intersecting MAFLD targets, enriching the AMPK pathway. QZF significantly reduced liver steatosis, inflammation, necrosis, serum liver enzymes, lipids, glucose, IL-6, IL-1β, TNF-α, FFA, MDA, and increased SOD in MAFLD rats. QZF upregulated hepatic p-AMPK protein and downregulated SREBP1C, FASN, and ACC1 mRNA/protein. QZF serum reduced lipid droplets in cells, most effectively at 24h, increasing p-AMPK and decreasing SREBP1C/FASN/ACC1 protein. AMPK inhibitor abolished QZF serum's effects. Discussion: QZF's AMPK-mediated lipid suppression advances TCM mechanism validation, though unexamined pathways and compound synergies require exploration. Conclusion: QZF ameliorates MAFLD by improving serum profiles, inhibiting lipid synthesis (via AMPK activation, suppressing SREBP1C/FASN/ACC1), reducing inflammation, and attenuating liver injury. Its "multi-target-multi-pathway" action supports its potential as a novel MAFLD treatment.