Study on the Mechanism of Rhodiola crenulata Against Exercise-induced Fatigue Based on Network Pharmacology and Metabolomics

Objective: To investigate the mechanism of Rhodiola crenulata in alleviating exercise-induced fatigue (EIF) through integrated network pharmacology, animal experiments, and metabolomics. Methods: Network pharmacology was employed to screen potential targets of active components and EIF-related disease targets. Protein-protein interaction (PPI) networks of overlapping targets were constructed, followed by GO and KEGG pathway enrichment analyses. C57BL/6N mice were divided into control, EIF model, and low, medium, and high-dose Rhodiola crenulata groups. The EIF model was established via exhaustive swimming and rotarod tests. The body weight, exhaustive swimming time, and other physiological signs were recorded. Histological examination of skeletal muscle was conducted using HE staining. The concentrations of serum lactate, muscle glycogen, and liver glycogen, as well as the levels of IL-6 and TNF-α in skeletal muscle, were measured using assay kits. Additionally, the expression levels of VEGFA and AKT1 proteins in skeletal muscle were assessed using Western Blotting and qRT-PCR. Metabolomic profiling of skeletal muscle was performed to identify differential metabolites and associated pathways, which were further integrated with network pharmacology results. Results: Network pharmacological identified 110 overlapping targets between Rhodiola crenulata and EIF. Key targets were identified as TNF, IL-6, VEGFA, and AKT1, while bioactive constituents included epicatechin, quercetin, epigallocatechin gallate and others. Compared to the model group, administration of medium and high doses of Rhodiola crenulata extremely significantly prolonged the exhaustive swimming time (P<0.001), improved skeletal muscle fiber alignment, reduced serum lactate (P<0.001), elevated glycogen levels (P<0.001), and decreased IL-6 and TNF-α levels (P<0.001). High-dose administration markedly downregulated the protein expression of VEGFA and upregulated that of AKT1 (P<0.001). Metabolomics revealed 46 differential metabolites in skeletal muscle, primarily enriched in glycerophospholipid metabolism, α-linolenic acid metabolism, linoleic acid metabolism, and arachidonic acid metabolism. Integrated analysis highlighted glycerophospholipid and arachidonic acid metabolism as pivotal pathways. Conclusion: Rhodiola crenulata alleviates EIF by ameliorating skeletal muscle pathology, promoting glycogen synthesis, reducing lactate accumulation, suppressing inflammation, and modulating lipid metabolism. This effect is potentially mediated via TNF, IL-6, VEGFA, AKT1, and the glycerophospholipid and arachidonic acid metabolism pathways.