This study aims to investigate the therapeutic effects and underlying mechanisms of astilbin on high-fat diet (HFD)-induced bone loss, focusing on its regulation of the UPP1-Mitogen-Activated Protein Kinase (MAPK) axis, metabolic reprogramming, and gut microbiota modulation. Male C57BL/6 mice were fed an HFD for 10 weeks to induce obesity and bone loss, followed by astilbin intervention at different doses (25, 50, 100 mg/kg/d) for 8 weeks. Body weight, serum biochemical parameters, bone microstructure (via micro-CT), and bone metabolism markers were assessed. Transcriptomic, 16S rRNA sequencing, and serum metabolomics analyses were performed to explore the molecular mechanisms. In vitro experiments using bone marrow-derived mesenchymal stem cells (BMSCs) were conducted to evaluate osteogenic and adipogenic differentiation under palmitic acid and astilbin treatment. Astilbin significantly reduced HFD-induced weight gain, dyslipidemia, and bone loss, as evidenced by improved bone mineral density and trabecular bone structure. It upregulated osteogenic markers (RUNX2, alkaline phosphatase) while downregulating adipogenic markers (PPAR-γ) and inflammatory signals (p38MAPK). Transcriptomic analysis revealed that astilbin restored UPP1 expression, which was downregulated in HFD mice, and modulated the MAPK signaling pathway. Metabolomic analysis showed that astilbin downregulated proinflammatory lipids (e.g., prostaglandin F2α) and upregulated anti-inflammatory metabolites (e.g., sphingolipids and 4-hydroxyindole). Gut microbiota analysis demonstrated that astilbin restored microbial diversity, reduced the Firmicutes/Bacteroidetes ratio, and suppressed proinflammatory genera while promoting beneficial bacteria. In vitro, astilbin enhanced osteogenic differentiation and inhibited adipogenic differentiation in bone marrow mesenchymal stem cells by regulating the UPP1-MAPK axis and reducing oxidative stress. Astilbin ameliorates HFD-induced bone loss by targeting the UPP1-MAPK axis, modulating lipid metabolism, reducing inflammation, and restoring gut microbiota homeostasis. These findings provide a comprehensive understanding of the multi-target mechanisms of astilbin in metabolic bone diseases and highlight its potential as a therapeutic agent for osteoporosis. The present study demonstrates significant novelty and innovation by elucidating astilbin's multi-target therapeutic mechanism in HFD-induced bone loss, integrating for the first time its regulation of the UPP1-MAPK signaling axis with metabolic reprogramming and gut microbiota modulation, which has not been previously reported. Unlike existing literature focusing on isolated pathways, this work reveals astilbin's unique capacity to simultaneously restore UPP1 expression, downregulate proinflammatory MAPK signaling, reshape gut microbiota composition (reducing Firmicutes/Bacteroidetes ratio), and modulate osteogenic-adipogenic differentiation through metabolomic regulation of sphingolipids and indole derivatives, providing a comprehensive "gut-bone axis" perspective that advances the field beyond conventional anti-inflammatory or antioxidant approaches for metabolic bone diseases.
