Antipyretic effects of Xiangqin Jiere granules on febrile young rats revealed by combining pharmacodynamics, metabolomics, network pharmacology, molecular biology experiments and molecular docking strategies.

Xiangqin Jiere granules (XQJRG) is a proprietary Chinese medicine treating children's colds and fevers, but its mechanism of action is unclear. The aim of this study was to explore the antipyretic mechanisms of XQJRG based on pharmacodynamics, non-targeted metabolomics, network pharmacology, molecular biology experiments, molecular docking, and molecular dynamics (MD) simulation. Firstly, the yeast-induced fever model was constructed in young rats to study antipyretic effect of XQJRG. Metabolomics and network pharmacology studies were performed to identify the key compounds, targets and pathways involved in the antipyretic of XQJRG. Subsequently, MetScape was used to jointly analyze targets from network pharmacology and metabolites from metabolomics. Finally, the key targets were validated by enzyme-linked immunosorbent assay (ELISA), and the affinity and stability of key ingredient and targets were evaluated by molecular docking and MD simulation. The animal experimental results showed that after XQJRG treatment, body temperature of febrile rats was significantly reduced, 13 metabolites were significantly modulated, and pathways of differential metabolite enrichment were mainly related to amino acid and lipid metabolism. Network pharmacology results indicated that quercetin and kaempferol were the key active components of XQJRG, TNF , AKT1 , IL6 , IL1B and PTGS2 were core targets. ELISA confirmed that XQJRG significantly reduced the plasma concentrations of IL-1β, IL-6, and TNF-α, and the hypothalamic concentrations of COX-2 and PGE2. Molecular docking demonstrated that the binding energies of kaempferol to the core targets were all below -5.0 kcal/mol. MD simulation results showed that the binding free energies of TNF-kaempferol, IL6-kaempferol, IL1B-kaempferol and PTGS2-kaempferol were -87.86 kcal/mol, -70.41 kcal/mol, -69.95 kcal/mol and -106.67 kcal/mol, respectively. In conclusion, XQJRG has antipyretic effects on yeast-induced fever in young rats, and its antipyretic mechanisms may be related to the inhibition of peripheral pyrogenic cytokines release by constituents such as kaempferol, the reduction of hypothalamic fever mediator production, and the amelioration of disturbances in amino acid and lipid metabolism.Communicated by Ramaswamy H. Sarma.