Integrating network analysis and pharmacokinetics to investigate the mechanisms of Danzhi Tiaozhi Decoction in metabolic-associated fatty liver disease (MAFLD).

Ethnopharmacological Relevance: Based on ancient classics, Danzhi Tiaozhi Decoction has been successfully used to treat nonalcoholic fatty liver disease for decades. However, its therapeutic mechanisms remain unclear.

Aim Of The Study: This study aimed to investigate the effects of Danzhi Tiaozhi Decoction (DZTZD) on metabolic-associated fatty liver disease (MAFLD).

Materials And Methods: First, we identified the active ingredients of DZTZD and their potential targets in the Traditional Chinese Medicine System Pharmacology database. Using the overlapped genes, we selected the key MAFLD-associated genes, then conducted GO and KEGG pathway enrichment analyses. Furthermore, DZTZD was administered orally to rats, and their serum and liver tissues were examined for absorbed compounds using pharmacochemistry. UPLC-Q-Exactive Orbitrap/MS was used to determine the main compounds. Then, we validated the binding association of the key targets with their active compounds with AutoDock Tools and other software. Finally, the predicted hub targets were experimentally validated.

Results: We found 254 active compounds in DZTZD corresponding to 208 targets. Sixteen key genes were identified, and the enrichment analysis revealed multiple signaling pathways, including the AGE-RAGE pathway in diabetic complications and the lipid and atherosclerosis signaling pathway. Next, 160 absorbed components and metabolites were characterized in vivo, and 53 absorbed components and metabolites were characterized in liver tissue. Thirteen parent compounds were identified, including coptisine, quercetin, luteolin, and aloe-emodin. The molecular docking data demonstrated the strongest binding between the active compounds and the core proteins. Moreover, the animal experiments showed that DZTZD decreased body weight, liver weight, lipid accumulation, and ALT, AST, CRP, FFA, IL-6, PEPCK, G6P, TG, TC, and LDL-c serum levels, and increased serum HDL-c levels compared to high-fat induced rats. Besides, the RT-PCR and Western blot showed that DZTZD inhibited the SREBP1c and FAS and increased hyperlipidemia-induced CPT-1A levels. In the high-fat group, JNK phosphorylation increased, and AKT protein phosphorylation decreased, while DZTZD reversed these effects.

Conclusion: Based on the pharmacological network analysis, pharmacochemistry, and experimental validation, DZTZD can potentially improve MAFLD via the JNK/AKT pathway.